Degradation Pathway for Eplerenone by Validated Stability Indicating UP-LC Method

Abstract

Degradation pathway for eplerenone is established as per ICH recommendations by validated and stability-indicating reverse phase liquid chromatographic method. Eplerenone is subjected to stress conditions of acid, base, oxidation, and thermal and photolysis. Significant degradation is observed in acid and base stress conditions. Four impurities are studied and the major degradant (RRT about 0.31) was identified by LC-MS and spectral analysis. The stress samples are assayed against a qualified reference standard and the mass balance is found close to 99.5%. Efficient chromatographic separation is achieved on a Waters symmetry C18 stationary phase with simple mobile phase combination delivered in gradient mode and quantification is carried at 240 nm at a flow rate of 1.0 mL min(-1). In the developed LC method the resolution between eplerenone and four potential impurities (imp-1, imp-2, imp-3, and imp-4) is found to be greater than 4.0. Regression analysis shows an r value (correlation coefficient) of greater than 0.999 for eplerenone and four potential impurities. This method is capable to detect the impurities of eplerenone at a level of 0.020% with respect to test concentration of 1.0 mg mL(-1) for a 20 μL injection volume. The developed UPLC method is validated with respect to specificity, linearity and range, accuracy, precision, and robustness for impurities and assay determination.

Figure 1

Chemical structures and names of eplerenone and its impurities. (a) Eplerenone: pregn-4-ene-7, 21-dicarboxylic acid, 9, 11-epoxy-17-hydroxy-3-oxo, γ-lactone, methyl ester (7α, 11α, 17α) (molecular weight 414.49), (b) impurity -1: (11a,17a)-11,17-dihydroxy-3-oxo-pregna-4,6-diene-21-carboxylic acid (molecular weight 356.46), (c) impurity-2 : 5′R(5′alpha),7′beta-20′-amino hexadecahydro-11′beta-hydroxy-10′a,13′alpha-dimethyl-3′,5′-dioxospiro[furan-2(3H),17′alpha(5′H)-[7,4]methano[4H[cyclopenta[a]phenathrene]-5′-carbonitrile (molecular weight 410), (d) impurity-3 : 4′S(4′alpha),7′alpha-hexadecahydro-11′alpha-hydroxy-10′beta,13′beta-dimethyl-3′,5,20′-trioxospiro [furan-2(3H),17′beta-[4,7]methanol[17H]cyclopenta[a]phenanthrene]-5′beta(2′H) carbonitrile (molecular weight 411), (e) impurity-4: Methyl hydrogen 11 alpha, 17alpha-dihydroxy-3-oxopregn-4-ene-7 alpha, 21- dicarboxylate, gamma-lactone (molecular weight 416), and (f) 0.31& 0.86 RRT degradation impurity: methyl hydrogen 9,11 dihydroxy,17-α-hydroxy-3-oxopregn 7-α-carbonate, 21-α-carboxylic acid. (molecular weight 451).

Figure 2

Typical chromatogram from the method development trials optimized conditions and stressed eplerenone samples.

Figure 3

LC-mass chromatograms.

Figure 4

Linearity chart for eplerenone impurities.