, 15:116 | Cite as

Combining untargeted and targeted metabolomics approaches for the standardization of polyherbal formulations through UPLC–MS/MS

  • Saeedur Rahman
  • Faraz Ul Haq
  • Arslan Ali
  • Muhammad Noman Khan
  • Syed Muhammad Zaki Shah
  • Achyut Adhikhari
  • Hesham R. El-Seedi
  • Syed Ghulam MusharrafEmail author
Original Article



Polyherbal formulations are an integral part of various indigenous medicinal systems such as Traditional Chinese Medicine (TCM) and Ayurveda. The presence of a very large number of compounds makes the quality control of polyherbal formulations very difficult.


To overcome this problem, we have developed a comprehensive strategy for the dereplication of natural products in polyherbal formulations by using Adhatoda vasica as a case study.


The strategy is based on five major steps: the collection of plant samples from different locations to observe the effects of environmental variables; LC–ESI–MS/MS-based untargeted metabolite profiling of the plant samples to identify marker compounds using extensive chemometric analysis of the obtained data; the identification of marker compounds in polyherbal products; the isolation, purification and characterization of the marker compounds; and MRM-based quantitative analysis of the isolated marker compounds using LC–ESI–MS/MS.


Using this strategy, we identified a total of 51 compounds in the methanolic extract of A. vasica plants from 14 accessions. Chemical fingerprinting of the plant led to the identification of characteristic peaks that were used to confirm the presence of A. vasica in complex polyherbal formulations. Four quinazoline alkaloids (marker compounds) were isolated, purified and quantified in various herbal formulations containing A. vasica.


This method demonstrates a comprehensive strategy based on untargeted and targeted metabolite analysis that can be used for the standardization of complex polyherbal formulations.


Chemical profiling Standardization Identification Quantification Polyherbal formulation 



The authors express gratitude to Mr. Arsalan Tahir and Mr. Junaid Ul Haq for technical assistance in UHPLC–MS/MS analyses. Mr. Faraz Ul Haq would also like to acknowledge the Higher Education Commission (HEC), Pakistan for an Indigenous PhD Fellowship.

Author contributions

SGM proposed the subject, designed the study and actively participated in manuscript reviewing. SR, FUH, MNK and SMZS performed the experiments and actively involved in the write-up of the manuscript. AA, AA and HRES assisted in reviewing the manuscript. All authors reviewed the manuscript.


This work was supported by the Higher Education Commission (HEC), Pakistan under the Industry Academic & Government Linkages program (HEC/R&D/UITSP/03/2016).

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Ethical approval

No animals were used/harmed in this study. This study did not involve any human participants.

Supplementary material

11306_2019_1582_MOESM1_ESM.docx (3.4 mb)
Supplementary material 1 (DOCX 3445 kb)


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Saeedur Rahman
    • 1
  • Faraz Ul Haq
    • 1
  • Arslan Ali
    • 2
  • Muhammad Noman Khan
    • 1
  • Syed Muhammad Zaki Shah
    • 1
  • Achyut Adhikhari
    • 1
  • Hesham R. El-Seedi
    • 3
    • 4
  • Syed Ghulam Musharraf
    • 1
    • 2
    Email author
  1. 1.H.E.J. Research Institute of Chemistry, International Center for Chemical and Biological SciencesUniversity of KarachiKarachiPakistan
  2. 2.Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological SciencesUniversity of KarachiKarachiPakistan
  3. 3.Pharmacognosoy Group, Department of Medicinal ChemistryUppsala University, Biomedical CentreUppsalaSweden
  4. 4.Alrayan Medical CollegeMedinaKingdom of Saudi Arabia

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