Improving the activity and stability of Yarrowia lipolytica lipase Lip2 by immobilization on polyethyleneimine-coated polyurethane foam

In this study, polyurethane foam (PUF) was used for immobilization of Yarrowia lipolytica lipase Lip2 via polyethyleneimine (PEI) coating and glutaraldehyde (GA) coupling. The activity of immobilized lipases was found to depend upon the size of the PEI polymers and the way of GA treatment, with best results obtained for covalent-bind enzyme on glutaraldehyde activated PEI-PUF (MW 70,000 Da), which was 1.7 time greater activity compared to the same enzyme immobilized without PEI and GA. Kinetic analysis shows the hydrolytic activity of both free and immobilized lipases on triolein substrate can be described by Michaelis–Menten model. The K_m for the immobilized and free lipases on PEI-coated PUF was 58.9 and 9.73 mM, respectively. The V_max values of free and immobilized enzymes on PEI-coated PUF were calculated as 102 and 48.6 U/mg enzyme, respectively. Thermal stability for the immobilization preparations was enhanced compared with that for free preparations. At 50 °C, the free enzyme lost most of its initial activity after a 30 min of heat treatment, while the immobilized enzymes showed significant resistance to thermal inactivation (retaining about 70% of its initial activity). Finally, the immobilized lipase was used for the production of lauryl laurate in hexane medium. Lipase immobilization on the PEI support exhibited a significantly improved operational stability in esterification system. After re-use in 30 successive batches, a high ester yield (88%) was maintained. These results indicate that PEI, a polymeric bed, could not only bridge support and immobilized enzymes but also create a favorable micro-environment for lipase. This study provides a simple, efficient protocol for the immobilization of Y. lipolytica lipase Lip2 using PUF as a cheap and effective material.     

Characterization of the lipase immobilized on Mg–Al hydrotalcite for biodiesel

Immobilization of Saccharomyces cerevisiae lipase by physical adsorption on Mg–Al hydrotalcite with a Mg/Al molar ratio of 4.0 led to a markedly improved performance of the enzyme. The immobilized lipase retained activity over wider ranges of temperature and pH than those of the free lipase. The immobilized lipase retained more than 95% relative activity at 50 °C, while the free lipase retained about 88%. The kinetic constants of the immobilized and free lipases were also determined. The apparent activation energies (E_a) of the free and immobilized lipases were estimated to be 6.96 and 2.42 kJ mol^?1, while the apparent inactivation energies (E_d) of free and immobilized lipases were 6.51 and 6.27 kJ mol^?1, respectively. So the stability of the immobilized lipase was higher than that of free lipase. The water content of the oil must be kept below 2.0 wt% and free fatty acid content of the oil must be kept below 3.5 mg KOH g [oil]^?1 in order to get the best conversion. This immobilization method was found to be satisfactory to produce a stable and functioning biocatalyst which could maintain high reactivity for repeating 10 batches with ester conversion above 81.3%.     

Biodiesel-fuel production in a packed-bed reactor using lipase-producing Rhizopus oryzae cells immobilized within biomass support particles

A packed-bed reactor (PBR) system using fungus whole-cell biocatalyst was developed for biodiesel fuel production by plant oil methanolysis. Lipase-producing Rhizopus oryzae cells were immobilized within 6 mm × 6 mm × 3 mm cuboidal polyurethane foam biomass support particles (BSPs) during batch cultivation in a 20-l air-lift bioreactor. Emulsification of the reaction mixture containing soybean oils and water improved the methanolysis reaction rate. Using a high flow rate for the reaction mixture in the PBR caused exfoliation of the immobilized cells from the BSPs, while the inefficient mixing of the reaction mixture at low flow rates allowed the BSPs to be covered with a hydrophilic layer of high methanol concentration, leading to a significant decrease in lipase activity. A high methyl ester content of over 90% was achieved at a flow rate of 25 l/h in the first cycle of repeated batch methanolysis and a high value of around 80% was maintained even after the tenth cycle. Comparison with methanolysis reaction in a shaken bottle suggested that the PBR enhances repeated batch methanolysis by protecting immobilized cells from physical damage and excess amounts of methanol. The process presented here is therefore considered to be promising for industrial biodiesel-fuel production.     

An improved method of lipase preparation incorporating both solvent treatment and immobilization onto matrix

A simple and effective preparation of lipases for use in organic solvents is hereby proposed. Lipases in aqueous solution were treated with isopropanol, immediately followed by immobilization onto a commercially available macroporous resin CRBO2 (crosslinked polystyrene with N-methylglucamine as a functional group). The dual modification of lipases by (1) isopropanol treatment and (2) immobilization improved the activity and stability of lipases more significantly than either of the two treatments alone. The degree of lipase activation was dependent on isopropanol–buffer (v/v) ratio and the source of lipase used. Among the lipases tested, Rhizopus oryzae lipase was more significantly activated. The maximum specific activity of R. oryzae lipase after dual modification was 94.9 mmol h⁻¹ g⁻¹, which was, respectively, 3.3-, 2.5- and 1.5-fold of untreated free, untreated immobilized and treated free lipases. The conformations of the treated and untreated free lipases were investigated by circular dichroism (CD) measurement. Changes in the far- and near-UV CD spectra of lipase indicate that lipase activation is accompanied by changes in secondary and tertiary structures of lipases. The increase in negative molar elipticity at 222 nm suggests that the α-helical content of lipase increase after pretreatment.     

Improving esterification activity of Burkholderia cepacia lipase encapsulated in silica by bioimprinting with substrate analogues

Bioimprinting is a promising, though relatively unexplored, approach to improving the performance of enzymes. In this study, bioimprinting with substrate analogues of fatty acids was systematically conducted to improve the esterification activity of Burkholderia cepacia lipase that had undergone a sol–gel immobilization procedure with methyltrimethoxysilane (MTMS) and tetramethoxysilane (TMOS) as the precursors. The specific activity of the bioimprinted lipases was 3682.0 μmol h^?1 mg protein, which was a 47.9- and 2.5-fold increase over the free and non-imprinted immobilized lipases, respectively. Compared to the free and non-imprinted immobilized lipases, bioimprinted lipases exhibited better thermal stability, and their activity did not change after being incubated at 60 °C for 12 h. Bioimprinted lipases were more easily affected by alcohol than the non-imprinted ones, whose specific activity could be markedly enhanced by ethanol, isopropanol and n-butanol by factors of 1.23-, 1.28- and 1.12-fold, respectively. The reasons for the improvement of imprinted enzyme activity are also discussed based on the surface structure, specific surface area and average pore diameter of the silane particles.     

Combination of two lipases more efficiently catalyzes methanolysis of soybean oil for biodiesel production in aqueous medium

A combination of two lipases was employed to catalyze methanolysis of soybean oil in aqueous medium for biodiesel production. The two lipase genes were cloned from fungal strains Rhizomucor miehei and Penicillium cyclopium, and each expressed successfully in Pichia pastoris. Activities of the 1,3-specific lipase from R. miehei (termed RML) and the non-specific mono- and diacylglycerol lipase from P. cyclopium (termed MDL) were 550 U and 1545 U per ml respectively, and enzymatic properties of these supernatant of fermentation broth (liquid lipase) were stable at 4 °C for >3 months. Under optimized conditions, the ratio of biodiesel conversion after 12 h at 30 °C, using RML alone, was 68.5%. When RML was assisted by addition of MDL, biodiesel conversion ratio was increased to >95% under the same reaction conditions. The results suggested that combination of lipases with different specificity, for enzymatic conversion of more complex lipid substrates, is a potentially useful strategy for biodiesel production.     

Development of a monolith based immobilized lipase micro-reactor for biocatalytic reactions in a biphasic mobile system

This paper reports a simple method for producing macroporous silica-monoliths with controllable porosity that can be used for the immobilization of lipases to generate an active and stable micro-reactor for biocatalysis. A range of commercially available lipases has been examined using the hydrolysis reactions of 4-nitrophenyl butyrate in water–decane media. The kinetic studies performed have identified that a similar value for k_cat is obtained for the immobilized Candida antarctica lipase A (0.13 min⁻¹) and the free lipase in solution (0.12 min⁻¹) whilst the immobilized apparent Michaelis constant K_m (3.1 mM) is 12 times lower than the free lipase in solution (38 mM). A 96% conversion was obtained for the immobilized C. antarctica lipase A compared to only 23% conversion for the free lipase. The significant higher conversions obtained with the immobilized lipases were mainly attributed to the formation of a favourable biphasic system in the continuous flowing micro-reactor system, where a significant increase in the interfacial activation occurred. The immobilized C. antarctica lipase A on the monolith also exhibited improved stability, showing 64% conversion at 80 °C and 70% conversion after continuous running for 480 h, compared to 40 and 20% conversions under the same temperature and reaction time for the free lipase.     

The study on effective immobilization of lipase on functionalized bentonites and their properties

Three different functionalized bentonites including acid activated bentonite (B_a), organically modified bentonite with cetyltrimethyl ammonium bromide (B_CTMAB) and the composite by acid activation and organo-modification (B_a-CTMAB) were prepared, and used for immobilization of lipase from bovine pancreatic lipase by adsorption. The amount of lipase adsorbed on the functionalized bentonites was in the following sequence: B_a > B_CTMAB > B_a-CTMAB, showing the strongest affinity of B_a for lipase among the three supports. However, the immobilized lipase on B_a-CTMAB showed the highest activity in the hydrolysis of olive oil by 1.67 times of activity of free lipase due to the hydrophobically interfacial activation and enlarged catalytic interface. While, the activity of immobilized lipase on B_a was lower than 20% of free lipase’s activity due to the absence of hydrophobic activation and negative impact of excessive hydrogen ions on the surface. The K_m values for the immobilized lipase on B_a-CTMAB (0.054 g/mL) and B_CTMAB (0.074 g/mL) were both lower than that of free lipase (0.115 g/mL), and the V_max values were higher for the immobilized lipases, exhibiting a higher affinity of the immobilized lipase toward olive oil than free lipase. In comparison to free lipase, the better resistance to heating inactivation, storage stability and reusability of the immobilized lipases on B_a-CTMAB and B_CTMAB were also obtained. The results show that the efficient and stable biocatalysts for industrial application can be prepared by using the low-cost bentonite mineral as the supports.     

Effect of high salt concentrations on the stability of immobilized lipases: Dramatic deleterious effects of phosphate anions

We have analyzed the effects of the buffer nature on the stability of immobilized lipases. Commercial phospholipase Lecitase Ultra (LU), lipase B from Candida antarctica (CALB) and lipase from Thermomyces lanuginosus (TLL) have been immobilized on octyl-glyoxyl agarose beads. The enzymes were readily inactivated using 4 M sodium phosphate but 6 M NaCl did not inactivate them. Using 2 M of sodium phosphate, the inactivation of the 3 immobilized enzymes still was very significant even at 25 °C but at lower rate than with higher phosphate concentration. Thermal stress inactivations of the immobilized enzymes revealed that even 100 mM sodium phosphate produced a significant decrease in enzyme stability; this effect was less pronounced for Lecitase but dramatic for CALB. While 6 M NaCl presented slightly positive (LU) or negative (TLL) effects on their thermal stabilities of, CALB was thermally stabilized under the same conditions. Results were very different using free enymes. Fluorescence spectroscopy revealed dramatic structural rearrangements of the immobilized enzymes in the presence of high phosphate concentration. From these results, the use of sodium phosphate does not seem to be recommended for studies on thermal stability of lipases, although this should be verified for each enzyme and immobilized preparation.     

Production of human milk fat substitutes enriched in omega-3 polyunsaturated fatty acids using immobilized commercial lipases and Candida parapsilosis lipase/acyltransferase

In human milk fat (HMF), palmitic acid (20–30%), the major saturated fatty acid, is mostly esterified at the sn-2 position of triacylglycerols, while unsaturated fatty acids are at the sn-1,3 positions, conversely to that occurring in vegetable oils.This study aims at the production of HMF substitutes by enzyme-catalyzed interesterification of tripalmitin with (i) oleic acid (system I) or (ii) omega-3 polyunsaturated fatty acids (omega-3 PUFA) (system II) in solvent-free media. Interesterification activity and batch operational stability of commercial immobilized lipases from Rhizomucor miehei (Lipozyme RM IM), Thermomyces lanuginosa (Lipozyme TL IM) and Candida antarctica (Novozym 435) from Novozymes, DK, and Candida parapsilosis lipase/acyltransferase immobilized on Accurel MP 1000 were evaluated. After 24-h reaction at 60 °C, molar incorporation of oleic acid was about 27% for all the commercial lipases tested and 9% with C. parapsilosis enzyme. Concerning omega-3 PUFA, the highest incorporations were observed with Novozym 435 (21.6%) and Lipozyme RM IM (20%), in contrast with C. parapsilosis enzyme (8.5%) and Lipozyme TL IM (8.2%). In system I, Lipozyme RM IM maintained its activity for 10 repeated 23-h batches while for Lipozyme TL IM, Novozym 435 and C. parapsilosis enzyme, linear (half-life time, t_1/2 = 154 h), series-type (t_1/2 = 253 h) and first-order (t_1/2 = 34.5 h) deactivations were respectively observed. In system II, Lipozyme RM IM showed linear deactivation (t_1/2 = 276 h), while Novozym 435 (t_1/2 = 322 h) and C. parapsilosis enzyme (t_1/2 = 127 h), presented series-type deactivation. Both activity and stability of the biocatalysts depended on the acyl donor used.     

High activity and selectivity immobilized lipase on Fe3O4 nanoparticles for banana flavour synthesis

Lipase (E.C.3.1.1.3) from Thermomyces lanuginosus (TL) was directly bonded, through multiple physical interactions, on citric acid functionalized monodispersed Fe₃O₄ nanoparticles (NPs) in presence of a small amount of hydrophobic functionalities. A very promising scalable synthetic approach ensuring high control and reproducibility of the results, and an easy and green immobilization procedure was chosen for NPs synthesis and lipase anchoring. The size and structure of magnetic nanoparticles were characterized by transmission electron microscopy (TEM) and X-ray diffraction (XRD). The samples at different degree of functionalization were analysed through thermogravimetric measurements. Lipase immobilization was further confirmed by enzymatic assay and Fourier transform infrared (FT-IR) spectra. Immobilized lipase showed a very high activity recovery up to 144% at pH = 7 and 323% at pH = 7.5 (activity of the immobilized enzyme compared to that of its free form). The enzyme, anchored to the Fe₃O₄ nanoparticles, to be easy recovered and reused, resulted more stable than the native counterpart and useful to produce banana flavour. The immobilized lipase results less sensitive to the temperature and pH, with the optimum temperature higher of 5 °C and optimum pH up shifted to 7.5 (free lipase optimum pH = 7.0). After 120 days, free and immobilized lipases retained 64% and 51% of their initial activity, respectively. Ester yield at 40 °C for immobilized lipase reached 88% and 100% selectivity.     

Lipase-catalyzed hydrazinolysis of phenyl benzoate: Kinetic modeling approach

Immobilized lipase-catalyzed synthesis of benzoic acid hydrazide from hydrazine and phenyl benzoate is reported in this work. A series of immobilized lipases such as Candida antarctica lipase B, Mucor miehei lipase and Thermomyces lanuginosus lipase were screened to establish that C. antarctica lipase B was the best lipase for hydrazinolysis. When phenyl benzoate (0.01 mol) and hydrazine (0.02 mol) in toluene (15 ml) were reacted with C. antarctica lipase B (Novozym 435) at 50 °C, 95% of phenyl benzoate was converted to benzoic acid hydrazide after 2 h. The effects of various parameters such as speed of agitation, concentration of the substrates, temperature, enzyme concentration, and reusability of the enzyme were studied to deduce kinetics and mechanism of the reaction. A mechanism based on an ordered bi–bi dead end complex with hydrazine was found to fit the data. Systematic deactivation studies indicated that the enzyme was deactivated due to the hydrazine and phenol, enzyme deactivation obeys first-order series model. The kinetic parameters deduced from these models were used to simulate the lipase activity. There was a very good agreement between the simulated and experimental values.     

Hen egg white as a feeder protein for lipase immobilization

Enzyme stabilization via immobilization is one of the preferred processes as it provides the advantages of recovery and reusability. In this study, Thermomyces lanuginosus lipase has been immobilized through crosslinking using 2% glutaraldehyde and hen egg white, as an approach towards CLEA preparation. The immobilization efficiency and the properties of the immobilized enzyme in terms of stability to pH, temperature, and denaturants was studied and compared with the free enzyme. Immobilization efficiency of 56% was achieved with hen egg white. The immobilized enzyme displayed a shift in optimum pH towards the acidic side with an optimum at pH 4.0 whereas the pH optimum for free enzyme was at pH 6.0. The immobilized enzyme was stable at higher temperature retaining about 83% of its maximum activity as compared to the free enzyme retaining only 41% activity at 70 °C. The denaturation of lipase in free form was rapid with a half-life of 2 h at 60 °C and 58 min at 70 °C as compared to 12 h at 60 °C and 2 h at 70 °C for the immobilized enzyme. The effect of denaturants, urea and guanidine hydrochloride on the free and immobilized enzyme was studied and the immobilized enzyme was found to be more stable towards denaturants retaining 74% activity in 8 M urea and 98% in 6 M GndHCl as compared to 42% and 33% respectively in the case of free enzyme. The apparent K_m (2.08 mM) and apparent V_max (0.95 μmol/min) of immobilized enzyme was lower as compared to free enzyme; K_m (8.0 mM) and V_max (2.857 μmol/min). The immobilized enzyme was reused several times for the hydrolysis of olive oil.     

Immobilization of lipase in organic solvent in the presence of fatty acid additives

In this study porcine pancreatic lipase (PPL) was covalently immobilized on cross-linked polyvinyl alcohol (PVA) in organic media in the presence of fatty acid additives in order to improve its immobilized activity. The effects of fatty acid additions to the immobilization media were investigated choosing tributyrin hydrolysis in water and ester synthesis by immobilized PPL in n-hexane. Various fatty acids which are also the substrates of lipases in esterification reactions were used as active site protecting agents during the immobilization process in an organic solvent. The obtained results showed that covalent immobilization carried out in the presence of fatty acids as protective ligands improved the hydrolytic and esterification activity of immobilized enzyme. A remarkable increase in activity of the immobilized PPL was obtained when octanoic acid was used as an additive and the hydrolytic activity was increased from 5.2 to 19.2 μmol min⁻¹ mg⁻¹ as compared to the non-additive immobilization method. With the increase of hydrolytic activity of immobilized lipase in the presence of octanoic acid, in an analogous manner, the rate of esterification for the synthesis of butyl octanoate was also increased from 7.3 to 26.3 μmol min⁻¹ g⁻¹ immobilized protein using controlled thermodynamic water activities with saturated salt solutions. In addition, the immobilized PPL activity was maintained at levels representing 63% of its original activity value after 5 repeated uses. The proposed method could be adopted for a wide variety of other enzymes which have highly soluble substrates in organic solvent such as other lipases and esterases.     

Immobilization of urease on vapour phase stain etched porous silicon

Porous silicon layers fabricated by the reaction-induced vapor phase stain etch method were coated with 5% polyethylenimine. Urease from Canavalia brasiliensis beans was immobilized on this support through covalent linking with 2.5% glutaraldehyde. The pH and temperature profile of the immobilized and free urease exhibited higher activity at pH 6.5 and 37 °C. After being stored for 30 days at 4 °C, the immobilized enzyme had 75% of the initial activity. The maximum apparent Michaelis constant for free urease (K_m) was 94.33 mM whereas for immobilized urease was 53.04 mM. The maximum reaction velocity (V_max) for free urease was 3.51 mmol/min and for immobilized urease was 1.57 mmol/min.     

Kinetics modeling of the acidolysis with immobilized Rhizomucor miehei lipases for production of structured lipids from sunflower oil

Sunflower oil modification for production of semisolid fats was carried out via acidolysis using palmitic and stearic acids (P + St), hexane and a developed biocatalyst from Rhizomucor miehei lipases. Its kinetic behavior was studied by employing three mathematical models proposed in the literature. Furthermore, a new model was proposed to describe not only the variation of triacylglycerols (TAG), diacylglycerols (DAG), and free fatty acids groups but also the acyl migration reaction occurrence. The effect of the reaction temperature on the kinetic and equilibrium parameters, as well as TAG and reaction intermediates profiles was analyzed. Increasing reaction temperature generated major changes in the overall composition of acylglycerols and gave rise to the highest composition of P + St in the obtained structured lipids (58%, 70 h, 60 °C). P + St incorporation was successfully adjusted by an empirical model (Model I) and a lumped parameter model (Model II) for all the studied reaction times, while the model based on a Ping Pong Bi Bi mechanism (Model III) was only able to describe the kinetics behavior (through the variation of reactant saturated fatty acids concentration) until 24 h. Experimental data were fit satisfactorily by the proposed model (Model IV), showing that the increment in the disaturated TAG formation achieved by the increment in temperature was principally related to the favored DAG formation from triunsaturated TAG.     

Construction of a whole-cell catalyst displaying a fungal lipase for effective treatment of oily wastewaters

To combine the advantage of the oleaginous yeast Yarrowia lipolytica with the high activity of some fungal lipases for oily wastewater treatment, an effective lipase-displaying arming yeast was constructed using the flocculation functional domain of Saccharomyces cerevisiae as the protein anchor. To estimate the effect of the whole-cell oily wastewater treatment, the lipase-displaying arming yeast was added into an open activated sludge bioreactor. Within 72 h of whole-cell treatment, 96.9% of oil and 97.6% of chemical oxygen demand (COD) were removed, while only 87.1% of oil and 91.8% of COD were removed in control A (Y. lipolytica Polg was added), 45.1% of oil and 67.5% of COD were removed in control B (no cell was added) in 72 h. The lipase-displaying arming yeast exhibited remarkable oil removal and COD degradation effect compared with the control samples, exemplifying its application potential.     

Immobilization of alkaline serine endopeptidase from Bacillus licheniformis on SBA-15 and MCF by surface covalent binding

An industrial enzyme, alkaline serine endopeptidase, was immobilized on surface modified SBA-15 and MCF materials by amide bond formation using carbodiimide as a coupling agent. The specific activities of free enzyme and enzyme immobilized on SBA-15 and MCF were studied using casein (soluble milk protein) as a substrate. The highest activity of free enzyme was obtained at pH 9.5 while this value shifted to pH 10 for SBA-15 and MCF immobilized enzyme. The highest activity of immobilized enzymes was obtained at higher temperature (60 °C) than that of the free enzyme (55 °C). Kinetic parameters, Michaelis–Menten constant (K_m) and maximum reaction velocity (V_max), were calculated as K_m = 13.375, 11.956, and 8.698 × 10^?4 mg/ml and V_max = 0.156, 0.163 and 0.17 × 10^?3 U/mg for the free enzyme and enzyme immobilized on SBA-15 and MCF, respectively. The reusability of immobilized enzyme showed 80% of the activity retained even after 15 cycles. Large pore sized MCF immobilized enzyme was found to be more promising than the SBA-15 immobilized enzyme due to the availability of larger pores of MCF, which offer facile diffusion of substrate and product molecules.     

Immobilization of α-galactosidase on galactose-containing polymeric beads

Industrial application of α-galactosidase requires efficient methods to immobilize the enzyme, yielding a biocatalyst with high activity and stability compared to free enzyme. An α-galactosidase from tomato fruit was immobilized on galactose-containing polymeric beads. The immobilized enzyme exhibited an activity of 0.62 U/g of support and activity yield of 46%. The optimum pH and temperature for the activity of both free and immobilized enzymes were found as pH 4.0 and 37 °C, respectively. Immobilized α-galactosidase was more stable than free enzyme in the range of pH 4.0–6.0 and more than 85% of the initial activity was recovered. The decrease in reaction rate of the immobilized enzyme at temperatures above 37 °C was much slower than that of the free counterpart. The immobilized enzyme shows 53% activity at 60 °C while free enzyme decreases 33% at the same temperature. The immobilized enzyme retained 50% of its initial activity after 17 cycles of reuse at 37 °C. Under same storage conditions, the free enzyme lost about 71% of its initial activity over a period of 7 months, whereas the immobilized enzyme lost about only 47% of its initial activity over the same period. Operational stability of the immobilized enzyme was also studied and the operational half-life (t_1/2 was determined as 6.72 h for p-nitrophenyl α-d-galactopyranoside (PNPG) as substrate. The kinetic parameters were determined by using PNPG as substrate. The K_m and V_max values were measured as 1.07 mM and 0.01 U/mg for free enzyme and 0.89 mM and 0.1 U/mg for immobilized enzyme, respectively. The synthesis of the galactose-containing polymeric beads and the enzyme immobilization procedure are very simple and also easy to carry out.     

Lipase immobilized in ordered mesoporous silica: A powerful biocatalyst for ultrafast kinetic resolution of racemic secondary alcohols

Although Burkholderia cepacia lipase (BCL) has been proved to be a potential catalyst for chiral resolution, it is rarely applied in industry because of the low catalysis activity and poor stability of the free form. In this article, BCL was immobilized on the phenyl-modified ordered mesoporous silica (Ph-OMMs) to obtain a novel immobilized lipase. Benefits from the bottle-neck mesoporous structure, high loading of BCL could be completed within only 15 min. When BCL@Ph-OMMs was used as a catalyst for the resolution of 1-phenylethanol, up to 50% conversion with more than 99% ee_s was obtained within only 25 min, which is about 65-folds faster than that of the free lipase. Stabilized BCL@Ph-OMMs was successfully used for the ultrafast resolution of six secondary alcohols by selectivity transesterification, which reached high conversion (50%) and high enantioselectivity (≥99%) within 20–180 min. The activity of BCL@Ph-OMMs was kept relatively constant in 50 consecutive cycles, which is the best result among the reported immobilized lipases. The study suggests that BCL@Ph-OMMs is an attractive catalyst in industrial applications.