ABSTRACT The Copaifera species are traditionally used for their anti‐inflammatory, antiseptic and healing properties. The objective of this study was to compare the chromatographic profiles of seven samples of copaiba oil‐resin with respect to chromatographic and physicochemical properties and to evaluate their biological activities. In addition, an HPLC method for quantifying β‐caryophyllene in the samples was developed and validated. The analysis revealed that most samples exhibited similar chromatographic profiles in TLC and HPLC‐DAD, while GC–MS identified β‐caryophyllene in all the samples. The analytical method presented adequate linearity ( R 2 = 0.9996), with no significant matrix effect. The β‐caryophyllene content varied from 3.36% to 37.12% across samples. A substantial majority of samples exhibited antimicrobial activity, with similar cytotoxicity observed in RAW 264.7 cells. In silico predictions indicated a potential for toxicity in the oil‐resin. The results highlight the importance of physicochemical and chromatographic analyses for quality control of copaiba oil‐resin.