Enantiomeric ratios of 2-methylbutanoic acid and its methyl ester: Elucidation of novel biogenetic pathways towards (R)-methyl 2-methylbutanoate in a beverage fermented with shiitake

Abstract

Up to 35% of (R)-methyl 2-methylbutanoate (M2MB) was observed in a beverage fermented with shiitake. As M2MB naturally occurs typically in high excesses of the (S)-enantiomer, the origin of the (R)-ester was elucidated by stable isotope labeled precursor-feeding studies. (R)-2-Methylbutanoic acid was identified as the main precursor in the substrate wort. Trace amounts of (R)-M2MB were produced by transformation of unsaturated secondary metabolites (tiglic aldehyde and tiglic acid) derived from l-isoleucine. Surprisingly, shiitake esterified (R)-2-methylbutanoic acid faster to (R)-M2MB than the corresponding (S)-enantiomer. Concurrently, spontaneous non-enantioselective degradation of M2MB occurred in shiitake. This explains diverse enantiomeric ratios of M2MB and different enantiomeric ratios of 2-methylbutanoic acid and M2MB in the beverage. As the odor threshold values of (R)-and (S)-M2MB differ significantly, these findings are of high relevance for the overall flavor of the fermented beverage and elucidate the discrepancy of enantiomeric ratios of 2-alkyl-branched acids and esters reported in nature.

Keywords: (R)-2-Methylbutanoic acid (PubChem CID: 6950479); (R)-Methyl 2-methylbutanoate; (S)-2-Methylbutanoic acid (PubChem CID: 448893); 2-Methylbutanal (PubChem CID: 7284); 2-Methylbutanoic acid; 2-Methylbutanoic acid (PubChem CID: 8314); 2-Methylbutanol (PubChem CID: 8723); Beverage; Ethyl tiglate (PubChem CID: 5281163); Fermentation; Methyl 2-methylbutanoate (PubChem CID: 13357); Methyl tiglate (PubChem CID: 5323652); Odor threshold; Pathway; Shiitake; Tiglic acid (PubChem CID: 125468); Tiglic aldehyde (PubChem CID: 5321950); l-Isoleucine.