Validation of HPLC-UV Method for Analysis of 4-Pyridoxic Acid in Human Urine | Abstract

International Journal of Medical Research & Health Sciences (IJMRHS)
ISSN: 2319-5886 Indexed in: ESCI (Thomson Reuters)


Validation of HPLC-UV Method for Analysis of 4-Pyridoxic Acid in Human Urine

Author(s):Mohammad Shawaqfeh, Wenchen Zhao and Suhaib Muflih

We present a simple and validated HPLC method for measuring Vitamin B6 urinary metabolite 4-pyridoxic acid (4-PA). Urine samples were treated with 6% perchloric acid. HPLC separation was achieved using Waters column (Symmetry® C18 250 mm × 4.6 mm i.d (5 μM). The mobile phase was a mixture of methanol and 35 mM sodium phosphate buffer containing 2.5 mM sodium heptane polysulfonate at pH=3.5 with 85% O-phosphoric acid. Retention times were 8.0 min for 4-pyridoxic acid. Detection (UV at 302 nm) of analyte was linear in a range from 0.0125 to 0.8 μM. This method is stable, reproducible and has practical advantages such as ease and low cost.

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