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  • P-ISSN 2233-4203
  • E-ISSN 2093-8950

Simultaneous Determination of Five Porphyrins in Human Urine and Plasma Using High Performance Liquid Chromatography-Tandem Mass Spectrometry

Mass Spectrometry Letters / Mass Spectrometry Letters, (P)2233-4203; (E)2093-8950
2014, v.5 no.2, pp.42-48
https://doi.org/10.5478/MSL.2014.5.2.42
Hur Yeoun (International Scientific Standards Co. Ltd.)
Tae Sookil (International Scientific Standards Co. Ltd.)
Koh Yun-Joo (The Korea Institute for children's Social Development and Rudolph Child Research Center)
Hong Sung-Hyun (Institute for EONE Laboratories)
Yoon Young Ho (Institute for EONE Laboratories)
Jang Haejong (International Scientific Standards Co. Ltd.)
Kim Sooji (International Scientific Standards Co. Ltd.)
Kim Kyeong Ho (Kangwon National University)
Kang Seung Woo (International Scientific Standards Co. Ltd.)
Lee Youngshin (International Scientific Standards Co. Ltd.)
Han Sang Beom (Chung-Ang University)
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Abstract

A specific and sensitive liquid chromatography-electrospray ionization tandem mass spectrometry method (LC-ESIMS/MS) was developed and validated for the simultaneous quantification of porphyrins (coproporphyrin, pentacarboxylporphyrin,hexacarboxylporphyrin, heptacarboxylporphyrin, and uroporphyrin) in human plasma and urine. Acidified plasma samplesand urine samples were prepared by using liquid-liquid extraction using ethyl acetate and protein precipitation with acetonitrile,respectively. The separation was achieved onto a Synergi Fusion RP column (150 mm × 2.0 mm, 4 μm) with a gradient elutionof mobile phase A (0.1% formic acid in 2 mmol/L ammonium acetate, v/v) and mobile phase B (20% methanol in acetonitrile, v/v) at a flow rate of 450 μL/min. Porphyrins and the internal standard (IS), coproporphyrin I-15N4, were detected by a tandemmass spectrometer equipped with an electrospray ion source operating in positive ion mode. Multiple reaction monitoring(MRM) transitions of the protonated precursor ions and the related product ions were optimized to increase selectivity and sensitivity. The proposed method was validated by assessing selectivity, linearity, limit of quantification (LOQ), precision, accuracy,recovery, and stability. The calibration curves were obtained in the range of 0.1-100 nmol/L and the LOQs were estimated as0.1 nmol/L for all porphyrins. Results obtained from the validation study of porphyrins showed good accuracy, precision, recovery,and stability. Finally, the proposed method was successfully applied to clinical studies on the autism spectrum disorder(ASD) diagnosis of 203 Korean children.

keywords
Porphyrins, tandem mass spectrometry, autism, diagnosis, plasma, urine


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Submission Date
2014-05-01
Revised Date
2014-05-01
Accepted Date
2014-06-01
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