miR­494 inhibits cancer­initiating cell phenotypes and reverses resistance to lapatinib by downregulating FGFR2 in HER2­positive gastric cancer.

In gastric cancer, >15% of cases are associated with the amplification of human epidermal growth factor receptor 2 (HER2), which leads to poor clinical outcomes. Lapatinib, a potent ATP­competitive inhibitor, is a small, orally active molecule, which inhibits the tyrosine kinases of HER2 and epidermal growth factor receptor type 1. The activation of receptor tyrosine kinases can contribute to lapatinib resistance in HER2­positive gastric cancer. The aim of the present study was to explore the effects of miR­494 and FGFR2 in regulation of cancer­initiating cell phenotypes and therapeutic efficiency of lapatinib in HER2­positive gastric cancer. Western blot analysis was used to identify that the expression of fibroblast growth factor receptor 2 (FGFR2), a receptor tyrosine kinase, was upregulated in gastric cancer tissues. Formation of cancer initiating cells (CICs) and resistance to lapatinib were determined using sphere growth assay and MTT assay, respectively. The overexpression of FGFR2 promoted the generation of cancer­initiating cells (CICs) and resistance to lapatinib in HER2­positive gastric cancer YCC1 cells. In addition, it was observed that overexpression of microRNA (miR)­494 downregulated the protein expression of FGFR2, inhibited the formation of CICs and reversed lapatinib resistance in YCC1­F cells (HER2­positive, FGFR2 overexpressing and lapatinib­resistant gastric cancer cells). Therefore, it was concluded that miR­494 inhibited the CIC phenotype and reversed resistance to lapatinib by inhibiting FGFR2 in HER2­positive gastric cancer.

Characterization and application of D-amino acid oxidase and catalase within permeabilized Pichia pastoris cells in bioconversions.

The high-density fermentation of recombinant Pichia pastoris was carried out in a 1-L fermentor. After 60 h of fermentation, the activities of D-amino acid oxidase (DAAO) and catalase assayed with the permeabilized cells attained 12,532 and 684,800 U/L, respectively. Additionally, the stability of DAAO and catalase within the permeabilized cells was relatively high. The half-life of the two enzymes reached 14.5 and 4.0 d at 30 degrees C, respectively. Furthermore, these permeabilized cells could convert D-phenylalanine into 99% phenylpyruvate within 100 min and could be efficiently reused up to 13 cycles. After being treated with base and heating, these treated permeabilized cells could be reused up to three cycles in a batchwise conversion of cephalosporin C, and about 90% 7-beta-(4-carboxybutanamido)-cephalosporanic acid was ultimately obtained at each cycle.

Production of Gluconobacter oxydans cells from low-cost culture medium for conversion of glycerol to dihydroxyacetone.

Gluconobacter oxydans could be immobilized as a biocatalyst for the conversion of glycerol to dihydroxyacetone. To reduce the production cost, the cells were produced from agricultural byproducts. Corn meal hydrolysate and corn steep liquor were employed to replace of sorbitol and yeast extract as medium for G. oxydans cell production. The optimal medium contained 80 g/L reducing sugar, 25 g/L corn steep liquor, and 10 g/L glycerol. The cell mass was about 4.22 g/L and the glycerol dehydrogenase activity was about 5.23 U/mL. For comparison, the cell mass was about 4.0 g/L and the glycerol dehydrogenase activity was about 5.35 U/mL cultured in sorbitol and yeast extract medium. These studies shown the corn meal hydrolysate and corn steep liquor medium was similar in performance to a nutrient-rich medium, but the cost of production was only 15% of that cultured in sorbitol and yeast extract medium. It was an economical process for the production of G. oxydans cells as biocatalyst for the conversion of glycerol to dihydroxyacetone in industry.

Study of ibuprofen glucopyranoside derivative synthesis by Candida antarctica lipase in organic solvent.

The direct esterification of ibuprofen and methyl alpha-D-glucopyranoside in organic solvent by Novozym 435 was investigated in terms of the main variables controlling the process, including initial water activity (a(w), 0.05-0.75), incubation time, (0-168 h) and substrate concentration. The results showed that the lower initial aw values resulted in higher enzymatic activity and bioconversion yield. The most appropriate initial aw and incubation time were 0.06 and 144 h, respectively. The results also showed that the optimal ratio of ibuprofen to methyl alpha-D-glucopyranoside was 2.0. By optimizing these parameters, the yield increased about 50%. In addition, the product was confirmed to be methyl 6-O-(2'-(4'-isobutylphenyl) propionyl) D-alpha-glucopyranoside.

Repeated use of immobilized Gluconobacter oxydans cells for conversion of glycerol to dihydroxyacetone.

Dihydroxyacetone (DHA) is of great interest in the fine chemical and pharmaceutical industry; therefore, the discovery of suitable biocatalysts for the efficient production of it is very necessary. In the experiment, Gluconobacter oxydans was immobilized in polyvinyl alcohol (PVA). Various parameters of the immobilized cells were investigated. The results have shown that the optimal conversion conditions by the immobilized cells were at 30 degrees C and pH 6.0. The immobilized cells remained very active over the period of 14 days for storage and only lost 10% of its original activity. Repeated use of immobilized cells for conversion of glycerol to DHA was carried out in a 1.5 L stirred tank reactor, the average conversion rate was about 86%. Despite the high shear stress, bead shape was not affected, even after five consecutive conversion cycles. The regenerated biocatalyst could recover 90% of its initial activity.

Penicillin acylase catalysis in the presence of ionic liquids.

Several ionic liquids were used as reaction media for penicillin G acylase catalysis. In all the assayed ionic liquids, [bmim]PF6 proved good media for PGA-catalyzed hydrolysis. A novel [bmim]PF6/water two-phase system is provided for 6-aminopenicillanic acid (APA) production, which will be more benefical than aquous batch systems used widely in industrial production of APA.

Pharmacological activity and hydrolysis behavior of novel ibuprofen glucopyranoside conjugates.

Novel ester prodrugs (II, III and IV) of ibuprofen (I) were synthesized using alpha-methyl, ethyl and propyl glucopyranoside as promoieties and tested for their anti-inflammatory, analgesic and ulcerogenic activities. Study of their chemical hydrolysis in aqueous buffer (pH 3.0-10.0) showed that these compounds acted as true prodrugs of ibuprofen, giving the ibuprofen and alkyl glucopyranoside. Additionally, all the derivatives studied did cleave rapidly inside the biological system and on oral administration did elicit a pharmacological profile quite similar to that of ibuprofen, but, unlike this drug, they displayed reduced gastric ulceration. In conclusion, these alkyl glucopyranoside esters have promising properties as prodrugs for oral delivery of ibuprofen.

Immobilization of permeabilized whole cell penicillin G acylase from Alcaligenes faecalis using pore matrix crosslinked with glutaraldehyde.

The activity of penicillin G acylase from Alcaligenes faecalis increased 7.5-fold when cells were permeabilized with 0.3% (w/v) CTAB. The treated cells were entrapped by polyvinyl alcohol crosslinked with boric acid, and crosslinked with 2% (v/v) glutaraldehyde to increase the stability. The conversion yield of penicillin G to 6-aminopenicillanic acid was 75% by immobilized system in batch reaction. No activity was lost after 15 cycles and about 65% enzyme activity was retained at the end of the 31th cycle.

Preparation of optically pure tert-leucine by penicillin G acylase-catalyzed resolution.

Penicillin G acylase, from Kluyvera citrophila, was used in kinetic resolution of DL-tert-leucine. N-phenylacetylated-DL-tert-leucine, chemically synthesized from DL-tert-leucine, was enantioselctively hydrolyzed by penicillin G acylase to obtain L-tert-leucine, D-tert-leucine was prepared by acid-catalyzed hydrolysis of the remaining substrate. The total yields of D-tert-leucine and L-tert-leucine are 80.6% and 83.1%, respectively. The enantiomeric excess of the two products, D-tert-leucine and L-tert-leucine, are 98.5% and 99%. This is a practical way for the preparation of D-tert-leucine and L-tert-leucine.

Coupling of enantioselective biooxidation of DL-1,2-propanediol and bioreduction of pinacolone via regeneration cycle of coenzyme.

Enantioselective biotransformation of DL-1,2-propanediol to D-2-hydroxypropanic acid was first reported by the authors. In the biooxidation process, there were some by-product formed and thus influenced the e.e. value and output of the acid. Restricting oxygen in the reaction system and offering additional proton receptor to the system displayed approving effect. The latter method constructed regeneration cycle system of coenzyme. In the article, the bioreduction of pinacolone was coupled to the enantioselective oxidation. Yield of the acid was increased by 36% and e.e. value of the product approached 99%.

Enzymatic synthesis of L-ascorbyl linoleate in organic media.

A novel L-ascorbyl fatty acid ester, L-ascorbyl linoleate was successfully prepared by enzymatic esterification and transesterification in a non-aqueous medium using immobilized lipase as biocatalyst. Changes in enzymatic activity and product yield were studied for the following variable: the nature of the fatty acid, the fatty acid concentration and water content. The yield of synthesis for the C18 unsaturated fatty acids were higher than for the C18 saturated fatty acid. Initial enzyme concentration does not affect the equilibrium of the reaction. And the product yield (33.5%) in the transesterification was higher than that of the esterification (21.8%) at a high-substrate concentration 0.3 M. The medium water content was found to have a distinct influence on the L-ascorbyl linoleate synthesis.

Effect of organic cosolvent on kinetic resolution of tert-leucine by penicillin G acylase from Kluyvera citrophila.

Penicillin G acylase (PGA) from Kluyvera citrophila immobilized on Amberzyml was used for enantioselective hydrolysis of N-phenylacetylated-DL-tert-leucine (N-Phac-DL-Tle) to produce L-tert-leucine (L-Tle). The effects of various organic cosolvents on hydrolysis of N-Phac-DL-Tle have been investigated in aqueous-cosolvent medium. It was founded that the rate of PGA-catalyzed reaction was significantly affected by the presence of 2% (v/v) organic cosolvent concentration. The initial rate fell with increasing logP of the cosolvent, but for logP values less than -0.24 the rate was faster than in purely aqueous medium. Additionally, the relative rate increases with the increase of dielectric constant (epsilon) of organic cosolvents. The yields of L-Tle in all aqueous-cosolvent systems were above 95% with the enantiomeric excess (ee) of >99%.

Studies on a novel carbon source and cosolvent for lipase production by Candida rugosa.

Oleic acid esters were shown to be the best carbon source for both cell growth and lipase production by Candida rugosa. Use of a cosolvent, dodecane, in fermentations improved the solubility of solid substrates and increased oxygen solubility. This resulted in the highest lipase activity in batch fermentation with glycerol trioleate and dodecane. Lipase activity reached 77.1 units ml(-1).

Enzymatic synthesis and antioxidant properties of L-ascorbyl oleate and L-ascorbyl linoleate.

L-Ascorbyl oleate and L-ascorbyl linoleate were synthesized by an immobilized lipase from Candida antarctica with yields of 38% and 44%, respectively. L-Ascorbyl oleate was stable in sterile culture medium over 12 h at 37 degrees C but L-ascorbyl linoleate degraded by 17%. Ascorbyl oleate had a better protective effect on human umbilical cord vein endothelial cells treated with H2O2 than of L-ascorbic acid-2-phosphate-6-palmitate (Asc2P6P).