Candida Cloacae长链脂肪酸醇氧化酶的结构域谱分析

目的分析酵母Candida Cloacae中长链脂肪酸醇氧化酶的酶活性结构域.方法利用RT-PCR技术,从酵母Candida cloacae细胞中克隆长链脂肪酸醇氧化酶的5个不同长度的cDNA片段.扩增这些片段的上游引物5'端引入了NheI限制性内切酶位点和起始密码ATG,下游引物5'端引入了NheI限制性内切酶位点.扩增产物经NheI消化后,被克隆进T7启动子控制下的表达质粒pET-17b中,转化大肠杆菌BL21(DE3)菌株.结果转化大肠杆菌BL21(DE3)菌株表达出5个不同长度的长链脂肪酸醇氧化酶的羧基端肽段,其氨基酸残基数分别为535AA(50KD)、473AA(43KD)、411AA(38KD)、306AA(26KD)和216AA(18KD).与完整的长链脂肪酸醇氧化酶(697AA)相比较,相当于它们分别从氨基端被切除162、224、288、391和481个氨基酸,其酶活性分别下降了18.8%、20.6%、88.6%、91.4%和99.7%.结论结果表明随着长链脂肪酸醇氧化酶的氨基端被切除的氨基酸数增加,其酶活性下降,尤其是切除224-288氨基酸序时,酶活性下降更明显,推测此肽段对长链脂肪酸醇氧化酶酶活性起重要作用.     

人肝谷胱甘肽过氧化物酶的动力学性质

从人肝中分离获得谷胱甘肽过氧化物酶，其最适pH为8．5，最适温度为37℃。该酶以H2O2或GSH为鹿物时表观Km值分别为0．27mmol／L和1．3mmol／L。酶与多分子鹿物作用的动力学研究表明人肝谷胱甘肽过氧化物酶的作用为乒乓机制。并观察了一些金属离子和烷化剂对该酶具有不同程度的抑制作用。     

SHV-4型ESBL在pET26b/BL(DE3)中的表达纯化及其特性研究

目的:表达SHV-4型超广谱β-内酰胺酶,对纯化的SHY-4酶进行特性研究.方法:将SHV-4基因克隆到pET26b表达质粒,重组质粒转入E.coli BL21(DE3)构建工程菌,重组工程菌经摇瓶发酵后获得种子菌,种子菌转入发酵罐发酵,当菌体密度OD600为0.8时,加入诱导剂IPTG至终浓度为1.0 mmol/L,于37℃培养6 h.培养液离心收获菌体沉淀,其超声裂解上清液进行Ni2 亲和层析,获取的SHV-4酶进行分子量、纯度、等电点、最适条件、米氏常数、催化常数等特性进行分析.结果:SDS-PAGE电泳,等电聚焦电泳及系列的酶动力学实验表明SHV-4酶分子量约为30 kD,纯度大于95%,等电点为7.9,最适pH值为6.0～8.0,最适温度为37℃,以头孢硝噻吩为底物测定的Km值为2.61×10-6 mol/L,Kcat为258 s,Kcat/Km为9.88×107 mol-1·s-1·vL.结论:重组的SHV-4酶初步满足制成商品酶的条件.     

甲壳糖固定血管紧张素转换酶的制备及其性质

目的 将血管紧张素转换酶(ACE)固定化并测定其动力学性质。方法 以甲壳糖为载体，碳二亚胺(EDC)为交联剂，与分离纯化的ACE混合,使ACE固定化；并以马尿酰甘氯酰甘氨酸(HCG)为底物，分别测定了其游离酶和固定化酶的动力学性质。结果 固定化酶活力110u／g湿重，固定化率为22％，与游离酶比较，固定化酶稳定且具良好的耐热性，在50—80℃固定化酶较游离酶稳定。在pH7．1的底物缓冲体系中，37℃，游离酶Km=0.026mol／L；在间歇振摇下固定化酶的表现Km=0．040mol／L。游离酶最适pH为、8.0固定化酶最适pH为9．0。结论 用甲壳糖固定ACE方法简便可行，且固定后的甲壳糖性能稳定，有广阔的应用前景。     

人肝谷胱甘肽过氧化物酶的动力学性质

从人肝中分离获得谷胱甘肽过氧化物酶，其最适ｐＨ为８．５，最适温度为３７℃。该酶以Ｈ２Ｏ２或ＧＳＨ为底物时表现Ｋｍ值分别为０．２７ｍｍｏｌ／Ｌ和１．３ｍｍｏｌ／Ｌ。酶与多分子底物作用的动力学研究表明人肝谷胱甘肽过氧化物酶的作用为乒乓机制。并观察了一些金属离子和烷化剂对该酶具有不同程度的抑制作用。     

篱栏网化学成分研究

从篱栏网的全草中分离鉴定了二十六碳酸长链脂肪醇酯系列，三十烷醇和三十二烷醇的混合物，Ｃ２０－Ｃ３０的长链脂肪酸系列以及Δ＾７－豆甾烯醇，皆为首次从该植物中分得。     

长链脂肪酸醇氧化酶基因在油菜不同组织中的表达

目的：研究长链脂肪酸醇氧化酶基因在油菜不同组织中的表达水平。方法利用RT-PCR方法从油菜RNA中扩增长链脂肪酸醇氧化酶基因的cDNA。并用[α-^32P]dCTP标记该cDNA作为探针。应用Northern blot杂交技术分析长链脂肪酸醇氧化酶基因在油菜根、茎、叶及花等组织中的表达水平。结果：Northern blot杂交所检测的油菜根、茎、叶及花组织中均有位于2．4kb处的杂交带，但表达水平有差异，在油菜叶中表达量最高。结论：长链脂肪酸醇氧化酶基因表达于所有被检测的油菜组织中，可能为一种组成性表达的基因。     

C型产气荚膜梭菌重组菌株B121（DE3）pXETAB2B1表达条件的优化

目的优化C型产气荚膜梭菌重组菌株B1221（DE3）pXETAB2B1融合蛋白表达的条件。方法以IPTG（异丙基硫代-β—D-半乳糖苷）为诱导剂，通过聚丙烯酰胺凝胶电泳比较pH值、培养温度、诱导时间、菌体生长密度等因素对重组α-β2-β1融合蛋白表达进行表达条件的影响。结果C型产气荚膜梭菌重组菌株BL21（DE3）pXETAB2B1的表达优化条件是：最适培养基pH值是7．5，最适诱导时间是5小时，最适表达温度是37℃，最适菌体生长密度是OD6001．0，IPTG最适诱导浓度是0．4mmol／L。结论获得C型产气荚膜梭菌重组菌株BL21（DE3）pXETAB2B1融合蛋白的最佳表达条件。     

SMB-1型金属β-内酰胺酶的原核表达及酶活性研究

目的原核表达SMB-1型金属β-内酰胺酶,并研究其水解β-内酰胺类抗生素的活性。方法利用化学合成方法合成blaSMB-1基因,经PCR扩增后克隆至pET28a载体,构建重组的原核表达质粒pET28a-SMB-1,转化至E.coli BL21(DE3),诱导表达。表达的重组蛋白经Ni2+-NTA纯化后,测定其水解底物时的酶促反应动力学参数及最适pH值和温度。结果重组质粒经测序证明其正确性;表达的重组蛋白相对分子质量约为33 000,纯化后蛋白的质量浓度为0.20 mg/mL,该酶对本实验中除氨曲南外的所有β-内酰胺类抗生素均具有较高水解活性,其最适pH值为8.0,最适温度为40℃。结论利用工程菌E.coliBL21(DE3)-pET28a-SMB-1成功实现了SMB-1型金属β-内酰胺酶的原核表达,为深入研究其生物学功能及作用机制提供了物质基础,同时也为研发该酶的抑制剂提供了靶标物质。     

壳聚糖的酶法降解

用壳聚糖酶降解壳聚糖,探讨了壳聚糖降解过程中温度、pH值、底物浓度和金属离子对酶促反应的影响。结果表明:酶促反应进行到5 h左右时,即可得到聚合度小于10的壳寡糖。该酶促反应的最适温度为50℃,最适pH=5.5;最适底物浓度为0.02 g/mL;金属离子Ca2+和Mg2+对酶降解有一定的促进作用,而Zn2+、Cu2+对酶降解有较强的抑制作用。该酶促反应符合米氏动力学方程,米氏常数Km=7.80 g/L,最大反应速率Vmax=7.72 g/(min.L)。     

APPLICATION OF LORNOXICAM TO PATIENT-CONTROLLED ANALGESIA IN PATIENTS UNDERGOING ABDOMINAL SURGERIES

FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…     

Dr.Zhang Ren''''s Experience in the Acupuncture Treatment of Different Diseases with the Same Therapeutic Principle

Dr.Zhang Ren,the chief physician,is the chairman of Shanghai Acupuncture and Moxibustion Association.Having been engaged in medicine for about 40 years,he is experienced in treating various intractable diseases.In his long years of clinical practice,he advocates taking the TCM differentiation as the basis to seek for the acupuncture method for treatment of modern intractable diseases.The author of this essay had the fortune to follow Dr.Zhang in study.The following is a summary of Dr.Zhang's experience in the acupuncture treatment for different intractable diseases with the same therapeutic principle.     

Luo Lingjie''''s Experience in Treating Chronic Nephropathy

In treating chronic nephropathy,Luo Lingjie,a chief physician,pays attention to regulating the balance between yin and yang,treating infection if present,and removing pathogenic factors.He prescribes gentle drugs and uses carefully strongly warming-tonifying ones,emphasizes the importance of persuading the patient to persist in treatment with medication and nurse one's health for recuperation,and is good at combined use of TCM and western medicine therapy and brings the merits of various therapies into full play,with obvious theraoeutic effects.     

Acupuncture Combined with Spinal Tui Na for Treatment of Primary Dysmenorrhea in 30 Cases

Objective: To observe the therapeutic effects in acupunture treatment of primary dysmenorrhea combined with spinal Tui Na, and study its mechanism. Methods: Thirty cases of the treatment group were treated by acupuncture combined with spinal Tui Na, and thirty cases in the control group were treated by routine acupuncture. Results: The total effective rate was 93.3% in the treatment group, and 73.3% in the control group, with a significant difference between the two groups (P<0.05). Conclusions: Acupuncture combined with spinal Tui Na has good prospects for treatment of primary dysmenorrhea.     

猪肺磷脂注射液联合经鼻持续气道正压通气对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性的影响

目的 探讨猪肺磷脂注射液联合经鼻持续气道正压通气(NCPAP)对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性(CK-MB)的影响.方法 选取呼吸衰竭早产儿80例,分为观察组和对照组各40例.对照组采用NCPAP给氧治疗,观察组给予NCPAP给氧联合猪肺磷脂气管内给药.观察两组患儿治疗前及治疗12h、24 h后PaO2、PaCO2、血氧饱和度(SaO2)、pH的变化情况,检测治疗前及治疗5d后血清CK-MB水平;评估两组患儿的临床治疗效果.结果 两组患儿PaO2、PaCO2、SaO2、pH比较,差异均有统计学意义(P＜0.05),其中观察组治疗后的PaO2、SaO2、pH均高于对照组,PaCO2则低于对照组.两组的PaO2、SaO2、pH均随观察时间延长而升高(P＜0.05),PaCO2均随观察时间的延长而降低(P＜0.05).观察组治疗有效率为87.5％,显著高于对照组的70.0％ (P ＜0.05).治疗5d后两组患儿血清CK-MB水平均较前降低(P＜0.05),且观察组明显低于对照组(P＜0.05).结论 猪肺磷脂注射液气管内给药联合NCPAP可以显著降低呼吸衰竭早产儿CK-MB的含量,提高治疗有效率,起到很好的呼吸循环支持作用.     

Survey and practice of reporting quality of randomized controlled clinical trials on traditional Chinese medicine

Evidence obtained from randomized controlled trials （RCTs） has been generally accepted as the gold standard in the evaluation of clinical effectiveness. Readers need to understand the trial design, implementation, results, analysis and interpretation, so as to fully Jnderstand the results of RCTs. Thus, the investigators of RCTs have to report these items in a complete, accurate and clear manner. Since 1998, we have conducted several evaluations on the reporting quality of RCTs published in Chinese journals on traditional Chinese medicine （TCM） and results have shown that there is an urgent need for higher quality RCTs on TCM.     

TCM Treatment of Ankylosing Spondylitis by Tonifying the Kidney and Strengthening the Governor Vessel

Ankylosing spondylitis is a chronic and progressive disorder with inflammation mainly involving the central axis joints. It mainly affects the cervical spine and the lumbosacral area, with the pathogenesis closely related to the kidney and the Governor Vessel (GV). TCM holds that the syndrome is deficiency in origin and excess in superficiality, which is due to insufficiency of the kidney, deficiency of GV, and blocking of the channels with the invasion of exogenous evil, leading to poor circulation of qi and blood and malnutrition of the bones, muscles and joints. The TCM method of tonifying the kidney and strengthening GV to regulate circulation of qi and blood and check the arthralgia pain should be adopted, with the Kidney-Tonifying and GV Strengthening Decoction (益肾强督汤) prescribed.     

IMPROVING P-gp EXPRESSION IN HUMAN MONONUCLEAR CELLS IN VITRO TRANSFECTED BY MULTIDRUG RESISTANCE-1 mRNA

CHEMOTHERAPY playsa greatrolein the treat- ment of malignanttumors,especiallyingynecolo- gicalones.But inanticancerchemotherapy,leuko-cytopeniaisfrequentlytheprimarydose-limitingsideeffect factor.Moreover,cancersarefrequentlychemoresistantbe-causeof overexpressionof P-glycoprotein(P-gp), which isencodedby multidrugresistancegene (MDR1 ) and detectableinup to50% ofhuman cancersand renderscellsresistancetoanticancerdrugs.The safetyand potentialtherapeuticbenefitof mdr1 gene transferredto h…