Immobilized Lipase Reactors

Malcata, F. X., H. Reyes, H. Garcia, C. Hill, and C. Amundson. 1990. Immobilized Lipase Reactors for Modification of Fats and Oils—A Review. Journal of the American Oil Chemists Society 67 (12) 890-910. 

Malcata, FX, HR Reyes, HS Garcia, CG HiU Jr, CH Amundson. (1990). Immobilized lipase reactors for modifications of fats and oils A review. J Am Oil Chem Soc 67 890-910. 

The catalysts was added after the reactants were fed in the tank reactor and pressure and temperature were set to the target values [84]. The study was performed using an immobilized lipase, Novozym-435 , as biocatalyst. The temperature was set to 65-75 °C and the pressure was reduced (60 mmHg). A catalyst concentration of 1-5% with an acid alcohol ratio of 1 3, 1 1 or 3 1 was used. 

One of the most promising applications of enzyme-immobilized mesoporous materials is as microscopic reactors. Galameau et al. investigated the effect of mesoporous silica structures and their surface natures on the activity of immobilized lipases [199]. Too hydrophilic (pure silica) or too hydrophobic (butyl-grafted silica) supports are not appropriate for the development of high activity for lipases. An adequate hydrophobic/hydrophilic balance of the support, such as a supported-micelle, provides the best route to enhance lipase activity. They also encapsulated the lipases in sponge mesoporous silicates, a new procedure based on the addition of a mixture of lecithin and amines to a sol-gel synthesis to provide pore-size control. 

Monoglyceride (MG) is one of the most important emulsifiers in food and pharmaceutical industries [280], MG is industrially produced by trans-esterification of fats and oils at high temperature with alkaline catalyst. The synthesis of MG by hydrolysis or glycerolysis of triglyceride (TG) with immobilized lipase attracted attention recently, because it has mild reaction conditions and avoids formation of side products. Silica and celite are often used as immobilization carriers [281], But the immobilized lipase particles are difficult to reuse due to adsorption of glycerol on this carriers [282], PVA/chitosan composite membrane reactor can be used for enzymatic processing of fats and oils. The immobilized activity of lipase was 2.64 IU/cm2 with a recovery of 24%. The membrane reactor was used in a two-phase system reaction to synthesize monoglyceride (MG) by hydrolysis of palm oil, which was reused for at least nine batches with yield of 32-50%. 

Bouwer, S.T., Cuperms, F.P. and Derksen, J.T.P. (1997) The performance of enzyme-membrane reactors with immobilized lipase. Enzyme and Microbial Technology, 21, 291-296. 

A lipase from Rhizomucor miehei is immobilized onto a porous polyamide support with a bead diameter of 0.3 mm and used for the hydrolysis of 100 mM caprylin at pH 7.0 and 30°C, in a continous plug flow reactor. Calculate the amount of immobilized lipase needed to achieve a conversion degree of 97% with an inlet flow rate of 1 l.min ... 

A lipase-immobilized membrane reactor was applied for the optical resolution of racemic naproxen, lipase stability was enhanced by the EMR set-up to > 200 h in comparison with a half-life of 2 h in a stirred tank. Only pure lipase gave the best enantioselectivity (Sakaki, 2001). 

To understand the potential of pressure application to enzyme processes and to help elucidate the reaction mechanism as well as a rational design of alcoholysis reactors for future scale-up, we investigated the influence of temperature, pressure, exposure times, and decompression rates on the activity of a commercial immobilized lipase (Novozym 435) activity in high-pressure C02 medium. 

Esterification reactions were carried out in a closed reactor with 10 mL of dried n-heptane containing suitable amounts of alcohol and acid. A molecular sieve (aluminum sodium silicate, type 13X BHD Chemicals) was used to removal water. The mixture was incubated at 37°C for 24 h with continuous shaking at 150 rpm. The effects of concentration of immobilized lipase (5-50 mg/mL) molar ratio of reactants (0.5-2.0), acid chain length... 

The enzymatic esterification of oleic acid and oleyl alcohol to obtain oleyl oleate, which is a synthetic analogue of jojoba oil, was studied. The reaction was catalyzed by a commercially available immobilized lipase from Rhizomucor miehei. As solvents, carbon dioxide and liquid n-butane were used. Reactions were performed in a batch and in continuously operating high pressure reactors. 

Reaction performance. Supercritical carbon dioxide was used as a reaction media for the enzymatic synthesis of oleyl oleate directly from oleic acid and oleyl alcohol. Reaction was catalyzed by immobilized lipase from Rhizomucor miehei-Ltpozyme IM. Reactions were carried out in the high pressure batch and continuous reactor. 

Immobilization of lipases on hydrophobic supports has the potential to (1) preserve, and in some cases enhance, the activity of lipases over their free counterparts (2) increase their thermal stability (3) avoid contamination of the lipase-modified product with residual activity (4) increase system productivity per unit of lipase employed and (5) permit the development of continuous processes. As the affinity of lipases for hydrophobic interfaces constitutes an essential element of the mechanism by which these enzymes act, a promising reactor configuration for the use of immobilized lipases consists of a bundle of hollow fibers made from a microporous hydrophobic polymer (137). 

Shukla A and Kumar A. Experimental studies and mass-transfer analysis of the hydrolysis of ohve oil in a biphasic zeohte-membrane reactor using chemically immobilized lipase. Ind Eng Chem Res 2004 43(9) 2017-2029. 

Membrane reactors using biological catalysts can be used in enantioselective processes. Methodologies for the preparation of emulsions (sub-micron) of oil in water have been developed and such emulsions have been used for kinetic resolutions in heterogeneous reactions catalyzed by enantioselective enzyme (Figure 43.4). A catalytic reactor containing membrane immobilized lipase has been realized. In this reactor, the substrate has been fed as emulsion [18]. The distribution of the water organic interface at the level of the immobUized enzyme has remarkably improved the property of transport, kinetic, and selectivity of the immobilized biocatalyst. 

Kinetic resolution of chiral alcohols by selective esterification has also been performed in SCCO2 by several groups. A glass-immobilized lipase from Candida cylindracea was found to catalyze the stereoselective esterification of only the S isomer from racemic citronellol (Eq. (3)] in a continuous-flow reactor (78) ... 

Hollow-fiber membrane reactors with immobilized lipases have been used for the continuous hydrolysis of triglycerides188 and in the esterification of fatty acids.189 There was no deactivation of the enzyme in the former case in 16 days. In a comparable run in solution, the enzyme lost 80% of its activity in 2 days of operation. The latter case used dodecanol and decanoic acid in hexane to give the ester in 97% yield. The half-life of the immobilized enzyme was 70 days. The integration of reaction and separation can decrease product inhibition, increase selectivity, shift equilibria, and reduce the number of downstream operations.190... 

In this example, lipase is immobilized on different carbon monoliths and applied in a transesterification reaction in toluene. The biocatalysts are compared in terms of carrier preparation, enzyme immobilization, and performance. A commercially available immobilized lipase is used as a comparison. A convenient tool to compare monolithic biocatalysts is the monolithic stirrer reactor (MSR), consisting of two monoliths that have the catalyst immobilized on the wall of their channels. These monoliths work as stirrer blades that can easily be removed from the reaction medium, thereby eliminating the need for a filtration step after reaction [37]. 

Catalytic tests with the lipase-monolithic catalysts were performed in a monolithic stirrer reactor consisting of a glass vessel equipped with a stirrer motor (V = 2.5 dm ). 1-Butanol and vinyl acetate concentrations were 0.6 M and 1 M, respectively. Activity tests with immobilized lipase Candida antarctica) were performed at varying stirrer rates and temperatures. Carbon monoliths (Westvaco integral carbon monoliths, with a loading of 30 wt% of microporous activated carbon, wall thickness 0.3 mm) were used as a reference material. 

During research on enzymatic acylation, Gelo-Pujic et al. [87] performed Novo-zym immobilized lipase-catalyzed esterification of 131a,b and 7 with dodecanoic acid in dry media both with microwave irradiation and with classical heating, using the same conditions of time and temperature (Scheme 12.54). Esterification of 131a by use of a focused Synthewave reactor and oil bath, both at 95 °C, afforded 132 in 95% and 55% yield, respectively. 

Laudani CG, Habulin M, Knez Z et al. (2007) Immobilized lipase-mediated long-chain fatty acid esterification in dense carbon dioxide bench scale packed-bed reactor study. J Supercrit Eluids 41 74-81... 

Continuous Flow Reaction. Two fixed-bed bioreactors packed with 2.0 g immobilized lipase were prepared. The first-step reaction was conducted by continuously feeding a mixture of DHA/ethanol (1 1, mol/mol) into one reactor at 30°C at a flow rate of 4.5 mL/h (4.0 g/h). The reaction mixture that flowed out of the above reactor was collected once each week and dehydrated. Five molar equivalents of ethanol was added to the dehydrated mixture, and the resulting substrates were fed into the second reactor at the same temperature and flow rate as those in the first-step reaction. The contents of DHAEE in the first- and second-step reaction mixtures are plotted in... 

An example of an industrial membrane bioreactor is the hollow-fiber membrane system for the production of (-)-MPGM (1), which is an important intermediate for the production of diltiazem hydrochloride [130, 131]. For the enantiospecific hydrolysis of MPGM a hoUow-fiber ultrafiltration membrane with immobilized lipase from Serratia marcescens is used. (-i-)-MPGM is selectively converted into (2S,3R)-(-i-)-3-(4-methoxy-phenyl)glycidic acid and methanol. The reactant is dissolved in toluene, whereas the hydrophilic product is removed via the aqueous phase at the permeate side of the membrane (see Fig. 5.17). Enantiomerically pure (-)-MPGM is obtained from the toluene phase by a crystallization step. In cooperation with Sepracor Inc., a pilot-plant membrane reactor has been developed, which produces annually about 40 kg (-)-MPGM per m of membrane surface. 

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