丁醇萃取发酵耦联生产改良型生物柴油过程的性能优化

传统丙酮丁醇发酵的产物浓度太低,蒸馏回收发酵产品大量耗能.为研究探讨直接利用发酵产物的可能性,在15%高初始玉米醪浓度条件下、以地沟生物柴油为萃取剂,探讨了生物柴油添加量,萃余液回用率、和添加微量电子供体对丁醇发酵耦联生产改良型生物柴油过程各主要性能指标的影响.通过环境条件优化,地沟生物柴油的质量显著提高,16烷值由51.4提高至54.4;添加微量中性红后"丁醇实质性得率"可以达到18%,耗能的发酵产品回收过程有望省去;萃余液回用率超过50%,有望逐步向国家所大力倡导的"节能减排"的工业生产模式靠近.     

使用硅藻土处理发酵废液、提高丁醇发酵废液回用率

以生物柴油为萃取剂耦联丁醇发酵,能以最为节能的方式生产"改良型"生物柴油、提高发酵性能,但却也产生了大量发酵废液、萃余液回用率超过50%次轮发酵就无法正常进行.探讨了使用硅藻土处理发酵废液、提高废液回用率的可能性和最适条件,在使用40目硅藻土、用量3%(w/v)的条件下,废液回用率可以提高至75%;对提高废液回用率的原因进行了初探,利用少量硅藻土可以吸附发酵残液中的丁醇,减少因美拉德反应所生成的不同分子量的增殖抑制型色素物质.原位添加微量硅藻土有利于丁醇发酵的进行,在使用油醇的萃取发酵条件下,丁醇总生产强度提高了8%.     

添加表面活性剂改善丁醇萃取发酵性能

研究了各种表面活性剂对丁醇萃取发酵的影响。丁醇发酵中有大量H2、CO2气体生成,生成的气泡携带发酵溶剂产物（丁醇、丙酮）进入萃取液相,促进了水相中发酵毒性产物向萃取液相的移动。研究发现,表面活性剂可以降低气-液膜的表面张力,促使大气泡破碎,从而使发酵产气以较小气泡的形式穿过萃取液相。添加表面活性剂可以强化发酵溶剂产物从水相到萃取相的移除速度,缩短发酵产物在油水两相中达到平衡的时间。有利于提高发酵生产强度。以地沟生物柴油为萃取剂,吐温-80的添加量为质量分数0．140％时,与对照相比（无表面活性剂的萃取发酵）,相同发酵时间内萃取相中丁醇体积分数提高了21．2％．总溶剂生产强唐也提高了16．5％．     

生物柴油耦联丙酮丁醇发酵的初步研究

以4种生物柴油(原料为地沟油、菜籽油、棕榈油和废肯德基油)作为萃取剂,开展了丙酮丁醇静态萃取发酵。通过分析发酵过程中的产气量及发酵40 h后油水两相中的溶剂浓度,发现生物柴油对丙丁梭菌有毒性。另外,静置条件下丁醇在不同油水两相中的液液平衡系数大致相同。在发酵24 h时加入棕榈生物柴油(油水体积比为0.4∶1),丁醇发酵强度达到最大值0.213 g.(L.h)-1、比对照(传统发酵)提高10.9%,且生物柴油中的丁醇质量浓度达到6.44 g.L-1。     

萃取耦合技术对玉米秸秆水解液发酵产丁醇的影响

本研究以玉米秸秆水解液为原料,通过萃取发酵技术生产燃料丁醇,以提高丁醇产量,降低生产成本。通过对萃取剂的筛选与条件优化,确定纤维丁醇发酵的萃取剂为油醇,添加时间为发酵0 h,添加比例为1:1 (V/V)。该条件下发酵32 g/L糖浓度的玉米秸秆水解液,丁醇和总溶剂产量分别为3.28 g/L和4.72 g/L,比对照分别提高958.1%和742.9%。以D301树脂脱毒后5%总糖浓度的玉米秸秆水解液进行丁醇萃取发酵,丁醇和总溶剂产量分别达到10.34 g/L和14.72 g/L,发酵得率为0.31 g/g,与混合糖发酵结果相当。研究结果表明萃取发酵技术能够显著提高原料的利用率和丁醇产量,为纤维丁醇工业化生产提供了技术支撑。     

萃取耦合发酵工艺生产丁醇研究

萃取耦合发酵可有效减弱产物抑制和提高底物利用效率,本文就萃取耦合发酵生产丁醇工艺中的萃取剂的选择、萃取剂加入量、底物浓度等发酵条件进行了研究。结果表明:最佳萃取剂为大豆油生物柴油,油水比为3∶5,发酵过程无需搅拌,静置发酵为宜,在发酵之初加入萃取剂。分别以玉米和木薯为发酵底物,确定其最适底物浓度为100 g/L,以玉米为原料萃取耦合发酵中丁醇和总溶剂产量分别为18.17 g/L和29.31 g/L。以木薯为原料萃取耦合发酵生产丁醇及总溶剂产量比传统发酵分别提高了48.69%和51.80%。     

木薯发酵产丁醇的研究

对丙酮丁醇梭菌发酵木薯产溶剂进行研究,分别考察了N源、木薯含量、酶处理条件和培养基pH对发酵产丁醇的影响。结果表明：最佳的产丁醇发酵培养基为木薯粉120g／L,乙酸铵6g／L;木薯粉先用高温淀粉酶按酶量20U／g、90℃水解60min,再糊化30min;发酵初始pH为6．0,发酵96h。在此条件下,5L发酵罐中丁醇产量达到13．5g／L,总溶剂达到22．8g／L。     

大蒜超临界萃余物提取SOD的工艺研究

大蒜超临界萃取大蒜油后有大量的萃余物,本文对大蒜超临界萃余物提取SOD工艺进行了初步研究,并对所提萃余物中的SOD酶活性与从新鲜原料大蒜提取SOD进行了比较,实验结果表明：超临界萃取工艺对大蒜原料中SOD酶活力损失很少,从大蒜萃余物中提取的SOD酶活力比直接从新鲜大蒜中提取的SOD酶活力相差不到10％,且粗酶提取物已显著脱臭。说明萃余物有很大的再利用价值,对萃余物进行深度开发具有重要意义。     

酵母浸粉刺激以木薯为原料的丁醇生产的发酵相转型

在7L静态厌氧发酵罐下,使用"非粮"作物木薯替代玉米淀粉开展丁醇发酵。无论是传统发酵还是油醇萃取发酵,木薯粉丁醇发酵的性能均远不及以玉米淀粉为原料时的水平,主要体现在发酵产酸相向溶剂生产相的转型严重迟延或无法转型、发酵时间长、丁醇生产效率低。实验结果表明,当发酵相转型迟延出现后,添加2.5g/L的酵母浸粉,可以刺激丁酸/乙酸向丁醇/丙酮的转化、转型延迟时间缩短10~30h左右。在此条件下,传统和萃取发酵方式下的丁醇总产量分别达到12.95g/L和29.81g/L,丁醇生产效率与使用玉米淀粉为原料时基本持平。     

高浓度丁醇浸泡及N＋束注入诱变双重作用筛选丁醇高产菌

通过高浓度丁醇浸泡处理丙酮丁醇梭菌（Clostridiumacetobutylicum）CL-2,筛选得到一株丁醇耐受能力提高并溶剂产量增加的菌株BR30—2,丁醇产量达11．77g/L,比CL-2提高了16．65％。以BR30—2作为出发菌株,进行N＋束注入诱变,筛选得到高产菌株BH．9,丁醇产量达14．5g/L,总溶剂为23．14g／L。在BH-9发酵过程中添加0．1％丁酸钠,丁醇产量达到16．59g/L,丁醇比例提高至67．38％。     

木薯和玉米原料丁醇发酵中丁醇丙酮质量比的图论理论计算及其验证

在丁醇发酵产溶剂阶段,乙酸和丁酸的生成途径、消耗途径同时存在,各自形成一个闭环路径。本研究利用图论对丁醇发酵中丁醇丙酮质量比进行了理论计算,并对以木薯和玉米为原料的丁醇发酵进行了模拟计算,结果表明:丁酸闭环路径(L2环)的代谢强度是影响丁醇丙酮质量比的主要因素,并且L2环的代谢强度越弱,丁醇丙酮质量比越高;与玉米原料丁醇发酵相比,木薯原料发酵的m(丁醇)/m(丙酮)提高了16.7%。实验结果证实了以上计算结果:在传统发酵、油醇萃取发酵和生物柴油萃取发酵中,以木薯(适时添加酵母浸粉)为原料的发酵批次与以玉米为原料的发酵批次相比,由于其丁酸闭环路径代谢强度较弱,相应发酵方式下丁醇丙酮质量比分别提高了12.9%、61.4%和6.7%,而且两种原料相应发酵方式的丁醇总产量和生产效率基本持平。另外,高丁醇丙酮质量比的木薯发酵所得改良型生物柴油中丁醇浓度与玉米发酵的相比提高了16%,性能得到进一步提高。     

In-situ recovery of butanol during fermentation

End product inhibition can be reduced by the in situ removal of inhibitory fermentation products as they form. Extractive fermentation, in which an immiscible organic solvent is added to the fermentor in order to extract inhibitory products, was applied to the acetone-butanol fermentation. Six solvents or solvent mixtures were tested in batch extractive fermentations: kerosene, 30 wt% tetradecanol in kerosene, 50 wt% dodecanol in kerosene, oleyl alcohol, 50 wt% oleyl alcohol in a decane fraction and 50 wt% oleyl alcohol in benzyl benzoate. The best results were obtained with oleyl alcohol or a mixture of oleyl alcohol and benzyl benzoate. In normal batch fermentation of Clostridium acetobutylicum, glucose consumption is limited to about 80 kg/m³ due to the accumulation of butanol in the broth. In extractive fermentation using oleyl alcohol or a mixture of oleyl alcohol and benzyl benzoate, over 100 kg/m³ of glucose can be fermented. Removal of butanol from the broth as it formed also increased the rate of butanol production. Maximum volumetric butanol productivity was increased by as much as 60% in extractive fermentation compared to batch fermentation. Butanol productivities obtained in extractive fermentation compare favorably with other in situ product removal fermentations.     

拜氏梭菌Clostridium beijerinckii NCIMB 8052发酵玉米皮生产丁醇

玉米皮作为玉米淀粉加工的副产物,是一种可用于生产液体燃料的潜在廉价优质的生物质资源。本文以玉米皮为原料,对拜氏梭菌发酵生产丁醇进行了研究。实验结果表明,玉米皮首先在最优的预处理温度140℃下使用0.5%硫酸水溶液以固液比1∶8处理20 min,再添加200 IU/g底物糖化酶、1.0 IU/g底物木聚糖酶进行酶解,可以使原料中的淀粉和半纤维素转化为可发酵糖,此时水解液中的总糖浓度为50.46 g/L。然后使用1.0%的活性炭对水解液进行脱毒处理以去除发酵抑制物,再进行丁醇发酵,丁醇产量为9.72 g/L,总溶剂产量可达14.09 g/L,糖醇转化率为35.1%。上述研究结果证明玉米皮作为一种粮食加工废弃物用于液体燃料丁醇的生产在技术上是完全可行的。     

Enhancement of Butanol Formation by Clostridium acetobutylicum in the Presence of Decanol-Oleyl Alcohol Mixed Extractants

Extractive fermentation has been proposed to enhance the productivity of fermentations that are end product inhibited. Unfortunately, good extractants for butanol, such as decanol, are toxic to Clostridium acetobutylicum. The use of mixed extractants, namely, mixtures of toxic and nontoxic coextractants, was proposed to circumvent this toxicity. Decanol appeared to inhibit butanol formation by C. acetobutylicum when present in a mixed extractant that also contained oleyl alcohol. However, maintenance of the pH at 4.5 alleviated the inhibition of butanol production and the consumption of butyrate during solventogenesis. A mixed extractant that contained 20% decanol in oleyl alcohol enhanced butanol formation by 72% under pH-controlled conditions. The production of acetone and acetoin was also increased, even though these two products were not extractable. The enhancement of butanol formation was not limited by the toxicity of decanol. Supplementation of glucose and butyrate in the extractive fermentation yielded a 47% increase in butanol. The enhancement of butanol formation appeared to be dependent on the presence of dissolved decanol in the broth but was not observed unless an organic phase was present to extract butanol. A mechanism for the effects of decanol on product formation is proposed.     

Evaluation of high butanol/acetone ratios in ABE fermentations with cassava by graph theory and NADH regeneration analysis

Higher butanol/acetone ratio is always desirable in ABE fermentation, and this ratio is closely associated with the complicated patterns of metabolic reactions and NADH generation rate. The patterns of acetate/butyrate formation and re-assimilation in multiple closed reaction loops, as well as NADH regeneration in ABE fermentation using different substrates varies. In this study, we evaluated butanol/acetone ratio in ABE fermentations utilizing cassava and corn based media by graph theory and NADH regeneration analysis. The theoretical calculations and experimental data revealed that a lower metabolic strength in butyrate loop and enhanced NADH generation rate were responsible for the achievement of higher butanol/acetone ratio when fermenting cassava based substrate. In traditional fermentations and extractive fermentations with oleyl alcohol/bio-diesel as the extractants when using cassava based substrate, butanol/acetone ratios reached 2.24, 2.84, and 2.19 with the increasing increments of 14.9, 61.4, and 6.8% respectively, while butanol productivities stayed at comparably high levels as compared with those of the fermentations when cultivating on corn based substrate.     

铁改性沸石对Clostridium acetobutylicum XY16固定效率及发酵性能的影响

考察4种无机铁盐改性沸石对丁醇生产菌Clostridium acetobutylicum XY16的固定效率及其发酵产丁醇性能的影响。结果表明:铁改性沸石对菌体的固定效率均优于未改性沸石,而Fe3+改性效果优于Fe2+,经FeCl3改性的沸石对菌体具有良好的吸附作用,当Fe3+-zeolite用量为180 g/L时,细胞的固定效率达到87%。在此基础上,比较了沸石负载的铁离子量对丁醇发酵性能的影响,沸石负载的铁离子量为6.0 mg/g时可显著提高丁醇发酵性能,当葡萄糖质量浓度为60 g/L时进行发酵,丁醇产量为13.5 g/L,总溶剂可达20 g/L,总溶剂的生产速率为0.385g/(L.h),比游离细胞发酵分别提高了9.5%、10.3%和40%。     

固定化植物乳杆菌的制备及其发酵条件优化

采用海藻酸钠包埋植物乳杆菌并通过测定固定化细胞发酵清液的抑菌效果,优化得到的固定化最佳工艺条件为:海藻酸钠浓度为3%,CaCl2浓度为1.5%,菌悬液体积为3.5 mL(4.0×108 cfu/mL).固定化细胞重复发酵多批次效果良好.固定化细胞发酵条件优化结果表明:最适pH为7.0,最适温度为36℃,培养基中添加0....     

Enhancement of butanol tolerance and butanol yield in Clostridium acetobutylicum mutant NT642 obtained by nitrogen ion beam implantation

As a promising alternative biofuel, biobutanol can be produced through acetone/butanol/ethanol (ABE) fermentation. Currently, ABE fermentation is still a small-scale industry due to its low production and high input cost. Moreover, butanol toxicity to the Clostridium fermentation host limits the accumulation of butanol in the fermentation broth. The wild-type Clostridium acetobutylicum D64 can only produce about 13 g butanol/L and tolerates less than 2% (v/v) butanol. To improve the tolerance of C. acetobutylicum D64 for enhancing the production of butanol, nitrogen ion beam implantation was employed and finally five mutants with enhanced butanol tolerance were obtained. Among these, the most butanol tolerant mutant C. acetobutylicum NT642 can tolerate above 3% (v/v) butanol while the wide-type strain can only withstand 2% (v/v). In batch fermentation, the production of butanol and ABE yield of C. acetobutylicum NT642 was 15.4 g/L and 22.3 g/L, respectively, which were both higher than those of its parental strain and the other mutants using corn or cassava as substrate. Enhancing butanol tolerance is a great precondition for obtaining a hyper-yield producer. Nitrogen ion beam implantation could be a promising biotechnology to improve butanol tolerance and production of the host strain C. acetobutylicum.     

In-situ recovery of butanol during fermentation

End-product inhibition in the acetone-butanol fermentation was reduced by using extractive fermentation to continuously remove acetone and butanol from the fermentation broth. In situ removal of inhibitory products from Clostridium acetobutylicum resulted in increased reactor productivity; volumetric butanol productivity increased from 0.58 kg/(m³h) in batch fermentation to 1.5 kg/(m³h) in fed-batch extractive fermentation using oleyl alcohol as the extraction solvent. The use of fed-batch operation allowed glucose solutions of up to 500 kg/m³ to be fermented, resulting in a 3.5- to 5-fold decrease in waste water volume. Butanol reached a concentration of 30–35 kg/m³ in the oleyl alcohol extractant at the end of fermentation, a concentration that is 2–3 times higher than is possible in regular batch or fed-batch fermentation. Butanol productivities and glucose conversions in fed-batch extractive fermentation compare favorable with continuous fermentation and in situ product removal fermentations.List of Symbols C _g kg/m³ concentration of glucose in the feed - C _w dm³/m³ concentration of water in the feed - F(t) cm³/h flowrate of feed to the fermentor at time t - V(t) dm³ broth volume at time t - V _i dm³ initial broth volume - V _si dm³ volume of the i-th aqueous phase sample - effective fraction of water in the feed Part 1. Bioprocess Engineering 2 (1987) 1–12