Time-controlled online SPE: strategic approach for N-acylethanolamines quantification in complex matrices.

Area of Science:

• Analytical Chemistry
• Biochemistry
• Veterinary Science

Background:

• Accurate quantification of N-acylethanolamines (NAEs) in biological fluids is vital for understanding their health and disease roles.
• Biological matrices present challenges for NAE analysis, often requiring improved pretreatment methods.
• N-acylethanolamines (NAEs) are key lipid signaling molecules implicated in various physiological processes.

Purpose of the Study:

• To develop and validate a novel analytical method for quantifying N-acylethanolamines (NAEs) in equine plasma.
• To specifically measure Palmitoyl Ethanolamide (PEA) and Oleoyl Ethanolamide (OEA) levels in horses.
• To advance the understanding of the endocannabinoid system in equines.

Main Methods:

• Utilized toluene liquid-liquid extraction for initial lipid isolation.
• Implemented Time-Controlled Online Solid Phase Extraction (SPE) coupled with HPLC-MS/MS.
• Developed a temporal SPE strategy to retain interferents while eluting target NAEs.

Main Results:

• Achieved excellent linearity, high recovery, and suitable matrix effects during method validation.
• Established low limits of detection (LOD) and quantification (LOQ) for PEA and OEA.
• Successfully quantified endogenous PEA and OEA levels in equine plasma, providing novel data.

Conclusions:

• The novel online SPE method enables reliable quantification of NAEs in complex equine plasma matrices.
• This method offers potential for analyzing other challenging biological samples with similar interferents.
• The findings significantly contribute to equine endocannabinoid research and future studies.