温度对甲烷产生和氧化的影响

综述了温度对土壤产甲烷和氧化甲烷的影响及其机制．温度主要通过土壤中产甲烷菌的优势菌发生更替来改变土壤的产甲烷能力．较高温条件下产甲烷菌以乙酸和H2／CO2都能利用的甲烷八叠球菌(Methanosarcinaceae)为主,使得土壤处于较高的产甲烷状态．较低温条件下产甲烷菌以只能利用乙酸的甲烷毛菌(Methanosaetaceae)为主,土壤形成甲烷的能力相对较弱．温度提高可以显著地增加甲烷的产生,Q10为1．5-28,平均4．1,但是温度效应明显受控于底物浓度,提高底物浓度降低了产甲烷菌对底物的亲和力,相应地增加了度效应,因此在较低温条件下提高底物浓度可以促进甲烷的产生．温度对大气甲烷氧化的影响弱于产甲烷,甲烷氧化菌较少受温度变化的影响,即便在较低温条件下,土壤也具有一定的氧化大气甲烷能力,原因尚不清楚,可能与甲烷氧化菌对大气甲烷具有较高的亲和力有关,有待进一步研究．     

用木瓜蛋白酶及固定化木瓜蛋白酶拆分DL-苯丙氨酸

为了将手性化合物D-苯丙氨酸和L-苯丙氨酸进行分离,利用木瓜蛋白酶及固定化木瓜蛋白酶催化的方法对其拆分．试验结果表明,用DL-苯丙氨酸合成N-乙酰-DL-苯丙氨酸,得率为88.7％．木瓜蛋白酶、海藻酸钠 壳聚糖固定化木瓜蛋白酶(IPSAC)、尼龙布固定化木瓜蛋白酶(IPN)催化合成N-乙酰-L-苯丙氨酰苯胺时,对催化合成过程影响最大的因素分别是溶液中的离子强度、溶液中的离子强度、反应温度;溶液中的离子强度与pH对合成的影响较大．本试验得出分别用木瓜蛋白酶、IPSAC、IPN催化合成N-乙酰-L-苯丙氨酰苯胺的3个最佳方案;用此3个方案合成时,产率分别为61.2%、54.7%、36.3%．N-乙酰-L-苯丙氨酰苯胺水解生成L-苯丙氨酸,产率59.2%,光学纯度为96.6％．N-乙酰-D-苯丙氨酸水解生成D-苯丙氨酸,产率61.7%,光学纯度为95.7％．     

培养基组成对树状多节孢(Nodulisporium sylviforme)紫杉醇产量的影响

采用正交试验法 ,研究了醋酸钠、苯丙氨酸、酪氨酸、亮氨酸对紫杉醇产生菌HQD3 3 产生紫杉醇的影响。结果表明 ,它们之间的协同作用对提高紫杉醇产量有显著影响。在改良的S - 7培养基基础上 ,再加入醋酸钠 1 0g/L、苯丙氨酸 5 0mg/L、酪氨酸 1 5mg/L、亮氨酸 6 0mg/L ,可以使紫杉醇产生菌HQD3 3 产生紫杉醇产量提高到 2 0 3 6 5 6 μg/L。     

荧光假单胞菌TM5-2中D-型氨基酸脱氢酶的鉴定和表达

从荧光假单胞菌TM5-2中得到一个含丙氨酸消旋酶基因的DNA片段(8.8kb),相邻的一个开读框(ORF)与甘氨酸/D-型氨基酸氧化酶基因相似。该ORF经过克隆、表达,并没有检测到甘氨酸/D-型氨基酸氧化酶的活性,推导而得的氨基酸序列与D-型氨基酸脱氢酶序列比较发现,ORF含有D-型氨基酸脱氢酶的所有重要的保守序列。经TTC培养基鉴定,其具有D-型氨基酸脱氢酶的活性,并对一系列D-型氨基酸有作用,最佳作用底物是D-组氨酸。     

粘质沙雷氏菌H3010发酵型D-乳酸脱氢酶基因的克隆表达、纯化及酶学性质研究

通过PCR技术从粘质沙雷氏菌H3010基因组DNA中扩增出该D-乳酸脱氢酶基因,连接至pET-28a（＋）表达载体,转入大肠杆菌BL21（DE3）中进行了重组表达,优化了酶纯化的条件,并对其酶学性质进行初步研究。结果表明,获得的该酶编码基因全长993bp,编码330个氨基酸,大小为37kDa。经优化表达及纯化条件后重组酶纯度可达90％。酶学性质研究发现,该重组酶最适反应温度为60℃,最适酶促反应pH为7．5（O．2mol／L磷酸盐缓冲液）,37℃下测得对底物丙酮酸的动力学参数Km=3．39mmol／L,Vmax=6．87mmol／（mg·min）,对辅酶NADH的动力学参数Km=1．43mmol／L,Vmax=1．61mmo]／（mg·min）。为酶法生产D-乳酸及利用代谢工程构建产D-乳酸的基因工程菌打下基础。     

一株食用菌生物转化富集γ-氨基丁酸条件研究

从本实验室保存的食用菌中筛选到菌株CSP501,其菌丝体可以谷氨酸钠为底物转化富集γ-氨基丁酸。通过单因素试验和正交试验,菌株CSP501转化富集γ-氨基丁酸的最适条件：转化温度45℃,转化时间5h,体系pH4．0,底物浓度1％。γ-氨基丁酸的含量可以达到干重的1．05％,达到较高的含量。     

儿童细菌性腹泻病原菌分布及耐药性分析

目的 了解小儿细菌性腹泻的病原菌分布及耐药情况。方法 对773例临床上疑为细菌性腹泻病的患儿的粪便标本进行培养、鉴定及药物敏感试验、超广谱β-内酰胺酶（ESBLs）检测。结果 773例标本中分离出阳性菌株671株,总检出率为86．6％。其中埃希菌属、志贺菌属及耶尔森菌3种致病性细菌占阳性菌株的54．5％,肠杆菌属、非发酵菌群、酵母样真菌、肠球菌属、变形杆菌属、克雷伯菌属和枸橼酸杆菌属等条件致病菌占阳性菌株的45．5％。埃希菌属及志贺菌属对头孢他啶及氨曲南耐药率低（16．4％以下）,而肠杆菌属等条件致病菌对常用抗生素耐药率均较高。G^-杆菌中ESBLs的检出率为38．3％,其中大肠埃希菌占66．7％。结论 埃希菌属是儿童细菌性腹泻病的主要致病菌（50．2％）,且耐药率及ESBLs产生率高,但肠杆菌属等条件致病菌检出率也较高（45．5％）,且耐药率高,可能与抗生素广泛使用,肠杆菌群失调有关,应引起高度重视。     

蝙蝠蛾拟青霉菌丝体发酵的研究

用20t发酵罐培养蝙蝠蛾拟青霉。菌粉收率达2．5％,D-甘露醇含量大于8％,腺苷含量大于0．2％．对发酵过程中pH、总糖、还原糖、得率、折光等的变化规律进行了考察．确定以发酵液pH的升高和还原糖含量作为综合判断发酵终点的依据．     

紫外诱变原生质体选育D-核糖生产菌株

以D-核糖产生菌枯草芽孢杆(Bacillus subtilis)B941为出发菌株,采用紫外诱变原生质体的方法,获得了4株可以在含有6．0％D-核糖的培养基上生长的D-核糖高产菌株,其摇瓶发酵产糖达55．0g／L左右。通过摇瓶发酵试验,研究了D-核糖高产菌株Buvp-24的遗传稳定性。研究结果表明,经多次传代,菌株Buvp-24的发酵产糖能力及对发酵产物D-核糖的耐受性没有改变,有望应用于工业生产中。     

D-氨基酰化酶拆分D,L-苯丙氨酸制备D-苯丙氨酸

进行了以D,L-苯丙氨酸为原料经D-氨基酰化酶制备D-苯丙氨酸的研究。乙酰-D,L-苯丙氨酸浓度为0.5mol.L-1,给酶量为3×104U.L-1时,24 h拆分率可达到97%。采用阳离子交换树脂进行了拆分液中的D-苯丙氨酸的分离,D-苯丙氨酸的收率为95.4%。采用醋酸酐作为催化剂,在145℃的条件下,乙酰-L-苯丙氨酸可以消旋成乙酰-D,L-苯丙氨酸继续拆分。     

梨头霉11α—羟基化制备16β—甲基—11α,17α21—三烃基孕甾—1, …

选育到一株对１６β－甲基－１７α,２１－二羟基孕甾－１,４＝－二烯－３,２０－二酮（Ⅱａ）１１α－羟基化活性强的梨头霉Ａ２８菌株,并发现底物２１－乙酰化（Ⅱｂ）可明显提高１１α－羟工 能力。在适宜的转化条件下,１１ｂ投料浓度０．５％,产物１６β－基１１α,１７α,２１－三羟基孕甾－１,４－二烯－３,２０－二酮（Ⅲ）收率为７３％,结构经波谱分析确认。     

Growth of Agrobacterium tumefaciens under octopine limitation in chemostats.

Agrobacterium tumefaciens B6 and ATCC 15955 were grown under octopine or glutamate limitation in chemostats. Examination of the maximum specific growth rate (mu max) and substrate affinity (KS) for each strain indicated that strain B6 was highly inefficient in its use of octopine as either a nitrogen or carbon source compared with strain ATCC 15955. Examination of the yield coefficients showed that in both strains octopine was used more efficiently as a nitrogen source than as a carbon source. The data permitted predictions to be made concerning the outcome of competition for a single limiting substrate. Under octopine limitation, strain ATCC 15955 should dominate; under glutamate limitation, strain B6 should dominate. The results of an observed competition with glutamate as the limiting substrate confirmed the latter prediction, although B6 did dominate at a rate faster than was predicted from simple competition theory. B6 displayed higher growth rates and substrate affinities than ATCC 15955 on all concentrations of glutamate. The yield of B6 on octopine was also considerably higher. This latter attribute could provide an ecological advantage to B6 because of the importance of yield in the fate of competitions under multisubstrate regimens. These will be the most prevalent regimens in the area around the tumor (tumorosphere) or the rhizosphere. The increased performance on glutamate could provide an advantage in an opine-free environment when B6 is growing as a saprophyte.     

Growth of Agrobacterium tumefaciens under octopine limitation in chemostats

Agrobacterium tumefaciens B6 and ATCC 15955 were grown under octopine or glutamate limitation in chemostats. Examination of the maximum specific growth rate (mu max) and substrate affinity (KS) for each strain indicated that strain B6 was highly inefficient in its use of octopine as either a nitrogen or carbon source compared with strain ATCC 15955. Examination of the yield coefficients showed that in both strains octopine was used more efficiently as a nitrogen source than as a carbon source. The data permitted predictions to be made concerning the outcome of competition for a single limiting substrate. Under octopine limitation, strain ATCC 15955 should dominate; under glutamate limitation, strain B6 should dominate. The results of an observed competition with glutamate as the limiting substrate confirmed the latter prediction, although B6 did dominate at a rate faster than was predicted from simple competition theory. B6 displayed higher growth rates and substrate affinities than ATCC 15955 on all concentrations of glutamate. The yield of B6 on octopine was also considerably higher. This latter attribute could provide an ecological advantage to B6 because of the importance of yield in the fate of competitions under multisubstrate regimens. These will be the most prevalent regimens in the area around the tumor (tumorosphere) or the rhizosphere. The increased performance on glutamate could provide an advantage in an opine-free environment when B6 is growing as a saprophyte.     

一株产电菌Nitratireductor sp. WJ5-4的筛选及产电分析

【目的】从生物垃圾燃料电池阳极淋洗液中分离一株产电菌WJ5-4,研究其产电特性。【方法】根据菌株的形态、生理生化性质及16S r RNA基因测序分析确定其种属,以该菌株为产电菌,以生物垃圾为底物,构建微生物燃料电池(Microbial fuel cell,MFC),研究在不同接种浓度和底物固含量条件下菌株的产电性能。【结果】菌株WJ5-4被初步鉴定属于Nitratireductor属,当接种量200 m L时可获得最大功率密度135.16 m W/m2、稳定电压370 m V和总有机碳(Total organic carbon,TOC)降解率41.46%。当底物固含量为23%时,可获得最大功率密度163.69 m W/m2、稳定电压434 m V和TOC降解率46.29%。【结论】WJ5-4菌能够利用较高固含量的生物垃圾产电,产电周期较长,为下一步微生物燃料电池处理生物垃圾提供科学依据。     

一株高效转化Phytosterol为雄甾烯酮菌株的筛选与鉴定

从保藏的200多株菌中筛选出1株高效转化植物甾醇为4-烯-雄甾-3,17-二酮和1,4-二烯-雄甾-3,17-二酮的菌株,并对该菌进行了形态、生理生化的研究。结果发现菌株ST06可以利用多种碳源,可以水解淀粉,但不利用纤微素。用16SrDNA的方法对其进行鉴定,发现与Bacillus属Bacillus amyloiquefaciens的相似性最高,达到99．9％,将该菌株命名为Bacillus amyloiquefaciens ST06。该菌在培养温度30℃,pH7．0,转速220r／min,转化时间7d,底物添加量为0．3％时,ADD与AD的总得率高达40％以上,此时底物转化率高达93．7％。     

Oxytetracycline production in solid state and submerged fermentation by protoplast fusants of Streptomyces rimosus

Streptomyces rimosus TM-55 was treated with 3% EMS, and 29 auxotrophic mutants (AM-1 - AM-29) were isolated from 5457 colonies at a survival rate between 6.6-66.7%. Three sets of the auxotrophic mutants, AM-3 and AM-27, AM-7 and AM-28, and AM-3 and AM-21, were chosen for protoplast fusion with 50% PEG 1000 for 30 minutes at 25 degrees C, and 25 fusants were isolated (f-1 - f-25). In solid substrate, oxytetracycline production of 20% fusants was higher than that of the wild strain, while in submerged fermentation, it was 44%. Oxytetracycline productions of fusants f-1, f-6, f-11, f-12, f-20 and f-21 were lower than that of the wild strain in solid substrate, but this was reversed in submerged fermentation. On the other hand, OTC production of fusant f-8 was higher than that of the wild strain in solid substrate, and this was reversed in submerged fermentation.     

Dependence of the genotypic characteristics of Acidithiobacillus ferrooxidans on the physical, chemical, and electrophysical properties of pyrites

Comparison of Acidithiobacillus ferrooxidans strains TFV-1 and TFBk with respect to their capacity to oxidize pyrite 1, with hole-type (p-type) conductivity, or pyrite 2, with an electron-type (n-type) conductivity, showed that, at a pulp density of 1%, both before and after its adaptation to the pyrites, strain TFBk, isolated from a substrate with a more complex mineral composition, grew faster and oxidized the pyrites of both conductivity types more efficiently than strain TFV-1, which was isolated from a mineralogically simple ore. At a pulp density of 3-5%, the oxidation of pyrite 1 by strain TFV-1 and both of the pyrites by strain TFBk began only after an artificial increase in Eh to 600 mV. If the pulp density was increased gradually, strain TFBk could oxidize the pyrites at its higher values than strain TFV-1, with the rate of pyrite 2 oxidation being higher than that of pyrite 1. During chemical oxidation of both of the pyrites, an increase was observed in the absolute values of the coefficients of thermoelectromotive force (KTEMF); during bacterial-chemical oxidation, the KTEMF of pyrite 1 changed insignificantly, whereas the KTEMF of pyrite 2 decreased.     

Enzymatic saccharification of pretreated rice straw and biomass production

A comparative study on the saccharification of pretreated rice straw was brought about by using cellulase enzyme produced by Aspergillus terreus ATCC 52430 and its mutant strain UNGI-40. The effect of enzyme and substrate concentrations on the saccharification rate at 24 and 48 were studied. A syrup with 7% sugar concentration was obtained with a 10% substrate concentration for the mutant case, whereas a syrup with 6.8% sugar concentration was obtained with 3.5 times concentrated enzyme from the wild strain. A high saccharification value was obtained with low substrate concentration; the higher the substrate concentration used, the lower the percent saccharification. The glucose content in the hydrolysate comprised 80-82% of total reducing sugars; the remainder was cellobiose and xylose together. The hydrolysate supported the growth of yeasts Candida utilis and Saccharomyces cerevisiae ATCC 52431. A biomass with a 48% protein content was obtained. The essential amino acid composition of yeast biomass was determined.     

高对映选择性环氧化物水解酶产生菌的筛选及特性研究

从土壤中分离的芽孢杆菌Bacillus megaterium ECU1001所产五氧化物水解酶能高对映选择性水解缩水甘油苯基醚（对映选择率E值可达47.8）,当转化率为55.9%时,剩余的（S）－缩水甘油苯基醚的光学纯度（对映体过量值,ee）可达99.5%;当底物浓度提高到60mmol/L时,光学纯（S）－缩水基油苯基醚的收率达到25.6%。     

Production of a microbial lipase by Staphylococcus carnosus (pLipMut2) in a bubble column reactor and a centrifugal field bioreactor

Extracellular lipase production by the recombinant strain Staphylococcus carnosus (pLipMut2) has been studied. First substrate optimization was carried out in shaken cultures. As a result, the best substrate yield of 20 units/g (peptone + yeast extract) and maximum lipase activity in the culture supernatant of 1.7 units/cm³ could be obtained by a nutrient rich complex medium consisting of 75 kg/m³ yeast extract, 15 kg/m³ tryptone, 5 kg/m³ glucose and 0.5 kg/m³ K₂HPO₄. Higher initial substrate concentration caused inhibition of growth. Antifoam agent at higher levels than 1 cm³/ dm³ resulted in a negative influence on lipase yield. Comparative fermentation studies have been carried out in a bubble column reactor and in a centrifugal field bioreactor. Direct proportionality between growth, lipase production and oxygen consumption was observed. In the bubble column reactor usual superficial air velocities (4 cm/s) caused intensive foam generation, thus fermentation was only possible after installation of a broader column head to allow coalescence. In the centrifugal field bioreactor higher productivities were obtained without foam problems at superficial gas velocities which were one order of magnitude lower than in the bubble column. Fermentations have been performed batchwise and without holding pH constant. Neither pH control nor glucose feeding could improve the substrate yield further. Compared to former fermentation studies with the strain S. carnosus (pLipPS1) lipase yield (lipase activity/cell density) could be improved by 300% and substrate yield (lipase activity/substrate concentration) by 600%.