制备高活力胰酶的安全低污染新工艺

目的建立安全、低污染制备高活力胰酶的方法。方法胰脏自溶水提取,以一定浓度的蛋白质沉淀剂P60在最佳筛选条件下沉淀6h,室温干燥得胰酶粉,对上清液中的P60进行回收再利用。结果收率达15%以上,胰蛋白酶、胰脂肪酶、胰淀粉胰三酶活力分别为3.70×103、2.80×104、5.30×104U·g-1,为《中国药典》要求的6～8倍。结论所建立的方法能安全简便的制备新型高活力胰酶。     

CM-纤维素固定化脂肪酶的研究

目的 　研究 CM-纤维素为载体的固定化脂肪酶的制备方法及比较固定化脂肪酶与游离酶在酶学特性上的变化。方法 　制备固定化脂肪酶的最适条件是 p H5 .5 ,离子浓度为 0 .0 1mol· L- 1 的柠檬酸缓冲液 ,加酶量为 10 0μ· g- 1 载体 ,固定化反应时间为 8h。结果 　所得的固定化酶酶活力为 49.6μ·g- 1载体 ,回收率为 45 % ,最适作用 p H值为 10 .6,最适作用温度为 46℃ ,其催化油脂水解操作半衰期为 2 2 5 h。结论 　本固定化酶制备简便 ,可重复使用 ,稳定性较高。     

亲和层析提取抑肽酶的研究

用4％琼脂糖,在碱性条件下,用双功能试剂β-硫酸醋乙砜基苯胺(SESA)连接于琼脂糖上,制备得到对氨基苯磺酰乙基(ABSE)琼脂糖。然后经重氮化制备得到固定化胰蛋白酶,装柱亲和层析分离抑肽酶,连续使10次,亲和效率维持不变,取得了令人满意的结果。和原工艺分级沉淀法比较,生产周期可从原来的15天缩短到2天,产品收得率从原来的15％提高到90％左右,比活从原来的350单位/毫克提高到1200单位/毫克。本文还研究了不同载体和方法制得的固定化胰蛋白酶的比较,发现对氨基苯磺酰乙基琼脂糖制备的固定化胰蛋白酶;活力回收高、非专一吸附小,分离效果好等优点。经推测SESA和琼脂糖连接,使酶分子和大分子载体之间形成了一定的长度,从而减少了大分子载体造成的空间障碍,使酶和抑制剂的亲和力更大。实践证明:亲和层析提取纯化生物物质不失为一种理想的好方法。     

固定化酵母不对称水解布洛芬乙酯制备S（＋）—布洛芬

用多种固定化载体对含有不对称水解布洛芬酯的脂肪酶的丝孢酵母进行了固定化，海藻酸钙，卡拉胶，聚丙烯酰胺凝胶和ＰＶＡ固定化酵母后，酶活力回收分别为６％，１６％，１８％和７％用自制的高脱乙酰度，高分子量的壳聚糖，通过成囊装置地母细胞，并用戊二醛交联，制得直径１．０ｍｍ左右的固定化酵母细胞壳聚糖多孔微球，酶活力回收达８０％以上，操作半衰期７５天以上，而且每次反应所产生的布洛芬的比旋光度均为（α）＾２０ｂ＝     

固定化超氧化物歧化酶的制备

目的 研究固定化超氧化物歧化酶(SOD)的制备方法。方法 分别以不同方法对铜锌超氧化物歧化酶(CuZnSOD)进行固定并比较其活力,对固定化方法进行相应的考察和优化。结果 以壳聚糖为载体,戊二醛交联法制备固定化SOD酶,酶活力和酶活回收率均较理想,固定化过程中的pH在6.0-8.2范围内,对固定化效果无明显影响;优化条件下制备的固定化铜锌超氧化物歧化酶,所得壳聚糖酶粉活力可达141 U·g-1,酶活回收率大于60％。结论 壳聚糖-戊二醛交联法可用于固定化超氧化物歧化酶粉的制备。     

利用固定化酯酶制备(R)-氟比洛芬

本研究利用固定化重组酯酶由氟比洛芬乙酯拆分制备（R）-氟比洛芬。将基因工程菌经高压破壁的裂解液直接固定到含Ni^2＋的载体上，所得固定化酯酶的比活力为340u／g。反应液分离纯化后（R）-氟比洛芬的光学纯度达99．6％，收率为30．7％。     

从生产胰岛素的胰渣中制备胰酶、胰蛋白酶及糜蛋白酶

目的 从中性乙醇提取胰岛素后的残渣中提取制备高活性的胰酶,并从中分离纯化出胰蛋白酶(T)、糜蛋白酶(CT).方法 在胰渣中加入活化剂活化后,用PEG-6000沉淀、丙酮脱脂干燥得到胰酶;胰酶经CM-纤维素纯化得到胰糜蛋白酶混合制剂,再经CM-sepharose FF层析分离得胰蛋白酶和糜蛋白酶.结果 胰酶回收率为12.1％,胰蛋白酶、胰脂肪酶、胰淀粉酶的活性分别为5.40、39.72、76.72 U·mg-1,3种酶的活性比例达1∶7.4∶14.2.胰蛋白酶、糜蛋白酶的效价分别为2505、1003 U·mg-1;相对于胰酶,二者的活性回收分别为48.93％、61.73％.结论 从胰渣中制得的胰酶活性较高(约9倍于《中国药典》2010年版中的标准);制得的T及CT的活力也达到《中国药典》2010年版中的标准.     

NTE酯酶活力域真核表达载体的构建及其表达

目的构建人神经病变靶标酯酶（NTE）酯酶活力域（NESP）真核表达载体，并在细胞中表达。方法利用含有特定酶切位点的引物，通过PCR扩增出其酯酶活力域，经T载体克隆测序正确后，双酶切回收cDNA片段定向插入到pcDNA3．1（＋）中，通过酶切鉴定其真核表达载体的构建。然后将其转染到真核细胞中，通过NTE活力测定，检测其表达效果。结果经PCR获得了NTE的1．6kb酯酶活力域的cDNA片段，T载体克隆后测序证实完全正确，然后克隆到真核表达载体中获得了NTE酯酶活力域真核表达载体pcDNA—NEST。瞬时转染到COS7和SH-SY5Y细胞中，NTE的活力与对照细胞相比显著提高。结论成功构建了NEST真核表达载体，并在哺乳动物细胞中表达。     

牛胰蛋白酶制备工艺的改进及其对基因工程人胰岛素前体的酶切作用

目的对牛胰蛋白酶制备工艺进行改进,制备高纯度牛胰蛋白酶,并对其酶切基因工程人胰岛素前体的作用进行考察。方法采用粗制后先色谱分离再活化的方法对传统工艺进行改进,对传统工艺制备样品和改进工艺制备样品中的胰蛋白酶和胰凝乳蛋白酶的比活力进行测定和比较;进一步采用这2种样品对基因工程人胰岛素前体进行酶切,对酶切作用进行比较。结果传统工艺制备样品中胰蛋白酶比活力为212.5 U·mg-1,胰凝乳蛋白酶比活力为20.4 U·mg-1;而改进工艺样品中胰蛋白酶比活力为240.4 U·mg-1,胰凝乳蛋白酶比活力为0.14 U·mg-1。传统工艺制备样品酶切基因工程人胰岛素前体目标产物纯度质量分数为16.3%,改进工艺制备样品酶切基因工程人胰岛素前体目标产物纯度质量分数为32.2%。结论粗制后先进行SP色谱可实现胰蛋白酶原和胰凝乳蛋白酶原较好的分离,该方法适合于高纯度胰蛋白酶的制备,制备的胰蛋白酶适合作为工具酶进行基因工程人胰岛素前体的酶切。     

用CHITOSAN固定化青霉素酰化酶

本文报道以Chitosan为载体制备固定化青霉素酰化酶的研究结果。1.4％ Chitosan醋酸溶液先用12.5％戊二醛交联,洗涤,研磨,得颗粒。再用1.5％pH5.5和1.5％pH8.5多聚磷酸处理,得机械强度良好的载体。此载体先后经0.5％戊二醛和对甲苯磺酰氯双活化后,再与青霉素酰化酶偶联。所得固定化酶的活力达20.8u/g。固定化青霉素酰化酶的反应最适温度略高于游离酶,最适pH略低于游离酶;其Km值基本接近游离酶。固定化酶经反复使用,活力基本不变。     

Design, Synthesis, and Antitumor Evaluation of Novel Pyrazolo[3,4-d]pyrimidine Derivatives

A new series of pyrazolo[3,4-d]pyrimidines has been synthesized. The new compounds were tested for their antitumor activity on 60 different cell lines, and some of the compounds were found to have potent antitumor activity. In particular, 2-hydroxybenzaldehyde [1-(4-chlorophenyl)-3-methyl-1H-pyrazolo-[3,4-d]pyrimidin-4-yl]hydrazone (VIIa) was found to be the most effective among the other derivatives, showing IC50 values of 0.326 to 4.31 μM on 57 different cell lines.     

Effects of isolation and fighting in mice on the rate of synthesis of noradrenaline,dopamine and 5-hydroxytryptamine in the brain

Male mice were isolated or housed in groups for 6–8 weeks. Some of the isolated mice were thereafter brought together in groups of 25–30 animals for 30 min, during which period they fought each other intensively. The brain concentration of 5-HIAA was lower in isolated than in grouped animals. 30 min after administration of the aromatic amino acid decarboxylase inhibitor 3-hydroxybenzyl-hydrazine HCl (NSD 1015) the amount of Dopa accumulated in the brain was lower in isolated than in grouped animals. There was no corresponding difference in the accumulation of 5-HTP. The accumulation of Dopa and 5-HTP after NSD 1015 was accelerated during fighting. The effects of isolation and fighting on DOPA accumulation were similar in NA- and DA-dominated areas of the brain. The brain concentrations of tryptophan and tyrosine were increased and the concentration of NA was decreased in fighting NSD 1015 treated animals. The brain concentrations of HVA was lower in isolated and higher in fighting animals than in grouped animals.The present results indicate that the rate of synthesis of brain CA is lower in isolated than in grouped animals and that intensive fighting rapidly accelerates the synthesis of CA and 5-HT in the brain.     

纤维状青霉素酰化酶在制备7-ADCA上的应用

用纤维状青霉素酰化酶替代进口酶,裂解头孢G制备7-ADCA,质量、收率均高于替代前,成本降低64.6元.     

南蛇藤属植物化学成分及活性研究进展

南蛇藤属植物在我国分布广泛，近年来从中分得了多种β-二氢沉香呋喃型倍半萜和friedlane等型三萜，其中一些具有昆虫拒食、抗肿瘤和细胞毒活性。本文综述了近10年来从南蛇藤属植物中分得的化学成分及其活性的研究进展。     

Combined effects of carbon monoxide and ethanol on the activities of leucine aminopeptidase and glutamic-pyruvic transaminase in the plasma of rats

An investigation was undertaken to determine the influence of single doses of carbon monoxide plus single doses of ethanol, repeated simultaneous doses of carbon monoxide plus ethanol, and repeated dosage of carbon monoxide (ethanol) plus single doses of ethanol (carbon monoxide) on the activities of leucine amino-peptidase and glutamic-pyruvic transaminase in the plasma of rats. Combinations of carbon monoxide and ethanol were found to produce additive effects with respect to the increase of indicator enzymes if at least 50 % of the hemoglobin was saturated with carbon monoxide and if one of the noxious agents was administered in large doses.This paper is dedicated to Professor Dr. Konrad Pohle on the occasion of his 75th birthday.     

~(59)Fe摄入法和ELISA法测定重组人红细胞生成素活性的比较

对重组人红细胞生成素（rhEPO）活性测定的两种方法进行了比较。认为标准方法（~(59)Fe摄入法）结果准确,但操作复杂。而ELISA法必须与其它纯度检查法如SDS-PAGE、IEF、HPLC等相结合才能得出准确结果。对于高纯度结构完整的rhEPO分子,两种方法结果应一致。     

两种蛇毒制剂的类凝血酶活性及精氨酸酯酶活性比较

去纤酶、东菱精纯克栓酶两种蛇毒制剂在类凝血酶活力和精氨酸酯酶活力方面表现出较大的差异。东菱精纯克栓酶类凝血酶活性强而酯酶活性相对较低，去纤酶则与之相反。     

Effects of diazepin derivatives on cholinesterase activity

The authors have investigated the effect of therapeutic doses of diazepin derivatives to the cholinesterase activity of healthy persons. All of the diazepin derivatives significantly increased the cholinesterase activity.     

有机磷中毒患者血胆碱酯酶活性的变化规律及临床意义

目的研究急性有机磷中毒（AOPP）患者全血及血清胆碱脂酶（ChE）活性的变化规律。方法所有51例患者入院后均给予清除毒物、应用双复磷或氯磷定、阿托品及积极的对症支持治疗。分别于治疗前、治疗后6～12、24、48、72、96、120h及出院时抽末梢血测定全血及血清ChE活性,并于同期测定37名均为健康者血ChE活性作为正常对照。结果全血、血清ChE活性在中毒患者治疗前均较正常对照组明显降低（P〈0.01）,治疗后轻度患者的全血ChE活性恢复较快,至出院时基本恢复正常;中、重度患者的全血ChE活性于治疗后24h降至最低,于48h缓慢回升,至出院时仍未恢复至正常水平;3例死亡患者全血ChE活性持续不回升。血清ChE活性值波动较大,分布较离散,恢复相对较快。结论全血ChE活性是对急情有机磷中毒进行早期诊断,判断疗效和估计预后的重要指标,在治疗上具有重要的指导意义。     

Parameters influencing the effect of delta9-THC on activity wheel behavior

In 2 studies the effect of Δ⁹-THC on activity wheel behavior in rats was examined. The amount of laboratory acclimation prior to testing was manipulated and either 4 mg/kg or 8 mg/kg Δ⁹-THC was given intraperitoneally. Activity counts were taken 15 minutes, 1, 6, 24, 48 or 72 hr after the injection. Those animals that received 4 mg/kg Δ⁹-THC and had little laboratory acclimation were significantly more active than their controls during the first 15 min but, after 1 hr were, like the other 3 experimental groups, less active than the appropriate controls. The time course for the depressant action of the Δ⁹-THC at both dose levels was quite similar and lasted for approximately 24 hr.