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Clinical Chemistry 47: 1627-1632, 2001;
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(Clinical Chemistry. 2001;47:1627-1632.)
© 2001 American Association for Clinical Chemistry, Inc.

Articles

Quantitative Measurement of Porphobilinogen in Urine by Stable-Isotope Dilution Liquid Chromatography-Tandem Mass Spectrometry

Roddey E. Ford1, Mark J. Magera1, Karen M. Kloke1, Paul A. Chezick1, Abdul Fauq2 and Joseph P. McConnell1a

1 Biochemical Genetics Laboratory, Department of Laboratory Medicine and Pathology, Mayo Clinic, 200 First St. SW, Rochester, MN 55905.

2 Pharmacology Research, Mayo Clinic Jacksonville, Jacksonville, FL 32224.

aAuthor for correspondence. Fax 507-284-1399;

Background: Measurement of porphobilinogen (PBG) is useful in the diagnosis of the acute neurologic porphyrias. Currently used colorimetric assays lack analytical and clinical sensitivity and specificity.

Methods: We developed a liquid chromatography-electrospray tandem mass spectrometry (LC-MS/MS) method for the measurement of PBG in 1 mL of urine, using 5-(aminoethyl)-4-(carboxymethyl) 1H-2,4-[13C]pyrolle-3-propanoic acid ([2,4-13C]PBG; 2.75 µg) as internal standard. After solid-phase extraction, LC-MS/MS analysis was performed in the selected-reaction monitoring (SRM) mode. PBG and [2,4-13C]PBG were monitored through their own precursor and product ion settings (m/z 227 to 210 and m/z 229 to 212, respectively). The retention time of PBG and [2,4-13C]PBG was 1.0 min in a 2.3-min analysis.

Results: Daily calibrations (n = 6) between 0.1 and 2.0 mg/L were linear and reproducible. Inter- and intraassay CVs were 3.2–3.5% and 2.6–3.1%, respectively, at mean concentrations of 0.24, 1.18, and 2.15 mg/L. The regression equation for the comparison between an anion-exchange column method (y) and the LC-MS/MS method (x) was: y = 0.84x + 0.74 (Sy|x = 5.8 mg/24 h; r = 0.85; n = 100). In 47 volunteers, PBG excretion was 0.02–0.42 mg/24 h, lower than reported reference intervals (up to 2.0 mg/24 h) based on colorimetric methods. In 85 samples with PBG <=0.5 by LC-MS/MS, 8 (9.4%) had values >=2.0 mg/24 h by the anion-exchange method (mean ± SD, 4.3 ± 1.8 mg/24 h). In 11 patients with confirmed diagnoses of acute porphyria and increased PBG by LC-MS/MS, 2 had values within the reported reference intervals by a quantitative anion-exchange method.

Conclusions: The quantitative LC-MS/MS method for PBG measurement exhibits greater analytical specificity and improved clinical sensitivity and specificity than currently available methods.




The following articles in journals at HighWire Press have cited this article:


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M. Roshal, J. Turgeon, and P. M. Rainey
Rapid Quantitative Method Using Spin Columns to Measure Porphobilinogen in Urine
Clin. Chem., February 1, 2008; 54(2): 429 - 431.
[Abstract] [Full Text] [PDF]

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Y. Floderus, E. Sardh, C. Moller, C. Andersson, L. Rejkjaer, D. E.H. Andersson, and P. Harper
Variations in Porphobilinogen and 5-Aminolevulinic Acid Concentrations in Plasma and Urine from Asymptomatic Carriers of the Acute Intermittent Porphyria Gene with Increased Porphyrin Precursor Excretion
Clin. Chem., April 1, 2006; 52(4): 701 - 707.
[Abstract] [Full Text] [PDF]

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J. J. Pitt, M. Eggington, and S. G. Kahler
Comprehensive Screening of Urine Samples for Inborn Errors of Metabolism by Electrospray Tandem Mass Spectrometry
Clin. Chem., November 1, 2002; 48(11): 1970 - 1980.
[Abstract] [Full Text] [PDF]

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C. A. Kroll, M. J. Magera, J. K. Helgeson, D. Matern, and P. Rinaldo
Liquid Chromatographic-Tandem Mass Spectrometric Method for the Determination of 5-Hydroxyindole-3-acetic Acid in Urine
Clin. Chem., November 1, 2002; 48(11): 2049 - 2051.
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