A simplified low volume colorimetric assay for rapid comparison of cyanogenic glycoside production between barley genotypes

Abstract

Ethyl carbamate (EC), a genotoxic and carcinogenic compound, is an undesirable trace component in spirits and whisky. The precursor of EC produced from barley (Hordeum vulgare L.) malt is primarily a cyanogenic glycosidic nitrile (GN) known as epiheterodendrin (EPH). EPH production is genetically controlled in barley, low or non-GN varieties exist and EPH production can be controlled by using low or non-GN barley varieties for malt production. Here, we report on a rapid and inexpensive colorimetric assay modified from Cook and Oliver (in: European Brewery Convention (eds) Proceedings of the 23rd Congress, Lisbon. European Brewery Convention, Zoeterwoude, Netherlands, 1991). The new low-volume, high-throughput, semi-quantitative test method can be used for the selection of low or non-GN breeding lines through samples of acrospires. This method is based on the detection of cyanide by the reaction with chloramine-T followed by the addition of the reagent (pyridine-barbituric acid) to form a soluble violet-blue product measured at 590 nm. Absorbance measurements and the visual color are used to estimate the presence of EPH production in acrospires. In this report, the level of EPH production was compared among some commercial European non-GN lines, Canadian malt varieties and advanced barley breeding lines.