Metabolism of Radiolabeled β-Guaiacyl Ether-Linked Lignin Dimeric Compounds by Phanerochaete chrysosporium

Phanerochaete chrysosporium metabolized the radiolabeled lignin model compounds [γ-¹⁴C]guaiacylglycerol-β-guaiacyl ether and [4-methoxy-¹⁴C]veratrylglycerol-β-guaiacyl ether (VI) to ¹⁴CO₂ in stationary and in shaking cultures. ¹⁴CO₂ evolution was greater in stationary culture. ¹⁴CO₂ evolution from [γ-¹⁴C]guaiacyl-glycerol-β-guaiacyl ether and [4-methoxy-¹⁴C]veratrylglycerol-β-guaiacyl ether in stationary cultures was two- to threefold greater when 100% O₂ rather than air (21% O₂) was the gas phase above the cultures. ¹⁴CO₂ evolution from the metabolism of the substrates occurred only as the culture entered the stationary phase of growth. The presence of substrate levels of nitrogen in the medium suppressed ¹⁴CO₂ evolution from both substrates in stationary cultures. [¹⁴C]veratryl alcohol and 4-ethoxy-3-methoxybenzyl alcohol were formed as products of the metabolism of VI and 4-ethoxy-3-methoxyphenylglycerol-β-guaiacyl ether, respectively.     

Transesterification/acetylation of long chain alcohols with alkyl acetate

Gas chromatographic characterizations of fatty alcohols are generally carried out as the free alcohols, trimethyl silyl or acetyl derivatives. In this study, transesterification/acetylation of long chain fatty alcohols is simply carried out by dissolving the alcohol in ethyl/methyl acetate and passing through a micro-column packed with solid NaOH. Reaction times are slightly different for alcohols of different chain length. Rice bran alcohols of 24–34 carbon atom are successfully acetylated. Also, castor oil methyl ester can be interesterified but with longer reaction time.     

A Phanerochaete chrysosporium mutant defective in lignin degradation as well as several other secondary metabolic functions

A pleiotropic mutant of Phanerochaete chrysosporium 104-2 lacking phenol oxidase and unable to form fruit bodies and a revertant strain 424-2 were isolated after UV mutagenesis. Strains 104-2 and 424-2 had no apparent dysfunction in primary metabolism with glucose as a carbon source. Unlike the wild type strain and strain 424-2, strain 104-2 was unable to evolve ¹⁴CO₂ from ¹⁴C ring, side chain and 3-O-¹⁴C-methoxy labeled lignin. In addition, strain 104-2 was unable to evolve ¹⁴CO₂ from a variety of lignin model compounds including ¹⁴C-4-methoxy labeled veratrylglycerol--guaiacyl (V) ether, -¹⁴C-guaiacylglycerol--guaiacyl ether (VI), as well as 1-(¹⁴C-4-methoxy, 3-methoxyphenyl)1,2 propene (III) and 1-(¹⁴C-4-methoxy-3-methoxyphenyl) 1,2 dihydroxypropane (IV). The addition of peroxidase/H₂O₂ to cultures of strain 104-2 did not alter its capacity to degrade the labeled lignins. A variety of unlabeled lignin model compounds previously shown to be degraded by the wild type organism including -aryl ether dimers and diaryl propane dimers were also not degraded by the mutant 104-2. The revertant strain 424-2 regained the capacity to degrade these compounds. The substrates described are degraded by oxygen requiring system(s) expressed during the secondary phase of growth, suggesting this pleiotropic mutant is possibly defective in the onset of postprimary metabolism. The inability of the mutant to produce the secondary metabolite veratryl alcohol and to elaborate enzymes in the veratryl alcohol biosynthetic pathway supports this hypothesis.Abbreviations GLC gas liquid chromatography - TMSi trimethylsilyl - MS mass spectrometry - LDS lignin degrading system     

Studies on the mechanism of 3-ketosphinganine synthetase.

The biosynthesis of sphinganine and 4-D-hydroxysphinganine was studied in rat liver microsomes and whole cells of yeast (Hansenula ciferri). It was shown in both cases that the condensation of [2,3,3-2H3]serine and palmitic acid yielded long chain bases containing only two deuterium atoms, both of which were located on the terminal (C-1) carbon atom by combined gas-liquid chromatography/mass spectrometry. When the reaction with the liver microsomal system was carried out in 2H2O with the protium species of serine, the sphinganine contained a deuterium atom on C-2. These results suggest that the synthesis of 3-ketosphinganine involves the replacement of the alpha-hydrogen atom and the carboxyl group of serine by a proton from the medium and a palmitoyl group, rather than a previously proposed mechanism in which the alpha-hydrogen of serine is retained. Some stereochemical requirements of 3-ketosphinganine synthetase are discussed.     

Peroxidase catalysed oxidative decarboxylation of vanillic acid to methoxy-p-hydroquinone

Horseradish peroxidase catalysed the oxidative decarboxylation of vanillic acid to methoxy-p-hydroquinone and subsequent oxidation of the hydroquinone to methoxy-p-benzoquinone. Peroxidase also catalysed the oxidation of vanillyl alcohol to vanillin and vanillic acid; however, neither vanillyl alcohol nor vanillin appeared to give rise to methoxyhydroquinone directly. Correspondingly, peroxidase catalysed the oxidative decarboxylation of syringic acid to 2,6-dimethoxy-p-hydroquinone and subsequent oxidation of the hydroquinone to 2,6-dimethoxy-p-benzoquinone.     

Metabolism of Radiolabeled beta-Guaiacyl Ether-Linked Lignin Dimeric Compounds by Phanerochaete chrysosporium

Phanerochaete chrysosporium metabolized the radiolabeled lignin model compounds [gamma-C]guaiacylglycerol-beta-guaiacyl ether and [4-methoxy-C]veratrylglycerol-beta-guaiacyl ether (VI) to CO(2) in stationary and in shaking cultures. CO(2) evolution was greater in stationary culture. CO(2) evolution from [gamma-C]guaiacyl-glycerol-beta-guaiacyl ether and [4-methoxy-C]veratrylglycerol-beta-guaiacyl ether in stationary cultures was two- to threefold greater when 100% O(2) rather than air (21% O(2)) was the gas phase above the cultures. CO(2) evolution from the metabolism of the substrates occurred only as the culture entered the stationary phase of growth. The presence of substrate levels of nitrogen in the medium suppressed CO(2) evolution from both substrates in stationary cultures. [C]veratryl alcohol and 4-ethoxy-3-methoxybenzyl alcohol were formed as products of the metabolism of VI and 4-ethoxy-3-methoxyphenylglycerol-beta-guaiacyl ether, respectively.     

Mass spectra of various deuterium-labelled forms of bis-O-trimethylsilyl-N-acetylsphinganine

The mass spectrum of the trimethylsilyl derivative of N-acetylsphinganine (dihydrosphingosine) has been studied in detail. Origins, structures and mechanisms of formation of the principal ions are supported by deuterium labelling and exact masses. The ions formed on electron impact ionization at 70 eV can be divided into two main categories with respect to electron abstraction from one of the oxygen atoms or the nitrogen atom on the long-chain carbon backbone.