Documenti di Didattica
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Editorial Board
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Dr. Ann Magoufis Director, Ariston College, Shannon, Ireland
Dr. Pongsak Faculty, Ubon Ratchathani University, Warin
Rattanachaikunsopon Chamrap, Ubon Ratchathani, Thailand
Dr. Chellappan Dean, School of Pharmacy, Masterskill
Dinesh University College of Health Sciences, Cheras,
Malaysia
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Dr. R. O. Ganjiwale HOD, Department of Pharmacognosy, I.P.E.R.
Wardha, Maharashtra
Dr. Shailesh Wader HOD, Department of Pharmaceutical Chemistry,
IPER, Wardha, MH, India
Dr. Alabi Olufemi Faculty, Bowen University, Iwo, Osun-State,
Mobolaji Nigeria
Dr. Joshua Danso Faculty, University of Cape Coast, Cape Coast,
ISSN 0975-5241 Owusu-Sekyere Ghana
Dr. Okorie Faculty, University of Nigeria Nsukka, Enugu
IC Value of Journal: 4.18 Ndidiamaka Hannah State
Dr. Parichat Faculty, Ubon Ratchathani University, Warin
“Let the science be your passion” Phumkhachorn Chamrap, Ubon Ratchathani, Thailand
Dr. Manoj Charde Dean, NRI Group of Post Graduate Studies,
Bhopal
Dr. Shah Murad HOD, Pharmacology and Therapeutics, Lahore
Vol 3 / Issue 7 / July 2011
Mastoi Medical and Dental College, Lahore, Pakistan
International Journal of Current Research and Review (ijcrr) is one of the popular
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Aims and Scope:
ijcrr is a monthly indexed international journal publishing the finest peer-reviewed
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Mission Statement:
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To set a landmark by encouraging and awarding publication of quality research and
review in all streams of Medical and Paramedical sciences.
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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
C
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
R
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
R
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
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.
__________________________________________________________________________
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)
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
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
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.
Table 3 The inter planar spacing, d for Sb-Bi-S compound from XRD analysis
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
( b )
I n t e n s i t y ( a. u )
( a )
(a)
0 10 20 30 40 50 60
( b )
I n t e n s i t y ( a. u )
( a )
(c)
10 20 30 40 50
(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
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)
(a)
60
(a)
(b)
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)
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)
60
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
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 )
(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
(a)
Relative Intensity (cps)
Bi ( 4 d 5/2 )
(b)
S(2p)
Relative Intensity (cps)
(c)
(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
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
improvement of glutathione metabolizing membrane structure causally involved in
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
these oxidative changes wherein Chloroform Gerontol A Biol Sci Med Sci. 2001;
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
charge levels during ageing. The outcome of the aggregation. Colloids Surf. Physiochem.
present study thus indicates, Chloroform extract 2000; 164:95–104.
of Solanum trilobatum supplementation could be 9. Aminoff D. Methods for the quantitative
an exact therapy for minimizing the age related estimation of N-acetylneuraminic acid and
alterations in erythrocytes for the existing their application to hydrolysates of
population, as the health care and welfare of the sialomucoids.. Biochem J. 1961; 81:384-
aged have been attracting considerably as a 92.
social problem. 10. Wautier JL, Paton RC, Wautier MP,
Pintigny D, Abadie E, Passa P, Caen J.
Increased adhesion of erythrocytes to
REFERENCES endothelial cells in diabetes mellitus and its
1. Kowald A. Theoretical Gompertzian relation to vascular complications. N Engl J
implications on life span variability among Med. 1981; 305: 237-42.
genotypically identical animals. Mech 11. Beppu M, Hayashi T, Hasegawa T,
Ageing Dev. 1999; 110:101-7. Kikugawa K. Recognition of
sialosaccharide chain of glycophorin on
44 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
damaged erythrocytes by macrophage terminal kinase (JNK), c-Jun and caspase-
scavenger receptors. Biochim. Biophys. 3. Biochem J. 2001; 358:547–57.
Acta. 1995; 1268: 9–19. 20. Hininger JF, Dragst LO, Daneshvar B,
12. Ando K, Kikugawa K, Beppu M. Binding Lauridsen ST, Hansen, Sandstrom MB. The
of anti-band 3 autoantibody to sialylated effect of grape-skin extract on oxidative
poly-N-acetyllactosaminyl sugar chains of status. Br J Nutr. 2000; 84:505–13.
band 3 glycoprotein on polyvinylidene 21. Abu-Amsha Caccetta R, Burke V, Mori
difluoride membrane and sepharose gel: TA, Beilin LJ, Puddey IB, Croft KD. Red
further evidence for anti-band 3 wine polyphenols, in the absence of
autoantibody binding to the sugar chains of alcohol, reduce lipid peroxidative stress in
oxidized and senescent erythrocytes. J smoking subjects. Free Rad Biol Med.
Biochem. 1996; 119:639-47. 2001; 30:636–42.
13. Chien S, Weinburg R, Li S, Li Y. Endo- 22. Natella F, Ghiselli A, Guidi A, Ursini F,
beta-N-acetylglucosaminidase from fig Scaccini C. Red wine mitigates the
latex. Biochem Biophys Res Commun. postprandial increase of LDL susceptibility
1976; 76:317-23. to oxidation. Free Rad Biol Med. 2001;
14. Sangeetha P, Balu M, Haripriya D, 30:1036–44.
Panneerselvam C. Age associated changes 23. Breinholt V, Lauridsen ST, Dragsted LO.
in erythrocyte membrane surface charge: Differential effects of dietary flavonoids on
Modulatory role of grape seed drug metabolizing and antioxidant enzymes
proanthocyanidins. Exp Gerontol. 2005; in female rat. Xenobiotica. 1999; 29:1227–
40: 820-8. 40.
15. Cutler RG, Rodriguez H. (Eds.). Critical 24. Monavalli B, Raja Rajeswari A, Gowri V,
Reviews of Oxidative Stress and Aging: Kanchana A. Invitro Antioxidant Activity
Advances in Basic Science, Diagnostics of Methanolic Extract of Solanum
and Intervention. World Scientific trilobatum Leaves. J. of Natural Science
Publishing, Singapore. 2003; 1523. and Technology - Life Sciences and
16. Duthie GG, Duthie SJ, Kyle JAM. Plant Bioinformatics. 2010; 2:168-174.
polyphenols in cancer and heart disease: 25. Mohanan PV, Devi KS. Effect of Sobatum
implications as nutritional antioxidants. on radiation-induced toxicity in mice.
Nutr Res Rev. 2000; 13:79–106. Cancer Lett. 1998; 123:141-5.
17. Trichopoulou A, Vasilopoulou E. 26. Dodge JT, Mitchell C, Hanahan DJ. The
Mediterranean diet and longevity. Br J preparation and chemical characteristics of
Nutr. 2000; 84:205–9. hemoglobin-free ghosts of human
18. Youdim KA, Martin A, Joseph JA. erythrocyte. Arch. Biochem. Biophys.
Incorporation of the elderberry 1963; 100:119–130.
anthocyanins by endothelial cells increases 27. Drabkin DL, Austin JH.
protection against oxidative stress. Free Spectrophotometric studies,
Rad Biol Med. 2000; 29:51–60. spectrophotometric constants for common
19. Schroeter H, Spencer JPE. Rice-Evans C, hemoglobin derivatives in human, dog and
Williams RJ. Flavonoids protect neurons rabbit blood. J. Biol. Chem. 1932; 98:719–
from oxidized lowdensity-lipoprotein- 733.
induced apoptosis involving c-Jun N-
45 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
28. Walter H. Surface properties of cells component of glutathione peroxidase
reflected by partitioning red blood cells as a purification and assay. Science 1973;
model. In: Walter H, Brooks, D.E., Fisher, 179:588–590.
D. (Eds.), Partitioning in Aqueous Two- 38. Staal GE, Visser J, Veeger C. Purification
Phase Systems. Academic, Orlando, FL, and properties of glutathione reductase of
1985; 327–376. human erythrocytes. Biochim. Biophys.
29. Lowry OH, Rosebrough NJ, Farr AL, Acta 1969; 185:39–48.
Randall RJ. Protein measurement with the 39. Moron MS, Depierre JW, Mannervik B.
Folin phenol reagent. J. Biol. Chem. 1951; Levels of glutathione, glutathione reductase
193:265– 275. and glutathione S-transferase activities in
30. Levine RL, Williams JA, Stadtman ER, and rat lung and liver. Biochim. Biophys. Acta.
Shacter E. Carbonyl assay for 1979; 582:67–78.
determination of oxidatively modified 40. Habig WH, Pabst MJ, Jakoby WB.
proteins. Methods Enzymol. 1994; Glutathione S-transferases, the first
233:346—357. enzymatic step in mercapturic acid
31. Folch J, Less M, Sloane Stanley GH. A formation. J. Biol. Chem. 1974; 249:7130–
simple method for the isolation and 7139.
purification of total lipids from animal 41. Zinkham WH, Lenhard RE, Childs B. A
tissues. J. Biol. Chem. 1957; 226:497–509. deficiency of glucose-6-phosphate
32. Niebes P. Determination of enzymes and dehydrogenase activity in erythrocytes
degradation products of from patients with favism. Bull. Johns
glycosaminoglycans metabolism in the Hopkins Hosp. 1958; 102:169–175.
serum of healthy and varicose subjects. 42. Tietze F. Enzymic method for quantitative
Clin. Chim. Acta. 1972; 42:399–408. determination of nanogram amounts of
33. Wagner WD. A more sensitive assay total and oxidized glutathione: applications
discriminating galactosamine and to mammalian blood and other tissues.
glucosamine in mixtures. Anal. Biochem. Anal. Biochem. 1969; 27:502–522.
1979; 94:394–396. 43. Samoilov MV, Mishnev OD, Kudriavtser
34. Warren L. The thiobarbituric acid assay of IV, Naumor AG, Daniltov AP.
sialic acid. J. Biol. Chem. 1959; 234:1971– Morphofunctional characteristics of
1975. erythrocytes in chronic kidney failure and
35. Marklund S, Marklund G. Involvement of purulent intoxication. Klin. Lab. Diagn.
the superoxide anion radical in the 2002; 6:18–23.
autooxidation of pyrogallol and a 44. Stadtman ER. Protein oxidation in aging
convenient assay for superoxide dismutase. and age-related diseases. Ann N Y Acad
Eur. J. Biochem. 1974; 47:469–474. Sci. 2001; 928, 22-38.
36. Beers RF, Seizer IW. A spectrophotometric 45. Beal MF. Oxidatively modified proteins in
method for measuring breakdown of aging and disease. Free Radic Biol Med.
hydrogen peroxide by catalase. J. Biol. 2002; 32: 797-803.
Chem. 1952; 195:133–140. 46. Hyun DH, Lee M, Halliwell B, Jenner P.
37. Rotruck JT, Pope AL, Ganther HE, Proteasomal inhibition causes the
Swanson AB, Hafeman DG, Hoekstra formation of protein aggregates containing
WG,. Selenium: biochemical role as a
46 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
a wide range of proteins, including nitrated reaction between desferal and the
proteins. J Neurochem. 2003; 86:363-73 superoxide radical. Biochem Pharmacol.
47. Arivazhagan P, Panneerselvam C. 1985; 342:229–233.
Favorable effect of L-Carnitine
supplementation on glycoprotein status in 55. Davis TA, Anderson EC, Ginsburg AV,
aged rats. J. Clin. Biochem. Nutr. 1999; Goldberg AP. Weight loss mproves
26:193–200. lipoprotein lipid profiles in patients with
48. Carney JM, Starke-Reed PE, Oliver CN, hypercholesterolemia. J Lab Clin Med.
Landum RW, Chen MS, Wu JF, 1985; 106:447-54
Froemming GR, O‘Brien NM. U937 cells 56. Inal MG, Kanbak G, Sunal. Antioxidant
as a model to study the effect of enzume activities and malondialdehyde
phytochemicals on superoxide anion levels related to ageing. Clin. Chim.
production. Nutr Res. 1997; 17:1091–1103. Actam. 2001; 305:75-80
49. Leb L, Snyder LM, Fortier NL, Anderson 57. Sini N, Devi KS. Pharmaceutical biology.
M. Antiglobulin serum mediated Antioxidant activities of the chloroform
phagocytosis of normal senescent and extract of Solanum trilobatum. 2004;
oxidized RBC: role of anti-IgM 959:30-44.
immunoglobulins in phagocytic 58. Pastore A, Federici G, Bertini E, Piemonte
recognition. Br. J. Haematol. 1987; F. Analysis of glutathione: implication in
66:565–570. redox and detoxification. Clin Chim Acta.
50. Vomel T. Properties of ATPases and 2003; 333:19-39.
energy-rich phosphate in erythrocyte of 59. John S, Kale M, Rathore N, Bhatnagar D.
young and old individuals. Gerontology. Protective effect of vitamin E in dimethoate
1984; 30:22–25. and malathion induced oxidative stress in
51. Sato M, Maulik N, Das DK. rat erythrocytes. J Nutr Biochem. 2001;
Cardioprotection with alcohol: role of both 12:500-504.
alcohol and polyphenolic antioxidants. Ann 60. Sastre J, Asensi M, Gasco E, Pallardo FV,
N Y Acad Sci. 2002; 957:122-35. Ferrero JA, Furukawa T, Vina J.
52. Kalyan Reddy, Manda Craig Adams, Nuren Exhaustive physical exercise causes
Ercal. Biologically important thiols in oxidation of glutathione status in blood:
aqueous extracts of spices and evaluation prevention by antioxidant administration.
of their in vitro antioxidant properties. Am J Physiol. 1992; 263:992-5.
Food Chemistry. 2010; 118:589-593. 61. Sah NK, Taneja TK, Pathak N, Begum R,
53. VanDuijn MM, Van den Zee J, Athar M and Hasnain SE. The baculovirus
Vansteveninck J, Van den Broek PJ. antiapoptotic p35 gene also functions via
Ascorbate stimulates ferricyanide reduction an oxidant-dependent pathway. Proc. Natl.
in HL-60 cells through a mechanism Acad. Sci. USA. 1999; 96: 4838–4843.
distinct from the NADH-dependent plasma 62. Choe M, Jackson C, Yu BP, 1995. Lipid
membrane reductase. J. Biol. Chem. 1998; peroxidation contributes to age-related
273:13415–13420. membrane rigidity. Free Radic Biol Med.
54. Halliwell B. Use of desferrioxamine as a 18:977-84.
'probe' for iron-dependent formation of 63. Ishige K, Schubert D, Sagara Y. Flavonoids
hydroxyl radicals. Evidence for a direct protect neuronal cells from oxidative stress
Table 1: Levels of GSH, GSSG, GSH/GSSG and redox state in control and CST treated young and
aged rats
Parameters
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
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
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
600
b*
500
a* b*
400
a*
300
200
100 b*
a*
0
Group Ia Group Id Group IIa Group IId
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
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
53 International Journal of Current Research and Review www.ijcrr.com
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
54 International Journal of Current Research and Review www.ijcrr.com
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
55 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
serum is α1-antitrypsin. It has the highest molar oligosaccharides of lysosomal enzymes. J
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
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.
______________________________________________________________________________
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.
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
Writing expanded field notes on same day (in the local language-Gujarati)
Data entry
Data analysis
No Treatment
Home Remedies
NGO/GO
General Practitioner
Consultant
Government Hospital
VCTC
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
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
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
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
MALES FEMALES
DISTANCE (meters) 560±61 514±32
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:
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)
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
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
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)
3
5
90 0 0 0
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.
______________________________________________________________________________
(c) (d)
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)
(c) (d)
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
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.
______________________________________________________________________________
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
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.
Fig. 11 Lathe bed Fig .12 Tool post guide way Fig. 13 Tool post
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.
Micro-Lathe
DAQ Card
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.
110 International Journal of Current Research and Review www.ijcrr.com
Vol. 03 issue 07 July 2011
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
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
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;.
Motor
Nut