Validated HPLC Methodology for Pregabalin Quantification in Human Urine Using 1-Fluoro-2,4-dinitrobenzene Derivatization

Mohamed S. Mostafa; Hosam A. Mohamed; Olfat E. Mostafa; Shimaa Talaat; Mohamed A. Abdelaziz

doi:10.22456/2527-2616.143059

Authors

Mohamed S. Mostafa Consultant of Biochemistry Poison Control Center - Ain Shams University hospitals
Hosam A. Mohamed Assistant Consultant of Biochemistry Poison Control Center - Ain Shams University hospitals
Olfat E. Mostafa Fellow of Biochemistry, Poison Control Center - Ain Shams University hospitals
Shimaa Talaat Fellow of Biochemistry, Poison Control Center - Ain Shams University hospitals https://orcid.org/0000-0002-2701-8097
Mohamed A. Abdelaziz Fellow of Biochemistry, Poison Control Center - Ain Shams University hospitals https://orcid.org/0000-0001-5964-8256

DOI:

https://doi.org/10.22456/2527-2616.143059

Keywords:

Pregabalin, 1-Fluoro-2,4-dinitrobenzene, derivatization, validation, HPLC analysis

Abstract

Pregabalin (PGB) is a synthetic drug used for the treatment of central nervous system disorders and neuropathic pain. PGB is metabolized to N-Methyl pregabalin while the rest is excreted virtually unchanged in the urine. Numerous analytical techniques for measuring pregabalin have been documented. This study aimed to validate a simple, sensitive, and accurate method for PGB quantification in human urine using the HPLC technique with 1-Fluoro-2, 4-dinitrobenzene used as a derivatizing agent. One hundred and twenty urine samples were analyzed by a reversed-phase (C18) column and a mixture of acetonitrile and 50 mM KH2PO4 (pH 2.5) (60:40, v/v) as mobile phase and the flow rate was 1 ml/min and the UV detector wavelength was set to 360nm. The procedure was linear within the 10-1000 μg/ml range of PGB in urine (r > 0.99). Intraday and interday RSD precision values fell between 2.8% and 5.9%. 2.5 and 1.5 μg/ml, respectively, were determined to be the method's limits of quantification and detection. The recovery (90.8%) and statistical characteristics show that the suggested method has excellent accuracy and precision. The method is accurate, precise, reproducible, and specific, and it can be applied to regular examinations of pregabalin in urine samples.

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References

Chalabianloo F, and Schjott J. Pregabalin and its potential for abuse. Tidsskr Nor Laegeforen. 2009; 129(3):186-187.

Gahr M, Freudenmann RW, Hiemke C, Kolle MA, Schonfeldt-Lecuona C. Pregabalin abuse and dependence in Germany: results from a database query. Eur J Clin Pharmacol. 2013; 69(6):1335-42. https://doi:10.1007/s00228-012-1464-6.

Schwan S, Sundstrom A, Stjernberg E, Hallberg E, Hallberg P. A signal for an abuse liability for Pregabalin results from the Swedish spontaneous adverse drug reaction reporting system. Eur J Clin Pharmacol. 2010; 66(9):947-53. https://doi:10.1007/s00228-010-0853-y

Gujral RS, Haque SM, Kumar S. A novel method for the determination of pregabalin in bulk pharmaceutical formulations and human urine samples. Afr. J. Pharm. Pharmacol. 2009; 3(6):327- 334. http://www.academicjournals.org/ajpp

Pauly C, Yegles M, Schneider S. Pregabalin determination in hair by ultra-high-performance liquid chromatography – tandem mass spectrometry. J Anal Toxicol. 2013; 37(9):676-9. http://doi:10.1093/jat/bkt081

Priez-Barallon C, Carlier J, Boyer B, Benslima M, Fanton L, Mazoyer C, Gaillard Y. Quantification of pregabalin using hydrophilic interaction HPLC-HighResolution MS in postmortem human samples: Eighteen case reports. J Anal Toxicol. 2014; 38(3):143-8. http://doi:10.1093/jat/bku004.

Ahmadkhaniha R, Mottaghi S, Zargarpoor M, Souri E. Validated HPLC Method f or Quantification of Pregabalin in Human Plasma U sing 1-Fluoro-2,4- dinitrobenzene as Derivatization Agent. Chromatography Research International. Article ID 450461, 2014 (1) 6 pages. http//:doi:10.1155/2014/450461.

Ben-Menachem E. Pregabalin pharmacology and its relevance to clinical practice. Epilepsia. 2004;45(6):13-18. http//:doi:10.1111/j.0013- 9580.2004.455003.x

Piskorska B, Miziak B, Czuczwar SJ, Borowicz KK. Safety issues around misuse of antiepileptics. Expert Opin Drug Saf. 2013;12(5):647-57. http//:doi:10.1517/14740338.2013.796363.

Loftus H, Wright A. Potential misuse of pregabalin and gabapentin. BMJ. 2014; 5:348:g1290. http//:doi:10.1136/bmj.g1290

Kolocouri F, Dotsikas Y, Loukas YL. Dried plasma spots as an alternative sample collection technique for the quantitative LC–MS/MS determination of gabapentin. Anal Bioanal Chem. 2010; 398(3):1339- 47. doi:10.1007/s00216-010-4048-2.

Wattananat T, Akarawut, W. Validated LC–MS–MS method for the determination of gabapentin in human plasma: application toa bioequivalence study. J Chromatogr Sci. 2009; 47(10):868-71. http//:doi:10.1093/chromsci/47.10.868.

Oertel R, Arenz N, Pietsch J, Kirch W. Simultaneous determination of three anticonvulsants using hydrophilic interaction LC–MS. J Sep Sci. 2009;32(2):238-43. http//:doi:10.1002/jssc.200800461.

Peterson BL. Prevalence of gabapentin in impaired driving casesinWashington State in 2003–2007. J Anal Toxicol. 2009; 33(8):545-9. http//:doi:10.1093/jat/33.8.545.

Kushnir MM, Crossett J, Brown PI, Urry FM. Analysis of gabapentin in serum and plasma by solidphase extraction andgas chromatography–mass spectrometry for therapeutic drug monitoring. J Anal Toxicol. 1999; 23(1):1-6. http//:doi:10.1093/jat/23.1.1

Wolf CE, Saady JJ, Poklis A. Determination of gabapentinin serum using solid-phase extraction and gas–liquid chromatography. J Anal Toxicol. 1996; 20(6):498-501. http//:doi:10.1093/jat/20.6.498.

Vermeij TA, Edelbroek PM. Simultaneous highperformanceliquidhromatographic analysis of pregabalin, gabapentin and vigabatrinin human serum by precolumn derivatization with ophtaldialdehydeand fluorescence detection. J Chromatogr B Analyt Technol Biomed Life Sci. 2004; 810(2):297-303. http//:doi:10.1016/j.jchromb.2004.08.018.

Lensmeyer GL, Kempf T, Gidal BE, Wiebe DA. Optimizedmethod for determination of gabapentin in serum by highperformanceliquid chromatography. Ther Drug Monit. 1995; 17(3):251-8.

Chang SY, Wang FY. Determination of gabapentin in humanplasma by capillary electrophoresis with laserinduced fluorescencedetection and acetonitrile stacking technique. J Chromatogr B Analyt Technol Biomed Life Sci. 2004; 799(2):265-270. http//:doi:10.1016/j.jchromb.2003.10.052.

Rastkari N, Khoobi M, Shafiee A, Khoshay MR, khaniha RA. Development and validation of a simple andsensitive HPLC-UV method for the determination of captoprilin human plasma using a new derivatizing reagent 2-naphthylpropiolate. J Chromatogr B Analyt Technol Biomed Life Sci. 2013; 932(1):144-151. http//:doi:10.1016/j.jchromb.2013.06.019.

Rastkari N, khaniha RA. Magnetic solidphaseextraction based on magnetic multi-walled carbon nanotubes for the determination of phthalate monoesters in urine samples. J Chromatogr A. 2013; 1286(19):22-28. http//:doi:10.1016/j.chroma.2013.02.070.

Nirogi R, Kandikere V, Mudigonda K, Komarneni P, Aleti R. Liquid chromatography atmospheric pressure chemical ionization tandem mass spectrometry method for the quantification of pregabalin in human plasma. J Chromatogr B Analyt Technol Biomed Life Sci. 2009; 877(30):3899-3906. http//:doi:10.1016/j.jchromb.2009.10.004.

Martinc B, Grabnar I, Mrhar A, Vovk T. Rapid highperformance liquid chromatography method for determination of pregabalin in a pharmaceutical dosage form following derivatization with fluorescamine. J AOAC Int. 2010; 93(4):1069-76.

Dousa M, Gibala P, Lemr K. Liquid chromatographic separation of pregabalin and its possible impurities with fluorescence detection after postcolumn derivatization with o-phtaldialdehyde. J Pharm Biomed Anal. 2010; 53(3):717-722. http//:doi:10.1016/j.jpba.2010.04.008.

Souri E, Eskandari M, Tehrani MB, Adib N, Ahmadkhaniha R. HPLC Determination of Pregabalin in Bulk and Pharmaceutical Dosage Forms After Derivatization with 1-Fluoro-2,4-dinitrobenzene. Asian J. Chem. 2013; 25 (13), 7332-7336. https://doi.org/10.14233/ajchem.2013.14624.

Report from the Scientific Working Group for Forensic Toxicology (SWGTOX).Standard Practices for Method Validation in Forensic Toxicology. J Anal Toxicol. 2013; 37(7):452-74. doi:10.1093/jat/bkt054.

Indupriya M, Chandan RS, Gurupadayya BM, Sowjanya K. Quantitative determination of levetiracetam by gas chromatography using ethyl chloroformate as a derivatizing reagent in pure and pharmaceutical preparation. IJPT. 2011; 3(1):1694- 1701.

Berry D, Millington C. Analysis of pregabalin at therapeutic concentrations in human plasma/serum by reversed-phase HPLC. Therapeutic Drug Monitoring. 2005; 27(4) 451-456. http//:DOI:10.1097/01.ftd.0000158874.54100.1a.

Furniss BS, Hannaford AJ, Rogers V, Smith PWG, Tatchell AR. (eds.) Qualitative organic analysis; the preparation of derivatives. In Vogel's, Textbook of practical organic chemistry, 4th edition, chapter 7, Longman group Ltd, England; 1986;1088-1142.

Midik I, Tagliaro F, Tesarova E, Deyl Z. Advanced Chromatographic and Electro migration Methods in BioSciences (Deyl Z, Miksik I, Tesarova E. eds). Journal of Chromatography Library, 1st edition, volume 60, chapter 4, Elsevier Science. 1998.