Enzymatic production of Volatile Organosulphur Compounds for the Flavour and Fragrance Industry

Volatile organosulphur compounds (VOSC) are a diverse group of chemical entities, which are among the highest impact aroma chemicals (HIACs) in the flavour and fragrance (F&F) industry. These compounds may be synthesized by both chemical processes, or through the use of biological pathways referred to 'natural production' methods. The overall aim of this investigation was to establish methods for the enzymatic production of the VOSCs methanethiol, hydrogen sulphide, thiomenthone and other valuable F&F compounds from natural precursors. The first part of this project was focussed on identifying novel bacterial and plant C-S lyase enzymes, which can cleave the bond between sulphur and carbon in methionine, cysteine and related conjugates synthesised through Michael addition reaction. Methods for the optimal production of recombinant C-S lyases were developed using E. coli as a bacterial host. The collection of C-S lyases selected were cloned and then used for in vitro natural production studies. A comprehensive understanding of the factors controlling their effective use in biotransformation studies included the dependence and optimisation of cofactor, temperature and pH in order to choose the most appropriate enzymes for the production of each VOSC. In addition, methods for the enzymatic methylation of VOSC and other free thiols were developed using an S-methyl transferase derived from Arabidopsis thaliana. Finally, building on these single-step reactions, the concept of linked enzymatic reactions in vivo and in vitro to generate S-methylated VOSCs directly from amino acid derivatives was investigated.