Urinary neopterin quantification by reverse-phase high-performance liquid chromatography with ultraviolet detection.
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 (n= 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 micromol/ml, respectively. Recovery from 79.5% to 82% was observed throughout the assay's linear range. Subjects with active (874.2 +/- 165.38 micromol/mol creatinin) and highly active SLE (1753.8 +/- 453.9 micromol/mol creatinin) showed three- and sixfold increased neopterin levels, respectively, compared to subjects with inactive SLE (314.3 +/- 121.3 micromol/mol creatinin) and healthy subjects (294.6 +/- 178.6 micromol/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.
de Castro
Marcel Ribeiro
MR
Department of Medicine, Nephrology Division, Universidade Federal de São Paulo, Escola Paulista de Medicina, Rua Botucatu, 740, 04023-900 São Paulo, SP, Brazil.
Di Marco
Giovana Seno
GS
Arita
Danielle Yuri
DY
Teixeira
Luciana Cristina
LC
Pereira
Aparecido Bernardo
AB
Casarini
Dulce Elena
DE
D015415:Biomarkers / Q000652:urine*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D008180:Lupus Erythematosus, Systemic / Q000175:diagnosis* / Q000652:urine*; D019798:Neopterin / Q000652:urine*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet / Q000379:methods*; D016482:Urinalysis / Q000379:methods*
D015415:Biomarkers; D019798:Neopterin