BE184P Exercise 1.1a Spectrophotometry

BE184P Instrumentation for Biological Engineering 2
Exercise 1.1a UV-visible absorption spectroscopy and IR spectroscopy

Instructions:
• Open the links to each simulations.

• Perform the required virtual experiment simulations.
• Gather the data generated (if any) and save a photo or file.
• After performing all the required simulations, answer the guide questions.
Simulation 1: Spectrophotometry
Go to https://vlab.amrita.edu/?sub=2&brch=190&sim=338&cnt=2.
Sign up if you don’t have an account yet.
Instructions:
A. For each known sample, determine at which wavelength (from 350nm to 700nm) will each sample
maximally absorb. Do this by getting the absorbance at each wavelength and then plotting it in a graph.
Do this for all the samples. Place your data in the report form.
B. Determine the molar absorptivity constant for each sample. Using another sample, get its absorbance
and using the data of the known sample compute for the unknown concentration of this other sample.
Place all your answers on the table provided in the report form.
Simulation 2: Instrumentation and working principles of IR Spectroscopy

http://ccnsb06-
iiith.vlabs.ac.in/exp6/Introduction.html?domain=%20Chemical%20Sciences&lab=Physical%20Chemistry
%20Lab
http://ccnsb06-
iiith.vlabs.ac.in/exp7/Introduction.html?domain=Chemical%20Sciences&lab=Physical%20Chemistry%20
Lab
Instructions:
For each spectra, identify wavenumber and the possible functional groups present in the sample
compound.
Name: Den Mark Celestra Student No. 2019102287 Program/Year: BE/3 Date: March 7, 2022
EXERCISE 1.1A
DATA REPORT FORM
SIMULATION 1: UV-VIS ABSORPTION SPECTROSCOPY
Data Gathered:
A. Wavelength at Maximum Absorbance at path length = 1.00cm
Sample Concentration Wavelength Maximum Absorbance

Cobalt chloride 0.4 (510 nm) 6.644
Hexaaquacobalt (II) ion 0.4 (510 nm) 4.048
Ferrocene 0.4 (445 nm) 12.0927
Crystal violet 0.4 (590 nm) 10.2759
Rose Bengal 0.4 (560 nm) 9.5286
Show the plot of each sample, x = wavelength and y = absorbance.
Cobalt Chloride
7
0
350 400 450 500 550 600 650 700 750
Hexaaquacobalt(II)Ion
4.5
4
3.5
3
2.5
2
1.5
1
0.5
0
350 400 450 500 550 600 650 700 750
Ferrocene
15
10
0
350 400 450 500 550 600 650 700 750
-5
-10
-15
-20
Crystal violet
15
10
0
350 400 450 500 550 600 650 700 750
-5
-10
-15
-20
Rose Bengal
15
10
0
350 400 450 500 550 600 650 700 750
-5
-10
-15
-20
Coumarin
15
10
0
350 400 450 500 550 600 650 700 750
-5
-10
-15
-20
B. Calculate the value of molar absorption coefficient of the known sample and the concentration of
each sample with respect to the known sample.
Sample Molar absorption Unknown Sample Concentration of

coefficient Absorbance Unknown Sample
Cobalt chloride 166.1 3.9366 0.0237
Hexa aquacobalt (II) 101.2 1.5079 0.0149
ion
Ferrocene 302.3175 13.8462 0.0458
Crystal violet 256.8975 5.6261 0.0219
Rose Bengal 238.215 4.2879 0.018
Coumarin 300.835 5.3850 0.0179
Provide solutions/computations in the determination of molar absorption coefficient and in the
determination of unknown concentration of sample.
Molar absorbance coefficient
Cobalt Chloride:
6.644
= 𝟏𝟔𝟔. 𝟏
0.04
Hexaaquacobalt (II) ion:
4.048
= 𝟏𝟎𝟏. 𝟐
0.04
Ferrocene:
12.0927
= 𝟑𝟎𝟐. 𝟑𝟏𝟕𝟓
0.04
Crystal violet:
10.2759
= 𝟐𝟓𝟔. 𝟖𝟗𝟕𝟓
0.04
Rose Bengal:
9.5286
= 𝟐𝟑𝟖. 𝟐𝟏𝟓
0.04
Coumarin:
12.0334
= 𝟑𝟎𝟎. 𝟖𝟑𝟓
0.04
Concentration of Unknown Sample
Cobalt Chloride:
3.9366
= 𝟎. 𝟎𝟐𝟑𝟕
166.1
Hexaaquacobalt (II) ion
1.5079
= 𝟎. 𝟎𝟏𝟒𝟗
101.2
Ferrounce
13.8462
= 𝟎. 𝟎𝟒𝟓𝟖
302.3175
Crystal Violet
5.6261
= 𝟎. 𝟎𝟐𝟏𝟗
256.8975
Rose Bengal
4.2879
= 𝟎. 𝟎𝟏𝟖
238.215
Coumarin
5.3850
= 𝟎. 𝟎𝟏𝟕𝟗
300.835
B. IR Spectroscopy
a. Asprin IR Spectra
Identify four (4) distinct IR absorption peaks of aspirin and assign its probable functional group
Wavenumber Functional Group

Absorption (cm-1)
3000 Phenol (O-H)
2700 Carboxylic Acid (OH)
1700-1800 Ester (C=O)
1100-1200 Anhydride (C-O)
b. Caffeine IR Spectra
Identify four (3) distinct IR absorption peaks of caffeine and assign its probable functional group

Absorption (cm-1)
3100 Carboxylic acid
1700 Carbonyl
c. IR spectra of 2-nitrophenol
Identify four (4) distinct IR absorption peaks of 2-nitrophenol and assign its probable functional
group

Absorption (cm-1)
1600 C=C
1300 N=O
d. IR Spectra of 2-methyl benzoic acid

Identify four (4) distinct IR absorption peaks of 2-methyl benzoic acid and assign its probable
functional group

Absorption (cm-1)
2400 Alkene
1600 Ester
800 Alkene

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BE184P Instrumentation for Biological Engineering 2

Exercise 1.1a UV-visible absorption spectroscopy and IR spectroscopy

• Open the links to each simulations.

Simulation 2: Instrumentation and working principles of IR Spectroscopy

SIMULATION 1: UV-VIS ABSORPTION SPECTROSCOPY

A. Wavelength at Maximum Absorbance at path length = 1.00cm

Sample Concentration Wavelength Maximum Absorbance

Sample Molar absorption Unknown Sample Concentration of

Molar absorbance coefficient

Wavenumber Functional Group

Wavenumber Functional Group

Wavenumber Functional Group

d. IR Spectra of 2-methyl benzoic acid

Wavenumber Functional Group

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