EFFECT OF DIFFERENT ANTICOAGULANTS IN DETERMINING METFORMIN HYDROCHLORIDE LEVELS IN HUMAN PLASMA USING HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

  • Yahdiana Harahap
  • Lista Roro Marsudi
  • Sunarsih .

Abstract

Objective: This study aimed to develop and validate an analytical method and to determine the effect of different anticoagulants for analyzing
metformin HCl level in human plasma.
Methods: Analysis was performed with high-performance liquid chromatography method using a photodiode array detector set at 234 nm, a C18
SunFire™ column (5 μm, 250 mm×4.6 mm), a temperature of 40°C, a mobile phase comprising 10 mM sodium dodecyl sulfate and 10 mM phosphate
buffer in water-acetonitrile (60:40 v/v) at pH 7.0, with a flow rate of 1.0 mL/min, and atorvastatin calcium as the internal standard. Plasma
extraction was performed through protein precipitation using human plasma (300 μL) and acetonitrile (600 μL; 1:2 v/v). The method was linear at a
concentration range of 20.0–5000.0 ng/mL, with r>0.9998.
Results: The stability and recovery of metformin HCl in human plasma were not different between citrate with ethylenediaminetetraacetic acid
(EDTA) and heparin with EDTA as anticoagulants. However, a significant difference in the peak area response ratio (p<0.05) was observed between
the three anticoagulants.
Conclusion: The validation results for the three types of plasma anticoagulants met validation requirements based on the European Medicines
Agency bioanalytical guidelines of 2011 for accuracy and precision, selectivity, calibration curve linearity, dilution integrity, carry-over, and stability.

Keywords: Anticoagulant, Citrate, Ethylenediaminetetraacetic acid, Heparin, Human plasma, Validation.

References

1. Dipiro J, Cecily V, Barbara GW, Terry LS. Pharmacotherapy:
A Pathophysiologic Approach. 9th ed. United States: The McGraw-Hill
Companies, Inc; 2015.
2. Goodman L, Gilman A, Brunton L, Lazo J, Parker K. Goodman and
Gilman’s the Pharmacological Basis of Therapeutics. 1st ed. New York:
McGraw-Hill; 2006.
3. Food and Drug Administration. Metformin Hydrochloride Tablets. United
States of America: Department of Health and Human Services; 2002.
4. Heinig K, Bucheli F. Fast liquid chromatographic-tandem mass
spectrometric (LC-MS-MS) determination of metformin in plasma
samples. J Pharm Biomed Anal 2004;34:1005-11.
5. Zarghi A, Foroutan S, Shafaati A, Khoddam A. Rapid determination
of metformin in human plasma using ion-pair HPLC. J Pharm Biomed
Anal 2003;31:197-200.
6. Shargel L, Wu-Pong S, Yu ABC. Applied Biopharmaceutics and
Pharmacokinetics. 5th ed. United States of America: Appeton and
Lange; 2004.
7. Harahap Y. The Role of Bioanalysis in Drug Quality Assurance and
Improving Quality of Patient Life. Jakarta: UI-Press; 2010.
8. Bowen RA, Hortin GL, Csako G, Otañez OH, Remaley AT. Impact of
blood collection devices on clinical chemistry assays. Clin Biochem
2010;43:4-25.
9. Gonzalez-Covarrubias V, Dane A, Hankemeier T, Vreeken RJ. The
influence of citrate, EDTA, and heparin anticoagulants to human
plasma LC-MS lipidomic profiling. Metabolomics 2012;9:337-48.
10. Ludong MM. Blood Transfusion. Jakarta: Faculty of Medicine,
Universitas Tarumanegara; 2012. p. 1-11.
11. Barri T, Dragsted LO. UPLC-ESI-QTOF/MS and multivariate
data analysis for blood plasma and serum metabolomics: Effect
of experimental artefacts and anticoagulant. Anal Chim Acta
2013;768:118-28.
12. European Medicines Agency. Guideline on Bioanalytical Method
Validation. Vol. 44. London: An Agency of the European Union; 2011.
13. AbuRuz S, Millership J, McElnay J. Determination of metformin in
plasma using a new ion pair solid phase extraction technique and ion
pair liquid chromatography. J Chromatogr B Analyt Technol Biomed
Life Sci 2003;798:203-9.
14. Haque A, Priya S, Mohanty D, Bakshi V, Boggula N. Analytical method
development and validation of amlodipine in human plasma using
liquid chromatography-mass spectrometry/mass spectrometry. Asian J
Pharm Clin Res 2017;11:393-7.
15. Evans G. A Handbook of Bioanalysis and Drug Metabolism. United
States of America: CRC Press; 2004.
16. Pawar AK. Development and validation of UV spectrophotometric
and reversed phase-high performance liquid chromatography-PDA
methods for the estimation of alogliptin benzoate. Asian J Pharm Clin
Res 2017;9:282-7.
17. Porwal P, Talele G. Development of validated HPLC-UV method for
simultaneous determination of metformin, amlodipine, glibenclamide
and atorvastatin in human plasma and application to protein binding
studies. Bull Fac Pharm 2016;55:129-139.
18. Porta V, Schramm S, Kano E, Koono E, Armando Y, Fukuda K, et al.
HPLC-UV determination of metformin in human plasma for application
in pharmacokinetics and bioequivalence studies. J Pharm Biomed Anal
2008;46:143-7.
19. Chhetri H, Thapa P, Van SA. Simple HPLC-UV method for the
quantification of metformin in human plasma with one step protein
precipitation. Saudi Pharm J 2013;22:483-7.
20. Alakhali KM. Validation method for measuring simvastatin in human
plasma by HPLC-UV and its application ii study simvastatin stability in
plasma and working solution. Asian J Pharm Clin Res 2017;7:131-3.
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How to Cite
Harahap, Y., Marsudi, L. R., & ., S. (2018). EFFECT OF DIFFERENT ANTICOAGULANTS IN DETERMINING METFORMIN HYDROCHLORIDE LEVELS IN HUMAN PLASMA USING HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY. International Journal of Applied Pharmaceutics, 10(1), 148-151. https://doi.org/10.22159/ijap.2018.v10s1.31
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