The Study Of Glycerol Fermentation Produced 1,3 - Propanediol

A strain was isolated according to glycerol induced from Woodland of Hunan Academy of Forestry, and it was named 2e. The strain could produce 1,3-propanediol from glycerol disproportionation. Combination of morphological, cultural characteristics, physiological and biochemical characteristics and phylogenetic analysis of 16S rRNA genes were identified on the strain. The results showed that this strain was Klebsiella pneumoniae.The optimum conditions were obtained though the process of fermentation. The conditions was as flows: the glycerol concentration was 40g/L, the nitrogen source was 3g/L (NH₄)₂HPO₄, pH 7.5, temperature was 30℃, liquid volume in flask was 40%, Rotation speed 100rpm, fermentation time 24h. Orthogonal test were made based on the important factors according to single factor experiments. Optimization results was that the glycerol concentration was 40g/L, (NH₄)₂HPO₄ was 4g/L, K₂HPO₄/KH₂PO₄ was 4g/L temperature was 30℃, Although the orthogonal test and optimize the results of single factor experiments are different, but the main factors and its amount were the same.The further optimized were made by response surface method fermentation, The optimization were carried out in the 5L fermenter. The results was that (NH₄)₂HPO₄ 2.76g/L, K₂HPO₄/KH₂PO₄ 3.98 g/L, pH 7.58, and examined the above factors on 1,3-PDO yield. The results show that pH >(NH₄)₂HPO₄> K₂HPO₄/KH₂PO₄, and the impact of after the interaction of two factors was more significant. The quadratic polynomial regression model of 1,3-PDO yield was abtained by the design of quadratic regression. It was that Y=25.86+2.5X₁-0.71X₂+2.54X₃+1.35X₁X₂-0.61X₁X₃ -1.83X₂X₃-2.32X₁²-8.7X₂²-7.01X₃². The model predicted the experimental results were verified and the result was that the final yield of 1,3-PDO was 27.597g/L, compared with 19.02% before optimization.Through the study of glycerol fermentation of 2e bacteria, The entire fermentation process was divided into four stages: I(0h-8h),II(8h-12h),III(12h-24h),IV(after24h). In stage II controled glycerol concentration as 10g/L, and III controlled it to 20g/L, fermentation 26h. The final yield of 1,3-PDO was up to 69.8g/L, that increased 28.5% than constant concentration of substrate fermentation.The optimal separation conditions was abtained by means the study of 1,3-PDO broth separation conditions. The result was that acetaldehyde/PDO was 3:1, temperature was 30℃, strong acid resin and sulfuric acid catalyst in two steps, reacted 15min. the final conversion of 1,3-PDO was 97.1%, and the yield of 2-methyl-1,3-dioxane (2MD) was 92.5%.