Comparative GC-MS based nutrients profiling of less explored legume seeds of Melilotus, Medicago, Trifolium, and Ononis analysed using chemometric tools.

Exploring novel sources of plant protein for nutrition of both humans and animals is motivated mainly by its growing demand worldwide, besides identifying healthy alternatives for animal protein. The present study evaluates metabolome diversity within 15 legume seed species. The examined samples comprised three Melilotus, four Medicago, four Trifolium, and four Ononis seed species. A holistic approach for metabolites profiling using gas chromatography-mass spectrometry (GC-MS) led to the annotation and quantification of 87 metabolites comprising alcohols, free amino acids, aromatics, fatty acids/esters, nitrogenous compounds, organic acids, sugar alcohols, sugars, terpenes, and steroids. Fatty acids represented the major metabolite class represented by palmitic, stearic, oleic, linoleic, and linolenic acids. Sucrose and pinitol were the major sugars and sugar alcohols among seeds. Ononis seeds (OR, OS and OA) were the most abundant in fatty acids, sugars, sugar alcohols, and free amino acids, whereas Melilotus species (MO and MS) were least enriched in these key nutrients posing Ononis as potential food source for humans and animals. The examined seeds were generally low in sulfur-containing free amino acids and lacking many of the essential free amino acids. Multivariate data analysis aided in the identification of Ononis metabolite markers belonging to various classes i.e., (alcohol) glycerol, (sugar) allofuranose, and (sugar alcohol) pinitol, although the differentiation between Medicago, Melilotus, and Trifolium genera was not attained suggestive for other analytical platforms for its classification.

NMR Profiling of Ononis diffusa Identifies Cytotoxic Compounds against Cetuximab-Resistant Colon Cancer Cell Lines.

In the search of new natural products to be explored as possible anticancer drugs, two plant species, namely Ononis diffusa and Ononis variegata, were screened against colorectal cancer cell lines. The cytotoxic activity of the crude extracts was tested on a panel of colon cancer cell models including cetuximab-sensitive (Caco-2, GEO, SW48), intrinsic (HT-29 and HCT-116), and acquired (GEO-CR, SW48-CR) cetuximab-resistant cell lines. Ononis diffusa showed remarkable cytotoxic activity, especially on the cetuximab-resistant cell lines. The active extract composition was determined by NMR analysis. Given its complexity, a partial purification was then carried out. The fractions obtained were again tested for their biological activity and their metabolite content was determined by 1D and 2D NMR analysis. The study led to the identification of a fraction enriched in oxylipins that showed a 92% growth inhibition of the HT-29 cell line at a concentration of 50 µg/mL.