Simplified method for quantitation of angiotensin peptides in tissue

A simple method for extraction, separation, identification and quantitation of angiotensin-like immunoactivity from tissue is described. Homogenized acetic acid extracts of tissue samples were lyophilized and reconstituted in mobile phase. Separation was performed by reversed-phase high-performance liquid chromatography on a phenyl silica gel column with an eluent consisting of 20% acetonitrile in 0.1 M aqueous ammonium phosphate buffer, pH 4.9. Elution of standard peptides under isocratic conditions revealed clear resolution of angiotensin I, II and III and the (1-7) and (3-8) peptides. Recoveries of labeled angiotensin peptide standards from the extraction step were > 90%. Radioimmunoassay of relevant peaks revealed detectable levels of angiotensin I-, II- and III-like immunoactivity in single rat hypothalami and brain stems.