1-氨基-2-丙醇的拆分

(2S,3S)-(-)-酒石酸于水中拆分消旋1-氨基-2-丙醇,然后用强酸性阳离子树脂分离,得到核苷类药物合成中间体(R)-(-)-1-氨基-2-丙醇,收率66.6%,拆分剂可直接回收利用.     

溶媒耐受性绿脓假单胞菌PseA产生耐溶媒蛋白酶的培养条件优化及产物鉴定

据报道，从源自Pseudomonas aeruginosa的溶媒耐受性绿脓假单胞菌株（PseA）中可获得耐溶媒蛋白酶。本研究所获得产生该酶的优化培养条件：以0．7％甘油为碳源，0．4％酪蛋白和0．6％酵母抽提物作为氮源，pH7．0；种子液培养36h后以2．5％的接种量接种；30℃振荡（250r／min）培养24h。在该条件下，菌体生长最好，酶活力最高（160lu／ml）。该蛋白酶的最适pH与温度为9．0和55℃。     

186株铜绿假单胞菌β-内酰胺酶检测及药敏分析

目的:了解铜绿假单胞菌对抗生素的耐药情况,为临床治疗提供依据。方法:收集临床不同感染部位标本中分离到的186株铜绿假单胞菌进行β-内酰胺酶的检测及药敏测定结果分析。结果:186株铜绿假单胞菌对所测试的11种抗生素耐药5种以上的有72株,占38.7%,其中β-内酰胺酶阳性的60株,占32.4%,β-内酰胺酶阴性的有12株,占6.3%。结论:产β-内酰胺酶的铜绿假单胞菌的耐药性更加严重,目前亚胺培南是治疗铜绿假单胞菌感染的首选药物,其次是丁胺卡那和头孢他啶。     

β-内酰胺类抗生素对铜绿假单胞菌耐药性的影响

目的研究派拉西林/他唑巴坦、头孢他啶、头孢噻肟、头孢曲松及亚安培南对铜绿假单胞菌耐药性的影响。方法以对临床分离的34株铜绿假单胞菌为研究对象,分别用肉汤渐进稀释法进行诱导,K-B法筛选,E-TEST法检测菌株的MIC。结果经头孢曲松诱导后,铜绿假单胞菌抗药敏感性下降,甚至出现耐药。其余四种β-内酰胺类抗生素对铜绿假单胞菌抗药敏感性影响不明显。结论各类β-内酰胺类抗生素对铜绿假单胞菌耐药性的影响不同。在治疗由铜绿假单胞菌引起的感染时,应根据药敏试验选择具有高抗菌活性、低选择能力的抗生素。     

352株铜绿假单胞菌的耐药性分析

目的分析铜绿假单胞菌的临床分布特点和耐药性,为临床合理用药提供指导。方法采用回顾性分析方法,352株铜绿假单胞菌均由检验科分离得到。细菌鉴定采用法国生物梅里埃公司VITEK-32系统,药敏试验采用K-B纸片法。结果 352株铜绿假单胞菌以痰（78.4%）和尿液（9.9%）培养为主,科室分布以外科（44.6%）、内科（23.3%）、儿科（17.6%）和妇科（14.5%）为主。铜绿假单胞菌对哌拉西林、头孢曲松、头孢唑啉、头孢哌酮、氨曲南、环丙沙星和左氧氟沙星的耐药性较高,对头孢他啶、头孢吡肟、庆大霉素、阿米卡星、亚胺培南、美罗培南、头孢哌酮/舒巴坦和哌拉西林/他唑巴坦耐药性较低。结论铜绿假单胞菌以呼吸道痰标本培养为主,在多科室都常见,临床耐药性极高。应控制抗生素药物的临床合理应用、加强动态监测,对治疗和预防感染具有重要指导意义。     

248例铜绿假单胞菌的分布及耐药分析

目的研究铜绿假单胞菌(PAE)感染现状及耐用性,指导临床合理用药。方法收集我院2011年1月至12月临床各种标本中分离的248株PAE,用VITEK 2 COMPACT进行鉴定及药敏分析。结果 248株铜绿假单胞菌对20种抗菌药物体外敏感试验结果显示:头孢唑林耐药率最高为100%,氨苄西林、呋喃妥因、头孢替坦、头孢呋辛钠、氨苄西林/舒巴坦耐药率也高达97%以上,其他阿米卡星、左氧氟沙星、哌拉西林/他唑巴坦耐药率分别为13.51%、13.95%、14.38%,亚胺培南的耐药率达18.47%,而美罗培南耐药率最低12.5%。结论铜绿假单胞菌耐药已非常严重,不断出现对几乎所有抗生素全部耐药的泛耐药菌株,这给临床治疗、院感控制带来了极大的困难。应加强对铜绿假单胞的耐药性监测,减少耐药菌的产生。     

铜绿假单胞菌的多重耐药机制及抗生素应用

铜绿假单胞菌（Pseudomonas aeruginosa，Pa）为条件致病菌，是医院感染的主要病原菌之一。2002年度全国细菌耐药性监测网所属57家三级甲等医院调查发现，Pa临床分离率为10．3％，仅次于大肠埃希菌的14．2％列第2位。Pa具有多重耐药特性，能天然抵抗多种抗生素。Kolar报道Pa多重耐药株占其临床分离菌株的4．0％～26．3％。常在治疗过程中通过突变发生耐药。     

铜绿假单胞菌耐药机制的分析

目的了解铜绿假单胞菌对常用抗菌药物的耐药机制。方法采用纸片K-B琼脂扩散法,参照CLSI2011年版标准判读结果。结果 154株铜绿假单胞菌中来自呼吸内科的52株,内分泌科6株,康复科46株,干部科11株,外科16株,儿科7株,ICU10株,五官科6株。可见,铜绿假单胞菌主要分布于呼吸内科,其次是康复科的慢性病患者。铜绿假单胞菌对米诺环素、头孢曲松、头孢唑肟耐药率达到50%以上,对氟喹诺酮类、氨基糖苷类均低于20%,对碳青霉烯类的美洛培南耐药率达到了7.8%。结论细菌产生头孢菌素酶(Ampc酶)是引起头孢类药物耐药的主要机制;产金属B-内酰胺酶是引起碳青霉烯类抗生素耐药的原因;拓扑异构酶的突变,改变了三元复合物的亲和力,抑制了氟喹诺酮的活性而产生耐药。     

226株铜绿假单胞菌的耐药性分析

目的了解铜绿假单胞菌的临床分离情况及耐药现状,指导临床合理用药。方法采用美国临床标准化研究所(CLSI)推荐的标准纸片扩散法测定226株铜绿假单胞菌对头孢他啶、阿米卡星和亚胺培南等16种抗菌药物的体外敏感性。结果铜绿假单胞菌感染以下呼吸道、泌尿道为主,耐药率最低的是多黏菌素B(1.3%)其次是头孢吡肟(8.4%)、头孢哌酮/舒巴坦(8.5%)。结论铜绿假单胞对多种抗生素呈现不用程度的耐药,临床医生应根据实验室的体外药敏结果合理选择抗生素。     

抗MRSA链霉菌NJ0510的抑菌活性及发酵影响因素的研究

从土壤中分离到一株有效抑制耐甲氧西林金黄色葡萄球菌(MRSA)的链霉菌NJ0510。考察了抗菌谱及各种发酵影响因素对链霉菌NJ0510抑菌活性物质产生的影响,通过正交实验对培养基中碳、氮源的组成及发酵天数进行了优化。实验表明该菌的最适培养条件为:培养40h的种子培养液以10%的接种量接入到含1%葡萄糖、2%糊精和4%蛋白胨,40mg/LFeCl3、20mg/LCuSO4.5H2O和1mg/LCoCl2.6H2O的发酵培养基中,在200r/min振荡条件下,培养5d。     

头孢霉AL031菌株液体培养工艺的研究

研究产生异香豆素类化合物的头孢霉（Cephalosporium sp.)AL031菌株的培养基和摇瓶培养条件。并通过正交试验进行优化。结果表明，其适宜的培养基组成（％）：马铃薯（煮汁）20，葡萄糖和蔗糖（1：2）2．0，KH2PO40．1，MgSO4.7H2O 0.05;Vitamin B10.01,摇瓶培养条件为：培养基的起始pH为7，500ml三角瓶装量200ml，接种量10％，24℃振汇培养4d，细胞生物量为10．8mg/ml发酵液。     

黄嘌呤氧化酶产生菌的产酶条件优化

通过恒化富集培养分离得到一株黄嘌呤氧化酶（XOD）产量较高的节杆菌Arthrobacter sp．。采用响应面分析法优化培养基组成，XOD比活力较初始水平提高了1．9倍。利用正交试验研究摇瓶发酵条件，结果发现，当培养温度为32℃、培养基初始pH为8．0、转速为230r/min时，产酶水平又提高50％，XOD比活力达0．083u（mg蛋白）^-1。在此基础上，绘制菌株产酶曲线，确定最佳产酶时间为45h。     

北极放线菌Streptomyces sp. MLA-21发酵条件优化

对北极放线菌Streptomyces sp.MLA-21的发酵条件进行优化,提高发酵液中抗肿瘤活性部位的产量。以发酵液对肺癌A549细胞的抑制率（IR）作为观测指标,利用响应面分析法对通过单因素实验和Plackett-Burman设计筛选出培养基初始pH值、黄豆浸出物含量,可溶性淀粉含量等关键因素进行优化。建立二次回归模型。得到菌株MLA-21发酵的最优条件为KNO₃1.0 g/L,可溶性淀粉23.04g/L,黄豆浸出物7.1g/L,K₂HPO₄ 0.5g/L,MgSO₄ 0.5g/L,NaCl 0.5g/L,pH 7.67,接种量:7.5%,培养温度:28℃,摇床转速:150 r/min。对优化结果进行验证发现优化后的培养基能够显著提高发酵液对A549细胞的抑制率和活性部位的产量。     

Isolation and identification of antifungal N-butylbenzenesulphonamide produced by Pseudomonas sp. AB2

An antifungal bacterial strain, isolated from a greenhouse soil sample, inhibits growth of microflora nearby. It was selected for further studies of bacterial antifungal properties. This isolate was identified as a Pseudomonas sp. based on carbohydrate utilization, and other biochemical and physiological tests. Petri plate assay revealed that the Pseudomonas sp. exhibited antifungal activity against the plant pathogens, Pythium ultimum, Rhizoctonia solani, Phytophthora capsici, Botrytis cinerea and Fusarium oxysporum. Using direct inhibition bioassay on TLC plates after ethyl acetate extraction of the culture filtrate, we correlated antifungal activity with production of antifungal compounds. An antifungal antibiotic was isolated from the culture filtrate and was identified as N-butylbenzenesulphonamide. ED50, values of the N-butylbenzenesulphonamide against P. ultimum, P. capsici, R. solani, and B. cinerea were 73, 41, 33 and 102 ppm, respectively.     

假单胞菌发酵制备精氨酸脱亚胺酶

采用假单胞菌（Pseudomonas sp．）发酵制备精氨酸脱亚胺酶，优化发酵培养基配方及培养条件，以葡萄糖为碳源，酵母膏和蛋白胨为氮源，30℃振荡培养20h时，发酵液中精氨酸脱亚胺酶的酶活力可达2．17u／ml。向HepG2肝癌细胞培养液中加入酶粗品0．5u／ml，对肿瘤细胞的生长抑制率为93．4％。     

A comparison of the binding characteristics of recombinant P2X1 and P2X2 purinoceptors.

1. We have recently provided evidence that [35S]-adenosine 5'-O-[3-thiotriphosphate] ([35S]-ATP gamma S) can label the human bladder recombinant P2X1 purinoceptor (human P2X1 purinoceptor). In this study we have characterized the binding of [35S]-ATP gamma S to a second P2X purinoceptor subtype, the rat PC12 phaeochromocytoma cell recombinant P2X2 purinoceptor (rat P2X2 purinoceptor), and compared its binding properties with those of both endogenous and recombinant P2X1 purinoceptors. 2. Infection of CHO-K1 cells with the rat P2X2 purinoceptor using Semliki forest virus (SFV) resulted in the expression of high affinity (pKd = 9.3; Bmax = 18.1 pmol mg-1 protein) binding sites for [35S]-ATP gamma S but not for [3H]-alpha, beta-methylene ATP ([3H]-alpha beta meATP). Since functional P2X purinoceptors could be detected electrophysiologically in these cells, but not in non-infected or CHO-K1 cells infected with SFV containing the LacZ gene, these results suggest that the rat P2X2 purinoceptor can be labelled using [35S]-ATP gamma S. 3. The binding characteristics of the rat P2X2 purinoceptor were compared with those of the human P2X1 purinoceptor, which was also expressed in the CHO-K1 cells using SFV. A major difference between the two recombinant P2X purinoceptor types was in the binding characteristics of alpha, beta-methylene ATP (alpha beta meATP). Thus, in the absence of divalent cations, alpha beta meATP possessed low affinity for both the human P2X1 purinoceptor (pIC50 = 7.2) and rat P2X2 purinoceptor (pIC50 = 7.1) labelled using [35S]-ATP gamma S. However, when the recombinant P2X purinoceptors were labelled with [3H]-alpha beta meATP in the presence of 4 mM CaCl2, the affinity of alpha beta meATP for the human P2X1 purinoceptor increased (pIC50 for alpha beta meATP = 8.2), while the affinity of the rat P2X2 purinoceptor for alpha beta meATP did not change (pIC50 for alpha beta meATP = 6.8). 4. Affinity estimates of 15 other nucleotide analogues for the [35S]-ATP gamma S binding sites on the two recombinant P2X purinoceptor subtypes were surprisingly similar (less than 5 fold difference), the only exception being 2'-deoxy ATP which possessed 8 fold higher affinity for rat P2X2 than for human P2X1 purinoceptors. In contrast dextran sulphate and the P2 purinoceptor antagonists, pyridoxalphosphate-6-azophenyl-2', 4'-disulphonic acid and 4,4'-diisothiocyanatostilbene-2,2' disulphonic acid, possessed 7 to 33 fold higher affinity for the human P2X1 than for the rat P2X2 purinoceptor. These data provide a correlation coefficient (r) of 0.894. 5. There was some evidence for species differences in the P2X1 purinoceptor. Thus, most nucleotides possessed slightly greater (up to 9-10 fold), while the P2 purinoceptor antagonists possessed slightly lower (up to 7-16 fold), affinity for the endogenous rat vas deferens and rat bladder P2X1 purinoceptors than for the human recombinant P2X1 purinoceptor. These differences were reflected in a slightly lower correlation coefficient, when comparing across species between the human recombinant P2X1 purinoceptor and the endogenous P2X1 purinoceptors labelled in either the rat deferens (r = 0.915) or the rat bladder (r = 0.932), than when comparing within species between the endogenous rat vas deferens and rat bladder P2X1 purinoceptors (r = 0.995). 6. In summary, [35S]-ATP gamma S can be used to label the recombinant P2X1 and P2X2 purinoceptors. Despite the marked differences reported between these two forms of P2X purinoceptor in functional studies, the differences in binding studies were more limited. However, a number of antagonists could discriminate between the P2X purinoceptor subtypes in the binding studies raising expectations that selective antagonists for these receptors can be developed.     

假单胞菌产壳聚糖酶突变菌株的筛选

以假单胞菌Y1为出发菌株 ,分别通过亚硝基胍 (NTG) ,Co6 0 ,UV(紫外 )诱变 ,采用透明圈法筛选 ,获得了产壳聚糖酶较好的突变菌株假单胞菌Y8,其所产酶活力达到 3.0U·ml- 1,酶活力提高近 6倍 ,并具有较好的遗传稳定性。 3种诱变方法中 ,UV诱变效果最好。     

重组人源白介素-1β发酵条件的优化及其产品的纯化

目的初步建立重组人源IL-1β巴斯德毕赤酵母的培养体系;进一步优化大规模培养重组人源IL-1β巴斯德毕赤酵母的条件;探讨大规模纯化重组人源IL-1β巴斯德毕赤酵母表达产物的条件;鉴定酵母表达产物及纯化样品。方法在菌体湿重达190g.L-1左右时开始甲醇诱导表达;选用了SP Sepharose XL阳离子交换层析和反相疏水柱层析进行纯化。结果在pH4.5的含0.5%蛋白胨的FM21培养基中发酵时,放罐时间在30h较为适宜;经过纯化蛋白纯度即可达95%以上。结论最终经过鉴定,表达和纯化的样品就是我们想要得到的IL-1β。     

在大肠杆菌中表达重组CotA蛋白的培养条件优化

为了提高大肠杆菌对重组CotA蛋白的表达量,对本实验室构建的含CotA基因表达载体的工程菌株的培养条件进行优化。通过单因素试验及正交试验L18（3 7）优化了产酶的最佳培养条件为：摇床转速为180r／min、装液量为100mL／500mL锥形瓶、接种量为5％、诱导温度25℃、诱导剂IPTG浓度为1．0mmol／L、诱导时间为12h,以及加诱导剂时菌体OD600值为1．0,其中诱导剂IPTG浓度和菌体OD600值对CotA蛋白表达影响显著。