Urinary neopterin quantification by reverse-phase high-performance liquid ultraviolet chromatography with detection

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Castro, M. R. de
Di Marco, G. S.
Arita, D. Y.
Teixeira, L. C.
Pereira, A. B.
Casarini, D. E.
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Neopterin plays an important role in the malignant disease diagnostics. However, the methods employed in neopterin determination are generally difficult and/or time consuming. the aim of this work was to standardize a practical method to quantify neopterin using high-performance liquid chromatography-ultraviolet (HPLC-UV) and quantify it in patients with systemic lupus erythematosus (SLE). Urine was collected from healthy subjects (n=49), patients with inactive (n = 15), active (n = 28), and highly active SLE (it = 6). the HPLC was performed using two coupled reverse-phase columns eluted with 150 mM sodium phosphate, pH 4.0, under a flow rate of 0.8 ml/ min, with UV detector set at 353 nm and 100-fold diluted urines. the inter- and intra-assay studies presented an imprecision of 12.5% and 12.9% for quality controls of 3.94 and 1.1 mumol/ml, respectively. Recovery from 79.5% to 82% was observed throughout the assay's linear range. Subjects with active (874.2 +/- 165.3 8 mumol/mol creatinin) and highly active SLE (1753.8 +/- 453.9 mumol/mol creatinin) showed three- and sixfold increased neopterin levels, respectively, compared to subjects with inactive SLE (314.3 mu 121.3 mumol/mol creatinin) and healthy subjects (294.6 +/- 178.6 mumol/mol creatinin) (P > 0.05). Briefly, the proposed method was precise, specific, and reproducible, not invasive and allows the urinary neopterin quantification only with UV detection, (C) 2004 Elsevier B.V. All rights reserved.
Journal of Biochemical and Biophysical Methods. Amsterdam: Elsevier B.V., v. 59, n. 3, p. 275-283, 2004.