Year: 2025
Type: article
Abstract: High Resolution Image Download MS PowerPoint Slide Thiaminase is an enzyme that destroys thiamine (vitamin B1) and is present in various fishes, mussels, plants, and bacteria. A sensitive and versatile assay is needed to measure thiaminase activity in complex biological samples using common reagents and variable reaction conditions. We developed a simple assay that uses fluorescence spectrophotometry and a microplate reader to measure thiaminase activity in fish tissue extracts via the destruction of added thiamine over time. Thiamine concentration, cosubstrate choice and concentration, and pH can be varied according to the needs of the investigator. Using this assay, we successfully measured common enzyme kinetic constants for thiaminase from a Pacific herring extract. The enzyme exhibited Michaelis–Menten kinetics ( V max = 13.8 nmol T g –1 min –1, K m = 27.4 μM) with respect to thiamine and showed positive cooperativity for nicotinic acid as a cosubstrate (Hill equation, V max = 12.1 nmol T g –1 min –1, K half = 20.3 mM, n = 3.2). The thiaminase pH optimum of a rainbow smelt extract was also successfully measured (pH = 5.06 ± 0.06). Thiaminase activities of common forage fishes from the northern Bering Sea (2024) were compared using this fluorescence-based assay and the conventional 4-nitrothiophenol assay, marking the first reported measurements of thiaminase activity in this ecosystem. This new fluorescence-based assay is a tool that can be used for studying thiaminase dynamics in specimens under a variety of reaction conditions, enabling studies that will improve the understanding of factors driving thiamine deficiency in consumers.
Source: ACS Measurement Science Au
Authors: Drew Porter, Cody Pinger
Language: English
Subfield: Neurology
Field: Medicine
Domain: Health Sciences
Open Access status: gold
Estimated APC paid: $3,500
OpenAlex ID: W4414352452