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Dr. Prof. Dato‘ Deputy Vice Chancellor, Research & Innovation


Proom Promwichit Division, Masterskill University College of
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Dr. R. O. Ganjiwale HOD, Department of Pharmacognosy, I.P.E.R.
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IPER, Wardha, MH, India
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Dr. Shah Murad HOD, Pharmacology and Therapeutics, Lahore
Vol 3 / Issue 7 / July 2011
Mastoi Medical and Dental College, Lahore, Pakistan

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Vol. 03 issue 07 July 2011
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Vol. 03 issue 07 July 2011
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N.
Title
Index
Authors Page
No.

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1 Isolation and Characterization of Smiline Girija,
Lolduvin-S: A Novel Antimicrobial J.Vijayshree 4
Protein from the Ink Of Indians Quid Priyadharshini, Pandi
Loligo Duvauceli Suba K., Hariprasad
G., Raghuraman R.
2 Raj Kaushal, Nitesh
An Insight into Metal Based Anti- Kumar, Rajeev 15
Cancer Drugs Kaushal, Pamita

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Awasthi
3 Effect of Thickness and Annealing Jessy Mathew N.,
Temperatures on Structural and Rachel Oommen, C.
Morphological Properties Of (Sb2s3)1- Sanjeeviraja , Usha 24
X (Bi2s3)X Thin Films Rajalakshmi P
4 Effect of Solanum Trilobatum Extract Amarnath Kanchana,
on Erythrocyte Membrane Surface Chinnakannu 37
Charge in Aged Rats Panneerselavam

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5 Lysosomal Enzymes Jayamathi G, Vishnu
Priya V 52
6 Evaluation of Anthelmintic Activity of Shripad M.Bairagi,
Momordica Charantia Fruit Extract Ritesh S. Mantri, 57
Nitin Nema
7 Health Seeking Behavior of HIV Shaikh Mohsin, Patil
Positive Patients Attending Voluntary Rajkumar, Pathan 61

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Counseling and Testing Center – A Sameer
Gender Perspective
8 A Comparative Study on Extended Sankar Sahayaraj
Timed Get Up and Go (ETGUG) Test Muthukaruppan, 70
Between Right and Left Hemiplegics RavindraSubbanna,
Harilal
Bapurajapanicker
9 Six Minute Walk Distance in Healthy Prem.V, R.D.
“Let the science be your passion” Adults Aged Between 20-30 Years Chakravarty, 78
Karvannan H.,
Rimmi, Vinita Arya,
Manikankanna
Vol 3 / Issue 7 / July 2011 10 Sea Water Absorption Phenomenon in Rajesh Ghosh, A
Banana Fibre Reinforced Vinyl Ester Ramakrishna, 84
Composites G.Reena
11 Period-Doubling Phenomena in a G. Kandiban, V.
Simple-3D Chaotic Oscillator with a Balachandran, S. 89
Diode Pair Manimaran
12 Hourly Ozone Concentration Prediction G.Geetha 96
Using Neural Network Model
13 A Study on the Design of Micro-Lathe S.Syath Abuthakeer,
for Education And Application P.V. Mohanram, G. 100
Mohan Kumar

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Vol. 03 issue 07 July 2011
ISOLATION AND CHARACTERIZATION OF LOLDUVIN-S: A
NOVEL ANTIMICROBIAL PROTEIN FROM THE INK OF
INDIANS QUID LOLIGO DUVAUCELI
Smiline Girija1, J.Vijayshree Priyadharshini1, Pandi Suba K.1,
Hariprasad G.1, Raghuraman R.1
ijcrr 1
Department of Microbiology, Meenakshi Ammal Dental College and
Vol 03 issue 07
Hospitals, Meenakshi University, Madhuravoyal, Chennai
Category: Research
Received on:14/04/11
Revised on:29/04/11 E-mail of Corresponding Author: smilinejames25@gmail.com
Accepted on:07/05/11

ABSTRACT
Objectives: Bioactive compounds from the marine habitat have been always represented as the
greatest unexploited source of potentially active pharmaceutical agents. Squid ink, being
reported for its various therapeutic applications has not been extensively studied for its
bioactive molecules. Thus an attempt has been made in this study to isolate and characterize a
novel antimicrobial protein from the ink of Loligo duvauceli.
Methods: The fresh squids were decapitated and the ink sac was dissected to collect the ink.
Melanin free ink was obtained by ultracentrifugation. The ink was then subjected to SDS-
PAGE to analyze the proteins present in it and a new protein with a molecular weight of
approximately 60 Kda was selected as the target protein. Native PAGE was then performed to
isolate the protein in an original form and was eluted using passive gel elution method. The
eluted protein was then subjected to an enzyme linked coupled assay for the L-amino oxidase
[LAO] activity. The Km and Vmax values for the L-amino acids utilized as substrates for the
assay were determined by Lineweaver Burk Plots. The eluted protein was sterilized by syringe
filtration after which the antimicrobial potential of the protein was checked by agar well
diffusion method against the drug resistant pathogens such as ESBL, MRSA and the pathogenic
yeast C.albicans. The MIC value of the protein was determined by Microbroth dilution method.
Results: A new protein of approximately 60 Kda was successfully isolated and was
characterized to exhibit the L-amino acid oxidase activity. The protein was scored to possess a
promising antibacterial and antifungal activity against the test pathogens. The MIC value was
determined as 25 µg/ml for ESBL producing strains of E.coli and K.pneumoniae. MIC value
for the methicillin resistant S.aureus and C.albicans was deduced as 12.5 µg/ml. This new
protein exhibiting a LAO activity was further named as Lolduvin – S.
Conclusion: This study was concluded by stating that Lolduvin-S exhibits the LAO activity
and had been reported to possess a promising antimicrobial activity against the dreadful drug
resistant pathogens. Lolduvin-S could be employed in near future as a novel therapeutic agent
to treat various systemic ailments.
__________________________________________________________________________

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Vol. 03 issue 07 July 2011
Keywords: Antimicrobial, drug resistant bacteria, Staphylococcus aureus4, clinical
pathogens, Microbroth dilution method. bacterial isolates and pathogenic yeasts5,
preservative property6, antioxidant values7
INTRODUCTION and anti-retroviral activity8. The ink has
Natural bioactive substances have the least also been studied for its various bioactive
quantum of side effects when compared proteins possessing cytolytic property9,
with the synthetic products. Although most antitumour fractions10 and antimicrobial
antibiotics were derived from terrestrial potential11. With these review background
life, it is the marine world that may provide this study has been aimed at isolating and
the pharmaceutical industry with the next characterizing a new and novel protein
generation of medicines. At present, a from the ink of the Indian squid Loligo
number of marine natural products are in duvauceli and to study its antimicrobial
the market or in clinical trials. Due to the potential against the drug resistant
huge diversity of marine bioactive pathogens such as MRSA and ESBL
compounds with respect to their chemical producing strains of E.coli and
structure, mode of action and applicability, K.pneumoniae.
the extreme concern has been increased
towards the screening of marine natural METHODS
products for their biomedical potential1. Collection and dissection of squids:
In spite of incredible evolution in medicinal Fresh squids were caught from the south
field, some deadly diseases are not curable. west coast of Chennai and were decapitated
The emergence and spread of antimicrobial by the fishermen. The squid was fixed in
resistance is now threatening to undermine 4% formalin (4ml formalin in 96ml of
our ability to treat infections and save lives. water) and was identified as Loligo
Drug resistant bacteria such as extended duvauceli by an eminent Zoologist based
spectrum beta lactamase (ESBL) producing on the Zoological taxonomy. The
strains of Escherichia coli, Klebsiella dissection of the squids was performed as
pneumoniae, methicillin resistant per the instructions of the Zoologist. After
Staphylococcus aureus (MRSA) continue dissection of the ink sac the surface of the
to cause a large number of infections and gland was sterilized with ethanol and was
deaths particularly in developing blotted with sterile cotton. The ink duct
countries2. Thus a novel therapeutic was cut with sterile scissors and the sac
compound from an alternate source with was gently squeezed and the excreted ink
unique metabolic and physiological was collected in sterile brown bottles. The
capabilities producing different kinds of ink was stored at 4ºC until use.
metabolites must be employed for future
therapeutic scenario globally. Preparation of melanin free ink:
Squids under cephalopod family, is known The extracted fresh ink was subjected to
to produce a black pigmented ink as a centrifugation under cold condition at
defensive ploy against its predators. This 15,000 rpm for 30 min to remove melanin.
ink has already been reported to exhibit The supernatant was used as melanin free
various therapeutic values3. The crude ink ink. The samples were frozen and kept at -
extracts from various species of 80°C until use.
cephalopods have already been studied for
its antimicrobial potential against biofilm

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Vol. 03 issue 07 July 2011
Estimation of protein Native Polyacrylamide gel electrophoresis:
The protein estimation was done by Native – PAGE was performed without
Lowry‘s method12, a protein assay that is SDS using the same methodology as
based on the reaction where the peptide described above. Staining was not done.
bonds of proteins react with copper under The unstained gel was then carefully
alkaline conditions producing Cu+ ions that removed and the target protein band
react with the Folin‘s reagent producing portion corresponding to target protein
blue colour. band size was excised in sterile phosphate
buffer and was eluted as follows.
SDS Polyacrylamide gel electrophoresis: Passive gel elution of the target protein:
SDS-PAGE was carried out using an Elution of the protein was performed by
acrylamide concentration of 7.5% in the passively eluting the proteins from
resolving gel13. Stock solutions were polyacrylamide gel pieces15. The excised
prepared as recommended14 and stored in gel pieces corresponding to the target
brown bottles at 4◦C. The separating gel protein band were placed in a clean
mix (12%) was prepared in a 25ml conical microcentrifuge tubes. The elution buffer
flask using acrylamide monomer 2.8ml, was prepared using Tris HCl 50 mM, NaCl
separating gel buffer (4x) 1.750 ml, 10 % 150 mM and EDTA 0.1 mM (pH 7.5). 0.5-
SDS 0.070 ml, 10% APS 0.035 μl and 1ml of cold elution buffer is added so that
distilled water 2.345 ml (Final volume – the gel pieces were completely immersed.
7ml). The solution was de-gased for 5 min The gel pieces were crushed using a sterile
and 3 μl of TEMED was added with gentle scalpel and was incubated in a rotary
mixing and was poured into the glass shaker at 30ºC overnight. Centrifugation
plates. 100 μl of N-butanol was added over was done at 5000-10000g for 10 minutes
the gel mix and the arrangement was left and the supernatant was carefully pipetted
undisturbed for 20 min. The stacking gel into a new microcentrifuge tube. An aliquot
mix (5%) was prepared by mixing of the supernatant might be tested for the
acrylamide monomer 0.333 ml, 4x presence of the same protein by subjecting
Stacking Gel Buffer 0.500 ml, 10 % SDS it to SDS-PAGE.
0.020 ml, 10% APS 0.010 μl and distilled Enzymatic assay for L – amino acid
water 1.137 ml (Final volume – 2.0 ml). oxidase [LAO] activity:
The electrophoretic run was performed as LAO activity of the protein was performed
per standard protocols by adding the by an enzyme coupled assay16. Purified
samples to the wells along with a molecular protein was prepared for the assay in 50
weight marker. Staining was done using mM/L phosphate buffer and 150 mM/L
Coomasie brilliant blue and destaining was KCl. The reaction mixture was prepared as
done with ethanol and acetic acid. The gel follows: Tris HCl 0.1 mM/L (pH 7.6),
was then analysed in the documentation Horseradish peroxide 10 μg, O-dianisidine
unit. The molecular weight of the unknown 0.2 mM/L and L-amino acids 2 mM/L. The
peptides was thus determined by test sample was mixed with 100 μl of the
comparing it with the standard molecular reaction mixture. Reaction was performed
weight marker bands. at room temperature for 1-60 min. The
activity was monitored by absorbance at
436 nm. The increase in absorbance was
transformed into molar concentration of the

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Vol. 03 issue 07 July 2011
end product o-dianisidine (8.31 x 103 37 C for 24 hrs for sterility checking. A 50
mol/L). The Km and Vmax values of the L- μl volume of the eluted protein in
amino acids were determined by phosphate buffered saline was propelled
Lineweaver-Burk plots. directly into the wells of the inoculated
Antimicrobial bioassay: specific media agar plates for each test
The antimicrobial assay was performed by organism. The plates were then allowed to
Nathan‘s agar well diffusion method17 with stand for 10 minutes for diffusion of the
the drug resistant strains that included the protein to take place and incubated at 37ºC
extended spectrum beta lactamase (ESBL) for 24 hrs. Sterile DMSO served as
producing strains of E.coli and Negative control. Erythromycin 30 µg (for
K.pneumoniae, methicillin resistant bacteria) and Amphotericin B 100 U (for
S.aureus (MRSA) and Amphotericin B yeast) were included as positive controls.
resistant C.albicans isolated from various After incubation the plates were observed
clinical specimens from the patients for the zone of inhibition around the wells
visiting the outpatient unit of Meenakshi and the zone size was measured using an
General Hospital (Data not given). The antibiotic sensitivity measuring scale
control strains include E.coli (ATCC (Himedia). The antimicrobial efficacy was
25922), K.pneumoniae (ATCC 10031), graded based on the zone diameter as high
S.aureus (ATCC 25923) and C.albicans activity (> 15 mm), moderately active (10-
(ATCC 10231). 14 mm), trace activity (5-9 mm) and no
A minimum of four colonies of the test activity (< 4 mm)18.
organisms were touched with a sterile loop Determination of MIC value for the protein
and transferred into Sterile Mueller Hinton MIC value for the purified protein was
broth and C.albicans into Saboraud‘s determined by Microbroth dilution
dextrose broth under aseptic conditions and method19. Serial dilutions of the eluted
was incubated for two hours at 37ºC. After protein were done in a 96 well microtitre
incubation the density of each microbial plate with sterile phosphate buffer. The
suspension was adjusted equal to that of dilution factor was 5, 2.5, 1.25, 0.625,
106 c.f.u/ml (standardized by 0.5 0.312 and 0.156 μg/100 μl. To each
MacFarland standards) and was used as the dilution 100 μl of the culture broths of the
inoculum. test strains and control strains were added
in their respective wells and the plate was
Agar well diffusion assay incubated at 37 C for 24 hrs. After
Fifty microlitres (50μl) of inoculum of each incubation the spectrophotometric analysis
test and control organism was spread as was performed and the OD values were
lawn cultures onto sterile Mueller Hinton recorded. The MIC value was also
agar plates using L-rods to achieve a confirmed with Microbial Spot Checker
confluent growth. The agar plates were board method20 where 3 μl of each dilution
allowed to dry and wells or cups of 8 mm was spotted onto Mueller Hinton agar
were made with a sterile agar borer in the plates and incubated at 37 C for 24 hrs.
inoculated agar plates. The eluted protein After incubation the spot showing the
was subjected to syringe membrane complete absence of microbial growth
filtration prior to the bioassay and an indicates the minimum inhibitory
aliquot of the same was streaked onto concentration value.
nutrient agar plates and was incubated at

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Vol. 03 issue 07 July 2011
RESULTS value of 20 mm for E.coli (ESBL), 21 mm
Centrifugation of the crude ink at 15,000 for K.pneumoniae (ESBL), 22 mm for
rpm for 15 min yielded melanin free ink. S.aureus (MRSA) and 20 mm for
Using Lowry‘s method, a standard curve of C.albicans. The zone size was compared
absorbance was obtained as a function of with the control strains (Figure 3). The
initial protein concentration and it was used MIC value was determined as 25 µg/ml for
to determine the unknown protein E.coli and K.pneumoniae, 12.5 µg/ml for
concentration. Thus the total protein in the S.aureus and C.albicans. The previous
melanin free ink was determined as 65.5 dilution that showed the visible decrease in
mg/ml. the number of colonies was determined as
the bacteriostatic dose and was deduced as
SDS PAGE showed a major protein band 12.5 µg/ml for E.coli and K.pneumoniae,
with a molecular weight of 60 KDa. Two 6.25 µg/ml for S.aureus and C.albicans.
weak bands were also obtained and their Thus a novel protein called Lolduvin-S has
molecular weights were determined as 100 been isolated and characterized to possess a
KDa and 80 KDa (Figure 1). The 60 Kda potent LAO activity from the ink of Loligo
band was the target protein of the study as duvauceli. The protein had also a
it has not been reported earlier. The protein promising antimicrobial activity against the
was successfully isolated by Native Gel dreadful resistant pathogens like ESBL and
Electrophoresis (Figure 2) and was eluted MRSA organisms.
by passive gel elution technique.
DISCUSSION
L-amino acid oxidase (LAO) assay The marine ecosystem offers rich niche for
performed showed that when the purified marine flora and fauna. The protein
protein and 2 mM/L amino acids were structures, metabolic pathways,
incubated for 1 min, L-lysine and L- reproductive systems, sensory and defense
Arginine proved to serve as excellent LAO mechanisms developed by marine
substrates. Km and Vmax values of lysine microorganisms and other lifeforms are
were 33-87 µmol and 1.93-2.71 µmol/sec distinctive in nature. During the last few
respectively and Km and Vmax values for decades several novel compounds in point
arginine is 25-125 µmol and 1.56-3.30 of view of potential drug development have
µmol/sec. The reactions were completed been isolated from marine habitat with
within 30 secs at room temperature for various therapeutic applications21. The
lysine and arginine at a concentration of antimicrobial property of the squid ink is
0.02-2 mM/L. Thus in this study we report already a known fact and the ink has been
a novel protein with LAO activity from the found to posses a promising antimicrobial
ink of Loligo duvauceli which has been effect in our previous studies. The proteins
named as Lolduvin-S. present in various species of gastropods
The protein after membrane filtration and cephalopods have been characterized
yielded no growth on nutrient agar plate; earlier22. This study is thus undertaken to
thus it was sterile and was used for characterize a novel antimicrobial protein
bioactivity assays. Lolduvin-S had scored from the ink of Loligo duvauceli. The
to possess high antibacterial activity melanin free ink has been used in various
against the drug resistant test strains (Table studies for the analysis of the protein and it
1). The zone size ranges from a mean holds suitable for various protein analysis.

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Vol. 03 issue 07 July 2011
The total protein estimated in this study been reported from the ink of a sea hare
from the squid ink suggests the availability under the Phylum Mollusca earlier27.
of the bioactive protein for various Escapin is known for its L-amino oxidase
pharmaceutical applications. activity which is a hydrogen peroxide
induced destruction of the target cells (such
Determination of the molecular weight of as micro-organisms) in the presence of L-
the proteins is best achieved by the amino acids. With this view an attempt to
bioinstrumentational tool such as the SDS check the LAO activity of Lolduvin-S
PAGE. The electrophoretic separation of protein is done by Enzyme linked coupled
the proteins from other species of squid has assay. Lolduvin-S has been found to
been achieved with 2.5% and 7.5% gel possess an enzymatic L-amino acid oxidase
resolution23. In this study we employed activity. Hydrogen peroxide is sufficient to
12.5% gel that is suitable for the separation cause bacteriostasis and does so with a
of the protein from Loligo duvauceli‘s ink. concentration dependency and threshold
Apart from the major protein band at 60 similar to that of amino acids as substrates
Kda, two other weak bands at 100 Kda and for the enzymatic activity. Both lysine and
80 Kda have also been observed. This arginine are substrates for LAO activity
shows the presence of tyrosinase24 and from the squid ink. It shows strong and
peroxidase25 respectively which has been rapid activity in this study when using
already reported from the ink of various arginine or lysine as substrates, with
cephalopods and squids. The 60 Kda reactions being completed within 30 secs at
protein in Loligo duvauceli has not been room temperature.
reported anywhere earlier and thus it has
been considered as the target protein. A similar protein has been reported to be
bacteriostatic and bactericidal from the
Native PAGE separates the protein in a purple gland of the sea hare Aplysia
pure form without denaturation though dactylomela28 and the substrates determine
effective and high amount of proteins can the bactericidal or bacteriostatic effects.
be obtained with gel filtration Another protein called achacin possesses a
26
chromatography methods . Due to high bacteriostatic effect that seems to be
cost, gel filtration chromatography is not mediated through its oxidation of the L–
done for the isolation of the proteins: thus amino acids lysine and/or arginine and the
the passive gel elution protocol suggested consequent subsequent production of
by the Pierce technologies, USA is hydrogen peroxide29. Another antimicrobial
followed for the same. It is a simple and protein has been named as escapin from the
rapid protocol for the elution of target Aplysia sp., which has been cloned and
protein. The elution protocol is slightly characterized. Thus this study reports the
modified by performing the elution under new protein Lolduvin-S with a potent LAO
cold condition. The protein eluted is activity from melanin free ink of Loligo
rechecked by subjecting the protein for duvauceli.
SDS-PAGE and by obtaining the same
band at 60 Kda. Lolduvin-S from the melanin free ink
sample has been found to be more effective
A similar type of protein called Escapin against the Gram positive bacteria, gram
with a molecular weight of 60 Kda has negative bacteria and the pathogenic yeast.

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Vol. 03 issue 07 July 2011
In an earlier study, a protein in low Lolduvin-S from Loligo duvauceli‘s ink
concentration with potent L-amino acid possesses a potent antifungal activity.
oxidase activity showing a broad
antibacterial activity against Gram-positive In conclusion, Lolduvin–S, a new and
and Gram-negative bacteria by H2O2 novel protein from the ink of the Indian
generation has been reported from the skin squid Loligo duvauceli with a potent
mucous of the rock fish30. In correlation antimicrobial and LAO activity, can be
with this, the MIC value is determined at considered in near future as a valuable
low concentration range of 6.25µg/ml. A pharmaceutical antimicrobial agent from
similar protein has also been isolated from the marine habitat. However further
two purple pigmented bacteria from characterization and structural analysis of
Mediterranean sea whose enzymatic the same is under progress to acquire a
activity is potent in generating the detailed understanding of this novel
hydrogen peroxide in the presence of L- protein. Lolduvin-S being found to be
amino acids31. A BLAST search suggests bactericidal against the ESBL and MRSA
that the cDNA encoded a novel pathogens, it can definitely aid in the
antibacterial protein sharing identity with a control and emergence of drug resistant
number of L-amino acid oxidases is strains globally.
effective against MRSA strain32. In
correlation with these reports, this study ACKNOWLEDGEMENT
states that the new protein Lolduvin-S from We are thankful to Dr.Mehar
Loligo duvauceli‘s ink possess a promising Sultana.,M.Sc.,M.Phil.,Ph.D., Professor,
antibacterial property against the drug Department of Zoology, Presidency
resistant pathogens. College, Chennai, Tamilnadu for speciating
the squid selected for our study.
The bioactive protein Lolduvin-S also
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2003; 308: 293-299. 19. Mcginnis, Ronaldi MG, Lorian V,
10. Takaya Y, Uchisawa H, Matsue H, Williams, Wilkins. Antibiotics in
Okusaki B. An investigation of the laboratory medicine (4th Edition),
antitumour peptide form squid ink. Baltimore 1996; 176-211.
Biol Pharm Bull 1994; 17: 846-849. 20. Nkere CK, Iroegbu C. Antibacterial
11. Charles Derby, Hsiuchin Yang, Paul screening of the root, seed and
Micah, Ko-Chun Ko & Phang. stembark extracts of Picralima nitida.
Cloning, characterization and Afri J of Biotech 2005; 4(6): 522-526.
expression of escapin, a broadly 21. Epel D. Frontiers in squid production:
antimicrobial FAD – containing L- prospecting for new antibiotics. Marine
amino acid Oxidase from ink of the sea natural products. Pub No CSG-MP-02-
hare Aplysia californica. J of Expe Biol 001. (Sea grant, California), 2002.
2005; 208: 3609-3622. 22. Kirsten & Benkendorff. 2005.
12. Lowry OH, Rosebrough NJ, Farr AL, Molluscan biological and chemical
Randall RJ. Estimation of proteins. J of diversity: secondary metabolites and
Biol Chem1951; 193: 265-266. medicinal resources produced by
13. Nagashima Y, Kikuchi N, Shimakura marine molluscs. Bio Rev. 7: 917-921.
K, Shiomi K. Purification and 23. Anna Palumbo, Gabriella Fiore,
characterization of an antibacterial Annarita Poli, Anna Di Cosmo, Marco

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d‘Ischia. Dopamine in the ink defence mucus of rockfish Sebastes schlegelii.
system of Sepia officinalis: Comparitive Biochem & Physiol 2008;
biosynthesis, vesicular 149(2): 394-400.
compartmentation in mature ink gland 31. Daniel Gomez, Elena Espinosa,
cells, nitric oxide (NO)/cGMP-induced Marcelo Bertazzo, Patricia Lucas,
depetion and fate in secreted ink. Francisco Solano, Antonio Sanchez.
Biochem J 2004; 378: 785-791. The macromolecule with antimicrobial
24. Gian Luigi, Elio Nisco, Gabriella, activity synthesized by
Paola, Anna Palumbo. Toxicity of Pseudoalteromonas luteoviolacea
melanin free ink of Sepia officinalis to strains is an L-amino acid oxidase.
transformed cell lines: identification of Appl Microbio and Biotec 2007; 79(6):
the active factor as tyrosinase. 925-930.
Biochem & Biophy Resear Comm 32. Kosuke Kasai, Takashi Ishikawa,
2003; 308: 293-299. Takafumi Komata, Kaori Fukuchi &
25. Ida Gesualdo, Franscesco, Margherita. Mitsuru Chiba. Novel L-amino acid
Molecular cloning of a peroxidase oxidase with antibacterial activity
mRNA specifically expressed in the against methicillin-resistant
ink gland of Sepia officinalis. Staphylococcus aureus isolated from
BioChemica et Biophysica Acta 1997; epidermal mucus of the flounder
1353: 111-117. Platichthys stellatus. FEBS J 2009;
26. Wang JF, Lei M, Wang YM, Pang L, 277(2) 453–465.
Xu W, Xue CH. Study of the radio- 33. Gomes VM, Carvalho AO, Da Cunha
protective effect of cuttlefish ink on M, Keller MN, Bloch C Jr, Deolindo P,
hemopoietic injury. Asia Pac J Clin Alves EW. Purification and
Nutr 2007; 16: 239-243. characterization of a novel peptide with
27. Johnson PM, Yang H, Tai PC, Derby antifungal and antiviral activity from
CD. Escapin: an antipredator protein in Bothrops jararaca venom. Toxicon
the defensive secretion of Aplysia 2005; 45(7): 817-827.
californica. Chem Senses 2003; 28: 34. Ryosuke Iijima, Jun Kisugi &
A24 (abstract). Masatoshi Yamazaki. A novel
28. Melo V, Duarte A, Carvalho A, Siebra antimicrobial peptide from the sea hare
E , Carvalho. Purification of a novel Dolabella auricularia. Develop &
antibacterial and haemagglutinating Comp Immunol 2003; 27(4): 305-311.
protein from the purple gland of the 35. Mitsuru Jimbo, Fumie Nakanishi,
sea hare Aplysia dactylomela. Toxicon Ryuichi Sakai, Koji Muramoto, Hisao
2000; 38: 1415-1427. Kamiya. Characterization of L-amino
29. Ehara T, Kitajima S, Kanzawa N, acid oxidase and antimicrobial activity
Tsuchiya T. Antimicrobial action of of Aplysianin-A, a sea hare-derived
achacin is mediated by L-amino acid antitumor-antimicrobial protein.
acid oxidase activity. FEBS Lett 2002; Fisheries Sci 2003; 69(6): 1240–1246.
531: 509-512. 36. Andre Gustavo, Marcia Souza,
30. Yoichiro, Nobuyo, GuoHua, Shoichiro, Carvalho Melhem, Frederico Oliveira
Kuniyoshi, Kazuo Shiomi, Yuji Prado, Gabriela, Roberto Mitsuyoshi.
Nagashima. Antibacterial action of L- Amphibian secretions for drug
amino acid oxidase from the skin discovery studies: a search for new

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Vol. 03 issue 07 July 2011
antiparasitic and antifungal acid oxidases from several marine
compounds. Letters in Drug Design & phytoplanktons. Marine Eco Prog
Discovery 2007; 4: 67-73. Series 2000; 59: 195-201.
37. Brian Palenik, Francois MM. Morel
Comparison of cell-surface L-amino

Table 1: Antimicrobial potential of Lolduvin-S protein isolated from the melanin free ink
of the Indian squid Loligo duvauceli

S.No Strains used for the study Zone size in mm MIC Value
[μg/ml]
REFERENCE PATHOGENS [ATCC]
E.coli[ATCC 25922] 21 6.25
K.pneumoniae [ATCC 10031] 20 6.25
S.aureus [ATCC 25923] 24 3.125
C.albicans [ATCC 10231] 23 3.125
TEST PATHOGENS
E.coli[ESBL] 20 25
K.pneumoniae[ESBL] 21 25
S.aureus [MRSA] 22 12.5
C.albicans [Germ tube positive, Amphotericin 20 12.5
B resistant]

Figure 1: SDS-PAGE gel show ing a prominent band (60 KDa) and tw o
w eak bands (100 KDa and 80 KDa)

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Vol. 03 issue 07 July 2011
Figure 2: 60 KDa protein band on NATIVE gel

Figure 3: Lolduvin-S protein showing the zone of inhibition against the test and control
pathogens by agar well diffusion method

E.coli K.pneumoniae

S.aureus C.albicans

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Vol. 03 issue 07 July 2011
AN INSIGHT INTO METAL BASED ANTI-CANCER
DRUGS

Raj Kaushal1, Nitesh Kumar1, Rajeev Kaushal2, Pamita Awasthi1


1
ijcrr 2
Department of Chemistry, National Institute of Technology, Hamirpur, H.P.
Department of Experimental Medicine & Biotechnology, PGIMER,
Vol 03 issue 07
Chandigarh
Category: Review
Received on:22/04/11
Email of Corresponding Author: kaushalraj7823@rediffmail.com
Revised on:02/05/11
Accepted on:09/05/11
____________________________________________________________________

ABSTRACT
Metal complexes play critical role in the treatment of cancer. Cis-platin which is model anti-
cancer drug has been used in the treatment of various types of cancer. But due to its side effects
and resistance phenomenon efforts have been made to explore the possibility of synthesizing
novel non-platinum based anti-cancer drugs. In addition to platinum based drugs, complexes of
other transition metals like titanium, ruthenium, palladium and gold etc. also show pronounced
anti-cancer activity. The Complexes with titanium and ruthenium have already been evaluated
in phase I and phase II clinical trials. Some transition metal complexes show good anticancer
activity against cis-platin resistant cell lines. This review will provide an insight into various
platinum as well as non-platinum based anti-cancer drugs.
______________________________________________________________________

Key words: Transition metal, cis-platin, electron rich biological components like
Nucleic acid, protein, platinum, titanium, proteins and nucleic acid because metal
ruthenium, palladium, gold, anti-cancer, centers are positively charged and favored
drug, amines, biological activity. to attack on negatively charged
3-6
biomolecules . Various transition metals
INTRODUCTION such as platinum(Pt), titanium(Ti),
Metals play an important role in our daily ruthenium(Ru), rhodium(Rh), iridium(Ir),
life due to their incorporation in our diet in molybdenum(Mo), copper(Cu) and
varying quantities1,2. Due to potential gold(Au) in their complex form are
pharmacological applications of transition effective against solid tumors in animals
metal based complexes such as anti- and human beings. The first metal complex
diabetic, anti-neurological, anti-bacterial, discovered to exhibit anti-cancer activity
anti-fungal, anti-cancer agents, metal was cis-platin (cis-
complexes have been used in medicinal diaminedichloroplatinum(II)). This drug is
chemistry since sixteenth century. considered best for treatment of certain
Transition metals have tendency to form types of cancers but due to its toxicity, its
variety of complexes due to the presence of utilization has been limited at broad range7-
9
vacant d-orbitals in their valence shell. . In coming era, interest has been growing
They can take part in various biological in developing non-platinum based anti-
processes which show their interaction with cancer drugs due to their less toxicity.
15 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
Also, non-platinum compounds may various types of cancers such as testicular,
provide different oxidation states, ovarian, lung, neck, and head cancers. This
coordination geometries, and affinities for metal complex used in the treatment of
certain types of biological ligands10. It has various cancerous malignancies and is one
been established that ligands having O, N, of the best-selling anti-cancer drug all over
S in their stem showed increased biological the world. Cis-platin has several
activity due to increase in coordination disadvantages some of which may include
capacity11,12. It has been reported in that by treating the cells with cis-platin,
literature that due to the symmetry of necrotic and apoptic cell death occur
ligand(s) uptake of drug by cancerous cells simultanesly. Also, it has limited solubility
has been increased13. The necessary in water hence it is given intravenously to
conditions for a complex to have anti reduce the harm to the kidney. Other side
cancer activities are as i) Complex should effects of using cis-platin are emesis,
be neutral so that it can diffuse through the nausea, vomiting, nephrotoxicity,
hydrophobic cell membrane. ii) Complex neurotoxicity, myelosuppression, oto-
should have square planer structure i.e. toxicity14. Also only a limited number of
leaving group should be at cis-position. iii) tumors can be treated with platinum based
Leaving groups should be labile, so that drugs. In addition to cis-platin many other
they can be easily substituted. iv) Groups platinum based drugs (Carboplatin,
which are not substituted should have low Oxaliplatin, nedaplatin and lobaplatin)
trans effect like NH3, heterocyclic amines passed for current tumor therapy1. The new
or diamines14. Amine ligands influence the platinum complexes of the formula [Pt(2,2'-
anti-cancer property, because non leaving bipyridine)amino acid]n+ where n =1-2 and
amine ligands are the reason for anti-cancer amino acid is an anion of L-histidine, L-
property15. Recently, some non metallocene Lysine, L-asparagine, L-tryptophan, or L-
titanium complexes having oxygen based tyrosine, had been prepared by interaction
ligands have been synthesized and it has of [Pt(2,2'-bipyridine)Cl2] and an
been established that ligand lability is not appropriate amino acid (sodium salt) in
essential to show anti-tumor activity16. water or water-methanol mixture which
are highly negatively charged molecules. In
Platinum Complexes as anti cancer case of Histidine the Platinum atom binds
agents: with –NH2 group of Histidine and in case
The first metal based anticancer drug of other amino acids Platinum binds by
discovered was Cis- NH2 and COO- groups and these complexes
diaminedichloroplatinum(II) (cis-platin) by were used against P-388 Lymphocytic
Rosenberg et al8,9.Cis-platin acts by leukemic cells18. An octahedral complex of
interacting with DNA (Deoxy ribo nucleic Platinum(IV) with adamantylamine of
acid) via cross linking with two adjacent composition bis(acetato)(1-
guanine molecules, followed by the adamantylamine)amminedichloro
replacement of two chloride groups by platinum(IV) had been prepared and
water molecules and form aquated cis- showed resistance factor 2.84 fold lower
platin which stops the replication of DNA than cis-platin because adamantylamine is
and obstruct the cell growth which is the a bulky hydrophobic ligand and the use
ultimate aim of anti- cancer drugs14. Cis- increase the uptake of compound by the
platin has been used in the treatment of cancerous cells and able to overcome the

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Vol. 03 issue 07 July 2011
acquired cis-platin resistance13. Pt(IV) [Au(cyclam)](ClO4) 2Cl, [Au(terpy)Cl]Cl2
complex with adamantylamine penetrate as (terpy=terpyridine), and [Au(phen)Cl2]Cl
a whole complex inside the cell membrane. (phen=phenanthroline) were characterized
It may be due to hydrophobicity of ligand. in solid state and in solution. These
The symmetry of hydrophobic compounds of gold were tested on human
adamntylamine ligand lightens the ovarian cell line A2780,which were either
penetration of whole complex inside the sensitive or resistance to cis-platin21. A new
cell membrane. The penetration of Pt(IV) compound Chloro-glycylhistidinate
complex with adamantylamine had been gold(III) (GHAu) had good biological
improved and facilitates transport across property and tested for cytotoxic properties
cell membrane. This hydrophobic ligand in vitro against MOLT-4 (human leukemia)
enhances accumulation inside cancer cell and C2C12 (mouse tumor) cell lines22,23.
and trigger rapid cell death in both cis- Nowadays, gold(III) compounds are good
platin sensitive and cis-platin resistant cell cytotoxic agents. Plenty of gold(III)
lines15. The Pt(II) Complexes bearing compounds were characterized and
pyridine carboxyldimines containing bulky synthesized in the last fifteen years24. Gold
aromatic groups examined for their complexes containing bipyridine ligands of
potential cytotoxicites against human general formula [Au(NΛN)Cl2]PF6 where
ovarian carcinoma and cis-platin resistant (NΛN) = 2,2'-bipyridine, 4,4'-dimethyl
cell line19. A series of trans-platinum(IV) bipyridine, 4,4'-dimethoxy-2,2'-bipyridine,
complexes with functionalized aromatic 4,4'-diamino-2,2'-bipyridine showed
carboxylate ligands cis, cis, trans- moderate to good cytotoxicity in vitro
Pt(NH3)2Cl2(CO2C6H4R)2 where R may be towards human ovarian carcinoma cell line
H, p-vinyl, p-methoxy, p-iodo, p-cyano, p- and cis-platin resistance variant25.
carboxyl had been synthesized and due to
presence of aryl groups uptake of drugs had RUTHENIUM COMPLEXES:
been improved and facilitate transport Ruthenium complexes have attracted much
across cell membrane. These complexes attention as building blocks for new
were evaluated for cellular uptake and transition metal based anti-tumor agents.
inhibition of cell proliferation against a Ruthenium compounds offer the potential
panel of lung, colon and breast carcinoma over anti-tumor platinum(II) complexes
cell lines20. currently used in the clinic because of
reduced toxicity, a novel mechanism of
Gold Compounds having anti-cancer action and the prospect of non cross
activities: resistance26-29. Organo ruthenium
The interaction of cytotoxic gold(III) complexes due to presence of lipophilic
compounds with DNA is weak than that of arene can interact better than that of cis-
platinum analogues but gold(III) complexes platin inside the cell, by causing chlorine
have good interaction with model proteins dissociation which is an important factor
and target proteins. The mode of action of for cell death. Ruthenium(III) complexes of
gold(III) compounds is significantly general formula [Ru(η6-
different than that of cis-platin. Some arene)Cl2(NC5H4OOC-C5H4 FeC5H5)]
compounds like [Au(en2)]Cl3 (en=ethylene where arene may be C6H6, C6H5Me, p-
diamine), [Au(dien)Cl]Cl2 PrC6H4Me, and C6Me6 and of formula
(dien=diethylene diamine), [Ru(η6-arene)Cl2]2 (NC5H4OOC-

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Vol. 03 issue 07 July 2011
C5H4FeC5H4-COOC5H4N), where arene=p- trans-PdCl2L2(where L=3-
PrC6H4Me and C6Me6 were cytotoxic hydroxypyridine, 2-hydroxypyridine, 4-
against A2780 and cis-platin resistant hydroxypyridine) had been synthesized and
human ovarian carcinoma cell lines30. It it had been found that the Solubility,
had been studied that the interaction of reactivity, electronic and steric properties
[(η6-p-cymene)Ru(ATSC)Cl)]PF6 complex can be modified by varying the geometry
(where ATSC=9-Anthraldehyde and ligands around the metal center. Out of
thiosemicarbazone ) with human serum these the compound of 2-Hydroxy pyridine
albumin protein can result into anti-cancer was found to be most active against A2780,
activity as biological activity is not always A2780cisR and A2780ZD0473R ovarian cancer
related to their DNA binding ability. So, cell lines. It had been found that both metal
protein could be the biological target of and ligand take part in biological activity of
these compounds31. The four cationic the complex but due to the rapid hydrolysis
ruthenium(II) complexes with formula of palladium complexes (105 faster that
[Ru(η5-C5H5)(pph3)2]+ with L=5-phenyl-1- Platinum analogues), they dissociate easily
H-tetrazole (TzH), imidazole (ImH) , benzo before reaching their pharmacological
[1,2-b;4,3-b'] dithio-phen-2-carbonitrile target35. The platinum(II) and palladium(II)
(Bzt), and [5-(2-thio phen-2-yl)-vinyl] complexes of 2,2'-bipyridine (bipy) with
thiophene-2-carbonitrile] (Tvt) were ethyl dithio carbamate (Et-dtc) in which
evaluated as anti-tumor agents. Out of the di thio carbamate ligand coordinate
these the first three compounds show much with pt(II) or pd(II) center as bidentate
higher toxicity than cis-platin against with two sulphur atoms were water soluble
human leukemic cancer cells (HL-60 and were tested for their in vitro anti-
cells)32. The advantages in using ions of tumor activity against chronic myllogenous
transition metals other than platinum leukemia cell line K562. But these
include the availability of additional complexes show cytotoxic concentration
coordination sites in octahedral complexes (Cc50) values lower than that of cis-platin.
and altered shape of the complex, changes The mode of interaction of these complexes
in ligand affinity and substitution kinetics, were investigated by circular-dichromism,
change in oxidation state, and ultraviolet difference and flouroscence
33
photodynamic approaches to therapy . spectroscopy. The interaction of Pd(II)
complex with DNA and its anti- tumor
activity against K562 is more than that of
Palladium Complexes: its Pt(II) analog36. The series of palladium
The marginal anti-tumor activities of the complexes with Salicyldamine thio
palladium complexes were explained on semicarbazone having formula [pd(salt
the basis of fast reactivities of leaving scR)PPh3], {H2Salt scR = Salicyldehyde
groups as the reactivity of palladium(II) thio semicarbazone R=H, 3-tert butyl, 3-
complexes is much higher as compared to methoxy, 5-chloro} were prepared by
platinum(II) complexes e.g. the reactivity reaction of appropriate salicyldamine
of palladium(II) complexes having 1,2- thiosemicarbazone with Pd(PPh3)2 in which
Diaminocyclohexne and dicarboxylate thiosemicarbazone coordinate to the
ligands was 105 times more than that of palladium in a tridentate manner that is
platinum(II) complexes having similar through phenolic oxygen, imine nitrogen
ligands34. The Pd(II) complexes of the form and thionate sulphur forming five or six

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Vol. 03 issue 07 July 2011
membered chelate rings with in the ascites and solid tumor. This drug had
structure and the fourth coordination site is maximum tolerable dose of 230 mg on two
occupied by PPh3. The biological activity week schedule with side effects of cardiac
of thiosemicarbazone ligand and palladium arrhythmia. It had been reported that doses
complexes were investigated towards higher than maximum tolerable result in
WHC01 oesophageal cancer cell line and liver and kidney toxicity52. Erich Dubler
against two strains of malarial parasite had synthesized and crystallized di-chloro
plasmodium Falciparum W2 derivatives of budotitane and found that
37
(Chloroquinone resistant) . Another anti-tumor activity appear due to
palladium(II) compound with 5-methyl unsubstituted phenyl rings, if phenyl rings
uracil of the general formula PdL2Cl2 get replaced by methyl groups, activity
where L= 5-methyl uracil was prepared by totally disappears53. The first metallocene
Anshu Srivastava et. al. and it had been i.e. titanocene dichloride show anti-tumor
found that this light brown compound was activity against colon, lung and breast
hygroscopic and had thermal stability up to cancer although the mechanism of
260˚C with anti-tumor activity38. cytotoxicity is not clear yet. This complex
also exhibits antiproliferative activity
Titanium based anti-cancer drugs: against solid ascite tumor. Titanocene
In the last few years there has been dichloride show anti-tumor activity against
growing interest in developing non- doxorubicin and cis-platin resistant ovarian
platinum based anti-cancer agents due to carcinoma cells and also less toxic effect
their pronounced biological activity39-51. than cis-platin. The advantage of this
After the discovery of cis-platin, the first complex is that no evidence of
non-platinum anti-cancer drugs were nephrotoxicity or myolotoxicity had been
budotitane and titanocene dichloride which reported. It has been found in literature that
are titanium based anti cancer drugs. Since studies on chemistry of titanium as anti
titanium is second most abundant transition tumor agent are more limited52. Titanocene
metal and ninth of all the elements on earth dichloride are proved to be superior
and pure titanium and titanium alloy are compounds of its derivatives since in
widely used for orthopedic and dental addition to anti-tumor properties titanocene
implants. Titanium is present in many dichloride exhibits anti-viral54, anti-
biomaterials such as food in the form of arithmetic, and anti-inflammatory
whitening pigment. So it may incorporated 55
activities . This compound exhibits higher
in drugs and in to living systems with low toxicity than cis-platin, doxorubicin,
toxicity10. Also Ti(IV) is an oxophillic mitoxantrone and vinblastine in human
metal and form strong bond with acidic renal cell carcinoma. The other halides or
DNA as well as other biological molecules. pseudohalides of Cp2TiX2 (where X=F,
Titanium as a metal posseses a wide
Br, I, NCS, N3) were tested for ehrlich
spectrum of anti-tumor properties.
Titanium based compounds i.e. bis (β- ascite tumor in mice and show anti
diketonato)titanium(IV) [Budotitane] and proliferative activity similar to Cp2TiCl2.
titanocene derivatives offer an alterative for Budotitane and titanocene dichloride
cancer chemotherapy. The anti-tumor possess same limitation that they have low
activity of budotitane was reported in hydrolytic stability at physiological pH52.
1982.This was first non-platinum complex In this respect titanium(IV) complexes
tested in clinical trials and used against offer a new outlook for chemotherapy. The
19 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
novel titanocene compounds are better than position show more cytotoxicity than
cis-platin for apoptic effect in vitro and titanocene dichloride due to –OCF3 group
they can induce more apoptosis than cis- on Para position and more stability in
platin. TitanoceneY (bis-[(p- aqueous solution. Different compounds
methoxybenzyl) cyclopentadienyl titanium were synthesized by replacing –OCF3 by
dichloride) had better effect in prostate, another groups and these were found active
pancreas, breast and ovarian cell lines and against breast cancer cell line MCF-759. In
in uterine and renal cancer cells56. Michael addition to these, titanium alkoxide
shavit et al studied Ti(IV) complexes of complex show toxicity in breast, colon and
oxygen-based ligands. They had prepared pancreatic cancer cell lines but molecular
the homoleptic complex of hydroxyamino mechanism yet to be elucidated60.
1,3,5 triazine ligands. These triazine
ligands possess mild reactivity despite SUMMARY
having no labile groups. This complex was Cis-Platin is the model drug used against
effective against colon and ovarian cancer mainly testicular and ovarian cancer cell
cells16. Since titanocene dichloride is active lines but many other non-platinum based
against colorectal, lung and breast drugs have been synthesized which are the
carcinomas, new derivatives may have anti- alternatives of cis-platin. Other transition
tumor activity profile. These complexes metal complexes of titanium, ruthenium,
have advantage that they do not show palladium and gold etc. have also been
common side effects such as emesis, used for the chemotherapy. Due to less side
alopecia, or bone marrow impairment52. effects of titanium metal complexes, they
These features make titanium compounds can be used for the synthesis of new metal
interesting for combined therapy and based anti-cancer drug. Specifically amine
further study16. The novel achiral based ligands which are highly charged
titanocene (Titanocene C and Titanocene have been used for the treatment of cancer
Y) anti-cancer drug are almost ten times whose ultimate target is DNA.
less toxic than cis-platin. The
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Vol. 03 issue 07 July 2011
EFFECT OF THICKNESS AND ANNEALING TEMPERATURES
ON STRUCTURAL AND MORPHOLOGICAL PROPERTIES
OF (SB2S3)1-X (BI2S3)X THIN FILMS
Jessy Mathew N1., Rachel Oommen1, C. Sanjeeviraja 2, Usha Rajalakshmi P1

ijcrr 1
Department of Physics, Avinashilingam Deemed University for Women, Coimbatore
Vol 03 issue 07 2
Category: Research Department of Physics, Alagappa University, Karaikkudy, Tamil Nadu
Received on:25/04/11
Revised on:10/05/11 E-mail of corresponding author: sisjees@yahoo.co.in
Accepted on:21/05/11
______________________________________________________________________________

ABSTRACT
(Sb2S3)1-x (Bi2S3)x (with x= 0.03, 0.05, 0.1) thin films were prepared on chemically well cleaned
glass substrates by thermal evaporation technique for two thickness. The orthorhombic crystal
structure of the (Sb2S3)1-x (Bi2S3)x powder samples and thin films have been found out from XRD
powder data and confirms the transition of films from amorphous to polycrystalline with thermal
treatment at 523K. Also lattice parameters, crystallite sizes and interplanar spacings for the
newly prepared ternary compounds (Sb-Bi-S) have been calculated. The XPS spectra were
carried out to identify the surface compositions. The elemental percentage in each compound
was verified by EDAX. Increase of crystallite size with thickness and temperature was proved by
XRD and the influences of thickness and annealing temperature on roughness and grain size was
observed from AFM measurements.

Keywords: Thermal evaporation technique, the applications in photosensitivity,


Ternary compounds, Amorphous, photoconductivity and thermoelectric power1.
Polycrystalline, Orthorhombic crystal structure Antimony sulphide and Bismuth sulphide are
layer structured direct band gap semiconductors
1. INTRODUCTION with orthorhombic crystal structure have been
The ternary semiconductor compounds in thin received considerable interest because of their
film form have become the focus of attention structural, morphological, optical and electrical
because of their important physical and chemical properties which allow their wide use in many
properties since these properties could be devices. A number of methods have been used
changed by varying the annealing temperature for the preparation of V-VI compound thin films
and thickness. Thin films of semiconductor by the thin film investigators like, N.S.
chalcogenides of the chemical formula m2V-B- Yesugade et al.2 prepared Sb2S3 and Bi2S3 thin
n3VI-B basically have the structure of antimony films by electrodeposition technique, Gang Xie
sulphide which are most important materials for et al.3 synthesized M2S3 (M = Sb, Bi) via a
24 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
hydrothermal treatment, Pawan Kumar et al.4 cleaned substrates were placed in a rotative
prepared Bi2S3 thin films by spray pyrolysis sample-holder to get uniformly coated films.
technique, Siham Mahmoud et al.5 produced thin Deposition rate and film thickness were
films of Bi2S3 by thermal evaporation technique, controlled during deposition by quartz oscillator
I.K. El Zawawi et al.6 prepared Sb2S3 thin films thickness monitor. The substrate temperature
by thermal vacuum evaporation technique, I. was maintained at 300K during the deposition.
Grozdanov et al.7 fabricated thin films of Sb2S3 The deposition chamber was evacuated to a
by chemical deposition technique etc. Pawar et residual pressure of about 10-5 torr. The
al.2 obtained Bi2S3, Sb2S3 and As2S3 films by a elemental compositions of the prepared powder
solution-gas interface technique. and thin films were determined using an Energy
This paper reports the investigations on the Dispersive X-ray Analysis (EDAX) (model:
effect of thickness and annealing temperature on INCA Oxford) based on SEM images. X-ray
the structural and morphological properties of Diffractograms of the prepared material and the
thermally evaporated (Sb2S3)1-x (Bi2S3)x (with x= investigated films of as-deposited and annealed
0.03, 0.05, 0.1) thin films from newly prepared (473K, 523K) samples were carried out by using
powder samples. Characterizations were done by an X-ray Diffractometer (XRD) (model: D8
means of Energy Dispersive X-ray Analysis Advance XRD) with CuKα (λ=1.5406Å)
(EDAX), X-ray Diffraction (XRD) technique, radiation. To analyse the surface compositions
X-ray Photoelectron Spectroscopy (XPS) and and purity, the x-ray photoelectron spectroscopy
Atomic Force Microscopic (AFM) analysis. To (XPS) data were recorded with the specimen
the best of our knowledge nobody has performed mounted on a specially designed sample holder8
the work on the preparation and characterization using an AlKalpha laboratory x-ray source that
of (Bi2S3)x doped (Sb2S3)1-x (with x= 0.03, 0.05, was operated at 150 watts and an electron
0.1) thin films from the freshly prepared powder energy analyser with five channeltrons from
samples and thermal vacuum evaporation Specs GmbH, Germany. The data were recorded
technique as the method of deposition. with 20 eV pass energy with 1 eV energy
resolution. The chamber base pressure was
2. MATERIALS AND METHODS 6×10-11 mbar. Surface morphological studies
Bulk thin film samples of (Sb2S3)1-x (Bi2S3)x were done before and after heat treatment by
(with x= 0.03, 0.05, 0.1) were prepared from means of Atomic Force Microscope (AFM) and
powder materials of Sb2S3 and Bi2S3 (99.999% surface roughness and grain size were estimated.
purity, Sigma-aldrich) on chemically cleaned
glass substrates by thermal vacuum evaporation 3. RESULTS
technique. The conventional thermal evaporation 3.1. Compositional analysis
method is being widely used for the growth of Fig. 1 shows the Energy Dispersive X-ray
binary and ternary compounds because of its Analysis (EDAX) spectra for the powder
simplicity and this method supports high quality samples and the corresponding thin films of
and pure films for thin film applications. For a (Sb2S3)1-x(Bi2S3)x, with x= 0.03, 0.05, 0.1. The
particular composition, the constituent calculated and observed percentages of the
compounds Sb2S3 and Bi2S3 were weighed and elements in different compositions from EDAX
grinded for several hours and sintered. The analysis are shown in table 1 and are in good
newly prepared powder samples were used as agreement each other.
the source material. During deposition the pre- (Figure 1, Table 1)
25 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
D=Kλ (1)
3.2. Structural analysis
β cosθ
The X-ray diffractograms of the as-deposited,
annealed (473K, 523K) thin films and powder where K is the shape factor (0.94), β is the full
samples of (Sb2S3)1-x(Bi2S3)x with x= 0.03, 0.05, width at half maximum (FWHM) of the
0.1 are shown in figures (2-13). It is clear from diffraction expressed in radians. The estimated
figs. (2-7) the as-deposited films and the films lattice parameters for both powder and thin films
annealed at 473K are amorphous in nature9-10. and crystallite size for the annealed film samples
As reported by F. Perales et al.11, it was proved of (Sb2S3)1-x(Bi2S3)x with x= 0.03, 0.05, 0.1 of
that the effect of annealing at 473K cannot different thickness are recorded in table 2.
change the amorphous nature. Thin films have Using Bragg‘s relation14, the interplanar spacing
been annealed for 1hr at 523K to investigate the dhkl was also calculated from powder diffraction
effect of higher annealing temperature, and the data:
growth of polycrystalline from amorphous dhkl = nλ (2)
phase. This observation is in agreement with the 2sinθ
results of M. S. Droichi et al.12 for Sb2S3 and
Mahmoud et al.10 for Bi2S3 thin films. where λ is the X-ray wavelength, n is the order
(Fig. 2 Fig. 3) number and θ is the Bragg‘s angle. Table 3
(Fig. 4 Fig. 5) shows the calculated dhkl values of Sb-Bi-S
(Fig. 6 Fig. 7) compound in comparison with the standard
From figs. (8-10), it is clear that the increase of powders of Sb2S3 (JCPDS card No. 42-1393)
thermal treatment improves crystallinity of Sb- and Bi2S3 (JCPDS card No. 17-0320) and in
Bi-S films with orthorhombic structure. The X- agreement with the recorded values by A.A. El-
ray diffractograms of (Sb2S3)1-x(Bi2S3)x with x= Shazly et al.15 and Siham Mahmoud et al.5. The
0.03, 0.05, 0.1 for powder samples are shown in surface composition and purity of the as-
figures (11-13). deposited (Sb2S3)0.95 (Bi2S3)0.05 thin film were
(Fig.8 Fig. 9) studied by XPS analysis and the core level
The orthorhombic crystal structure of these spectra are given in figs. (14-16). The
compounds, both in powder and thin film forms, photoelectron spectra of the Sb(3d3/2), Bi(4d5/2)
with cell parameters have been found out from and S(2p) show the peaks at the binding energy
XRD powder data. The position of peaks 537.61eV, 440.01eV and 162. 21eV
appeared in the diffractograms of the powder respectively.
sample lie very near to those listed in the PDF (Fig. 14)
for Sb2S3 (PDF 42-1393) and Bi2S3 (PDF 17- (Table 2)
0320). The estimated values of the lattice 3.3 Morphological analysis
parameters lie close to the reported values of The surface topographical images for Sb-Bi-S
Sb2S3 and Bi2S311, 13. thin films, deposited at room temperature and
(Fig. 10 Fig.11) annealed at 523K for 1 hr, of film thickness
(Fig.12 Fig.13) 900nm and 1200nm recorded from atomic force
The crystallite size D estimated using the microscope (AFM) are shown in fig.15. This
Sherrer‘s formula14: measurements have been taken for films from
the compound with x=0.05. The measured grain
size and RMS roughness from AFM images for

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Vol. 03 issue 07 July 2011
thickness 900nm and 1200nm are given in table 4. CONCLUSIONS
4. It is observed from the AFM images that the
average grain size and roughness increase with The thermal evaporation technique has been
increasing thickness and temperature16. used to prepare (Sb2S3)1-x(Bi2S3)x with x=0.03,
(Fig. 15) 0.05, 0.1 thin films on glass substrate for two
thickness. The measurements obtained from
(Table 3) EDAX showed the elemental percentages of
(Table 4) respective compounds. The orthorhombic crystal
structure of all compounds with x=0.03, 0.05
3. DISCUSSIONS and 0.1 and lattice parameters and the
As the crystallization process proceeds with dependence of crystallite size on thickness were
thickness at temperature 523K, the intensity of showed by X-ray Diffraction (XRD)
the (310) peak increases for x=0.03 compound, measurements. The as-deposited amorphous thin
(120) peak increases for x=0.05 compound and films transformed to polycrystalline films during
(130) peak increases for x=0.1 compound, the thermal treatment at 523K. The XPS core
indicating further orientation of the film level spectra of Sb(3d3/2), Bi(4d5/2) and S(2p)
crystallites in these directions. And also, in all showed the peaks of corresponding binding
the three compounds, more peaks appeared in energies. Dependence of grain size and RMS
films of highest thickness (1200nm). Some roughness on thickness and annealing
peaks appeared, which are common to both temperature was proved by atomic force
Sb2S3 and Bi2S3 like, (120) and (250) for x=0.03, microscopic (AFM) photographs.
(120) and (240) for x=0.05, (120), (130), (240)
and (250) for x=0.1. The calculated values of the ACKNOWLEDGEMENT
lattice parameters for the three compounds lie The measurements have been taken in the UGC-
close to the reported values of Sb2S3 and Bi2S3. DAE Consortium for Scientific Research,
The calculated interplanar spacing dhkl values of Indore. The authors are much grateful to the
Sb-Bi-S compound lie close to the standard supporting authorities, especially to Prof. Ajay
powders of Sb2S3 (JCPDS card No. 42-1393) Gupta- the centre Director of Indore. The
and Bi2S3 (JCPDS card No. 17-0320). corresponding author is also much grateful to the
financial support given by DST- CURIE Project.
As K.Y. Rajpure et al.17 point out, annealing
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3. Gang Xie, Zheng-Ping Qiao, Ming-Hua 11. Perales F, Lifante G, Agullo-Rueda F, de las
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Mitreski M. Fabrication of amorphous The effect of deposition rate and heat
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D‘Souza S W, Nayak J, Barman S R. An 16. Kasturi L Chopra. Thin Film Phenomena.
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Vol. 03 issue 07 July 2011
Table1. Percentages of the elements in (Sb2S3)1-x(Bi2S3)x with x= 0.03, 0.05 and 0.1 from EDAX
analysis
Percentage of element at
Element calculated observed
x
Powder Film
Sb 38.8 35.97 32.9
Bi 1.2 1.04 0.92
0.03
S 60 62.99 66.18
Sb 38 42.71 42.73
Bi 2 1.98 1.39
0.05
S 60 55.31 55.88
Sb 36 37.19 32.49
Bi 4 3.82 3.27
0.1
S 60 58.99 64.24

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Vol. 03 issue 07 July 2011
Table 2 Cell parameters for both powder and film samples and crystallite sizes for thin films of
thickness 900nm and 1200nm of (Sb2S3)1-x(Bi2S3)x with x= 0.03, 0.05 and 0.1 from XRD
measurements

Cell parameters (nm)


x Powder Film Thickness Crystallite
(thickness=1200nm) (nm) size (nm)
a b c a b c
0.03 1.124 1.126 0.381 1.124 1.127 0.381 900 57
1200 64
0.05 1.11 1.131 0.386 1.120 1.131 0.384 900 61
1200 73

0.1 1.126 1.129 0.383 1.128 1.130 0.382 900 59


1200 74

Table 3 The inter planar spacing, d for Sb-Bi-S compound from XRD analysis

Experimental JCPDS- d (Å)


data - d (Å) hkl

Sb2S3 Bi2S3
5.653 5.660 5.654 020
5.040 5.057 5.040 120
3.986 3.987 3.969 220
3.570 3.575 3.569 130
3.128 3.131 - 211
2.765 2.765 2.812 221
2.680 2.681 2.717 301
2.607 2.609 2.641 311
2.521 2.525 - 240
2.425 2.428 - 231
2.277 2.232 2.258 141
1.992 1.994 1.985 440
1.884 1.886 1.857 060

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Vol. 03 issue 07 July 2011
Table 4 The measured grain size and RMS roughness from AFM images of (Sb2S3)0.95(Bi2S3)0.5
compound

Grain size (nm) Roughness (nm)


Thickness as- Annealed as- Annealed
(nm) deposited at 523K deposited at 523K
900 49 55 2.496 2.582

1200 61 68 2.591 2.696

Fig. 1. EDAX spectra of (Sb2S3)1-x(Bi2S3)x


with (a) x=0.03, (b)x=0.05 and (c)x=0.1
Fig. 2 XRD patterns of as-deposited (Sb2S3)0.97 (Bi2S3)0.03
thin films of thickness a) 900nm and b) 1200nm

( b )
I n t e n s i t y ( a. u )

( a )
(a)

0 10 20 30 40 50 60

NB: Figure nos. (2-13) - a.u (on Y-axis) : aurbitrary unit

Fig. 3 XRD patterns of as-deposited (Sb2S3)0.95 (Bi2S3)0.05


thin films of thickness a) 900nm and b) 1200nm
(b)

( b )
I n t e n s i t y ( a. u )

( a )

(c)

10 20 30 40 50

31 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
(b)

(b )

I n t e n s i t y ( a. u )
I n t e n s i t y ( a. u )

(a)
(a)

0 10 20 30 40 50 60 0 10 20 30 40 50 60

2 2

Fig. 4 XRD patterns of as-deposited (Sb2S3)0.90 (Bi2S3)0.1 Fig.5 XRD patterns of annealed (473K) (Sb2S3)0.97 (Bi2S3)0.03
thin films of thickness a) 900nm and b) 1200nm thin films of thickness a) 900nm and b) 1200nm

(b)
(b)
I n t e n s i t y ( a. u )

I n t e n s i t y ( a. u )

(a)
(a)

10 20 30 40 50 0 10 20 30 40 50 60

2 2

Fig.6 XRD patterns of annealed (473K) (Sb2S3)0.95 (Bi2S3)0.05 Fig.7 XRD patterns of annealed (473K) (Sb2S3)0.90 (Bi2S3)0.1
thin films of thickness a) 900nm and b) 1200nm thin films of thickness a) 900nm and b) 1200nm

32 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
I n t e n s i t y ( a. u ) I n t e n s i t y ( a. u )

33
0

0
10

10
(020) (020)
(020) (020) ( 1 2 0 ) ( S b2 S3 , B i2 S3 ) (120) (Sb S , Bi S )
2 3 2 3

20
( 1 2 0 ) ( S b2 S3 , B i2 S3 ) ( 1 2 0 ) ( S b2 S3 , B i2 S3 ) (220)
(220)

20
(220) (220) (310) (310)
(130) (130) (320) (320)

2
(211) (211)

30
( S b S , B i2 S3 ) ( S b S , B i2 S3 )
(230) 2 3 (230) 2 3
(211) (221) (211)

30
(211) (301) (301)
(221) (240)
(301) ( 3 0 1()2 2 1 ) (231) (240)

40
( 2 4 0 )( S b2 S3 , B i2 S3 ) ( 2 3 1 )( 2 4 0 )( S b2 S3 , B i2 S3 ) (411) (411)

40
(141) (250) (Sb S , Bi S ) ( 2 5 0 ) ( S b2 S3 , B i2 S3 )
2 3 2 3
( 2 5 0 ) ( S b2 S3 , B i2 S3 ) ( 2 5 0 ) ( S b2 S3 , B i2 S3 ) (530) (501)

50
(151)(Bi S ) ( 1 5 1 ) ( B i2 S3 )
2 3

50
(061)
(132)

thin films of thickness a) 900nm and b) 1200nm

thin films of thickness a) 900nm and b) 1200nm


60
(b)

(a)

60
(a)
(b)

Fig. 10 XRD patterns of annealed (523K) (Sb2S3)0.90 (Bi2S3)0.1


I n t e n s i t y ( a. u )
I n t e n s i t y ( a. u )

0
100
200
300

0
10

( 1 1 0 ) ( S b2 S3 , B i2 S3 )

10
( S b2 S3 , B i2 S3 )
(020) (020) (120)
(120)
(020) ( S b2 S3 , B i2 S3 )
20

( 1 2 0 ) ( S b2 S3 , B i2 S3 ) (220) (220)

20
(130) (130)

powder sample
(220) (310)
(111) ( 1 1 1 )( 3 1 0 )
(310)

Vol. 03 issue 07 July 2011


(111) (320) (230)
2

(320) (211) (211)


30

2
(211) (221)

30
(221)
(221) (301)
(301) ( 3 1 1 () 3 0 1 )
(311) (420) ( 2 4 0 ) ( S b2 S3 , B i2 S3 )
(231) ( 2 4 0 ) ( S b2 S3 , B i2 S3 )
(041) (231)
40

40
(411)
( 2 5 0 ) ( S b2 S3 , B i2 S3 ) (250)
(501) (501)
(511)

50
50

( 5 3 0 ) ( B i2 S3 )
(531)
(360)
(b)

(a)

thin films of thickness a) 900nm and b) 1200nm

60

Fig.11 XRD pattern of (Sb2S3)0.97 (Bi2S3)0.03


Fig.8 XRD patterns of annealed (523K) (Sb2S3)0.97 (Bi2S3)0.03 Fig. 9 XRD patterns of annealed (523K) (Sb2S3)0.95 (Bi2S3)0.05

International Journal of Current Research and Review www.ijcrr.com


Intensity (a.u.)

34
0
5000
10000
15000
20000

10
( 1 1 0 ) ( S b2 S3 , B i2 S3 )

(020)
( 1 2 0 ) ( S b2 S3 , B i2 S3 )

20
(220)
( S b2 S3 , B i2 S3 )

2
(111) (130)
(230)
(211)

30
(221)
(301)
(311)

( deg )
( 2 4 0 ) ( S b2 S3 , B i2 S3 )
(231)
(041)
40
(141)
(250)
(440)
( 1 5 1 ) ( B i2 S3 )
(060)
50

Fig.12 XRD pattern of (Sb2S3)0.95 (Bi2S3)0.05 powder sample ( S b2 S3 , B i2 S3 )

I n t e n s i t y ( a. u )
0
100
200
300

0
10

(020)
( 1 2 0 ) ( S b2 S3 , B i2 S3 )
20

(220) ( S b2 S3 , B i2 S3 )

Vol. 03 issue 07 July 2011


(111) (230) (130)
2

(211)
30

(221)
(301)
(231) ( 2 4 0 ) ( S b2 S3 , B i2 S3 )
40

(141)
( 2 5 0 ) ( B i2 S3 )
( 1 5 1 ) ( B i2 S3 )
50

(531)
(132)
60

Fig.13 XRD pattern of (Sb2S3)0.90 (Bi2S3)0.1 powder sample

International Journal of Current Research and Review www.ijcrr.com


Relative Intensity (cps) Sb ( 3d 3/2 )

532 536 540 544

Binding Energy (eV)

(a)
Relative Intensity (cps)

Bi ( 4 d 5/2 )

436 440 444

Binding Energy (eV)

(b)
S(2p)
Relative Intensity (cps)

Fig. 14 XPS spectra of (Sb2S3)0.95 (Bi2S3)0.05 thin film of


thickness 1200nm. Core level spectra of (a) Sb(3d 3/2),
(b) Bi(4d5/2) and (c) S(2p)

160 164 168

Binding Energy (eV)

(c)

35 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
(a) (b)

(c) (d)

Fig.15 AFM images of the (Sb2S3)0.95(Bi2S3)0.5 thin films for thickness 900nm [as-deposited (a), annealed (b)] and

for 1200nm [as-deposited (c), annealed (d)]

36 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
EFFECT OF SOLANUM TRILOBATUM EXTRACT
ON ERYTHROCYTE MEMBRANE SURFACE CHARGE
IN AGED RATS
Amarnath Kanchana1 , Chinnakannu Panneerselavam1
ijcrr 1
Vol 03 issue 07 Deprtment of Biochemistry, Dr. ALM Post Graduate Institute of Basic Medical
Category: Research Sciences, Taramani, Chennai, India
Received on:19/04/11
Revised on:06/05/11 E-mail of corresponding author: kanchibms@yahoo.co.in
Accepted on:15/05/11

ABSTRACT
Objectives: Oxidative stress an unavoidable consequence of oxygen metabolism in aerobic cells is
postulated to be one of the most important causes of age related changes. The changes induced by free
radicals are believed to be the major source of ageing, disease development, destruction of normal cell
function and membrane rigidity. The present study was intended to determine the effect of chloroform
extract of Solanum trilobatum (CST) on membrane surface charge density in aged rat erythrocytes upon
oxidative stress.
Methods: The leaves of Solanum trilobatum were subjected to chloroform and the extract was prepared.
Chloroform extract of Solanum trilobatum (CST) (150 mg/day/kg body weight) was administered orally
for 90 days to young and aged rats. The erythrocyte membrane was isolated and was used to perform the
studies such as estimation of protein carbonyls, glycoproteins, surface charge and enzymatic and non
enzymatic levels.
Key findings: A noteworthy decline in surface charge levels with simultaneous increase in protein
carbonyls and shrink in glycoprotein and antioxidants status was prominent in erythrocytes of aged rats
contrast to young rats. Administration of CST improved the erythrocyte surface charge density to near
normalcy in aged rats. In addition a decrease in protein carbonyls level and increase in glycoproteins as
well as antioxidant status was observed in aged rat erythrocytes on CST therapy.
Conclusions: Therefore the polyphenols and flavanoids are the phytochemical compounds in CST that
might play a central role in defending the oxidative stress related loss of membrane surface charge in a
way maintaining the erythrocyte membrane integrity and functions in aged individuals.
______________________________________________________________________________

Keywords: Aging; Erythrocyte surface charge; age-related diseases with a progressive increase
Protein carbonyls; Glycoprotein; Antioxidants; in the chance of morbidity and mortality. At the
Solanum trilobatum; Phytochemicals. cellular level, the reactive oxygen species
inclination due to oxidative stress damages vital
INTRODUCTION cell components like poly unsaturated fatty
Oxidative damage apparently increases with age acids, protein and nucleic acids2. Free radical
and thus may overwhelm the natural repair attack on cellular proteins, damages the proteins3
system in the organism1leading to the onset of leading to oxidation of side chains of lysine,
37 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
arginine, proline and threonine thereby yield complications10and sequestration of circulating
protein carbonyl derivatives the markers of erythrocytes by macrophages11. Age associated
protein oxidation4. These oxidative oxidative changes in the glycomoieties of
modifications cause loss of structural and erythrocyte glycoprotein affects the surface
catalytic functions of the cell contributing to charge including aggregation and binding of
serious deleterious changes affecting cell erythrocytes to macrophages12 and found to be
survival5. associated with cardiovascular risk factors such
The cell membrane is a structural barrier that as, hypertension, hyperlipoproteinemia and
plays an essential role in protecting cellular myocardial ischemia13and thus crucial for
integrity and underlying cause of ageing erythrocyte survival14.
process6. Changes in the macromolecules of During lifetime, an antioxidant network
membrane are one of the earliest signs of counteracts the deleterious action of free radicals
erythrocytes membrane alterations during and reactive species on macromolecules. Cells
ageing. Reactive oxygen species generated in the synthesize some of their antioxidant enzymes, as
aqueous or lipid phase can attach to the the superoxide dismutase, catalase, and
erythrocyte membrane and can induce oxidation glutathione peroxidase, as well as the peptides
of lipids, proteins and carbohydrates, triggering with thiol groups, as glutathione, while other
disruption in membrane7and further enhances non-enzymatic antioxidants from nature through
the development of one or better-recognized age nutrition, as vitamin C, vitamin E, and
related modifications, such as alteration of carotenoids. Several repair systems help
intrinsic membrane properties (fluidity, ion antioxidant action by the recovery of damaged
transport, loss of enzyme activity, protein macromolecules15. Together, these systems play
crosslinking), inhibition of protein synthesis and an important role in the ability of the body to
DNA damage4, ultimately resulting in the cell respond to the oxidant challenge of using
death. molecular oxygen to drive reactions that yield
Carbohydrates in erythrocytes have many the necessary energy for life processes.
important functions that are necessary for the However various anti ageing agents are being
proper functioning of the cell to survival. ushered into the field of medicine to conflict
Carbohydrate is thought to be highly necessary ageing and age-associated disorders. Flavonoids,
for cell surface negative charges and have a the potential dietary antioxidant found
crucial role in the clearance of senescent cells ubiquitously in plants, attracted global interest in
and essentially to prevent aggregation of combating the devasting effects of oxidation in
erythrocytes from each other8. Like most cells and tissues of an organism. Dietary
biological surfaces, erythrocytes exhibit a antioxidants are considered beneficial because of
negative surface charge that is mainly attributed their potential protective role against oxidative
to sialic acid residues located on the stress, which is involved in the pathogenesis of
glycoproteins in the membrane surface9. multiple diseases such as cancer and coronary
Oxidative stress or other damaging effects to heart disease16,17. The potential antioxidant
surface sialosaccharide may itself play role in effect in vivo of individual food polyphenols
aggregation of erythrocytes, increasing the (PP) or concentrated extracts has been widely
adhesiveness to endothelial cells contributing for investigated in cultured cells18,19 live animals20
the development of various pathologies and men21,22,23. Herbal remedies as antioxidant
including diabetes mellitus, atherothrombotic supplement is thus globally growing up owing to
38 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
broad spectrum of beneficial biological concentrator. Then it was frozen and subjected
properties. Owing to the presence of phenolic to lyophilization.
compounds and the free radical scavenging 2.3. Animals, treatment and fixation of
properties24 of an anti cancer herabal extract dosage and duration
Solanum trilobatum25, the present study is Male albino rats of Wistar strain, weighing
carried out to study the effect of Chloroform approximately 120–160 g (young) and 380–420
extract of Solanum trilobatum (CST) on g (old) were used. The animals were obtained
erythrocyte membrane surface charges during from King Institute of Preventive Medicine,
animal aging. The study also explores the effect Chennai. The animals were divided into two
of CST on erythrocyte membrane protein major groups namely, Group Ia: normal young
carbonyl levels and the antioxidant status in rats (3–4 months old) and Group IIa: normal
aged rats. aged rats (about 24 months old) and two
experimental group Id (Young rats) and group
2. MATERIALS AND METHODS IId (Aged rats) on CST administration for 90
days. Each group consisted of six animals. The
2.1. Chemicals animals were maintained on commercial rat feed
Dextran 500 (500,000 molecular weight), containing 5% fat, 21% protein, 55% nitrogen
Polyethylene glycol (PEG, 8000 molecular free extract, 4% fiber (w/w) with adequate
weight), Bovine serum albumin (BSA), and 2,4- mineral and vitamin contents and had access to
dinitro-phenylhydrazine (DNPH) were water ad libitum. CST (Chloroform extract of
purchased from Sigma Chemical and Company, Solanum trilobatum) was supplemented orally
Saint Louis, MO, USA. All other chemicals (oral gavage) at the dosage of 150 mg/kg body
were of reagent grade. weight/day for 90 days, whereas young control
2.2. Preparation of Chloroform extract of and aged control rats received vehicle alone in a
Solanum trilobatum (CST) similar manner. On completion of 90 days of
Extraction of plant materials CST supplementation, the blood was collected
The leaves of Solanum trilobatum were with 3.8% sodium citrate from jugular vein and
collected from the local market and samples of used for the isolation of erythrocytes and
the plant were identified and authenticated by erythrocyte membranes.
Dr. Brindha, Botanist, Dept. of Pharmacognosy, Fixation of dose and duration: Trial studies were
Captain Srinivasa Murti Drug Research Institute carried out with different concentrations of
for Ayurveda (CCRAS, New Delhi), Chloroform extract of Solanum trilobatum (50,
Arumbakkam, Chennai, Tamil Nadu. The fresh 100, 150, 200 and 250 mg) dissolved in
leaves were shad dried, powdered and extracted physiological saline and supplemented at various
successively with 1.2 L of chloroform, in a durations (30, 60, 90 and 120 days) orally to
Soxhlet extractor for 18-20 h. The extracts were rats. The effectual dosage (150 mg/kg body
concentrated to dryness under reduced pressure weight/day) was selected on the basis of the
and controlled temperature (40o-50oC). The concentration at which Chloroform extract of
chloroform, extract yielded a brown semisolid Solanum trilobatum was capable of inhibiting
residue weighing 6.2 g (2.39% w/w). The lipid peroxidation significantly above which
extract was then filtered through Whatmann there was (200 and 250 mg/kg body weight) no
No.1 filter paper and concentrated using significant inhibition of lipid peroxidation.
Similarly the effectual duration was found to be
39 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
90 days above which there was insignificant 2.5. Determination of erythrocyte surface
inhibition of lipid peroxidation. As per the charge
results obtained, the forthcoming biochemical Erythrocyte partition coefficients were
and molecular parameters were carried out with determined at room temperature by a two-phase
Chloroform extract of Solanum trilobatum aqueous system containing 5% Dextran 500 and
supplementation for 90 days alone. 4.3% PEG 8000 in isotonic phosphate buffer
The body weight of the animals were increased (pH 6.8) by the method of Walter (1985)29. The
at the end of experiment as in group Ia (from two-phase system used is ‗charge sensitive‘
74.50 ± 5.6 to 80.40 ± 6.7), group Id (from whose partition coefficient ratio indicates the
74.50 ± 5.6 to78.30 ± 3.3), group IIa (from level of erythrocyte surface charge.
120.4 ± 4.8 to 155.9 ± 5.0) and group IId (from 2.6. Determination of protein carbonyls
126.2 ± 5.0 to 162.0 ± 5.3) in respect to their Protein carbonyls content was determined by the
initial body weight. reliable method based on the reaction of
carbonyl groups with 2, 4- dinitro-
2.4. Preparation of erythrocytes and phenylhydrazine to form 2, 4-dinitro-
erythrocyte membranes phenylhydrazone as suggested by Levine et al.
Isolation of erythrocytes and erythrocyte (1994)30.
membranes was done according to Dodge et al. 2.7. Determination of glycoproteins
(1963)26 with slight modifications. Briefly, Lipids were extracted from the erythrocyte
blood collected from animals with 3.8% sodium membrane pellets according to the method of
citrate was subjected to centrifugation at 3000 Folch et al. (1957)31using chloroform–methanol
rpm for 10 min at 40C. The plasma and buffy mixture (2:1 v/v). The resulting defatted residue
coat were removed by aspiration and the was suspended in sodium acetate buffer
erythrocytes were washed three times with cold (containing 2 mg cysteine HCl/ml, final pH 7.0)
(40C) phosphate-buffered saline (PBS: 0.15 M and deproteinized by 4–5 volumes of ethanol,
NaCl, 1.9 mM NaH2PO4, 8.1 Mm Na2HPO4, pH evaporated to dryness in the cold under a
7.4). Washed erythrocytes were hemolyzed in 40 vacuum and subjected to hydrolysis by heating
volumes of 5 mM sodium phosphate buffer (pH with 2 ml of 4 N HCl for 4–6 h. The hydrolyzed
8.0) (containing 1 mM EDTA and 0.5 mM material was neutralized with 4 N sodium
phenylmethylsulfonyl fluoride, as protein hydroxide and used for estimating erythrocyte
inhibitor), and centrifuged for 20 min at 40C at hexose (Niebes, 1972)32, hexosamine (Wagner,
15,000 rpm. The supernatant (or hemolysate) 1979)33 and sialic acid (Warren, 1959)34.
was decanted carefully and saved, while the 2.8. Determination of antioxidants
pellet were washed repeatedly and incubated for Superoxide dismutase (SOD) in hemolysate was
45 min at 370C to reseal the membrane in the assayed by the method of Marklund and
presence of 0.8 Mm MgCl2-ATP solutions, for Marklund (1974)35, while erythrocyte
hemoglobin free ghosts. Hemoglobin amount membranes were used to estimate catalase
was estimated by Drabkin and Austin (1932)27 (CAT) (Beers and Seizer, 1952)36. Hemolysate
method and erythrocyte membrane protein glutathione peroxidase (GPx) was determined by
content by Lowry et al. (1951)28. Rotruck et al. (1973)37 method, glutathione
reductase (GR) by the procedure of Staal et al.
(1969)38, glucose-6-phosphate dehydrogenase
(G6PD) by Zinkham et al. (1958)39 and
40 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
glutathione-S-transferase (GST) according to treatment to aged rats showed significant
Habig et al. (1974)40. Hemolysate reduced decrease in protein carbonyl levels by 1.6 fold in
glutathione (GSH) was estimated by Moron et erythrocytes and 1.5 fold in plasma. The
al. (1979)41 and oxidized glutathione (GSSG) observed partition coefficient ratio and protein
was assayed according to the method of Tietze carbonyls level were normal in erythrocytes of
(1969)42. Erythrocyte redox state was control and Chloroform extract of Solanum
determined by the redox index: (GSH + 2 X trilobatum treated young rats. Thus the results
GSSG) / (2 X GSSG / 100). reveal the membrane integrity and functions are
maintained at normal levels in erythrocytes of
2.9. Statistical analysis young rat.
The results are expressed as mean standard Figure 3 depicts the levels of glycoproteins like
deviation (SD) for six rats in each group. hexose, hexosamine and sialic acid in
Differences between groups were assessed by erythrocyte membrane of control and
one-way ANOVA using the SPSS version 7.5 Chloroform extract of Solanum trilobatum
software packages for Windows, USA. Post hoc treated young and aged rats. The glycoprotein
testing was performed for inter-group levels in erythrocytes of young rats were at
comparisons using the least significance normal level and did not show any significant
difference (LSD) test; statistical significance at alterations on Chloroform extract of Solanum
p-values<0.05, have been given respective trilobatum treatment. Profound decrease in the
symbols in the figures and tables. levels of hexose, hexosamine and sialic acid by
23%, 33% and 22 % respectively in erythrocytes
3. RESULTS of aged rats was observed. However
Figure 1 illustrates surface charge in supplementation of Chloroform extract of
erythrocytes of young and experimental rats in Solanum trilobatum enhanced the level of
terms of partition coefficient ratio. Significant (p hexose by 30%, hexosamine by 40% and sialic
< 0.05) decrease in surface charge level in acid by 23% in aged rat erythrocytes.
erythrocytes was noticed in aged rats when Figure 4 highlights the activities of enzymatic
compared to young rats. CST supplementation to antioxidants SOD, CAT, GPx, GST, GR and
aged rats showed increment in surface charge G6PD in erythrocytes of young and aged rats.
(p<0.05) when compared to control aged rats. Young control rats had normal level of
The result illustrated significant surface charge enzymatic antioxidants and showed insignificant
loss and increased free radical attack on changes on CST supplementation. The activities
membrane proteins in aged animal. of SOD, CAT GPx, GR, GST and G6PD were
Figure 2 shows age related changes in protein decreased significantly by 33%, 47% 38% and
carbonyls in erythrocytes of control and 39% 39% and 45%respectively in aged rats
Chloroform extract of Solanum trilobatum when compared to young rats. However CST
treated young and aged rats. Increase in protein supplementation to aged rats improved the
carbonyl by 1.7 fold was observed in erythrocyte activities of SOD by 31%, CAT by 68%. GPx
membrane and plasma of aged rats when by 39%, GST by 43%, GR by 44% and G6PD
compared with young rats. Such an elevation in by 59%.
protein carbonyl content indicates increased Table 1 demonstrates the glutathione status in
protein oxidation with advancement of animal erythrocytes of control and CST treated young
age. Chloroform extract of Solanum trilobatum and aged rats. Erythrocyte glutathione status was
41 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
observed to be at normal levels in control and Sangeetha et al., 200514 who also specifies
treated young rats. Remarkable decline in GSH modifications in the surface charge due to an
level (38%) with increase in GSSG level (66%) increase in the protein carbonyl levels in aged
was noted in aged rat erythrocytes. These rat erythrocytes. Furthermore, enhanced level of
alterations indicate increased oxidation of carbonyl formation can be endorsed to age
glutathione in erythrocytes with advancement of dependent changes in the rate of oxidized
age. Further significant decrease in both protein degradation44, 45. Addition reactions by
GSH/GSSG ratio and redox index by 2.7 fold highly reactive intermediate products of LPO or
was also observed in aged rat erythrocytes. CST and glycosidation, and direct oxidative
treatment to aged rats increased the GSH level modification of macromolecules during
by 41% and reduced GSSG levels by 40% with oxidative stress may probably be the reasons for
subsequent increase in GSH/GSSG ratio by 2.3 the modification of proteins46. Though several
fold and redox index by 1.4 fold. antioxidant defense systems have evolved to
prevent free radical mediated damage, oxidized
4. DISCUSSION proteins appear to accumulate with animal age47
Cell membrane is an important target for all and may represent 30-50% of the total protein in
radical damages and blood can reflect the old cells48. Thus, increased oxidative damage on
liability of the whole animal to oxidative membrane proteins during animal aging may
conditions, erythrocytes and erythrocyte lead to the attenuation of negative surface charge
membranes have been used extensively for in erythrocytes.
determining the effects of ageing. Change in Additionally, the observed decrease in
erythrocyte membrane accompanying red cell glycoproteins may also be attributed to the
ageing, lead to certain disorders such as - surface charge determination with advancement
thalasemia, sickle cell anemia, and hereditary of animal age. It has been publicized that
spherocytosis and therefore responsible for the sialosaccharide chains of glycophorin A changes
selective removal of erythrocytes prematurely the sialic acid levels upon oxidation that
from circulation. Preservation of the cell eventually decreases membrane surface charges
membrane structure and suitable charge levels thereby contributing to rouleaux formation11.
on its surface decides the accurate course of Earlier studies have made known, decline in
many processes. Alteration in erythrocyte sialic acid levels on erythrocytes decreases
membrane surface charge being a measure of surface charge level leading to the appearance of
cell condition indicated the augmented risk of non-IgG covered epitopes on the surface of
erythrocytes aggregation involving in the oxidized erythrocytes49, therefore allowing the
changes in erythrocyte structure, functions and recognition of erythrocytes by macrophages11.
pathologic conditions such as chronic renal Furthermore, Vomel (1984)50 has confirmed the
insufficiency and purulent intoxication43. In decrease of sialic acid in old individuals result in
accordance the present study demonstrated a surface charge modifications.
significant reduction of surface charge in aged Chloroform extract of Solanum trilobatum
rats when compared with young rats. This supplementation lead to an increase in
decrease could be possibly due to the increased glycoproteins especially sialic acid level and
protein oxidation leading to carbonyl formations ultimately increased the membrane surface
in aged rat erythrocytes. Consequently, our charge. A significant increase in surface charge
results are in accordance with the work of noted in erythrocytes of Chloroform extract of
42 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
Solanum trilobatum treated rats would have house the production apparatus of these radicals
been possibly due to the preservation of protein and since membranes suffer great damage from
carbonyl levels by polyphenolic compounds these radicals, modification of macromolecules
functioning as an in vivo antioxidant that has been proposed to play a major role in the
prevents protein oxidation51 with advancement process of animal ageing57. Significant increase
of age. These alterations may perhaps be due to in SOD and CAT activity on supplementation of
the free radical scavenging and reduced thiol Chloroform extract of Solanum trilobatum to
group elevating properties of near normalcy in aged rat erythrocyte may be
52
phytochemicals that protects the membrane due to the potential quenching of free radicals24
proteins from oxidative insults. Moreover the by phenolic acids present in it. Further reports
ability to maintain redox state of sulphydryl have suggested that polyphenols in CST are
groups in membrane proteins by effectual scavenger of superoxide and hydroxyl
53
flavonoids would has contributed to the radicals58 thereby sparing the antioxidant
maintenance of glycoprotein levels and thus the enzymes valuable in protecting erythrocytes
membrane surface charge in erythrocytes of from oxidative damages.
aged rats. Glutathione, a tripeptide containing -glutamic
The inequity flanked by protective antioxidant acid, cysteine and glycine, provides the first line
defense and increased radical production during of defense against ROS and protects
aging would transform the red cells towards erythrocytes from oxidative damage that acts as
oxidative stress resulting in amendment of a ―radical quencher‖. The glutathione redox
membrane properties and cell dysfunction. The enzymes, glutathione peroxidase, glutathione-s-
biological effects of reactive oxygen species are transferase, glutathione reductase and glucose-6-
tightly controlled by a wide spectrum of phosphate dehydrogenase provide the second
enzymatic antioxidant defense systems54. line of defense through detoxification of noxious
Erythrocytes being regularly exposed to free by-products and alleviation of free radicals
radicals are however equipped with antioxidants mediated macromolecular damages in
59
far in excess of normal requirement and also erythrocytes . An intense increase in the level
function as an effective oxidative sink in the of GSSG with a concomitant depletion in the
organism55. Enzymatic antioxidants present in concentration of GSH, the redox status and the
the cells significantly delay and prevent glutathione metabolizing enzymes may represent
oxidation and related damages. Cu-Zn SOD, a an imbalance in prooxidant and antioxidant
major antioxidant enzyme in erythrocytes, status in aged rats. Significant decrease in the
protects against oxygen free radicals by activity of glutathione metabolizing enzymes is
catalyzing the removal of superoxide radical well correlated with the declined availability of
(O2 -) and catalase, (CAT) a haemoprotein, its substrate, glutathione with advancement of
primarily works to catalyze the decomposition age14. The elevated exposure of erythrocytes to
of hydrogen peroxide to water. In particular, the free radicals may likely be the motivation for
oxidation or autooxidation of hemoglobin (Hb- GSH depletion6. Further studies suggested that
- enhanced utilisation of glutathione by enzymes
the continuous formation of superoxide radicals such as GPx, GST and reduced activity of
56
which is reflected as declined activities of glutathione regenerating enzyme G6PD, due to
SOD and CAT in erythrocytes of aged rat in the the upsurge of reactive oxygen species
present study. Because cellular membranes production may decrease the GSH status in
43 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
erythrocytes of aged animals61. Thus fall in the 2. Halliwell B, The antioxidant paradox.
activity of G6PD essential for an adequeate Lancet 2000; 355:1179 – 1180.
supply of NADPH (enzymes involved in GSH 3. Sinclair AJ, Barnett AH, Lunec J. Free
regeneration) creates an imbalance in radicals and antioxidant systems in health
GSH/GSSG ratio62 and thereby a shift in the and disease. Br J Hosp Med. 1990; 43: 334-
redox state of cells with advancement of age63. 44.
The flavanoids and polyphenols in Chloroform 4. Bandyopadhyay U, Das D, Banerjee RK.
extract of Solanum trilobatum protected the Reactive oxygen species, oxidative stress
sulphydryl groups and thiols from oxidative and pathogenesis. Curr. Sci. 1999; 77:658–
damage thereby elevating their levels64. Also the 666.
metal chelating action of Chloroform extract of 5. Levine RL, Williams JA, Stadtman ER,
Solanum trilobatum improved the activities of Shacter E. Carbonyl assays for
glutathione metabolizing enzymes (GPx, GST, determination of oxidatively modified
GR, and G6PD) in aged rats65. Moreover the proteins. Methods Enzymol. 1994; 233:
decrease in free radical levels by polyphenols in 346-57.
CST might also have contributed to the 6. Spitelle,r G. Are changes of the cell
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enzymes66. the aging process? Ann N Y Acad Sci.
2002; 959:30-44.
5. CONCLUSION 7. Grune T, Shringarpure R, Sitte N, Davies
Ageing can thus be viewed as an irreversible K. Age-related changes in protein oxidation
process associated with the accumulation of and proteolysis in mammalian cells. J
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extract of Solanum trilobatum potentially 11:459-67.
enhances the antioxidants levels by protecting 8. Jovtchev S, Djenev I, Stoeff S, Stoylov S.
erythrocyte membrane from free radical attack Role of electrical and mechanical
and eventually preventing the loss of surface properties of red blood cells for their
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aged have been attracting considerably as a 92.
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Pintigny D, Abadie E, Passa P, Caen J.
Increased adhesion of erythrocytes to
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Table 1: Levels of GSH, GSSG, GSH/GSSG and redox state in control and CST treated young and
aged rats

Young rats Aged rats

Parameters

Group Ia Group Id Group IIa Group IId

GSH 3.12 ± 0.37 3.20 0.39 1.95 ± 0.21a* 2.76 ± 0.34b*

GSSG 0.06 ± 0.006 0.06 ± 0.007 0.10 ± 0.01a* 0.06 ± 0.006b*

GSH/GSSG Ratio 52.00 ± 5.03 53.33 ± 4.18 19.50 ± 2.15a* 46.33 ± 5.12b*

Redox State 0.27 ± 0.02 0.27 ± 0.03 0.17 ± 0.00a* 0.24 ± 0.02b*

Group Ia – Young control, Group Id– Young CST treated, Group IIa – Aged control, Group IId –
Aged CST treated Units: GSH, GSSG: µmoles/g Hb. Values are expressed as Mean ± SD for six
rats.'a' - Group IIa compared with Group Ia, 'b' - Group IIb compared with Group IIa. *
represents p < 0.05

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Figure 1 Level of surface charge in erythrocytes of control and CST treated
young and aged rats. The surface charge was analyzed using two
phase aqueous system and expressed in terms of partition co-
efficient ratio

1
Partition co-efficient ratio

b*
0.8
a*
0.6

0.4

0.2

0
Group Ia Group Id Group IIa Group IId

Group Ia – Young control, Group Id– Young CST treated, Group IIa – Aged control, Group IId – Aged CST
treated.Values are expressed as Mean + for six rats. 'a' - Group Ia compared with Group IIa, 'b' - Group IIa
compared with Group IId * represents p < 0.05

Figure 2: Levels of erythrocyte membrane and plasma protein carbonyls in


control and CST treated young and aged rats

Group Ia – Young control, Group Id– Young CST treated, Group IIa – Aged control, Group IId – Aged CST treated.Values a
as Mean + for six rats. 'a' - Group Ia compared with Group IIa, 'b' - Group IIa compared with Group IId * represents p < 0.0

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Vol. 03 issue 07 July 2011
Figure 3 Levels of Glycoprotein in erythrocytes and plasma of
control and CST treated young and aged rats

600
b*

500
a* b*

400

a*
300

200

100 b*
a*

0
Group Ia Group Id Group IIa Group IId

Hexose Hexose amine Sialic acid

Group Ia – Young control, Group Ib – Young CST treated, Group IIa – Aged control, Group IIb – Aged CST
treated Units: µg/mg protein Values are expressed as Mean ± SD for six rats.'a' - Group IIa compared with
Group Ia, 'b' - Group IId compared with Group IIa * represents p < 0.05

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Vol. 03 issue 07 July 2011
51 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
LYSOSOMAL ENZYMES

Jayamathi G1, Vishnu Priya V2


1
Meenakshi Ammal Dental College, Meenakshi Academy of Higher education and
ijcrr Research, Alapakkam Main Road, Chennai. Tamil Nadu
2
Vol 03 issue 07 Department of Biochemistry, Saveetha Dental College & Hospital, Saveetha
Category: Review University, Velappanchavadi, Chennai, Tamil Nadu
Received on:14/04/11
Revised on:03/05/11 E-mail of corresponding author: gjayamathe@gmail.com
Accepted on:10/05/11

ABSTRACT
Lysosomal enzymes are implicated in tissue remodeling and in regulating the immune responses.
Lysosomal enzymes can be incorporated into the explanation of mechanisms of development of various
diseases and give scientific grounds for prevention of inflammatory disease. This review highlights
synthesis, functions and regulation of lysosomal enzymes.
______________________________________________________________________________

Key words: Phagocytosis, endocytosis, acid intended to degrade ingested microbes, could
hydrolases, lysosomes, alpha1 –antitrypsin also lead to tissue destruction and amplification
of inflammatory response with continued
INTRODUCTION recruitment of new leukocytes. Altered
Lysosomes are small intracellular organelle lysosomal membrane stability leads to release
present in all animal cells. They destroy any lysosomal hydrolases, ensuing altered
foreign material which enters the cell such as metabolism of different connective tissue
bacteria or virus. Lysosomes also remove the constituents including collagen and also
worn out and poorly working cellular organelles involved in the destruction of non - collagenous
by digesting them to make way for their new components of the extracellular matrix. Hence
replacements. Since they remove cell debris, the present study is designed to give precise
they are also known as scavengers, cellular account on lysosomal enzymes, may open up
2
housekeepers or demolition squads. Lysosomes new horizons in the research field.
form a kind of garbage disposal system of cell.
During breakdown of cell structure, when the Synthesis of lysosomal enzymes
cell gets damaged, lysosomes burst and the More than 50 hydrolases involved in the
enzymes eat up their own cells. So, lysosomes lysosomal degradation of protein, carbohydrate,
are also known as suicide bags of a cell. The lipids and nucleic acids have been identified.
major function of lysosomes and lysosomal The hydrolases are enclosed by a membrane
proteases is not to kill the cell but to take care of containing a set of highly glycosylated
cellular homeostasis and possibly differentiation lysosomal membrane proteins. The targeting of
1
by recycling cellular components. acid hydrolases depends on the presence of
The release of lysosomal enzymes are normally mannose-6-phosphate (M6P) residues that are
52 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
recognized by specific receptors mediating the modified by the addition of complex sugar
intracellular transport to an endosomal or residues , sulfate groups and by the formation of
4
prelysosomal compartment. The lysosomal M6P recognition marker.
apparatus is responsible for the intracellular Enzymology of lysosomes: Some important
digestion of externally and internally generated enzymes found within lysosomes include:
macromolecules. Coated vesicles internalize Lipase, which digests lipids
most extracellular macromolecules by
Amylase, which digests amylose, starch, and
endocytosis to form early endosomes, which
maltodextrins
move from the plasma membrane towards the
cell nucleus. They become acidic and give rise Proteases, which digest proteins
to 'late' endosomes. This increasing acidity leads Nucleases, which digest nucleic acids
to the dissociation of lysosomal enzymes. Late
endosomes also fuse with primary lysosomes Phosphoric acid monoesters
(which contain lysosomal hydrolases and bud The proteolytic capacity of lysosomes comprises
from the Golgi) to form secondary lysosomes. a mixture of endo- and exo-peptidases, called
Secondary lysosomes might remain in the cell cathepsins, which act in concert to degrade
and become residual bodies, or be transported to proteins to a mixture of amino acids and
5&6
the cell surface, where they fuse with the plasma dipeptides. Some cathepsins, for example, G
membrane and exocytose their digested and E, also function outside the lysosome. All of
3
materials. the proteases are active at an acidic pH, although
Lysosomal enzymes are synthesized with an N- this may not be their pH-optimum. They are
terminal sequence of 20-25 aminoacids synthesized in the form of inactive precursors,
recognized by signal recognition particle which preproenzymes, which are transported to the
enable the nascent polypeptide to be transferred lysosome by the mannose-6-phosphate pathway
across the membrane of endoplasmic reticulum. like other lysosomal hydrolases. Proteases are
Signal peptidase removes signal peptide. classified by the catalytic residue in the active
Preformed oligosaccharides undergo N- site involved in the mechanism of peptide bond
glycosylation with asparagine residue. cleavage. Cathepsins with a serine (cathepsins A
Furthermore sulfatase family members are and G), cysteine (B, C, F, H, K, L, O, S, and W)
formed from sulfated mono and polysaccharides, or an aspartic acid (D and E) residue in the
6
glycolipid and hydroxyl steroids, and are active site have been characterized.
modified in endoplasmic reticulum. Endopeptidases
Lysosomal enzymes are synthesized and are Cysteine proteases: cathepsins B, C, H, K, L,
glycosylated in the rough endoplasmic O, S, and W
reticulum. They are then transferred to the Golgi Aspartyl proteases: cathepsins D and E
bodies, where they acquire mannose-6- Serine proteases: cathepsin G in azurophil
phosphate (M6P) residues on their high- granules of neutrophils
mannose and hybrid-type oligosaccharide
chains. This recognition marker is specific to Exopeptidases
lysosomal hydrolases and allows these Carboxypeptidases: lysosomal
hydrolases to be sorted from other proteins. carboxypeptidase (cathepsin A or protective
Upon arrival of golgi the oligo saccharide chain protein)—serine protease; cathepsin B
of lysosomal enzymes are further trimmed and
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Vol. 03 issue 07 July 2011
(dipeptidase); cathepsin X, mono- or consistent with the major role of this cell in
dipeptidase; lysosomal carboxypeptidase B; chronic inflammation.
prolylcaboxypeptidase; peptidyl dipeptidase B In contrast, mast cells, granulocytes and
Aminopeptidases: cathepsin H—true platelets, which contribute primarily to acute
aminopeptidase; dipeptidyl peptidase I inflammation, exhibit rapid degranulation
(cathepsin C); dipeptidyl peptidase II; tripeptidyl processes. Platelets secrete lysosomal enzymes
peptidase (TPP-I) during the "platelet release reaction" early in clot
formation. During the "platelet release reaction"
Lysosomal enzymes in various cells: induced by thrombin or collagen, mammalian
Lysosomes are subcellular organelles which blood platelets secrete lysosomal enzymes into
8&9
perform many important cellular functions. For the surrounding medium. Finally,
example, lysosomes digest foreign material and inflammatory substances may leak from cells
engulfed viruses and bacteria presenting in simply as a result of cell death due to plasma
phagosomes during the process of phagocytosis. membrane injury . A number of animal,
The influx of neutrophils and mononuclear bacterial, and chemical toxins, as well as
phagocytes into tissues may be seen as the synthetic detergents may cause such lysis of the
hallmark of inflammation and significantly outer cell membrane.
contributes to both the injury and the subsequent 10
FUNCTIONS OF LYSOSOMES
repair seen in the normal tissues.
Cellular Digestion: Lysosomal enzymes degrade
Lysosomes are found in all eukaryotic cells, but
proteins into dipeptides and carbohydrates into
are most numerous in disease-fighting cells,
7 monosaccharides. Sucrose and polysaccharides
such as leukocytes found that peritoneal are not digested and remain in the lysosomal
macrophages in culture release lysosomal vacuoles.
enzymes in response to phagocytic, but the Autophagy: By the process of autophagy,
mechanisms that regulate macrophage lysosomal lysosomes constantly remove cellular
enzyme secretion are not fully understood. components like mitochondria etc. Cytoplasmic
Polymorphonuclear leukocytes also release organelles become surrounded by smooth
7
lysosomal enzymes during phagocytosis and the endoplasmic reticulum and lysosomes attach
mechanisms that control this secretary process with it and discharge their contents into
are well documented. autophagic vacuole and the organelle is digested.
Macrophage lysosomal enzyme release has Autophagy is a general property of eukaryotic
many similarities to secretion from cells.
polymorphonuclear leukocytes and it is tempting Exocytosis: Contents of the primary lysosome
to suggest that the processes might be regulated mat be released into the medium by exocytosis
in the same way. Thus, macrophage lysosomal and it occurs during replacement of cartilage by
enzyme release is not controlled by the same bone during development where osteoclasts
regulatory mechanisms as the degranulation release lysosomal enzymes . It can also occur in
processes in polymorphonuclear leukocytes, bone remodeling under influence of parathyroid
platelets, and mast cells. Lysosomal enzyme hormone. Crinophagy refers to the process by
release from macrophages is a much slower which secretary granules produced in excess are
process than secretion from the other cell types removed by lysosomes.
and this may be functionally very important.
Prolonged enzyme release from macrophages is
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Vol. 03 issue 07 July 2011
Endocytosis: Lysosomes may fuse with vesicles phagosomes open at their external border to
or vacuoles formed by endocytosis and release tissue space while joined at their internal border
their enzymes into it for digestion. The material with granules discharging acid hydrolases into
for digestion may be food (protozoa) or a the vacuole (phagolysosome). Under such
foreign body like parasite .The products of circumstances lysosomal enzymes are
digestion are absorbed and assimilated leaving selectively released to the outside of the cell
undigested which are released outside by without necessarily causing cytoplasmic
12
exocytosis. damage. Reverse Endocytosis: It and may be
Role in germ cells and fertilization: The pertinent to the pathogenesis of tissue injury.
acrosome in spermatozoa may be considered as When leukocytes encounter immune complexes
a special lysosome containing protease and which have been dispersed along a
hyaluronidase along with acid phosphatase .The nonphagocytosable surface,there is similar,
lysosome in ova help in digestion of stored food selective release of lysosomal enzymes directly
Role of lysosomes in diseases: Lysosomes are to the outside of the cell.
involved in many diseases like rheumatoid Perforation from within: Another mechanism for
arthritis, silicosis, acute inflammatory responses, lysosomal enzyme release followed
anorexia, myocardial infarction, different phagocytosis of crystals was due to "perforation
storage diseases etc. from within" of the lysosomal membrane, rather
Lysosomal enzymes release than lysis by crystals of the plasma membrane.
Although a series of studies have indicated that Enzyme release occurs when certain materials
mechanism which account for release of gain access to the vacuolar system wherein they
lysosomal enzymes can provoke acute interact with, and finally rupture, lysosomal
inflammation, may progressed to chronic state membranes . A wave of membrane damage
also. results with release of cytoplasmic and
Regurgitation during feeding: Human lysosomal enzymes followed by cell and tissue
neutrophils release lysosomal hydrolases during 11
death.
phagocytosis. Microtubules were more
Regulation of lysosomal proteases
prominent in phagocytosing than in resting cells,
Selective secretion of lysosomal enzymes from
and were observed near primary lysosomes and
neutrophils during acute inflammation.
forming phagosomes. ―Regurgitation during
Discharge of lysosomal content is requires
feeding‖ resulted from degranulation of primary
extracellular calcium and can be modulated by
lysosomes into newly formed phagosomes
several different classes of hormones, protease
which were still open to the extracellular space
inhibitors such as α2 -macroglobulin and α1-
as well as from the ingestion of additional
antiprotease, drugs and other agents which
material directly into already loaded secondary
11
results in the provocation of acute inflammation
lysosomes. and connective tissue degradation. These
Phagocytosis: When cells engage in antiproteases are present in serum and synovial
phagocytosis (leukocytes which engulf immune fluids. They are thought to function by binding
complexes in the synovial fluid of patients with to and covering the active sites of proteases.
rheumatoid arthritis) they release a portion of Protease-antiprotease imbalance is probably
their lysosomal hydrolases into the surrounding important in the pathogenesis of emphysema.
medium . This effect appears due to extrusion of The most prominent protease inhibitor in human
lysosomal materials from incompletely closed
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Vol. 03 issue 07 July 2011
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concentration of all inhibitors and is responsible Bio sci. 1983; 5: 101–104.
for approximately 90% of the total trypsin- 4. Dell'Angelica EC, Mullins C, Caplan S and
inhibiting activity of normal serum α1- Bonifacino JS. Lysosomal related
antitrypsin is a glycoprotein with a carbohydrate hydrolases. FASEB J. 2000;14: 1265-1278.
portion of 12.4% containing galactose, 5. Barrett AJ, Rawlings ND and JF. Jr.
mannose,fucose, acetyl hexosamine, and sialic Woessner, 1998. Handbook of Proteolytic
acid. Its amino acid composition is Enzymes. Academic Press, London.
unremarkable except perhaps for the content of 6. Mason RW, 1996. Lysosomal Metabolism
13&14
only two cysteine residues of Proteins. In: Subcellular Biochemistry:
Biology of Lysosome, Lloyd, J.B. and R.W.
CONCLUSION Mason (Eds.). Vol. 27, Plenum Press, New
Concluding, the present study, the lysosomal York, pp: 159–190.
enzymes are crucial for the degradation of 7. Weissmann G, Zurier RB, Spieler PJ and
numerous macromolecular substrates and have Goldstein IM. Mechanisms of lysosomal
been involved in many inflammatory responses. enzyme release from leucocytes exposed to
Our understanding of the importance of immune complexes and other particles. J
lysosomal cysteine proteases has advanced Exp Med. 1971;134: 149s-165s
considerably in recent years. It is now evident 8. Davey MG and Luscher EF. Release
that they regulate biological processes such as reactions of human platelets induced by
matrix remodeling and the immune response. thrombin and other agents. Biochiini.
Although their exact roles in the pathobiology of Biophys. Acta. 1968;165: 490-506
various diseases are uncertain, continued 9. Holmsen H and Day HJ. The selectivity of
research should clarify their roles in various the thrombin-induced platelet release
grounds. Accurate knowledge of lysosomal reaction: Subcellular localization of released
enzyme is essential to expand our current and retained constituents. J Lab Clin Med;
understanding of intracellular proteolysis that 1970;75: 840-855.
plays important role in health and disease. 10. Luzio JP, Pryor PR and Bright NA.
Lysosomes: Fusion and function. Nat Rev
REFERENCES Mol Cell Bio. 2007; 8: 622-632.
1. Turk B and Turk V. Lysosomes as Suicide 11. Weissmann G. Lysosomes and joint disease.
Bags in cell death: Myth or reality? J Biol Arth Rheum.1966; 9: 834-840.
Chem 2009; 284: 21783-21787 12. Becker EL,. Some interrelations of
2. Layik MN, Yamalik F, Caglayan K, Kilinc neutrophil chemotaxis, lysosomal enzyme
I, Etikan and Eratalay K. Analysis of human secretion and phagocytosis as revealed by
gingival tissue and gingival crevicular fluid synthetic peptides. Am J Pathol 1976; 85:
beta-glucuronidase activity in specific 385-394.
periodontal diseases. J Periodontol. 2000; 13. Parrott DP and Lewis DA. Protease and
71: 618-624. antiprotease levels in blood of arthritic rats.
3. Varki AP, Reitman Ml, Tabas I and S. Ann Rheumatic Dis. 1977;36: 166-169.
Kornfeld,. Studies of the synthesis, structure 14. Kueppers F. α1,-Antitrypsin. Am J Hum
and function of the phosphorylated Genet 1973; 25: 677-686

56 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
EVALUATION OF ANTHELMINTIC ACTIVITY OF
MOMORDICA CHARANTIA FRUIT EXTRACT
Shripad M.Bairagi1, Ritesh S. Mantri2, Nitin Nema3
1
Department of Pharmacology, MES College of Pharmacy, Sonai, Newasa,
ijcrr Ahmednagar, Maharashtra
2
Vol 03 issue 07 MES College of Pharmacy, Sonai, Newasa, Ahmednagar, Maharashtra
3
Category: Research Department of Pharmaceutical Sciences, Dr.H.S.Gaur Central University,
Received on:28/04/11 Sagar(M.P.)
Revised on:16/05/11 E-mail of corresponding author: ssanandss@yahoo.com
Accepted on:27/05/11

ABSTRACT
The present study is an attempt to explore the anthelmintic activity of Methanolic extract of fruits of
Momordica charantia. Various doses of methanolic extract were evaluated for their anthelmintic activity
on adult Indian earthworms,Pheretima posthuma. All extract able to show anthelmintic activity at 25, 50
and 100 mg/ml. The activities are comparable with standard drugs, piperazine citrate and albendazole. All
doses of momordica charantia showed dose dependent anthelmintic activity in comparison to standard
drug. The data were found statistically significant. It is concluded that methanolic extract of M.charantia
is having anthelmintic activity.
______________________________________________________________________________

Key words: Momordica charantia, The hypoglycemic chemical includes mixture of


Cucurbitaceae, Anthelmintic, Piperazine citrate, steroidal saponin known as charantins, insulin
Albendazole. like peptide and alkaloid. M.charantia fruits and
seeds has been shown to reduce total cholesterol
INTRODUCTION and triglycerides in both the presence and
Momordica charantia (Fam: Cucurbitaceae) absence of dietary cholesterol [3,4].Noval
grows in tropical areas. The herbaceous, tendril- phytochemical in M.charantia demonstrated the
bearing vine grows to 5 m. It bears simple, ability to inhibit an enzyme named guanylate
alternate leaves 4–12 cm across, with 3–7 deeply cyclase. This enzyme thought to be linked to
separated lobes. Each plant bears separate pathogenesis and replication of not-only
yellow male and female flowers. Fruit has a psoriasis but leukemia and cancer as well. [5-
7]
distinct warty looking exterior and an oblong The phytochemical momordin has clinically
shape, it is hollow in cross section, with a demonstrated cytotoxic activity against
relatively thin layer of flesh surrounding central Hodgkin‘s lymphoma in vivo. [8]

seed cavity filled with large flat seed and pith.[1]


M.charantia contain novel and biologically MATERIAL AND METHOD
active phytochemicals including triterpens, The fruit of Momordica charantia were
proteins and steroids. In numerous studies, at collected from M.P.K.V.Rahuri, Maharashtra,
least three different group of constituent found India. And was authenticated by botanical
in all parts of M.charantia have clinically survey of India, Pune, Maharashtra.
demonstrated hypoglycemic properties. [2]
57 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
Preparation of Extract: variance (Anova) at 5% level significance was
The methanolic extract of air dried fruit powder employed. P values < 0.05 were considered
(500 gm) was prepared by using soxhlet significant.
apparatus, concentrated and vaccum dried which
give dark a brownish mass (62.20gm). RESULTS
Anthelmintic Bioassay: The methanolic extract of M.charantia produced
Healthy Indian earthworms (Pheretima a significant anthelmintic activity in dose
posthuma) selected due to its anatomical and dependent manner as shown in Table-1. Normal
physiological resemblance with the intestinal saline water used as control. The activity shown
round worm parasites of human being were used by methanolic extract is of considerable
in the present study.[9-11].All the earthworms importance. All data were found to be
were of approximately equal size. They were statistically significant at 5% level of
procured from local supplier and maintained at significance (P<0.05) when subjected to one
MES College of Pharmacy, Sonai. way ANOVA. The extent of activity shown by
The methanolic extract of Momordica charantia the extract was found to be dose dependent and
was tasted in various doses in each group. same effect as that of piperazine citrate (10
Normal saline water used as control. Piperazine mg/ml) and albendazole (10mg/ml) was seen as
citrate and albendazole were used as standard shown in Fig.1.
drug for comparative study with methanolic
extract.The anthelmintic activity was assessed DISCUSSION
using earthworm by the Naragund [12] reported The anthelmintic of the methanolic extract was
method. Earthworm divided into six groups. comparable with that of standard drugs.
Each group containing five earthworms. First Albendazole and piperazine citrate are used as
group (I) serve as normal control which receive anthelmintic drugs but they having some adverse
saline water only. Second (II) group receive effect such as bronchospasm, GIT disturbance
standard piperazine citrate 10 mg/ml, third group and they are contraindicated in pregnant
(III) receive standard albendazole 10 woman.So if we formulate the anthelmintic drug
mg/ml.Group four (IV), group five (V) and from M.charantia will show very less adverse
group six (VI) receive dose of methanolic effect because of herbal formulation. And the
extract of 25 mg/ml, 50 mg/ml and 100 mg/ml M.charantia easily available in local market so
respectively. the formulation cost will be low. More work is
Observations were made for the time taken to required to identify the main active principle
cause paralysis and death of individual worm for responsible for anthelmintic activity.The above
two hours. Paralysis was confirmed when the result reveled that methanolic extract of
worm did not revive even in normal saline M.charantia is having anthelmintic activity.
water. Death was concluded when worms lost
their motility followed by fading away of their CONCLUSION
body colour. [13] The fruit extract of M.charantia has activity
against Pheretima posthuma. However the
Statistical analysis: activity observed in this study, would appear to
The data expressed as mean ± standard deviation justify the ethonomedical use in the recipes for
(n=5). For determining the statistical anthelmintic activity. Further studies would
significance, standard error mean and analysis of
58 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
focus on isolation of the bioactives, biological 6. Claflin AJ,Inhibition of growth and
and chemical characterization. guanylate cyclase activity of an
undifferentiated prostate adenocarcinoma by
ACKNOWLEDGEMENT an extract of the balsam pear (momordica
The authors are grateful to Dr.V.K.Deshmukh, charantia abbreviata),J. Proc Nati.Acad.
Principal MES College of Pharmacy, Sonai, Sci.,75(2),1978,989-93.
Ahmednager for providing the lab facilities for 7. Vesely DL, Isolation of a guanylate cyclase
the research work. inhibitor from the balsam pear(momordica
charantia
REFERENCES abbreviate)J.Biochem.Res.Commun.77(4),1
1. Grover JK,Yadav SP,Pharmacological 977,1294-99.
actions and potential uses of momordica 8. Terenzi A, Anti-CD30 (BER=H2)
charantia,J.Ethnopharmacol,93 immunotoxins containing the type-1
(1),2004,123-32. ribosome-inactivating proteins momordin
2. Raza H,Moduation of xenobiotic and PAP-S (pokeweed antiviral protein from
metabolism and oxidative stress in chronic seeds) display powerful anitumour activity
streptozotocin induced diabetic rat fed with against CD30+tumor cells in vitro and in
momordica charantia fruit extract,J.Biochem SCID mice.Br.J.Haematol,92(4),1996,872-
Mol.Toxicol.14(3),2000,131-39 79.
3. Jayasooyiya,AP, Effect of momordica 9. Vidyaethi RD,A textbook of zoology,Chand
charantia powder on serum glucose level and Co.Press,New delhi, 14th edn, 1977,329.
and various lipid parameters in rats fed with 10. Thron GW,Harrison‘s principles of internal
cholesterol free and cholesterol rich medicine, MC Grew Hill,New
diets,J.Ethnopharmacol,72(1-2),2000,331-36 York,1977,1088.
4. Ahmed I,Hypotriglyceridemic and 11. Vigear Z, Atlas of medical parasitology,
hypocholesterolemic effect of anti-diabetic Publishing House,Singapore,2nd
n
momordica charantia (karela) fruit extract in ed ,1984,216.
streptozotocin induced diabetic rat, 12. Jayachandran E,Bhatia K,Naraguand V and
J.Diabetis Res.Clin.Pract,51(3),2001,155-61 Ray A,Anthelmintic activity of 2[3-amino,5-
5. Takemoto,DJ,Partial purification and 8 methyl-6-carboxamidepyrazol-1-yl] 6-
characterization of a guanylate cyclase fluro-7-substituted (1,3) benzothiazoles on
inhibitor with cytotoxic properties from the pheritima postuma, Indian
bitter melon(momordica drugs,40(7),2003,408.
charantia),J.Biochem.Biophys.Res.Commun 13. Mali RG,Hundiwale JC,Sonawane RS,Patil
,94(1),1980,332-39. RN,Indian J.Nat.Prod.,20(4),2004,10.

59 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
Table-1 Anthelmintic activity of methanolic extract of Momordica charantia

Group Treatment Dose (mg/ml) Time taken for Time take for death
paralysis (min) (min) Mean±S.D.
Mean± S.D.
I Control (Normal ….. ….. …..
saline water)
II Standard-1 (Piperazine 10 28.2±0.30 46.4±0.81
citrae)
III Standard-2 (Albendazole) 10 26.3±0.61 39.7±0.61
IV Metahnolic Extract 25 27.2±0.31 48.3±0.38
V Metahnolic Extract 50 25.1±0.63 40.2±0.71
VI Metahnolic Extract 100 23.6±0.74 36.4±0.51

Each value represented as mean±standard deviation. When compared with standard drug using one way ANOVA.

Fig-1 Anthelmintic activities of methanolic extract of fruits of Momordica charantia on Indian


earthworms (Pheretima posthuma).

Group-I control (normal saline water), group-II-standard-1(piperazine citrate), group-III-standard-


2(albendazole),group-IV to VI methanolic extract of dose 25,50,100 mg/ml respectively .

60 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
HEALTH SEEKING BEHAVIOR OF HIV POSITIVE PATIENTS
ATTENDING VOLUNTARY COUNSELING AND TESTING
CENTER – A GENDER PERSPECTIVE
Shaikh Mohsin, Patil Rajkumar, Pathan Sameer
1
Department of Preventive and Social Medicine, Government Medical College
ijcrr Baroda, Vadodara, Gujarat
2
Department of Preventive and Social Medicine,Mahatama Gandhi Medical
Vol 03 issue 07 College and Research Institute, Pondicherry
Category: Research 3
Indian Institute of Public Health - Delhi (IIPH-D), Public Health Foundation of
Received on:03/05/11 India (PHFI), New Delhi
Revised on:16/05/11
E-mail of corresponding author: dr_mohsinshaikh@yahoo.com
Accepted on:26/05/11

ABSTRACT
The epidemic of Acquired Immuno Deficiency Syndrome (AIDS) has emerged as a serious public health
problem in many parts of the world and a gender based difference in the health seeking behavior has
significantly precipitated to it. Aims: To study the health seeking behavior of HIV positive cases and
impact of gender discrimination over it. Settings & Design: It is a cross sectional study conducted in
Voluntary Counseling and Testing Center (VCTC) – Baroda. Methods: A semi-structured and pretested
proforma was used to interview HIV positive patients attending VCTC located at Sayaji Hospital,
Vadodara. With the help of VCTC counselors, In-depth interview of all patients were arranged to collect
the detailed information on health seeking behavior. Prior verbal and written consent was taken before
starting each interview. This study included 100 HIV positive cases (>13years) attending VCTC during
April-December 2007. Results: The present study included 100 individuals with equal ratio of male and
female, 73 % were in age group 21-40 years, 92% were literate and 60 % were married. 54 % patients
consulted private clinic for their health problem while 30 % went to government hospital of which
majority were females (70 %). None of the female patients contacted VCTC initially for counseling
purpose, while 13 % patients didn‘t consult any health care providers before reaching VCTC. 54 %
patients consulted private GP initially of which 21 % didn‘t satisfied and visited Government hospital
later. Over all 48 % patients reached SSG hospital and were referred to skin, TB, urology and general
medicine before reaching VCTC.
______________________________________________________________________________

Key Message: Study of gender based health many parts of the world. Estimates at the end of
behavior is the vital link to control the spread of 2006 suggest that 39.5 million men, women and
HIV. The need of the hour is to strengthen the children are living with HIV/AIDS worldwide
health services with the focus of gender. and almost 22 million have already lost their
lives1. In sub-Saharan Africa, young women
INTRODUCTION (aged 15-24 years) are infected more frequently
The epidemic of Human Immunodeficiency than young men. In 2001, the estimated infection
Virus (HIV) infection that causes Acquired rates for young women were 6-11% compared to
Immuno-Deficiency Syndrome (AIDS) has 3- 6% for young men1.
emerged as a serious public health problem in

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Vol. 03 issue 07 July 2011
Focus of Gender in HIV/AIDS: Of the 37.2 2. Economic barriers: different access to
million adults living with HIV/AIDS at the end resources, 3.Cultural barriers: social status of
of 2006, 17.7 million or nearly 50 percent were women which situates them in socially inferior
women2. Rather than representing equity positions, 4. Education barriers: women having
between the sexes, the one-to-one ratio of male less access to education.
to female HIV infections demonstrates the way
in which gender inequalities affect HIV/AIDS Objective: To study the health seeking behavior
incidence rates. Since the HIV/AIDS pandemic of HIV positive cases and impact of gender
began more than 20 years ago, infection rates discrimination over it.
among women have accelerated. Traditional
Need of the study
gender roles held by many of the world‘s
societies definitely affects the health seeking Women are most vulnerable and likely to get
behavior of women, which continued spread of discriminated with HIV infection, having so
HIV, particularly from men to women. In most many reasons in the background than men who
societies, women also carry a disproportionate plays dominant bread earner role in the society,
amount of the burden of caring for family there was a need of gender based research to go
members living with HIV/AIDS, and experience through the health seeking behavior of the HIV
the brunt of the stigma associated with HIV positive patients to understand the
infection. Correcting the gender imbalance that discrimination. It would also help in policy
contributes to and is exacerbated by the designing for national AIDS control Program.
HIV/AIDS pandemic will depend upon
improving women‘s social and economic status, METHODOLOGY
and increasing men's responsibility for HIV This is VCTC based cross sectional study
prevention and care, so as to provide better consisting of qualitative data collection of HIV
health care access. Unequal social roles and positive patients conducted between April ‘07
vulnerability to men‘s demands prevent women to December ‘07. Total number of 50 male and
to have better health care access. Number of 50 female HIV positive patients who visited
facts associated with HIV transmission like VCTC, Vadodara during the study period in age
illiteracy, employment, gender discrimination group >13 years were purposively selected for
poor health infrastructure etc., stimulates already this research. The following selected criteria was
prevalent stigma related to HIV/AIDS. Finally adopted for inclusion of HIV positive patients
all these factors such as gender roles, economic, attending VCTC in this research, 1) 50 adult
cultural and social factors and stigma related to male and 50 adult female >13 years of age. 2)
HIV/AIDS are likely to influence the motivation HIV positive patients diagnosed minimum 2
of HIV infected people to practice safer sex3. months back. 3) Patient who voluntarily took
United Nations for HIV/AIDS has reported 5.7 part in the research and visited VCTC – SSGH
million people in India with HIV/AIDS this after HIV infection. 4) Patients with HIV
year, which is almost 15 % of the global burden antibody test positive according to NACO
of HIV/AIDS, and 37 % of all the infected guidelines6. (ELISA & Rapid test).
people in India are women4. Nash Ojanuga & Information on Health seeking behavior and
Gilbert (1992) systematized the obstacles which access to health care of HIV positive patient was
women face into four categories5: 1.Institutional collected through semi-structured, pre-designed
barriers: unequal treatment by health providers, and pre-tested proforma following in-depth

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Vol. 03 issue 07 July 2011
interview technique with the help of health confidentiality. Prior permission from the
seeking behavior pathway (Annexure). In-depth Ethical committee of the university has been
interview of each and every HIV positive patient taken to carry out whole research. Analysis of
was done with the help of expert VCTC the study was done using Epi Info 6.04 d
counselors, through previously prepared Statistical Package and Chi square test was
questionnaire at VCTC in a private room with applied wherever needed.
prior informed, written and verbal consent. Following steps were involved in documentation
Proper care has been taken to maintain the and data management after data collection.

Collecting ‗raw‘ field notes

Writing expanded field notes on same day (in the local language-Gujarati)

Translating field notes (in English)

Coding/Organizing field notes by themes

Data entry

Data analysis

Limitations of study: group among both the genders. Almost 60 % of


We could have been covered VCTCs of Gujarat HIV positive patients were married with
but we didn‘t get the permission to do so and significantly higher percentage of males (58 %)
also could have consulted the other high risk as compared to females (42 %). Whereas total
groups like male and female sex workers, number of widow/widower found is 23 % and
homosexual group, or truck drivers to more than two third of them were females as
understand their health seeking behavior. compared to males. Majority of female patients
(80 %) were housewives while majority of males
RESULTS were involved in high risk occupations like truck
The study included 50 male and 50 female HIV driving, auto driving, call center job and sales
positive (diagnosed minimum 2 months back) work. Of total patients, 8 % were illiterate, 43 %
cases who visited VCTC. Over two third of the patients were educated upto 7th standard, 30 %
patients (73%) were within 21-40 years of age untill 10th standard, 14 % untill 12th standard and
63 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
only 5 % had studied untill graduation or patients who had contacted VCTC initially were
higher; without much gender difference. Of the males (100) as compared to females which may
total economically self dependant patients be due to the fact that males have better
(49%), more than two third were males. Out of awareness towards their health or females don‘t
total 66 % of patients, 42 % had TB infection in have control over resources, while 3 female
the past while 30 % had STD (with equal gender patients were forced to consulted quack either by
distribution) history. their spouse of family members, during their
All HIV positive patients had undergone CD4 initial visit. More females consulted NGOs
count testing of whom 49 % were females and initially (80 %) as compared to males (20%).
51 % were males (Table – 1). Out of 37 patients Out of 100 HIV positive patients studied 13 %
on ART, 78 % (n=29) cases experienced at least patients did not contact any health care provider
one ADR related to ART use. 86 % cases had before reaching VCTC. They either reached
history of ADR in the form of nausea, vomiting VCTC directly or went through NGOs. 72 %
and in 31 % cases, in the form of skin reaction patients consulted less or equal to 3 health care
with no gender difference. providers while 15 % consulted >3 health care
Out of 100 patients, 8 directly went to VCTC providers before reaching VCTC.
without getting referred by anyone and all of Out of 100 patients studied, 8 directly contacted
them were males, while 62 % patients were VCTC voluntary bases immediately. 79 %
referred to VCTC by Doctors ( Private & patients had first health care contact within 6
Government). 7 % patients were referred by the months of observing first sign/symptom by
family members, 6 % by their friends, 5 % by themselves, while 13 % patients took >6 months
NGOs and 9 % of patients by their relatives to contact health care providers. There is no
other than their family members. There was no significant gender difference seen (Table-2). 13
statistically significant gender difference (Table % visited VCTC because of Tuberculosis
2). infection, 10 % due to skin problems, 9 % cases
Table 2, shows the distribution of HIV positive visited VCTC because of their high risk
patients by appearance of first sign/symptoms behavior, 7 % gave a reason of their spouse
noticed by them when asked. HIV positive being HIV positive, 4 % due to penile ulcer and
patients had history of single or multiple same percentage of patients visited VCTC as a
symptoms during initial health problems. part of pre operative investigation. 2 % had
Majority of patients observed weight loss (26 %) pneumonia and were consulted VCTC for HIV
followed by fever (23 %), cough (22 %), skin testing while 2 % were consulted during visa
problems (17 %), diarrhea (10 %), burning procedure for HIV testing and others during
micturition (6 %) and genital ulcer (5 %). There ANC check-ups, for insurance purpose and some
is no statistically significant gender difference MSM behavior. 46 % patients gave a reason that
seen. they visited VCTC for other health problems
54 % patients initially contacted private like low grade fever, chronic cough, diarrhea
clinic/hospital for their symptomatic problems of and weight loss.
whom 59 % were males as compared to 41 % According to health care seeking pathway,
females. 30 % patients contacted government Initially 3 patients directly went to Quack with
hospital initially and 70 % of them were females the hope of getting treated – but finally
as compared to 30 % males, which is statistically consulted GP/SSGH for their treatment. 8
significant (P<0.05, Chi square=19.45). All the Patients directly went to VCTC - Baroda for
64 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
testing and for counseling purpose. 54 Patients problems by their spouse. This was because of
directly went to Private General Practitioner out the male dominance to their decision where as
of which 9 patients changed GP for the 3rd time female has to depend on others physically as
and 2 patients changed GP treatment for the 4th well as economically. Almost 15 % cases
time and finally went to VCTC. 21 Patients who contacted more than three health care providers
consulted private GP for the first time - were not as they were not improved and 13 % patients
satisfied and moved to SSG Hospital/Govt. took more than 6 months to have first health care
CHC/PHC for further testing and treatment contact. 8 patients contacted VCTC directly and
purpose. 38 Patients directly consulted all of them were males, none of them were
SSGH/Govt. CHC/PHC without going to Private females.
GP. 5 Patients went to NGO directly with their A Study on Gender impact of HIV/AIDS in
suspicion of having HIV infection and after India shows that as compared to men, women
counseling they were sent to VCTC. 16 patients have poorer access to healthcare. The financial
consulted Govt. CHC/PHC of which 6 patients constraint is also a big impediment for getting
went to private GP for the first and then to Govt. proper healthcare and it is more so for women7.
CHC/PHC. 8 patients out of those 16 went to Women may be more affected by stigma and
SSGH because of their unsatisfactory treatment discrimination than men because of social norms
and counseling, finally all were guided to concerning acceptable sexual behavior in
VCTC. Over all 48 patients reached SSGH after women, and that‘s why women are often more
consulting one or more GP of whom 15 economically vulnerable than men. Our study
patients reached OPD-1 (Skin and VD OPD) for also reported higher number of economically
skin lesions over body parts and private parts vulnerable females. Gender differences in
while 16 patients reached OPD-18 (General decision-making may also affect access to health
Medicine OPD) for their generalized health facilities. For example, a study conducted in
problems and 8 patients reached OPD-17 (TB developing countries found that while men made
and Chest diseases OPD) for their suspected independent decisions to seek voluntary
koch‘s infection. 8 Patients consulted OPD-25 counselling and testing services, women felt
(Urology OPD) for suspected UTI and 2 patients obliged to discuss testing with their partners
visited OPD -4 (Orthopedic OPD) for their before accessing the service5. Another report by
orthopedic problems. center for disease control and prevention shows
that as far as possible, women would try to avoid
CONCLUSION going to the doctor, in particular, avoid getting
Majority of male patients (59 %) had contacted hospitalized as the household routine is likely to
private clinic/hospital initially for their health be disrupted more by the sickness of women of
problems as compared to 41 % females because the household. In households where both men
of fear of social stigma related to HIV/AIDS and and women are sick, more importance is given
belief of more privacy at private setup. Nearer to to get the man who is the breadwinner, the same
two third (70 %) female patients contacted may be the reason in our study as we found less
government hospital initially for their problems number of women who accessed treatment.7
as compared to 30 % males (P<0.05), as they
could not afford to go to private hospital due to
their economic dependency on others. 3 females
were forced to contact quack for their health
65 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
REFERENCE 4. United Nations Program on HIV/AIDS
1. Joint United Nations Programme on (2007). www.unaids.org.in . Accessed on
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Vol. 03 issue 07 July 2011
Pathway to Health Seeking Behavior

No Treatment

Home Remedies

Primary Health Center

NGO/GO

General Practitioner

Consultant

Government Hospital

VCTC

Patients visiting VCTC following different AIDS – Acquired Immunodeficiency Syndrome


pathways for health care were evaluated through ANC- Ante Natal Care
this flowchart. Whether they directly contacted ARV – Anti Retroviral Drug
VCTC without starting any treatment or ART – Anti Retroviral Therapy
contacted few health care providers before FGD – Focal Group Discussion
reaching VCTC (PHC MO / NGO / GO / Private GO – Government Organization
Practitioner / Consultant / Government GP – General Practitioner
Hospital). The entire interview was focused and GSACS – Gujarat State AIDS Control Society
in depth. HIV – Human Immunodeficiency Virus
Abbreviations: IDI – In-depth Interview
ADR – Adverse Drug Reaction MO – Medical Officer

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Vol. 03 issue 07 July 2011
MSM- Men having sex with Men UNAIDS – United Nations
NGO – Non Government Organization UNDP – United Nations Development Program
OPD – Out Patient Department UTI- Urinary Track Infection
PHC – Primary Health Centre VCTC – Voluntary Counseling and Testing
PLHIV – People living with HIV Centre
SSGH – Sir Sayajirao General Hospital VDRL – Venereal Disease Research Laboratory
STD – Sexually Transmitted Disease WHO – World Health Organization
STI – Sexually Transmitted Infection
TB – Tuberculosis

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Vol. 03 issue 07 July 2011
Pathway Of Health Seeking Behavior Of HIV Positive Cases Studied

QUAKE
03

35+3 PATIENT
54

21 PRIVATE GP -1

10 PRIVATE GP - 2

01 PRIVATE GP -3

PRIVATE GP -4 02 VCTC

16 GOVT. CHC/PHC

01
15 OPD – 1 (SSGH)

01
02 OPD – 4 (SSGH)

08 OPD – 17 (SSGH)

06
16 OPD – 18 (SSGH)

08 OPD – 25 (SSGH)

05 NGO

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Vol. 03 issue 07 July 2011
A COMPARATIVE STUDY ON EXTENDED TIMED GET UP
AND GO (ETGUG) TEST BETWEEN RIGHT AND LEFT
HEMIPLEGICS
Sankar Sahayaraj Muthukaruppan1, RavindraSubbanna2, Harilal
Bapurajapanicker1
ijcrr 1
Asst. Prof., Dept of Neuro Physiotherapy, KJPCP, Sumandeep Vidyapeeth,
Vol 03 issue 07
Piparia, Vadodara
Category: Research 2
Professor, 527, Ananya, 3rd Cross, Dollors Colony, Bangalore
Received on:05/03/11
Revised on:01/04/11 E-mail of corresponding author: subasankar@live.com
Accepted on:10/05/11

ABSTRACT
Objective: The purpose of the study is to enlighten the role of limb dominance in gait training and for
tailoring the gait training programme in physical therapy treatment between the categorical (temporal
parameters) and the representational (visuo-spatiial parameters) hemispheres.
Methods: between 45-55 years old left hemisphere dominant sixty males with hemiplegic (30 right side
& 30 left side), satisfying the inclusion criteria were chosen for the study. Extended Timed Get Up & Go
(ETGUG) test was used to assess the duration of sit to stand, gait initiation, walking, turn around, sitting
down, and speed of the walk. Independent‗t‘ test was used to compare the components of both the groups.
Results: The findings showed that there were significant durational changes in the various components of
ETGUG test between right side and left side hemiplegic subjects.
Conclusion: The gait rehabilitation should be emphasized on standing up, turning and sitting down for
left sided hemiplegics and gait initiation, walking & speed for right sided hemiplegics. Temporal and the
spatial parameters should be considered during gait training and gait training programme should be given
for right and left side hemiplegic subjects differently.
______________________________________________________________________________

Key words: Hemiplegic patient, ETGUG test, lower limb contributed mainly to body weight
limb dominance, gait training. transfer during walking; whereas the right lower
limb was responsible for propulsion.2 The
INTRODUCTION leading limb mainly contributes to forward
Hemispheric specialization is related to progression, where as the trailing limb provides
handedness. Handedness appears to be control and contributes to propulsion to a lesser
genetically determined. 96% of right-handers extent.3 The concept of cerebral dominance and
had left cerebral dominance, and the remainder a dominant and non dominant hemisphere has
had right cerebral dominance. Left dominance been replaced by a concept of complementary
was observed in 70% of left-handers, bi- specialization of the hemispheres, One for
laterality in 15% and right-dominance in 15%1. sequential analytic processes (the categorical)
There are anatomic differences between the two and the other one for visuo-spatiial relations (the
hemispheres that may correlate with the representational hemisphere).4
functional differences. During gait the left

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In India the incidence of cerebrovascular disease caused by increasing life expectancy resulted in
was found to be 13/1,00,000 population/year in a a burgeoning aging population in India. The
study conducted in 1969 - 71 at Vellore5, previous urban community-based studies had
31/1,00,000 population/year in a study documented an age adjusted incidence rate of
conducted at Rohtak6 and 124/1,00,000 stroke from 13 in 1970 to 105 per 100 000
population / year in a study conducted at persons per year in 2001.5,13 Stroke
Baruipur, West Bengal7. A WHO study in 1990 rehabilitation is a long drawn process and the
quoted incidence of mortality due to stroke in knowledge about the contributing factors might
India to be 73/1,00,000 per year. 8 help in tailoring the treatment and make it cost
Hand dominance affects an individual‘s effective by reducing treatment duration. The
functional ability after a stroke; yet current ability to walk is the prime factor that
constrained induced movement therapy studies determines whether a patient will go home or to
have not sufficiently addressed dominance and a nursing home and whether he or she will return
its role in functional recovery9. People with non to the previous level of productivity after a
dominant could possibly returns to a previous stroke. 14,15,16,17 This study aimed to provide an
level of functioning at a quicker phase compared opportunity for Neuro - physical therapist as
with an individual with a dominant side stroke. they would be able to scientifically understand
Gait asymmetry in temporal and kinematics and assess the role of dominant hemispheres
variables challenging the assumption of that during the gait training programme, the role of
right and left limbs are functioning parametric screening tool utilizing in physical
symmetrically10. Gait asymmetry in people therapy assessment, and the role of visuo-spatial,
without impairments can be explained in terms cognitive, psychosomotor and limb dominance
of actions taken by the lower limb to propel the in gait rehabilitation programme in stroke.
body segments and to control their forward
progression.11 Subjects with right hemisphere MATERIALS AND METHODS
lesions had greater troubles than those with left It was an observational study. 30 right side
hemisphere lesions in regaining the ability to affected and 30 left side affected hemiplegic
stay seated unassisted.12 Walking speed, step subjects (Middle Cerebral Artery infarcts &
length, and double support ratio, have been Brunstorm stage V), taking treatment in the
varied more in cognitive impairments than in physiotherapy department were taken for this
healthy older adults. study.
The objectives of the study were to investigate Subjects included in the study were 45-55 years
8
the duration of various components of ETGUG and having the ability to walk 20 meters
Test between right side hemiplegic subjects, and independently without walking aids. Subjects
left side hemiplegic subjects to modify the were excluded if they had deteriorating medical
treatment protocol for the right and left side conditions, structural and functional orthopedic
stroke subjects. problems of lower limb, and neurological
problems like cerebellar lesions, basal ganglion
Need of the Study lesions and previous history of head injury. Also
Changing life styles, and habitual intake of junk subjects under sedative therapy, visually
food and the added stress increase the incidence impaired subjects and non co-operative subjects
of stroke in developing countries like India. were excluded.
Over the past few decades, demographic shift
71 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
The Extended Timed Get Up & Go (ETGUG) training. It better isolates functional deficits
test is a potential, objective, assessment tool that thereby aiding the therapist in devising
can be used in almost any clinical setting with prevention strategies and guiding both treatment
minimal equipment, professional expertise or and further testing.

FIGURE I ETGUG TEST

Multi-memory stopwatch is the basics of Statistical Methods


ETGUG and measures the total time to complete
a task, as well as the time intervals for Data were statistically analyzed by using
components within that task. The laps and the Independent t test for comparing each
phases of test with which they correspond are as components of ETGUF between right and left
follows: Lap task: Sit to stand, Gait initiation, sided hemiplegics by SPSS version 16 for
Walk 1, Turn around, Walk 2, Slow down, stop, windows.
turn around and sit down. Since the distance for RESULTS
walk 1 and walk 2 are to know walking speeds, Stand up, Turning, Sit down and speed, were
can be calculated from the times taken for these increased in left hemiplegic subjects is showed
components of the ETGUG test. This is Figure 2. It showed they took maximum time to
important objective measure of functional ability complete the tasks than right hemiplegic
in the stroke subjects. In order to avoid errors; subjects. So these components are difficult for
subjects were oriented about the study prior to left hemiplegic subjects.
the conduct of the test. Standard protocol for
ETGUG test was followed. 18

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Vol. 03 issue 07 July 2011
Stand up is 0.000. Thus there is a significant difference
Group I right hemiplegics has the mean of between the two groups and result indicates that
1.5540 with SD of 0.12227 and Group II left right group is able to sit in the chair easily than
hemiplegics has the mean of 1.6230 with a SD left group.
of 0.11114. The mean difference is 0.690 and Speed
the t calculated value of 2.287 df= 58 at 5%. P Group I right hemiplegics has the mean of
value is 0.026. Thus there is a significant 0.5200 with SD of 0.2387 and left hemiplegics
difference between the two groups and result group has the mean of 0.5724 with a SD of
indicates that right group is able to stand up 0.3101. The mean difference is .0525 and the t
easily than left group calculated value of 7.343, df= 58 at 5%. P value
Turning is 0.000. Thus there is a significant difference
between the two groups and result indicates that
Group I right hemiplegics has the mean of right group is able to walk faster than left group.
7.6170 with SD of 0.43560 and left hemiplegics Gait initiation, Walk I and Walk II took longer
group has the mean of 8.7253 with a SD of duration to complete the task in right hemiplegic
0.95102. The mean difference is 1.1083 and the t subjects than left side hemiplegic subjects are
calculated value of 5.803, df= 58 at 5%. P value shown in Figure 3. So these components are
is 0.000. Thus there is a significant difference difficult for right hemiplegics than left
between the two groups and result indicates that hemiplegics
right group is able to turn easily than left group.

Sitting down
Group I right hemiplegics has the mean of
3.3410 with SD of 0.15103 and left hemiplegics
group has the mean of 3.6940 with a SD of
0.13361. The mean difference is 0.3530 and the t
calculated value of 9.588, df= 58 at 5%. P value

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Vol. 03 issue 07 July 2011
Gait initiation Figure 3 showed the time duration to complete
Group I right hemiplegics has the mean of the task is longer in right hemiplegic subjects
3.2377with SD of 0.66638 and left hemiplegics than left hemiplegic subjects. Also the left
group has the mean of 2.6910 with a SD of hemiplegic subjects were able to finish the task
0.18894. The mean difference is 0.5467 and the t faster than in right hemiplegic subjects.
calculated value of 4.323, df= 58 at 5%. P value Total time
is 0.000. Thus there is a significant difference Group I right hemiplegics has the mean of
between the two groups and result indicates that 39.0800 with SD of 2.08339 and left
right group is easily able to perform gait hemiplegics group has the mean of 37.9000 with
initiation than left group a SD of 1.52131. The mean difference 1.1800
Walk I and the t calculated value of 2.505, df= 58 at
Group I right hemiplegics has the mean of 5%. P value is 0.015. Thus there is a significant
11.547 with SD of 0.5708 and left hemiplegics difference between the two groups and result
group has the mean of 10.503 with a SD of indicates that right group requires maximum
0.5425. The mean difference is 1.045and the t time to finish the task than left group.
calculated value of 7.266, df= 58 at 5%. P value
is 0.000. . Thus there is a significant difference DISCUSSIONS
between the two groups and result indicates that The absence of well recognized measures to
right group is walk faster than the left group. evaluate functional recovery after a stroke and
Walk II the large variety of rehabilitation techniques and
Group I right hemiplegics has the mean of protocols add credence to the questioning of the
11.5540 with SD of 0.48604 and left value of rehabilitation in terms of costs and
hemiplegics group has the mean of 10.4987 with psychosocial benefits.19, 20, 21 The maximum
a SD of 0.58557. The mean difference is 1.0553 weight-bearing difference between the 2 lower
and the t calculated value of 7.596 df= 58 at 5%. extremities during the Sit To Stand task was
P value is 0.000. Thus there is a significant highest for the subjects with stroke who had the
difference between the two groups and result lowest scores on the Functional Independence
indicates that right group is more faster walk Measure.22
than the left group.

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Vol. 03 issue 07 July 2011
The present study analyzed various components side hemiplegics it was 0.5200 ± 0.2387 m/s and
of functional mobility in right and left side left side hemiplegics group it was 0.5724 ±
hemiplegic subjects. In our study, Standing up 0.3101. The right side affected group was able
(Sit to stand), turning and sitting down require to walk faster than left side affected group. The
maximum balance, which is significantly ability to complete the task is faster in left sided
reduced in trailing limb (left limb) affected hemiplegic subjects. But both the groups took
subjects. Leading limb (right limb) affected more than 30 seconds to complete the task. So
subjects took longer duration for gait initiation they were under the risk category, since the
and finishing the task (walking) than the trailing falling criteria is more than 30 sec for ETGUG
limb affected subjects. This implies right side test.
affected stroke subjects (right handed) had more CONCLUSIONS
difficulties with gait initiation and walking and ETGUG test components were not in similar. In
left side affected subjects had more difficulties stroke patient‘s conventional rehabilitation
in sit to stand, turning and sitting downGait program will not provide additional benefit in
performance in subjects with stroke is terms of lower extremity motor recovery and
characterized by slower gait velocity and mobility. So the training of functional mobility
residual spatial and temporal left-right and gait training should be concentrated
asymmetry, compared with that in healthy differently for right side affected hemiplegics
adults. The average walking speed of stroke and left side affected hemiplegics in
subjects is lower than that of healthy controls physiotherapy. Training should not be
but the values vary depending on the severity of concentrated only on temporal parameters. The
the stroke, the time post-stroke and the age of temporal and spatial parameters as well as
the subjects23. In this study these variables were hemisphere dominance should be considered for
controlled. Previous studies suggested that the gait training protocols.
gait velocity of patients with stroke of varying
severity ranges from approximately 0.18 to ACKNOWLEDGEMENT
1.03m/s, 24,25,26,27 whereas that of healthy adults The author expressing his sincere thanks to
of similar ages averages about 1.4m/s28 in our Professor James C. Wall, PhD Department of
present study also suggested the same. The Physical Therapy, University of South Alabama
speed was less than the normal and for the Right for his advices in ETGUT Test.

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Vol. 03 issue 07 July 2011
REFERENCE symmetry. Phys Ther 1989; 69:640-650
1. Annett, M. (2002). Handedness and brain (medlins).
asymmetry the right shift theory. East 12. Wade DT, Hewer C, Wood VA, Stroke;
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6:32-44. 13. Banerjee TK, Mukherjee CS, Sarkhel A.
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Hall international, Inc; 1997. p.257. gait in patients after stroke. Scandinavian J.
5. Abraham J, Rao PSS, Inbaraj SG, Shetty G, Rehab. Med. 1981; 13:109-115
Jose CJ, An epidemiological study of 15. Shiavi, R.; Bugle, H. J.; and Limbird, T.:
hemiplegia due to stroke in South India. Electromyographic gait assessment, part 1:
Stroke 1970; 1: 477-81. adult EMG profiles and walking speed. J.
6. Bansal BC, Parkash C, Jain AC, Rehab. Res. and Devel 1987; 24:13-23
Brahmanandan KRV. Cerebrovascular 16. Shiavi, R.; Bugle, H. J.; and Limbird, T.:
disease in young individuals below the age Electromyographic gait assessment, part2:
of 40 years. Neurol India 1973; 21: 11-8. preliminary assessment of hemiparetic
7. Bhattacharya S, Saha SP, Basu A, Das SK. synergy patterns. J. Rehab. Res. and Devel.
A 5-year prospective study of incidence, 1987; 24:24-30
morbidity and mortality profile of stroke in a 17. Wagenaar, R. C.; Meijer, O. G.; van
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Med Assoc 2005; 103: 655-9 Hazenberg, G. J.; Lindeboom, J.; Wichers,
8. Prasad K: Epidemology of cerebrovasular F.; and Rijswijk, H.: The functional
disorders in India. In: Recent concepts in recovery of stroke: a comparison between
stroke by Bansal BC (ed) Indian college of neurodevelopmental treatment and the
physicians, New Delhi, 199; 4-19. Brunnstrom method. Scandinavian J. Rehab.
9. Hellige J. Hemispheric Asymmetry: What‘s Med. 1990;22:1-8
Right and What‘s Left? Cambridge, Mass: 18. Wall C. James, Bell Churan, Campbell
Harvard University Press; 1993. Stewart; Davis Jennifer the Timed get-up
10. SadeghiH, Allard P, Duhaime M. Functional and-go Test Revisited: measurement of the
gait asymmetry in able bodied subjects. component tasks. J Rehabil Res
Human Movement science. 1997; 16:243- January/February 2000; 37(1): 109-114.
258. 19. Brunnstrom, S.: Movement Therapy in
11. Gunderson LA, Valle DR, Barr AE, et al, Hemiplegia: a Neurophysiological
Bilateral analysis of the knee and ankle Approach. Hagerstown, Maryland, Harper
during gait: an examination of the and Row, 1970
relationship between lateral dominance and 20. Kaplan, P. E.: Hemiplegic: rehabilitation of
the lower extremity. In Stroke
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Rehabilitation, pp. 119-146. Edited by P. E. 25. Knutsson E, Richards C. Different types of
Kaplan and L. J. Cerullo. Boston, disturbed motor control in gait of hemiplegic
Butterworth, 1986 patients. Brain 1979; 102: 405-30.
21. Stevens, R. S.; Ambler, N. R.; and Warren, 26. Olney SJ, Griffin MP, McBride ID.
M. D.: A randomized controlled trial of a Temporal, kinematic, and kinetic variables
stroke rehabilitation ward. Age and Ageing related to gait speed in subjects with
1984; 13:65-75 hemiplegia: a regression approach. Phys
22. Lee MY, Wong MK, Tang FT, et al. Ther 1994; 74: 872-85.
Comparison of balance responses and motor 27. Wade DT, Heller WA, Maggs J, Hewer RL.
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functional mobility in stroke patients. Am J 1987;19:25-30
Phys Med Rehabil 1997; 76: 401– 410. 28. Murray MP, Kory RC, Clarkson SB.
23. Olney SJ, Richards C. Hemiparetic gait Walking patterns in healthy old men. J
following stroke. Part I: Characteristics. Gait Gerontol 1969; 24: 164-78
Posture 1996; 4: 136-148.
24. Brandstater ME, de Bruin H, Gowland C,
Clark BM. Hemiplegic gait: analysis of
temporal variables. Arch Phys Med Rehabil
1983; 64: 583-7.

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Vol. 03 issue 07 July 2011
SIX MINUTE WALK DISTANCE IN HEALTHY ADULTS
AGED BETWEEN 20-30 YEARS
Prem.V1, R.D. Chakravarty2, Karvannan H.1, Rimmi3, Vinita Arya3, Manikankanna3
1
Department of physiotherapy, Manipal Hospital, Manipal College of Allied Health
Sciences, Manipal University, Bangalore
2
ijcrr 3
Head of the department, Orthopaedics, Manipal hospital, Bangalore
Vol 03 issue 07 Department of physiotherapy, kasturba medical college, Manipal University,
Mangalore
Category: Research
Received on:19/05/11 E-mail of corresponding author: prem.v@manipal.edu
Revised on:25/05/11
Accepted on:29/05/11

ABSTRACT
OBJECTIVE: To find the normal range of distance covered during six minute walk distance (6MWD) in
healthy adults aged between 20-30 years.
METHODS & RESULTS: Six minute walk distance was performed in a 100 feet hallway by 50 males
and 50 females healthy adults ranging in age from 20-30 years. Each subject underwent a thorough
physical examination including weight, standing height and BMI. Any subject with underlying
cardiopulmonary, neurological or musculoskeletal pathology or conditions that could interfere with the
walk test was excluded. All subjects underwent 6MWT and pre and post measurements such as distance,
heart rate were recorded. Males walked 45 m more than females. Height of females is significantly
correlated with 6MWD. There was significant increase in heart rate and respiratory rate following six
minute walk test.
CONCLUSION:
Six minute walk distance covered in healthy individuals of 20-30 years is 560±61 in males and 514±32 in
females.

______________________________________________________________________________

KEY WORDS: Six minute walk distance, Heart submaximal levels of exertion and therefore it
rate, Healthy adults . has been proposed that submaximal functional
tests are better reflection of physical capabilities.
INTRODUCTION The ability to work a set distance is a quick,
An individual‘s response to exercise is an safe, easy and inexpensive way to assess
important clinical assessment tool, since it physical function. It is an important component
provides a composite assessment of their of quality of life, since it reflects the capacity to
respiratory, cardiac and metabolic system. undertake day to day activities. According to the
The current goal standard for assessing a American Thoracic Society (ATS), the most
person‘s aerobic response is the maximum precise indication for the performance of the six
incremental cardiopulmonary exercise test. minute walk test (6MWT) is mild or moderate
However, most daily activities are performed at lung or heart disease, in which it is used in order
78 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
to measure treatment response, as well as to more acceptable and provides a better reflection
predict morbidity and mortality. of activities of daily living than other walk test.5
Balke developed a simple test for examining
functional capacity, measuring the distance The normal six minute walk distance has been
walked during a definite period of time. reported in western countries 1, 5-7, but there is no
A twelve minute performance test was then normative data for Indian population. Hence the
developed to evaluate the physical fitness of present study aims to generate a normative data
healthy individuals. This test was subsequently for Indian population.
modified for use in patients with chronic
bronchitis. In order to allow the test to be used in METHODOLOGY
patients with respiratory diseases, for whom Subjects
twelve minute walking was too demanding, a The study was carried out at Kasturba Medical
shortened version six minute walk test was College, Mangalore. There were 100 healthy
developed and found to perform equally as volunteered subjects comprising 50 males and
well.1-3 50 females aged 20-30 years. Each subject
Twelve minute walk test (12MWT) is a practical underwent a thorough physical examination
guide to everyday disability nevertheless it is including weight, standing height and BMI(
both time consuming for investigator and Appendix I). Any subject with underlying
exhausting for patients, therefore the possibility cardiopulmonary, neurological or
of using walking test of shorter duration to musculoskeletal pathology or conditions that
assess exercise tolerance was explored.4 could interfere with the walk test was excluded.
Compared to traditional laboratory index of All subjects underwent 6MWT and pre and post
exercise capacity such as cycle, treadmill and measurements such as distance, heart rate were
step ergo meter; walk test require less technical recorded. The healthy adults included were age
expertise and equipment making them group between 20-30 years and adults excluded
inexpensive and easy to administer. More were, Underlying cardiopulmonary,
importantly, they employ an activity that neurological, musculoskeletal pathology,
individuals perform on a daily basis that is Smoker for more than one year, Alcoholic for
walking. more than one year (occasional alcoholic can be
6MWT has been frequently used to measure included), Cognitively impaired subject,
outcomes before and after treatment, in patients Uncooperative subjects.
with moderate to severe heart and lung diseases
and in their prognosis. It has also been used to PROCEDURE
measure functional status and for Baseline measurements such as pulse rate,
epidemiological research purposes. The distance respiratory rate and dyspnoea level using
covered in 6MWD has been showed to Modified Borg 0-10 scale were taken. The
accurately predict morbidity and mortality from materials required were 100 Ft. hallway,
cardiopulmonary diseases. In healthy elderly Sphygmomanometer, Stethoscope, Timer,
subjects, 6MWT represents submaximal Watch, Pen and relevant paper work, Cones,
exercise, but at almost 80% of the VO2 max. A Chair, Inch tape. We instructed the subject that
recent review of functional walking test objective of this test is to walk as far as possible
concluded that 6MWT is easier to carry out, for six minutes and were told to ―walk back and
forth in this hallway as quickly as you can so
79 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
that you cover as much ground as possible‖. average the 6MWD was 560±61m in males and
They were informed that they could slow down 514±32m in females. An important part of the
or rest if necessary. We demonstrated way of variability is 6MWD was explained by height,
walking and turning around the cones placed at sex, age and weight as independent variables.
the two ends of the hallway. They were also Distance walked in males and females is directly
instructed not to run and jog. proportional to height and age and it is
We positioned the subject at the starting line. As significant in females.
soon as subject started walking, timer was Study done by T.Troosters et al. showed
started. At the end of each minute subject were considerable variability in 6MWD of healthy
given feedback on the elapsed time and subjects aged 50-85years,ranging 383-820m on
standardized encouragement in the form of an average 6MWD was 631±93m and it was
statements such as ―you are doing well, keep up 84m greater in males as compared to female
the good work‖ and ―do your best‖. Test was subjects. An important part of the variability in
terminated at the end of six minutes.1 6MWD was explained by height, sex, age and
weight as independent variable.8
Post test measurement The greatest 6MWD from among several
Heart rate was recorded using three finger repetition in a wide age range of healthy
palpation method at rest and at the end of test. volunteers showed that the distance walked after
Respiratory rate was recorded by observation the first walk was the best and in 86%
method prior to and upon completion of 6MWT. individuals an average increase of 43m was
At the end of the test distance was measured and observed from first to best 6MWD.Best 6MWD
dyspnoea was rated using modified Borg 0-10 average 698±96m and was inversely related to
scale. age, directly to height and greater in male then
female.9
DATA ANALYSIS The present study shows the correlation between
Test used for the study is student unpaired t-test. height of the subject and the distance walked
Result has been analyzed by SPSS.vers.14.0 which is directly proportional to each other.
Because taller individuals have greater stride
RESULTS length and so the distance covered is more.
The baseline characteristics of males and When compared between males and females,
females are demonstrated in table 1.Mean males cover longer distance because of greater
distance covered in males is 45m greater than functional capacity and more muscle mass.
females table 2.There was a significant increase Moreover, distance walked is also directly
between pre test and post test heart rate table 3. proportional to age of the subject because the
There was a significant increase between pre test present study covers young population in a
and post test respiratory rate table 4. There was narrow range. In this age group as the as the age
no difference in pre test and post test Borg Scale increases muscle mass increases.
table 5. Study done in Chinese population shows
marked similarity with the independent variables
DISCUSSION – height and sex. Height is another important
The present study showed considerable determinant of 6MWD. This is not surprising as
variability in the 6MWD of the healthy subjects taller people have, in theory, a larger stride
aged 20-30 years; ranging 455 to 600m.On length and, thus, greater 6MWD.Young males
80 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
are also found to have greater exercise capacity needed for varying age groups for eg.30-70
and 6MWD than young females, probably as a years.
result of their greater muscle mass.10
In the present study, subjects were able to reach CONCLUSION
50% HRmax which was 101 for males and 100 Six minute walk distance covered in healthy
for females. The less distance walked during individuals of 20-30 years is 560±61 in males
6MWD could be due to different persons and 514±32 in females.
administering the test and variability in
understanding the instructions by the subjects. REFERENCES
Though there is not much difference in mean
height of the subjects as compared to study done 1. ATS Statement Guidelines for Six Minute
by Bernadine Camarri et al., the lesser distance Walk Test American Journal of Respiratory
walked by subjects may be because of lower and Critical Care Medicine vol 166, pp 111-
HRmax as compared to 80% of HRmax in their 117
study.11 2. Sherra Solway, Lina Brooks, Yves Lacasse,
The functional status and capacity can be Scott Thomas A Qualitative Systematic
effectively measured by functional walk test, the Overview Of The Measurement Properties
best being 6MWT.The measurement properties Of Functional Walk Test used in the
of the 6MWT have been the most extensively Cardiorespiratory Domain chest
researched and established. In addition 6MWT is 2001;119:256-270
easy to administer, better tolerated and more 3. Kervio, Gaelle, Carre, Francois,Ville,
reflective of activities of daily living than the Nathlie Reliability and Intensity of the
other walk tests. Therefore the 6MWT is 6MWT in healthy Elderly subjects Med. Sci.
currently the test of choice when using a Sports Exerc, vol 35, no 1,pp. 169-174,2003
functional walk test for clinical or research 4. CR Mcgavin, S P Gupta 12 Minute walking
purposes. test for assessing disability in chronic
Reference Equations bronchitis. British Medical Journal
Males 1976,1,822-823
6MWD = 89.455 + (4.303 age) + (2.491 5. Two, Six, Twelve min walking test in
height) - (0.877 weight) respiratory diseases British Medical
Journal,vol284,29 may 1982
Females 6. Fryderyk Prochaczek, Hanna Winiarska1 et
6MWD = 120.129 + (5.320 × age) + (2.080 × al. Six-minute walk test on a special
height) – (1.002 × weight) treadmill: Primary results healthy
Limitations of the Study volunteers. Cardiology Journal 2007, Vol.
There are certain limitations to present study. 14, No. 5, pp. 447–452.
only 100 subjects were recruited, the sample size 7. Paul L. Enright and Duane L. Sherrill
was inadequate for establishment of more Reference Equations for The Six Minute
accurate value of distance walked in Indian walk in Healthy Adults. AMJ Respir J
population. 1999;14:270-274
Future Studies 8. T.Troosters, R.Gooselink, M.Decramer Six
Future studies need to be done with larger Minute Walking Test in healthy elderly
sample size with more trials. Studies are also subjects. Eur Respir J 1999;14:270-274
81 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
9. Gibbons, William J, Fruchter, Nadine, 11. Bernadine Camarri, Peter R. Eastwood, Nola
Sloan, Sherry et al Reference Values for a M.Cecins, Philip J. Thompson, Sue Jenkins
multiple repetition Six Minute Walk Test in Six minute walk distance in healthy subjects
healthy adults older than 20 years. Journal of aged 55-75 years. Respiratory Medicine
Cardiorespiratory Rehabilitation 21(2):87-93 (2006)100,658-665
March/April 2001.
10. A.M. Li, J.Yin, C.C.W.Yu, T.Tsang,
H.K.So, E.Wong et al The 6 MWT in
healthy children; reliability and validity Eur
Respir J 2005;25:1057-1060

Table 1 – Baseline characteristics of males and females

CHARACTERISTIC MALES ( n=50) FEMALES ( n=50)


AGE 23 ±2 22 ±1
HEIGHT 172±8 159±6
WEIGHT 66±9 54±7
BMI 22.39±2.7 21.29±2.3

Table 2 – Mean distance covered by males and females

MALES FEMALES
DISTANCE (meters) 560±61 514±32

Table 3 – Change in heart rate pre and post test

HRbpm MALES FEMALES


PRE HRbpm 82±12 78±11
POST HRbpm 101 100

Table 4 – Change in respiratory rate pre test and post test

RRbpm MALES FEMALES


PRE RRbpm 20±3 20±3
POST RRbpm 27±5 28±4

Table 5 – Change in dyspnoea scale pre test and post test

BORG SCALE MALES FEMALES


PRE BORG SCALE 0 0
POST BORG SCALE 0.2±0.3 0.4±0.4

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Appendix 1. ASSESSMENT PERFORMA

NAME: AGE: SEX: M ( ) F( )

HEIGHT:
BMI: UNDERWEIGHT ( ) <18.5-
NORMAL ( ) UNDERWEIGHT
WEIGHT: OVERWEIGHT ( ) 18.5-25-NORMAL
OBESE ( ) 25-30-
OVERWEIGHT
>30- OBESE
FINDINGS PRE TEST POST TEST

PULSE RATE

RESPIRATORY RATE
BORG’S SCALE

NO. OF LAPS:

TOTAL DISTANCE COVERED:

ANY SYMPTOMS:

BORG’S SCALE:

0- nothing at all
0.5-very very slight (just noticeable)
1- very slight
2- slight
3- moderate
4- somewhat severe
5- severe
6-
7- very severe
8-
9-10- very very severe(maximal)

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Vol. 03 issue 07 July 2011
SEA WATER ABSORPTION PHENOMENON IN BANANA
FIBRE REINFORCED VINYL ESTER COMPOSITES
Rajesh Ghosh1, A Ramakrishna2, G.Reena3
1
ijcrr Mechanical Engineering Department, GIT – GITAM University, Visakhapatnam
2
Vol 03 issue 07 Mechanical Engineering Department, Andhra University, Visakhapatnam
3
Category: Research Onsite ETL Coordinator, Mahindra Satyam, Singapore
Received on:19/05/11 E-mail of corresponding author: rajesh_ghosh@yahoo.com
Revised on:23/05/11
Accepted on:28/05/11

ABSTRACT
The ageing response of banana fibre reinforced vinyl ester composites in sea water environment
is investigated. The main objective was to evaluate the effects of sea water on the mechanical
properties. Fibre mats were reinforced into vinyl ester matrix and composite laminates were
made. These were then subjected to sea water ageing and water absorption and mechanical
properties were tested. It is observed that during the initial stages there was increased rate of
water absorption following the Fickian law. On prolonged time of immersion, moisture
absorption led to plasticization of the matrix and also reduced the mechanical properties of the
specimen.
______________________________________________________________________________

Keyword: Sea water, absorption, banana fibre, by a hydrolysis reaction of the unsaturated
vinyl ester. groups within the resin [3]. Vinyl ester
composites show superior chemical stability in
INTRODUCTION sea water atmosphere [4, 5]. The absorption of
Fibre reinforced polymers are increasingly water into the macromolecular network of a
shown interest from the engineering and thermoset matrix causes swelling and
structural view point. A lot of work is done and plasticization of the matrix [6- 8]. Researchers
investigated on glass fibre reinforced have reported that absorption of water (distilled
composites. But glass fibre has a detrimental or sea water) causes changes in the
effect on the environment and hence there is a thermophysical and mechanical properties by
growing interest on bio fibres on the possibility plasticization and hydrolysis [9, 10]. It is
of replacing glass fibres. These composites are reported that degree of degradation depends on
being used for marine applications such as water the degree of crosslinking of the polymers [11].
storage vessels, pipelines, small boats etc. Moisture may also affect the fibers.
Researchers found that prolonged exposure of Experimental work is done on glass fibre
glass fibre composites caused degradation in reinforced in epoxy or vinyl ester resin. But
flexural strength and modulus [1, 2]. It is much work is not done on banana fiber
observed that the polymer matrix gets degraded reinforced in polymer matrix and the composite

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Vol. 03 issue 07 July 2011
exposed to sea water environment. In the present between the fibre and the resin and also through
work banana fibre reinforced vinyl ester the fiber, and transport of water through the
composite specimens were immersed in sea microcracks in the matrix.
water and the moisture gain is noted and The diffusion, in most of the cases, follows the
compared with distilled water moisture gain. equation

EXPERIMENTAL Mt
Banana fibers were procured from Tamil Nadu - kt n - (1)
M m ax
India. The fibers were knitted in the form of
where Mt is the moisture content at time t, Mmax
mats. The mats were alkali treated. Vinyl ester is
is the maximum moisture content at saturation
procured from ECMAS India pvt ltd.
and k and n are constants.
Composites were made by hand layup process in
The diffusion coefficient is an important
an MS die with inner cavity of dimensions
parameter in Fick‘law. This can be found out
200mm x 200mm x 10mm. These laminates
from the following equation
were later oven cured. Composite specimens
were later cut to size as per ASTM standards.
The cut edges of the specimens were coated with 4 M m ax Dt
Mt - (2)
a thin layer of adhesive. h
Sea water absorption tests: These specimens
were first weighed and then immersed in sea where Mt , Mmax and t are as denoted above, and
water taken from the coast of Visakhapatnam – h is the specimen thickness.
India (Bay of Bengal). Specimens were The diffusion coefficient can be found out by
periodically taken out of the water; the surface is considering the slope of the first portion of the
wiped with a tissue paper and weighed in an curve between moisture gain and square root of
electronic balance. The water uptake was plotted time by the following equation.
against square root of immersion time. 2
kh
The moisture absorbed M (in %) is calculated D -(3)
using 4M m ax
where k is the initial slope of the plot.
Mt Mo Fig.1 represents the percentage moisture gain
M% 100 plotted against square root of time.
Mo
The analysis of diffusion mechanism and
where Wt is the measured weight of the
kinetics can be performed by modifying eqn.(1)
specimen at time t and Wo is the initial dry
as shown below.
weight of the specimen.
Mt
log log k n log t - (3)
RESULTS AND DISCUSSION M max
The water absorption into the composite
specimen may be considered to be following
three different modes. The principal mode being Fig.2 shows the graph plotted against log(Mt/
the diffusion of water molecules into the Mmax) against log(t).
microgaps of the resin, while the other processes
being capillary action through the interfacial gap

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Vol. 03 issue 07 July 2011
12.19 %. There is a decrease in the tensile
The straight line in Fig.2 shows the fitting of the modulus and flexural modulus of the specimen
experimental data to eqn. (3). by 12.32% and 10.09%.
The values of k and n resulting from the graph in
fig.3 is found to be 0.001254 and 0.59 REFERENCE
respectively. 1. Kootsokoos A, Mouritz AP, ―Sea water
The value of ‗n‘ suggests that the initial durability of glass and carbon polymer
diffusion follows Fick‘s law. composites. Compos. Sci. Technol. 64,
The diffusion coefficient was found to be 2.67 x 1503-1511 (2004)
10-6 mm2/sec. 2. Gellert EP, Turley DM, ―Seawater
immersion ageing of glass fibre
Effect of sea water on mechanical properties: reinforced polymer laminates for marine
Tensile properties: Tensile test is done applications.‖ Composites 30A, 1259
according to ASTM D638. The tests were (1999)
carried out in a Hounsfield tensometer model 3. Srinivas MV, Dvorak GJ and Prochazka
H20KW. The cross head speed is 1mm/min. P, ―Design and fabrication of submerged
Flexural properties: The flexural test is done cylindrical laminates – II, Effects of
according to ASTM D790 in a universal testing fibre pre-stress. International journal of
machine by UNITED calibration corp. with a solids and structures. 36, 3945 – 3976
cross head speed of 0.5mm/min. (1999).
Fig.4 shows the values of tensile strength, tensile 4. Apicella A, Migliaresi C, Nicholais L
modulus, flexural strength and flexural modulus and Roccotelli S, ―The water ageing of
of the composite specimens before and after unsaturated polyester based composites:
immersion in sea water. Influence of resin chemical structure,
Sea water diffuses into the composite specimen Composites. 14, 4, 387 – 392 (1983).
following the Fick‘s law of diffusion initially. 5. Dvorak GJ, Prochazka P, and Srinivas
With elapsing time, the salts get deposited into MV, ―Design and fabrication of
the microcracks preventing further seepage. submerged cylindrical laminates – I,
Hence the rate of water absorption slows down International journal of solids and
with time. With the ingress of water, the structures. 36, 3917 – 3943 (1999).
mechanical properties decrease as understood 6. Bradley WL, (1995). J. Mater. Sci
from the graphs. 30:5537. doi:10.1007/BF00351570.
7. Pomies F, Carlsson LA, Gillespie JW Jr
CONCLUSION (1995) ASTM STP 1230. philadelphia.
The water uptake increases rapidly during the 8. Springer GS (ed) (1981) Environmental
initial stages. The absorption process follows the effects on composite materials,
Fickian diffusion process during the initial Technomic, CA.
stages but later it follows non-fickian diffusion 9. Ellyin F, Rohrbacher C, ―Effect of
process due to the deposition of the salts present aqueous environment and temperature
in the water. Absorption of sea water causes the on glass fibre epoxy resin composites‖.
deterioration of mechanical properties. The J. Reinf. Plast. Compos. 19(17), (2000)
tensile strength is found to decrease by 6.46 % 10. Wood C, Bradley WL, ―Determination
and the flexural strength is found to decrease by of the effect of sea water on the
86 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
interfacial strength of an interlayer e- 11. Rutowska M, Krasowska K, Heimwoska
glass/graphite/epoxy composite by in A, Steinka E, Janik H, ―Degradation of
situ observation of transverse cracking polyurethanes in sea water‖. Polym.
in an environmental SEM. Compos. Sci. Degrad. Stab. 76, 233 – 239 (2002)
Technol. 57, 1033 – 1043, (1997)

12

10
% water gain

0
0 5 10 15 20 25 30

√t̅

0
0 1 2 3 4 5 6 7
-0.1

-0.2
log(Mt /Mmax )

-0.3

-0.4

-0.5

-0.6

-0.7
log(t)

Fig.2. Diffusion case fitting plot

87 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
120 6 160 3.5

3
5

flexural strength (MPa)

flexural modulus (GPa)


tensile strength (MPa)

tensile modulus (GPa)


120
2.5
110 4
2
3 80
1.5
100 2
1
40
1 0.5

90 0 0 0

Fig.3. comparison of mechanical properties of dry and saturated composite specimens.

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Vol. 03 issue 07 July 2011
PERIOD-DOUBLING PHENOMENA IN A SIMPLE-3D
CHAOTIC OSCILLATOR WITH A DIODE PAIR
G. Kandiban1, V. Balachandran2, S. Manimaran3

ijcrr 1
2
Department of Physics, Thanthai Hans Roever College, Perambalur
Vol 03 issue 07 Department of Physics, A.A Government Arts College, Musiri, Tiruchirappalli
3
Department of Physics, Roever Engineering College, Perambalur
Category: Research
Received on:25/04/11 E-mail of corresponding author: brsbala@rediffmail.com
Revised on:15/05/11
Accepted on:27/05/11

ABSTRACT
In this paper, in order to show some interesting phenomena of third-order chaotic oscillator circuit with a
smooth cubic nonlinearity, different kinds of attractors, time waveforms and corresponding power spectra
of systems are presented, respectively. The perturbation transforms an unpredictable chaotic behavior into
a predictable chaotic or periodic motion via stabilization of unstable, aperiodic, or periodic orbits of the
strange chaotic attractor. One advantage of the method is its robustness against noise. A theoretical
analysis of the circuit equations is presented, along with experimental and numerical results.
______________________________________________________________________________

Keywords: Autonomous third-order chaotic In this work we introduce an autonomous third-


circuit; Smooth cubic nonlinearity; chaos. order chaotic oscillator circuit that realizes
period-doubling route to chaos followed by
INTRODUCTION periodic window and then to lower dimensions
In the present report the behavior of a third- of strong chaos through boundary crisis etc. We
order autonomous chaotic oscillator circuit has consider that such complicated chaotic time
been studied. This circuit consists of two active waveforms are expected to be utilized for
elements, one linear negative conductance and realization of several chaotic applications such
diode pair exhibiting symmetrical piecewise- as chaos communication system with robustness
linear v-i characteristics, two linear capacitances against various interferences including multi
(C1 and C2) and one linear inductor (L) is also user access.
included in the circuit, serves as the control
parameters [1-5]. EXPERIMENTAL REALIZATION OF THE
Most chaotic and bifurcation effect cited in the AUTONOMOUS THIRD-ORDER
literature have been observed in electrical CHAOTIC OSCILLATOR CIRCUIT
circuits. They include the period-doubling route The experimental realization of the autonomous
to chaos [6], the intermittency route to chaos [7], third-order chaotic oscillator is shown in Figure
and the quasi-periodicity route to chaos and of 1. It consists of two active elements, one linear
course the crisis [8-10]. This popularity is negative conductance (G1) using op-amp and
attributed to the advantages which electric smooth cubic nonlinearity with an odd
circuits offer to experimental chaos studies, such symmetric piecewise-linear v-i characteristic
as robustness and convenient implementation. [11-12]. The characteristics of the negative
89 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
conductance are mathematically represented doubling window, lower dimensions of strong
by i G1 V2 . chaos have not been reported.
The symmetrical cubic nonlinear is designed by
the help of two signal diodes. The v-i EXPERIMENTAL OBSERVATIONS:
characteristics of the global non-linearity which PERIOD-DOUBLING ROUTE TO CHAOS
can therefore be approximated by a cubic VIA PERIOD-DOUBLING WINDOW
function of the form with a < 0 and b > 0. For our present experimental study we have
chosen the following typical values of the circuit
f (V ) v aV bV 3
in Fig.1: C1 = 10nF, C2 = 100nF. The negative
The constant term v describes the input current conductance G1 = -0.5mS and cubic nonlinear
offset of the op-amp which can be practically resistance a < 0 and b > 0. Here the variable
adjusted to zero using the potentiometer ( v = 0). inductor (L) is assumed to be the control
By applying Kirchhoff‘s laws to the equivalent parameter.
circuit of (Figure 1) we obtain the following set By increasing the value of L from 100mH to
of differential equations: 220mH, the circuit behavior of Fig. 1 is found to
dV1 transmit from a period doubling route to chaos,
C1 iL f (V1 )
dt and then to period doubling window through
dV lower dimensions of strong chaos followed by
C2 2 G1V2 iL (1)
dt boundary crisis etc.,. The projection of the
di attractors on the (V1 V2 ) plane and current
L L V2 V1 iL R
dt sensing resistor with voltage plane of Cathode
Ray Oscilloscope is shown in Fig. 2. For various
While V1 and V2 are the voltages across the values of control parameter L. Fig. 3 shows the
capacitors C1 and C2, and iL denote the current experimental chaotic time series were registered
through the inductance (L) respectively, the term using a Cathode Ray Oscilloscope for discrete
f(V1) representing the characteristics of the values of L serving as the control parameter.
symmetrical cubic nonlinearity can be expressed The third-order autonomous chaotic oscillator
mathematically as circuit with the symmetrical cubic nonlinearity
3 can produce lower dimensions of strong chaos
f (V1 ) aV1 bV1 (1.1)
see in Fig. 4, from which we observe clearly that
there are chaotic power spectrums. The power
The autonomous simple-3D chaotic oscillator
spectrum corresponding to the voltages V1 (t)
circuit is also truly chaotic oscillator. This is
and V2(t) waveform across the capacitors C1 and
because, there is no locally active resistance (R)
C2 respectively, which resembles broad-band
in this circuit, only varying that the inductance
spectrum noise.
(L) value, this circuit exhibits a very interesting
dynamical phenomena like period-doubling
NUMERICAL REALIZATION OF THE
bifurcation sequence leading to chaos, period-
AUTONOMOUS THIRD-ORDER
doubling window and then to lower dimensions
CHAOTIC OSCILLATOR CIRCUIT
of strong chaos through boundary condition [13-
For a convenient numerical analysis of the
14]. However in the parameter regimes
experimental systems given by Eqns.(1), we
investigated, important features like period-

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Vol. 03 issue 07 July 2011
x1 discrete value of L serving as the control
rescale the parameters as V1 ,
parameter.
bR
x2 x3 C2
V2 , iL , t R C2 , , RESULTS AND DISCUSSION
bR bR 3 C1 We have introduced period-doubling phenomena
C2 R 2 in a simple-3D chaotic oscillator with a diode
, a R, G1 R and then pair. Figure 2 shows an experimental attractor
L
redefine τ as t. Then the normalized equations of (projected onto the four planes formed by the V1,
the third-order autonomous chaotic oscillator V2, iC1 and iC2 axes), corresponding to the
circuit (Figure 1) are following component values: C1 = 10nF, C2 =
100nF, R = 100Ω, L = 200mH and R1 = R2 = R3
3
x1 ( x3 x1 x1 ) = 2KΩ. As voltage op-amp, TL082CD type ICs
biased with ±9V, have been used. It is gratifying
x2 x2 x3 (2)
to note that the experimental results agree
x3 ( x2 x1 x3 ) qualitatively very well with that of the numerical
results are shown in Figure 5.
3
Where f ( x1 ) x1 x1 (2.1)
CONCLUSION
The dynamics of Eqns. (2) now depends on the It appears that the autonomous chaotic oscillator
parameter α, β, γ and υ. The experimental results circuit presented in this paper is one of the
have been verified by computer simulation of simplest third-order systems reported so far. Its
the normalized Eqns. (2) using the standard simplicity arises from the fact that (i) the
fourth-order Runge-Kutta method for a specific negative conductance is a simple op-amp
choice of system parameters employed in the impedance converter; (ii) the symmetrical cubic
laboratory experiments. nonlinearity is synthesized from mere two signal
That is, in the actual experimental set up the diodes and (iii) the circuit equations are the
inductor L is varied from L = 100mH upwards to most simple because the inductance (L) as the
220mH. Therefore in the numerical simulation control parameter. The attractive features of this
we study the corresponding Eqns.(2) for L in the circuit are the presence of period-doubling route
range L = (100mH, 220mH). From our to chaos, period-doubling window through lower
numerical investigations, we find that for the dimensions of strong chaos followed by
value of L above 100mH, limit cycle motion is boundary condition etc. It is of further interest
obtained, when the value of L is increased, to study these aspects also in this system as well
particularly in the range L = 200mH the system as the intermittency route to chaos and
displays a double band chaotic motion and then synchronization of coupled chaotic circuits of
to period-doubling window through lower the present system for improved high security
dimensions of strong chaos followed by communication systems.
boundary crisis etc.,. These numerical results are
summarized in the phase portraits given in ACKNOWLEDGEMENT
the ( x1 x2 ) , and ( x2 x3 ) planes are shown The authors are thankful to the Management of
in Figure 5. Figure 6 show the numerical Thanthai Hans Roever College of Arts and
chaotic time series were registered using a Science, Perambalur, Tamil Nadu, India for
providing necessary facility for the work.
91 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
REFERENCES 8. A. Tamasevicius A. Namajunas and A.
1. T. Matsumoto IEEE Trans. Circuits Syst. I Cenys Electron. Lett. 32 (1996)957.
31 (1984) 1055. 9. H. B. Fostin and P. Woafo Chaos, Solitons
2. G. Q. Zhong and F Ayrom IEEE Trans. and Fract. 24 (2005) 1363.
Circuits Syst. I 32 (1985) 501. 10. C. C. Hwang H. Y. Chow and Y. K. Wang
3. M. P. Kennedy IEEE Trans. Circuits Syst. I. Physica D 92 (1996) 95.
41 (1994) 771. 11. G. P. King and S. T. Gaito Phys. Rev. E 46
4. A. Namajunas and A. Tamasevicius (1992) 3093.
Electron. Lett. 32 (1996) 945. 12. K. Murali and M. Lakshmanan Phys. Rev. E
5. A. Namajunas and A. Tamasevicius 45 (1993) 1624.
Electron. Lett. 31 (1995) 335. 13. L. O. Chua and G. N. Lin IEEE Trans.
6. M. P. Kenndy Freq. 46 (1992)66. Circuits Syst. 37 (1990) 885.
7. T. Matsumoto L. O. Chua and K. Kobayashi 14. V. Balachandran and G. Kandiban Indian J
IEEE Trans. Circuits Syst. I 33 (1986) Pure & Appl Phy. 47 (2009) 823.
1143.

Fig. 1 Third-order autonomous chaotic oscillator circuit.

92 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
(a) (b)

(c) (d)

Fig 2. Typical experimental phase portraits of the system corresponding to


different regimes.

Fig. 3 Time-domain measurements of the proposed autonomous


third-order chaotic oscillator.

93 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
(a) (b)

Fig. 4 Power spectrum of the signals V1(t) and V2(t) from the circuit of
autonomous third-order chaotic oscillator.
2 "kandiban.dat"u 2:3 0.5 "kandiban.dat"u 2:4
1.5 0.4
0.3
1
0.2
0.5 0.1
0 0
-0.5 -0.1
-0.2
-1
-0.3
-1.5 -0.4
-2 -0.5
-1 -0.8-0.6-0.4-0.2 0 0.2 0.4 0.6 0.8 1 -1 -0.8-0.6-0.4-0.2 0 0.2 0.4 0.6 0.8 1

(a) (b)

0.5 "kandiban.dat"u 3:4 2 "kandiban.dat"u 4:3


0.4 1.5
0.3
1
0.2
0.1 0.5
0 0
-0.1 -0.5
-0.2
-1
-0.3
-0.4 -1.5
-0.5 -2
-2 -1.5 -1 -0.5 0 0.5 1 1.5 2 -0.5-0.4-0.3-0.2-0.1 0 0.1 0.2 0.3 0.4 0.5

(c) (d)

Fig. 5 Typical numerical phase portraits of the system corresponding to


different regimes.

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Vol. 03 issue 07 July 2011
1 "kandiban.dat"u 1:2 2 "kandiban.dat"u 1:3
0.8 1.5
0.6
1
0.4
0.2 0.5
0 0
-0.2 -0.5
-0.4
-1
-0.6
-0.8 -1.5
-1 -2
200 400 600 800 1000 1200 200 400 600 800 1000 1200
(a) (b)

Fig. 6 Numerical time-domain measurements of the proposed autonomous third-


order chaotic oscillator.

95 International Journal of Current Research and Review www.ijcrr.com


Vol. 03 issue 07 July 2011
HOURLY OZONE CONCENTRATION PREDICTION USING
NEURAL NETWORK MODEL
G.Geetha
ijcrr Research Department of Physics, Presidency College, Chennai , Tamil Nadu
Vol 03 issue 07
Category: Research E-mail of corresponding author: gpsgeetha@gmail.com
Received on:01/05/11
Revised on:20/05/11
Accepted on:28/05/11

ABSTRACT
The aim of the present work is to provide a methodological procedure to forecast hourly Ozone
concentrations using Artificial Neural Networks (ANNs). The study area is the urban center of Chennai,
and the results are presented here. The model can predict the mean surface ozone based on the parameters
like concentration of Nitrogen-di-oxide, temperature, relative humidity, sun spot number, wind direction
and wind speed. The model can perform well both in training and independent periods. The achieved
results were satisfactory.

______________________________________________________________________________

Key words: Artificial Neural Networks surface Area of Study and data
ozone Air pollution Ground level ozone concentration and nitrogen
dioxide measurements were carried out in the
INTRODUCTION urban site Chennai, Tamil Nadu, India. India is a
Ground-level ozone air pollution is of great tropical country urbanized district. Its irregular
concern because of its adverse effects on human shape covers about 174 Km2 (M.Pulikesi et al.,
health and ecosystems (Poupkou et al.2008, 2005). It is geographically positioned between
Cristofanelli & Bonasoni 2009). Ground-level 12º9´ and 13º9´ of the Northern latitude and
ozone is not emitted directly into the 80º12´ and 80º19´of the eastern longitude.
atmosphere. It results from photochemical
reactions between oxides of nitrogen (NOx) and MATERIALS AND METHODS
volatile organic compounds (VOCs) in the A portable Aeroqual series S200 ozone monitor
presence of sunlight (Pudasaine etal2006, was used. An Aeroqual series S200 ozone
Vingarzan & Taylor 2003, Clapp & Lenkin monitor is constructed to measure low and high
2001, Sillman 1999). ozone levels. Its ultra low concentration ozone
Present paper aims to develop a simple model head measures the ozone concentration from
using neural network technique based on the 0.000 to 0.500 ppm, and a high concentration
data which are easily available. The performance ozone head measures the ozone Concentration
of the model is satisfactory both in training and from 0.50 to 20.00 ppm. Accuracy of a low
independent period. concentration ozone head is ± 0.001 ppm (from

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Vol. 03 issue 07 July 2011
0 to 0.100 ppm); ± 10% (0.100 to 0.500 ppm), structure of the network established during the
while that of a high concentration ozone head is learning period.
± 10% (from 0.20 to 2.00 ppm); ± 15% (from The transfer function used here is the sigmoidal
2.00 to 20.00 ppm), the measurement unit being function. The ANN‘s are product of the artificial
either ppm or μg/m3. The operating intelligence,which miming the neurons
Temperature range is from – 5o C to 50o C, networks, allow expert systems and learning
relative humidity limits are 5% and 95%. skills..(Benvenuto et al,2000)
Similar kind of NO2 sensor has been used for The process of learning the training set of
nitrogen dioxide measurement. patterns means the determination of the
A gas sensitive semiconductor (GSS) type is optimum weights which minimize the mean
used to measure the ozone and nitrogen dioxide square error between the outputs in the output
values. Gas sensitive sensor head is layer and the desired values. Most commonly
interchangeable and replaceable. LCD type used ―back-propagation learning algorithm‖
display. The temperature range is from -400C to [Rumbelhart et al., (1986)] is used for the
1240C humidity range 0 to 100%. training. Initially random weights between ±0.5
Wind speed is calculated using a wind vane. are assigned to each weight as initial guesses.
Wind velocity is measured using AM- The weights are learned through an iterative
4201digital Anemometer. Measurement in range process. During learning the weights are
0.4 – 3 m\s has resolution 0.1 m\s of accuracy ± updated. When the network learns the training
(2% + 0.2 m\s). The ambient temperature and set of patterns well enough it can be used for
humidity are measured by Thermo Hydrometer. determining the output values for the pattern
Temperature accuracy ± 0.1oCand humidity with unknown outputs (Test period or prediction
accuracy ± 5%. period).
Sampling was carried out for five days from 13-
11-2010 to 17-11-2010 for every 1 hour interval. RESULTS AND DISCUSSION
The data is separated for training of the network
Brief review of neural network technique and the network was trained. The weight values
Neural Networks are signal processing systems were fixed. Remaining data was used for testing
that attempt to emulate the behavior of of the network. The results suggest that the
biological nervous systems by providing a Sunspot number is one of the best predictors for
mathematical model of combination of ozone prediction.
numerous basic blocks called neurons connected Table 1 gives the correlation coefficient values
in a network. It is remotely analogous to living of the set of values. The correlation coefficient
nervous system and hence its name. values give the relationship among the various
As input to the model, a historical set of predictors.
significant meteorological data is used, whereas The result obtained is shown in the Fig.
the output, ozone concentration is predicted by 1.Surface ozone - predicted versus actual values.
the model. The network is trained with the past As indicated by the results it provided the
data. By the proper choice of training sets, after highest performance. This was due to enabling
the learning process, the trained network is of the ozone by Sunspot number and the input
capable of predicting the ozone concentrations parameters, resulting in improved training and
as an output according to the inputs and internal thus improved prediction.

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Vol. 03 issue 07 July 2011
Root Mean Square Error (RMSE) training data 3. Cristofanelli P., and Bonasoni P.,
set 0.176988 % Background ozone in the southern Europe
Root means Square Error (RSME) for testing and Mediterranean area: Influence of the
data 21.168520 % transport and processes. Environmental
Pollution, 2009, Vol. 157, p. 1399-1406.
The above results validate the proposed model. ELSEVIER. ISSN 0269-7491.
4. Poupko A., Symeonidis P., Lisaridis I.,
CONCLUSION Melas D., Ziomas I., Yay O.D., Blis D.
Our results suggest that neural network Effects of anthropogenic emission sources
technique is promising tools for modeling and on maximum ozone concentration over
prediction of ozone. Results can be summarized Greece. Atmospheric Research, 2008, Vol.
as follows; 89, p. 374-381. ELSEVIER.ISSN 1352-
(i) The ANN constructed models suggests high 2310.
feasibility of the application of ANN 5. Pudasainee D., Sapkota B., Manohar L.S.,
technique for the prediction of hourly ozone Kaga A., Kondo A., Inoue Y. Ground
within reasonable error bounds. levels ozone concentration and its
(ii) The predictive analysis suggest a association with NOx and meteorological
considerable link among solar/ Sunspot parameters in Katmandu valley, Nepal.
cycle, related variability and ozone Atmospheric Environment, 2006, Vol. 40,
formation. p. 8081-8087 ELSEVIER.ISSN 1352-2310.
(iii) It is concluded that the above model can be 6. Pulikesi M, P.Baskaralingam, D.Ilango,
used for Predicting surface ozone V.N.Raidu, V.Ramamoorthy, S.Sivanesan,
concentration with nitrogen dioxide, Air quality monitoring in Chennai, India, in
temperature, % relative humidity, Sunspot the summer of 2005. J.Hazard.Mater.
number, wind direction and wind speed as 136(2006)589-596.
predictors. 7. Rumbelhart, D.; Hinton, G.E.; Wiliams,
R.J. (1986): Learning internal
representation by error propagation, In
REFERENCES parallel distributed processing Exploration
1. Chudzynski S., Czyzewski A., Ernst K., in the Microstructure of Cognition, Vol I,
Pietruczuk A., Skubiszak W., Stacewicz T., Cambridge.
Stelmaszczk K., Szymanski A., Sowka I., 8. Sillman S., The relation between ozone,
Zwozdziak A., Zwozdziak J., Observation NOx and hydrocarbons in urban and
of ozone concentration during the solar polluted rural environments. Atmospheric
eclipse. Atmospheric Research, 2001, Vol. Environment, 1999, Vol. 33, P. 1821-1845.
57, p. 43-49. ELSEVIER . ISSN 0169- ELSEVIER.ISSN 1352-2310.
8095. 9. Vingarzan R., Taylor B. Trend analysis of
2. Clapp I., J., Jenkin M., E., Analysis of the ground levels ozone in the greater
relationship between ambient levels of O3 . Vancouver/Fraser Valley area of British
NO2 and NOx as a function of NOx in the Columbia. Atmospheric Environment,
UK Atmospheric Environment. 2001. Vol. 2003, Vol. 37, p. 2159-2171.
35 p.0401-0405 ELSEVIER ISSN 1352- ELSEVIER.ISSN 1352-2310.
2310.
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Vol. 03 issue 07 July 2011
10. Westmoreland E. J., carslaw N., Carslaw 2007, Vol. 41, p. 9195-9205.
D., Gillah A., Bates E Analysis of air ELSEVIER.ISSN 1352-2310.
pollution within a street canyon using 11. www.aeroqual.com/gas sensitive
statistical and dispersion modeling sensor/14-10-2010
techniques. Atmospheric Enviroment,

Table-1 Correlation Coefficient table

O3 P/O3 NO2 WS WD RH TEMP


TEMP 0.506668 0.54723 -0.1864 -0.06267 -0.25268 -0.48434 1
RH -0.17293 -0.23388 0.327279 0.021949 0.053008 1 -0.48434
WD -0.12948 -0.12302 -0.09453 0.474686 1 0.053008 -0.25268
WS -0.0086 0.082046 0.1633 1 0.474686 0.021949 -0.06267
NO2 -0.07775 -0.08468 1 0.1633 -0.09453 0.327279 -0.1864
P/O3 0.677217 1 -0.08468 0.082046 -0.12302 -0.23388 0.54723
O3 1 0.677217 -0.07775 -0.0086 -0.12948 -0.17293 0.506668

Fig.1 Performance of the neural network model

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Vol. 03 issue 07 July 2011
A STUDY ON THE DESIGN OF MICRO-LATHE FOR
EDUCATION AND APPLICATION
S.Syath Abuthakeer1, P.V. Mohanram1, G. Mohan Kumar2
1
Department of Mechanical Engineering, PSG College of Technology, Coimbatore
ijcrr 2
Park college of Engineering, Coimbatore
Vol 03 issue 07
Category: Research
Received on:01/03/11 E-mail of corresponding author: syathpsgtech@gmail.com
Revised on:11/03/11
Accepted on:20/03/11

ABSTRACT
As the factory automation progresses, the number of specialized product is increasing rapidly. Previous
years have been characterized by the growth of 3-D micro-components production. Now-a-days,
machined parts are becoming progressively smaller. So, production of machinery which remains in a
conventional size is often inappropriate for such products. The term ―micro factory‘‘ represents an
entirely new approach to design and manufacture which minimizes production systems to match the size
of the parts they produce. The micro-lathe was one of key components in "Micro-factories" claiming
"small machine tools for small mechanical parts‖. There is an alternative to manufacture micro-
components by micro-machine tools and micro-manipulators using conventional mechanical techniques.
In India, Robot, Micro-factory, several prototypes of micro-machine tools (MMTs) and micro-
manipulators (MMs) have been developed. Furthermore, there is an increased need for engineers trained
in micro-machine tool design and operation. This development necessitates thorough and systematic
education in both industry and education institution. Inexpensive educational micro-machine tools will
facilitate the required education in India. In this study we design and manufacture a prototype of an
inexpensive LabVIEW controlled micro-lathe. This will facilitate the micro-machine tool education in
India as these activities become active.
______________________________________________________________________________

Keywords: Micro-machine, micro-lathe, equipped with more gadgets as miniaturization


LabVIEW goes further. Miniaturization process of
mechanical components started with micro
1 INTRODUCTION fabricated sensors and was followed by micro
1.1. RATIONALE AND STIPULATION OF fabricated parts and micro actuators. In recent
THE MICRO-MACHINE TOOL years integration of micro components such as
There is a great effort towards the precision mechanisms, sensors, actuators and
miniaturization in the last few decades. We can embedded electronic circuits into micro systems
see the effects of that trend in every aspect of has become one of the most prominent research
our lives. From the laptops to the cellular areas all over the world. When the micro
phones, we always prefer the smallest one since components were first introduced, they were
the idea of ―the smaller the better‖ has simple and could be naturally integrated directly
penetrated into our minds and one can be into the product. However, developments in
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micro system technology resulted in a large represents an entirely new approach to design
variety of micro components made from and manufacture that minimize production
dissimilar materials and technologies. These systems to match the size of the parts they
miniaturized products use even smaller produce. In the earliest attempt to turn the
components, and in more and more cases they concept of micro-factory [4] in to a reality a
are micro components with sizes of components micro-lathe smaller than a human palm was
less than one millimeter. developed in 1996[5].
A new bid to manufacture pieces with overall And it was the first big success for the further
sizes smaller than 1mm using conventional step into the concentration on process physics of
mechanical technology was made in[1]. This micromachining including materials and micro
proposal was based on the development of structural effects, machine tools, tooling and
micro-factories (composed of micro-machine sensing, workpiece and design issues, software
tools, micro-manipulators (MMs), assembly and simulation tools, and other issues [6] micro
devices, etc.) to manufacture and assemble 3-D factory. The first micro press was developed in
micro-devices employing conventional the year 2000[7].
mechanical techniques. The micro-factories can In the micro-world, the error sources of MMTs
help to reduce the consumption of resources can be reduced by reducing their sizes
(energy, materials and space); and can help to [8],[9],[10] and [11]. Some prototypes were
increase the productivity[2]. The micro-machine made in order to demonstrate the advantages of
tools (MMTs) in such micro-factories must be this proposal. Countries such as Japan, Ukraine
sufficiently precise to produce components and Mexico have developed micro-machines
according to industrial and research demands. tools with overall sizes from 130 × 160 × 85 mm
The conservation of energy has been the slogan to 32 × 28 × 30 mm[12],[13], and [14]. In
for the past decade in order to reduce energy Mexico, the research in this area began in 1999
consumption. The conservation of energy by and the main goal was to develop micro-
reducing the machine tool size for machining mechanical technology for automated
micro components is attaining popularity as production systems based on low-cost and high
micro-factory. Micro-factory can be said that it efficiency equipment and instrumentation
is a small manufacturing system for achieving [15].In the low-cost micro-equipment
higher throughput with less space and reduced development, the principal challenge is to obtain
consumption of both resource and energy via high precision employing low-cost components.
downsizing of production processes. Advanced countries like Japan, Taiwan, Korea,
Europe, Ukraine ,Mexico, Gemany and USA
The energy-saving effect not only manufacture production micro-
factoriesareminiaturizedto1/2size.Theenergy- machine tool but also educational micro-
saving effect is large when the size of the machine tool in a balanced manner. As the
processing and assembly Equipment is economic development of India progresses
extremely large compared to the dimensions of toward becoming an advanced nation there is a
the products. As for watch manufacturing, the need for an effective policy for science,
amount of Energy consumption may be reduced engineering and technology and their education.
to approximately 30 Percent of the conventional Particularly, that for the machinery, automobile
factory by the half-miniaturization of the and electrical industries is more urgent. Because
production systems [3].The term micro-factory the resources, fund and technology for the
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Vol. 03 issue 07 July 2011
domestic industry are not sufficient, and also electronic technology allows developing micro-
technology protection policy in the level of mechanical components with simple shapes (two
advanced nation is nonexistent, there is a great and a half dimensions), and the materials
deal of problems associated with the Indian employed in this technology are silicon, silicon
Machine tool Industry. oxide, metallic films (mainly aluminum), and
In particular, Micro-machine tool systems which piezoelectric materials like quartz. These micro-
are based on an industry are less manufactured devices applications are encountered in many
in India. Therefore, it is an urgent problem that industries such as the automotive, the
the Indian industry develops capability to biomedical, electronics, computer, etc.
manufacture integrated special tool system, such The manufacture of micro-components
as Micro-factory ,Micro-machine tool, micro- employing micro-mechanical systems (micro-
manipulators, Robots; Educational high level factories) was proposed in the 90‘s as a new
technicians for the special fields. Sufficient alternative to cover some of the micro-world
equipment and other necessary materials are applications where MEMS could not be applied.
needed for experiments essential for effective A research group from Japan proposed the
education. It is more suitable to use specially development of tools that allow generating other
built micro machine tool for educational uses. kind of application which can be made from
In our country, certain educational institutes different materials and can have 3-D geometry
recognize this problem and utilize educational shapes. The main goal was to create MMTs,
micro-machine tool, but this machinery depend micro-manipulators (MMs), etc. at a scale
on total import. Therefore it is expected that this comparable with the size of the produced micro-
research contributes towards micro-equipment components. Their proposal consisted of
education by domestic production of educational transferring the conventional mechanical
MICRO LATHE so it can reduce the import and methods to the micro-world and to develop
foreign exchange and eventually manufacture of micro-factories able to produce micro-devices.
Micro-lathe for production. A micro-factory contains several systems: a
The necessity of educational Micro-machine tool manufacturing system, an assembly system, a
is considered by this method: and an economical quality control system, a transport system, a
prototype micro-lathe is designed, manufactured maintenance system, and others. The micro-
and studied. In the low-cost micro-equipment factories allow a decrease in the consumption of
development, the principal challenge is to obtain energy, space, and resources[16]. The produced
high precision employing low-cost components. micro-components can be used in the watch
For this reason, we have proposed to use the industry, the automotive industry, medical
labVIEW control systems to increase the micro- facilities, biology investigations, etc. [16] and
machine tools accuracy without increasing [17]. For example, in the medical field, the
significantly the total device cost. micro-equipment demands are: microscopy,
1.2 DEVELOPMENT OF MICRO-LATHE diagnosis, non-invasive surgery, etc. in the
An interest to produce mechanical parts with industrial field for the development of micro-
sizes less than 1 mm arose worldwide in the robots to inspect inaccessible or dangerous
80‘s. Some methods based on micro-electronic places, pipe inspection, transportation
technology were proposed; nowadays, these machinery, archeological research, etc. Another
developments are called Micro Electro interesting application field is the development
Mechanical Systems (MEMS) [8]. The micro- of micro-actuators, for example: micro-grippers
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Vol. 03 issue 07 July 2011
for manipulation with living cells, micro- machines with overall sizes of some
generators, micro-motors, etc.[13]. micrometers have been obtained [14]. Based on
The first micro-machine tool was developed in this study, the prototype of LabVIEW controlled
the National Institute of Advance Industrial Micro-lathe is developed for production
Science and Technology of Japan in 1996 [12]. maximized education efficiency, it can be
Nowadays, there are many groups in different learned easily and a disjointing and assembling
countries around the world such as Germany, are possible.
Korea, Switzerland, Mexico, USA, etc. 2 .0 DESIGN OF HEADSTOCK
interested in this field [18], [19] and [20]. Headstock is the heart of any machine tool.
Researchers from the National Institute of Design of head stock covers motor selection,
Advance Industrial Science and Technology of estimation of cutting forces, spindle design,
Japan developed an automated micro-factory to bearing selection, belt drive selection and
produce components for micro-bearings in 2000 headstock housing. It is necessary to justify
[21]. The assembly of micro-bearings was made developed cutting forces with available motor
in the same micro-factory with a semi automated power. Headstock housing has to do properly to
process. prevent transmission of vibration to other
Particularly, in Mexico, investigation in micro- machine elements.
mechanics began in 1999. The main goal is to 2.1 MOTOR SELECTION
create technology for automated micro- DC motors are electric motors that use
mechanical devices for production based on electricity and a magnetic field to produce
low-cost and high efficiency equipment and torque, which turns the motor. To drive the
instrumentation. To achieve this goal, it was spindle, Permanent Magnet DC motor (PMDC),
proposed to work out the micro-equipment as rated power of 1.6 W is selected. Characteristic
sequence of generations where the first curve of the motor is shown in figure 1. Torque
generation of micro-equipment is produced by varies greatly with the speed ranging from a
conventional machine tools. The micro- maximum (stall) torque at zero speed to Zero
equipment of this generation will be able to torque at maximum (no load) speed. This
produce the second generation of micro- relationship comes from the fact that Power =
equipment having smaller overall sizes than the torque X angular velocity. Since the power
previous one. Employing the second generation, available from the motor is limited to some
it would be possible to produce the third finite value, an increase in torque requires a
generation of micro-equipment, and so on. The decrease in angular velocity and vice versa. Its
sizes of each new generation devices are smaller torque is maximum at stall which is typical of all
than the sizes of the prec++edent ones. This electric motors. This is an advantage when
process can be repeated until the micro- starting heavy loads.

Fig 1 Characteristic curve of PMDC


(Source: Igarashi Motors Ltd, Chennai) Fig .2. 2-D Diagram of motor

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From figure 1 load lines represent a time varying are characterized by having only one cutting
load applied to the driven mechanism. The force during machining. But that force is
problem comes from the fact that as the required resolved into two or three components for ease
load torque increases the motor must reduce of analysis and exploitation. Figure 3 visualizes
speed to supply it. Thus input speed will vary in how the single cutting force in turning is
response to load variations in most motors, resolved into three components along the three
regardless of their design.It is a PMDC Motor as orthogonal directions X, Y and Z. The resolution
shown in figure 2 which runs at 10,000 rpm of the force components in turning can be more
(without load condition). conveniently understood from their display in 2-
2.2 ESTIMATION OF CUTTING FORCES D as shown in Figure 4.
The single point cutting tools being used for
turning, shaping, planning, slotting, boring etc.

Fig .3 Cutting forces in turning operation Fig. 4 Turning force resolved into PZ, PX
and PY
Where, PZ = tangential component taken in the direction of Z axis
PX = axial component taken in the direction of longitudinal feed or X axis
PY = radial or transverse component taken along Y axis.
In figure 3 and figure 4 the force components belt-driven spindle or integral motor-spindle
are shown to be acting on the tool. A similar design is required. This depends upon the
set of forces also act on the job at the cutting requirements of the machine tool which also
point but in opposite directions as indicated include the maximum speed, power and stiffness
by PZ', PXY', PX' and PY' in figure 4. required and also cost. Based on these factors
2.3 DESIGN OF SPINDLE belt driven spindle is chosen. Reaction of cutting
Spindles are rotating drive shafts that serve as forces transferred to the spindle through Collet
axes for cutting tools or hold cutting instruments chuck. Based on the reaction forces, shear force
in machine tools. Spindles are essential in and bending moment diagram has been drawn
machine tools and in manufacturing because for different load condition. Optimum space
they are used to make both parts and the tools between bearing supports is given by thumb rule
that make parts, which in turn strongly influence L ≤ (Ds4/3 / k1/3) (1)
production rates and parts quality. To obtain the
desired result, a normal spindle design must take Where L = Optimum span between bearing
into consideration the required power, torque, supports. By logically assume L = 20 mm,
tooling system used, Speed, Accuracy and life. reaction forces at supports are calculated.
2.3.1 SPINDLE STYLE -To get a typical style DS = Average diameter of the supported length
of a spindle, the first thing to decide is whether a of the spindle, 5 mm

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k = 0.1 for precision machine tools factor of safety 3. Mechanical properties of
Belt force at the pulley end is calculated by spindle material are given below:
assuming pulley diameter is equal to 16 mm and Yield strength = 490 MPa; Shear strength =
the belt force = 0.4 N. Radial force acting at the 280 MPa;Young‘s Modulus = 210GPa
centre of pulley is = 1.5 X 0.4 = 0.6 N. Brinell hardness no HB= 192 ;Te = (π/16) X ζs
2.3.2 MATERIAL SELECTION X Ds3
Distortion is the function of Young‘s modulus of Diameter of the spindle = 3mm
the material. Hence to satisfy first requirement However, to prevent angular deflection
the spindle material should have high Young‘s below 250 the following equation is used:
modulus. Costly high tensile steel doesn‘t have
significant stiffness when compared to structural KW = 4mm (2)
Ds 1284
and alloy steel. In our precise machining Rpm
application any alloy steel is suitable. For
calculation purpose, AISI 1040 cold rolled Power is in kW and operational speed is in rpm.
carbon steel properties are taken as it is widely Diameter of the spindle is varying from 3 to
used as spindle material. 5mm. For calculation purpose let us take Ds=
2.3.3 Spindle Design Calculations 5mm. Deflection of the spindle can be calculated
Equivalent bending moment and twisting by using McCauley‘s method. After derivation
moment is already found by using equations the final equation of deflection of the spindle is
8 and 9. Now Spindle has to design to given by
withstand these load conditions by applying
1
y 0.05x 3 153.75x 2137.5 0.1375( x 15) 3 0.485( x 35) 3 (3)
EI
Maximum deflection of the spindle at the machining end ymax = 2.12 microns
2.4 BEARING SELECTION selected base on spindle diameter. After that
Deep groove, ―Conrad‖, ball bearings are the bearing selection is justified with given load
most common form of ball bearings used for conditions. NSK miniature ball bearings
supporting radial load and bidirectional axial specification is used for calculation purpose.
loads equal to the radial load because all Samples of bearings are got from KHF
balls share the load. Balls have to roll from Components private limited, Bangalore.
one side of the contact groove to the other, so They are exporting bearings to NSK
bidirectional stiffness is non-linear. Since in bearings. Table 1 gives specification of
our application radial loads are major loads, miniature ball bearings.
deep groove miniature ball bearings are

Table 1 Specification of miniature ball bearing

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Vol. 03 issue 07 July 2011
BASIC BASIC
OUTSIDE INSIDE LIMITING DYNAMIC STATIC
MODEL THICKNESS
DIAMETER DIAMETER SPEED LOAD LOAD
NO. (mm)
(mm) (mm) (rpm) RATING RATING
(kgf) (kgf)
MR84 8 4 3 56000 40 14
DDL1040 10 4 3 54000 73 40

2.5 CIRCLIP SELECTION Thickness T = 0.4 mm; Clearance diameter C


To prevent axial movement of bearings, = 8.64 mm; Groove width W = 0.64 mm;
Circlip or locknut can be used. Standard Groove diameter G = 3.8 mm; Material =
Circlip dimensions are shown in figure 5. to Stainless steel
meet with spindle dimensions. Specification
of Circlip is given below:

Fig .5 Circlip (Source: Reliance Precision Mechatronics Pvt Ltd)

2.6 MODEL OF SPINDLE in spindle. Step on spindle has given to


3- D model of spindle is shown in figure 6. prevent axial movement bearings.
Grooves have given to accommodate Circlip

Fig .6. 3-D model of Spindle


2.7 BELT AND PULLEY SELECTION belt an amount equal to 1/64th of the span
Timing Belt is used to transmit the power centers ―a‖.
from motor to spindle. Timing belt efficiency The belt must be rigidly mounted. Variations
ranges from 95 -98% better than flat or vee in center distance can lead to premature
belts which rely on friction to transmit wear. The belt and pulley system must be
power. Timing belt drives do not require as assembled loose to prevent over stretching.
much tension as other belt drives which The belts are guided on the pulleys by
depend on friction to transmit the load. The flanges. To minimize belt fatigue pulleys of
belt should be installed with a snug fit, minimum 20 teeth are recommended.
neither taut nor loose. As a general guide the Knowing the centre distance the belt length
correct level of tension can be determined by can be calculated from the following:
measuring the force necessary to deflect the
For ratios = 1:1
LB = (Z * t) + 2a
Where, LB = Length of belt, mm
Z = No. of teeth on pulley = 12 (from standards)
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Vol. 03 issue 07 July 2011
t = Belt Pitch = 2.5 mm (from standards)
a =Centre distance between spindle and motor shaft = 30 mm
Therefore, Length of belt = 90 mm. The standard 2.5 mm pitch timing belt is shown in figure 7

Fig. 7 Timing belt (Source: Reliance Precision Mechatronics Pvt Ltd) Fig . 8 Pulley
Material: High Tensile steel reinforced Polyurethane;Width: 6 mm;Maximum Peripheral load: 65 N
Maximum Peripheral speed: 80m/s;Temperature range: -300 to 800
The pulley corresponding to timing belt is shown in figure 8. It is made of Aluminium pulley and
flange is made of Zinc coated plates. Bore diameter = 3mm; Hub diameter = 12mm.

2.8 MATERIAL SELECTION OF 2.8.1 MODEL OF HEADSTOCK


HEADSTOCK HOUSING HOUSING
The weight of the structure is inversely Typical 3-D model of headstock housing is
proportional to the quantity of unit stiffness. shown in figure 9 and 10. It is split type
Unit stiffness is the ratio of young‘s modulus housing and it has top and bottom casing.
and specific weight. The larger the unit Since it is the first prototype development,
stiffness of the material the smaller is the there will be lot of inspection has to do inside
weight of the structure required to ensure that the microlathe. There will not be much
the deflection of the structure due to a intention given to functionality. It is just
particular given load does not exceed a claiming for operation. Hence split type
specified value. Mild steel has unit stiffness housing is selected. Top casing will be
of 2.69 X108 and cast iron has a unit assembled with lathe bed. The detailed 2-D
stiffness of 1.66 X108. Hence Mild steel is diagram is available in appendix.
selected as material for headstock housing.

Fig 9 Bottom casing Fig 10 Top casing


2.9. MATERIAL SELECTION OF slide ways. This, in turn, results in an
SLIDEWAY DESIGN inevitable loss of accuracy of travel. The
The wear of slideways depends to wear resistance of slideways is determined
considerable extent upon what materials are primarily by the physic-mechanical
used to make the ways of the bed and of the properties of their material. In most cases it is
travelling unit. An inexpedient selection of more expedient to use the harder material for
these materials may lead to premature wear the stationary slideways since their shape is
which is not uniform along the length of the copied in travel of the moving unit.
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The most common slideway materials are Slideways are designed for the following two
cast-iron and low carbon or alloyed steel. If parameters:
the slideway is integral with the bed, it is Wear resistance and Stiffness
almost exclusively made from gray cast iron. The wear resistance of slideways depends
This material is cheap but does not have upon various conditions,
good wear resistance when subjected heavy
p m ax [ p m ax ]
one of the most important
loading. The wear resistance can be being as uniform as possible distribution of
improved through proper heat treatment. If the pressure over the way surfaces, the
both slideways are hardened the average specific pressure not exceeding a
improvement may be up to four times. certain definite value established on the basis
Therefore, gray cast-iron is chosen as of experience in machine tool operation. The
slideway material for this application. specific pressure is determined by checking
2.9.1 SHAPE OF SLIDEWAYS calculations based on the assumption that the
The commonly used shapes of slideways specific pressure is distributed according to
used in machine tools are flat, V, dovetail, linear function lengthwise along the
and cylindrical. They may be closed or open. slideway, across the width of each face of the
Closed flat and closed V is employed for slideway; the specific pressure is considered
precision application. The included angle of to be distributed uniformly.
V, both in closed and open version is The wear resistance of slideways is
generally 900. For proper functioning of governed mainly by the maximum pressure
slideways it is imperative that the friction be acting on the mating surfaces. This condition
kept as low as possible by ensuring that a may be written down as
certain minimum amount of lubricant is
always present between sliding surfaces.
2.9.2 DESIGN CRITERIA AND
CALCULATIONS
(4)

Here, Pmax = Maximum pressure acting on the mating surfaces


[Pmax]= Permissible value of the maximum pressure
It will be seen during the subsequent analysis that slideway design in terms of maximum pressure is
quite complicated. Sometimes, this design is replaced by a simple procedure based upon the average
pressure acting on the mating surfaces. The condition is that:
(5)
p av [ p av ]
Here, Pav = Average pressure acting on the mating surfaces
[Pav]= Permissible value of the average pressure
The design condition for stiffness stipulates that the deflections of the cutting edge in directions that
significantly influences the machining accuracy should not exceed certain permissible values. The
condition may be expressed as follows:
[ ] (6)
i i

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2.10 MODEL OF GUIDEWAYS
Typical 3-D model of lathe bed, tool post guideway and tool post is shown in figure 11,12 and 13
respectively.

Fig. 11 Lathe bed Fig .12 Tool post guide way Fig. 13 Tool post

2.11 DESIGN OF NUT


Nut is designed based on the lead screw profile. There is a relationship available to find out the length of
nut. The relationship is given below:
λ’= L/dp (7)
Where λ‘ – Constant, it varies between 3 and 5. Let us assume λ‘ = 3
dP- Pitch diameter of thread = 4 mm
From the equation 7 length of nut can be found.
Length of nut (L) = 12 mm.
For support, miniature ball bearings are used with simply supported as shown in figure 14. Bearing inner
bore diameter is 3mm.

Fig 14 Bearing supports for lead screw

2.12MODEL OF LEAD SCREW AND NUT designed based on load conditions that act on
Typical 3- D model of lead screw and nut is the lathe bed. Hence the same lead screw
shown in figure 15 and 16. The lead screw is profile is used for both lathe bed and tool
post guideway.

Fig 15 Lead screw Fig 16 Nut


Since run out leads to efficiency of the lead screw, it has to maintain with high accuracy. For
precision applications, the value is 4 microns.
3. RESULTS AND DISCUSSIONS electronic devices. The two main functions of
The operating features of machine tool and, in machine tool control system are changing
particular, its production capacity, convenience speeds, feeds and providing the working and
and ease in servicing and its reliability in auxiliary motions in the desired sequence
operation, depend to great extent upon how well necessary for machining a particular part.In this
its control system has been designed. The microlathe, LabVIEW software is used to
control system of a machine tool is often control the feed movement of lead screw
combination of mechanical, electrical and coupled with stepper motor. Stepper motor is
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interfaced with LabVIEW software through Data The spindle is driven by a PMDC motor. The
Acquisition Card. The operative member of the linear axis is controlled by stepper motor is
machine tool is directed to move to a certain shown in figure 25.The experimental setup of
position. However, whether it precisely arrives micro lathe is shown in figure 17.
at the desired position or not is not ascertained.
For simplified design, in this application open
loop control system is collected.
Stepper motor

Micro-Lathe

DAQ Card

Fig.17 Experimental setup of the microlathe with stepper motor

A stepper motor, as its name suggests, moves incrementally that many number of steps and
one step at a time, unlike those conventional stops.
motors, which spin continuously. If we figure 18 shows 2-D diagram of stepper
command a stepper motor to move some motor.
specific number of steps, it rotates

Fig 18 Stepper motor (Source: ARK Motion Controls Pvt Ltd., Kochin)
Table 3 describes specifications of stepper and Z drives can be controlled at the time
motor. Maximum pulling force is only 5N. thus making operations taper turning
The feed movement also will be small. To possible. The linear axis drive contains two
suit this requirement, stepper motor is stepper motors. These two stepper motors are
selected. connected through LabVIEW, the linear axis
The linear axis drive is used for controlling drive can be controlled.
the X and Z axis of the Micro lathe. The
linear axis drive is a combination of two Virtual instruments get their name because of
stepper motors with slides mounted on them. the reason they imitate physical instruments
The tool post is mounted on the drive. By such as oscilloscopes and multimeters. The
controlling the linear axis drive through VI software used in the project is LabVIEW
LabVIEW software, the feed and depth of cut 8.5. LabVIEW stands for Laboratory Virtual
of the operations are controlled. Both the X Instrumentation Engineering Workbench.
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The LabVIEW software is used to control the and Z axis coordinates and their direction
PMDC motor, and stepper motors that already extracted are then passed on to the
operate the spindle and lead screws sub VI called stepper VI.
respectively. The program terminates whenever it
The program consists of two VIs, one for encounters the code M30.
reading the CNC codes and another to The linear axis drive contains two stepper
control the stepper motors according to the motors. By interfacing these stepper motors
codes. with virtual instruments, the linear axis can
The sequence of tasks done in the first decoder be controlled. The driver circuit for stepper
VI is explained below. motor consists of a adaptor (transformer) to
The file containing the CNC codes is given convert 230V into 12V since the rating of the
as input to the VI [10] i.e. the operator has to motor is 12 V. There are four MOSFETs, one
specify the location of the file containing the for each coil. The LEDs indicate which coil
required codes. is being energized at that particular time. The
The VI splits the program into individual stepper motor can be controlled by
lines using the ― ; ― as the separator energizing its four coils in a sequence.
The individual lines are then split into The sequence in which the stepper motor
separate words using the blank space as the must be energized can be entered into an
separator. array of LEDs (Boolean array). If the motor
The words are arranged into an string array. operates in full step mode, it has four
The Different parameters such as spindle energizing steps i.e. each of the four coils is
speed, feed, X and Z axis coordinates and executed one after the other.
their direction are extracted from the words.
The string array is read by a separate loop
containing a case structure. Different
parameters such as spindle speed, feed, X

Fig.19 Boolean array in full step mode

In the above figure 19 each column represents a are executed simultaneously in the coil. The half
coil of the stepper motor and each row denotes step mode gives very precise movement
an energizing steps. The glowing LED denotes compared to the full step mode. Hence full step
the excitation of that particular coil. Similarly in mode of operation has been adopted for the
the half step mode there are eight energizing purpose of controlling the linear tool movement.
steps. In half step mode not only single coils are
executed separately but also consecutive coils

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Delay The stepper motor is controlled by generating a
The delay is the amount of time (in milliseconds series of digital pulses in the sequence
the computer waits between the subsequent mentioned above. The DAQ is placed inside the
executions of the energizing steps. Delay is used for loop. The DAQ passes the signals from the
to control the speed of the motor. More the delay computer to the motor through the driver circuit
lower is the speed. after amplification.

The stepper motor is driven with the help of the


No of Steps parameters obtained from the previous decoder
It is a factor that denotes the angle the motor VI.
must rotate. Motor in half step mode turns 0.9 The X/Z coordinate is converted into no of
rotations for every step. steps, which the motor has to execute to
Direction move the tool to the desired location
The motor can be made to rotate in both (coordinates). The conversion is obtained
clockwise and anti clockwise directions. using the relation
Anticlockwise direction is obtained by giving X/Z Coordinate No. of steps required for
the above mentioned energizing sequence in moving the tool by 1 mm
reverse.

Since pitch of the lead screw is the linear tool Direction i.e. positive or negative X/Z
movement obtained for one full rotation of the coordinate is passed as a Boolean signal from
lead screw and hence for one full rotation of the the decoder VI.
motor (the motor is coupled directly to the lead If coordinates are positive then true signal is
screw). obtained and the motor rotates in clockwise
The feed obtained from the decoder VI is direction, on the other hand false value is passed
converted to DELAY. if the coordinates are negative and the motor
DELAY is the amount of time the computer rotates in anti clockwise direction.The final
must wait before giving the next pulse to the Exploded and assembly view of Micro-lathe is
motor or in other words the time the computer shown in fig 20 and 21
waits after every execution of a step;.

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Top casing
Spindle

Tool Post Guideway

Motor
Nut

Bottom casing Lathe Bed Tool Post Lead screw

Fig 20 Exploded view of microlathe

Fig 21 Assembly view of Microlathe

4. CONCLUSION Symposium on Micromechatronics and


In this research we developed Micro-lathe for Human Science, Japan; 1997. p. 13–20.
education institute by using LabVIEW software 3. N.Kawahara,T.Suto,T.Hirano,Y.Ishikawa,T.
.This developed Micro-Lathe will contribute to Kitahara,N.Ooyama,T.Ataka, Technical
Micro-factory education and Micro-machine papers ;Micro system Technologies Micro
domestic production for education, and to factories; new applications of micro
prototype FMS for education by combining machine technology to the Manufacture of
Micro-Lathe, Micro Milling, Micro grinding , small products (1997) 37—41 Springer-
Micro cutting, micro-manipulator, micro- Verlag 1997
machine tool and Micro-Factory system. 4. Kitahara.T, Ishikawa.Y, Terada.K,
Nakajima.N, and Furuta K., ―Development
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