Arch. Pharm. Res. DOI 10.1007/s12272-015-0590-y
RESEARCH ARTICLE
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride using a fluidized bed coater Yong-Il Kim1,2 • Roshan Pradhan1 • Bijay K. Paudel1 • Ju Yeon Choi1 Ho Taek Im2,3 • Jong Oh Kim1
•
Received: 3 January 2015 / Accepted: 7 March 2015 Ó The Pharmaceutical Society of Korea 2015
Abstract In this study, the enteric-coated delayed-release pellets of duloxetine hydrochloride (DLX) were formulated using a fluidized bed coater. Three separate layers, the drug layer, the barrier layer, and the enteric layer, were coated onto inert core pellets. Among the three formulations (F1– F3), the dissolution profiles of formulation F2 were most similar to those of the marketed product, with similarity and difference factors of 83.99 and 3.77, respectively. In addition, pharmacokinetic parameters of AUC, Cmax, Tmax, t1/2, Kel, and MRT of DLX for the developed formulation (F2) did not differ significantly from those for the marketed product in beagle dogs, suggesting that they were bioequivalent. Our results demonstrated that the in vitro dissolution data resembled the in vivo performance of the drug. Therefore, this study has a positive scope for further scale up and development of the formulation for achievement of the generic product. Keywords Duloxetine hydrochloride Delayed-release Enteric coated Pellet Pharmacokinetics
Yong-Il Kim and Roshan Pradhan contributed equally to this study. & Ho Taek Im [email protected] & Jong Oh Kim [email protected] 1
College of Pharmacy, Yeungnam University, 214-1, Dae-Dong, Gyongsan 712-749, Republic of Korea
2
Pharm. R&D Institute, Hanmi Pharm. Co., Ltd., Hwasung 445-913, Republic of Korea
3
School of Pharmacy, Sungkyunkwan University, Jangan-gu, Suwon 440-746, Gyeonggi-do, Republic of Korea
Introduction Duloxetine hydrochloride (DLX) is a novel dual serotonin and norepinephrine reuptake inhibitor, approved for treatment of major depressive disorders, diabetic peripheral neuropathic pain (Fishbain et al. 2008), fibromyalgia (Mease et al. 2010), generalized anxiety, and woman stress urinary incontinence (Robinson and Cardozo 2010). The claimed mechanism of action of the drug is based on the specific inhibition of both serotonin and norepinephrine reuptake while it weakly inhibits dopamine reuptake and has no significant affinity for histaminergic, dopaminergic, cholinergic or adrenergic receptors (Goldstein et al. 2004). DLX is a newer and more preferable antidepressant because of its favorable pharmacodynamic features viz. dual inhibition, tolerability, safety, faster recovery, fewer side effects, and low affinity for other neuronal receptors (Bymaster et al. 2001; Sharma et al. 2000; Westanmo et al. 2005). However, the drug was found to be acid labile, which results in its degradation in a gastric environment, thus necessitating the need for development of an entericcoated system (Shravani et al. 2011). Enteric-coated formulations are suitable vehicles for modification of the release of drug or active pharmaceutical ingredients at specific target areas within the gastrointestinal tract. Enteric coating is an effective method of protecting the drug against a gastric environment and preventing the release of the encapsulated particles or the drug before reaching the target site. DLX is unstable in solution at pH values less than 2.5, therefore, enteric polymer-coated formulations have been developed to prevent its acid degradation in the stomach and to provide for its subsequent release in the small intestine (Jansen et al. 1998; Sinha et al. 2009). Hydroxy propyl methyl cellulose acetate succinate (HPMC-AS) and hydroxy propyl methyl
123
Y. Kim et al.
cellulose phthalate (HPMC-P) are the major polymers used as an enteric-coating for DLX capsules. However, it has been reported that reaction of DLX with HPMC-AS and HPMC-P forms impurities like succinamide and phthalamide (Jansen et al. 1998). In order to prevent impurity formation between the core and the surrounding enteric polymer, the separate layer or middle barrier layer is coated. The type of coating technique strongly affects the film microstructure and thus affects the release mechanism and rate from pellets coated with polymer blends (Lecomte et al. 2004). Pellets have increased in importance over the years due to their distinctive advantages in both technological and therapeutic aspects (Korakianiti et al. 2000). When administered orally, therapeutic advantages include modification of drug release, division of dose strength, and free dispersion in the gastro intestinal tract (Abdul et al. 2010). In this study, DLX pellets were prepared with three separate coatings, consisting of the inner drug layer, middle barrier layer or sub-layer, and the outer enteric layer (Fig. 1). All three layers were coated using a fluidized bed coater. HPMC was used for the drug and barrier layering, while HPMC-P was used as the enteric polymer. The effect of the concentration of the enteric polymer on the dissolution of prepared formulations was examined. The in vitro release of DLX from the developed formulations was compared with that from the marketed CymbaltaÒ capsules (Lilly), and pharmacokinetic studies were performed in beagle dogs after oral administration of formulated DLX capsule and the marketed product.
Materials and methods Materials DLX was purchased from Zydus Pharmaceuticals (New Jersey, USA). Hydroxypropyl methylcellulose (HPMC 2910, PharmacoatÒ 606) and hydroxypropyl methylcellulose phthalate (HPMCP, HP-50) were obtained from ShinEtsu Chemical Co. (Tokyo, Japan), respectively. Fig. 1 Schematic illustration of DLX pellets
123
Fluoxetine, as an internal standard, was purchased from Sigma (St. Louis, MO, USA). Sugar beads (25–30 sieve size) were purchased from Hanns G. Werner GmbH Co. (Holstein, Germany). Sucrose, talc and triethyl citrate (TEC) were supplied by BASF Chemical Co. (Ludwigshafen, Germany). Solvents such as ethanol and acetone were purchased from OCI Company Ltd. (Seoul, South Korea). The commercial product (CymbaltaÒ) was purchased from a market. All other chemicals used were of reagent grade and used without further purification. Preparation of DLX pellets DLX pellets were prepared using a fluidized bed coater (XP-70). Using this technique, the polymeric solutions containing drug and other excipients were deposited onto the inert sugar beads. Details regarding the parameters of the fluidized bed coating process are shown in Table 1. The sugar beads were coated in three layers viz., the inner drug layer, middle barrier layer, and the outer enteric layer. HPMC was used both as the binding and the barrier polymer because of its low molecular weight and use as a good pelletization aid (Chatlapalli and Rohera 1998), while HP-50 was used as the enteric polymer. Inner drug layer Initially, the sugar beads, 42 mg per capsule (cap) were loaded into the fluidized bed and warmed before treatment with drug solution. The coating solution was prepared by first dissolving the HPMC (11 mg/cap) in distilled water (300 ml) followed by addition of DLX (33.7 mg/cap) with constant stirring. Talc (4.47 mg/cap) was dispersed and stirred in order to obtain a homogeneous dispersion. This coating solution was sprayed over the sugar beads at a spray rate of 5 ml/min. Middle barrier layer The primary function of this layer is to minimize the interaction between the inner drug layer and the outer enteric
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride... Table 1 Process parameters of the fluidized bed coating
Parameters
Inner drug layer
Middle barrier layer
Outer enteric layer
Nozzle size (mm)
1.0
1.0
1.0
Inlet temperature (°C)
55
45
31
Outlet temperature (°C)
40
30
29
Bed temperature (°C)
25
25
25
Spray rate (ml/min)
5
6
8
1.2
1.0
0.8
2
Automizing air pressure (kgf/cm )
Table 2 Composition of the outer enteric layer of DLX pellets Ingredients
F1 (mg)
F2 (mg)
F3 (mg)
DLX (barrier coated pellets)
121.17
121.17
121.17
HP-50
20
25
30
Triethyl citrate
4
5
6
Talc
2.4
3
3.6
Ethanol
100
125
150
Acetone
200
250
300
Total
147.57
154.17
160.77
index (CI) were calculated according to the equations given below: qtapped 0 Hausner s ratio ¼ qbulk ! qtapped qbulk 0 100% Carr s index ¼ qtapped where qbulk and qtapped are bulk and tapped densities of the enteric-coated DLX pellets. In vitro dissolution test
polymer. HPMC (6 mg/cap) was dissolved in distilled water (190 ml) followed by addition of sucrose (9.6 mg/cap) with continuous stirring. Talc (14.4 mg/cap) was dispersed and stirred until a homogeneous dispersion was obtained. This coating solution was sprayed over the drug layer at a spray rate of 6 ml/min. Outer enteric layer HP-50 (20–30 mg/cap), was dissolved in an ethanol/acetone (1:2 v/v) mixture. In order to improve the elasticity of the coating film, and to minimize film-forming temperature, TEC (5 mg/cap) was added as a plasticizer to the above mentioned solution (Frohoff-Hulsmann et al. 1999). Talc (3 mg/cap) was used as the anti-adherent. The coating solution was applied over the barrier layer at a spray rate of 8 ml/min. The optimized weight ratio of HP-50:TEC:talc was 5:1:0.6. The detail compositions of formulations with varying amounts of enteric polymer (HP-50) are shown in Table 2. Determination of flow properties of enteric-coated DLX pellets The flow property of the pellets is an important parameter for hard gelatin capsule filling. The formulated pellets were tested for bulk and tap density using the AS-100 tap density tester (AimSizer). Then, Hausner’s ratio (HR) and Carr’s
A dissolution test was performed on six capsules from each formulation containing different amounts of enteric polymer (HP-50). Dissolution profiles of the prepared formulations and the marketed product were determined in 0.1 N HCl and phosphate buffer (pH 6.8). The dissolution apparatus consisted of the USP XXIII, apparatus 1 (basket method) (VanKel VK 7000; Varian Inc., Palo Alto, CA, USA) set at a speed of 100 rpm, with 900 ml of dissolution medium, pre-warmed at 37 ± 0.5 °C. Initially, the capsules were exposed to 0.1 N HCl (pH 1.2) for 120 min, followed by replacement of the medium with phosphate buffer (pH 6.8). 2 ml of dissolution samples were collected after 5, 10, 15, 30, 45, 60, 90 and 150 min, followed immediately by addition of an equal volume of fresh dissolution media maintained at the same temperature to keep the volume of dissolution media constant and to maintain the sink condition (Choi et al. 1998; Yong et al. 2005). The withdrawn samples were filtered through a membrane filter (0.45 lm) and after suitable dilution with distilled water, analyzed by injecting 50 ll into the HPLC, as described below. The percent drug release was then graphed against time and the release profiles were studied. HPLC conditions The HPLC analysis system (Hitachi Co., Tokyo, Japan) consisted of a Hitachi D-7000 interface, a pump (Model L-7100), ultraviolet detector (Model L-7400), autosampler (Model L-7200), and a column oven (Model L-7300). A C8 analytical column (ACEÒ (ACT): 3 lm, 75 mm 9 4.6 mm,
123
Y. Kim et al.
GL Sciences Inc., Tokyo, Japan) was used. The mobile phase consisted of tetrahydrofuran (THF): methanol: pH 5.5 phosphate buffer (9:32.3:58.7, volume ratio). The effluent was monitored at a UV absorption wavelength of 288 nm, and a flow rate of 1.5 ml/min. The column oven was maintained at 45 °C. The analytical method was validated according to ICH recommendations for validation of analytical methods (ICH Q2A, Q2B 1995). Similarity factor (f2) and difference factor (f1) Similarity (f2) and difference factors (f1) measure the closeness between the two dissolution profiles. f2 and f1 were calculated according to the equations given below: 9 8" #0:5 n = < 1X f2 ¼ 50 log 1 þ ðRj TjÞ2 100 ; : n t¼1 2P n
R j Tj
6t¼1 f1 ¼ 6 n 4 P
Rj
3 7 7 100 5
t¼1
where n is the number of time points, Rj and Tj are the dissolution values of the reference product and the test product, respectively, at each time point j. In order to consider the dissolution profiles similar, f1 values should be close to 0 and f2 values should be close to 100. In general, f1 values lower than 15 (0–15) and f2 values higher than 50 (50–100) indicate similarity of the dissolution profiles (Costa et al. 2001). Pharmacokinetic study In vivo experiments A pharmacokinetic study was carried out in ten beagle dogs, which were subjected to fasting overnight at least 10 h prior to dosing with DLX. One capsule (optimized formulation) or capsule (CymbaltaÒ) containing DLX 30 mg was administered orally. 5 ml of blood was collected into heparin-vacutainer tubes from the forearm vein before administration (at 0 h) and at predetermined time intervals of 0.5, 1, 2, 3, 4, 5, 6, 8, 12, 18, and 24 h after oral administration. Blood samples were centrifuged at 30009g for 10 min and 0.2 ml aliquots of plasma were stored at -80 °C until further analysis. All animal-handling procedures were in accordance with the protocols approved by the Institutional Animal Ethical Committee, Yeungnam University, South Korea. Blood sample analysis using LC–MS/MS The concentrations of DLX in dog plasma were analyzed by the reported LC–MS/MS method (Zhao et al. 2009).
123
Briefly, 20 ll of acetonitrile containing the working internal standard (fluoxetine, 40 ng/ml) was added to a 100 ll aliquot of plasma sample, followed by vortexing for 2 min. After centrifugation of the sample at 12,0009g for 10 min, the organic layer was collected and then separated on an Atlantis C18 column (3 lm, 50 mm 9 2.1 mm, Waters, Milford, MA, USA) using isocratic elution with acetonitrile and 30 mM ammonium formate solution maintained at pH 5 (90:10, volume ratio) at a flow rate of 0.3 ml/min. The column and autosampler tray temperatures were 35 and 15 °C, respectively. The eluent was introduced directly into the positive ionization electrospray source of a tandem quadrupole mass spectrometer (Quattro Micro; Micromass Manchester, UK). The ion source and desolvation temperatures were held at 100 and 400 °C, respectively. The optimum cone voltages and collision energy were 10 V and 6 eV, and 15 V and 18 eV for duloxetine and fluoxetine, respectively. Selected reaction monitoring (SRM) using precursor product ion transitions of m/z 298.08 [ 154 and m/z 310.02 [ 148.07 were used to quantify duloxetine and fluoxetine, respectively. Linear calibration curves were obtained for the concentration range of 0.1–100 ng/ml. The lower limit of quantification (LOQ) of DLX was 0.1 ng/ml. The within-day and between-day coefficients of variation were generally less than 10 %. Data acquisition and processing were accomplished using MasslynxTM 4.1 with a QuanLynxTM application manager. Pharmacokinetic analysis Standard methods (Gibaldi and Perrier 1982) were used in calculation of the pharmacokinetic parameters such as area under the drug-concentration time-curve (AUC), half-life (t1/2), elimination rate constant (Kel), and mean residence time (MRT) using non-compartmental analysis (WinNonlin 3.1TM; Pharsight Co., Mountain View, CA, USA). The maximum plasma concentration of drug (Cmax) and time to reach Cmax (Tmax) were directly computed from the plasma concentration versus time plot. All data are expressed as mean ± SD. Furthermore, the relative bioavailability (F) with reference to the AUC of the developed formulation compared to the marketed product was calculated using the equations given below: AUC A F¼ 100 % AUC B where F is the relative bioavailability, AUCA and AUCB are the area under drug-concentration time-curve of developed formulation (test) and marketed product (reference) respectively.
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride...
Statistical analysis Non-linear regression analysis was performed using the Statistical Package for Social Sciences (SPSS) computer program to compare the mechanism of drug release of the different formulations. Comparison of the pharmacokinetic parameters of the developed formulation and the marketed product were made using Student’s t test at a 95 % level of confidence using Microsoft Office Excel 2007. All data were expressed as the mean ± SD.
Results and discussion Preparation and characterization of enteric-coated DLX pellets Different formulations of enteric-coated DLX pellets (F1– F3) with different amounts of HP-50, as the enteric coating polymeric agent were designed and prepared using the fluidized bed coating method, based on the composition shown in Table 2. All of the formulated products lay within the limit of 98–102 % for the drug content (data not shown). To ensure good flow property, bulk density and tap density were measured, and HR and CI were calculated. The results obtained are presented in Table 3. Although both the HR and CI indicate the flow characteristics of the powder, granules or pallets, specifically CI is a measure of powder bridge strength and stability, while the HR is a measure of the interparticulate friction. Materials exhibiting lower values of CI or HR indicate better flow properties than higher values (Hausner 1967). In our study, HR of all of the formulated products was within 1.0 to 1.11 and CI was less than 10 %. Thus, all the pellet formulations showed excellent flow characteristics indicating uniformity of size and good sphericity of the pellets (Goyanes and Martinez-Pacheco 2013). The sieve analysis revealed that almost 95 % of pellets were distributed between 800–900 lm so pellets size distribution was found uniform. Since the size of the pellets depends upon the coating thickness, the average size of pellets (F1–F3) was found to be 820 ± 4 lm, 860 ± 5 lm 890 ± 8 lm respectively, which was similar with pellet size of the commercial product. Eventually, the free flowing enteric-coated pellets
Table 3 Flow properties of enteric-coated DLX pellets
equivalent to 30 mg of DLX were accurately weighed and filled into the hard gelatin capsules and were used for in vitro dissolution study. In vitro dissolution studies of enteric-coated DLX pellets The in vitro dissolution profiles of DLX from all formulations (enteric-coated DLX pellets) and the marketed product in 0.1 N HCl (pH 1.2) and phosphate buffer (pH 6.8) are shown in Fig. 2. In 0.1 N HCl (pH 1.2), formulation F1 exhibited a release of approximately 3 % in 120 min as well as a faster release pattern (approximately 80 % in 60 min) of DLX in phosphate buffer (pH 6.8) as compared to formulations F2, F3, and the marketed product, respectively. This may be attributed to the low amount of enteric polymers present in the formulation (Cao et al. 2013; Mohamed et al. 2013; Shravani et al. 2011), which could not resist the release of drug in the acidic dissolution medium. On the other hand, formulations F2, F3, and the marketed product showed no release of drug in 0.1 N HCl (pH 1.2) in 120 min indicating the effectiveness of the enteric coating polymer. Therefore, the result was in close agreement with the statement that the concentration of enteric polymer plays a primary role in acid-resistant pellets since there was an increased resistance to acid with an increase in the concentration of the enteric polymer. In pH 6.8, formulations F2 and F3 showed drug release of approximately 70 and 65 % in 60 min, respectively, which may be attributed to the higher amount of enteric polymers present in the formulation. These results indicated that as the amount of HP-50 increased, the release rate showed a gradual decrease. In fact, the extent of dissolution depends on the degree of ionization. The polymer should be ionized to be dissolved. Once the enteric polymer gets dissolved, the barrier polymer is another factor that affects the drug release from the pellets since it is the next layer after enteric polymer that obstructs the drug release. In addition, the increase in the concentration of the barrier polymer may increase the extent of obstruction of drug release. Furthermore, in order to investigate the effect of the rate of rotation on the drug release profile, formulations F1, F2, F3 and the marketed product were subjected to paddle speeds of 50, 100, and 150 rpm in 0.1 N HCl (pH 1.2) and
Formulations
Bulk density (g/cm3)
Tapped density (g/cm3)
Hausner ratio (HR)
Carr’s index (CI) (%)
F1
0.86 ± 0.02
0.88 ± 0.04
1.02
2.27
F2
0.87 ± 0.03
0.91 ± 0.03
1.04
4.39
F3
0.89 ± 0.02
0.94 ± 0.04
1.06
5.32
Each value represents the mean. (n = 3)
123
Y. Kim et al.
results, formulation F2 was considered the best formulation and was selected for the pharmacokinetic study in beagle dogs. Pharmacokinetic study of enteric-coated delayedrelease DLX pellets
Fig. 2 Dissolution profiles of DLX from different formulations (F1– F3) of enteric-coated DLX pellets and marketed product in 0.1 N HCl (pH 1.2) for 120 min and subsequently in phosphate buffer (pH 6.8) for 150 min. Dissolution was performed at 37 ± 0.5 °C and 100 rpm. Each value represents the mean ± SD (n = 6)
subsequently in phosphate buffer (pH 6.8) as the dissolution media; the comparative results are shown in Fig. 3. As reported, in vitro drug release for a polymeric system is affected by the dissolution medium and the rate of rotation (Costa and Sousa Lobo 2001). Our results showed that the drug release from all three formulations was not similar, but in fact increased with increasing rate of rotation (similarity factor \50; difference factor [15), indicating that the drug release from the enteric-coated pellets was affected by the rate of rotation. The similarity factor (f2) and the difference factor (f1) were calculated for all formulations using the release profile of the marketed product as a reference (Table 4). Also the f2 and f1 values of all three formulations at different rates of rotation with reference to 50 rpm are shown in Table 5. As shown in Table 4, the f2 value of formulation F1 was 46.67 (\50), and the f1 value was 20.75 (greater than 15), respectively which suggested that the dissolution profile of F1 was not similar to that of the marketed product. The dissimilarity may be attributed to the presence of a low amount of HP-50 in formulation F1, which resulted in more rapid release of dissolution profiles compared to the reference product. In contrast, formulations F2 and F3 had f2 values greater than 50 (83.99 and 65.48, respectively) and f1 values less than 15 (3.77 and 8.78, respectively), indicating that their dissolution profiles were similar to that of the marketed product (Costa et al., 2001; Yuksel et al., 2000). Among all formulations tested, formulation F2 showed the highest f2 value (83.99) and the lowest f1 value (3.77), corresponding to the highest similarity between its release profile and that of the reference product (CymbaltaÒ capsules). Considering all of the
123
The pharmacokinetic study in beagle dogs (n = 10) after oral administration of formulation F2 and the marketed product was performed and the mean plasma concentration–time curve was constructed. The dose administered was 30 mg DLX. As shown in Fig. 4, the developed formulation F2 exhibited almost similar pharmacokinetic profile with almost overlapping mean plasma concentration–time curves. A list of the corresponding pharmacokinetic parameters is shown in Table 6. The AUC (ng.h/ ml) and Cmax (ng/ml) values of developed formulation F2 were 147.2 ± 79.2 and 45.5 ± 17.9, respectively, while those of marketed product were 148.61 ± 88.16 and 46.80 ± 23.11, respectively. When these values were evaluated using student’s t test, they were not significantly different with each other (p \ 0.05). In addition, other pharmacokinetic parameters including Tmax, t1/2, elimination rate constant (kel), and mean retention time (MRT) were not significantly different. These results indicated that the pharmacokinetic profiles of the two formulations (test and reference) were very similar with each other. In addition, when the relative bioavailability (F) was calculated with reference to the AUC of the developed formulation F2 and the marketed product, it was found to be 99.0 %, which further indicated that the two formulations were bioequivalent (Mastan et al. 2011; Valizadeh et al. 2014). Therefore, our results demonstrated that there was a good correlation between the in vitro dissolution and the in vivo pharmacokinetic study with reference to the enteric-coated delayed-release DLX pellets. In conclusion, the enteric-coated delayed-release pellets of DLX (equivalent to 30 mg of DLX) were successfully prepared by a fluidized bed coating process, using HPMC as the binding and barrier coating and HP-50 as the enteric coating polymer. Among the three formulations (F1-F3), the dissolution profile of formulation F2 was most similar to the marketed product with similarity and difference factors of 83.99 and 3.77, respectively. Moreover, results of the pharmacokinetic study performed in beagle dogs after oral administration of prepared enteric-coated DLX pellets and the marketed product showed that the pharmacokinetic parameters, including AUC, Cmax, Tmax, t1/2, Kel, and MRT were not significantly different from those of the marketed product, suggesting that they were bioequivalent. Therefore, this study has a positive scope for further scale up and development of the formulation for achievement of the generic product.
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride...
Fig. 3 Effect of rate of rotation on the drug release profile of formulations F1, F2, F3, and marketed product in 0.1 N HCl (pH 1.2) and subsequently in phosphate buffer (pH 6.8) at a 50 rpm, b 100 rpm, and c 150 rpm. Dissolution was performed at 37 ± 0.5 °C. Each value represents the mean ± SD (n = 6)
Table 4 Comparison of dissolution profiles of enteric-coated DLX pellets with the profile of the marketed product Formulations
Similarity factor (f2)
Difference factor (f1)
F1
46.67
20.75
F2
83.99
3.77
F3
65.48
8.78
Table 5 Comparison of dissolution profiles of enteric-coated DLX pellets at different rates of rotation with reference to 50 rpm Formulations
100 rpm
150 rpm
f2
f1
f2
f1
F1
47.62
19.10
34.66
30.98
F2
48.34
21.58
33.89
36.09
F3
43.14
30.26
33.24
41.93
Fig. 4 Pharmacokinetic profile of DLX in beagle dogs after oral administration of marketed product (filled circle) and developed formulation (F2) (empty circle) at a single dose of 30 mg of DLX. Each value represents the mean ± SD (n = 10)
123
Y. Kim et al. Table 6 Pharmacokinetic parameters of DLX delivered by marketed formulation and developed formulation F2 after oral administration to beagle dogs Parameters
Marketed product
Developed formulation (F2)
AUC (ng.h/ml)
148.61 ± 88.16
147.18 ± 79.16
46.80 ± 23.11
45.54 ± 17.99
2.00 ± 0.48 3.16 ± 1.14
1.86 ± 0.69 3.74 ± 0.97
Cmax (ng/ml) Tmax (h) t1/2 (h) Kel (h-1)
0.25 ± 0.09
0.19 ± 0.06
MRT (h)
3.60 ± 0.82
3.67 ± 1.11
Each value represents the mean ± SD (n = 10)
Acknowledgments This study was supported by a grant from the Medical Cluster R&D Support Project of Daegu Gyeongbuk Medical Innovation Foundation, Republic of Korea (2013) (No. HT13C0011).
References Abdul, S., A.V. Chandewar, and S.B. Jaiswal. 2010. A flexible technology for modified-release drugs: Multiple-unit pellet system (MUPS). Journal of Controlled Release 147: 2–16. Bymaster, F.P., L.J. Dreshfield-Ahmad, P.G. Threlkeld, J.L. Shaw, L. Thompson, D.L. Nelson, S.K. Hemrick-Luecke, and D.T. Wong. 2001. Comparative affinity of duloxetine and venlafaxine for serotonin and norepinephrine transporters in vitro and in vivo, human serotonin receptor subtypes, and other neuronal receptors. Neuropsychopharmacology 25: 871–880. Cao, Q.R., J.S. Choi, Y. Liu, W.J. Xu, M. Yang, B.J. Lee, and J.H. Cui. 2013. A formulation approach for development of HPMCbased sustained release tablets for tolterodine tartrate with a low release variation. Drug Development and Industrial Pharmacy 39: 1720–1730. Chatlapalli, R., and B.D. Rohera. 1998. Physical characterization of HPMC and HEC and investigation of their use as pelletization aids. International Journal of Pharmaceutics 161: 179–193. Choi, H., Y. Oh, and C. Kim. 1998. In situ gelling and mucoadhesive liquid suppository containing acetaminophen: enhanced bioavailability. International Journal of Pharmaceutics 165: 23–32. Costa, P., J.M. Sousa Lobo, M.C. Gohel, and M.K. Panchal. 2001. Modeling and comparison of dissolution profiles. European Journal of Pharmaceutical Sciences 13: 123–133. Costa, P., and J.M. Sousa Lobo. 2001. Influence of dissolution medium agitation on release profiles of sustained-release tablets. Drug Development and Industrial Pharmacy 27: 811–817. Fishbain, D.A., J. Hall, A.L. Meyers, J. Gonzales, and C. Mallinckrodt. 2008. Does pain mediate the pain interference with sleep problem in chronic pain? Findings from the studies for management of diabetic peripheral neuropathic pain with duloxetine. Journal of Pain and Symptom Management 36: 639–647. Frohoff-Hulsmann, M.A., B.C. Lippold, and J.W. McGinity. 1999. Aqueous ethyl cellulose dispersion containing plasticizers of different water solubility and hydroxy propyl methyl-cellulose as coating material for diffusion pellets II: properties of sprayed films. European Journal of Pharmaceutics and Biopharmaceutics 48: 67–75. Gibaldi, M., and D. Perrier. 1982. Pharmacokinetics, 2nd ed. New York: Marcel Dekker.
123
Goldstein, D.J., Y. Lu, M.J. Detke, C. Wiltse, C. Mallinckrodt, and M.A. Demitrack. 2004. Duloxetine in the treatment of depression: A double-blind placebo-controlled comparison with paroxetine. Journal of Clinical Psychopharmacology 24: 389–399. Goyanes, A., and R. Martinez-Pacheco. 2013. New co-processed MCC-based excipient for fast release of low solubility drugs from pellets prepared by extrusion-spheronization. Drug Development and Industrial Pharmacy (Early online:1–7). Hausner, H.H. 1967. Friction conditions in a mass of metal powder. International Journal of Powder Metallurgy 3: 7–13. International Conference on Harmonization (ICH Q2A, Q2B), 1995. Guidelines on validation of analytical procedures. Jansen, P.J., P.L. Oren, C.A. Kemp, S.R. Maple, and S.W. Beartschi. 1998. Characterization of impurities formed by interaction of duloxetine HCl with enteric polymers hydroxypropyl methylcellulose acetate duccinate and hydroxypropyl methylcellulose phthalate. Journal of Pharmaceutical Sciences 87: 81–85. Korakianiti, E.S., D.M. Rekkas, P.P. Dallas, and N.H. Choulis. 2000. Optimization of the pelletization process in a fluid- bed rotor granulator using experimental design. AAPS Pharm SciTech 1: 71–75. Lecomte, F., J. Siepmann, M. Walther, R.J. MacRae, and R. Bodmeier. 2004. Polymer blends used for the coating of multiparticulates: Comparison of aqueous and organic coating techniques. Pharmaceutical Research 21: 882–890. Mastan, S., T.B. Latha, and S. Ajay. 2011. The basic regulatory considerations and prospects for conducting bioavailability/ bioequivalence (BA/BE) studies—An overview. Comparative Effectiveness Research 1: 1–25. Mease, P.J., I.J. Russell, D.K. Kajdasz, C.G. Wiltse, M.J. Detke, M.M. Wohlreich, D.J. Walker, and A.S. Chappell. 2010. Long term safety, tolerability, and efficacy of duloxetine in the treatment of fibromyalgia. Seminars in Arthritis and Rheumatism 39: 454–464. Mohamed, F.A., M. Roberts, L. Seton, J.L. Ford, M. Levina, and A.R. Rajabi-Siahboomi. 2013. The influence of HPMC concentration on release of theophylline or hydrocortisone from extended release mini-tablets. Drug Development and Industrial Pharmacy 38: 1167–1174. Robinson, D., and L. Cardozo. 2010. New drug treatments for urinary incontinence. Maturitas 65: 340–347. Sharma, A., M.J. Goldberg, and B.J. Cerimele. 2000. Pharmacokinetics and safety of duloxetine, a dual-serotonin and norepinephrine reuptake inhibitor. The Journal of Clinical Pharmacology 40: 161–167. Shravani, D., P.K. Lakshmi, and J. Balasubramaniam. 2011. Preparation and optimization of various parameters of enteric coated pellets using the Taguchi L9 orthogonal array design and their characterization. Acta Pharmaceutica Sinica B 1: 56–63. Sinha, V.R., R. Kumria, and J.R. Bhinge. 2009. Stress degradation studies on duloxetine hydrochloride and development of an RPHPLC method for its determination in capsule formulation. Journal of Chromatographic Science 47: 589–593. Valizadeh, H., P. Nayyeri-Maleki, S. Ghanbarzadeh, A. Sheikhloo, H. Servat, M. Nemati, and P. Zakeri-Milani. 2014. Pharmacokinetics and bioequivalence of two brands of metformin 500 mg tablets in Iranian healthy volunteers. Journal of Pharmaceutical Investigation 44: 61–68. Westanmo, A.D., J. Gayken, and R. Haight. 2005. Duloxetine: A balanced and selective norepinephrine-and serotonin-reuptake inhibitor. American Journal of Health-System Pharmacy 62: 2481–2490. Yong, C.S., D.X. Li, D.H. Oh, J.A. Kim, B.K. Yoo, J.S. Woo, J.D. Rhee, and H.G. Choi. 2005. Retarded dissolution of ibuprofen in
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride... gelatin microcapsule by cross-linking with glutaraldehyde. Archives of Pharmacal Research 29: 431–534. Yuksel, N., A.E. Kanik, and T. Baykara. 2000. Comparison of in vitro dissolution profiles by ANOVA-based, model-dependent and independent methods. International Journal of Pharmaceutics 209: 57–67.
Zhao, R.K., G. Cheng, J. Tang, J. Song, and W.X. Peng. 2009. Pharmacokinetics of duloxetine hydrochloride enteric-coated tablets in healthy Chinese volunteers: A randomized, open-label, single- and multiple-dose study. Clinical Therapeutics 31: 1022–1036.
123
RESEARCH ARTICLE
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride using a fluidized bed coater Yong-Il Kim1,2 • Roshan Pradhan1 • Bijay K. Paudel1 • Ju Yeon Choi1 Ho Taek Im2,3 • Jong Oh Kim1
•
Received: 3 January 2015 / Accepted: 7 March 2015 Ó The Pharmaceutical Society of Korea 2015
Abstract In this study, the enteric-coated delayed-release pellets of duloxetine hydrochloride (DLX) were formulated using a fluidized bed coater. Three separate layers, the drug layer, the barrier layer, and the enteric layer, were coated onto inert core pellets. Among the three formulations (F1– F3), the dissolution profiles of formulation F2 were most similar to those of the marketed product, with similarity and difference factors of 83.99 and 3.77, respectively. In addition, pharmacokinetic parameters of AUC, Cmax, Tmax, t1/2, Kel, and MRT of DLX for the developed formulation (F2) did not differ significantly from those for the marketed product in beagle dogs, suggesting that they were bioequivalent. Our results demonstrated that the in vitro dissolution data resembled the in vivo performance of the drug. Therefore, this study has a positive scope for further scale up and development of the formulation for achievement of the generic product. Keywords Duloxetine hydrochloride Delayed-release Enteric coated Pellet Pharmacokinetics
Yong-Il Kim and Roshan Pradhan contributed equally to this study. & Ho Taek Im [email protected] & Jong Oh Kim [email protected] 1
College of Pharmacy, Yeungnam University, 214-1, Dae-Dong, Gyongsan 712-749, Republic of Korea
2
Pharm. R&D Institute, Hanmi Pharm. Co., Ltd., Hwasung 445-913, Republic of Korea
3
School of Pharmacy, Sungkyunkwan University, Jangan-gu, Suwon 440-746, Gyeonggi-do, Republic of Korea
Introduction Duloxetine hydrochloride (DLX) is a novel dual serotonin and norepinephrine reuptake inhibitor, approved for treatment of major depressive disorders, diabetic peripheral neuropathic pain (Fishbain et al. 2008), fibromyalgia (Mease et al. 2010), generalized anxiety, and woman stress urinary incontinence (Robinson and Cardozo 2010). The claimed mechanism of action of the drug is based on the specific inhibition of both serotonin and norepinephrine reuptake while it weakly inhibits dopamine reuptake and has no significant affinity for histaminergic, dopaminergic, cholinergic or adrenergic receptors (Goldstein et al. 2004). DLX is a newer and more preferable antidepressant because of its favorable pharmacodynamic features viz. dual inhibition, tolerability, safety, faster recovery, fewer side effects, and low affinity for other neuronal receptors (Bymaster et al. 2001; Sharma et al. 2000; Westanmo et al. 2005). However, the drug was found to be acid labile, which results in its degradation in a gastric environment, thus necessitating the need for development of an entericcoated system (Shravani et al. 2011). Enteric-coated formulations are suitable vehicles for modification of the release of drug or active pharmaceutical ingredients at specific target areas within the gastrointestinal tract. Enteric coating is an effective method of protecting the drug against a gastric environment and preventing the release of the encapsulated particles or the drug before reaching the target site. DLX is unstable in solution at pH values less than 2.5, therefore, enteric polymer-coated formulations have been developed to prevent its acid degradation in the stomach and to provide for its subsequent release in the small intestine (Jansen et al. 1998; Sinha et al. 2009). Hydroxy propyl methyl cellulose acetate succinate (HPMC-AS) and hydroxy propyl methyl
123
Y. Kim et al.
cellulose phthalate (HPMC-P) are the major polymers used as an enteric-coating for DLX capsules. However, it has been reported that reaction of DLX with HPMC-AS and HPMC-P forms impurities like succinamide and phthalamide (Jansen et al. 1998). In order to prevent impurity formation between the core and the surrounding enteric polymer, the separate layer or middle barrier layer is coated. The type of coating technique strongly affects the film microstructure and thus affects the release mechanism and rate from pellets coated with polymer blends (Lecomte et al. 2004). Pellets have increased in importance over the years due to their distinctive advantages in both technological and therapeutic aspects (Korakianiti et al. 2000). When administered orally, therapeutic advantages include modification of drug release, division of dose strength, and free dispersion in the gastro intestinal tract (Abdul et al. 2010). In this study, DLX pellets were prepared with three separate coatings, consisting of the inner drug layer, middle barrier layer or sub-layer, and the outer enteric layer (Fig. 1). All three layers were coated using a fluidized bed coater. HPMC was used for the drug and barrier layering, while HPMC-P was used as the enteric polymer. The effect of the concentration of the enteric polymer on the dissolution of prepared formulations was examined. The in vitro release of DLX from the developed formulations was compared with that from the marketed CymbaltaÒ capsules (Lilly), and pharmacokinetic studies were performed in beagle dogs after oral administration of formulated DLX capsule and the marketed product.
Materials and methods Materials DLX was purchased from Zydus Pharmaceuticals (New Jersey, USA). Hydroxypropyl methylcellulose (HPMC 2910, PharmacoatÒ 606) and hydroxypropyl methylcellulose phthalate (HPMCP, HP-50) were obtained from ShinEtsu Chemical Co. (Tokyo, Japan), respectively. Fig. 1 Schematic illustration of DLX pellets
123
Fluoxetine, as an internal standard, was purchased from Sigma (St. Louis, MO, USA). Sugar beads (25–30 sieve size) were purchased from Hanns G. Werner GmbH Co. (Holstein, Germany). Sucrose, talc and triethyl citrate (TEC) were supplied by BASF Chemical Co. (Ludwigshafen, Germany). Solvents such as ethanol and acetone were purchased from OCI Company Ltd. (Seoul, South Korea). The commercial product (CymbaltaÒ) was purchased from a market. All other chemicals used were of reagent grade and used without further purification. Preparation of DLX pellets DLX pellets were prepared using a fluidized bed coater (XP-70). Using this technique, the polymeric solutions containing drug and other excipients were deposited onto the inert sugar beads. Details regarding the parameters of the fluidized bed coating process are shown in Table 1. The sugar beads were coated in three layers viz., the inner drug layer, middle barrier layer, and the outer enteric layer. HPMC was used both as the binding and the barrier polymer because of its low molecular weight and use as a good pelletization aid (Chatlapalli and Rohera 1998), while HP-50 was used as the enteric polymer. Inner drug layer Initially, the sugar beads, 42 mg per capsule (cap) were loaded into the fluidized bed and warmed before treatment with drug solution. The coating solution was prepared by first dissolving the HPMC (11 mg/cap) in distilled water (300 ml) followed by addition of DLX (33.7 mg/cap) with constant stirring. Talc (4.47 mg/cap) was dispersed and stirred in order to obtain a homogeneous dispersion. This coating solution was sprayed over the sugar beads at a spray rate of 5 ml/min. Middle barrier layer The primary function of this layer is to minimize the interaction between the inner drug layer and the outer enteric
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride... Table 1 Process parameters of the fluidized bed coating
Parameters
Inner drug layer
Middle barrier layer
Outer enteric layer
Nozzle size (mm)
1.0
1.0
1.0
Inlet temperature (°C)
55
45
31
Outlet temperature (°C)
40
30
29
Bed temperature (°C)
25
25
25
Spray rate (ml/min)
5
6
8
1.2
1.0
0.8
2
Automizing air pressure (kgf/cm )
Table 2 Composition of the outer enteric layer of DLX pellets Ingredients
F1 (mg)
F2 (mg)
F3 (mg)
DLX (barrier coated pellets)
121.17
121.17
121.17
HP-50
20
25
30
Triethyl citrate
4
5
6
Talc
2.4
3
3.6
Ethanol
100
125
150
Acetone
200
250
300
Total
147.57
154.17
160.77
index (CI) were calculated according to the equations given below: qtapped 0 Hausner s ratio ¼ qbulk ! qtapped qbulk 0 100% Carr s index ¼ qtapped where qbulk and qtapped are bulk and tapped densities of the enteric-coated DLX pellets. In vitro dissolution test
polymer. HPMC (6 mg/cap) was dissolved in distilled water (190 ml) followed by addition of sucrose (9.6 mg/cap) with continuous stirring. Talc (14.4 mg/cap) was dispersed and stirred until a homogeneous dispersion was obtained. This coating solution was sprayed over the drug layer at a spray rate of 6 ml/min. Outer enteric layer HP-50 (20–30 mg/cap), was dissolved in an ethanol/acetone (1:2 v/v) mixture. In order to improve the elasticity of the coating film, and to minimize film-forming temperature, TEC (5 mg/cap) was added as a plasticizer to the above mentioned solution (Frohoff-Hulsmann et al. 1999). Talc (3 mg/cap) was used as the anti-adherent. The coating solution was applied over the barrier layer at a spray rate of 8 ml/min. The optimized weight ratio of HP-50:TEC:talc was 5:1:0.6. The detail compositions of formulations with varying amounts of enteric polymer (HP-50) are shown in Table 2. Determination of flow properties of enteric-coated DLX pellets The flow property of the pellets is an important parameter for hard gelatin capsule filling. The formulated pellets were tested for bulk and tap density using the AS-100 tap density tester (AimSizer). Then, Hausner’s ratio (HR) and Carr’s
A dissolution test was performed on six capsules from each formulation containing different amounts of enteric polymer (HP-50). Dissolution profiles of the prepared formulations and the marketed product were determined in 0.1 N HCl and phosphate buffer (pH 6.8). The dissolution apparatus consisted of the USP XXIII, apparatus 1 (basket method) (VanKel VK 7000; Varian Inc., Palo Alto, CA, USA) set at a speed of 100 rpm, with 900 ml of dissolution medium, pre-warmed at 37 ± 0.5 °C. Initially, the capsules were exposed to 0.1 N HCl (pH 1.2) for 120 min, followed by replacement of the medium with phosphate buffer (pH 6.8). 2 ml of dissolution samples were collected after 5, 10, 15, 30, 45, 60, 90 and 150 min, followed immediately by addition of an equal volume of fresh dissolution media maintained at the same temperature to keep the volume of dissolution media constant and to maintain the sink condition (Choi et al. 1998; Yong et al. 2005). The withdrawn samples were filtered through a membrane filter (0.45 lm) and after suitable dilution with distilled water, analyzed by injecting 50 ll into the HPLC, as described below. The percent drug release was then graphed against time and the release profiles were studied. HPLC conditions The HPLC analysis system (Hitachi Co., Tokyo, Japan) consisted of a Hitachi D-7000 interface, a pump (Model L-7100), ultraviolet detector (Model L-7400), autosampler (Model L-7200), and a column oven (Model L-7300). A C8 analytical column (ACEÒ (ACT): 3 lm, 75 mm 9 4.6 mm,
123
Y. Kim et al.
GL Sciences Inc., Tokyo, Japan) was used. The mobile phase consisted of tetrahydrofuran (THF): methanol: pH 5.5 phosphate buffer (9:32.3:58.7, volume ratio). The effluent was monitored at a UV absorption wavelength of 288 nm, and a flow rate of 1.5 ml/min. The column oven was maintained at 45 °C. The analytical method was validated according to ICH recommendations for validation of analytical methods (ICH Q2A, Q2B 1995). Similarity factor (f2) and difference factor (f1) Similarity (f2) and difference factors (f1) measure the closeness between the two dissolution profiles. f2 and f1 were calculated according to the equations given below: 9 8" #0:5 n = < 1X f2 ¼ 50 log 1 þ ðRj TjÞ2 100 ; : n t¼1 2P n
R j Tj
6t¼1 f1 ¼ 6 n 4 P
Rj
3 7 7 100 5
t¼1
where n is the number of time points, Rj and Tj are the dissolution values of the reference product and the test product, respectively, at each time point j. In order to consider the dissolution profiles similar, f1 values should be close to 0 and f2 values should be close to 100. In general, f1 values lower than 15 (0–15) and f2 values higher than 50 (50–100) indicate similarity of the dissolution profiles (Costa et al. 2001). Pharmacokinetic study In vivo experiments A pharmacokinetic study was carried out in ten beagle dogs, which were subjected to fasting overnight at least 10 h prior to dosing with DLX. One capsule (optimized formulation) or capsule (CymbaltaÒ) containing DLX 30 mg was administered orally. 5 ml of blood was collected into heparin-vacutainer tubes from the forearm vein before administration (at 0 h) and at predetermined time intervals of 0.5, 1, 2, 3, 4, 5, 6, 8, 12, 18, and 24 h after oral administration. Blood samples were centrifuged at 30009g for 10 min and 0.2 ml aliquots of plasma were stored at -80 °C until further analysis. All animal-handling procedures were in accordance with the protocols approved by the Institutional Animal Ethical Committee, Yeungnam University, South Korea. Blood sample analysis using LC–MS/MS The concentrations of DLX in dog plasma were analyzed by the reported LC–MS/MS method (Zhao et al. 2009).
123
Briefly, 20 ll of acetonitrile containing the working internal standard (fluoxetine, 40 ng/ml) was added to a 100 ll aliquot of plasma sample, followed by vortexing for 2 min. After centrifugation of the sample at 12,0009g for 10 min, the organic layer was collected and then separated on an Atlantis C18 column (3 lm, 50 mm 9 2.1 mm, Waters, Milford, MA, USA) using isocratic elution with acetonitrile and 30 mM ammonium formate solution maintained at pH 5 (90:10, volume ratio) at a flow rate of 0.3 ml/min. The column and autosampler tray temperatures were 35 and 15 °C, respectively. The eluent was introduced directly into the positive ionization electrospray source of a tandem quadrupole mass spectrometer (Quattro Micro; Micromass Manchester, UK). The ion source and desolvation temperatures were held at 100 and 400 °C, respectively. The optimum cone voltages and collision energy were 10 V and 6 eV, and 15 V and 18 eV for duloxetine and fluoxetine, respectively. Selected reaction monitoring (SRM) using precursor product ion transitions of m/z 298.08 [ 154 and m/z 310.02 [ 148.07 were used to quantify duloxetine and fluoxetine, respectively. Linear calibration curves were obtained for the concentration range of 0.1–100 ng/ml. The lower limit of quantification (LOQ) of DLX was 0.1 ng/ml. The within-day and between-day coefficients of variation were generally less than 10 %. Data acquisition and processing were accomplished using MasslynxTM 4.1 with a QuanLynxTM application manager. Pharmacokinetic analysis Standard methods (Gibaldi and Perrier 1982) were used in calculation of the pharmacokinetic parameters such as area under the drug-concentration time-curve (AUC), half-life (t1/2), elimination rate constant (Kel), and mean residence time (MRT) using non-compartmental analysis (WinNonlin 3.1TM; Pharsight Co., Mountain View, CA, USA). The maximum plasma concentration of drug (Cmax) and time to reach Cmax (Tmax) were directly computed from the plasma concentration versus time plot. All data are expressed as mean ± SD. Furthermore, the relative bioavailability (F) with reference to the AUC of the developed formulation compared to the marketed product was calculated using the equations given below: AUC A F¼ 100 % AUC B where F is the relative bioavailability, AUCA and AUCB are the area under drug-concentration time-curve of developed formulation (test) and marketed product (reference) respectively.
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride...
Statistical analysis Non-linear regression analysis was performed using the Statistical Package for Social Sciences (SPSS) computer program to compare the mechanism of drug release of the different formulations. Comparison of the pharmacokinetic parameters of the developed formulation and the marketed product were made using Student’s t test at a 95 % level of confidence using Microsoft Office Excel 2007. All data were expressed as the mean ± SD.
Results and discussion Preparation and characterization of enteric-coated DLX pellets Different formulations of enteric-coated DLX pellets (F1– F3) with different amounts of HP-50, as the enteric coating polymeric agent were designed and prepared using the fluidized bed coating method, based on the composition shown in Table 2. All of the formulated products lay within the limit of 98–102 % for the drug content (data not shown). To ensure good flow property, bulk density and tap density were measured, and HR and CI were calculated. The results obtained are presented in Table 3. Although both the HR and CI indicate the flow characteristics of the powder, granules or pallets, specifically CI is a measure of powder bridge strength and stability, while the HR is a measure of the interparticulate friction. Materials exhibiting lower values of CI or HR indicate better flow properties than higher values (Hausner 1967). In our study, HR of all of the formulated products was within 1.0 to 1.11 and CI was less than 10 %. Thus, all the pellet formulations showed excellent flow characteristics indicating uniformity of size and good sphericity of the pellets (Goyanes and Martinez-Pacheco 2013). The sieve analysis revealed that almost 95 % of pellets were distributed between 800–900 lm so pellets size distribution was found uniform. Since the size of the pellets depends upon the coating thickness, the average size of pellets (F1–F3) was found to be 820 ± 4 lm, 860 ± 5 lm 890 ± 8 lm respectively, which was similar with pellet size of the commercial product. Eventually, the free flowing enteric-coated pellets
Table 3 Flow properties of enteric-coated DLX pellets
equivalent to 30 mg of DLX were accurately weighed and filled into the hard gelatin capsules and were used for in vitro dissolution study. In vitro dissolution studies of enteric-coated DLX pellets The in vitro dissolution profiles of DLX from all formulations (enteric-coated DLX pellets) and the marketed product in 0.1 N HCl (pH 1.2) and phosphate buffer (pH 6.8) are shown in Fig. 2. In 0.1 N HCl (pH 1.2), formulation F1 exhibited a release of approximately 3 % in 120 min as well as a faster release pattern (approximately 80 % in 60 min) of DLX in phosphate buffer (pH 6.8) as compared to formulations F2, F3, and the marketed product, respectively. This may be attributed to the low amount of enteric polymers present in the formulation (Cao et al. 2013; Mohamed et al. 2013; Shravani et al. 2011), which could not resist the release of drug in the acidic dissolution medium. On the other hand, formulations F2, F3, and the marketed product showed no release of drug in 0.1 N HCl (pH 1.2) in 120 min indicating the effectiveness of the enteric coating polymer. Therefore, the result was in close agreement with the statement that the concentration of enteric polymer plays a primary role in acid-resistant pellets since there was an increased resistance to acid with an increase in the concentration of the enteric polymer. In pH 6.8, formulations F2 and F3 showed drug release of approximately 70 and 65 % in 60 min, respectively, which may be attributed to the higher amount of enteric polymers present in the formulation. These results indicated that as the amount of HP-50 increased, the release rate showed a gradual decrease. In fact, the extent of dissolution depends on the degree of ionization. The polymer should be ionized to be dissolved. Once the enteric polymer gets dissolved, the barrier polymer is another factor that affects the drug release from the pellets since it is the next layer after enteric polymer that obstructs the drug release. In addition, the increase in the concentration of the barrier polymer may increase the extent of obstruction of drug release. Furthermore, in order to investigate the effect of the rate of rotation on the drug release profile, formulations F1, F2, F3 and the marketed product were subjected to paddle speeds of 50, 100, and 150 rpm in 0.1 N HCl (pH 1.2) and
Formulations
Bulk density (g/cm3)
Tapped density (g/cm3)
Hausner ratio (HR)
Carr’s index (CI) (%)
F1
0.86 ± 0.02
0.88 ± 0.04
1.02
2.27
F2
0.87 ± 0.03
0.91 ± 0.03
1.04
4.39
F3
0.89 ± 0.02
0.94 ± 0.04
1.06
5.32
Each value represents the mean. (n = 3)
123
Y. Kim et al.
results, formulation F2 was considered the best formulation and was selected for the pharmacokinetic study in beagle dogs. Pharmacokinetic study of enteric-coated delayedrelease DLX pellets
Fig. 2 Dissolution profiles of DLX from different formulations (F1– F3) of enteric-coated DLX pellets and marketed product in 0.1 N HCl (pH 1.2) for 120 min and subsequently in phosphate buffer (pH 6.8) for 150 min. Dissolution was performed at 37 ± 0.5 °C and 100 rpm. Each value represents the mean ± SD (n = 6)
subsequently in phosphate buffer (pH 6.8) as the dissolution media; the comparative results are shown in Fig. 3. As reported, in vitro drug release for a polymeric system is affected by the dissolution medium and the rate of rotation (Costa and Sousa Lobo 2001). Our results showed that the drug release from all three formulations was not similar, but in fact increased with increasing rate of rotation (similarity factor \50; difference factor [15), indicating that the drug release from the enteric-coated pellets was affected by the rate of rotation. The similarity factor (f2) and the difference factor (f1) were calculated for all formulations using the release profile of the marketed product as a reference (Table 4). Also the f2 and f1 values of all three formulations at different rates of rotation with reference to 50 rpm are shown in Table 5. As shown in Table 4, the f2 value of formulation F1 was 46.67 (\50), and the f1 value was 20.75 (greater than 15), respectively which suggested that the dissolution profile of F1 was not similar to that of the marketed product. The dissimilarity may be attributed to the presence of a low amount of HP-50 in formulation F1, which resulted in more rapid release of dissolution profiles compared to the reference product. In contrast, formulations F2 and F3 had f2 values greater than 50 (83.99 and 65.48, respectively) and f1 values less than 15 (3.77 and 8.78, respectively), indicating that their dissolution profiles were similar to that of the marketed product (Costa et al., 2001; Yuksel et al., 2000). Among all formulations tested, formulation F2 showed the highest f2 value (83.99) and the lowest f1 value (3.77), corresponding to the highest similarity between its release profile and that of the reference product (CymbaltaÒ capsules). Considering all of the
123
The pharmacokinetic study in beagle dogs (n = 10) after oral administration of formulation F2 and the marketed product was performed and the mean plasma concentration–time curve was constructed. The dose administered was 30 mg DLX. As shown in Fig. 4, the developed formulation F2 exhibited almost similar pharmacokinetic profile with almost overlapping mean plasma concentration–time curves. A list of the corresponding pharmacokinetic parameters is shown in Table 6. The AUC (ng.h/ ml) and Cmax (ng/ml) values of developed formulation F2 were 147.2 ± 79.2 and 45.5 ± 17.9, respectively, while those of marketed product were 148.61 ± 88.16 and 46.80 ± 23.11, respectively. When these values were evaluated using student’s t test, they were not significantly different with each other (p \ 0.05). In addition, other pharmacokinetic parameters including Tmax, t1/2, elimination rate constant (kel), and mean retention time (MRT) were not significantly different. These results indicated that the pharmacokinetic profiles of the two formulations (test and reference) were very similar with each other. In addition, when the relative bioavailability (F) was calculated with reference to the AUC of the developed formulation F2 and the marketed product, it was found to be 99.0 %, which further indicated that the two formulations were bioequivalent (Mastan et al. 2011; Valizadeh et al. 2014). Therefore, our results demonstrated that there was a good correlation between the in vitro dissolution and the in vivo pharmacokinetic study with reference to the enteric-coated delayed-release DLX pellets. In conclusion, the enteric-coated delayed-release pellets of DLX (equivalent to 30 mg of DLX) were successfully prepared by a fluidized bed coating process, using HPMC as the binding and barrier coating and HP-50 as the enteric coating polymer. Among the three formulations (F1-F3), the dissolution profile of formulation F2 was most similar to the marketed product with similarity and difference factors of 83.99 and 3.77, respectively. Moreover, results of the pharmacokinetic study performed in beagle dogs after oral administration of prepared enteric-coated DLX pellets and the marketed product showed that the pharmacokinetic parameters, including AUC, Cmax, Tmax, t1/2, Kel, and MRT were not significantly different from those of the marketed product, suggesting that they were bioequivalent. Therefore, this study has a positive scope for further scale up and development of the formulation for achievement of the generic product.
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride...
Fig. 3 Effect of rate of rotation on the drug release profile of formulations F1, F2, F3, and marketed product in 0.1 N HCl (pH 1.2) and subsequently in phosphate buffer (pH 6.8) at a 50 rpm, b 100 rpm, and c 150 rpm. Dissolution was performed at 37 ± 0.5 °C. Each value represents the mean ± SD (n = 6)
Table 4 Comparison of dissolution profiles of enteric-coated DLX pellets with the profile of the marketed product Formulations
Similarity factor (f2)
Difference factor (f1)
F1
46.67
20.75
F2
83.99
3.77
F3
65.48
8.78
Table 5 Comparison of dissolution profiles of enteric-coated DLX pellets at different rates of rotation with reference to 50 rpm Formulations
100 rpm
150 rpm
f2
f1
f2
f1
F1
47.62
19.10
34.66
30.98
F2
48.34
21.58
33.89
36.09
F3
43.14
30.26
33.24
41.93
Fig. 4 Pharmacokinetic profile of DLX in beagle dogs after oral administration of marketed product (filled circle) and developed formulation (F2) (empty circle) at a single dose of 30 mg of DLX. Each value represents the mean ± SD (n = 10)
123
Y. Kim et al. Table 6 Pharmacokinetic parameters of DLX delivered by marketed formulation and developed formulation F2 after oral administration to beagle dogs Parameters
Marketed product
Developed formulation (F2)
AUC (ng.h/ml)
148.61 ± 88.16
147.18 ± 79.16
46.80 ± 23.11
45.54 ± 17.99
2.00 ± 0.48 3.16 ± 1.14
1.86 ± 0.69 3.74 ± 0.97
Cmax (ng/ml) Tmax (h) t1/2 (h) Kel (h-1)
0.25 ± 0.09
0.19 ± 0.06
MRT (h)
3.60 ± 0.82
3.67 ± 1.11
Each value represents the mean ± SD (n = 10)
Acknowledgments This study was supported by a grant from the Medical Cluster R&D Support Project of Daegu Gyeongbuk Medical Innovation Foundation, Republic of Korea (2013) (No. HT13C0011).
References Abdul, S., A.V. Chandewar, and S.B. Jaiswal. 2010. A flexible technology for modified-release drugs: Multiple-unit pellet system (MUPS). Journal of Controlled Release 147: 2–16. Bymaster, F.P., L.J. Dreshfield-Ahmad, P.G. Threlkeld, J.L. Shaw, L. Thompson, D.L. Nelson, S.K. Hemrick-Luecke, and D.T. Wong. 2001. Comparative affinity of duloxetine and venlafaxine for serotonin and norepinephrine transporters in vitro and in vivo, human serotonin receptor subtypes, and other neuronal receptors. Neuropsychopharmacology 25: 871–880. Cao, Q.R., J.S. Choi, Y. Liu, W.J. Xu, M. Yang, B.J. Lee, and J.H. Cui. 2013. A formulation approach for development of HPMCbased sustained release tablets for tolterodine tartrate with a low release variation. Drug Development and Industrial Pharmacy 39: 1720–1730. Chatlapalli, R., and B.D. Rohera. 1998. Physical characterization of HPMC and HEC and investigation of their use as pelletization aids. International Journal of Pharmaceutics 161: 179–193. Choi, H., Y. Oh, and C. Kim. 1998. In situ gelling and mucoadhesive liquid suppository containing acetaminophen: enhanced bioavailability. International Journal of Pharmaceutics 165: 23–32. Costa, P., J.M. Sousa Lobo, M.C. Gohel, and M.K. Panchal. 2001. Modeling and comparison of dissolution profiles. European Journal of Pharmaceutical Sciences 13: 123–133. Costa, P., and J.M. Sousa Lobo. 2001. Influence of dissolution medium agitation on release profiles of sustained-release tablets. Drug Development and Industrial Pharmacy 27: 811–817. Fishbain, D.A., J. Hall, A.L. Meyers, J. Gonzales, and C. Mallinckrodt. 2008. Does pain mediate the pain interference with sleep problem in chronic pain? Findings from the studies for management of diabetic peripheral neuropathic pain with duloxetine. Journal of Pain and Symptom Management 36: 639–647. Frohoff-Hulsmann, M.A., B.C. Lippold, and J.W. McGinity. 1999. Aqueous ethyl cellulose dispersion containing plasticizers of different water solubility and hydroxy propyl methyl-cellulose as coating material for diffusion pellets II: properties of sprayed films. European Journal of Pharmaceutics and Biopharmaceutics 48: 67–75. Gibaldi, M., and D. Perrier. 1982. Pharmacokinetics, 2nd ed. New York: Marcel Dekker.
123
Goldstein, D.J., Y. Lu, M.J. Detke, C. Wiltse, C. Mallinckrodt, and M.A. Demitrack. 2004. Duloxetine in the treatment of depression: A double-blind placebo-controlled comparison with paroxetine. Journal of Clinical Psychopharmacology 24: 389–399. Goyanes, A., and R. Martinez-Pacheco. 2013. New co-processed MCC-based excipient for fast release of low solubility drugs from pellets prepared by extrusion-spheronization. Drug Development and Industrial Pharmacy (Early online:1–7). Hausner, H.H. 1967. Friction conditions in a mass of metal powder. International Journal of Powder Metallurgy 3: 7–13. International Conference on Harmonization (ICH Q2A, Q2B), 1995. Guidelines on validation of analytical procedures. Jansen, P.J., P.L. Oren, C.A. Kemp, S.R. Maple, and S.W. Beartschi. 1998. Characterization of impurities formed by interaction of duloxetine HCl with enteric polymers hydroxypropyl methylcellulose acetate duccinate and hydroxypropyl methylcellulose phthalate. Journal of Pharmaceutical Sciences 87: 81–85. Korakianiti, E.S., D.M. Rekkas, P.P. Dallas, and N.H. Choulis. 2000. Optimization of the pelletization process in a fluid- bed rotor granulator using experimental design. AAPS Pharm SciTech 1: 71–75. Lecomte, F., J. Siepmann, M. Walther, R.J. MacRae, and R. Bodmeier. 2004. Polymer blends used for the coating of multiparticulates: Comparison of aqueous and organic coating techniques. Pharmaceutical Research 21: 882–890. Mastan, S., T.B. Latha, and S. Ajay. 2011. The basic regulatory considerations and prospects for conducting bioavailability/ bioequivalence (BA/BE) studies—An overview. Comparative Effectiveness Research 1: 1–25. Mease, P.J., I.J. Russell, D.K. Kajdasz, C.G. Wiltse, M.J. Detke, M.M. Wohlreich, D.J. Walker, and A.S. Chappell. 2010. Long term safety, tolerability, and efficacy of duloxetine in the treatment of fibromyalgia. Seminars in Arthritis and Rheumatism 39: 454–464. Mohamed, F.A., M. Roberts, L. Seton, J.L. Ford, M. Levina, and A.R. Rajabi-Siahboomi. 2013. The influence of HPMC concentration on release of theophylline or hydrocortisone from extended release mini-tablets. Drug Development and Industrial Pharmacy 38: 1167–1174. Robinson, D., and L. Cardozo. 2010. New drug treatments for urinary incontinence. Maturitas 65: 340–347. Sharma, A., M.J. Goldberg, and B.J. Cerimele. 2000. Pharmacokinetics and safety of duloxetine, a dual-serotonin and norepinephrine reuptake inhibitor. The Journal of Clinical Pharmacology 40: 161–167. Shravani, D., P.K. Lakshmi, and J. Balasubramaniam. 2011. Preparation and optimization of various parameters of enteric coated pellets using the Taguchi L9 orthogonal array design and their characterization. Acta Pharmaceutica Sinica B 1: 56–63. Sinha, V.R., R. Kumria, and J.R. Bhinge. 2009. Stress degradation studies on duloxetine hydrochloride and development of an RPHPLC method for its determination in capsule formulation. Journal of Chromatographic Science 47: 589–593. Valizadeh, H., P. Nayyeri-Maleki, S. Ghanbarzadeh, A. Sheikhloo, H. Servat, M. Nemati, and P. Zakeri-Milani. 2014. Pharmacokinetics and bioequivalence of two brands of metformin 500 mg tablets in Iranian healthy volunteers. Journal of Pharmaceutical Investigation 44: 61–68. Westanmo, A.D., J. Gayken, and R. Haight. 2005. Duloxetine: A balanced and selective norepinephrine-and serotonin-reuptake inhibitor. American Journal of Health-System Pharmacy 62: 2481–2490. Yong, C.S., D.X. Li, D.H. Oh, J.A. Kim, B.K. Yoo, J.S. Woo, J.D. Rhee, and H.G. Choi. 2005. Retarded dissolution of ibuprofen in
Preparation and evaluation of enteric-coated delayed-release pellets of duloxetine hydrochloride... gelatin microcapsule by cross-linking with glutaraldehyde. Archives of Pharmacal Research 29: 431–534. Yuksel, N., A.E. Kanik, and T. Baykara. 2000. Comparison of in vitro dissolution profiles by ANOVA-based, model-dependent and independent methods. International Journal of Pharmaceutics 209: 57–67.
Zhao, R.K., G. Cheng, J. Tang, J. Song, and W.X. Peng. 2009. Pharmacokinetics of duloxetine hydrochloride enteric-coated tablets in healthy Chinese volunteers: A randomized, open-label, single- and multiple-dose study. Clinical Therapeutics 31: 1022–1036.
123
