A Large-Scale Detection, Identification and Quantification of Target Metabolites using dMRM-MS Combined with Transcriptome of Two Rheum Species Focused on Anthraquinone and Flavonids Biosynthesis

Abstract Background Rheum emodi ( R. emodi ) is a perennial herb as well as an important medicinal plant. The main active compounds in Rhubarb are anthraquinones, but also contains flavonoids. As a class of important active components in rhubarb, anthraquinones have many pharmacological effects. However, due to the lack of rhubarb genome, it is difficult to mine functional genes involved in the biosynthesis of anthraquinones in rhubarb. Results Based on the a large-scale quantification of target metabolites combined with the transcriptome data of Rheum Palmatum L . (RPL) and Rheum Officinale Bails (ROB), in this study we aimed to firstly mine out anthrquinones and other pharmacological metabolites in rhubarb, then search genes which may related to the biosynthesis of anthraquinones and other metabolites. Via dynamic-multiple reaction monitoring (dMRM) of triple quadrupole mass spectrometry (QqQ-MS) technical means, 62 compounds including 21 anthraquinones, 17 flavonoids, 6 stilbenes, 12 gallate esters, 3 tannins, and 3 others were simultaneous qualitatively and quantitatively assessed. Furthermore, a total of 180,689 transcripts and 90,242 unigenes were generated by the de novo transcriptome assembly. Down-stream bioinformatic analyses annotated the unigenes with BLAST against seven public databases including KEGG and KOG. GO and KEGG pathway analyses further characterized the unigenes, resulting in 21,691 unigenes with metabolic pathway annotations. There were 121 genes annotated in the flavonoid pathways, and 38 of them showed a significant difference in expression level between the RPL and ROB. According to the difference in the content of emodin and physcion in the two kinds of rhubarb and the expression level of key enzyme genes, we further search 7 differentially expressed genes annotated with the activity of caffeoyl coenzyme methyltransferase. RT-qPCR results showed that the expression of the cluster-14354.38156 in the ROB was approximately 14-fold higher than that observed in the RPL, which may participate in the process of O-methylation from emodin to physcion. Conclusion Combining metabolome and transcriptome analysis, a key enzyme gene was identified. This study supplied a useful resource for further study of metabolite in rhubarb.