Nonspecific Lipase Research Articles

Combined cross-linking of Rhizomucor miehei lipase and Candida antarctica lipase B for the effective enrichment of omega-3 fatty acids in fish oil

The incorporation of a non-specific lipase and a sn-1,3 specific one in a single immobilized system can be a promising approach for the exploitation of both lipases. A one-step immobilization platform mediated by an isocyanide-based multi-component reaction was applied to create co-cross-linked enzymes (co-CLEs) of lipases from Rhizomucor miehei (sn-1,3 specific) and Candida antarctica (non-specific). Glutaraldehyde was found to be effective cross-linker by producing specific activity of 16.9 U/mg and immobilization yield of 99 %. High activity recovery of up to 404 % was obtained for immobilized derivatives. Leaking experiment showed covalent nature of the cross-linking processes. BSA had considerable effect on the immobilization process, providing 87–100 % immobilization yields and up to 10 times improvement in the specific activity of the immobilized derivatives. Scanning electron microscopy images showed flower-like and rod-like structures for the CLEs prepared by glutaraldehyde and undecanedicarboxylic acid, respectively. The prepared co-CLEs were examined in non-selective enrichment of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) from fish oil, showing capability of releasing up to 100 % of both omega-3 fatty acids within 8 h of the reaction. The reusability of co-CLEs in five successive cycles presented retaining 63–72 % of their initial activities after the fifth reuse cycle in the hydrolysis reaction.

Design of a New Chemoenzymatic Process for Producing Epoxidized Monoalkyl Esters from Used Soybean Cooking Oil and Fusel Oil

The aim of this study was to produce epoxidized monoalkyl esters (EMAE), a valuable class of oleochemicals used in a wide range of products and industries, from used soybean cooking oil (USCO) and fusel oil via a three-step chemoenzymatic process. This process consists of a first enzymatic hydrolysis of USCO to produce free fatty acids (FFA). Here, five microbial lipases with different specificities were tested as biocatalysts. Full hydrolysis of USCO was obtained after a 180 min reaction time under vigorous stirring (1500 rpm) using a non-specific lipase from Candida rugosa (CRL). Then, monoalkyl esters (MAE) were produced via the esterification of FFA and fusel oil in a solvent-free system using the lipase Eversa® Transform 2.0 (ET2.0) immobilized via physical adsorption on poly(styrenene-divinylbenzene) (PSty-DVB) beads as a biocatalyst. Different water removal strategies (closed and open reactors in the presence or absence of molecular sieves at 5% m.m−1) on the reaction were evaluated. Maximum FFA conversions of 64.3 ± 2.3% (open reactor after a 30 min reaction time) and 73.5 ± 0.4% (closed reactor after a 45 min reaction time) were observed at 40 °C, using a stoichiometric FFA:fusel oil molar ratio (1:1), without molecular sieves, and 5 mg of immobilized protein per gram of reaction mixture. Under these conditions, maximum FFA conversion was only 30.2 ± 2.7% after a 210 min reaction time in a closed reactor using soluble lipase. Reusability tests showed better retention of the original activity of immobilized ET2.0 (around 82%) after eight successive batches of esterification reactions conducted in an open reactor. Finally, the produced MAE was epoxidized via the Prilezhaev reaction, a classical chemical epoxidation process, using hydrogen peroxide and formic acid as a homogeneous catalyst. The products were characterized by standard methods and identified using proton nuclear magnetic resonance (1H NMR). Maximum unsaturated bond conversions into epoxy groups were at approximately 33%, with the experimental epoxy oxygen content (OOCexp.) at 1.75–1.78%, and selectivity (S) at 0.81, using both MAEs produced (open or closed reactors). These results show that this new process is a promising approach for value-added oleochemical production from low-cost and renewable raw materials.

Fatty acid production from coffee waste

The valorization of coffee processing waste by producing products with high added value is a promising approach toward a circular bioeconomy. Crude coffee silverskin oil can be directly used in enzymatic hydrolysis catalyzed by non-specific lipases or sn-1,3 regioselective lipases. Converting coffee silverskin oil into free fatty acids or enriched polyunsaturated fatty acids in the form of acylglycerols with promoting properties can reduce the environmental impact and increase the profitability of the coffee industry.

Lipase catalysed transesterification of palm stearin with ferulic acid in solvent-free media

Recently, several scientific groups investigated the suitable enzymatic route for synthesis of structured phenolic lipids for cosmetic and nutritive purposes. The feruloylated palm stearin (FPS) was obtained from enzymatic transesterification reactions of palm stearin with ferulic acid in solvent-free media with 1 mmol of glycerol or without it, using non-specific lipases, NovoCor ADL (Candida antarctica lipase A); Novozyme435® (Candida antarctica lipase B), and 1,3-specific lipase Lipozyme TL 100L from Thermomyces lanuginosus. The individual lipases and their combinational systems were examined. ESI+-MS/MS confirmed the formation of 19 feruloylated compounds. FPSs were produced by all lipases and their combinations. The positive synergetic effect occurred for 1,3-specific lipase and unspecified lipases with mass ratio 8:2, respectively. The antioxidant activities of all FPSs were examined. FPS produced by NovoCor ADL lipase in presence of free glycerol showed an antioxidant activity 10.3-times higher. In conclusion, the two low-cost lipases NovoCor ADL and Lipozyme TL 100L and its combi-lipase system could be considered suitable low-cost alternative to the expensive wide use N435 lipase in the solvent free production process of FPS for food and cosmetic applications.

Lipase from Rhizopus oryzae R1: in-depth characterization, immobilization, and evaluation in biodiesel production

BackgroundRhizopus species is among the most well-known lipase producers, and its enzyme is suitable for use in many industrial applications. Our research focuses on the production of lipase utilizing waste besides evaluating its applications. ResultsAn extracellular lipase was partially purified from the culture broth of Rhizopus oryzae R1 isolate to apparent homogeneity using ammonium sulfate precipitation followed by desalting via dialysis. The partially purified enzyme was non-specific lipase and the utmost activity was recorded at pH 6, 40 °C with high stability for 30 min. The constants Km and Vmax, calculated from the Lineweaver-Burk plot, are 0.3 mg/mL and 208.3 U/mL, respectively. Monovalent metal ions such as Na+ (1 and 5 mM) and K+ (5 mM) were promoters of the lipase to enhance its activity with 110, 105.5, and 106.5%, respectively. Chitosan was used as a perfect support for immobilization via both adsorption and cross-linking in which the latter method attained immobilization efficiency of 99.1% and reusability of 12 cycles. The partially purified enzyme proved its ability in forming methyl oleate (biodiesel) through the esterification of oleic acid and transesterification of olive oil. ConclusionThe partially purified and immobilized lipase from Rhizopus oryzae R1 approved excellent efficiency, reusability, and a remarkable role in detergents and biodiesel production.

Immobilized Lipase in the Synthesis of High Purity Medium Chain Diacylglycerols Using a Bubble Column Reactor: Characterization and Application.

Novozym® 435, an immobilized lipase from Candida antarctica B. (CALB), was used as a biocatalyst for the synthesis of high purity medium chain diacylglycerol (MCD) in a bubble column reactor. In this work, the properties of the MCD produced were characterized followed by determining its practical application as an emulsifier in water-in-oil (W/O) emulsion. Two types of MCDs, namely, dicaprylin (C8-DAG) and dicaprin (C10-DAG), were prepared through enzymatic esterification using the following conditions: 5% Novozym® 435, 2.5% deionized water, 60°C for 30 min followed by purification. A single-step molecular distillation (MD) (100–140°C, 0.1 Pa, 300 rpm) was performed and comparison was made to that of a double-step purification with MD followed by silica gel column chromatography technique (MD + SGCC). Crude C8-DAG and C10-DAG with DAG concentration of 41 and 44%, respectively, were obtained via the immobilized enzyme catalyzing reaction. Post-purification via MD, the concentrations of C8-DAG and C10-DAG were increased to 80 and 83%, respectively. Both MCDs had purity of 99% after the MD + SGCC purification step. Although Novozym® 435 is a non-specific lipase, higher ratios of 1,3-DAG to 1,2-DAG were acquired. Via MD, the ratios of 1,3-DAG to 1,2-DAG in C8-DAG and C10-DAG were 5.8:1 and 7.3:1, respectively. MCDs that were purified using MD + SGCC were found to contain 1,3-DAG to 1,2-DAG ratios of 8.8:1 and 9.8:1 in C8-DAG and C10-DAG, respectively. The crystallization and melting peaks were shifted to higher temperature regions as the purity of the MCD was increased. Dense needle-like crystals were observed in MCDs with high purities. Addition of 5% C8-DAG and C10-DAG as emulsifier together in the presence of 9% of hydrogenated soybean oil produced stable W/O emulsion with particle size of 18 and 10 μm, respectively.

Co-immobilization of Rhizomucor miehei lipase and Candida antarctica lipase B and optimization of biocatalytic biodiesel production from palm oil using response surface methodology

Lipases from Candida antarctica B (nonspecific lipase) and Rhizomucor miehei (1,3-specific lipase) were simultaneously immobilized on epoxy functionalized silica gel under mild conditions. The results showed rapid and simple immobilization of 4–15 mg of CALB:RML (different ratios 4:1, 2:1, 1.5:1, 1:1) on 1 g of support after 6 h. The thermal stability of derivatives and also their stability in methanol were greatly improved compared to the single immobilized enzyme. All the derivatives were also used to catalyze the transesterification of palm oil with methanol to produce fatty acid methyl esters (FAMEs). Response surface methodology (RSM) and a central composite rotatable design (CCRD) was used to study the effects of five factors, reaction temperature, methanol/oil ratio, reaction time, t-butanol concentration and CALB:RML ratio on the fatty acid methyl esters (FAME) yield. A quadratic polynomial equation was obtained for methanolysis reaction by multiple regression analysis. The optimum combinations for the reaction were CALB:RML ratio (2.5:1), t-butanol to oil (39.9 wt%), temperature (35.6 °C), methanol:oil ratio (5.9), reaction time 33.5 h. FAME yield of 78.3.5%, which was very close to the predicted value of 75.2%, was obtained. Verification experiment confirmed the validity of the predicted model.

Prediabetes Directly Deteriorates into Diabetic Ketoacidosis and Hyperosmolar Hyperglycemic Syndrome Triggered by Acute Pancreatitis: A Case Report Illustrating a “Chicken and Egg” Paradigm in Ketosis-Prone Diabetes

IntroductionDiabetic crises occur most often in patients with type 1 diabetes and occasionally in type 2 diabetes, especially under stressful conditions. However, a diabetic crisis occurring directly from prediabetes is an unusual phenomenon.Case ReportA 45-year-old woman presented with postprandial left upper quadrant abdominal pain, nausea, and vomiting. She had a past medical history of prediabetes with impaired fasting glucose and HbA1c 6.4%. On admission, routine laboratory tests showed high anion gap metabolic acidosis (pH 6.92), anion gap 41 mmol/L, blood glucose 931 mg/dL, beta-hydroxybutyrate 28 mmol/L, and calculated effective osmolarity 322 mOsm/kg; she was diagnosed with diabetic ketoacidosis (DKA) and hyperosmolar hyperglycemic syndrome (HHS), and DKA-related abdominal pain. Later, the patient was found to have elevated lipase and amylase, and diagnosed with acute pancreatitis. Since DKA can induce abdominal pain and nonspecific lipase elevation, both of which are characteristics of acute pancreatitis, while acute pancreatitis can conversely trigger DKA, there exists a “chicken and egg” paradigm. Therefore, the differential diagnosis is discussed.ConclusionIt is important to differentiate DKA from concomitant causes of abdominal pain to avoid missing the underlying etiology, which can be the trigger for DKA. During diabetic crises, treating the underlying trigger is just as important as managing metabolic derangements in order to achieve favorable outcomes; meanwhile, managing acute pancreatitis-associated hyperglycemia can promote recovery. Additionally, diabetic crisis that directly evolves from prediabetes illustrates an atypical form of diabetes called ketosis-prone diabetes; we briefly discuss its clinical characteristics, classification, and follow-up.

ENZYMATIC BIODIESEL SYNTHESIS FROM ACID OIL USING A LIPASE MIXTURE

The conventional biodiesel production process has some disadvantages. It is necessary to use refined vegetable oils with low free fatty acids (FFAs) content. An alternative route is to use low-cost acid oils in an enzymatic process. The use of lipases allows simultaneous esterification of FFAs and transesterification of triglycerides present in raw material forming alkyl esters. The aim of this work was to study the production of biodiesel using soybean oils with different acid contents (Acid Value of 8.5, 50, 90) and ethanol catalyzed by commercial immobilized lipases (Novozym 435, Lipozyme RM IM and Lipozyme TL IM). A significant decrease of acid value was observed mainly with Novozym 435 and Lipozyme RM IM. The use of a mixture of two immobilized lipases was also investigated to decrease catalyst cost and increase the amount of ester produced. The three commercial immobilized lipases were mixed in a dual system and tested for biodiesel synthesis from acid oil (AV of 8.5, 50 and 90). A positive synergistic effect occurred mainly for Lipozyme TL IM (1,3-specific lipase) and Novozym 435 (non-specific lipase) blend. The ester content doubled when this lipase mixture was used in ethanolysis of acid oil with AV of 90.

Process optimization for biodiesel production from waste cooking oil using multi-enzyme systems through response surface methodology

Lipase from Rhizomucor miehei (RML) and lipase B from Candida antarctica (CALB) were covalently immobilized onto epoxy-functionalized silica. In this study, we developed a multi-enzyme system to produce biodiesel with waste cooking oil and methanol. To increase the biodiesel production yield, a mixture of 1,3-specific lipase (RML) and nonspecific lipase (CALB) was used. Response Surface Methodology (RSM) and a central composite rotatable design (CCRD) was used to study the effects of four factors, CALB:RML ratio, ratio of t-butanol to oil (wt.%), water adsorbent content (wt.%) and reaction time on the fatty acid methyl esters (FAME) yield. A quadratic polynomial equation was obtained for methanolysis reaction by multiple regression analysis. The optimum combinations for the reaction were CALB:RML ratio (3:1), t-butanol to oil (10 wt%), water adsorbent content (22.5 wt%) at the reaction time of 10 h. FAME yield of 91.5%, which was very close to the predicted value of 95.6%, was obtained. Verification experiment confirmed the validity of the predicted model.

Maximización del contenido de Omega-3 (EPA y DHA) en el proceso de acidólisis enzimática de aceite de canola y concentrado de ácidos grasos poliinsaturados de cadena larga (AGPICL), en condiciones de CO2 supercrítico

The aim of this study was to optimize the content of EPA and DHA in the process of enzymatic acidolysis of canola oil and concentrated of long-chain polyunsaturated fatty acids (LCPUFA) in structured triacylglycerols (TAGs). For this purpose, nonspecific lipase B from Candida antarctica immobilized in a supercritical CO2 was used. Crude salmon oil obtained from the industrial byproducts was used to obtain LCPUFA concentrate. Initially, a LCPUFAs concentrate was obtained by basic hydrolysis and posterior complexation with urea. Subsequently the process variables were optimized enzymatic acidolysis were optimized using a central composite rotational design 25-1 + star, with 5 factors and 30 experimental trials, based on the response surface methodology. The optimal conditions that maximized the content of EPA and DHA to 3.92 g/100 g TFA and 9.09 g/100 g TFA, respectively in the purified TAGs corresponded to a LCPUFA percentage 71.71% and canola oil percentage 28.29%, temperature 57.8 °C, pressure 172.0 bar, time 23.97 h enzyme percentage of 7.74%.

Green synthesis of isopropyl myristate in novel single phase medium Part I: Batch optimization studies

Isopropyl myristate finds many applications in food, cosmetic and pharmaceutical industries as an emollient, thickening agent, or lubricant. Using a homogeneous reaction phase, non-specific lipase derived from Candida antartica, marketed as Novozym 435, was determined to be most suitable for the enzymatic synthesis of isopropyl myristate. The high molar ratio of alcohol to acid creates novel single phase medium which overcomes mass transfer effects and facilitates downstream processing. The effect of various reaction parameters was optimized to obtain a high yield of isopropyl myristate. Effect of temperature, agitation speed, organic solvent, biocatalyst loading and batch operational stability of the enzyme was systematically studied. The conversion of 87.65% was obtained when the molar ratio of isopropyl alcohol to myristic acid (15:1) was used with 4% (w/w) catalyst loading and agitation speed of 150rpm at 60°C. The enzyme has also shown good batch operational stability under optimized conditions.

Maximización del contenido de Omega-3 (EPA y DHA) en el proceso de acidólisis enzimática de aceite de canola y concentrado de ácidos grasos poliinsaturados de cadena larga (AGPICL), en condiciones de CO2 supercrítico

The aim of this study was to optimize the content of EPA and DHA in the process of enzymatic acidolysis of canola oil and concentrated of long-chain polyunsaturated fatty acids (LCPUFA) in structured triacylglycerols (TAGs). For this purpose, nonspecific lipase B from Candida antarctica immobilized in a supercritical CO2 was used. Crude salmon oil obtained from the industrial byproducts was used to obtain LCPUFA concentrate. Initially, a LCPUFAs concentrate was obtained by basic hydrolysis and posterior complexation with urea. Subsequently the process variables were optimized enzymatic acidolysis were optimized using a central composite rotational design 25-1 + star, with 5 factors and 30 experimental trials, based on the response surface methodology. The optimal conditions that maximized the content of EPA and DHA to 3.92 g/100 g TFA and 9.09 g/100 g TFA, respectively in the purified TAGs corresponded to a LCPUFA percentage 71.71% and canola oil percentage 28.29%, temperature 57.8 °C, pressure 172.0 bar, time 23.97 h enzyme percentage of 7.74%.

Starch and lipid in diets for douradoSalminus brasiliensis(Cuvier 1816): growth, nutrient utilization and digestive enzymes

Carbohydrates and lipids are the major and cheaper non-protein energy sources for fish diets. This study aimed at determining ideal relationship between dietary starch and lipids and their influence on digestive enzyme profile of juvenile dourado. Trials were set up in a closed-loop system with controlled temperature, dissolved oxygen, pH and total ammonia. Experimental extruded diets formulated to contain different starch (St) to lipids (Lip) ratio (St : Lip = 0 : 1, 0.3 : 1, 0.7 : 1, 1.0 : 1, 1.4 : 1, 2.0 : 1, 2.3 : 1 to 3.3 : 1) were fed to juvenile dourado (3.34 ± 0.16 g) and stocked into 30 polyethylene tanks (330 L, 30 fish per tank) in a completely randomized design with unbalanced repetition (n = 3 or 4), in two daily meals (1000 and 1530) until satiation, for 95 days. Dietary starch contents higher than 145.2 g kg-1 (St : Lip > 1.0 : 1.0) hampered the performance of fish, but, paradoxically, the inclusion of carbohydrates in the diet elicited better utilization of dietary protein and energy by the species. Fish adapted digestive enzyme profile to dietary starch contents through increasing secretion of alkaline proteases and non-specific lipases, but secretion of amylase was not altered. Physiological responses show the limitation of species on the use of dietary carbohydrates.

PH values and activity of digestive enzymes in the gastrointestinal tract of fish in Lake Chany (West Siberia)

Optimum pH of various digestive enzymes and physiological pH values were determined in the gastrointestinal tract of fish species, such as goldfish Carassius auratus gibelio, ide Leuciscus idus, wild carp Cyprinus carpio, perch Perca fluviatilis, and zander Sander lucioperca. Physiological values of pH in the stomach of perch and zander and in different parts of the intestine of all studied fish species range within 3.5–4.5 and 6.5–7.2, respectively. The maximum values of the activity of alkaline protease and nonspecific lipase and esterase are recorded at pH 8–9; the maximum acid protease activity is recorded at pH 2–3. The optimum pH activity of alkaline protease and nonspecific lipase and esterase do not correlate with actual pH value in the digestive tract of fish. Two peaks of α-amylase activity are observed at pH 7 and 9.

Combi-protein coated microcrystals of lipases for production of biodiesel from oil from spent coffee grounds

Abstract Background Replacing chemical catalysts with biocatalysts is a widely recognized goal of white biotechnology. For biocatalytic processes requiring low water containing media, enzymes for example commercial preparations of lipases, show low catalytic efficiencies. Some high activity preparations for addressing this concern have been described. Protein coated microcrystals (PCMC) constitute one such preparation. The present work describes a Combi-PCMC for synthesis of biodiesel from the oil extracted from spent coffee grounds. Results Different lipases were screened for biodiesel synthesis from crude coffee oil out of which Novozym 435 gave the best conversion of 60% in 4 h. Optimization of reaction conditions i.e. % water, temperature and purification of coffee oil further enhanced conversion upto 88% in 24 h. A mixture of Novozym 435 and a cheap commercially available 1,3-specific lipase RMIM (from Mucor miehei) was used in different ratios and 1:1 was found to be the best trade-off between conversion and cost. The commercial preparations then were replaced by a novel biocatalyst design called Combi-Protein coated microcrystals (Combi-PCMC) wherein CAL B and Palatase were co-immobilized with K2SO4 as the core and this performed equivalent to the commercial preparations giving 83% conversion in 48 h. Conclusion Coffee oil extracted from spent coffee grounds could be used for the synthesis of biodiesel by using appropriate commercial preparations of lipases. The expensive commercially immobilized preparations can also be replaced by a simpler and inexpensive immobilization design called combi-PCMC which synergizes the catalytic action of a nonspecific lipase CAL B and a free form of 1,3-specific lipase from Mucor miehei.

Comparative transcriptome analysis of Gossypium hirsutumL. in response to sap sucking insects: aphid and whitefly

BackgroundCotton (Gossypium hirsutum L.) is a major fiber crop that is grown worldwide; it faces extensive damage from sap-sucking insects, including aphids and whiteflies. Genome-wide transcriptome analysis was performed to understand the molecular details of interaction between Gossypium hirsutum L. and sap-sucking pests, namely Aphis gossypii (Aphid) and Bemisia tabacci (Whiteflies). Roche’s GS-Titanium was used to sequence transcriptomes of cotton infested with aphids and whiteflies for 2 h and 24 h.ResultsA total of 100935 contigs were produced with an average length of 529 bp after an assembly in all five selected conditions. The Blastn of the non-redundant (nr) cotton EST database resulted in the identification of 580 novel contigs in the cotton plant. It should be noted that in spite of minimal physical damage caused by the sap-sucking insects, they can change the gene expression of plants in 2 h of infestation; further change in gene expression due to whiteflies is quicker than due to aphids. The impact of the whitefly 24 h after infestation was more or less similar to that of the aphid 2 h after infestation. Aphids and whiteflies affect many genes that are regulated by various phytohormones and in response to microbial infection, indicating the involvement of complex crosstalk between these pathways. The KOBAS analysis of differentially regulated transcripts in response to aphids and whiteflies indicated that both the insects induce the metabolism of amino acids biosynthesis specially in case of whiteflies infestation at later phase. Further we also observed that expression of transcript related to photosynthesis specially carbon fixation were significantly influenced by infestation of Aphids and Whiteflies.ConclusionsA comparison of different transcriptomes leads to the identification of differentially and temporally regulated transcripts in response to infestation by aphids and whiteflies. Most of these differentially expressed contigs were related to genes involved in biotic, abiotic stresses and enzymatic activities related to hydrolases, transferases, and kinases. The expression of some marker genes such as the overexpressors of cationic peroxidase 3, lipoxygenase I, TGA2, and non-specific lipase, which are involved in phytohormonal-mediated plant resistance development, was suppressed after infestation by aphids and whiteflies, indicating that insects suppressed plant resistance in order to facilitate their infestation. We also concluded that cotton shares several pathways such as phagosomes, RNA transport, and amino acid metabolism with Arabidopsis in response to the infestation by aphids and whiteflies.

Potential use of cross-linked enzyme aggregates of nonspecific lipase as biocatalyst to produce structured lipids with commercial application

Potential use of cross-linked enzyme aggregates of nonspecific lipase as biocatalyst to produce structured lipids with commercial application

Characterization of an extracellular lipase and its chaperone from Ralstonia eutropha H16

Lipase enzymes catalyze the reversible hydrolysis of triacylglycerol to fatty acids and glycerol at the lipid-water interface. The metabolically versatile Ralstonia eutropha strain H16 is capable of utilizing various molecules containing long carbon chains such as plant oil, organic acids, or Tween as its sole carbon source for growth. Global gene expression analysis revealed an upregulation of two putative lipase genes during growth on trioleate. Through analysis of growth and activity using strains with gene deletions and complementations, the extracellular lipase (encoded by the lipA gene, locus tag H16_A1322) and lipase-specific chaperone (encoded by the lipB gene, locus tag H16_A1323) produced by R. eutropha H16 was identified. Increase in gene dosage of lipA not only resulted in an increase of the extracellular lipase activity, but also reduced the lag phase during growth on palm oil. LipA is a non-specific lipase that can completely hydrolyze triacylglycerol into its corresponding free fatty acids and glycerol. Although LipA is active over a temperature range from 10°C to 70°C, it exhibited optimal activity at 50°C. While R. eutropha H16 prefers a growth pH of 6.8, its extracellular lipase LipA is most active between pH7 and 8. Cofactors are not required for lipase activity; however, EDTA and EGTA inhibited LipA activity by 83%. Metal ions Mg(2+), Ca(2+), and Mn(2+) were found to stimulate LipA activity and relieve chelator inhibition. Certain detergents are found to improve solubility of the lipid substrate or increase lipase-lipid aggregation, as a result SDS and Triton X-100 were able to increase lipase activity by 20% to 500%. R. eutropha extracellular LipA activity can be hyper-increased, making the overexpression strain a potential candidate for commercial lipase production or in fermentations using plant oils as the sole carbon source.

ChemInform Abstract: Preparation of Optically Pure 2-Aryl- and 2-Aryloxy-propionates by Selective Enzymatic Hydrolysis.

Selective enzymatic hydrolysis of racemic esters is an attractive method for the resolution of 2-aryl- and 2-aryloxy-propionates. The non-steroidal antiinflammatory drugs, naproxen, ibuprofen, flurbiprofen and the herbicides, fluazifop and diclofop, can be obtained in high optical purity with the aid of carboxylesterase NP. This enzyme was brought to high production levels by cloning it into Bacillus subtilis 1-85. The unfavourable influence of a non-specific lipase, present in the wild type rain, was also completely eliminated by cloning. The ability of the esterase under application conditions was greatly improved by chemical modification, the be results being obtained with formaldehyde, glutaraldehyde and glyoxal.