Electromembrane extraction of human chorionic gonadotropin: A case study on mass transfer of intact protein versus signature peptide

For the first time, we present targeted protein detection by tryptic digestion of human chorionic gonadotropin (hCG) followed by electromembrane extraction (EME). Operational parameters were optimized, and urine and serum samples spiked with hCG underwent tryptic digestion followed by EME of the βT5 signature peptide. The liquid membrane comprised nitrophenyl octyl ether (NPOE), carvacrol, and di(2-ethyl hexyl) phosphate (DEHP) at ratios of 49:49:2 (w/w/w). Extractions were performed in a conductive vial format for 45 min at 5 V. Even from highly complex digested samples of serum and urine, the signature peptide βT5 was extracted by EME and detected by LC-MS/MS. While attempts to extract intact hCG protein were unsuccessful, the extraction of the signature peptide was efficient. The extraction recovery from undigested and digested urine was 71 % (RSD = 17 %) and 116 % (RSD = 17 %), respectively. For serum, the extraction recoveries were 11 % (RSD = 23 %) for undigested samples and 110 % (RSD = 14 %) for digested samples. This study demonstrates both the potential and challenges of EME for protein analysis. Experiments regarding EME of intact proteins provided new insights into protein phase distribution. This fundamental case study underscores the potential of EME as a sample preparation technique for the targeted determination of protein biomarkers and drugs.