• Biotransformation of aromatic hydrocarbons and organic sulphides by fungi

      Munoz, Benito; Department of Chemistry (Brock University, 1987-10-02)
      Toluene is converted to benzyl alcohol by the fungi Mortierella isabellina and Helminthosporium species; in the latter case, the product is further metabolized. Toluene-a -d 1 , toluene-a,a-d2, and toluene-a,a,a-d 3 have been used with Mortierellaisabellina in a series of experiments to determine both primary and secondary deuterium kinetic isotope effects for the enzymic benzylic hydroxylation reaction. The values obtained, intermolecular primary kH/kD = intramolecular p rim a r y kH r kD = 1. 0 2 + O. 0 5, and sec 0 n dar y k H I kD = 1. 37 .:!. 0.05, suggest a mechanism for the reaction involving benzylic proton removal from a radical intermediate in a non-symmetrical transition state. 2H NMR (30.7 MHz) studies using ethylbenzene-l,1-d 2 , 3 -fluoroethylbenzene-l,1-d 2 , 4 -fluoroethylbenzene-l,1-d 2 , and toluene-dB as substrates with Mortierella isabellina suggest, based on the observable differences in rates of conversion between the substrates, that the hydroxylation of hydrocarbons at the benzylic position proceeds via a one electron abstraction from the aromatic ring, giving a radical cation. A series of 1,3-oxathiolanes (eight) were incubated with Mortierella isabellina , Helminthosporium , Rhizopus arrhizus , and Aspergillus niger . Sulphoxides were obtained from Mortierella isabellina and Rhizopus arrhizus using the substrates 2-phenyl-, 2-methyl-2-phenyl-, and 2-phenyl-2-tert. butyl-l,3-oxathiolane. The relative stereochemistry of 2-methyl-2-phenyl-l,3-oxathiolan-l-oxide was assigned based on lH decoupling, n.O.e, 1 and H NMR experiments. The lH NMR (200 MHz) of the methylene protons of 2-methyl-2-phenyl-l,3-oxathiolan-l-oxide was used as a diagnostic standard in assigning the relative stereochemistry of 2-phenyl-l,3-oxathiolan-l-oxide and 2-phenyl-2-tert. butyl-l,3-oxathiolan-l-oxide. The sulphoxides obtained were consistent with an oxidation occurring from the opposite side of the molecule to the phenyl substituent.
    • Biotransformation of polycylic aromatic compounds by fungi and an investigation into the oxidation of alkylbenzenes by Mortierella isabellina NRRL 1757

      Khan, Shaheer Hasan.; Department of Chemistry (Brock University, 1985-07-09)
      Incubations of several polycyclic heteroaromatic compounds and two polycyclic aromatic hydrocarbons with a series of common fungi have been performed. The fungi Cunninghamella elegans ATCC 26269, Rhizopus arrhizus ATCC 11145, and Mortierella isabellina NRRL 1757 were studied in this regard. Of the aza heteroaromatics, only dibenzopyrrole gave a ring hydroxylated product following the incubation with C. elegans. From the thio heteroaromatics studied, dibenzothiophene was metabolized by all the three fungi and thioxanthone by C. elegans and M. isabellina giving sulfones and sulphoxides. Thiochromanone was metabolized stereoselectively to the corresponding sulphoxide by C. elegans. Methyl substituted thioxanthones on incubation with C. elegans produced oxidative products, arising from S -oxidation and hydroxylation at the methyl group. Of the cyclic ketones studied, only fluorenone was reduced to hydroxyfluorene and this metabolism is compared with that reported with cytochrome P-450 monooxygenases of hepatic microsomes. A series of para-substituted ethylbenzenes has been transformed stereoselectively to the 1-phenylethanols by incubation with M. isabellina. Comparisons of the enantiomeric purities obtained from products with their respective para substituent of the same steric size but different electronic properties indicate that the stereoselectivity of hydroxylation at benzylic carbon may be susceptible to electron donating or withdrawing factors in some cases, but that observation is not va lid in all the comparisons. The stereochemistry of the reaction is discussed in terms of three possible steps, ethylbenzene ---) 1-phenylethanol ---) acetophenone ---) 1-phenylethanol. This metabolic pathway could account for the inconsistencies observed in the comparisons of optical purities and electronic character of para substituents. Furthermore, formation of 2-phenylethanol (in some cases), l-(p-acetylphenyl)ethanol from p-diethylbenzene, and N-acetylation of p-ethylaniline was observed. n-Propylbenzene was also converted to optically active 1-phenylpropanol. Acetophenone, p-ethylacetophenone, and o(,~,~-trifluoroacetophenone were transformed to 1-phenylethanol, l-(p-ethylphenyl)ethanol, and 1-phenyl-2,2,2-trifluoroethanol, respectively, with high chemical and excellent optical yields. The 13 C NMR spectra of several substrates and metabolic products have been reported and assigned for the first time.
    • Host-parasite relationships in mycoparasite /

      Lee, Kwai Yiu Dennis.; Department of Chemistry (Brock University, 1972-06-15)
      A mycoparasite, Piptocephalis virginiana ^ shows a resemblance to fungal parasites of higher plants in the fine structure of hyphae and haustoria. The morphology and fine structure of host and parasitic fungi have been described. The mode of penetration of the host cell, Choanephora cucurbitarum , probably involves mechanical forces. Although the presence of cell wall degrading enzyme was not detected by conventional techniques, its role in penetration can't be ruled out. A collar around the haustorial neck is formed as an extension of the host cell wall. No papilla was detected although appressorixim was seen during penetration. The young haustorium is enclosed in highly invaginating plasmalemma of the host cell and n\imerous cisternae of endoplasmic reticulum. Appearance of an electron—dense sheath around the mature haustorium seems to coincide with the disappearance of cisternae of endoplasmic reticulum from the host cystoplasm in the vicinity of the haustorium. The role of host cytoplasm particularly of endoplasmic reticulum in the development of the sheath is discussed. Extensive accumulation of spherosomes-like bodies, containing lipids, is found in haustorium, parasite and host hypha. Electron microscope revealed the parasiticculture spore has more lipid content than the axenic culture spore of P. virginiana . The biochemical and cytochemical tests also support these results. The mature spore of C. cucurbitarum possesses a thick three-layered cell wall, different from the hyphal wall. Its germination is accompanied by the formation of an elastic thin inner layer which surrounds the emerging germ tube and the growing hypha. High resolution autoradiography showed that H N-acetyl-glucosamine , a precursor of chitin, was incorporated preferentially in the thin inner layer of the spore wall and also in the cell wall of the growing hypha. When the label was fed to the infected cells, at different intervals after inoculation, grains were observed on the sheath which developed around the haustorium of P. virginiana , 30 hours after inoculation. The significance of these results in relation to the origin and composition of the sheath is discussed.
    • Investigation of regio- and sterochemistries of microbial biotransformation /

      Zabic, Mirjana.; Department of Chemistry (Brock University, 1997-05-21)
      Epoxides can be hydrolyzed by fungi to produce chiral diols. The first part of this thesis presents an investigation of the microbial hydrolysis of aziridines comparable in structure to epoxide biotransformation substrates. Biotransformation of the aziridines 1 -methyl-2-phenyl aziridine, 2- phenylaziridine and N-methyl-7-aza bicyclo[4.1.0] heptane was studied using Beauveria sulfurescens, Aspergillus niger and Diplodia gossypina but no evidence for enzymic hydrolysis was obtained. The hydroxylation reaction performed by the fungus Beauveria sulfurescens ATCC 7159 has been studied for many years and several models describing the hydroxylating pattern exhibited by this fungus have been proposed. The second part of this thesis presents a test of the proposed models. The ability of Beauveria sulfurescens to hydroxylate thirty potential substrates was examined, and the data suggest that none of the earlier proposed models accounts for all of the bioconversion results. A possible explanation is proposed, suggesting that there is more than one enzyme responsible for the hydroxylation reactions performed by Beauveria sulfurescens.