Abstract
Maximal productivity of arbutin per cell at various culture stages was examined. Catharanthus roseus cells from early log phase to stationary phase were collected on nylon mesh, then aliquots of cells were suspended in fresh medium or the original medium from which the cells had been removed, for comparison, and hydroquinone was added at 2mM every day until the cells died, in order to obtain the maximal yield. Biotransforming ability based on dry weight of cells in fresh medium dramatically increased with advancing cell stage. In the original medium, however, the biotransforming ability decreased in cells older than day 5. When sucrose (1%) was added to 6-day cells in the original medium, arbutin yield drastically increased. These results suggest that older cells possess higher ability to produce arbutin, but can not manifest this in batch culture because of depletion of essential component (s) in the medium; one of these components was sucrose.
UDPG (uridine 5′-diphosphate glucose) was strongly induced in 6-day cells in fresh medium after the addition of hydroquinone even though it was barely detected before. UDPG had originally been supplied on the 3rd day. In those young cells, the hydroquinone supplied UDPG molecules were presumed to be used competitively both for the glucosylation of hydroquinone and for synthesizing cell wall polysaccharide. The enzyme activity to glucosylate hydroquinone was sufficient in cells on the 3rd, 6th and 8th day. Cell volume of 8-day cells was 1.4-fold as much as that of 3-day cells with the increament of vacuole volume; this might be related to arbutin accumulation.
In conclusion, older cells have a greater ability to produce arbutin by biotransformation. The larger amount of UDPG which is available for the glucosylation of hydroquinone, as well as the larger vacuole, contribute to its ability at least partly.
