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

Marcel Ribeiro de Castro1, Giovana Seno Di Marco, Danielle Yuri Arita

  • 1Department of Medicine, Nephrology Division, Universidade Federal de São Paulo, Escola Paulista de Medicina, Rua Botucatu, 740, 04023-900 São Paulo, SP, Brazil.

Journal of Biochemical and Biophysical Methods
|May 29, 2004
PubMed
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This study presents a practical high-performance liquid chromatography-ultraviolet (HPLC-UV) method for measuring neopterin (a biomarker for disease) in urine. The new method shows elevated neopterin levels in patients with active systemic lupus erythematosus (SLE).

Area of Science:

  • Biochemistry
  • Clinical Chemistry
  • Analytical Chemistry

Background:

  • Neopterin is a significant biomarker in disease diagnostics, particularly for malignant conditions.
  • Current methods for neopterin determination are often complex and time-consuming.
  • Systemic lupus erythematosus (SLE) is an autoimmune disease where monitoring disease activity is crucial.

Purpose of the Study:

  • To standardize a practical and efficient method for quantifying urinary neopterin using high-performance liquid chromatography-ultraviolet (HPLC-UV).
  • To assess neopterin levels in patients with varying degrees of systemic lupus erythematosus (SLE) activity.
  • To establish the precision, specificity, and reproducibility of the developed HPLC-UV assay for neopterin.

Main Methods:

  • Urine samples were collected from healthy individuals and patients with inactive, active, and highly active SLE.

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  • High-performance liquid chromatography (HPLC) was performed using coupled reverse-phase columns with UV detection at 353 nm.
  • Assay validation included inter- and intra-assay variability, recovery studies, and linear range assessment.

    • Main Results:

    • The developed HPLC-UV method demonstrated good precision (imprecision < 12.9%) and recovery (79.5%–82%).
    • Patients with active and highly active SLE exhibited significantly elevated urinary neopterin levels compared to those with inactive SLE and healthy subjects.
    • Specifically, highly active SLE patients showed a sixfold increase in neopterin levels compared to healthy controls.

    Conclusions:

    • The proposed HPLC-UV method is a precise, specific, and reproducible assay for urinary neopterin quantification.
    • This non-invasive method simplifies neopterin measurement, relying solely on UV detection.
    • Elevated urinary neopterin levels correlate with SLE disease activity, suggesting its potential as a biomarker for monitoring SLE.