UNIVERSITY OF ZIMBABWE

2016 Nov/Dec Examinations

Science
Faculty:...................................................................................................................................
Biochemistry
Department:............................................................................................................................
HBC201 : Practical Techniques in Biochemistry 2
Paper code and Title:............................................................................................................
2 hours
Duration:.................................................................................................................................
Mr A. Kasiyamhuru
Examiner:................................................................................................................................

Authorized Materials: Calculator, Graph Paper, Ruler

INSTRUCTIONS:

1. This paper contains 6 Questions

2. Answer 4 questions
3. Start each question on a new page
4. This question paper comprises 2 printed pages

NB: DO NOT TURN OVER THE QUESTION PAPER OR COMMENCE WRITING UNTIL
INSTRUCTED TO DO SO.
1. Describe paper chromatography including choice of paper, solvent, application,
advantages and disadvantages. [25 marks]

2. With the aid of a labelled diagram, describe the principles and application of gas
liquid chromatography (GLC). [25 marks]

3.
(a) Draw a block diagram of an HPLC system. [8 marks]

(b) With respect to High Performance Liquid Chromatography (HPLC), write notes
on the following:

(i) normal phase chromatography [5 marks]

(ii) reversed-phase chromatography [5 marks]
(iii) HPLC columns [7 marks]

4.
(a) Describe the principles of sodium dodecyl sulphate-polyacrylamide gel
electrophoresis (SDS-PAGE). [13 marks]

(b) A mixture of 3 amino acids lysine, glycine and aspartic acid was subjected
to paper electrophoresis in buffer pH 6.0. If the sample was applied at the middle
of the paper, predict with reasons, the final positions of the amino acids at the
end of the run. Draw and label the positive and negative ends of the
electrophoretogram.

Isoelectric point (pI); lysine = 9.7, glycine (pI) = 6.0 and aspartic acid (pI) = 3.0
[12 marks]

5.
(a) Give the meaning of the following terms in relation to gel filtration
chromatography;

(i) non-adsorptive technique [3 marks]

(ii) void volume [3 marks]
(iii) retention time [3 marks]
(iv) exclusion limit [4 marks]

(b) Briefly explain the basis of the use of gel filtration chromatography in the:

(i) purification of biological macromolecules [4 marks]

(ii) determination of the relative molecular mass of an analyte [4 marks]
(iii) desalting of a protein solution [4 marks]

PLEASE TURN TO THE NEXT PAGE

HBC 201 PAGE TWO

6. Describe the principle behind ion-exchange chromatography, pointing out its

limitations and the appropriate situations in which the technique can be used.
[25 marks]

END OF QUESTION PAPER