Biomarker Profiling for Pyridoxine Dependent Epilepsy in Dried Blood Spots by HILIC-ESI-MS

International Journal of Analytical Chemistry
Elizabeth Mary MathewPragna Rao

Abstract

Pyridoxine dependent epilepsy is a condition where the affected infant or child has prolonged seizures (status epilepticus), which are nonresponsive to anticonvulsant therapy but can be treated with pharmacological doses of pyridoxine. If identified earlier and treated prophylactically with pyridoxine, severe brain damage due to seizures can be prevented. Alpha-amino adipic semialdehyde (AASA), piperidine-6-carboxylic acid (P6C), and pipecolic acid (PA) are known biomarkers of pyridoxine dependent epilepsy. We report the development and validation of a hydrophilic interaction liquid chromatography (HILIC) hyphenated with mass spectroscopy for the quantification of the above analytes from dried blood spot samples. The samples were extracted using methanol and analysed on a iHILIC fusion plus column with formic acid buffer (pH 2.5): acetonitrile (20:80) at a flow rate of 0.5 mL/min within 3 minutes. The method demonstrated a LOD of 10 ng/mL, LOQ of 50 ng/mL, linearity of r2 ≥ 0.990, and recovery of 92-101.98% for all analytes. The intra- and interday precision CVs were < 8% and 6%, respectively. Extensive stability studies demonstrated that the analytes were stable in stock solution and in matrix when stored at -80°C. We performe...Continue Reading

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