In vitro cytocompatibility assessment and antibacterial effects of quercetin encapsulated alginate/chitosan nanoparticle

Highlights

• Significantly high level of antioxidant property for nanoencapsulated quercetin

• In vitro cytotoxicity assay exposed biocompatibility of nanoformulations.

• Q-ALG/CSNPs exposed enhanced antibacterial activity.

Abstract

The present work aims at evaluating the in vitro biocompatibility, antibacterial activity and antioxidant capacity of the fabricated and optimized Alginate/Chitosan nanoparticles (ALG/CSNPs) and quercetin loaded Alginate/Chitosan nanoparticles (Q-ALG/CSNPs) with an improved biological efficacy on the hydrophobic flavonoid.The physicochemical properties were determined by TEM and FTIR analysis. The nanoparticles evaluated for the encapsulation of quercetin exerted % encapsulation efficiency (EE) that varied between 76 and 82.4 % and loading capacity (LC) from 31 to 46.5 %. Potential cytotoxicity of the ALG/CSNPs and Q-ALG/CSNPs upon L929 fibroblast cell line was evaluated by MTT reduction Assay and expressed as % cell viability. The in vitro antibacterial property was studied by well diffusion method against gram-positive bacteria Staphylococcus aureus (ATCC 25925) and gram-negative bacteria Escherichia coli (ATCC 25923). The inhibitory efficacy by scavenging free radical intermediates was evaluated by 1,1, diphenyl 2-picrylhydrazyl (DPPH) assay. The results of in vitro cytotoxicity showed biocompatibility towards L929 cells. Quercetin loaded Alginate/Chitosan nanoparticles inhibited the growth of microorganisms than pure quercetin. The 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging results have shown a high level of antioxidant property for encapsulated Quercetin in Alginate/Chitosan nanoparticles compared to free Quercetin. The findings of our study suggest that the developed ALG/CSNPs and Q-ALG/CSNPs possess the prerequisites and be proposed as a suitable system for delivering quercetin with enhanced therapeutic effectuality.

Introduction

In recent years, advances in nanotechnology and its utilization in applications like drug delivery, targeted therapy, gene delivery, molecular imaging and tissue regeneration have enabled the researchers to find alternative approaches to attain the desired improvement in the pharmacokinetics of bioactive with poor dissolution [1], [2]. The insight of encapsulating hydrophobic compounds in appropriately designed nanoscale drug delivery system represents one of the eminently productive strategies facilitating its improved bioavailability and therapeutic effectuality [3]. Biomaterials pose significant challenges in fabricating stable delivery systems that could maintain their structural integrity under extreme pH conditions and digestive enzymes, as well as high ionic strength in the gastrointestinal environment. Lipids, protein, and polysaccharides are the three major natural food colloids that have been extensively studied to fabricate nanoscale oral delivery systems [4].When nanoformulations are to be considered biopolymers are at the forefront and another major direction in nanoscale drug formulations owing to their biochemical properties such like nontoxicity, biocompatibility, and biodegradability representing a suitable vehicle for efficient delivery of drugs. Polycarbohydrates might be formulated from pectin, cellulose, gum, chitosan, starch, gelatin [5], [6], [7]. Nanosystems applying herbal remedies show many advantages, including improvements in the solubility of poorly water soluble drugs, can deliver the active constituent at an adequate concentration throughout the treatment period, their nano range feature imparts the ability directing it to the desired site of action circumventing all the deterrents in the drug metabolism [8], [9]. The ability for controlled release of drugs from the nanoparticles makes it possible to tailor the pharmacokinetics of drugs and reduce their toxic side effects. Enhanced bio-distribution and expanded intracellular penetration improve the effectiveness of drugs [10].

Among several nano delivery systems such as micelles, dendrimers, nanofibers, niosomes and liposomes, polymeric nanoparticles are extensively employed biomaterials in both therapeutic and diagnostic applications due to their desirable characteristics in terms of colloidal stability in physiological medium, targeting specific cells or tissues and noticeable bio-imitative characteristics [11], [12], [13]. The physicochemical properties, effectiveness to condense lively agents such as hydrophilic/hydrophobic drugs, peptides, proteins, DNA, RNA, releasing mechanisms of bioactives, protecting the drug from degradation, dose reduction had also prioritized its attention in the easy adaptability for enhancement of efficacy and bioavailability, management. Thus, the pattern of delivery of polymeric nanoparticles certainly be different through different routes, and the physical and chemical properties might have a strong impact on the therapeutic outcome [14], [15]. Most importantly drugs encapsulated within the polymeric nanoparticles are released by diffusion through the polymeric matrices facilitating controlled drug release and efficient absorption associated without any subsequent toxic side effects [16]. With the aforementioned advantages, the implementation of nanotechnology in drug delivery would enable the use of lower doses of the drug, contributing to a betterment in the action of pathologies as well as a inferior environmental impact [17], [18].

It has been recognized that Quercetin is a major flavonoid among the group of secondary metabolites found in numerous fruits, vegetables, rhizomes, tree barks, flowers, tea and is a most active promising component responsible for diverse beneficial pharmacological effects including DNA protection, antioxidative, antimicrobial and anticarcinogenic effect [19]. Studies have proposed that an increase in the intake of dietary quercetin can help prevent the risk of cardiac diseases, neurodegenerative diseases, aging predominantly relating to its ability to scavenge free radicals [20], [21]. Outstandingly the antihypertensive effects of quercetin and ameliorative endothelial function, its anti-thrombotic, anti-inflammatory and anti-obesity effects seem to be the most relevant [22]. However, the hydrophobicity, low oral bioavailability and instability in the physiological medium of quercetin limits its biomedical application [23]. The outcome of flavonoids is claimed to be ameliorated when they are in nanoformulations [24]. Quercetin was encapsulated into polymeric monomethyl poly(ethylene glycol)–poly(3-caprolactone) micelles using a simple solid dispersion method [25].Chitosan/ZnO bio-nanocomposite hydrogel beads as controlled drug delivery system was studied by Mehdi Yadollahi et al., [26]. Accordingly to make quercetin available at the spot of physiological activity for a longer period of time and to increase its bioactivity, in our previous study we reported a nanosystem wherein quercetin had been effectively loaded and encapsulated in optimized Alginate/Chitosan nanoparticles (ALG/CSNPs) in nanoregimen with good stability and sustained drug release efficacy [27].

Biocompatibility, a unifying property of biomaterials involves that the biomaterials should not directly or indirectly be the cause of any local or systemic effects and cause no adverse carcinogenic, reproductive and developmental effects effects [28]. The development of nanoparticles made from food biopolymers (i.e., protein and polysaccharide)complex is to develop safe and effective oral delivery systems for food bioactive compounds using food derived biomaterials to improve the bioavailability of nutrients and thus the overall quality and health benefits of functional food [4], [29], [30]. Similarly in our study we envisioned to design novel nanocarriers using polysaccharide based alginate-chitosan nanoparticles to encapsulate the phytoconstituent quercetin for biomedical applications. In the current nanotoxicity criterion, in vitro biocompatibility assays are mainly the first to be conducted wherein monocultures of cancer or long-lived cell lines are the choice in demonstrating the specific behavior of the nanoparticles in culture media. The primary objective of this present study was to investigate the in vitro cytocompatibility using MTT cell viability assay, antibacterial activity using gram-positive (S. aureus), gram-negative (E. coli) pathogens, antioxidant capacity of the fabricated blank and quercetin loaded nanoparticles.