Effects of different combinations of nanocrystallization technologies on avanafil nanoparticles: in vitro, in vivo and stability evaluation

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Abstract

The study investigated the effects of different combined top-down and bottom-up nanocrystallization technologies on particle size and solid state of avanafil nanoparticles. Combined antisolvent precipitation-ultrasonication (sonoprecipitation) technique was adopted to prepare 18 formulas according to 32.21 factorial design using 3 stabilizers; Tween 80, polyvinyl alcohol (PVA) and Pluronic F68 at different concentrations with different cryoprotectants. Particle size analysis of the lyophilized formulas showed that Tween 80 was an effective nanoparticles stabilizer in contrast to Pluronic F68 and PVA which failed to prevent nanoparticles flocculation when they were used at high concentration. The combined effects of nanonization and amorphism contributed to the improvement in solubility. Further processing of the sonoprecipitated formulas by high pressure homogenization (HPH) (modified NANOEDGE™ technology) resulted in further size reduction of PVA-stabilized particles, while it stimulated flocculation of Tween-stabilized nanoparticles. Nevertheless, all of the homogenized formulas partially retrieved their crystallinity which reduced their solubility. Non-homogenized formula 2E composed of 1:2 (avanafil: Tween) with glucose as cryoprotectant, exhibited 13.68- and 2.59-fold improvement in solubility and in vitro dissolution, respectively. This formula had oral bioavailability of 137.02% relative to Spedra® tablets and it maintained its nanosize, amorphism and dissolution behavior over 6 months of storage under stress conditions.

Introduction

Avanafil is an inhibitor of phosphodiesterase enzyme type 5 (PDE5). It was approved by US Food and Drug Administration (FDA) (2012) followed by European Medicines Agency (2013) for treatment of erectile dysfunction. It is characterized by its faster onset of action and its lower visual side effects than other PDE5 inhibitors (Alwaal et al., 2011, Limin et al., 2010). However, avanafil is poorly soluble in water which contributes to its low bioavailability (38–41%) (Burke and Evans, 2012). Formulation of poorly water soluble drugs as nanoparticles – which were defined by Junghanns and Müller (2008) as particles of size <1-μm – is a traditional method used for improvement of their oral bioavailability (Mudshinge et al., 2011). Several techniques have been used for preparation of nanoparticles including top-down technologies, bottom-up technologies and combinations of both (Müller et al., 2011). The top-down approach refers to particle size reduction by wet milling, ultrasonication and high-pressure homogenization (HPH) (Keck and Müller, 2006, Moschwitzer, 2013). However, the bottom-up approach refers to methods that create small drug particles from drug molecules dissolved in an organic solvent either by anti-solvent precipitation or evaporation (Sinha et al., 2013). Kipp et al. (2003) invented NANOEDGE™ technology which employed combination of microprecipitation and HPH for preparation of stable crystalline nanoparticles of poorly soluble drugs. Later on, Dhumal et al. (2008) devised a combined top-down and bottom-up technique called sonoprecipitation which involved anti-solvent precipitation followed by ultrasonication. The technique was applied for preparation of nanoparticles of cefuroxime axitel which were resolved in an amorphous form. The solid state of the prepared nanoparticles is a crucial factor affecting their oral bioavailability. Nonetheless, the use of amorphous nanoparticles in the pharmaceutical market is limited by physical stability considerations (Sun and Yeo, 2013). It is very challenging to prepare amorphous nanoparticles that can maintain their nanosize and solid state over their shelf-life which is considered a prerequisite for the use of this product in pharmaceutical industries (Junghanns and Müller, 2008). This study aimed at preparation of stable amorphous nanoparticles of avanafil. The study introduced a modified NANOEDGE™ technology which is composed of three subsequent processes; anti-solvent precipitation, ultrasonication and finally HPH. The rationale of the study is comparing the particle size, zeta potential and solid state of the nanoparticles prepared by sonoprecipitation technique to those prepared by modified NANOEDGE™ technology. The oral bioavailability and shelf-life stability of a selected formula were investigated.

Section snippets

Materials

Avanafil and dapoxetine hydrochloride were purchased from Arcadia biotechnology (Shanghai, China), Tween 80 (polyoxyethylene sorbitan monooleate, hydrophilic-lipophilic balance (HLB) = 15), polyvinyl alcohol 89–98 KDa (PVA) (HLB = 18), dimethylsuphoxide (DMSO), methanol, absolute alcohol, high-performance liquid chromatography (HPLC) grade acetonitrile, hydrochloric acid, glucose and trehalose were purchased from Sigma-Aldrich (Taufkirchen, Germany), Pluronic F68 (polyoxyethylene-polyoxypropylene

Drug content

Mean drug loading values ranged from 97.8 ± 0.3% to 101.5 ± 1.7% for the prepared formulas. Two-sided t-test did not identify any significant differences between the actual mean drug contents and the hypothetical value in all formulas.

DLS

Table 1 shows that the prepared formulas exhibited wide variation of Z-average and PDI. Formula 2B had the smallest Z-average (128.9 ± 6.9 nm) with PDI (0.47), while formula 3F had the largest Z-average (4868.0 ± 250.5 nm) with PDI (1.00). The control formula had a

Discussion

Sonoprecipitation technique succeeded in preparing amorphous nanoparticles of avanafil with significant improvement in aqueous solubility. Formulas 2B, 2C, 1D, 2D, 3D, 2E and 2F had Z-average values in the nano-range which contributed to the improvement in solubility (Hecq et al., 2005). The amorphous nature of these formulas was inferred from their DSC thermograms and X-ray diffraction patterns. The improvement in solubility due to combined effects of nanonization and amorphism was also

Conclusions

PVA-stabilized particles prepared by modified NANOEDGE™ technology had smaller size than those prepared by sonoprecipitation. However, the homogenized formulas exhibited lower solubility due to partial retrieval of avanafil crystallinity on HPH. Although HPH had non-significant effect on zeta potential, it stimulated flocculation of Tween-stabilized formulas. Further studies will be required to explore this effect in a more comprehensive way.

Disclosure of interest

The authors report no declarations of interest.

Authors contribution

The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. All the authors contributed equally.

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