氢化可的松和表氢化可的松的密度泛函理论研究

采用密度泛函理论B3LYP/6-31G和B3LYP/6-311G*方法优化了氢化可的松和表氢化可的松的几何结构,利用优化的结构得到了氢化可的松和表氢化可的松的原子净电荷、总能量及前沿分子轨道组成.基于简谐振动分析求得了氢化可的松和表氢化可的松的红外光谱频率和强度,由统计热力学分析得到了热力学函数;进而确定了氢化可的松和...     

有机溶剂与氢化可的松的溶剂—溶质作用

通过测定氢化可的松在多种溶剂／水体系中的分配系数研究了氢化可的松与溶剂的相互作用机制．其中醇、酮与氢化可的松有较强的氢键作用，酯、醚、氯代烷烃等溶剂与氢化可的松的氢键作用较弱，四氯化碳、苯、环己烷不能与氢化可的松形成氢键．     

氢化可的松催化超氧阴离子歧化及反应动力学

在0.1mol·L^-1NaHCO3介质中,用伏安法研究了超氧阴离子O2^.-与糖皮质类甾体氢化可的松的化学反应。实验表明,氢化可的松清除O2^.-的化学作用机制为氢化可的松催化O2^.-的歧化反应,氢化可的松是O2^.-的清除剂。氢化可的松催化O2^.-歧化反应的速率对O2^.-为零级表观反应,对氢化可的松则为二级表观反应,求得20℃时氢化可的松催化O2^.-歧化反应表观速率常数k为8.76×10^5L·mol^-1·s^-1。本结果为医学组织研究结果提供了新的实验证据。在抗炎作用方面,氢化可的松除抑制磷脂酶A2的活性从而间接阻止O2^.-的产生外,还能直接化学清除产生的O2^.-,认为氢化可的松的抗炎作用应是这种生物和化学的综合作用结果。     

高分子支撑液膜固定化细胞甾体△1-脱氢转化的研究

研究了高分子支撑液膜固定化细胞的制备、性质以及影响其催化氢化可的松→去氢氢化可的松生物转化的各种因素,并与经典珠状包埋法固定细胞的转化作用进行了比较。结果表明前者的△¹-脱氢速率和产物转化率均高于后者。在所使用的条件下二者未见机械强度方面明显差别。     

氢化可的松的热力学函数和晶体结构——密度泛函理论分析

采用密度泛函理论B3LYP/6-31G和B3LYP/6-311G*方法优化了气相状态氢化可的松的几何结构,利用优化的结构得到了氢化可的松各原子净电荷及前沿轨道分布;基于简谐振动分析求得氢化可的松的红外光谱频率和热力学函数;并计算了环己烷、乙酸丁酯、二氯乙烷、异丙醇、甲醇、水六种不同极性的溶剂对氢化可的松几何构型、电荷分布及能量的影响.结果表明,由理论计算得到的氢化可的松的晶体结构与实测的晶体结构接近.     

场放大进样-胶束毛细管电泳法测定化妆品中3种糖皮质激素

采用场放大进样-胶束毛细管电泳法对化妆品中氢化可的松、泼尼松和乙酸氢化可的松3种糖皮质激素进行了分离测定。电泳介质为0.20mmol.L-1硼砂缓冲溶液(pH 9.0),运行电压为-20kV,进样电压-20kV,进样时间45s,进水压力3kPa,进水时间20s,检测波长250nm。在优化试验条件下,氢化可的松、泼尼松和乙酸氢化可的松的检出限分别(3S/N)为0.015,0.017,0.017mg.L-1。应用此方法分析了化妆品样品,测得回收率在93.8%～107%之间,测定值的相对标准偏差(n=5)均小于5.1%。     

HPLC法测定氯霉素氢化可的松滴耳液中两组分的含量

采用反相高效液相色谱法,以醋酸泼尼松为内标,ＵＶ检测波长为２４４ｎｍ,以ＹＷＧ－Ｃ１８为固定相,以甲醇－水（６０∶４０）为流动相,同时测定了氯霉素氢化可的松滴耳液中氯霉素和氢化可的松的含量,并进行了线性范围和回收率测定,平均回收率（ｎ＝６）和相对标准偏差分别为：９９．９１％和０．８６％（氯霉素）,９９．５８％和１．３４％（氢化可的松）。     

反相流动注射-化学发光法测定氢化可的松

在碱性介质中,氢化可的松可被Fe(CN)6^3-氧化产生化学发光,奎宁对此发光反应有显著的增敏作用。据此,建立了反相流动注射-化学发光法测定氢化可的松的新方法。方法的线性范围和检出限分别为0．05．-50mg／L和0．02mg／L,相对标准偏差(n=11,C=1．0mg／L)为0．9％。方法用于药物中氢化可的松含量的测定,结果与标准方法分光光度法测定结果吻合。     

阴离子选择性耗尽进样-胶束扫集毛细管电泳法对痕量醋酸氢化可的松的测定

联用阴离子选择性耗尽进样和胶束扫集两种在线富集技术,建立了胶束毛细管电泳方法测定化妆品中醋酸氢化可的松的方法。讨论了SDS浓度、样品基体、进样电压、进水时间和进样时间对富集和分离的影响。优化的实验条件:以120 mmol/L SDS-20 mmol/L NaH2PO4(pH2.2)-10%(体积分数)甲醇为缓冲体系,分离电压-20 kV,进样电压-20 kV,进样时间80 s,进水时间200 s,测量波长250 nm。在该实验条件下,醋酸氢化可的松的富集倍数比普通毛细管电泳法提高了约173倍。方法的线性范围为0.05~5.0 mg/L,检出限为12.6μg/L。该方法用于化妆品中醋酸氢化可的松含量的测定,回收率为98%~105%,相对标准偏差均小于4.0%(n=4)。     

氢化可的松氨基糖衍生物的合成

描述了一条简单有效的合成氢化可的松氨基糖衍生物的合成路线．实验中首先 将氢化可的松转化为琥珀酸酯衍生物2，葡萄糖、2—乙酰氨基葡萄糖、半乳糖、乳 糖及氨基葡萄糖盐酸盐转化成为相应的氨基糖7a-7d和12，然后2与7a-7d和12在 DCC的作用下通过酰胺键进行偶联，得到一系列氢化可的松糖氨基糖衍生物13a- 13e．上述目标化合物均为新化合物，其结构分别通过红外光谱(IR)、核磁共振波 谱(^1H NMR,^13C NMR)及HMRS进行了确证。     

Biotransformation of tetrahydro-alpha-santonins by Absidia coerulea

The fungus, Absidia coerulea was employed to bioconvert tetrahydro-alpha-santonins, 1,2,4alpha,5alpha-tetrahydro-alpha-santonin (1), and its 4-epimer (2), from which 10 products (3-12) were obtained. Furthermore, their structures were determined, based on their chemical and spectroscopic data analyses. Among them, 3-5, 7, 9, 11 and 12 were observed to be seven new compounds. The reactions mainly involved in these bio-process included hydroxylation(s) (C-4, C-11, and C-1), reduction (C-3 ketone to alcohol).     

Optimisation of Hydrocortisone Production by Curvularia lunata

A new method for breeding the hydrocortisone overproducing strain Curvularia lunata by screening ketoconazole-resistance mutant was developed. A hydrocortisone overproducing mutant C. lunata KA-91 with ketoconazole-resistance marker was obtained from protoplasts treated with ultraviolet radiation. The hydrocortisone conversion rate of C. lunata KA-91 was increased by 42.1% compared to the original strain CL-114 at the substrate 17α-hydroxypregn-4-en-3, 20-dione-21-acetate addition concentration of 1.0 g/L. The by-products produced by KA-91 were fewer than those of the original strain. It was assumed that the higher cytochrome P450 content of ketoconazole-resistance mutant resulted in the increase of 11β-hydroxylation capacity. The culture conditions for biotransformation of 17α-hydroxypregn-4-en-3, 20-dione-21-acetate to hydrocortisone were optimized by response surface methodology. Plackett–Burman design was applied to elucidate the key factors affecting the hydrocortisone production, and the results indicated that glucose, initial pH, and glucose to total nitrogen sources ratio (ω) had significant effects on hydrocortisone production. Box–Behnken design was employed to search for the optimal parameters of those three key factors. According to the model, the trial checking at the optimal conditions showed a high hydrocortisone conversion rate of 82.67%.     

Supported liquid extraction in combination with LC‐MS/MS for high‐throughput quantitative analysis of hydrocortisone in mouse serum

A high‐throughput LC–MS/MS bioanalytical method was developed and validated for the determination of hydrocortisone in mouse serum via supported liquid extraction (SLE) in a 96‐well plate format. Although sample extracts from SLE result in similar matrix effects compared with conventional liquid–liquid extraction (LLE), greater analyte extraction recovery and much higher analysis throughput for the quantitative analysis of hydrocortisone in mouse serum were obtained. The current LC‐MS/MS method was validated for a concentration range of 2.00–2000 ng/mL for hydrocortisone using a 0.100 mL volume of mouse serum. The intra‐ and inter‐day precision and accuracy of the quality control samples at low, medium and high concentration levels showed ≤12.9% CV and ?3.4–6.2% bias for the analyte in mouse serum. Copyright © 2009 John Wiley & Sons, Ltd.     

Capillary electrophoresis and hollow fiber liquid-phase microextraction for the enantioselective determination of albendazole sulfoxide after biotransformation of albendazole by an endophytic fungus

Hollow fiber liquid-phase microextraction and CE were applied for the determination of albendazole sulfoxide (ASOX) enantiomers in liquid culture medium after a fungal biotransformation study. The analytes were extracted from 1 mL of liquid culture medium spiked with the internal standard (rac-hydroxychloroquine) and buffered with 0.50 mol/L phosphate buffer, pH 10. The analytes were extracted into 1-octanol impregnated in the pores of the hollow fiber, and into an acid acceptor solution inside the polypropylene hollow fiber. The electrophoretic separations were carried out in 0.05 mol/L tris(hydroxymethyl)aminomethane buffer, pH 9.3, containing 3.0% w/v sulfated-β-CD (S-β-CD) with a constant voltage of +15 kV and detection at 220 nm. The method was linear over the concentration range of 250-5000 ng/mL for each ASOX enantiomer. Within-day and between-day assay precision and accuracy for the analytes were studied at three concentration levels and the values of RSD% and relative error % were lower than 15%. The developed method was applied for the determination of ASOX after a biotransformation study employing the endophytic fungus Penicillium crustosum (VR4). This study showed that the endophytic fungus was able to metabolize the albendazole to ASOX enantioselectively. In addition, it was demonstrated that hollow fiber liquid-phase microextraction coupled to CE can be an excellent and environmentally friendly technique for the analysis of samples obtained in biotransformation studies.     

Development of a simple HPLC method for determination of paeoniflorin-metabolizing activity of intestinal bacteria in rat feces

A simple and reproducible HPLC method for the determination of paeoniflorin (PF)-metabolizing activity of intestinal bacteria in rat feces was developed and validated. Orally administered PF, a major active constituent of Paeoniae Radix, is metabolized into a bioactive compound, paeonimetabolin I (PM-I) by intestinal bacteria. Direct determination of the PF-metabolizing rate into PM-I is hard to achieve by HPLC due to the lack of intense chromophore in PM-I. However, when PF was incubated with Lactobacillus brevis, an intestinal bacterium, in the presence of phenylmercaptan, the metabolizing rate of PF into 8-phenylthio-paeonimetabolin I (PT-PM-I) was found to be equivalent to that of PF into PM-I. Thus, the PF-metabolizing activity of intestinal bacteria in rat feces was determined by measuring the rate of biotransformation of PF into PT-PM-I, which was detected by HPLC at 255 nm. This method can be utilized in the biopharmaceutical study of traditional Chinese formulations containing Paeoniae Radix.     

High-performance liquid chromatographic assay of betamethasone 17-valerate and its degradation products

A high-performance liquid chromatographic assay of betamethasone 17-valerate is described. The procedure may be use for quantitative assay of the degradation products, betamethasone 21-valerate and betamethasone, and the application to the analysis of ointments is described. The method is also suitable for the determination of the kinetics of decomposition from one experimental run, and the determination of rate constants from a four-compartment sequential reaction is described. The procedure is also applicable to other corticosteroids, and hydrocortisone 17-butyrate, hydrocortisone 21-butyrate, and hydrocortisone may similarly be determined without modification to the method.     

高效阴离子交换色谱-脉冲安培检测法同时测定生物转化样品中的海藻糖、葡萄糖和麦芽糖

建立了高效阴离子交换色谱-脉冲安培电化学检测法同时测定生物转化样品中海藻糖、葡萄糖和麦芽糖的分析方法。选用CarboPac^TM10色谱柱(250 mm×2 mm)对分离条件进行优化,使用标准样品测定了线性范围和工作曲线,柱温为30 ℃,流速为0.30 mL/min,以氢氧化钠溶液和醋酸钠溶液为流动相进行梯度洗脱,脉冲安培法进行检测。研究结果表明,该方法可在15 min内实现海藻糖生物转化液中3种糖的快速定量分析。海藻糖、葡萄糖和麦芽糖峰面积与质量浓度的线性关系良好,检出限为0.010~0.100 mg/L。将此方法用于酶法制备海藻糖的检测,加标回收率为89.40%~103.2%。在生物转化样品中检测到海藻糖浓度为101.084 g/L,转化率达到了50.5%。该方法灵敏度高,简便快速,可应用于海藻糖制备样品中各种成分的分离和定量检测。     

Development and validation of a reversed-phase HPLC method for quantitative determination of ginsenosides Rb1, Rd, F2, and compound K during the process of biotransformation of ginsenoside Rb1

A simple and rapid gradient RP-HPLC method for simultaneous separation and determination of related ginsenosides during the process of biotransformation of ginsenoside Rb1 has been developed. As many as four process ginsenosides have been separated and identified on an Eclipse XDB C(18) column (4.6 mm x 150 mm, 5 mum) with gradient elution using water and ACN as a mobile phase. The column was maintained at 30 degrees C and the eluents were monitored with diode array detection at 203 nm. The method was validated in terms of linearity, sensitivity, precision, and accuracy. The correlation coefficients (r) for calibration curves of ginsenosides were in the range of 0.9996-1.0000. The proposed RP-HPLC method was successfully applied to the analysis of fermentation broth and the recoveries of ginsenosides were in the range of 94.4-103.1% with RSD <2.87%. The method could be of use for rapid and routine evaluation of the quantity of ginsenosides during the biotransformation process of ginsenoside Rb1.     

高碘酸钾-鲁米诺体系中一些后化学发光反应的研究

研究了葡醛内酯、盐酸美司坦和重酒石酸去甲肾上腺素等10种物质在高碘酸钾-鲁米诺体系中的后化学发光现象、后化学发光反应的动力学、化学发光光谱、荧光光谱以及其它相关性质, 提出了其可能的发光机理; 在优化的分析条件下建立了这10种物质的后化学发光分析方法, 初步构建了高碘酸钾-鲁米诺后化学发光分析体系.