Division Note 2 MONITORING LIPASE ACTIVITYF1

Prepare an olive oil emulsion as follows:

In a mixer stir for 10 minutes a mixture of 165 ml of a 100 g/l gum arabic solution, 15 g of crushed ice and 20 ml of a previously neutralised olive oil.

Pour 10 ml of the emulsion into a 50 ml beaker, then 0,3 ml of a 0,2 g/ml sodium cholate solution and then 20 ml of distilled water.

Put the beaker in a thermostat set at 37 °C; introduce the electrodes of the pH meter and the screw agitator.

Using a burette, add a 0,1 N sodium hydroxide solution drop by drop until a pH of 8,3 is obtained.

Add an aliquot of the lipase powder suspension in water (0,1 g/ml of lipase). As soon as the pH meter reads 8,3, start the chronometer and add the sodium hydroxide solution drop by drop at a rate which maintains the pH at 8,3. Note every minute the volume of solution consumed.

Record the data on an x/y graph with the time on the x-axis and millilitres of 0,1 N alkaline solution consumed to keep a constant pH on the y-axis. A linear graph should be obtained.

Lipase activity, expressed in lipase units per mg, is given by the following formula:

$A=\frac{V\times N\times 100}{m}$

where:

A

is activity in lipase units/mg

V

is the number of millilitres of 0,1 N sodium hydroxide solution per minute (calculated on the basis of the graph)

N

is the titre of the sodium hydroxide solution

m

is the mass in mg of the test lipase.

A lipase unit is defined as the quantity of enzyme which releases 10 micro-equivalents of acid per minute.