The Study On Quality Control For The Innovative Drug Fluoropaclitaxel And Technology Of Pharmceutial Analysis

Paclitaxel was a compound extracted from the skin of taxus brevifolia. It could act on microtubule of cells, disturbing cell functions, so as to inhibit cell division. By structure modifications, the new compound could show comparative or even stronger anti-cancer activities, and properties could be better than paclitaxel. Innovative drug fluoropaclitaxel was synthesis by pharmaceutical chemistry of Fudan University following new synthesis route, and was planned to apply for state class new medicine according to FDA requests.The study on quality control and stability of drugs is an important content of drug pre-clinical studies. During the study on drug quality standards, a thorough analysis of drug quality should be done to guarantee the safety and reliability of drugs, and to guarantee the smooth progress of pre-clinical research, as well as the safety of post-marketing drugs. Based on the study on drug quality standards and stability, the contents and methods of the quality standards and stability of fluoropaclitaxel and the draft quality standards of fluoropaclitaxel were established.By elemental analysis, UV, IR, NMR (13C,1H) and mass spectrometry for structural confirmation of fluoropaclitaxel, reliable raw material for the study on quality standards and stability was provided. The dissertation was divided into the following two parts:the first part is as the main part titled the study on quality control and stability of the innovative drug fluoropaclitaxel, the second part as the deputy papers titled determination of Leu-enkephalin drugs by a novel high performance liquid chromatographic method with fluorimetric detection.The first partThe study on quality control and stability of the innovative drug fluoropaclitaxel1、The quality control of fluoropaclitaxelThe quality standard of fluoropaclitaxel was investigated according to Chinese Pharmacopoeia and technical specifications of the drug quality standards. The item of shape and properties, physical constant, identification, examination and assay were studied to determine the appropriate test methods and limitation requests. Stability was studied on this basis.The method of residual organic solvents inspection was established. And differences between two injector types were compared. The result indicated that organic solvent residues of fluoropaclitaxel met Chinese Pharmacopoeia requirements.The method of the related substances inspection was established. And differences between four types of columns were compared. C8column was finally chosen for the inspection. The results of related substances inspection were between0.49-1.26%.The method of the assay was established. The method was optimized. The assay was performed in HITACHI LaChrom-C18column(4.6×250mm,5μm) with linear gradient elution using acetonitrile-water(60:40(v/v)), the flowing rate was1ml/min, the column temperature was30℃, the UV detection wavelength was227nm, the sample volume was20μl. The result of method evaluation indicated the method was accurate and exclusive. The fluoropaclitaxel content determined by nonaqueous UV and high performance liquid chromatography were unanimous.2. The stability study of fluoropaclitaxelStability of fluoropaclitaxel was conducted according to the guiding principles of drug stability test, including impact factors test, accelerated test and long-term test. The sample characteristics, the main component content and related substances of stability samples were analyzed. It was found that fluoropaclitaxel was insensitive to light, but sensitive to temperature. It should be stored under the condition of low temperature. Also, for that fluoropaclitaxel had a little moisture absorption, it couldn’t be stored in humidity.The accelerated test indicated that fluoropaclitaxel was unstable at40℃, humidity of75%, while stable at30℃, humidity of65%for6months. And the six-months-long-term test indicated that fluoropaclitaxel was more stable at6℃, humidity of60%, than at25℃, humidity of60%. The long-term test is still carrying.For statistical analysis on the validity, if main component content of fluoropaclitaxel becomes98.5%, the retention time is13.7months. So the fluoropaclitaxel is valid for13.7months.According to the stability study, fluoropaclitaxel was suggested to be stored under the condition of low temperature, dark and dry.3. The draft quality standards of fluoropaclitaxel and drafting instructionsThe draft quality standards of fluoropaclitaxel and drafting instructions were formulated and drafted according to the first two chapters of the research content. This chapter provides norms and basis for quality control of fluoropaclitaxel. The second partDetermination of Leu-enkephalin drugs by a novel high performance liquid chromatographic method with fluorimetric detectionIn this study, we establish a novel high performance liquid chromatographic method with fluorimetric detector (HPLC-FLU), using pre-column redox derivatization for the determination of Leu-enkephalin (L-EK) drugs. After mixing L-EK with0.012mol/L sodium periodate,0.01mol/L catecholamine and0.3mol/L boric acid (2:2:1:1, v/v), mixture was heated at120℃for10minutes. The chromatographic separation was performed on an ODS column with0.25mol/L borate buffer (pH7.6)-methanol as mobile phases, using gradient elution. The derivatized compounds were detected with fluorimetric detector. L-EK was detected by this method in vitro and in human plasma, compared with HPLC-UV and HPLC-FLU with pre-column OPA derivatization. Results indicated that the linear range of HPLC-FLU with pre-column redox derivatization was0.5-50.0mg/L (r=0.9998) in human plasma and the limit of detection was0.1mg/L. This method was better than the other two in not disturbed by endogenous substances. HPLC-FLU with pre-column redox derivatization has advantages in detecting L-EK in human plasma.