Use Of Lipase Research Articles

Application of Lipases for Obtaining Optically Active Organic Compounds

Among the diversity of biotechnological processes that are used in the chemical industry, particular attention is drawn to the production of chiral compounds, both as the target and intermediate products in the synthesis of drugs. This review highlights recent advances in the use of lipases for the production of optically active organic compounds.

Microbial cell factory for butyl butyrate production: Knowledges and perspectives.

Butyl butyrate is a short-chain fatty acid ester (C8) with a fruity aroma. It has broad prospects in the fields of foods, cosmetics and biofuels. At present, butyl butyrate is produced by chemical synthesis in the industry, but it is highly dependent on petroleum-based products. The growing concerns regarding the future scarcity of fossil fuels have been strongly promoted the transition from traditional fossil fuels and products to renewable bioenergy and biochemicals. Therefore, it is necessary to develop a green biochemical technology to replace traditional petroleum-based materials. In recent years, microorganisms such as Escherichia coli and Clostridium have been engineered to serve as cell factories for the sustainable one-pot production of short-chain fatty acid esters, including butyl butyrate. This opinion highlights the recent development in the use of lipases and alcohol acyltransferases (AATs) for butyl butyrate production in microbial fermentation, as well as future perspectives.

Use of hydroxyapatite as a support in the immobilization of Thermomyces lanuginosus lipase for application in the production of biodiesel using a by-product as lipid raw material.

The use of new materials in the field of biofuel production has been represented as a step in the development of remarkable catalysts. The use of lipases in the production of biodiesel is often seen as a cost-limiting step, as the operating expenses in recovering such catalysts can lead to unfeasible market expectations. In this study, hydroxyapatite (HAp) particles were evaluated as a support to immobilize commercial lipase, following application in ethyl ester synthesis. First, hydroxyapatite was synthesized through the co-precipitation method at constant pH and selected as a support to be used in enzyme immobilization. The characterization of the biocatalyst support materials produced was carried out using DRX, BET, FTIR, TGA, and SEM analysis. The lipase from Thermomyces lanuginosus was then immobilized in the matrices, and, subsequently, there was transesterification of the vegetable oil deodorization distillate (VODD). The biodiesel samples generated showed that they were within commercial standards, achieving ester conversion greater than 96.5%. Other properties such as density (0.87 g.cm-3) and viscosity (4.36 mm2.s-1) meet the specifications required by ASTM to be used as a biofuel. In the experiment planning technique, the results revealed an experimental trend and a defined behavior: a higher lipase loading in the immobilization and the use of temperatures in the range of 40-50 °C favor high conversions of ethyl esters. Thus, this confirms that the enzymatic chemical catalyst was able to form the main fatty acid esters even using a residual lipid raw material.

A New Approach in Lipase-Octyl-Agarose Biocatalysis of 2-Arylpropionic Acid Derivatives.

The use of lipase immobilized on an octyl-agarose support to obtain the optically pure enantiomers of chiral drugs in reactions carried out in organic solvents is a great challenge for chemical and pharmaceutical sciences. Therefore, it is extremely important to develop optimal procedures to achieve a high enantioselectivity of the biocatalysts in the organic medium. Our paper describes a new approach to biocatalysis performed in an organic solvent with the use of CALB-octyl-agarose support including the application of a polypropylene reactor, an appropriate buffer for immobilization (Tris base-pH 9, 100 mM), a drying step, and then the storage of immobilized lipases in a climatic chamber or a refrigerator. An immobilized lipase B from Candida antarctica (CALB) was used in the kinetic resolution of (R,S)-flurbiprofen by enantioselective esterification with methanol, reaching a high enantiomeric excess (eep = 89.6 ± 2.0%). As part of the immobilization optimization, the influence of different buffers was investigated. The effect of the reactor material and the reaction medium on the lipase activity was also studied. Moreover, the stability of the immobilized lipases: lipase from Candida rugosa (CRL) and CALB during storage in various temperature and humidity conditions (climatic chamber and refrigerator) was tested. The application of the immobilized CALB in a polypropylene reactor allowed for receiving over 9-fold higher conversion values compared to the results achieved when conducting the reaction in a glass reactor, as well as approximately 30-fold higher conversion values in comparison with free lipase. The good stability of the CALB-octyl-agarose support was demonstrated. After 7 days of storage in a climatic chamber or refrigerator (with protection from humidity) approximately 60% higher conversion values were obtained compared to the results observed for the immobilized form that had not been stored. The new approach involving the application of the CALB-octyl-agarose support for reactions performed in organic solvents indicates a significant role of the polymer reactor material being used in achieving high catalytic activity.

Application of Goat and Lamb Lipases on the Development of New Immobilized Biocatalysts Aiming at Fish Oil Hydrolysis.

The use of lipases from animal sources for the synthesis of new biocatalysts is barely studied in the literature. The present work focused on the immobilization of lipases from kid goat's and lamb's epiglottis in different ionic supports. For this, anionic supports (monoaminoethyl-N-aminoethyl-agarose (MANAE) and diethylaminoethyl-agarose (DEAE)) and cationic supports (carboxymethyl-agarose and sulfopropyl-agarose) were used. The immobilization parameters were evaluated, as well as the thermal stability of the immobilized enzymes and their stability at different values of pH. Then, the performance of the biocatalysts was evaluated in hydrolysis reactions for obtaining omega-3 fatty acids from fish oil (eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)). Values of 100% of recovered activity were obtained for lipase from goats, indicating that it was possible to maintain all the enzymatic activities of the immobilized enzymes on the supports. The immobilized enzymes were more stable in different pH conditions and at a temperature of 50°C, reaching values of stabilization factor of 12.17 and t1/2 of 9.86h-1, for lamb lipase immobilized in sulfopropyl agarose. In general, the anionic supports led to lower Km values and the cationic ones to a higher Vmax. Lamb lipase showed the highest selectivity values for EPA/DHA, reaching values of 6.43 using MANAE. Thus, the high potential for using such biocatalysts from animal sources in the food or pharmaceutical industries is observed.

A decade development of lipase catalysed synthesis of acylglycerols using reactors: a systematic review

Abstract Glycerol esterification produces a variety of valuable chemicals and has been a subject of great interest in recent years. Lipase-catalysed synthesis of acylglycerols reactions have many potential applications, including in the production of structured lipids, biofuels and pharmaceuticals. It offers several benefits over traditional chemical methods, including higher selectivity, milder reaction conditions and increased sustainability. In this systematic review, we summarize 52 primary research of lipase catalysed synthesis of acylglycerol published from 2012 to 2022. Different types of reactions employed in the synthesis of acylglycerols using various lipases are thoroughly discussed. The use of lipase in as biocatalyst in the synthesis of acylglycerols is safe for consumption and sustainable as it can reduce the utilization of toxic and hazardous organic solvents and chemicals that cause harm to human health and the environment. This review serves as a reference for the research and development of acylglycerols using biocatalysts and reactors. It could be an interest to the food and pharmaceutical industries or individuals who are interested to explore this field. Several limitations associated with the overall research on production of acylglycerols are presented in the conclusion section.

Use of Lipases as a Sustainable and Efficient Method for the Synthesis and Degradation of Polymers

Use of Lipases as a Sustainable and Efficient Method for the Synthesis and Degradation of Polymers

Effectiveness of different immobilized Candida antarctica lipase b (iCaLB) in the production of biobased oligoester derivatives from poly(butylene succinate) (PBS) and poly(butylene adipate) (PBA)

The use of lipases (Candida antarctica Lipase B, CaLB) immobilized on different solid supports for the preparation of biodegradable aliphatic polyesters poly(butylene adipate) (PBA) and poly(butylene succinate) (PBS) by esterification of unactivated dicarboxylic acids with 1,4-butanediol is presented. The immobilization of CaLB through absorption or a covalent attachment to different solid supports enhances their use as a catalyst for polycondensation reactions both in solution and in bulk. The results indicated that the different immobilized CaLBs were able to catalyze succesfully the polycondensation of both succinic and adipic acid with 1,4-butanediol, yielding the linear aliphatic polyesters PBS and PBA, respectively, by using a two-step polymerization process. Techniques such as nuclear magnetic resonance (NMR), relative molar mass obtained by gel permeation chromatography (GPC) and differential scanning calorimetry (DSC) were applied to compare the influence of the solid support on the obtention of PBA and PBS. Herein, it is shown that the nature of the solid support employed on each immobilized CaLB enzyme modifies the activity and effectivity towards polycondensation of PBA and PBS having a strong influence on the molecular weights and crystallinity.

Evaluation of Lipase from an Indigenous Isolated Bacillus Strain for Biodiesel Production

Lipases are utilized in biodiesel production utilizing various types of substrates. The use of lipase in bioenergy production aims to reduce energy crises and environmental pollution. Lipase-producing indigenous bacteria Bacillus licheniformis (Accession no. OP56979) and Bacillus rugosus (Accession no. OP56980) were isolated from various oil-contaminated sites. The isolated potential lipolytic bacteria were screened for maximum lipase production. Then, the bacteria showing the highest lipolytic activity were subjected to identification using the 16s rRNA technique while other isolated were identified biochemically. Lipase [LipBL-WII(c)] from Bacillus licheniformis having the highest lipolytic activity expressed various characteristics. Characterization of crude LipBL-WII(c) expressed that it showed stability in a wide range of pH (4 to 10) with optimum lipolytic activity observed at pH 8. It was then found to be active at a temperature range from 20°C to 80°C with optimal at 50°C. Lipase activity was also stimulated in metal ions such as Ca+1, Mg2+, and Zn2+ the most. Furthermore, LipBL-WII(c) retained lipolytic activity in the presence of various organic solvents and surfactants. The kinetic parameters (Km and Vmax) for LipBL-WII(c) were ascertained using Lineweaver- Burk plot. LipBL-WII(c) showed a potential for biodiesel production using olive oil as a source. Lipase gave 84% yield of biodiesel production from olive oil. Thus, it could be employed as a potential candidate for green biodiesel production using oil sources.

Trends in the Use of Lipases: A Systematic Review and Bibliometric Analysis.

Scientific mapping using bibliometric data network analysis was applied to analyze research works related to lipases and their industrial applications, evaluating the current state of research, challenges, and opportunities in the use of these biocatalysts, based on the evaluation of a large number of publications on the topic, allowing a comprehensive systematic data analysis, which had not yet been conducted in relation to studies specifically covering lipases and their industrial applications. Thus, studies involving lipase enzymes published from 2018 to 2022 were accessed from the Web of Science database. The extracted records result in the analysis of terms of bibliographic compatibility among the articles, co-occurrence of keywords, and co-citation of journals using the VOSviewer algorithm in the construction of bibliometric maps. This systematic review analysis of 357 documents, including original and review articles, revealed studies inspired by lipase enzymes in the research period, showing that the development of research, together with different areas of knowledge, presents good results related to the applications of lipases, due to information synchronization. Furthermore, this review showed the main challenges in lipase applications regarding increased production and operational stability; establishing well-defined evaluation criteria, such as cultivation conditions, activity, biocatalyst stability, type of support and reactor; thermodynamic studies; reuse cycles; and it can assist in defining goals for the development of successful large-scale applications, showing several points for improvement of future studies on lipase enzymes.

Lipase-catalyzed glycerolysis of technical lignin towards high-density polyurethane foams

The lignin can be applied in the synthesis of several products, including polymers. Nevertheless, its direct application can affect negatively the mechanical properties of the material, since it presents a highly cross-linked and complex structure. In this scenario, some chemical methods have been explored to increase the reactivity of lignin to produce of polyurethane. Although the literature reports methods intended the modification and incorporation of the lignin in the monomers production, to the best of our knowledge, there are no reports on the use of lipases in transesterification using this biomass. Thus, in this study, biopolyols were synthetized by enzyme-catalyzed transesterification using an ester (castor oil) and an alcohol (glycerol) in the presence of low-sulfur alkaline lignin as substrates, to produces a monomer. Then, high-density rigid foams were obtained via bulk polymerization. An increase in the apparent density and mechanical strength was observed with increasing lignin content. By Scanning Electron Microscopy (SEM) it was possible to observe more elongated and denser mixed cells, associated with the incorporation of lignin. Moreover, the compressive strength reached values of 0.14–0.52 MPa and polymer insoluble fraction were higher than 80 % for all the foams. Then, through this study, it was possible to obtain rigid and high density lignopolyurethane for industrial applications derived from enzymatic glycerolysis and renewable resources.

Lipase-catalyzed Production of Sugar Esters in Pressurized Fluid Media: A Review

Sugar esters (SE) are amphiphilic compounds containing hydrophilic moieties linked to hydrophobic parent structures with environmental-friendly features regarding non-toxicity, odorless, tasteless, and biodegradability as well as functional, emulsifying, stabilizing, dispersant, and detergent functions. The use of lipases as alternative biocatalysts to the alkaline compounds used industrially to produce SEs introduces interesting points to the process regarding energy consumption reduction, high selectivity, purer products production, and minimization of fossil-based solvents utilization. However, the usual organic solvents required in the reaction are harmful to enzymes, affecting their catalytic capacity. Accordingly, supercritical fluids (SCFs) are interesting options for the enzymatic process, especially carbon dioxide, which reaches such conditions under non-severe circumstances. SCFs have characteristics that make them attractive solvents for SEs production, allowing the manipulation of their properties to increase substrate solubility and reduce mass transfer limitations, intensifying process productivity. However, issues concerning with the strict operating conditions required by lipases and costs linked to project and equipments are shortcomings that must be considered. Therefore, this review gathers the main guidelines and opportunities that drive SEs production through enzymatic catalysis under supercritical conditions, as well as the challenges faced by this technology. Furthermore, technical aspects of properties and characteristics that justify the application of these compounds as well as considerations about how the main process parameters should be managed to enhance the substrates’ conversion are also discussed. Future perspectives indicate that the design of cost-effective bioreactors to improve performance as well as protein engineering to enhance lipases’ stability are points to be explored.

Bio-Approach for Obtaining Enantiomerically Pure Clopidogrel with the Use of Ionic Liquids.

Clopidogrel is a chiral compound widely used as an antiplatelet medication that lowers the risk of blood clots, strokes, and heart attacks. The main aim of the study presented herein was to obtain (S)-clopidogrel, which is commercially available in treatments, via the kinetic resolution of racemic clopidogrel carboxylic acid with the use of lipase from Candida rugosa and a two-phase reaction medium containing an ionic liquid. For this purpose, the enantioselective biotransformation of clopidogrel carboxylic acid and chiral chromatographic separation with the use of a UPLC-MS/MS system were optimized. The best kinetic resolution parameters were obtained by using a catalytic system containing lipase from Candida rugosa OF as a biocatalyst, cyclohexane and [EMIM][BF4] as a two-phase reaction medium, and methanol as an acyl acceptor. The enantiomeric excess of the product was eep = 94.21% ± 1.07 and the conversion was c = 49.60% ± 0.57%, whereas the enantioselectivity was E = 113.40 ± 1.29. The performed study proved the possibility of obtaining (S)-clopidogrel with the use of lipase as a biocatalyst and a two-phase reaction medium containing an ionic liquid, which is in parallel with green chemistry methodology and does not require environmentally harmful conditions.

Lipases and their Applications in Biomedical Field

Abstract: Lipases are carboxylic ester hydrolase enzymes, constituting the class of serine hydrolases, requiring no cofactor for their action. They have various substrates and produce glycerol and free fatty acids through the hydrolysis of fats and oils. Owing to their wide applications and the simplicity with which they can be mass-produced, these are a significant group of biotechnologically important enzymes. In addition, lipases have the special characteristic of operating at a lipid/ water interface. Present review focuses on the medical and therapeutic use of lipases. These enzymes and their inhibitors have applications with remarkable success for managing or even treating diseases such as cancer, obesity, atherosclerosis, and Alzheimer’s. Lipases have also been used to produce healthier fatty acids and low-fat cheese. The characteristic physicochemical and catalytic properties of enzymes make them ideal for biosensors and digestive aids in individuals with exocrine pancreatic inefficiency. Newer avenues open as deeper and more relevant studies are being conducted on newer lipases.

The Recent Advances in the Utility of Microbial Lipases: A Review

Lipases are versatile biocatalysts and are used in different bioconversion reactions. Microbial lipases are currently attracting a great amount of attention due to the rapid advancement of enzyme technology and its practical application in a variety of industrial processes. The current review provides updated information on the different sources of microbial lipases, such as fungi, bacteria, and yeast, their classical and modern purification techniques, including precipitation and chromatographic separation, the immunopurification technique, the reversed micellar system, aqueous two-phase system (ATPS), aqueous two-phase flotation (ATPF), and the use of microbial lipases in different industries, e.g., the food, textile, leather, cosmetics, paper, and detergent industries. Furthermore, the article provides a critical analysis of lipase-producing microbes, distinguished from the previously published reviews, and illustrates the use of lipases in biosensors, biodiesel production, and tea processing, and their role in bioremediation and racemization.

Resolution of Racemic Aryloxy-Propan-2-yl Acetates via Lipase-Catalyzed Hydrolysis: Preparation of Enantiomerically Pure/Enantioenriched Mexiletine Intermediates and Analogs

The lipase kinetic resolution (KR) of aryloxy-propan-2-yl acetates, via hydrolysis, produced enantiomerically pure/enantioenriched mexiletine intermediates and analogs. Racemic acetates rac-1-(2,6-dimethylphenoxy)propan-2-yl acetate (rac-5a), rac-1-(2,4-dimethylphenoxy)propan-2-yl acetate (rac-5b), rac-1-(o-tolyloxy)propan-2-yl acetate (rac-5c) and rac-1-(naphthalen-1-yloxy)propan-2-yl acetate (rac-5d) were used as substrates. A preliminary screening (24 h, phosphate buffer pH 7.0 with 20% acetonitrile as co-solvent, 30 °C and enzyme:substrate ratio of 2:1, m:m) was carried out with twelve lipases using acetate 5a as substrate. Two enzymes stood out in the KR of 5a, the Amano AK lipase from Pseudomonas fluorescens and lipase from Thermomyces lanuginosus (TLL) immobilized on Immobead 150. Under these conditions, both the (R)-1-(2,6-dimethylphenoxy)propan-2-ol [(R)-4a] and the remaining (S)-1-(2,6-dimethylphenoxy)propan-2-yl acetate [(S)-5a] were obtained with enantiomeric excess (ee) > 99%, 50% conversion and enantiomeric ratio (E) > 200. The KR study was expanded to racemic acetates 5b-d, leading to the corresponding chiral remaining acetates with ≥95% ee, and the alcohols 4b-d with ≥98% ee, and conversion values close to 50%. The best conditions for KRs of rac-5b-d involved the use of lipase from P. fluorescens or TLL immobilized on Immobead 150, 24 or 48 h and 30 °C. These intermediates had their absolute configurations determined using 1H NMR spectroscopy (Mosher’s method), showing that the KRs of these acetates obeyed the Kazlauskas’ rule. Molecular docking studies corroborated the experimental results, indicating a preference for the hydrolysis of (R)-5a-d.

An Appraisal on Prominent Industrial and Biotechnological Applications of Bacterial Lipases.

Microbial lipases expedite the hydrolysis and synthesis of long-chain acyl esters. They are highly significant commercial biocatalysts to biotechnologists and organic chemists. The market size of lipase is anticipated to reach $590 million by 2023. This is all owing to their versatility in properties, including stability in organic solvents, interfacial activation in micro-aqueous environments, high substrate specificity, and activity in even non-aqueous milieu. Lipases are omnipresent and synthesized by various living organisms, including animals, plants, and microorganisms. Microbial lipases are the preferred choice for industrial applications as they entail low production costs, higher yield independent of seasonal changes, easier purification practices, and are capable of being genetically modified. Microbial lipases are employed in several common industries, namely various food manufactories (dairy, bakery, flavor, and aroma enhancement, etc.), leather tanneries, paper and pulp, textiles, detergents, cosmetics, pharmaceuticals, biodiesel synthesis, bioremediation and waste treatment, and many more. In recent decades, circumspection toward eco-friendly and sustainable energy has led scientists to develop industrial mechanisms with lesser waste/effluent generation, minimal overall energy usage, and biocatalysts that can be synthesized using renewable, low-cost, and unconventional raw materials. However, there are still issues regarding the commercial use of lipases which make industrialists wary and sometimes even switch back to chemical catalysis. Industrially relevant lipase properties must be further optimized, analyzed, and explored to ensure their continuous successful utilization. This review comprehensively describes the general background, structural characteristics, classifications, thermostability, and various roles of bacterial lipases in important industries.

Preclinical safety evaluation of Lipase OF from Candida cylindracea.

Triacylglycerol lipases are enzymes commonly used to process foods and beverages such as oils, wines, and cheeses through catalyzation of long-chain triglyceride hydrolysis. Lipase OF derived from Candida cylindracea (strain MS-5-OF) is only intended for use as a processing aid in food production applications; however, it may be present at trace levels in some products. As such, the safety of Lipase OF was evaluated in this study that included a bacterial reverse mutation assay, an in vitro chromosome aberration test, and a 90-day subchronic toxicity study in rats. In the in vitro bacterial reverse mutation and chromosome aberration assays, Lipase OF was not observed to be mutagenic at concentrations up to 5000 μg/plate and 50 μg/ml, respectively, in the presence or absence of metabolic activation. Results from the 90-day subchronic toxicity study indicated only minimal adverse effects (i.e., increased platelet count and prothrombin time) in male rats from the high-dose group following administration of Lipase OF via the diet at levels of 0, 1.0, 2.5, and 5.0 w/w%. The no-observed-adverse-effect level (NOAEL) for Lipase OF was therefore considered 2.5 w/w% (1597.6mg/kg body weight/day [1027.3mg TOS/kg body weight/day]) in males and 5.0 w/w% (3700.4mg/kg body weight/day [2379.4mg TOS/kg body weight/day]) in females under the test conditions. Thus, the evidence presented within this study supports the safe use of Lipase OF as a processing aid in various food production applications for human consumption.

Lipase-mediated Baeyer–Villiger oxidation of benzylcyclopentanones in ester solvents and deep eutectic solvents

This work presents the chemo-enzymatic Baeyer–Villiger oxidation of α-benzylcyclopentanones in ester solvents as well as deep eutectic solvents (DES). In the first part of the work the effect of selected reaction conditions on the reaction rate was determined. The oxidation process was most effective in ethyl acetate at 55 °C, with the use of lipase B from Candida antarctica immobilized on acrylic resin and UHP as oxidant. Ultimately, these preliminary studies prompted the development of an effective method for the implementation of lipase-mediated Baeyer–Villiger oxidation of benzylcyclopentanones in DES. The highest conversion was indicated when the oxidizing agent was a component of DESs (minimal DESs). The fastest conversion of ketones to lactones was observed in a mixture of choline chloride with urea hydrogen peroxide. In this case, after 3 days, the conversion of the ketones to lactones products exceeded 92% for all substrates. As a result, two new lactones were obtained and fully characterized by spectroscopic data.

Design and development of a new static mixing bioreactor for enzymatic bioprocess: Application in biodiesel production

Mass transfer efficiency plays an important role in enzymatic biological processes, especially for non-aqueous biocatalytic processes. In these processes, low interphase mass transfer rates result in relatively long reaction residence time in traditional batch reactors. Therefore, the design and development of new bioreactors is crucial. In this study, a novel static mixing bioreactor was designed and applied to enzymatically catalyzed biodiesel production, which is a typical interfacial reaction system. In order to achieve good mixing effect and low pressure drop, the structure of bioreactor, including unit configuration, aspect ratio and rotation angle was designed and optimized by computational fluid dynamics. The realizable k-epsilon model was used, it was proved to be grid independent with 833678 hexahedral unstructured grids that were adopted to improve the structure for material mixing. After modification of the reactor, the final optimized structure is an R-R configuration with an aspect ratio of 1.5 and a rotation angle of 150°. Based on the above improved bioreactor, the process conditions for enzymatic production of biodiesel were optimized. The optimized reaction conditions were as follows: the length of the static mixer was 30 cm, the material flow rate was 1.77 m/s, the reaction temperature was 40 °C, the amount of methanol added was 3 times, and molar ratio of 3.4 alcohol to oil and 10% lipase dose were used. Under the optimal conditions, the yield of biodiesel in 1.5 h could reach 81.28%, and it could still be above 70% after 12 times of repeated use of lipase. Under the same reaction conditions, the reaction time for the static mixer to reach the same conversion rate (81.28%) was 1/3 of that of the stirred reactor, confirming the excellent mass transfer performance of the static mixer bioreactor.