Measurement of proenkephalin A-derived peptides in biological tissues by high pressure liquid chromatography coupled with amperometric electrochemical detection.
We describe an analytic method for the separation and quantitation of a number of proenkephalin A-derived peptides using high pressure liquid chromatography coupled with amperometric electrochemical detection (HPLC-AECD). Initially, we coupled our HPLC separation system with AECD in series with a UV detector for additional confirmation of peak specificity. AECD provided a 10(6)-fold increase in sensitivity over UV detection for these peptides. In addition to Met-enkephalin (ME), ME-Arg, ME-Arg-Phe, ME-Arg-Gly-Leu, Leu-enkephalin (LE) and LE-Arg (Dyn 1-6), we separated and detected the sulfoxides of ME and its extended peptides. Subsequently, we used minor modifications of the isocratic mobile phase to separate and detect enkephalin-related peptides with greater sensitivity and shorter chromatographic run times; each of these mobile phases was used to separate and detect two to three peptides. We have applied this HPLC-AECD methodology to quantitate ME, ME-Arg-Phe, ME-Arg-Gly-Leu and LE in pheochromocytoma tumors.
D000310:Adrenal Gland Neoplasms / Q000032:analysis; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004563:Electrochemistry; D004745:Enkephalins / Q000032:analysis*; D006801:Humans; D010673:Pheochromocytoma / Q000032:analysis; D011498:Protein Precursors / Q000032:analysis*
D004745:Enkephalins; D011498:Protein Precursors; C029992:proenkephalin