University of North Florida
Browse the Citations

Contact Info

Stuart Chalk, Ph.D.
Department of Chemistry
University of North Florida
Phone: 1-904-620-1938
Fax: 1-904-620-3535
Website: @unf

View Stuart Chalk's profile on LinkedIn

Mahi mahi

Classification: Marine -> fish -> mahi mahi

Citations 1

"Flow Injection Assay Of Enzyme Inhibition In Fish Using Immobilized Diamine Oxidase"
Anal. Chim. Acta 1992 Volume 261, Issue 1-2 Pages 351-359
James M. Hungerford* and Aleksei A. Arefyev

Abstract: The sample is merged with 0.04 M sodium phosphate buffer of pH 7.2, each flowing at 0.8 mL min-1, and two portions of the mixture are injected, one upstream and one downstream of a reactor containing the cited enzyme immobilized on CNBr-activated Sepharose, into a stream (0.4 mL min-1) of 0.02 M sodium phosphate buffer of pH 7.2. The resulting stream is treated with 0.24 mM phthalaldehyde in 0.3 M NaOH and, after reaction in a 1-m knitted coil, with 0.37 M H3PO4 (each at 0.4 mL min-1), and the fluorescence due to histamine in each sample plug is measured at 450 nm (excitation at 365 nm). The ratio of the peak heights for the solution that has passed through the reactor and the one that has not corresponds to the fraction of endogenous histamine (the enzyme substrate) remaining. This fraction increases with increasing concentration. of inhibitors of the cited enzyme in the sample. The mechanism of inhibition is discussed. The inhibition caused by anserine is sigmoidally related to concentration. up to ~10 µM; inhibition by aminoguanidine, cadaverine and tyramine is similar, but that by putrescine is much weaker. The method also allows the determination of histamine. Fish samples are extracted by homogenization with 0.02 M HCl and the extracts are diluted with 1 mM HCl for analysis. A flow injection assay for inhibition of diamine oxidase (DAO) is described. Histamine in decomposed fish is detected and used as the enzyme substrate. Sample extracts are doubly injected; 1 of 2 resulting sample zones is passed through a reactor containing DAO immobilized to Sepharose. Histamine is detected sequentially in both the pristine and enzymatically treated zones. The detection chemical (kinetics-optimized condensation with phthalic dicarboxaldehyde) allows the selective detection of histamine. Subsequent calibration with histamine standard and comparison of the 2 peak heights allows generic detection of compounds inhibiting the DAO-mediated conversion of histamine. Preliminary data show that the system can detect inhibitors in extracts of decomposed Mahi-Mahi (associated with a Scombroid poisoning outbreak) and that far lower levels of inhibition are observed in fresh fish. Several known inhibitors and competitive substrates of DAO are also tested.
Enzyme, inhibition Sample preparation Fluorescence Interferences Immobilized enzyme Knotted reactor