土霉素残留对蔬菜发酵过程微生物群落演替及代谢产物的影响

【目的】探究土霉素残留对蔬菜自然发酵过程中微生物群落演替和代谢产物动力学的影响,为评估抗生素残留对蔬菜发酵的影响提供理论基础。【方法】超高效液相色谱-串联质谱法测定土霉素残留;高效液相色谱法测定有机酸、电子鼻和气相色谱-质谱联用测定挥发性成分和高通量技术测定微生物种类。【结果】蔬菜自然发酵过程中,土霉素残留从4.00 mg/L下降到2.53 mg/L;不含抗生素残留的蔬菜发酵含有同型和异型乳酸发酵,而土霉素残留的蔬菜发酵仅含有同型乳酸发酵;同时,其特征微生物由Lactobacillus pentosus和Lactobacillus plantarum转变为Lactobacillus paratarrginis、Lactobacillus buchneri和Lactobacillus kisonensis;土霉素残留明显影响了乳酸、柠檬酸、乙酸、香茅醇、3-辛醇、异硫氰酸烯丙酯、乙酸香叶酯、乙烯基硬脂醚和异硫氰酸苯乙酯等代谢产物的含量。【结论】土霉素残留影响了蔬菜乳酸发酵的类型、微生物群落的演替、有机酸和挥发性化合物的形成过程,因此应将抗生素残留纳入发酵蔬菜原料的质量控制指标。     

反相高效液相色谱法测定肠球菌FQ15发酵液中的有机酸

目的研究肠球菌FQ15发酵液中的有机酸在不同发酵时间的变化趋势。方法建立反相高效液相色谱法测定此株益生菌发酵液中有机酸的主要成分及其在发酵不同时间的变化趋势。结果丙酸、丙酮酸、乳酸、乙酸的浓度都是在菌体生长处于衰亡期时达到最大值,乳酸随发酵时间的延长浓度明显下降,而乙酸在发酵后期含量呈上升趋势。结论此方法重现性好,精密度高,为研究微生物合成中有机酸种类及变化趋势提供了一种可供参考的快捷分析手段。     

高效液相色谱法在测定丁二酸厌氧发酵体系中有机酸的应用

建立了一种利用高效液相色谱法定量分析丁二酸厌氧发酵体系中多种有机酸的方法。利用Alltech反相Prevail有机酸色谱柱,以25 mmol.L-1KH2PO4(pH2.5)作为流动相,流速1 mL.min-1,采用紫外检测器,于215 nm处检测,能将丁二酸厌氧发酵体系中多种有机酸完全分离并准确定量。有机酸的回收率均在99%~103%之间。本方法能够快速、精确测定丁二酸厌氧发酵体系中多种有机酸含量,并初步应用于该发酵体系培养基成分优化方面,对于指导厌氧代谢调控生产丁二酸具有重要意义。     

一株太子参内生真菌Aspergillus terreus TZS-201607中抗肿瘤活性代谢产物研究

为研究太子参内生真菌Aspergillus terreus TZS-201607所产的次级代谢产物,利用硅胶柱层析、Sephadex LH-20凝胶柱层析、反相柱层析及半制备高效液相色谱等技术,从其PDB培养基发酵产物的乙酸乙酯萃取物中分离纯化得到16个单体化合物.利用波谱学方法结合文献数据分析分别鉴定为柄曲霉素(1)...     

海洋链霉菌Streptomyces sp. SCSIO 1672及其代谢产物水杨酸的分离鉴定

从南海海洋沉积物中分离得到1株海洋放线菌,鉴定为链霉菌Streptomyces sp. SCSIO 1672。通过优化发酵条件,采用海虾生物致死活性和高效液相色谱追踪,利用有机溶剂萃取、正相硅胶、反相硅胶等各种色谱层析方法分离出活性化合物,通过波谱数据解析出海洋放线菌SCSIO 1672次级代谢产物中的该活性化合物为水杨酸。     

高效液相色谱测定微量胆固醇氧化产物

介绍一种高效液相色谱测定胆固醇氧化产物的方法,以苯甲酰氯为衍生剂,将胆固醇及其氧化产物衍生成苯甲酸酯,反相高效液相色谱分离,紫外检测,内标法定量．本法灵敏度高,重现性好,可应用于胆固醇纯度标准物质的杂质分析及血清中的胆固醇氧化产物测定。     

气相色谱检出和鉴定厌氧菌方法的改进

用程序升温毛细管气相色谱法分析厌氧菌代谢产物中的有机酸丁酯,有可能代替常规方法中的两步色谱分析,简化了实验程序,缩短了时间,初步验证结果表明改进方法的可行性。     

一株放线菌次生代谢产物化学成分的研究

目的：研究放线菌这类重要的可再生资源的次生代谢产物的化学成分。方法：通过对一株采集于云南西部土壤放线菌的发酵培养,发酵液经TLC分离纯化、高效液相分析检测、核磁共振和质谱测定。结果：从其次生代谢产物中分离获取了8个化合物,其中4个化合物的结构已经初步确定。结论：4个化合物中两个具有抗肿瘤活性,两个具有抑菌活性。     

重组大肠杆菌YK537/pSB┐TK高密度培养过程中有机酸的RP┐HPLC快速分析

采用高压液相色谱法（ＨＰＬＣ）,利用Ｃ１８反相柱快速检测重组大肠杆菌ＹＫ５３７／ｐＳＢ－ＴＫ高密度发酵过程中细菌代谢产生的各种有机酸。分析表明,经乙醚和水两次萃取处理后能去除样品中大部分的干扰峰,可获得理想的分离效果,并能定量测出发酵过程中各种有机酸的积累。不同培养方式的结果表明用甘油代替葡萄糖作为细菌生长的碳源可减少乙酸的积累,降低其对细菌生长和外源蛋白表达的抑制作用,从而更易实现重组大肠杆菌的高表达、高密度培养。     

2种检测灯盏花素血药浓度方法的比较

目的：建立测定Beagle犬血浆中灯盏花素浓度的反相高效液相色谱法和液相-质谱联用法,并将其进行多参数的两两比较。方法：采用液-液萃取法处理血清,反相高效液相色谱法测定,流动相为甲醇-水（pH3．0）=42：58,EclipseXDB-C18为固定相;将同样的样品用液相-质谱法开展平行检测;将所测得的数据做两两对比,采用SPSS统计软件进行分析。结果：2种方法测定的数据具有良好的相关性（r=0．982,P〉0．05）;最低检测限前者为0．05μg／mL,后者为0．01μg／mL,反相高效液相色谱法的最低检测限和灵敏度不及液相-质谱联用法。结论：反相高效液相色谱法可用于灯盏花素毒代动力学研究,与液相-质谱联用法检测结果无显著差异,且费用较低,不失为-种低成本且行之有效的检测方法。     

Anaerobic digestion: microbial and biochemical aspects of volatile acid production

Summary The production of organic acids has been tested with bacterial flora selected from a municipal sludge digestor. In order to elucidate the basic mechanisms by which glucose is converted to volatile fatty acids, the examination of non-methanogenic bacteria was attempted. Both lactate-producers and lactate-utilizers were found among these bacteria. When mixed isolates were used as the inoculum, the accumulation of lactic acid and its further conversion to propionic and butyric acids was demonstrated at a carbon conversion rate of about 0.75. It is therefore suggested that this metabolic sequence may occur as a normal process in acidogenic fermentation, which is the first step in anaerobic digestion.     

pH-dependent uptake of fumaric acid in Saccharomyces cerevisiae under anaerobic conditions

Microbial production of C(4) dicarboxylic acids from renewable resources has gained renewed interest. The yeast Saccharomyces cerevisiae is known as a robust microorganism and is able to grow at low pH, which makes it a suitable candidate for biological production of organic acids. However, a successful metabolic engineering approach for overproduction of organic acids requires an incorporation of a proper exporter to increase the productivity. Moreover, low-pH fermentations, which are desirable for facilitating the downstream processing, may cause back diffusion of the undissociated acid into the cells with simultaneous active export, thereby creating an ATP-dissipating futile cycle. In this work, we have studied the uptake of fumaric acid in S. cerevisiae in carbon-limited chemostat cultures under anaerobic conditions. The effect of the presence of fumaric acid at different pH values (3 to 5) has been investigated in order to obtain more knowledge about possible uptake mechanisms. The experimental results showed that at a cultivation pH of 5.0 and an external fumaric acid concentration of approximately 0.8 mmol · liter(-1), the fumaric acid uptake rate was unexpectedly high and could not be explained by diffusion of the undissociated form across the plasma membrane alone. This could indicate the presence of protein-mediated import. At decreasing pH levels, the fumaric acid uptake rate was found to increase asymptotically to a maximum level. Although this observation is in accordance with protein-mediated import, the presence of a metabolic bottleneck for fumaric acid conversion under anaerobic conditions could not be excluded.     

Selective production of organic acids in anaerobic acid reactor by pH control

The selective production of organic acids by anaerobic acidogenesis with pH control was examined using a chemostat culture. The results showed that the product spectrum in the acid reactor strongly depended on the culture pH. Under acidic and neutral conditions, the main products were butyric acid, while acetic and propionic acids were the main products under the basic condition. This phenomenon was reversible between the acidic and basic conditions, and was not affected by the dilution rate. The change in the main products was caused by the change in the dominant microbial populations, from butyric acid-producing bacteria to propionic acid-producing bacteria in the acid reactor due to the pH shift. The control of culture pH was considered to be a useful way for controlling the product spectrum in the anaerobic acid reactor.     

Nitrogenous compounds stimulate glucose‐derived acid production by oral Streptococcus and Actinomyces

Both Streptococcus and Actinomyces can produce acids from dietary sugars and are frequently found in caries lesions. In the oral cavity, nitrogenous compounds, such as peptides and amino acids, are provided continuously by saliva and crevicular gingival fluid. Given that these bacteria can also utilize nitrogen compounds for their growth, it was hypothesized that nitrogenous compounds may influence their acid production; however, no previous studies have examined this topic. Therefore, the present study aimed to assess the effects of nitrogenous compounds (tryptone and glutamate) on glucose‐derived acid production by Streptococcus and Actinomyces. Acid production was evaluated using a pH‐stat method under anaerobic conditions, whereas the amounts of metabolic end‐products were quantified using high performance liquid chromatography. Tryptone enhanced glucose‐derived acid production by up to 2.68‐fold, whereas glutamate enhanced Streptococcus species only. However, neither tryptone nor glutamate altered the end‐product profiles, indicating that the nitrogenous compounds stimulate the whole metabolic pathways involving in acid production from glucose, but are not actively metabolized, nor do they alter metabolic pathways. These results suggest that nitrogenous compounds in the oral cavity promote acid production by Streptococcus and Actinomyces in vivo.     

谷氨酸棒杆菌生产琥珀酸的代谢工程研究进展

琥珀酸是一种具有重要应用价值的四碳平台化合物。微生物法发酵生产琥珀酸以其社会、环境和经济优势展现出良好的发展前景。谷氨酸棒杆菌被广泛应用于氨基酸、核苷酸等高附加值化学品的工业化生产,在厌氧条件下细胞处于生长停滞状态,但仍能高效利用碳源合成有机酸,通过代谢工程改造的谷氨酸棒杆菌有望成为理想的琥珀酸生产菌株。结合近年来谷氨酸棒杆菌生产琥珀酸取得的最新成果,本文综述了构建高产琥珀酸工程菌株的代谢工程策略、底物的扩展利用,并展望了将来的研究方向。     

Gas chromatographic detection of sulphur compounds as methyl esters after growth of anaerobic bacteria in broth media

The use of a highly polar capillary column permits gas chromatographic analysis of organic acids as methyl esters. This method has found use in the study of end products of anaerobic bacteria. In addition, it also permits the detection of nonvolatile sulphur compounds which are neglected on the usual packed columns. These compounds have been identified by gas chromatography-mass spectrometry as methyl esters of 3-(methylthio)-propanoic acid and 4-(methylthio)-butanoic acid. When certain species were grown in a medium supplemented with 0·4% (w/v) DL-methionine, the relative amounts of both acids increased significantly. These nonvolatile sulphur compounds may serve as markers for specific bacteria.     

Expression of Different Levels of Ethanologenic Enzymes from Zymomonas mobilis in Recombinant Strains of Escherichia coli

The expression of Zymomonas mobilis genes encoding pyruvate decarboxylase and alcohol dehydrogenase II in Escherichia coli converted this organism from the production of organic acids to the production of ethanol. Ethanol was produced during both anaerobic and aerobic growth. The extent to which these ethanologenic enzymes were expressed correlated with the extent of ethanol production. The replacement of organic acids with ethanol as a metabolic product during aerobic and anaerobic growth resulted in dramatic increases in final cell density, indicating that these acids (and the associated decline in pH) are more damaging than the production of ethanol. Of the plasmids examined, the best plasmid for growth and ethanol production expressed pyruvate decarboxylase and alcohol dehydrogenase II at levels of 6.5 and 2.5 IU/mg of total cell protein, respectively.     

城市污泥中邻苯二甲酸酯(PAEs)的厌氧微生物降解

邻苯二甲酸酯(PAEs)是城市污泥中普遍存在的一类具有内分泌干扰性作用的有机污染物.研究污泥厌氧生物处理过程中PAEs的微生物降解对保障污泥农用的安全性十分必要.本文以污泥中两种主要的PAEs——邻苯二甲酸二丁酯(DBP)和邻苯二甲酸(2-乙基己)酯(DEHP)为研究对象,通过比较PAEs在污泥厌氧消化系统与发酵产氢系统中降解过程的差异及系统污泥特性的变化,分析不同污泥厌氧生物处理系统中影响PAEs降解的可能因素.结果表明: 在污泥厌氧发酵系统中,DBP在6 d内降解率达99.6%, DEHP在整个14 d的培养期间也降解了46.1%;在发酵产氢系统中,在14 d培养过程DBP的降解率仅为19.5%,DEHP则没有明显的降解.与厌氧消化系统相比,PAEs在发酵产氢系统中的降解受到明显抑制,这与发酵产氢过程中微生物量下降、革兰氏阳性菌/革兰氏阴性菌(G⁺/G^-)和真菌/细菌变小及挥发性脂肪酸(包括乙酸、丙酸及丁酸)浓度升高有关.     

Anaerobic Phenol Degradation by Microorganisms of Swine Manure

Swine manure contains diverse groups of aerobic and anaerobic bacteria. An anaerobic bacterial consortium containing sulfate-reducing bacteria (SRB) and acetate-utilizing methanogenic bacteria was isolated from swine manure. This consortium used phenol as its sole source of carbon and converted it to methane and CO₂. The sulfate-reducing bacterial members of the consortium are the incomplete oxidizers, unable to carry out the terminal oxidation of organic substrates, leaving acetic acid as the end product. The methanogenic bacteria of the consortium converted the acetic acid to methane. When a methanogen inhibitor was used in the culture medium, phenol was converted to acetic acid by the SRB, but the acetic acid did not undergo further metabolism. On the other hand, when the growth of SRB in the consortium was suppressed with a specific SRB inhibitor, namely, molybdenum tetroxide, the phenol was not degraded. Thus, the metabolic activities of both the sulfate-reducing bacteria and the methanogenic bacteria were essential for complete degradation of phenol. Received: 31 January 1997 / Accepted: 7 March 1997     

Changes in lactic acid bacteria and components of Awa-bancha by anaerobic fermentation

ABSTRACT

Awa-bancha is a post-fermented tea produced in Naka and Kamikatsu, Tokushima, Japan. We investigated the lactic acid bacteria in each stage of production of Awa-bancha and evaluated the relationships with the components. Lactic acid bacteria were isolated from tea leaves cultured with de Man, Rogosa, and Sharpe (MRS) agar plates, and the species were identified by homology of the 16 S rRNA gene and multiplex polymerase chain reaction (PCR) of the recA gene to distinguish the Lactobacillus plantarum group. As a result, a variety of species were isolated from the raw tea leaves, and Lactobacillus pentosus was isolated most frequently after anaerobic fermentation. Regarding the tea leaf components, organic acids, such as lactic acid, increased, free amino acids decreased, and catechins changed owing to anaerobic fermentation. Our results suggest that the microbial flora mainly composed of L. pentosus is important in the anaerobic fermentation process for flavor formation of Awa-bancha.