Production of limonene and its derivative in Saccharomyces cerevisiae via metabolic engineering / 生物工程学报

Limonene and its derivative perillic acid are widely used in food, cosmetics, health products, medicine and other industries as important bioactive natural products. However, inefficient plant extraction and high energy-consuming chemical synthesis hamper the industrial production of limonene and perillic acid. In this study, limonene synthase from Mentha spicata was expressed in Saccharomyces cerevisiae by peroxisome compartmentalization, and the yield of limonene was 0.038 mg/L. The genes involved in limonene synthesis, ERG10, ERG13, tHMGR, ERG12, ERG8, IDI1, MVD1, ERG20ww and tLS, were step-wise expressed via modular engineering to study their effects on limonene yield. The yield of limonene increased to 1.14 mg/L by increasing the precursor module. Using the plasmid with high copy number to express the above key genes, the yield of limonene significantly increased up to 86.74 mg/L, which was 4 337 times higher than that of the original strain. Using the limonene-producing strain as the starting strain, the production of perillic acid was successfully achieved by expressing cytochrome P450 enzyme gene from Salvia miltiorrhiza, and the yield reached 4.42 mg/L. The results may facilitate the construction of cell factory with high yield of monoterpene products by S. cerevisiae.

A new monoterpene glycoside compound from roots of Paeonia lactiflora / 中草药

Objective: To study the constituents from the roots of Paeonia lactiflora. Methods: The chemical constituents were isolated by various chromatographic techniques and their structures were elucidated by physicochemical properties and NMR data. Results: Eleven compounds were obtained and characterized as (4S)-perillic acid 6-O-α-L-arabinopyranosyl-(1'→6'')- β-D-glucopyranosyl (1), 4,9-dihydroxy-8,10-dehydrothymol-1-O-β-D-glucoside (2), emodin-8-O-β-D-glucoside (3), resveratrol (4), β-D-glucopyranosyl benzoate (5), ilexperphenoside A (6), catechol (7), methyl gallate (8), ethyl gallate (9), 3-methoxygallic acid (10), 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranoside (11). Conclusion: Compound 1 is identified as a new compound, named perillic acid glycoside, compounds 2 and 5 are identified from the genus Paeonia for the first time.

Bioconversion of R-(+)-limonene to perillic acid by the yeast Yarrowia lipolytica

Perillyl derivatives are increasingly important due to their flavouring and antimicrobial properties as well as their potential as anticancer agents. These terpenoid species, which are present in limited amounts in plants, may be obtained via bioconversion of selected monoterpene hydrocarbons. In this study, seventeen yeast strains were screened for their ability to oxidize the exocyclic methyl group in the p-menthene moiety of limonene into perillic acid. Of the yeast tested, the highest efficiency was observed for Yarrowia lipolytica ATCC 18942. The conversion of R (+)-limonene by Y. lipolytica was evaluated by varying the pH (3 to 8) and the temperature (25 to 30 ºC) in a reaction medium containing 0.5% v/v limonene and 10 gµL of stationary phase cells (dry weight). The best results, corresponding to 564 mgµL of perillic acid, were obtained in buffered medium at pH 7.1 that was incubated at 25 ºC for 48 h. The stepwise addition of limonene increased the perillic acid concentration by over 50%, reaching 855 mgµL, whereas the addition of glucose or surfactant to the reaction medium did not improve the bioconversion process. The use of Y. lipolytica showed promise for ease of further downstream processing, as perillic acid was the sole oxidised product of the bioconversion reaction. Moreover, bioprocesses using safe and easy to cultivate yeast cells have been favoured in industry.