Research On Quality Standard,UPLC Fingerprints And Chemical Pattern Recognition Of Trollius Chinensis Bunge

Trollius chinensis Bunge consists of the dried flowers of Trollius chinensis Bunge(Ranunculaceae), was first recorded in Supplement to Compendium of Materia Medica during the Qing Dynasty. Trollius chinensis Bunge has antibacterial, antiviral, antitumor and antioxidant activity, has been used treat tract infection, pharyngitis, tonsillitis and bronchitis. The Flora of China project documented and described 16 Trollius species. Among these plants, Trollius ledebouri Reichb, Trollius altaicus C. A. Mey, Trollius macropetalus Fr. Schmidt,Trollius buddae Schipcz, Trollius farreri Stapt and Trollius ranunculoides Hemsl. were also used as Chinese folk medicines. No information about Trollius chinensis Bunge has been found in Ch.P.2010, EP, USP, JP and Hong Kong Chinese Medicinal Material Standards, while Trollius chinensis Bunge in Heilongjiang Chinese Medicinal Material Standards 2001 edition, Shanghai Chinese Medicinal Material Standards 1994 edition and Mongolian Medicine of Ministerial Standards without TLC identification and composition. Thus quantity control of Trollius chinensis Bunge and its related TCMPs is great important. The constituents of Trollius chinensis Bunge have been demonstrated to be mainly composed of flavonoids, phenolic acids, and alkaloids. Flavonoids are the most abundant compounds among them and have significant bioactivities, Orientin and vitexin are the two most important flavone C-glycosides, and they and their derivatives comprise almost two-thirds of the total flavonoids. Both orientin and vitexin were reported to have significant pharmacological activities such as antibacterial, antiviral, anticancer, anti-oxidative, anti-inflammatory, and analgesia activity. Therefore, orientin and vitexin can be used to control the quality of Trollius chinensis Bunge. The TLC method of identification and HPLC method of determination of orientin and vitexin was developed, and provide scientific basis for quality control of Trollius chinensis Bunge. Using ultra-performance liquid chromatography(UPLC) method to establish fingerprint of Trollius chinensis Bunge, comprehensive reflected different main components and content of Trollius chinensis Bunge from different base source, and provided the basis for classification and evaluation of intrinsic quality characteristics for Trollius. Part one Research on quality standard of Trollius chinensis BungeObjective: To establish the quality standard of Trollius chinensis Bunge.Methods: The orientin and vitexin of Trollius chinensis Bunge were identified by TLC and determined by HPLC. The method of HPLC used Shiseido Capcell pak C18 MGⅡ column(4.6×250mm; 5 μm) with a mixed mobile phase of 0.1% phosphoric acid and acetonitrile in a gradient mode. The flow rate was 1.0 m L?min-1and the wavelength of measurement was 340 nm. The temperature was 40℃.Results: The TLC identification exhibited an intense fluorescent band, corresponding in color to the band in the chromatogram of Standard. Orientin showed a good linear relationship in the range of 7.59~379.62μg/ml, the regression equation was Y=25548.9728X+34229.8504,R2=0.9995. Vitexin showed a good linear relationship in the range of 2.31~115.50μg/ml, the regression equation was Y=27760.6117X+10636.2063, R2= 0.9996.The precision, repeatability and stability of the method were satisfying. The average recovery was in the range of 98~102%, RSD<1.45%.Conclusion: This quality standard can be used to control the quality o f Trollius chinensis Bunge. Part two UPLC fingerprints and chemical pattern recognition ofTrollius chinensis BungeObjective: The paper is to report the establishment of a method of fingerprints analysis for the quality control of Trollius chinensis Bunge.Methods: Application of UPLC techniques was connected in series and applied. The method was performed on a ACQUITY UPLC BEH C18(100 mm × 2.1 mm ID, 1.7 μm) with a mixed mobile phase of 0.1% phosphoric acid and acetonitrile in a gradient mode. The flow rate was 0.3 m L?min-1and the wavelength of measurement was 340 nm. The temperature was 45℃.Results: Thirty-eight batches of the Trollius chinensis Bunge were determined. The UPLC fingerprints of Trollius chinensis Bunge was developed. The 2″-O-β-L-galactopyranosyl orientin, orientin and vitexin were identified. Hierarchical cluster analysis(HCA) and principal component analysis(PCA) were applied to study on the UPLC fingerprints and chemical pattern recognition. The Trollius chinensis Bunge, T. ledebouri Reichb and T. altaicus C. A. Mey were very good to distinguish.Conclusion: This method is accurate and reliable, and it can be used to control the quality of the Trollius chinensis Bunge.