Reliable and reproducible techniques for identification and quantification of bioactive compound usually require derivatization. However, techniques such as LC–MS/MS may perhaps side line the derivatization with significant accuracy. Several physiological processes apparently requires that the pool be studied in the seed itself. The present analysis revealed a higher concentration of bioactive compound especially Auranofin, Beauvericin, β-Carotein, conessine, Hyperforin, Hypericin, Morphine, Panaxydol, Primaquine, ropivacaine, Siphonaxanthin, Sphingosine, Strophanthidin, Sulfamerazine, syringin, Trimethoprim,Tuftsin, vincristine bioactive compound. The study constitutes first report of use of LC–MS/MS method for analysing the bioactive compound of amaranth grain.