EXPERIMENT 1 chm260

EXPERIMENT 1: The Visible Spectra of Soft drink
Objective:
1. To determine the origin of the colour of soft drink sample from its absorption
spectrum.
2. To determine the wavelength at maximum absorbance (ƛmax) for a soft drink sample.
3. To determine the unknown concentration of a soft drink from the calibration curve.
Introduction:
Spectroscopy is the study of the relationship between electromagnetic radiation and matter,it
is one of the technique that are widely used nowadays,such as UV,X-rays, and infrared. The
word spectroscopy is used to refer to the broad area of science dealing with the absorption,
emission or scattering of electromagnetic radiation by molecules, ions, atoms or nuclei.The
interaction between light and matter are one of chemical analysis that are important towards
the instrumental method.
White light is visible light that contains all the colors of the rainbow. If white light hits a
colored solution,the electrons in the solution will absorb some of that light, while the rest will
be transmitted through the solution. It is the transmitted light that we see with our eyes. It’s
important to understand that a red solution does not absorb red light; it actually absorbs
blue-green light. The solution appears red because of the intensity of the blue-green light in
the original white light has been decreased. The following table show the relationship
between the color absorbed by a solution, and the color we observe.
Table 1.1: Relationship between absorbed and observed colour
Absorbed wavelength in nm(Absorbed Observed colour

colour)
400(violet) Greenish yellow
450(blue) Yellow
490(blue-green) Red
570(yellow-green) Violet
580(yellow) Dark blue
600(orange) Blue
650(red) Green
One of the most convenient, accurate and sensitive methods for measuring the
concentration of dilute solutions is by colorimetry or absorption spectrophotometry. The
techniques is based upon the measurement of the amount of light energy a solution absorbs
from a beam of light of a certain wavelength. The wavelength chosen is usually that one at
which the absorbance, of the species to be analysed, is at maximum.
The Spectronic 20 instrument you will use can correctly be called a colorimeter, because it
measures the absorption of light in the visible spectrum that we perceive as color, and the
technique used is said to be colorimetric. Although the instruments that are used to measure
the interaction of various regions of electromagnetic radiation with matter differ a great deal
in design and operation, they all contain the same basic components. A schematic diagram
of a simple instrument that is used to measure the absorption of visible light is shown in
figure1.1.
Figure 1.1: Main components of an absorption spectrophotometer
All the parts of the instrument work together as follows (Figure 1.1): Light from the source
passed through the monochromator producing a beam with a single energy or a narrow
band of energies. The intensity of this beam, Io is measured by the detector. The sample is
then placed in the beam between the monochromator and the detector. If some of the light is
absorbed by the sample, the intensity of the beam reaching the detector, I, will be less than
Io. The detector compares the two intensities and reports the result as either percent
transmittance(T%) or absorbance(A). These terms are defined to be
%T = I / Io x 100 (Equation 1)
(The fraction of Io that gets through the samples is called Transmittance)
A = -log T = -log(I/I0) = 2 -log(%T) (Equation 2)
The amount of light absorbed is dependent on how well the substance absorb light,the path
length of the light, and the concentration. These parameters are combined into a
mathematical relationship known as follows:
A = ƹbC (Equation 3)
In equation 3, ƹ (epsilon) is the molar absorptivity,b is the path length, and c is the
concentration. The molar absorptivity is a characteristic of the absorbing species; it tells us
how much light absorbed at a particular wavelength. Since the units on c are typically moles
per liter (M) and the path length is usually given in centimeters,the units on ƹ are most
commonly expressed as M-1 cm-1 to make A dimensionless. Equation 3 is known as the Beer-
Lambert Law, or simply Beer’s Law. It is important because it shows that absorbance is
directly proportional to the concentration of the absorbing species.
However, if the concentration is described in units other than moles per liter, then ƹ is
replaced with the symbol a or absorptivity, and Beer’s Law is written as follows:
A = abc (Equation 4)
The picture below is a representation of a spectronic 20-D colorimeter which is the

instrument you will be using in this experiment. Use of the instrument is easy and results are
excellent provided you follow all directions carefully
Figure 1.2 : Diagram showing the components of a spectronic 20 spectrophotometer
Two types of analysis can be done with the visible absorption measurements:
1. Qualitative analysis: To determine the wavelengths of maximum absorbance from the

absorption spectrum as well as absorptivity value of a specific molecule.
2. Quantitative analysis: To determine the concentration of an unknown solution from
the standard calibration curve.
Although,there a limitations to the applicability of Beer’s Law.This is because of the

deviation occurs by certain factors.Deviations are frequently observed from the direct
proportionality between absorbance,A and concentration,c when pathlength,b is
constant.it may be due to:
-Real deviations
Fundamental deviations due to the limitations of the law itself
-Instrument deviation
Due to polychromatic radiation:light exhibit more than one colour,where it contains

radiation of more than one wavelength.
Due to presence of stray radiation:define as radiation from the instrument that is

outside the nominal wavelength band selected.
Mismatched cells:The cells hold the analyte and the blank solution are having
different path-length.
-Chemical deviation
Occur when the analyte undergo dissociation,association or reaction with the

solvent,let the products that absorb differently than the analyte.
Apparatus:
Beaker,Pipette,Glass rod,Volumetric flask 50mL
Chemical:
Soft drink,(orange colour),Distilled water

Procedure
Fundamentals of spectroscopy was introduced to us in the first part of this experiment.
Instrument was properly learned by us and by that we were able to find the wavelength ( ƛmax
) at which absorbance of light by a Soft Drink solution that has a maximum value (ƛmax ). The
absorption spectrum was obtained by measuring the absorbance the absorbance of a
sample of the Soft Drink at different wavelengths using spectrophotometer. A plot of
absorbance vs wavelength was shown.
The law of spectroscopy, termed Beer-lambert’s Law or also known as Beer’s Law was
learned in the second part where we able to determine the concentration of an unknown
solution. A calibration curve was prepared by measuring the absorbance of different dilutions
of Soft Drinks at a single wavelength using ƛmax. The absorbance of the unknown was then
measured at the same wavelength and the calibration curve was compared to determine it’s
concentration.
A) Preparation of ‘Standard’ Solutions of Soft Drink ( Known Concentrations)
1) Soft drink was poured into a beaker and was stirred to remove the carbonation
2) Soft drink was pipette 5.00 mL into 50.0 mL of volumetric flask and then was diluted
to calibration mark with distilled water. Then it was covered and shake to make a
homogenous solution. It was then stored into a small beaker.
3) Step was repeated from step 2 by using 10 mL, 15 mL, 20 mL and 25 mL of soft
drink.
B) Operation of the Spectronic 20 and Determination of ƛmax
Operating Instruction
1) Spectronic was turned on and was given 15 minutes(minimum) to warm up.
2) Wavelength was set to 600 nm
3) 0% transmittance (% T) was adjusted. ( Adjusting dark current – nothing was in the
sample compartment)
4) Cuvette was obtained and by that it was cleaned and rinsed with distilled water and
then the tube was filled about ¾ full ‘blank’ solution (Distiled water). The surface of
the cuvette was wiped to remove any fingerprint and solution by using tissue.
5) 0 absorbance and 100% absorbance were adjusted with the cuvette that contained
the ‘blank’ in the sample holder. Cuvette was then removed and set aside without
emptying the distilled water.
6) Another cuvette was cleaned and rinsed with small amount of ‘standard’ soft drink
whose absorbance was to be measured. Tube was filled with ¾ of solution and was
wiped with tissue to remove any fingerprints and other solution before it was placed
in the sample holder with hash mark aligned. Absorbance was recorded.
7) Cuvette was remove and the top was closed and wavelength was set 20 nm lower.
8) 0% transmittance was reset for it had changed.
9) Cuvette was insert and 100% T was reset. Cuvette removed.
10) Cuvette that contained soft drink solution was insert as in step 6.
11) Absorbance was read and recorded at table 1.2.
12) Steps 8 through 11 was repeated until 360 nm and absorbance was taken at each 20
nm interval.
13) Graph paper was used to plot the absorption spectrum of our soft drink and ƛmax was
determined.
C) Preparation of ‘Unknown’ Soft Drink Sample
1) Some soft drink was poured into a beaker and was stirred to remove any
carbonation.
2) Soft drink was poured into 50 mL volumetric flask without measured the volume and
was diluted to mark with distilled water. By using stopper, flask was shake to make
the solution homogenize. Unknown solution was recorded where it is not darker than
the most concentrated standard solution.
3) ‘Unknown sample’ was placed into a cuvette until it filled ¾ of it.
D) Quantitative Analysis Of The Soft Drink Solution.
1) Spectronic 20 was set at wavelength maximum (ƛmax ) that was obtained at Part B
2) 0 and 100% T was set at given procedure above.
3) Absorbance of each 5 ‘standard’ soft drink solution was measureed and recorded.
4) Absorbance of ‘unknown solution’ was measured and recorded.
E) Cleaning Up
1) Since only soft drink was used in the experiment, we can dispose the waste down the
drain.
2) Cuvette was cleaned and was dried
DATASHEET EXPERIMENT 1
THE VISIBLE SPECTRA OF SOFT DRINKS
Table 1.2: The wavelength and absorbance of soft drink
Wavelength (nm) Absorbance

600 0.013
580 0.049
560 0.064
540 0.091
520 0.242
500 0.337
480 0.358
460 0.299
440 0.234
420 0.212
400 0.206
380 0.186
360 0.159
ƛmax = 480 nm
Table 1.3: Table of soft drink concentration (volume %) and absorbance
Solutions Concentration(volume %) Absorbance

1 5/50 x 100 = 10% 0.358
2 10/50 x 100 = 20% 0.713
3 15/50 x 100 = 30% 1.057
4 20/50 x 100 = 40% 1.406
5 25/50 x 100 = 50% 1.753
Unknown sample 0.528
Sample calculation for the concentration of solution
Y = mx – C
Concentration of unknown:
Lecturer’s Signature,
_____________________
Question
1. Based on Beer’s Law,when the concentration of an analyte increases, how will

the following be affected(increase,decrease,no change)?
i. Absorbance:When concentration increase,absorbance also increase

ii. Transmittance:when concentration increase,the transmittance decrease
2. Why is it important to first obtain the absorption spectrum of the soft drink
before making a calibration curve?
This is because we need to plot a graph of absorbance against concentration.It is
use to determine the concentration of the soft drink.it can be determine by cross the
calibration line of the graph,from the equation y=mx+c
3. What is the purpose of using the ‘blank’ solution?

It doesn’t contain any analyte,and is used to calibrate instrument.for
example,colorimeter is used to measure the absorbance of light.
4. What colour(s) is/are being absorbed in your soft drink?

Blue.

EXPERIMENT 1 chm260

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What is the relationship between concentration, absorbance and transmittance according to Beer's Law?

What is the relationship between concentration, absorbance and transmittance according to Beer's Law?

Why is it important to obtain the absorption spectrum of the soft drink before making a calibration curve?

Why is it important to obtain the absorption spectrum of the soft drink before making a calibration curve?

EXPERIMENT 1: The Visible Spectra of Soft drink

Absorbed wavelength in nm(Absorbed Observed colour

Figure 1.1: Main components of an absorption spectrophotometer

(The fraction of Io that gets through the samples is called Transmittance)

A = -log T = -log(I/I0) = 2 -log(%T) (Equation 2)

The picture below is a representation of a spectronic 20-D colorimeter which is the

1. Qualitative analysis: To determine the wavelengths of maximum absorbance from the

Although,there a limitations to the applicability of Beer’s Law.This is because of the

Fundamental deviations due to the limitations of the law itself

Due to polychromatic radiation:light exhibit more than one colour,where it contains

Due to presence of stray radiation:define as radiation from the instrument that is

Occur when the analyte undergo dissociation,association or reaction with the

Beaker,Pipette,Glass rod,Volumetric flask 50mL

Soft drink,(orange colour),Distilled water

THE VISIBLE SPECTRA OF SOFT DRINKS

Table 1.2: The wavelength and absorbance of soft drink

Wavelength (nm) Absorbance

Solutions Concentration(volume %) Absorbance

Sample calculation for the concentration of solution

1. Based on Beer’s Law,when the concentration of an analyte increases, how will

i. Absorbance:When concentration increase,absorbance also increase

3. What is the purpose of using the ‘blank’ solution?

4. What colour(s) is/are being absorbed in your soft drink?

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