The marine bacterium Beneckea natriegens was grown in batch culture on a glucose/ NH,+/salts medium; growth terminated due to either carbon or nitrogen depletion from the medium. Nitrogen-limited cultures converted part of the excess glucose into glycogen whereas the carbon-limited cultures formed little glycogen. Glycogen-rich cultures survived longer than glycogen-poor cultures during starvation. Little protein was utilized during starvation and RNA was degraded as the primary endogenous source of energy. Glycogen was consumed only when the RNA content had decreased to about a third of the growth value.The adenine nucleotide content of nitrogen-limited cultures increased at the start of the stationary phase but the energy charge remained at the growth value of 0.9 to 0.95. The maximum size of the adenine nucleotide pool depended on the concentration of glucose remaining in the medium at the start of the stationary phase but a limiting value of about 60 pmol ATP (g protein)-l was attained, compared with 12 to 14 pmol ATP (g protein)-l in exponentially growing cultures. During extended starvation of both glycogen-rich and glycogen-poor cultures, there was a large decrease in adenine nucleotide content, but the energy charge remained above 0-6 even when viability was very low. I N T R O D U C T I O NThere are many factors that can cause loss of viability of bacteria (Dawes, 1976). During long-term starvation, endogenous metabolites such as non-essential RNA, protein and reserve polymers (glycogen, poly-P-hydroxybutyrate, triacylglycerols and polyphosphate) act as energy sources permitting essential maintenance processes to occur, thereby preserving viability (Dawes & Senior, 1973). Under these circumstances loss of viability may be related to the inability to regenerate ATP due to depletion of endogenous metabolites and reserve polymers. This would be seen as a decrease in the adenine nucleotide content or as a decrease in the ATP : ADP and ATP : AMP ratios, which may be measured as a decrease in energy charge (Atkinson, 1968(Atkinson, , 1978Chapman & Atkinson, 1977;Knowles, 1977Knowles, ,1979.There have been few studies on the adenylate pools of starving bacteria. Chapman et uZ. (1 97 1) showed that, following nitrogen depletion from a glucose-containing growth medium, there was a gradual decrease in the intracellular energy charge of Escherichia cofi strain B from a growth value of about 0-8 to between 0.6 and 0-5 after 60 to 80 h, without loss of viability. The intracellular adenine nucleotide content also decreased during this period. After 60 to 80 h, a rapid decrease in energy charge occurred, coincident with a loss of viability. Montague & Dawes (1974) showed a similar relationship of decrease in energy charge and loss of viability during starvation of Peptococcus pre'votii, which does not form reserve polymers but utilizes RNA as the sole endogenous source of energy. 7 Present address: The College of the Resurrection, Mirfield, West Yorkshire WF14 OBN.

393The effect of dissolved oxygen concentration on glucose metabolism, poly-P-hydroxybutyrate (PHB) synthesis and respiration of nitrogen-fixing Azotobacter beijerinckii was investigated over a range of values spanning oxygen limitation and sufficiency (nitrogen limitation) in continuous culture. The activities of the Entner-Doudoroff enzymes decreased with increasing oxygen supply while glucose-6-phosphate dehydrogenase was unaffected; /3-ketothiolase and acetoacetyl-CoA reductase decreased more markedly, reflecting the fall in PHB content. The metabolic quotients for 0, and CO, increased rapidly with increasing oxygen supply, in keeping with the respiratory protection of nitrogenase. The specific activities of all these enzymes, and also of fructose-1,6-bisphosphate aldolase, increased with increasing dilution rate under either oxygen or nitrogen limitation, except that both /3-ketothiolase and acetoacetyl-CoA reductase activities fell at the highest dilution rate (0.23 h-l) under oxygen limitation. It was concluded that the role of the Entner-Doudoroff sequence as the major route of glucose metabolism in A. beijerinckii was unaffected by oxygen concentration and growth rate.

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