Quantitative determination of rhamnolipid using HPLC‐UV through carboxyl labeling

Abstract

Rhamnolipid, as a low‐toxic, biodegradable and environmentally friendly biosurfactant, has broad application prospects in many industries. However, the quantitative determination of rhamnolipid is still a challenging task. Here, a new sensitive method for the quantitative analysis of rhamnolipid based on a simple derivatization reaction was developed. In this study, 3‐[3′‐(l‐rhamnopyranosyloxy) decanoyloxy] decanoic acid (Rha‐C10‐C10) and 3‐[3′‐(2′‐O‐α‐l‐rhamnopyranosyloxy) decanoyloxy] decanoic acid (Rha‐Rha‐C10‐C10) were utilized as the representative rhamnolipids. Liquid chromatography–mass spectrometry and high‐performance liquid chromatography‐ultra violet results showed that these two compounds were successfully labeled with 1 N1‐(4‐nitrophenyl)‐1,2‐ethylenediamine. There was an excellent linear relationship between rhamnolipid concentration and peak area of labeled rhamnolipid. The detection limits of the Rha‐C10‐C10 and Rha‐Rha‐C10‐C10 were 0.018 mg/L (36 nmol/L) and 0.014 mg/L (22 nmol/L), respectively. The established amidation method was suitable for the accurate analysis of rhamnolipids in the biotechnological process. The method had good reproducibility with the relative standard deviation of 0.96% and 0.79%, respectively, and sufficient accuracy with a recovery of 96%–100%. This method was applied to quantitative analysis of 10 rhamnolipid homologs metabolized by Pseudomonas aeruginosa LJ‐8. The single labeling method was used for the quantitative analysis of multiple components, which provided an effective method for the quality evaluation of other glycolipids with carboxyl groups.