Enhanced antifungal activity of voriconazole-loaded nanostructured lipid carriers against Candida albicans with a dimorphic switching model

Abstract

Candida commonly adheres to implanted medical devices and forms biofilms. Due to the minimal activity of current antifungals against biofilms, new drugs or drug-delivery systems to treat these persistent infections are urgently needed. In the present investigation, voriconazole-loaded nanostructured lipid carriers (Vrc-NLCs) were formulated for enhanced drug-delivery efficiency to C. albicans to increase the antifungal activity of Vrc and to improve the treatment of infectious Candida diseases. Vrc-NLCs were prepared by a hot-melt, high-pressure homogenization method, and size distribution, ζ-potential, morphology, drug-encapsulation efficiency, drug loading, and physical stability were characterized. The antifungal activity of Vrc-NLCs in vitro was tested during planktonic and biofilm growth in C. albicans. The mean particle size of the Vrc-NLCs was 45.62±0.53 nm, and they exhibited spheroid-like morphology, smooth surfaces, and ζ-potential of -0.69±0.03 mV. Encapsulation efficiency and drug loading of Vrc-NLCs were 75.37%±2.65% and 3.77%±0.13%, respectively. Physical stability results revealed that despite the low measured ζ-potential, the dispersion of the Vrc-NLCs was stable during their 3-week storage at 4°C. The minimum inhibitory concentration of Vrc-NLCs was identical to that of Vrc. However, the inhibition rate of Vrc-NLCs at lower concentrations was significantly higher than that of Vrc during planktonic growth in C. albicans in yeast-extract peptone dextrose medium. Surprisingly, Vrc-NLCs treatment reduced cell density in biofilm growth in C. albicans and induced more switches form hyphal cells to yeast cells compared with Vrc treatment. In conclusion, Vrc-NLCs maintain antifungal activity of Vrc and increase antifungal drug-delivery efficiency to C. albicans. Therefore, Vrc-NLCs will greatly contribute to the treatment of infectious diseases caused by C. albicans.

Keywords: Candida albicans; antifungal activity; biofilms; nanostructured lipid carriers; voriconazole.

Figure 1

Characterization of NLCs. Notes: (A) Particle-size distribution spectrum of Vrc-NLCs; (B) ζ-potential of Vrc-NLCs; (C) morphology of Vrc-NLCs determined by TEM. Abbreviations: NLCs, nanostructured lipid carriers; Vrc, voriconazole; TEM, transmission electron microscopy.

Figure 2

Inhibition rates of Vrc-NLCs against Candida albicans in YPD. Notes: **P<0.01; *P<0.05. Cells were incubated with different concentrations of Vrc-NLCs and Vrc for 24 hours. Values given in reference to control group, which was cultured without Vrc-NLCs or Vrc. Antifungal activity of Vrc-free NLCs against C. albicans, was also tested, showing no activity at the evaluated concentrations. It was considered to have 100% viability in the control group. Vertical bars represent mean ± SD obtained from three experiments for each concentration. Abbreviations: Vrc, voriconazole; NLCs, nanostructured lipid carriers; YPD, yeast-extract peptone dextrose.

Figure 3

Confocal laser-scanning microscopy showing inhibition of Candida albicans biofilm and dimorphic switching by Vrc-NLCs and Vrc. Notes: (A) C. albicans biofilm treated with 0.05 μg/mL Vrc; (B) C. albicans biofilm treated with 0.05 μg/mL Vrc-NLCs; (C) C. albicans biofilm treated with 4 μg/mL Vrc; (D) C. albicans biofilm treated with 4 μg/mL Vrc-NLCs; (E) C. albicans biofilm treated with 8 μg/mL Vrc; (F) C. albicans biofilm treated with 8 μg/mL Vrc-NLCs; (G) C. albicans biofilm treated with 32 μg/mL Vrc; (H) C. albicans biofilm treated with 32 μg/mL Vrc-NLCs; (I) C. albicans biofilm treated with 1% dimethyl sulfoxide; (J) C. albicans biofilm treated with Vrc-free NLCs. Bar 25 μm. Magnification 600×. Abbreviations: Vrc, voriconazole; NLCs, nanostructured lipid carriers.