Department of Chemical Engineering, Indian Institute of Technology

CLL780: Bioprocessing and Bioseparations

Academic Year: 2020-2021
Major Examination

Date: 6th December 2020 Closed book and closed notes Maximum marks: 30
Please note: A. Cell phones are banned from exam rooms. B. Marks are not only for the final
answers but also for the intermediate steps. C. Be concise in your answers.

Question 1: 1 Mark: You are the Vice President of manufacturing in a major biopharmaceutical
company. Your CEO has just tasked you to get the facility ready for continuous processing of
biopharmaceuticals. What are the top 3 hurdles you see to achieve this?
Question 2: 1 Mark: Perfusion cell culture seems to offer significant advantages over the
conventional fed batch cell culture. What are the 3 biggest challenges that once faces when
doing large scale perfusion cell culture?
Question 3: 1 Mark: State 3 major differences with respect to prices of biotherapeutic products
in the US vs India.
Question 4: 1 Mark: What role do biosimilars play with respect to affordability of
biotherapeutic products? What impact have they had in the US vs. Africa?
Question 5: 1 Mark: Make a plot to show how cost of materials, labor cost and indirect cost
vary as a function of annual production capacity and bioreactor volume?
Question 6: 1 mark: State major advantages that Model Predictive Control offers vs. PID control.
Question 7: 1 Mark: State 3 advantages and 3 limitation of PAT in bioprocessing.
Question 8: 2 marks: What would be your tool of choice for assessing the secondary structure
and for assessing the tertiary structure of a protein? In 2 sentences each state the working
principle of the tool.
Question 9: 2 marks: Why do some cells die when exposed to very high temperatures and
others when they are exposed to lower (but greater than zero) temperatures?
Question 10: 1 mark: What is the Dittus-Boelter and Desalius’ expression in filtration? Write the
equation as well as the expressions used. What does it relate?
Question 11: 1 mark: In a few sentences state the three mechanisms that govern plugging of
filters and state their applicability with respect to particle to pore size ratio and particle
concentration.
Question 12: 2 marks: State any 3 approaches for scaling down a microbial fermentation.
Write the pertinent expressions as well.
Question 13: 3 marks: (1+0.5+0.5+1 marks) Pseudomonas sp. has a mass doubling time of 2.4 h
when grown on acetate. The saturation constant using this substrate is 1.3 g/l (which is unusually
high), and cell yield on acetate is 0.46 g cell/g acetate. If we operate a chemostat on a feed stream
containing 38 g/l acetate, find the following:
a. Cell concentration when the dilution rate is one-half of the maximum
b. Substrate concentration when the dilution rate is 0.8 Dmax
c. Maximum dilution rate
d. Cell productivity at 0.8 Dmax

Question 14: 3 marks: (1+1+1 marks) A stirred-tank reactor is to be scaled down from 10 m3 to
0.1 m3. The dimensions of the large tank are: Dt = 2 m; Di = 0.5 m; N = 100 rpm.
a. Determine the dimensions of the small tank (Dt, Di, H) by using geometric similarity?
b. What would be the required rotational speed of the impeller in the small tank if the following
criteria were used?
(i) Constant tip speed
(ii) Constant impeller Re number

Question 15: 3 marks: At a constant vacuum pressure of 60 kPa, a cell broth is required to be
filtered at a rate of 1000 L/h using a rotary vacuum filter. The total cycle time for the drum is 50 s
with the cake formation time as 15 s. Determine the area of the filter that will be required if the
broth has a viscosity of 0.003 Pa.s with cake solids (dry basis) formation per volume of filtrate
given as 10 kg/m3. The specific cake resistance is 9 x 102 m/kg. The resistance of the filter medium
can be neglected.

Question 16: 3 marks: (1+1+1 marks) The following data were obtained by gas-liquid
chromatography on a 30-cm packed column:
Compound tR (min) W1/2 (min)

Air 1.9 -

Methylcyclohexane 10.0 0.76

Hexane 10.9 0.82

Toluene 11.4 1.06

Calculate:
(a) an average number of plates from the data
(b) an average plate height for the column
(c) resolution between methylcyclohexane and toluene
Question 17: 3 marks: (1+1+1 marks)

The process flow diagram for a continuous process for production of a mAb therapeutic molecule
is shown in the figure. Given the following information:

1. Outlet flowrate from bioreactor = 100 mL/min

2. Titer of mAb in bioreactor harvest = 0.6 g/L
3. Binding capacity of Protein A resin:

Loading flowrate (mL/min) Concentration in load Binding capacity (mg mAb

(mg/mL) /mL resin)
5 0.5 40
5 5 55
10 0.5 35
10 5 50
30 0.5 15
30 5 25

4. Height of each Protein A column = 20 cm

5. Loading time for each Protein A column = 30 min
6. Yield of protein A step = 92%
7. Elution flowrate of each Protein A column = 10 mL/min
8. Loading time for each CEX column = 45 min
9. Volume of each CEX column = 60 mL
10. Maximum binding capacity of CEX resin = 60 mg/mL
11. Elution time for each CEX column = 25 min

(a) The titer of the mAb in the bioreactor is relatively low. Therefore, an in-line concentrator (ILC) is
placed before the Protein A step. The ILC has a concentration factor of 8.3x. Find the optimum
column diameter for the Protein A columns under the given conditions. (Hint: ensure that the
column is loaded up to its binding capacity under the given conditions).
(b) The elution from the Protein A column is collected in a surge tank. The duration of each elution is
20 minutes, with 10 minutes pause between elutions. Determine the loading flowrate for the CEX
column to ensure that the surge tank never overflows or runs dry. (Assume constant loading into
CEX with no pause time.)
(c) What is the total amount of mAb (in mg) loaded onto each CEX column? (Hint: find the
concentration of the Protein A elute). What percentage of the binding capacity of the CEX resin is
utilized? Suggest a reason why this is not 100%.