Susumu Kyotani

Culture Conditions for Intracellular Lipase Production by Rhizopus chinensis and Its Immobilization within Biomass Support Particles

Acetone-dried cells of Rhizopus chinensis (with a 1,3-positional specificity lipase) were investigated for the interestierification reaction of olive oil and methyl stearate. First, the culture conditions for intracellular lipase production were examined, and then the activities of dried cells obtained from immobilization in Biomass Support Particles (BSPs) were compared with those of freely suspended cells. It was clear from cultivation of freely suspended cells that intracellular lipase activity for the interesterification reaction was enhanced sifnificantly by the presence of oleic acid, oil, and tea oil, but that the presence of glucose reduced the activity. The specific activity of dried cells within BSPs increased 7-fold compared with that obrained from freely suspended cells. The process presented here, using immobilization within BSPs, can provide cells directly as a catalyst with high activity, where cells become immobilized simply during batch operation, and no special preparation of cells is necessary. Therefore, the reaction system using dried cells immobilized within BSPs is a promising interesterifcation process for industrial applications.

Interesterification of fats and oils by immobilized fungus at constant water concentration

Abstract The kinetics of enzymatic interesterification of oils and fats, using acetone-dried cells of Rhizopus chinensis immobilized on biomass support particles as a lipase catalyst, were investigated in batch operations at several constant water concentrations. Even under microaqueous (i.e., low-water-content) conditions, not only interesterification but also hydrolysis occured, and the water content in the reaction system decreased. The reaction rates of interesterification and hydrolysis at constant water concentrations were determined. For the reactions between olive oil and methyl stearate at several water concentrations, the parameters involved in the reaction model were determined by a trial-and-error method so as to make the calculated results correlate with the experimental data. The relationship between the parameters obtained and water concentration were examined. The rate constants involved in the reaction model of both interesterification and hydrolysis increased or decreased monotonically with the increasing water content, while the apparent activity of the lipase catalyst for interesterification had a maximum value at a water concentration of about 50 ppm. This suggests that when the water content is excessive the hydrolysis activity of lipase is accelerated more than its interesterification activity, and that when the water content is too little lipase activity can not be activated for either hydrolysis or interesterification.

Kinetic studies on the interesterification of oils and fats using dried cells of fungus

Abstract The kinetics of enzymatic interesterification of oils and fats, using acetone dried cells of Rhizopus chinensis as a lipase catalyst, have been investigated in a batch operation. To clarify the mechanism of this reaction, several models are discussed under various conditions in terms of the ratio of triglyceride (TG)/fatty acid (FA) and of the water content. First, in the reaction between olive oil and methyl stearate, the parameters involved in each model were determined by the trial-and-error method so as to make the calculated results fit with the experimental data. Then, the models were compared with the experimental data obtained from the reactions with a mixture of stearic and palmitic acid methyl esters, where the proportions of (TG)/(FA) and water content were varied. From the results, the model based on either first order kinetics or on the formation of the glyceride-enzyme complex was confirmed to fit best with the data under a wide range of reaction conditions. This suggests that the fatty acid moiety of TG seems to be exchanged through the glyceride-enzyme complex in the enzymatic interesterification of oils and fats.

Cell aggregation as a trigger for enhancement of intracellular lipase production by a Rhizopus species

Abstract Immobilization of Rhizopus chinensis cells in various Biomass support particles (BSPs) made from polyurethane, nylon, polyester, stainless steel, polyvinyl alcohol, and cellulose enhanced intracellular lipase activity as much as several-fold that of freely suspended cells irrespective of the BSP material. In freely suspended cultivation the overall intracellular lipase activity of free cells, consisting of pellet cells and pulp-like cells, increased with an increase in the ratio of pellet cells to pulp-like cells. It was thus considered that cell aggregation seems to be a principal factor in the enhancement of intracellular lipase formation by Rh. chinensis . Also, other Rhizopus species such as Rh. javanicus, Rh. delemar, Rh. oligosporus, Rh. niveus, Rh. japonicus and Rh. oryzae were immobilized in both polyurethane and cellulose BSPs, In all species used, intracellular lipase production was significantly enhanced compared to results without immobilization. Cell aggregation therefore seems to be a common trigger for the enhancement of intracellular lipase production by Rhizopus species.

Continuous interesterification of oils and fats using dried fungus immobilized in biomass support particles

Abstract The industrial feasibility of an interesterification process using acetone-dried fungus (as a lipase catalyst) immobilized in biomass support particles (BSPs) was examined by continuous interesterification between olive oil and methyl stearate, where the water content of the reaction mixture (Cw) was controlled at a given value. The Cw affected not only the inactivation rate of lipase in the cells but also the production rate of the by-product (diglycerides). The optimal Cw was determined as about 100 ppm. The half-life of lipase in the cells was about 1200 h at the optimal Cw, suggesting that the interesterification process using the immobilized fungus is industrially feasible.

A Novel Interesterification Process for Fats and Oils Using Acetone-Dried Fungus Immobilized in Biomass Support Particles

Lipase not only hydrolyzes triglycerides but also catalyzes transesterification under micro-aqueous condition. Among lipase reactions the production of cocoa butter substitute by transesterification has recently drawn industrial attention. Since lipases are expensive as compared with amylases and proteases, however, the development of a more economical transesterification process is desirable.