己酸菌的分离筛选及发酵条件的研究

己酸菌所生成的己酸是生产浓香型白酒主体香气己酸乙酯的主要成分,将其应用于白酒的生产,可提高白酒的香气和质量．从我省富裕酒厂的窖泥中分离到一株高产己酸的菌株,通过发酵条件的研究,该菌株己酸最高产量可达540mg／100ml。     

α-淀粉酶高产菌株选育方法的研究

在含有氨苄青霉素1—10μg/ml的LB液体培养基中,接入枯芽孢杆菌BF7658菌悬液,在37℃下振荡培养3—5天,经平板分离单菌落以及摇瓶发酵试验,初筛获得2213、2104和2120等α-淀粉酶高产菌株。将2213菌株的菌悬液涂布于含有4-6μg/ml氨苄青霉素的2%淀粉培养基上,复筛得到4213、4218、4237等α-淀粉酶高产菌株。4213菌株再经4次亚硝基胍诱变,未能得到蛋白酶活性低、α-淀粉酶活性高的突变株。     

α-淀粉酶高产菌株选育方法的研究

在含有氨苄青霉素1—10μg/ml的LB液体培养基中,接入枯芽孢杆菌BF7658菌悬液,在37℃下振荡培养3—5天,经平板分离单菌落以及摇瓶发酵试验,初筛获得2213、2104和2120等α-淀粉酶高产菌株。将2213菌株的菌悬液涂布于含有4-6μg/ml氨苄青霉素的2%淀粉培养基上,复筛得到4213、4218、4237等α-淀粉酶高产菌株。4213菌株再经4次亚硝基胍诱变,未能得到蛋白酶活性低、α-淀粉酶活性高的突变株。     

以掷孢酵母作为伴生菌产生VC前体KGA的研究

以掷孢酵母作为伴生菌与氧化葡萄糖酸杆菌组成新混菌体系,通过测定生长代谢曲线,对其产酸性能和特点进行了研究。结果表明：相同条件下,新菌系产酸能力高于现有菌系,酸量增加5-7mg/ml,发酵周期缩短6-8h,酸转化率提高3-4%,最高产酸点pH值下降约0.5个单位,表现出较大的产酸潜力和可修饰性。     

用固定化细胞发酵生产己酸的研究

固定于海藻酸钙、琼脂、卡拉胶、聚丙烯酰胺凝胶,魔芋葡苷露聚糖等几种载体上的己酸菌株(Doseridium sp．WI)批次发酵表明,海藻酸钙包埋己酸菌活性最高。在最适条件下,己酸产量最高可达15mg／ml,经18批次(200余天)的批式发酵,固定化己酸菌产己酸活性稳定性较好,4℃储存二月后的固定化细胞,其发酵产己酸活性与储前基本相同。短暂的与空气接触对固定化己酸菌的活性几乎没受影响。与游离已酸菌比较,固定化细胞的己酸生成速度加快,己酸产量明显提高,单位体积内的细胞数目可高出游离培养的近10倍。     

衣康酸高产菌株的选育及发酵工艺研究

用多种诱变法对衣康酸产生菌原始菌株进行诱变,得到高产菌株16株,采用“正交试验-中心试验-正交试验”的策略优化培养基,对影响衣康酸生产的原料、水质等进行了研究。所得高产菌株生产适应性强,产酸水平为65g/100ml,残还原糖小于01g/100ml,摇床发酵周期小于96h,500L发酵罐发酵周期小于70h。     

高产吲哚乙酸及高泌氨巴西固氨螺菌的筛选与鉴定

目的：从玉米根际和土壤中分离具有高产吲哚乙酸较强的泌氨能力的巴西固氮螺菌。方法：分别通过半固体NFb培养基、CR培养基、LB培养基分离培养固氮菌株,并经过一系列菌落菌体形态特征、生理生化特性和16S rDNA序列测定等试验对其进行鉴定。结果：经分离纯化获得10株固氮菌,并鉴定均为巴西固氮螺菌（Azospirillum brasilense）,其中菌株R7在甘油半固体培养基上能分泌约14mmol/L的氨,在添加了色氨酸的培养基中能够合成58.8μg/ml的吲哚-3-乙酸（IAA）。结论：成功筛选得到一株既高产吲哚乙酸又有较强的泌氨能力的巴西固氮螺菌。     

具有差向选择性还原(R)-6-氰基-5-羟基-3-羰基己酸叔丁酯活性的微生物菌株筛选和鉴定

从土样中分离得到一株具有差向选择性还原(R)-6-氰基-5-羟基-3-羰基己酸叔丁酯活性的微生物菌株ZJB-09225,经生理生化特征鉴定和18S rDNA测序后鉴定为卡里比克毕赤酵母(Pichia caribbic ZJB-09225)。研究结果发现,在最适发酵条件下培养32 h,生物量为8.8 g/L,体积酶活达7.2 U/L;P.caribbic ZJB-09225最适作用温度、最适作用pH值分别为35℃和7.5。在最适的催化条件下,P.caribbic ZJB-09225细胞催化50.0 g/L(R)-6-氰基-5-羟基-3-羰基己酸叔丁酯3 h后,产物6-氰基-(3R,5R)-二羟基己酸叔丁酯得率3.4%,产物d.e.值99.5%以上。     

羰基还原酶不对称还原（R）-6-氰基-5-羟基-3-羰基己酸叔丁酯

选择R-羰基还原酶和葡萄糖脱氢酶双酶,协同催化（R）-6-氰基5-羟基-3-羰基己酸叔丁酯不对称还原制备阿托伐他汀关键手性合成子6-氰基-（3R,5R）-二羟基已酸叔丁酯。转化条件优化结果显示：在不添加外源性辅酶NADP（H）、菌体用量15．0g/L、147．0g/L（R）-6-氰基-5-羟基-3-羰基己酸叔丁酯、128．2g／L葡萄糖,30℃、pH6．5条件下反应6h后,底物转化率达到100％,产物d．e．值大于99．5％。     

紫外线诱变原生质体选育苏氨酸高产菌株

以黄色短杆菌T6-13的苏氨酸产生菌GSR8-25为出发菌株,利用溶菌酶制备其原生质体后进行紫外线(UV)诱变处理,从再生株中筛选出苏氨酸高产菌株。经多次连续诱变处理得到一苏氨酸高产菌株25-20,在含10%葡萄糖的培养基中摇瓶发酵60小时可积累苏氨酸19.5mg/ml。分离纯化后得菌株25-202,其苏氨酸产量可达21.5mg/ml,比出发菌株提高43.3%。     

光合细菌H3菌株的分离及其生物学特性研究

H3菌株系由盐田微生物垫中分离获得的光合细菌株。革兰氏阴性杆菌,0.5-0.7×15-2.5μm,在培养条件改变时呈多形态,其膨大细胞可达4.0-7.0μm.单根极生鞭毛,二均分裂。菌落及液体培养物呈深紫红色。菌体蛋白质含量约50%,各种必需氨基酸齐全,并含有丰富的细菌叶绿素a和类胡萝卜素。H3菌株在光照和黑暗条件下均可生长,能利用多种有机和无机碳、氮源,并能固N2.在含0.5-7.0%NaCI培养基中均可生长。但光照、盐浓度、温度、pH和通气等条件对其生长量有明显的影响。作者对其分类地位和应用潜力作了讨论。       

一株瘤胃源乳酸利用菌的分离鉴定及其体外代谢特性

【目的】从饲喂高精料的本地山羊瘤胃内分离到一株利用乳酸并能产生大量丙酸的菌株L9,并进一步研究了该菌在调控瘤胃微生物发酵中的作用。【方法】采用厌氧培养技术,结合形态、生理生化特性和16SrRNA基因序列分析结果。【结果】该菌株被鉴定为反刍兽新月形单胞菌（Selenomonas ruminantium）。该菌株体外代谢特性研究表明,L9可利用乳酸作为唯一碳源,该菌在24h内可对90mmol／L的乳酸完全降解。体外摸拟瘤胃急性酸中毒的发酵试验结果表明,以淀粉为底物时,与对照组相比,添加菌株L9可显著降低瘤胃微生物体外培养体系中乳酸浓度,提高pH值,提高总挥发性脂肪酸和丙酸浓度,并显著降低乙酸与丙酸的浓度比（P〈0．05）。【结论】结果显示,菌株L9是一株可代谢乳酸,促进丙酸生成,提高总挥发性脂肪酸浓度的有益瘤胃细菌。     

Determination of the Chemical Form of Fluorine in Tea Infusions by 19F-NMR

A novel marine ice-nucleating bacterium, KUIN-5, was isolated from a marine algae, Monostroma latissum. Strain KUIN-5 was identified as a Pseudomonas sp. from its characteristics and taxonomies; the optimum temperature and pH for its growth were 25°C and 6.0, respectively. When strain KUIN-5 was aerobically cultured in Carlucci-Pramer medium (pH 6.0) for 50 h at 25°C, the highest ice-nucleating activity of the cells among the media for marine bacteria was obtained, and the ice-nucleating temperature, T₅₀, was indicated to be ? 3.2°C. Also, the optimum concentration of NaCl for the growth in this medium, which was prepared with distilled water instead of seawater, was 2.0% (w/v) and then the ice-nucleating activity was inversely proportional to the NaCl concentration. Moreover, when strain KUIN-5 was cultured in Davis medium under optimum conditions, it produced insoluble polysaccharide (IPS) in the culture. The maximum amount of IPS production by strain KUIN-5 was 84.5 mg/ml of medium under optimum conditions. Therefore, this IPS was isolated and could be identified as cellulose, based on TLC or HPLC of the acid hydrolysate, and GC-MS of the acetylated polyalcohol prepared by periodate oxidation and Smith degradation of this polysaccharide. This is the first report of cellulose production by a marine ice-nucleating bacterium.     

Inhibition of Listeria monocytogenes and Listeria innocua by hexanoic and octanoic acids

Hexanoic acid and octanoic acid inhibited growth of 10 strains of Listeria monocytogenes and two strains of L. innocua at pH 5·0 and pH 5·5 and 20°C. Octanoic acid was more inhibitory than hexanoic acid and both were more inhibitory at pH 5·0 than at pH 5·5. The minimum inhibitory concentrations (MICs) were comparable with the concentrations of these acids that have been reported in Danish Blue cheese, where they were probably formed by the metabolism of Penicillium roquefortii . Thus hexanoic and octanoic acids may contribute to the inhibition of listerias in some cheeses.     

A mesophilic Clostridium species that produces butanol from monosaccharides and hydrogen from polysaccharides

A unique mesophilic Clostridium species strain BOH3 is obtained in this study, which is capable of fermenting monosaccharides to produce butanol and hydrolyzing polysaccharides to produce hydrogen (H(2)) and volatile fatty acids (VFAs). From 30 g/L of glucose and xylose each, batch culture BOH3 was able to produce 4.67 and 4.63 g/L of butanol. Enhancement treatments by increasing the inoculated cells improved butanol production to 7.05 and 7.41 g/L, respectively. Hydrogen production (2.47 and 1.93 mmol) was observed when cellulose and xylan (10 g/L each) were used, suggesting that strain BOH3 possesses xylanolytic and cellulolytic capabilities. These unique features reveal the strain's novelty as most wild-type solventogenic strains have not been reported to have such properties. Therefore, culture BOH3 is promising in generating butanol and hydrogen from renewable feedstock.     

极地适冷菌Pseudoalteromonas sp. QI-1产适冷蛋白酶发酵条件的优化

对极地适冷菌Pseudoalteromonas sp. QI-1产适冷蛋白酶的发酵条件进行优化。结果表明:菌株QI-1的最适生长和产酶温度均为5℃;最佳接种量为1%;发酵培养基的最适初始pH和最佳装样量分别为5和10%;盐度为2%时对菌株的生长和产酶最为有利;麸皮和醋酸钠分别为最佳N源和C源;添加0.75%酪蛋白时菌株QI-1胞外蛋白酶的活性最高;10 mmol/L Mg2+和0.5%Tween-80有利于产酶。正交试验结果表明:菌株Pseudoalteromonassp. QI-1产蛋白酶较佳培养基配方(g/L)为麸皮5,酵母粉2.5,酪蛋白3,MgCl2.6H2O 3,KCl 1.5;发酵液比酶活为166.20 U/mL,较优化前提高了约56%。     

Determination of the growth stages of Lactobacillus paracasei subsp. paracasei M3 from Bulgarian yellow cheese by electroconductivity.

The growth curve and the bacteriocin production by Lactobacillus paracasei subsp. paracasei M3-a strain isolated from Bulgarian yellow cheese, was studied using electroconductivity measurements in the culture medium (MRS). The bacteriocin produced by this bacterium inhibits strains belonging to the Candida species as well as Bacillus subtilis ATCC 6633.The use of the direct impedance technique coupled with statistical treatment of the results for the strain's growth curve allowed the detection of the growth phase-related bacteriocin production. Definition of the average particle size and aggregation estimations for the cells of the bacteriocin-producing strain was also possible. The comparison of results of impedance measurements with results obtained by classic microbiological methods and microscopic observations confirms the good correlation between these methods. In addition, the advantages of an impedance method such as rapidity, simplicity and higher sensitivity are highlighted.     

Lipids of the marine bacterium Flexibacter polymorphus

This paper reports on the total fatty acid composition of a marine bacterium representative of the genus Flexibacter. Flexibacter polymorphus is unusual in containing a high proportion of the polunsaturated acid C20:53 whilst the level of branched fatty acids is low. These facts suggest that the membrane flexibility necessary for its gliding motility is a function of the polyunsaturated fatty acid composition. Biosynthetic precursors to the C20:5 acid are present which are characteristic of an oxygen-dependent pathway. The fatty acid composition of the structural lipids is influenced by changes in the culture medium. Na₂S inhibits production of the C20:5 acid at levels much lower than that at which it is known to inhibit growth. The intracellular granules observable in F. polymorphus do not contain elemental sulphur, in contrast to Beggiatoa sps., but instead probably contain lipids.     

南极交替单胞菌R11-5产卡拉胶酶的发酵条件优化

【背景】极地寒冷环境中发现了大量具有潜在应用前景的冷适应酶,同时也存在种类繁多的海藻多糖降解菌,因此极端环境微生物是筛选获得新颖、高效多糖降解酶的重要新源泉。由于筛选培养基通常并非野生菌发酵产酶的最优条件,为了使野生菌的产酶效率达到最高,需要对其培养条件进行优化,从而为其深入研究及开发利用提供依据。【目的】对一株产卡拉胶酶的南极菌株进行种属鉴定,并采用响应面法对该菌的发酵产酶条件进行优化。【方法】通过16SrRNA基因对产卡拉胶酶的南极菌株进行种属鉴定,采用响应面法优化南极菌株产酶发酵条件。【结果】该南极菌属于交替单胞菌属(Alteromonas),命名为交替单胞菌R11-5。发酵条件优化结果显示,7个环境因子影响交替单胞菌R11-5的产酶量。利用Design-Expert软件中的Plackett-Burman设计实验,筛选出影响交替单胞菌R11-5产酶量的4个主要因素分别为培养温度、牛肉膏浓度、卡拉胶浓度和Ca~(2+)浓度。通过Box-Behnken设计和响应面分析得到交替单胞菌R11-5最佳产酶发酵条件为:温度15.0°C,牛肉膏浓度11.0 g/L,卡拉胶浓度3.0 g/L,Ca~(2+)浓度5.0 mmol/L。优化后发酵上清液酶产量达到87.193 U/mL,与优化前相比提高了1.8倍。【结论】响应面法提高了南极交替单胞菌R11-5卡拉胶酶的产量,为其开发应用提供了科学依据。     

A beta-galactoside alpha2,6-sialyltransferase produced by a marine bacterium, Photobacterium leiognathi JT-SHIZ-145, is active at pH 8

A gene encoding a sialyltransferase produced by Photobacterium leiognathi JT-SHIZ-145 was cloned, sequenced, and expressed in Escherichia coli. The sialyltransferase gene contained an open reading frame of 1494 base pairs (bp) encoding a predicted protein of 497 amino acid residues. The deduced amino acid sequence of the sialyltransferase had no significant similarity to mammalian sialyltransferases and did not contain sialyl motifs, but did show high homology to another marine bacterial sialyltransferase, a beta-galactoside alpha2,6-sialyltransferase produced by P. damselae JT0160. The acceptor substrate specificity of the new enzyme was similar to that of the alpha2,6-sialyltransferase from P. damselae JT0160, but its activity was maximal at pH 8. This property is quite different from the properties of all mammalian and bacterial sialyltransferases reported previously, which have maximal activity at acidic pH. In general, both sialosides and cytidine-5'-monophospho-N-acetylneuraminic acid, the common donor substrate of sialyltransferases, are more stable under basic conditions. Therefore, a sialyltransferase with an optimum pH in the basic range should be useful for the preparation of sialosides and the modification of glycoconjugates, such as asialo-glycoproteins and asialo-glycolipids. Thus, the sialyltransferase obtained from P. leiognathi JT-SHIZ-145 is a promising tool for the efficient production of sialosides.