Study On The Impurity Profiles Of Quinolones On The Basisi Of LC-MS/MS Method

The impurity profiles of quinolones were systematically investigated using liquid chromatography coupled with tandem mass spectrometry. In order to investigate the relationship between the structural characteristics and the mass spectrometric behavior of quinolones, a number of quinolones with several structural skeletons were studied in detail by electrospray ionization tandem mass spectrometry in positive ion mode. An integrated approach on basis of liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed for the rapid, automatic and efficient detection and identification of impurities from quinolone antibiotics using the data acquisition method and automatic impurity screening method. Some process-related impurities and degradation products were found by analyzing the different batches of drug substances and degradation samples under various stress tests using the integrated approach, and the degradation pathway was also elucidated by analyzing the trends of degradation products with time. A validated quantitative method for the impurities in the quinolone formulations was established, it was estimated to be sensitive, stable and accurate for the detection and monitoring of trace impurities. The impurities in different formulations from different pharmaceutical plants were determined using this approach, and the degradation products were the major impurities in the formulations which were generated by the degradation of drug ingredient. Some process-related impurities and degradation products in the drug substances and formulations were synthesized, their quality and cytotoxity were evaluated.1. The fragmentation behavior of quinolones using electrospray ionization tandem mass spectrometryThe relationship between the structural characteristics and the fragmentation behavior of quinolones were investigated in positive ion mode using QTRAP and Q-TOF mass spectrometers. It was found that the quinolone moiety was stable in collision induced dissociation (CID) mode, and the fragment ions were generated by the fragmentation of the substituents. The product ions were generated by the fragmentation of the substituent at N-1through the loss of alkyl radical or the neutral loss of alkene. The product ions were formed by the loss of H2O or CO2due to the fragmentation of carboxyl group at C-3and were always observed in the MS/MS spectra. Dehydrohalogenation can take place when the halogen atom is at C-6or C-8.7-piperazinyl-quinolones could generate characteristic ions by the neutral loss of43Da,57Da and so on, or the fragment ions at m/z70, et al, which can be used for the sensitive class-specific screening of7-piperazinyl-quinolones. The present study supplemented the gas-phase ion chemistry of quinolones and should be valuable in the structural identification of impurities.2. Integrated LC-MS/MS approach for the identification and characterization of the trace impurity of quinolonesBased on liquid chromatography-tandem mass spectrometry (LC-MS/MS), an approach integrating efficient data acquisition method and automatic impurity screening method using MMDFs and BS was developed. This approach was used to acquire the structural and semi-quantitative information related to UV and MS data in a single sample run, and even to discover the impurity signals submerged by background and drug ions. Structural characterization was performed through accurate mass measurement and comparison of fragmentation behaviors of impurities with the parent drug. This approach was illustrated by the comprehensive impurity analysis of levofloxacin, and was demonstrated to be rapid, sensitive and automatic for the detection, characterization and monitoring of impurities, especially those unknown or at trace levels.Process-related impurities and degradation products were screened from the test samples including different batches of quinolone drug substances and degradation samples using the integrated approach. And the degradation pathway was postulated by analyzing the trends of degradation products with time under various chemistry or environmental factors. The approach was estimated to have the widespread application by determining other quinolone antibiotics drug substances and degradation samples to find the impurities.3. A LC-MS/MS quantitative method for the impurities in quinolone antibiotics and the evaluation of cytotoxcity of impuritiesA validated quantitative LC-MS/MS method for the impurities in the formulations of quinolones was developed. The performance characteristics of the method in terms of robustness and information content clearly showed that this approach was adequate for the reliable detection, identification, and quantitative determination of trace levels of impurities in quinolones. Degradation products were the major impurities in formulations and there was remarkable difference in the impurity contents among the different pharmaceutical plants. The pure impurities were synthesized to obtain more enough to perform the quality evaluation and the evaluation of cytotoxicity. The cytotoxicity test was performed using L929mouse fibroblasts.