Studies of the Phospholipase A₂ Activity of Rat Ileal Mucosa

Abstract

A rapid and simple procedure has been used to determine phospholipase A₂ activity (EC 3.1.1.4) in rat ileal mucosa. We used ¹⁴C-oleate-labeled Escherichia coli as substrate for the phospholipase activity and a 0.45-um Millipore filter to separate the product of hydrolysis–the ¹⁴C-oleic acid–from the unhydrolyzed substrate. The phospholipase A₂ activity was optimal at pH 9.8 and at 2 mM Ca²⁺ but another peak of activity appeared at pH 7.2. In addition, cell fractionation revealed yet another phospholipase A₂ activity at pH 5.0 in the absence of Ca²⁺ These findings suggest the presence of more than one phospholipase A₂ in the ileal mucosa and points to the possible use of a simple procedure for studying their distribution and properties.

Key Words:

• Ileal mucosa
• Millipore filtration
• phospholipase A₂ assay
• radiolabeled E. coli
• subcellular fractionation