Simultaneous quantitative determination of multiple bioactive markers in Ocimum sanctum obtained from different...

Ocimum sanctum L., with phenolic acids, flavonoids, propenyl phenols and terpenoids as active pharmacological constituents, is a popular medicinal herb and is present as an ingredient in many herbal formulations. Therefore, development of a reliable analytical method for simultaneous determination of the pharmacologically active constituents of O. sanctum is of high importance.

To develop and validate a new, rapid, sensitive and selective UPLC-ESI/MS/MS method for simultaneous determination of 23 bioactive markers including phenolic acids, flavonoids, propenyl phenol and terpenoid in the leaf extract and marketed herbal formulations of O. sanctum.

An UPLC-ESI/MS/MS method using negative electrospray ionisation (ESI) in multiple-reaction-monitoring (MRM) mode was used for simultaneous determination. Chromatographic separation was achieved on an Acquity UPLC BEH C18 -column using a gradient elution with 0.1% formic acid in water and 0.1% formic acid in acetonitrile. Principal component analysis (PCA) was applied to correlate and discriminate eight geographical collections of O. sanctum based on quantitative data of the analytes.

The developed method was validated as per International Conference on Harmonization guidelines and found to be accurate, with overall recovery in the range 95.09-104.84% (RSD ≤ 1.85%), precise (RSD ≤ 1.98%) and linear (r(2)  ≥ 0.9971) over the concentration range of 0.5-1000 ng/mL. Ursolic acid was found to be the most abundant marker in all the samples investigated, except for the marketed tablet.

The method established is simple, rapid and sensitive, hence it can be reliably utilised for the quality control of O. sanctum and derived herbal formulations.