Inhibition of alternative respiration system of Scheffersomyces stipitis and effect on glucose or xylose fermentation.

Scheffersomyces stipitis is a Crabtree-negative pentose fermenting yeast, which shows a complex respiratory system involving a cytochrome and an alternative salicylhydroxamic acid (SHAM)-sensitive respiration mechanism that is poorly understood. This work aimed to investigate the role of the antimycin A (AA) sensitive respiration and SHAM-sensitive respiration in the metabolism of xylose and glucose by S. stipitis, upon different agitation conditions. Inhibition of the SHAM-sensitive respiration caused a significant (P < 0.05) decrease in glycolytic flux and oxygen consumption when using glucose and xylose under agitation conditions, but without agitation, only a mild reduction was observed. The combination of SHAM and AA abolished respiration, depleting the glycolytic flux using both carbon sources tested, leading to increased ethanol production of 21.05 g/L at 250 rpm for 0.5 M glucose, and 8.3 g/L ethanol using xylose. In contrast, inhibition of only the AA-sensitive respiration, caused increased ethanol production to 30 g/L using 0.5 M glucose at 250 rpm, and 11.3 g/L from 0.5 M xylose without agitation. Results showed that ethanol production can be induced by respiration inhibition, but the active role of SHAM-sensitive respiration should be considered to investigate better conditions to increase and optimize yields.

Effect of cytochrome bc1 complex inhibition during fermentation and growth of Scheffersomyces stipitis using glucose, xylose or arabinose as carbon sources.

Scheffersomyces stipitis shows a high capacity to ferment xylose, with a strong oxygen dependence to allow NAD+ regeneration. However, without oxygen regeneration of NADH occurs by other metabolic pathways like alcoholic fermentation. There are few reports about inhibitors of mitochondrial respiration and their effects on growth and fermentation. This work aimed to explore the effect of cytochrome bc1 complex inhibition by antimycin A (AA), on growth and fermentation of S. stipitis using glucose, xylose and arabinose as carbon sources, at three agitation levels (0, 125 and 250 rpm). It was possible to discriminate between respiratory and fermentative metabolism in these different conditions using xylose or arabinose. Despite the inhibition of mitochondrial respiration, the glycolytic flux was active because S. stipitis metabolized glucose or xylose to produce ATP; on 0.5 M glucose the cells yielded 17-33 g L-1 ethanol. However, more complex results were obtained on xylose, which depended upon agitation conditions where ethanol production without agitation increased up to 11 g L-1. Inhibition of respiratory chain in S. stipitis could therefore be a good strategy to improve ethanol yields.

Optimization of nisin production by Lactococcus lactis UQ2 using supplemented whey as alternative culture medium.

Lactococcus lactis UQ2 is a nisin A-producing native strain. In the present study, the production of nisin by L. lactis UQ2 in a bioreactor using supplemented sweet whey (SW) was optimized by a statistical design of experiments and response surface methodology (RSM). In a 1st approach, a fractional factorial design (FFD) of the order 2(5-1) with 3 central points was used. The effect on nisin production of air flow, SW, soybean peptone (SP), MgSO(4)/MnSO(4) mixture, and Tween 80 was evaluated. From FFD, the most significant factors affecting nisin production were SP (P = 0.011), and SW (P = 0.037). To find optimum conditions, a central composite design (CCD) with 2 central points was used. Three factors were considered, SW (7 to 10 g/L), SP (7 to10 g/L), and small amounts of added nisin as self-inducer (NI 34.4 to 74.4 IU/L). Nisin production was expressed as international units (IU). From RSM, an optimum nisin activity of 180 IU/mL was predicted at 74.4 IU/L NI, 13.8 g/L SP, and 14.9 or 5.11 g/L SW, while confirmatory experiments showed a maximum activity of 178 +/- 5.2 IU/mL, verifying the validity of the model. The 2nd-order model showed a coefficient of determination (R(2)) of 0.828. Optimized conditions were used for constant pH fermentations, where a maximum activity of 575 +/- 17 IU/mL was achieved at pH 6.5 after 12 h. The adsorption-desorption technique was used to partially purify nisin, followed by drying. The resulting powder showed an activity of 102150 IU/g. Practical Application: Nisin production was optimized using supplemented whey as alternative culture medium, using a native L. lactis UQ2 strain. Soybean peptone, SW, and subinhibitory amounts of nisin were successfully employed to optimize nisin production by L. lactis UQ2. Dried semipurified nisin showed an activity of 102150 IU/g.