Rapid screening for bisbibenzyls in bryophyte crude extracts using liquid chromatography/tandem mass spectrometry

A simple and rapid qualitative liquid chromatography-diode-array detection/tandem mass spectrometry (LC-DAD/MS/MS) method was developed and validated for screening bisbibenzyl compounds in bryophyte crude extracts at sub-ppm levels. After simple extraction with ethanol and analyte concentration with diethyl ether, the extracts were subjected to LC-DAD/MS/MS analysis. The overall instrument turnaround time was 50 min to obtain baseline separation of bisbibenzyl isomers in bryophytes. MS full scan, MS/MS precursor ion scan and MS/MS product ion scan modes were used for the screening. The bisbibenzyl standards studied gave limits of detection (LODs) at or below 10 ng/mL. The results also indicated that the method had acceptable precision to be used on a day-to-day basis for qualitative identification. The bisbibenzyl types, i.e. one biphenyl ether bond (A-type), two biphenyl ether bonds (B-type), one biphenyl ether and one biphenyl bond (C-type), or other biphenyl types can be differentiated by their ESI-MS/MS product profiles, and the number of alkoxyl substituents can also be identified. The linkage sites of biphenyl and biphenyl ether bonds cannot be identified for an unknown bisbibenzyl solely from its mass spectra. This system was used to support three screening assays of bryophytes including Marchantia polymorpha L., Ptagiochasm intermedium L. and Asterella angusta, which were collected from different places in China. From them, 7/12, 8/5 and 8/9 confirmed/unconfirmed bisbibenzyls were identified, respectively, based on their MS/MS data, UV spectra and the retention behavior. The screening method considerably reduced the time and the cost for the qualitative analyses, and the structure-fragmentation-UV relationships will facilitate the high-throughput screening (HTS) of bisbibenzyl compounds in bryophytes. It is also intended as a simple and convenient way for the determination of other structural families of natural products.