羟基喜树碱静脉注射乳剂的研究
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摘要
本文建立了羟基喜树碱的体外HPLC分析方法，测定了羟基喜树碱在正辛醇／水中的表观分配系数及在不同pH值下内酯形式与羧酸形式转变后的百分比，为羟基喜树碱静脉注射乳剂处方设计、含量测定及体内分析方法的建立提供了依据。
以大豆卵磷脂(SPC)和泊洛沙姆F68为乳化剂，注射用大豆油为油相，采用超声分散-过纳米机法制备了羟基喜树碱静脉注射乳剂，并以乳剂的外观、离心稳定性及灭菌稳定性为指标，在单因素考察的基础上，采用正交设计优化处方，确定了羟基喜树碱静脉注射乳剂的处方和制备工艺。建立了羟基喜树碱静脉注射乳剂的含量分析方法，日内和日间精密度的标准偏差均小于2.5％，平均回收率为94.5％，说明该方法测定含量准确度高。
考察了羟基喜树碱静脉注射乳剂的理化性质。乳剂粒度分布呈为单峰分布，平均粒径约为399.6nm，无粒径大于1μm的粒子。乳剂ζ电位平均值为-26.4mV，pH平均值为6.50。羟基喜树碱静脉注射乳剂与5％葡萄糖注射液配伍良好而与生理盐水配伍稳定性差，用5％葡萄糖稀释20倍后渗透压平均值为297mmol／kg，基本与血浆等渗；无刺激性，无溶血，满足静脉注射制剂要求。利用Sephadex G-50凝胶过滤色谱法，将乳滴中的药物和游离于外水相中的药物分离，计算非游离型药物占制剂中药物含量的78％。由羟基喜树碱静脉注射乳剂和羟基喜树碱钠盐注射液在磷酸盐缓冲液(PBS)与血浆中内酯环的稳定性实验可以看出乳剂可以显著的延缓药物的水解，对药物起到保护作用。乳剂的释放实验验证了羟基喜树碱与血浆蛋白有较强的结合力。
以外观色泽、药物含量、pH值和粒度分布为指标，考察了羟基喜树碱静脉注射乳剂在影响因素实验、加速实验和长期实验中的稳定性，结果表明羟基喜树碱静脉注射乳剂稳定性较好，但对光不稳定，宜避光贮存。
大鼠尾静脉注射给药后用HPLC法测定了血浆中不同时间点的羟基喜树碱浓度，并与羟基喜树碱钠盐注射液进行了比较。药物动力学实验结果表明，两种制剂均符合双隔室模型，T_(1/2(a))分别为7.910min和8.938min，T_(1／2(β))分别为62.481min和87.133min，AUC分别为40.698(μg／mL)·min和39.096(μg／mL)·min。
小鼠尾静脉注射给药后，利用HPLC法测定了各组织中不同时间点的羟基喜树碱浓度，并与羟基喜树碱钠盐注射液进行了比较。组织分布实验结果表明，乳剂中的羟基喜树碱在小鼠肝、脾、肺、肾中的滞留时间及药物量均有所增加，其中以肝组织增加最显著，说明羟基喜树碱制成乳剂后可提高药物肝靶向性。
The method of high performance liquid chromatography (HPLC) was set up to determine the concentration of 10-hydroxyeamptothecin(HCPT). The apparent partition coefficient of HCPT in octanol/distilled water was determined. HCPT was dissolved in PBS of varying pH and the ratio of lactone form HCPT was quantified by HPLC. It will direct the preformulation, assaying and analytical method in vivo.
The oily phase was soybean oil and Soya lecithin and poloxamer-188 were used as emulsifier, so the HCPT intraeenous injection emulsions were prepared by through nanomizer system after ultrasonic dispersion. The orthogonal design was adopted to obtain the optimized prescription and preparation, based on single-factor investigation. The appearance and the stability after centrifuging and sterilizing were as the evaluated index. Another method was established to determine the content of HCPT in the emulsions. The precision and recovery was fit for the standard, so the method was feasible.
The physicochemical properties of HCPT emulsions were studied. The particle size distribution displayed single-peaked and no particle was larger than 1μm. The mean droplet size and zeta potential of the emulsions were 399.6nm and-26.4mV, respectively. The pH of the emulsions was 6.50. They were steady with 5% Glucose, but unsteady with physiological saline. Irritable and hemolytic experiments showed the preparation was fit for the regulations of the I.V. Sephadex G-50 filtration chromatography was employed to separate the free drug from HCPT emulsions. The HCPT incorporated into the emulsion was 78%. The stability of lactone ring in the HCPT emulsions was a bit more than that of in HCPT injection, both in PBS and human plasma. It was indicated that HCPT emulsions could delay the hydrolytic rate of HCPT and protect the lactone ring. Release curse illustrated HCPT was set free from the emulsion sufficiently and it also verified that HCPT had strong affinity to serum albumin.
The results of influencing factor tests explained that the HCPT emulsions should be tightly preserved from light. In the long-term testing and acceleration testing, the content of HCPT emulsion was also stable in six months.
After the rats were administrated HCPT injection and emulsions intravenously, the drug concentrations in plasma were monitored by HPLC. The pharmacokinetic processes in rats for the two preparations were both in accorde with double compartmental models. The distribution half-life was 7.910min and 8.938min and the elimination half-life was 62.481min and 87.133min individually. The AUC were 40.698(μg/mL)·min and 39.096 (μg/mL). min, separately.
After the mice were administrated HCPT Injection and emulsions intravenously, the drug concentrations in different tissues were detected by HPLC. The levels of HCPT in spleen, lung and kidney in emulsions groups were slightly higher than that of injection groups. Especially, the concentration of drug in liver increased greatly compared with that of injection groups.
以大豆卵磷脂(SPC)和泊洛沙姆F68为乳化剂，注射用大豆油为油相，采用超声分散-过纳米机法制备了羟基喜树碱静脉注射乳剂，并以乳剂的外观、离心稳定性及灭菌稳定性为指标，在单因素考察的基础上，采用正交设计优化处方，确定了羟基喜树碱静脉注射乳剂的处方和制备工艺。建立了羟基喜树碱静脉注射乳剂的含量分析方法，日内和日间精密度的标准偏差均小于2.5％，平均回收率为94.5％，说明该方法测定含量准确度高。
考察了羟基喜树碱静脉注射乳剂的理化性质。乳剂粒度分布呈为单峰分布，平均粒径约为399.6nm，无粒径大于1μm的粒子。乳剂ζ电位平均值为-26.4mV，pH平均值为6.50。羟基喜树碱静脉注射乳剂与5％葡萄糖注射液配伍良好而与生理盐水配伍稳定性差，用5％葡萄糖稀释20倍后渗透压平均值为297mmol／kg，基本与血浆等渗；无刺激性，无溶血，满足静脉注射制剂要求。利用Sephadex G-50凝胶过滤色谱法，将乳滴中的药物和游离于外水相中的药物分离，计算非游离型药物占制剂中药物含量的78％。由羟基喜树碱静脉注射乳剂和羟基喜树碱钠盐注射液在磷酸盐缓冲液(PBS)与血浆中内酯环的稳定性实验可以看出乳剂可以显著的延缓药物的水解，对药物起到保护作用。乳剂的释放实验验证了羟基喜树碱与血浆蛋白有较强的结合力。
以外观色泽、药物含量、pH值和粒度分布为指标，考察了羟基喜树碱静脉注射乳剂在影响因素实验、加速实验和长期实验中的稳定性，结果表明羟基喜树碱静脉注射乳剂稳定性较好，但对光不稳定，宜避光贮存。
大鼠尾静脉注射给药后用HPLC法测定了血浆中不同时间点的羟基喜树碱浓度，并与羟基喜树碱钠盐注射液进行了比较。药物动力学实验结果表明，两种制剂均符合双隔室模型，T_(1/2(a))分别为7.910min和8.938min，T_(1／2(β))分别为62.481min和87.133min，AUC分别为40.698(μg／mL)·min和39.096(μg／mL)·min。
小鼠尾静脉注射给药后，利用HPLC法测定了各组织中不同时间点的羟基喜树碱浓度，并与羟基喜树碱钠盐注射液进行了比较。组织分布实验结果表明，乳剂中的羟基喜树碱在小鼠肝、脾、肺、肾中的滞留时间及药物量均有所增加，其中以肝组织增加最显著，说明羟基喜树碱制成乳剂后可提高药物肝靶向性。
The method of high performance liquid chromatography (HPLC) was set up to determine the concentration of 10-hydroxyeamptothecin(HCPT). The apparent partition coefficient of HCPT in octanol/distilled water was determined. HCPT was dissolved in PBS of varying pH and the ratio of lactone form HCPT was quantified by HPLC. It will direct the preformulation, assaying and analytical method in vivo.
The oily phase was soybean oil and Soya lecithin and poloxamer-188 were used as emulsifier, so the HCPT intraeenous injection emulsions were prepared by through nanomizer system after ultrasonic dispersion. The orthogonal design was adopted to obtain the optimized prescription and preparation, based on single-factor investigation. The appearance and the stability after centrifuging and sterilizing were as the evaluated index. Another method was established to determine the content of HCPT in the emulsions. The precision and recovery was fit for the standard, so the method was feasible.
The physicochemical properties of HCPT emulsions were studied. The particle size distribution displayed single-peaked and no particle was larger than 1μm. The mean droplet size and zeta potential of the emulsions were 399.6nm and-26.4mV, respectively. The pH of the emulsions was 6.50. They were steady with 5% Glucose, but unsteady with physiological saline. Irritable and hemolytic experiments showed the preparation was fit for the regulations of the I.V. Sephadex G-50 filtration chromatography was employed to separate the free drug from HCPT emulsions. The HCPT incorporated into the emulsion was 78%. The stability of lactone ring in the HCPT emulsions was a bit more than that of in HCPT injection, both in PBS and human plasma. It was indicated that HCPT emulsions could delay the hydrolytic rate of HCPT and protect the lactone ring. Release curse illustrated HCPT was set free from the emulsion sufficiently and it also verified that HCPT had strong affinity to serum albumin.
The results of influencing factor tests explained that the HCPT emulsions should be tightly preserved from light. In the long-term testing and acceleration testing, the content of HCPT emulsion was also stable in six months.
After the rats were administrated HCPT injection and emulsions intravenously, the drug concentrations in plasma were monitored by HPLC. The pharmacokinetic processes in rats for the two preparations were both in accorde with double compartmental models. The distribution half-life was 7.910min and 8.938min and the elimination half-life was 62.481min and 87.133min individually. The AUC were 40.698(μg/mL)·min and 39.096 (μg/mL). min, separately.
After the mice were administrated HCPT Injection and emulsions intravenously, the drug concentrations in different tissues were detected by HPLC. The levels of HCPT in spleen, lung and kidney in emulsions groups were slightly higher than that of injection groups. Especially, the concentration of drug in liver increased greatly compared with that of injection groups.
引文
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