Purification and Characterization of Highly Alkaline Lipase from Bacillus licheniformis MTCC 2465: and Study of its Detergent Compatibility and Applicability

An extracellular alkaline lipase from Bacillus licheniformis MTCC 2465 has been studied and analyzed for its applicability as a detergent additive. The lipase obtained from Bacillus licheniformis MTCC 2465 was purified by ammonium sulfate fractionation and gel filtration chromatography. The enzyme was precipitated with a 60 % saturated ammonium sulfate fraction showed 6.73 fold purification with the fold purification of 10.4 and specific activity of 398 U/mg of protein with gel filtration chromatography. The optimal pH and temperature for activity were 10 and 60 °C respectively. The enzyme was found to be stable in the pH range of 8–11 with 90 % retention of activity at pH 11. The enzyme retained 90 % activity at 60 °C and 70 % of activity at 70 °C for 1 h. The enzyme activity was maximally enhanced by Hg²⁺ followed by Co²⁺ and Fe²⁺. The lipase was found to be stable in organic solvents with maximum stability in acetone followed by ethanol. The lipase exhibited remarkable stability in the presence of commercial detergents and found to be stable in bleaching agents. Wash performance analysis resulted in improvement of 10 % more grease removal ability of the present lipase in comparison to commercially available lipase.     

The Use of Boron Compounds for Stabilization of Lipase from Pseudomonas aeruginosa ES3 for the Detergent Industry

This study aimed to characterize a lipase that is highly active and stable under typical washing conditions for use as a detergent ingredient by investigating the effects of various boron compounds on lipase stabilization under different conditions. In addition, the antimicrobial activity of the boron compounds used in enzyme stabilization was examined in order to obtain an effective antimicrobial detergent. A lipase‐producing bacterium was isolated from kitchen wastewater samples using Rhodamine‐B Agar medium and identified as Pseudomonas aeruginosa based on 16S rDNA sequence analysis. The ES3 lipase obtained from P. aeruginosa was purified, and the purified enzyme was found to have a molecular mass of 40 kDa. The enzyme showed optimal activity at pH 9.0–10.0 and 40 °C and remained stable in the presence of various metal ions, surfactants and oxidizing agents. Moreover, the pH stability and thermostability of the enzyme was improved by the addition of boron compounds, which, when used as stabilizers in the incubation media, also increased the stability of the enzyme towards commercial detergents. Furthermore, the enzyme displayed properties comparable with the commercial product Lipolase^®, which has shown excellent stability towards various commercial detergents. Finally, boron compounds used to stabilize the lipase were found to possess antimicrobial properties, suggesting that detergents incorporating these compounds will also exhibit antimicrobial activity when washing clothes and dishes.     

Production and Characteristics of an Enantioselective Lipase from Burkholderia sp. GXU56

The lipase production of Burkholderia sp. GXU56 was influenced by carbon and nitrogen sources, inorganic salts, initial pH of the medium and cultivation temperature. The maximum lipase production was 580.52 U/mL and reached 5 times the level of the basic medium in the optimum medium at pH 8.0, 32 °C, 200 rpm and 40–48 h. The lipase was purified 53.6 fold to homogeneity and the molecular weight was 35 KDa on SDS‐PAGE. The optimum pH and temperature of the lipase were 8.0 and 40 °C, respectively, and it was stable in the range of pH 7–8.5 and at temperatures below 45 °C. The lipase activity was strongly inhibited by Zn²⁺, Cu²⁺, Co²⁺, Fe²⁺, Fe³⁺ ions and SDS, while it was stimulated by Li⁺ and Ca²⁺ ions and in presence of 0.1 % CTAB, 0.1 % Triton X‐100 and 10 % DMSO. K_m and V_max of the lipase were calculated to be 0.038 mmol/L, and 0.029 mmol/L min^–1, respectively, with PNPB as the substrate. The GXU56 lipase showed enantioselective hydrolysis of (R,S)‐methyl mandelate to (R)‐mandelic acid, which is an important intermediate in the pharmaceutical industry.     

Identification and Characterization of a Novel Thermophilic,Organic Solvent Stable Lipase of <Emphasis Type="Italic">Bacillus</Emphasis> from a Hot Spring

A novel lipase gene lip256 was cloned and identified from the genomic library of hot spring strain Bacillus sp. HT19. The deduced amino acid sequence of lip256 has less than 32% identity to a predicted esterase (Cog1752) from Photobacterium leiognathi lrivu.4.1 and contains a novel motif (GTSAG) that differs from other clusters in the lipase superfamily. Following purification, a single band was obtained with a molecular mass of 33 kDa by SDS-PAGE, and the optimal temperature and pH for lipolytic activity of Lip25 were 70 °C and 9.0, respectively. Lip256 exhibited high activity at high temperatures, with 40% maximum activity at 80 °C and good stability at temperatures ranges between 50 and 80 °C. Additionally, the enzyme was highly stable in the presence of butyl-alcohol, glycerol, acetonitrile, pyridine, and urea. However, the presence of acetone, methanol, trichloromethane, petroleum ether, hexane, tert-butanol, isopropanol, dithiothreitol, ethylenediaminetetraacetic acid, polyhexamethylene biguanide, dimethyl sulfoxide, benzene, Triton X-100, Tween-20, Tween-80, and sodium dodecyl sulfate suppressed or absolutely inhibited enzyme activity. Furthermore, Ca²⁺, Mg²⁺, and Cu²⁺ suppressed enzyme activity, whereas Na⁺, Fe³⁺, K⁺, Fe²⁺, and Sr²⁺ enhanced enzyme activity. The unique characteristics of novel lipase Lip256, including its thermo-alkaliphilic performance, high tolerance toward metal ions, inhibitors, and detergents, and high stability in organic solvents, implied that this enzyme might be an interesting candidate for industrial processes.     

Purification and Characterization of a Lipase from Candida curvata Lodder and Kreger — van Rij CBS 570

The purification of the lipase from Candida curvata CBS 570 was achieved steps. Its optimum pH is 6 and its optimum temperature range is Iff C to 60°C. This enzyme is thermoresistant and only loses 20% of its activity when heated at 50°C during 30 minutes. Its activation energy is 144 kcal/mole and its inactivation energy 22 kcal/mole. Its molecular weight was determined to be 195000. EDTA, p-chloromercuri benzoate, N-ethyl- and iodoacetamide have no influence on the activity of this enzyme, whereas Cu⁺⁺ and Zn⁺⁺ show strong inhibitory effects. The lipase activity is induced by the presence of triglycerides and inhibited by the presence of glucose. This enzyme strongly attacks triolein and trilinolein molecules, however it only hydrolyzes a little tristearin and trilinolenin.     

Detergent Compatible Extracellular Lipase from Streptomyces cellulosae AU‐10: A Green Alternative for the Detergent Industry

Enzymes can decrease the environmental and economic load of detergent products by reducing the amount of chemicals used in detergents and by allowing washing at ambient temperatures. In this study, Streptomyces cellulosae AU‐10 (GenBank accession number: MG780240) lipase was purified 7.08‐fold with 68% yield using an aqueous 2‐phase system. The Streptomyces sp. AU‐10 lipase showed maximal activity at pH 9.0 and 40 °C. Hundred percent activities were measured in the pH range from 9.0 to 11.0 for 1 h. The enzyme was also highly stable at 30–50 °C. The values of K_m and V_max were calculated as 0.34 mM and 0.83 mM min^?1, respectively. The lipase has high hydrolytic activity for olive oil and sunflower oil. The effect of ethylenediamine tetraacetic acid on the enzyme has shown that the lipase is a metalloenzyme. The activity increased in the presence of Fe²⁺, Cu²⁺, and various boron compounds. The enzyme has shown a good stability not only with surfactants but also with oxidizing agents. In addition, activities in the presence of Omo, Ariel, Tursil, Pril, and Fairy were measured as 108.8%, 115.6%, 98.35%, 140.4%, and 107.6%, respectively. Considering its remarkable ability, the S. cellulosae AU‐10 lipase can be considered as a potential additive in the detergent industry.     

Purification and Biochemical Characterization of Lipase from Tunisian Euphorbia peplus Latex

Lipases from vegetable sources have been the focus of intense and growing research. The use of enzymes from plants has the advantage of employing industrial waste products. The lipase activity of Euphorbia peplus L. (Euphorbiaceae) was investigated for the first time. The Euphorbia peplus latex lipase (EpLL) was purified after ammonium sulfate fractionation and anion exchange chromatography on a DEAE-Cellulose column leading to 12.57-fold purification. The EpLL displayed a probable molecular weight of about 40 kDa. The lipase activity was optimum at a temperature of 40 °C and pH 8, the specific activities of EpLL were found to be 249 ± 12.45 and 161.4 ± 8.07 U/mg when tributyrin (TC₄) and olive oil were used as substrate respectively. The enzyme retained 80 % of its activity when incubated for 1 h at 50 °C. The EpLL was strongly destabilised by divalent metal ions (Fe²⁺, Mg²⁺, Zn²⁺ and Cu²⁺). Lipase was slightly stimulated by Triton X-100 and Tween-80, while strongly inhibited by sodium dodecyl sulfate. A good stability of the enzyme in the presence of organic solvents was reveled suggesting its industrial utility.     

Production of thermostable alkaline lipase on vegetable oils from a thermophilic Bacillus sp. DH4, characterization and its potential applications as detergent additive

BACKGROUND: The alkaline lipase production on vegetable oils as sole carbon source, its characterization and effect of different commercial detergents and surfactants on enzymatic activity from thermophilic Bacillus sp. DH4 was investigated. RESULTS: The organism grew on mannose, but the amount of lipase secreted was significantly less than on vegetable oils. This study identified a simple substrate for lipase production and established the utility of groundnut oil for increasing the lipase yield. The enzyme was compatible with various ionic and non‐ionic surfactants as well as commercial detergents. Lipase activity was strongly inhibited by sodium dodecyl sulfate (SDS), but not by Triton X‐100 or Triton X‐114. The best assay conditions observed for this lipase was found to be pH 9.0 and 50 °C. The enzyme was stable at alkaline pH and considerable activity was observed at pH 10 and it retained 93% of the residual activity at 60 °C. The lipase showed a novel property of marked activation at alkaline pH. Wash performance analysis of commercial detergent for removal of fatty stains improved upon addition of lipase. CONCLUSION: The production on cheap vegetable oils, novel properties and resistance towards various surfactants and tolerance to commercial detergents make this lipase a potential additive for detergent formulations. Significance and impact of the study: Bacillus sp. produces alkaline and thermostable lipase on cheap vegetable oils and its compatibility can find use in the detergent industry. Copyright © 2008 Society of Chemical Industry     

Methacrylic acid and dodecyl methacrylate (MAc‐DMA) hydrogel for enhanced catalytic activity of lipase of Bacillus coagulans MTCC‐6375

An alkaline thermotolerant bacterial lipase of Bacillus coagulans MTCC‐6375 was purified and immobilized on a methacrylic acid and dodecyl methacrylate (MAc‐DMA) hydrogel. The lipase was optimally bound to the matrix after 20 min of incubation at 55°C and pH 9 under shaking conditions. The matrix‐bound lipase retained approximately 50% of its initial activity at 70–80°C after 3 h of incubation. The immobilized lipase was highly active on medium chain length p‐nitrophenyl acyl ester (C: 8, p‐nitrophenyl caprylate) than other p‐nitrophenyl acyl esters. The presence of Fe³⁺, NH₄⁺, K⁺, and Zn²⁺ ions at 1 mM concentration in the reaction mixture resulted in a profound increase in the activity of immobilized lipase. Most of the detergents partially reduced the activity of the immobilized lipase. The immobilized lipase performed ～62% conversion in 12 h at temperature 55°C. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 100: 1420–1426, 2006     

Purification and biochemical characterization of an intracellular lipase by Rhizopus chinensis under solid‐state fermentation and its potential application in the production of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)

BACKGROUND: Purification and characterization of an intracellular lipase produced by Rhizopus chinenesis cultured in solid‐state fermentation was investigated. The potential application in concentrating eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) from fish oil by the pure enzyme was also studied. RESULTS: Through four successive purification steps, the enzyme was purified to homogeneity with an apparent molecular mass of 36 kDa. The lipase was active for pH between 7.0 and 9.0 and temperatures 20–45 °C. Lipase activity was slightly increased in the presence of Ca²⁺ and Mg²⁺, but strongly inhibited by Hg²⁺ and SDS. The pure enzyme was most active on medium chain p‐nitrophenol esters, with the highest activity towards pNP‐caprylate (C8). The enzyme is a non‐specific lipase, because it cleaved not only the 1,3‐positioned ester bonds but also the 2‐positioned bond in triolein. High EPA (17.6%) and DHA (32.9%) contents were achieved using the pure lipase (100 U) within 10 h. CONCLUSION: The enzymatic activity of the lipase on a wide variety of substrates and its stability in the presence of some organic solvents suggest that the lipase should be investigated for a range of commercial applications. The pure lipase was proved to possess potential ability for the production and concentration of EPA and DHA from fish oil. Copyright © 2008 Society of Chemical Industry     

Fatty Acid and Position Specificities of an Intracellular Lipase from Geotrichum candidum

An intracellular lipase was isolated and purified to homogeneity from mycelia of Geotrichum candidum. The lipase showed maximum activity at 35° C and pH 7.5. The enzyme preferentially hydrolyzed oleic acid glycerol ester bonds on using mono-acid triglycerides (C_10:0, C_18:0) as substrates. However the results indicated that the enzyme is position unspecific.     

Purification of Bacillus licheniformis Lipase and its Application as an Additive in Detergent for Destaining

In this study, we aimed to optimize the nutritional and environmental conditions for the production of a novel lipase (LBL) from Bacillus licheniformis (GenBank accession no. MT118724). This strain was characterized by morphological and biochemical assays and Sanger sequencing of 16S rDNA. The crude lipolytic activity reached a maximum level 7.5 U mL⁻¹ at 40 °C and pH 8.0 using olive oil as substrate. Additionally, the crude enzyme maintained 100% of its initial activity after incubation for 1 h at 50 °C and pH 9.0. It is mandatory to note that LBL lipase displayed appreciable stability over a wide pH range and extreme temperatures. After purification, the optimal lipolytic activity was observed at pH 8.0 and 40 °C. LBL was shown to be a monomeric protein with an estimated molecular weight of 40 kDa. This novel lipase exhibited high stability and excellent compatibility compared to lipase extracted from Thermomyces lanuginosa (Lipolase® from Novozymes, Denmark) toward various detergents. Washing performance analysis revealed that it efficiently removes tomato sauce stain from cotton cloth. All these interesting enzymatic properties favor this new lipase as a potent candidate for applications in detergent formulations.     

Catalytic potential of a poly(AAc‐co‐HPMA‐cl MBAm)‐matrix‐immobilized lipase from a thermotolerant Pseudomonas aeruginosa MTCC‐4713

A purified alkaline thermo‐tolerant lipase from Pseudomonas aeruginosa MTCC‐4713 was immobilized on a series of five noble weakly hydrophilic poly(AAc‐co‐HPMA‐cl MBAm) hydrogels. The hydrogel synthesized by copolymerizing acrylic acid and 2‐hydroxy propyl methacrylate in a ratio of 5 : 1 (HG_5:1 matrix) showed maximum binding efficiency for lipase (95.3%, specific activity 1.96 IU mg^?1 of protein). The HG_5:1 immobilized lipase was evaluated for its hydrolytic potential towards p‐NPP by studying the effect of various physical parameters and salt‐ions. The immobilized lipase was highly stable and retained ～92% of its original hydrolytic activity after fifth cycle of reuse for hydrolysis of p‐nitrophenyl palmitate at pH 7.5 and temperature 55°C. However, when the effect of pH and temperature was studied on free and bound lipase, the HG_5:1 immobilized lipase exhibited a shift in optima for pH and temperature from pH 7.5 and 55°C to 8.5 and 65°C in free and immobilized lipase, respectively. At 1 mM concentration, Fe³⁺, Hg²⁺, NH₄⁺, and Al³⁺ ions promoted and Co²⁺ ions inhibited the hydrolytic activities of free as well as immobilized lipase. However, exposure of either free or immobilized lipase to any of these ions at 5 mM concentration strongly increased the hydrolysis of p‐NPP (by ～3–4 times) in comparison to the biocatalysts not exposed to any of the salt ions. The study concluded that HG_5:1 matrix efficiently immobilized lipase of P. aeruginosa MTCC‐4713, improved the stability of the immobilized biocatalyst towards a higher pH and temperature than the free enzyme and interacted with Fe³⁺, Hg²⁺, NH₄⁺, and Al³⁺ ions to promote rapid hydrolysis of the substrate (p‐NPP). © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 100: 4252–4259, 2006     

Biochemical and structural characterization of triacylglycerol lipase from Penicillium cyclopium

An extracellular lipase, active on water-insoluble triacylglycerols, has been isolated from Penicillium cyclopium. The purified enzyme has a molecular mass of 29 kDa by gel filtration and SDS-polyacrylamide gel electrophoresis. It hydrolyzes emulsions of tributyrin, trioctanoin, and olive oil at the same rate as pancreatic lipase and shows very low activity against partial acylglycerols (monooctanoin and dioctanoin) and methyl esters. It is stable at 35°C for 60 min and has maximal activity in a pH range of 8–10. Hydrolysis of triacylglycerols by P. cyclopium lipase is inhibited by detergents such as Triton X-100. Comparison of the sequence of the 20 first amino acid residues of P. cyclopium triacylglycerol lipase with other Penicillium lipases indicates a high homology with previously characterized lipases produced by P. expansum and P. solitum which are enzymes of comparable size and substrate specificity. Conversely, homology between P. cyclopium lipase and P. simplicissimum lipase, a nonspecific lipolytic enzyme, is low. Penicillium cyclopium triacylglycerol lipase shows no homology with P. camembertii lipase which is specific to monoacylglycerol and diacylglycerol.     

Characterization of Crude and Partially Purified Lipase from Geotrichum candidum Obtained with Different Nitrogen Sources

Lipases from Geotrichum candidum were produced in two different medium: A = 12 % (w/v) clarified corn steep liquor (CCSL) + 0.6 % (w/v) soybean oil (SO) and B = 3.5 % (w/v) yeast hydrolysate (YH) + 0.7 % (w/v) SO. Lipases were partially purified from both media by hydrophobic interaction chromatography using 3.0 mol L^?1 of NaCl as mobile phase, and they were characterized in the crude and partially purified forms. The recovery of lipase activity from CCSL and YH via HIC were 96 and 94.3 %, and the purification factors were 44.3 and 86.7‐fold, respectively. All evaluated lipases had similar optimum pH (7.0–7.7), but, for the CCSL crude lipase, optimum temperature (47 °C) was 10 °C higher than others lipases evaluated. CCSL crude lipase possessed a higher thermo stability than YH crude lipase, e.g., at 37 °C (pH 7.0) the half‐life of CCSL crude lipase was 19.25 h and at pH 8.0 (30 °C) the half‐life was 48 h, which are five and ten times higher than with YH crude lipase, respectively. On the other hand, the YH crude lipase possessed a higher catalytic constant (k_cat = 2.3 min^?1) but with almost the same catalytic efficiency (K_m/k_cat = 32.12 mg mL min^?1) in relation to CCSL crude lipase. The lipases differ in biocatalytic properties between substrates, suggesting that the two lipases can be employed for different applications.     

Some Biochemical Properties of Lipase from Bay Laurel (<Emphasis Type="Italic">Laurus nobilis</Emphasis> L.) Seeds

Lipase was isolated from bay laurel (Laurus nobilis L.) seeds, some biochemical properties were determined. The bay laurel oil was used as the substrate in all experiments. The pH optimum was found to be 8.0 in the presence of this substrate. The temperature optimum was 50 °C. The specific activity of the lipase was found to be 296 U mg protein⁻¹ in optimal conditions. The enzyme activity is quite stable in the range of pH 7.0–10. The enzyme was stable for 1 h at its optimum temperature, and retained about 68% of activity at 60 °C during this time. K _m and V _max values were determined as 0.975 g and 1.298 U mg protein⁻¹, respectively. Also, storage stability and metal effect on lipolytic activity were investigated. Enzyme activity was maintained for 9, 12, and 42 days at room temperature, 4 and −20 °C, respectively. Ca²⁺, Co²⁺, Cu²⁺, Fe²⁺, and Mg²⁺ lightly enhanced bay laurel lipase activity.     

Biocatalytic Properties of Lipase from Walnut Seed (<Emphasis Type="Italic">Juglans regia</Emphasis> L.)

Lipase (E.C. 3.1.1.3) from walnut seed was purified 28.6-fold with 31% yield using Sephadex G-100 gel chromatography. Olive oil served as good substrate for the enzyme. The optimum pH and temperature were 9.0 and 70 °C, respectively. The lipase was stable between 30 and 80 °C for 5 min. K _m and V _max values were determined as 48 mM and 23.06 × 10⁻³ U/min mg for triolein as substrate. Lipase activity was slightly reduced by Cu²⁺, Ca²⁺, Hg²⁺, Mn²⁺, and Ni²⁺ ions, while Mg²⁺ and Zn²⁺ had no effects. Anionic surfactant sodium dodecyl sulfate stimulated lipase activity while non-ionic surfactants Tween-80 and Triton X-100 had negligible effects on enzymatic activity. The enzyme activity was not affected by 50 mM urea and thioacetamide. Potassium ferricyanide, n-bromosuccinamide and potassium cyanide reduced the enzyme activity. The enzyme showed a good stability in organic solvents, the best result being in n-hexane (113% residual activity). The activity of dialysate was maintained approximately 80% for 1 year at −20 °C.     

Biochemical Characterization of Purified Esterase from Soybean (Glycine max) Seed

Alkaline esterase (carboxylic‐ester hydrolases; EC 3.1.1.1) extracted from germinated soybean seeds (Glycine max) was purified approximately 3.6 times by chromatography in a DEAE‐cellulose anion exchange column and filtration in Sephadex G100 gel. The molecular mass of the enzyme was estimated at 45 kDa by gel electrophoresis (SDS‐PAGE). The purified enzyme showed a specific activity of 5.6 U mg^?1 using p‐nitrophenyl butyrate as substrate. The esterase showed optimal activity at 47 °C in moderately alkaline pH, low stability in temperatures higher than 50 °C, and high stability at pH values between 6 and 9.5. The Ca²⁺ and Co²⁺ ions proved to have a positive effect on enzyme activity; however, Hg²⁺ completely inhibited esterase activity. Using p‐nitrophenyl butyrate as substrate, the enzyme showed a K_m of 0.39 mM, V_max of 31.5 mM mg^?1 min^?1 and k_cat 7.60 × 10⁶ s^?1. Regarding substrate affinity, the enzyme showed greater activity for substrates containing short‐chain fatty acids, especially p‐nitrophenyl acetate. Such characteristics give the enzyme great potential for application in the production of low molecular weight esters, in the food industry, and in chemical products. This enzyme is another new member of the family of lipases and esterases from vegetable seeds with high activity and stability in alkaline pH.     

Burkholderia multivorans SB6 Lipase as a Detergent Ingredient: Characterization and Stabilization

In this study, 265 bacterial isolates were collected from kitchen wastewater samples using Rhodamine B agar medium. Of these, 115 isolates were found to respond positively to the addition of commercial detergents. Using 16S rRNA sequence analysis, the isolate demonstrating the high stability towards commercial detergents was identified as Burkholderia multivorans. An SB6 lipase with a molecular mass of 70 kDa was purified from B. multivorans. The purified enzyme showed optimal activity at pH 9.0 and 40 °C and remained stable in the presence of various metal ions, surfactants, and oxidizing agents. The addition of boron compounds improved the pH stability and thermostability of the enzyme, which displayed stability against some commercial detergents; moreover, this stability increased when boron compounds were added to the incubation medium as stabilizers. These properties make SB6 lipase an ideal choice as an additive in detergent formulations.     

Purification and characterization of thermophilic and alkalophilic tributyrin esterase fromBacillus strain A30-1 (ATCC 53841)

An extracellular esterase (EC 3.1.1.1) from a thermophilicBacillus A30-1 (ATCC 53841) was purified 139-fold to homogeneity by sodium chloride (6 M) treatment, ammonium sulfate fractionation (30–80%) and phenyl-Sepharose CL-6B column chromatography. The native enzyme was a single polypeptide chain with a molecular weight of about 65,000 and an isoelectric point at pH 4.8. The optimum pH for esterase activity was 9.0, and its pH stability range was 5.0–10.5. The optimum temperature for its activity was 60°C. The esterase had a half-life of 28 h at 50°C, 20 h at 60°C and 16 h at 65°C. It showed the highest activity on tributyrin, with little or no activity toward long-chain (12–20 carbon) fatty acid esters. The enzyme displayed K_m and K_cat values of 0.357 mM and 8365/min, respectively, for tributyrin hydrolysis at pH 9.0 and 60°C. Cyclodextrin (α, β, and γ), Ca²⁺, Co²⁺, Mg²⁺ and Mn²⁺ enhanced the esterase activity, and Zn²⁺ and Fe²⁺ acted as inhibitors of the enzyme activity. The enzyme activity was not affected by ethylenediaminetetraacetic acid, p-chloromercuribenzoate andN-bromosuccinimide. This paper was presented in part at the 82nd Annual Meeting and Exposition of the American Oil Chemists’ Society, held May 12–15, 1991, in Chicago, Illinois.