Nicotinic acid, N-methylpyridinium ion, and trigonelline are well studied nutritional biomarkers present in coffee, and they are indicators of thermal decomposition during roasting. However, no method is yet available for their simultaneous determination. This paper describes a rapid and validated HPLC-diode array detector method for the simultaneous quantitation of caffeine, trigonelline, nicotinic acid, N-methylpyridinium ion, 5-caffeoylquinic acid, and 5-hydroxymethyl furfural that is applicable to three coffee matrixes: green, roasted, and instant. Baseline separation among all compounds was achieved in 30 min using a phenyl-hexyl RP column (250 × 4.6 mm, 5 μm particle size) , 0.3% aqueous formic buffer (pH 2.4) -methanol mobile phase at a flow rate of 1 mL/min, and a column temperature at 30°C. The method showed good linear correlation (r² > 0.9985) , precision (less than 3.9%) , sensitivity (LOD = 0.023-0.237 μg/mL; LOQ = 0.069-0.711 μg/mL) , and recovery (84-102%) for all compounds. This simplified method is amenable for a more complete routine evaluation of coffee in industry.