Stability study of nanoemulsions containing zinc phthalocyanine for photodynamic therapy of cancer

Malahat Rezaee,1 Mahnoosh fatemi,2,* Fereshte ghandehari,3 Mojdeh dinarvand,4

1. Department of Biochemistry, Falavarjan Branch, Islamic Azad University, Isfahan, Iran
2. Department of Biology, Falavarjan Branch, Islamic Azad University, Isfahan, Iran
3. Department of Microbiology, Falavarjan Branch, Islamic Azad University, Isfahan, Iran
4. Microbial Pathogenesis and Immunity Group, Department of Infectious Disease and Immunology, Faculty of Medicine and Hea

Abstract


Introduction

Photodynamic therapy (pdt) is a clinically approved, minimally invasive therapeutic procedure that can exert a selective cytotoxic activity toward malignant cells. pdt combines light, molecular oxygen and a photosensitizer (ps) to induce oxidative stress in target cells. the oxidative stress has the potential to induce cell death or a therapeutically significant stress response. certain hydrophobic photosensitizers, such as zinc-phthalocyanine (znpc), have significant potential for antitumor pdt applications. however, hydrophobic molecules of ps often require drug-delivery systems, such as nanoemulsions, to improve their pharmacokinetic properties and to prevent aggregation, which has a quenching effect on the photoemission properties in aqueous media. however, in spite of the nanoemulsions advantages, they usually face some physical unstability during the time, due to changes in temperature and pressure and addition of additives that can affect the system and prevent effective drug delivery. the homogeneity of the particle size, surface charge (zeta potential) of the particles, ratio of co-surfactants and surfactant(s) are reported to have a role in determining the stability of nanoemulsion formulations. as a result, this work aims to test the physical stability of znpc nanoemulsions to choose the best formulation that use in anticancer photodynamic therapy.

Methods

The oil-in-water nanoemulsions were prepared from a mixture of oil and surfactants in different s/o ratio by low energy emulsification method. at first, znpc (0.005 wt%) were solved in hazel nut oil using sonicator for 30 minutes. lecithin and cremophore el were applied as mixed surfactants in the three s/o ratios of 1.45, 1.75 and 2.25 in the formulations of f1, f2 and f3, respectively. the amount of added water kept constant at 73 g wt%. emulsion droplet size and polydispersity index (pdi) were determined by dynamic light scattering. the surface charge of nanoemulsion droplets was determined using a zeta potential analyzer. stability evaluation of the nanoemulsions were performed using centrifugation, freeze-thaw cycle and storage stability tests. the formulations were centrifuged at 4000 rpm for 1 h and observed for phase separation. the stable formulations were tested for the freeze-thaw cycle test. for freeze-thaw test, the formulations were subjected to six freeze-thaw cycles which consisted freezing at -18.0°c for 16 h followed by thawing at 25.0°c for 8 h. the stability of the nanoemulsions were evaluated at 4.0°c, 25.0°c and 45.0° for three months. the physical appearance of the formulations (sedimentation, creaming and phase separation) was observed, and the changes in particle size, pdi and surface charge of the formulations were recorded over the storage period of three months.

Results

The particle size of nanoemulsions were 133.9, 164.4 and 215.9 nm for the formulations of f1, f2 and f3, respectively. pdi was determined 0.349, 0.411 and 0.517 for the f1, f2 and f3, respectively. it means that with increasing the s/o ratio particle size and pdi were decreased. in addition, zeta potential of particles for f1, f2 and f3 was calculated -66.6,-65.2 and -75.9 mv, respectively. also, all formulations showed a good physical stability after centrifugation at 4000 rpm for 1 hour. endurance of an emulsion against centrifugation is related to the difference of density between the oily and aqueous phases and on the resistance on the interfacial film. f1 and f2 showed good stability against freeze-thaw test. this result could be due to the sufficient amount of surfactants in these formulations, which can provide a thicker layer supporting the droplets to protect them against the ice region during the freezing. in addition, both f1 and f2 showed good stability in three months storage at 4.0°c and 25.0°c, however the changes in particle size, pdi and zeta potential for f1 was not considerable and significant. at 45.0°c just f1 was remained stable. ostwald ripening plot was constructed to evaluate and compare the stability of the formulations.

Conclusion

The current study demonstrated that the selected oil and surfactants were adequate for formulation of nanoemulsion systems. the stability evaluations showed that s/o ratio had impact effect on size, pdi and zeta potential and finally on the nanoemulsions stability. the best formulation with high stability was f1 with s/o of 1.45 which can select for incorporation of znpc in nanoemulsions for treatment of cancer.

Keywords

Photodynamic therapy, nanoemulsions stability, anticancer