芒柄花素磺酸钠在脑缺血再灌注大鼠体内的药动学研究
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摘要
目的建立芒柄花素磺酸钠血浆样品的高效液相色谱(HPLC)分析方法,并应用该方法对正常大鼠和模型大鼠体内的芒柄花素磺酸钠进行药动学分析;明确芒柄花素磺酸钠在正常、模型大鼠血浆中的吸收分布情况。方法采用Venusil MP C_(18)色谱柱(250 mm×4.6 mm,5μm),流动相乙腈–水(30∶70,水相中加0.1%甲酸),柱温30℃,进样量20.0μL,体积流量1.0 mL/min,检测波长250 nm。制备大鼠脑缺血再灌注模型,将SD大鼠随机分为10组,每组5只大鼠。于大鼠脑缺血1 h及再灌注3.5 h按20 mg/kg ip芒柄花素磺酸钠注射液,并于给药后15、30、45、60、90、120、150、180、240、300 min腹主动脉取血。正常组大鼠分别于给药后5、10、15、30、45、60、90、120、150、180、240、300、480、720min目内眦取血。样品经处理后,采用HPLC法检测各时间点血浆中的芒柄花素磺酸钠浓度。采用药动学软件WinNonLin 6.3进行数据分析,计算药动学参数。结果芒柄花素磺酸钠在0.25～100.0μg/mL线性关系良好。定量下限RSD值为12.15%。日内精密度RSD值均<5.0%,日间精密度RSD值均<10.0%。提取回收率均>80%,RSD值均<5.0%。芒柄花素磺酸钠在正常大鼠、模型大鼠血浆内药动学参数为:t_(1/2)分别为(173.13±13.12)min、(117.86±45.05)min,C_(max)分别为(67.23±4.04)μg/mL、(75.22±10.57)μg/mL,AUC_(0-t)分别为(2 493.69±216.98)min·μg/mL、(6 094.52±250.81)min·μg/mL,Vd分别为(1 981.97±134.12)mL/kg、(484.56±167.93)mL/kg,Cl分别为(7.96±0.73)mL·min/kg、(2.87±0.14)mL·min/kg,MRT_(0-t)分别为(76.92±1.54)min、(89.06±0.77)min。结论应用HPLC法测定芒柄花素磺酸钠的血药浓度定量准确、灵敏,芒柄花素磺酸钠在模型大鼠血浆中吸收的量多,单位时间内消除的量少,滞留时间长。
Objective To establish an HPLC analysis method for sodium formononetin-3'-sulphonate plasma samples, and the method was applied to the pharmacokinetic analysis of sodium formononetin-3'-sulphonate in normal rats and model rats. The absorption and distribution of sodium formononetin-3'-sulphonate in plasma of normal and model rats were determined. Methods The separation was performed on Venusil MP C_(18) column(250 mm × 4.6 mm, 5 μm). The mobile phase consisted of acetonitrile-water(30∶70, add 0.1% formic acid to the aqueous phase). Temperature of column was set at 30 ℃, and the injection volume was 20.0 μL. The flow rate was 1.0 mL/min, and the detective wavelength was set at 250 nm. Rat cerebral ischemia reperfusion models were established. SD rats were randomly divided into 10 groups, and each group had 5 rats. At the time point of 1 h after cerebral ischemia and 3.5 h after reperfusion, rats were ip administered with sodium formononetin-3'-sulphonate injection 20 mg/kg, and the abdominal aorta blood samples were taken at 15, 30, 45, 60, 90, 120, 150, 180, 240, and 300 min after administration. Rats in the normal group were collected the inner canthus blood at the time of 5, 10, 15, 30, 45, 60, 90, 120, 150, 180, 240, 300, 480, and 720 min after the administration.Samples were treated, and HPLC method was used to detect sodium formononetin-3'-sulphonate concentration in plasma at different time points. Pharmacokinetic data were analyzed by WinNonLin 6.3 pharmacokinetic software, and pharmacokinetic parameters were calculated. Results A good linearity was shown in the concentration ranges of 0.25 —100.0 μg/mL for sodium formononetin-3'-sulphonate, and RSD value of the quantitative limits was 12.15%. The RSD value of inter-day precisions were less than 5.0%, and the intra-day precisions were less than 10.0%. The recovery rate of extraction was over 80%, and RSD value was less than 5.0%. The pharmacokinetic parameters of sodium formononetin-3'-sulphonate in plasma of normal rats and model rats were as following: t_(1/2) were(173.13 ± 13.12) and(117.86 ± 45.05) min, C_(max) were(67.23 ± 4.04) and(75.22 ± 10.57) μg/mL, AUC_(0-t) were(2 493.69 ±216.98) and(6 094.52 ± 250.81) min·μg/mL, Vd were(1 981.97 ± 134.12) and(484.56 ± 167.93) mL/kg, Cl were(7.96 ± 0.73) and(2.87 ± 0.14) mL·min/kg, MRT_(0-t) were(76.92 ± 1.54) and(89.06 ± 0.77) min. Conclusion The determination of sodium formononetin-3'-sulphonate in blood by HPLC is accurate and sensitive. And the amount of sodium formononetin-3'-sulphonate absorbed in the plasma of model rats is large, the amount eliminated in unit time is small, and the retention time is long.
Objective To establish an HPLC analysis method for sodium formononetin-3'-sulphonate plasma samples, and the method was applied to the pharmacokinetic analysis of sodium formononetin-3'-sulphonate in normal rats and model rats. The absorption and distribution of sodium formononetin-3'-sulphonate in plasma of normal and model rats were determined. Methods The separation was performed on Venusil MP C_(18) column(250 mm × 4.6 mm, 5 μm). The mobile phase consisted of acetonitrile-water(30∶70, add 0.1% formic acid to the aqueous phase). Temperature of column was set at 30 ℃, and the injection volume was 20.0 μL. The flow rate was 1.0 mL/min, and the detective wavelength was set at 250 nm. Rat cerebral ischemia reperfusion models were established. SD rats were randomly divided into 10 groups, and each group had 5 rats. At the time point of 1 h after cerebral ischemia and 3.5 h after reperfusion, rats were ip administered with sodium formononetin-3'-sulphonate injection 20 mg/kg, and the abdominal aorta blood samples were taken at 15, 30, 45, 60, 90, 120, 150, 180, 240, and 300 min after administration. Rats in the normal group were collected the inner canthus blood at the time of 5, 10, 15, 30, 45, 60, 90, 120, 150, 180, 240, 300, 480, and 720 min after the administration.Samples were treated, and HPLC method was used to detect sodium formononetin-3'-sulphonate concentration in plasma at different time points. Pharmacokinetic data were analyzed by WinNonLin 6.3 pharmacokinetic software, and pharmacokinetic parameters were calculated. Results A good linearity was shown in the concentration ranges of 0.25 —100.0 μg/mL for sodium formononetin-3'-sulphonate, and RSD value of the quantitative limits was 12.15%. The RSD value of inter-day precisions were less than 5.0%, and the intra-day precisions were less than 10.0%. The recovery rate of extraction was over 80%, and RSD value was less than 5.0%. The pharmacokinetic parameters of sodium formononetin-3'-sulphonate in plasma of normal rats and model rats were as following: t_(1/2) were(173.13 ± 13.12) and(117.86 ± 45.05) min, C_(max) were(67.23 ± 4.04) and(75.22 ± 10.57) μg/mL, AUC_(0-t) were(2 493.69 ±216.98) and(6 094.52 ± 250.81) min·μg/mL, Vd were(1 981.97 ± 134.12) and(484.56 ± 167.93) mL/kg, Cl were(7.96 ± 0.73) and(2.87 ± 0.14) mL·min/kg, MRT_(0-t) were(76.92 ± 1.54) and(89.06 ± 0.77) min. Conclusion The determination of sodium formononetin-3'-sulphonate in blood by HPLC is accurate and sensitive. And the amount of sodium formononetin-3'-sulphonate absorbed in the plasma of model rats is large, the amount eliminated in unit time is small, and the retention time is long.
引文
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[17]Ma S W,Zhao M,Liu H X,et al.Pharmacokinetic effects of baicalin on cerebral ischemia-reperfusion after iv administration in rats[J].Chin Herb Med,2012,4(1):53-57.
[2]幸奠霞,刘志高,薛存宽,等.刺芒柄花素雌激素样作用及其与血脂相关性研究[J].中国老年学杂志,2009,29(18):2340-2342.
[3]顾民华,洪文,唐传其,等.芒柄花素保护前脑缺血再灌注损伤中的血脑屏障并抑制神经炎症[J].暨南大学学报:自然科学与医学版,2015,36(1):34-39.
[4]王秋亚,孟庆华,张尊听,等.芒柄花素磺化物的合成、溶解性能及降脂保肝活性[J].药学学报,2009,44(4):386-389.
[5]张淑敏.Sul-F对心肌损伤的保护作用及其机制研究[D].青岛:中国海洋大学,2011.
[6]杨彬,余丽娟,王佳,等.全脑缺血/再灌注大鼠皮层PGI2和TXA2时程变化特征[J].中国药理学通报,2013,29(12):1667-1671.
[7]郑莜萸.化学药品和治疗用生物制品研究指导原则(试行)[M].北京:中国医药科技出版社,2005:1-21.
[8]王本祥.现代中药药理与临床[M].天津:天津科技翻译出版公司,2004:963.
[9]陈奇.中药药理研究方法学[M].北京:人民卫生出版社,2006:34-36.
[10]崔治家,李富云,候嘉,等.甘肃不同产地黄芪芒柄花素的含量测定[J].甘肃中医学院学报,2012,29(1):56-58.
[11]周广为,慕慧,李岳锋,等.测定红车轴草中刺芒柄花素含量的新方法[J].中药材,2007,30(1):54-56.
[12]李莹,陈晓辉,张天虹,等.RP-HPLC法测定鸡血藤中芒柄花素的含量[J].沈阳药科大学学报,2009,26(12):975-977.
[13]海力茜,张烜,徐建军,等.HPLC测定新疆红芪中芒柄花素的含量[J].华西药学杂志,2005,20(5):433-434.
[14]张君仁,藏恒昌,等.体内药物分析[M].北京:化学工业出版社,2002:7.
[15]He X,Xing D,Ding Y,et al.Effects of cerebral ischemiareperfusion on pharmacokinetic fate of paeoniflorin after intravenous administration of Paeoniae Radix extract in rats[J].J Ethrnopharmacol,2004,94(2-3):339-344.
[16]Yan B,Xing D M,Ding Y,et al.HPLC method for the determination and pharmacokinetic studies on puerarin in cerebral ischemia reperfusion rat plasma after intravenous administration of puerariae radix isoflavone[J].J Pharm Biomed Anal,2005,37(2):297-301.
[17]Ma S W,Zhao M,Liu H X,et al.Pharmacokinetic effects of baicalin on cerebral ischemia-reperfusion after iv administration in rats[J].Chin Herb Med,2012,4(1):53-57.