甘蔗渣纤维素固定化木瓜蛋白酶及其应用研究

匀浆状甘蔗渣纤维素用NaOH活化后，顺次经过高碘酸钠、尿素、甲醛处理，成为高反应性固定化酶载体，可用于木瓜黄蛋白酶的固定化，其酶蛋白的结合量与甲醛处理时间、酶固定时间、溶液酶浓度等有关，甲醛处理最适时间为14h，酶固定8h后，载体结合酶量趋向稳定；溶液酶最佳浓度范围为1－2．0mg/ml粗酶，固定化酶活力回收达34．5％，半衰期35天，固定化酶和溶液酶的Km值（底物酪蛋白，w/v,%)分别为0．12％和0．26％；固定化酶和溶液酶的最适PH分别为PH8．0和PH8．5，二者的最适温度均为60－70℃。固定化酶和溶液酶一样有底物抑制现象，固定化酶在6mol/L脲中处理6h活力趋向稳定，其活力为原有活力的46．75，用固定化酶处理啤酒，啤酒浊度下降了2－9．25倍，蛋白质含量下降了78．8％，冷藏120天，处理啤酒无冷混浊现象发生，同时啤酒原有风味和其它理化指标保持不变。     

胞外青霉素酰化酶产生菌的筛选及产酶条件

采用青霉素梯度琼脂平皿法，从土壤中快速筛选出３５株产胞外青霉素酰化酶的菌株，经复筛有５株酶活力较高，经鉴定均为芽胞杆菌，Ｂ３５－１７菌株酶活力最高．研究了Ｂ３５－１７菌株的胞外青霉素酰化酶产生条件，该菌株在最适产酶条件下，酶活力达２．６７ｕ／ｍＬ．在发酵培养基中添加０．２％的玉米浆能降低Ｆｅ２＋对酶合成的抑制作用．     

色氨酸酶基因工程菌固定化及其培养条件的研究

对色氨酸酶基因工程菌 ＷＷ－ １１进行固定化及培养条件研究,为工业化生产 Ｌ－色氨酸奠定基础。方法;通过色氨酸酶活力测定,考察三种固定化材料及温度、 ｐＨ、单价阳离子和乙醇对固定化 ＷＷ－ １１色氨酸酶活力的影响。结果：以聚乙烯醇作为ＷＷ－１１的固定化载体,其活力回收为６０．９％。固定化ＷＷ－１１色氨酸酶降解反应最适ｐＨ为９．０、最适温度为５０℃;固定化ＷＷ－１１色氨酸酶合成反应最适ｐＨ为７．５、最适温度为４５℃。Ｋ＋、ＮＨ＋对固定化工程菌色氨酸酶有明显激活作用,而Ｎａ＋则有一定的抑制作用。结论：固定化工程菌色氨酸酶对温度、乙醇等的稳定性比游离菌有显著提高。     

聚二醇修饰牛胰核糖核酸酶

采用N－羟基珀酰亚胺活化酯法活化单甲氧基聚乙二醇,测定了乙二醇（PEG）的活化度为86．2％。以活化的PEG对牛胰核糖核酸酶进行化学修饰;分析了蛋白质被修饰程度。用毛细管电泳法给出了被修饰蛋白的修饰度与修饰蛋白分布的定量结果。比较了被修饰产物对大分子底物（酵母RNA）与小分子底物（2‘,3‘－环磷酸胞嘧啶）的降解活力,其表观酶活力分别保留了52．8％和66．3％。结合毛细管电泳定量分析得到的修正酶活力略低于表观酶活力。     

电位滴定法测定胰激肽释放酶活力测试条件探讨

本文用ＦＩＰ推荐的方法，以具有国际单位的参照品为标样，滴定液为０．０１ｍｏｌ／ＬＮａＯＨ标准溶液，考查了以ＢＡＥＥ为底物的电位滴定法测定胰激肽释放酶活力的实验条件，通过正交试验得出直观和方差分析的结果，依据酶活力单位高、ＳＤ和ＲＳＤ％较小的原则，求出酶活力测定的最适条件是：酶反应温度为２５℃；缓冲液为０．００１５ｍｏｌ／ＬＮａ２Ｂ４Ｏ７－０．２５ｍｏｌ／ＬＮａＣＬ－２×１０（－４）ｍｏｌ／ＬＥＤＴＡ，ｐＨ为８．００；反应液中酶浓度为０．４８ＩＵ／ｍｌ；底物浓度为５×１０（－３）ｍｏｌ／Ｌ；胰蛋白酶抑制剂浓度为１２５μｇ／ｍｌ。胰激肽释放酶活力测定的实验数据的相对标准偏差小于５％。     

AS1.398中性蛋白酶固定化条件的初步研究

考察壳聚糖、卡拉胶、海藻酸钠、琼脂等化载体对ＡＳ１．３９８中性蛋白酶固定化的 影响，确定０．３％戊二醛在３０℃、ＰＨ８．０的条件下处理壳聚糖８－１０ｈ，加酶液，固定８ｈ，酶活力回收约为７７％，固定化酶最适作用温度为５５℃，最适作用ＰＨ为８．０；固定化酶的稳定性比游离酶的稳定性有显著提高。     

醛糖还原酶抑制剂筛选模型的建立和初步应用

本研究从牛晶状体中提取醛糖还原酶（AR），以DL－甘油醛为底物、还原型辅酶II（NADPH）为辅酶，应用96孔石英板建立醛糖还原酶抑制剂（ARI）微量高效筛选模型。酶促反应体系体积为200μl，其组成成分为：50mmol/L磷酸盐缓冲液（pH6.2)、400mmol/L硫酸锂、5mmol/L 2－硫基乙醇、0．24mmol/L NADPH 1.2mg粗酶、2mmol/L DL-甘油醛，用蒸馏水补足体积。反应温度为37℃，规定反应体系吸光度每分钟下降0．001为一个酶活力单位。以不含底物的样品为空白对照。开展了从微生物代谢产物中筛选ARI的工作，已经累计筛选真菌和稀有放线菌发酵样品约1500个，其中获得阳性样品6个，阳性率约为0．4％。     

重组E.coli L-天冬酰胺酶在大鼠中的药物代谢动力学

应用放射性核素示踪技术研究^125I重组E.coliL－天冬酰胺酶在大鼠体内的药物代谢动力学。^125I重组L－天冬氨酶静脉注射后24h内，在尿、粪、胆汁中的排泄量分别占注射剂量的68．95％，4．44％和5．36％。测定血浆中^125I重组E.coliL－天冬酰胺酶浓度，应用聚丙烯酰胺凝胶电泳和生物成像分析系统结合方法评价原药水平，由房室模型评价药物动力学参数，静注后，浓度时间曲线符合二房室模型，初期和末端的t1/2分别为0．52－0．63h和2．39－2．76h，AUFC与剂量成正比。重组E.coliL－天冬酰胺酶的分解代谢产物主要随尿液排泄，重组E.coliL－天冬酰胺酶在大鼠中的药物热力学参数为临床试验提供了有用依据。     

固定化胰凝乳蛋白酶拆分L—苯丙氨酸

用酶活力为２．５ｍｇ／ｇ的多孔硅球固定化胰凝乳蛋白酶可拆分ＤＬ－苯丙氨酸甲酯得Ｌ－苯丙氨酸，拆分收率９２．３％，光学纯度９６．２％。     

α－双炔失碳酯对红细胞膜和钙调蛋白功能的影响

探讨α－双炔失碳酯对细胞膜功能的影响．结果表明，α－双炔失碳酯对红细胞溶血有很强的保护作用．１ｍｇ·Ｌ－１时保护率达７０％。该作用与α－双炔失碳酯对钙调蛋白功能的抑制有关。α－双炔失碳酯对钙调蛋白激活的Ｃａ２＋．Ｍｇ２＋－ＡＴＰ酶和钙调蛋白依赖性磷酸二酯酶的活力都有较明显的抑制作用，提示α－双炔失碳酯对钙信号系统的干扰可能与其抗肿瘤作用密切相关。     

Synthesis,Cytotoxicity and Antileishmanial Activity of Some N-(2-(indol-3-yl)ethyl)-7-chloroquinolin-4-amines

We report herein the condensation of 4,7-dichloroquinoline (1) with tryptamine (2) and D-tryptophan methyl ester (3) . Hydrolysis of the methyl ester adduct (5) yielded the free acid (6) . The compounds were evaluated in vitro for activity against four different species of Leishmania promastigote forms and for cytotoxic activity against Kb and Vero cells. Compound (5) showed good activity against the Leishmania species tested, while all three compounds displayed moderate activity in both Kb and Vero cells.     

Lung disease and PKCs

The lung offers a rich opportunity for development of therapeutic strategies focused on isozymes of protein kinase C (PKCs). PKCs are important in many cellular responses in the lung, and existing therapies for pulmonary disorders are inadequate. The lung poses unique challenges as it interfaces with air and blood, contains a pulmonary and systemic circulation, and consists of many cell types. Key structures are bronchial and pulmonary vessels, branching airways, and distal air sacs defined by alveolar walls containing capillaries and interstitial space. The cellular composition of each vessel, airway, and alveolar wall is heterogeneous. Injurious environmental stimuli signal through PKCs and cause a variety of disorders. Edema formation and pulmonary hypertension (PHTN) result from derangements in endothelial, smooth muscle (SM), and/or adventitial fibroblast cell phenotype. Asthma, chronic obstructive pulmonary disease (COPD), and lung cancer are characterized by distinctive pathological changes in airway epithelial, SM, and mucous-generating cells. Acute and chronic pneumonitis and fibrosis occur in the alveolar space and interstitium with type 2 pneumocytes and interstitial fibroblasts/myofibroblasts playing a prominent role. At each site, inflammatory, immune, and vascular progenitor cells contribute to the injury and repair process. Many strategies have been used to investigate PKCs in lung injury. Isolated organ preparations and whole animal studies are powerful approaches especially when genetically engineered mice are used. More analysis of PKC isozymes in normal and diseased human lung tissue and cells is needed to complement this work. Since opposing or counter-regulatory effects of selected PKCs in the same cell or tissue have been found, it may be desirable to target more than one PKC isozyme and potentially in different directions. Because multiple signaling pathways contribute to the key cellular responses important in lung biology, therapeutic strategies targeting PKCs may be more effective if combined with inhibitors of other pathways for additive or synergistic effect. Mechanisms that regulate PKC activity, including phosphorylation and interaction with isozyme-specific binding proteins, are also potential therapeutic targets. Key isotypes of PKC involved in lung pathophysiology are summarized and current and evolving therapeutic approaches to target them are identified.     

Gender-related symptoms and correlates of alcohol dependence among men and women with a lifetime diagnosis of alcohol use disorders

This study explored gender-related symptoms and correlates of alcohol dependence in a crosssectional study of 150 men and 150 women with a lifetime diagnosis of alcohol use disorders (AUD). Participants were recruited in equal numbers from treatment settings, correctional centres and the general community. Standardized measures were used to determine participants' use of substances, history of psychiatric disorders and psychosocial stress, their sensation seeking and family history of substance use and mental health disorders. Multivariate analyses were used to detect patterns of variables associated with gender and the lifetime severity of AUD. Men had a longer history of severe AUD than women. Women had similar levels of alcohol dependence and medical and psychological sequelae as men, despite 6 fewer years of AUD. More women than men had a history of severe psychosocial stress, severe dependence on other substances and antecedent mental health problems, especially mood and anxiety disorders. There were differences in family history of alcohol-related problems approximating same-gender aggregation. The severity of a lifetime AUD was predicted by its earlier age at onset and the occurrence of other disorders, especially anxiety, among both men and women. The limitations in the generalizability of these findings due to sample idiosyncrasies are discussed.     

Biochemical and molecular characterisation of cubozoan protein toxins

Class Cubozoa includes several species of box jellyfish that are harmful to humans. The venoms of box jellyfish are stored and discharged by nematocysts and contain a variety of bioactive proteins that are cytolytic, cytotoxic, inflammatory or lethal. Although cubozoan venoms generally share similar biological activities, the diverse range and severity of effects caused by different species indicate that their venoms vary in protein composition, activity and potency. To date, few individual venom proteins have been thoroughly characterised, however, accumulating evidence suggests that cubozoan jellyfish produce at least one group of homologous bioactive proteins that are labile, basic, haemolytic and similar in molecular mass (42-46 kDa). The novel box jellyfish toxins are also potentially lethal and the cause of cutaneous pain, inflammation and necrosis, similar to that observed in envenomed humans. Secondary structure analysis and remote protein homology predictions suggest that the box jellyfish toxins may act as α-pore-forming toxins. However, more research is required to elucidate their structures and investigate their mechanism(s) of action. The biological, biochemical and molecular characteristics of cubozoan venoms and their bioactive protein components are reviewed, with particular focus on cubozoan cytolysins and the newly emerging family of box jellyfish toxins.     

Dose tolerability of chronically inhaled voriconazole solution in rodents

Invasive pulmonary aspergillosis (IPA) is a fungal disease of the lung associated with high mortality rates in immunosuppressed patients despite treatment. Targeted drug delivery of aqueous voriconazole solutions has been shown in previous studies to produce high tissue and plasma drug concentrations as well as improved survival in a murine model of IPA. In the present study, rats were exposed to 20 min nebulizations of normal saline (control group) or aerosolized aqueous solutions of voriconazole at 15.625 mg (low dose group) or 31.25 mg (high dose group). Peak voriconazole concentrations in rat lung tissue and plasma after 3 days of twice daily dosing in the high dose group were 0.85 ± 0.63 μg/g wet lung weight and 0.58 ± 0.30 μg/mL, with low dose group lung and plasma concentrations of 0.38 ± 0.01 μg/g wet lung weight and 0.09 ± 0.06 μg/mL, respectively. Trough plasma concentrations were low but demonstrated some drug accumulation over 21 days of inhaled voriconazole administered twice daily. Following multiple inhaled doses, statistically significant but clinically irrelevant abnormalities in laboratory values were observed. Histopathology also revealed an increase in the number of alveolar macrophages but without inflammation or ulceration of the airway, interstitial changes, or edema. Inhaled voriconazole was well tolerated in a rat model of drug inhalation.