Esterification of corn and sunflower acid oils with straight‐ and branched‐chain alcohols were conducted using lipase B from Candida antarctica (Novozym 435) in n‐hexane. Sunflower acid oil consisted of 55.6% free fatty acids and 24.7% triacylglycerols, while the free fatty acids and triacylglycerols contents of corn acid oil were 75.3% and 8.6%, respectively. After 1.5 h of methanolysis of sunflower acid oil, the highest fatty acid methyl ester content (63.6%) was obtained at 40 °C and the total fatty acid/methanol molar ratio was 1/1, using 15% enzyme based on acid oil weight. The conversion of both acid oils with straight‐ and branched‐chain alcohols was not significantly affected by the chain length of the alcohols. However, the lowest fatty acid methyl ester content (50%) was obtained in the reaction of corn acid oil with methanol. Sunflower acid oil was converted to fatty acid esters using primer alcohols such as n‐propanol, i‐ and n‐butanol, n‐amylalcohols, n‐octanol, and a mixture of amylalcohol isomers, resulting in a fatty acid ester content of about 70% at 40 °C. 相似文献
A number of model structures of the CalA suggested by comparative modeling were tested by site-directed mutagenesis. Enzyme variants were created where amino acids predicted to play key roles for the lipase activity in the different models were replaced by an inert amino acid (alanine). The results from activity measurements of the overproduced and purified mutant enzymes indicate a structure where the active site consists of amino acid residues Ser184, His366, and Asp334 and in which there is no lid. This model can be used for future targeted modifications of the enzyme to obtain new substrate acceptance, better thermostability, and higher enantioselectivity. 相似文献
The aim of this study is to pursue the identification and characterization of different CAL‐A variants displaying higher specificity toward erucic acid than CAL‐A wild type (wt). A careful analysis of the data generated from previously created site‐directed saturation libraries reveals several variants that display a higher preference for the hydrolysis of p‐nitrophenyl (pNP)‐erucate over pNP‐oleate than the wt. The best three candidates (CAL‐A V238D, V238Y, and V286N) are applied in biocatalysis using both Crambe oil and ethyl ester derivatives. When acting on Crambe oil, these CAL‐A variants are as efficient as CAL‐A wt in terms of C22:1 enrichment and product recovery independently of the temperature (enrichment and recovery values between 70–76% and 67–79% at 37 °C, and between 71–73% and 61–75% at 50 °C). In contrast, hydrolysis of Crambe ethyl esters leads to substantially increased accumulations of C22:1 and recovery values (V238Y: 78% enrichment and 92% recovery; V286N: 83% enrichment and 91% recovery) when using CAL‐A V238Y and CAL‐A V286N compared to CAL‐A wt (78% enrichment, 60% recovery) in the free fatty acid fraction. Practical Applications: This study describes the enhancement of lipase CAL‐A selectivity for the isolation and recovery of erucic acid (C22:1) from plant oil or its ethyl ester derivatives. Hence, this approach could represent a more eco‐friendly alternative for its application in processes where the erucic acid is used as building block, such as the production of surfactants or polymers. 相似文献
The effects of high-speed homogenization, high-intensity ultrasound, and their combination were evaluated for the reduction of the particle size of sucrose crystals to enhance solvent-free lipase-catalyzed synthesis of sucrose oleate at 65 °C. The combination of homogenization and ultrasound greatly decreased the particle size of suspended sucrose crystals in mixtures of oleic acid/sucrose oleate (86 wt% monoester and 14 wt% diester at a ratio of 90/10 w/w) from 88 to 18 μm. The suspension-based medium was charged to a stirred tank bioreactor that also contained immobilized lipase from Rhizomucor miehei or Candida antarctica (Lipozyme®IM and Novozym® 435, respectively; Novozymes, Franklinton, NC, USA), that was pre-incubated in oleic acid for several different temperatures (23–60 °C), durations (4–24 h), and stir rates (50–400 rpm, radius of 3 cm), prior to use. The optimal performance was achieved using C. antarctica lipase (83.3 wt% ester, consisting of 46 wt% monoester) in the presence of molecular sieves (18 wt%). The low water concentration (~0.12 wt%) did not affect the activity of C. antarctica lipase. 相似文献
Seven different reaction products were prepared via enzymatic interesterification of extra‐virgin olive oil (EVOO) and fully hydrogenated palm oil (FHPO), by varying the initial weight ratio of EVOO to FHPO from 80 : 20 to 20 : 80. The chemical, physical and functional properties of both the semi‐solid reaction products and the corresponding physical blends of the precursor starting materials were characterized. Fats prepared using large proportions of FHPO contained high levels of TAG species containing only saturated fatty acid residues. By contrast, high levels of TAG species containing both saturated and unsaturated fatty acid residues were found in fat products obtained with the lowest proportions of FHPO. Independently of the initial weight ratio of EVOO to FHPO, the interesterified products were characterized by a higher molar ratio of unsaturated to saturated fatty acid residues at the sn‐2 position, were softer over a wide temperature range, exhibited lower oxidative stabilities and were completely melted at lower temperatures than the corresponding physical blends. Potential applications of the reaction products range from margarines (highest weight ratios of EVOO to FHPO) to frying fats (lowest weight ratios of EVOO to FHPO). 相似文献
Candida antarctica lipase B (CALB) is one of the most extensively used biocatalysts in both academia and industry and exhibits remarkable (R)‐enantioselectivity for various chiral sec‐alcohols. Considering the significance of tailor‐made stereoselectivity in organic synthesis, a discovery of enantiocomplementary lipase mutants with high (R)‐ and (S)‐selectivity is valuable and highly desired. Herein, we report a highly efficient directed evolution strategy, using only 4 representative amino acids, namely, alanine (A), leucine (L), lysine (K), tryptophan (W) at each mutated site to create an extremely small library of CALB variants requiring notably less screening. The obtained best mutant with three mutations W104V/A281L/A282K displayed highly reversed (S)‐selectivity towards a series of sec‐alcohol with E values up to 115 (conv. 50%, ee 94%). Compared with the previously reported (S)‐selective CALB variant, W104A, a single mutation provided less selectivity, while the synergistic effects of three mutations in the best variant endow better (S)‐selectivity and a broader substrate scope than the W104A variant. Structural analysis and molecular dynamics simulation unveiled the source of reversed enantioselectivity.
A facile approach for the synthesis of 3,4‐dihydropyrimidin‐2(1H )‐ones was developed by a tandem multi‐component reaction (MCR) in one pot. This approach involves two steps, lipase‐catalyzed in situ generation of acetaldehyde and the Biginelli reaction in turn. Several control experiments were performed using acetaldehydes directly to explore the possible mechanism of this procedure. Moreover, owing to the distinct modularity and highly efficient features of the MCR, it can assemble libraries of structurally diverse products (yields up to 98% under the optimized conditions in this paper) and provides an exceptional synthesis tool for the discovery of the minimal deep‐blue luminogen in the solid state, namely, a single ring.
In the present study, Candida antarctica lipase B was immobilized on amine-functionalized silica microspheres as cross-linked enzyme aggregates (CLEA) and utilized for the biomanufacturing of rhamnolipids (RL). Lipase CLEA synthesized under optimized conditions of 2.0:1.0 by volume of silica microsphere/enzyme concentration, a 1.0:2.5 (v/v) ratio of enzyme/2-propanol, 7 mM glutaraldehyde concentration, when incubated at pH 9.0 and 40 °C, for a cross-linking time of 30 min were observed to exhibit superior biocatalytic properties and a maximum enzyme load of 770 U g−1. Lipase CLEA exhibited enhanced pH stability in acidic and alkaline media and increased temperature resistance as compared to free lipase. Both free and CLEA lipases were used to synthesize RL in different solvent systems. After 12 h, from initiation of the esterification, the degree of esterification (molar conversion yield) reached 46% and 71% in the batch mode. 1H and 13C nuclear magnetic resonance (NMR) and high-performance liquid chromatographic (HPLC) analysis confirm RL production by CLEA lipase. The CLEA showed greater confrontation to enzyme-mediated bioprocess approach as compared to its soluble counterpart and exhibited excellent RL production and catalytic activity even after its tenth successive reuse. 相似文献
A fully enzymatic methodology for the resolution of chiral amines has been demonstrated. Candida antarctica lipase B (CaLB)‐catalyzed acylation with N‐methyl‐ and N‐phenylglycine, as well as analogues having the general formula R1 X CH2CO2R2 (R1=Me, Ph; X=O, S) afforded the corresponding enantioenriched amides, which were subsequently enzymatically hydrolyzed. Surprisingly, CaLB also proved to be the catalyst of choice for this latter step. The heteroatom in the acyl donor profoundly influences both the enzymatic acylation and deacylation; the O‐substituted reagents performed best with regard to enantioselectivity as well as reaction rate in synthesis and hydrolysis. 相似文献
