EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

Bachelor of Chemical Engineering (Hons)

LAB REPORT

EKB 3171 ANALYTICAL TECHNIQUES AND

INSTRUMENTATIONS LAB

EXPERIMENT TITLE
NAME OF CANDIDATE
WITH REG NO.

ASSAY OF ASPIRIN BY UV VISIBLE

SPECTROPHOTOMETER
ZANDRA LAVANYA
1000338
22nd March 2016, Tuesday

SESSION DATE
DATE OF SUBMISSION

11th April 2016, Tuesday

NAMES OF GROUP MEMBERS

No.
1

REG No.
1000048

NAME
DENESH A/L A.MOHAN

1000337

YOGENDRAN A/L GANESAN

1000329

KARTHIKEYAN A/L SUBRAMANIAN

SIGNATURE

Page 1

ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

INCHARGE
(Ms.Preeti Lokesh)

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

TABLE OF CONTENTS

S. NO.

DESCRIPTION
Abstract

PAGE NO.
3

Introduction

3-5

Materials and Methods

5-6

Results and Discussions

6-8

Conclusions and Recommendations

References

Appendix

10-12

Page 2

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

ASSAY OF ASPIRIN BY UV VISIBLE SPECTROPHOTOMETER

ABSTRACT
To understand the ultraviolet-visible spectroscopy technique and its importance in qualitative
and quantitative analysis. . Standard aspirin was used determine absorbance of standard aspirin
which was diluted in HCl and 5mg of potassium dichromate dissolved in 100ml of 0.005H 2SO4
was used as the blank solution. 5g sample aspirin that was prepared was in 10ml of ethanol and the
absorbance value of the sample aspirin was obtained The standard aspirins were prepared from 1ml
to 8ml. The concentration of the aspirin was being determined for the absorbance of 1.2678 which
was 0.0681mol/dm3 . The percentage of aspirin which was determined was 24.5%. In order to get
better results, ensure all the equipment used are being sterilised in order to avoid the presence of
impurities.

1. INTRODUCTION
Ultravioletvisible

spectroscopy or ultraviolet-visible

spectrophotometry refers

to spectroscopy or reflectance spectroscopy in the ultraviolet-visible spectral region. This means it

Page 3

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

uses light in the visible and adjacent (near-UV and near-infrared [NIR]) ranges. The absorption or
reflectance in the visible range directly affects the perceived colour of the chemicals involved. In
this region of the electromagnetic spectrum, molecules undergo electronic transitions. This
technique is complementary to fluorescence spectroscopy, in that fluorescence deals with transitions
from the excited state to the ground state, while absorption measures transitions from the ground
state to the excited state. [1]
An obvious difference between certain compounds is their colour. Thus, quinone is yellow,
chlorophyll is green, the 2,4-dinitrophenylhydrazone derivatives of aldehydes and ketones range in
colour from bright yellow to deep red depending on double bond conjugation and aspirin is
colourless.. The component colours of the visible portion can be separated by passing sunlight
through a prism, which acts to bend the light in differing degrees according to
wavelength. Wavelength is defined as the distance between adjacent peaks or troughs, and may be
designated in meters, centimeters or nanometers (10-9 meters). Frequency is the number of wave
cycles that travel past a fixed point per unit of time, and is usually given in cycles per second, or
hertz (Hz). Visible wavelengths cover a range from approximately 400 to 800 nm. The longest
visible wavelength is red and the shortest is violet. [2]
When a beam of electromagnetic radiation is passed through the sample solution, some of it is
absorbed. UV-VIS spectrophotometer is an instrument used to measure the intensity of light passing
through the sample solution (I), and compares it to the intensity of light before it passes through the
sample solution (Io).
The ratio I / Io is called the transmittance T:
T = I / Io

(1)

The absorbance, A, is the logarithm for the reciprocal of T:

A = log(1/T)

(2)

Page 4

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

The Beer-Lambert law gives an empirical relationship that relates with the absorption of
light of an analyte and which is measured quantitatively and states that the absorbance of a solution is
directly proportional to the concentration (c) of the analyte in solution and the path length of the
solution (l) at a certain wavelength. Thus absorption spectroscopy can be used to determine the
concentration of a solution by following formula:

A = klc

(3)

Where k is a constant, l is the path length in cm, c is the concentration (mole/liter or gm/100 ml)
Note:
If the concentration of the solution is in moles/liter, the constant k becomes the molar absorptivity ().
In analytical chemistry, the concentration is expressed in gm/100ml (%) and thus the constant k
becomes A (1%, 1 cm) the specific absorbance (Extinction coefficient).Then the Beer-Lambert Law
can be expressed as:
A = A (1%, 1 cm) x b x c

(4)

Where A is the absorbance, b the path length of the cell (in cm), c the concentration in gm/100
ml and A1%1cm is the specific absorbance(extinction coefficient).A1%1cm values, individual
characteristics of a compound and express the absorbance reading obtained from a 1 M or a 1%
solution given a path length of 1 cm.
The Beer-Lambert law can be used to assay the drug content of a solution. If and A 1%1cm are
known, the concentration (in % or mol / l) can be calculated as

C = A / [A (1%, 1cm) x b ] or c = A / ( x b)

Page 5

(5)

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

If or A1%1cm is unknown, they must be determined first by analyzing a standard solution of the
drug with a known concentration.

= A / (b x c) or A(1%, 1cm) = A / (b x c) (6)

or A (1%, 1cm) is calculated from a set of several standards with known concentrations to
establish a calibration curve. Then the values of and A (1%, 1cm) can then be calculated and the
average value may be obtained. Alternatively, the values of and A (1%, 1cm) can be estimated
graphically as the slope of the calibration curve in a standard plot of concentration(C) on X-axis and
absorbance (A) on Y-axis.

2. MATERIALS AND METHODS

2.1 MATERIALS
0.005M H2SO4, potassium dichromate, 0.1M HCl, Aspirin(pure sample), Aspirin tablets
2.2 APPARATUS
Ultraviolet spectrophotometer, Volumetric flask, 250 ml, (1), volumetric flask, 100
ml (5), volumetric flask, 10 ml (12), Pipette and bulb,1 ml (1) , pipette, graduated, 10 ml
(1), Pipette graduated 1ml, (2); Beaker or conical flask, 500 ml (1),Test-tube 20 , 1 testtube rack

2.3 EXPERIMENTAL PROCEDURE

2.3.1 Determination of aspirin concentration in an unknown samples

Page 6

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

Firstly a sample of 5 ml was being chosen. The sample was then diluted using 6.3g of
K2Cr2O7 in 100ml of 0.005 H2SO4 and the blank was taken. There were 8ml of standard aspirin
( 1ml to 8 ml) therefore it was all diluted with 100 ml of 0.1M HCl in order to determine the
absorbance range. The absorbance were then determined for all the 8 samples. The absorbance
value for the 5 ml prepared aspirin was determined and the concentration of the aspirin was
determined by interpreting from the graph. Graph of absorbance against the concentration of
aspiring was plotted.

3. RESULTS AND DISCUSSION

A frequent analytic application of UV-vis spectroscopy is the precise determination of
concentration. This experiment is conducted to ensure the understanding of the ultraviolet-visible
spectroscopy technique and its importance in qualitative and quantitative analysis.Quantitative
analysis is simply a way of measuring things while qualitative analysis deals with the identification
of elements or grouping of elements present in a sample. The techniques employed in qualitative
analysis vary in complexity, depending on the nature of the sample.
A total of 8ml of standard aspirin was prepared, (1ml to 8ml). 0.75g of standard aspirin was
then dissolved in 100 ml of 0.1M HCL, and the standard aspirin prepared (1ml to 8ml) of the
solution was then diluted in 100ml of HCL in order to determine the concentration of standard
aspirin for each of the solution. The blank space used in the UV spectrometer was when 6.3mg of
K2Cr2O7 was diluted in 0.005M. The absorbance was determined from the UV spectrometer for the
solution (1ml to 8ml). The sample aspirin, 0.05g which was prepared was then dissolved in 100cm3
of 0.1HCl and was put into UV spectrometer and the absorbance value was recorded for all. Besides
that, the wavelength used was 257 throughout, initially the wavelength was changed to 300, but
there were no proper readings being indicated, therefore the wavelength was changed back. Below

Page 7

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

in table 1 is the absorbance and the concentration which was obtained for the dilution of the sample
aspirin from 1ml to 8ml the sample aspirin.
Table 1 : Absorbance and Concentration for 1ml to 8ml l of dilution for standard aspirin and
sample aspirin
Number

Wavelength

Temperature

B
1ml
B
2ml
B
3ml
B
4ml
B
5ml
B
6ml
B
7ml
B
8ml
B
Sample

(nm)
257
257
257
257
257
257
257
257
257
257
257
257
257
257
257
257
257
257

(oC)
32.2
32.2
32.1
32.1
32.2
32.2
32.2
32.2
32.2
32.2
30.4
30.4
30.7
30.7
30.9
30.9
31.3
31.3

%T
99.92
85.74
100.1
69.11
99.98
56.29
99.98
46.47
100.05
33.25
100.02
48.25
99.94
27.72
99.98
21.72
100.09
5.39

aspirin

ABS(AU)
0
0.067
0
0.16
0
0.25
0
0.333
0
0.478
0
0.316
0
0.557
0
0.663
0
1.268

Concentration
(mol/dm3)
0
0.000416
0
0.000832
0
0.001248
0
0.001664
0
0.002080
0
0.002496
0
0.002912
0
0.003328
0
Unknown

Co-relation with the table above, a graph of absorbance against the concentration of aspirin
which was obtained from the UV spectrometer was plotted. Below in figure 1, is the graph which
was being plotted.

Page 8

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

0.7
0.6

f(x) = 186.01x + 0
R = 0.88

0.5
0.4
Absorbance

0.3
0.2
0.1
0

Concentration of aspirin mol/DM3

Figure
1 : Absorbance against the concentration of aspirin which was obtained from the UV
spectrometer

Based on figure 1 above, it indicates that the rate of absorbance tends to increase as the
concentration of the aspirin increases. This is because, as the concentration of aspirin tends to increase,
it will absorb more of the UV light, therefore the absorbance also increases co relation with the
concentration of the aspirin. Based on the equation above, the linear equation was taken and then the
concentration of the aspirin was being determined for the absorbance of 1.2678 which was
0.0681mol/dm3 . The percentage of aspirin which was determined was 24.5%, this could be due to the
impurities present while conducting the experiment.
The blank solution was formed by 5mg potassium dichromate in 100ml of 0.005M H 2SO4 while
the standard aspirin absorbance was determined together with the concentrations.

Page 9

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

4. CONCLUSIONS AND RECOMMENDATIONS

In this experiment conducted it is to understand the ultraviolet-visible spectroscopy technique
and its importance in qualitative and quantitative analysis. The standard aspirins were prepared from
1ml to 8ml. The concentration of the aspirin was determined by the absorbance and also the standard
aspirin. Based on the linear equation was taken and then the concentration of the aspirin was being
determined for the absorbance of 1.2678 which was 0.0681mol/dm3 . The percentage of aspirin
which was determined was 24.5%. In order to get a better result, when carrying out the experiment,
ensure to have all the equipment used fully sterilised in order to prevent impurities.

5. REFERENCES
1. Lynch, David K.; Livingston, William Charles (2001). Color and Light in Nature (2nd ed.).
Cambridge, UK: Cambridge University Press. p. 231. ISBN 978-0-521-77504-5.
2. Open Access Journal, Dr. Nagham Mahmood Aljamali (January 2015), International Journal of
Medical Research and Pharmaceutical Sciences (ISSN: XXX-XXX) Volume 2(Issue 1)
3. M. Hesse, H. Meier, B. Zeh, Spektroskopische Methoden in der organischen Chemie, 3. Aufl.,
Thieme Verlag, Stuttgart (1987)

Page
10

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

APPENDIX
Concentration
Standard aspirin concentration of = 0.75g in 100ml of 0.1 HCl
ml
d m3
mass of sample aspirin (g)
mol

3
g
molar mass of aspirin (
) 100 ( volume of HCl ( ml ) ) d m
mol
1000

Standard aspirin concentration =

0.75 1000 mol

180
100 d m3

( )

= 0.0416mol/dm3
Dilution
For 1 ml of standard aspirin
V1 = 1ml
M1 = 0.0416mol/dm3
M2 = x(unknown)
V2 = 100ml
M1V1 = M2V2
0.0416(1) = x(100)
x = 0.000416mol/dm3

The step above in continued using the different ml of standard aspirin

Page
11

( )

( )

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

Table 1 : Absorbance and Concentration for 1ml to 8ml l of dilution for standard aspirin and
sample aspirin
Num

Wavelength(nm)

Temperature(oC)

%T

257

32.2

99.92

1ml

257

32.2

85.74

0.067

0.000416

257

32.1

100.1

2ml

257

32.1

69.11

0.16

0.000832

257

32.2

99.98

3ml

257

32.2

56.29

0.25

0.001248

257

32.2

99.98

4ml

257

32.2

46.47

0.333

0.001664

257

32.2

100.05

5ml

257

32.2

33.25

0.478

0.002080

257

30.4

100.02

Page
12

ABS(AU) Concentration(mol/dm3)

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

6ml

257

30.4

48.25

0.316

0.002496

257

30.7

99.94

7ml

257

30.7

27.72

0.557

0.002912

257

30.9

99.98

8ml

257

30.9

21.72

0.663

0.003328

Absorbance of sample aspirin prepared: 1.268

Based on the linear equation (tabulated on the graph), Concentration of sample aspirin = 0.00681

Concentration of sample aspirin=

1000

( dmlm )
3

mass of sample aspirin ( g)

mol

3
g
molar mass of aspirin (
) 10 ( volume of ethanol ( ml ) ) d m
mol

( )
0.00681

1000

( dmlm )
3

mol
d m3

mass of sample aspirin ( g)

mol

3
g
( )
molar mass of aspirin (
) 10 (volume of ethanol ml ) d m
mol
0.00681 =

( )

ml
3
dm
mass of sample aspirin
mol

180 g /mol
10( volume of ethanol ( ml )) d m3
1000

( )

Mass of sample aspirin = 0.012258 g in 10cm3 of ethanol

Compared with 0.05g in 10cm3 of ethanol to get the percentage of aspirin

Page
13

( )

EKB 3171 ANALYTICAL TECHNIQUES AND INSTRUMENTATIONS LAB

Percentage of aspirin =

0.012258 g
100 =24. 5
0.05 g

Page
14