Specific Activity Of Lipase Research Articles

Production and characterization of extracellular lipase secreted by Mucor geophillus

Nowadays, the growing interest in microbial lipase production by using the cheapest sources of carbon source, and its potential applications in biotechnological field has shown a remarkable place of prominence among the enzymes, owing to their significant ability to catalyze a wide variety of reactions in aqueous and non aqueous media. The purpose of this study was to enhance the production of lipase by Mucor geophillus in submerged culture condition, using molasses mineral medium as a low cost substrate and to increase the economical feasibility attractiveness. M. geophillus lipase was also assessed and characterized on the basis of some kinetic parameters. M. geophillus secreted higher yield of lipase at approximately 44.56 U.ml -1 at 72 h cultivation time. The optimal assay conditions were found at 60 min incubation of 2.0 and 8% enzyme and substrate concentration, respectively. However, lipase was active at 35°C and pH 6.5, but enzyme was found to be stable at about 70°C. From the range of pH 4.0 to 8.0, KCl enhanced the lipase activity, and had higher specific lipase activity towards the olive oil than other substrates. Key words : Lipase, Mucor geophillus , submerged fermentation, characterization.

Lipase and esterase: to what extent can this classification be applied accurately?

Enzyme technology is an ever-growing field of knowledge and, in recent years, this technology has raised renewed interest, due to the search for new paradigms in several productive processes. Lipases, esterases and cutinases are enzymes used in a wide range of processes involving synthesis and hydrolysis reactions. The objective of this work was to investigate and compare the specific lipase and esterase activities of five enzymes - four already classified as lipases and one classified as cutinase - in the presence of natural and synthetic substrates. All tested enzymes presented both esterase and lipase specific activities. The highest specific esterase activity was observed for Aspergillus 1068 lipase in natural substrate and for F. oxysporum cutinase in synthetic substrate, while the highest specific lipase activity was observed for Geotrichum sp. lipase in natural substrate and for F. oxysporum cutinase in synthetic substrate. These results display some interface-independent lipolytic activity for all lipases tested. This is in accordance with the rationale that a new and broader definition of lipases may be necessary.

Lipase production and growth modeling of a novel thermophilic bacterium: Aneurinibacillus thermoaerophilus strain AFNA

800x600 Normal 0 21 false false false ES-CL X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tabla normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman","serif";} Aneurinibacillus thermoaerophilus strain AFNA as a novel isolated extracellular thermostable organic solvent tolerant lipase producing bacterium was employed in the present study. The lipase production of strain AFNA and its correlation with bacterial growth was studied via a modeling assessment by response surface methodology (RSM) and artificial neural network (ANN) techniques. The best achieved models were multilayer full feed forward incremental back propagation network and modified cubic response surface model (mRSM) using backward elimination. The highest lipase specific activity (13.1 Umg -1 ) and bacterial growth (OD 600 = 3.0) were obtained at technically similar: growth temperature (53 and 53ºC), inoculum size (2.6 and 3.0%), agitation rate (118 and 115 rpm) and initial pH (7.0 and 7.2) but different medium volume (139 and 87 ml) and incubation period (48 and 38 hrs), respectively. In addition, the importance of effective parameters on the bacterial growth and lipase production was studied where pH and inoculum size were the most and the least effective factors, respectively. Significant correlation between lipase production and bacterial growth was observed when Bivariate correlation was employed to analyse the data. As a conclusion, lipase production was the result of a synergistic combination of effective parameters interactions and these parameters were in equilibrium.

Optimization of biomass production of a mutant of Yarrowia lipolytica with an increased lipase activity using raw glycerol.

The yeast Yarrowia lipolytica accumulates oils and is able to produce extracellular lipases when growing in different carbon sources including glycerol, the principal by-product of the biodiesel industry. In this study, biomass production of a novel mutant strain of Y. lipolytica was statistically optimized by Response Surface Methodology in media containing biodiesel-derived glycerol as main carbon source. This strain exhibited distinctive morphological and fatty acid profile characteristics, and showed an increased extracellular lipase activity. An organic source of nitrogen and the addition of 1.0 g/l olive oil were necessary for significant lipase production. Plackett-Burman and Central Composite Statistical Designs were employed for screening and optimization of fermentation in shaken flasks cultures, and the maximum values obtained were 16.1 g/l for biomass and 12.2 Units/ml for lipase, respectively. Optimized batch bioprocess was thereafter scaled in aerated bioreactors and the values reached for lipase specific activity after 95 % of the glycerol had been consumed, were three-fold higher than those obtained in shaken flasks cultures. A sustainable bioprocess to obtain biomass and extracellular lipase activity was attained by maximizing the use of the by-products of biodiesel industry.

Effect of Three Probiotics Administered Through Live Feed on Digestive Enzyme Activity in California Halibut, Paralichthys californicus, Larvae

The objective of this study was to evaluate the effect of probiotics as digestive system maturation promoters in larvae of the California halibut, Paralichthys californicus, during ontogeny. Three commercial probiotics: Levucell®, Bactocell®, and live yeast, were supplied at a concentration of 50 mg/L, using enriched rotifers and Artemia nauplii as vectors. Total and specific activities of trypsin, leucine‐aminopeptidase (LAP), lipase, chymotrypsin, and pepsin were assessed throughout the development. Growth in terms of length was not significantly different between treatments. Total pepsin and chymotrypsin activity were significantly higher at the end of the experiment, 46‐d posthatch (dph) in larvae supplied with Bactocell. However, no significant effect of the probiotic was detected for trypsin, lipase, and LAP. On the basis of the results, California halibut larvae seemed to have adequate alkaline proteolytic activity early in development, whereas acid proteolytic activity was not detected until the complete formation of a functional stomach. The feeding protocol and probiotics evaluated were unable to improve the morphological development of digestive system or the digestive enzyme activity early on. Nevertheless, the increase in activity of pepsin and chymotrypsin observed at 46 dph suggests a potential use of these probiotics once the metamorphosis is completed, and thus warrants further evaluation.

Developmental changes of digestive enzymes in Persian sturgeon ( Acipenser persicus) during larval ontogeny

The development of digestive enzymes from the stomach (pepsin), pancreas (trypsin, chymotrypsin, α-amylase and lipase) and intestine (alkaline phosphatase) was studied in Persian sturgeon ( Acipenser persicus) from hatching to the juvenile stage at 40 days post hatching (dph). Larvae were obtained from artificial propagation of one male and one female and transferred to larval culture tanks where, after yolk sac absorption (9 dph at 17–18 °C), they were fed with Artemia urmiana and Daphnia sp. The assessment of the activity of digestive enzymes showed that at the onset of exogenous feeding, gastric glands were already functional as indicated by the increase in pepsin specific activity. In contrast, alkaline proteases like trypsin and chymotrypsin decreased their specific activity after the onset of exogenous feeding, indicating the importance of these types of enzymes in the cleavage of yolk proteins during the endogenous feeding phase and the replacement of the larval alkaline-type digestion by a juvenile-type acid digestion. After the first feeding, amylase and lipase specific activities increased. Such increments in the activity of amylase might be genetically programmed to better digest carbohydrates in diets with the goal of sparing proteins during the larval stage, whereas the increase in lipase was related to changes in the lipid content of live prey and the progressive maturation of the pancreatic function during larval development. Changes in enzyme activities from the stomach and pancreas were coupled with that in the intestine (brush border membrane), where the specific activity of alkaline phosphatase progressively increased until 19–24 dph and remained constant thereafter, indicated the maturation of the intestine and the achievement of a juvenile-like mode of digestion. Considering these data on the digestive enzymes from the pancreas, stomach and intestine, Persian sturgeon larvae might be weaned around 19–24 dph, as larvae have achieved the complete maturation of their digestive functions by this date. This developmental process, and particularly for the digestive functions, can be considered as a reference to evaluate the effect of a formulated micro diets feeding on larvae.

Digestion of lipids and carbohydrates from microalgae (Chaetoceros muelleri Lemmermann and Isochrysis aff. galbana clone T-ISO) in juvenile scallops (Pecten maximus L.)

The digestion of laminaran and neutral lipids from the microalgae Chaetoceros muelleri or mixtures of C. muelleri and Isochrysis galbana clone T-ISO in 4-mm scallops Pecten maximus were studied. The microalgae were cultivated at fixed low and high growth rate (5% and 60% of μmax). The cultivation method clearly affected the lipid and carbohydrate contents, as well as their enzyme activity. Chaetoceros muelleri and T-ISO grown at low growth rate exhibited a considerably higher content of lipids and carbohydrates than those algae grown at high growth rate. The specific activity of laminaranase and neutral lipase was the highest in C. muelleri grown at the low growth rate. Growth rate of scallops was best when they were fed a mono-species diet of C. muelleri grown at low growth rate. The growth rate of the scallops was positively correlated to the dietary carbohydrate content as well as to the specific activity of neutral lipase and laminaranase in the diet. The capacity to digest carbohydrates (laminaran) might be limiting for the young scallops, and the content of carbohydrates and enzymatic activities of laminaran in the diet should be considered when formulating an optimal diet, both when the diet consist of live microalgae and formulated diets.

Improving catalytic performance of Burkholderia cepacia lipase immobilized on macroporous resin NKA

Abstract The lipase from Burkholderia cepacia adsorbed on macroporous resin NKA was investigated by combined strategies of bioimprinting and interfacial activation to enhance its catalytic performance. The specific activity of the derivative lipase was 211,733.3 U/g-protein, which was 21.7-fold, 19.4% and 47% enhancement over the free lipase powder, non-bioimprinted and non-interfacial activation lipase, respectively. The derivative lipase exhibited a satisfactory thermal stability over a wide range of temperature (from 30 °C to 70 °C) and a strong tolerance to organic solvents such as methanol, ethanol and acetone with 50% concentration. After being used of 50 successive batches (400 h), the derivative lipase still retained over 92% of its original activity (methyl esters yield decreased from 98% to 90%). Circular dichroism analysis indicated that the activity enhancement of the derivative lipase was ascribed to the secondary structure changes. The derivative lipase preparation in this work was probably a promising alternative to produce a biocatalyst of satisfactory thermo-stability, strong solvents tolerance and high operational reusability.

Dietary soybean phosphatidylcholine affects growth performance and lipolytic enzyme activity in Caspian brown trout (Salmo trutta Caspius) alevin

The effects of supplemental dietary phosphatidylcholine (PtCho) on the growth performance, survival and digestive enzyme activity of Caspian brown trout (Salmo trutta caspius Kessler 1877) alevins were investigated in this study. Alevin (initial weight 0.8 ± 0.12 g) was fed for 5 weeks with an isoproteic- and isolipidic-formulated diet with increased levels of PtCho from 0 to 60 g kg−1 dry matter and decreasing levels of soybean oil. The increase in dietary PtCho up to 4% led to an increase in alevin final weight, suggesting that moderate PtCho levels are needed during this stage of Caspian brown trout. Survival was not affected by the dietary PtCho level. Phosphatidylcholine incorporation into the diet caused higher phospholipase A2-specific activity. Phosphatidylcholine did not show a beneficial effect on the specific activity of amylase and protease. Lipase-specific activity was significantly higher in the PtCho groups compared with the control group. The hepatosomatic index (HSI) was significantly influenced by the dietary PtCho level. The results of the present study indicated that the dietary supplementation of PtCho in the diet of Caspian brown trout alevin improved growth and lipolytic enzyme activity.

Polycationic amino acid tags enhance soluble expression of Candida antarctica lipase B in recombinant Escherichia coli

Lipase (EC 3.1.1.3) is a popular enzyme used as an ingredient in detergents and biocatalyst in many biochemical reactions. Lipase is usually expressed in Escherichia coli as an inactive inclusion body and at a low level. In this study, Candida antarctica lipase B (CalB) was fused with various polycationic amino acid tags and expressed in E. coli in order to increase a soluble expression level. By induction with 1.0 mM IPTG, the authentic and fused CalBs were expressed at 27-56% of total protein. The 10-arginine and 10-lysine tags fused at the C-terminal of CalB significantly increased the solubility of CalB by five- to ninefold, relative to the case of the authentic CalB expressed in a recombinant E. coli Origami 2™ (DE3) strain. Among a series of the C-terminal poly-arginine tags, the recombinant CalB combined with the 10-arginine tag (CalB-R10) possessed the highest lipase specific activity of 9.5 ± 0.03 U/mg protein, corresponding to a fourfold enhancement compared with the authentic CalB.

Expression, purification of a novel alkaline Staphylococcus xylosus lipase acting at high temperature

A new Staphylococcus xylosus strain was isolated. The extracellular lipase of S. xylosus (wt-SXL2) was purified to homogeneity from the culture medium. The specific activity of the purified enzyme, measured at pH 8.5 and 55 °C using tributyrin or olive oil emulsion, reached, respectively, 6300 U/mg or 2850 U/mg. The sequenced 18 N-terminal amino acid showed a high degree of identity with known staphylococcal lipase sequences. The gene encoding the mature lipase was cloned and sequenced. The deduced amino acid sequence showed a significant similarity with various staphylococcal lipases. The highest overall identity (98.74%) was found with S. xylosus lipase (SXL1). The mature part of the lipase was expressed in Escherichia coli. The recombinant lipase was purified by affinity chromatography. The specific activity of the recombinant lipase was 4100 or 1500 U/mg using tributyrin or olive oil emulsion as substrate, respectively, at pH 8.5 and 55 °C. The wild type and recombinant lipases presented a quite interesting thermal stability, after an incubation of 60 min at 55 °C and they are found to be highly stable at a pH ranging from 4 to 11. Due to its stability at high temperature and in organic solvent, the wt-SXL2 was used as biocatalyst to synthesise a high added value molecules.

판크레아틴 소화효소의 활성화 조건 연구

돼지 췌장에서 추출한 판크레아틴 효소들의 활성화 조건에 대해 조사하였다. 십이지장의 첨가는 췌장의 단백질분해효소와 지방분해효소의 활성화를 유도하였다. 췌장액에 10% 십이지장을 첨가하여 $30^{\circ}C$ 에서 90분간 반응시키거나 $25^{\circ}C$ 에서 4시간 반응시켰을때 췌장의 단백질분해효소와 지방분해효소의 활성화는 정점에 도달하였고, 전분분해효소의 활성에는 거의 영향을 주지 않았다. 2 $5^{\circ}C$ 에서 4시간의 효소활성화, 원심분리, 아세톤침전 및 동결건조의 연속 공정으로 제조한 판크레아틴 효소들의 비활성도는 단백질분해효소 136 U/mg, 지방분해효소 170 U/mg 및 전분분해효소 400 U/mg으로 나타났다. 확립된 공정에 의해 제조된 판크레아틴은 USP 기준의 5.4배의 단백질분해효소, 58배의 지방분해효소 및 16배의 전분분해효소 활성을 보유한 고활성의 제품이었다. 【This study investigated the activation conditions of pancreatic enzymes from porcine pancreas. Duodenum induced the activation of pancreatic protease and lipase in pancreas. When 10% duodenum was added to pancreatic juice and the mixture was incubated at $30^{\circ}C$ for 90 min or at $25^{\circ}C$ for 4 hrs, the activities of pancreatic protese and lipase reached the peak. When the pancreatin was prepared by sequential process of enzymatic activation at $25^{\circ}C$ for 4 hrs, centrifugation, acetone precipitation and freeze-drying, the specific activities of pancreatic protease, lipase and amylase were 136, 116 and 400 U/mg-protein, respectively. The protease, lipase and amylase activities of the prepared pancreatin were 5.4, 58.0 and 16.0 times higher than those of USP standard, respectively.】

Optimization and Modeling of Process Parameters for Lipase Production by Bacillus brevis

Statistical evaluation of fermentation conditions and nutritional factors by Plackett–Burman two-level factorial design followed by optimization of significant parameters using response surface methodology for lipase production by Bacillus brevis was performed in submerged batch fermentation. Temperature, glucose, and olive oil were found to be the significant factors affecting lipase production. Maximum lipase activity of 5.1 U ml−1 and cell mass of 1.82 g l−1 at 32 h were obtained at the optimized conditions of temperature, 33.7 °C; initial pH, 8; and speed of agitation, 100 rpm, with the medium components: olive oil, 13.73 ml l−1; glucose, 13.98 g l−1; peptone, 2 g l−1; Tween 80, 5 ml l−1; NaCl, 5 g l−1; CH3COONa, 5 g l−1; KCl, 2 g l−1; CaCl2·2H2O, 1 g l−1; MnSO4·H2O, 0.5 g l−1; FeSO4·7H2O, 0.1 g l−1; and MgSO4·7H2O, 0.01 g l−1. The lipase productivity and specific lipase activity were found to be 0.106 U (ml h)−1 and 2.55 U mg−1, respectively. Unstructured kinetic models and artificial neural network models were used to describe the lipase fermentation. The kinetic analysis of the lipase fermentation by B. brevis shows that lipase is a growth-associated product.

Nutritional evaluation of mysids Mesopodopsis orientalis (Crustacea:Mysida) as live food for grouper Epinephelus fuscoguttatus larvae

The potential of mysids Mesopodopsis orientalis as live food source for grouper Epinephelus fuscoguttatus larvae was investigated. In comparison with Artemia biomass, a common live food source in larviculture, mysids contained significantly higher levels of protein, total lipid, eicosapentaenoic acid (EPA, C 20:5 n−3), and docosahexaenoic acid (DHA, C 22:6 n−3). DHA was not detected in Artemia biomass. Grouper larvae fed mysids from 35 to 55 days after hatching (DAH) had 2-fold and 3-fold higher specific growth rates and survival, respectively, than those fed Artemia biomass. DHA levels increased 6-fold while EPA levels remained constant in mysid-fed grouper larvae. In contrast, DHA and EPA significantly decreased in Artemia biomass-fed grouper larvae. Furthermore, the specific activities of amylase, lipase and protease generally significantly increased ( P < 0.05) in both mysid-fed and Artemia biomass-fed grouper larvae from 35 to 55DAH. A marked increase in the specific activity of amylase was seen in mysid-fed than in Artemia biomass-fed grouper larvae. Results of the nutritional evaluation suggest that mysids are superior live food organisms than Artemia biomass for grouper larvae and could significantly improve production of grouper juveniles in the nursery phase.

Effects of different sources of dietary non-starch polysaccharides on the growth performance, development of digestive tract and activities of pancreatic enzymes in goslings

1. The purpose of this study was to investigate the effects of different sources of dietary non-starch polysaccharides (NSP) on growth performance, development of gastrointestinal tract, and activities of pancreatic enzymes in goslings from 0 to 21 d of age. 2. A total of 100 one-day-old White Roman female goslings were selected and randomly divided into 5 dietary treatment groups. Each group had 4 replicate cages (100 × 60 × 58 cm) of 5 goslings. Goslings in each group were given one of 5 isoenergetic and isonitrogenous semi-purified diets. Experimental diets were designed to contain 90 g/kg of different sources of dietary NSP from maize (control), barley hull, rice bran, wheat bran, or pectin, respectively, and 97·4 g/kg NSP were supplied by soybean meal. 3. The goslings receiving the pectin diet had significantly higher viscosity in intestinal digesta than the other treatment diets. However, the daily feed intake, daily weight gain, and feed conversion ratio of goslings in the pectin group were significantly lower than those given the other diets. 4. The relative weights of proventriculus, gizzard, liver and pancreas, as well as the relative weights and lengths of various intestinal segments in the goslings that received the pectin diet, were significantly higher than those receiving the other diets. The specific activities (SA) of amylase, lipase, trypsin, and chymotrypsin in the pancreatic tissue of goslings in the pectin group were significantly lower than those in the other groups. 5. Goslings given a soluble NSP source (pectin) diet tended to exhibit inhibited growth and decreased activity of pancreatic enzymes. However, compared with maize, the inclusion of insoluble NSP sources from barley hull, rice bran, and wheat bran did not negatively affect growth and also were not significantly different among the treatment diets.

Surfactant-modified yeast whole-cell biocatalyst displaying lipase on cell surface for enzymatic production of structured lipids in organic media

The cell surface engineering system, in which functional proteins are genetically displayed on microbial cell surfaces, has recently become a powerful tool for applied biotechnology. Here, we report on the surfactant modification of surface-displayed lipase to improve its performance for enzymatic synthesis reactions. The lipase activities of the surfactant-modified yeast displaying Rhizopus oryzae lipase (ROL) were evaluated in both aqueous and nonaqueous systems. Despite the similar lipase activities of control and surfactant-modified cells in aqueous media, the treatment with nonionic surfactants increased the specific lipase activity of the ROL-displaying yeast in n-hexane. In particular, the Tween 20-modified cells increased the cell surface hydrophobicity significantly among a series of Tween surfactants tested, resulting in 8-30 times higher specific activity in organic solvents with relatively high log P values. The developed cells were successfully used for the enzymatic synthesis of phospholipids and fatty acid methyl esters in n-hexane, whereas the nontreated cells produced a significantly low yield. Our results thus indicate that surfactant modification of the cell surface can enhance the potential of the surface-displayed lipase for bioconversion.

Synthesis of thermo-sensitive copolymer with affinity butyl ligand and its application in lipase purification

In this study, thermo-sensitive N-alkyl substituted polyacrylamide polymer P(NNB) was synthesized by using N-hydroxymethyl acrylamide(NHAM), N-isopropyl acrylamide (NIPA) and butyl acrylate (BA) as monomers, and its low critical solution temperature (LCST) was controlled to be 28 degrees C. The recovery of the thermo-sensitive polymer was over 98%. Butanol as a hydrophobic ligand was covalently attached onto polymer P(NNB) and butyl ligand density was 80 micromol g(-1) polymer. The affinity polymer was used for purification of lipase from crude material. Optimized condition was pH 7.0, 35 degrees C adsorption temperature, 120 min adsorption time and 0.5 mg ml(-1) initial concentration of lipase. The adsorption isotherm accords with a typical Langmuir isotherm. The maximum adsorption capacity (Q(m)) of the affinity polymer for lipase was 24.8 mg g(-1)polymer. The affinity copolymer could be recycled by temperature-inducing precipitation and there was only about 6% loss of adsorption capacity after five recyclings. Specific activity of lipase was improved from 14IU mg(-1) to 506 IU mg(-1) protein, and its recovery achieved 82%. The affinity polymer is suitable for the purification of target proteins from the crude material with large volume and dilute solution.

Influence of supplying bullfrog tadpoles with feed containing 28% crude protein on performance and enzymatic activities

The objective of this study was evaluate the influence of feeding bullfrog tadpoles on commercial feed containing 28% crude protein (CP), on their digestive enzyme performance and activities. The experiment lasted 60 days, at the density of one tadpole/L in boxes containing 30 L water. A hundred and twenty tadpoles at the 25 Gosner stage averaging weight and length was 0.046 g and 6.22 mm, respectively, were used. Survival rate, length, final weight, weight gain, feed consumption, apparent food conversion, specific growth rate and activities of chime, amylase, lipase and trypisin were the parameters evaluated, biweekly, in five biometries. Quadratic effect was observed for the length and the weight. There was larger growth of the tadpoles from the 15th to the 45th day (19.82 mm). On the 15th day, the tadpoles presented the largest specific growth rate (16.93%/day), and the largest weight gain (5.460 g), feed intake (14.099 g) and the best apparent food conversion (2.46) was from the 45th to the 60th day. The specific activity of amylase was 205 times greater at 60 days when compared to the beginning of the experiment. The results demonstrated that, for the three enzymes studied, the action capacity over the tadpole chime increased significantly after the 30th experimental day. Moreover, they suggested a greater capacity of tadpoles to digest carbohydrates in detriment to proteins, and this fact was accentuated in the initial phase of the exogenous feeding of this amphibian. The commercial feed with 28% CP provide good performance in the bullfrog tadpoles, indicating the juvenile formation within commercial bullfrog farming standards.

Enhancing performance of lipase immobilized on methyl-modified silica aerogels at the adsorption and catalysis processes: Effect of cosolvents

Hydrophobic silica aerogels modified with methyl group were applied as support to immobilize Candida rugosa lipase (CRL). At the adsorption process, different alcohols were used to intensify the immobilization of CRL. The results showed that n-butanol wetting the hydrophobic support prior to contacting with enzyme solution could promote lipase activity, but the adsorption quantity onto the support decreased. Based on this, a novel immobilization method was proposed: the support contacted with enzyme solution without any alcohols, and then the immobilized enzymes were activated by 90% (V) n-butanol solution. The experimental results showed that this method could keep high adsorption quantity (413.0 mg protein/g support) and increase the lipase specific activity by more than 50%. To improve the stability of immobilized lipase, the support after adsorption was contacted with n-octane to form an oil layer covering the immobilized lipases, thus the leakage can be decreased from over 30–4% within 24 h. By utilizing proper cosolvents, a high enzyme activity and loading capacity as well as little loss of lipase was achieved without covalent linkage between the lipase and the support. This is known to be an excellent result for immobilization achieved by physical adsorption only.

Influence of cultivation conditions on the production of a thermostable extracellular lipase from Amycolatopsis mediterranei DSM 43304

Among several lipase-producing actinomycete strains screened, Amycolatopsis mediterranei DSM 43304 was found to produce a thermostable, extracellular lipase. Culture conditions and nutrient source modification studies involving carbon sources, nitrogen sources, incubation temperature and medium pH were carried out. Lipase activity of 1.37 +/- 0.103 IU/ml of culture medium was obtained in 96 h at 28 degrees C and pH 7.5 using linseed oil and fructose as carbon sources and a combination of phytone peptone and yeast extract (5:1) as nitrogen sources. Under optimal culture conditions, the lipase activity was enhanced 12-fold with a twofold increase in lipase specific activity. The lipase showed maximum activity at 60 degrees C and pH 8.0. The enzyme was stable between pH 5.0 and 9.0 and temperatures up to 60 degrees C. Lipase activity was significantly enhanced by Fe(3+) and strongly inhibited by Hg(2+). Li(+), Mg(2+) and PMSF significantly reduced lipase activity, whereas other metal ions and effectors had no significant effect at 0.01 M concentration. A. mediterranei DSM 43304 lipase exhibited remarkable stability in the presence of a wide range of organic solvents at 25% (v/v) concentration for 24 h. These features render this novel lipase attractive for potential biotechnological applications in organic synthesis reactions.