【作者】 刘书妤；

【导师】 邵荣光； 胡昌勤； 宋丹青；

【作者基本信息】 中国协和医科大学 ， 微生物与生化药学， 2008， 博士

【摘要】 抗生素标准物质是抗生素质量分析中使用的实物对照,是保证抗生素药品质量的关键,其量值必须准确可靠,即具有溯源性。虽然标准物质在标定过程中向国际基本单位的溯源是最终确定药品标准物质的量值和量值传递的基础,但现阶段国内抗生素标准物质并不能完全实现对每个化学对照品、生物标准品都直接溯源至国际单位,为实现溯源这一目标,我们针对抗生素标准物质研制过程中的定值和选材这两个重要环节,将核磁定量法应用到标准物质的定值中,探讨核磁定量方法的可行性;分析头孢菌素类抗生素标准物质原料的理化性质,建立头孢菌素类抗生素标准物质原料选择的指导原则。在对抗生素标准物质的定值研究中,首先采用核磁定量法和质量平衡法两种不同原理的方法,对盐酸头孢唑兰基准物质进行标定,确定其量值,验证量值的准确性,归纳总结标准物质定值的一般方法。同时针对国产盐酸头孢哗兰比旋度普遍不在日本药典标准中间值的现象,对盐酸头孢唑兰比旋度测定方法的准确性进行分析,通过对比旋度测定中的两个影响因素（溶剂和样品纯度）的分析,发现日本药典的比旋度测定条件不能保证头孢唑兰以单一构型存在,因此改进了测定方法;通过圆二色谱、紫外光谱、质谱、液核联用、核磁碳谱等多种分析技术研究盐酸头孢唑兰变构现象,确定了盐酸头孢唑兰两种构型的结构。其次,采用核磁定量法对常见的大环内酯类抗生素标准物质进行定值,建立了大环内酯类抗生素核磁定量分析的一般方法,通过与质量平衡法的标定值和不确定度进行比较,证明核磁定量法定值结果准确可靠,可以作为质量平衡法标定的重要补充。第三,对不适合采用质量平衡法定值的红霉素和琥乙红霉素及部分纯度相对较低的十六元环大环内酯类抗生素进行核磁定量分析,通过与质量平衡法的测定结果及不确定度,或者与高效液相法（HPLC）的测定结果进行比较,进一步评价了核磁定量方法的适用性;虽然十六元环大环内酯类抗生素的不同核磁定量峰的响应因子不完全相同,但当选择适宜的定量峰,核磁定量法仍能得到准确的标定结果。核磁定量法为标准物质定的值提供了新的思路。在对头孢菌素类抗生素标准物质的选材研究中,以头孢硫脒和头孢呋辛钠二个头孢菌素为例:（1）通过对不同工艺生产的头孢硫脒的外观、晶型,结晶水等方面的研究,发现头孢硫脒Ⅰ型结晶的热稳定性最好,对湿度和温度变化不敏感,因此其化学稳定性最好,据此确定了头孢硫脒标准物质原料选择的理化指标。在对不同晶型的头孢硫脒化学稳定性的研究中,对头孢硫脒降解过程中的杂质谱变化进行分析,寻找杂质的变化规律,确定杂质的来源和主要杂质的的结构,进而确定表征头孢硫脒对照品稳定性的信号杂质。（2）通过粉末X-射线衍射、扫描电镜、固体Na²³NMR等方法对不同来源的头孢呋辛钠晶体结构进行分析:发现不同厂家生产的头孢呋辛钠晶型相同,结晶度不同;头孢呋辛钠固体是头孢呋辛钠晶体和无定形粉末的混合物;通过分析头孢呋辛钠结晶度与其稳定性的关系,发现结晶度越高,稳定性越好。据此确定了头孢呋辛钠标准物质原料选择的理化指标。更多还原

【Abstract】 The antibiotic reference standards served as the reference object play a very important role in the quality control of antibiotics. Therefore, the purity of reference standards must be accurate and reliable. The reference standards must be traceable. Although it is elementary that the reference standards trace to international system of units in the calibration process, not all chemical reference standards of antibiotics and biological standards of antibiotics can achieve the traceability at present in China. So we researched on the evaluation of reference standards and the selection of raw material in the preparation of antibiotic reference standards. First, We applied the quantitative nuclear magnetic resonance （qNMR） to evaluate reference standards and analyzed the possibility of qNMR method. Second, We researched on the physico-chemical property of raw material cephem antibiotic raw material and established the direction for the selection of raw material.In the study of evaluation of antibiotic reference standards, firstly, we calibrated the cefozopran primary reference standards by two different principle methods including the qNMR method and the mass balance method, validated the accuracy of cefozopran purity and summarized the general method for the calibration of reference standards. To solve the problem that the specific rotation of cefozopran produced in China was not generally in the middle of limitation of specific rotation method in Japanese Pharmacpoeia, we analyzed the accuracy of specific rotation method. In the research on two influencing factors including solvent and purity of sample, we found cefozopran did not exist as single configuration in the condition of specific rotation method. So we modified specific rotation method and determined two different configuration structures by circular dichroism spectra, ultraviolet spectra, mass spectra, HPLC-NMR and C¹³-NMR. Secondly, we calibrated the frequent macrolide antibiotic reference standards by the qNMR method and established the general qNMR method for macrolide antibiotics. It was proved that the qNMR method was accurate and reliable by comparison with the purity and uncertainty of 7mass balance method. Therefore the qNMR method could be complementary with the mass balance method for the assay of standard references. Thirdly, we calibrated the erythromycin and erythromycin ethylsuccinate which were not suitable for the mass balance method by the qNMR method. We also calibrated the 16-membered ring macrolide antibiotics of low purity by the qNMR method. The qNMR method was evaluated again by comparision with the results of mass balance method or of HPLC method. Although the response factor of different qNMR peak was not absolutely same, the qNMR method was still accurate when we selected the suitable quantitative peak. The qNMR method provided a new thought for the calibration of reference standards.In the study of raw material selection of cephem antibiotic reference standards, we taked the cefathiamidine and cefuroxime sodium for example: （1） Researching on appearance, crystal form, water of crystallization and so on, We found that the thermal stability of the crystal formⅠof cefathiamidine was best in all of the cefathiamidine produced by different technology. The crystal formⅠof cefathiamidine was not sensitive to humidity and temperature. So its chemical stability was best. Therefore, we determined the physicochemical index for the raw material selection of cefathiamidine reference standard. In the study of stability of different crystal form of cefathiamidine, we analyzed the changes of impurities, searched the change regularity, found the source of different impurities and determined the structure of major impurities. Finally we obtained some signal impurities which could monitor the stability of cefathiamidine. （2） In the research on crystal structure of cefuroxime sodium coming from different source, we found that the cefuroxime sodium produced by different manufacturers had the same crystal form and the different crystallinity by X-ray powder diffractometer, scanning electron microscope, solid Na²³ NMR and other methods. The powder of cefuroxime was the mixture of crystal and amorphous. In the research on the relationship of crystallinity and stability for the cefuroxime, we found that the higher crystallinity possesed better stability. So we determined the physicochemical index for raw material selection of cefuroxime sodium reference standard.更多还原