Spectrophotometric assay and electrophoretic detection of trans-feruloyl esterase activity

• Published in: Analytical biochemistry Vol. 196; no. 2; p. 360
• Main Authors: McCallum, J A, Taylor, I E, Towers, G H
• Format: Journal Article
• Language: English
• Published: United States 01-08-1991
• Subjects: Arabinose - analogs & derivatives; Arabinose - isolation & purification; Carboxylic Ester Hydrolases - metabolism; Cell Wall - enzymology; Coumaric Acids - isolation & purification; Electrophoresis - methods; Hydrogen-Ion Concentration; Isoenzymes - metabolism; Spectrophotometry, Ultraviolet - methods; Triticum - enzymology; Triticum - ultrastructure
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(91)90479-D

Wheat bran cell walls were subjected to mild acid hydrolysis and the major phenolic product was purified and identified as 5-O-(trans-feruloyl)-arabinofuranose. Sensitive continuous and stopped, microtiter plate-based spectrophotometric assays for trans-feruloyl esterase activity were developed using this compound as substrate. Procedures were also developed for the detection of trans-feruloyl esterase activities on gels following electrophoresis using this compound. These procedures are applicable to other natural feruloyl esters derived from plant cell walls by enzymatic hydrolysis. The extracellular trans-feruloyl esterases of Aspergillus niger 814 grown on 1% wheat bran were fractionated by anion-exchange chromatography and isoelectric focusing. These studies indicate that there are multiple forms of trans-feruloyl esterase but that most activity is associated with a major isozyme with a pI of 3.2.