Determination of Lipase Activity by Reginald M. Archibald

Heretofore most methods for the determination of lipase activity have employed emulsions of fatty acid esters in water (1). These emulsions have been stabilized by use of such agents as gum acacia. The disadvantages of such methods are that (1) the emulsions break up to a considerable and variable extent during the course of the incubation, and (2) the enzyme is presumed to be in the aqueous phase, whereas the substrate is only dispersed (not in solution) in the aqueous phase. Attempts to study the relation of substrate concentration to rate of hydrolysis are complicated by the fact that the substrate and enzyme are in different phases. The rate of hydrolysis in these cases is dependent partly on the degree of dispersion of the substrate rather than on its concentration. Some of the commercially available esters’ of sorbitan and fatty acids are completely soluble in water. Use of solutions of these esters permits one to study the effect of substrate concentration on enzyme activity and eliminates the complications which attend the use of emulsions. Gomori (2) has employed esters with the trade names Tween 40 and Tween 601 as substrates to detect, and to localize histologically, lipase activity in sections of tissue. This paper outlines a method of measuring quantitatively lipase activity in which an aqueous solution of a polyoxyalkylene derivative of sorbitan monolaurate is used as substrate.’ For convenience this substrate will be designated hereafter by its trade name, Tween 20. ApparatusBurette, 10 cc. It is convenient to employ a burette connected with a reservoir for NaOH. Both burette and reservoir should be fitted with soda lime tubes to protect the NaOH from atmospheric COZ. 30 cc. glass-stoppered bottles (Pyrex) or 30 cc. heavy walled test-tubes with footed stirring rods 2 inches longer than the test-tubes.