Abstract During the cultivation of microorganisms environmental conditions are known to exert an strong influence on metabolism, morphology and chemical composition of microbial cells. These phenotypic variations are also concerned with the chemical properties and function of the outer membrane. In the case of gram-negative bacteria lipopolysaccharides represent the main antigens of the bacterial cell-surface. At the same time lipopolysaccharides are the endotoxins of these bacteria, their toxicity residing in the lipid part of the molecule, the lipid A. It is therefore interesting whether the conditions of cultivation would also influence the amount of lipopolysaccharide synthesized, or even the biosynthesis of individual antigenic determinants, and finally the chemical composition of the lipid A. Differences in environmental conditions occur, for example, in mass-cultures at the different phases of growth. The present study deals with the cultivation of 5 Salmonella R-mutants of different chemotypes (Ra to Re) in complex medium under constant conditions of temperature, pH and aeration. From the growth curves thus obtained it may be seen that the time required by the various chemotypes to reach the exponential phase increases in the order of Ra to Re ( Download : Download full-size image ). Correspondingly the deep R-mutants require a longer time of cultivation before reaching the stationary phase of growth. Despite the above differences in growth-behaviour the overall yield of bacteria obtained with the 5 mutants was comparable (4.2–4.6 g/l). In the course of cultivation all mutants showed differences in the LPS content of the bacterial mass. A temporary stagnation in LPS biosynthesis, frequently found previously during cultivation of Salmonella S-forms (S. Schlecht, Zbl. Bakt. Hyg., I. Abt. Orig. A 232 [1975] 61–72), was observed here only in the case of an Ra mutant. Highest LPS yields were obtained in the stationary phase (207–334 mg/l culture fluid). Here, the content of extractable LPS was 5.0–6.2% of the bacterial masses. The true LPS content, estimated by measuring the amount of 3-hydroxymyristic acid present in the cells was 8.5–10% indicating that about 40% of the LPS was either not extractable or was lost during the isolation procedure. In the R mutants the molecular distribution of LPS was found to be 2–3 times higher on the bacterial surface than in the S forms, the density increasing from the higher to the lower R-chemotypes (Ra to Re). A small portion of the LPS was found extracellularly. Thus at the end of cultivation the culture medium contained about 5% of the total LPS, as determined in the case of the deeper R-mutants ( Download : Download full-size image ). Chemical analysis on LPS from cultures at different stages of growth, revealed no significant differences in the fatty acid composition of the lipid A or in the sugar composition of the R oligosaccharide ( Download : Download full-size image ). Only in the case of one R-strain at the end of cultivation a low increase in the phosphate content was found. Further, the substitution of lipid A with core oligosaccharide was the same at all stages of growth as indicated by the constant ratio of heptose: β-hydroximyristic acid. Hemagglutination-inhibition studies showed, however, that in the case of a leaky mutant (RcRb2) the Rb2 specificity was expressed more strongly in the exponential phase than at later stages of growth. This suggests, that the biosynthesis of the R-oligosaccharide may, in principle, be influenced by the changing culturing conditions. With all other mutants tested no differences were seen in the serological activity of the LPS at different stages of growth.