摘 要
目的：建立以气相谱法同时测定常用药用辅料吐温80中残留溶剂二氯甲烷、、三氯乙烯的含量以及吐温80中脂肪酸组成分析的方法，以提高吐温80的质量标准及使用安全性；建立以高效液相谱法测定中药生脉注射液中辅料吐温80含量的方法。 方法：药用辅料吐温80中残留溶剂二氯甲烷、、三氯乙烯的含量以及脂肪酸组成分析及中药生脉注射液中辅料吐温80含量测定的方法如下：（1） 吐温80中二氯甲烷、、三氯乙烯的含量测定：谱柱为AT-1000（30 m × 0.32 mm × 0.25 μm）弹性石英毛细管柱。采用程序升温的方法，初温
℃－1的速度升温至200 ℃，保持17 min。进样口温40 ℃保持10 min，以50·min
∶，进样量1 μL，柱流量恒定，流度200 ℃，检测器温度250 ℃，分流比为201
速2 mL·min－1。顶空平衡温度80 ℃，平衡时间45 min。
（2） 吐温80中脂肪酸的组成分析：谱柱为：AT-Wax（30 m × 0.32 mm × 0.25 μm）弹性石英毛细管柱。采用程序升温的方法，初温140 ℃保持0 min，以2 ℃·min －1的速度升温至190 ℃，保持10 min，再以10 ℃·min－1的速度升温至245 ℃，保持20 min。进样口温度260 ℃，检测器温度270 ℃，分流比为10∶1，进样量1 μL，柱压力恒定，柱流速3 mL·min－1。
（3） 中药生脉注射液中辅料吐温80的含量测定：谱柱：TSKgel G3000
SW XL(7.8 mm × 30 cm,5 μm)；流动相：20 mmo l·L-1醋酸铵-乙腈（90:10），流速：0.6 mL·min-1；柱温：30 ℃。ESLD检测条件：漂移管温度110 ℃，氮气流速：2.3 L·min-1；进样体积10 μL。
结果：
（1） 二氯甲烷、及三氯乙烯的线性范围分别为49.9~2395.2 μg/mL
挠度计算公式（r=0.9999），5.3~255.3 μg/mL（r=0.9998）及9.1~108.6 μg/mL（r=0.9950），平均回收率分别为98.3%（RSD＝8.34％），96.5%（RSD＝9.58％）及96.1%（RSD ＝9.30％）。
（2） 采用面积归一化法定性并定量分析吐温80中的脂肪酸组成。所建立的分析方法测得各组分含量分别为十四烷酸甲酯0.54%，棕榈酸甲酯0.82%，棕榈酸油酸甲酯1.24%，硬脂酸甲酯0.18%，油酸甲酯97.22%，亚油酸甲酯未检出，亚麻酸甲酯未检出。十四烷酸甲酯、棕榈酸甲酯、棕榈油酸甲酯、硬脂酸甲酯、油酸甲酯、亚油酸甲酯、亚麻酸甲酯分离度均大于1.5；十四烷酸甲酯、棕榈酸甲
酯、棕榈油酸甲酯、硬脂酸甲酯、油酸甲酯的RSD%分别为1.77%，1.97%，1.98%，1.94%，2.00%，亚油酸甲酯及亚麻酸甲酯均未检出。
高行建（3） 吐温80在1～25 mg·mL-1范围内呈良好的线性关系，线性方程为
Y=1.4471X+2.6408 ，r=0.9989，RSD为1.48%。信噪比（S/N）为3时，吐温80的最低检出限为0.10 mg·mL-1。低、中、高浓度的平均加样回收率（n=3）分别为97.78%（RSD＝0.20％）、97.71%（RSD＝0.25％）、100.84%（RSD＝0.65％）。
质量对企业的重要性结论：论文建立了以气相谱法测定常用药用辅料吐温80中残留溶剂二氯甲烷、、三氯乙烯的含量以及吐温80中脂肪酸组成分析的方法，方法简便、快速，结果准确、可靠，重复性好。方法的建立对吐温80的质量标准的提高提供了一定的参考。论文同时建立了以高效液相谱法测定中药生脉注射液中辅料吐温80含量的方法，方法简便、准确快速，可用于中药生脉注射液中辅料吐温80的含量测定。
关键词：吐温80，二氯甲烷，，三氯乙烯，溶剂残留，脂肪酸组成，生脉注射液
Abstract
Objective: To establish a GC method for the determination of methylene chloride, trichloromethane, trichloroethylene and the composition of fatty acid in tween 80. And to establish a RP－HPLC method applicable to determine the content of tween 80 in traditional Chinese medicine Sheng mai injection.
Methods:The Methods of the determination of methylene chloride, trichloromethane, trichloroethylene and the composition of fatty acid, and the method of the analysis of tween 80 in traditional Chinese medicine Sheng mai injection were as follows:
中国课堂教学网植兰(1)The analysis of methylene chloride, trichloromethane and trichloroethylene in Tween 80 was carried out on an AT-1000 capillary gas chromatography column（30 m×0.32 mm×0.25 μm）, the temperature of injection port and detector were 200 ℃and 250 ℃, respectively. Maintain the temperature of the column at 40 ℃ for 5 minutes, then raise the temperature at a rate of 50 ℃·min-1 to 200 ℃ for 17 minutes. The Split ratio is 20:1, the volume of injection is 1 μL and the rate of flow is 2 mL·min－1.
(2) The analysis of the composition of fatty acid in Tween 80 was carried out on an AT-Wax capillary gas chromatography column（30 m × 0.32 mm × 0.25 μm）, the temperature of injection port and detector were 260 ℃ and 270 ℃, respectively. Maintain the temperature of the column at 140 ℃ for 0 minutes, raise the temperature at a rate of 2 ℃·min-1 to 190 ℃ for 10 minutes, then raise the temperature at a rate of 10 ℃·min-1 to 245 ℃ for 20 minutes. The Split ratio is 10:1, the volume of injection is 1 μL and the rate of flow is 3 mL·min－1.
(3)Method of the determination of Tween 80 in traditional Chinese medicine injection: The chromatographic analysis were carried out on a TSKgel G3000 SW XL column(7.8 mm×30 cm,5 μm), and the mobile phase consisted of 20 mmol·L-1 ammon -ium acetate and acetonitrile (90:10). Flow rate was 0.6mL·min-1, and the column temperature was 30 ℃.The flow rate N2 in ELSD was 2.3 L·min-1.The temperature of drift tube was 110 ℃.
Results: (1) A good resolution was obtained by this method between the peaks due to methylene chloride, trichloromethane and trichloroethylene. The linearity equation of methylene chloride was Y=0.5379X-14.344(r=0.9999), the average
recovery was 98.3%, the precision was 8.34%. The linearity equation of trichloro -methane was Y=0.2118X-0.3240 (r=0.9998), the average recovery was 96.5%, the precision was 9.58%. The linearity equation of trichloroethylene was Y=0.91327X- 3.8996 (r=0.9950), the average recovery was 96.1%, the precision was 9.30%.
(2) The resolution of methyl myristate, methyl palmitate,methyl palmitoleate, methyl stearate, methyl oleate, methyl linoleate and methyl linolenate were all more than 1.5. The precision of methyl myristate , methyl palmitate,methyl palmitoleate, methyl stearate, methyl oleate was 1.77%, 1.97%,
1.98%, 1.94%, 2.00%. Methyl linoleate and methyl linolenate were not detected. And the content of methyl myristate , methyl palmitate,methyl palmitoleate, methyl stearate, methyl oleate was 0.54%, 0.82%, 1.24%, 0.18%, 97.22%. Methyl linoleate and methyl linolenate were not detected.
(3)A good resolution was obtained by this method between the peaks due to Tween 80. The linearity equation of methylene chloride was Y=1.4471X+2.6408 (r=0.9989), the precision was 1.48%,and the linear range was 1～25 mg·mL-1. Signal to noise ratio (S / N) is 3, the minimum detection limit of tween 80 was 0.10 mg • mL-1. The average recoveries of low, medium and high concentrations(n = 3), were 97.78%, 97.71%, 100.84%.
Resolution: Methods of GC were established for the determination of methylene chloride, trichloromethane, trichloroethylene and the composition of fatty acid in tween 80 in the thesis.The method is simple, rapid, accurate, reliable and repeatable. The establishment of the method provides a reference for the improvement of quality standards of tween 80 in Chinese Pharmacopoeia of 2010. The method of HPLC to determine the content of tween 80 in traditional Chinese medicine Sheng mai injection was also established.The method is simple, accurate and fast, which can be used in the detection of the content of tween 80 in traditional Chinese medicine Sheng mai injection.
Key words：polysorbate 80, methylene chloride, trichloromethane, trichloroethy -lene, residual solvent, the composition of fatty acid, sheng mai injection
目录
第一章 前言 (1)
1.1 吐温80简介 (1)
1.2 吐温80的合成工艺 (1)
1.3 吐温80的药理及毒理活性 (3)
1.3.1 急性超敏反应 (3)
1.3.2 外周神经毒性 (3)
1.3.3 抑制P2糖蛋白活性 (3)
1.3.4 内在抗肿瘤效应 (3)
1.3.5 肝毒性及其它 (3)
1.4 吐温80引起不良反应可能的因素 (4)
1.4.1 吐温80中含有的有机挥发性杂质 (4)
1.4.2 吐温80中含有的有机不挥发性杂质 (4)
1.4.3 吐温80的氧化降解 (4)
1.5 吐温80在实际应用中存在的问题 (4)
介休话1.6 各国药典对吐温80的标准规定 (5)
1.7 本文的研究内容 (6)
第二章 实验部分 (8)
2.1 二氯甲烷、、三氯乙烯残留量测定的方法学考察 (8)
2.1.1 仪器与试剂 (8)
2.1.2 方法与结果 (8)
2.2脂肪酸组成测定方法学考察 (11)
2.2.1 仪器与试剂 (11)
2.2.2 方法与结果 (11)
2.3 中药注射液中吐温80含量的测定 (13)
2.3.1 仪器与试剂 (13)
2.3.2 谱条件 (13)
2.3.3 方法学考察 (13)
2.3.4 样品中吐温80的测定 (15)
第三章 总结与展望 (16)
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