肠道内产丁酸细菌及其产物丁酸生理功能的研究进展

产丁酸细菌是利用糖类发酵产生丁酸的一类细菌,代表种是丁酸梭菌。在动物及人类肠道内存在的产丁酸细菌主要是梭菌属、柔嫩梭菌属、罗斯式菌属、真菌属及丁酸弧菌属。本文一方面介绍部分肠道内产丁酸细菌的种类、特点及膳食纤维的摄入和肠道益生菌对产丁酸细菌的影响,另一方面对其主要代谢产物丁酸在体内的生理功能进行探讨,以期为产丁酸细菌的应用及产品开发提供理论依据。     

丁酸梭菌活菌制剂对肠道菌群影响的研究

通过动物实验和人体试食试验,对丁酸梭菌活菌制剂一泰平扯对肠道菌群的影响进行了研究。结果表明：受试对象肠道内的双歧杆菌的数量增加,差异有极显著性（Ｐ≤０．０１）,乳杆菌的数量增加,差异有显著性（Ｐ≤０．０５）。说明丁酸梭菌活菌制剂一泰平片具有一定的增殖肠道内有益菌和抑制有害菌的作用。     

一株产丁酸羊源拜氏梭菌的筛选及其培养条件优化

【背景】人类和动物消化道内栖息着极其复杂和多样化的微生物群落,这些微生物群落分布在肠道的不同位置并执行着特定的功能。近年来,产丁酸菌逐渐成为微生物领域的研究热点,产丁酸菌主要为产芽孢革兰氏阳性厌氧菌,对肠道健康有重要意义。【目的】从反刍动物瘤胃中筛选出产丁酸菌株并研究其生长特性,进一步优化其培养条件,从而提高产丁酸菌的丁酸产量。【方法】以绵羊瘤胃内容物为样品,运用稀释涂布法进行产丁酸菌的筛选,通过形态学观察和16S rRNA基因序列分析等方法对菌株进行鉴定。通过单因素试验与Box-Behnken design试验相结合,对培养条件进行优化,确定筛选菌株在梭菌增殖培养基(reinforced clostridium medium,RCM)中的最佳产酸培养条件。【结果】经过筛选鉴定得到的菌株为梭菌属的拜氏梭菌(clostridium beijerinckii,CB),命名为拜氏梭菌R8(CB.R8)。对拜氏梭菌R8的培养条件进行优化,得出该菌株在接种量为1.22%、温度为38.45℃、pH6.08和培养时间为64.67h的条件下丁酸产量为2.48g/L。【结论】筛选到1株拜氏梭菌R8,该菌能够在RCM培养基中生长并代谢产生丁酸,具备较高的应用价值。     

肠道菌群相关代谢物丁酸的量效和时效对人脐带间充质干细胞增殖的影响

目的探讨人肠道菌相关代谢物丁酸对人脐带间充质干细胞(human umbilical cord mesenchymal stem cells,hUC-MSCs)增殖的影响。方法用含不同浓度丁酸(0.02、0.08、0.32、1.25和5.00 mmol/L)的干细胞培养基培养hUC-MSCs,采用CCK-8增殖实验检测24、48和72 h细胞增殖情况,并与正常对照组比较。结果 hUC-MSCs的增殖与丁酸的量效和时效均有关系,作用24 h内抑制hUC-MSCs增殖,到48 h则表现出低浓度促进细胞增殖、高浓度抑制细胞增殖的作用;当时间达到72 h,则低浓度丁酸对hUC-MSCs的促增殖作用减弱,高浓度组对其增殖抑制的作用有所增强。结论丁酸对hUC-MSCs增殖的影响存在量效和时效的共同作用,适宜的丁酸浓度和作用时间对机体肠道的功能维持及平衡有重要影响。     

微生物催化生产丁酸研究进展

丁酸传统上可用于制造丁酸纤维素、合成丁酸酯、食品香料等,近年来研究发现丁酸是肠道上皮细胞的优选能量来源,能抑制组蛋白去乙酰化酶,具有抗癌作用.随着丁酸在生物相关领域功用的不断发现和实际应用,而消费者又日趋青睐生物源制品,微生物催化生产丁酸将越来越具有竞争力.产物浓度低、选择性差是当前丁酸发酵的主要限制因素.许多研究从选用廉价培养基料、优化发酵工艺、简化提取步骤、遗传改造生产菌株等方面来提高生产效率、降低成本,并取得一定进展.今后这些方面更多的突破,都将使微生物催化生产丁酸的产业化成为可能.     

酪酸梭状芽胞杆菌研究进展

酪酸梭状芽胞杆菌（Clostridium butyricum）是属于硬壁菌门（Firmicutes）梭菌纲（Clostridia）梭菌目（Clostridiales）梭菌科（Clostridiaceae）梭菌属（Clostridium）的一种革兰阳性菌（以下简称酪酸梭菌）。早在1877年就开始了对酪酸梭菌的研究。近130年以来,不同国别的不同研究者曾经先后用了10来种不同的名称记录或描述这个菌种（表1）,1980年学术界规范地将其统一命名。尽管早已有文献报道酪酸梭菌是人的正常肠道菌,但是在i999年2月由联合国粮农组织（FAO）和世界卫生组织（WHO）联合在罗马召开的第52次食品添加剂专家委员会的会议文件中还是再次指出了酪酸梭菌是人的正常肠道菌这一事实。酪酸梭菌是严格的厌氧菌,产生丁酸,所以也有人将其称为丁酸梭菌。它们不仅存在于人的肠道中,也存在于动物肠道和土壤中。在一定条件下形成孢子。     

丁酸梭菌在医学领域中的研究进展

肠道益生菌是定植于人体肠道内,能产生确切健康功效从而改善宿主微生态平衡,对宿主发挥有益作用的一类活性微生物总称。最近越来越多的研究表明肥胖、糖尿病、肠易激综合征、骨质疏松等与肠道菌群密切相关,而丁酸梭菌由于能够促进其他有益菌（乳杆菌、双歧杆菌）生长,具有其他有益生菌没有的特性,受到越来越多研究者的重视。本文通过综述近几年有关丁酸梭菌与神经、内分泌、消化等系统中部分疾病的关系,总结了丁酸梭菌可能具有的治疗机制,并将其与益生菌在治疗骨质疏松症时所发挥的机制进行对比,以展望丁酸梭菌可能具有治疗骨质疏松的潜力。     

丁酸梭菌及产丁酸代谢改造

丁酸梭菌(Clostridium butyricum)是一种专性厌氧菌,可通过多基因过表达、同源重组、基于非复制型质粒和非复制型质粒的同源重组等多种遗传操作方式对其进行改造。丁酸是丁酸梭菌发酵的产物之一,丁酸用途广泛,用于饲料添加剂,可提高动物抵抗力,减少抗生素的使用。用丁酸梭菌发酵产丁酸,其产量仍然较低,不利于工业化生产,有必要通过代谢工程对丁酸梭菌产丁酸的途径进行优化。对丁酸梭菌的主要代谢途径、遗传操作体系及丁酸合成途径的优化等研究进展进行了综述。在此基础上,对丁酸梭菌进一步改造的思路和想法进行了展望。     

短链脂肪酸丁酸抑制动脉粥样硬化形成及其分子机制

本研究通过观察丁酸对动脉粥样硬化斑块形成以及肠道组织结构和功能的影响,探讨丁酸防治动脉粥样硬化的效应及可能机制.选取8周龄雄性载脂蛋白E基因敲除(apolipoprotein E-knockout,ApoE-/-)小鼠,随机分成对照组(高脂高胆固醇饲料+饮水中给予200 mmol/L氯化钠,n = 10)和丁酸组(高脂...     

特基拉芽孢杆菌XK29挥发物2-甲基丁酸对甘薯长喙壳菌的抑制作用研究

【目的】研究特基拉芽孢杆菌挥发物2-甲基丁酸对甘薯长喙壳菌的抑制作用,评价2-甲基丁酸对甘薯黑斑病的防治效果。【方法】采用I-分隔平皿和气相抑菌体系,研究不同剂量的2-甲基丁酸对甘薯长喙壳菌菌丝生长和孢子萌发的抑制作用;使用乳酸酚棉蓝染色观察2-甲基丁酸对甘薯长喙壳菌显微形态的影响;利用荧光探针钙荧光白和溴化丙锭检测2-甲基丁酸对甘薯长喙壳菌细胞壁结构与细胞膜通透性的影响;使用荧光探针2,7-二氯荧光素二乙酸酯检测甘薯长喙壳菌胞内活性氧含量变化;测定谷胱甘肽含量分析病原菌应对氧化损伤能力的改变;通过线粒体脱氢酶活力和丙酮酸含量的检测,分析2-甲基丁酸对甘薯长喙壳菌线粒体功能和能量代谢的影响;评价2-甲基丁酸在甘薯黑斑病防治中的使用效果。【结果】2-甲基丁酸显著抑制甘薯长喙壳菌菌丝生长和孢子萌发,降低其产孢能力,导致菌丝折叠弯曲并形成不连续的空腔。2-甲基丁酸使甘薯长喙壳菌的细胞壁结构改变,细胞膜通透性增加,胞内活性氧含量升高,谷胱甘肽含量显著降低,使病原菌应对氧化损伤的能力下降,线粒体脱氢酶活力和丙酮酸含量显著降低,诱发线粒体功能障碍,干扰细胞能量代谢,最终导致细胞死亡。此外,2-甲基丁酸对甘薯黑斑病也具有良好的防治作用。【结论】2-甲基丁酸对甘薯长喙壳菌具有显著的抑制作用,可作为安全高效的气相抑菌材料用于新型熏蒸制剂的研发。     

Butyric acid: Applications and recent advances in its bioproduction

Butyric acid is an important C4 organic acid with broad applications. It is currently produced by chemosynthesis from petroleum-based feedstocks. However, the fermentative production of butyric acid from renewable feedstocks has received growing attention because of consumer demand for green products and natural ingredients in foods, pharmaceuticals, animal feed supplements, and cosmetics. In this review, strategies for improving microbial butyric acid production, including strain engineering and novel fermentation process development are discussed and compared regarding product yield, titer, purity and productivity. Future perspectives on strain and process improvements for butyric acid production are also discussed.     

Efficient production of butyric acid from Jerusalem artichoke by immobilized Clostridium tyrobutyricum in a fibrous-bed bioreactor

Butyric acid is an important specialty chemical with wide industrial applications. The feasible large-scale fermentation for the economical production of butyric acid requires low-cost substrate and efficient process. In the present study, butyric acid production by immobilized Clostridium tyrobutyricum was successfully performed in a fibrous-bed bioreactor using Jerusalem artichoke as the substrate. Repeated-batch fermentation was carried out to produce butyric acid with a high butyrate yield (0.44 g/g), high productivity (2.75 g/L/h) and a butyrate concentration of 27.5 g/L. Furthermore, fed-batch fermentation using sulfuric acid pretreated Jerusalem artichoke hydrolysate resulted in a high butyric acid concentration of 60.4 g/L, with the yield of 0.38 g/g and the selectivity of ∼85.1 (85.1 g butyric acid/g acetic acid). Thus, the production of butyric acid from Jerusalem artichoke on a commercial scale could be achieved based on the system developed in this work.     

Effect of butyric acid on the germination of the seeds of Phacelia tanacetifolia

The effect of a weak acid (butyric acid) on the germination of seeds of Phacelia tanacetifolia Benth. (cv. Bleu Clair) has been studied. Butyric acid inhibited the early phase of germination, and the inhibition was correlated to the amount of the per-meant undissociated form present in the incubation medium. The inhibition by butyric acid in the dark was also correlated to a decrease of dark fixation of CO₂ and to a more pronounced decrease in malic acid levels during the early phase of germination; suggesting that the uptake of the uncharged form of this weak acid was followed by a release of H+ into the cytoplasm, leading to a decrease in its pH. The inhibitory effect of butyric acid in the dark on many metabolic events (rise in respiratory activity, levels of reducing sugars, glucose-6–phosphate and malic acid, dark CO₂ fixation, transport activities and macromolecular synthesis) appeared similar to the one of light. Fusicoccin (FC), which directly stimulates the H⁺ pump at the plasmalemma level, ameliorated the effect of butyric acid, as well as that of light, on germination. The bulk of these data is interpreted as suggesting that the mechanism of light inhibition of germination of Phacelia tanacetifolia seeds might be the consequence of a general block of metabolic reactivation due to the presence of an unfavourable (acidic) cytoplasmic pH; which might be explained with the lack of the phytochrome-dependent activation of the H⁺ pump at the plasmalemma level.     

Butyric acid retention in gingival tissue induces oxidative stress in jugular blood mitochondria

Butyric acid (BA) is a major extracellular metabolite produced by anaerobic periodontopathic bacteria and is commonly deposited in the gingival tissue. BA induces mitochondrial oxidative stress in vitro; however, its effects in vivo were never elucidated. Here, we determined the effects of butyric acid retention in the gingival tissues on oxidative stress induction in the jugular blood mitochondria. We established that BA injected in the rat gingival tissue has prolonged retention in gingival tissues. Blood taken at 0, 60, and 180 min after BA injection was used for further analysis. We isolated blood mitochondria, verified its purity, and measured hydrogen peroxide (H₂O₂), heme, superoxide (SOD), and catalase (CAT) to determine BA effects. We found that H₂O₂, heme, SOD, and CAT levels all increased after BA injection. This would insinuate that mitochondrial oxidative stress was induced ascribable to BA.     

生物技术生产丁酸的研究进展

丁酸作为一种重要的化工原料,已经广泛应用于食品添加剂与医药等领域。目前,工业上生产丁酸主要是从石油中提取有机化合物进行化学合成。与有机化合物合成法相比,微生物发酵产丁酸的优势有：所用的原料来源非常广,发酵过程低能耗,不污染环境,而且可以持续添加原料发酵生产丁酸。因此,通过生物技术发酵生产丁酸越来越受到人们的重视。介绍了丁酸的性质、产丁酸菌株的特点、微生物发酵产丁酸的细胞代谢途径及其调控、发酵法生产丁酸的工艺运行方式和产丁酸菌株及其代谢产物的生理功能这五部分内容,以期为今后开展发酵法产丁酸的微生物基因工程改造以及生产工艺的优化提供参考。     

Increased susceptibility to tumor necrosis factor-α in butyric acid-induced apoptosis is caused by downregulation of cFLIP expression in Jurkat T cells

Butyric acid is one of the major extracellular metabolites of periodontopathic Gram-negative bacteria. We previously demonstrated that butyric acid induced apoptosis in human T cells. In the present study, we examined the interaction between butyric acid and TNF-α in Jurkat T-cell apoptosis. Simultaneous treatment with TNF-α enhanced butyric acid-induced apoptosis by promoting caspase activity more than was achieved by either reagent alone. We examined which genes were associated with the increased susceptibility to TNF-α caused by butyric acid, and revealed that expression of cFLIP decreased with increased concentrations of butyric acid. Furthermore, exogenous expression of cFLIP protein suppressed the enhancing effect by TNF-α in the apoptosis. These results suggest that butyric acid downregulates cFLIP expression and increases the susceptibility to TNF-α by activating caspases via the death receptor signal.     

Candida albicans and Saccharomyces cerevisiae induce interleukin-8 production from intestinal epithelial-like Caco-2 cells in the presence of butyric acid

Intestinal epithelial cells (IEC) are important in initiation and regulation of immune responses against numerous foreign substances including food, microorganisms and their metabolites in the intestine. Since the responses of IEC against yeasts have not yet been well understood, we investigated the effects of Candida albicans, Saccharomyces cerevisiae, and their cell wall components on interleukin-8 (IL-8) secretion by the IEC-like Caco-2 cells. Live cells of both yeast species stimulated Caco-2 cells to produce IL-8 only in the presence of butyric acid, which is a metabolite produced by intestinal bacteria. S. cerevisiae zymosan and glucan also enhanced IL-8 secretion. Treatment of Caco-2 cells with butyric acid increased the expression of mRNAs coding for Toll-like receptor 1 (TLR1), TLR6 and dectin-1, which recognize zymosan. C. albicans induced more IL-8 secretion and also decreased transepithelial electrical resistance more rapidly than S. cerevisiae. These results suggest that both yeasts in the intestine stimulate the host's mucosal immune systems by interacting with IEC.     

Increased susceptibility to tumor necrosis factor-alpha in butyric acid-induced apoptosis is caused by downregulation of cFLIP expression in Jurkat T cells

Butyric acid is one of the major extracellular metabolites of periodontopathic Gram-negative bacteria. We previously demonstrated that butyric acid induced apoptosis in human T cells. In the present study, we examined the interaction between butyric acid and TNF-alpha in Jurkat T-cell apoptosis. Simultaneous treatment with TNF-alpha enhanced butyric acid-induced apoptosis by promoting caspase activity more than was achieved by either reagent alone. We examined which genes were associated with the increased susceptibility to TNF-alpha caused by butyric acid, and revealed that expression of cFLIP decreased with increased concentrations of butyric acid. Furthermore, exogenous expression of cFLIP protein suppressed the enhancing effect by TNF-alpha in the apoptosis. These results suggest that butyric acid downregulates cFLIP expression and increases the susceptibility to TNF-alpha by activating caspases via the death receptor signal.     

Metabolic engineering of Clostridium acetobutylicum for butyric acid production with high butyric acid selectivity

A typical characteristic of the butyric acid-producing Clostridium is coproduction of both butyric and acetic acids. Increasing the butyric acid selectivity important for economical butyric acid production has been rather difficult in clostridia due to their complex metabolic pathways. In this work, Clostridium acetobutylicum was metabolically engineered for highly selective butyric acid production. For this purpose, the second butyrate kinase of C. acetobutylicum encoded by the bukII gene instead of butyrate kinase I encoded by the buk gene was employed. Furthermore, metabolic pathways were engineered to further enhance the NADH-driving force. Batch fermentation of the metabolically engineered C. acetobutylicum strain HCBEKW (pta⁻, buk⁻, ctfB⁻ and adhE1⁻) at pH 6.0 resulted in the production of 32.5 g/L of butyric acid with a butyric-to-acetic acid ratio (BA/AA ratio) of 31.3 g/g from 83.3 g/L of glucose. By further knocking out the hydA gene (encoding hydrogenase) in the HCBEKW strain, the butyric acid titer was not further improved in batch fermentation. However, the BA/AA ratio (28.5 g/g) obtained with the HYCBEKW strain (pta⁻, buk⁻, ctfB⁻, adhE1⁻ and hydA⁻) was 1.6 times higher than that (18.2 g/g) obtained with the HCBEKW strain at pH 5.0, while no improvement was observed at pH 6.0. These results suggested that the buk gene knockout was essential to get a high butyric acid selectivity to acetic acid in C. acetobutylicum.