Metabolic activity of Corynebacterium glutamicum grown on l-lactic acid under stress

Abstract

Respiration measurement in shake flasks is introduced as a new method to characterize the metabolic activity of microorganisms during and after stress exposure. The major advantage of the new method is the possibility to determine the metabolic activity independent of manual sampling without the necessity to change the culture vessel or the cultivation medium. This excludes stress factors, which may be induced by transferring the microorganisms to plates or respirometers. The negative influence, which interruptions of the shaker during sampling times may have on the growth of microorganisms was demonstrated. The applicability of the method was verified by characterizing the behavior of Corynebacterium glutamicum grown on the carbon source L: -lactic acid under stress factors such as carbon starvation, anaerobic conditions, lactic acid, osmolarity, and pH. The following conditions had no effect on the metabolic activity of C. glutamicum: a carbon starvation of up to 19 h, anaerobic conditions, lactic acid concentrations up to 10 g/l, 3-(N-morpholino) propanesulfonic acid buffer concentrations up to 42 g/l, or pH from 6.4 to 7.4. Lactic-acid concentrations from 10 to 30 g/l lead to a decrease of the growth rate and the biomass substrate yield without effecting the oxygen substrate conversion. Without adaptation, the organism did not grow at pH< or =5 or > or =9.

Scale‐up from shake flasks to fermenters in batch and continuous mode with Corynebacterium glutamicum on lactic acid based on oxygen transfer and pH

“…All batch media did not contain buffer to use the same medium for shake flask and fermenter cultures and fermenter media usually do not contain buffer. This is possible, because C. glutamicum can grow between pH 5 and pH 9 (Seletzky et al, 2006). Further, the large pH shift of the batch cultures aloud a better demonstration of the possibilities of the pH model.…”

“…4a ba ) or from the slope of the oxygen transfer rate (OTR) (Fig. 4b ba ) (Seletzky et al, 2006;Stöckmann et al, 2003). Independent of the scale, three growth phases can be identified from the OTR curves of the batch cultures (Fig.…”

“…The influence of the pH on our model organism Corynebacterium glutamicum has been described by Seletzky et al (2006), Shah et al (2002), Bröer and Krämer (1991). Thus, the scale-up from shake flasks to fermenters can only succeed, if the pH shift is similar.…”

“…At a pH between 5 and 10, where C. glutamicum may be viable (Seletzky et al, 2006), salts, strong acids, and strong bases are regarded as completely dissociated . Acids and bases are considered as strong, when having a dissociation constant smaller than 10 À3 or larger than 10 À12 .…”

“…For biomass measurements from shake flask cultures 1.5 mL culture broth was used. A more detailed description of the biomass measurements is given by Seletzky et al (2006). All biomass concentrations were determined twice.…”