JOtrRNAL or FERMENTATION AND BIOENGINEERING Vol. 78, No. 4, 293-297. 1994 Hydrolysis of Olive Oil with Lipase in a "VibroMixer" YOSHINOBU KAWANO, 1. SHIRO KIYOYAMA, 1 KOICHIRO SHIOMORI, 1 YOSHINARI BABA, 1 AND TADASHI HANO 2 Department of Materials Science, Miyazaki University, Miyazaki 889-211 and Department of Applied Chemistry, Oita University, Oita 870--11, 2 Japan Received 26 May 1994/Accepted 27 July 1994 Hydrolysis of olive oil in i-octane with Candida cylindracea lipase in aqueous solution was performed in a VihroMixer reactor. The kinetic data were interpreted by the interracial reaction model, whose rate was controlled by the desorption of the product as shown in a previous study of the Lewis-type transfer cell. The desorption rate constant of the product and the equilibrium constant of the reaction between iipase and olive oil were evaluated using the adsorption parameters of lipase. The desorption rate constant was 1.3 x 103 moi/m2h, which was about 3 × 103 times greater than that observed in the Lewis cell. The desorption rate was suspected to be facilitated by the repeated dispersion and coalescence of the oil phase. The equilibrium constant of the interracial reaction was nearly of the same order as that obtained in the Lewis cell. The hydrolysis of lipids with enzyme has been carried out in two phases systems under mild conditions (1, 2, 5, 6, 9). Unfortunately, however the rate of hydrolysis is usually very slow (2, 5, 6, 9). A number of reactors for enzymatic reaction has been developed (1, 2, 4--6), but most of them have a problems under the optimum opera- tion for the hydrolysis (4-6, 9). In order to obtain high lipid conversion either a large reactor volume or long residence time is necessary in these reactors. Acceleration of the reaction and rapid separation of oil and water are required to establish an industrial process. Increasing the interfacial area by strong mixing in a stirred tank is one useful technique for heterogeneous reactions. However, there are problems such as a long separation time for the oil and aqueous solutions, a small surface renewal rate, or discontinuous operation (5). Here, we report on the heterogeneous reaction of olive oil with lipase performed in a VibroMixer, in which vibration and mixing facili- tated the reaction with satisfactory separation of the two phases. The results obtained were analyzed and com- pared with those for the Lewis-type transfer cell. EXPERIMENTAL Reagent Olive oil of analytical grade and i-octane, the solvent for olive oil, of guaranteed reagent grade were used without further purification. Candida cylin- dracea lipase was 905 units/mg-solid purchased from Sigma Co., St. Louis, MO, USA. Experimental apparatus and procedures A VibroMixer, a gift from Reika Co., Tokyo, was used as the reactor. This consists of glass tube 12.9 mm in inter- nal diameter and 190mm in height (Fig. 1) containing vibration plates of ethylene tetrafluoride. The vibration plates (Fig. 2) were connected to the crankshaft of a motor and oscillated at 0 to 3,000 strokes per minute (spm). A buffer solution of pH 7.7-7.8 prepared by aque- ous disodium hydrogenphosphate and potassium hydro- genphosphate was used for the aqueous solution. An aqueous solution of lipase and i-octane solution of olive oil were introduced at a fixed flow rate into the bottom * Corresponding author. of the reactor. The two solutions were mixed by the vibration of the plates. The emulsified effluent from the top of the reactor was centrifuged (4,000 rpm and 2 min) and was quickly separated into the aqueous and i-octane solutions. The concentrations of free fatty acids in the organic solution were determined by titration with alkali solution. In tracer-response runs to clarify the character- istics of the mixing in the reactor, the concentrations of the tracer (aqueous H2SO4) in the effluent were measured under various conditions. A part of the emulsified effluent was collected in 3% aqueous polyvinyl alcohol (PVA) solution. The drop sizes of i-octane were ob- served under a microscope to obtain the interracial area in the emulsified fluid. EXPERIMENTAL RESULTS Characteristics of mixing in the reactor The age distribution, E, of the tracer in a vessel, the residence time distribution, is defined as a function of the concen- tration of the tracer in the effluent as follows (3): ..... l ......... i ............. 1 90x10 m . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I I ~ ~ ""~ Outlet --,- 1.29 X 102m Organic ~ - Aqueous phase phase FIG. 1. VibroMixerreactor. 293