Use of the Hydroxamic Acid Reaction for Determining Pectinesterase Activity

Abstract

The technique is: React pectinesterase contained on filter paper discs on a pectin-agar plate, flood the plate with a solution of hydroxylamine hydrochloride and then a solution of sodium hydroxide, acidify with hydrochloric acid, and then add a ferric chloride solution. The areas of substrate, desterified by the enzyme, appear as clear zones on a red background. The formation of the insoluble, red-colored ferric-pectin hydroxamic acid is characteristic of the presence of pectin carbomethoxy (methyl ester). Areas which are not colored are indicative of pectinesterase activity and this test can be used for qualitative and semi-quantitative test for approximately 1 × 10-3 to 1 × 10-4 pectinesterase units.