Development of biodegradable nanoparticles for delivery of quercetin

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Abstract

The antioxidant molecule quercetin has been encapsulated on poly-d,l-lactide (PLA) nanoparticles by solvent evaporation method for the improvement of its poor aqueous solubility and stability. The surface morphology and average size of PLA and quercetin loaded PLA nanoparticles are 170 ± 25 and 130 ± 30 nm respectively. The antioxidant activities of the PLA encapsulated quercetin nanomedicine are identical to free quercetin. The nanoencapsulation efficiency of quercetin evaluated by HPLC and antioxidant assay is 96.7%. The in vitro release kinetics under physiological condition show initial burst release followed by slow and sustained release. The complete release and maximum retention of quercetin is 72 and 96 h respectively. The less fluorescence quenching efficiency of quercetin–PLA nanoparticles than free quercetin on BSA confirms the controlled release of quercetin from PLA nanoparticles. These properties of PLA encapsulated quercetin molecule pave way for encapsulating various therapeutically less useful highly active antioxidant molecules towards the development of better therapeutic compounds.

Introduction

Quercetin, 3,3′,4′,5′-7-pentahydroxy flavone (Fig. 1) is one of the most abundant flavonoid in plants. It is abundantly found in varying concentrations in berries between 53 and 153 mg kg−1of dry weight of plant material. This molecule is an important constituent of wine and its concentration varies from 1 to 33 μM. The eating of fried onions (equivalent to 225 μmol quercetin) and apples (equivalent to 325 μmol quercetin) increases the peak plasma levels of quercetin up to 0.74 and 0.30 μM respectively [12]. The antioxidant activity of this molecule is higher than well-known antioxidant molecules ascorbyl, trolox and rutin [14]. This is due to the number and position of the free hydroxyl groups in the quercetin molecule [1]. The flavonoid glycosides are rapidly hydrolysed in the small intestine or by bacterial activity in the colon to generate the quercetin aglycones, which is further metabolized into the glucuronidated or sulfated form of quercetin [6]. This molecule is retained in the large intestine for approximately 6 h after oral administration. However, it is chemically unstable, especially in aqueous alkaline medium, which possibly involves attack of hydroxyl ions on the C-ring of quercetin [11]. Apart from the antioxidant activity, this molecule shows anticancer and antiviral activities also [2], [20], [29]. In spite of this wide spectrum of pharmacological properties, the use of quercetin in pharmaceutical field is limited due to its low aqueous solubility and instability in physiological medium [16]. These properties of quercetin result in poor bioavailability, poor permeability, instability and extensive first pass metabolism before reaching the systemic circulation [18]. One-way to circumvent these problems are to entrap/adsorb these molecule into biodegradable polymeric nanoparticles. Among the biodegradable polymeric nanoparticles [8]. PLA is extensively used for the encapsulation of many therapeutic agents due to its high hydrophobicity, biodegradability, biocompatibility, low toxicity, strong mechanical strength and slow drug release [9]. Quercetin (synthetic) molecule has been successfully encapsulated into liposomes [17] and chitosan nanoparticles [27]. However, the detailed characterisations of liposome and chitosan nanoencapsulated quercetin molecule are not reported. In this study, we have investigated the feasibility of encapsulating synthetic quercetin molecule into PLA nanoparticles. The solvent evaporation method has been used for the encapsulation of this molecule on polymeric PLA nanoparticles. The quercetin loaded PLA nanoparticles have been characterised by scanning electron microscope, atomic force microscope, UV–vis spectrophotometer. Effect of quercetin loaded PLA nanoparticles on fluorescence quenching of BSA protein has also been evaluated. The quantification of encapsulation efficiency, antioxidant activity and in vitro release was also carried to enhance its application in pharmaceutical field. PLA encapsulated quercetin molecule shows higher aqueous solubility and sustained release. Thus it is speculated that the PLA nanoencapsulation may improve the bioavailability and stability of quercetin and other similar small molecular drugs.

Section snippets

Materials

Poly-d,l-lactide (PLA) (MW = 75,000–120,000) and polyvinyl alcohol (PVA) were purchased from Sigma–Aldrich and used as received. Quercetin was purchased from Merck, and used as received. Dichloromethane (DCM) was purchased from Merck. HPLC grade acetonitrile (ACN), water, ethanol and trifluoroacetic acid (TFA) were procured from Sigma. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) was purchased from Fluka. Solutions were prepared using water filtered through a Milli-Q water system (Millipore, Bedford, MA).

Synthesis and encapsulation of quercetin in PLA nanoparticles

The antioxidant molecule quercetin is hydrophobic in nature. Thus for encapsulation of this molecule PLA has been found to be the best biodegradable polymer among other known polymeric molecules used for nanoparticles synthesis. This molecule was encapsulated on PLA by solvent evaporation method in its fully active forms. This method is based on the emulsification of organic phase (DCM + PLA + quercetin) and aqueous phase (PVA solution). The emulsion droplets were stabilised by the surfactant

Conclusion

Quercetin, a lipophilic drug, was successfully encapsulated on PLA nanoparticles using solvent evaporation method with encapsulation efficiency of 96.7% and 19.4% actual drug loading. The mean diameter of PLA nanoparticles and quercetin loaded PLA nanoparticles was ∼170 ± 25 and ∼130 ± 30 nm respectively. Antioxidant activity assay revealed that the functional activity of quercetin was retained after nanoencapsulation. The biphasic release profile includes initial burst effect followed by sustained

Acknowledgements

We are grateful to Dr. P.S. Ahuja, Director, IHBT for providing necessary facilities for carrying out this work. The IHBT communication number of this article is 2009. Financial assistance to Avnesh Kumari, from Council of Scientific and Industrial Research (CSIR) and Women Scientists Scheme (A) (SR/WOS-A/CS-61/2008), Department of Science and Technology (DST), Government of India is truly acknowledged.

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