Thermostable, alkaline and detergent-tolerant lipase from a newly isolated thermophilic Bacillus stearothermophilus.

Lipases are the enzymes of choice for laundry detergent industries, owing to their triglyceride removing ability from the soiled fabric, which eventually reduces the usage of phosphate-based chemical cleansers in the detergent formulation. In this study, a novel thermo-alkaline lipase-producing strain identified as Bacillus stearothermophilus was isolated from the soil samples of olive oil mill. Enhanced lipase production was observed at 55°C, pH 11 and after 48 h of incubation. Among the substrates tested, xylose (a carbon source), peptone (a nitrogen source) and olive oil at a concentration of 1% were suitable substrates for enhancing lipase production. MgSO4 and Tween-80 were suitable substrates for maximizing lipase production. The enzyme was purified to homogeneity by a single CM-Sephadex column chromatography and revealed molecular mass of 67 kDa. The enzyme (BL1) was active over a wide range of pH from 9.0 to 13.0, with an optimum at pH 11.0, exhibited maximal activity at 55°C and retained more than 70% of its activity after incubation at 70°C or pH 13 for 0.5 h or 24 h, respectively. The enzyme hydrolyzed both short and long-chain triacylglycerols at comparable rates. BL1 was studied in a preliminary evaluation for use in detergent formulation solutions. This novel lipase showed extreme stability towards non-ionic and anionic surfactants after pre-incubation for 1 h at 40°C, and good stability towards oxidizing agents. Additionally, the enzyme showed excellent stability and compatibility with various commercial detergents, suggesting its potential as an additive in detergent formulations.

An oxidant and organic solvent tolerant alkaline lipase by P. aeruginosa mutant: downstream processing and biochemical characterization

An extracellular alkaline lipase from Pseudomonas aeruginosa mutant has been purified to homogeneity using acetone precipitation followed by anion exchange and gel filtration chromatography and resulted in 27-fold purification with 19.6% final recovery. SDS-PAGE study suggested that the purified lipase has an apparent molecular mass of 67 kDa. The optimum temperature and pH for the purified lipase were 45°C and 8.0, respectively. The enzyme showed considerable stability in pH range of 7.0-11.0 and temperature range 35-55 °C. The metal ions Ca2+, Mg2+ and Na+ tend to increase the enzyme activity, whereas, Fe2+ and Mn2+ ions resulted in discreet decrease in the activity. Divalent cations Ca+2 and Mg+2 seemed to protect the enzyme against thermal denaturation at high temperatures and in presence of Ca+2 (5 mM) the optimum temperature shifted from 45°C to 55°C. The purified lipase displayed significant stability in the presence of several hydrophilic and hydrophobic organic solvents (25%, v/v) up to 168 h. The pure enzyme preparation exhibited significant stability and compatibility with oxidizing agents and commercial detergents as it retained 40-70% of its original activities. The values of Km and Vmax for p-nitrophenyl palmitate (p-NPP) under optimal conditions were determined to be 2.0 mg.mL-1 and 5000 μg.mL-1.min-1, respectively.

Computation of interactive effects and optimization of process parameters for alkaline lipase production by mutant strain of Pseudomonas aeruginosa using response surface methodology

Alkaline lipase production by mutant strain of Pseudomonas aeruginosa MTCC 10,055 was optimized in shake flask batch fermentation using response surface methodology. An empirical model was developed through Box-Behnken experimental design to describe the relationship among tested variables (pH, temperature, castor oil, starch and triton-X-100). The second-order quadratic model determined the optimum conditions as castor oil, 1.77 mL.L-1; starch, 15.0 g.L-1; triton-X-100, 0.93 mL.L-1; incubation temperature, 34.12 ºC and pH 8.1 resulting into maximum alkaline lipase production (3142.57 U.mL-1). The quadratic model was in satisfactory adjustment with the experimental data as evidenced by a high coefficient of determination (R²) value (0.9987). The RSM facilitated the analysis and interpretation of experimental data to ascertain the optimum conditions of the variables for the process and recognized the contribution of individual variables to assess the response under optimal conditions. Hence Box-Behnken approach could fruitfully be applied for process optimization.

Optimization of extracellular thermophilic highly alkaline lipase from thermophilic Bacillus sp isolated from Hotspring of Arunachal Pradesh, India

Studies on lipase production were carried out with a bacterial strain (Bacillus sp LBN 2) isolated from soil sample of hotspring of Arunachal Pradesh, India. The cells were cultivated in a mineral medium with maximum production at 1% groundnut oil. The optimum temperature and initial medium pH for lipase production by the organism were 50ºC and 9.0 respectively. The molecular mass was found to be 33KDa by SDS PAGE. The optimal pH and temperature for activity were 10 and 60ºC 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 60ºC and 70% of activity at 70ºC for 1h. The lipase was found to be stable in acetone followed by ethanol. The present findings suggested the enzyme to be thermophilic alkaline lipase.

Screening and characterization of a novel alkaline lipase from Acinetobacter calcoaceticus 1-7 isolated from Bohai Bay in China for detergent formulation

A novel alkaline lipase-producing strain 1-7 identified as Acinetobacter calcoaceticus was isolated from soil samples collected from Bohai Bay, China, using an olive oil alkaline plate, which contained olive oil as the sole carbon source. The lipase from strain 1-7 showed the maximum activity at pH 9.0 under 40ºC. One interesting feature of this enzyme is that it exhibits lipase activity over a broad range of temperatures and good stability. It is also stable at a broad range of pHs from 4.0 to 10.0 for 24 h. Its catalytic activity was highly enhanced in the presence of Ca2+, Mg2+ and K+, but partially inhibited by Cu2+, Al3+, Fe3+ , Ba2+and Zn2+. The fact that it displays marked stability and activity in the presence of TritonX-100, Tween-20, Tween-80, SDS, Hydrogen peroxide, Sodium perborate, Sodium hypochlorite, Sodium citrate, Sodium taurocholate, Glycerine and NaCl suggests that this lipase is suitable as an additive in detergent formulations.

Optimization of physical parameters for lipase production from Arthrobacter sp. BGCC#490.

The physical parameters for the production of thermostable, alkaline lipase from Arthrobacter sp. BGCC# 490 were optimized using response surface methodology (RSM), employing face centered central composite design (FCCCD). The design was employed by selecting pH, temperature and incubation period as the model factors and to achieve maximum yield, interaction of these factors was studied by RSM. A second-order quadratic model and response surface method showed that the optimum conditions for lipase production (pH 10.0, temperature 40oC and incubation period 48 h) resulted in 1.6-fold increase in lipase production (13.75 EUml-1), as compared to the initial level (8.6 EUml-1) after 48 h of incubation, whereas its value predicted by the quadratic model was 12.8 EUml-1. Lipase showed stability in the pH range 8-10 and temperature range 40-60oC, with maximum activity at pH 9.0 and temperature 50oC. Lipase activity was enhanced in the presence of K+, Ca2+ and Mg2+ ions, but inhibited by Hg2+ ions. The enzyme exhibited high activity in the presence of acetone, isopropanol and ethanol, but was unaffected by methanol. These properties suggest that the lipase may find potential applications in the detergent industry. The present work also demonstrated the feasibility of using experimental design tools to optimize physical parameters for lipase production by an indigenous Arthrobacter sp.

Effect of mandur bhasma on lipolytic activities of liver, kidney and adipose tissue of albino rat during CCl4 induced hepatic injury.

'Mandur bhasma', an ayurvedic preparation of iron is used in traditional medicine against hepatitis. In the present study the hepatoprotective property of this drug was tested in albino rats during CC14 induced hepatic injury. The effect of mandur bhasma on the activities of the lipolytic enzymes of rat liver, kidney and adipose tissue were studied during hepatitis induced by CCl4. The activities of acid lipase, alkaline lipase, lipoprotein lipase and hormone sensitive lipase exhibited significant alterations during CCl4 induced hepatic injury, indicating a role for these enzymes in the mobilization of fat from adipose tissue and accumulation of fat in liver and kidney. Simultaneous treatment with mandur bhasma prevented the paraffin mediated and CC14 mediated changes in the enzyme activities. These results suggest the hepatoprotective role of mandur bhasma during CC14 induced hepatic injury.