0 7 0 0 4 62 2 ± 2 6 62 4 47 1 ± 2 8 47 9 29 1 ± 2 5 27 5 11 0 +1

0 7.0 0.4 62.2 ± 2.6 62.4 47.1 ± 2.8 47.9 29.1 ± 2.5 27.5 11 0 +1 3.7 5.0 7.0 0.4 125.3 ± 0.8 123.7 54.1 ± 0.2 53.5 44.9 ± 2.9 51.9 12 +1 +1 7.4 5.0 7.0 0.4 140.2 ± 8.0 140.7 78.8 ± 0.5 78.5 61.1 ± 1.9 55.6 *The cultivations were performed in triplicate, with the exception of cultivation at condition (0,0) performed in quadruplicate; SD = standard deviation. Results and discussion Individual effect of diamines and precursors on cephamycin C production For this study, two concentrations for each

diamine were defined based on literature data obtained for other beta-lactam antibiotic producing microorganisms [32, 33, 35, 42]. Cephamycin C biosynthesis precursors lysine and alpha-aminoadipic acid were tested at several concentrations in order to define ranges of adequate values

for the experimental designs. Cephamycin C production and cell growth obtained at see more 48 h and 72 h cultivations in basal medium without additives and supplemented with putrescine, 1,3-diaminopropane, and cadaverine are shown in Figure 1. Leitão et al. [32] found that all three diamines promoted cephamycin C production by N. lactamdurans, albeit at different levels. The largest increase was observed in culture media containing 2.5 or 5.0 g l-1 of 1,3-diaminopropane. In this study, this diamine also produced a similar effect: a 100% increase in volumetric production was observed after the addition of 5.0 g l-1 of the compound as compared to selleck screening library that of the culture medium with no additive. Also, the addition of Guanylate cyclase 2C 1,3-diaminopropane alone promoted higher specific production than that obtained at the control condition (Figure 1C). Similarly, Martín et al. [42] observed that adding 5.0 mM (0.37 g l-1) or 10 mM (0.74 g l-1) of 1,3-diaminopropane enhanced Penicillium chrysogenum beta-lactam antibiotic production by approximately

100%. It is likely that one of the effects of 1,3-diaminopropane is to maintain high mRNA transcript levels during the production phase [43]. Figure 1 Effect of biomass and cephamycin C with different diamines. Biomass (A), cephamycin C concentration (CephC) (B), and specific production (C) obtained in shake-flasks cultivations of basal medium with no antibiotic-production enhancing compound (control condition) and with putrescine (Put), 1,3-diaminopropane (1,3D), and cadaverine (Cad), at two concentration values (in parentheses); the cultures were performed in triplicate. In the present work, putrescine did not affect antibiotic production by S. clavuligerus (Figure 1B), as Martín et al. [42, 43] observed with P. chrysogenum. However, Leitão et al. [32] observed positive effects on cephamycin C production with N. lactamdurans when 0.20 g l-1 of putrescine was added. With regard to cadaverine, volumetric production almost doubled by adding 7.0 g l-1 of this diamine (Figure 1B). However, specific production was not higher than that obtained in media without additives (Figure 1C). For cultivations with N. lactamdurans, a threefold increase was obtained using 5.

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