Identification, characterization and distribution of monoterpene indole alkaloids in Rauwolfia species by Orbitrap Velos Pro mass spectrometer

• LC/MS Orbitrap Velos Pro method was developed for metabolite profiling of Rauwolfia species.
• CID mode provide more structural information than HCD.
• Fragment pathways/diagnostic fragment ions were identified via MSn of standards for MIAs.
• Ten bioactive MIAs were identified and characterized in six Rauwolfia species.

Monoterpene indole alkaloids (MIAs) are medicinally important class of compounds abundant in the roots of Rauwolfia species (Apocynaceae). MIAs such as yohimbine (aphrodisiac agent) and reserpine (antihypertensive, tranquilizer) are the official drugs included in Model List of Essential Drugs of World Health Organization (WHO). Therefore, we have attempt to identify and characterize the MIAs in the crude extracts of six Rauwolfia species using ultrahigh-performance liquid chromatography coupled with Orbitrap Velos Pro hybrid mass spectrometer. The identity of the MIAs were construed using the high resolution tandem mass spectrometry (HRMS/MS) spectra of standard compounds ‘yohimbine’ and ‘reserpine’ in higher energy collisional dissociation (HCD) and collision-induced dissociation (CID) modes. The diagnostic fragment ions found in HCD mode was highly affected by variation of normalized collision energy (NCE) and gave few product ions (‘C–F’) while CID produced intense and more diagnostic product ions (‘A–F’). Consequently, CID-MS/MS mode provided significantly more structural information about basic skeleton and therefore the recommended mode for analysis of MIAs. Furthermore, six diagnostic fragmentation pathways were established by multi-stage mass analysis (MSn (n = 5)) analysis which gave information regarding the substitution. Fragment ions ‘A–F’ revealed the number and position of substituents on indole and terpene moieties. The proposed diagnostic fragmentation pathways have been successfully applied for identification and characterization of MIAs in crude root extracts of six Rauwolfia species. Ten bioactive reserpine class of MIAs were tentatively identified and characterized on the basis of chromatographic and mass spectrometric features as well as HRMS/MS an MSn (n = 4) analysis.

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