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.
Highlights
Autism Spectrum Disorder (ASD), which begins in infancy or before the first 3 years old, is neurodevelopmental disorders characterized by impaired social relatedness, and communication, repetitive behaviors, abnormal movement
Endogenous porphyrins have been a problem for porphyrins analysis, and make it difficult to determine the limit of quantif-ication
In this study, activated charcoal was used to remove the endogenous porphyrins from the pooled urine and plasma
Summary
Autism Spectrum Disorder (ASD), which begins in infancy or before the first 3 years old, is neurodevelopmental disorders characterized by impaired social relatedness, and communication, repetitive behaviors, abnormal movementAll MS Letters content is Open Access, meaning it is accessible online to everyone, without fee and authors’ permission. 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. Results obtained from the validation study of porphyrins showed good accuracy, precision, recovery, and stability.
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