Studies On The Methodology And Application Of Direct Analysis In Real Time Mass Spectrometrv For The Quality Control Of Traditional Chinese Medicine

Mass spectrometry is widely used in the quality control of traditional Chinese medicine (TCM). The advantage of this technique lies in its strong specificity and high sensitivity. However, the mass spectrometric analysis of complex sample is generally time-consuming. Direct analysis in real time mass spectrometry (DART-MS) can obtain the mass spectrum within just one minute, which significantly increases the efficiency of mass spectrometric analysis. The aim of this dissertation is thus to study the application of DART-MS on the quality control of TCM. The main innovation and academic contributions of this study are summarized as follows:1A rapid quality assessment method based on DART-MS was developed for TCM injection. DART-MS was employed to identify analytical markers from Danshen injection.The optimal analysis conditions were achieved by measuring the contribution of various experimental parameters to the mass spectrum. Salvianolic acids and saccharides were simultaneously determined within a single1-min DART-MS run. The spectra of Danshen injections supplied by five manufacturers were processed with principal component analysis (PCA). Obvious clustering was observed in the PCA score plot, and candidate markers were recognized from the contribution plots of PCA. The suitability of potential markers was then confirmed by contrasting with the results of traditional analysis methods. Using this strategy, fructose, glucose, sucrose, protocatechuic aldehyde and salvianolic acid A were rapidly identified as the markers of Danshen injection. Furthermore, two novel compounds in analytical markers were separated by semi-preparative liquid chromatography and then identified by nuclear magnetic resonance. Finally, DART-MS was successfully applied to the fingerprint analysis of Danhong injection. DART-MS was found to be a reliable technique for assessing the quality of TCM injection.2The application of DART-MS to monitor TCM manufacturing process was presented. Danshen alkaline precipitation was taken as an example. A total of15batches run under normal operating conditions were used to develop a PCA model. Two kinds of multivariate control charts (Hotelling T2and DModX charts) were applied to examine the batch-to-batch reproducibility of12test batches. Artificial variations including starting material changes and process faults were identified. Further understanding of the deviating process behavior was achieved by investigating the contribution plot of abnormal batches. The results showed that DART-MS, in conjunction with multivariate analysis, provided a useful tool for monitoring TCM manufacturing processes.3A surface flowing mode sample holder was designed for DART. The online coupling of macroporous resin column chromatography with DART-MS was then achieved through the sample holder. This new hyphenated system was employed to monitor the elution process of Panax notoginseng’s column chromatography. The diluted eluent from macroporous resin column was firstly mixed with derivatization reagent, then transferred into the ionization region of DART by the sample holder. Notoginsenosides were methylated and ionized in metastable helium gas stream, and then led into MS for the detection. The alteration of chemical composition was rapidly and accurately identified, which agreed well with the offline determination result. This study suggested that the surface flowing mode DART-MS could be used in the online monitoring of TCM manufacturing process.4It was studied that the effects of key ion source operating parameters on DART-MS analysis of various TCM active components. A total of18active compounds from phenylpropanoids, alkaloids, saponins, flavones, volatile oils and glycosides were investigated. For each substance, the peak area and signal to noise of its characteristic ions under different discharge gas and heater temperature were compared. Based on the comparison, the relationships among chemical structure, ion source parameters and instrument responses were revealed. Finally, some suggestions about choosing discharge gas and heater temperature were proposed for types of TCM active substance, which offered a reference for the application of DART-MS on TCM analysis.