With an attempt to synthesize high-value isoquercitrin (quercetin-3- O - β -D-glucopyranoside), we carried out the biotransformation of quercetin ( 1 ) by Gliocladium deliquescens NRRL 1086. Along with the aimed product quercetin 3- O - β -D-glycoside ( 2 ), three additional metabolites, 2-protocatechuoyl-phlorogucinol carboxylic acid ( 3 ), 2,4,6-trihydroxybenzoic acid ( 4 ), and protocatechuic acid ( 5 ), were also isolated. The time-course experiments revealed that there were two metabolic routes, regio-selectivity glycosylation and quercetin 2,3-dioxygenation, co-existing in the culture. Both glycosylation and oxidative cleavage rapidly took place after quercetin feeding; about 98% quercetin were consumed within the initial 8 h and the oxdized product (2-protocatechuoyl-phlorogucinol carboxylic acid) was hydrolyzed into two phenolic compounds (2,4,6-trihydroxybenzoic acid and protocatechuic acid). We also investigated the impact of glucose content and metal ions on the two reactions and found that high concentrations of glucose significantly inhibited the oxidative cleavage and improved the yield of isoquercitrin and that Ca 2+ , Fe 2+ , Mn 2+ , Mg 2+ , and Zn 2+ inhibited glycosylation. To test the promiscuity of this culture, we selected other four flavonols as substrates; the results demonstrated its high regio-selectivity glycosylation ability towards flavonols at C-3 hydroxyl. In conclusion, our findings indicated that the versatile microbe of G. deliquescens NRRL 1086 maitained abundant enzymes, deserving further research.
Fan N.,Du C. H.,Xu J. Q.,Xu Y. X.,Yu B. Y.,Zhang J.
The aim of the study was to expand the substrate scope of Gliocladium deliquescens NRRL 1086 and generate coumarin glycosides using 4-methylumbelliferone (4-MU) as a substrate. The obtained results indicated that 4-MU can be metabolized to its glucoside (M1) and sulfate conjugate (M2), and the structures of the metabolites were elucidated by spectroscopic or enzymatic methods. Time course experiments detected that nearly 90% of 4-MU could be metabolized within 24 h, and the maximum yield of M1 could reach as high as 32%. Further tests revealed that the glucose concentration in the medium had little effect on the yield of M1 but time of 4-MU-adding could markedly affect the glycosylation procedure, and the favorable time to accumulate M1 was in the 12 or 24 h-old stage II culture, while in the 36 h-old or even older stage II culture, the substrate was almost metabolized to M2. The attempts to alter the ratio between M1 and M2 were performed by addition of quercetin and S-tetrahydroberberrubine or reduction of the sulfate concentration in the culture medium. Herein we describe, to the best of our knowledge, the first example of simultaneously microbial glycosylation and sulfation of coumarins.
This study explored the substrate scope and regio-selectivity of Gliocladium deliquescens NRRL 1086 on hydroxylated benzophenones (HBPs), which was the follow-up investigation on the glycosylation of phenolic hydroxyl compounds by this unique microbe. Five HBPs substrates were used and three glycosylated metabolites, iriflophenone 2- O - β - d -glycopyranoside ( 6 , 40%), riflophenone 4- O - β - d -glycopyranoside( 7 , 21%) and 2,6,4′-trihydroxy-4-methoxybenzophenone-2- O - β - d -glycopyranoside ( 8 , 65%) were isolated and structurally identified unambiguously by ESI-MS, 1 H NMR, 13 C NMR, and 2D NMR spectroscopy. The versatile glycosylation ability of G. deliquescens NRRL 1086 on phenolic hydroxyl compounds may provide an alternative and convenient method to enable high-efficiency synthesis of various HBPs glycosides.
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