Pharmaceutical NanotechnologyMarked effects of combined TPGS and PVA emulsifiers in the fabrication of etoposide-loaded PLGA-PEG nanoparticles: In vitro and in vivo evaluation
Graphical abstract
Using TPGS in combination with PVA as an emulsifier in fabrication of etoposide (ET) loaded nanoparticles (NPs) markedly increased encapsulation efficiency, sustained drug release and resulted in nanoparticles with noticeable sustainable in vivo disposition.
Introduction
In recent years, research on colloidal drug carriers has intensified. In particular, polymeric nanoparticles (NPs) are thought to be a suitable way to control drug release and targeting (Park et al., 2009, Peracchia et al., 1997). Drug carriers can modify the original pharmacokinetics and biodistribution of an active substance, thereby raising the drug concentration in targeted tissue while improving efficacy and reducing systemic side effects (Moghimi et al., 2001). In order to reach these important objectives, NPs must be composed of a biodegradable and biocompatible polymer. NPs must also have a rigorously restricted diameter and size distribution, sufficient reservoir amount and sustained release properties (Panyam and Labhasetwar, 2003). These characteristics of NPs have to be optimized during the preparation process depending on their physicochemical properties (Sant et al., 2005). Any fabrication parameters or additives used in the formulation could alter the characteristics of the resulting NPs. During the preparation process of NPs, emulsifiers play a key role in separating the polymer containing phase (oil) from the dispersion medium (water) (Soppimath et al., 2001). Emulsifiers also stabilize the produced emulsion, decrease particle aggregation and affect the particle size and distribution, morphological properties, encapsulation efficiency and drug release characteristics of NPs (Coombes et al., 1998, Mitra and Lin, 2003, Mu and Feng, 2002). Polyvinyl alcohol (PVA) is the most commonly used emulsifier during the formulation of poly (lactic-co-glycolic acid) (PLGA) NPs because this emulsifier generates particles that are relatively uniform, small and easy to redisperse in aqueous medium (Sahoo et al., 2002).
TPGS (d-alpha tocopheryl polyethylene glycol 1000 succinate), a water-soluble derivative of natural vitamin E, contains both a hydrophilic moiety and a lipophilic moiety, and therefore is similar to conventional surfactants. TPGS is also a p-glycoprotein efflux inhibitor (Collnot et al., 2006, Dintama and Silverman, 1999). Accordingly, it has been effectively applied to advance NP properties for controlled delivery. Feng and coworkers applied TPGS as a surfactant stabilizer for the first time to fabricate paclitaxel-loaded PLGA nanospheres using solvent evaporation/extraction. They indicated that TPGS could be an ideal and effective emulsifier in NP formulation (Mu and Feng, 2002). In addition to the other applications of TPGS in drug delivery systems, this group of researchers has successfully prepared and evaluated various TPGS-emulsified NPs, such as PLGA (Feng et al., 2007a, Mu and Feng, 2003a), MPEG-SS-PLA (disulfide bond-containing poly(ethylene glycol)-b-poly(lactic acid)) (Song et al., 2011;) and PLA based NPs (Zhang et al., 2008). Notably, the majority of these studies have focused on paclitaxel delivery (Feng et al., 2004, Feng et al., 2007a, Feng et al., 2007b, Mu and Feng, 2002, Mu and Feng, 2003a, Mu et al., 2004, Song et al., 2011, Zhao and Feng, 2010). However, different results have been reported when other drugs were investigated. Sengel et al. (2011) applied TPGS as an emulsifier for the preparation of meloxicam loaded PLGA NPs using ultrasonication-solvent evaporation. In that study, TPGS was not shown to be an effective emulsifier since it did not result in a very high drug encapsulation of the drug meloxicam in the NP formulation. Moreover, for amphotericin B and atorvastatin calcium-loaded PLGA nanoparticles in which TPGS was used as the emulsifier, the encapsulation efficiency was not high (∼34.5%) (Italia et al., 2009, Meena et al., 2008). Thus, the efficiency of TPGS as an emulsifier for encapsulating various drugs in polymeric NPs under different preparation conditions needs to be further evaluated.
To the best of our knowledge, there has been no systematic study on the influence of TPGS on the in vitro characteristics of PEGylated PLGA NPs.
Etoposide is a widely used anticancer drug and is prescribed alone or in combination with other anticancer drugs to treat both solid tumors and hematological malignancies (Montecucco and Biamonti, 2007). However, etoposide administration is limited because of its strong lipophilicity and chemical instability (Hande, 1992, O’Dwyer and Weiss, 1984, Shah et al., 1989). Etoposide also causes dose-limiting hematological toxicity (Sinkule, 1984). In order to overcome these disadvantages, drug-loaded nanoparticles fabricated from biodegradable polymers might represent an ideal alternative dosage form.
The main objective of this study was to evaluate the effect of TPGS as an emulsifier, both alone and in combination with several commonly used emulsifiers on particle size, encapsulation efficiency and release behavior of PLGA-PEG NPs containing etoposide. Specifically, this study evaluated etoposide loaded PLGA-PEG NPs prepared via single-emulsion solvent evaporation method. Afterward, the same factors were examined on NPs formed by nanoprecipitation, which is a frequently used, simple one-step preparation technique. Finally, the in vivo disposition of the optimal formulation was evaluated after intravenous (IV) administration of free and encapsulated drugs in rats. The results showed that employing TPGS in combination with PVA markedly increased the encapsulation efficiency, sustained drug release and improved the circulation of NPs in vivo.
Section snippets
Materials
PLGA-PEG 5% (RGP d 5055) was obtained from Boehringer Ingelheim (Ingelheim, Germany). Etoposide was kindly provided by Cipla (Mumbai, India). Polyvinyl alcohol (PVA, Mowiol®, MW 30,000 Da, 87% hydrolyzed) was a gift from Hoechst (Frankfurt, Germany). TPGS, sodium cholate and pluronic F68 (F68) were provided by Sigma–Aldrich (Steinheim, Germany). Analytical grade acetone and dichloromethane (DCM) were purchased from Merck (Darmstadt, Germany). HPLC grade methanol and acetonitrile were obtained
Drug content
Our preliminary studies showed, apart from the type of surfactant used, at low drug percentage in the internal phase (below 0.07% w/v), very little drug was encapsulated into the prepared NPs in both preparation methods (EE < 10%). Above that, an increase in the drug amount led to a significant increase in the EE until the optimal drug amount provided the highest EE. Based on these results, drug percentage of approximately 0.15% (w/v) and 0.3% (w/v) gave the maximum EE for the nanoprecipitation
Discussion
In both solvent evaporation and nanoprecipitation preparation methods, the most important factor controlling the EE was the drug percentage in the internal phase. A likely reason for this observation could be due to drug partitioning between oil and water phases during NP formation. When the drug amount is greater, the aqueous concentration approaches the saturation limit, which results in its entrapment in organic phase and consequently higher encapsulation in produced NPs.
Conclusion
In this investigation, PLGA-PEG NPs loaded with etoposide were prepared by nanoprecipitation and emulsion-solvent evaporation methods, and then the effect of different surfactants (PVA, F68, sodium cholate and TPGS) and combinations of TPGS and PVA on in vitro characteristics was evaluated. The results showed a clear influence of the surfactant type on the particle size, EE and release behavior in both preparation methods. However the NPs produced by nanoprecipitation did not provide proper
Conflict of interest
The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.
Acknowledgments
This research was supported by a grant from Shahid Beheshti University of Medical Sciences, Tehran, Iran. The authors wish to thank Mrs. Zahra Abbasian for her technical assistance.
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