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Experiment 9

1. Basic principles of Beers Law


Beer-Lamberts Law, also known as Beers Law, quantitatively indicates the amount of
attenuation in relation to the concentration of the absorbing molecules and the path length
at which absorption occurs. The law states that the amount of the colored concentration in
a solution is directly proportional to its absorbance as given by the formula:
A = bc (1)
Where:
A = absorbance
a = specific absorptivity constant
b = path length
c = concentration of solution
2. Why Beers Law is expressed in terms of absorbance instead of transmittance.
It is expressed in absorbance, because this is directly proportional to the concentration,
while transmittance is directly proportional to light intensity.
3. Limitations of Beers Law
Deviations from direct proportionality can be observed between the solutions absorbance
and concentration. According to Skoog et al., these can be classified into real, chemical,
and instrumental deviations, which act as limits to the law. The real limit to the law is that
it describes the absorption behavior only of dilute solutions (concentration not exceeding
0.01M). The molar absorptivity of the analyte can be altered due to the electrostatic
interactions, caused by the close proximity of the ions, which leads to deviations from the
law. Deviation from the law can also occur due to the chemical reactions absorbing
species undergo such as dissociation, association, and reaction with the solvent which
creates products that have a different absorptivity.
4. Significance of spectral scanning
Spectral scanning can indicate the sample composition and its purity. Also, this
determines the calibration curve and max. Through manipulation of Beers Law and the
slope generated, the concentration of the analyte in the three unknown solutions can be
determined.
5. Reaction between iron and hydroxylamine hydrochloride
4Fe3+ (aq) + 2NH2OH (aq) 4Fe2+ (aq) + N2O (g) + 4H+ (aq) +H2O
6. Absorbing species and the spectral region at which the complex strongly absorbs
Fe(II) and 1,10-phenanthroline produces a red-orange complex. The
maximum absorption of the solution is 509.20 nm. The solution is nearer
to the color red and the wavelength which it absorbs can be obtained by
looking at its complementary color in the color wheel, in this case it
absorbs green (480-560nm).
7. Significance of following the sequence of addition of reagents.

This is in order to reduce the uncertainty of the absorbance.


8. Importance of calibration procedure.
This procedure is done to make correct measurements and also to obtain the maximum
wavelength absorption. In calibration a blank solution is used which contains the
substances causing absorbance except the analyte itself.
9. Importance of adding phenanthroline in excess.
The rationale behind this is that phenanthroline is used for quantitative determination and
is the main component for the color and complex formation. Color development is rapid
in the presence of excess phenanthroline.
10. Requirements for species to be analyzed using the spectrophotometric method.
A species can be analyzed using spectrophotometry if it has color and absorbs light at
wavelengths from a certain spectrum range. In addition, this absorption should be
distinguishable from that due to other substances in the sample, according to Skoog et al.
11. Importance of determining the max and the baseline
Obtaining the max will help in determining the absorbance spectrum, since light
absorbance is dependent on the wavelength. This also provides information on the
electronic structure of the analyte. The baseline will ensure the total absorption of the
species studied.
12. Possible sources of errors and their effect on caculated parameters.
Impurities in the reagents used and improper cleaning of the cuvette may trap more light
and increase the detected absorbance. Error in reading the volumes of the reagents to be
mixed may have altered the concentration, thus the absorbance as well.

References:
Biochrom.co.uk, (2015). Glossary of UV Vis Spectrophotometry - Biochrom. [online]
Available
at:
http://www.biochrom.co.uk/content/1/91/glossary-of-uv-visspectrophotometry.html [Accessed 5 May 2015].
Blauch,
D.
(2014).
Spectrophotometry:
Absorbance
Spectrum.
[online]
Chm.davidson.edu.
Available
at:
http://www.chm.davidson.edu/vce/Spectrophotometry/AbsorbanceSpectrum.html
[Accessed 5 May 2015].
Harris, D. (2010). Quantitative Chemical Analysis. 8th ed. New York: W.H. Freeman and
Co., p.400.
Sciencebuddies.org, (n.d.). Color Wheel. [image] Available at:
http://www.sciencebuddies.org/Files/4261/5/Chem_img103.jpg [Accessed 5 May 2015].
Skoog, D.; West, D.; Holler, F.; Crouch, S. (2014) Fundamentals of Analytical Chemistry;
9th ed.; Mary Finch: Belmont, California, pp.658-665.

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