UBC Theses and Dissertations
The cinnamic acid pathway and hispidin biosynthesis in cultures of Polyporus hispidus Fries Perrin, Peter William
The biosynthesis of hispidin, 6-(3,4-dihydroxystyryl)-4-hydroxy-2-pyrone, was examined in cultures of Polyporus hispidus Fr. Cultural studies were undertaken to determine the most suitable medium for investigating the biosynthesis of this pigment. These studies showed that light was necessary for hispidin formation and that the development of basidiocarps with viable spores could be achieved on agar media. On the liquid medium employed for biochemical studies, the maximum rate of hispidin production was observed to lag the maximum rate of growth by about five days. Trimethylhispidin, 4-methylhispidin and yangonin were synthesized for comparative purposes and for dilution in tracer experiments. These and numerous other phenolic and aromatic compounds were employed as references in examinations of nonradioactive culture extracts. P-coumaric, caffeic, p-hydroxybenzoic, protocatechuic and o- and p-hydroxyphenyl-acetic acids were detected in extracts of the culture medium. Bis-noryangonin (6-(4-hydroxystyryl)-4-hydroxy-2-pyrone) and other possible styrylpyrones were detected in extracts of the mycelium. Tracer experiments established that phenylalanine was metabolized to cinnamic, benzoic, p-hydroxybenzoic and protocatechuic acids. The incorporation of radioactivity into phenyllactic, phenylpyruvic, phenylacetic and p-hydroxy-phenylacetic acids also was observed. Furthermore, p-coum-aric acid, caffeic acid and hispidin incorporated radioactivity from phenylalanine. Hispidin also was shown to incorporate radioactivity from tyrosine, cinnamic acid, p-coum-aric acid, caffeic acid, malonic acid and sodium acetate. Degradation of the labelled hispidin obtained from these precursors confirms the hypothesis that this molecule is bio-synthesized from a phenylpropanoid moiety with the addition of two equivalents of acetate. Crude and partially purified preparations of several enzymes related to aromatic metabolism were obtained. Phenylalanine and tyrosine ammonia-lyase activity were demonstrated in cell-free preparations. Maximum phenylalanine ammonia-lyase activity was obtained from cultures during the logarithmic phase of growth. Enzymes capable of hydroxylating cinnamic acid, benzoic acid and bis-noryangonin also were obtained in vitro.
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