Pharmaceutical Nanotechnology
Evaluation of ciprofloxacin-loaded Eudragit® RS100 or RL100/PLGA nanoparticles

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Abstract

The objective of present study was to prepare positively charged ciprofloxacin-loaded nanoparticles providing a controlled release formulation. The particles were prepared by water-in-oil-in-water (w/o/w) emulsification and solvent evaporation, followed by high-pressure homogenisation. Two non-biodegradable positively charged polymers, Eudragit® RS100 and RL100, and the biodegradable polymer poly(lactic-co-glycolic acid) or PLGA were used alone or in combination, with varying ratios. The formulations were evaluated in terms of particle size and zeta potential. Differential scanning calorimetry measurements were carried out on the nanoparticles and on the pure polymers Eudragit® and PLGA. Drug loading and release properties of the nanoparticles were examined. The antimicrobial activity against Pseudomonas aeruginosa and Staphylococcus aureus was determined. During solvent evaporation, the size and zeta potential of the nanoparticles did not change significantly. The mean diameter was dependent on the presence of Eudragit® and on the viscosity of the organic phase. The zeta potential of all Eudragit® containing nanoparticles was positive in ultrapure water (around +21/+25 mV). No burst effect but a prolonged drug release was observed from all formulations. The particles’ activity against P. aeruginosa and S. aureus was comparable with an equally concentrated ciprofloxacin solution.

Introduction

Most ocular infections are treated by topical application of antibiotic solutions administered as aqueous eye drops. A low bioavailability is observed due to rapid and extensive precorneal loss; consequently, instillation of highly concentrated solutions or frequent administration is required, resulting in poor patient compliance. New drug delivery systems for ophthalmic administration, such as microparticles, liposomes or nanoparticles, have been developed and studied over the last decades and were designed to combine prolonged ophthalmic action with the ease of application of liquid eye drops (Zimmer and Kreuter, 1995, Le Bourlais et al., 1998). After administration, colloidal drug carriers can remain at the application site (cul-de-sac) and the prolonged release of the active ingredient starts by particle degradation or erosion, drug diffusion, or a combination of both, depending on the biodegradable or inert nature of the polymer (Soppimath et al., 2001).

Poly(dl-lactic-co-glycolic acid) or PLGA, a copolymer of lactic and glycolic acid is one of the most biocompatible, biodegradable and non-toxic materials used for preparing nano- and microparticles (Zimmer and Kreuter, 1995, Bala et al., 2004). Blends of Eudragit® and PLGA have been described in the preparation of heparin-loaded nanoparticles as potential oral carriers (Hoffart et al., 2002, Jiao et al., 2002).

Eudragit® RS100 (RS) and RL100 (RL) polymers have been proposed as ocular delivery systems with prolonged release and improved ocular availability, but mainly for non-steroidal anti-inflammatory drugs (Pignatello et al., 2002a, Pignatello et al., 2002b). Such drug-loaded carrier systems showed good stability properties and size distribution and a positive surface charge, which makes them potential ocular drug delivery systems. This charge can allow a prolonged residence time of the nanoparticles on the corneal surface because of interactions with anionic mucins present in the tear film. Microspheres made of RS or RL containing gentamicin or acetazolamide have also been described as ophthalmic delivery systems (Safwat and Al-Kassas, 2002, Haznedar and Dortunç, 2004).

Desgouilles et al. (2003) proposed two hypotheses according to the mechanism of particle formation during polymer precipitation: one nanoparticle arising from one emulsion droplet, after shrinkage, or from fusing of several emulsion droplets. The physicochemical parameters size and zeta potential were followed during the process of solvent evaporation to determine whether a reassembling of the polymers at the particles’ surface occurred. Such a rearrangement could occur in Eudragit® RS or RL containing nanoparticles, due to their amphiphilic properties.

Ciprofloxacin, a powerful broad-spectrum fluoroquinolone antibiotic, useful in the treatment of infections of the outer eye such as bacterial conjunctivitis and keratitis, has been chosen as a model drug for incorporation in the formulation (Blondeau, 2004). It has been reported that fluoroquinolones possess an in vitro efficacy against gram-negative as well as gram-positive ocular pathogens, superior to other antibiotics tested (Armstrong, 2000, Egger et al., 2001). Moreover, the frequency of spontaneous resistance to ciprofloxacin is very low (Hwang, 2004). Besides providing a controlled release system, nanoparticles can be a promising alternative for eye drops, since during intensive dosing of ciprofloxacin containing eye drops or ointments, corneal precipitates and crystalline deposits at eyelashes and in dropper bottle tips may occur (Wilhelmus and Abshire, 2003).

Pseudomonas aeruginosa and Staphylococcus aureus, some of the most common ocular pathogens causing bacterial infection of the human cornea (keratitis and cornea ulceration), especially ulcers underneath contact lenses, were selected as test microorganisms during the determination of antimicrobial activity of the formulations prepared (Armstrong, 2000, Bourcier et al., 2003).

The aim of present study was to evaluate the particle formation process, the physicochemical properties and activity of ciprofloxacin·HCl-loaded nanoparticles prepared from mixtures of poly(lactide-co-glycolide) (PLGA) and Eudragit® in different ratios.

Moreover, the objective was to prepare positively charged nanoparticles which could interact with the anionic mucins present in the mucus layer of the tear film.

Section snippets

Materials

The PLGA polymer Resomer® RG 503 was obtained from Boehringer Ingelheim (Ingelheim am Rhein, Germany) and Eudragit® RS100 and RL100 from Röhm Pharma (Darmstadt, Germany). Poly(vinylalcohol) or PVA (MW 30,000–70,000) was supplied by Sigma Chemicals Co. (St. Louis, USA), ciprofloxacin hydrochloride monohydrate by Roig Farma (Barcelona, Spain) and d-mannitol by Federa Co. (Brussels, Belgium). Dichloromethane was purchased from Sigma–Aldrich (Steinheim, Germany) and acetonitrile (HPLC grade) from

Physical characterisation: particle size and zeta potential measurements

All batches showed a small mean size, well suited for possible ocular application to prevent patient discomfort and provide a good drug diffusional release. Particle size for ophthalmic applications should not exceed 10 μm because with larger sizes a scratching feeling might occur (Zimmer and Kreuter, 1995). Therefore, a reduced particle size improves the patient comfort.

In Fig. 1, an overview is shown of particle sizes of nanoparticles consisting of Eudragit® and/or PLGA, before and after

Conclusions

Although PLGA nanoparticles are biodegradable, they possess a negative zeta potential and probably low interactions with anionic mucus. Therefore, the possibility of producing positively charged nanoparticles by adding Eudragit to these formulations was investigated. Since the zeta potential of Eudragit®/PLGA mixtures was independent of the percentage of Eudragit® incorporated, the lowest concentration Eudragit (25%) could be selected for further research, since it contains the lowest amount of

Acknowledgements

K. Dillen is a Research Assistant of the Fund for Scientific Research-Flanders (Belgium) (F.W.O. Vlaanderen). The authors wish to thank Prof. D. Vanden Berghe, Dr. P. Cos and Mrs. R. Vingerhoets, Laboratory of Microbiology, University of Antwerp, for the assistance with the microbiological tests and Dr. Pierre Dardenne (Sterigenics, IBA-Mediris S.A., Fleurus, Belgium) for gamma-sterilisation of the nanoparticles and polymers.

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