Formulation and characterization of propolis and tea tree oil nanoemulsion loaded with clindamycin hydrochloride for wound healing: In-vitro and in-vivo wound healing assessment

Abstract

This study aimed to develop propolis and tea tree oil nanoemulsion loaded with clindamycin hydrochloride to heal wound effectively. Nanoemulsion formulae were prepared and characterized by droplet size analysis, zeta potential, viscosity, ex-vivo permeation, and skin deposition. The optimal formula was evaluated in terms of morphology, cytotoxicity, and in-vitro wound healing assay. Also, the efficacy of the optimal formula was evaluated by in-vivo wound healing and histopathological studies. The optimal formula (F3) was composed of 9% tea tree oil and 0.4% propolis extracts with mean droplet size 19.42 ± 1.7 nm, zeta potential value -24.5 ± 0.2 mV, and viscosity 69.4 ± 1.8 mP. Furthermore, the optimal formula showed the highest skin deposition value 550.00 ± 4.9 µg/cm² compared to other formulae. The TEM micrograph of the optimal formula showed that the nanoemulsion droplet has an almost spherical shape. Also, the optimal formula did not show noticeable toxicity to the human skin fibroblast cells. The in-vitro and in-vivo wound healing assay showed unexpected results that the un-loaded drug nanoemulsion formula had a comparable wound healing efficacy to the drug-loaded nanoemulsion formula. These results were confirmed with histopathological studies. Our results showed that the propolis and tea tree oil nanoemulsion, whether loaded or unloaded with an antibiotic, is an efficient local therapy for wound healing.

Keywords: Clindamycin hydrochloride; Nanoemulsion; Propolis; Tea tree oil.

Fig. 1

The images of scratch wound assay of different formulae at different time intervals.

Fig. 2

Ex-vivo permeation profile of different formulae.

Fig. 3

Transmission electron microscope (TEM) image of the optimal NE formula (F3).

Fig. 4

The percentage of viability was determined by SRB assay after treating human skin fibroblast (HSF) cells with F3.

Fig. 5

In-vitro wound healing activity of different formulae presented as a percentage of wound closure at various time intervals.

Fig. 6

Percentage of wound closure of different formulae (In-vivo wound healing assay).

Fig. 7

Histopathological examination of tissue samples from the skin wound collected on day 5 from the different groups (H&E).

Fig. 8

Charts present histological scores among different groups on day 5. Data Expressed as means ± standard error. A significant difference is considered at p < 0.05.

Fig. 9

Histopathological examination of tissue samples from the skin wound collected on day 14 from the different groups (H&E).

Fig. 10

Charts present histological scores among different groups on day 14. Data Expressed as means ± standard error. A significant difference is considered at p < 0.05.

Fig. 11

Masson's trichrome staining of skin sections derived from different groups on days 5 and 14 post-injury. Charts present area % of bluish stained sections in different groups on days 5 and 14. Data Expressed as means ± standard error. A significant difference is considered at p < 0.05.