黄杨宁缓释胶囊释放度的研究

目的 研究自制的黄杨宁缓释胶囊的体外释放度。方法 采用酸性染料染色分光光度法测定黄杨宁缓释胶囊的释放度,0.05 mol·L^-1的磷酸二氢钠缓冲溶液为释放介质,转速为100 r·min^-1。结果 平均回收率98.5%,RSD=3.39%,3批样品在1,4,8 h的平均累计释放度分别为23.10%,57.39%和85.86%,释放度符合要求。结论 自制黄杨宁缓释胶囊有较好的释药性能。本试验建立的释放度测定方法可以作为该制剂的质控方法。     

黄杨宁缓释胶囊在犬体内的药动学研究

目的评价自制黄杨宁缓释胶囊在犬体内的药动学。方法Beagle犬分别单剂量口服给药6mg自制黄杨宁缓释胶囊和市售黄杨宁普通片,两次交叉试验间隔两周。采用反相高效液相色谱法,测定犬给药后不同时间的血药浓度,用以估算药动学参数和相对生物利用度。结果黄杨宁缓释胶囊和黄杨宁普通片的Cmax分别为（5．28±0．81）ng·mL^-1和（11．70±2．01）ng·mL^-1,tmax分别为（4．3±0．5）h和（1．0±0．1）h。缓释胶囊相对于普通片的生物利用度为96．23％。结论黄杨宁缓释胶囊具有显著的缓释效果。     

黄杨宁缓释胶囊在犬体内的药动学研究

目的 评价自制黄杨宁缓释胶囊在犬体内的药动学。方法 Beagle犬分别单剂量口服给药6 mg自制黄杨宁缓释胶囊和市售黄杨宁普通片,两次交叉试验间隔两周。采用反相高效液相色谱法,测定犬给药后不同时间的血药浓度,用以估算药动学参数和相对生物利用度。结果 黄杨宁缓释胶囊和黄杨宁普通片的C_max分别为(5.28±0.81) ng·mL^-1和(11.70±2.01) ng·mL^-1,t_max分别为(4.3±0.5) h和(1.0±0.1) h。缓释胶囊相对于普通片的生物利用度为96.23%。结论 黄杨宁缓释胶囊具有显著的缓释效果。     

维生素C微丸缓释胶囊的体外释放特征

目的建立维生素C微丸缓释胶囊体外释放度检测方法,评价其体外释放特性。方法以0．01mol·L^-1盐酸-0．03％乙二胺四乙酸二钠-0．05％L-半胱氨酸溶液为释放介质,介质体积为1000mL,转速为50r·min^-1,采用紫外-可见分光光度法于243nm波长处测定吸光度,转篮法评价体外释放特性。结果维生素C质量浓度在1．028～12．336mg·L^-1内与其吸收度之间呈良好线性关系（r=0．9999）;平均回收率为99．56％,RSD为0．86％。结论维生素C微丸缓释胶囊具有较好的释药性能和良好的缓释效果,符合《中国药典》要求。     

己酮可可碱缓释胶囊的制备及体外释放度检测

目的研制己酮可可碱缓释胶囊,确定其体外释放度测定方法.方法采用挤出滚圆法制取含药小丸,再对小丸进行缓释包衣,制得缓释小丸.采用紫外分光光度法测定其体外释放度,并进行方法验证.结果自制缓释胶囊粒间、批间差异小;采用紫外分光光度法测定其体外释放度,线性范围4～20μg·ml-1,线性方程A=0.036 1C 0.000 9(r=1.000 0).平均回收率为98.8%,RSD1.4%(n=6).结论采用紫外分光光度法测定其体外释放度方便、快速、准确.     

磷酸苯丙哌林缓释胶囊的释放度测定

目的：建立了UV法测定磷酸苯丙哌林缓释胶囊的释放度。方法：采用《中国药典》2005年版二部附录XD第二法，转速为50r·min^-1，检测波长为270nm。结果：磷酸苯丙哌林在0．03～0．15mg·ml^-1浓度范围内线性线性关系良好（r=0．9999），平均回收率为99．9％，RSD为1．1％（n=9）。结论：用UV法测定磷酸苯丙哌林缓释胶囊释放度方法简单，易于控制其释放度。     

罗红霉素缓释胶囊含量和释放度测定方法研究

目的：建立罗红霉素缓释胶囊含量和释放度测定方法。方法：采用高效液相色谱法,色谱柱为ZORBAX SB-C18（4.6 mm ×150 mm,5μm）,0.067 mol· L-1磷酸二氢铵溶液（三乙胺含量0.3％,调pH值至7.5）-乙腈（60∶40）为流动相,检测波长210 nm,用外标法计算罗红霉素缓释胶囊含量;释放度测定以磷酸盐缓冲液（pH 6.8）为溶剂,溶出2,4,8 h时取样测定。结果：含量测定在0.22～2.00 mg· mL-1（ R2＝0.9999）浓度范围内呈良好的线性关系,平均回收率为101.50％（ RSD＝1.07％）;样品在2,4,8 h的释放率分别相应为标示量的15％～45％、40％～70％和70％以上。结论：该方法简便、准确,适用于罗红霉素缓释胶囊含量和释放度的测定。     

盐酸二甲双胍缓释片释放度的比较

目的制备盐酸二甲双胍缓释片,并与国外相同缓释片的释放度进行比较。方法以HPMC（K4M）和CMC-Na为缓释材料制备盐酸二甲双胍缓释片,采用相似因子法对实验室自制的和国外进口的盐酸二甲双胍缓释片的释放度进行考察。结果自制缓释片和进口缓释片1h的累计释放百分率分别为27．71％±0．97％和25．30％±2．03％,6h的累计释放百分率分别为67．47％±1．45％和70．34％±1．17％,12h的累计释放百分率分均大于85％;两种样品释放曲线的f2为（84．18±4．67）。结论自制的缓释片具有12h缓释效果;两种制剂的缓释曲线经相似因子法f2比较不存在显著性差异。     

曲尼司特缓释胶囊的研制

本文用颗粒包衣方法制备了曲尼司特缓释胶囊，缓释胶囊含有两种释放度不同的颗粒，再将颗粒混合，最后达到较理想的释放水平．采用高效液相色谱法测定了家犬的生物利用度，曲尼司特缓释胶囊较普通胶囊的达峰时间延迟5．64h，相对生物利用度为113％．     

醋氯芬酸缓释微丸在犬体内的药动学及相对生物利用度

目的:评价自制醋氯芬酸缓释微丸在犬体内的药动学和相对生物利用度。方法:3只犬分别单剂量服用自制醋氯芬酸缓释胶囊和市售醋氯芬酸普通胶囊,剂量均为200 mg。1周后两组交叉用药。采用HPLC法,以乙萘酚为内标测定犬给药后不同时间的血药浓度,用3P87软件估算药动学参数和相对生物利用度。结果:参比制剂和测试制剂的Tmax分别为(3．33±0．58)和(7．33±0．58)h,Cmax分别为(17．12±0．48)和(12．26±0．27)μg·mL-1, AUC0～24h分别为(96．26±21．70)和(133．22±24．50)μg·h·mL-1。统计分析显示,各主要药动学参数均有显著性差异。醋氯芬酸缓释微丸相对于普通胶囊的平均生物利用度为142．6％。结论:醋氯芬酸缓释微丸具有明显的缓释特征,其生物利用度优于醋氯芬酸普通胶囊。     

分泌人共刺激分子B7-2的重组卡介苗的构建及表达与鉴定

目的构建分泌表达人共刺激分子B7-2（hB7-2）的重组卡介苗（rBCG）。方法采用聚合酶链反应（PCR）以含hB7-2的质粒为模板扩增出hB7-2活性序列,利用基因重组技术插入pYL-MCS质粒构建出分泌型卡介苗穿梭表达载体pYL—hB7-2。用电穿孔方法将该质粒转入卡介苗得到rBCG,利用PCR扩增和测序检测其hB7-2的DNA表达,分别SDS-PAGE和ELISA法测定rBCG上清液中和菌内hB7-2蛋白。结果酶切鉴定、PCR扩增和DNA测序均表明rBCG含有的hB7-2DNA与文献结果一致,SDS—PAGE检测出有相应蛋白表达,ELISA法测定rBCG上清液中分泌的B7-2蛋白为3．8U／ml。结论构建的重组BCG可以分泌表达人共刺激分子hB7-2,能够提供激活免疫反应的共刺激信号,将会成为膀胱肿瘤免疫治疗的一种新型方法。     

RMP-7及其衍生物对脂质体跨血脑屏障作用的影响

目的研究RMP-7及其衍生物对脂质体跨血脑屏障的促进作用。方法测定RMP-7与DSPE-PEG-NHS偶联的物质的量比；用血脑屏障的体外模型验证其生物活性；观察脑组织切片中伊文思蓝的荧光位置和强度，测定脑组织中伊文思蓝的浓度。结果RMP-7与DSPE-PEG-NHS偶联物质的量比为1∶1；RMP-7和DSPE-PEG-RMP-7能使过氧化合物酶跨血脑屏障体外模型的转运速率增加2～3倍，修饰脂质体后能使所包封的伊文思蓝在脑中的浓度增加。结论RMP-7与DSPE-PEG-RMP-7对脂质体跨血脑屏障均有促进作用，且后者效果更好。     

人乳腺癌细胞MCF-7紫杉醇耐药株的建立及其生物学特性研究

目的建立紫杉醇(Taxol)的人乳腺癌耐药细胞株MCF-7/Taxol,并初步研究其生物学特性。方法采用低浓度加量持续诱导法建立MCF-7/Taxol;细胞模型的多药耐药性以SRB法检测;流式细胞术分析细胞周期分布;免疫组化法检测细胞表型变化;高效液相鉴定细胞中药物蓄积;光镜和电子扫描显微镜观察细胞形态。结果SRB法检测,MCF-7/Taxol细胞的紫杉醇半数抑制浓度(IC50)是亲代MCF-7细胞的525倍,撤药3mon后细胞的耐药指数保持在150倍以上,并对羟喜树碱、表柔比星、多柔比星、米托蒽醌、硫酸长春新碱、博来霉素和丝裂霉素等多种化疗药物产生交叉耐药性。MCF-7/Taxol细胞分化程度低于同步传代的MCF-7细胞,倍增时间明显高于亲本细胞,S期细胞明显增加,G1期细胞减少;随撤药时间的延长,细胞的增殖速度加快。MCF-7/Taxol细胞P-gp、LRP和GSTπ的表达水平较亲本细胞有明显增加,ER、PR阳性表达丢失;光镜下MCF-7/Taxol细胞明显变大并且形态不规则;电镜下其表面纤绒毛成小球状隆起和絮状突起;在稳定生长的耐药细胞和撤药培养的细胞中都有Taxol蓄积。结论该MCF-7/Taxol模型具有多药耐药细胞的基本生物学特性,可用于肿瘤多药耐药机制药的研究,推测耐药细胞株中含有天然MCF-7肿瘤干细胞。     

HPLC法测定7-氨基-去乙酰氧基头孢烷酸中有关物质含量

目的建立高效液相色谱法测定7-氨基-3-脱乙酰氧基头孢烷酸(7-ADCA)中有关物质[Δ-2-7-氨基-3-脱乙酰氧基头孢烷酸(Δ-2-7-ADCA)、苯乙酸、扩环酸]。方法色谱柱:Hypersil ODS柱(250 mm×4.6mm,10μm);流速:2.0 mL/min;柱温:30℃。DAD二极管矩阵检测器检测,检测波长:220 nm;进样量:20μL。梯度洗脱:流动相A:0.05 mol/L磷酸盐缓冲液(pH=6.0);流动相B:乙腈。结果Δ-2-7-ADCA、苯乙酸和扩环酸相对于7-ADCA的响应因子为0.56、0.83和1.1;7-ADCA的线性范围为0.59~5.88μg/mL(r=0.999 7),最低检测浓度:0.12μg/mL。结论该方法操作简便、结果准确、稳定,可用于7-ADCA中有关物质含量的测定。     

叶酸受体及二氢叶酸还原酶与人乳腺癌细胞MCF-7/ADR多药耐药的关系

目的以人乳腺癌多药耐药细胞系MCF-7/ADR及其敏感亲本系MCF-7为对象,探讨叶酸受体(FOLRα)及其下游基因二氢叶酸还原酶(DHFR)的表达与乳腺癌细胞多药耐药的关系。方法 MTT法测定阿霉素的细胞毒性作用及DHFR抑制剂对MCF-7/ADR细胞多药耐药的逆转作用;RT-PCR检测细胞FOLRα和DHFR mRNA的表达水平;免疫细胞化学检测FOLRα、P-gp的表达水平。结果 MCF-7细胞增殖速度快于MCF-7/ADR细胞,MCF-7细胞FOLRα mRNA转录水平较MCF-7/ADR细胞高,而MCF-7/ADR细胞DHFR mRNA较MCF-7细胞转录水平高;免疫细胞化学显示MCF-7细胞FOLRα的表达高于MCF-7/ADR细胞,而MCF-7/ADR细胞P-gp的表达较MCF-7细胞高;MCF-7/ADR对甲氨蝶啶无耐药性,甲氨蝶啶对MCF-7/ADR细胞多药耐药有逆转作用。结论 MCF-7/ADR细胞FOLRα的表达水平下调可能与细胞增殖水平有关,其下游基因DHFR表达水平与MCF-7/ADR细胞的MDR可能存在相关性。     

Combined delivery of temozolomide and the thymidine kinase gene for treatment of glioblastoma

Glioblastoma is the most malignant form of brain tumor. In this study, combination therapy with temozolomide (TMZ) and the herpes simplex virus thymidine kinase (HSVtk) gene was evaluated in glioblastoma models. The R7L10 peptide was used as a carrier of TMZ and the HSVtk gene. TMZ was loaded into R7L10 micelles using the oil-in-water emulsion/solvent evaporation method. The TMZ-loaded R7L10 (R7L10-TMZ) micelles formed a complex with the HSVtk gene, pHSVtk. The formation of the R7L10-TMZ/pHSVtk complex was confirmed by gel retardation and heparin competition assays. An in vitro transfection assay demonstrated that the transfection efficiency of R7L10-TMZ was similar to that of R7L10 in C6 glioblastoma cells. R7L10-TMZ had greater anti-tumor effects than TMZ alone in C6 cells in vitro, suggesting that R7L10 is an efficient carrier of TMZ. The in vivo efficacy of the R7L10-TMZ/pHSVtk complex was evaluated in the intracranial glioblastoma model. HSVtk expression in tumors was confirmed by immunohistochemistry. Furthermore, a greater anti-tumor effect was observed in the R7L10-TMZ/pHSVtk group compared with the TMZ or R7L10/pHSVtk single injection group. In conclusion, combined delivery of TMZ and the HSVtk gene using R7L10 peptides may be useful for the treatment of glioblastoma.     

Tissue distribution of coumarin, 7-hydroxycoumarin and their 7-hydroxy metabolites following parenteral administration of 14C-labeled compound in the DBA/lac mouse

The time related tissue distributions of coumarin (C), 7-hydroxycoumarin (7HC) and their metabolites were studied in DBA/lac mice following retro-orbital injection of 14C-labeled compound. The activity was determined as dpm/ml or g wet weight over a period of 120 min. C was found in blood, kidney, liver, muscle, brain, heart, lung and fat but not in the testes. Upon C dosing, 7HC was found in kidney, liver and lung; 7-hydroxycoumarin glucuronide (7HCG) was found in blood, kidney and liver; 7-hydroxycoumarin sulfate (7HCS) was found in the blood, kidney and lung. Unidentified glucuronide and sulfate metabolites were detected in blood, kidney, liver and lung. Two unidentified unconjugated metabolites were detected in the kidney and liver. Unidentified water soluble metabolites were detected in kidney, liver, muscle, heart and testes. Upon 7HC dosing only 7HC, 7HCG and 7HCS were detected in any organ. 7HC and 7HCG were found in blood, kidney, liver, muscle and lung. 7HCS was detected in blood, kidney, liver and lung. Upon both C and 7HC dosing the kidney and liver contained the highest levels of radioactivity.     

耐药人乳腺癌细胞中葡萄糖神经酰胺合成酶的表达及意义

目的:探讨葡萄糖神经酰胺合成酶(GCS)基因在人乳腺癌细胞株(MCF-7)和耐阿霉素人乳腺癌细胞株(MCF-7/ADR)中的表达及意义。方法:采用MTT法检测阿霉素对MCF-7/ADR和MCF-7的半数抑制浓度(IC50),用不同浓度的GCS抑制剂PDMP预处理MCF-7/ADR不同时间后检测IC50;运用流式细胞术检测MCF-7及PDMP预处理MCF-7/ADR前、后GCS蛋白的表达水平;应用实时荧光定量PCR法检测MCF-7及PDMP作用前、后MCF-7/ADR中GCS基因的表达(GCSN)水平。结果:阿霉素对MCF-7/ADR和MCF-7的IC50分别为(18.95±0.54)、(0.84±0.07)μg·mL-1;MCF-7/ADR对MCF-7的耐药倍数为22.69倍,PDMP作用后阿霉素对MCF-7/ADR的IC50下降至(5.63±0.58)μg·mL-1。MCF-7/ADR中GCS蛋白及GCSN的表达均高于MCF-7(P<0.01),PDMP使MCF-7/ADR中GCSN表达水平从0.0048±0.0017下降至0.0021±0.0004。结论:GCS可能参与肿瘤耐药的发生过程,并在MCF-7/ADR多药耐药中起重要作用。     

The Effect of RMP-7 and its Derivative on Transporting Evens Blue Liposomes into the Brain

To investigate the effect of RMP-7 and its derivative on drug transport across blood brain barrier (BBB), RMP-7 and DSPE-PEG-NHS [1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-n-[poly(ethyleneglycol)]-hydroxy succinamide, PEG M 3400] were conjugated under mild conditions and the reaction ratio was determined using MALDI–TOF-MS (matrix-assisted laser desorption-ionization time-of-flight mass spectrometry). An endothelial cell monolayer in vitro BBB model was established and used to determine the bioactivity of RMP-7 and its derivative “opening BBB.” Horse radish peroxide (HRP), liposome (HRP-L-PEG), and Evens blue (EB) liposome (EB-L-PEG) were prepared using the reverse-phase evaporation method. HRP-L-PEG-RMP-7 and EB-L-PEG-RMP-7 were obtained by inserting DSPE-PEG-RMP-7 into the surface of liposome. The bioactivity of RMP-7 and DSPE-PEG-RMP-7 opening BBB were evaluated to determine their effect on the permeation ratio of HRP and HRP liposome across the in vitro BBB model. To evaluate the in vivo bioactivity of RMP-7 and DSPE-PEG-RMP-7 on EB transport across BBB into the brain, the indicated compounds were administered to rats. Then, brain slices were analyzed using confocal laser scanning microcopy and the EB concentration in the brain, liver, spleen, lung, and kidney was determined using the formamide–extraction–ultraviolet-spectrophotometric method. The results demonstrated that RMP-7 was conjugated with DSPE-PEG-NHS at the molecular ratio of 1:1 and the product is DSPE-PEG-RMP-7. Compared with adding HRP alone, RMP-7 and DSPE-PEG-RMP-7 improved 2- to 3-fold the transport of HRP in the in vitro BBB model. The in vivo experiments showed that DSPE-PEG-RMP-7 was better at facilitating EB transport into brain than RMP-7. The reason may be that DSPE-PEG-RMP-7 can “open BBB” as soon as the EB-L-PEG-RMP-7 reaches BBB.     

Enzymatic oxidation of 7-hydroxylated delta 8-tetrahydrocannabinol to 7-oxo-delta 8-tetrahydrocannabinol by hepatic microsomes of the guinea pig

Hepatic microsomes of the guinea pig converted delta 8-tetrahydrocannabinol (delta 8-THC) to various oxidized metabolites, including 7 alpha-hydroxy-delta 8-THC (7 alpha-OH-delta 8-THC), 7 beta-OH-delta 8-THC, and 7-oxo-delta 8-THC. The enzyme which mediates biotransformation of 7-OH-delta 8-THCs to 7-oxo-delta 8-THC was characterized in the present study. The oxidative activity was mainly located in microsomes. The microsomal reaction required NADPH and oxygen and showed an optimal pH around 7.5. The reaction was inhibited by beta-diethylaminoethyl diphenylpropylacetate (SKF 525-A), an inhibitor of cytochrome P-450, but not by pyrazole, a specific inhibitor of alcohol dehydrogenase. However, 7-oxo-delta 8-THC formation was not affected by carbon monoxide or by pretreatment of animals with cobaltous chloride (40 mg/kg, ip, once a day for 3 days). Atmospheric oxygen was incorporated into 7-oxo-delta 8-THC formed from 7 alpha-OH-delta 8-THC, but not into that from 7 beta-OH-delta 8-THC. Further, 7-oxo-delta 8-THC formed from 7 alpha-18OH-delta 8-THC released about half of 18O at the 7-position, whereas the 7-oxo metabolite from 7 beta-18OH-delta 8-THC lost little of the isotope at the 7 beta-position during the oxidative reaction. From these results, it is likely that hepatic microsomal monooxygenase (probably cytochrome P-450) plays a main role in the oxidation. In addition, mechanisms for 7-oxo-delta 8-THC formation from 7 alpha-OH-delta 8-THC or 7 beta-OH-delta 8-THC are different.