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Sensitive determination of (−)‐isoproterenol and (−)‐( R )‐1‐(3,4‐dihydroxyphenyl)‐2‐[(3,4‐dimethoxyphenethyl)amino] ethanol (T‐0509), a cardiotonic agent, in rat plasma utilizing a fully automated catecholamine analyser
Author(s) -
Prados Pablo,
Higashidate Sakae,
Imai Kazuhiro,
Sato Yoji,
Nagao Taku
Publication year - 1994
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.1130080113
Subject(s) - chemistry , ethanol , catecholamine , chromatography , analyser , biochemistry , medicine
(−)‐Isoproterenol (ISO), used as bronchodilator, and (−)‐( R )‐1‐(3,4‐dihydroxyphenyl)‐2‐ [(3,4‐dimethoxyphenethyl)amino]ethanol (T‐0509), a new cardiotonic agent, were determined in rat plasma (only 25 μL) using a fully automated catecholamine (CA) analyser that includes in‐line pre‐extraction of CAs, coupled with an HPLC–ethylenediamine condensation reaction and a peroxyoxalate chemiluminescence detection system (HPLC–ED–PO‐CL). The chromatographic conditions were: precolumn, SERUMOUT‐CEX; buffer for delivering CAs in the precolumn, 10 m M potassium phosphate buffer (pH 7.5)/ethanol (92 + 8, v/v) (1 mL/min); precolumn clean‐up solution, 4% phosphoric acid/acetonitrile (50 + 50, v/v) (1 mL/min); analytical column, TSKgel ODS‐80Ts, 250 × 4.6 mm i.d.; eluent, 75 m M potassium acetate buffer (pH 3.2)/50 m M potassium phosphate buffer (pH 3.2)/acetonitrile (83.6 + 4.4 + 12, v/v/v for ISO and 76 + 4 + 20 v/v/v for T‐0509) (1 mL/min); fluorogenic reagent solution, 105 m M ED and 175 m M imidazole in acetonitrile/ethanol (90 + 10, v/v) (0.25 mL/min); chemiluminogenic reaction solution, 0.25 m M TDPO, 150 m M hydrogen peroxide and 110 m M TFA in dioxane/ethyl acetate (50 + 50, v/v) (1.4 mL/min). Colterol, (±)‐1‐(3,4‐dihydroxyphenyl)‐2‐[(1,1‐dimethyl)ethylamino]ethanol, (±)‐ N ‐ t ‐butylnorepinephrine was used as an internal standard (IS) for ISO, and (±)‐1‐(3,4‐dihydroxyphenyl)‐2‐ [(3,4,5‐trimethoxyphenethyl)amino]ethanol, (±)‐(3,4,5‐trimethoxyphenethylaminomethyl)‐3,4‐dihydroxybenzylalcohol (T‐1583) for T‐0509. The detection limits for ISO and T‐0509 were 1.3 and 0.9 fmol on injection, respectively.