Non-water miscible ionic liquid improves biocatalytic production of geranyl glucoside with Escherichia coli overexpressing a glucosyltransferase.

Whole cells of Escherichia coli overexpressing a glucosyltransferase from Vitis vinifera were used for the glucosylation of geraniol to geranyl glucoside. A high cell density cultivation process for the production of whole-cell biocatalysts was developed, gaining a dry cell mass concentration of up to 67.6 ± 1.2 g L(-1) and a glucosyltransferase concentration of up to 2.7 ± 0.1 g protein L(-1) within a process time of 48 h. Whole-cell batch biotransformations in milliliter-scale stirred-tank bioreactors showed highest conversion of geraniol at pH 7.0 although the pH optimum of the purified glucosyltransferase was at pH 8.5. The biocatalytic batch process performance was improved significantly by the addition of a water-immiscible ionic liquid (N-hexylpyridinium bis(trifluoromethylsulfonyl)imid) for in situ substrate supply. The so far highest final geranyl glucoside concentration (291 ± 9 mg L(-1)) and conversion (71 ± 2 %) reported for whole-cell biotransformations of geraniol were achieved with 5 % (v/v) of the ionic liquid.

Four new constituents from the fruits of Cyclocodon lancifolius (Roxburgh) Kurz.

Phytochemical analysis of the fruits of Cyclocodon lancifolius led to the isolation of two new phenylpropanoid-derived glycosides (1-2), two new geranyl glucosides (3-4), and nine known compounds (5-13). Their chemical structures were elucidated by extensive spectroscopic data. The absolute configuration of the sugar moiety was determined by hydrolysis and derivatization. All compounds were evaluated for their xanthine oxidase (XO) and α-glucosidase inhibitory activities, and four compounds showed weak inhibitory activity towards XO.

Variation patterns in the content of glycosides during green tea manufacturing by a modification-specific metabolomics approach: Enzymatic reaction promoting an increase in the glycosidically bound volatiles at the pan firing stage.

The glycosides are presumed to influence the quality of green tea but the molecular mechanism behind remains unclear. To elucidate the contribution of glycosides to the flavor formation of green tea, changes of both glycosidically bound non-volatiles (GBNVs) and glycosidically bound volatiles (GBVs) during the manufacturing of green tea were investigated using a modification-specific metabolomics method. A total of 64 glycosides (47 GBNVs and 17 GBVs) were identified and their contents mainly changed during the pan firing and drying stages of green tea manufacturing. Notably, the contents of GBVs significantly increased by 1.12-4.46-fold during pan firing. Correlation analysis showed that the GBVs contents were negatively related to the contents of volatiles and glucose. Model experiments revealed that enzymatic synthesis contributed to the increase in the content of GBVs during the pan firing. This comprehensive study on the glycosides changes revealed the molecular bases for GBVs increments during the pan firing.

Rational selection of biphasic reaction systems for geranyl glucoside production by Escherichia coli whole-cell biocatalysts.

Geranyl glucoside, the glucosylated, high-value derivative of the monoterpenoid geraniol, has various applications in the flavor and fragrance industry and can be produced through whole-cell biotransformation of geraniol with Escherichia coli whole-cell biocatalysts expressing the glucosyltransferase VvGT14a. However, the low water solubility and high cytotoxicity of geraniol require the design of a proper biphasic system where the second, non-aqueous phase functions as an in-situ substrate reservoir. In this work, a rational selection strategy was applied for choosing suitable sequestering phases for geranyl glucoside production by whole-cell biotransformation of geraniol. Hansen solubility parameters and octanol/water distribution coefficients were used as first principle methods in combination with extensive database research to preselect 12 liquid and 6 solid sequestering phases. Subsequently, experimental approaches were applied to determine physicochemical characteristics and the distribution of geraniol and geranyl glucoside between the phases. Moreover, the effects of the sequestering phases on the whole-cell biocatalysts and on the produced geranyl glucoside concentration were measured during parallel biotransformations in milliliter-scale stirred-tank bioreactors. The fatty acid ester isopropyl myristate emerged as the best choice due to its low viscosity, very poor water solubility, low price and compatibility with the whole-cell biocatalyst. The biphasic system containing 20% (v/v) of this solvent boosted geranyl glucoside production (4.2-fold increase of geranyl glucoside concentration in comparison to aqueous system) and exhibits advantageous partitioning of geraniol into the organic phase (logD of 2.42±0.03) and of geranyl glucoside into the water phase (logD of -2.08±0.05). The systematic selection of a suitable biphasic system constitutes basic groundwork for the development of new bioprocesses involving geraniol. Moreover, this study can serve as a guideline for selecting sequestering phases for other whole-cell biotransformation processes.