A continuous spectrophotometric method for monitoring phospholipase D-catalyzed reactions of physiological substrates

A continuous spectrophotometric method for monitoring phospholipase D-catalyzed hydrolysis of long acyl chain phosphatidylcholines has been formulated at pH 8.0 in a mixed detergent system using the coupling enzymes choline oxidase and peroxidase. Standard curves for phosphatidylcholine determination in both end-point and rate modes are presented and applied to the estimation of that phospholipid in a solubilized human erythrocyte membrane sample. In rate mode the method is suitable for kinetic study of phospholipase D with physiological substrates in micellar form.