Kinetics and specificity of Lipozyme-catalysed oil hydrolysis in supercritical CO2
Sustainable process-product development & green chemistry
SCF as Solvent Substitutes (T1-8)
Keywords: blackcurrant oil, hydrolysis, supercritical carbon dioxide, lipase, kinetics
Blackcurrant seed oil was dissolved in wet carbon dioxide and continuously passed through a bed of Lipozyme® where its partial hydrolysis to free fatty acids took place under the pressure of 15-28 MPa and temperature of 40 ºC. Lipozyme® is a 1,3-specific lipase from Mucor miehei immobilised on a macroporous ion-exchange resin. The amount of enzyme in the reactor was varied between 15 and 808 mg.
The reaction kinetics was studied in dependence on pressure, flow rate, amount of enzyme and content of water in the reactor. Two analytical methods were used to measure the degree of conversion. The content of free fatty acids in the reaction mixture was evaluated using colorimetric method and the mass fractions of tri-, di-, monoacylglycerols (TG, DG, MG) and free fatty acids (FFA) in the mixture were determined using preparative TLC method. Due to a large stoichiometric excess of water the reaction was assumed to follow irreversible, one-substrate Michaelis-Menten kinetics. The values of Michaelis constant Km and maximum reaction rate were estimated and the rate expressions for TG hydrolysis to DG, MG, and glycerol were integrated for a plug-flow reactor. The enzyme activity was stable unless an increase in water concentration in CO2 solvent above the saturation value caused its partial inactivation.
The composition of TG and DG was measured using reversed-phase HPLC and the fatty acid composition in TG, DG, MG and FFA was determined by GC after their ethylesterification. The composition was affected by the enzyme regiospecificity. The major fatty acids in blackcurrant seed oil are linoleic, oleic, alpha- and gamma-linolenic, palmitic and stearic acids. As saturated fatty acids in triacylglycerols of vegetable oils are preferably at sn-1 and sn-3 positions, the mixture of free fatty acids liberated from the oil was enriched with palmitic and stearic acids and the content of these acids in di- and monoacylglycerols obtained by hydrolysis was, in comparison to oil, decreased. It is interesting that the mixture of free fatty acids was also enriched with alpha-linolenic acid, while gamma-linolenic acid was more concentrated in di- and monoacylglycerols.
Mathematical model for stereospecific hydrolysis was derived and applied to simulate the experimental results.
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Presented Monday 17, 15:00 to 15:20, in session SCF as Solvent Substitutes (T1-8).
