Green synthesis, formulation and biological evaluation of a novel ZnO nanocarrier loaded with paclitaxel as drug delivery system on MCF-7 cell line

https://doi.org/10.1016/j.colsurfb.2019.110686Get rights and content

Highlights

  • Green synthesis of zinc oxide nanoparticles (ZnO NPs) were prepared by ethanolic extract of Camellia sinensis L.

  • Paclitaxel (PTX) as an important anticancer drug was successfully loaded on the chitosan-coated ZnO nanoparticles (ZnO-Ch NPs).

  • Formulation and biological evaluation of a novel ZnO-nanocarrier loaded with paclitaxel as drug delivery system on MCF-7 cell line successfully was done.

  • The ZnO-Ch NPs could be used as a promising drug delivery platform for PTX with low side effect on normal cells but high cytotoxic effect on cancer cell line.

Abstract

In this study we design green synthesis of a novel ZnO nanocarrier loaded with paclitaxel as a drug delivery system with high cytotoxicity against breast cancer cell line (MCF-7) and low side effects on the normal cell line (fibroblast). Paclitaxel is formulated in high concentration in Cremophor EL because of its low solubility. Zinc oxide nanoparticles (ZnO NPs) were prepared by the ethanolic extract of Camellia sinensis L., then coated with chitosan (Ch) and loaded with paclitaxel (PTX) to improve drug delivery. The physicochemical properties were observed by transmission electron microscopy (TEM), thermo-gravimetric analysis (TGA), field emission scanning electron microscopy (FESEM), and Fourier transform infrared spectroscopy (FTIR). Drug loading on ZnO-Ch NPs was measured by high performance liquid chromatography (HPLC). In vitro apoptosis assay was assessed by flow cytometry. The cytotoxic effect of the nanocarrier drug was investigated using MTT assay in cancerous and normal cell lines. The PTX-loaded ZnO-Ch NPs showed cytotoxic effects on MCF-7 cells, with minimal detrimental effects on normal fibroblasts. The results of apoptosis assay were compliant with MTT findings. Generally, ZnO-Ch NPs could be used as a promising drug delivery platform for PTX with low side effect on normal cell line and high cytotoxic effect on breast cancer cell line.

Introduction

Drug delivery systems (DDS) based on nanocarrier (NCs) can improve solubility, stability and chemotherapeutic effects of drug, and reduce its toxicity [1]. In nanomedicine, various types of NCs such as liposomes, dendrimers, micelles, hydrogel and inorganic or semiconductor nanoparticles (NPs) were used. Semiconductor nanoparticles with unique physical and chemical characteristics have extensively been used in biomedical areas, such as bioimaging, gene delivery, cancer therapy and drug delivery [[2], [3], [4], [5], [6]]. Biocompatibility, high selectivity, and stability are the main characteristics of ZnO nanoparticles, which result in being promising potential in bioimaging and drug delivery [3]. Zinc as an important trace element in the human body plays different roles including the co-factor of variety enzymes in mammals contributing of DNA replication, DNA repair, and cell cycle progression [7].

There are various biosynthetic methods, among of which the use of plant extract being more effective due to their easy accessibility and various metabolites. This extraction helps the reduction of zinc ions and improves the rate of synthesis. Physical and chemical procedures for synthesis of NPs are expensive and consequently its biosynthesis has attracted much attention [[8], [9], [10], [11]]. The synthesis of nano-particles is not the final aim because these nanoparticles can be easily agglomerated into large-size particles again. Therefore, it is necessary that this agglomeration process be prevented and nanoparticles be stabilized using surfactant, polymer molecules, or any organic molecules bound to the surface of nanoparticles (e.g., chitosan) [7]. Chitosan is a natural polymer with positive charge, derived from the deacetylation of chitin. Regarding solid tumors having negative charge, the chitosan with positive charge can be attracted by these cells and facilitate drug delivery [12,13]. In addition to favorable biocompatibility, chitosan has the potential to enhance the cell membrane permeability both in vitro and in vivo. Some characteristics of chitosan including high potential in film-forming and chemical modifications, mechanical properties, biocompatibility, safety, high permeability towards water, and cost-effectiveness have introduced this polymer as a stabilizing agent in nanoparticles [[14], [15], [16]].

Paclitaxel (PTX), a plant-derived alkaloid, is a chemotherapeutic compound with antitumor activity in breast, prostate and cervical cancer. However, its application in cancer therapy is restricted partly because this alkaloid is not completely solved in water and thus the use of Cremophor EL formulation is necessary. The multiple doses of the drug are required to prevent the nonspecific toxicity due to high drug concentration, and maintain the therapeutic drug concentration in the tumor. The mechanism playing role in this action is binding to microtubules, stabilizing them, and ultimately causing apoptosis [[17], [18], [19], [20], [21]]. In this case, the development of ZnO NPs based DDS for PTX would be a promising approach which can increase the solubility of PTX leading to its enhanced bio-availability. Moreover, ZnO NPs – the NCs in this DDS – can also take part in the overall therapeutic efficacy of this system.

In this study, ZnO NPs was prepared using ethanolic extract from Camellia sinensis L. at 550 °C calcination temperature. Then, a novel PTX loaded chitosan coated ZnO NPs (ZnO-Ch-PTX) and its pharmacologic effect on breast cancer cell line (MCF-7) was evaluated. Moreover, a domestically established human foreskin fibroblast cell line was used to compare its side effects on normal tissues. In vitro MTT assay of the NPs and cell death pathways (necrosis/apoptosis) were also studied. Drug loading in the ZnO-Ch NPs was measured by high performance liquid chromatography (HPLC).

Section snippets

Preparation of ZnO NPs

The leaves of Camellia sinensis L. were collected from tea garden of Gilan province, North of Iran. It was authenticated by the Agricultural Research Center and Natural Resources of Gilan province. Dried leaves (2 g) of Camellia sinensis L. was mixed to ethanol (70 mL) for 72 h to obtain the ethanolic extract. Then, 2 g of zinc nitrate hexahydrate (Zn (NO3)2·6H2O (0.268 M) was added to the ethanolic extract (25 mL), and stirred at 60 °C. The obtained sample was incubated at 60 °C for 90 min,

Microstructure analysis

One of the most useful techniques for studying the phase structure and purity of a sample is the X-ray diffraction (XRD) spectroscopy. In this study, the XRD patterns of ZnO NPs which were prepared by Green method and through the ethanolic extraction from Camellia sineisis L. were studied. The ZnO NPs were calcinated at 550 °C and the result of XRD analysis is shown in Fig. 1a. It is clear that the ZnO NPs have a pure hexagonal wurtzite structure. The peaks in Fig. 1a. correspond to (100),

Conclusions

It can be concluded that the green synthetized ZnO-nanocarriers loaded with paclitaxel as selective drug delivery system in MCF-7 cell line had been successful in terms of improving the PTX efficacy by minimizing its cytotoxic effects on normal cells (fibroblast). Bearing in mind that both ZnO and chitosan have natural origin and are degradable, this drug delivery system (ZnO-Ch-PTX NPs) could be a promising candidate for further studies, including in-vivo pharmacological researches.

Declaration of Competing Interest

All authors declare that there is no conflict of interest about their role in the current study.

References (38)

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