OPTICAL METHOD OF ANALYSIS
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Transcript of OPTICAL METHOD OF ANALYSIS
Reporter: John Kevin G. San Jose Instructor: Prof Virgilio Agbayani
Section: 2ChEC February 15, 2011
Group No.: 5
EXPERIMENT NO. 9
OPTICAL METHOD OF ANALYSIS
USE OF BEER’S LAW ON A KMnO4 SOLUTION
Abstract
Spectrophotometry is a method of chemical analysis based on the absorption or attenuation by matter of electromagnetic radiation of a specified wavelength or frequency.
1 Four solutions with different
concentrations were prepared. Absorbance of the four known concentration was measured and the gathered data was plotted in a graph in the ratio of absorbance with concentration. As measured from the spectrophotometer, the absorbance of the unknown solution is 0.198. Through this numbers and the plotted graph, the solution was found out to have a 7.66 X 10
-5 molar
concentration.
Introduction
A study of the interaction of light or other electromagnetic radiation with matter is an
important and versatile tool. Indeed, much of our knowledge of chemical substances comes
from their specific absorption or emission of light.
As the color of the solution deepens its concentration also increases. This is an
underlying principle of spectrophotometry. The intensity of color is a measure of the amount of
a material in solution. A second principle of spectrophotometry is that every substance absorbs
or transmits certain wavelengths of radiant energy but not other wavelengths. The light energy
absorbed or transmitted must match exactly the energy required to cause an electronic
transition (a movement of an electron from one quantum level to another) in the substance
under consideration. Only certain wavelength photons satisfy this energy condition. Thus, the
absorption or transmission of specific wavelengths is characteristic for a substance, and a
spectral analysis serves as a “fingerprint” of the compound.
In recent years, spectrophotometric methods have become the most frequently used
and important methods of quantitative analysis. They are applicable to many industrial and
clinical problems involving the quantitative determination of compounds that are colored or
that react to form a colored product. 2
An application of this is by the use of Beer-Lambert’s Law. The Beer-Lambert law (also
known as Beer's law) (as it applies to solutions of light-absorbing substances) states that the
absorbance is directly proportional to the path length, l of the sample and its concentration, c:
A = ecl
Where e is the MOLAR EXTINCTION COEFFICIENT (with dimensions of dm3.mol-1cm-1)
of the solute, c is the molar concentration (in moles.dm-3), and l, the path length, is measured in
centimeters.
The Beer-Lambert law is readily applicable to the determination of the concentration of
numerous substances, provided that (i) the molecular extinction coefficient e for the subtance is
known at the wavelength at which the measurements are carried out, and (ii), that the path
length of the solution is known accurately. Commonly, cuvettes with a path length of 1 cm are
used, then, the molar concentration c is simply: 3
c = A/e
The Beer-Lambert Law (A = εlc) implies that when concentration is equal to zero (c = 0),
absorbance must also be zero (A = 0). In other words, the calibration line must pass through the
origin.
A major source of error in spectrophotometric analysis is applying the Beer-Lambert
Law at inappropriate concentrations. The Beer-Lambert Law is strictly applicable only for dilute
solutions. It becomes less and less accurate as the concentration of the solution increases.
Once you have the calibration curve set up, you can measure the absorbance of any
unknown solution at the same wavelength and read off its concentration from the graph or
calculate from the slope. 4
In this experiment, absorbance of KMnO4 will be measured. KMnO4 is a compound
forming purple crystals with a metallic sheen, soluble in water (intense purple solution),
acetone, and methanol, but decomposed by ethanol; r.d. 2.70; decomposition begins slightly
above 100°C and is complete at 240°C. The compound is prepared by fusing manganese(IV)
oxide with potassium hydroxide to form the manganate and electrolysing the manganate
solution using iron electrodes at about 60°C. An alternative route employs production of sodium
manganate by a similar fusion process, oxidation with chlorine and sulphuric acid, then
treatment with potassium chloride to crystallize the required product.
Potassium manganate(VII) is widely used as an oxidizing agent and as a disinfectant in a
variety of applications, and as an analytical reagent. 5
Review of Related Literature
Beer-Lambert Law, more commonly known as Beer's Law, states that the optical
absorbance of a chromophore in a transparent solvent varies linearly with both the sample cell
pathlength and the chromophore concentration. Beer's Law is the simple solution to the more
general description of Maxwell's far-field equations describing the interaction of light with
matter. In practice, Beer's Law is accurate enough for a range of chromophores, solvents and
concentrations, and is a widely used relationship in quantitative spectroscopy.
Absorbance is measured in a spectrophotometer by passing a collimated beam of light
at wavelength λ through a plane parallel slab of material that is normal to the beam. For liquids,
the sample is held in an optically flat, transparent container called a cuvette. Absorbance (Aλ) is
calculated from the ratio of light energy passing through the sample (I0) to the energy that is
incident on the sample (I):
Aλ = -log (I/I0)
Beer's Law follows:
Aλ = ελbc
ελ = molar absorptivity or extinction coefficient of the chromophore at wavelength
λ (the optical density of a 1-cm thick sample of a 1 M solution). ελ is a property
of the material and the solvent.
b = sample pathlength in centimeters
c = concentration of the compound in the sample, in molarity (mol L-1)
In an absorbance experiment, light is attenuated not only by the chromophore, but also
by reflections from the interface between air and the sample, the sample and the cuvette, and
absorbance by the solvent. These factors can be quantified separately, but are often removed
by defining I0 as the light passing through a sample "blank" or "baseline" or reference sample
(for example, a cuvette filled with solvent but zero concentration of the chromophore is used as
the blank).
Many factors can affect the validity of Beer's Law. It is usual to check for the linearity of
Beer's Law for a chromophore by measuring the absorbance of a series of standards. This
"calibration" can also remove errors in the experiment, the equipment, and the batch of
reagents (such as cuvettes of unknown pathlength). 6
One of the most common applications of spectrophotometry is to determine the
concentration of an analyte in a solution. The experimental approach exploits Beer's Law, which
predicts a linear relationship between the absorbance of the solution and the concentration of
the analyte (assuming all other experimental parameters do not vary).
In practice, a series of standard solutions are prepared. A standard solution is a solution
in which the analyte concentration is accurately known. The absorbances of the standard
solutions are measured and used to prepare a calibration curve, which is a graph showing how
the experimental observable (the absorbance in this case) varies with the concentration. For this
experiment, the points on the calibration curve should yield a straight line (Beer's Law). The
slope and intercept of that line provide a relationship between absorbance and concentration:
A = slope c + intercept
The unknown solution is then analyzed. The absorbance of the unknown solution, Au, is
then used with the slope and intercept from the calibration curve to calculate the concentration
of the unknown solution, cu. 7
cu = Au - intercept
slope
Calibration curve or calibration plot is a plot of absorbance versus concentration for a
series of standard solutions whose concentrations are accurately known.
Because calibration curves are used in reading off the unknown concentrations, their
accuracy is of absolute importance. Therefore, make the standard solutions as accurately as
possible and measure their absorbances carefully. Each standard solution should be prepared in
identically the same fashion, the only difference between them being their concentrations. 8
Methodology
I. Preparation of Spectrophotometer
The spectrophotometer was turned on for 20 minutes before use. It was calibrated
by the use of water and adjusting its wavelength by 460 and its absorbance by 0.
II. Preparation of Solutions with Different Concentration
The 250mL of the assigned concentration (1 x 10-4) was prepared by adding the
computed volume of the distilled water with the standardized KMnO4 solution prepared
from experiment no. 8.
Four flasks were prepared and labelled as solution 1, 2, 3 and 4. In flask 1, a
small amount of the prepared aliquot solution was transferred. 30mL of the same
solution was diluted with 10mL distilled water and was placed on flask 2. On the third
flask, 10mL of the solution was mixed with 10mL distilled water. 30mL of distilled water
was added to a 10mL solution and was placed on flask 4. And fifth flask was given to the
instructor for the unknown solution.
III. Determination of Absorbance
The absorbance of the five solutions was read with the use of a
spectrophotometer.
After calibrating the spectrophotometer, a cuvette was washed then filled with
the first solution making sure that it reached the half of the circle marker.
The line in the cuvette was placed aligned on the line pointer of the spectrophotometer.
The absorbance of the solution was recorded. The same step was followed with solution
2, 3, 4 and the unknown solution, calibrating the spectrophotometer with water before
reading the next solution.
IV. Determination of the Unknown Concentration
A graph of the ratio between the absorbance and the concentration of the
solution was drawn. Using the recorded absorbance of the unknown solution and the
graph, the concentration of the unknown was traced.
Discussion of Data and/or Result
The concentration of every solution was determined by the volume used and the known
concentration of the aliquot solution (1 x 10-4) and the total volume of the desired solution
(Refer to appendix for computation).
The concentration of the unknown was identified using the measured absorbance and
the curve generated from the Beer’s Law equation. A graph of the absorbance versus the
concentration of the solution was plotted. Solution 1 with a concentration of 1.0 X 10-4 has an
absorbance of 0.277. In addition, Solution 2 with 7.5 X 10-5 molar concentration reads 0.193
absorbance. Moreover, Solution 3 with 5.0 X 10-5 concentration was found out to have an
absorbance of 0.115. And finally, solution 4 with 2.5 X 10-5 molar concentration has an
absorbance of 0.027.
Using the gathered data and its measured absorbance, the concentration of the
unknown solution was determined by plotting it on the graph. The concentration was known to
have a concentration of 7.66 X 10-5 and an absorbance of 0.198 as seen on Figure 1.0.
0.027
0.115
0.193
0.277y = 0.0828x - 0.054
R² = 0.9995
0
0.05
0.1
0.15
0.2
0.25
0.3
2.5 5 7.5 10
AB
SOR
BA
NC
E
CONCENTRATION (10-5M)
ABSORBANCE VS. CONCENTRATION
UNKNOWN SOLUTION
0.198
Figure 1.0 Graph shows the ratio between the absorbance and the molar concentration of the four different solution and the unknown.
7.66
Conclusion and Recommendations
When light passes through a colored solution, some of it is absorbed and some are
transmitted. These effects depend upon the transparency of the solution. More light is absorbed
and less is transmitted in opaque solutions while less light is absorbed and more is transmitted
in transparent ones.
Having the gathered data and the computed results, it can be said that the higher the
concentration of the solution, the higher its absorption. This is due to the number of particles in
the solution. If there are many particles present, more light will be absorbed and less will pass
through.
Percentage errors can be acquired through the calibration of the spectrophotometer.
Therefore, the absorbance and wavelength reading should be adjusted to the proper value using
a blank solution. In addition, errors can be obtained by the incorrect preparation of
concentration of the solution to be measured.
Bibliography
1. (n.a.). Spectrophotometry. Retrieved March 14, 2011 from
http://www.answers.com/topic/spectrophotometry .
2. (n.a.). Spectrophotometry: Absorption measurements & their application to quantitative
analysis. Retrieved March 14, 2011 from
http://employees.oneonta.edu/kotzjc/LAB/Spec_intro.pdf
3. (n.a.). South african structural biology initiative: The beer-lambert law. Retrieved March
14, 2011 from
http://sbio.uct.ac.za/Sbio/postgrad/modules/GRD/spectrophotometry/beer1.php
4. (n.a.). Plotting calibration graph. Retrieved Marach 15, 2011 from
http://employees.oneonta.edu/kotzjc/LAB/Spec_intro.pdf
5. (n.a.). Pottasium permanganate. Retrieved March 14, 2011 from
http://www.answers.com/topic/potassium-permanganate
6. (n.a.). Beer-lambert law. Retrieved March 15, 2011 from
http://www.oceanoptics.com/technical/beerslaw.asp
7. (n.a.). Determination of analyte concentration. Retrieved March 15, 2011 from
http://www.chm.davidson.edu/vce/spectrophotometry/UnknownSolution.html
8. (n.a.). Spectrophotometric analysis. Retrieved March 15, 2011 from
http://employees.oneonta.edu/kotzjc/LAB/Spec_intro.pdf
Appendix
SOLUTION MOLARITY ABSORBANCE
SOLUTION NO. 1 1.0 X 10-4 0.277
SOLUTION NO. 2 7.5 X 10-5 0.193
SOLUTION NO. 3 5.0 X 10-5 0.115
SOLUTION NO. 4 2.5 X 10-5 0.027
UNKNOWN 7.66 X 10-5 0.198
FORMULA: M1 V1 = M2 V2
SOLUTION NO.1: 1 X 10-4
SOLUTION NO.2: 1 X 10-4 30 = X2 40
X2 = 7.5 X 10-5
SOLUTION NO.3: 1 X 10-4 10 = X3 20
X3 = 5.5 X 10-5
SOLUTION NO.4 : 1 X 10-4 10 = X4 40
X4 = 2.5 X 10-5
Table 1.0 The computed molarity and measured absorbance of different solutions.