A purified alkaline thermo-tolerant bacterial lipase from Pseudomonas aeruginosa BTS-2 was immobilized on a poly (AAc-co-HPMA-cl-MBAm) hydrogel network. The hydrogel showed approximately 95% binding efficiency for lipase (specific activity 1.96 U mg )1 ). The immobilized enzyme achieved 65.1% conversion of ethanol and propionic acid (100 mM each) into ethyl propionate in n-nonane at 65°C in 9 h. When alkane of C-chain length lower than n-nonane was used as the organic solvent, the conversion of ethanol and propionic acid into ethyl propionate decreased with a decrease in the log P value of alkanes. The immobilized lipase retained approximately 30% of its original catalytic activity after five cycles of reuse for esterification of ethanol and propionic acid into ethyl propionate at temperature 65°C in 3 h. Addition of a molecular sieve (3 Å ) to the reaction mixture enhanced the formation of ethyl propionate to 89.3%. Moreover, ethanol and propionic acid when taken a molar ratio of 3:1 further promoted the conversion rate to 94%. However, an increase in the molar ratio of propionic acid with respect to ethanol resulted in a decline of ethyl propionate synthesis.

Lipase (EC 3.1.1.3) is a tri-acylglycerol ester hydrolase, catalysing the hydrolysis of tri-, di-, and mono-acylglycerols to glycerol and fatty acids. To study the effect of adsorption of a lipase obtained from Bacillus coagulans BTS-1, its lipase was immobilized on native and activated (alkylated) matrices, i.e. silica and celite. The effect of pH, temperature, detergents, substrates, alcohols, organic solvent etc. on the stability of the immobilized enzyme was evaluated. The gluteraldahyde or formaldehyde (at 1% and 2% concentration, v/v) activated matrix was exposed to the Tris buffered lipase. The enzyme was adsorbed/entrapped more rapidly on to the activated silica than on the activated celite. The immobilized lipase showed optimal activity at 50 degrees C following one-hour incubation. The lipase was specifically more hydrolytic to the medium C-length ester (p-nitro phenyl caprylate than p-nitro phenyl laurate). The immobilization/entrapment enhanced the stability of the lipase at a relatively higher temperature (50 degrees C) and also promoted enzyme activity at an acidic pH (pH 5.5). Moreover, the immobilized lipase was quite resistant to the denaturing effect of SDS.

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Extracellular lipase from Bacillus coagulans BTS-3 was immobilized on activated (alkylated, 2.5% glutaraldehyde) and native (nonactivated) polyethylene powder, and its thermostability and esterification efficiency were studied. Immobilization on activated support was found to enhance thermostability as well as esterification efficiency. The optimum time for immobilization on activated (AS) and nonactivated (NS) polyethylene support was found to be 10 min, and the binding of the lipase was markedly higher on AS. Lipase was more efficiently bound to AS (64%) than to NS (30%) at an optimum temperature of 378C. The pH and temperature optima for AS-and NS-bound lipase were 9.0 and 558C and 8.5 and 558C respectively. At 558C the free lipase, which had a half-life of 2 h, lost most of its activity at elevated temperatures. In contrast, AS-bound lipase retained 60%-80% of its original activity at 558C, 608C, 658C, and 708C for 2 h. Exposure to organic solvents resulted in enhanced lipase activity in n-hexane (45%) and ethanol (30%). Both AS-and NS-bound biocatalysts were recyclable and retained more than 85% of their initial activity up to the fourth cycle of hydrolysis of p-nitrophenyl palmitate. The AS-bound lipase efficiently performed maximum esterification (98%) of ethanol and propionic acid (300 mM each, 1 : 1) in n-hexane at 558C. With free or NS-bound lipase in similar conditions, the conversion of reactants into ester was relatively low (40%).