A rapid and simple ultra high performance liquid chromatography method for the simultaneous determination of methoxyphenol derivatives involved in the eugenol catabolic pathway

An efficient ultra high performance liquid chromatography method of separation was developed for the analysis of six important methoxyphenol derivatives involved in the eugenol catabolic pathway. In the present study, an Acquity UPLC BEH C18 column was used for the chromatographic separation of the industrially important phenolic compounds such as vanillin, vanillic acid, ferulic acid, coniferyl alcohol, and coniferyl aldehyde obtained during microbial transformation of eugenol. Eluted components were identified using the dual wavelength (254 and 310 nm) UV detector. A gradient method of elution using mobile phase of aqueous 1 mM trifluoroacetic acid (Solvent A) and methanol (Solvent B) at a flow rate of 0.3 mL/min separated all the five intermediate methoxyphenol derivatives along with their precursor eugenol within 15 min with stable baseline resolution. Method validation was performed for the accurate quantification of vanillin, coniferyl aldehyde, and eugenol using the parameters of linearity, specificity, precision, limit of detection, limit of quantification, and robustness. The developed method would be helpful for clear separation and identification of the five most important intermediate metabolites of the eugenol catabolism pathway.