Resting Cell Suspensions Research Articles

Aerobic biodegradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) as a nitrogen source by a Rhodococcus sp., strain DN22

An aerobic, Gram-positive bacterium was isolated from explosives-contaminated soil by enrichment culture, using the nitramine explosive, hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), as the sole added N source. The organism, identified by 16S rDNA analysis as a Rhodococcus sp., strain DN22, grew exponentially with a mean generation time of 6.5 h at 25°C in minimal medium containing RDX as the sole N source. The growth medium was depleted of RDX 24 h after inoculation but growth did not cease until 20 h later. It was concluded that strain DN22 was using NO − 2, released from RDX, as a growth substrate because, following inoculation of strain DN22 into RDX-containing minimal medium, the concentration of NO − 2 increased during the first 10 h of incubation and declined to undetectable amounts 20 h later. The ratio of the growth yields of strain DN22 grown on RDX or NO − 3 as N sources indicated that three of the six N atoms of RDX were being incorporated into biomass. Increased concentrations of NH + 4 in the growth medium reduced the extent of RDX degradation. Nitrite was produced from RDX by resting cell suspensions grown on RDX, NO − 3, NO − 2 or glutamine as N sources, but not by cells grown on NH + 4–N or in peptone-yeast extract medium. Resting cells grown on RDX showed the highest degradative activity, compared to cells grown on alternative N sources, indicating that the RDX degradation system is inducible. When soil was inoculated with RDX-grown cells of strain DN22rr, a rifampicin-resistant derivative of strain DN22, there was significant biodegradation of RDX. The addition of oat chaff to soil stimulated the growth of strain DN22rr and enhanced biodegradation of RDX, resulting in 90% degradation of the explosive in 21 d.

Conversion of chlorinated propanes by Methylosinus trichosporium OB3b expressing soluble methane monooxygenase

Chlorinated propanes are important pollutants that may show persistent behaviour in the environment. The biotransformation of 1-chloropropane, 1,2-dichloropropane, 1,3-dichloropropane and 1,2,3-trichloropropane was studied using resting cell suspensions of Methylosinus trichosporium OB3b expressing soluble methane monooxygenase. The transformation followed first-order kinetics. The rate constants were in the order 1-chloropropane > 1,3-dichloropropane > 1,2-dichloropropane > 1,2,3-trichloropropane, and varied from 0.07 to 1.03 ml min−1 mg of cells−1 for 1,2,3-trichloropropane and 1-chloropropane respectively. Turnover-dependent inactivation occurred for all of the chloropropanes tested. The inactivation constants were lower for 1-chloropropane and 1,2-dichloropropane than for 1,2,3-trichloropropane and 1,3-dichloropropane. Not all the chloride was released during cometabolic transformation of the chlorinated propanes and production of monochlorinated- and dichlorinated propanols was found by gas chromatography. The reaction pathway of 1,2,3-trichloropropane conversion was studied by mass spectrometric analysis of products formed in 2H2O, which indicated that 1,2,3-trichloropropane was initially oxidized to 2,3-dichloropropionaldehyde and 1,3-dichloroacetone, depending on whether oxygen insertion occurred on the C-3 or C-2 carbon of 1,2,3,-trichloropropane, followed by reduction to the corresponding propanols. The results show that chloropropanes are susceptible to cometabolic oxidation by methanotrophs, but that the transformation kinetics is worse than with cometabolic conversion of trichloroethylene.

Acid tolerance and acid-neutralizing activity of Porphyromonas gingivalis, Prevotella intermedia and Fusobacterium nucleatum.

The tolerance to acid and the acid-neutralizing activity of three important periodontopathic bacteria, Porphyromonas gingivalis, Prevotella intermedia and Fusobacterium nucleatum were studied. P. gingivalis strains grew only at neutral pH and did not utilize glucose, whereas strains of P. intermedia and F. nucleatum could grow under acidic conditions and increased their growth by utilizing glucose. P. gingivalis tended to raise the culture pH during growth. P. intermedia and F. nucleatum raised the culture pH during growth in the absence of glucose, while in the presence of glucose they decreased the pH. Resting cell suspensions of all the bacteria raised the pH in the presence of tryptone and casamino acids. Acid-neutralizing activity was confirmed by measuring base production at a fixed pH with a pH-stat. During neutralization, the cells produced cytotoxic substrates, ammonia and organic acids (butyric, isobutyric and isovaleric acids by P. gingivalis; isovaleric and succinic acids by P. intermedia; propionic and butyric acids by F. nucleatum). These findings suggest that deamination of amino acids into ammonia and organic acids occurs simultaneously with base production, resulting in acid neutralization. These results could partially explain the survival of P. intermedia and F. nucleatum in both supragingival and subgingival plaque and the apparent restriction of P. gingivalis to subgingival plaque. The former bacteria may aid in creation of an environment fostering colonization of subgingival plaque by P. gingivalis.

Biodegradation of 2,4-Dichlorophenol through a Distal meta-Fission Pathway

Alcaligenes eutrophus JMP222, a derivative of A. eutrophus JMP134 which has lost plasmid pJP4 (encoding the tfd genes for the ortho fission pathway), was induced for the meta fission pathway when grown on o-cresol. Resting cell suspensions, grown on o-cresol, oxidized 2,4-dichlorophenol (2,4-DCP), a degradation product of 2,4-dichlorophenoxyacetic acid, to 3,5-dichlorocatechol. Further degradation of 3,5-dichlorocatechol was observed by the production of a yellow ring fission product with liberation of chloride. Oxidation of 2,4-DCP (305 (mu)M) in 47 hs resulted in 69% dehalogenation through this pathway. The ring fission product was characterized as 2-hydroxy-3,5-dichloro-6-oxo-hexa-2,4-dienoic acid by gas chromatography-mass spectrometry and gas chromatography-Fourier transform infrared spectroscopy. These data indicate that 2,4-DCP is degraded through a distal meta ring fission pathway, in contrast to either a suicidal proximal fission or the standard ortho fission pathway.

Substrate selectivity pattern of Comamonas testosteroni strain B-356 towards dichlorobiphenyls

In this work, we have investigated the substrate selectivity pattern of strain B-356 resting cell suspensions and cell lysates towards selected chlorobiphenyl congeners. The strain showed a preference for the double meta-substituted congener 3,3′-dichlorobiphenyl over the double ortho-substituted congener 2,2′-dichlorobiphenyl and the double para-substituted congener 4,4′-dichlorobiphenyl. The results are discussed with reference to the substrate selectivity pattern reported for Pseudomonas sp. strain LB400.

Isolation of Alcaligenes sp. strain L6 at low oxygen concentrations and degradation of 3-chlorobenzoate via a pathway not involving (chloro)catechols

Isolations of 3-chlorobenzoate (3CBA)-degrading aerobic bacteria under reduced O2 partial pressures yielded organisms which metabolized 3CBA via the gentisate or the protocatechuate pathway rather than via the catechol route. The 3CBA metabolism of one of these isolates, L6, which was identified as an Alcaligenes species, was studied in more detail. Resting-cell suspensions of L6 pregrown on 3CBA oxidized all known aromatic intermediates of both the gentisate and the protocatechuate pathways. Neither growth on nor respiration of catechol could be detected. Chloride production from 3CBA by L6 was strictly oxygen dependent. Cell-free extracts of 3CBA-grown L6 cells exhibited no catechol dioxygenase activity but possessed protocatechuate 3,4-dioxygenase, gentisate dioxygenase, and maleylpyruvate isomerase activities instead. In continuous culture with 3CBA as the sole growth substrate, strain L6 demonstrated an increased oxygen affinity with decreasing steady-state oxygen concentrations.

Biodehalogenation: Oxidative and Hydrolytic Pathways in the Transformations of Acetonitrile, Chloroacetonitrile, Chloroacetic Acid, and Chloroacetamide by Methylosinus trichosporium OB-3b

Resting cell suspensions of the soil methylotroph Methylosinus trichosporium OB-3b rapidly metabolize acetonitrile (ACN) and chloroacetonitrile (CCN) to bicarbonate. Formate, a natural substrate for this organism, is a major metabolite in both cases. The first step in these processes, established with 13C-labeled substrates and NMR analysis, is an oxygen insertion into the C−H bond to yield the corresponding cyanohydrins. With acetonitrile, the loss of HCN from the initially formed cyanohydrin (HOCH2CN) produces formaldehyde. The latter is rapidly converted to formate and CO2. With chloroacetonitrile, the first formed cyanohydrin [HOCHCl(CN)] may lose HCN to produce formyl chloride or lose HCl to yield formyl nitrile. The generation of CO, in addition to formate, implicates formyl chloride and/or formyl nitrile as intermediates. With neither ACN nor CCN is the nitrile moiety directly attacked or hydrolyzed. In particular, the nitrile hydrolysis products from CCN, chloroacetamide (CAM), and chloroacetic acid (CA) are not detected. In addition, hydrolysis of the C−Cl moiety of CCN does not occur. In contrast, incubation of CAM and CA with this organism does result in the corresponding hydroxy amide and acid directly. The slower dehalogenation of the amide and acid entail a direct microbial hydroxylation of the C−Cl bond. Based upon this study and related work with methylene and ethylene halides, it may be concluded that in the series ClCH2X (X = Cl, Br, CN) oxygen insertion is the initial step in dehalogenation by this organism. However, in the series ClCH2Y (Y = CH2Cl, CO2H, CONH2), the dehalogenation is the result of a direct hydrolysis of the C−Cl bond. This organism also transforms cyanide ion, but incubations with K13CN resulted in no detectable products.

Nitrile hydrolysis by Rhodococcus erythropolis BL1, an acetonitrile-tolerant strain isolated from a marine sediment.

A number of physiologically different nitrile-hydrolysing bacteria were isolated from coastal marine sediments in Denmark by enrichment culture. One strain, BL1, identified as Rhodococcus erythropolis, grew on acetonitrile as sole carbon and nitrogen source in a defined medium. Growth occurred between 0 and 8% NaCl with an optimum around 2%, thus reflecting the marine origin of the isolate. Intact cells of R. erythropolis BL1 could hydrolyse a large variety of saturated and unsaturated aliphatic nitriles to their corresponding acids. Benzonitrile and benzylcyanide were not hydrolysed, whereas some aromatic compounds containing a -CN group attached to a C3 or C4 aliphatic side chain were accepted as substrates. The substrate spectrum of R. erythropolis BL1 was thus markedly different from those of other Grampositive nitrile-hydrolysing bacteria isolated from non-marine environments. Nitrile hydrolysis during growth and in resting cell suspensions usually occurred without intermediate accumulation of amide outside the cells. Detailed studies, however, showed that nitrile hydrolysis by strain BL1 was due to a nitrile hydratase/amidase enzyme system. Nitrile hydratase activity was found to be inducible whereas amidase activity was constitutive. The amidase activity of cells could, however, be enhanced manyfold by growth in media containing acetamide or acetonitrile. In most cases amides were hydrolysed at a much higher rate than the corresponding nitriles, which explained why amides were rarely detected in the surrounding medium during nitrile hydrolysis. R. erythropolis BL1 exhibited the highest tolerance towards acetonitrile ever reported for a nitrile-hydrolysing bacterium, as demonstrated by its ability to grow exponentially in the presence of 900 mM acetonitrile.

Degradation of Polynuclear Aromatic Hydrocarbons bySphingomonas paucimobilis

The ability of Sphingomonas paucimobilis, strain EPA 505 (a soil bacterium capable of utilizing fluoranthene as the sole source of carbon and energy for growth) to metabolize a variety of high molecular weight polynuclear aromatic hydrocarbons (PAHs) was investigated. After 16 h of incubation with 10 ppm of a PAH, a resting cell suspension (1 mg wet cells/mL) of S. paucimobilis grown on fluoranthene degraded 80.0, 72.9, 31.5, 33.3, 12.5, and 7.8% of pyrene, benz[a]anthracene (B[a]A), chrysene, benzo[a]pyrene (B[a]P), benzo[b]fluoranthene (B[b]F), and dibenz[a,h]anthracene (DB[a,h]A), respectively. No degradation of dibenzo[a,l]pyrene was detected under these conditions. 1-Nitropyrene was degraded at a lower rate than pyrene. The extent of degradation of the PAHs increased with an increase in cell density. Studies with [7-14C]B[a]P and [5,6,11,12-14C]chrysene showed that, after 48 h of incubation, the cells degraded nearly 28 and 42% of [14C]B[a]P and [14C]chrysene to 14CO2, respectively, suggesting that the bacterium is able to metabolize B[a]P and chrysene via ring cleavage. No evolution of 14CO2 was detected from cultures incubated with [4,5,9,10-14C]pyrene or [1,2,3,4,4a,4b-U-14C]dibenz[a,l]pyrene. Analysis of the ethyl acetate extracts of the culture medium by reverse-phase HPLC showed that B[a]P, B[b]F, and B[a]A were each degraded to the major, high polar metabolite(s). The degradation of B[a]P with S. paucimobilis significantly reduced the mutagenic activity associated with the hydrocarbon. The addition of solubilizing agents such as Tween 80 or cyclodextrin to the incubation medium did not enhance the biodegradation of B[a]P by EPA 505. The addition of 5 ppm of B[a]A, chrysene, fluoranthene, or DB[a,h]A to the incubation medium containing 5 ppm of B[a]P had no effect on the degradation of B[a]P by EPA 505. However, the biodegradation of B[a]P was reduced by nearly 30% in the presence of 5 ppm of B[b]F. The results demonstrate that S. paucimobilis EPA 505 has the ability to degrade several four- and five-ring PAHs ranging in molecular size, shape, and chemical structure.

Thermophilic biodegradation of BTEX by two Thermus Species

Two thermophilic bacteria, Thermus aquaticus ATCC 25104 and Thermus species ATCC 27978, were investigated for their abilities to degrade BTEX (benzene, toluene, ethylbenzene, and xylenes). Thermus aquaticus and the Thermus sp. were grown in a nominal medium at 70 degrees C and 60 degrees C, respectively, and resting cell suspensions were used to study BTEX biodegradation at the same corresponding temperatures. The degradation of BTEX by these cell suspensions was measured in sealed serum bottles against controls that also displayed significant abiotic removals of BTEX under such high-temperature conditions. For T. aquaticus at a suspension density of only 1.3 x 10(7) cells/mL and an aqueous total BTEX concentration of 2.04 mg/L (0.022 mM), benzene, toluene, ethylbenzene, m-xylene, and an unresolved mixture of o-and p-xylenes were biodegraded by 10, 12, 18, 20, and 20%, respectively, after 45 days of incubation at 70 degrees C. For the Thermus sp. at a suspension density of 1.1 x 10(7) cells/mL and an aqueous total BTEX concentration of 6.98 mg/L (0.079 mM), benzene, toluene, ethylbenzene, m-xylene, and the unresolved mixture of o-and p-xylenes were biodegraded by 40, 35, 32, 33, and 33%, respectively, after 45 days of incubation at 60 degrees C. Raising the BTEX concentrations lowered the extents of biodegradation. The biodegradations of both benzene and toluene were enhanced when T. aquaticus and the Thermus sp. were pregrown on catechol and o-cresol, respectively, as carbon sources. Use of [U-(14)C]benzene and [ring-(14)C]toluene verified that a small fraction of these two compounds was metabolized within 7 days to water-soluble products and CO(2) by these nongrowing cell suspensions. Our investigation also revealed that the nominal medium can be simplified by eliminating the yeast extract and using a higher tryptone concentration (0.2%) without affecting the growth and BTEX degrading activities of these cells. (c) 1995 John Wiley & Sons, Inc.

Monitoring trichloroethylene mineralization by Pseudomonas cepacia G4 PR1

To analyze the extent of mineralization of trichloroethylene (TCE) without disturbing an actively growing biofilm, a minimal growth medium was formulated that reduces the concentration of chloride ions to the extent that the chloride ions generated from TCE mineralization may be detected with a chloride-ion-specific electrode. By substituting chloride salts with phosphates and nitrates, a chloride-free minimal medium was produced that yields a specific growth rate for Pseudomonas cepacia G4 PR1 which was 93% of that in chloride-ion-containing minimal medium. Furthermore, TCE degradation by resting cell suspensions was similar in both media (85% of 75 μM TCE degraded in 6 h), and complete mineralization of TCE was slightly superior in the chloride-free minimal medium (77% compared to 60% of 75 μM TCE mineralized in 6 h). In addition, indole-containing, minimal-medium agar plates were developed to indicate the presence of the TCE-degrading enzyme toluene ortho-monooxygenase (fire-engine-red colonies) as well as to distinguish this enzyme from other TCE-degrading enzymes (toluene dioxygenase and toluene para-monooxygenase).

Biotransformation of tributyltin chloride in the presence of resting-cell suspensions of pure strains of microorganisms

We describe the degradation of tributyltin chloride by several strains of fungi, yeasts and bacteria under resting-cell conditions in phosphate buffer, with low initial concentrations of substrate. Yields of biotic conversion of tributyltin ranging from 10 to 77% were observed after a five-day incubation at 28 °C. In most cases, dibutyltin and monobutyltin compounds and a fraction of volatile products were formed. Volatile tin compounds essentially included derivatives of monobutyltin and traces of other organomethyltins (mono-, di-, and tri-methyltins; di- and tri-butyltins), probably as the corresponding organostannanes. Compared with conditions in which the substrate was incubated with growing microorganisms, higher yields of degradation and substantial amounts of volatile products were obtained.

Microbial degradation op dibenzothiophene bynocardioides

Abstract The atmospheric sulfur pollution is one of the major problem world is facing today. Combustion of low quality fossil fuels emitt sulfur dioxide gas. Precombustion desulfurization of fuels is a solution for reduction of atmospheric sulfur pollution. An actinomycetes, Nocardioides sp has been isolated from soil by enrichment culture technigue for the biodegradation of heterocyclic sulfur compound, dibenzothiophene (DBT). Resting cell suspension can degrade DBT with the release of sulfate. The investigation on DBT degradation by Nocardioidea has shown that the organism has got the potential for deaulfurization of petroleum crude.

Growth, induction, and substrate specificity of dehydroabietic acid-degrading bacteria isolated from a kraft mill effluent enrichment.

We investigated resin acid degradation in five bacteria isolated from a bleach kraft mill effluent enrichment. All of the bacteria grew on dehydroabietic acid (DHA), a resin acid routinely detected in pulping effluents, or glycerol as the sole carbon source. None of the strains grew on acetate or methanol. Glycerol-grown, high-density, resting-cell suspensions were found to undergo a lag for 2 to 4 h before DHA degradation commenced, suggesting that this activity was inducible. This was further investigated by spiking similar cultures with tetracycline, a protein synthesis inhibitor, at various times during the DHA disappearance curve. Cultures to which the antibiotic was added prior to the lag did not degrade DHA. Those that were spiked with the antibiotic after the lag phase (4 h) degraded DHA at the same rate as did controls with no added tetracycline. Therefore, de novo protein synthesis was required for DHA biodegradation, confirming that this activity is inducible. The five strains were also evaluated for their ability to degrade other resin acids. All strains behaved in a similar fashion. Unchlorinated abietane-type resin acids (abietic acid, DHA, and 7-oxo-DHA) were completely degraded within 7 days, whereas pimarane resin acids (sandaracopimaric acid, isopimaric acid, and pimaric acid) were poorly degraded (25% or less). Chlorination of DHA affected biodegradation, with both 12,14-dichloro-DHA and 14-chloro-DHA showing resistance to degradation. However, 50 to 60% of the 12-chloro-DHA was consumed within the same period.

Dehalogenation of haloalkanes by Rhodococcus erythropolis Y2. The presence of an oxygenase-type dehalogenase activity complements that of an halidohydrolase activity.

Rhodococcus erythropolis Y2 produced two types of dehalogenase: a hydrolytic enzyme, that is an halidohydrolase, which was induced by C3 to C6 1-haloalkane substrates, and at least one oxygenase-type dehalogenase induced by C7 to C16 1-haloalkanes and n-alkanes. The oxygenase-type activity dehalogenated C4 to C18 1-chloroalkanes with an optimum activity towards 1-chlorotetradecane. The halidohydrolase catalysed the dehalogenation of a wide range of 1- and alpha,omega-disubstituted haloalkanes and alpha,omega-substituted haloalcohols. In resting cell suspensions of hexadecane-grown R. erythropolis Y2 the oxygenase-type dehalogenase had a specific activity of 12.9 mU (mg protein)-1 towards 1-chlorotetradecane (3.67 mU mg-1 towards 1-chlorobutane) whereas the halidohydrolase in 1-chlorobutane-grown batch cultures had a specific activity of 44 mU (mg protein)-1 towards 1-chlorobutane. The significance of the two dehalogenase systems in a single bacterial strain is discussed in terms of their contribution to the overall catabolic potential of the organism.

Metabolic pathway to propionate of Pectinatus frisingensis, a strictly anaerobic beer-spoilage bacterium

Pectinatus frisingensis, a recently described species of anaerobic mesophilic beer-spoilage bacteria, grows by fermenting various organic compounds, and produces mainly propionate, acetate, and succinate. Although acrylate and succinate were both dismutated by dense resting-cell suspensions, propionate production proceeded through the succinate pathway: [3-13C]pyruvate consumption led to equal 13C-labeling of propionate on methyl and methylene groups. Growth on glucose or glycerol led to a similar propionate to acetate ratio, suggesting dihydroxyacetone phosphate as being a common metabolic intermediate. Diacetyl, 1,3-propanediol, and 2,3-butanediol were not growth substrates or fermentation products, but they were all dismutated by dense resting-cells suspensions to acetate and propionate. Acetoin was a minor fermentation product. The consumption of [2-13C] or [3-13C]pyruvate by dense resting-cell suspensions demonstrated the involvement of two equivalent pyruvate molecules during acetoin production. Key enzymes involved in this metabolism were measured in anoxic cell-free extracts. A tentative metabolic pathway to the main fermentation products was proposed from the above results.

Biodegradation of sulphosuccinate: direct desulphonation of a secondary sulphonate.

The bacterial biodegradation of a secondary sulphonate, sulphosuccinate, has been shown to occur by direct desulphonation. A bacterium, designated Pseudomonas sp. BS1, was isolated from activated sewage sludge, for its capacity to grow on sulphosuccinate as the sole source of carbon and energy. Cultures grown on sulphosuccinate were able to convert this substrate to sulphite which was subsequently oxidized rapidly to sulphate. The sequence of desulphonation and carbon-chain catabolism of sulphosuccinate was determined from measurements of the kinetics of sulphite and 14CO2 release from specifically radiolabelled sulpho[1,4-14C]succinate and sulpho[2,3-14C]succinate, which were synthesized from the corresponding maleic anhydrides. When each radiolabelled compound was incubated separately with washed-cell suspensions of Pseudomonas BS1, sulphite was released before 14CO2, as shown by chemical assay and radiorespirometry, respectively. Differences in the kinetics and extent of 14CO2 release from the 1,4- and 2,3-labelled substrates were consistent with entry of the intact C4 chain into the citric acid cycle. When carrier oxaloacetate was added to incubation mixtures containing resting-cell suspensions and radiolabelled sulphosuccinate, a radiolabelled metabolite with the same HPLC retention time as oxaloacetate accumulated. No radioactive metabolites accumulated when carrier oxaloacetate was replaced with succinate, fumarate or malate. Collectively, the data indicated co-production of sulphite and oxaloacetate from sulphosuccinate, which is interpreted in terms of an oxidative desulphonation mechanism.

Production of 10-Ketostearic Acid from Oleic Acid by Flavobacterium sp. Strain DS5 (NRRL B-14859)

A microbial isolate, Flavobacterium sp. strain DS5, produces 10-ketostearic acid (10-KSA) from oleic acid in 85% yield. This is the first report on this type of reaction catalyzed by a Flavobacterium strain. The product was purified to give white, plate-like crystals melting at 79.2 degrees C. The optimum time, pH, and temperature for the production of 10-KSA are 36 h, 7.5, and 30 degrees C, respectively. A small amount of 10-hydroxystearic acid (10-HSA) (about 10% of the amount of the main product, 10-KSA) is also produced during the bioconversion. 10-KSA is not further metabolized by strain DS5 and accumulates in the medium. In contrast to growing cells, a resting-cell suspension of strain DS5 produces 10-HSA and 10-KSA in a ratio of 1:3. The crude cell extract obtained from ultrasonic disruption of the cells converts oleic acid mainly to 10-HSA (10-HSA/10-KSA ratio, 97:3). This result strongly suggested that oleic acid is converted to 10-KSA via 10-HSA. Enzymes catalyzing the hydration and secondary alcohol dehydrogenation are cell associated. Product 10-HSA from strain DS5 is 66% enantiomeric excess in the 10(R) form. The oleic acid conversion enzyme(s) reacts with unsaturated fatty acids in the order oleic acid > palmitoleic acid > linoleic acid > linolenic acid > gamma-linolenic acid > myristoleic acid.

Batch cultivation of Xanthobacter Py2 on 1-pentene

The propene isolated strain Xanthobacter Py2 was able to grow on 1-pentene. The biomass yield for growth under 1-pentene limiting conditions was 0.48 (Ceq/Ceq). Upon storage at both +4°C and -20°C no loss of enzymatic epoxide degrading activity in resting cell suspensions was observed after a month. However, activity decay was pronounced during the stationary phase of growth as well as under reaction conditions.

Glucose fermentation to acetate and alanine in resting cell suspensions ofPyrococcus furiosus: Proposal of a novel glycolytic pathway based on13C labelling data and enzyme activities

Abstract Suspensions of maltose-grown cells of the hyperthermophilic archaeon Pyrococcus furiosus , when incubated at 90°C with 35 mM [1- 13 C]glucose or [3- 13 C]glucose, consumed glucose at a rate of about 10 nmol min −1 (mg protein) −1 . Acetate (10 mM), alanine (3 mM), CO 2 and H 2 were the fermentation products. The 13 C-labelling pattern in alamine and acetate were analyzed. With [1- 13 C]glucose the methyl group of both alanine and acetate was labelled; with [3- 13 C]glucose only the carboxyl group of alanine was labelled whereas acetate was unlabelled. Extracts of maltose-grown cells contained glucose isomerase (12.8 U mg −1 , 100°C), ketohexokinase (0.23 U mg −1 , 100°C), and fructose 1-phosphate aldolase (0.06 U mg −1, 100°C). Enzymes catalyzing the formation of fructose 1,6-bisphosphate from fructose 1-phosphate or fructose 6-phosphate could not be detected. As publihed previously by our group and other authors P. furiosus also contains enzymes of glyceraldehyde conversion to 2-phosphoglycerate according to a non-phosphorylated Entner-Doudoroff pathway, of dihydroxyacetone phosphate conversion to 2-phosphoglycerate according to the Embden-Meyerhof pathway, and of 2-phosphoglycerate conversion - via pyruvate - to acetate and alanine. Based on the enzyme activities in P. furiosus , the following pathway for glucose degradation to alanine and acetate in cell suspensions is proposed which can explain the [ 13 C]glucose labelling data: glucose→ fructose → fructose 1- phosphate → dihydroxyacetone phosphate + glyceraldehyde and further conversion of both trioses to alanine and acetate via pyruvate.