Application Of Impurity Profiling Technology In Control Of Impurities In Chemical Injections

Drug safety is directly related to human health,which is a major problem of people's livelihood and public safety.Injection takes action faster,which is one of the most extensive and significant formulation clinically.Currently,a large number of injections have been included in the National Essential Drugs.However,in recent years,due to the phenomenon that injection adverse events caused by the production technology and the sterilization processoccur occasionally,such as Clindamycin Phosphate Glucose Injectionand Acanthopanax Injection,the society began to question the safety of the infection.These adverse reactions are not only related to the pharmacological activity of drug itself,but also related to the presence of impurities,especially toxic impurities.Because of the complications of the synthetic process,formulation and sterilization process,and a wide variety of impurities,it is difficult for traditional chromatographic analysis methods to provide accurate and complete analytical information.Therefore,to establish a set of systematic,comprehensive,and accurate method for injection impurity analysis is a meaningful topic vital to national well-being and the people's livelihood.Recently,drug impurity research generally adopts the concept of impurity profile control.The key of impurity profile control is to separate all the impurities in drug,understand the source,structures and activities of impurities,and have good ruggedness.However,current quality control method of impurity and the control limits of certain impurities and impurities were far from effective control the drug safety.Therefore,it needs to further improve standards.On the basis of the Twelfth Five-Year Plan of National Drug Safety,using impurity profiling technology,through the combination of chemical and biological technologies,we carried out a series of research including industrial analysis,impurity screening,targeted synthesis,structure confirmation,separation and determination,toxicity evaluation and strived to carry out a comprehensive and in-depth analysis of composition of drug impurities and relationship of relevant toxicity and effectiveness,which provided a basis for the improvement of the quality of drugs and the clinical medication safety.This dissertation studied and developed traditional impurity profiling technology,and on the basis of the liquid-phase separation technology,established and optimized high performance liquid chromatography(HPLC)methods with effective combination of two dimension liquid chromatography quadrupole time of flight mass spectrometry(2DLC-QTOF MS)and Mass Frontier,and applied these methods in the separation and analysis of injection impurities.Additionally,the Derek,Sarah software and Ames method were applied to evaluate for biological properties of impurities,especially genotoxicity,which provided supporting for national standard setting of kinds of injection impurities and offer new idea and methods for the development of impurity profiling technology.The research work in this thesis could be divided into three sections,which were summarized as follows:Section One:IntroductionThis chapter introduced the research background and effect of impurities on drug safety.Besides,we summarized universal method of impurity analysis,and generalized current research in impurity as well as impurity profiling technology.It described the meaning and thoughts of this dissertation.Section Two:The Impurity Research of Milrinone Injection.Based on the national pharmaceutical sample platform,through the comparison of the difference in quality of Milrinone injection and the production process together with factors of experiment and using newly established liquid chromatography method to comprehensively analyze impurities of Milrinone Injection,the impurity spectrum of Milrinone Injection was developed.In addition,two impurities were synthesized and separated by liquid chromatography,and structures were confirmed using LC-MS,1H-NMR as well as ESI-MS.In particular,impurity ?,a new structure,was evaluated for biological properties by Derek and Sarah,especially genotoxicity.Results showed that impurity ? had no genotoxicity,but it had skin irritation.Therefore,in order to reduce the adverse events,we should view impurity ? as risk impurity to control.In addition,on the basis of ICH and National Drug Standards,the quality standards of Milrinone and Milrinone Injection were revised and were recorded as mandatory standard by Chinese Pharmacopoeia,which were used for the quality control of Milrinone Injection nationwide.Furthermore,impurity ?has attained the authority of Chinese National Invention Patent,and research results have been applied.Section Three:The Impurity Research of Aminobutyric Acid Injection.This section described a systematic study of impurities of the aminobutyric acid and aminobutyric acid injection using impurity profiling technology.The impurities in aminobutyric acid sample were determined by the combination of preparation technology and results of domestic typical sample,and speculated by 2D-HPLC-QEplus as well as Mass Frontier.In addition,impurities were synthesized and confirmed using UV,IR,MS,!H-NMR and LC-MS.In particular,compound 2 that is first mentioned has attained the authority of Chinese National Invention Patent,WO:ZL201510077458.1.Moreover,impurity 2 was evaluated for their biological properties,especially genotoxicity.The QSAR and Sarah soft analysis showed that it has positive test results.In order to further investigate,we used Ames test to study whether impurity 2 has genotoxicity.Results confirmed that impurity 2 has no genotoxicity.However,it may lead to the nervous system adverse reaction.Therefore,impurity 2 should be control as risk impurity.In addition,the impurity inspection of government standard of aminobutyric acid and aminobutyric acid injection were proposed,which has been adopted by Chinese Pharmacopoeia Commission,meanwhile,the research results have been applied to the mass control by dongbei Pharmaceutical Co Ltd.Section Four:The Impurity Research of BC-02 for Injection.On the technical level of the screening of separation,qualitative analysis and toxic assessment of impurities,impurities in BC-02 were analyzed and studied systematically.We developed a liquid chromatography method suitable to the separation and analysis of BC-02,and established an impurity spectrum which comprehensively reflected the process and impact factor of BC-02.The impurity analysis method has attained the authority of Chinese National Invention Patent(WO:ZL201510077487.8).Besides,the structures of seven impurities were speculated by 2D-HPLC-QEplus and Mass Frontier and four impurities were synthesized.Among them,impurities 5 and 6 were two novel structures which were separated by liquid chromatography and confirmed using MS and elemental analysis.Although other impurities are not new compounds,these impurities belong to the first study in BC-02.These seven impurities were also evaluated for genotoxicity by Derek and Sarah.Results showed that except impurity 7.impurities 1-6 had no genotoxicity.However,further studies showed impurity 7 is an unstable compound,which can not be detected in samples.Besides,these six impurities had other adverse reactions,such as hepatotpxicity,ocular toxicity,skin sensibility and so on.Thus,impurity 1-6 should be regarded as a risk impurity to control.In accordance with ICH and CDE's requirements,we proposed and established the quality control standards of impurity in BC-02,which provided a good example for impurity profiling technology,in the field for new drug research.Section Five:Conclusion and ProspectNew development and systematic analytical method of impurity profiling have been concluded in this dissertation.Results revealed that this method could be used as a general technical method for chemical analysis of impurities.Meanwhile,we summarized the research course and results of impurities in Milrinone Injection,Aminobutyric Acid Injection and BC-02 for injection.In addition,we also analyzed existing problems and shortcomings,which pointed out the direction for further studies.The starting point of this dissertation is to analyze impurities in injection;the ways to the discovery are the combination of process analysis and experimental screening;the method of analysis are modern chromatography and hyphenated technologies;the methods of biological characteristics assessment are the combination of the software toxicity evaluation technology and Ames test.All of these provide a systematic and comprehensive thoughts and approaches of study of discovery,separation,identification and determination of drug impurities.Moreover,we summarized research process of impurity and results including Milrinone Injection,Aminobutyric acid injection and BC-02 for injection,which point the way to the further study.