Simultaneous separation of angiotensin and endothelin peptide families by high-performance liquid chromatography: application to the specific radioimmunoassay measurement of angiotensin II or endothelin-1 from tissue.
Currently available measurements of endogenous angiotensin II (ANG II) and endothelin-1 (ET-1) concentrations by radioimmunoassay (RIA) lack specificity to ANG II or ET-1. ANG II and ET-1 antibodies cross-react with immuno-reactive angiotensin and endothelin family members, respectively. We have therefore developed an ion-pair reversed-phase high-performance liquid chromatography (HPLC) for simultaneously separating angiotensin and endothelin peptides and enhancing RIA specificity in the measurement of ANG II and ET-1. The developed HPLC separation was applied to canine myocardium extracts; ANG II or ET-1 fractions were collected and quantified by RIA. Elution times for both peptide families, ANG I, ANG II, ANG III, ANG IV, ANG II(4-8), bET-1, ET-1, ET-2 and ET-3 were within 25 min. In normal canine myocardium from the right atrium, right ventricle, left atrium and left ventricle, ANG II concentrations were 39+/-11, 28+/-21, 31+/-11 and 21+/-8 fmol/g and ET-1 concentrations were 43+/-16, 42+/-19, 55+/-21 and 57+/-34 fmol/g (mean+/-SD, N=7), respectively. The combination of HPLC with RIA renders the measurement of ANG II or ET-1 specific and convenient, and saves time. This HPLC separation may be applied to the specific measurement of other immuno-reactive angiotensin and endothelin peptides.
Naik
G O
GO
St. Michael's Hospital and the Department of Medicine, University of Toronto, ON, Canada. george.naik@utoronto.ca
Moe
G W
GW
D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification*; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004285:Dogs; D019332:Endothelin-1 / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D008297:Male; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry; D011863:Radioimmunoassay / Q000379:methods*; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet
D019332:Endothelin-1; D000804:Angiotensin II