Objectives:

To study and understand the principle of the BeerLambert law.

Introduction
Beer-Lambert Law is a common law which is used to determine the concentration of a sample by
measuring the degree of the absorption of radiation of that particular sample and comparing it to
standard curve constructed with a set sample with known concentration. Samples can include
chemical and biological solution. The basic principle of Beer-Lambert Law is a combination of
Lamberts law and Beers law. In Beers law, the absorption of light is proportional to the
concentration of sample. Beers law was combined with Lamberts law which state that a
transparent medium absorbed a fraction of incident radiation is independence to the intensity of
the incident radiation and every continuous layer of the medium will absorb equal fraction of
incident radiation. The ratio of transmitted intensity and incident radiation intensity is expressed
by formula below.
T = I/I0
I0 is incident radiation intensity while I is transmitted intensity. The number of the photons
interacted in unit time is equal to the number of intensity. A total transparent substance will have
no absorbed radiation while a zero reading of transmittance means the sample is opaque.
Absorbance (A) and extinction (E) can be

determined by using the logarithm of the

transmittance.

c is molar concentration of the absorbing

species while l is the thickness of the

absorbing layer and is molar

absorptivity. The formula below states

that the absorbance is proportional to the

concentration of samples, the diameter

of transmission and also the molar absorbance coefficient.

Beer-Lambert Law although useful, but it has its limitation which includes instrumental factors
and chemical factors. For example, reading might be caused by stray light from the source, nonmonochromatic radiation, particulate which scatters the light, phosphorescence of the sample and
equilibriation which occurs in the sample, molecular dissociation of the solute and ionization of
the solute. This limitation will cause the intensity of the radiation to reach the detector in a
deviated way from the real intensity of the radiation transmitted by the sample.

Results
1. Graphical determination of unknown benzenes concentration
Concentration of benzene in stock solution:
Molar weight of benzene = 78.11 g/mol
mol of benzene=

0.5 g
=0.006 mol
78.11 g/mol

Concentration of benzenefirst dilution=

0.006 mol
=0.24 mol/ L
0.025 L

C1V1=C2V2
(0.24mol/L)(0.005L) = C2 (0.025L)
C2 = 0.0048mol/L
*Concentration of benzene in stock solution = 0.0048 mol/L = 0.0048 mol dm-3
Standard

Volume of

Volume of

Concentration

solution

stock

diluent (dm3)

(mol/dm3)

0.002

0.0038

Absorbance

solution
1

added (dm3)
0.008

1.300

2
3
4
5

0.006
0.004
0.002
0.001

0.004
0.006
0.008
0.009

0.0028
0.0019
0.0009
0.0004
Unknown
Table 1: Benzene concentration in each standard solution.

1.050
0.650
0.320
0.175
0.200

The calibration curve of the standard solution was constructed and the unknown sample
concentration was determined using the curve below.
1.4

f(x) = 342.76x + 0.03

1.2
1
0.8

Absorbance

0.6
0.4
0.2
0

concentration (mol/dm3)

Unknown sample concentration :

Discussion
Beer-Lambert law is used as a measure of the absorption in this experiment.
A = lc
The equation tells us that absorbance depends on the total quantity of the absorbing compound in
the light path through the cuvette. A set of data was plotted in order to generate the straight line
that passed through the origin (0, 0) as shown in Graph 1. The gradient of the straight line
generated should be 0.99 as shown in Graph 1.
Benzene is a hydrocarbon which cannot dissolve in water but dissolve in organic solvent. Thus,
cyclohexane is used as the solvent to dissolve the benzene in dilution. Benzene has wavelength
in the range 230 to 270 nm while the cyclohexane cannot be detect in this range. Benzene acted
as the stock solution and the decrement of the volume of stock solution, the absorbance value
obtained decreased due to decreasing concentration in the standard solution. The concentration of
the unknown solution can be obtained from the graph by substituting the x value from the
equation obtained with the absorbance of the unknown sample. The concentration of the
unknown is 0.0005 mol/dm3.
The reaction between ferric sulphate solution (Fe3+) and potassium thiocyanate (SCN-) solution
involves the formation of thiocyanatoiron (III) complex which is in red colour. The half equation
of the reaction is shown below:

The formation of the iron complex was carried under two distinct conditions. First, medium used
for the solvation of ferric sulphate solution and potassium thiocyanate solution, namely water
and acetone. Next, absorbance of the iron complex in both water and acetone was measured after
exposure to sunlight at 5 minutes intervals for a total of 30 minutes.
The initial absorbance reading before exposure to sunlight and the absorbance readings
corresponded to each time intervals after exposure to the sunlight were tabulated (Table 2). A
distinct trend observed is that the value of absorbance readings declined over time for both water

and acetone medium (Figure 2). In other words, the longer the solution mixture exposure to
sunlight, the lower the absorbance reading and the intensity of the coloured complex also fades
with time. However, there was a variation in the absorbance readings, whereby the solution
mixture in acetone medium had a higher absorbance values compared to solution mixture in
water medium. This observation also was applied to the initial absorbance reading before
exposure to sunlight.
Water and acetone have different effect on ferric sulphate and potassium thiocyanate solution
mixture. The former hinders the formation of thiocyanatoiron (III) complex while the latter
promotes the complex formation. The water introduces a complication because of its reaction
with Fe3+ ion to form iron hydroxide, which is insoluble in water:
Fe3+ (aq) + 3 H2O (l) <==> Fe (OH) 3 (s) + 3 H+ (aq)
As a result, precipitation of iron (III) hydroxide is more favoured than the formation of
thiocyanatoiron (III) complex which absorbs light at = 447 nm (Z. & M. , 2000). On the
contrary, the presence of organic solvents like acetone normally favours the formation of
complexes due to their well-known solvation effects (Martins et al., 2005). The other advantage
of acetone in this system is the fact that it acts as auxiliary oxidizing agent and complex
stabilizer allowing very sensitive spectrophotometric determinations (Martins et al., 2005). This
explains the higher absorbance reading account for solution mixture in acetone which is 2X
higher compared to solution mixture in water regardless of exposure to the sunlight.
Another trend noticed in this experiment was, the absorbance readings of both solution mixture
in water and acetone medium before sunlight exposure was higher compared to after exposure to
sunlight. One possible explanation is exposure to UV light will cause photodecomposition of
some metal cyanide complexes and reduce their concentration (A.Meech, n.d.). Iron complexes
will be decomposed if sunlight is sufficient (B.M.Tissue, 2005). The absorbance reading after
exposure to sunlight is 3X lower than the initial absorbance reading before exposed to sunlight.
However, the solution mixture in acetone still acquired higher absorbance readings compared
solution mixture in water. This is due to the one of the properties of acetone in which acetone
suppresses the dissociation of iron complex (R. M. & I. W. , 2013). The absorbance difference
between water and acetone after exposure to sunlight was high, approximately 9 %.

Photochemical reactions or photodecomposition causes derivations in Beer-Lamberts law

References:
A.Meech, J., n.d. Cyanide Destruction Methods. [Online] Available at:
http://www.belgeler.com/blg/2ng4/chemistry-and-treatment-of-cyanidation-wastes-by-terry-imudder# [Accessed 8 March 2015].
B.M.Tissue, 2005. Introduction to Spectrophotometry. [Online] Available at:
http://www.chem.vt.edu/chem-ed/spec/uv-vis/singlebeam.html%20accessed%208/12/05
[Accessed 8 March 2015].
Martins, F.G. et al., 2005. Spectrophotometric study of iron oxidation in the
iron(II)/thiocyanate/acetone system and some analytical applications. Scientific Electronic
Library Online - SciELO, 30(3), pp.63-71.
Mudakavi, P.J.R., n.d. Modern Instrumental Methods of Analysis. [Online] Available at:
http://textofvideo.nptel.iitm.ac.in/103108100/lec7.pdf [Accessed 8 March 2015].
R. M., J. & I. W. , S., 2013. The Determination of Organic Peroxides: Monographs in Organic
Functional Group Analysis. Elsevier, 2013.
Z., & M. , B., 2000. Separation, Preconcentration and Spectrophotometry in Inorganic Analysis.
Elsevier, 2000.