Development and Validation of an HPLC Method for Simultaneous Determination of Capsaicinoids and Camphor in Over-the-Counter Medication for Topical Use

A reverse-phase high-performance liquid chromatography method was developed to determine and quantify capsaicin (trans-8-methyl-N-vanillyl-6- nonenamid), dihydrocapsaicin (8-methyl-N-vanillylnonanamide), and camphor (trimethylbicyclo[2.2.1]heptan-2-one). It is applicable in analyses of over-the-counter (OTC) medications for topical use and raw materials such as chili pepper oleoresin. Chromatographic separation was carried out on a C18 column using an isocratic flow of the mobile phase containing acetonitrile and ultrapure water in a ratio of 2:3, with pH adjusted to 3.2 using glacial acetic acid, and a flow rate of 1.5 mL/min. The concentration of the eluting compounds was monitored by a diode-array detector at a wavelength of 281 nm. The method was evaluated for several validation parameters, including selectivity, accuracy (confidence intervals < 0.05%), repeatability, and intermediate precision. The limit of detection (LOD) was determined to be 0.070 mu g/mL for capsaicin, 0.211 mu g/mL for dihydrocapsaicin, and 0.060 mu g/mL for camphor. The limit of quantification (LOQ) was determined to be 0.212 mu g/mL for capsaicin, 0.640 mu g/mL for dihydrocapsaicin, and 0.320 mu g/mL for camphor. Linearity was set in the range of 2.5-200 mu g/mL for capsaicin and dihydrocapsaicin and 25-2000 mu g/mL for camphor. The suggested analytical method can be used for quality control of formulated pharmaceutical products containing capsaicinoids, camphor, and propolis.

Automated summary

A reverse-phase high-performance liquid chromatography method was developed for the determination and quantification of capsaicin, dihydrocapsaicin, and camphor. The method is applicable for the analysis of over-the-counter medications and raw materials such as chili pepper oleoresin. Various validation parameters were evaluated, demonstrating good selectivity, accuracy, repeatability, and intermediate precision.