Exploration of interfacial dynamics in squaraine based nanohybrids for potential photodynamic action

Highlights

• Various sizes of ZnO nanoparticles are synthesized and the surface is modified using Squaraine as ligand molecule.

• Nanohybrids show molecular level interaction described using FRET.

• Photoinduced charge transfer enhances its activity.

• ZnO-SQ nanohybrids is used for potential photodynamic cancer therapy.

Abstract

Photodynamic therapy (PDT) is a clinically approved, minimally invasive therapeutic procedure exhibiting cytotoxic effects toward malignant cells. Hydrophobic nature of most photosensitizers used in PDT with lower light absorption ability restricts practical use of PDT. Herein, we have employed a squaraine drug (SQ) with 665 nm absorbance peak maxima as the photosensitizing agent and evaluate its photo-physical properties. The tendency of aggregation formation in aqueous media limits its practical usefulness. Thus, we have synthesised wide band gap semiconductor zinc oxide (ZnO) nanoparticles and functionalized the surface using squaraine molecules. The molecular cross-talking was evaluated using excited state fluorescence lifetime decay profiles and by employing Fӧrster resonance energy transfer (FRET) technique. The nanohybrids show improvement in three aspects compared to bare SQ molecule such as lesser aggregate formation in aqueous media, pH responsive precipitation of the drug and improvement of photo-induced reactive oxygen species (ROS) generation. Ultrafast dynamical study at the inorganic (ZnO) - organic (SQ) interface depicts presence of photo-induced charge transfer process at the junction which indeed improves the ROS generation capability. Finally, the photodynamic action has been evaluated in human breast cancer cell line MCF-7. The nanohybrids depict enhanced photo toxicity in cancer cell with loss of adherence and typical morphology of cancerous cell depicting controlled cell death. The present study employ characterisation of nanohybrids for potential use in PDT for cancer treatment.

Introduction

Squaraine, condensation products of electron-rich substrates and squaric acid derivatives, with a four-membered squaraine ring present into the polymethine chain structure consist of absorption maxima in longer wavelength region [[1], [2], [3]]. Besides large applications in dye sensitized solar cell [4], squaraine dyes impose greater light-absorption ability at low-energy region and higher photo-stability with respect to other available commercial dye and can be considered as photomedicine [5]. Recently, photodynamic therapy (PDT) substantiates its effectivity towards destruction of malignancy in a minimally invasive approach [6]. PDT involves a photosensitizer (PS) molecule that can be excited from the ground state to excited state upon irradiation of light of specific wavelength similar to the band-energy gap of the molecule [7], producing active molecular species, such as free radicals and reactive oxygen species (ROS) following cell death [8]. The efficacy of PDT depends on several factors as light and drug dosimetry, drug localization to the diseased site, light exposure and its penetration capability through biological tissues [9,10]. In this view, SQ can be considered as a new age photosensitizer molecule [11]. However, low water solubility and aggregation in biological fluid restrict their applications [12]. The aggregated forms include deactivation of excited state through various non-radiative process which lowers the ROS generation capability [13]. Thus, several drug delivery agents are employed to enhance the effectivity of squaraine.

Earlier it was reported that chitosan, a biopolymer based drug-delivery system with hydrophilicity, biocompatibility, biodegradability properties can serve well as a platform for squaraine delivery [14]. The incorporation of dyes onto suitable surfaces improve molecular rigidification by diminishing the photoisomerization process which enhance dye photostability and solubility [15]. On the other hand, inorganic nanoparticles (NPs) with wide band gap often establish themselves as a candidate for better drug loading as well as an active component for enhancing the drug activity [16]. In this purpose, ZnO NPs with a wide band gap of3.3 eV is a promising nanomaterial. ZnO itself often depicts photodynamic action in presence of UV irradiation [17] and generates ROS that can kill the cancer [18]. The pH responsive precipitation and dissolution of ZnO NPs can magnify its potential as cancer therapeutics [19]. However, UV light is itself very much harmful for human tissue often affecting the normal tissues significantly [20]. Thus, surface functionalization of ZnO NPs with photosensitizer molecules can lead to nanohybrids formation with alternate properties that will lead to a new avenue of nanoparticles based PDT systems [21,22].

Herein, we have examined a squaraine dye (SQ) with absorption at longer wavelength region for potential PDT application. The dye depicted absorbance at 665 nm and emission at 675 nm and thus can be consider as a novel PDT agent. Next, we have synthesised the surface modified ZnO NPs using the dye as the sensitizer. To check the integrity of nanohybrids formation, we have followed Fӧrster resonance energy transfer (FRET) from donor ZnO to acceptor SQ molecule. The photo-induced electron transfer process is estimated using time-resolved fluorescence decay transients. The enhanced water stability, lesser aggregation, pH responsive delivery of drug in the nanohybrids has been monitored. The ROS generation was assessed using dichlorofluorescin (DCFH) assay. We have tested and checked the light penetration efficacy through a mimic human tissue and evaluated the cytotoxicity of ZnO-SQ nanohybrids on human breast cancer cell line MCF-7. The present study depicts physico-chemical insight of an inorganic-organic hybrid material for potential biological effectivity.