Liquid chromatographic fluorescence determination of amino acids in plasma and urine after derivatization with phanquinone.
Phanquinone (4,7-phenanthroline-5,6-dione) has been investigated as a pre-column derivatization fluorogenic reagent for liquid chromatographic determination of primary amino acids in biological samples. The derivatization reaction was carried out at 68 degrees C both in the presence of aqueous phosphate buffer (pH 8) for 30 min and without buffer for 60 min to allow the determination of basic amino acids (Orn, Lys, Arg). The resulting derivatives were separated under reversed-phase HPLC and detected at lambda(em) = 460 nm with lambda(ex) = 400 nm. The proposed method was validated and applied to the determination of a variety of amino acids directly in urine and after deproteinization with 5-sulfosalicylic acid in plasma samples. The detection and quantitation limits were found in the range 10-450 and 35-1400 fmol, respectively.
Gatti
Rita
R
Dipartimento di Scienze Farmaceutiche, Alma Mater Studiorum-Università di Bologna, Via Belmeloro 6, 40126 Bologna, Italy. rita.gatti2@unibo.it
Gioia
Maria Grazia
MG
D000596:Amino Acids / Q000097:blood* / Q000302:isolation & purification / Q000652:urine*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D007202:Indicators and Reagents; D010618:Phenanthrolines; D013050:Spectrometry, Fluorescence / Q000379:methods*
D000596:Amino Acids; D007202:Indicators and Reagents; D010618:Phenanthrolines; C004902:phanquinone