Eplerenone nanocrystals engineered by controlled crystallization for enhanced oral bioavailability

Abstract

Poor aqueous solubility of eplerenone (EPL) is a major obstacle to achieve sufficient bioavailability after oral administration. In this study, we aimed to develop and evaluate eplerenone nanocrystals (EPL-NCs) for solubility and dissolution enhancement. D-optimal combined mixture process using Design-Expert software was employed to generate different combinations for optimization. EPL-NCs were prepared by a bottom-up, controlled crystallization technique during freeze-drying. The optimized EPL-NCs were evaluated for their size, morphology, thermal behavior, crystalline structure, saturation solubility, dissolution profile, in vivo pharmacokinetics, and acute toxicity. The optimized EPL-NCs showed mean particle size of 46.8 nm. Scanning electron microscopy revealed the formation of elongated parallelepiped shaped NCs. DSC and PXRD analysis confirmed the crystalline structure and the absence of any polymorphic transition in EPL-NCs. Furthermore, EPL-NCs demonstrated a 17-fold prompt increase in the saturation solubility of EPL (8.96 vs. 155.85 µg/mL). The dissolution rate was also significantly higher as indicated by ∼95% dissolution from EPL-NCs in 10 min compared to only 29% from EPL powder. EPL-NCs improved the oral bioavailability as indicated by higher AUC, C_max, and lower T_max than EPL powder. Acute oral toxicity study showed that EPL-NCs do not pose any toxicity concern to the blood and vital organs. Consequently, NCs prepared by controlled crystallization technique present a promising strategy to improve solubility profile, dissolution velocity and bioavailability of poorly water-soluble drugs.

Keywords: Eplerenone; acute toxicity study; controlled crystallization; dissolution rate and bioavailability; nanocrystals; poorly aqueous solubility.

Figure 1.

2D contour plots presenting the effects of independent variables on size (A, B) and dissolution efficiency (C, D) of EPL-NCs.

Figure 2.

Particle size distribution curve (A) and scanning electron microscopic image (B) of the optimized EPL-NCs.

Figure 3.

Chemical structure of EPL (A) and FTIR spectra (B) of EPL (a), mannitol (b), physical mixture (c), and EPL-NCs (d).

Figure 4.

DSC thermograms (A) and PXRD patterns (B) of EPL (a), mannitol (b), physical mixture (c), and EPL-NCs (d).

Figure 5.

Saturation solubility (A) and dissolution profile (B) of the optimized EPL-NCs. Data are presented as mean ± S.D. (n = 3). *p<.001 vs. EPL powder.

Figure 6.

Average plasma level vs. time curve after oral administration of EPL-NCs and EPL powder to rats at a dose equivalent to 10 mg/kg of EPL. Data are expressed as mean ± S.D. (n = 3).

Figure 7.

Organ-to-body weight index (A) and histological microphotographs of liver, kidney and heart tissues of mice (B) at day 14 after treatment with EPL-NCs. In histological micrographs, (1) hepatocytes, (2) hepatic artery, (3) sinusoids, (4) central vein, (5) bile duct, (6) bowman capsule, (7) glomerulus, (8) renal tubule, (9) proximal tubule, (10) distal tubule, and (11) myocardial fibers. Organ-to-body weight indices are presented as mean ± S.D. (n = 6).

Figure 8.

Blood hemolysis by the optimized EPL-NCs indicated with real-time photographs of RBCs samples (A) and quantified with spectroscopic analysis of hemoglobin at 541 nm (B). The RBCs were mixed with Triton X-100 solution (positive control), isotonic PBS (negative control), 20, 40, 60, 80, and 100 µg/mL of EPL (D-1 to D-5) and 20, 40, 60, 80, and 100 µg/mL of EPL-NCs (F-1 to F-5), respectively. For the quantification of hemolysis (%), data are presented as mean ± S.D. (n = 3).

Figure 9.

DSC thermograms (A) and PXRD patterns (B) of EPL-NCs at day 0 (a) and after 90 days (b) of storage stability period.