Introduction Astragalus anthylloides, A. dipsaceus, A. karamasicus, A. lycius, A. sigmoideus and A. xylobasis var. angustus are an endemic and generally grow in the Irano‐Turanian phytogeographic region of Turkey. Astragalus species contain saponins, polysaccharides, and phenolics, while the toxic compounds include imidazoline alkaloids, nitro toxins, and selenium derivatives. Objectives To apply a combined metabolomic fingerprinting approach by nuclear magnetic resonance (NMR) and gas chromatography‐mass spectrometry (GC‐MS) of endemic six Astragalus species extract. Methodology The whole plant collected in Turkey of six endemic Astragalus subsp. were dried and then extracted with hexane, chloroform, ethylacetate, n‐butanol and methanol solvents, respectively. The hexane extracts were analyzed by GC‐MS. Carbon‐13 (13C)‐NMR analyzes of all extracts were performed. In both analyses, a biomarker was obtained. Results The hexane extracts were determined as palmitic acid, arachidic acid, behenic acid, and linolenic acid as the main components. As a result of 13C‐NMR analyzes, in hexane, chloroform, and ethylacetate the extracts detected were palmitic acid, arachidic acid, behenic acid, and linolenic acid. d‐Pinitol was obtained using 13C‐NMR analyzes with n‐butanol and methanol extracts. Conclusion This study demonstrated that d‐pinitol is a biomarker for the endemic six Astragalus subsp.