PHAYG062 – Preformulation

Partitioning: Consolidation questions

The questions below are designed to help you consolidate your learning. If you can answer them
all correctly, then you will do very well in the course. If not, then consult your lecture notes, the
course text (“Essentials of Pharmaceutical Preformulation” by Gaisford and Saunders), or me
(g.williams@ucl.ac.uk)! Please note that there is no obligation to do these questions: they do not
need to be handed in and will not be marked, but they offer you the opportunity to test your
understanding of the material covered.

1. Explain why we measure partioning rather than permeability.

The biopharmaceutical classification system requires knowledge of both solubility and

permeability. The former is easy to measure, but the latter is much harder – permeability is defined
as the ratio of drug absorbed into the blood from an oral dose relative to that given by IV. To
measure it would require extensive in vivo experiments, which would be very expensive and would
yield very variable results (because different people have different physiologies). Essentially then,
we measure partitioning because it is easy to measure, but permeability is hard! Although
partitioning does not give us results completely representative of in vivo conditions, it does enable
us to develop self-consistent sets of data for comparing drug performance.

2. Give the expression for Po,w. What values will Po,w have if a drug is (a) lipophilic, and (b)
hydrophilic?

Po,w = Co / Cw. For a lipophilic drug Po,w is > 1; for a hydrophilic drug Po,w will be < 1.

3. 107 mg of a drug D is dissolved in 50 mL of water, and 50 mL octanol added. The shake flask
method is then used to determine the partition coefficient. The concentration of drug in the
water layer at the end of the experiment is found to be 0.754 mg/mL. Assuming that the drug
is unionisable, calculate Po,w and log P.

Co + Cw = 107 mg

Cw = 0.754 mg/mL, so total amount of drug in water layer is 50 x 0.754 = 37.7 mg.

Hence, amount of water in octanol layer s 107 - 37.7 = 69.3 mg. Co = 69.3 / 50 = 1.386 mg/mL.

Po,w = Co / Cw = 1.386 / 0.754 = 1.84 (3 s.f.).

log P = log 1.84 = 0.264 (3 s.f.)

4. log Po,w for an unionisable drug is determined to be 0.566. A shake flask experiment was
performed with 100 mL water and 100 mL octanol, and after the experiment the drug
concentration in the water layer was found to be 0.356 mg/mL. Calculate the total amount of
drug used for the experiment.

log P = 0.566, and hence P = 3.681.

Amount of drug in water layer = 0.356 x 100 =35.6 mg.

Conc of drug in octanol layer = 0.356 x 3.681 = 1.310539 mg/mL

Amount of drug in octanol layer = 1.310… x 100 = 131.0539 mg

Total amount of drug = amount in water layer + amount in octanol layer = 35.6 +131.0539 =
166.6539 mg = 167 mg (3 s.f.)

5. An ionisable drug is analysed in a shake-flask experiment. At the end of the experiment, it is

determined that the concentration of drug in the water layer is 0.259 mg/mL, and the
concentration in the octanol layer 0.0897 mg/mL. In a separate experiment, the ratio of
ionised to unionised drug is calculated as 5.667 : 1. Calculate (a) Do,w, and (b) Po,w.

a) Do,w = Co / Cw = 0.0897 / 0.259 = 0.346 (3 s.f.).

b) Do,w = Po,w x funionised and so Po,w = Do,w / funionised

funionised = 1/ 6.667 = 0.14999… and hence Po,w = 0.346 / 0.14999… = 2.3089958 = 2.31 (3 s.f.)

6. Explain what is meant by hyper- and hypo-discriminating solvents.

A solvent which is more polar than n-octanol is hypo-discriminating; one which is less polar than n-
octanol is hyper-discriminating.

7. What effect does salt formation have on log P?

Salt formation causes the production of ionised species, and hence more of the drug will be found
in the water layer. This will cause log P to reduce in value, and frequently it becomes negative (salt
formation causes materials to become more hydrophilic, and in the limit through salt formation we
can convert a lipophilic material into a hydrophilic species).

8. Detail the differences between normal phase and reverse phase chromatography.

Normal phase chromatography has a polar stationary phase and a non-polar mobile phase;
reverse phase chromatography has a non-polar stationary phase and a polar mobile phase.

9. Draw a diagram to illustrate how the retention factor may be determined in TLC.
This gives you the resolution factor, R f. We then convert this to the retention factor using:

10. What factors need to be considered when developing an HPLC method for analysis of an API?

A range of factors must be considered, and extensive optimisation is likely to be required. We will
need to select:

- The column;
- The solvent system;
- The column pressure;
- The column temperature;
- The flow rate;
- An appropriate drug concentration range;
- A suitable detection technology (UV, mass spec, etc).

These parameters need to be optimised to ensure good resolution between analytes, but also the
effective use of machine time and reagents, solvents, etc.