Enzymatic synthesis of terpenyl esters by transesterification with fatty acid vinyl esters as acyl donors by Trichosporon fermentans lipase

Enzymatic synthesis of terpenyl esters by esterification or transesterification with fatty acid vinyl esters as acyl donors by celite-adsorbed lipase of Trichosporon fermentans was investigated. In direct esterification of geraniol, the lipase showed high reactivity toward fatty acids with carbon chains longer than C-8, but little reactivity toward fatty acids with shorter chains. With fatty acid vinyl esters as acyl donors, the lipase catalysed the synthesis of geranyl and citronellyl esters with carbon chains shorter than C-6 in with yields of >90% molar conversion. Time course, effects of added water, temperature and substrate concentration were studied for the synthesis of geranyl acetate. Molar conversion yield reached 97.5% after 5 h incubation at 30–40°C with the addition of 3% water. In this reaction, no inhibition by substrates such as geraniol and vinyl acetate was observed.     

Increase of catalytic activity of α-chymotrypsin in organic solvent by co-lyophilization with cyclodextrins

-Chymotrypsin was lyophilized in the presence of 2,3,6-tri-O-methyl -cyclodextrin, and it displayed activity 40 fold higher than free -chymotrypsin for transesterification in acetonitrile. -Chymotrypsin which was co-lyophilized with hydroxypropylated - or -cyclodextrins retained more than 98% of its initial activity after 6 h incubation in acetonitrile.     

Peptide sequencing and site-directed mutagenesis identify tyrosine-319 as the active site tyrosine of Escherichia coli DNA topoisomerase I

Tyrosine 319 of E. coli topoisomerase I is shown to be the active site tyrosine that becomes covalently attached to a DNA 5' phosphoryl group during the transient breakage of a DNA internucleotide bond by the enzyme. The tyrosine was mapped by trapping the covalent complex between the DNA and DNA topoisomerase I, digesting the complex exhaustively with trypsin, and sequencing the DNA-linked tryptic peptide. Site-directed mutagenesis converting Tyr-319 to a serine or phenylalanine completely inactivates the enzyme. The structure of the enzyme and its catalysis of DNA strand breakage, passage, and rejoining are discussed in terms of the available information.     

Potential biotechnological application of microalgae: a critical review

Microalgae are diverse microorganisms inhabiting a wide range of habitats with only a small fraction being cultivated for human use. Recently, interest in microalgal research has increased in the quest for alternative renewable fuels due to possible depletion of fossil fuels in the near future. However, costly downstream processing has hampered the commercialization of biofuels derived from microalgae. Several value added products of industrial, pharmaceutical and agricultural relevance could be simultaneously derived from microalgae during bioenergy production. Despite these value-added products having the potential to offset the high cost of downstream processing of renewable fuels, their production has not been explored in-depth. This review presents a critical overview of the current state of biotechnological applications of microalgae for human benefit and highlights possible areas for further research and development.     

Six‐membered cyclic carbonates from trimethylolpropane: Lipase‐mediated synthesis in a flow reactor and in silico evaluation of the reaction

Six‐membered cyclic carbonates with hydroxyl and methoxycarbonyloxy functional groups were prepared by transesterification of trimethylolpropane (TMP) with dimethylcarbonate (DMC) by solvent‐free lipase‐mediated flow reaction followed by thermal cyclization. The flow reaction efficiency was evaluated using different configurations of reactor consisting of packed beds of Novozym®435 (immobilized Candida antarctica lipase B—CalB—a.k.a. N435) and molecular sieves, flowrate, and biocatalyst loads. The mixed column of the biocatalyst and molecular sieves, allowing rapid and efficient removal of the by‐product—methanol—was the most efficient setup. Higher conversion (81.6%) in the flow reaction compared to batch process (72%) was obtained using same amount of N435 (20% (w/w) N435:TMP) at 12 h, and the undesirable dimer and oligomer formation were suppressed. Moreover, the product was recovered easily without extra separation steps, and the biocatalyst and the molecular sieves remained intact for subsequent regeneration and recycling. The reaction of CalB with DMC and the primary transesterification product, monocarbonated TMP, respectively, as acyl donors was evaluated by in silico modeling and empirically to determine the role of the enzyme in the formation of cyclic carbonates and other side products. DMC was shown to be the preferred acyl donor, suggesting that TMP and its carbonated derivatives serve only as acyl acceptors in the lipase‐catalyzed reaction. Subsequent cyclization to cyclic carbonate is catalyzed at increased temperature and not by the enzyme. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:375–382, 2017     

A simplified and efficient method for the analysis of fatty acid methyl esters suitable for large clinical studies

Conventional sample preparation for fatty acid analysis is a complicated, multiple-step process, and gas chromatography (GC) analysis alone can require >1 h per sample to resolve fatty acid methyl esters (FAMEs). Fast GC analysis was adapted to human plasma FAME analysis using a modified polyethylene glycol column with smaller internal diameters, thinner stationary phase films, increased carrier gas linear velocity, and faster temperature ramping. Our results indicated that fast GC analyses were comparable to conventional GC in peak resolution. A conventional transesterification method based on Lepage and Roy was simplified to a one-step method with the elimination of the neutralization and centrifugation steps. A robotics-amenable method was also developed, with lower methylation temperatures and in an open-tube format using multiple reagent additions. The simplified methods produced results that were quantitatively similar and with similar coefficients of variation as compared with the original Lepage and Roy method. The present streamlined methodology is suitable for the direct fatty acid analysis of human plasma, is appropriate for research studies, and will facilitate large clinical trials and make possible population studies.     

Lipase-catalyzed ethanolysis of fish oils: multi-response kinetics

The kinetics of the lipase-catalyzed (Pseudomonas cepacia) ethanolysis of fish oil has been studied in a batch reactor using menhaden oil, tuna oil, and acylglycerol mixtures derived from menhaden oil. Multi-response models derived from a generalized Michaelis-Menten mechanism were developed to describe the rates of formation of ethyl esters of the primary fatty acids present in the precursor oil. A first-order model for deactivation of the lipase was fit simultaneously to one of the data sets.     

A novel enzymatic route for biodiesel production from renewable oils in a solvent-free medium

A new enzymatic route for biodiesel production from soybean oil was developed using methyl acetate as a novel acyl acceptor. Novozym 435 (immobilized Candida antarctica lipase) gave the highest methyl ester (ME) yield of 92%. The optimum conditions of the transesterification were 30% enzyme based on oil weight; a molar ratio of methyl acetate/oil of 12:1; temperature 40 °C and reaction time 10 h. Since no glycerol was produced in the process, this method is very convenient for recycling the catalyst and by-product triacetylglycerol showed no negative effect on the fuel property.     

Regioselective acylation of flavonoids catalyzed by immobilized Candida antarctica lipase under reduced pressure

A single-step acylation of rutin and naringin, catalyzed by immobilized Candida antarctica lipase B in 2-methyl-2-butanol, occurred preferentially on the primary hydroxyl group. Using palmitic methyl ester as acyl donor, the acylation rate of naringin was 10-fold higher than that of rutin. Under optimal conditions, i.e. a molar ratio acyl donor/naringin of 7:1 and 200 mbar, 92% naringin was acylated.     

有机相脂肪酶催化合成乙酸肉桂酯

对有机相中酶法催化合成乙酸肉桂酯的转酯化反应进行研究。结果发现:Candida anatarctic脂肪酶(Novozyme435)、根霉脂肪酶(Rhizopus niveus lipase)和荧光假单胞菌脂肪酶(Pseudomonas fluore lipase)均有较好的催化活性。同时考察各反应参数(温度、反应溶剂、体系水活度、酰化剂类型、肉桂醇与酰化剂摩尔比、肉桂醇浓度等)对脂肪酶Novozyme435合成乙酸肉桂酯反应的影响,确定了反应体系最优工艺条件:在10 mL甲基叔丁基醚中,肉桂醇200 mmol/L,n(肉桂醇)∶n(乙酸乙烯酯)=1∶1.5,初始水活度αw=0.84,温度35℃,酶加量0.02 g,反应3 h后肉桂醇转化率可达到99%,产物经质谱(MS)鉴定。固定化酶经过10个批次反应,反应转化率都保持在90%以上。