Enzymatic synthesis optimization of isoamyl butyrate from fusel oil

Many food, cosmetic and pharmaceutical industries have increased their interest in short-chain esters due to their flavor properties. From the industrial standpoint, enzyme reactions are the most economical strategy to reach green products with neither toxicity nor damage to human health. Isoamyl butyrate (pear flavor) was synthesized by isoamyl alcohol (a byproduct of alcohol production) and butyric acid with the use of the immobilized lipase Lipozyme TL IM and hexane as solvents. Reaction variables (temperature, butyric acid concentration, isoamyl alcohol:butyric acid molar ratio and enzyme concentration) were investigated in ester conversion (%), concentration (mol L-1) and productivity (mmol ester g-1 mixture . h), by applying a sequential strategy of the Fractional Factorial Design (FFD) and the Central Composite Rotatable Design (CCRD). High isoamyl butyrate conversion of 95.8% was achieved at 24 hours. At 3 hours, the highest isoamyl butyrate concentration (1.64 mol L-1) and productivity (0.19 mmol ester g-1 mixture . h) were obtained under different reaction conditions. Due to high specificity and selectivity of lipases, process parameters of this study and their interaction with the Lipozyme TL IM are fundamental to understand and optimize the system so as to achieve maximum yield to scale up. Results show that fusel oil may be recycled by the green chemistry process proposed by this study.

Lipase-catalyzed transesterification of medium-long-medium structured lipid (MLM-SL) using palm olein and tricaprylin in packed-bed reactor (PBR)

Lipase-catalyzed transesterification between refined bleached deodorized palm olein (RBDO) and tricaprylin to produce medium-long-medium structured lipid (MLM-SL) in a packed bed reactor (PBR) has been investigated. A specific sn-1,3 commercial Lipozyme TL IM was used as biocatalyst. Within this study, the progress of transesterification was monitored especially for triacylglycerol (TAG) formation with equivalent carbon number (ECN) of 32, presumably 1,3-dicapryoyl-2-oleoyl-sn-glycerol (COC). Transesterification conditions investigated were residence times (i.e., 15, 30, and 60 min) and enzyme loadings (2.0 and 4.5 g). The highest yield of ECN 32 (13%) and transesterification degree (71%) were obtained at residence time of 15 mins for both enzyme loadings. Longer residence time seemed to facilitate lipid hydrolysis over transesterification. This was indicated by the number of peaks appearing in the high-performance liquid chromatography (HPLC) chromatograms and the reduction of fat slip melting point (SMP). Additionally, the highest productivity was obtained at 2.0 g enzyme loading. Conclusively, this study has demonstrated the potential use of packed-bed reactor with immobilized Lipozyme TL IM for continuous synthesis of MLM-SLs especially TAG with ECN32.

Synthesis of Dietetic Structured Lipids from Spent Coffee Grounds Crude Oil Catalyzed by Commercial Immobilized Lipases and Immobilized Rhizopus oryzae Lipase on Biochar and Hybrid Support

The aim of this study was the valorization of coffee industry residues, namely spent coffee grounds (SCG) as a source of oil, and silverskin (CS) as a source of both oil and biomass, under the concept of the circular economy. Therefore, crude oil from SCG was used to produce low-calorie structured lipids (SL) for food and pharmaceutical industries, and CS to produce biochar by pyrolysis for biotechnological uses. SL were obtained by acidolysis with caprylic or capric acid, or interesterification with ethyl caprylate or ethyl caprate, in solvent-free media, catalyzed by immobilized sn-1,3 regioselective lipases. Silverskin biochar (BIO) was directly used as enzyme carrier or to produce hybrid organic-silica (HB) supports for enzyme immobilization. Rhizopus oryzae lipase (ROL) immobilized on Amberlite (AMB), silica (SIL), BIO or HB, and the commercial immobilized Thermomyces lanuginosus (Lipozyme TL IM) and Rhizomucor miehei (Lipozyme RM IM) lipases were tested. Lipozyme RM IM showed better results in SL production than Lipozyme TLIM or ROL on BIO, SIL or HB. About 90% triacylglycerol conversion was attained after 7 h acidolysis or interesterification. Lipozyme RM IM was more stable in interesterification (80% and 65% activity with ethyl caprylate or ethyl caprate) than in acidolysis (first-order decay) after 10 reuses.

Lipase-Catalysed In Situ Transesterification of Waste Rapeseed Oil to Produce Diesel-Biodiesel Blends

Rapeseed oil of high acidity, an agricultural industry by-product unsuitable for food, was used as an inexpensive raw material for the production of biodiesel fuel. The use of rapeseed oil that is unsuitable for food and lipase as a catalyst makes the biodiesel production process environmentally friendly. Simultaneous oil extraction and in situ transesterification using diesel as an extraction solvent was investigated to obtain a diesel-biodiesel blend. The diesel and rapeseed oil blend ratio was 9:1 (w/w). The enzymatic production of biodiesel from rapeseed oil with high acidity and methanol using eleven different lipases as biocatalysts was studied. The most effective biocatalyst, lipase—Lipozyme TL IM (Thermomyces lanuginosus), which is suitable for in situ transesterification—was selected, and the conversion of rapeseed oil into fatty acid methyl ester was evaluated. The influence of the amount of methanol and lipase, the reaction temperature and the reaction time were investigated to achieve the highest degree of transesterification. The optimal reaction conditions, when the methanol to oil molar ratio was 5:1, were found to be a reaction time of 5 h, a reaction temperature of 25 °C and a lipase (Lipozyme TL IM) concentration of 5% (based on oil weight). Under these optimal conditions, 99.90% (w/w) of the rapeseed oil was extracted from the seed and transesterified. The degree of transesterification obtained was 98.76% (w/w). Additionally, the glyceride content in the biodiesel fuel was investigated and met the requirements perfectly.

The synthesis of medium-long-medium structured lipid (MLM-SL) by lipase-catalyzed transesterification using palm olein and tricaprylin in packed-bed reactor (PBR)

Lipase-catalyzed transesterification between refined bleached deodorized palm olein (RBDO) and tricaprylin to produce medium-long-medium structured lipid (MLM-SL) in a packed bed reactor has been investigated. A specific sn -1,3 commercial Lipozyme TL IM was used as biocatalyst. Within this study, the progress of transesterification was monitored especially for triacylglycerol (TAG) formation with equivalent carbon number (ECN) of 32, presumably 1,3-dicapryoyl-2-oleoyl- sn -glycerol (COC). Transesterification conditions investigated were residence times ( i.e ., 15, 30, and 60 min) and enzyme loadings (2 and 4.5 g). The highest yield of ECN 32 (13%) and transesterification degree (71%) were obtained at residence time of 15 mins for both enzyme loadings. Longer residence time seemed to facilitate lipid hydrolysis over transesterification. This was indicated by the number of peaks appearing in the high-performance liquid chromatography (HPLC) chromatograms and the reduction of of fat slip melting point (SMP). Additionally, at 4.5 enzyme loading the highest productivity was obtained for one-cycle reaction. Conclusively, this study has demonstrated the potential use of packed-bed reactor with immobilized Lipozyme TL IM for continuous synthesis of MLM-SLs especially TAG ECN32.

Incorporation of Caprylic Acid into a Docosahexaenoic Acid Single Cell Oil for the Production of Specialty Lipids

Research background. New sources of docosahexaenoic acid have recently been investigated aiming at infant formula fortification and dietary supplementation, among which the docosahexaenoic acid single cell oil, which contains 40-50 % of this acid. Experimental approach. For this purpose, such an oil was blended with caprylic acid in molar proportions ranging from 1:1 to 5:1 and the blends were interesterified using either Novozym 435 or Lipozyme TL IM as the catalyst. The influence of the amount of excess free caprylic acid in the substrate, as well as the type of enzyme on the triacylglycerol rearrangement resulting from the synthesis of the structured lipids were evaluated. Results and conclusions. The regiospecific lipase Lipozyme TL IM seemed to induce transesterification among single cell oil triacylglycerols preferably to acidolysis with caprylic acid, which was directly proportional to the ratio of this acid in the substrate. In reactions catalyzed by the non-regiospecific lipase Novozym 435, a higher incorporation of caprylic acid by single cell oil triacylglycerols was observed when compared with Lipozyme TL IM, independently of the oil:caprylic acid molar ratio. Novelty and scientific contribution. These results revealed the importance of combining the choice of the type of lipase, either regiospecific or not, with the proportions of excess free fatty acids in acidolysis reactions when aiming to produce structured lipids as a source of docosahexaenoic acid.

A sustainable innovation for the tandem synthesis of sugar-containing coumarin derivatives catalyzed by lipozyme TL IM from Thermomyces lanuginosus in continuous-flow microreactors

An effective and environmentally friendly two-step tandem protocol for the synthesis of sugar-containing coumarin derivatives catalyzed by lipozyme TL IM in continuous-flow microreactors has been developed.

Synthesis and Characterization of N-Methyl Fatty Hydroxamic Acids from Ketapang Seed Oil Catalyzed by Lipase

N-methyl fatty hydroxamic acid (N-MFHA), which is a derivative of hydroxamic acid (HA), was synthesized from ketapang seed oil (Terminalia catappa L.). In general, HAs have wide applications due to their chelating properties and biological activities. N-MFHAs were synthesized using immobilized lipase (Lipozyme TL IM) in biphasic medium which was the ketapang seed oil dissolved in hexane and N-methylhydroxylamine dissolved in water. The products were characterized through color testing and FT-IR spectroscopy after purification. Various factors affecting the enzyme activity investigated in the study included the effect of incubation time, the amount of lipase used, and the temperature. On the basis of the results, the optimum conditions for the synthesis of N-MFHA obtained are 25 h of incubation time, a temperature of 40 °C, and a ratio of 1:100 for the amount of enzyme (g)/oil (g). At the optimum conditions of the reaction, 59.7% of the oils were converted to N-MFHA.

Enzymatic Reetherification in the Production of Butterfat Substitutes

Enzymatic reetherification of fats has numerous technological and economic advantages, which makes its large-scale implementation highly efficient. Unlike chemical modification, enzymatic reetherification demonstrates a greater specificity, typical of the catalytic action of lipase, and a higher controllability. Lipases with positional specificity cause redistribution of fatty acids to occur only in extreme provisions of triglycerides. In addition, this method is 1.5 times lower than hydrogenation of fats. The authors used the facilities of an innovative laboratory provided by JSC Eurasian Foods Corporation to conduct practical research on reetherification of fatty mixes. The main objective was to study the effect of the fats obtained by fermental reetherification on the quality indicators of butterfat substitutes. The research featured the input products to be used in the formula of reetherified fat and prepared fat mixes for butterfat substitutes. The paper describes the process of enzymatic reetherification of mixes of oils and fats, prepared reesterified fats, and buttermilk substitutes obtained from reetherified fats. The process involved a sequence of reactors filled with Lipozyme TL IM, a granulated substance of a microbic 1.3-specific lipase. The lipase was obtained from Thermomyces Lanuginosus, which had been immobilized with silica gel. The obtained products conformed to the butterfat standards in that they contained 16–2% of polynonsaturated fatty acids, no transisomers of fatty acids, ≤ 38% of palmitiny acid, and ≤ 5% of solid triglycerides at 35 of °C. The melting temperature was under body heat. The resulting characteristics of butterfat substitutes make them high-quality dairy products.

The convenient Michael addition of imidazoles to acrylates catalyzed by Lipozyme TL IM from Thermomyces lanuginosus in a continuous flow microreactor

A fast and green Michael addition based protocol in a continuous flow microreactor was developed, an innovation which may open up the use of enzymatic microreactors in imidazole analogue biotransformations.