Metabolite variation and discrimination of five licorice (Glycyrrhiza) species: HPTLC and NMR explorations

Authors: ZHAO, JIANPING - University Of Mississippi; Wang, Mei; SABASTIAN, JOHN - University Of Mississippi; LEE, JOSEPH - University Of Mississippi; CHITTIBOYINA, AMAR - University Of Mississippi; AVULA, BHARATHI - University Of Mississippi; ALI, ZULFIQAR - University Of Mississippi; LI, JING - Food And Drug Administration(FDA); WU, CHARLES - Food And Drug Administration(FDA); KHAN, IKHLAS - University Of Mississippi

Submitted to: Journal of Pharmaceutical and Biomedical Analysis
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/17/2022
Publication Date: 8/19/2022
Citation: Zhao, J., Wang, M., Sabastian, J.A., Lee, J., Chittiboyina, A.G., Avula, B., Ali, Z., Li, J., Wu, C., Khan, I. 2022. Metabolite variation and discrimination of five licorice (Glycyrrhiza) species: HPTLC and NMR explorations. Journal of Pharmaceutical and Biomedical Analysis. https://doi.org/10.1016/j.jpba.2022.115012.
DOI: https://doi.org/10.1016/j.jpba.2022.115012

Interpretive Summary: Licorice, the roots and rhizomes of several species of genus Glycyrrhiza Tourn. ex L. (Fabaceae family), is widely used as a sweetening and flavoring agent in food. It is also one of the most popular herbal medicines in the world. Glycyrrhiza species can be differentiated based on the morphologic features of their aerial parts, i.e. leaves, flowers, and fruits. But for the medicinal/commercial part of the plant - the root, the accurate identification of its species source is challenging due to the similarity in morpho-anatomical features of the roots derived from the five Glycyrrhiza species, viz. G. uralensis, G. glabra, G. inflata, G. echinata, and G. lepidota. It is not surprising that much of the commercially available licorice material is often mislabeled since raw materials from different species of Glycyrrhiza are sold on the market with the same common name "licorice roots". To assure the safe and effective utilization of licorice, it is important to develop simple and effective methods for determining the identity of different Glycyrrhiza species. This study investigated the metabolite differences of five Glycyrrhiza species using HPTLC and NMR analytical techniques. It was concluded that both the HPTLC and NMR fingerprinting approaches could be used to identify and discriminate licorice species. Particularly, as a reliable, rapid, and comparably simple analytical tool, the developed HPTLC method could be useful for species authentication and quality evaluation of licorice raw materials and products.

Technical Abstract: The roots and rhizomes of several Glycyrrhiza species are widely used as sweetening and flavoring agents in food, as well as important ingredients in formulations of traditional medicines. Five Glycyrrhiza species, G. uralensis, G. glabra, G. inflata, G. echinata, and G. lepidota, often share the name "licorice roots" in the botanicals’ marketplace. Unfortunately, misidentification/mislabelling is very common due to their similarities in morpho-anatomical features. Significant metabolite alterations among the different Glycyrrhiza species and their hybrids have been reported, suggesting that the biological activities could vary with the licorice roots or products derived from different species. Development of simple, effective methods for species identification and differentiation is of key importance. In this study, 78 licorice samples were investigated using HPTLC and NMR as analytical tools. Significant metabolite variations were observed between the five species. The species-specific fingerprint patterns for the five Glycyrrhiza species were determined with HPTLC and NMR; then applied to the sample identification and discrimination. The results obtained from these two orthogonal analytical methods agreed with each other. Furthermore, the NMR signals and the species-specific constituents that made significant contributions to the differentiation of the five Glycyrrhiza species were confirmed based on the multivariate analysis of the NMR spectral data. Using the established OPLS-DA models, the classification of hybrids was evaluated and confirmed. The developed methods, particularly the HPTLC method with its simplicity and low cost, could be used as a rapid and reliable approach for the authentication of licorice species and quality control of licorice raw material and products.