Showing papers on "Lipase published in 1998"
TL;DR: Reactions based on lipase catalysis usually proceed with high regio- and enantioselectivity, and the Ca2+ antagonist diltiazem was obtained with lipase from Serratia marcescens.
Abstract: Unusually versatile substrate specificity is shown by lipases. Not only do they hydrolyze triacylglycerols-for example, in the stomach and intestine during digestion of dietary fat-and various synthetic esters and amides, but their high stability in organic solvents permits their use in transesterification reactions and ester synthesis as well. Reactions based on lipase catalysis usually proceed with high regio- and enantioselectivity. Thus, the Ca2+ antagonist diltiazem (1) was obtained with lipase from Serratia marcescens. Over 30 lipases have been cloned in the last few years. Since the tertiary structure of 12 lipases is known, there are presently significant efforts to improve this class of enzymes by protein engineering techniques, in view of their use in detergents and other fields of industrial application.
1,145 citations
TL;DR: The C. antarctica B-lipase is an example of an enzyme for which its specificity has been predicted based on the crystal structure and modeling of the active site region and a very close correlation is found.
Abstract: The application of the B-component lipase from the yeast Candida antarctica in organic synthesis is reviewed. This enzyme has been found to be a particularly efficient and robust lipase catalyzing a surprising diversity of reactions including many different regio- and enantio-selec-tive syntheses. Furthermore, the C. antarctica B-lipase is an example of an enzyme for which its specificity has been predicted based on the crystal structure and modeling of the active site region. This prediction is compared to experimental observations and a very close correlation is found.
642 citations
TL;DR: The model identifies the residues which mediate chain length specificity and thus may guide protein engineering of lipases for changed chain lengthspecificity and was supported by published experimental data on the chainlength specificity profile of various lipases and on mutants of fungal lipases with changed fatty acid chain length Specificity.
421 citations
TL;DR: A review of reaction systems employed in ester production catalyzed by lipases can be found in this paper, where water has a profound effect on the lipase behavior either directly by affecting the hydration of the enzyme or indirectly by changing the nature of the reaction media and enzyme support materials.
273 citations
TL;DR: Results strongly suggest that the enzyme purified from the hepatopancreas of red sea bream is homologous to mammalian bile salt-activated lipase.
Abstract: A lipase was purified from the extract of the delipidated powder of red sea bream hepatopancreas to nar homogeneity by fractional precipitation with ammonium sulfate and sequential chromatography on first anion-exchange-, hydrophobic- and second anion-exchange columns followed by gel filtration and anion-exchange HPLC. The final enzyme preparation showed a single band with an apparent molecular mass of approx. 64 kDa by sodium dodecyl sulfate-polyacrylamid e gel electrophoresis. The purified enzyme had a pH optimum in the range of pH 7.0–9.0. Using ρ-nitrophenyl myristate or triolein as a substrate, the enzyme required the presence of sodium taurocholate or sodium cholate for its activity. No activity was observed in the presence of sodium deoxycholate. The enzyme preferentially hydrolyzed ethyl esters of polyunsaturated fatty acid, such as arachidonic acid and eicosapentaenoic acid which were resistant to porcine pancreatic lipase. These results strongly suggest that the enzyme purified from the hepatopancreas of red sea bream is homologous to mammalian bile salt-activated lipase.
266 citations
TL;DR: The enzyme showed a 1,3-positional specificity toward triolein and was stable between pH 6 and 9, and the optimal pH for the enzymatic hydrolysis of tributyrin was around 8.2, while the enzyme was unstable at temperatures higher than 45°C.
Abstract: A psychrotrophic bacterium producing a cold-adapted lipase upon growth at low temperatures was isolated from Alaskan soil and identified as a Pseudomonas strain. The lipase gene (lipP) was cloned from the strain and sequenced. The amino acid sequence deduced from the nucleotide sequence of the gene (924 bp) corresponded to a protein of 308 amino acid residues with a molecular weight of 33,714. LipP also has consensus motifs conserved in other cold-adapted lipases, i.e., Lipase 2 from Antarctic Moraxella TA144 (G. Feller, M. Thiry, J. L. Arpigny, and C. Gerday, DNA Cell Biol. 10:381–388, 1991) and the mammalian hormone-sensitive lipase (D. Langin, H. Laurell, L. S. Holst, P. Belfrage, and C. Holm, Proc. Natl. Acad. Sci. USA 90:4897–4901, 1993): a pentapeptide, GDSAG, containing the putative active-site serine and an HG dipeptide. LipP was purified from an extract of recombinant Escherichia coli C600 cells harboring a plasmid coding for the lipP gene. The enzyme showed a 1,3-positional specificity toward triolein. p-Nitrophenyl esters of fatty acids with short to medium chains (C4 and C6) served as good substrates. The enzyme was stable between pH 6 and 9, and the optimal pH for the enzymatic hydrolysis of tributyrin was around 8. The activation energies for the hydrolysis of p-nitrophenyl butyrate and p-nitrophenyl laurate were determined to be 11.2 and 7.7 kcal/mol, respectively, in the temperature range 5 to 35°C. The enzyme was unstable at temperatures higher than 45°C. The Km of the enzyme for p-nitrophenyl butyrate increased with increases in the assay temperature. The enzyme was strongly inhibited by Zn2+, Cu2+, Fe3+, and Hg2+ but was not affected by phenylmethylsulfonyl fluoride and bis-nitrophenyl phosphate. Various water-miscible organic solvents, such as methanol and dimethyl sulfoxide, at concentrations of 0 to 30% (vol/vol) activated the enzyme.
250 citations
TL;DR: A method for the preparation of polyionic hydrogels from the complexation of chitosan and xanthan is reported, and electronic microscopy studies have shown the formation of a fibrilar structure.
186 citations
TL;DR: The gene coding for an extracellular lipase of Bacillus stearothermophilus L1 was cloned in Escherichia coli and showed highest activity toward p-nitrophenyl caprylate among the synthetic substrates and tripropionin among the triglycerides.
Abstract: The gene coding for an extracellular lipase of Bacillus stearothermophilus L1 was cloned in Escherichia coli. Sequence analysis showed an open reading frame of 1254 bp, which encodes a polypeptide of 417 amino acid residues. The polypeptide was composed of a signal sequence (29 amino acids) and a mature protein of 388 amino acids. An alanine replaces the first glycine in the conserved pentapeptide (Gly-X-Ser-X-Gly) around the active site serine. The expressed lipase was purified by hydrophobic interaction and ion exchange chromatography using buffers containing 0.02% (v/v) Triton X-100. The lipase was most active at 60-65°C and in alkaline conditions around pH 9-10. The lipase had highest activity toward p-nitrophenyl caprylate among the synthetic substrates and tripropionin among the triglycerides. It hydrolyzed beef tallow and palm oil more rapidly than olive oil at 50°C.
184 citations
TL;DR: The Aspergillus niger strain MTCC 2594 lipase was optimally active at pH 7·0 and 37°C, and was found to be stable between pH 4·0–10–0 and 4–50°C and it could be effectively used for the removal of triglyceride soils in laundry.
177 citations
TL;DR: The data suggest that genetic differences in hepatic lipase activity contribute to the differences in plasma HDL-C concentrations between African American men and white American men.
168 citations
TL;DR: The evidence presented here agrees with a more specific role for calcium ions, i.e., the formation of a new catalytically active enzyme complex, (enzyme)-(mixed micelle)-(calcium ion).
TL;DR: The 3D structure of the GPLRP2 chimera and the model of DolmI PLA1 reveal a higher hydrophilic/lipophilic balance (HLB) of the surface loops (beta5 loop, beta9 loop, lid domain) surrounding the active site, as compared to the homologous loops in HPL.
TL;DR: The discovery that lipases can also catalyze ester syntheses and transesterification reactions in organic solvent systems has opened up the possibility of enzyme catalyzed production of biodegradable polyesters.
TL;DR: In this paper, trimethylolpropane [2-ethyl-2-(hydroxymethyl)-1,3-propanediol] esters of rapeseed oil fatty acids were synthesized by transesterification with rapeseedoil methyl ester both by enzymatic and chemical means, both in bench and pilot scales.
Abstract: Biodegradable trimethylolpropane [2-ethyl-2-(hydroxymethyl)-1,3-propanediol] esters of rapeseed oil fatty acids were synthesized by transesterification with rapeseed oil methyl ester both by enzymatic and chemical means, both in bench and pilot scales. Nearly complete conversions were obtained with both techniques. A reduced pressure of about 2 to 5 kPa, to remove the methanol formed during transesterification, was critical for a high product yield. The quantity of added water was also critical in the biocatalysis. Candida rugosa lipase was used as biocatalyst and an alkaline catalyst in chemical transesterifications. In biocatalysis the maximum total conversion to trimethylolpropane esters of up to 98% was obtained at 42°C, 5.3 kPa, and 15% added water. The maximum conversion of about 70% to the tri-ester was obtained at the slightly higher temperature of 47°C. The reaction time was longer in the biocatalysis, but considerably higher temperatures were required in chemical synthesis. In the chemical synthesis tri-ester yields increased when the temperature was first held at 85 to 110°C for 2.5 h and subsequently increased to up to 120°C for 8 h. The trimethylolpropane esters obtained were tested as biodegradable hydraulic fluids and compared to commercially available hydraulic oils. The hydraulic fluids based on trimethylolpropane esters of rapeseed oil had good cold stability, friction and wear characteristics, and resistance against oxidation at elevated temperatures.
TL;DR: The enzyme, a 34 kDa monomeric protein, was demonstrated to be a B'-type carboxylesterase (EC 3.1.1) on the basis of substrate specificity and the action of inhibitors, and supported the previous suggestion of a catalytic triad made up of Ser-His-Asp.
Abstract: We previously purified a new esterase from the thermoacidophilic eubacterium Bacillus acidocaldarius whose N-terminal sequence corresponds to an open reading frame (ORF3) reported to show homology with the mammalian hormone-sensitive lipase (HSL)-like group of the esterase/lipase family. To compare the biochemical properties of this thermophilic enzyme with those of the homologous mesophilic and psychrophilic members of the HSL group, an overexpression system in Escherichia coli was established. The protein, expressed in soluble and active form at 10 mg/l E. coli culture, was purified to homogeneity and characterized biochemically. The enzyme, a 34 kDa monomeric protein, was demonstrated to be a B'-type carboxylesterase (EC 3.1.1.1) on the basis of substrate specificity and the action of inhibitors. Among the p-nitrophenyl (PNP) esters tested the best substrate was PNP-exanoate with Km and kcat values of 11+/-2 microM (mean+/-S.D., n=3) and 6610+/-880 s-1 (mean+/-S.D., n=3) respectively at 70 degreesC and pH7.1. In spite of relatively high sequence identity with the mammalian HSLs, the psychrophilic Moraxella TA144 lipase 2 and the human liver arylacetamide deacetylase, no lipase or amidase activity was detected. A series of substrates were tested for enantioselectivity. Substantial enantioselectivity was observed only in the resolution of (+/-)-3-bromo-5-(hydroxymethyl)-Delta2-isoxazoline, where the (R)-product was obtained with an 84% enantiomeric excess at 36% conversion. The enzyme was also able to synthesize acetyl esters when tested in vinyl acetate and toluene. Inactivation by diethylpyrocarbonate, diethyl-p-nitrophenyl phosphate, di-isopropylphosphofluoridate (DFP) and physostigmine, as well as labelling with [3H]DFP, supported our previous suggestion of a catalytic triad made up of Ser-His-Asp. The activity-stability-temperature relationship is discussed in relation to those of the homologous members of the HSL group.
TL;DR: The results show that lipase production using thermostable fungal cultures of Rhizomucor pusillus and Rhizopus rhizopodiformis compared favourably with most of the activities reported for other lipase hyperproducing microorganisms.
Abstract: Olive oil cake (OOC) from Morocco and sugar cane bagasse (SCB) were used for lipase production using thermostable fungal cultures of Rhizomucor pusillus and Rhizopus rhizopodiformis. The maximum production of lipase by Rhizomucor pusillus and Rhizopus rhizopodiformis in solid state fermentation (SSF) using SCB, was 4.99 U/g DM equivalent to 1.73 U/ml and 2.67 U/g DM equivalent to 0.97 U/ml, respectively. However, the mixture of OOC and SCB, 50% each, increased the lipase activity as high as 79.6 U/g DM equivalent to 43.04 U/ml and 20.24 U/g DM equivalent to 10.83 U/ml obtained by Rhizopus rhizopodiformis and Rhizomucor pusillus, respectively. These data compare favourably with most of the activities reported for other lipase hyperproducing microorganisms.
TL;DR: In this paper, the authors evaluated the regulation of lipoprotein lipase activity, protein mass, and messenger ribonucleic acid by estradiol and found that estrogen decreases lip-protein lipase activation by a post-transcriptional modification of protein levels.
TL;DR: To investigate the enantioselectivity of Pseudomonas cepacia lipase, inhibition studies were performed with Sc- and Rc-(Rp,Sp)-1,2-dialkylcarbamoylglycero-3-O-p-nitrophenyl alkylphosphonates of differentAlkyl chain lengths.
Abstract: To investigate the enantioselectivity of Pseudomonas cepacia lipase, inhibition studies were performed with S(c)-and R(c)-(R(p),S(p))-1,2-dialkylcarbamoylglycero-3-O-p-nitrophenyl alkylphosphonates of different alkyl chain lengths. P. cepacia lipase was most rapidly inactivated by R(c)-(R(p),S(p))-1,2-dioctylcarbamoylglycero-3-O-p-nitrophenyl octylphosphonate (R(c)-trioctyl) with an inactivation half-time of 75 min, while that for the S(c)-(R(p),S(p))-1,2-dioctylcarbamoylglycero-3-O-p-nitrophenyl octyl-phosphonate (S(c)-trioctyl) compound was 530 min. X-ray structures were obtained of P. cepacia lipase after reaction with R(c)-trioctyl to 0.29-nm resolution at pH 4 and covalently modified with R(c)-(R(p),S(p))-1,2-dibutylcarbamoylglycero-3-O-p-nitrophenyl butyl-phosphonate (R(c)-tributyl) to 0.175-nm resolution at pH 8.5. The three-dimensional structures reveal that both triacylglycerol analogues had reacted with the active-site Ser87, forming a covalent complex. The bound phosphorus atom shows the same chirality (S(p)) in both complexes despite the use of a racemic (R(p),S(p)) mixture at the phosphorus atom of the triacylglycerol analogues. In the structure of R(c)-tributyl-complexed P. cepacia lipase, the diacylglycerol moiety has been lost due to an aging reaction, and only the butyl phosphonate remains visible in the electron density. In the R(c)-trioctyl complex the complete inhibitor is clearly defined; it adopts a bent tuning fork conformation. Unambiguously, four binding pockets for the triacylglycerol could be detected: an oxyanion hole and three pockets which accommodate the sn-l, sn-2, and sn-3 fatty acid chains. Van der Waals' interactions are the main forces that keep the radyl groups of the triacylglycerol analogue in position and, in addition, a hydrogen bond to the carbonyl oxygen of the sn-2 chain contributes to fixing the position of the inhibitor.
TL;DR: In this article, the enantioselective esterification of ibuprofen (2-(4-iso-butylphenyl)propionic acid), catalyzed by Candida antarctica lipase (type B), was performed in various organic solvents.
TL;DR: In this article, the effects of oil/ethanol molar ratio, temperature, added water content, and amount of enzyme were analyzed for sunflower oil (SFO) with Lipozyme (immobilized 1,3-specific Mucor miehei lipase).
Abstract: The ethanolysis of sunflower oil (SFO) with Lipozyme (immobilized 1,3-specific Mucor miehei lipase) in a medium solely composed of substrates was investigated. The effects of oil/ethanol molar ratio, temperature, added water content, and amount of enzyme were analyzed. The optimal values were, respectively, 1:3, 50°C, 0% (vol/vol), and 0.4 g of Lipozyme per 5.7 mmol of SFO. The use of immobilized lipase made the reuse of enzyme feasible, and the enzyme could be recovered easily from the reaction mixture and recycled to reduce the cost of catalyst. In the last three consecutive runs of enzyme reuse, the final conversion yield of SFO from ethanolysis with added silica gel support was higher than that obtained from ethanolysis under standard conditions. The lipase-catalyzed alcoholytic reaction is potentially useful in the production of alkyl esters of specific interest.
TL;DR: Of seven lipases used to esterify functionalized phenols and fatty acids to synthesize lipophilic antioxidants, that from Candida antarctica lipase B (CAL-B) had the highest catalytic activity.
Abstract: Of seven lipases used to esterify functionalized phenols and fatty acids to synthesize lipophilic antioxidants, that from Candida antarctica lipase B (CAL-B) had the highest catalytic activity. Esterification yields of benzoic acid derivatives catalyzed by CAL-B are below 2% after 7 days. In contrast, relatively high yields were obstained for the esterification of (poly)phenols bearing primary hydroxy groups and aliphatic acids (85% yield within 15 hours). Crude products were purified and used as lipophilic antioxidants in sunflower oil.
TL;DR: Optimization of process selectivity considers the solubilities of the sugar and its monoesters in acetone at different temperatures, the percentage of this organic solvent in the reaction mixture, and the reaction temperature to select emulsifiers from glucose and fatty acids.
Abstract: Selective production of emulsifiers from glucose and fatty acids has been achieved using an immobilized Candida antarctica lipase. Optimization of process selectivity considers the solubilities of the sugar and its monoesters in acetone at different temperatures, the percentage of this organic solvent in the reaction mixture, and the reaction temperature. The solvent (acetone) is both easily eliminated and accepted by the European Community for use in the manufacture of foods and/or food additives. Different fatty acids with a longer length chain than that of caprylic acid may be employed. For saturated fatty acids longer than lauric acid, continuous precipitation of the monoester as it is formed at 40 degrees C permits nearly complete conversion (98%) of glucose to the monoester within 2-3 days. The procedure does not require total dissolution of the sugar, and precipitation of the monoester permits selective conversion of charges of glucose higher than 100 mg/mL solvent. A scaleup of the process under the optimum conditions gives high yields of 6-O-lauroyl glucose, which may be readily prepared on a gram scale. Copyright 1998 John Wiley & Sons, Inc.
TL;DR: The relationships between reaction time and water content are inverse and give a quantitative prediction of incorporation and acyl migration in selected reaction conditions and vice versa.
Abstract: Effects of water content, reaction time, and their relationships in the production of two types of specific-structured lipids (sn-MLM- and sn-LML-types: L-long chain fatty acids; M-medium chain fatty acids) by lipase-catalyzed interesterification in a solvent-free system were studied. The biocatalyst used was Lipozyme IM (commercial immobilized lipase). The substrates used for sn-MLM-type were fish oil and capric acid, and medium chain triacylglycerols and sunflower free fatty acids for sn-LML-type. The observed incorporation with the time course agrees well with the Michaelis-Menten equation, while the acyl migration is proportional to time within the range of 20 mol% acyl migration (MLM-type: M
f
=0.2225 T, R2=0.98; LML-type: M
f
=0.5618 T, R2=0.99). As water content (wt%, on the enzyme basis) increased from 3.0 to 11.6% for MLM-type and from 3.0 to 7.2% for LML-type in the solvent-free systems, the incorporation rates in the first 5 h increased from 3.34 to 10.30%/h, and from 7.29 to 11.12%/h, respectively. However, the acyl migration rates also increased from 0.22 to 1.12%/h and from 0.56 to 1.37%/h, respectively. Different effects in the production of two totally position-opposed lipids can be observed. Presumably these are caused by the different chain length of the fatty acids. The relationships between reaction time and water content are inverse and give a quantitative prediction of incorporation and acyl migration in selected reaction conditions and vice versa. The acyl migration can not be totally avoided in present systems, but can be reduced to a relatively low level. Acyl migration during the downstream processing has also been observed and other factors influencing the acyl migration are briefly discussed.
TL;DR: CC lipase appears to be most suitable for preparation of n-3 PUFA in the acylglycerol form from marine oils.
Abstract: Preparation of n-3 polyunsaturated fatty acid (PUFA) concentrates from seal blubber oil (SBO) and menhaden oil (MHO) in the form of acylglycerols was carried out by hydrolysis with a number of commercial microbial lipases. The lipases tested were Aspergillus niger, Candida cylindracea (CC), Chromobacterium viscosum, Geotrichum candidum, Mucor miehei, Pseudomonas sp., Rhizopus oryzae, and Rhizopus niveus. After lipase-assisted hydrolysis of oils, free fatty acids were removed, and fatty acid composition of the mixture containing mono-, di-, and triacylglycerols was determined. All lipases were effective in increasing the n-3 PUFA content of the remaining acylglycerols of both SBO and MHO. The highest concentration of n-3 PUFA was provided by CC lipase; 43.5% in SBO [9.75% eicosapentaenoic acid (EPA), 8.61% docosapentaenoic acid (DPA), and 24.0% docosahexaenoic acid (DHA)] and 44.1% in MHO (18.5% EPA, 3.62% DPA, and 17.3% DHA) after 40 h of hydrolysis. Thus, CC lipase appears to be most suitable for preparation of n-3 PUFA in the acylglycerol form from marine oils.
Journal Article•
TL;DR: It is suggested that structural gene defects are also present in CESD cells, however, the reason (s) for the clinical difference between Wolman disease and CESD remain (S) to be studied.
Abstract: Wolman disease and cholesteryl ester storage disease (CESD) are caused by a deficiency of lysosomal acid lipase activity, resulting in massive accumulation of cholesteryl ester and triglycerides. Wolman disease occurs in infancy, with hepatosplenomegaly, steatorrhea and adrenal calcification. It is fatal before the age of 1 year. In CESD, hepatomegaly may be the only clinical abnormality, although lipid deposition is widespread. Lysosomal acid lipase hydrolyzes both triaclyglycerols and cholesteryl esters, and the enzyme plays an important role in the cellular processing of plasma lipoproteins, and contributes to homeostatic control of lipoprotein levels in blood and prevention of cellular lipid overloading. The gene encoding lysosomal acid lipase was cloned and characterized in 1994, and two mutations of acid lipase gene were found in a patient with Wolman disease, as a compound heterozygote. It is suggested that structural gene defects are also present in CESD cells. However, the reason (s) for the clinical difference between Wolman disease and CESD remain (s) to be studied.
TL;DR: In this paper, the authors showed that the simultaneous entrapment of a lipase such as Pseudomonas cepacia or Candida antarctica and nanostructured superparamagnetic magnetite (Fe3O4) in hydrophobic sol-gel materials derived from CH3Si(OCH3)3 (MTMOS) or other hydrophilic precursors leads to catalytically active, mechanically stable and magnetically separable heterogeneous biocatalysts.
Abstract: The simultaneous entrapment of a lipase such as Pseudomonas cepacia or Candida antarctica and nanostructured superparamagnetic magnetite (Fe3O4) in hydrophobic sol–gel materials derived from CH3Si(OCH3)3 (MTMOS) or other hydrophobic precursors leads to catalytically active, mechanically stable and magnetically separable heterogeneous biocatalysts. The relative enzyme activity in the test reaction involving the esterification of lauric acid by n-octanol in isooctane is typically 200–300%, with respect to the same reaction using a conventional suspension of the non-immobilized enzyme. Separation of the catalyst by applying a simple magnet poses no problems. In the kinetic resolution of 2-pentylamine, enantioselectivity is essentially complete (ee=97–99%).
TL;DR: External mass transfer was not a major problem in the linear flow range, but internal mass transfer did impose some transfer limitations, and residence time, water content, and effects of mass transfers were also investigated.
Abstract: Production of specific-structured lipids (interesterified lipids with a specific structure) by enzymatic interesterification was carried out in a continuous enzyme bed pilot scale reactor. Commercial immobilized lipase (Lipozyme IM) was used and investigations of acyl migration, pressure drop, water dependence, production efficiency, and other basic features of the process were performed. The extent of acyl migration (defined as a side reaction) occurring in the present enzyme bed reactor was compared to that in a pilot batch reactor. The continuous enzyme bed reactor was better than the batch reactor in minimizing acyl migration. Generally the former produced about one-fourth the acyl migration produced by the latter at a similar extent of incorporation. Pressure drop and production efficiency were evaluated in order to obtain a suitable yield in one reaction step. High incorporation was favored by high substrate ratios between acyl donors and oils, requiring long reaction times on the enzyme bed. Under these conditions, the pressure drop of the reactor was modeled statistically and theoretically. Residence time, water content, and effects of mass transfers were also investigated. Incorporation of medium-chain fatty acids increased with increased residence time. Approximately 40% of lipase activity was lost after a 4-wk run. External mass transfer was not a major problem in the linear flow range, but internal mass transfer did impose some transfer limitations.
TL;DR: Direct evolution based on error prone PCR, the enantioselectivity of the hydrolysis of a chiral ester, catalyzed by the lipase from P. aeruginosa, can be increased from ee 2 to ee 81% in just four mutagenesis cycles.
TL;DR: Structured triacylglycerides with medium-chain fatty acids (caprylic acid) in sn1- and sn3-positions and a long-chain unsaturated fatty acid in the sn2-position of glycerol (MLM) were synthesized by lipase catalysis in a two-step process.
Abstract: Structured triacylglycerides with medium-chain fatty acids (caprylic acid) in sn1- and sn3-positions and a long-chain unsaturated fatty acid (oleic or linoleic acid) in the sn2-position of glycerol (MLM) were synthesized by lipase catalysis in a two-step process. First, pure 2-monoacylglycerides (2-MG) were synthesized by alcoholysis of triacylglycerides (triolein, trilinolein, or peanut oil) in organic solvents with 1,3-regiospecific lipases (from Rhizomucor miehei, Rhizopus delemar, and Rhizopus javanicus). The 2-MG were purified by crystallization and obtained in up to 71.8% yield. These 2-MG were esterified in a second reaction with caprylic acid in n-hexane to form almost pure MLM. For 2-MG obtained from peanut oil, the final product contained more than 90% caprylic acid in the sn1- and sn3-positions, whereas the sn2-position was composed of 98.5% unsaturated long-chain fatty acids. Reaction conditions for both steps were optimized with respect to source and immobilization of lipase, water activity, and solvent.