Method development for the determination of digoxin using fluorescence quenching

PERALTA CECILIA MARIANA; VICARIO ANA; FERNÁNDEZ LILIANA PATRICIA; DI CHIACCHIO GABRIELA; GÓMEZ ROXANA; SAIDMAN ELBIO; ACOSTA MARÍA GIMENA

Rosario

Congreso; VII Reunión Internacional de Ciencias Farmacéuticas; 2023

Digoxin (DG) is a cardiac glycoside steroid that is commonly used to cure congestive heart failure and arrhythmias. Several studies support that the effective range of plasma concentration is between 0.5 and 2.0 ng/mL, being it toxic at higher plasma concentration (>2.0 ng/mL). It is also well stablished that the effective plasma drug concentration is close to the toxic, and large individual differences in the effects of the drug have been observed [1,2]. The official methodology for the quality control of solid oral forms uses derivatization using hydrogen peroxide/methanol and fluorescence detection.[3] The main objective of this study was the design of new derivatization strategies for automatization of analytical methods that allow the DG determination by molecular fluorescence. A further aim of this work was to validate methods to better the analytical performance currently achieved with fluorescence. In this way, reaching detection limits compatible with the DG concentrations in biological and pharmaceutical samples was a goal to achieve. In this research, a standard solution of DG (Sigma Chemical Co.) was taken up in ethanol (1 mg mL-1). The reagents used (rodhamine B (RhB), eosyne, ethanol, mili Q water) were analytical grade. Bovine seroalbumine (BSA) was acquired of Lab. Alimentos-UNSL, San Luis, Argentina. All experiments were carried out in a 1 cm quartz cell, using a Shimadzu RF-5301PC spectrofluorimeter, equipped with a Xenon discharge lamp. Several factors that affect the derivatization reaction were studied i.g., type and concentration of fluorophore (BSA, RhB, eosyne, quinoleine), pH, ionic strength, organic solvents. Also, the effect of different natural and synthetic surfactants was evaluated as improvement fluorescence signal (SDS, Bile salt, HTAB), etc. The spectral behavior observed after the derivatization were evaluated to establish the experimental conditions that allowed reaching an adequate sensitivity (excitation and emission wavelength: 280 nm and 340 nm (BSA) and 555 nm and 575 nm (RhB), slits: 5/5). The attenuation in the fluorescent signal (quenching effect) with increase DG concentration on both fluorophores (BSA and RhB) was observed. Adequate dynamic calibration range was obtained over a concentration interval of 0.2 – 20.0 ng mL-1. The limits of detection (LOD) and quantification (LOQ) were 0.1 and 0.2 ng mL-1, respectively. The studied strategies will be incorporated to schemes of high-performance liquid chromatography associated with fluorescent detection to automate the methodology and allow the determination of DG in different samples.References[1] S.H. Yao, H.-T. Tsai, W.L. Lin, Y.C. Chen, C. Chou, H.W. Lin, Predicting the serum digoxin concentrations of infants in the neonatal intensive care unit through an artificial neural network, BMC Pediatr. 19 (2019) 1-11.[2] D.J. Joshi, N.I. Malek, S.K. Kailasa, Fluorescence OFF-ON-OFF mechanism for the detection of digoxin, La3+, and epinephrine using bovine serum albumin functionalized molybdenum oxide quantum dots, Materials Today Chemistry 27 (2023) 101291.[3] Farmacopea Argentina Séptima edición, Vol. III, DIGOXINA COMPRIMIDOS.