Abstract:

Pentose and hexose hydrolyzed from crop straw with acid and enzyme were determined by HPLC.Based on the discussion of corresponding product and enzyme, the metabolic pathway was analyzed.For interdicting lactic acid metabolic pathway and acetic acid metabolic pathway, the parent strain was mutated by soft X-ray of synchronous radiation.In order to improve the balance of H reducing power, the flux ratio of HMP to EMP was modulated by adding citrate.Then the xylulokinase activity was regulated for the co-fermentation of pentose and hexose.The results showed that about 57 g glucose and 11 g xylose were hydrolyzed from 200 g crop straw.Metabolic flux analysis of the parent strain indicated that the flux of succinic acid,lactic acid and acetic acid were 2.547 mmol•g^-1•h^-1, 0.726 mmol•g^-1•h^-1 and 0.611 mmol•g^-1•h^-1, respectively.Lactic acid flux of Ldh mutant strain HF-1 decreased markedly to 0.0296 mmol•g^-1•h^-1.Acetic acid flux of anti-fluoroacetic mutant strain HF-2 decreased distinctly to 0.100 mmol•g^-1•h^-1.However, the decrease of acetic acid resulted in not strengthening but weakening the metabolism of succinic acid.By adding citrate the flux ratio of HMP to EMP increased from 1.389∶0.389 to 1.684∶0.330 and thus the H reducing power was better balanced, then the succinic acid flux increased from 3.005 mmol•g^-1•h^-1 to 3.468 mmol•g^-1•h^-1 accordingly.Substituting MgCO₃ for CaCO₃ as neutralizer increased xylulokinase specific activity to 560 U•mg^-1 from 76 U•mg^-1, and thus the consumed xylose flux increased from 0.0444 mmol•g^-1•h^-1 to 0.453 mmol•g^-1•h^-1 accordingly, then pentose and hexose were co-fermented and the succinic acid yield reached 68.7 g•L^-1.