In Silico Study, Physicochemical, and In Vitro Lipase Inhibitory Activity of α, β-Amyrenone Inclusion Complexes with Cyclodextrins

Abstract

α,β-amyrenone (ABAME) is a triterpene derivative with many biological activities; however, its potential pharmacological use is hindered by its low solubility in water. In this context, the present work aimed to develop inclusion complexes (ICs) of ABAME with γ- and β-cyclodextrins (CD), which were systematically characterized through molecular modeling studies as well as FTIR, XRD, DSC, TGA, and SEM analyses. In vitro analyses of lipase activity were performed to evaluate possible anti-obesity properties. Molecular modeling studies indicated that the CD:ABAME ICs prepared at a 2:1 molar ratio would be more stable to the complexation process than those prepared at a 1:1 molar ratio. The physicochemical characterization showed strong evidence that corroborates with the in silico results, and the formation of ICs with CD was capable of inducing changes in ABAME physicochemical properties. ICs was shown to be a stronger inhibitor of lipase activity than Orlistat and to potentiate the inhibitory effects of ABAME on porcine pancreatic enzymes. In conclusion, a new pharmaceutical preparation with potentially improved physicochemical characteristics and inhibitory activity toward lipases was developed in this study, which could prove to be a promising ingredient for future formulations.

Keywords: amyrenone; cyclodextrins; inclusion complexes; lipase activity; triterpenes.

Figure 1

Solubility profiles of α- (A) and β- (B) amyrenones. Lipo (Lipid affinity), Size (Molecular structure size), Polar (Polarity of the molecule), Insolu (Compound water insolubility), Unsatu (Compound unsaturations), Flex (Structural flexibility).

Figure 2

Similarity between the caulophyllogenin and α,β-amyrenone triterpenes.

Figure 3

Energetic simulation of the interaction between α,β-amyrenone and β-cyclodextrin (βCD) in the molar ratios of 1:1 (A,B) and 2:1 (C,D). (A) α-amyrenone: βCD, coupling with the face containing hydroxyl groups inside the βCD cavity, (B) β-amyrenone: βCD, coupling with the face containing the hydroxyl groups outside the βCD cavity, (C) α-amyrenone: βCD in complete coupling, (D) β-amyrenone: βCD, in complete coupling.

Figure 4

Energetic simulation of the interaction between α,β-amyrenone and β-cyclodextrin (γCD) in the molar ratios of 1:1 (A,B) and 2:1 (C,D). (A) α-amyrenone: γCD, coupling with the face containing hydroxyl groups inside the γCD cavity, (B) β-amyrenone: γCD, coupling with the face containing the hydroxyl groups outside the γCD cavity, (C) α-amyrenone: γCD in complete coupling, (D) β-amyrenone: γCD, in complete coupling.

Figure 5

X-ray diffraction profiles of individual compounds, namely α,β-amyrenone (ABAME), β-cyclodextrin (βCD) and γ-cyclodextrin (γCD), as well as inclusion complexes (ICs). ICs prepared with ABAME and βCD at 1:1 (A) and 2:1 (B) molar ratios using physical mixture (PMB1 and PMB2), kneading (KNDB1 and KNDB2), and rotary evaporation (EVB1 and EVB2), respectively. ICs prepared with ABAME and γCD at 1:1 (C) and 2:1 (D) molar ratios using physical mixture (PMG1 and PMG2), kneading (KNDG1 and KNDG2), and rotary evaporation (EVG1 and EVG2), respectively.

Figure 6

FTIR-ATR spectra of individual compounds, namely α,β-amyrenone (ABAME), β-cyclodextrin (βCD) and γ-cyclodextrin (γCD), as well as inclusion complexes (ICs). ICs prepared with ABAME and βCD at 1:1 (A) and 2:1 (B) molar ratios using physical mixture (PMB1 and PMB2), kneading (KNDB1 and KNDB2), and rotary evaporation (EVB1 and EVB2), respectively. ICs prepared with ABAME and γCD at 1:1 (C) and 2:1 (D) molar ratios using physical mixture (PMG1 and PMG2), kneading (KNDG1 and KNDG2), and rotary evaporation (EVG1 and EVG2), respectively.

Figure 7

SEM micrographs of individual compounds, namely α,β-amyrenone (ABAME), β-cyclodextrin (βCD) and γ-cyclodextrin (γCD), as well as inclusion complexes (ICs). ICs prepared with ABAME and βCD at 1:1 (A) and 2:1 (B) molar ratios using physical mixture (PMB1 and PMB2), kneading (KNDB1 and KNDB2), and rotary evaporation (EVB1 and EVB2), respectively. ICs prepared with ABAME and γCD at 1:1 (C) and 2:1 (D) molar ratios using physical mixture (PMG1 and PMG2), kneading (KNDG1 and KNDG2), and rotary evaporation (EVG1 and EVG2), respectively.

Figure 8

Differential Scanning Calorimetry curves of individual compounds, namely α,β-amyrenone (ABAME), β-cyclodextrin (βCD) and γ-cyclodextrin (γCD), as well as inclusion complexes (ICs). ICs prepared with ABAME and βCD at 1:1 (A) and 2:1 (B) molar ratios using physical mixture (PMB1 and PMB2), kneading (KNDB1 and KNDB2), and rotary evaporation (EVB1 and EVB2), respectively. ICs prepared with ABAME and γCD at 1:1 (C) and 2:1 (D) molar ratios using physical mixture (PMG1 and PMG2), kneading (KNDG1 and KNDG2), and rotary evaporation (EVG1 and EVG2), respectively.

Figure 9

Thermogravimetry curves of individual compounds, namely α,β-amyrenone (ABAME), β-cyclodextrin (βCD) and γ-cyclodextrin (γCD), as well as inclusion complexes (ICs). ICs prepared with ABAME and βCD at 1:1 (A) and 2:1 (B) molar ratios using physical mixture (PMB1 and PMB2), kneading (KNDB1 and KNDB2), and rotary evaporation (EVB1 and EVB2), respectively. ICs prepared with ABAME and γCD at 1:1 (C) and 2:1 (D) molar ratios using physical mixture (PMG1 and PMG2), kneading (KNDG1 and KNDG2), and rotary evaporation (EVG1 and EVG2), respectively.

Figure 10

Rate of inhibition of lipase activity (%) exerted by different samples at a concentration of 1 µg mL⁻¹. Data, worked out by the statistical software GraphPad Prism (version 6.0), are expressed as means ± SD. p < 0.05 was considered statistically significant when compared to the control group.