Hydrolysis of long-chain monoglycerides in micellar solution by pancreatic lipase

Abstract

1. 1. Certain long-chain monoglycerides have a high solubility in bile-salt solutions forming mixed monoglyceride-bile-salt micelles. Monoglycerides in such an optically clear, micellar solution are readily hydrolyzed by pancreatic lipase (glycerol ester hydrolase, EC 3.1.1.3). Using a rat-pancreatic-juice preparation containing negligible esterase activity, the hydrolysis of a number of pure monoglycerides in micellar solution has been studied. 2. 2. The rate of hydrolysis was essentially linear with time and enzyme concentration up to a level of around 60–70% hydrolysis of the substrate. The extent of hydrolysis fell when the concentration of micelles of identical composition was increased. There was a broad pH optimum 5.5–7.5. Bile salts greatly enhanced the rate of hydrolysis at low concentrations, but this effect was possibly chiefly due to their dispersing effect; at higher concentrations, they caused a marked inhibition of hydrolysis. Typical anionic detergents influenced hydrolysis in the same manner as bile salts. With a cationic detergent the enzyme concentration had to be considerably increased to obtain the same rate of hydrolysis. 3. 3. 2-Monoolein was not attacked to any appreciable extent by pancreatic lipase; thus the specificity of lipase for the 1-ester position could be directly shown. Glycol monooleate was readily hydrolyzed; 1-α,α-dimethyl-monodecanoin was not split. 4. 4. Under experimental conditions where 1-monoolein was extensively hydrolyzed, middle-chain saturated 1-monoglycerides were hydrolized to a very limited extent. Extensive hydrolysis could be obtained by increasing the micellar amphiphile to micellar bile-salt ratio, by adding other amphiphiles such as fatty acid, or by using a two-phase heptane-buffer system where the monoglyceride was concentrated at the interface. The results of these and other experiments were interpreted as showing the tightness of packing of the micelle to be a critical factor influencing the extent of hydrolysis. 5. 5. Titration of the fatty acid of a mixed bile salt-monoglyceride-fatty acid micelle indicated a pK a of 6.9 with the experimental conditions present.