Sodium Chloride → Oleic Acid

Participants

Reaction Steps

Purified human synovial fluid PLA2 (purified by the procedure of Fawzy, A. A. and Franson, R. C. Biophys. J. 49, 533a 1986!) was obtained from R. C. Franson (Medical College of Virginia, Richmond, Va.). PLA2 activity was measured by using 14C!-oleate labeled autoclaved E. coli as substrate as described by Franson, R. C., Patriarca, P., and Elsbach, P. J., Lipid Res. 15, 380-388 (1974). The assay was performed at 37° C for 30 min. in a final volume of 100 μL 50 mM Hepes buffer (pH 7.0) containing 150 mM NaCl, 5 mM CaCl2, and E. coli cells (corresponding to 10 mmol phospholipid). The test compound or control vehicle was preincubated with PLA2 for 5 min., followed by adding E. coli to initiate the reaction. The reaction was terminated by adding 2 mL tetrahydrofuran. The reaction product, 14C!-oleic acid, was isolated using a 1 mL Bond Elute--NH2 column and counted by liquid scintillation spectrometry.

Provenance & Evidence