1.1类新药“富马酸替诺福韦双特戊酯”和鹰嘴豆总生物碱抗肝炎药理学研究
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摘要
本课题包括上篇“1.1类新药富马酸替诺福韦双特戊酯抗肝炎临床药理学研究”和下篇“鹰嘴豆生物碱体外抗HBV活性研究”两部分。
上篇：1.1类新药富马酸替诺福韦双特戊酯抗肝炎临床药理学研究
富马酸替诺福韦双特戊酯(Tenofovir dipivoxil fumarate)是江苏正大天晴药业开发的新型替诺福韦酯类前药，用于治疗慢性乙型肝炎。该药按化学药品1.1类注册，本研究严格遵循“化学药物临床药代动力学研究技术指导原则”以及“2010版药典二部附录”的规定进行试验设计，以全面、系统的阐明其在人体内的安全性、吸收、分布、代谢、排泄的规律，为其II期临床试验的给药方案提供依据。本研究从以下五方面获得了具有科学意义和应用价值的研究结果：
1.药物分析测定方法的建立
目的：建立快速、灵敏的液相色谱-串联质谱(LC-MS/MS)检测血浆和尿液中替诺福韦的方法，LC-MS/MS检测血浆中左卡尼汀的方法。
方法：1.采用LC-MS/MS检测血浆和尿液中替诺福韦，三重四极杆质谱以多反应监测方式进行检测。检测离子为：替诺福韦，[M+H]+离子，m/z288.1→176.0；恩替卡韦，[M+H]+离子，m/z278.1→152.0。
2.采用LC-MS/MS检测血浆中左卡尼汀：三重四极杆质谱以多反应监测方式进行检测。检测离子为：左卡尼汀，[M+H]+离子，m/z162→84.65；托特罗定，[M+H]+离子，m/z326.2→146.9。
结果：血浆中替诺福韦线性范围为2~(-1)200ng/mL，批内和批间精密度分别为1.1~4.8%、8.4~12.9%，绝对回收率>90%；尿液中替诺福韦线性范围为800-64000ng/mL，批内和批间精密度分别为0.9~3.4%、0.8~2.6%，绝对回收率接近100%；血浆中左卡尼汀线性范围为2-20μg/mL，批内和批间精密度分别为2.7~3.0%、6.6~8.9%，绝对回收率>90%。
结论：所建方法灵敏、快速、准确，能够满足血浆和尿液中替诺福韦、以及血浆中左卡尼汀的检测。
2.药代动力学研究
目的：评价富马酸替诺福韦双特戊酯人体内吸收、分布和消除的特点。
方法：24名健康受试者随机均分为2组进行药动学试验，用LC/MS/MS法测定血浆和尿液中替诺福韦浓度，计算药动学参数。
结果：口服富马酸替诺福韦双特戊酯150mg、300mg和600mg后，替诺福韦的消除半衰期t1/2为14.06±1.83h、17.00±0.84h、16.32±2.29h，AUC0-72为1410.05±200.42ng·h·mL~(-1)、2872.94±335.99ng·h·mL~(-1)、5728.67±1056.59ng·h·mL~(-1)，达峰时间Tmax为1.5±0.8h、0.8±0.3h以及1.5±0.5h和达峰浓度Cmax分别为209.23±54.47ng·mL~(-1)、511.82±67.78ng·mL~(-1)以及959.91±275.03ng·mL~(-1)，0～48小时的尿累积排泄率为29.58±2.84%，28.05±3.55%，30.81±7.35%。多次口服富马酸替诺福韦双特戊酯，稳态平均血药浓度Cssav为156.81±33.0ng·mL~(-1)，稳态血药浓度波动度DF为3.94±0.5，蓄积常数R为1.59±0.15，消除半衰期t_(1/2)为19.58±2.11h。
结论：150~300mg剂量范围内替诺福韦AUC(0-72)、C_(max)均与剂量呈线性关系。连续7天给药，体内出现蓄积。
3.饮食影响研究
目的：考察饮食对于口服富马酸替诺福韦双特戊酯吸收、分布、代谢和排泄的影响。
方法：12名健康受试者，采用标准两阶段自身对照试验设计，分别空腹及餐后口服受试制剂。
结果：12名健康受试者单剂量餐后口服富马酸替诺福韦双特戊酯片300mg，测得血浆中替诺福韦主要药代动力学参数： AUC_(0-72)为3270.37±290.52h·ng/ml，C_(max)为432.62±85.02ng·mL~(-1)，T_(max)为1.5±0.5h，t_(1/2)为15.48±2.04h。
结论：空腹和餐后给药的AUC、C_(max)、T_(max)、t_(_(1/2))均有显著性差异，可见饮食对替诺福韦的药动学有显著影响，高脂高热量饮食后，替诺福韦生物利用度与空腹给药相比提高14.7%，表明饮食有利于药物吸收。
4.相对生物利用度研究
目的：评价口服等剂量新药富马酸替诺福韦双特戊酯与已上市同类药韦瑞德相比在人体内的相对生物利用度。
方法：选择18名男性健康受试者，采用标准两阶段自身对照试验设计，分别口服受试制剂和参比制剂。
结果：18名健康男性受试者，单剂量口服富马酸替诺福韦双特戊酯与韦瑞德分别300mg，测得血浆中替诺福韦的主要药动学参数分别为t_(1/2)15.74±1.68h、16.75±2.11h，AUC_(0-72)2695.62±483.42ng·h·mL~(-1)、2250.41±387.31ng·h·mL~(-1)，T_(max)1.1±0.5h、0.5±0.2h和C_(max)466.72±162.55ng·mL~(-1)、445.54±109.44ng·mL~(-1)。
结论：受试者单次口服富马酸替诺福韦双特戊酯片的平均相对生物利用度为120.1±12.5%
5.富马酸替诺福韦双特戊酯对内源性左卡尼汀的影响
目的：考察富马酸替诺福韦双特戊酯对内源性左卡尼汀水平的影响。
方法：临床试验中随行监测受试者体内左卡尼汀水平，观察各受试者临床试验前后血浆左卡尼汀水平变化。
结果：人体对替诺福韦吸收、分布、代谢和消除行为会对人体内源性左卡尼汀水平造成一定影响。替诺福韦血药浓度升高到一定程度就会导致人体左卡尼汀水平降低，药物消除后左卡尼汀水平会逐渐恢复正常。连续多次给药后，人体左卡尼汀会维持在较低水平。
结论：进行富马酸替诺福韦双特戊酯连续给药时，建议加服左卡尼汀口服液。
下篇：鹰嘴豆生物碱提取分离与体外抗HBV活性研究
目的：对鹰嘴豆进行提取分离，将分得总生物碱成分进行体外抗HBV活性研究。
方法：采用硅胶柱层析，氯仿-甲醇-水梯度洗脱对鹰嘴豆正丁醇层进行系统分离，将分得到的总生物碱部分进行体外抗肝炎药理学研究。
结果：首次从鹰嘴豆中分离到生物碱成分，研究发现鹰嘴豆总生物碱具有体外抗乙型肝炎病毒活性，可以明显抑制HepG2.2.15细胞上清液中的HBeAg、HBsAg、HBV DNA水平，抑制作用具有浓度和时间依赖性。进一步对总生物碱分离得到生物碱单体化合物，并通过理化性质和现代波谱学手段(UV、IR、MS、~1H-NMR、~13C-NMR、2D-NMR)鉴定了它们的结构。通过单晶培养得到了化合物1的单晶结构，明确了该化合物的立体构型。
结论:鹰嘴豆生物碱对乙肝病毒具有体外抑制活性。
Part Ⅰ: Clinical pharmacology study of tenofovir dipivoxilfumarate
Tenofovir dipivoxil fumarate is a new prodrug of tenofovir, exploited byJiangsu Chia-Tai Tianqing Pharmaceutical Co. Ltd., used for the treatment ofchronic hepatitis B. It’s registered for category1.1of chemical drugs accordingto Drug Registration Regulation. The clinical pharmacology study was designedstrictly following “Technical guidance for chemical drugs in clinicalpharmacokinetic studies” and “The appendix Ⅱ of Chinese pharmacopoeia” inorder to comprehensively and systematically clarify the laws of safety,absorption, metabolic, distribution, excretion of the new drug in human body,and to provide the evidence of dosage regimen for Phase II clinical trials. Thisstudy obtained scientific significance and application values in following fiveaspects:
1. Establishment of drug analysis methods.
Objective: To establish a rapid, specific and sensitive LC-MS/MS methodto detect tenofovir in plasma and urine and levocarnitine in plasma: Applicationto a clinical pharmacology study of tenofovir dipivoxil fumarate in human.
Methods:1. Detection of tenofovir in plasma and urine: Massspectrometric detection was performed on a triple quad LC–MS/MS inelectrospray positive ionization using multiple reaction monitoring. The masstransition was m/z288.1→176for tenofovir, and m/z278.1→152.0forentecavir, respectively.
2. Detection of levocarnitine in plasma: Mass spectrometric detection wasperformed on a triple quad LC–MS/MS in electrospray positive ionization usingmultiple reaction monitoring. The mass transition was m/z162→84.65forlevocarnitine, and m/z326.2→146.9for tolterodine, respectively.
Result: The linear range of tenofovir in plasma is2~(~(~(~(-1))))200ng/mL, inter andintra precision is in the range of1.1~4.8%and8.4~12.9%, respectively.Absolute recovery is greater than90%. The linear range of tenofovir in urine is800-64000ng/mL, inter and intra precision is in the range of0.9~3.4%and0.8~2.6%, respectively. Absolute recovery is nearly100%. The linear range oflevocarnitine in plasma is2-20μg/mL, inter and intra precision is in the range of2.7~3.0%and6.6~8.9%. Absolute recovery is greater than90%.
Conclusion: The simple, specific and rapid LC-MS/MS method foranalysis of tenofovir in human plasma and urine and levocarnitine in plasmahave been developed and validated.
2. The study of pharmacokinetics
Objective: To detect tenofovir in plasma and urine and evaluate the singleand multiple pharmacokinetic characteristics for absorption, distribution andeliminate of tenofovir dipivoxil fumarate in human.
Methods:24healthy volunteers are divided into two groups randomly. Todetect tenofovir in plasma and urine by LC-MS/MS and calculate thepharmacokinetic parameters.
Result: After administration of single150,300or600mg of tenofovirdipivoxil fumarate, elimination half-life (t_(1/2)) of tenofovir are14.06±1.83h,17.00±0.84h,16.32±2.29h, AUC0-721410.05±200.42ng·h·mL~(-1),2872.94±335.99ng·h·mL~(-1),5728.67±1056.59ng·h·mL~(-1), T_(max)1.5±0.8h、0.8±0.3h and1.5±0.5h, C_(max)209.23±54.47ng·mL~(-1),511.82±67.78ng·mL~(-1)and959.91±275.03ng·mL~(-1), urine cumulative excretion rates of0～48h is29.58±2.84%,28.05±3.55%,30.81±7.35%. After administration of multiple oral doses of tenofovirdipivoxil fumarate, the steady average concentration (Cssav) is156.81±33.0ng·mL~(-1), fluctuations degrees (DF) is3.94±0.5, cumulative constant (R) is1.59±0.15, t_(1/2)is19.58±2.11h.
Conclusion: AUC(0-72)、C_(max)were found to be a linear relationship withdose in the range dose of150~300mg. Accumulation appears in human aftergiven tenofovir for7days.
3. The study of food effect
Objective: To investigate the effect of high fat and calorie diet on theabsorption, distribution, metabolism and excretion after administration ofmiddle doses of tenofovir dipivoxil fumarate300mg.
Methods:12healthy volunteers were oral administered300mg of dipivoxilfumarate in fed or fast state in a single-dose, randomized,2-way, open-label,crossover study.
Result: The main pharmacokinetic parameters of tenofovir in fed statewere AUC0-723270.37±290.52h·ng/ml, C_(max)432.62±85.02ng/ml, T_(max)1.5±0.5h,t_(1/2)15.48±2.04h.
Conclusion: The AUC, C_(max), T_(max), and t_(1/2)have the significantdifferences in fasted or fed, and the bioavailability increased14.7%after a highfat and calorie diet.
4. The study of relative bioavailability
Objective: To evaluate of the relative bioavailability between tenofovir dipivoxil fumarate and viread after administration a comparative same dose.
Methods:18healthy male volunteers were administered test and reference,respectively, in a single-dose, randomized,2-way, open-label, crossover study.
Result: The main pharmacokinetic parameters of tenofovir afteradministration test or reference were t1/215.74±1.68h,16.75±2.11h, AUC0-722695.62±483.42ng·h·mL~(-1),2250.41±387.31ng·h·mL~(-1), T_(max)1.1±0.5h,0.5±0.2hand C_(max)466.72±162.55ng·mL~(-1),445.54±109.44ng·mL~(-1), respectively.
Conclusion: The relative bioavailability of tenofovir dipivoxil was20%higher compared with tenofovir disoproxil fumarate.
5. The effect of tenofovir dipivoxil fumarate on the endogenouslevocarnitine
Objective: To evaluate the effect of tenofovir dipivoxil fumarate on theendogenous levocarnitine.
Methods: To determine levels of levocarnitine before and after the clinicaltrials, and to observe changes of levocarnitine before and after the clinical trials.
Result: The process of absorption, distribution, metabolism andelimination of tenofovir dipivoxil fumarate could affect the levels of endogenouslevocarnitine in human. The level of levocarnitine could decrease whenconcentration of tenofovir rise to a certain degree. Levocarnitine will graduallyreturn to normal after drug elimination of tenofovir. After multiple doses,levocarnitine will keep in a low level.
Conclusion: Levocarnitine was advised to be administered whenadministration of multiple doses of tenofovir dipivoxil fumarate.
Objective: To extract and separate of chickpeas and conduct anti-HBVexperiments in vitro for total alkaloids separated from chickpeas.
Methods: To separate the part of N-butyl alcohol systematically extractedfrom chickpea using silica gel column chromatography method, the chloroform-methanol-water was used for gradient elution. The sub-total alkaloids wereconducted in vitro anti-hepatitis pharmacology study.
Results: For the first time, alkaloids were isolated from the chickpeas, andthe pharmacology study found that the total alkaloids had in vitro anti-hepatitisB virus activity, which could significantly inhibit HBsAg, HBeAg, HBV DNAin the supernatant of HepG2.2.15cells dependent on the dose and time. Furtherto isolate the alkaloid monomers from the total alkaloids, and identify theirstructure by physicochemical properties and the means of modern spectroscopic(UV, IR, MS,1H-NMR,13C-NMR and2D-NMR) methods. Crystal singlestructure of compound1was obtained by single crystal cultivation to clarify thestereochemistry of the compound1.
Conclusion: Chickpeas alkaloids have in vitro anti-hepatitis B virus activity.
上篇：1.1类新药富马酸替诺福韦双特戊酯抗肝炎临床药理学研究
富马酸替诺福韦双特戊酯(Tenofovir dipivoxil fumarate)是江苏正大天晴药业开发的新型替诺福韦酯类前药，用于治疗慢性乙型肝炎。该药按化学药品1.1类注册，本研究严格遵循“化学药物临床药代动力学研究技术指导原则”以及“2010版药典二部附录”的规定进行试验设计，以全面、系统的阐明其在人体内的安全性、吸收、分布、代谢、排泄的规律，为其II期临床试验的给药方案提供依据。本研究从以下五方面获得了具有科学意义和应用价值的研究结果：
1.药物分析测定方法的建立
目的：建立快速、灵敏的液相色谱-串联质谱(LC-MS/MS)检测血浆和尿液中替诺福韦的方法，LC-MS/MS检测血浆中左卡尼汀的方法。
方法：1.采用LC-MS/MS检测血浆和尿液中替诺福韦，三重四极杆质谱以多反应监测方式进行检测。检测离子为：替诺福韦，[M+H]+离子，m/z288.1→176.0；恩替卡韦，[M+H]+离子，m/z278.1→152.0。
2.采用LC-MS/MS检测血浆中左卡尼汀：三重四极杆质谱以多反应监测方式进行检测。检测离子为：左卡尼汀，[M+H]+离子，m/z162→84.65；托特罗定，[M+H]+离子，m/z326.2→146.9。
结果：血浆中替诺福韦线性范围为2~(-1)200ng/mL，批内和批间精密度分别为1.1~4.8%、8.4~12.9%，绝对回收率>90%；尿液中替诺福韦线性范围为800-64000ng/mL，批内和批间精密度分别为0.9~3.4%、0.8~2.6%，绝对回收率接近100%；血浆中左卡尼汀线性范围为2-20μg/mL，批内和批间精密度分别为2.7~3.0%、6.6~8.9%，绝对回收率>90%。
结论：所建方法灵敏、快速、准确，能够满足血浆和尿液中替诺福韦、以及血浆中左卡尼汀的检测。
2.药代动力学研究
目的：评价富马酸替诺福韦双特戊酯人体内吸收、分布和消除的特点。
方法：24名健康受试者随机均分为2组进行药动学试验，用LC/MS/MS法测定血浆和尿液中替诺福韦浓度，计算药动学参数。
结果：口服富马酸替诺福韦双特戊酯150mg、300mg和600mg后，替诺福韦的消除半衰期t1/2为14.06±1.83h、17.00±0.84h、16.32±2.29h，AUC0-72为1410.05±200.42ng·h·mL~(-1)、2872.94±335.99ng·h·mL~(-1)、5728.67±1056.59ng·h·mL~(-1)，达峰时间Tmax为1.5±0.8h、0.8±0.3h以及1.5±0.5h和达峰浓度Cmax分别为209.23±54.47ng·mL~(-1)、511.82±67.78ng·mL~(-1)以及959.91±275.03ng·mL~(-1)，0～48小时的尿累积排泄率为29.58±2.84%，28.05±3.55%，30.81±7.35%。多次口服富马酸替诺福韦双特戊酯，稳态平均血药浓度Cssav为156.81±33.0ng·mL~(-1)，稳态血药浓度波动度DF为3.94±0.5，蓄积常数R为1.59±0.15，消除半衰期t_(1/2)为19.58±2.11h。
结论：150~300mg剂量范围内替诺福韦AUC(0-72)、C_(max)均与剂量呈线性关系。连续7天给药，体内出现蓄积。
3.饮食影响研究
目的：考察饮食对于口服富马酸替诺福韦双特戊酯吸收、分布、代谢和排泄的影响。
方法：12名健康受试者，采用标准两阶段自身对照试验设计，分别空腹及餐后口服受试制剂。
结果：12名健康受试者单剂量餐后口服富马酸替诺福韦双特戊酯片300mg，测得血浆中替诺福韦主要药代动力学参数： AUC_(0-72)为3270.37±290.52h·ng/ml，C_(max)为432.62±85.02ng·mL~(-1)，T_(max)为1.5±0.5h，t_(1/2)为15.48±2.04h。
结论：空腹和餐后给药的AUC、C_(max)、T_(max)、t_(_(1/2))均有显著性差异，可见饮食对替诺福韦的药动学有显著影响，高脂高热量饮食后，替诺福韦生物利用度与空腹给药相比提高14.7%，表明饮食有利于药物吸收。
4.相对生物利用度研究
目的：评价口服等剂量新药富马酸替诺福韦双特戊酯与已上市同类药韦瑞德相比在人体内的相对生物利用度。
方法：选择18名男性健康受试者，采用标准两阶段自身对照试验设计，分别口服受试制剂和参比制剂。
结果：18名健康男性受试者，单剂量口服富马酸替诺福韦双特戊酯与韦瑞德分别300mg，测得血浆中替诺福韦的主要药动学参数分别为t_(1/2)15.74±1.68h、16.75±2.11h，AUC_(0-72)2695.62±483.42ng·h·mL~(-1)、2250.41±387.31ng·h·mL~(-1)，T_(max)1.1±0.5h、0.5±0.2h和C_(max)466.72±162.55ng·mL~(-1)、445.54±109.44ng·mL~(-1)。
结论：受试者单次口服富马酸替诺福韦双特戊酯片的平均相对生物利用度为120.1±12.5%
5.富马酸替诺福韦双特戊酯对内源性左卡尼汀的影响
目的：考察富马酸替诺福韦双特戊酯对内源性左卡尼汀水平的影响。
方法：临床试验中随行监测受试者体内左卡尼汀水平，观察各受试者临床试验前后血浆左卡尼汀水平变化。
结果：人体对替诺福韦吸收、分布、代谢和消除行为会对人体内源性左卡尼汀水平造成一定影响。替诺福韦血药浓度升高到一定程度就会导致人体左卡尼汀水平降低，药物消除后左卡尼汀水平会逐渐恢复正常。连续多次给药后，人体左卡尼汀会维持在较低水平。
结论：进行富马酸替诺福韦双特戊酯连续给药时，建议加服左卡尼汀口服液。
下篇：鹰嘴豆生物碱提取分离与体外抗HBV活性研究
目的：对鹰嘴豆进行提取分离，将分得总生物碱成分进行体外抗HBV活性研究。
方法：采用硅胶柱层析，氯仿-甲醇-水梯度洗脱对鹰嘴豆正丁醇层进行系统分离，将分得到的总生物碱部分进行体外抗肝炎药理学研究。
结果：首次从鹰嘴豆中分离到生物碱成分，研究发现鹰嘴豆总生物碱具有体外抗乙型肝炎病毒活性，可以明显抑制HepG2.2.15细胞上清液中的HBeAg、HBsAg、HBV DNA水平，抑制作用具有浓度和时间依赖性。进一步对总生物碱分离得到生物碱单体化合物，并通过理化性质和现代波谱学手段(UV、IR、MS、~1H-NMR、~13C-NMR、2D-NMR)鉴定了它们的结构。通过单晶培养得到了化合物1的单晶结构，明确了该化合物的立体构型。
结论:鹰嘴豆生物碱对乙肝病毒具有体外抑制活性。
Part Ⅰ: Clinical pharmacology study of tenofovir dipivoxilfumarate
Tenofovir dipivoxil fumarate is a new prodrug of tenofovir, exploited byJiangsu Chia-Tai Tianqing Pharmaceutical Co. Ltd., used for the treatment ofchronic hepatitis B. It’s registered for category1.1of chemical drugs accordingto Drug Registration Regulation. The clinical pharmacology study was designedstrictly following “Technical guidance for chemical drugs in clinicalpharmacokinetic studies” and “The appendix Ⅱ of Chinese pharmacopoeia” inorder to comprehensively and systematically clarify the laws of safety,absorption, metabolic, distribution, excretion of the new drug in human body,and to provide the evidence of dosage regimen for Phase II clinical trials. Thisstudy obtained scientific significance and application values in following fiveaspects:
1. Establishment of drug analysis methods.
Objective: To establish a rapid, specific and sensitive LC-MS/MS methodto detect tenofovir in plasma and urine and levocarnitine in plasma: Applicationto a clinical pharmacology study of tenofovir dipivoxil fumarate in human.
Methods:1. Detection of tenofovir in plasma and urine: Massspectrometric detection was performed on a triple quad LC–MS/MS inelectrospray positive ionization using multiple reaction monitoring. The masstransition was m/z288.1→176for tenofovir, and m/z278.1→152.0forentecavir, respectively.
2. Detection of levocarnitine in plasma: Mass spectrometric detection wasperformed on a triple quad LC–MS/MS in electrospray positive ionization usingmultiple reaction monitoring. The mass transition was m/z162→84.65forlevocarnitine, and m/z326.2→146.9for tolterodine, respectively.
Result: The linear range of tenofovir in plasma is2~(~(~(~(-1))))200ng/mL, inter andintra precision is in the range of1.1~4.8%and8.4~12.9%, respectively.Absolute recovery is greater than90%. The linear range of tenofovir in urine is800-64000ng/mL, inter and intra precision is in the range of0.9~3.4%and0.8~2.6%, respectively. Absolute recovery is nearly100%. The linear range oflevocarnitine in plasma is2-20μg/mL, inter and intra precision is in the range of2.7~3.0%and6.6~8.9%. Absolute recovery is greater than90%.
Conclusion: The simple, specific and rapid LC-MS/MS method foranalysis of tenofovir in human plasma and urine and levocarnitine in plasmahave been developed and validated.
2. The study of pharmacokinetics
Objective: To detect tenofovir in plasma and urine and evaluate the singleand multiple pharmacokinetic characteristics for absorption, distribution andeliminate of tenofovir dipivoxil fumarate in human.
Methods:24healthy volunteers are divided into two groups randomly. Todetect tenofovir in plasma and urine by LC-MS/MS and calculate thepharmacokinetic parameters.
Result: After administration of single150,300or600mg of tenofovirdipivoxil fumarate, elimination half-life (t_(1/2)) of tenofovir are14.06±1.83h,17.00±0.84h,16.32±2.29h, AUC0-721410.05±200.42ng·h·mL~(-1),2872.94±335.99ng·h·mL~(-1),5728.67±1056.59ng·h·mL~(-1), T_(max)1.5±0.8h、0.8±0.3h and1.5±0.5h, C_(max)209.23±54.47ng·mL~(-1),511.82±67.78ng·mL~(-1)and959.91±275.03ng·mL~(-1), urine cumulative excretion rates of0～48h is29.58±2.84%,28.05±3.55%,30.81±7.35%. After administration of multiple oral doses of tenofovirdipivoxil fumarate, the steady average concentration (Cssav) is156.81±33.0ng·mL~(-1), fluctuations degrees (DF) is3.94±0.5, cumulative constant (R) is1.59±0.15, t_(1/2)is19.58±2.11h.
Conclusion: AUC(0-72)、C_(max)were found to be a linear relationship withdose in the range dose of150~300mg. Accumulation appears in human aftergiven tenofovir for7days.
3. The study of food effect
Objective: To investigate the effect of high fat and calorie diet on theabsorption, distribution, metabolism and excretion after administration ofmiddle doses of tenofovir dipivoxil fumarate300mg.
Methods:12healthy volunteers were oral administered300mg of dipivoxilfumarate in fed or fast state in a single-dose, randomized,2-way, open-label,crossover study.
Result: The main pharmacokinetic parameters of tenofovir in fed statewere AUC0-723270.37±290.52h·ng/ml, C_(max)432.62±85.02ng/ml, T_(max)1.5±0.5h,t_(1/2)15.48±2.04h.
Conclusion: The AUC, C_(max), T_(max), and t_(1/2)have the significantdifferences in fasted or fed, and the bioavailability increased14.7%after a highfat and calorie diet.
4. The study of relative bioavailability
Objective: To evaluate of the relative bioavailability between tenofovir dipivoxil fumarate and viread after administration a comparative same dose.
Methods:18healthy male volunteers were administered test and reference,respectively, in a single-dose, randomized,2-way, open-label, crossover study.
Result: The main pharmacokinetic parameters of tenofovir afteradministration test or reference were t1/215.74±1.68h,16.75±2.11h, AUC0-722695.62±483.42ng·h·mL~(-1),2250.41±387.31ng·h·mL~(-1), T_(max)1.1±0.5h,0.5±0.2hand C_(max)466.72±162.55ng·mL~(-1),445.54±109.44ng·mL~(-1), respectively.
Conclusion: The relative bioavailability of tenofovir dipivoxil was20%higher compared with tenofovir disoproxil fumarate.
5. The effect of tenofovir dipivoxil fumarate on the endogenouslevocarnitine
Objective: To evaluate the effect of tenofovir dipivoxil fumarate on theendogenous levocarnitine.
Methods: To determine levels of levocarnitine before and after the clinicaltrials, and to observe changes of levocarnitine before and after the clinical trials.
Result: The process of absorption, distribution, metabolism andelimination of tenofovir dipivoxil fumarate could affect the levels of endogenouslevocarnitine in human. The level of levocarnitine could decrease whenconcentration of tenofovir rise to a certain degree. Levocarnitine will graduallyreturn to normal after drug elimination of tenofovir. After multiple doses,levocarnitine will keep in a low level.
Conclusion: Levocarnitine was advised to be administered whenadministration of multiple doses of tenofovir dipivoxil fumarate.
Objective: To extract and separate of chickpeas and conduct anti-HBVexperiments in vitro for total alkaloids separated from chickpeas.
Methods: To separate the part of N-butyl alcohol systematically extractedfrom chickpea using silica gel column chromatography method, the chloroform-methanol-water was used for gradient elution. The sub-total alkaloids wereconducted in vitro anti-hepatitis pharmacology study.
Results: For the first time, alkaloids were isolated from the chickpeas, andthe pharmacology study found that the total alkaloids had in vitro anti-hepatitisB virus activity, which could significantly inhibit HBsAg, HBeAg, HBV DNAin the supernatant of HepG2.2.15cells dependent on the dose and time. Furtherto isolate the alkaloid monomers from the total alkaloids, and identify theirstructure by physicochemical properties and the means of modern spectroscopic(UV, IR, MS,1H-NMR,13C-NMR and2D-NMR) methods. Crystal singlestructure of compound1was obtained by single crystal cultivation to clarify thestereochemistry of the compound1.
Conclusion: Chickpeas alkaloids have in vitro anti-hepatitis B virus activity.
引文
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