From heartbreak to happiness with in-house MS database

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  • Published: Jun 15, 2017
  • Author: Ryan De Vooght-Johnson
  • Channels: Laboratory Informatics / Chemometrics & Informatics
thumbnail image: From heartbreak to happiness with in-house MS database

So-called ‘heartbreak grass’ contains a complex mixture of natural products

Gelsemium elegans is used in traditional Chinese medicine, and there is some interest in its possible use to treat inflammation, cancer, pain and anxiety.

Gelsemium elegans, known as ‘heartbreak grass’, is a toxic climbing plant rather than a grass. It is found in several Southeast Asian countries and has been implicated in various suicides and murders. In small doses, it is used in traditional Chinese medicine, and there is some interest in its possible use to treat inflammation, cancer, pain and anxiety.

Many different natural products have been isolated from Gelsemium elegans and other Gelsemium species by different researchers. These include the toxic alkaloid gelsemine. Often, complicated and lengthy extraction and separation techniques have been employed, which don’t necessarily give a good overview of the compounds present in the fresh plant material.

The Hunan researchers decided to first of all compile a database of the accurate masses of all the isolated compounds from Gelsemium species reported in the literature. Few standards were available for Gelsemium compounds, so comparison with existing samples was not an option. The researchers then aimed to use LC-MS/MS to give a good indication of the compounds present in a crude extract, matching the accurate masses from the database with the experimental results. Quadrupole time-of-flight mass spectrometry (QqTOF MS) was employed, where the initial quadrupole separation of ions is followed by time-of-flight separation.

Gelsemium elegans LC-QqTOF MS profile compared to new database

Ground roots of Gelsemium elegans were extracted with 95% ethanol; the resulting solution was filtered and evaporated. The residue was dissolved in methanol, which was filtered prior to LC-MS. HPLC was carried out using an Agilent 1290 Infinity instrument fitted with an ODS-C18 column. Acetonitrile and 0.1% aqueous formic acid were used for gradient elution, with the volume of the former going from 10 to 90%. Mass spectrometry employed an Agilent 6530 Q-TOF instrument using electrospray ionisation (ESI) in positive mode and Agilent Mass Hunter software.

An in-house database of accurate masses was compiled using Agilent PCDL Management software from 204 compounds previously reported from Gelsemium species. For each entry, the molecular formula, accurate mass and chemical structure were recorded. The authors noted that an in-house database was less likely to give false positives than general mass spectral databases, such as MassBank, which contain many compounds unlikely to be present in herbal extracts.

Mass spectrometry data from the QqTOF instrument was examined, with the auto MS/MS function being used to find compounds. Initially, 397 ‘compounds’ were detected (although some of these were duplicates or artefacts), 91 of which were matched with compounds from the in-house database. The matching was carried out by means of extracted ion chromatograms (EIC, also known as XIC) of compounds with the correct mass for those in the database, in conjunction with the MS/MS function.

The accurate mass measurements allowed isobaric species that were not isomers to be distinguished (e.g. C10H8O4 and C11H12O3). Examination of the fragmentation patterns allowed for some isomeric species to be distinguished, particularly in cases where comparison with other related compounds was possible. Various duplicate compounds were discarded, leading to a total of 60 compounds, 57 of which were tentatively identified. Many of these were alkaloids (43 out of 57), such as gelsemine, but non-alkaloids were also present, such as the iridoid gelsemide.

Complex mixture sorted with in-house database and LC-QqTOF-MS

The use of an in-house database based on previous studies allowed many compounds from a complex mixture of natural products to be identified by LC-QqTOF-MS. This method avoids complex and time-consuming isolation procedures, so that decomposition and rearrangements during ‘work-up’ are likely to be minimal. The method could be adapted for quality control of Gelsemium extracts isolated for medicinal or research purposes. Doubtless similar techniques could be employed for other natural products where there is an extensive, but fragmented, literature.

Related Links

J. Mass Spectrom., 2016, Early View Paper. Liu et al., Comprehensive identification and structural characterization of target components from Gelsemium elegans by high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry based on accurate mass databases combined with MS/MS spectra.

Wikipedia, Gelsemium

Article by Ryan De Vooght-Johnson

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

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