Formulation and pharmacokinetics of multi-layered matrix tablets: Biphasic delivery of diclofenac Academic research paper on "Chemical sciences"

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Formulation and pharmacokinetics of multi-layered matrix tablets: Biphasic delivery of diclofenac

Ehab Mostafa Elzayat, Ali Abdelzaher Abdel-Rahman, Sayed Mohamed Ahmed, Fars Kaed Alanazi, Walid Abdulazim Habib, Hisham Suliman Abou Auda, Adel Sakr

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S1319-0164(16)30109-8 http://dx.doi.org/10.1016/j.jsps.2016.10.004 SPJ 525

Saudi Pharmaceutical Journal

14 December 2015 31 May 2016 7 October 2016

Pharmaceutical Journal

Please cite this article as: Mostafa Elzayat, E., Abdelzaher Abdel-Rahman, A., Mohamed Ahmed, S., Kaed Alanazi, F., Abdulazim Habib, W., Suliman Abou-Auda, H., Sakr, A., Formulation and pharmacokinetics of multi-layered matrix tablets: Biphasic delivery of diclofenac, Saudi Pharmaceutical Journal (2016), doi: http://dx.doi.org/ 10.1016/j.jsps.2016.10.004

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Formulation and pharmacokinetics of multi-layered matrix tablets: Biphasic delivery of diclofenac

Ehab Mostafa Elzayata1, Ali Abdelzaher Abdel-Rahman2, Sayed Mohamed Ahmed2, Fars Kaed Alanazia,b, Walid Abdulazim Habiba, Hisham Suliman Abou-Auda3, Adel Sakr4

a Kayyali chair, Department of Pharmaceutics, College of Pharmacy, King Saud University, Riyadh 11451, P.O. Box: 2457, Saudi Arabia.

b Supervisor of Kayyali Research Chair for Pharmaceutical Industries, Riyadh, Saudi Arabia.

i.edu.sa. Cellular no.: +966 (54) 8783477, Fax no.:

2 Department of Industrial Pharmacy. College of Pharmacy. Assiut University. Assiut. Egypt.

3 Department of Clinical Pharmacy. College of Pharmacy. King Saud University. Riyadh. KSA.

4 Department of Industrial Pharmacy. College of Pharmacy. Future University in Egypt. Cairo.

Abstract

The rapid availability of the drug at the site of action followed by maintaining its effect for a long period of time is of great clinical importance. Thus, the purpose of the present study was to prepare and evaluate multi-layered matrix tablets of diclofenac using Eudragit blend to achieve both immediate and sustained therapeutic effects. Diclofenac potassium (25mg) was incorporated in an outer immediate release layer to provide immediate pain relief whereas; diclofenac sodium (75mg) was incorporated in the inner core to provide extended drug release. Wet granulation was employed to prepare the inner core of the tablets which were further layered with an immediate release drug layer in the perforated pan coater. The in-vitro and in-vivo performance of the developed formulation were compared with the marketed products Voltaren® SR 75mg and Cataflam® 25mg. The in-vitro drug release of the prepared formulation showed similarity (f2 = 66.19) to the marketed product. The pharmacokinetic study showed no significant difference (p>0.05) in AUCo-24 and Cmax between the test and reference formulations. The AUC0-24 values were 105.36±83.3 and 92.87±55.53 ^g.h/ml whereas; the Cmax values were 11.25±6.87 and 12.97±8.45 ^g/ml, for the test and reference, respectively. The multi-layered tablets proved to be bioequivalent with the commercially available tablets and in agreement with the observed in-vitro drug release results. Stable physical characteristics and drug release profiles were observed in both long term and accelerated conditions stability studies.

KEYWORDS: Diclofenac; Matrix; Pharmacokinetic; Bi-phasic release; In-vivo; Multi-layered tablets

1. Introduction

Diclofenac Sodium (DS) (sodium;2-[2-(2,6-dichloroanilino) phenyl]acetate) is a potent non-steroidal anti-inflammatory drug (NSAID) and one of the most useful pain killers; used clinically in treatment of rheumatic or non-rheumatic inflammatory diseases. Diclofenac inhibit the conversion of arachidonic acid to prostaglandins which are responsible for the inflammatory process (Fini et al., 2007; Kovala-Demertzi, 2006).

The extended release (ER) tablets containing 75mg of diclofenac sodium are administered as once daily to guarantee less frequent dosing and diminished occurrence of gastro-intestinal side effects (Shaw et al., 2005). Different release matrices have been proposed for the preparation of diclofenac ER formulations (González--RodríGuez et al., 2003; Su et al., 2003) using different polymers including HPMC with various grades (Mourao et al., 2010) and Eudragits (González-RodríGuez et al., 2003; Pignatello et al., 2002). In most therapies, such ER preparations are desirable. However, in certain cases, rapid availability of the drug as a loading dose is needed to relieve the symptoms of the disease, followed by a maintenance dose of an effective drug plasma level for the continuation of the clinical effects. The combination of different drug delivery forms (immediate, enteric or sustained) of diclofenac in one pharmaceutical formulation has been explored (Lim et al., 2012; Sawayanagi and Otani, 1990; Shaw, 2012). Marketed pharmaceutical products, namely Diclofenac Duo® 75mg (PharmaSwiss)

or Divido® 75 mg (Tabuk Pharmaceuticals), contains two drug delivery forms of the drug (25 mg diclofenac sodium in gastro-resistant form, 50 mg diclofenac sodium in prolonged-release form) in each hard gelatin capsule. Fast release of the drug from such formulations is not substantial. Release of the drug from the prolonged release form may startin the stomach but the amount

released is not adequate to achieve a rapid pharmacologic response because of the low acidic solubility of DS. In addition, the lag time associated with the gastro-resistant form of the drug will contribute to the delayed response. Such preparations are not suitable for initiating treatment of conditions where rapid onset of action is needed. Accordingly, in this study, development of

multi-layered tablet that combines two drug delivery forms (immediate and sustained) using two salt forms of diclofenac was pursued. Diclofenac potassium (DP), with a relatively higher solubility in water and faster dissolution and absorption rate than observed with the sodium salt (Ahmad et al., 2010; Altman et al., 2015), was used to achieve prompt pain relief by incorporating it into an immediate release, fast dissolving outer layer, whereas the sodium salt with of its lower solubility was incorporated into an insoluble matrix inner core to be released slowly and provide a sustained analgesic effect. This kind of formulation is expected to add a clinical and therapeutic value, especially for patients taking multiple drugs. The multi-layered tablets were prepared and assessed for their in-vitro behavior, in-vivo performance and stability in this research.

2. Materials and methods 2.1. Materials

DS & DP were kindly supplied by TABUK Pharmaceuticals (Tabuk, Saudi Arabia). Eudragit RL PO and Eudragit RS PO were obtained from Evonik (Germany). Granulac 200® (lactose monohydrate) was kindly supplied by Meggle Pharma (Wassenberg, Germany). Vivapur® PH 101 (microcrystalline cellulose or MCC) was purchased from JRS Pharma (Weissenborn,Germany). All other chemicals were of pharmaceutical grade.

2.2. Methods

2.2.1. Preparation of the inner core matrix tablets

A high shear mixer was used to prepare tablets by a wet-granulation technique. In preliminary experiments, the polymer concentration and compression force were optimized using a two factor, three levels (32) full factorial design to obtain tablets matching the release profile of Voltaren® SR 75mg and at the same time matching USP acceptance criteria for extended release diclofenac tablets (Table 2). Each tablet of the optimized formulation (designated as Formulation E) contains 75mg of DS, 52.5mg of a 1:1 w/w blend of Eudragit RL and Eudragit RS, 3.5mg of magnesium stearate, 1.75mg of colloidal silicon dioxide and a sufficient quantity of a 70:30 w/w filler blend consisting of a mixture of lactose monohydrate and MCC to achieve a final tablet weight of 350mg. Each 600 g of the powder blend specified in the experimental designwas sieved through a 0.63mm mesh screen and then mixed in the high shear mixer (chopper speed = 700 rpm, impreller speed = 200 rpm) for 20 min. A 20% w/v ethanolic solution of a 1:1 w/w mixture of Eudragit RL and Eudragit RS was used for granulation of the powder blend in the high shear mixer with the chopper speed at 2500 rpm, the impeller speed at 200 rpm, the spray rate at 46 ml/min, and the wet massing time set at 5 min to obtain the desired mass consistency. The amount of the polymer in the granulating solution was subtracted from the total amount of the polymer assigned for the formulation. The wetted mass was sieved through a 1.25mm aperture mesh. Drying of the granules was carried out in a hot air oven (Memmert, Tv80ul, Germany) for 1 h at 40oC. The percent of the residual solvent, 2-3% was calculated using an infrared dryer (Mettler, LP16 Greifensee, Switzerland). The dried granules were passed through a 0.8mm mesh screen. A mixture of 1% w/w magnesium stearate and 0.5% w/w colloidal silicon

dioxide was added and mixed in the high shear mixer (chopper speed = 400 rpm, impeller speed = 200 rpm for 5 min). The tablets were compressed in Rotab T tablet press (Kg-Pharma, Germany) using standard concave punches of 10 mm diameter at 15 kN.

2.2.2. Drug layering and coating of the prepared matrix tablets

Layering of tablets with a layer of DP mixture was performed using a lab scale perforated

pan mini coater (Glatt, Pratteln, Switzerland). Each of the 450g batches of tablets was loaded into the coating pan. The drug layering solution consisted of 10 % w/w solid content of DP/Opadry II 3:1 in distilled water. The solution was sprayed using the following conditions: air volume 25 m3, inlet temperature 78oC, outlet temperature 43oC, spray rate 4 g/min, pump speed 25 rpm, atomization air 1.5 bar, air pattern 1.5 bar, pan speed 20 rpm and drying time 15 min. The spraying process continued until achieving a target tablet mass of approximately 383.3 mg, equivalent to a drug load of 25mg DP. An extra outer protective film using a 20% w/w solution of Opadry II was applied to add 5% w/w of the total tablet weight to each tablet to act as a taste masking and moisture protection layer. The coating conditions were the same as for the drug layering process except that the inlet temperature was 60oC and the outlet temperature was 45oC.

2.2.3. Assay for drug content

Drug content uniformity in the tablets was evaluated in accordance with USP guidelines. Ten tablets were taken individually and weighed, crushed, and then the drug was extracted with methanol. The solution was filtered, and samples were injected into an ACQUITY™ UPLC system (Waters Corp., Milford, MA, USA). Briefly, separation was accomplished using reversed-phase isocratic elution using a 50:50 v/v mixture of 0.05 M acetate buffer (pH 2.5) and acetonitrile as the mobile phase at 0.5 ml min-1 and an injection volume of 1 |il. The photodiode

array detector was set to acquire two-dimensional data from 210 to 280 nm; the second channel was recording at 254 nm. The column temperature was set at 50oC while sample temperature was kept at 10oC. This method was also used to measure DS content in DS-containing samples as well as dissolution samples (Elzayat et al., 2013).

To determine the amount of DP alone, analysis for the amount of potassium present was

e photo

carried out by flame photometry against a calibration curve of potassium chloride in aqueous solution. The exact amount of DP was back calculated from the flame photometry results.

2.2.4. In- vitro release studies

A USP Type II dissolution apparatus (LOGAN Instrument Corp., Somerset, NJ), was used to conduct the release studies. The dissolution medium was 900 ml 0.05 M phosphate buffer (pH = 7.5) kept at 37oC for 24 h with paddle rotating at 50 rpm. Samples of 5 ml each were collected at predetermined time intervals and replaced immediately with freshly prepared dissolution medium. The collected samples were filtered and analyzed by injection into the UPLC (see section 2.2.3). Release studies were conducted in triplicate and the average value of the cumulative % drug release was plotted against the time interval. The release profiles for the matrix tablets before coating were compared to that of Voltaren® 75mg. After drug layering and coating of these tablets, the release profiles obtained for these tablets were compared to Voltaren® 75mg and Cataflam® 25mg when these two commercial products were placed together

ne dissol

in one dissolution vessel.

2.2.5. Modeling of the in-vitro release data

The mechanism of drug release from formulations was elucidated by fitting the data to zero & first order, Higuchi and Korsmeyer-Peppas models (Elzayat et al., 2016).

The release efficiency (RE) was calculated from the fractional area under the curve at each time t, as determined using the trapezoidal rule (Khan, 1975). The mean dissolution times (MDT) were calculated according to the following equation (Costa and Sousa Lobo, 2001):

2?-! t/AM/

MD T = -j eqn. 1

2 % 1 ■ A M j

where j refers to the number of the sample, n refers to the number of release time points, tj refers to the time midway between tj _1 and tj , AMj refers to the extra amount of drug released between and .

The RE and the MDT values were analyzed by one-way analysis of variance (ANOVA) using SPSS 18 software® to compare their mean values. Fisher's least significant differenc e procedure was used to test for significance and the level of confidence was set at 95%.

The similarity factor among formulations, /2, was determined (Moore and Flanner, 1996)

/2 = 5 Ox 1 O g ( ■ 1 X 1 0 0 ) eqn. 2

:rs to th

+£)Z?=i(at-7t)2]

where n refers to the number of samples, R refers to the drug release from the reference material at time t, and T refers to the drug release from the test sample at time t.

'.2.6. Bioavailability study

2.2.6.1. Administration program and blood sampling

The administration of the prepared matrix tablets and the marketed product to six dogs was completed according to a two way crossover design with a one-week washout period

between each treatment phase. Dogs having weight of 12 Kg, age of 12-14 months and sex of 4 females and 2 males, were divided randomly into two groups and allocated to one of the two treatments according to the study design shown in Table 1. Each dog received the same water and meal prior to treatment. A single oral dose equivalent to a 100 mg diclofenac dose in one of

the formulations was administered after overnight fasting for at least 10 hours with free access to water. Multiple venous blood samples for analysis of diclofenac concentrations in plasma were collected prior to drug administration (0 hour) and at 0.5, 1, 2, 3, 4, 5, 6, 8, 10, 12 and 24 hours post-dose. The volume of blood taken for determination of diclofenac in plasma was 5 ml. The blood samples were placed in heparinized tubes, slightly shaken and centrifuged at 3000 rpm for 10 minutes. The plasma was then decanted and placed in coded polypropylene tubes and then stored at -70°C until drug analysis. The protocol of the study for animal experiments was approved by the Institutional Animals Ethical Committee of the department.

Table 1. Study design of the crossover bioequivalence study

Treatment

Sequence Dogs

Period 1 Period 2

1 1, 3, 5 a Rb

2 2, 4, 6 R T

aFormulatio

bVoltaren® 75 mg SR, Batch number: T1339, and Cataflam® 25 mg IR, Batch number:

Y0086, manufactured by Novartis Pharma AG, Switzerland)

2.2.6.2. Plasma sample preparation and drug analysis

One milliliter of the plasma sample was spiked with 100^1 of ibuprofen (internal

standard: 0.5mg/ml dissolved in methanol). The sample was vortexed for 30 sec, made

quantity 1ml of

acidic with 250 ^l 0.15M phosphoric acid, vortexed for another 30 sec and extracted with 5 ml diethyl ether with vortexing for 1 min. The extract was centrifuged at 3000 rpm for 5 min, and then 3 ml of the supernatant was decanted to a glass tube. The separated quantity was evaporated to dryness at 50oC and the remaining residue was dissolved with 1i mobile phase and vortexed for 1min. After centrifugation at 15000 rpm for 2 min, ten microliters was injected into the UPLC-UV system for analysis.

The ratio of the peak-area ratio of the drug to the peak area of the internal standard estimated by Masslynx Version 4.1 software (Waters, USA) allowed quantification of the DS present. The relationshipbetween concentration and peak area ratio was linear, with an r2 coefficient of not less than 0.99 within the range 0.1-30 ^g/ml for diclofenac. The lower limit of quantification (LLOQ) for diclofenac from 1ml of plasma was 0.5 ^g/ml. Intra- and inter day accuracy of the method ranged from 97.04 to 107.8% and from 99.2 to 114.7%; respectively, while the intra- and inter-day precision ranged from 0.29 to 9.3% and from 3.1 to 9.5%; respectively, for concentrations 0.5, 1, 8, 15 and 30 ^g/ml. The average absolute analytical recovery of diclofenac ranged from •>, and o:

93.69 to 107.2%, and of the internal standard (ibuprofen) was 96.31%.

)7.2%, a

2.2.6.3. Pharmacokinetic analysis

WinNonlin software (Version 4.1, Pharsight Corp., Palo Alto, CA, USA) was d to determine the pharmacokinetic parameters. Non-compartmental models (Yamaoka et al., 1978), were employed for the evaluation of these parameters. The terminal log-linear portion of the concentration-time curves in plasma was analyzed by

least squares regression analysis to determine the elimination rate constant and the half-

life of the drug. The trapezoidal method was employed to determine the area under the concentration-time curve in plasma from zero time to the last quantifiable concentration ( AUC0_t) and to infinity (). The ratio of C^/AUC^ was used as an estimate of the absorption rate constant (Endrenyi et al., 1991). The area under the moment curves from zero time to the last quantifiable concentration ( AUMC ) and to infinity ( AUMC^) were also calculated by the trapezoidal method. The following equation was used to estimate the mean residence time (MRT):

MR T = ^^ eqn. 3

AUC0- oo

The apparent volume of distribution at steady state (Vss) was estimated using the following relationship (Perrier and Mayersohn, 1982):

Vss/ D.AUMCQ—oq

where F refers to the drug bioavailability of the drug and D refers to the dose administered. In addition, the oral clearance of the drug was calculated by the following equation:

CL/ _ _d

i additi

eqn. 5

AUCq—oq

2.2.6.4. Statistical analysis

The Statistical Analysis System software package (SAS Institute, Cary, NC,

USA) was employed. The stu d ent's t-test for independent samples was employed to

compare the calculated pharmacokinetic parameters between test and reference assuming

homoscedasticity. It is well known that the time to peak plasma concentration (Tmx) data do not follow a Gaussian distribution; therefore, the nonparametric Mann-Whitney test was used. The results were expressed as mean±SD and comparisons were carried out at the 0.05 level of significance.

2.2.7. Effect of storage on physical properties and drug release from the tablets

The multi-layered tablet that demonstrated bioequivalence to the marketed product was assessed for the effect of long term and accelerated storage conditions on the physical characters and release profile, according to International Conference on Harmonization Guidelines (ICH, 2003). The tablets were stored in closed containers in stability cabinet (Binder Gmbh, Tuttlinger, Germany) at 25 ± 2°C and 60 ± 5% relative humidity (RH) for 12 months and 40 ± 2°C and 75 ± 5% (RH) for 6 months.

3. Results

3.1. Evaluation of the prepared tablets

The characteristics of the core matrix, drug layered and final coated tablets are listed in Table 2. The core tablets displayed acceptable weight variation, content uniformity and friability. Tablets were assessed after drug layering for the coating efficiency by determining the amount of diclofenac potassium loaded on each tablet. The

photometry analysis showed an average DP content of 100.5 ± 3.54 %. After the application of the final film coat of Opadry II, the average tablet weight was found to be 400 ± 3.72 mg, hardness 182.1 ± 6.33 N, thickness 4.83 ± 0.02 mm and overall diclofenac content 96.25 ± 1.64 %.

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flame ph appl

Table 2. Physical characterization of biphasic release matrix tablets of diclofenac in the three manufacturing stages (before coat, drug layered and film coated)

Formulations

Drug content (%) (RSD)

Tablet mass (mg) ± SD Hardness (N) ± SD Thickness (mm) ± SD Friability0 (% loss) ± SD

Formulation E DP layered (before coata)

Formulation E* (after final film coat)

94.91 (1.86) 96.25b (1.7) 96.25 (1.7)

351 ± 3.21 105.4 ± 5.17 4.46 ± 0.02 0.1 ± 0.03

385 ± 4.32 400 ± 3.72

129.3 ± 13.65 182.1

4.78 ± 0.02 4.83 ± 0.

0.02 ± 0.01 0.02 ± 0.01

a Core matrix tablets before drug layering with DP. b Content calculated based on total diclofenac content. c Initial mass of 6.5 g

3.2. In-vitro drug release

Figure 1 show that the release profile of formulation E (core matrix) and Voltaren® 75mg are similar to each other with f value of 79.2. The release efficiency (RE) and mean dissolution time (MDT) values were comparable where the RE was 80.15% and 83.5% and the MDT was 4.81h and 4.91h for formulation E and Voltaren® 75mg; respectively. Formulation E* (after DP layering and coating) showed an increased amount of drug released at each time point compared with previous formulations due to the additional drug load. This was also the case when monitoring the simultaneous release from the two tablets Voltaren® 75mg and Cataflam® 25mg as the reference product. The release efficiency values were comparable 86.38% vs 82.72% and the mean dissolution times were 4.05h vs 4.52h for formulation E* and the reference, respectively.

T3 dJ to

CD 80 _OJ

Formulation E*

Voltaren SR 75 mg & Cataflam 25 mg Formulation E Voltaren SR 75 mg Voltaren SR 100 mg

Time (h)

Figure 1

3.3. Pharmacokinetic study

The plasma concentration-time curves for the two formulations; formulation E* and the reference (Voltaren® 75mg & Cataflam® 25mg) are shown in Figure 2. Table 3 lists the pharmacokinetic parameters of diclofenac in both formulations. No significant difference between any of the mean values of the pharmacokinetic parameters calculated for the treatments was found at a 95% confidence interval (CI). The Cmax values were 11.29 ± 6.87 & 12.97 ± 8.45 ^g/L and the AUC0-24 values were 105.36 ± 83.3 & 92.87 ± 55.53 ^g.h/L for formulation E* and the reference; respectively. This indicates that the rate and extent of absorption were comparable between these two formulations. These results are in agreement with the observed in-vitro release data.

(O _TB

(C 8 +J

S 4 (0

■Formulation E*

■Reference product "Voltaren 75mg and Cataflam 25mg"

0 4 8 12 16 20 24

Time (h)

Figure 2

Table 3. Pharmacokinetic parameters (mean ± SD) for formulation E* and the reference (Voltaren® 75mg and Cataflam® 25mg) in 6 dogs

Formulation E* Reference ^-value

kel (h-1) 0.121 ± 0.05 0.12 ± 0.05 0.95

t1/2 (h) 6.63 ± 2.69 7.46 ± 5.34

Tmax (h) 4.83 ± 3.76 4.33 ± 0.82

Cmax (|g/l) 11.29 ± 6.87 12.97 ± 8.45

Clast/Cmax 0.13 ± 0.1 0.1 ± 0.12

AUC0-t (|g.h/l) 105.36 ± 83.3 92.87 ± 55.53

AUC0-^ (|g.h/l) 118.12 ± 85.8 104.05 ± 52.05

AUC ratio 0.88 ± 0.1 0.88 ± 0.19

Cmax/AUC0-» (h-1) 0.124 ± 0.1 0.127 ± 0.05

AUMC0-W (|g.h2/l) 1357±960 1199±809

MRT (h) 11.19 ± 4.12 12.12 ± 8.93

Vss/F (l/kg) 0.89 ± 0.54 1.08 ± 0.94

CL/F (l.kg-1.h-1) 0.096 ± 0.06 0.094 ± 0.04

0.74 0.94 0.71 0.65 0.77

0.91 0.77 0.81 0.67 0.94

3.4. Stability study

The moisture content as well as the rate of moisture uptake are affected by humidity and temperature and should be considered. Figure 6 shows the release profiles for the formulation E* after storage under long term and accelerated conditions. Table 4 shows the physical

ong ter rmulati

characteristics of formulation E* and the release profile similarity factor f2 after exposure to the long term and accelerated conditions as compared to the initial release profile. There was no significant increase in tablet mass or hardness. The f2 values for drug release, after long term and iccelerated conditions, ranged from 62 to 71 suggesting similarity to the initial release results.

accelera

Figure 3

Table 4. Effect of long term and accelerated storage conditions on the physical properties of formulation E* (mean ± SD) and the similarity factor f2

Stability status Drug content Tablet weight Hardness (N) Thickness Friabilitya f2

(%) (mg) ± SD ± SD (mm) ± SD (% loss)

Initial 96.3 (1.7)b 400 ± 3.7 182 ± 6.3 4.83 ± 0.02 0.02 N/A

Long term 3M 96.4 (1.64) 402 ± 3.2 166 ± 7.2 4.87 ± 0.02 0.02 71.74

6M 96.5 (1.64) 400 ± 4.7 167 ± 6.6 4.87 ± 0.02 0.03 64.14

12M 96.8 (1.37) 406 ± 3.6 189 ± 4.9 4.86 ± 0.03 0.01 62.02

Accelerated 3M 96.5 (1.2) 408 ± 4.3 194 ± 6.6 4.92 ± 0.01 0.01 66.2

conditions 6M 96.4 (1.21) 408 ± 4.5 192 ± 5.8 4.92 ± 0.01 0.01 63.31

Amount of tablets weighed equivalent to 6.5 g Values in parentheses are the RSD of the mean

4. Discussion

4.1. In-vitro drug release

Both formulation E* tablets and the commercially available formulations (Voltaren® SR 75mg and Cataflam® 25mg) exhibited higher initial concentrations when compared to results of formulation E or Voltaren® 75mg alone due to the rapid dissolution of the immediate release layer of DP from formulation E* or the contribution of the rapid disintegration followed by dissolution of diclofenac by the Cataflam® 25mg tablet. The remainder of the drug is released over a 16 hour period; however, the core matrix did not lose its integrity even after 24h. The drug release from formulation E* reached 42% in 1 h, which suggests that the pharmacologic effect can be attained rapidly. Analysis of drug release data revealed that release from formulation E* or the reference material occurred by diffusion (Higuchi model fir woth r2=0.99) where the release of diclofenac was dependent on the square root of time. The Krosymeyer-Peppas exponent n was 0.34 and 0.405 for formulation E* and the reference, respectively after fitting the dissolution profiles into the exponential equations. It can be inferred from n<0.45 that diffusion is the dominating mechanism in the release process from these matrix type formulations (Su et al., 2003). After exposure of the inner matrix to the aqueous medium, solvent penetrates into the voids between macromolecular chains of Eudragit RL more easily allowing solvation of the polymer chains. This might suggest polymer relaxation as a result of increased polymer dimensions due to the hydrostatic pressure built up by the

entered solvent (Apu et al., 2009). The similarity factor (f2) value was found to be 66.19 indicating comparable release profiles for diclofenac from the two formulations.

4.2. Pharmacokinetic study

The bioavailability study presents strong evidence for bioequivalence between the test and comparator formulations. Being a highly permeable drug, diclofenac is found to be absorbed throughout the gastrointestinal tract (Gleiter et al., 1985). After dosing of the two formulations, the plasma concentration of diclofenac was easily detectable in the blood after 30 minutes from each of the formulations. This corresponded well with the observed pulse effect of drug release that occurred due to the fast dissolution of DP layer

in formulation E* and the rapid drug release of DP from Cataflam® 25mg. The in-vivo

profiles can be explained on the basis of the three main stages. The first stage takes place

from 0 to 2 h where the extent and rate of absorption of formulation E* was higher than

that of the reference. This can be attributed to the rapid dissolution of the immediate

release coat containing DP. On the other hand, the Cataflam® 25mg tablet tends to

disintegrate first and then dissolution begins, and this decreased the absorption rate

relative to formulation E*. The second stage is from 2-7 h at which point the drug

reached the small intestine which has a high effective surface area that facilitates

absorption. Even though diclofenac solubility is higher with greater extent of absorption

in the small intestine than in the stomach, the absorption rate in case of formulation E* is

controlled by the core matrix which, according to Figure 1, releases drug at a slower rate

than did the outermost coat with its DP. And that is what is reflected in the 2-7 h

timeframe in Figure 2. Although the release from the core of Formulation E* is slightly

in th con

slower than observed with the rate provided by the reference combination of the commercial products, there is still no statistical difference between the two formulations at each point of the drug release during this stage which may be attributed to the wide scatter in the reference data. In the third stage from 8 to 24 h, a higher level of the drug in the blood was detected from formulation E* than the reference combination where the drug in formulation E* continued to be released from the tablets. Formulation E*, hence, keeps an elevated diclofenac concentration in the plasma for a longer time which is good for the patient.

4.3. Stability study

The drug release and the physical characteristics of the tablets remained unchanged after storage for 1 year under long term conditions and 6 months at accelerated conditions suggesting that DS is stable in the multi-layered tablets. Noticeably, there was an insignificant decrease in the drug release after 10h in both long and accelerated conditions. This effect can be linked with the slight increase in hardness of the tablets. The long exposure of the tablets to elevated humidity/temperature might promote fusion of the polymer particles due to increased mobility with possible redistribution of the pores within the matrix system (Grund et al., 2014). This resulted in a small retardation in the drug release in accordance with similar findings that have been reported previously (Grund et al., 2014). However, the overall assessment of the release profile indicates stability of the tablets according to ICH guidelines.

repo prof

Conclusion

Modification of Eudragit-based matrix systems made it possible to achieve a biphasic drug release patterns. A first pulse was obtained by incorporation of a percentage of the drug in an outer immediate release layer applied onto a release controlling matrix which would provide the sustained pharmacologic effect. The clin significance of such formulations would be profound with patients suffering from acute or long lasting pains. Based on its in-vitro, in-vivo and stability profiles of formulation E*, it is a system that can accomplish both immediate and sustained release of diclofenac for extended pain relief.

elease of

Acknowledgement

The project was financially supported by King Saud University, Vice Deanship of Research Chairs, Kayyali Chair for Pharmaceutical Industry ( Grant no. E-2015). Special thanks to Engineer Abdullah Bagshan Research Chair for Growth Factors and Bone Regeneration at King Saud University for their support towards this research with registration number F-750-03-07 and for allowing the in-vivo work to be conducted in their animal house.

Conflict of Interest

The authors have declared that no conflict of interest exists.

<eierei

rences

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Figure Captions

Figure 1. Comparative release profiles of diclofenac from different formulations at pH 7.5: Voltaren® 75 mg; Formulation E (before coat); Voltaren® 100 mg; Voltaren® 75 mg & Cataflam® 25 mg; Formulation E*. Each data point represents the average of a triplicate study.

Figure 2. Mean diclofenac plasma profiles from a single dose bioavailability study compared with the reference product (n = 6). Voltaren® SR 75 mg & Cataflam® 25 mg;

— .SE mgr

Figure 3. Release profiles after stability studies in long term (A) & accelerated (B) conditions for formulation E* according to ICH guidelines.

T3 dJ to CD

_CD 13

Figure 4

Formulation E*

Voltaren SR 75 mg & Cataflam 25 mg Formulation E Voltaren SR 75 mg Voltaren SR 100 mg

Time (h)

(O _T0

(C 8 +J

S 4 (0

Figure 5

■Formulation E*

■Reference product "Voltaren 75mg and Cataflam 25mg"

Time (h)

~i— 20

Table 1. Study design of the crossover bioequivalence study

Treatment

Sequence Dogs

Period 1 Period 2

1 1, 3, 5 rpa Rb

2 2, 4, 6 R T

b Voltaren® 75 mg SR, Batch number: T1339, and Cataflam®

Y0086, manufactured by Novartis

rmulation E* atch number: AG, Switzerland)

Table 2. Physical characterization of biphasic release matrix tablets of diclofenac in the three manufacturing stages (before coat, drug layered and film coated)

Formulations

Formulation E (before coata)

DP layered

Formulation E* (after final film coat)

Drug content (%) (RSD)

Tablet mass (mg) ± SD Hardness (N) ± SD Thickness (mm) ± SD Friability0 (% loss) ± SD

94.91 (1.86) 96.25b (1.7) 96.25 (1.7)

351 ± 3.21 385 ± 4.32 400 ± 3.72

105.4 ± 5.17 129.3 ± 13.65 182.1 ± 6.33

4.46 ± 0.02 4.78 ± 0.02 4.83 ± 0.02

0.1 ± 0.03 0.02 ± 0.01 0.02 ± 0.01

a Core matrix tablets before drug layering with DP. Content calculated based on total diclofenac content.

c Initial mass of 6.5 g

Table 3. Pharmacokinetic parameters (mean ± SD) for formulation E* and the reference (Voltaren 75mg and Cataflam 25mg) in 6 dogs

Formulation E* Reference

kel (h-1) 0.121 ± 0.05 0.12 ± 0.05

t1/2 (h) 6.63 ± 2.69 7.46 ± 5.34

Tmax (h) 4.83 ± 3.76 4.33 ± 0.82

Cmax (^g/l) 11.29 ± 6.87 12.97 ± 8.45

Clast/Cmax 0.13 ± 0.1 0.1 ± 0.12

AUC0-t (^g.h/l) 105.36 ± 83.3 92.87 ± 55.53

AUC0.^ (^g.h/l) 118.12 ± 85.8 104.05 ± 52.0:

AUC ratio 0.88 ± 0.1 0.88 ± 0.19

Cmax/AUC0_. (h-1) 0.124 ± 0.1 0.127 ± 0.05

AUMC0-» (^g.h2/l) 1357±960 1199± 809

MRT (h) 11.19 ± 4.12 12.12 ± 8.93

Vss/F (l/kg) 0.89 ± 0.54 1.08 ± 0.94

CL/F (l.kg-1.h-1) 0.096 ± 0.06 0.094 ± 0.04

^-value

0.95 0.74 0.

0.77 0.74 0.95 0.91 0.77 0.81 0.67 0.94

Table 4. Effect of long term and accelerated storage conditions on the physical properties of formulation E* (mean ± SD) and the similarity factor f2

Stability status Drug content Tablet weight Hardness (N) Thickness Friabilitya f2

(%) (mg) ± SD ± SD (mm) ± SD (% loss)

Initial 96.3 (1.7)b 400 ± 3.7 182 ± 6.3 4.83 ± 0.02 002 N/A

Long term 3M 96.4 (1.64) 402 ± 3.2 166 ± 7.2 4.87 ± 0.02 0.02 71.74

6M 96.5 (1.64) 400 ± 4.7 167 ± 6.6 4.87 ± 0.02 0.03 64.14

12M 96.8 (1.37) 406 ± 3.6 189 ± 4.9 4.86 ± 0.03 0.01 62.02

Accelerated 3M 96.5 (1.2) 408 ± 4.3 194 ± 6.6 4.92 ± 0.01 0.01 66.2

conditions 96.4 (1.21)

6M 408 ± 4.5 192 ± 5.8 4.92 ± 0.01 0.01 63.31

tablets weighed equivalent to 6.5 g

a Amount of tab Values in parentheses are the RSD of the mean