Further kinetic characterization of alginate-entrapped cells of Mucuna pruriens L.
Alginate-entrapped cells of Mucuna pruriens L. hydroxylate L-tyrosine, tyramine, para-hydroxyphenylpropionic acid, and para-hydroxyphenylacetic acid to their corresponding catechols, which were released into the incubation medium. Michaëlis-Menten kinetics was applied for each bioconversion. The apparent affinity constants were comparable with the affinity constants obtained with a homogenate directly prepared from the cells used for entrapment and with a derived partly purified phenoloxidase. The values found for the apparent maximum rates of bioconversion of the entrapped cells were ca. 50% of the values of the maximum rates of bioconversion of the cell homogenate, indicating that the entrapped cell system was not operating optimally. The effective diffusivities of the substrates and products were measured with alginate-entrapped, inactivated cells. From the five inactivation methods tested, glutaric aldehyde treatment was chosen as the general procedure. Calculated effective diffusivities for the monophenols and catechols demonstrated that these compounds could diffuse freely into and out of the beads. For each bioconversion, the observable modulus was calculated from the initial rate of bioconversion and the effective diffusivity of the substrate. The resulting values indicated that the diffusional supply rate of the substrates was not the limiting factor, except for the conversion of tyramine for which a modulus higher than one was obtained. Analogously, the observable moduli were calculated for oxygen, which was utilized for bioconversion and cell respiration, and these values pointed towards strong oxygen limitation in all cases. The bioconversion rates of the entrapped cells increased with decreasing cell aggregate size. Therefore, it was concluded that direct cell-matrix contact determined the amount of phenoloxidase involved in the bioconversions. The bioconversion rate on a protein basis was constant with enhancement of the bead charge and thus, in spite of limitations, the mixing conditions as such were relatively optimal. In conclusion, the nonoptimal efficiency of the plant cell system studied was caused by oxygen limitation and a partial phenoloxidase participation, but not by mass transfer limitations for substrates and products with the exception of the conversion of tyramine into dopamine.
Pras
N
N
Department of Pharmacognosy, University of Groningen, A. Deusinglaan 2, 9713 AW Groningen, The Netherlands.
Hesselink
P G
PG
Guikema
W M
WM
Malingré
T M
TM