**3. Conclusion**

Experimental parameters associated with cold plasma discharge at atmospheric air pressure for enhancing ethanol yield of *S. cerevisiae* has been successfully optimized in this research. The maximum theoretical ethanol yield of 0.49 g/g was predicted by the response model under three optimized parameters (1 min of exposure time, 26 V of power voltage and 9 mL of test sample volume), which was closely consistent with the experimental yield of 0.48 g/g. The model may be used as a reference for modulating the experimental parameters related with dielectric barrier discharge at air atmospheric pressure and a novel approach for improving ethanol yield in bio-manufacturing industry.

Furthermore, the potential mechanism that air cold plasma alters the cofactor metabolism of *S. cerevisiae* was explored by analyzing the changes in plasma membrane potential, cytoplasmic calcium concentration and the two cofactors of ATP and NADH. The sample of 1-min treatment presented a notable increment in plasma membrane potential, whereas the sample of 2-min treatment presented a distinct reduction in plasma membrane potential. In addition, the calcium concentrations for the samples treated by plasma for 1–5 min were remarkably improved prior to the beginning of the fermentation compared with that for the untreated sample. An increase of 7.0% in calcium concentration led to the remarkable reductions of 40% in ATP and 60% in NADH in the sample of 1-min treatment. At 9-h culture, the ATP concentration of treated sample for 1 min increased by 72%, whereas NADH concentration decreased by 88% relative to those of the control. Briefly, the mechanism that plasma promoted alterations in cofactor level in *S. cerevisiae* showed to be by improving the cell membrane potential, which then caused increases in cytosolic free Ca2+ concentrations within the cells, eventually enhancing microbial productivity. This may a potential and broad application in intensifying the biotransformation capability of microorganisms in the future.
