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Improved oral absorption of dutasteride via Soluplus®-based supersaturable self-emulsifying drug delivery system (S-SEDDS)
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IJP 14499 1–7 International Journal of Pharmaceutics xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
International Journal of Pharmaceutics journal homepage: www.elsevier.com/locate/ijpharm
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Pharmaceutical nanotechnology
Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS)
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Dong Hoon Lee a , Dong Woo Yeom a , Ye Seul Song a , Ha Ra Cho b , Yong Seok Choi b , Myung Joo Kang b, * , Young Wook Choi a, ** a b
College of Pharmacy, Chung-Ang University, 221 Heuksuk-dong, Dongjak-gu, Seoul 156-756, Republic of Korea College of Pharmacy, Dankook University, 119 Dandae-ro, Dongnam-gu, Cheonan, Chungnam 330-714, Republic of Korea
A R T I C L E I N F O
A B S T R A C T
Article history: Received 30 August 2014 Received in revised form 26 November 2014 Accepted 27 November 2014 Available online xxx
A novel supersaturable self-emulsifying drug delivery system (S-SEDDS) was formulated to improve the oral absorption of dutasteride (DTS), a 5a-reductase inhibitor that is poorly water-soluble. A supersaturable system was prepared by employing Soluplus1 (polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer) as a precipitation inhibitor with a conventional SEDDS vehicle consisted of CapryolTM 90, Cremophor1 EL and Transcutol1 HP (DTS:SEDDS vehicle: Soluplus1 = 1.0:67.6:10.0 w/v/w). In an in vitro dissolution test in a non-sink condition, the drug dissolution rate from SEDDS was rapidly increased to 72% for an initial period of 5 min, but underwent rapid drug precipitation within 2 h, decreasing the amount of drug dissolved to one-seventh of its original amount. On the other hand, S-SEDDS resulted in a slower crystallization of DTS by virtue of a precipitation inhibitor, maintaining a 3 times greater dissolution rate after 2 h compared to SEDDS. In an in vivo pharmacokinetic study in rats, the S-SEDDS formulation exhibited 3.9-fold greater area-under-curve value than that of the drug suspension and 1.3-fold greater than that of SEDDS. The maximum plasma concentration of S-SEDDS was 5.6- and 2.0-fold higher compared to drug suspension and SEDDS, respectively. The results of this study suggest that the novel supersaturable system may be a promising tool for improving the physicochemical property and oral absorption of the 5a-reductase inhibitor. ã 2014 Published by Elsevier B.V.
Keywords: Dutasteride Supersaturable self-emulsifying drug delivery system Soluplus1 Oral absorption Bioavailability
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1. Introduction Dutasteride ((5a,17b)-N-{2,5 bis(trifluoromethyl)phenyl}3oxo-4-azaandrost-1-ene-17-carboxamide, DTS) is a potent inhibitor of 5a-reductase types 1 and 2 (Roehrborn et al., 2002). It is prescribed as a once-daily oral therapy for the treatment of moderate to severe symptoms of benign prostatic hyperplasia (BPH) and to reduce the risk of acute urinary retention and surgery in patients with moderate to severe BPH (Clark et al., 2004). In spite of its attractive pharmacological effects, the absorption of DTS is quite challenged by its poor aqueous solubility, which is 0.038 ng/mL in water at 25 C (Avodart1 FDA Biopharmaceutics Review, 2001). The oral bioavailability (BA) of the marketed product (Avodart1, GlaxoSmithKline, UK), formulated with
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* Corresponding author. Tel.: +82 41 550 1446; fax: +82 41 550 7899. ** Corresponding author. Tel.: +82 2 820 5609; fax: +82 2 826 3781. E-mail addresses: [email protected] (M.J. Kang), [email protected] (Y.W. Choi).
monoglycerides and diglycerides of caprylic/capric acid, is still unsatisfactory, at about 40–60% in humans (Avodart1 Product Monograph, 2013). Several attempts to improve dissolution and/or oral absorption of the potent 5a-reductase inhibitor such as solid dispersion (Beak and Kim, 2012; Kim et al., 2013), polymeric nanoparticle (Park et al., 2013), cyclodextrin complex (Kim et al., 2013), and a self-emulsifying drug delivery system (SEDDS) (Choo et al., 2013) have been reported. Among them, SEDDS is an isotropic mixture of oil, surfactants, and/or co-solvent, which form fine oil-in-water (o/w) droplets upon dilution with aqueous medium in the gastrointestinal (GI) track (Constantinides,1995). In an earlier study, Choo et al. (2013) developed the SEDDS formulation of DTS consisting of CapryolTM 90 (propylene glycol monocaprylate), Cremophor1 EL (polyoxyl 35 hydrogenated castor oil), and Transcutol1 HP (purified diethylene glycol monoethyl ether), which exhibited a significantly higher plasma concentration profile of DTS compared to that of drug suspension (Choo et al., 2013). However, after long-term oral administration, the high concentration of surfactant (60 w/v%) in the formulation could lead to severe GI irritation.
http://dx.doi.org/10.1016/j.ijpharm.2014.11.060 0378-5173/ ã 2014 Published by Elsevier B.V.
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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In an attempt to reduce the side effects of surfactants and maximize the intestinal absorption of poorly soluble drugs, a supersaturable self-emulsifying drug delivery system (S-SEDDS) was proposed. The S-SEDDS formulations generally contain a lower concentration of surfactant and a polymeric precipitation inhibitor to yield and stabilize the drug in a temporary supersaturated state (Gao et al., 2003). The precipitation inhibitors thermodynamically and/or kinetically protract the supersaturated state of active molecules in aqueous medium by reducing the rate of drug nucleation and crystal growth through physical interactions with drug compounds or by changing the medium properties (Chauhan et al., 2013; Loftsson et al., 1996; Usui et al., 1997). Several previous studies with poorly water soluble drugs, such as paclitaxel (Gao et al., 2003), PNU-91325 (Gao et al., 2004), and AMG 517 (Gao et al., 2009), clearly demonstrated that the supersaturable systems provide higher oral absorption and fewer side effects as compared to that of the conventional SEDDS. Also, we previously reported that Soluplus1 (polyvinylcaprolactam–polyvinyl acetate–polyethylene glycol graft copolymer) significantly amplified the spring and parachute effect in S-SEDDS formulation of celecoxib (Song et al., 2013). Soluplus1 is an amphiphilic polymer designed to produce amorphous dispersions by hot melt extrusion due to its low glass transition temperature of 72 C (Hardung et al., 2010). It can enhance the solubility and the dissolution, and even retard drug precipitation of poorly watersoluble drug in aqueous medium (Zhang et al., 2013; Kim, 2013a,b). The present study was performed to construct a novel S-SEDDS to improve oral absorption of DTS. A supersaturable system of DTS was formulated by screening various kinds of hydrophilic and/or amphiphilic polymers including Soluplus1, as a precipitation inhibitor. Each formulation was evaluated for their physicochemical characteristic such as droplet size, homogeneity, drug content, and in vitro dissolution profile under non-sink condition to assess the supersaturation state of the 5a-reductase inhibitor. We subsequently compared the drug absorption of optimized S-SEDDS with drug suspension and conventional SEDDS in rats using a validated LC–MS/MS assay.
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2. Materials and methods
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2.1. Materials
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DTS was obtained from MSN Laboratories Limited (India, purity over 99.0 w/w%). Capryol 90TM and Transcutol1 HP were kindly
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supplied by Gattefosse (Saint Priest, France). Cremophor1 EL, Kollicoat1 MAE 30DP (methacrylic acid ethylacrylate copolymer), Kollidon1 90F (polyvinylpyrrolidone), and Soluplus1 were kindly supplied by BASF (Ludwigshafen, Germany). Hypromellose 2910 (Hydroxypropylmethyl cellulose, HPMC) was supplied by Shin-Etsu Chemical Co. (Tokyo, Japan). Finasteride (purity over 98 w/w%) was purchased from Sigma Chemical Co. (St. Louis, MO, USA). Acetonitrile and methanol of HPLC grade were purchased from J.T. Baker (Phillipsburg, NJ, USA). All other chemicals used were of analytical grade.
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2.2. Animals
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Sprague-Dawley rats (male; 200–250 g; 7–9 weeks) were purchased from Orient Bio (Kyungki-do, Korea). They were kept in specific pathogen-free conditions with food and water freely available. All animal experiments were performed in accordance with the NIH “Principles of laboratory animal care” guidelines (NIH publication No. 85-23, revised 1996) and were approved by the Institutional Animal Care and Use Committee of Chung-Ang University in Seoul, Korea.
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2.3. Preparation of SEDDS and S-SEDDS formulations
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The DTS-loaded SEDDS was prepared by dissolving the drug (50 mg) in the mixture of Capryol 90TM/Cremophor1 EL/Transcutol1 HP. The components were mixed by vortexing at room temperature until the DTS was completely dissolved. The compositions of each formulation are detailed in Table 1. The S-SEDDS formulation was acquired by the further addition of precipitation inhibitors to SEDDS F3 formula. Various polymeric excipients (Kollidon1 90F, HPMC, Kollicoat1 MAE 30DP, and Soluplus1) were added into the SEDDS solution, and then, the mixture was vortexed vigorously using magnetic stirrer at room temperature to obtain a uniform suspension. The S-SEDDS suspension was stored at room temperature until used.
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2.4. Physical characterization of SEDDS and S-SEDDS formulations
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2.4.1. Droplet size Each formulation was diluted with an appropriate volume of simulated gastric juice and particle size distribution. The droplet size was assayed using a dynamic light-scattering particle size analyzer (Zetasizer Nano-ZS, Malvern Instrument, Worcestershire,
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Table 1 Compositions and physicochemical characteristics of DTS-loaded SEDDS and S-SEDDS formulations.
Compositions DTS (mg) CapryolTM 90 (mL) Cremophor1 EL (mL) Transcutol1 HP (mL) Soluplus1 (mg) Kollidon1 90F (mg) HPMC 2910 (mg) Kollicoat1 MAE (mg)
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
F13
0.5 42.3 24.7 34.6 – – – –
0.5 21.2 12.4 17.3 – – – –
0.5 14.1 8.2 11.5 – – – –
0.5 14.1 8.2 11.5 5 – – –
0.5 14.1 8.2 11.5 – 5 – –
0.5 14.1 8.2 11.5 – – 5 –
0.5 14.1 8.2 11.5 – – – 5
0.5 14.1 8.2 11.5 1 – – –
0.5 14.1 8.2 11.5 2.5 – – –
0.5 14.1 8.2 11.5 10 – – –
0.5 14.1 8.2 11.5 15 – – –
0.5 14.1 8.2 11.5 5 – 5 –
0.5 14.1 8.2 11.5 5 – – 5
137.3 0.29 98.8 N.D.
119.9 0.07 97.6 25.9
99.7 0.26 105.1 65.8
96.1 0.18 105.7 34.5
96.9 0.13 101.5 20.1
95.9 0.18 100.4 N.D.
98.3 0.17 99.4 N.D.
108.3 0.25 101.0 N.D.
112.2 0.19 105.6 N.D.
95.9 0.14 103.0 N.D.
91.3 0.17 103.8 N.D.
97.4 0.16 102.9 N.D.
Physicochemical characteristics Size (nm)a 128.0 PDIa 0.26 Drug contents (%)a 100.3 DT50 (min)a,b N.D.c a
Data are expressed as mean values (n = 3). Their standard deviations were less than 10% of the mean values. Indicates the time to maintain the dissolution rate over 50%. DT50 is calculated by subtracting the initial time point from the last time point where over 50% of dissolution rate is reached in Fig. 2. c Not determined. b
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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UK) with a 50 mV laser at a scattering angle of 90 . All measurements were carried out in triplicate under ambient conditions. 2.4.2. Drug content Each formulation containing 0.5 mg of DTS was dissolved in 20 mL of acetonitrile and sonicated for 10 min. Samples were then centrifuged at 12,000 rpm for 10 min and the supernatants were analyzed by HPLC analysis. The quantitative determination of DTS was performed by HPLC using acetonitrile:water (70:30) as a mobile phase at a flow rate of 1 mL/min. The HPLC system consisted of a pump (L-2130), a UV detector (L-2400), a data station (LaChrom Elite, Hitachi, Japan), and a 15 cm C18 column (Kromasil, AkzoNobel, Sweden). The column eluant was monitored at 210 nm, and the DTS peak was separated at a retention time of 6.5 min. The calibration curve was linear in the DTS concentration range of 0.1–10 mg/mL (y = 210673x 7208.2, r2 = 0.999).
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2.5. In vitro dissolution test
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In vitro dissolution tests were performed according to the USP apparatus II (paddle) method with a Vision1 classic 6TM dissolution testing station and a Vision1 heater (Hanson, USA). Each formulation containing 0.5 mg of DTS was placed in the dissolution vessel containing 250 mL of simulated gastric juice maintained at 37.0 0.5 C and stirred at 50 rpm. The drug release experiment with F4 was further carried out in distilled water and intestinal fluid (pH 6.8). Then, 5 mL of samples were taken at various time intervals and replaced with fresh and pre-warmed dissolution medium. The samples were centrifuged at 12,000 rpm for 10 min and the supernatants diluted with methanol for HPLC analysis.
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2.6. In vivo oral absorption study
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2.6.1. Experimental Male Sprague-Dawley rats weighing approximately 250 g were fasted for 16 h prior to the experiment and divided into 3 groups. The rats in the first group were orally administered 1 mL of a 0.2% methylcellulose aqueous suspension containing powdered DTS, the rats in the second group received the conventional SEDDS formulation (F3), and the rats in the third group received optimized S-SEDDS (F4) at a dose of 2 mg/kg (Choo et al., 2013). Blood samples (0.3 mL) were then collected from the retro-orbital plexus with heparinized tubes at predetermined time intervals and centrifuged at 12,000 rpm for 10 min. Plasma samples were stored at 20 C until analysis by LC–MS/MS. 2.6.2. LC–MS/MS determination of DTS in rat plasma To isolate components in a sample, liquid-liquid extraction (LLE) was carried out prior to its analysis (Choo et al., 2013; Kang et al., 2014). Plasma (100 mL) was thawed and mixed with 20 mL of a finasteride (an internal standard, IS) solution (200 ng/ mL), 100 mL of an aqueous sodium hydroxide solution (39.997 mg/mL), and 600 mL of a LLE reagent (tert-butyl ether:methylene chloride, 70:30, v/v) for 10 min by using a rotator. The mixture was centrifuged at 12,000 rpm for 5 min and the resulting organic phase was dried under a gentle stream of nitrogen gas. Finally, the residue was reconstituted with 200 mL of a 50% aqueous methanol solution for introduction into the LC–MS/MS system. LC separation was performed by a LC-20 Prominence system (Shimadzu, Japan). The temperature of the autosampler was kept at 4 C and components in 30 mL of each reconstituted sample were separated on a Hydrosphere C18 (5 mm, 250 2.0 mm, YMC, Japan) column at 45 C. An isocratic mobile phase condition (0.1% aqueous formic acid solution:
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acetonitrile, 30:70%, v/v) was used at a flow rate of 0.250 mL/min and the analysis time per sample was 15 min. The components eluted from the column were delivered into an API 2000 triple quadruple mass spectrometer (AB/SCIEX, USA) through a Turbospray ion source (AB/SCIEX) for multiple reaction monitoring (MRM) assays of DTS (528.9/460.9/49, the m/z value of the precursor ion/the m/z value of the product ion/the collision energy, V) and IS (373.3/305.2/49, the m/z value of the precursor ion/the m/z value of the product ion/the collision energy, V) in positive ion mode. Additional mass spectrometer conditions are as follows: spray voltage at 5300 V; spray temperature at 400 C; nebulizer gas (gas 1) at 35; heater gas (gas 2) at 35; declustering potential at 70 V. The good linearity (r2 = 0.993) was observed at a DTS concentration range between 5 and 400 ng/mL.
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2.6.3. Pharmacokinetic analysis Pharmacokinetic analysis was performed using a BA Calc 2007 pharmacokinetic analysis program (Korea Food & Drug Administration, Korea). Area under the curve (AUC) from 0 to 24 h was calculated using the program’s linear trapezoidal rule. Maximum plasma concentration (Cmax) and the time needed to reach the maximum plasma concentration (Tmax) were determined directly from the concentration–time data. The relative BA was calculated as a percentage of the AUC of formulations to that of the drug suspension.
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2.7. Statistical analysis
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All data are expressed as the mean SD. Statistical significance was determined by Student’s t-test with a threshold of P < 0.05, unless otherwise indicated.
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3. Results
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3.1. Physicochemical characteristics of SEDDS and S-SEDDS formulations
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The physicochemical characteristics of DTS-loaded SEDDS and S-SEDDS formulations were evaluated by droplet size, homogeneity, and drug content (Table 1). The particle size of the SEDDS dispersions (F1–F3) was approximately 130 nm, with no significant differences between the SEDDS formulas. On the other hand, mean particle size of S-SEDDS (F4–F13) ranged from 90 to 110 nm, which is slightly smaller than SEDDS. The low polydispersity index (PDI) of less than 0.3 in all formulations indicates a narrow and homogeneous size distribution. The drug content in each formulation was almost equal (97.6–105.7%) with small standard deviations, indicating that the drug was uniformly distributed in the liquid formulations.
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3.2. In vitro dissolution study
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3.2.1. Effect of drug loading amount on drug dissolution from SEDDS formulations The dissolution patterns of DTS from the SEDDS formulations at different ratios of the drug to SEDDS vehicle (F1–F3) in simulated gastric fluid are shown in Fig. 1. Upon contact with the gastric fluid, F1–F3 formulations rapidly formed a fine emulsion with a bluish reflection, and over 70% the drug was released after 5 min. However, as time elapsed, the drug release from the F2 and F3 rapidly decreased to 24% and 11% at 2 h, while F1 maintained its dissolution state. In case of F3, the dissolved amount was reduced drastically to reach the equilibrium level of about 11% at 2 h. In the next study, precipitation inhibitors were incorporated in SEDDS F3 formulation to stabilize the supersaturated state of DTS and establish the S-SEDDS formulation.
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Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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DTS dissolved (%)
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Time (min) Fig. 1. Dissolution profiles of DTS from SEDDS formulations in simulated gastric juice prepared with different ratios of drug to SEDDS vehicle; 1:200 (F1, &); 1:100 (F2, 4), and 1:67 (F3, ^). Data represent mean SD (n = 3).
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3.2.2. Effect of various precipitation inhibitors on drug dissolution from S-SEDDS formulations The effects of hydrophilic and/or amphiphilic polymers for supersaturating F3 formulation were evaluated as depicted in Fig. 2. The amount of a precipitation inhibitor in each formulation was identically set to 5 mg, which is approximately 15% to SEDDS vehicle. In the case of the hydrophilic polymers Kollidon1 90F (F5), HPMC (F6), and Kollicoat1 MAE (F7), there was no increase in the dissolution level, even though F7 displayed a positive effect on decelerating precipitation. In contrast, the addition of Soluplus1 (F4), an amphiphilic polymer, to SEDDS formulation significantly increased the drug dissolution rate at 2 h (Fig. 2). Upon contact with the gastric fluid, almost 80% of the drug was released within 5 min and the DTS concentration in simulated gastric fluid was gradually decreased but maintained over 30% at 2 h, which drastically reduced the amount dissolved to about 10% in F3. For further quantitative observation, we established the term ‘DT50’ that indicates ‘the time to maintain the dissolution rate over 50% (Table 1). DT50 is calculated by subtracting the initial time point from the last time point where over 50% of dissolution rate is
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3.2.3. Effect of Soluplus1 amount on drug dissolution from S-SEDDS formulations In order to evaluate the influence the amount of Soluplus1 upon the duration of the supersaturated state, a series of S-SEDDS formulations containing the precipitation inhibitor (3, 7.5, 15, 30, and 45% to SEDDS vehicle) were evaluated (F4 and F8–11). As plotted in Fig. 3, as the proportion of the precipitation inhibitor increased, the drug concentration in aqueous medium at 2 h increased and plateaued for the F4 formula containing 15% of Soluplus1, dissolving over 30% of the drug dissolved at 2 h. The DTS concentration–time profiles suggest that the presence of 15%, 30% and 45% of Soluplus1 (F4, F10 and F11) appears sufficiently effective, yielding similar drug concentration–time profiles. The dissolution profile of DTS from the drug powder and physical mixture of drug and Soluplus1 in a weight ratio of 1:10 are further investigated (Fig. 3). In gastric juice, physical mixtures of drug with the polymer showed dissolution profiles similar to those of the intact drug; only about 10% of the drug was release for 2 h. The S-SEDDS F4 showed a consistently higher drug 7concentration–time profile as compared to the SEDDS F3 and physical mixtures, indicating a synergistic effect on the dissolution of the drug by virtue of the precipitation inhibitor with SEDDS.
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3.2.4. Effect of combination of Soluplus1 with hydrophilic polymers on drug dissolution from S-SEDDS formulations In order to produce the extended supersaturated state, some hydrophilic additives were combined with Soluplus1-based F4 formula (Fig. 4). However, both HPMC-added (F12) and Kollicoat1 MAE-added (F13) compositions did not produce a complementary precipitation-inhibiting effect with Soluplus1. Conversely, the other hydrophilic additives showed an antagonistic effect compared to Soluplus1 alone (F4), resulting in decreased supersaturation or an increase in drug precipitation from the
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100
100
80
DTS dissolved (%)
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DTS dissolved (%)
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reached in Fig. 2. Initial and last time points were calculated by drawing a horizontal line at 50% on the Y axis and dropping a vertical line form the point of intersection with the dissolution curve to the X axis in Fig. 2. DT50 value of F4 was the highest at about 65.9 min, among the various formulations. It was 2.6-, 2.0-, and 3.3-fold higher compared to the DT50 values for F3, F5, and F6, respectively (Table 1).
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Time (min) Fig. 2. Dissolution profiles of DTS from S-SEDDS formulations in simulated gastric juice prepared with different precipitation inhibitors; Soluplus1-added (F4, ^); Kollidon1 90F-added (F5, *), HPMC-added (F6, ~), and Kollicoat1 MAE-added (F7, &), and SEDDS (F3, ^). The amount of precipitation inhibitor in each formulation was identical to 5 mg, which is approximately 15% to SEDDS vehicle. Data are expressed as mean SD (n = 3).
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Time (min) Fig. 3. Dissolution profiles of DTS from S-SEDDS formulations according to the amount of Soluplus1 added; 1 mg (F8, ); 2.5 mg (F9, 4); 5 mg (F4, ^); 10 mg (F10, *), and 15 mg (F11, ~), physical mixture of drug with 5 mg of Soluplus1 (^), and drug powder () in simulated gastric juice. Data are expressed as mean SD (n = 3).
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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Plasma conc. (ng/mL)
DTS dissolved (%)
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250 200 150 100
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Time (h)
Time (min) Fig. 4. Dissolution profiles of DTS from S-SEDDS formulations after combination of hydrophilic polymers with Soluplus1; Soluplus1 alone-added (F4, ^); Soluplus1 and HPMC-added (F12, ~), and Soluplus1 and Kollicoat1 MAE-add (F13, &). Data are expressed as mean SD (n = 3).
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supersaturated state. From these findings, the optimized formula for the drug, SEDDS vehicle and the Soluplus1 at a ratio of 1:67.6:10 (F4) was finally established to as the supersaturable oral dosage form of DTS.
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3.2.5. Effect of dissolution medium on drug dissolution from the optimized S-SEDDS formulation Dissolution profiles of the optimized S-SEDDS (F4) were further evaluated in different dissolution media (Fig. 5). The release rate of the 5a-reductase inhibitor was pH-independent, yielding that over 30% of DTS was supersaturated at 2 h in media at pH 1.2, pH 6.8 and distilled water.
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3.3. In vivo absorption study in rats
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Fig. 6 shows the mean plasma concentration profiles of DTS in rats upon oral dosing of suspension, SEDDS (F3) or S-SEDDS (F4) formulations at a drug dose of 2 mg/kg. Mean plasma DTS levels of both SEDDS and S-SEDDS formulations were significantly higher than that of the drug suspension, which slowly increases until 12 h (Fig. 6). In both SEDDS and S-SEDDS formulations, however, the
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DTS dissolved (%)
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Fig. 6. Plasma concentration-time profiles of DTS in rats after oral administrations of drug suspension (), SEDDS (^), and S-SEDDS (^) formulations at a dose of 2 mg/kg. Data are expressed as mean SD (n = 3).
5a-reductase inhibitor is rapidly absorbed and shows a sharp increase in plasma DTS levels within 3 h. In particular, S-SEDDS F4 showed higher plasma levels than SEDDS for 24 h. The pharmacokinetic parameters, including Tmax, Cmax and AUC(0– 24 h), are listed in Table 2. As expected, S-SEDDS (F4) was superior to the other formulations in all parameters; the parameters were significantly different at P < 0.05. The AUC(0–24 h) value of S-SEDDS formula was 3.9-fold greater than that of the drug suspension and 1.3-fold greater than that of SEDDS. The Cmax value of S-SEDDS was about 5.6- to 2.0-fold higher compared to drug suspension and SEDDS, respectively. From these findings, we concluded that the oral BA of DTS was greatly improved by the supersaturable formulation.
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4. Discussion
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SEDDSs are widely used to enhance the oral absorption of poorly soluble drugs, including DTS. In our study, the SEDDS formulation consisted of Capryol 90TM, Cremophor1 EL, and Transcutol1 HP that was easily dispersed in aqueous dissolution medium to make fine o/w microemulsion spontaneously and exhibited high initial dissolution rate. However, the dissolution was time-dependent, in which the amount dissolved rapidly decreasing due to the loss of its solubilization capacity with lower amount of SEDDS vehicle (F3). It was reported that the solvent capacity of the lipid formulation falls logarithmically as the mixture is diluted into an aqueous medium (Pouton, 2000). On the other hand, the large amount of surfactant present in the SEDDS formulation could lead to GI side effects after daily intake. Especially Cremophor1 EL, which was reported to cause severe side effects such as hypersensitivity, nephrotoxicity and neurotoxicity (Danhier et al., 2009). Therefore, a novel supersaturable
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40 Table 2 Pharmacokinetic parameters of DTS from the drug suspension, SEDDS, and S-SEDDS formulations after a single 2 mg/kg oral administration in rats.
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Time (min) Fig. 5. Dissolution profiles of DTS from the optimized S-SEDDS (F4) formulation in the different pH media; gastric juice (pH 1.2, ^), intestinal fluid (pH 6.8, ), and distilled water (&). Data are expressed as mean SD (n = 3).
Parameters
Drug suspension SEDDS (F3)
S-SEDDS (F4)
AUC(0–24 h) (mg h/mL)a Cmax (mg/mL)a Tmax (h)a Relative BA (%)b
580.99 107.99 35.98 8.02 9.6 2.19 –
2282.50 460.98 183.30 38.81 1.92 0.67 393
1796.16 359.87 108.55 20.64 3.13 2.17 309
a
Values represent mean SD (n = 5–10). Calculated as percentage of the mean AUC(0–24 h) of each group to that of drug suspension. b
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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system containing a reduced amount of surfactant and a small amount of precipitation inhibitor was designed to generate and maintain a supersaturated state to improve oral absorption of DTS in GI track. In order to evaluate the drug concentration sustained in the supersaturated state and the degree of supersaturated state as a function of time, an in vitro dissolution test was designed as a non-sink condition. The total volume of the medium chosen here was 250 mL, based on physiological considerations of the total residual volume of stomach fluid. On the other hand, the solubility of DTS in aqueous solution was reported as an approximately 0.038 ng/mL in water at 25 C (Avodart1 FDA Biopharmaceutics Review, 2001). Thus, the experimental condition does not guarantee the sink conditions for DTS. In designing the supersaturable formulations, it is essential to include a precipitation inhibitor in the SEDDS vehicle to maintain drugs in supersaturated state in aqueous medium (Xu and Dai, 2013). Precipitation inhibitors inhibit drug nucleation and/or crystal growth by steric stabilization, surface stabilization by adsorption onto colloidal surfaces, and/or specific interactions with a drug (e.g., by hydrogen bonding or hydrophobic interactions) (Wen et al., 2005; Kojima et al., 2012; Song et al., 2013). For this purpose, in the present study, hydrophilic polymers such as HPMC, Kollidon1 90F, and Kollicoat1 MAE were screened as potential candidates for precipitation inhibitor. However, these polymers were not effective for stabilizing the supersaturated state of DTS with SEDDS formulation (Fig. 2). It is assumed that these hydrophilic polymers were not absorbed efficiently on the surface of the oily droplet or free drug molecules, but rather molecularly dissolved and freely diffused into the aqueous medium. Alternatively, Soluplus1, an amphiphilic polymer, significantly inhibited precipitation of DTS, preserving it in the supersaturated state (Fig. 2). The F4 formula showed a consistently higher apparent drug concentration as compared to F3, providing 2.6-fold higher DT50 value compared to F3. It was interpreted by our groups that a hydrophobic polycaprolactam moiety of Soluplus1 intercalates to hydrophobic tail packing of dispersed phase and thus, the interface becomes denser. Simultaneously, polyethylene glycol groups located on the surface of colloidal dispersion form a hydrophilic barrier, hindering aggregation and/or destruction of emulsion droplet in gastric fluid (Song et al., 2013). There was a critical amount of Soluplus1 for stabilizing the supersaturated state of DTS. In the S-SEDDS systems with Soluplus1, 5 mg of the precipitation inhibitor effectively retards DTS precipitation and sustains a higher apparent concentration in a non-sink condition (Fig. 3). This result is consistent with an earlier report that the S-SEDDS formulation containing 40 mg of Soluplus1 as a precipitation inhibitor exhibited the greatest dissolution of celecoxib in a non-sink condition with a medium volume of 1000 mL (Song et al., 2013). The concentration of Soluplus1 in the aqueous medium was quite comparable in two studies between 20 and 40 mg/mL, which is analogous with the critical micelle concentration of Soluplus1 (46.3 mg/L) at pH 1.2. It suggests that the optimal level of Soluplus1 in S-SEDDS is closely associated with the micelle formation of the amphiphilic polymer. Three types of different formulations including DTS suspension, SEDDS F3, and S-SEDDS F4, were administered orally to rats and their pharmacokinetic parameters were compared (Table 2). The relative BA of SEDDS and S-SEDDS was about 2.9- and 3.9-fold higher compared to drug suspension which was used as a reference. It indicates that self-emulsifying formulations spontaneously formed fine oil droplets and present DTS in a dissolved form, avoiding the dissolution step. Especially, S-SEDDS could remarkably improve the peroral absorption of DTS, providing even 30% greater BA than that of SEDDS. Supersaturable formulation is
able to induce a supersaturated drug concentration that absorbed in the desired time frame when exposed to the GI tract in vivo. In this study, both the in vitro dissolution test and the in vivo pharmacokinetic study clearly indicate that the S-SEDDS formulation is remarkably effective in inhibiting drug precipitation in the GI track and thus improving the rate and extent of DTS absorption. From these results, we conclude that S-SMEDDS is superior to SEDDS formulations for DTS oral delivery.
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5. Conclusion
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Novel DTS-loaded S-SEDDS formulation was successfully prepared by incorporating Soluplus1 as a precipitation inhibitor to SEDDS composed of Capryol 90TM as an oil, Cremophor1 EL as a surfactant, and Transcutol1 HP as a cosurfactant. The in vitro dissolution tests in a non-sink condition revealed that a small amount of Soluplus1 (15% w/v to SEDDS vehicle) effectively slowed down drug precipitation and played a critical role in maintaining a supersaturated state of DTS. Drug dissolution from S-SEDDS was pH-independent. The in vivo pharmacokinetic study in rats demonstrated that S-SEDDS formulation significantly improved oral absorption of DTS, providing 3.9-fold greater BA compared to drug suspension. This study demonstrates the potential use of the S-SEDDS formulation in the development of poorly water-soluble oral drugs.
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References
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Avodart1 FDA Biopharmaceutics Review, 2001. http://www.accessdata.fda.gov/ drugsatfda_docs/nda/2001/21319_Duagen_biopharmr_P1.pdf. Beak, I.H., Kim, M.S., 2012. Improved supersaturation and oral absorption of dutasteride by amorphous solid dispersions. Chem. Pharm. Bull. 60, 1468–1473. Chauhan, H., Hui-Gu, C., Atef, E., 2013. Correlating the behavior of polymers in solution as precipitation inhibitor to its amorphous stabilization ability in solid dispersions. J. Pharm. Sci. 102, 1924–1935. Choo, G.H., Park, S.J., Hwang, S.J., Kim, M.S., 2013. Formulation and in vivo evaluation of a self-microemulsifying drug delivery system of dutasteride. Drug Res. 63, 203–209. Clark, R.V., Hermann, D.J., Cunningham, G.R., Wilson, T.H., Morrill, B.B., Hobbs, S., 2004. Marked suppression of dihydrotestosterone in men with benign prostatic hyperplasia by dutasteride, a dual 5a-reductase inhibitor. J. Clin. Endocrinol. Metab. 89, 2179–2184. Constantinides, P.P., 1995. Lipid microemulsions for improving drug dissolution and oral absorption: physical and biopharmaceutical aspects. Pharm. Res. 12, 1561–1572. Danhier, F., Lecouturier, N., Vroman, B., Jérôme, C., Marchand-Brynaert, J., Feron, O., Préat, V., 2009. Paclitaxel-loaded PEGylated PLGA-based nanoparticles: in vitro and in vivo evaluation. J. Control. Release 133, 11–17. Gao, P., Rush, B.D., Pfund, W.P., Huang, T., Bauer, J.M., Morozowich, W., Kuo, M.S., Hageman, M.J., 2003. Development of a supersaturable SEDDS (S-SEDDS) formulation of paclitaxel with improved oral bioavailability. J. Pharm. Sci. 92, 2386–2398. Gao, P., Guyton, M.E., Huang, T., Bauer, J.M., Stefanski, K.J., Lu, Q., 2004. Enhanced oral bioavailability of a poorly water soluble drug PNU-91325 by supersaturatable formulations. Drug Dev. Ind. Pharm. 30, 221–229. Gao, P., Akrami, A., Alvarez, F., Hu, J., Li, L., Ma, C., Surapaneni, S., 2009. Characterization and optimization of AMG 517 supersaturable self-emulsifying drug delivery system (S-SEDDS) for improved oral absorption. J. Pharm. Sci. 98, 516–528. GlaxoSmithKline, Avodart1 Product Monograph, 2013. http://www.gsk.ca/english/ docs-pdf/product-monographs/Avodart.pdf, cited 1 December, 2012. Hardung, H., Djuric, D., Ali, S., 2010. Combining HME and solubilization: Soluplus1 – the solid solution. Drug Deliv. Technol. 10, 20–27. Kang, M.J., Cho, H.R., Lee, D.H., Yeom, D.W., Choi, Y.W., Choi, Y.S., 2014. A method to monitor dutasteride in rat plasma using liquid–liquid extraction and multiple reaction monitoring: comparisons and validation. MSL 5, 79–83. Kim, M.S., 2013a. Influence of hydrophilic additives on the supersaturation and bioavailability of dutasteride-loaded hydroxypropyl-b-cyclodextrin nanostructures. Int. J. Nanomed. 8, 2029–2039. Kim, M.S., 2013b. Soluplus-coated colloidal silica nanomatrix system for enhanced supersaturation and oral absorption of poorly water-soluble drugs. Artif. Cells Nanomed. Biotechnol. 41, 363–367. Kim, N.A., Choi, D.H., Lim, J.Y., Kim, K.H., Lim, D.G., Lee, E., Park, E.-S., Jeong, S.H., 2013. Investigation of polymeric excipients for dutasteride solid dispersion and its physicochemical characterization. Arch. Pharm. Res. 37, 1–11.
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Kojima, T., Higashi, K., Suzuki, T., Tomono, K., Moribe, K., Yamamoto, K., 2012. Stabilization of a supersaturated solution of mefenamic acid from a solid dispersion with EUDRAGIT1 EPO. Pharm. Res. 29, 2777–2791. Loftsson, T., Fririksdóttir, H., Gumundsdóttir, T.K., 1996. The effect of water-soluble polymers on aqueous solubility of drugs. Int. J. Pharm. 127, 293–296. Park, S.J., Choo, G.H., Hwang, S.J., Kim, M.S., 2013. Quality by design: screening of critical variables and formulation optimization of Eudragit E nanoparticles containing dutasteride. Arch. Pharm. Res. 36, 1–9. Pouton, C.W., 2000. Lipid formulations for oral administration of drugs: nonemulsifying, self-emulsifying and ‘self-microemusifying’ drug delivery systems. Eur. J. Pharm. Sci. 11, 93–98. Roehrborn, C.G., Boyle, P., Nickel, J.C., Hoefner, K., Andriole, G., 2002. Efficacy and safety of a dual inhibitor of 5-alpha-reductase types 1 and 2 (dutasteride) in men with benign prostatic hyperplasia. Urology 60, 434–441.
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Song, W.H., Park, J.H., Yeom, D.W., Ahn, B.K., Lee, K.M., Lee, S.G., Woo, H.S., Choi, Y.W., 2013. Enhanced dissolution of celecoxib by supersaturating self-emulsifying drug delivery system (S-SEDDS) formulation. Arch. Pharm. Res. 36, 69–78. Usui, F., Maeda, K., Kusai, A., Nishimura, K., Keiji, Y., 1997. Inhibitory effects of water soluble polymers on precipitation of RS-8359. Int. J. Pharm. 154, 59–66. Xu, S., Dai, W.G., 2013. Drug precipitation inhibitors in supersaturable formulations. Int. J. Pharm. 453, 36–43. Wen, H., Morris, K.R., Park, K., 2005. Hydrogen bonding interactions between adsorbed polymer molecules and crystal surface of acetaminophen. J. Colloid Interface Sci. 290, 325–335. Zhang, K., Yu, H., Luo, Q., Yang, S., Lin, X., Zhang, Y., Tian, B., Tang, X., 2013. Increased dissolution and oral absorption of itraconazole/Soluplus extrudate compared with itraconazole nanosuspension. Eur. J. Pharm. Biopharm. 85, 1285–1292.
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Contents lists available at ScienceDirect
International Journal of Pharmaceutics journal homepage: www.elsevier.com/locate/ijpharm
1
Pharmaceutical nanotechnology
Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS)
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Dong Hoon Lee a , Dong Woo Yeom a , Ye Seul Song a , Ha Ra Cho b , Yong Seok Choi b , Myung Joo Kang b, * , Young Wook Choi a, ** a b
College of Pharmacy, Chung-Ang University, 221 Heuksuk-dong, Dongjak-gu, Seoul 156-756, Republic of Korea College of Pharmacy, Dankook University, 119 Dandae-ro, Dongnam-gu, Cheonan, Chungnam 330-714, Republic of Korea
A R T I C L E I N F O
A B S T R A C T
Article history: Received 30 August 2014 Received in revised form 26 November 2014 Accepted 27 November 2014 Available online xxx
A novel supersaturable self-emulsifying drug delivery system (S-SEDDS) was formulated to improve the oral absorption of dutasteride (DTS), a 5a-reductase inhibitor that is poorly water-soluble. A supersaturable system was prepared by employing Soluplus1 (polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer) as a precipitation inhibitor with a conventional SEDDS vehicle consisted of CapryolTM 90, Cremophor1 EL and Transcutol1 HP (DTS:SEDDS vehicle: Soluplus1 = 1.0:67.6:10.0 w/v/w). In an in vitro dissolution test in a non-sink condition, the drug dissolution rate from SEDDS was rapidly increased to 72% for an initial period of 5 min, but underwent rapid drug precipitation within 2 h, decreasing the amount of drug dissolved to one-seventh of its original amount. On the other hand, S-SEDDS resulted in a slower crystallization of DTS by virtue of a precipitation inhibitor, maintaining a 3 times greater dissolution rate after 2 h compared to SEDDS. In an in vivo pharmacokinetic study in rats, the S-SEDDS formulation exhibited 3.9-fold greater area-under-curve value than that of the drug suspension and 1.3-fold greater than that of SEDDS. The maximum plasma concentration of S-SEDDS was 5.6- and 2.0-fold higher compared to drug suspension and SEDDS, respectively. The results of this study suggest that the novel supersaturable system may be a promising tool for improving the physicochemical property and oral absorption of the 5a-reductase inhibitor. ã 2014 Published by Elsevier B.V.
Keywords: Dutasteride Supersaturable self-emulsifying drug delivery system Soluplus1 Oral absorption Bioavailability
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1. Introduction Dutasteride ((5a,17b)-N-{2,5 bis(trifluoromethyl)phenyl}3oxo-4-azaandrost-1-ene-17-carboxamide, DTS) is a potent inhibitor of 5a-reductase types 1 and 2 (Roehrborn et al., 2002). It is prescribed as a once-daily oral therapy for the treatment of moderate to severe symptoms of benign prostatic hyperplasia (BPH) and to reduce the risk of acute urinary retention and surgery in patients with moderate to severe BPH (Clark et al., 2004). In spite of its attractive pharmacological effects, the absorption of DTS is quite challenged by its poor aqueous solubility, which is 0.038 ng/mL in water at 25 C (Avodart1 FDA Biopharmaceutics Review, 2001). The oral bioavailability (BA) of the marketed product (Avodart1, GlaxoSmithKline, UK), formulated with
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* Corresponding author. Tel.: +82 41 550 1446; fax: +82 41 550 7899. ** Corresponding author. Tel.: +82 2 820 5609; fax: +82 2 826 3781. E-mail addresses: [email protected] (M.J. Kang), [email protected] (Y.W. Choi).
monoglycerides and diglycerides of caprylic/capric acid, is still unsatisfactory, at about 40–60% in humans (Avodart1 Product Monograph, 2013). Several attempts to improve dissolution and/or oral absorption of the potent 5a-reductase inhibitor such as solid dispersion (Beak and Kim, 2012; Kim et al., 2013), polymeric nanoparticle (Park et al., 2013), cyclodextrin complex (Kim et al., 2013), and a self-emulsifying drug delivery system (SEDDS) (Choo et al., 2013) have been reported. Among them, SEDDS is an isotropic mixture of oil, surfactants, and/or co-solvent, which form fine oil-in-water (o/w) droplets upon dilution with aqueous medium in the gastrointestinal (GI) track (Constantinides,1995). In an earlier study, Choo et al. (2013) developed the SEDDS formulation of DTS consisting of CapryolTM 90 (propylene glycol monocaprylate), Cremophor1 EL (polyoxyl 35 hydrogenated castor oil), and Transcutol1 HP (purified diethylene glycol monoethyl ether), which exhibited a significantly higher plasma concentration profile of DTS compared to that of drug suspension (Choo et al., 2013). However, after long-term oral administration, the high concentration of surfactant (60 w/v%) in the formulation could lead to severe GI irritation.
http://dx.doi.org/10.1016/j.ijpharm.2014.11.060 0378-5173/ ã 2014 Published by Elsevier B.V.
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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In an attempt to reduce the side effects of surfactants and maximize the intestinal absorption of poorly soluble drugs, a supersaturable self-emulsifying drug delivery system (S-SEDDS) was proposed. The S-SEDDS formulations generally contain a lower concentration of surfactant and a polymeric precipitation inhibitor to yield and stabilize the drug in a temporary supersaturated state (Gao et al., 2003). The precipitation inhibitors thermodynamically and/or kinetically protract the supersaturated state of active molecules in aqueous medium by reducing the rate of drug nucleation and crystal growth through physical interactions with drug compounds or by changing the medium properties (Chauhan et al., 2013; Loftsson et al., 1996; Usui et al., 1997). Several previous studies with poorly water soluble drugs, such as paclitaxel (Gao et al., 2003), PNU-91325 (Gao et al., 2004), and AMG 517 (Gao et al., 2009), clearly demonstrated that the supersaturable systems provide higher oral absorption and fewer side effects as compared to that of the conventional SEDDS. Also, we previously reported that Soluplus1 (polyvinylcaprolactam–polyvinyl acetate–polyethylene glycol graft copolymer) significantly amplified the spring and parachute effect in S-SEDDS formulation of celecoxib (Song et al., 2013). Soluplus1 is an amphiphilic polymer designed to produce amorphous dispersions by hot melt extrusion due to its low glass transition temperature of 72 C (Hardung et al., 2010). It can enhance the solubility and the dissolution, and even retard drug precipitation of poorly watersoluble drug in aqueous medium (Zhang et al., 2013; Kim, 2013a,b). The present study was performed to construct a novel S-SEDDS to improve oral absorption of DTS. A supersaturable system of DTS was formulated by screening various kinds of hydrophilic and/or amphiphilic polymers including Soluplus1, as a precipitation inhibitor. Each formulation was evaluated for their physicochemical characteristic such as droplet size, homogeneity, drug content, and in vitro dissolution profile under non-sink condition to assess the supersaturation state of the 5a-reductase inhibitor. We subsequently compared the drug absorption of optimized S-SEDDS with drug suspension and conventional SEDDS in rats using a validated LC–MS/MS assay.
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2. Materials and methods
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2.1. Materials
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DTS was obtained from MSN Laboratories Limited (India, purity over 99.0 w/w%). Capryol 90TM and Transcutol1 HP were kindly
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supplied by Gattefosse (Saint Priest, France). Cremophor1 EL, Kollicoat1 MAE 30DP (methacrylic acid ethylacrylate copolymer), Kollidon1 90F (polyvinylpyrrolidone), and Soluplus1 were kindly supplied by BASF (Ludwigshafen, Germany). Hypromellose 2910 (Hydroxypropylmethyl cellulose, HPMC) was supplied by Shin-Etsu Chemical Co. (Tokyo, Japan). Finasteride (purity over 98 w/w%) was purchased from Sigma Chemical Co. (St. Louis, MO, USA). Acetonitrile and methanol of HPLC grade were purchased from J.T. Baker (Phillipsburg, NJ, USA). All other chemicals used were of analytical grade.
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2.2. Animals
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Sprague-Dawley rats (male; 200–250 g; 7–9 weeks) were purchased from Orient Bio (Kyungki-do, Korea). They were kept in specific pathogen-free conditions with food and water freely available. All animal experiments were performed in accordance with the NIH “Principles of laboratory animal care” guidelines (NIH publication No. 85-23, revised 1996) and were approved by the Institutional Animal Care and Use Committee of Chung-Ang University in Seoul, Korea.
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2.3. Preparation of SEDDS and S-SEDDS formulations
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The DTS-loaded SEDDS was prepared by dissolving the drug (50 mg) in the mixture of Capryol 90TM/Cremophor1 EL/Transcutol1 HP. The components were mixed by vortexing at room temperature until the DTS was completely dissolved. The compositions of each formulation are detailed in Table 1. The S-SEDDS formulation was acquired by the further addition of precipitation inhibitors to SEDDS F3 formula. Various polymeric excipients (Kollidon1 90F, HPMC, Kollicoat1 MAE 30DP, and Soluplus1) were added into the SEDDS solution, and then, the mixture was vortexed vigorously using magnetic stirrer at room temperature to obtain a uniform suspension. The S-SEDDS suspension was stored at room temperature until used.
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2.4. Physical characterization of SEDDS and S-SEDDS formulations
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2.4.1. Droplet size Each formulation was diluted with an appropriate volume of simulated gastric juice and particle size distribution. The droplet size was assayed using a dynamic light-scattering particle size analyzer (Zetasizer Nano-ZS, Malvern Instrument, Worcestershire,
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Table 1 Compositions and physicochemical characteristics of DTS-loaded SEDDS and S-SEDDS formulations.
Compositions DTS (mg) CapryolTM 90 (mL) Cremophor1 EL (mL) Transcutol1 HP (mL) Soluplus1 (mg) Kollidon1 90F (mg) HPMC 2910 (mg) Kollicoat1 MAE (mg)
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
F13
0.5 42.3 24.7 34.6 – – – –
0.5 21.2 12.4 17.3 – – – –
0.5 14.1 8.2 11.5 – – – –
0.5 14.1 8.2 11.5 5 – – –
0.5 14.1 8.2 11.5 – 5 – –
0.5 14.1 8.2 11.5 – – 5 –
0.5 14.1 8.2 11.5 – – – 5
0.5 14.1 8.2 11.5 1 – – –
0.5 14.1 8.2 11.5 2.5 – – –
0.5 14.1 8.2 11.5 10 – – –
0.5 14.1 8.2 11.5 15 – – –
0.5 14.1 8.2 11.5 5 – 5 –
0.5 14.1 8.2 11.5 5 – – 5
137.3 0.29 98.8 N.D.
119.9 0.07 97.6 25.9
99.7 0.26 105.1 65.8
96.1 0.18 105.7 34.5
96.9 0.13 101.5 20.1
95.9 0.18 100.4 N.D.
98.3 0.17 99.4 N.D.
108.3 0.25 101.0 N.D.
112.2 0.19 105.6 N.D.
95.9 0.14 103.0 N.D.
91.3 0.17 103.8 N.D.
97.4 0.16 102.9 N.D.
Physicochemical characteristics Size (nm)a 128.0 PDIa 0.26 Drug contents (%)a 100.3 DT50 (min)a,b N.D.c a
Data are expressed as mean values (n = 3). Their standard deviations were less than 10% of the mean values. Indicates the time to maintain the dissolution rate over 50%. DT50 is calculated by subtracting the initial time point from the last time point where over 50% of dissolution rate is reached in Fig. 2. c Not determined. b
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UK) with a 50 mV laser at a scattering angle of 90 . All measurements were carried out in triplicate under ambient conditions. 2.4.2. Drug content Each formulation containing 0.5 mg of DTS was dissolved in 20 mL of acetonitrile and sonicated for 10 min. Samples were then centrifuged at 12,000 rpm for 10 min and the supernatants were analyzed by HPLC analysis. The quantitative determination of DTS was performed by HPLC using acetonitrile:water (70:30) as a mobile phase at a flow rate of 1 mL/min. The HPLC system consisted of a pump (L-2130), a UV detector (L-2400), a data station (LaChrom Elite, Hitachi, Japan), and a 15 cm C18 column (Kromasil, AkzoNobel, Sweden). The column eluant was monitored at 210 nm, and the DTS peak was separated at a retention time of 6.5 min. The calibration curve was linear in the DTS concentration range of 0.1–10 mg/mL (y = 210673x 7208.2, r2 = 0.999).
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2.5. In vitro dissolution test
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In vitro dissolution tests were performed according to the USP apparatus II (paddle) method with a Vision1 classic 6TM dissolution testing station and a Vision1 heater (Hanson, USA). Each formulation containing 0.5 mg of DTS was placed in the dissolution vessel containing 250 mL of simulated gastric juice maintained at 37.0 0.5 C and stirred at 50 rpm. The drug release experiment with F4 was further carried out in distilled water and intestinal fluid (pH 6.8). Then, 5 mL of samples were taken at various time intervals and replaced with fresh and pre-warmed dissolution medium. The samples were centrifuged at 12,000 rpm for 10 min and the supernatants diluted with methanol for HPLC analysis.
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2.6. In vivo oral absorption study
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2.6.1. Experimental Male Sprague-Dawley rats weighing approximately 250 g were fasted for 16 h prior to the experiment and divided into 3 groups. The rats in the first group were orally administered 1 mL of a 0.2% methylcellulose aqueous suspension containing powdered DTS, the rats in the second group received the conventional SEDDS formulation (F3), and the rats in the third group received optimized S-SEDDS (F4) at a dose of 2 mg/kg (Choo et al., 2013). Blood samples (0.3 mL) were then collected from the retro-orbital plexus with heparinized tubes at predetermined time intervals and centrifuged at 12,000 rpm for 10 min. Plasma samples were stored at 20 C until analysis by LC–MS/MS. 2.6.2. LC–MS/MS determination of DTS in rat plasma To isolate components in a sample, liquid-liquid extraction (LLE) was carried out prior to its analysis (Choo et al., 2013; Kang et al., 2014). Plasma (100 mL) was thawed and mixed with 20 mL of a finasteride (an internal standard, IS) solution (200 ng/ mL), 100 mL of an aqueous sodium hydroxide solution (39.997 mg/mL), and 600 mL of a LLE reagent (tert-butyl ether:methylene chloride, 70:30, v/v) for 10 min by using a rotator. The mixture was centrifuged at 12,000 rpm for 5 min and the resulting organic phase was dried under a gentle stream of nitrogen gas. Finally, the residue was reconstituted with 200 mL of a 50% aqueous methanol solution for introduction into the LC–MS/MS system. LC separation was performed by a LC-20 Prominence system (Shimadzu, Japan). The temperature of the autosampler was kept at 4 C and components in 30 mL of each reconstituted sample were separated on a Hydrosphere C18 (5 mm, 250 2.0 mm, YMC, Japan) column at 45 C. An isocratic mobile phase condition (0.1% aqueous formic acid solution:
3
acetonitrile, 30:70%, v/v) was used at a flow rate of 0.250 mL/min and the analysis time per sample was 15 min. The components eluted from the column were delivered into an API 2000 triple quadruple mass spectrometer (AB/SCIEX, USA) through a Turbospray ion source (AB/SCIEX) for multiple reaction monitoring (MRM) assays of DTS (528.9/460.9/49, the m/z value of the precursor ion/the m/z value of the product ion/the collision energy, V) and IS (373.3/305.2/49, the m/z value of the precursor ion/the m/z value of the product ion/the collision energy, V) in positive ion mode. Additional mass spectrometer conditions are as follows: spray voltage at 5300 V; spray temperature at 400 C; nebulizer gas (gas 1) at 35; heater gas (gas 2) at 35; declustering potential at 70 V. The good linearity (r2 = 0.993) was observed at a DTS concentration range between 5 and 400 ng/mL.
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2.6.3. Pharmacokinetic analysis Pharmacokinetic analysis was performed using a BA Calc 2007 pharmacokinetic analysis program (Korea Food & Drug Administration, Korea). Area under the curve (AUC) from 0 to 24 h was calculated using the program’s linear trapezoidal rule. Maximum plasma concentration (Cmax) and the time needed to reach the maximum plasma concentration (Tmax) were determined directly from the concentration–time data. The relative BA was calculated as a percentage of the AUC of formulations to that of the drug suspension.
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2.7. Statistical analysis
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All data are expressed as the mean SD. Statistical significance was determined by Student’s t-test with a threshold of P < 0.05, unless otherwise indicated.
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3. Results
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3.1. Physicochemical characteristics of SEDDS and S-SEDDS formulations
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The physicochemical characteristics of DTS-loaded SEDDS and S-SEDDS formulations were evaluated by droplet size, homogeneity, and drug content (Table 1). The particle size of the SEDDS dispersions (F1–F3) was approximately 130 nm, with no significant differences between the SEDDS formulas. On the other hand, mean particle size of S-SEDDS (F4–F13) ranged from 90 to 110 nm, which is slightly smaller than SEDDS. The low polydispersity index (PDI) of less than 0.3 in all formulations indicates a narrow and homogeneous size distribution. The drug content in each formulation was almost equal (97.6–105.7%) with small standard deviations, indicating that the drug was uniformly distributed in the liquid formulations.
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3.2.1. Effect of drug loading amount on drug dissolution from SEDDS formulations The dissolution patterns of DTS from the SEDDS formulations at different ratios of the drug to SEDDS vehicle (F1–F3) in simulated gastric fluid are shown in Fig. 1. Upon contact with the gastric fluid, F1–F3 formulations rapidly formed a fine emulsion with a bluish reflection, and over 70% the drug was released after 5 min. However, as time elapsed, the drug release from the F2 and F3 rapidly decreased to 24% and 11% at 2 h, while F1 maintained its dissolution state. In case of F3, the dissolved amount was reduced drastically to reach the equilibrium level of about 11% at 2 h. In the next study, precipitation inhibitors were incorporated in SEDDS F3 formulation to stabilize the supersaturated state of DTS and establish the S-SEDDS formulation.
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Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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DTS dissolved (%)
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Time (min) Fig. 1. Dissolution profiles of DTS from SEDDS formulations in simulated gastric juice prepared with different ratios of drug to SEDDS vehicle; 1:200 (F1, &); 1:100 (F2, 4), and 1:67 (F3, ^). Data represent mean SD (n = 3).
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3.2.2. Effect of various precipitation inhibitors on drug dissolution from S-SEDDS formulations The effects of hydrophilic and/or amphiphilic polymers for supersaturating F3 formulation were evaluated as depicted in Fig. 2. The amount of a precipitation inhibitor in each formulation was identically set to 5 mg, which is approximately 15% to SEDDS vehicle. In the case of the hydrophilic polymers Kollidon1 90F (F5), HPMC (F6), and Kollicoat1 MAE (F7), there was no increase in the dissolution level, even though F7 displayed a positive effect on decelerating precipitation. In contrast, the addition of Soluplus1 (F4), an amphiphilic polymer, to SEDDS formulation significantly increased the drug dissolution rate at 2 h (Fig. 2). Upon contact with the gastric fluid, almost 80% of the drug was released within 5 min and the DTS concentration in simulated gastric fluid was gradually decreased but maintained over 30% at 2 h, which drastically reduced the amount dissolved to about 10% in F3. For further quantitative observation, we established the term ‘DT50’ that indicates ‘the time to maintain the dissolution rate over 50% (Table 1). DT50 is calculated by subtracting the initial time point from the last time point where over 50% of dissolution rate is
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3.2.3. Effect of Soluplus1 amount on drug dissolution from S-SEDDS formulations In order to evaluate the influence the amount of Soluplus1 upon the duration of the supersaturated state, a series of S-SEDDS formulations containing the precipitation inhibitor (3, 7.5, 15, 30, and 45% to SEDDS vehicle) were evaluated (F4 and F8–11). As plotted in Fig. 3, as the proportion of the precipitation inhibitor increased, the drug concentration in aqueous medium at 2 h increased and plateaued for the F4 formula containing 15% of Soluplus1, dissolving over 30% of the drug dissolved at 2 h. The DTS concentration–time profiles suggest that the presence of 15%, 30% and 45% of Soluplus1 (F4, F10 and F11) appears sufficiently effective, yielding similar drug concentration–time profiles. The dissolution profile of DTS from the drug powder and physical mixture of drug and Soluplus1 in a weight ratio of 1:10 are further investigated (Fig. 3). In gastric juice, physical mixtures of drug with the polymer showed dissolution profiles similar to those of the intact drug; only about 10% of the drug was release for 2 h. The S-SEDDS F4 showed a consistently higher drug 7concentration–time profile as compared to the SEDDS F3 and physical mixtures, indicating a synergistic effect on the dissolution of the drug by virtue of the precipitation inhibitor with SEDDS.
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3.2.4. Effect of combination of Soluplus1 with hydrophilic polymers on drug dissolution from S-SEDDS formulations In order to produce the extended supersaturated state, some hydrophilic additives were combined with Soluplus1-based F4 formula (Fig. 4). However, both HPMC-added (F12) and Kollicoat1 MAE-added (F13) compositions did not produce a complementary precipitation-inhibiting effect with Soluplus1. Conversely, the other hydrophilic additives showed an antagonistic effect compared to Soluplus1 alone (F4), resulting in decreased supersaturation or an increase in drug precipitation from the
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DTS dissolved (%)
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reached in Fig. 2. Initial and last time points were calculated by drawing a horizontal line at 50% on the Y axis and dropping a vertical line form the point of intersection with the dissolution curve to the X axis in Fig. 2. DT50 value of F4 was the highest at about 65.9 min, among the various formulations. It was 2.6-, 2.0-, and 3.3-fold higher compared to the DT50 values for F3, F5, and F6, respectively (Table 1).
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Time (min) Fig. 2. Dissolution profiles of DTS from S-SEDDS formulations in simulated gastric juice prepared with different precipitation inhibitors; Soluplus1-added (F4, ^); Kollidon1 90F-added (F5, *), HPMC-added (F6, ~), and Kollicoat1 MAE-added (F7, &), and SEDDS (F3, ^). The amount of precipitation inhibitor in each formulation was identical to 5 mg, which is approximately 15% to SEDDS vehicle. Data are expressed as mean SD (n = 3).
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Time (min) Fig. 3. Dissolution profiles of DTS from S-SEDDS formulations according to the amount of Soluplus1 added; 1 mg (F8, ); 2.5 mg (F9, 4); 5 mg (F4, ^); 10 mg (F10, *), and 15 mg (F11, ~), physical mixture of drug with 5 mg of Soluplus1 (^), and drug powder () in simulated gastric juice. Data are expressed as mean SD (n = 3).
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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Time (min) Fig. 4. Dissolution profiles of DTS from S-SEDDS formulations after combination of hydrophilic polymers with Soluplus1; Soluplus1 alone-added (F4, ^); Soluplus1 and HPMC-added (F12, ~), and Soluplus1 and Kollicoat1 MAE-add (F13, &). Data are expressed as mean SD (n = 3).
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supersaturated state. From these findings, the optimized formula for the drug, SEDDS vehicle and the Soluplus1 at a ratio of 1:67.6:10 (F4) was finally established to as the supersaturable oral dosage form of DTS.
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3.2.5. Effect of dissolution medium on drug dissolution from the optimized S-SEDDS formulation Dissolution profiles of the optimized S-SEDDS (F4) were further evaluated in different dissolution media (Fig. 5). The release rate of the 5a-reductase inhibitor was pH-independent, yielding that over 30% of DTS was supersaturated at 2 h in media at pH 1.2, pH 6.8 and distilled water.
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3.3. In vivo absorption study in rats
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Fig. 6 shows the mean plasma concentration profiles of DTS in rats upon oral dosing of suspension, SEDDS (F3) or S-SEDDS (F4) formulations at a drug dose of 2 mg/kg. Mean plasma DTS levels of both SEDDS and S-SEDDS formulations were significantly higher than that of the drug suspension, which slowly increases until 12 h (Fig. 6). In both SEDDS and S-SEDDS formulations, however, the
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Fig. 6. Plasma concentration-time profiles of DTS in rats after oral administrations of drug suspension (), SEDDS (^), and S-SEDDS (^) formulations at a dose of 2 mg/kg. Data are expressed as mean SD (n = 3).
5a-reductase inhibitor is rapidly absorbed and shows a sharp increase in plasma DTS levels within 3 h. In particular, S-SEDDS F4 showed higher plasma levels than SEDDS for 24 h. The pharmacokinetic parameters, including Tmax, Cmax and AUC(0– 24 h), are listed in Table 2. As expected, S-SEDDS (F4) was superior to the other formulations in all parameters; the parameters were significantly different at P < 0.05. The AUC(0–24 h) value of S-SEDDS formula was 3.9-fold greater than that of the drug suspension and 1.3-fold greater than that of SEDDS. The Cmax value of S-SEDDS was about 5.6- to 2.0-fold higher compared to drug suspension and SEDDS, respectively. From these findings, we concluded that the oral BA of DTS was greatly improved by the supersaturable formulation.
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4. Discussion
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SEDDSs are widely used to enhance the oral absorption of poorly soluble drugs, including DTS. In our study, the SEDDS formulation consisted of Capryol 90TM, Cremophor1 EL, and Transcutol1 HP that was easily dispersed in aqueous dissolution medium to make fine o/w microemulsion spontaneously and exhibited high initial dissolution rate. However, the dissolution was time-dependent, in which the amount dissolved rapidly decreasing due to the loss of its solubilization capacity with lower amount of SEDDS vehicle (F3). It was reported that the solvent capacity of the lipid formulation falls logarithmically as the mixture is diluted into an aqueous medium (Pouton, 2000). On the other hand, the large amount of surfactant present in the SEDDS formulation could lead to GI side effects after daily intake. Especially Cremophor1 EL, which was reported to cause severe side effects such as hypersensitivity, nephrotoxicity and neurotoxicity (Danhier et al., 2009). Therefore, a novel supersaturable
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40 Table 2 Pharmacokinetic parameters of DTS from the drug suspension, SEDDS, and S-SEDDS formulations after a single 2 mg/kg oral administration in rats.
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Time (min) Fig. 5. Dissolution profiles of DTS from the optimized S-SEDDS (F4) formulation in the different pH media; gastric juice (pH 1.2, ^), intestinal fluid (pH 6.8, ), and distilled water (&). Data are expressed as mean SD (n = 3).
Parameters
Drug suspension SEDDS (F3)
S-SEDDS (F4)
AUC(0–24 h) (mg h/mL)a Cmax (mg/mL)a Tmax (h)a Relative BA (%)b
580.99 107.99 35.98 8.02 9.6 2.19 –
2282.50 460.98 183.30 38.81 1.92 0.67 393
1796.16 359.87 108.55 20.64 3.13 2.17 309
a
Values represent mean SD (n = 5–10). Calculated as percentage of the mean AUC(0–24 h) of each group to that of drug suspension. b
Please cite this article in press as: Lee, D.H., et al., Improved oral absorption of dutasteride via Soluplus1-based supersaturable self-emulsifying drug delivery system (S-SEDDS). Int J Pharmaceut (2014), http://dx.doi.org/10.1016/j.ijpharm.2014.11.060
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system containing a reduced amount of surfactant and a small amount of precipitation inhibitor was designed to generate and maintain a supersaturated state to improve oral absorption of DTS in GI track. In order to evaluate the drug concentration sustained in the supersaturated state and the degree of supersaturated state as a function of time, an in vitro dissolution test was designed as a non-sink condition. The total volume of the medium chosen here was 250 mL, based on physiological considerations of the total residual volume of stomach fluid. On the other hand, the solubility of DTS in aqueous solution was reported as an approximately 0.038 ng/mL in water at 25 C (Avodart1 FDA Biopharmaceutics Review, 2001). Thus, the experimental condition does not guarantee the sink conditions for DTS. In designing the supersaturable formulations, it is essential to include a precipitation inhibitor in the SEDDS vehicle to maintain drugs in supersaturated state in aqueous medium (Xu and Dai, 2013). Precipitation inhibitors inhibit drug nucleation and/or crystal growth by steric stabilization, surface stabilization by adsorption onto colloidal surfaces, and/or specific interactions with a drug (e.g., by hydrogen bonding or hydrophobic interactions) (Wen et al., 2005; Kojima et al., 2012; Song et al., 2013). For this purpose, in the present study, hydrophilic polymers such as HPMC, Kollidon1 90F, and Kollicoat1 MAE were screened as potential candidates for precipitation inhibitor. However, these polymers were not effective for stabilizing the supersaturated state of DTS with SEDDS formulation (Fig. 2). It is assumed that these hydrophilic polymers were not absorbed efficiently on the surface of the oily droplet or free drug molecules, but rather molecularly dissolved and freely diffused into the aqueous medium. Alternatively, Soluplus1, an amphiphilic polymer, significantly inhibited precipitation of DTS, preserving it in the supersaturated state (Fig. 2). The F4 formula showed a consistently higher apparent drug concentration as compared to F3, providing 2.6-fold higher DT50 value compared to F3. It was interpreted by our groups that a hydrophobic polycaprolactam moiety of Soluplus1 intercalates to hydrophobic tail packing of dispersed phase and thus, the interface becomes denser. Simultaneously, polyethylene glycol groups located on the surface of colloidal dispersion form a hydrophilic barrier, hindering aggregation and/or destruction of emulsion droplet in gastric fluid (Song et al., 2013). There was a critical amount of Soluplus1 for stabilizing the supersaturated state of DTS. In the S-SEDDS systems with Soluplus1, 5 mg of the precipitation inhibitor effectively retards DTS precipitation and sustains a higher apparent concentration in a non-sink condition (Fig. 3). This result is consistent with an earlier report that the S-SEDDS formulation containing 40 mg of Soluplus1 as a precipitation inhibitor exhibited the greatest dissolution of celecoxib in a non-sink condition with a medium volume of 1000 mL (Song et al., 2013). The concentration of Soluplus1 in the aqueous medium was quite comparable in two studies between 20 and 40 mg/mL, which is analogous with the critical micelle concentration of Soluplus1 (46.3 mg/L) at pH 1.2. It suggests that the optimal level of Soluplus1 in S-SEDDS is closely associated with the micelle formation of the amphiphilic polymer. Three types of different formulations including DTS suspension, SEDDS F3, and S-SEDDS F4, were administered orally to rats and their pharmacokinetic parameters were compared (Table 2). The relative BA of SEDDS and S-SEDDS was about 2.9- and 3.9-fold higher compared to drug suspension which was used as a reference. It indicates that self-emulsifying formulations spontaneously formed fine oil droplets and present DTS in a dissolved form, avoiding the dissolution step. Especially, S-SEDDS could remarkably improve the peroral absorption of DTS, providing even 30% greater BA than that of SEDDS. Supersaturable formulation is
able to induce a supersaturated drug concentration that absorbed in the desired time frame when exposed to the GI tract in vivo. In this study, both the in vitro dissolution test and the in vivo pharmacokinetic study clearly indicate that the S-SEDDS formulation is remarkably effective in inhibiting drug precipitation in the GI track and thus improving the rate and extent of DTS absorption. From these results, we conclude that S-SMEDDS is superior to SEDDS formulations for DTS oral delivery.
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5. Conclusion
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Novel DTS-loaded S-SEDDS formulation was successfully prepared by incorporating Soluplus1 as a precipitation inhibitor to SEDDS composed of Capryol 90TM as an oil, Cremophor1 EL as a surfactant, and Transcutol1 HP as a cosurfactant. The in vitro dissolution tests in a non-sink condition revealed that a small amount of Soluplus1 (15% w/v to SEDDS vehicle) effectively slowed down drug precipitation and played a critical role in maintaining a supersaturated state of DTS. Drug dissolution from S-SEDDS was pH-independent. The in vivo pharmacokinetic study in rats demonstrated that S-SEDDS formulation significantly improved oral absorption of DTS, providing 3.9-fold greater BA compared to drug suspension. This study demonstrates the potential use of the S-SEDDS formulation in the development of poorly water-soluble oral drugs.
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References
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