Klebsiella aerogenes NCTC 418 was grown in chemostat cultures (D = 0.17 hr-1; pH 6.8;35 degrees C) that were, successively, carbon-, sulphate-, ammonia-, and phosphate-limited, and which contained as the sole carbon-substrate first glucose, then glycerol, mannitol and lactate. Quantitative analyses of carbon-substrate used and products formed allowed carbon balances to be constructed and direct comparisons to be made of the efficiency of substrate utilzation. With all sixteen cultures, carbon recoveries of better than 90% were obtained. Optimum utilization of the carbon substrate was invariably found with the carbon-limited cultures, the sole products being organisms and carbon dioxide. But the extent to which excess substrate was over-utilized varied markedly with both the nature of the growth-limitation and the identity of the carbon-substrate. In general, sulphate-, ammonia-, and phosphate-limited cultures utilized glycerol more efficiently than mannitol, mannitol better than lactate, and glucose least efficiently. Glucose-containing cultures also synthesized some extracellular polysaccharide. When the carbon source was in excess, a range of acidic compounds generally were excreted. Sulphate-limited cultures, growing on glucose, excreted much pyruvate and acetate, whereas similarly-limited cultures growing on glycerol, mannitol or lactate produced only acetate. Ammonia-limited cultures invariably excreted 2-oxoglutarate and acetate, whereas phosphate-limited cultures produced gluconic acid, 2-ketogluconic acid and acetate, when growing on glucose, but only acetate when growing on mannitol or lactate. From the rates of substrate and oxygen consumption, and the rates of cell synthesis, yield values for both substrate and oxygen were calculated. These showed different trends, but were similar in being highest under carbon-limitation and substantially lower under all other limitations. The physiological significance of these findings, and the probable nature of the regulatory mechanisms underlying "overflow metabolism" are discussed.

The regulation of carbohydrate metabolism in Klebsiella aerogenes NCTC 418 organisms, growing in chemostat culture.

Halobacteroides halobius gen. nov., sp. nov., a Moderately Halophic Anaerobic Bacterium from the Bottom Sediments of the Dead Sea

When grown at the expense of 3,4,5-trimethoxybenzoic acid, a strain of Pseudomonas putida oxidized this compound and also 3,5-dimethoxy-4-hydroxybenzoic (syringic) and 3,4-dihydroxy-5-methoxybenzoic (3-O-methylgallic) acids; but other hydroxy- or methoxy-benzoic acids were oxidized slowly or not at all. Radioactivity appeared exclusively in carbon dioxide when cells were incubated with [4-methoxyl-14C]trimethoxybenzoic acid, but was found mainly in methanol when[methoxyl-14C]3-O-methylgallic acid was metabolized. The identity of methanol was proved by analyzing the product from [methoxyl-13C]3-O-methylgallic acid by nuclear magnetic resonance spectroscopy and by isolating the labeled 3,5-dinitrobenzoic acid methyl ester, which was examined by mass spectrometry. These results, together with measurements of oxygen consumed in demethylations catalyzed by cell extracts, showed that two methoxyl groups of 3,4,5-trimethoxybenzoate and one of syringate were oxidized to give carbon dioxide and 3-O-methylgallate. This was then metabolized to pyruvate; the other product was presumed to be the 4-methyl ester of oxalacetic acid, for which cell extracts contained an inducible, specific esterase. P. putida did not metabolize the methanol released from this compound by hydrolysis. Support for the proposed reaction sequence was obtained by isolating mutants which, although able to convert 3,4,5-trimethoxybenzoic acid into 3-O-methylgallic acid, were unable to use either compound for growth.

Production of methanol from aromatic acids by Pseudomonas putida.

Isolation of an aspartate-fermenting, free-living campylobacter species

The products in commerce today that are produced by Bacillus fermentations include enzymes antibiotics , and insecticides. This chapter focuses on the basic principles necessary for an understanding of growth and product production in Bacillus spp. The authors show the important link between some very basic observations and how these can be (and need to be) applied in medium development processes, choice of equipment, and elimination of bottlenecks in production. Using protease production as a model, examples show the potential value of using controlled fermentations in identifying some of the limitations of product formation in Bacillus spp. Bacillus spp. as a class are known to secrete a large number of extracellular enzymes, including several proteases , amylase, cellulase, and other degradative enzymes. Costs of exoprotein production can be estimated from material balances, molecular formulae, and growth parameters. Results of these calculations can be a key element in choosing a fermentor type and mode of operation for desired product formation. The functions included under strain development are identification and isolation of an organism exhibiting the trait (or traits) of interest; analysis and optimization of cultural conditions that allow expression of the trait; and mutagenesis, selection, and screening for isolates that hyperproduce the trait. In addition, overexpression of these proteins may improve the secretion efficiency of heterologous proteins, as has been demonstrated for signal peptidase.

Fermentation of Bacillus

Publisher Summary The products of a fermentation process may be few, as in the case of the classical yeast fermentation, or they may be tremendously diverse, which is typical of much bacterial fermentation. Therefore, the analysis of fermentation presents tasks ranging in magnitude from the relatively simple to the highly complex, involving the separation, identification, and quantitation of a broad spectrum of compounds. The development of column, paper, and thin-layer chromatography has had a major impact not only on the separation and identification of fermentation products but also on their quantitative determination. This chapter discusses the analysis of fermentation products, the general principles of fermentation balances, and the apparatus and techniques for carrying out fermentation studies. The chapter also discusses the procedures involved in the separation, identification and quantitative analysis of the various compounds likely to be encountered in fermentation processes. The identification and determination of volatile fatty acids by gas-liquid chromatography is also described.

Chapter III Analysis of Fermentation Products

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Chapter III Analysis of Fermentation Products