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. 2017 Jun 6;12:4269-4283.
doi: 10.2147/IJN.S124264. eCollection 2017.

Preparation, Characterization, and Evaluation of Amphotericin B-loaded MPEG-PCL-g-PEI Micelles for Local Treatment of Oral Candida albicans

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Free PMC article

Preparation, Characterization, and Evaluation of Amphotericin B-loaded MPEG-PCL-g-PEI Micelles for Local Treatment of Oral Candida albicans

Li Zhou et al. Int J Nanomedicine. .
Free PMC article

Abstract

Fatal Candida albicans infections in the mucosal system can occur in association with immune-compromised diseases and dysbacteriosis. Currently, amphotericin B (AmB) is considered to be the most effective antibiotic in the treatment of C. albicans infections, but its clinical application is limited by side effects and poor bioavailability. In order to use AmB in the local treatment of oral C. albicans infections, AmB/MPEG-PCL-g-PEI (monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine, MPP) micelles were prepared. A series of characterizations were performed. The micelles allowed a sustained in vitro release in both normal oral conditions (pH 6.8) and C. albicans infection conditions (pH 5.8). Then, buccal tablets containing freeze-dried powder of AmB/MPP micelles were produced by direct compression process and evaluated as regards to weight variation, hardness, and friability. In vitro drug release of the buccal tablets was measured in both the United States Pharmacopeia dissolution apparatus and the dissolution rate test apparatus, which was previously designed for simulating in vivo conditions of the oral cavity. The buccal tablets could sustainably release within 8 h and meet the antifungal requirements. Regarding safety assessment of AmB/MPP micelles, in vivo histopathological data showed no irritation toward buccal mucosa of the rats in both optical microscopy and ultrastructure observation of the tissues. MTT experiment proved that AmB/MPP micelles reduced the cytotoxicity of AmB. The micelles delivered through the gastrointestinal route were also found to be non-systemic toxicity by liquid chromatography-mass spectrometry analysis. Furthermore, the antifungal action of AmB/MPP micelles was evaluated. Although AmB/MPP had no obvious improvement as compared to AmB alone in the antifungal effect on planktonic C. albicans, the micelles significantly enhanced the antifungal activity against the biofilm state of C. albicans. Thus, it was concluded that AmB/MPP micelles represent a promising novel drug delivery system for the local treatment of oral C. albicans infections.

Keywords: Candida albicans; amphotericin B; antifungal effect; buccal tablet; micelle; safety evaluation.

Conflict of interest statement

Disclosure The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
(A) Spontaneous AmB-loaded micelles formation from amphiphilic molecules in aqueous media; (B) the chemical formula and the 1H-NMR spectra (400 MHz, CDCl3) of MPEG-PCL-g-PEI; (C) buccal tablets containing AmB+DOC (T1), AmB+MPEG-PCL-g-PEI physical mixtures (T2), and AmB/MPEG-PCL-g-PEI micelles (T3). Abbreviations: AmB, amphotericin B; MPEG-PCL-g-PEI, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine; DOC, sodium deoxycholate.
Figure 2
Figure 2
(A) The particle size distribution spectrum of blank micelles and AmB-loaded micelles; (B) zeta potential of blank micelles and AmB-loaded micelles; (C) morphology of AmB-loaded micelles determined by TEM (scale bar 100 nm); (D) DSC analysis of (a) MPP, (b) AmB, (c) AmB/MPP micelles, and (d) AmB and MPP physical mixtures. Abbreviations: AmB, amphotericin B; TEM, transmission electron microscopy; DSC, differential scanning calorimetry; MPP, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine.
Figure 3
Figure 3
Release profile of AmB-loaded micelles and redissolved micelles in different mediums. Abbreviations: AmB, amphotericin B; SDS, sodium dodecyl sulfate.
Figure 4
Figure 4
The release profiles of AmB from buccal tablets using different dissolution apparatuses. Notes: (A) USP dissolution apparatus (in 0.5% SDS solution) and (B) the dissolution rate test apparatus (in pH 5.8 and pH 6.8 medium). T1, AmB+DOC; T2, AmB+MPP; T3, AmB/MPP micelles. Abbreviations: AmB, amphotericin B; SDS, sodium dodecyl sulfate; DOC, sodium deoxycholate; MPP, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine; USP, United States Pharmacopeia.
Figure 5
Figure 5
Cytotoxicity of AmB, Fungizone (AmB+DOC), AmB/MPP micelles, and MPP blank micelles incubation for (A) 24 h and (B) 12 h. Abbreviations: AmB, amphotericin B; DOC, sodium deoxycholate; MPP, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine.
Figure 6
Figure 6
Histopathological observation of oral buccal tissues through injection with 5% dextrose solution, Fungizone (AmB+DOC), AmB/MPP micelles, and MPP blank micelles under a light microscope (400×) after dosing for 5, 15, and 30 days. Abbreviations: AmB, amphotericin B; DOC, sodium deoxycholate; MPP, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine; d, days.
Figure 7
Figure 7
Ultrastructure observation of oral buccal tissues through injection with 5% dextrose solution, Fungizone (AmB+DOC), AmB/MPP micelles, and MPP blank micelles after dosing for 30 days under transmission electron microscope. Notes: (A) Epithelial surface layer of the oral buccal tissues (10,000×); (B) epithelial middle layer containing prickle cells (10,000×); (C) basement membrane (30,000×, orange arrows pointed to the disappeared parts in the basement membrane of the AMB+DOC group); (D) desmosome junctions between epithelial cells (30,000×, orange arrows pointed to the residual desmosome junctions in AmB+DOC group). Abbreviations: AmB, amphotericin B; DOC, sodium deoxycholate; MPP, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine.
Figure 8
Figure 8
(A) MIC values of AmB/MPP and AmB; (B) observation of Candida albicans mature biofilms under SEM; (C) C. albicans BBU, concentrations of drugs were μg/mL. Notes: *Statistical difference at P,0.05. All data represent the mean ± SD (n=6). Abbreviations: MIC, minimum inhibitory concentration; AmB, amphotericin B; MPP, monomethoxy poly(ethylene glycol)-poly(epsilon-caprolactone)-graft-polyethylenimine; SEM, scanning electron microscope; BBU, biofilm biomass unit; SD, standard deviation; C.A., Candida albicans.

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