(M1030-06-40) Development and Validation of UPLC Method for Digoxin and Degradants and It's Application to Assess the Effect of Process Variables on Quality Attributes of Digoxin Formulations

Purpose: Digoxin (DGX), one of the oldest cardiac therapies, was first approved by the FDA in 1954 under the brand name LANOXIN^® (NDA 020405). As a Biopharmaceutical Classification System Class IV drug characterized by low solubility, low permeability, and a narrow therapeutic index, Digoxin requires stringent formulation control to ensure consistent performance. The manufacturing processes and critical process variables significantly influence key attributes such as stability, dissolution, bioavailability, and ultimately, clinical efficacy. Current United States Pharmacopeia (USP) monograph methods for Digoxin utilize fluorescence detection, which, while sensitive, may not adequately distinguish the parent drug from its known degradation products. Although some analytical methods have been explored, there remains limited development of UPLC-based methods employing UV detection for the simultaneous quantification of Digoxin and its degradants. Therefore, the objective of this study was twofold: (1) to systematically investigate the impact of process parameters including particle size, binder type (lactose anhydrous versus lactose monohydrate), and granulating fluid (water versus ethanol) on the critical quality attributes of Digoxin immediate-release formulations, and (2) to develop and validate a novel stability-indicating UPLC-UV method capable of comprehensively assessing Digoxin and its degradation behavior. Eight formulations, produced by wet granulation, were developed and evaluated under accelerated stability conditions (40°C/75% RH) over a 10-day period.

Methods: Eight formulations were prepared using wet granulation, varying particle size (32 μm, 75 μm), lactose type (anhydrous, monohydrate), and granulating fluid (water, ethanol). A full factorial design assessed the impact of these variables on hardness, disintegration time, dissolution, assay, impurities, and crystallinity. Tablets were evaluated initially and after accelerated stability testing at 40°C/75% RH for 10 days. Assays, dissolution, and impurity profiling were performed using UPLC, while crystallinity changes were assessed using X-ray powder diffraction (XRPD). A stability-indicating UPLC method was developed to simultaneously quantify Digoxin and its known degradation products (digoxigenin, digoxigenin monodigitoxoside, and digoxigenin bisdigitoxoside). Chromatographic separation was achieved within 25 minutes using a Luna^® C8 column and UV detection at 220 nm. The method was validated over three days according to ICH Q2(R1) guidelines, demonstrating accuracy, specificity (baseline resolution between Digoxin and its degradants), linearity (r² > 0.999 across the tested range), precision (RSD < 2% for repeatability and intermediate precision), as well as appropriate limits of detection (LOD) and quantitation (LOQ) for all analytes. The validated method confirmed its suitability for stability monitoring of Digoxin formulations.

Results: Tablet hardness ranged from 3.0 to 6.75 KP initially, significantly influenced by particle size, lactose type, and granulating fluid. Disintegration time varied widely from 1.0 to 10.22 minutes. Formulation F7 (32 μm particle size, anhydrous lactose, ethanol) exhibited the best balance, with high initial dissolution (100% at 60 minutes) and low disintegration time (1.23 minutes). Upon stability testing, F5 and F8 formulations exhibited significant deterioration, with dissolution decreasing to 28.1% and 27.3%, respectively. These dissolution changes correlate with tablet hardness and disintegration time. Assay values remained within USP acceptance limits (95–105%) across all formulations initially. Impurity levels remained within acceptable ranges, with no new degradation products detected. Stability analysis showed that formulations with increased hardness after storage (F4 and F8) exhibited delayed disintegration and reduced dissolution, highlighting the critical impact of moisture and excipient interactions during storage. The stability-indicating UPLC-UV method successfully resolved Digoxin and its degradation products, demonstrating high specificity, sensitivity, and robustness for stability monitoring. The method's sensitivity also allowed the detection of variations in drug release profiles during dissolution testing, supporting the comprehensive evaluation of formulation performance. These findings collectively confirmed the chemical stability of Digoxin formulations under the tested conditions and validated the robustness of the analytical method for monitoring the impact of process variables and storage on product quality.

Conclusion: This study demonstrates that Digoxin tablet quality is highly dependent on formulation and process variables, with particle size, lactose type, and granulating fluid playing critical roles. Smaller particle size, the use of anhydrous lactose, and ethanol as a granulating fluid synergistically enhanced dissolution performance and stability by reducing crystallinity and promoting matrix porosity. Formulation F7 was identified as the optimal composition, maintaining robust dissolution and physical stability even under accelerated storage conditions. Additionally, a stability-indicating UPLC-UV method was successfully developed and validated, enabling simultaneous quantification of Digoxin and its major degradation products. The method demonstrated sufficient sensitivity to detect variations in drug release across formulations, supporting comprehensive evaluation of formulation stability. Together, these findings emphasize the importance of integrated formulation optimization and analytical method validation to ensure therapeutic efficacy and consistent quality of Digoxin immediate-release formulations.

References: 1. United States Pharmacopeia (USP-NF), Digoxin Tablets Monograph.
2. Naseem A. Charoo et al., " Quality by design approach for formulation development: A case study of dispersible tablets," International Journal of Pharmaceutics, 2011.
3. M. Draguet-Brughmans et al., " The physicochemical properties of digoxin," International Journal of Pharmaceutics, 2011.


Dissolution_Initial and Stability



Disintegration and Hardness