Microbial transformation of neutral fraction and upgraded neutral fraction of Polish tall oil

Summary Microbial transformations of neutral fraction (NF) and upgraded neutral fraction (UNF) of Polish tall oil byMycobacterium sp. MB 3683 were performed. Final metabolites and yields were compared to bioconversion of pure -sitosterol. Additionally, origin of a new metabolite —5-androsta-3,6,17-trione was proved by transformation of UNF in the presence of labeled -sitosterol.     

Effect of applied and internal hormones on vitrification and apical necrosis of different plants cultured in vitro

Development of vitrification and apical necrosis was followed in Camellia sinensis, Gerbera jamesonii, Malus domestica and hybrid Populus tremula x P. alba shoots cultured in vitro on Murashige & Skoog (MS) medium with different concentrations of growth regulators. High humidity in the culture vessels and excess of BA in the medium were found to be the major factors influencing vitrification. Lack of exogenous cytokinin in the medium during successive subcultures induced apical necrosis in poor-rooting species (Malus domestica, Camellia sinensis). The level of internal phytohormones (ABA, IAA, IPA, 2iP, Z, ZR) was determined in the apple shoots by means of ELISA. The content of internal cytokinins in the vitrified apple shoots was several times greater than in normal ones, which supports the hypothesis that excess of cytokinins, inducing rapid divisions of cells in meristems in the atmosphere with high humidity, is responsible for vitrification. Apical necrosis of the plantlets that appeared after cultivation on cytokinin-free medium is the result of deficiency in endogenous hormones in apple shoots and this being confirmed by analysis of endogenous hormones in apple shoots.Abbreviations BA benzyladenine - BHT butylated hydroxy-toluene - ABA abscisic acid - IAA indole-3-acetic acid - ELISA enzyme-linked immunosorbent assay - IPA isopentenyladenosine - 2iP isopentenyladenine - NAA naphthyl-3-acetic acid - TBS trishydroxymethylaminomethane buffered saline - TLC thin layer chromatography - Z zeatin - ZR zeatin riboside     

Quinolinate-like neurotoxicity produced by aminooxyacetic acid in rat striatum

Summary The endogenous tryptophan metabolite quinolinic acid elicits in rodent brain a pattern of neuronal degeneration which resembles that caused by L-glutamate. Its qualities as a neurotoxic agent raised the hypothesis that quinolinic acid might be involved in the pathogenesis of human neurodegenerative disorders. Kynurenic acid, another endogenous tryptophan metabolite and preferential N-methyl-D-aspartate (NMDA) antagonist, has been shown to block quinolinic acid neurotoxicity. Here we report that microinjections of aminooxyacetic acid (AOAA), an inhibitor of kynurenine transaminase and of other pyridoxal phosphate-dependent enzymes, into the rat striatum produce neuronal damage resembling that caused by quinolinic acid. AOAA-induced striatal lesions can be prevented by kynurenic acid and the selective NMDA antagonist 2-amino-7-phosphonoheptanoic acid. These results suggest that AOAA produces excitotoxic lesions by depleting brain concentrations of kynurenic acid (inhibition of synthetic enzyme) or due to impairment of intracellular energy metabolism (depletion of cell energy resources). The concept of deficient neuroprotection due to metabolic defects might help to clarify the pathogenesis of human neurodegenerative disorders and to develop strategies that may be useful in their treatment.This work was supported by research grant from the Polish Academy of Sciences.These data have been communicated to the International Congress on Amino Acid Research held in Vienna in August 7–12, 1989.     

Stabilization of steroid 11-hydroxylation activity ofCunninghamella elegans protoplasts in organic osmotic stabilizers

Protoplasts ofCunninghamella elegans, showing 11-, and 11-hydroxylating ability of Substance S, preserved high transformation activity when dispersed in glucose-enriched, organic osmotic stabilizers. A joint action of polyoxins and 2-deoxy-d-glucose was necessary to prevent regeneration of the cell wall in long-lasting experiments. Stabilized and active, dispersed protoplasts may be an alternative research model for studying the function of the cell wall and intracellular metabolic pool constituents in steroid hydroxylation.     

Cumulation of mycotoxins in maize cobs infected with<Emphasis Type="Italic">Fusarium gramihearum</Emphasis>

Maize cobs withFusarium ear rot were collected at 1986 season and five infected byFusarium graminearum were analyzed for presence of triohothecenes and zearalenone. Collected material was subsampled forFusarium damaged kernels and corresponding axial stems and healthy looking kernels. All investigated cobs contained deoxynivalenol (DON) (range 18.0–131.5 mg/kg) and zearalenone (ZEA) (range 0.38–2.17 mg/kg), in four cobs 15-acetyl-deoxynivalenol (15-AcDON) (range 5.2–6.2 mg/kg) was present and two cobs besides three all metabolites contained 3-acetyl-deoxynivalenol (3-AcD0N) (range 0.5–0.8 mg/kg).The average of individual toxins amount in axial stems: in mg/kg was equal to: DON — 110.36, ZEA — 4.57, 15-AcD0N — 16.66, and 3-AcD0N — 1.32.Fusarium damaged kernels contained in average the following amount (mg/kg) of: DON 77.00, ZEA 0.98, 15-AcD0N 3.78 and 3-AcD0N 0.06. Healthy looking kernels contained DON 1.96 mg/kg and ZEA 0.07 mg/kg only. Cooccurrence of 3-AcDON and 15-AcDON in two samples was an interesting finding. The amount of DON in total cob was highly correlated (r = 0.94) with percentage ofFusarium damaged kernels in given ear.     

Expedient syntheses of neoglycoproteins using phase transfer catalysis and reductive amination as key reactions

Starting from peracetylated chloro- or bromo-glycosyl donors ofN-acetylneurmainic acid,N-acetylglucosamine, glucose and lactose, the correspondingp-formylphenyl glycosides were synthesized stereospecifically under phase transfer catalysed conditions at room temperature in yields of 38–67%. After Zemplén de-O-acetylation, the formyl groups were directly and chemoselectively coupled to the lysine residues of bovine serum albumin by reductive amination using sodium cyanoborohydride. The conjugation reactions were followed as a function of time and under a series of different molar ratios of the reactants to provide glycoconjugates of varying degree of antigenicities. Thus, carbohydrate protein conjugates were made readily available using essentially two key reactions.Presented in part at the 15th International Carbohydrate Symposium, Yokohama, Japan, August 12–17, 1990.     

The effect of different krill meals fed to laboratory rats on their blood indices.

1. In 102 laboratory rats fed with (a) the krill standardized meal, (b) the krill meal with low chitin content, (c) the casein diet with D,L-methionine, (d) the casein diet with D,L-methionine supplemented with the krill carapace meal, (e) the casein diet with D,L-methionine supplemented with ash from the krill standardized meal and (f) the control diet--"Murigran" standard pelleted feed; the different blood indices were investigated. 2. The mean values of following indices: the number of erythrocytes and leucocytes, the percentage of leucocytes, the corpuscular haemoglobin concentration and red blood cell diameter were similar in all experimental and control groups. 3. The mean values of haematocrit and haemoglobin levels, the mean corpuscular thickness and volume were lower in rats fed with the casein diet with D,L-methionine supplemented with the krill carapace meal than in other groups. 4. All kinds of investigated indices were similar in rats fed with krill meal with low chitin contents, whose parents received the standardized krill meal and no sex differences have been shown here.     

CTP-dependent lipid kinases of yeast

Membrane fractions from yeast Saccharomyces cerevisiae catalyzed a transfer of gamma-phosphate from [gamma-32P]CTP into membranous lipids. Phosphorylated compounds were identified as phosphatidic acid and dolichyl phosphate (DolP). The membrane fraction also catalyzed phosphorylation of the exogenous dolichol. The activity of the phosphorylating enzymes could be modified by the yeast growing conditions; i.e., the enzyme from yeast grown aerobically favored the synthesis of phosphatidate over dolichyl phosphate in the ratio of 3:1, whereas the membrane fraction from anaerobically grown yeast synthesized PA and DolP in the ratio of 0.5:1. The activity of the phosphorylating enzymes could also be modified by divalent cations and the concentration of detergents. Phosphorylation of lipids does not occur in the presence of [gamma-32P]ATP and is not influenced by the presence of UTP or GTP. This result points to the specific role of CTP as a gamma-phosphate donor for the synthesis of phosphatidate and dolichyl phosphates in the yeast system.     

Non-protein-mediated transfer of phosphatidic acid between microsomal and mitochondrial membranes

The transfer of phosphatidic acid between rat liver microsomes loaded with [32P]-phosphatidic acid and rat liver mitochondria was studied in the absence of added lipid transfer proteins. It was found that during 1 h at 37 degrees C in the medium containing 100 mM KCl, 20-30% of phosphatidic acid but only 2.5% of phosphatidylcholine were transferred. This spontaneous transfer of phosphatidic acid remained the same after pretreatment of microsomes and mitochondria with 125 mM KCl or microsomes alone with 1 mM Tris, pH 8.6, procedures reported to remove adsorbed lipid transfer proteins. This transfer was insensitive to thiol-blocking reagents. The initial rate of this non-protein-mediated transfer of phosphatidic acid was virtually independent of the concentration of the acceptor membranes (mitochondria), thus indicating that it occurs by diffusion of the phospholipid through the aqueous phase rather than by membrane collision. About 80% of phosphatidic acid synthesized in the outer mitochondrial membrane was recovered in the inner membrane after a 1-h incubation, pointing to a high rate of the intermembrane transfer of this phospholipid within intact mitochondrion.     

Isolation and properties of glycerol-3-phosphate oxidoreductase from human placenta

Glycerol-3-phosphate oxidoreductase (sn-glycerol 3-phosphate: NAD+ 2-oxidoreductase, EC 1.1.1.8) from human placenta has been purified by chromatography on 2,4,6-trinitrobenzenehexamethylenediamine-Sepharose, DEAE-Sephadex A-50 and 5'-AMP-Sepharose 4B approximately 15800-fold with an overall yield of about 19%. The final purified material displayed a specific activity of about 88 mumol NADH min-1 mg protein-1 and a single protein band on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. The native molecular mass, determined by Ultrogel AcA 44 filtration, was 62000 +/- 2000 whereas the subunit molecular mass, established on polyacrylamide gel in the presence of 0.1% sodium dodecyl sulphate, was 38000 +/- 500. The isoelectric point of the enzyme protein, determined by column isoelectric focusing, was found to be 5.29 +/- 0.09. The pH optimum of the placental enzyme was in the range 7.4-8.1 for dihydroxyacetone phosphate reduction and 8.7-9.2 for sn-glycerol 3-phosphate oxidation. The apparent Michaelis constants (Km) for dihydroxyacetone phosphate, NADH, sn-glycerol 3-phosphate and NAD+ were 26 microM, 5 microM, 143 microM and 36 microM respectively. The activity ratio of cytoplasmic glycerol-3-phosphate oxidoreductase to mitochondrial glycerol-3-phosphate dehydrogenase in human placental tissue was 1:2. The consumption of oxygen by human placental mitochondria incubated with the purified glycerol-3-phosphate oxidoreductase, NADH and dihydroxyacetone phosphate was similar to that observed in the presence of sn-glycerol 3-phosphate. The possible physiological role of glycerol-3-phosphate oxidoreductase in placental metabolism is discussed.