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

JAGAPATHI RAJU VATSAVAYI; NALANDA BABY REVU

doi:10.22159/ijap.2024v16i5.51095

Authors

JAGAPATHI RAJU VATSAVAYI GITAM School of Pharmacy, GITAM (Deemed to be University), Visakhapatnam-530045, Andhra Pradesh (State), India
NALANDA BABY REVU GITAM School of Pharmacy, GITAM (Deemed to be University), Visakhapatnam-530045, Andhra Pradesh (State), India

DOI:

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

Keywords:

Semaglutide, Dapagliflozin, Electro spray 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 treating Diabetes.

Methods: An intricate protein precipitation extraction technique was employed using 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, with a dual polarity ionization Electro Spray Ionization (ESI) on a Liquid chromatograph Tandem Mass Spectrometry (LC-MS/MS) instrument. The 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 min.

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, and matrix effects were within acceptable limits. The stability was also established under various conditions as necessitated by the International Council for Harmonisation of Technical Requirements of Pharmaceuticals for Human Use (ICH) M10 guideline on Bioanalytical method validation.

Conclusion: This highly selective and sensitive method where 1.00 ng/ml for semaglutide and 2.00 ng/ml for dapagliflozin as the Lower Limit of Quantification (LLOQ) can be utilized for estimation in human plasma will facilitate the further application to pharmacokinetic and bioequivalence studies for combination of these two drugs in pharmaceutical dosage forms.

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