Qualitative Analysis Of Flavonoids In Phyllostachys Edulis And Pleioblastus Maculatus Leaves Using Mass Spectrometry

Flavonoids is main bioactive substance in bamboo leaf extract,qualitative research of BLF is of great significance to elucidate the structure-activity relationship of BLF.At present,mass spectrometry qualitative analysis technology has the advantages of high sensitivity,high resolution and rapid qualitative analysis of complex mixed samples.However,the mass spectrometry data of related compounds database is limited,so it is necessary to study the mass spectrometric fragmentation patterns of flavonoids to provide reference for qualitative analysis.Hereby,in this thesis,with Phyllostachys edulis leaves（5kg）and Pleioblastus maculatus leaves（5kg）are used as materials after ethanol extraction,petroleum ether extraction,macroporous absorption resin column chromatography purification,concentration drying to obtain bamboo leaves extract,qualitative analysis of flavonoids were performed using UHPLC-Q-TOF-MS.The main results of this thesis were as follows:1.UHPLC-QTOF-MS/MS was used to analyze the main chemical components in Phyllostachys edulis leaves extracts:Method is that the column is an C18 column（150×2.1mm,1.8μm）,the column temperature is 40℃,the mobile phase A is methanol,B is 0.1%formic acid/water,sample were eluted at a flow rate 1.4ml/min with gradient elution,and the injection volume of 10μL,Retention time 120min,ESI ion source,negative ion mode,mass-to-charge ratio scanning range is m/z 50-800.13 flavonoids were successfully separated and detected within 120 min in Phyllostachys edulis leaves extract.The mass spectrometry mechanism of the four standards were studied,the results showed that The characteristic ion of flavonoid c-glycoside is ^0,3X₀^-,[^0,3X₀-H₂O]^-,and the abundance of the two characteristic ions of flavonoid 6-C-glycoside is higher than that of flavonoid-8-C-glycoside.The mass spectrometry analysis of flavonoid-C-glycosides in the extract showed that the characteristic ions were ^0,3X₀^-,^0,2X₀^-,^0.1X₀^-and[Y₀-H]^-·,And the characteristic ion[Y₀-H]^-·of 3’-methoxy flavonoid-C-glycosides had a higher abundance than that of flavonoid-C-glycosides.Qualitative mass spectrometry analysis of flavonoid-O-glycosides in the extract showed that the characteristic ions were Y₀^-,Z₀^-,[M-H-CH₃]^-·and[M-H-2CH₃]^-.For the mass spectrometry analysis of flavonoid C,O-diglycoside in the extract,the characteristic ions are[Y₁-H₂O]^-,[Y₁-^0,3A₂]^-,[Y₁-^0,2A₂]^-,producing the characteristic ion[Y₁-H₂O]^-with high abundance.The qualitative mass spectrometry analysis of flavonoid di-C,O-glycoside in the extract showed that the characteristic ions were Y₁^-,[Y₁-^0,3A₂]^-,[Y₁-^0,2A₂]^-.Nine flavonoids were preliminarily identified by the above results.2.UHPLC-QTOF-MS/MS was used to analyze the main chemical components in Pleioblastus maculatus leaves extracts:Method is that the column is an C18 column（150×2.1mm,1.8μm）,the column temperature is 40℃,the mobile phase A is methanol,B is 0.1%formic acid/water,sample were eluted at a flow rate 2.5 ml/min with gradient elution,and the injection volume of 10μL,Retention time 80 min,ESI ion source,negative ion mode,mass-to-charge ratio scanning range is m/z 50-800.31 flavonoids were successfully separated and detected in Pleioblastus maculatus leaves extracts.The mass spectrometry mechanism of the ten standards were studied,The results show that flavonoid-di-C,C-glycosidic characteristic ions for[M-H-C₂H₄O₂]^-,[M-H-C₃H₆O₃]^-,[M-H-C₅H₁₀O₅]^-,[M-H-C₆H₁₂O₆]^-;The mass spectrometric analysis of flavonoids in the extracts of Pleioblastus maculatus leaves showed that the cracking rule and characteristic ions of mass spectrometry were consistent with those of flavonoids in the extracts of Phyllostachys edulis leaves,and 21 flavonoids were tentatively identified.In summary,an UHPLC-Q-TOF-MS/MS method was successfully applied for the structural identification of flavonoids in Phyllostachys edulis and Pleioblastus maculatus leaves.Research results of this thesis have reference value for the rapid identification and structural characterization of these compounds in crude bioactive extracts or mixtures.