Natural 2(E)-hexenal was produced in two steps from hydrolysed linseed oil, which contains the most linolenic acid among the available natural sources. In the first step 13-hydroperoxy-9(Z),11(E),15(Z)-octadecatrienoic acid (13-HPOT) was formed from linolenic acid (100 mM) by soybean lipoxygenase-1 (Lox-1) isoenzyme with oxygen as co-substrate. The reaction resulted in 57 mM 13-HPOT with a yield of 62%. In the second step 13-HPOT (20 mM) was cleaved by green bell pepper hydroperoxide lyase resulting in 1.6 mM 2(E)-hexenal and 5.9 mM 3(Z)-hexenal (37% yield for the hexenal isomers together). Hexenals were isolated from the reaction mixture by repeated steam distillations. During distillations the 2(E)-hexenal:3(Z)-hexenal isomer ratio was changed from 0.27 to 7.86 as a consequence of heat. (C) 2004 Elsevier Inc. All rights reserved.