Figure 4.

Nucleotide concentrations and their relative relations to each other in Penicillium ochrochloron CBS 123.824 during growth in chemostat cultures with different nutrient limitations at μ = 0.1 h⁻¹, 30°C and pH 7. Limiting nutrients were glucose (20 mM), ammonium (2 mM), and phosphate (0.1 mM). (A) Nucleotide levels in dependence of the nutrient limitation. (B) Sum parameters of all targeted nucleotides: adenine nucleotides (AXP), NADH, and NADH, NADP and NADPH, and GDP and GTP (GXP). (C) Relative ratios of nucleotides to each other expressed as Energy Charge (EC), Catabolic Reduction Charge (CRC), Anabolic Reduction Charge (ARC), NADH/NAD-ratio and NADPH/NADP-ratio. (D) Correlation between organic acid excretion and intracellular nucleotide concentration. Samples for intracellular nucleotide analysis were subjected to an immediate cold methanol stop and further processed using the newly developed FiltRes device (Vrabl et al., 2016). Nucleotides were extracted with hot ethanol and quantified by gradient reversed phase HPLC and CE. For method optimization see supplementary information, section metabolism. Nucleotide data for each nutrient limitation were derived from three subsequent days of steady state cultures and represent averages ± standard deviation of n = 14 (glucose limitation), n = 13 (ammonium limitation) and n = 12 (phosphate limitation) samples.