Investigation of bioactive fractions from fenugreek seeds (Trigonella foenum-graecum L.): chemical profiling, in vitro anti-inflammatory activity, and their potential against LPS-stimulated J774A.1 cells

Despite the known anti-inflammatory effect of fenugreek seeds ( Trigonella foenum-graecum L.), the specific mechanism and compounds responsible for this effect have been less studied so far. This research aimed to fractionate the total methanolic extract and investigate their impact on the iNOS expression in J774A.1 macrophage induced by lipopolysaccharide (LPS). The fractions of the total extract were obtained using semi-preparative HPLC. J774A.1 cells were treated with the highest non-toxic concentration of the samples and bacterial LPS (5 μg/ml). After 24 h, treated and untreated cells were used for RNA extraction, and mRNA levels were determined by qRT-PCR analysis. A total of 30 fractions were obtained with 9 main fractions (F3, F6, F9, F12, F15, F18, F21, F24, and F27) selected for cell studies. F9, total extract, and F15 significantly decreased iNOS gene expression compared to the LPS-treated group (up to 99.3%, 84.7% and 76.5% respectively). Overall, these results suggest that the anti-inflammatory effect of fenugreek seeds and/or fractions may be partially due to the suppression of iNOS. This effect can be attributed to various plant compounds, and further isolation may help identify novel anti-inflammatory compounds become a new inflammation. These findings support the potential of both the extract and the fractions as promising candidates for the treatment of inflammation-related diseases in future research.