Study On The Traceability Of Antibiotic Reference Standards

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 formâ… of cefathiamidine was best in all of the cefathiamidine produced by different technology. The crystal formâ… of 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.