Preparation and evaluation of an immunoaffinity sorbent for the analysis of opioid peptides by on-line immunoaffinity solid-phase extraction capillary electrophoresis–mass spectrometry

In this study, we explored a procedure for the preparation of an immunoaffinity (IA) sorbent for the analysis of opioid peptides by on-line immunoaffinity solid-phase extraction capillary electrophoresis–mass spectrometry (IA-SPE-CE–MS). We followed a site-specific antibody immobilization approach based on the covalent attachment of the oxidized antibodies through their carbohydrate moieties to hydrazide silica particles, using a polyclonal antibody against Endomorphin 1 and 2 (End1 and End2). The main features of the IA sorbent were studied, such as the amount of hydrazide groups and antibodies attached onto oxidized diol silica particles. Once the procedure was optimized, standard solutions of End1 and End2 were used in order to establish the IA-SPE-CE–MS methodology. Acceptable repeatability, reproducibility and linearity range values were obtained for the proposed methodology. The limits of detection (LODs) of 1 ng mL−1 were approximately 100-fold better than those obtained by CE–MS. Selectivity of the IA sorbent was good but some cross-reactivity against Dynorphin A (1–7) was observed when a mixture of several opioid peptides was analyzed. Human plasma samples spiked with End1 and End2 were also analyzed and both peptides could be detected down to 100 ng mL−1.

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► We prepared an IA sorbent for the analysis of opioid peptides by IA-SPE-CE–MS.
► We followed a site-specific antibody immobilization approach.
► The main features of the IA sorbent were studied.
► Endomorphins 1 and 2 were analyzed in standards and plasma samples to achieve the best LODs.