Tandem solid-phase extraction followed by HPLC-ESI/QTOF/MS/MS for rapid screening and structural identification of trace diterpenoids in flowers of Rhododendron molle.
BACKGROUND
'Naoyanghua', composed of the flowers of Rhododendron molle G. Don, is a traditional Chinese medicine that is widely known for its toxicity. Grayanane-type diterpenoids are the main active ingredients in R. molle, as well as possibly their toxicity: they are, however, difficult to isolate and analyse using common chromatographic methods, due to their small amounts and absence of conjugated groups, such as phenyl and α, β-unsaturated ketone.


OBJECTIVE
To establish a highly sensitive, selective and reliable method for the qualitative evaluation of trace diterpenoids in the flowers of R. molle by using tandem solid-phase extraction followed by high-performance liquid chromatography with electrospray ionisation quadrupole-time-of-flight mass spectrometry (HPLC-ESI/QTOF/MS/MS).


METHODS
Tandem solid phase extraction (SPE) was undertaken using a polyamide cartridge and a C18E cartridge in succession to enrich the trace diterpenoids. HPLC-ESI/QTOF/MS/MS was used to determine the fragmentation patterns of diterpenoids and to tentatively characterise their fragmentation pathways.


RESULTS
HPLC-ESI/QTOF/MS/MS detected a total of 14 diterpenoids, eight of which were identified by comparison with literature sources and six based on fragmentation analysis. Among the latter six, rhodojaponin VI-3-glucoside was tentatively identified as a new diterpenoid glycoside and rhodojaponin VII, rhodojaponin IV and rhodojaponin I were reported from R. molle for the first time.


CONCLUSIONS
By qualitative research of diterpenoids in this plant by HPLC-ESI/QTOF/MS/MS, a reliable methodology for the analysis of these active constituents of R. molle was established for the first time.
State Key Laboratory of Natural Medicines, Department of Natural Medicinal Chemistry, China Pharmaceutical University, 24 Tong Jia Xiang, Nanjing, 210009, People's Republic of China.
Hong-Yan
HY
Zou
Jun
J
Luo
De-Ran
DR
Xu
Ling-Yi
LY
Kong
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004224:Diterpenes / Q000737:chemistry / Q000302:isolation & purification*; D004365:Drugs, Chinese Herbal / Q000737:chemistry* / Q000302:isolation & purification; D035264:Flowers / Q000737:chemistry; D015394:Molecular Structure; D010946:Plants, Medicinal; D029793:Rhododendron / Q000737:chemistry*; D052616:Solid Phase Extraction; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013997:Time Factors
D004224:Diterpenes; D004365:Drugs, Chinese Herbal
HPLC-ESI/QTOF/MS/MS; Rhododendron molle; diterpenoids