| Butanol is an important platform chemical. It is also a clean and high power liquidbio-fuel. Bio-butanol production by Clostridia spp. with renewable biomass as the substrateshas become more and more attractive due to the price fluctuation/rise of petroleum and globalgreenhouse effect. Butanol fermentation is also referred as Acetone-Butanol-Ethanol (ABE)fermentation. Generally, in ABE fermentation, butanol, acetone and ethanol are produced at aratio of6:3:1. Improving butanol/acetone ratio and butanol concentration without sacrificingthe total solvent productivity has been one of the major objectives by many researchers. ABEfermentation is a process featured with extremely high energy and water consumption, whichhas limited the development of ABE industrial fermentation. In addition, bio-butanolproduction using corn based substrate has the problem of competition with foods for arableland. Focusing on those problems, in this dissertation, ABE fermentations were conducted in a7L anaerobic fermentor with the aids of directed signal flow diagram theory (DSFD), usingClostridium acetobutylicum ATCC824as the protocol strain and corn/cassava as substrates.Based on the DSFD model, co-culturing technology and novel integrated ABE extractivefermentation process were also developed to achieve the objects, and theoretical mechanismof butanol/acetone ratio was deeply interpreted. The target is to improve ABE fermentationperformance, to reduce distillation energy consumption, and to fully utilize fermentationwaste water. Major contents and results of the dissertation were summarized as follows:(1) In traditional and extractive fermentations with oleyl alcohol as the extractants whenusing cassava based substrate, butanol/acetone ratios increased14.9%and61.4%respectively,while butanol productivities stayed at comparably high levels as compared with thecorn-based ABE fermentations. The metabolic network based on directed signal flow diagrammodel was proposed to simulate the relationship between butanol/aceone ratio and metabolicstrength of the two organic acids closed-loops strength. The result indicated that weakeningmetabolic strength of butyrate closed formation/re-assimilation loop could largely increase thebutanol/acetone ratio. In addition, NADH regeneration rates and genes transcriptional levelsof key enzymes were analyzed in fermentations with cassava/corn based substrates, and theresults indicated that butanol/acetone ratio was enhanced by reduced butyrate loop strengthand enhanced NADH regeneration rate in cassava based ABE fermentations.(2) In corn-based ABE fermentations with a small amount of butyrate consecutiveaddition, butanol/acetone ratio was elevated by21.7%. The DSFD theory based non-linearmodel was proposed to calculate the formation/re-assimilation rates of butyrate/acetate. Theresults indicated that acids addition accelerated the re-assimilation of butyrate/acetate butrepress their synthesis. The calculation results of the DSFD model was in accordance with the results of experiments and relevant reports. This model could help to understand thedistribution of carbon flux when using butyrate or acetate as the supplemental co-substrate.(3) Co-culturing C. acetobutylicum and S. cerevisiae was conducted in corn-based ABEfermentation in a7L fermentor with the additions of small amount of butyrate/acetate. In thesolventogenesis phase, with2.0g-DCW·L-1S. cerevisiae broth and concentrated butyratesolution (4g·L-broth-1) addition, butanol/acetone ratio and final butanol concentrationachieved2.74and15.74g·L-1respectively, with the increment attitudes of38%and35%. Themeasurement results indicated that methionine, lysine and aromatic amino acids werenaturally released to the broth in the C.acetobutylicum/S.cerevisiae co-culturing system,which is both beneficial for butanol synthesis and C.acetobutylicum growth. The additions ofsmall amount of butyrate also reduced the metabolic strength of butyrate closed-loop. As aresult, butanol/acetone ratio and butanol concentration were largely increased.(4) An integrated water/energy saving ABE extractive fermentation processsimultaneously producing both properties-improved bio-diesel and butanol fuel was proposed.The bio-diesel was used as in-situ extractant (1:1, v/v) and bio-diesel extracting more than10g L-1butanol could be used as properties-improved bio-diesel directly, as its Cetane Numbercould be increased from51.4to54.4. The residual butanol in waste supernatant was firstlyextracted and concentrated by small amount of n-octanol (1:5, v/v) for future butanol fuelproduction. The supernatant was then treated by activated carbon to remove the fermentativeinhibitor-melanoidin, and the treated supernatant could be100%re-utilized for the nextfermentation run without performance decline. Consecutively recycling of waste supernatantfor ABE extractive fermentation could be continued for at least14runs without performancedeterioration in100mL anaerobic bottles.(5) The integrated water/energy saving corn/cassava based ABE extractive fermentationprocess was testified in a7L anaerobic fermentor. Neutral red and cassava substrate wereused to improve the overall fermentation performance. When1.0g·L-broth-1neutral red wasadded into the broth to increase reductive power,“aimed butanol” yield and butanolproductivity could further increase and eventually reach about24~26%and0.25~0.32g·L-1·h-1, respectively, and butanol in bio-diesel further increased. In addition, butanolconcentration in n-octanol could achieve a range of20~30g·L-1and ratio of butanol to totalsolvents in n-octanol exceeded90%with neutral red added, which is favorable for easingworking load of the subsequent distillation process. For fermentations with cassava basedsubstrate, butanol in bio-diesel further increased by16%when compared with corn basedfermentations and the quality of properties-improved bio-diesel was improved furthermore. |
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