Prepared by: Eng.

Elham Ghazi Al-Hamaideh

Beer’s law and spectrophotometric analysis

Abstract

The purpose of this experiment is to find concentration of unknown matter (solute) by

determining the light passing and by using DR- Absorption device to draw standard
linear equation. In this experiment, the extension coefficient of copper sulphate will
be measured. The main cause of error in this experiment comes from pipetting the
solutions.

Introduction

Some solutions absorb light at certain visible wavelengths more than others. This

property gives the solution a colour which can be observed by the eye. Copper

sulphate is blue in solution, since the Cu2+ ions absorb the red (longer wavelength)

light which pass through. The amount of light absorbed is proportional to the

concentration of the solution, and the path length through which the light passes. This

is shown by the Lambert-Beer law:

𝐴 = 𝜀 ∗ 𝑐 ∗ 𝑙

where A is the absorbance, c the concentration (in molar), l the length in cm, and ε is

a constant, known as the molar extinction coefficient. In order to measure absorbance,

a spectrophotometer is used. The spectrophotometer can be set to a precise light

wavelength, for example 600 nm for red light, and zeroed using a solution which does

not absorb at the wavelength of interest, for example water. To measure the

absorbance of a coloured solution, the solution is first transferred into a cuvette in

which the path length (l) is known precisely. The cuvettes we used were 1 cm in

length. Plastic cuvettes are OK when measuring using visible light, but for measuring

at UV wavelengths, special quartz glass cuvettes are required. The extinction

Prepared by: Eng. Elham Ghazi Al-Hamaideh

coefficient for a certain solution can be determined as follows. Using a solution of

known concentration, a series of known dilutions is made and the absorbance

measured. A plot of absorbance vs. concentration (in molar) gives a straight line

whose equation is given by the Lambert-Beer law. The slope of the line is the molar

extinction coefficient. The reason for taking many measurements rather than just one

is to reduce the possible errors in dilution and measurement.

Methods

The experiment is performed according to the instructions given in the lab

instructions. From the 0.4 M stock solution we made 10 dilutions by taking 1 ml, 2
ml, 3 ml, ..., 10ml and then making the total volume up to 10 ml using distillation
water. Different volumes of pipette are used in this experiment. The absorbance of
each solution is measured in a plastic cuvette with 1 cm path length using a
spectrophotometer (DR- Absorption device) at a wavelength of 600 nm. The
spectrophotometer is first zeroed at this wavelength using a water blank.
For preparing 0.4M of CuSO4, use the following equation:
𝑚
# 𝑜𝑓 𝑚𝑜𝑙𝑒𝑠 = =𝑀×𝑉
𝑚𝑤

Materials
 CuSO4 as solid matter
 Distilled water
 Bottles
 Pipettes and pipette wheel
 Cuvette
 Spectrophotometer
 Beakers
 Test tubes
 Magnetic stirrer
 Glass stirrer
 Analytical balance
Prepared by: Eng. Elham Ghazi Al-Hamaideh

Data sheet

Date Tasted:

Tested by:

Group Number:

Sample Number:

Sample Description:

 Table

V of CuSO4 V of
Tube # V of H2O (ml) C A
(ml) solution
1
2
3
4
5
6

 Sample calculations

- Mass of CuSO4 to prepare 0.4M in 25 ml

𝑔 𝑔 𝑔
 (molar mass for atoms are Cu (63.5 𝑚𝑜𝑙) , O (16 𝑚𝑜𝑙 ), S (32 𝑚𝑜𝑙)

- Concentrations ( 𝑀1 𝑉1 = 𝑀2 𝑉2)
Tube #1:
Tube #2:
Tube #3:
Tube #4:
Tube #5
Tube #6:
Prepared by: Eng. Elham Ghazi Al-Hamaideh

 Plot concentration vs. absorption

 Find beer's law equation (using standard curve of copper sulphate)

 Find concentration of copper in unknown sample of water (in mol/L)

Prepared by: Eng. Elham Ghazi Al-Hamaideh

 Questions

1. What is the definition of Beer's law? What is the importance of Beer's Law?

2. What is the relationship between absorbance (A) and transmittance (T)?

3. What are the main reasons of error in this experiment?

4. How does the percent of transmittance and absorbance of a solution vary

(differ) with (a) increasing concentration and (b) increasing path length?