Analysis of mixtures of fatty acids and fatty alcohols in fermentation broth

• A simple protocol for simultaneous extraction of fatty acids and fatty alcohols from fermentation broth.
• A novel method to simultaneously quantify fatty acids and fatty alcohols with HPLC-RID.
• Fast quantification of fatty acids and fatty alcohols in fermentation broth in less than 40 min.
• The methods are fast, economical and widely applicable.

Microbial production of fatty acids and fatty alcohols has attracted increasing concerns because of energy crisis and environmental impact of fossil fuels. Therefore, simple and efficient methods for the extraction and quantification of these compounds become necessary. In this study, a high-performance liquid chromatography-refractive index detection (HPLC-RID) method was developed for the simultaneous quantification of fatty acids and fatty alcohols in these samples. The optimum chromatographic conditions are C18 column eluted with methanol:water:acetic acid (90:9.9:0.1, v/v/v); column temperature, 26 °C; flow rate, 1.0 mL/min. Calibration curves of all selected analytes showed good linearity (r2 ≥ 0.9989). The intra-day and inter-day relative standard deviations (RSDs) of the 10 compounds were less than 4.46% and 5.38%, respectively, which indicated that the method had good repeatability and precision. Besides, a method for simultaneous extraction of fatty acids and fatty alcohols from fermentation broth was optimized by orthogonal design. The optimal extraction conditions were as follows: solvent, ethyl acetate; solvent to sample ratio, 0.5:1; rotation speed, 2 min at 260 rpm; extraction temperature, 10 °C. This study provides simple and fast methods to simultaneously extract and quantify fatty acids and fatty alcohols for the first time. It will be useful for the study of microbial production of these products.