| The energization of waste meets the national energy conservation,emissions reduction,and low carbon development strategy. The application of the abundant, low price, and renewable straw waste as hydrogen fermentation substrates is a new direction for hydrogen generation. Since agicultural waste could be effectively disposaled accompanied with the related environmental problems be effectively solved, so this method has became one of the main trends for future energy development. However, due to the complexity and recalcitrance of the straw waste, how to efficiently obtain the glucose and xylose riched in straw, and solve the common problem that microorganisems lack the ability to metabolize xylose is essential for the realization of high valued straw waste utilization. This research determines using Thermoanaerobacterium thermosaccharolyticum W16, which could ferment both glucose and xylose isolated by our lab, aimed to establish a cost-efficetive bioconverstion of straw waste to hydrogen system. The hydrogen producing partern of W16 by utilizing glucose and xylose was investigated, and then sequential batch fermentation process was proposed to resolve the problem of low xylose utilization rate in mixed substrates; the metabolic regulation mechanism of glucose and xylose were discussed based on the differences in expression levels of genes and proteins by application of transcriptomics and proteomics. Based on this, the biological pretreatment and enzymatic saccharification of cornstalk were applied, and the fermentative hydrogen producing efficiency of W16 was performed when cornstalk hydrolyate was used as substrate, results obtained in this research provides technical and theoretical proof for the high value use of cornstalk.When different mass ratios of glucose and xylose were used as substrate, strain W16 could utilize both of them for hydrogen production efficiently. However, compared to glucose utilization, xylose utilization rate was decreased with the increase of glucose content in the mixture. Node interpolation analysis showed that when glucose content in the mixed sugar is higher than 58.2%, its inhibitory effect on xylose utilization was increased, but when glucose levels fell below 21.7%, its utilization will be subject to a certain degree of feedback inhibition. To solve the low xylose utilization problem, continuous and sequential batch fermentation process with hydraulic retention time and dilution rate adjustment strategy was developed. Results showed that, by sequential batch fermentation process, under the HRT of 12 h, hydrogen production rate could reach the maximum of 8.9 mmol/L/h, the corresponding glucose and xylose utilization rate were achieved 92.2% and 82.2%, respectively.In addition, gene regulation and expression analysis of strain W16 under different carbon souces was determined by the high-throughput sequencing platform based on Illumina 2000. Results showed that strain W16 could encode more xylose transport protein genes would be the main reason that why xylose could be utilized accompanied with glucose. RNA-seq data also revealed the relative xylose ismerase genes active expression although large amount of glucose was existed in the culture, which might be attributed to its high affinity to glucose. We presume it would be the main explanation that why the expression of xylose transports protein genes was high expresed but still a certain degree of inhibition existed during the high proportion glucose and xylose mixture. At the same time, when xylose existed in the sugar mixture, express of genes related to glucose transport(phosphotransferase system) were down regulated, we speculated this would be the reason that glucose and xylose exist mutual influence and inhibition phenomenon with each other during the cofermentation process.On the basis of glucose and xylose coutilization characteristic of W16, pretreatment and enzymatic saccharification process was established. By comparing the chemical component and morphological structure of cornstalk under different pretreatments, biological pretreatment by Phanerochaete chrysosporium CGMCC 5.776 could remove 35.3% of lignin, whearas with only 9.5% of holocellulose loss. To reduce the cost of cellulase during cornstalk saccharification, extracellular crude cellulase produced from Trichoderma viride CGMCC 3.2876 was selected to hydrolyze pretreated cornstalk. Under the optimized conditions of 49.7°C, p H 5.0, cellulase concentration of 35.7 IU/g, and substrate concentration of 38.5g/L, the maximum saccharification efficiency reached 62.5%, in other words, by going through the pretreatment and enzymatic saccharification steps, a total of 27.2 g of reducing sugars were obtained from 100 g of cornstalk. At last, hydrogen production was carried out by batch, continuous and sequential batch fermentation, the maximum hydrogen production rate reached 9.1 mmol/L/h under sequential batch fermentation process. In order to further improve the hydrogen production efficiency, a simultaneous saccharifcation and fermentataion process was put forward. Experimental results showed that by adjusting the reaction temperature, substrate concentration, and enzyme dosage, the operataion time could be effectively shornten to 72 h, at the same time, hydrogen yield could reach 89.3 m L/g-corntalk. |
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