Absolute quantification of terpenes in conifer-derived essential oils and their antibacterial activity

Abstract

A simple, robust, and precise method for the absolute quantification of the terpenes α-pinene, camphene, β-pinene, 3-carene, limonene, bornyl acetate, β-caryophyllene, and borneol was developed using gas chromatography coupled to a flame ionization detector (GC-FID) and validated according to the International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use. The GC-FID method shows high accuracy (91–105%) and low imprecision (< 7.6%) for all terpenes at quality control (QC) low, medium, and high level. The curves are linear with strong correlation (R2 ≥ 0.999) for all terpenes. Additionally, the relative response factor (RRF) for each terpene is calculated. The method was validated in terms of specificity, linearity, accuracy, precision, LOD, LOQ, stability tests, and carry-over. The method was successfully applied to quantify the selected terpenes in conifer-derived essential oils (CEOs). The total amount of terpenes ranged from 6.3 to 11.8 mM. Additionally, the CEOs were screened in vitro for the antibacterial activity against E. coli and S. aureus using the broth microdilution method to determine the minimum inhibitory concentration (MIC). All the CEOs showed antibacterial activity in the concentration from 0.3 to 50 μg/mL (S. aureus) and 1.2 to 50 μg/mL (E. coli), respectively. CEO14 showed the most effective antibacterial activity of the entire tested CEOs (MIC values 0.3 μg/mL (S. aureus); 1.2 μg/mL (E. coli)). Our results suggest that the terpenoid profile should be considered for a comprehensive evaluation of the antibacterial activity, as none of the single compounds seems to be fully responsible for antibacterial activity.