Optimations On Parameters Control Of Pyruvate Fermentation And Studies On Rapid Detection Of Pyruvic Acid

Pyruvate is an important intermediate metabolite of glucose metabolism,which is the precursor of a variety of useful chemical compounds.it is widely used in pharmaceutical,agricultural,chemicals of industrial and scientific research, their commercial demand was continuously increasing in recent years. At present,the pyruvate production methods have begun to shift from original chemical synthesis to microbial fermentation.In this paper,Torulopsis glabrata was used as the fermentation strain.,Pyruvate was obtained through liquid fermentation . The main contents are as follows: Established the method of rapid detection of pyruvate. through experiments. The large amounts of by-productα-ketoglutarate which contained in the broth, will cause a great interference of detection accuracy, when routine chemical determination methods are used to detect pyruvate in the production. Optimized the traditional spectrophotometric determination method of pyruvic acid with iron(Ⅲ) nitrate,together with the dual-wavelength spectrophotometric method that can eliminate the interference of matrix background, color agents andα-ketoglutarate. Detection wavelength 510 nm, reference detection wavelength 540 nm.Recovery rate is 99. 02%, RSD was 0.48%, the method is simple, low cost, accurate and reliable. Optimized the fermentation parameters. With the optimization of culture conditions of the seed, 20 h was determined to be the best age. seed medium was: glucose 40 g / L, peptone 4 g / L, magnesium sulfate 0.3 g / L, potassium dihydrogen phosphate 1g/L. With the optimization of fermentation conditions, determine the optimal medium volume is 42mL/250 mL, the optimum addition level of calcium carbonate 32 g / L, the best temperature program is 0-8 h, 33℃culture,9-36 hours, 30℃culture,37-56 h, 28℃culture. Using Box-Behenken center response method of response surface analyze fermentation medium, determine that glucose, ammonium sulfate and peptone medium are main factors. The best combination of medium is ammonium sulfate 6.5 g / L, glucose 108.6 g / L, peptone, 1.92 g / L. Production was 63 g / L, compared with 17.53% before optimization.In this experiment,we use dual enzymatic hydrolysis starch, starch sugar was used to fermentate pyruvate. Single factor analysis of starch hydrolysis conditions were optimized to determine the best amylase hydrolysis conditions: pH is 6.3-6.5, temperature is 93℃, hydrolysis time is 20 min, enzyme dosage is 15 U / g, 20% starch concentration. Box-Behenken center response was used to determine the optimal conditions for glucoamylase hydrolysis: temperature is 62.8℃, hydrolysis time is 27 hours, enzyme dosage is 170 U / g, the final DE value is 91.59%.