Bee chitosan nanoparticles loaded with apitoxin as a novel approach to eradication of common human bacterial, fungal pathogens and treating cancer

Abstract

Antimicrobial resistance is one of the largest medical challenges because of the rising frequency of opportunistic human microbial infections across the globe. This study aimed to extract chitosan from the exoskeletons of dead bees and load it with bee venom (commercially available as Apitoxin [Api]). Then, the ionotropic gelation method would be used to form nanoparticles that could be a novel drug-delivery system that might eradicate eight common human pathogens (i.e., two fungal and six bacteria strains). It might also be used to treat the human colon cancer cell line (Caco2 ATCC ATP-37) and human liver cancer cell line (HepG2ATCC HB-8065) cancer cell lines. The x-ray diffraction (XRD), Fourier transform infrared (FTIR), and dynamic light scattering (DLS) properties, ζ-potentials, and surface appearances of the nanoparticles were evaluated by transmission electron microscopy (TEM). FTIR and XRD validated that the Api was successfully encapsulated in the chitosan nanoparticles (ChB NPs). According to the TEM, the ChB NPs and the ChB NPs loaded with Apitoxin (Api@ChB NPs) had a spherical shape and uniform size distribution, with non-aggregation, for an average size of approximately 182 and 274 ± 3.8 nm, respectively, and their Zeta potential values were 37.8 ± 1.2 mV and - 10.9 mV, respectively. The Api@ChB NPs had the greatest inhibitory effect against all tested strains compared with the ChB NPs and Api alone. The minimum inhibitory concentrations (MICs) of the Api, ChB NPs, and Api@ChB NPs were evaluated against the offer mentioned colony forming units (CFU/mL), and their lowest MIC values were 30, 25, and 12.5 μg mL^-1, respectively, against Enterococcus faecalis. Identifiable morphological features of apoptosis were observed by 3 T3 Phototox software after Api@ChB NPs had been used to treat the normal Vero ATCC CCL-81, Caco2 ATCC ATP-37, and HepG2 ATCC HB-8065 cancer cell lines for 24 h. The morphological changes were clear in a concentration-dependent manner, and the ability of the cells was 250 to 500 μg mL^-1. These results revealed that Api@ChB NPs may be a promising natural nanotreatment for common human pathogens.

Keywords: Apis mellifera; antimicrobial resistance; apitoxin; bee’s chitosan NPs; colon cancer; drug delivery system.

Figure 1

XRD patterns of extracted bees Apis mellifera (A) Chitin; and (B) honey bee chitosan (ChB), (C) Api; (D) ChB NPs; and (E) Api@ChB NPs; (F) FTIR spectra of extracted bees Apis mellifera pure chitosan and (G) FTIR spectra of Api, ChB NPs, and Api@ChB NPs.

Figure 2

Hydrodynamic size, polydispersity index (PDI), and ζ-potential (A,B) ChB NPs; and (C,D) Api@ChB NPs. The numbers are given as mean standard deviation (SD) for ζ-potential (n = 3) and for particle size and PDI (n = 4). Average particle size and data size distribution of prepared samples (E,F) ChB NPs and (G,H) Api@ChB NPs measured by TEM (scale bar = 100 nm).

Figure 3

The inhibition zone (A) different pathogenic bacteria strains E. faecalis, S. aureus, S. hominis, E. coli, K. pneumonia, and A. baumannii. (B) C. auris and A. niger against by Api, ChB NPs, and Api@ChB NPs. Values in each column followed by different letters are significantly different ^a,b, in Duncan test (P < 0.05); statically differences between positive control and tested samples using one-way ANOVA.

Figure 4

Cytotoxicity against Api, ChB NPs, and Api@ChB (A) normal Vero ATCC CCL-81 cells, (B) Caco2 ATCC ATP-37, and (C) HepG2 ATCC HB-8065 cancer cells. The numbers are given as mean standard deviation (SD) (n = 3).