SENSITIVE DETERMINATION OF GABAPENTIN IN HUMAN URINE BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY WITH UV–VIS AND FLUORESCENT DETECTION

Abstract

Two rapid, sensitive, and reproducible methods for the determination of gabapentin (GBP) in urine based on high-performance liquid chromatography (HPLC) with UV-Vis and fluorescent detection, respectively, were developed for the first time using a new synthesized fluorescent label, 6-oxy-(N-succinimidylacetate)-9-(2′-methoxycarbonyl) fluorescein (SAMF). The highest derivatization efficiency was obtained in borate buffer (pH 7.25) at room temperature for 10 min. The chromatographic separation was carried out using a mixture of methanol and water containing 5 mmol l⁻¹ sodium citrate buffer (pH 5) as a mobile phase with UV-Vis detection at λ = 455 nm and fluorescent detection (FLD) at λ_ex/λ_em = 488/520 nm, respectively. A calibration curve ranging from 1 × 10⁻⁸ mol l⁻¹ to 1 × 10⁻⁶ mol l⁻¹was shown to be linear. The concentration limit of detection (LOD) was 1 × 10⁻⁸ mol l⁻¹ and 2.5 × 10⁻¹⁰ mol l⁻¹(signal to noise ratio = 3) for HPLC-UV-Vis and HPLC-FLD, respectively. The proposed methods were applied to the determination of GBP in urine samples with satisfying results.

Keywords:

• fluorescent detection
• gabapentin
• high-performance liquid chromatography
• 6-oxy-(N-succinimidyl acetate)-9-(2′-methoxycarbonyl) fluorescein
• UV-Vis detection

ACKNOWLEDGMENTS

This work was supported by the Fundamental Research Funds for the Central Universities.

Notes

^a x is the concentration of GBP (mol l⁻¹) and y is the peak area of GBP derivatives in the chromatogram.

^b PA is the peak area and RT the retention time.

^a PA is the peak area and RT is the retention time.

^a OPA-MPA is o-phthalaldehyde-3-mercaptopropionic acid.