中国科技论文统计源期刊 中文核心期刊  
美国《化学文摘》《国际药学文摘》
《乌利希期刊指南》
WHO《西太平洋地区医学索引》来源期刊  
日本科学技术振兴机构数据库(JST)
第七届湖北十大名刊提名奖  
医药导报, 2019, 38(5): 641-645
doi: 10.3870/j.issn.1004-0781.2019.05.024
柱前衍生化反相高效液相色谱法测定去水卫矛醇的含量*
Determination of Dianhydrogalactitol by Pre-column Derivatization and RP-HPLC
王杏利1,2,, 孙丽涵1,3, 张雷4, 吴华晶1,3, 张峻颖2,, 吴春勇1,3,

摘要:

目的 建立测定去水卫矛醇含量的柱前衍生化反相高效液相色谱法(RP-HPLC)。方法 去水卫矛醇经二乙基二硫代氨基甲酸钠衍生化后进行RP-HPLC分析,采用Welch Ultimate XB-CN色谱柱(250 mm×4.6 mm,5 μm),流动相为乙腈-水(50:50),流速为 1.0 mL∙min-1,检测波长为 278 nm。结果 40 ℃下衍生化反应可在30 min内完成,去水卫矛醇衍生物在10 h内稳定。去水卫矛醇在0.100~20.0 μg∙mL-1范围内线性关系良好(r=0.999 8)。检测限(LOD)和定量限(LOQ)分别为30和100 ng·mL-1。方法专属性、准确度和精密度均符合要求。结论 该方法快速、准确、灵敏,具有良好的专属性和重复性,可用于去水卫矛醇的质量控制。

关键词: 去水卫矛醇 ; 柱前衍生化 ; 色谱法,反相高效液相 ; 含量测定

Abstract:

Objective To establish a reversed-phase high performance liquid chromatographic (RP-HPLC) method with pre-column derivatization for determination of dianhydrogalactitol. Methods After dianhydrogalactitol was derivatized with sodium diethyldithiocarbamate, the RP-HPLC separation was performed on a Welch Ultimate XB-CN column (250 mm×4.6 mm,5 μm) with the mixture of acetonitrile-water (50:50) as the mobile phase. The flow rate was 1.0 mL∙min-1 and the detection wavelength was set at 278 nm. Results The derivatization reaction could be finished within 30 min at 40 ℃ and the derivative was stable within 10 h. The calibration curve was linear over the range of 0.100-20.0 μg·mL-1 (r=0.999 8). The limit of detection (LOD) and limit of quantification (LOQ) was 30 and 100 ng·mL-1, respectively. The method showed good specificity, accuracy and precision. Conclusion The proposed method was fast, accurate and sensitive with high specific and repeatability. The study provides a new approach for the quality control of dianhydrogalactitol.

Key words: Dianhydrogalactitol ; Pre-column derivatization ; Reversed-phase high performance liquid chromatography ; Content determination

去水卫矛醇(dianhydrogalactitol)是以卫矛科植物蜜花美登木中提取的卫矛醇为原料,经溴化、消除等反应得到的细胞周期非特异性抗肿瘤药物[1],可在肿瘤组织中优先蓄积[2],具有毒性小的优点[3],对慢性粒细胞白血病及多种肿瘤均有良好疗效[4,5]。除单独使用外,去水卫矛醇还可与其他药物联合使用,并发挥协同作用[6]。去水卫矛醇具有中枢神经系统活性,可跨血-脑屏障进入脑部[7],使胶质瘤细胞增殖停滞在细胞周期的G2/M期,从而抑制其生长[8]。作为一种潜在的多靶点药物,去水卫矛醇作为治疗胶质瘤的孤儿药已在美国通过了Ⅱ期临床试验[9],具有较好的应用前景。

鉴于良好的临床应用前景,去水卫矛醇的新药开发和质量控制显得尤为重要。该品种原料[10]及制剂[11]的国家标准均采用滴定分析法测定含量,虽简便易行,但专属性较差。魏宁漪等[12]采用气相色谱(GC)法测定注射用去水卫矛醇的含量,但笔者在重现实验时发现药物在气化过程中存在降解的风险。吴先富等[13]采用磁共振法测定去水卫矛醇原料含量,但该法对仪器要求较高,应用受到较大限制。高效液相色谱法(HPLC)具有分析速度快、灵敏度高、专属性强的优点,是目前药品质量控制中最常用的方法之一。去水卫矛醇的化学结构式中缺乏生色团,无法采用紫外检测器测定其有关物质和含量,因此考虑采用柱前衍生化的方法改善其紫外吸收特性。二乙基二硫代氨基甲酸钠(DDTC)是一种便宜易得的衍生化试剂,可与环氧基反应生成稳定的具有紫外吸收的产物。笔者采用DDTC作为柱前衍生化试剂,选择使用广泛的紫外检测器,探索建立测定去水卫矛醇含量的衍生化反相HPLC法,并利用液相色谱-串联质谱联用技术(LC-MS/MS)对衍生物进行结构确认。

1 仪器与试药
1.1 仪器

日本岛津高效液相色谱仪(含二元高压输液泵、自动进样器、柱温箱、紫外检测器及Labsolution工作站);安捷伦6420 HPLC-MS/MS三重四级杆液质联用系统(含安捷伦1260 Infinity高效液相色谱仪、自动进样器、电喷雾离子化接口、四级杆质谱检测器、MassHunter数据分析处理软件);BT25-S型电子分析天平(赛多利斯科学仪器有限公司,感量:0.01 mg);XW-80A旋涡混合器(上海医大仪器有限公司);HH-W三用恒温水箱(常州市国立试验设备研究所);Mini-14K微型高速离心机(杭州奥盛仪器有限公司);ZLS-1真空离心浓缩仪(湖南赫西仪器装备有限公司)。

1.2 试剂

去水卫矛醇原料(批号170601)和对照品由广西梧州制药(集团)股份有限公司提供。乙腈(色谱纯,美国天地公司);DDTC(分析纯,上海阿拉丁生化科技股份有限公司);实验用水为去离子纯化水;其余试剂均为分析纯。

2 方法与结果
2.1 色谱条件

色谱柱为Welch Ultimate XB-CN(250 mm×4.6 mm,5 μm),流动相为乙腈-水(50:50),流速为1.0 mL∙min-1,柱温为30 ℃,检测波长为278 nm,进样量为20 μL。

2.2 质谱条件

电喷雾离子源,正离子检测模式,离子扫描范围m/z 100~500,干燥气温度350 ℃,干燥气流速9 L∙min-1,雾化气压力103.425 kPa,喷雾电压4500 V。二级质谱采用产物离子扫描模式,裂解电压110 V,碰撞能20 eV。

2.3 溶液的制备

2.3.1 供试品溶液和对照品溶液 取去水卫矛醇原料和对照品10 mg,精密称定,分别置100 mL量瓶中,加水溶解并稀释至刻度,摇匀,精密量取1 mL,置10 mL量瓶中,用水稀释至刻度,摇匀,作为供试品溶液和对照品溶液。

2.3.2 衍生化试剂溶液 精密称取DDTC适量,加水溶解并定量稀释制得浓度为5 %的衍生化试剂溶液。

2.3.3 磷酸盐缓冲液(pH 值7.0) 取三水合磷酸氢二钾约0.439 g及磷酸二氢钾0.419 g,精密称定,置50 mL量瓶中,加水溶解并稀释至刻度,摇匀。

2.4 衍生化方法

精密量取去水卫矛醇对照品溶液和供试品溶液各400 μL,依次精密加入磷酸盐缓冲液(pH值7.0)400 μL和衍生化试剂溶液200 μL,涡旋30 s,40 ℃水浴反应30 min后,冰水浴5 min终止反应。精密加入乙酸乙酯2 mL,涡旋3 min萃取衍生物,离心5 min,精密量取上清液1.8 mL,用氮气吹干后,精密加入流动相800 μL复溶,进行HPLC分析。

2.5 色谱方法的建立

2.5.1 色谱柱的选择 笔者在本实验比较了Hypersil-CN(250 mm×4.6 mm,5 μm)、Ultimate XB-CN(250 mm×4.6 mm,5 μm)、Ultimate XB-C18(250 mm×4.6 mm,5 μm)和Hedera ODS-2(250 mm×4.6 mm,5 μm)等不同品牌及填料的色谱柱对分析的影响,发现在Welch Ultimate XB-CN色谱柱上,分析物峰形对称且保留时间合适。

2.5.2 检测波长的选择 精密量取去水卫矛醇对照品溶液适量,按“2.4”项处理后,用流动相溶解并稀释制成适宜浓度的溶液,于200~350 nm波长范围内扫描紫外吸收光谱图(图1)。结果表明,去水卫矛醇衍生物在278 nm处有最大吸收。

图1 去水卫矛醇衍生物的紫外吸收光谱

Fig.1 Ultraviolet absorption spectrum of dianhydroga-lactitol derivative

2.5.3 流动相的选择 实验考察了甲醇-水和乙腈-水系统,发现采用乙腈-水系统作为流动相时,基线稳定且峰形尖锐,经系统摸索,最终确定流动相为乙腈-水(50:50,V/V)。

2.6 去水卫矛醇衍生物的鉴定

取去水卫矛醇对照品溶液,按“2.4”项处理后,进行LC-ESI-MS/MS分析,特征碎片离子以及质谱裂解规律见图2。结果表明,[M+H]+准分子离子峰为445.1,与衍生物中含偶数氮的情况相符,特征碎片离子包括m/z 116.1和427.2,因此鉴定该色谱峰对应的成分为去水卫矛醇衍生物。去水卫矛醇与DDTC的反应示意图见图3。

图2 去水卫矛醇衍生物的特征碎片离子谱

Fig.2 Ion mass spectra of distinctive product dianhydrogalactitol derivative

图3 去水卫矛醇与DDTC的化学衍生化反应示意图

Fig.3 Reaction pathway of derivatization of dianhydrogalactitol and DDTC

2.7 方法学考察

2.7.1 专属性实验 空白干扰实验:分别取空白溶剂、去水卫矛醇对照品溶液和供试品溶液,按“2.4”项操作后,进行HPLC分析,典型的色谱图见图4。结果表明,在本色谱条件下,去水卫矛醇衍生物峰形良好,保留时间约为5.7 min,空白溶剂对主峰的测定无干扰。

图4 衍生化去水卫矛醇的典型高效液相色谱图
A.空白溶液;B.对照品溶液;C.供试品溶液;1.衍生物

Fig.4 Representative HPLC chromatograms of dianhydrogalactitol derivative
A.blank solution;B.reference solution;C.sample solution;1.derivative

强制降解研究:取去水卫矛醇10 mg,精密称定,置100 mL量瓶中,加水溶解并稀释至刻度,摇匀,精密量取1 mL,置10 mL量瓶中,分别进行酸破坏(0.1 mol·L-1盐酸溶液1 mL,室温2 h)、碱破坏(0.1 mol·L-1氢氧化钠溶液1 mL,室温2 h)、氧化破坏(3%过氧化氢溶液1 mL,室温1 h)、高温破坏(60 ℃ 2 h)和光照破坏(紫外灯下2 h),用水稀释至刻度,摇匀,按“2.4”项处理后,进行HPLC分析,典型色谱图见图5。去水卫矛醇在酸和氧化条件下有明显降解,而在其他条件下较为稳定。本品进行制剂工艺改进、药效、药动学或毒理学研究时应注意药物的稳定性。

图5 去水卫矛醇强制降解实验典型色谱图
A.酸破坏 ;B.碱破坏 ;C.氧化破坏;D.高温破坏;E.光照破坏;F.未破坏;1.衍生物

Fig.5 Reprehensive HPLC chromatograms of forced degradation study on dianhydrogalactitol
A.acid degradation;B.alkaline degradation;C.oxidation degradation;D.heat degradation;E.light degradation;F.sample without destruction;1.derivative

2.7.2 检测限及定量限 精密称取对照品适量,用水溶解并定量稀释制成不同浓度的溶液,衍生化处理后进行HPLC分析,去水卫矛醇检测限(S/N=3)和定量限(S/N=10)分别为30和100 ng·mL-1(n=5)。

2.7.3 线性关系考察 精密称取对照品适量,加水溶解并定量稀释制成浓度分别为0.100,0.500,1.00,2.00,5.00,10.0及20.0 μg∙mL-1的系列标准溶液,按“2.4”项处理后,进行HPLC分析,以浓度(C)为横坐标,峰面积(A)为纵坐标进行线性回归。结果表明,去水卫矛醇在0.100~20.0 μg·mL-1范围内线性良好,典型回归方程为A=38 725C+755.19(r=0.999 8)。

2.7.4 加样回收率 精密称取已测定含量的去水卫矛醇原料约10 mg,置100 mL量瓶中,加水溶解并稀释至刻度,摇匀,精密量取1 mL,置20 mL量瓶中,分别精密加入质量浓度为100 μg·mL-1的去水卫矛醇对照品溶液0.6,1.0,1.4 mL,用水稀释至刻度,摇匀,作为浓度水平80%,100%和120%的回收率溶液,每个浓度水平配制3份。按“2.4”项处理后,进行HPLC分析。结果见表1,加样回收率为98.2%~101.3%,方法准确度良好。

表1 去水卫矛醇加样回收率结果
Tab.1 Results of recovery test of dianhydrogalactitol
浓度水平 原有量 加入量 测得量 回收率 RSD
μg %
99.8 60.1 159.5 99.0 0.97
99.9 60.1 158.9 98.2
100.0 60.1 160.1 100.0
100.1 100.1 198.9 98.7 1.31
100.2 100.1 199.3 99.0
99.9 100.1 201.1 101.1
100.0 140.1 240.6 100.4 0.90
99.9 140.1 241.8 101.3
100.1 140.1 239.5 99.5

表1 去水卫矛醇加样回收率结果

Tab.1 Results of recovery test of dianhydrogalactitol

2.7.5 精密度实验 供试品溶液经衍生化反应后,按上述色谱条件连续进样6次,峰面积的RSD为1.15%,表明进样精密度良好。

2.7.6 重复性实验 配制6份供试品溶液,按“2.4”项处理后,进行HPLC分析。结果表明,6份样品含量RSD为1.44%,重复性良好。

2.7.7 稳定性实验 将供试品溶液按“2.4”项处理后,分别于0,2,4,6,10 h进行HPLC分析,与0h比较,不同时间峰面积在0 h峰面积的99.5%~100.8%范围内,去水卫矛醇衍生物在10 h内稳定性良好。另将供试品溶液在室温下放置不同时间后,进行衍生化HPLC分析,不同时间测定的含量在98%~102%,表明去水卫矛醇水溶液在室温至少可放置4 h。

2.8 含量测定

取对照品溶液及供试品溶液,按“2.4”项处理后,进行HPLC分析,典型色谱图见图4。按外标法计算,样品的平均含量为100.5%。

3 讨论

去水卫矛醇具有环氧环的特征结构,可与DDTC发生衍生化反应,生成具有紫外吸收的衍生物,用广泛使用的紫外检测器检测。笔者考察不同温度(25,40,60 ℃)、加热时间(5,15,30,45,60,90和120 min)、衍生化试剂浓度(1%,2%,3%,5%和10%)以及不同反应体系pH值(6.0,7.0和8.0),发现药物与5% DDTC在pH 值7.0条件下,于40 ℃水浴加热30 min可衍生化完全。为了除去过量的衍生化试剂,避免其干扰衍生物的测定,实验比较了乙酸乙酯、三氯甲烷、二氯甲烷和乙醚等试剂的萃取效率,发现乙醚和乙酸乙酯的效果最好,考虑到操作的简便性和安全性,最终选择乙酸乙酯作为萃取剂。

笔者建立了衍生化HPLC法测定去水卫矛醇含量,该方法线性范围宽,重复性好,稳定性高,简单易行,易于在普通实验室操作,不仅可用于去水卫矛醇原料的质量控制,还可用于去水卫矛醇制剂和生物样品的分析,对去水卫矛醇的新药开发、生产和临床应用具有重要的参考价值。

The authors have declared that no competing interests exist.

参考文献

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【目的】探寻二去水卫矛醇(1,2:5,6-Dianhydrogalctitol,DAG)对人脑肿瘤细胞的抑制作用,为DAG抗肿瘤作用提供理论基础。【方法】通过体外实验,并采用噻唑蓝(MTT)比色法检测不同浓度DAG对11株人脑肿瘤细胞的体外抑制作用,观察DAG的半数抑制浓度。【结果】DAG对IMR-32、SH-SY5Y、U87、A172、BT325、T98G、SHG-44、U373、C6、SK-N-BE、U251等11株人脑肿瘤细胞作用72h后半数抑制浓度(72h-IC50)分别为24.28μg/mL、19.24μg/mL、21.55μg/mL、25.13μg/mL、20.63μg/mL、21.03μg/mL、24.91μg/mL、23.52μg/mL、22.06μg/mL、22.92μg/mL、23.44μg/mL。【结论】DAG在体外对上述11株人脑细胞均有抑制生长作用。
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[4] LIN B J,TAO R F.New drugs of dianhydrogalactitol to chronic granulocyte leukemia[J].New Medicine,1980,11(12):619-620.
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[5] 张慧玲,王稼农,梁霜,.二去水卫矛醇对四种肿瘤细胞的体外抑制作用[J].广西科学,2013,20(1):82-84.
采用噻唑蓝(MTT)比色法检测不同浓度二去水卫矛醇(DAG)对人胃癌细胞SGC-7901、人肺癌细胞H460、人鼻咽癌细胞CNE和人肝癌细胞BEL-7404的抑制率研究DAT对4种肿瘤细胞的体外抑制作用。结果表明,DAG对4种癌细胞:CNE、H460、SGC-7901和BEL-7404有较强的抑制作用,作用72h后半数抑制浓度(72h-IC50)分别为4.12μg·ml-1,15.98μg·ml-1,16.00μg·ml-1和16.73μg·ml-1。显微镜下可见经药物作用后细胞圆缩、胞质浓缩、胞浆内颗粒集中边缘化甚至细胞溶解等损伤现象。DAG在体外对上述4种癌细胞有明显的抑制生长作用。
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[6] 樊亦军,韩锐,周军,.抗癌药1,2:5,6-二去水卫矛醇与长春新碱联合用药的实验研究[J].药学学报,1987,22(2):98-102.
去水卫矛醇(DAG)与长春新碱(VCR)联合使用,对小鼠淋巴细胞白血病L_(1210)有协同疗效,疗效强度决定于给药方案和剂量。先给DAG,24 h后给VCR,协同疗效最高。DAG剂量是影响疗效的主要因素,最佳剂量为3.0 mg/kg。琼脂扩散合试验亦证明二药有方案依赖性协同作用。二药合用对小鼠骨髓干细胞仅有微弱的协同杀灭作用。
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[7] SUN L,WU H,REN L,et al.Development and validation of a highly sensitive LC-MS/MS method for determination of brain active agent dianhydrogalactitol in mouse plasma and tissues:Application to a pharmacokinetic study [J].J Chromatogr B Analyt Technol Biomed Life Sci,2018,1087-1088(11):90-97.
A sensitive and specific liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method was developed and validated for quantitative analysis of 1,2:5,6-dianhydrogalactitol (DAG) in mouse plasma and tissues. Sodium diethyldithiocarbamate (DDTC) was used as the derivatization reagent to improve its LC-MS/MS behavior. Analytes were separated on a Welch Ultimate XB-CN column with a mobile phase consisting of acetonitrile and 0.1% formic acid solution (65:35). The MS analysis was conducted by positive electrospray ionization in multiple-reaction monitoring (MRM) mode. Good linearity (r 2 62>620.9958) was observed over the concentration range of 1–100062ng/ml in plasma and tissue homogenates (brain, liver, heart, spleen, lung and kidney). The intra- and inter-batch precision and accuracy of DAG in plasma and brain samples were all within the acceptable limits. The extraction recovery was stable and no significant matrix effects were observed. The method was successfully applied to study the pharmacokinetic and tissue distribution of DAG in mice after intravenous administration. DAG could cross the blood-brain barrier and had limited liver distribution. Rat primary hepatocytes in vitro experiments demonstrated that DAG had a safe profile in liver.
DOI:10.1016/j.jchromb.2018.04.026      URL    
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[8] PENG C,QI X,MIAO L,et al.1,2:5,6-dianhydrogalactitol inhibits human glioma cell growth in vivo and in vitro by arresting the cell cycle at G2/M phase[J].Acta Pharmacol Sin,2017,38(4):561-570.
1,2∶5,6-dianhydrogalactitol (DAG) is a hexitol epoxide with marked antitumor activity against multiple types of cancer cells,but the molecular mechanisms by which DAG functions as an antitumor agent is largely unknown.In this study,we investigated the inhibitory effects of DAG on human glioma cell growth in vitro and in vivo and uncovered the underlying molecular mechanisms.Treatment with DAG (120 μmol/L) dose-dependently inhibited the proliferation and colony formation in human glioma cell lines LN229,U251,and U87MG in vitro.DAG (1,2,5 μmol/L) induced cell cycle arrest at G2/M phase in the 3 glioma cell lines in a dose-dependent manner.The signaling pathways involved in DAG-caused cell cycle arrest was further analyzed in LN229 cells,which revealed that DAG dosedependently activated two parallel signaling cascades,ie,the p53-p21 cascade and the CDC25C-CDK1 cascade.DAG also significantly enhanced the radiosensitivity of LN229 cells as shown in the clonogenic assay.In nude mice bearing subcutaneously xenografted LN229 glioma,administration of DAG (5 mg/kg,iv,twice per week for 6 weeks) effectively suppressed the growth of xenografted tumors:the relative tumor growth rate (T/C) was reduced to 22.38%,and the tumor growth inhibitory rate (TGI) was 83.58% (P<O.01).In addition,DAG administration significantly activated the CDC25C-CDK1 cascade in the xenografted tumors.In conclusion,DAG inhibited human glioma cell growth in vitro and in vivo by inducing cell cycle arrest at G2/M phase.Two parallel cascades are activated and involved in the cell cycle arrest.
DOI:10.1038/aps.2016.154      PMID:28216618      URL    
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[9] JIANG X,HUANG Y,WANG X,et al.Dianhydrogalactitol,a potential multitarget agent,inhibits glioblastoma migration,invasion,and angiogenesis[J].Biomed Pharmacother,2017,91(4):1065-1074.
Abstract The complexity of cancer has led to single-target agents exhibiting lower-than-desired clinical efficacy. Drugs with multiple targets provide a feasible option for the treatment of complex tumors. Multitarget anti-angiogenesis agents are among the new generation of anticancer drugs and have shown favorable clinical efficacy. Dianhydrogalactitol (DAG) is a chemotherapeutic agent for chronic myeloid leukemia and lung cancer. Recently, it has been tested in phase II trials of glioblastoma treatment; however, mechanisms of DAG in glioblastoma have not been elucidated. Here we show that DAG could inhibit the migration and invasion of U251 cell line by inhibiting matrix metalloproteinase-2 (MMP2) expression. Furthermore, DAG could also inhibit tumor angiogenesis in vitro as well as in the zebrafish model. Mechanistic studies reveal that DAG inhibited both VEGFR2 and FGFR1 pathways. Our results suggest that DAG may be a potential multitarget agent that can inhibit tumor migration, invasion, and angiogenesis, and the anti-angiogenic effects may be involved in dual-suppression VEGF/VEGFR2 and FGF2/FGFR1 signal pathways. Copyright 2017 The Authors. Published by Elsevier Masson SAS.. All rights reserved.
DOI:10.1016/j.biopha.2017.05.025      PMID:28525947      URL    
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[10] 国家药品标准.WS-10001-(HD-1416)-2003.化学药品地标升国标第15册[S].2003.
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[11] 国家药品标准.WS-10001-(HD-0723)-2002.化学药品地标升国标第15册[S].2002.
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[12] 魏宁漪,宁保明.气相色谱法测定注射用去水卫矛醇的含量及有关物质[J].中国药师,2011,14(9):1306-1307.
目的:建立注射用卫矛醇含量及有关物质的气相色谱法。方法:采用Agilent DB-1(30 m×0.53 mm,1.5μm);程序升温:起始温度为100℃,维持3 min,以10℃·min~(-1)升至200℃,维持5 min;进样口温度为250℃,检测器F1D温度为250℃;载气为氮气,流速为5 ml·min~(-1)。结果:去水卫矛醇与其相邻杂质峰能完全分离,去水卫矛醇在0.010 05~1.005 mg·ml~(-1)浓度范围内线性关系良好(r=0.999 1),平均回收率为101.1%(RSD=1.12%,n=9)。结论:本方法操作简单,精密度好,结果准确可靠,可用于该制剂的质量控制。
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[13] 吴先富,张琪,马玲云,.定量核磁共振法测定去水卫矛醇的含量[J].药物分析杂志,2017,37(1):181-184.
目的:建立定量核磁共振法(qNMR)测定去水卫矛醇的含量。方法:采用核磁共振仪测定一维定量氢谱,90°脉冲,谱宽7 500 Hz,驰豫延迟时间为20s,采样次数为64次测定温度25℃,以对苯二甲酸二甲酯为内标,氘代二甲基亚砜(DMSO-d_6)为溶剂,对去水卫矛醇进行定量研究。结果:qNMR法测定去水卫矛醇的含量为95.82%,与质量平衡法测定结果(96.22%)基本一致。结论:本文建立的qNMR法测定去水卫矛醇的含量准确可靠,简便快速,为该品种的质量控制和对照品赋值提供了新的测定方法。
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关键词(key words)
去水卫矛醇
柱前衍生化
色谱法,反相高效液相
含量测定

Dianhydrogalactitol
Pre-column derivatization
Reversed-phase high perfo...
Content determination

作者
王杏利
孙丽涵
张雷
吴华晶
张峻颖
吴春勇

WANG Xingli
SUN Lihan
ZHANG Lei
WU Huajing
ZHANG Junying
WU Chunyong