Functional characterization of UDP-glycosyltransferases from the liverwort Plagiochasma appendiculatum and their potential for biosynthesizing flavonoid 7-O-glucosides.

Flavonoid glucosides, typically generated from aglycones via the action of uridine diphosphate-dependent glycosyltransferases (UGTs), both contribute to plant viability and are pharmacologically active. The properties of UGTs produced by liverworts, one of the basal groups of non-vascular land plants, have not been systematically explored. Here, two UGTs potentially involved in flavonoids synthesis were identified from the transcriptome of Plagiochasma appendiculatum. Enzymatic analysis showed that PaUGT1 and PaUGT2 accepted various flavones, flavonols, flavanones and dihydrochalcones as substrates. A mutated form PaUGT1-Q19A exhibited a higher catalytic efficiency than did the wild type enzyme. When expressed in Escherichia coli, the yield of flavonol 7-O-glucosides reached to over 70 %. Co-expression of PaUGT1-Q19A with the upstream flavone synthase I PaFNS I-1 proved able to convert the flavanone aglycones naringenin and eriodictyol into the higher-yield apigenin 7-O-glucoside (A7G) and luteolin 7-O-glucoside (L7G). The maximum concentration of 81.0 µM A7G and 88.6 µM L7G was achieved upon supplementation with 100 µM naringenin and 100 µM eriodictyol under optimized conditions. This is the first time that flavonoids UGTs have been characterized from liverworts and co-expression of UGTs and FNS Is from the same species serves as an effective strategy to synthesize flavone 7-O-glucosides in E. coli.

An economical and environmentally friendly oxidative biaryl coupling promoted by activated MnO2.

An activated manganese dioxide (MnO2)-BF3·OEt2 oxidation system was developed to efficiently mediate the intramolecular as well as intermolecular biaryl coupling. The oxidative coupling proceeds smoothly at ambient temperature to deliver the corresponding five- to eight-membered tricyclic products in good to excellent yields. The employment of the combination of MnO2 and BF3·OEt2 is attractive on the basis of economical and environmental issues.

The combination effects of phenolic compounds and fluconazole on the formation of ergosterol in Candida albicans determined by high-performance liquid chromatography/tandem mass spectrometry.

A liquid chromatography/tandem mass spectrometry (LC/MS) with atmospheric pressure chemical ionization (APCI) for the quantification of ergosterol, lanosterol, and squalene was developed to evaluate the combination effects of phenolic compounds with fluconazole on ergosterol biosynthesis in Candida albicans. The three analytes were separated by a column of C18 and were quantified without interference with each other using positive mode tandem mass spectrometry (MS/MS). Molecular ions of ergosterol and lanosterol were detected as the [M+H-H2O]+ ion species at m/z 380 and 410, whereas squalene appeared as the [M+H]+ ion species at m/z 412. On fragmentation of ergosterol, lanosterol, and squalene, the product ions at m/z 69, 149, and 109, respectively, were present as major fragments. These product ions were used for the quantification of them in multiple reaction monitoring acquisition mode. The relationship between signal intensity and the analytes' concentration was linear over the concentration range of 0.05-10 microg/ml. Following the treatment of C. albicans with fluconazole in combination with albicanyl caffeate, resveratrol, and 3,4'-difluorostilbene, respectively, the content of ergosterol in both the sensitive and resistant C. albicans showed depletion, whereas the squalene showed accumulation especially in the sensitive isolates determined with the method developed.