Development of a validated UPLC–qTOF-MS/MS method for determination of bioactive constituent from Glycyrrhiza glabra

Gupta, D.K.; Verma, M.K.; Anand, R.; Khajuria, R.K.

Glycyrrhetic acid used as a standard in the study showed separate peaks for a and b isomers in the extracted ion chromatogram (EIC) (Fig. 5A) . Glycyrrhizin appeared in the EIC at 1.17 min (Fig. 5C ). a and b isomers of glycyrrhetic acid exhibited peaks at retention times of 1.75 and 2.91 mins in the total ion chromatogram (TIC; Fig. 5D ), respectively. As the b isomer was predominant in case of glycyrrhetic acid and it was quantified in the (Fig. 5D ). The quality of extraction procedure of a plant material is reflected in the percentage of a compound of interest extractable by a particular process. After extraction, the desired compound can be purified by normal reported methods of purification. Extraction values of glycyrrhizin from G. glabra with water and 50% aqueous ethanol were almost the same as with ASE and USW and were observed to be 3.50%, 3.48%, 3.40% and 4.12%, 4.00%, 2.82%, respectively ( Table 3 ). The comparison of extraction values in 50% aqueous ethanol by CE at ambient temperature with extraction under USW and ASE in the same solvent exhibited no significant increase in the yield of glycyrrhizin. However, in acidic conditions (pH 0.22) a maximum of 3.00% of glycyrrhizin was extracted, whereas under basic conditions (pH 11.85) the extractive value was to the tune of 12.70% (Table 3) . As G. glabra is one of the major plants used commercially, the ASE method with ammonium hydroxide is the most appropriate method of extraction. By this process the amount of extractable glycyrrhizin increases by more than 50% as compared to extraction by CE under similar conditions. The amount of glycyrrhizin and glycyrrhetic acid in the extract obtained by different extraction methods (Table 4) with 50% ethyl alcohol was 4.12% and 0.19% by ASE, and 4.00% and 0.17% by USW whereas 1.71% and 0.11% was observed by CE, respectively. It is clear from the results that ASE with liquid ammonia is the most suitable method for extraction of glycyrrhizin from the roots of G. glabra.