Determination of 1,3-Dioleic acid-2-palmitoyl triglyceride in Infant Formula by High Performance Liquid Chromatography

A method for the quantification of 1,3-dioleyl-2-palmitoyl-glycerol (OPO) in infant formula was developed. The samples were treated with ammonia and extracted with organic solvents. The fat containing OPO was purified on a NH2 solid-phase extraction (SPE) cartridge packed with aminopropyl as the sorbent. The eluate was separated by silver ion chromatography using 0.55% acetonitrile-hexane as the mobile phase. The detection was carried out with a high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD). This novel procedure enabled the complete separation of OPO and its isomer 1,2-dioleyl-3-palmitoyl-glycerol (OOP), thus allowing for the accurate quantification of OPO. The developed method showed the desired linearity in the concentration range of 25–500 μg/mL with a determination coefficient (R2) of 0.999 6. The limits of detection (LOD) and limits of quantification (LOQ) were 0.30 and 0.90 g/kg, respectively. At spiked concentrations from 1 to 96 g/kg, the average recoveries of OPO varied from 97.1% to 104.2% with relative standard deviations (RSD) between 1.2% and 2.9%. The precision and accuracy of this method met the relevant requirements, and it passed the inter-laboratory collaborative validation. Our investigation analyzed 39 commercial samples of OPO-fortified infant formula in China, revealing that the measured OPO content only accounted for 28.4% to 59.7% of the labelled value, which is mainly due to the inconsistency of detection methods.