A story, specific and fragile single-molecule checking (SIM) gas chromatography-mass spectrometry (GCMS) strategy was made and affirmed for the confirmation of imperativeness metabolites related to glycolysis, the tricarboxylic destructive (TCA) cycle, glutaminolysis, and unsaturated fat β-oxidation. The assessment was used to dismember liver tissue and serum from C57BL/6N bold mice dealt with a high-fat eating routine (HFD) and C57BL/6N mice dealt with common chow for around two months. HFD-dealt with mice serum demonstrated genuinely through and through diminished groupings of pyruvate, citrate, succinate, fumarate, and 2-oxoglutarate, with a raised union of pantothenic destructive. In liver tissue, HFD-dealt with mice indicated disheartened degrees of glycolysis conclusive outcomes pyruvate and lactate, glutamate, and the TCA cycle intermediates citrate, succinate, fumarate, malate, and oxaloacetate. Pantothenate levels were 3-overlay raised joined by an unassuming extended quality verbalization of Scl5a6 that encodes the pantothenate transporter SLC5A6. Since both glucose and unsaturated fats control coenzyme An association from pantothenate, it was assumed that these data were consistent with downregulated unsaturated fat β-oxidation, glutaminolysis, glycolysis, and TCA cycle development, on account of prevented anaplerosis. The epic SIM GCMS look at gave new bits of information into metabolic effects of HFD in mice.