DEVELOPMENT AND VALIDATION OF A SPECIFIC AND UNIQUE DUAL POLARITY ESI - LC-MS/MS METHOD FOR SIMULTANEOUS DETERMINATION OF SEMAGLUTIDE AND DAPAGLIFLOZIN IN HUMAN PLASMA

R Baby Nalanda; JAGAPATHI RAJU VATSAVAYI

doi:10.22159/ijap.2024v16i5.51095

Authors

R Baby Nalanda Assistant Professor
Jagapathi Raju Vatsavayi GITAM SCHOOL OF PHARMACY

DOI:

https://doi.org/10.22159/ijap.2024v16i5.51095

Keywords:

SEMAGLUTIDE, DAPAGLIFLOZIN, ELECTROSPRAY IONIZATION, METHOD VALIDATION, MASS SPECTROMETER

Abstract

Objective: To develop a method capable of simultaneous quantification and estimation of Semaglutide and Dapagliflozin which are being studied as a prospective combination therapy for the treatment of Diabetes.

Materials: An intricate protein precipitation extraction technique was employed after addition of Verapamil and Tolbutamide as Internal standards for Semaglutide and Dapagliflozin respectively. The two compounds were separated on a Kinetex C18 (50 mm x 2.1 mm, 5 µ Particle size) column, while a dual polarity ionization ESI mass detection was carried out with a multiple reaction monitoring (MRM) method and a gradient program utilizing Acetonitrile and water as mobile phases to achieve a separation in 3 minutes.

Results: The method established was proved linear over a working range of 1.00 ng/mL to 1000 ng/mL and 2.00 ng/mL to 2000 ng/mL for Semaglutide (r²>0.98) and Dapagliflozin (r²>0.98) respectively in human plasma. The accuracy, recovery, matrix effects were within acceptable limits. The stability was also established under various conditions as necessitated by the the ICH M10 guideline on Bioanalytical method validation.

Conclusion: The highly sensitive method where 1.00 ng/mL for Semaglutide and 2.00 ng/mL for Dapagliflozin as the lower limit of quantification can be utilized for estimation in human plasma will facilitate the further application to pharmacokinetic and bioequivalence studies for these two drugs in pharmaceutical dosage forms or their new pharmaceutical formulation. This extraction technique and the optimized chromatographic conditions can be applied to pivotal clinical studies to achieve the best outcomes. This method allows fast accurate analysis procedure which can be leveraged effectively for clinical subject sample analysis.

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References

Marso SP, Bain SC, Consoli A, Eliaschewitz FG, Jódar E, Leiter LA, Lingvay I, Rosenstock J, Seufert J, Warren ML, Woo V, Hansen O, Holst AG, Pettersson J, Vilsbøll T; SUSTAIN-6 Investigators. Semaglutide and Cardiovascular Outcomes in Patients with Type 2 Diabetes. N Engl J Med. 2016; 375(19): 1834-1844.

Goldenberg RM, Steen O. Semaglutide: Review and Place in Therapy for Adults with Type 2 Diabetes. Can J Diabetes. 2019; 43(2): 136-145.

Li Y, Hansotia T, Yusta B, Ris F, Halban PA, Drucker D. Glucagon-like Peptide-1 Receptor Signaling Modulates Cell Apoptosis. J. Biol. Chem. 2003; 278 (1): 471–478.

Meier JJ. Efficacy of Semaglutide in a Subcutaneous and an Oral Formulation. Front Endocrinol (Lausanne). 2021; 12: 645617.

Lee TS, Park EJ, Choi M, Oh HS, An Y, Kim T, Kim TH, Shin BS, Shin S. Novel LC-MS/MS analysis of the GLP-1 analog semaglutide with its application to pharmacokinetics and brain distribution studies in rats. J Chromatogr B Analyt Technol Biomed Life Sci. 2023; Apr 15; 1221:123688.

Gumieniczek A & Berecka A. (2016). Analytical tools for determination of new oral antidiabetic drugs, glitazones, gliptins, gliflozins and glinides, in bulk materials, pharmaceuticals and biological samples. Open Chem. 2016; 14. 10.1515.

DeFronzo RA, Ferrannini E, Groop L, Henry RR, Herman WH, Holst JJ, Hu FB, Kahn CR, Raz I, Shulman GI, Simonson DC, Testa MA, Weiss R. Type 2 diabetes mellitus. Nature Reviews: Disease Primers. 2015; 1:15019.

Scheen AJ, Paquot N. Metformin revisited: a critical review of the benefit-risk balance in at-risk patients with type 2 diabetes. Diabetes Metab. 2013; 39(3): 179-90.

Aubry AF, Gu H, Magnier R, Morgan L, Xu X, Tirmenstein M, Wang B, Deng Y, Cai J, Couerbe P, Arnold M. Validated LC-MS/MS methods for the determination of dapagliflozin, a sodium-glucose co-transporter 2 inhibitor in normal and ZDF rat plasma. Bioanal. 2010; 2(12): 2001-9.

Ji QC, Xu X, Ma E, Liu J, Basdeo S, Liu G, Mylott W, Boulton DW, Shen JX, Stouffer B, et al. Selective reaction monitoring of negative electrospray ionization acetate adduct ions for the bioanalysis of dapagliflozin in clinical studies. Anal Chem. 2015; 87: 3247–3254.

Shah PA, Shrivastav PS, Shah JV, George A. Simultaneous quantitation of metformin and dapagliflozin in human plasma by LC–MS/MS: Application to a pharmacokinetic study. Biomed Chromatogr. 2019; 33: e4453.

Van der Aart-Van der Beek AB, Wessels AMA, Heerspink HJL, Touw DJ. Simple, fast and robust LC-MS/MS method for the simultaneous quantification of canagliflozin, dapagliflozin and empagliflozin in human plasma and urine. J Chromatogr B Analyt Technol Biomed Life Sci. 2020; 1152: 122257.

U.S. Food and Drug Administration, Bioanalytical Method Validation: Guidance for Industry, Silver Spring, MD, 2018.

ICH guideline M10 on Bioanalytical method validation and study sample analysis 2023.

Matuszewski BK, Constanzer ML & Chavez-Eng CM. Strategies for the assessment of matrix effect in quantitative bioanalytical methods based on HPLC-MS/MS. Anal Chem. 2003; 75(13), 3019–3030.

Jensen L, Helleberg H, Roffel A, van Lier JJ, Bjørnsdottir I, Pedersen PJ, Rowe E, Derving Karsbøl J, Pedersen ML. Absorption, metabolism and excretion of the GLP-1 analogue semaglutide in humans and nonclinical species. Eur J Pharm Sci. 2017; 104: 31-41.

Kapitza C, Nosek L, Jensen L, Hartvig H, Jensen CB, Flint A. Semaglutide, a once-weekly human GLP-1 analog, does not reduce the bioavailability of the combined oral contraceptive, ethinylestradiol/levonorgestrel. J Clin Pharmacol. 2015; 55(5): 497-504.

Marbury TC, Flint A, Jacobsen JB, Derving Karsbøl J, Lasseter K. Pharmacokinetics and Tolerability of a Single Dose of Semaglutide, a Human Glucagon-Like Peptide-1 Analog, in Subjects With and Without Renal Impairment. Clin Pharmacokinet. 2017; 56(11): 1381-1390.

Overgaard RV, Navarria A, Ingwersen SH, Bækdal TA, Kildemoes RJ. Clinical Pharmacokinetics of Oral Semaglutide: Analyses of Data from Clinical Pharmacology Trials. Clin Pharmacokinet. 2021; 60(10): 1335-1348.

Jensen L, Kupcova V, Arold G, Pettersson J, Hjerpsted JB. Pharmacokinetics and tolerability of semaglutide in people with hepatic impairment. Diabetes Obes Metab. 2018; 20(4): 998-1005.

Baekdal TA, Thomsen M, Kupčová V, Hansen CW, Anderson TW. Pharmacokinetics, Safety, and Tolerability of Oral Semaglutide in Subjects with Hepatic Impairment. J Clin Pharmacol. 2018; 58(10): 1314-1323.