Enzymatic Interesterification Between Pine Seed Oil and a Hydrogenated Fat to Prepare Semi-Solid Fats Rich in Pinolenic Acid and Other Polyunsaturated Fatty Acids

Enzyme catalyzed interesterification (EIE) of pine seed oil (PSO) and a fully hydrogenated soybean oil (FHSBO) were studied in batch reactors in solvent-free media to prepare different semisolid fats rich in polyunsaturated fatty acids (PUFA). Optimal operation conditions found were: 10 % (w/w) enzyme loading, 75 °C and magnetic agitation at 300 rpm. Quasi-equilibrium conditions were reached after 2, 3 and 6 h, when immobilized lipases from Thermomyces lanuginosus (Lipozyme^® TL IM), Candida antarctica B. (Novozym^® 435) and Rhizomucor miehei (Lipozyme^® RM IM) from Novozymes A/S (Bagsvaerd, Denmark) were employed, respectively. Similar distributions of unsaturated to saturated fatty acid (UFA/SFA) residues along the glycerol backbone of the fat products were obtained with both non-selective and sn-1(3) regioselective lipases due to significant spontaneous acyl migration during the reaction. The products had higher UFA/SFA ratios at the sn-2 position (2.4–2.5, 1.4–1.7, and 0.5–0.8 for the trials involving 20, 40 and 70 % FHSBO, w/w, respectively) than the corresponding physical blends (0.8, 0.4 and 0.5, respectively). Fat products containing 3.1–11.6 % (w/w) pinolenic acid (Pn) and 16.1–35.7 % (w/w) linoleic acid (L) at the sn-2 position were prepared. The free acid contents of EIE products prepared with Lipozyme^® TL IM and Novozym^® 435 were 6.1–6.4 and 2.5–2.6, respectively. Residual activities of Lipozyme^® TL IM and Novozym^® 435 diminish by ca. 20 % after 9 reaction cycles.     

Application of Taguchi Method in the Enzymatic Modification of Menhaden Oil to Incorporate Capric Acid

Structured lipids (SL) were produced using menhaden oil and capric acid or ethyl caprate as the substrate. Enzymatic reaction conditions were optimized using the Taguchi method L9 orthogonal array with three substrate molar ratio levels of capric acid or ethyl caprate to menhaden oil (1:1, 2:1, and 3:1), three enzyme load levels (5, 10, and 15% [w/w]), three temperature levels (40, 50, and 60 °C), and three reaction times (12, 24, 36 hours). Recombinant lipase from Candida antarctica, Lipozyme^® 435, and sn‐1,3 specific Rhizomucor miehei lipase, Lipozyme^® RM IM (Novozymes North America, Inc., Franklinton, NC, USA), were used as biocatalysts in both acidolysis and interesterification reactions. Total and sn‐2 fatty acid compositions, triacylglycerol (TAG) molecular species, thermal behavior, and oxidative stability were compared. Optimal conditions for all reactions were 3:1 substrate molar ratio, 10% [w/w] enzyme load, 60 °C, and 16 hours reaction time. Reactions with ethyl caprate incorporated significantly more C10:0, at 30.76 ± 1.15 and 28.63 ± 2.37 mol% versus 19.50 ± 1.06 and 9.81 ± 1.51 mol%, respectively, for both Lipozyme^® 435 and Lipozyme^® RM IM, respectively. Reactions with ethyl caprate as substrate and Lipozyme^® 435 as biocatalyst produced more of the desired medium‐long‐medium (MLM)‐type TAGs with polyunsaturated fatty acids (PUFA) at sn‐2 and C10:0 at sn‐1,3 positions.     

Aspergillus oryzae Lipase-Catalyzed Synthesis of Dioleoyl; Palmitoyl-Rich Triacylglycerols in Two Reactors

Dioleoyl; palmitoyl‐rich triacylglycerols (OPO‐rich TAG) were synthesized through Aspergillus oryzae lipase (AOL)‐catalyzed acidolysis of palm stearin with commercial oleic acid by a one‐step process in a stirred tank reactor and continuous packed bed reactor to evaluate the feasibility of using immobilized AOL. AOL was found to be valuable for the synthesis of OPO‐rich TAG when compared with commercial lipase from Thermomyces lanuginose (Lipozyme^® TL IM; Novozymes A/S, Bagsvaerd, Denmark). The C52 (triglycerides with a carbon number of 52, stands for OPO, OPL, LPL and their isomers) content of AOL was higher (45.65 %), and the intensity of treatment (IOT: lipase amount × reaction time/TAG amount) of AOL was just 6.25 % of that of Lipozyme^® TL IM under similar reaction conditions in the stirred tank reactor. Response surface methodology were used to optimize the reaction conditions of the AOL‐catalyzed acidolysis is reaction in the packed bed reactor. The optimal point for the set of experimental conditions generated were as follows: residence time 3.0 h; temperature 62.09 °C; substrate molar ratio 7.13 mol/mol. The highest C52 content obtained was 48.60 ± 2.36 %, with 57.46 ± 1.72 % total palmitic acid at the sn‐2 position and 74.21 ± 2.45 % oleic acid at the sn‐1,3 positions. The half‐life of AOL was 24 h in the stirred tank reactor and 140 h in the packed bed reactor. The immobilized AOL achieved similar conversion and selectivity to commercial lipases for the catalyzed synthesis of OPO‐rich TAG and may offer a cheaper alternative.     

Enzymatic Synthesis of Infant Formula Fat Analog Enriched with Capric Acid

A structured lipid (SL) with a substantial amount of palmitic acid at the sn‐2 position and enriched with capric acid (C), was produced in two enzymatic interesterification stages by using immobilized lipase, Lipozyme^® TL IM (Novozymes North America Inc., Franklinton, NC, USA). The substrates for the reactions were high melting point palm stearin, high oleic sunflower oil and tricaprin. The SL was characterized for total and positional fatty acid profiles, triacylglycerol (TAG) molecular species, free fatty acid content, melting and crystallization profiles. The final SL contained 20.13 mol% of total palmitic acid, of which nearly 40 % was located at the sn‐2 position. The total capric acid content was 21.22 mol%, mostly at the sn‐1 and sn‐3 positions. The predominant TAGs in the SL were oleic–palmitic–oleic, POP and CLC. The melting completion and crystallization onset temperatures of the SL were 27.7 and 6.1 °C, respectively. The yield for the overall reaction was 90 wt%. This SL might be totally or partially used in commercial fat blends for infant formula.     

Application of lipases to the regioselective synthesis of sucrose fatty acid monoesters

A regioselective synthesis of 6′-O-acyl sucrose monoesters has been developed through the lipase-catalyzed esterification of sucrose acetals with fatty acids in both organic solvents and under solvent-free conditions. The products were obtained in overall yields of 20–27% after hydrolysis of the isopropylidene groups with aqueous acids. The strict selectivity of the enzymes used also enabled the preparation of a monoester fraction that was highly enriched in 6-O-acyl sucrose. This was accomplished by lipase-catalyzed transesterification of sucrose monoesters, prepared by conventional chemical methods, in propan-2-ol. After removal of the transesterification products (sucrose and fatty acid isopropyl esters) and column chromatography on silica gel, the obtained monoester product contained 80% of the single regioisomer, 6-O-acyl sucrose.     

Effects of Particle Size of Sucrose Suspensions and Pre-incubation of Enzymes on Lipase-Catalyzed Synthesis of Sucrose Oleic Acid Esters

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.     

Enzymatic production of human milk fat substitutes containing γ-linolenic acid: Optimization of reactions by response surface methodology

Structured lipids resembling human milk fat and containing GLA were synthesized by an enzymatic interesterification between tripalmitin, hazelnut oil FA, and GLA in n-hexane. Commercially immobilized 1,3-specific lipases, lipozyme^® RM IM and Lipozyme^® TL IM, were used as the biocatalysts. The effect of these enzymes on the incorporation levels was investigated. A central composite design with five levels and three factors—substrate ratio, reaction temperature, and time—were used to model and optimize the reaction conditions via response surface methodology. Good quadratic models were obtained for the incorporation of GLA (response 1) and oleic acid (response 2) by multiple regression and backward elimination. The determination coefficient (R ²) values for the models were found to be 0.92 and 0.94 for the reactions catalyzed by Lipozyme RM IM, and 0.92 and 0.88 for the reactions catalyzed by Lipozyme TL IM, respecitively. The optimal conditions generated from the models for the targeted GLA (10%) and oleic acid (45%) incorporation were 14.8 mol/mol, 55°C, and 24 h; 14 mol/mol, 55°C, and 24 h for substrate ratio (moles total FA/mol tripalmitin), temperature and time for the reactions catalyzed by Lipozyme RM IM and Lipozyme TL IM, respectively. Human milk fat substitutes containing GLA that can be included in infant formulas were success-fully produced using both Lipozyme RM IM and Lipozyme TL IM enzymes. The effect of the two enzymes on the incorporation of GLA and oleic acid were found to be similar.     

Lipase-Catalyzed Synthesis of Saccharide-Fatty Acid Esters Utilizing Solvent-Free Suspensions: Effect of Acyl Donors and Acceptors,and Enzyme Activity Retention

The effect of acyl donor (oleic, caprylic, lauric and myristic acids) and acceptor (fructose, sucrose, glucose and xylose) for synthesis of saccharide-fatty acid esters was conducted using solvent-free (50–200 μm sized) suspensions of saccharide crystals in a mixture of fatty acid/fructose oleate (90 wt% monoester and 10 wt% diester) at a ratio of 75/25 w/w initially, and a bioreactor system containing a packed bed bioreactor filled with immobilized Rhizomucor miehei lipase (Lipozyme^®IM, Novozymes, Franklinton, NC, USA) at 53 °C or a stirred tank bioreactor (STBR) at 65 °C. A nearly linear relationship between initial saccharide concentration and initial rate of reaction and final ester concentration was achieved which was independent of acyl donor or acceptor type. Slightly lower reaction rate and yield were obtained for operation in STBRs. Suspensions containing the highest saccharide concentration coincided with saccharide crystals of the smallest average size, since large-sized crystals sedimented out during the workup for formation of the suspensions. The best performance was achieved using fructose and oleic acid as substrates (92.3 wt% ester, consisting of 92 wt% monoester) in a packed-bed bioreactor (PBBR). The activity of RML did not decrease appreciably after four successive runs for solvent-free fructose-oleate esterification, or equivalently, a 22 day reaction period.     

Continuous lipase‐catalyzed synthesis of hexyl laurate in a packed‐bed reactor: optimization of the reaction conditions in a solvent‐free system

BACKGROUND: Hexyl laurate has been applied widely in cosmetic industries and is synthesized by chemical methods with problems of cost, environmental pollution, and by‐products. In this study, Lipozyme^® IM77 (from Rhizomucor miehei) was used to catalyze the direct‐esterification of hexanol and lauric acid in a solvent‐free system by utilizing a continuous packed‐bed reactor, wherein the aforementioned difficulties could be overcome. Response surface methodology (RSM) and three‐level‐three‐factor Box‐Behnken design were employed to evaluate the effects of synthesis parameters, such as reaction temperature (45–65 °C), mixture flow rate (0.25–0.75 mL min^?1) and concentration of lauric acid (100–300 mmol L^?1) on the production rate (µmol min^?1) of hexyl laurate by direct esterification. RESULTS: The production rate was affected significantly by the mixture flow rate and lauric acid concentration. On the basis of ridge‐max analysis, the optimum synthesis conditions for hexyl laurate were as follows: 81.58 ± 1.76 µmol min^?1 at 55 °C, 0.5 mL min^?1 flow rate and 0.3 mol L^?1 lauric acid. CONCLUSION: The lipase‐catalyzed synthesis of hexyl laurate by Lipozyme^® IM‐77 in a continuous packed‐bed bioreactor and solvent‐free system was successfully developed; optimization of the reaction parameters was obtained by Box–Behnken design and RSM. Copyright © 2008 Society of Chemical Industry     

Synthesis of Fatty Acid Ethyl Ester from Acid Oil in a Continuous Reactor via an Enzymatic Transesterification

Synthesis of a fatty acid ethyl ester via the lipase‐catalyzed transesterification of acid oil and ethanol was investigated in a continuous reactor. Lipozyme TL IM was employed as the immobilized lipase. This immobilized lipase derived from Thermomyces lanuginosus was purchased from Novozymes (Seoul, Korea). The acid oil was prepared by the acidification of soapstock formed as a by‐product during the refining of rice bran oil. The parameters investigated were water content, temperature, and molar ratio of substrates. The relative activity of Lipozyme TL IM was assessed during the repeated use of the immobilized lipase. The water content of the substrate had a considerable effect on the yield and the optimum water content was 4 %. The optimum temperature and molar ratio of acid oil to ethanol were 20 °C and 1:4, respectively. The maximum yield of approximately 92 % was achieved under the optimum conditions. The corresponding compositions were 92 % fatty acid ethyl esters, 3 % fatty acids, and 5 % acylglycerols. When glycerol formed during the reaction was removed by intermittent washing with ethanol, the relative activity of lipase was maintained over 82 % for a total usage of 27 cycles. For a mean residence time of 4 h, the half‐life times of Lipozyme TL IM on the control (unwashed) and treatment (washed) were 39 and 45 cycles, respectively.     

Identification of triacylglycerols in the enzymatic transesterification of rapeseed and butter oil

A blend of rapeseed and butter oil was transesterified using immobilized Thermomyces lanuginosus lipase (Lipozyme® TL IM) as catalyst. The reaction was followed by reversed-phase HPLC and the triacylglycerol peaks were tentatively identified from their elution times by calculating equivalent carbon numbers. Further identification was made using HPLC-electrospray tandem mass spectrometry. A few of the triacylglycerols detected were typical combinations of fatty acids originating from rapeseed oil, such as α-linolenic acid, and short-chain fatty acids from butter oil. Due to the regioselectivity of the lipase, the transesterification reaction involved mainly fatty acids in the sn-1 and sn-3 positions. However, significant changes in the fatty acid composition in the sn-2 position were detected after 6 h.     

Synthesis of Infant Formula Fat Analogs Enriched with DHA from Extra Virgin Olive Oil and Tripalmitin

Structured lipids (SLs) containing palmitic, oleic, and docosahexaenoic acids for possible use in infant formulas were synthesized by enzymatic acidolysis reactions. The substrates used were tripalmitin, extra virgin olive oil free fatty acids (EVOOFFA), and docosahexaenoic acid single cell oil free fatty acids (DHASCOFFA) in 1:1:1, 1:2:1, 1:3:2, 1:4:2, and 1:5:1 molar ratios. Reactions were carried out at 65 °C for 24 h using Lipozyme^® TL IM lipase. The products were analyzed for total and positional fatty acids by GC-FID, triacylglycerol (TAG) molecular species by HPLC-ELSD, and thermal behavior by DSC. The SLs, SL132, SL142, and SL151 had desirable fatty acid distribution for infant formula use with nearly 60 mol% palmitic acid at the sn-2 position and oleic acid predominantly at the sn-1,3 positions. The total DHA content of SL132, SL142, and SL151 were 7.54, 6.72, and 5.89 mol%, respectively. The major TAG molecular species in the SLs were PPP, OPO, and PPO. The melting completion temperature of SL132 was 37.1, 35.2 °C in SL142, and 32.9 °C in SL151. The SLs synthesized in this study have potential use in infant formulas.     

Production of Structured Triacylglycerol via Enzymatic Interesterification of Medium-Chain Triacylglycerol and Soybean Oil Using a Pilot-Scale Solvent-Free Packed Bed Reactor

Oils rich in medium- and long-chain triacylglycerols (MLCT) serve as functional oils to help reduce body fat accumulation and weight gain. However, most of the MLCT-rich products on the market are physical blends of medium- and long-chain triacylglycerols (MCT and LCT, respectively) that are not structured triacylglycerols (TAG). In this study, an efficient pilot-scale packed bed reactor (PBR) of immobilized lipase from Thermomyces lanuginosus (Lipozyme® TL IM, Novozymes, Bagsvaerd, Denmark) was employed for producing structured MLCT via 1,3-specific interesterification of TAG enriched in caprylic and capric acyl groups and soybean oil (SBO). The PBR was operated under continuous recirculation mode in the absence of solvent. Optimal reaction conditions were determined to be: caprylic/capric TAG: SBO ratio (45:55 w/w), reaction temperature (75 °C) and residence time (16.0 min) on MLCT production were studied. When employing a pilot-scale PBR (100 kg day⁻¹) under optimal conditions, a product containing 76.61 wt% MLCT was produced. Lipozyme TL IM was reused for 25 successive batch reactions (125 kg substrates) with no significant reduction in catalytic efficiency. The light yellow MLCT-enriched product had a high level of saturated fatty acids (SFA, 82.74 wt%) in its sn-2 position as a result of the enzyme's 1,3-positional specificity. One-stage molecular distillation and methanol extraction were used to remove the free fatty acids, mono-, and diacylglycerols generated from hydrolysis. With distillation temperature of 150 °C and oil-to-methanol ratio of 1:3 v/v, MLCT content was further increased to 80.07 wt%. The enzymatic PBR was therefore effective in producing structured MLCT at a pilot-scale under solvent-free conditions.     

Glucoside ester synthesis in microemulsions catalyzed by Candida antarctica component B lipase

The surfactant, ethyl 6-O-decanoyl glucoside, was synthesized in microemulsion systems by lipase catalysis. The microemulsions were based on the two substrates for the reaction, ethyl glucoside and fatty acid, and either the sodium salt of the fatty acid or the glucoside ester was used as surfactant. The lipase used was component B from Candida antarctica. Reduced pressure was employed to eliminate the water of condensation. The reaction yield was good, with conversion of fatty acid and ethyl glucoside reaching 77 and 96%, respectively.     

Synthesis and Properties of Ascorbyl Esters Catalyzed by Lipozyme TL IM using Triglycerides as Acyl Donors

Esters of l-ascorbic acid with long-chain fatty acids (E-304) are employed as antioxidants in foods rich in lipids. Although their enzymatic synthesis offers some advantages compared with the current chemical processes, most of the reported methods employ the immobilized lipase from Candida antarctica as biocatalyst and free fatty acids or activated esters as acyl donors. In order to diminish the cost of the process, we have investigated the synthesis of ascorbyl oleate and ascorbyl palmitate esters with the immobilized Thermomyces lanuginosus lipase Lipozyme TL IM—which is significantly less expensive than Novozym 435—and triglycerides as source of fatty acids. Lipozyme TL IM gave rise to a lower yield of 6-O-ascorbyl oleate than Novozym 435 when using triolein (64 vs. 84%) and olive oil (27 vs. 33%) as acyl donors. Both 6-O-ascorbyl oleate and 6-O-ascorbyl palmitate displayed excellent surfactant and antioxidant properties. The Trolox Equivalent Antioxidant Capability values for the oleate and palmitate were 71 and 84%, respectively, of those obtained with l-ascorbic acid; however, both derivatives were able to stabilize soybean oil towards peroxide formation.     

Production of Human Milk Fat Substitutes Catalyzed by a Heterologous Rhizopus oryzae Lipase and Commercial Lipases

This study aims to produce human milk fat substitutes by an acidolysis reaction between lard and the free fatty acids (FFA) from a fish oil concentrate rich in docosahexaenoic acid, in solvent-free media. The immobilized commercial lipases from (1) Rhizomucor miehei (Lipozyme RM IM), (2) Thermomyces lanuginosa (Lipozyme TL IM) and (3) Candida antarctica (Novozym 435) were tested as biocatalyst. Also, the heterologous Rhizopus oryzae lipase (rROL), immobilized in Accurel^® MP 1000, was tested as a feasible alternative to the commercial lipases. After 24 h of reaction at 50 °C, similar incorporations of polyunsaturated fatty acids (c.a. 17 mol%) were attained with Novozym 435, Lipozyme RM IM and rROL. The lowest incorporation was achieved with Lipozyme TL IM (7.2 mol%). Modeling acidolysis catalyzed by rROL and optimization of reaction conditions were performed by response surface methodology, as a function of the molar ratio FFA/lard and the temperature. The highest acidolysis activity was achieved at 40 °C at a molar ratio of 3:1, decreasing with both temperature and molar ratio. Operational stability studies for rROL in seven consecutive 24-h batches were carried out. After the fourth batch, the biocatalyst retained about 55 % of the original activity (half-life of 112 h).     

Optimization of the Solvent-Free Lipase-Catalyzed Synthesis of Fructose-Oleic Acid Ester Through Programming of Water Removal

Fructose oleate, an environmentally-friendly biobased surfactant, was prepared using solvent-free suspensions of saccharide in a mixture of acyl donor and monoester (the latter present at ≥5 wt% initially) continuously recirculated through a closed-loop packed bed bioreactor (PBBR)-based system at 53 °C, with the PBBR containing immobilized Rhizomucor miehei lipase (Lipozyme^®IM, Novozymes, Franklinton, NC, USA). To replenish the acyl acceptor consumed during the time course of reaction, the medium was isolated, fructose added, and a suspension formed by rigorous stirring at 80 °C for 6 h followed by centrifugation to remove larger particles, with the placement of the acyl acceptor replenishment treatments during the time course of reaction were optimized. Water removal via free evaporation was augmented during the latter portion of the time course (using a molecular sieve packed column, N₂ bubbling, vacuum pressure, or a combination of the latter two), with an optimal performance achieved when initiating N₂ + vacuum (\( 2. 1 6\,{\text{mg}}_{{{\text{H}}_{ 2} {\text{O}}}} \,{\text{h}}^{{^{ - 1} }} \) removal rate) upon reaching 60% ester, to maintain the liquid-phase water content near 0.40 wt%. When employing the above-mentioned conditions, 92.6 wt% fructose oleate was produced within 132 h, yielding a productivity of \( 0. 2 9 7\,{\text{mmol}}_{\text{Ester}} \,{\text{h}}^{ - 1} \, {\text{g}}_{\text{lipase}}^{ - 1} \).     

Designs of Bioreactor Systems for Solvent-Free Lipase-Catalyzed Synthesis of Fructose–Oleic Acid Esters

Fructose–oleic acid esters, biodegradable, biocompatible and biobased surfactants and value-added products were synthesized under solvent-free conditions at 65 °C in stirred-batch mode and using several different bioreactor systems. For a stirred-tank bioreactor (STBR) using fed-batch fructose addition and 5.0 wt.% immobilized Rhizomucor miehei lipase (Lipozyme^® IM, Novozymes, Franklinton, NC), the conversion yield was over 80%, and the initial rate of the reaction was comparable to previously obtained results using tert-butanol during the initial phase. The bioreactor systems contained a packed “desorption” column (DC) containing fructose crystals and silica gel for delivery of saccharide, and either a STBR or packed-bed bioreactor (PBBR). The liquid stream, initially containing oleic acid and a mixture of fructose–oleic acid esters at a ratio of 75/25 w/w, was continuously recirculated throughout the system. The PBBR system yielded the highest conversion (84.4%) and rate of reaction subsequent to the addition of 10 wt.% molecular sieves during the latter stage of reaction; however, the reaction rate was several-fold lower than the batch mode reactions due to the lower fructose concentrations provided by the DC.     

Optimization of the synthesis of lower glycerides rich in unsaturated fatty acid residues obtained via enzymatic ethanolysis of sesame oil

The lipases Novozym 435, Lipozyme TL IM and Lipozyme RM IM were employed in the production of lower acylglycerols (LG), i.e. mono‐ (MAG) and diacylglycerols (DAG), rich in unsaturated fatty acids from sesame oil in batch reactors. The effect of the molar ratio of ethanol to fatty acids on the reusability of these immobilized lipases was studied in detail. The effects of pretreatment on lipase activity for ethanolysis were investigated. Glycerol had a strong product inhibition effect on the ethanolysis reaction, and a relatively large excess of ethanol was necessary to remove the glycerol adsorbed on these biocatalysts. The enzymatic activity was drastically reduced by addition of water to the reaction medium. The presence of organic solvents (hexane and acetone) did not favor the production of LG. For the Novozym 435‐catalyzed reaction, optimum conditions were a molar ratio of ethanol to fatty acid residues of 5 : 1, 15 wt‐% lipase and 50 °C. For Lipozyme TL IM, the optimum conditions were a molar ratio of ethanol to fatty acid residues of 5 : 1, 20 wt‐% biocatalyst, and 30 °C. Novozym 435 and Lipozyme TL IM produced LG with molar ratios of unsaturated to saturated fatty acids of 20.4 in 1 h and 25.3 in 5 h, respectively. In the original oil, this ratio was 5. For trials conducted under optimum conditions, the products from the Novozym 435 trials contained 21.8 wt‐% triacylglycerols (TAG), 24 wt‐% DAG and 54.2 wt‐% MAG. The products of the Lipozyme TL IM trials consisted of 12.9 wt‐% DAG and 87.1 wt‐% MAG. No TAG species were detected.     

Production of margarine fats by enzymatic interesterification with silica-granulated Thermomyces lanuginosa lipase in a large-scale study

Interesterification of a blend of palm stearin and coconut oil (75∶25, w/w), catalyzed by an immobilized Thermomyces lanuginosa lipase by silica granulation, Lipozyme TL IM, was studied for production of margarine fats in a 1- or 300-kg pilot-scale batch-stirred tank reactor. Parameters and reusability were investigated. The comparison was carried out between enzymatic and chemical interesterified products. Experimentally, Lipozyme TL IM had similar activity to Lipozyme IM for the interesterification of the blend. Within the range of 55–80°C, temperature had little influence on the degree of interesterification for 6-h reaction, but it had slight impact on the content of free fatty acids (FFA). Drying of Lipozyme TL IM from water content 6 to 3% did not affect its activity, whereas it greatly reduced FFA and diacylglycerol contents in the products. Lipozyme TL IM was stable in the 1-kg scale reactor at least for 11 batches and the 300-kg pilot-scale reactor at least for nine batches. Due to regiospecificity of the lipase (sn-1,3 specific), enzymatically interesterified products had different fatty acid distribution at sn-2 position from the chemically randomized products, implying the potential nutritional benefits of the new technology. Presented at the 91st American Oil Chemists' Society Annual Meeting in San Diego, April 28, 2000.