QUANTITATIVE DETERMINATION OF COPPER

(II)
CONCENTRATION
BY
SPECTROPHOTOMETRY
J.G.K.B. CORTEZ1
1

COLLEGE OF PUBLIC HEALTH

UNIVERSITY OF THE PHILIPPINES, ERMITA, CITY OF MANILA 1000, PHILIPPINES
DATE SUBMITTED: JULY 16, 2015
DATE PERFORMED: JULY 14, 2015

ANSWERS TO QUESTIONS
1. What is the significance of the addition of ammonia to Cu (II) solutions.
The presence of ammonia aids in providing the solution with a more intense color to
be examined effectively through the spectrophotometer. With the addition of
ammonia to the Cu2+, a complex of the two is produced as seen in equation 1. This
complex is the source of the intense blue color of the solution. The color of the
solution makes it possible for the spectrophotometer to measure its absorbance (Birk,
2001).

(1)

2. Why is Beer-Lambert
transmittance?

Law

NH
( 3)4
Cu

2+

2++ 4 NH 3

Cu

expressed

in

terms

of

absorbance

instead

of

Absorbance is the measure of amount of light absorbed at a particular wavelength as

the light passes through a sample substance (Skoog, West, Holler, & Crouch, 2004).
The Beer-Lambert Law is the linear relationship between absorbance and
concentration of an absorbing species. Equation 2 shows the embodiment of BeerLambert Law to show this direct relationship (Kazakevich, 2010) .

A=abc
Wherein a a=absorptivity of the substance, b= path length, c= concentration of the
component of interest in the solution with units of ppm.
With respect to transmittance (T), which shows the fraction of light able to pass the
sample, absorbance values can be directly measured and is more definitive due to its
ability to display the simple dependence on the concentration and cell path length

(Skoog, West, Holler, & Crouch, 2004). This makes it easier for chemists to use
absorbance
3. What are the limitations of the Beers Law?
One of the most prominent limitations of Beers Law is that solutions should be
properly diluted with concentrations less than 0.01M. Any concentration greater
than 0.01M will exhibit electrostatic interactions between molecules in close
proximity which will result to deviations in absorptivity of the solution. Another effect
of higher concentrations is the slight increase in the refractive index of the particles
present in the solution (Kazakevich, 2010).
Another limitation is the introduction of stray light during spectrophotometry. When
stray light is introduced, the wavelength of the light source will disturbed by an
external source. This will result to a decrease in the absorbance value (Kazakevich,
2010).
4. Why is it significant to scan over a wavelength range? Why is the analytical
wavelength used in the determination of absorbance of the standard and sample
solutions?
The usage of a range is practical due to its ability to display the absorbance value
within the restricted limit of wavelengths. The change in absorbance per unit
concentration is greatest is usually observed at this range of values. The calibration
curve then becomes much wider and will be able to exhibit a much larger working
scale for the analysis. Because of this, there is a higher chance that the absorbance
will be observed in this range (Skoog, West, Holler, & Crouch, 2004).
Apart from this, certain phenomena affect the measurements of absorption of
substances. Molecular vibration and rotation are the reason absorbance readings are
usually observed as bands of absorption in a range of wavelengths and not as a
single sharp rise in the spectrum. This makes the usage of a wavelength range ideal
for solutions (Caprette, 1996).
5. Why do we have to measure absorbance reading against reagent blank solutions?
The usage of blank solutions is performed in order to eliminate the effects of certain
factors to the absorbance reading of the spectrophotometer. The blank transmits as
much light as possible to calibrate the machine before being subjected to the sample
solutions. This will prevent the effects of scattering particles, light refraction and
reflection of the cuvette, and the absorbance of ammonia solution (Caprette, 1996).
6. What is the significance of the y-intercept of your calibration curve?
deviation from the theoretical value.

Discuss its

The y-intercept of the calibration curve represents the absorbance capacity of the
blank solution. With regards to the computation of the researcher, the y-intercept for
our equation is 0.0197. This particular value is subtracted in order to get the exact
absorbance of the solution being measured to negate the influence of unwanted
factors in the sample. This value is the absorbance measured from the reagent blank
tube. (Caprette, 1996)
7. Cite other analytical applications of spectrophotometry.

As a public health student, the researcher has been exposed to a lot of

spectrophotometric methods in the field of healthcare. Determination of different
vitamins and other micronutrients are done through measurement of transmittance
of light through samples.
Additional to these methods is the significance of spectrophotometry in toxicology.
Toxins, if enough sample is present, can be identified with the use of spectrum
analysis. Apart from toxins, certain hormones are also measured with the use of this
method. Steroid hormones such as progesterone and estrogen values are monitored
through spectrophotometry. Apart from hormones, more biological markers in the
field of pathology are measured through spectrophotometry. (Rand, 1972)
8. What are the possible sources of errors and their effect on the calculated
parameters? Rationalize.
As the researchers have stated in question no. 3 and 5, sources of errors in
spectrophotometry can be attributed to concentration of the solution, the solution
itself, and the spectrometer itself. These sources affect the absorbance readings
differently depending on its discrepancy to the standard conditions used in
measuring for theoretical values.
Concentrations greater than 0.01 M are not practically used in spectrophotometry
due to the electrostatic between the molecules/particles, which will result to higher
absorbance values than usual (Kazakevich, 2010).
The solutions being used should also be stable. This means that there are no side
reactions happening when the method is being done. Precipitation and formation
reactions will be able to produce bigger particles thus blocking the light passing
through the sample. If such reactions are occurring, the values obtained will be
higher (Skoog, West, Holler, & Crouch, 2004).
Apart from this, if the spectrometer has defects such as stray light, light source
problems, or the uncontrolled polychromatic light (multiple wavelengths) , deviations
from the theoretical absorbance values will surely be observed (Kazakevich, 2010).
Failure to use reagent blank also opens the possibility of the effects of scattering
particles, light refraction and reflection through the cuvette, and the influence of
ammonias absorptive properties. These factors will increase the absorbance
readings (Kazakevich, 2010).

REFERENCES
Birk, J. (2001, December). Copper. Retrieved July 15, 2015, from General Chemistry
with Qualitative Analysis:
http://www.public.asu.edu/~jpbirk/qual/qualanal/copper.html
Caprette, D. (1996). Principles of Spectrophotometry. Retrieved 2015, from
Experimental Biosciences:
http://www.ruf.rice.edu/~bioslabs/methods/protein/spectrophotometer.html
Kazakevich, Y. (2010). Beer-Lambert Law. Retrieved July 15, 2015, from Analytical
Chemistry: https://hplc.chem.shu.edu/NEW/Undergrad/Molec_Spectr/Lambert.html
Rand, R. (1972). The Role of Spectrophotometric Standards in the Clinical Chemistry
Laboratory. Journal of Research of the National Bureau of Standards .

Skoog, D., West, D., Holler, J., & Crouch, S. (2004). Fundamentals of Analytic
Chemistry. In S. Kiselica (Ed.). Thomson Learning Academic Resource Center.

GRAPHS AND COMPUTATIONS

Concentration of Standard Cu (II), ppm

M Cu ( II )=

2500 x 2
=100 ppm
50

Linear Equation of Calibration Curve

y=0.000199x+0.0197

Figure1. Calibration Curve

Grubbs Test

Gexp=

X q X
S

Gexp=

0.0920.0843
=1.13
0.00681

Both the highest and lowest values are computed to be non-outliers at 95%
confidence level
Concentration of Stock Sample Cu(II) in ppm

0.092=0.000199 x +0.0197
x=363 ppm Cu ( II ) for Trial 1
Average ppm of Cu(II)

Average ppm=

363+313+298
=325 ppm Cu(II )
3