高效木薯渣分解复合菌群RXS的构建及其发酵特性研究

从富含腐烂纤维质的环境中取样,通过以木薯渣及滤纸为碳源的蛋白胨纤维素培养基不断地富集培养,构建了一组高效稳定的纤维质分解复合菌群.考察了该复合菌群对不同纤维质底物的分解性能及其在木薯渣水解过程中主要参数的变化.研究发现该复合菌群对滤纸、脱脂棉、微晶纤维素、麦秸秆和木薯渣等原料均能够进行有效的降解.在该复合菌群应用于木薯渣的水解过程中,监测发现纤维素酶、半纤维素酶、果胶酶等关键酶的酶活力分别在第2～3 d达到最大值34.4、90.5和15.8U;经过10 d的发酵后,木薯渣中的纤维素、半纤维素及木质素分别降解了79.8%、85.9%和19.4%,且木薯渣的失重高达61.5%;此外,代谢产物主要是乙酸、丁酸、己酸和甘油;而溶解性COD、总糖和总挥发酸的变化表明第2 d时木薯渣的水解率最高.上述结果表明,该复合菌群能够有效地水解木薯燃料酒精生产过程中的废弃物木薯渣,并有望用于木薯渣高效沼气发酵的前处理中.     

北方泥炭地泥炭土矿化速率、温度敏感性及其影响因素研究

北方泥炭地是全球重要的碳汇,也是全球变暖最为敏感的区域之一.然而,由于泥炭地表层和亚表层泥炭土碳排放过程对全球变暖的响应过程及机制仍存在一定争议,目前对全球变暖背景下泥炭地碳排放的认识仍存在一定不足.本研究于2019年8月在大兴安岭满归泥炭地采集表层（0~10 cm）和亚表层（15~30 cm）泥炭土进行室内增温模拟有氧培养,测定其矿化速率、有机质性质和水解酶活性.结果表明,表层泥炭土矿化速率在5、15、25 ℃下培养时（（142.8±66.9）~（545.3±30.6）、（575.0±62.1）~（1843.0±547.4） 、（888.4±123.9）~（3646.7±167.9） μg·g^-1·d^-1）均 高于亚表层（（113.0±41.5）~（367.1±64.1）、（357.4±52.3）~（1122.1±218.8）、（697.1±38.1）~（2336.4±150.6） μg·g^-1·d^-1）,但表层和亚表层矿化作用的温度敏感性不具有显著差异;培养过程中,表层与亚表层泥炭土β-1,4-N-乙酰葡糖胺糖苷酶（NAG）活性存在显著差异,但β-1,4-葡萄糖苷酶（BG）和酸性磷酸酶（AP）活性相似;表层和亚表层泥炭土总有机碳（TOC）、总氮（TN）、碳氮比（C/N）及纤维素含量均存在显著差异.在具体有机质组成方面,亚表层泥炭土中芳香族的惰性有机质含量较多,而表层泥炭土中多糖类的活性有机质含量较高,且芳香族化合物与泥炭土矿化速率呈显著负相关.综上,本研究认为全球变暖对表层和亚表层泥炭土矿化作用的影响没有显著性差异,常见的NAG、BG和AP酶并不是引起亚表层泥炭土矿化速率较慢的原因,而亚表层泥炭土中含有更多的惰性有机质可能是导致亚表层泥炭土矿化速率较低的主要因素.     

Review on the use of enzymes for the detection of organochlorine, organophosphate and carbamate pesticides in the environment

Pesticides are released intentionally into the environment and, through various processes, contaminate the environment. Three of the main classes of pesticides that pose a serious problem are organochlorines, organophosphates and carbamates. While pesticides are associated with many health effects, there is a lack of monitoring data on these contaminants. Traditional chromatographic methods are effective for the analysis of pesticides in the environment, but have limitations and prevent adequate monitoring. Enzymatic methods have been promoted for many years as an alternative method of detection of these pesticides. The main enzymes that have been utilised in this regard have been acetylcholinesterase, butyrylcholinesterase, alkaline phosphatase, organophosphorus hydrolase and tyrosinase. The enzymatic methods are based on the activation or inhibition of the enzyme by a pesticide which is proportional to the concentration of the pesticide. Research on enzymatic methods of detection, as well as some of the problems and challenges associated with these methods, is extensively discussed in this review. These methods can serve as a tool for screening large samples which can be followed up with the more traditional chromatographic methods of analysis.     

Properties of a Poly(3-hydroxybutyrate) Depolymerase from Penicillium funiculosum

A poly(3-hydroxybutyrate) (PHB) depolymerase was purified from a fungus, Penicillium funiculosum (IFO6345), with phenyl-Toyopearl and its properties were compared with those of other PHB depolymerases. The molecular mass of the purified enzyme was estimated at about 33 kDa by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis. The pH optimum and pI were 6.5 and 6.5, respectively. The purified protein showed affinity to Con A-Sepharose, indicating that it is a glycoprotein. Diisopropylfluorophosphate and dithiothreitol inhibited the depolymerase activity completely. The N-terminal amino acid sequence of the purified enzyme was TALPAFNVNPNSVSVSGLSSGGYMAAQL, which contained a lipase box sequence. This purified enzyme is one of the extracellular PHB depolymerase which belong to serine esterase. The purified enzyme showed relatively strong hydrolytic activity against 3-hydroxybutyrate oligomers compared with its PHB-degrading activity. PHB-binding experiments showed that P. funiculosum depolymerase has the weakest affinity for PHB of all the depolymerases examined.     

七株有机磷农药降解菌的降解特性比较

对分离自同一有机磷农药污染土壤的7株有机磷农药降解菌的降解特性进行了比较,7株降解菌都能利用甲基对硫磷为唯一碳源生长,并生成中间代谢产物对硝基苯酚.对硝基苯酚的降解经过一段延滞期,不同菌株降解对硝基苯酚的能力和延滞期有很大差异.降解菌株对多种有机磷类农药和芳香族化合物具有降解能力,其降解谱表现了一定的差异.用PCR方法从7株降解菌中克隆了有机磷农药水解酶基因.     

瘤胃微生物强化醋糟厌氧消化及其机制

针对醋糟中木质纤维素利用效率低的问题,通过接种瘤胃微生物可强化木质纤维素水解.采用逐步提升体系有机负荷的方式,考察瘤胃微生物生物强化对醋糟厌氧消化性能的提升效果,并运用绝对定量实时聚合酶链锁反应（Q-PCR）技术探究其微生物学强化机制.结果表明:长期连续运行成功塑造了高效的木质纤维素瘤胃强化体系.该体系的最高有机负荷达8.90 g/（L·d）（以VS计）,是强化前的1.53倍,该有机负荷下半纤维素和纤维素降解率分别达73.9%和40.1%,单位质量底物沼气和甲烷产量相应地分别达到451和261 mL/g（以VS计）,半纤维素和纤维素较高的降解率是该体系维持高产气性能的主要原因.生物相机制研究表明,瘤胃微生物强化体系中与木质纤维素水解密切相关的GH5（糖苷水解酶家族5）水解菌逐步富集,其基因拷贝数从初始的964×1010 copies/g升至最高有机负荷下的6.83×1011 copies/g,这是底物在高有机负荷下仍能被高效生物转化的根本原因.研究显示,瘤胃微生物的介入可有效强化体系底物的降解能力,促进醋糟产甲烷性能的提升.      

Insights into the structure of urea-like compounds as inhibitors of the juvenile hormone epoxide hydrolase (JHEH) of the tobacco hornworm Manduca sexta: analysis of the binding modes and structure-activity relationships of the inhibitors by docking and CoMFA calculations

Substituted urea compounds are well-known as potent inhibitors of juvenile hormone epoxide hydrolase (JHEH) of the tobacco hornworm Manduca sexta. Docking simulations of 47 derivatives inside JHEH were performed to gain insight into the structural characteristics of these complexes. The obtained orientations show a strong similitude with the observed in the known X-ray crystal structures of human soluble epoxide hydrolase (sEH) complexed with dialkylurea inhibitors. In addition, the predicted inhibitor concentration (IC₅₀) of the above-mentioned compounds as JHEH inhibitors were obtained by a quantitative structure-activity relationship (QSAR) method by using comparative molecular field analysis (CoMFA) applied to aligned dataset. The best models included steric and electrostatic fields and had adequate predictive abilities. In addition, these models were used to predict the activity of an external test set of compounds that was not used for building the model. Furthermore, plots of the CoMFA fields allowed conclusions to be drawn for the choice of suitable inhibitors.     

有机磷降解酶的固定化及其工业化应用初探

采用一种新型的酶固定化方法，在大孔微球中对有机磷降解酶（OPHC2）固定化，制备固相载体化的交联酶聚集体固定化酶。对具有实际应用意义的参数：热稳定性、牢固度和连续反应批次进行了重点研究。结果表明，有机磷降解酶固定化酶的半衰期在70℃时比游离酶提高了2.1倍。固定化酶分别在37℃的摇床中经14 h高速振荡，以及在pH 8、37℃条件下循环反应49个批次降解甲基对硫磷后，均未发现固定化酶酶活力下降。在37℃，有机磷降解酶固定化酶柱以每小时3倍柱床体积的速率对甲基对硫磷农药乳油的稀释液（甲基对硫磷含量为216 mg/L）进行降解处理，系统连续工作156 h，甲基对硫磷的降解率保持在98.4%～99.9%。     

Production, Purification and Activity of an Extracellular Depolymerase from Aspergillus fumigatus

A strain of Aspergillus fumigatus, which was observed to rapidly degrade poly-3-hydroxybutyrate (PHB) in a leaf compost, was found to secrete an extracellular hydrolase when grown on PHB as the sole carbon source. Isolation and characterization of the PHB hydrolase (depolymerase) from this fungus revealed that the enzyme had a molecular weight of 57 kDa, an isoelectric point of 7.2, and a PHB hydrolysis activity maxima which occurred at 70°C and pH 8.0. Affinity labeling experiments suggested that this fungal hydrolase is a type of serine esterase. The cyclic trimers of 3-hydroxybutyrate were found to reversibly inhibit the enzymes.     

一株高效降酚菌的分离鉴定及降酚特性研究

以苯酚为唯一碳源,采用逐量分批的方法从被酚类物质污染的土壤中驯化、筛选、分离降酚菌,利用形态学、培养特性、生理生化反应以及分子手段鉴定所得菌株,研究了该菌株生长与降酚的关系以及其降酚能力,并设计引物检测降酚酶基因。结果表明,获得的一株高效降酚菌GDYW-0027经鉴定为不动杆菌属(Acinetobacter),它的生长速率与苯酚的降解速率基本同步,48h内对高质量浓度(2.2g/L)苯酚的降解率为86.73%;在菌株GDYW-0027中检测到苯酚羟化酶及邻苯二酚2,3双加氧酶基因片段,推测其采用间位开环方式打开苯环。