Eur J Drug Metab Pharmacokinet DOI 10.1007/s13318-014-0194-9
ORIGINAL PAPER
Development of hollow/porous floating beads of metoprolol for pulsatile drug delivery Sangmesh S. Taranalli • Panchaxari M. Dandagi Vinayak S. Mastiholimath
•
Received: 24 April 2013 / Accepted: 25 March 2014 Ó Springer International Publishing Switzerland 2014
Abstract The purpose of this work was to develop hollow calcium pectinate beads for floating pulsatile release of metoprolol tartrate intended for chronopharmacotherapy. Floating pulsatile concept was applied to increase the gastric residence of the dosage form having lag phase followed by a burst release. To overcome limitations of various approaches for imparting buoyancy, hollow/porous beads were prepared by simple process of acid-base reaction during ionotropic cross-linking using low methoxy pectin, xanthan gum, sodium carboxy methyl cellulose, guar gum, locust bean, gellan gum and calcium chloride as a cross-linking agent. Based on the preliminary studies optimized polymers were selected for formulation design with different polymers ratio concentrations. The obtained floating beads were studied for entrapment efficiency, buoyancy study, swelling index, surface morphology, in vitro release, stability studies and in vivo floating study. The floating beads obtained were porous, float up to 12–24 h. The radiological studies (X-rays) pointed out the capability of the system to release drug in lower parts of GIT after a programmed lag time for hypertension. The floating beads provided expected two-phase release pattern with initial lag time during floating in acidic medium followed by rapid pulse release in phosphate buffer. From the accelerated stability studies, it was observed that the formulations are quite stable. All formulations followed firstorder release kinetics by diffusion mechanism. This S. S. Taranalli (&) P. M. Dandagi Department of Pharmaceutics, KLEU’s College of Pharmacy, Belgaum 590010, Karnataka, India e-mail: [email protected] V. S. Mastiholimath Department of Quality Assurance, KLEU’s College of Pharmacy, Belgaum, India
approach suggested the use of hollow calcium pectinate microparticles as promising floating pulsatile drug delivery system for site- and time-specific release of drugs acting as per chronotherapy of diseases. Keywords Hollow beads Calcium pectinate beads Floating pulsatile drug delivery Metoprolol tartrate Chronotherapy Radiology
1 Introduction A delivery system with a release profile that is characterized by a time period of no release (lag time) followed by a rapid and complete drug release (pulse release) can be called as an ideal pulsatile drug delivery system. Pulsatile delivery system provides one or more rapid release pulses at predetermined lag times or at specific sites resulting in better absorption of the drug, and there by providing more effective plasma concentration time profile (Venkatesh 2005). Natural biodegradable polysaccharides like pectin, locust bean, guar gum, chitosan, sodium alginate and gellan gum have been used in controlled drug delivery. Various approaches to induce buoyancy in cross-linked beads have been used (Whitehead et al. 2000; Iannuccelli et al. 1998; Sriamornsak and Nunthanid 1999). The use of sodium bicarbonate as buoyancy imparting agent to produce floating beads is simplest among the various approaches and has been attempted successfully by many workers (Bussmer et al. 2003). Their floating property was based on the evolution of CO2 when in contact with acidic environment followed by the ability of polymer gel to entrap it, which decreases their density below one. These beads have been used to achieve prolonged gastric residence time, for sustained release/stomach-specific drug delivery providing
Eur J Drug Metab Pharmacokinet
an opportunity for both local and systemic drug action (Rajnikanth et al. 2007). In cardiovascular disease, capillary resistance and vascular reactivity are higher in the morning and decreases latter in the day. Platelet agreeability is increased and fibrinolytic activity decreased in the morning, leading to a state of relative hypercoagulability of the blood. Therefore, the frequencies of myocardial infarction (MI) and of sudden cardiac death are more prone from morning to noon. Ambulatory blood pressure measurements show a significant circadian variation to characterize blood pressure. Increased heart rate, blood pressure, imbalanced autonomic tone, and circulating level of catecholamines controlling the cardiac arrhythmias show important circadian variation and trigger the genesis of the circadian pattern of cardiac arrhythmias. Atrial arrhythmias appear to exhibit circadian pattern usually with a higher frequency in the daytime and lower frequency in the night time with the abnormal foci under the same long-term autonomic regulation as normal pacemaker tissue. According to study, ventricular tachyarrhythmias show late morning peak in the patients with MI sometime in the distant past morning peak and afternoon peak in patients with recent MI. Both pharmacokinetics and pharmacodynamics of some oral nitrates, calcium channel blocker and b-adrenoceptor antagonist medications have been shown to be influenced by the circadian time of their administration (Mandal et al. 2010). Metoprolol is a cardioselective b1-adrenergic blocking agent used for acute MI, heart failure, angina pectoris and mild to moderate hypertension. It may also be used for supraventricular and tachyarrhythmias and prophylaxis for migraine headaches. Metoprolol competes with adrenergic neurotransmitters such as catecholamines for binding at beta (1)-adrenergic receptors in the heart. Beta (1)-receptor blockade results in a decrease in heart rate, cardiac output, and blood pressure (Metoprolol tartrate (internet) 2012). All beta blockers are nearly equally effective in decreasing frequency and severity of attacks and in increasing exercise tolerance in classical Angina, but cardioselective agents. Long-term b-blocker therapy lowers risk of sudden cardiac death among ischemic heart disease patients. In angina pectoris, beta blockers are to be taken on a regular schedule not on as and when required basis (Tripathi 2008). It is moderately lipophilic drug belonging to class I of BCS classification having high solubility and high permeability. Since the half-life of metoprolol tartrate is 3–4 h, two multiple doses are needed to maintain a constant plasma concentration for a good therapeutic response and improved patient compliance (Metoprolol tartrate (internet) 2012). Although it is well absorbed in the gastrointestinal tract, its bioavailability is 40–60 % as a result of extensive first pass metabolism (Metoprolol (internet) 2012). Thus, there is a strong clinical need and market potential for a
dosage form that will deliver metoprolol tartrate in a controlled manner to a patient needing this therapy, thereby resulting in a better patient compliance. Multi-particulate or multiple unit systems offer various advantages over single-unit systems. These include no risk of dose dumping, flexibility of blending units with different release patterns, relative merits of bioavailability more consistent blood levels, reproducible and avoid all or none effect. The aim of the study was to design and characterize hollow/porous floating beads of metoprolol tartrate for pulsatile drug delivery for the treatment of hypertension.
2 Materials and method 2.1 Materials Low methoxy pectin (LMP) was obtained as generous gift sample from Krishna Pectins Pvt. Ltd., Jalgaon, Maharashtra, India. Metoprolol tartrate was obtained as generous gift sample from Astrazeneca Pharmaceuticals Pvt. Ltd., Bangalore (India). Xanthan gum, Sodium CMC, Guar gum Locust bean, Gellan gum and Sodium bicarbonate from Himedia Laboratories Pvt. Ltd., Mumbai, and Calcium Chloride from Loba Chem Pvt. Ltd., Mumbai. All other chemicals used were of analytical grade. 2.2 Method 2.2.1 Formulation of metoprolol tartrate beads 2.2.1.1 Preliminary studies for selection of polymer combination Four formulations were prepared using different polymers as shown in Table 1. Evaluation parameters such as drug entrapment efficiency, buoyancy study of the beads and in vitro drug release for the formulated beads were done and the promising batch was selected on the basis of above parameters for the further study. 2.2.1.2 Ionotropic gelation/bead formation A polymeric solution was prepared by dissolving various amounts of polymers in 15 ml of deionized water. Then, gas-forming agent, such as sodium bicarbonate and metoprolol tartrate (100 mg), was added into the solution (Table 2). The dispersion was sonicated for 30 min to break the lumps and to remove air bubbles. The resultant dispersion was dropped via a 21-gauge syringe needle into 3 % w/v calcium chloride (CaCl2) solution containing 10 % (v/v) acetic acid. The distance between the tip of the needle and the surface of the CaCl2 medium was about 6 cm. The beads formed were stirred at 150 ± 5 rpm using magnetic stirrer for 15 min for curing. The beads were separated by
Eur J Drug Metab Pharmacokinet Table 1 Preliminary studies for selection of polymer combination Batch no.
Drug (mg)
Low methoxy pectin (mg)
Xanthan gum (mg)
Sodium CMC cellulose (mg)
Guar gum (mg)
FA
100
300
50
100
–
FB
100
300
50
–
FC
100
300
50
–
FD
100
300
50
–
Locust bean (mg)
Gellan gum (mg)
Sodium bicarbonate (mg)
–
–
150
100
–
–
150
–
100
–
150
–
–
100
150
Table 2 Formulation code for metoprolol tartrate floating beads Batch no.
Metoprolol tartrate (mg)
Low methoxy Pectin (mg)
FCE
100
300
FCF FCG
100 100
300 300
Xanthan gum (mg)
Locust bean (mg)
Sodium bicarbonate (mg)
Deionized water (ml)
30
120
150
15
60 120
90 30
150 150
15 15
filtration, washed three times with distilled water and subsequently oven dried at 45 °C for 6 h (Dupuis et al. 2006). 2.2.2 Characterization of beads 2.2.2.1 Particle size analysis and surface morphology 100 beads were analyzed for their size distribution by optical microscopy. The mean diameter was determined by measuring the number of divisions covered by beads using ocular micrometer previously calibrated using stage micrometer. The surface Morphology was analyzed with a scanning electron microscope (JEOL JSM-6360 SEM) operated at an acceleration voltage of 5 kV (Pornsak et al. 2005; Gadad et al. 2009). 2.2.2.2 Determination of drug entrapment efficiency Accurately weighed quantities (50 mg) of beads from each batch were placed in 100 ml phosphate buffer, pH 7.4 and mechanically agitated on shaker at 200 rpm for 24 h. The resultant dispersions were filtered and analyzed spectroscopically at 224 nm. The percentage entrapment efficiency was calculated using following equation (Claire et al. 2011). Drug entrapment efficiency % ¼ ðActual drug content in the beads =theoretical drug contentÞ 100: 2.2.2.3 Bead porosity and bulk density The bead porosity was assessed using mercury porosimetry (Autoscan 60 Porosimeter, Quantachrome software, USA). The pressure was applied from 0 to 6,000 psi. The mercury intrusion data were recorded and plotted against pressure. Standard values for the contact angle and surface tension of mercury were used for calculations. The bulk densities of the beads
were also measured using same mercury porosimeter (Bulgarelli et al. 2002). 2.2.2.4 Buoyancy study Floating property of beads was evaluated using USP XXIII type II dissolution test apparatus (Electrolab TDT-06P, Mumbai, India) filled with 900 ml in pH 1.2 containing 0.02 % w/v Tween 80, using paddle at a rotation speed of 100 rpm. The temperature of medium was maintained at 37 ± 2 °C. 100 beads of each batch were placed in the media. Floating ability was observed visually (Huimin et al. 2012; Mastiholimath et al. 2008). 2.2.2.5 Determination of swelling index The swelling behavior of the beads was studied in pH 7.4 phosphate buffer solution. Approximately 100 mg of beads was taken in a dissolution basket and weighed; the baskets along with the beads were immersed in 7.4 phosphate buffer solution. The weight of the basket along with the beads was determined after 1 h, and then every hour. The swelling index (SI) of each formulation was calculated using the following equation (Sandolo et al. 2011; Patel et al. 2011): %SI ¼
W2 W1 100 W1
where, W1 is weight of the dry beads and basket and W2 is weight of the swollen beads and basket. 2.2.2.6 In vitro drug release The dissolution study of beads equivalent to 100 mg of metoprolol tartrate was performed using a USP XXIII type 2 dissolution test apparatus. The drug release study was performed in 900 ml for 6 h in pH 1.2. Then the dissolution medium was replaced with phosphate buffer of pH 6.8 for 3 h, the average small intestinal transit time is about 3 h. After 9 h, the dissolution medium was replaced with phosphate buffer
Eur J Drug Metab Pharmacokinet
of pH 7.4 maintained at 37.0 ± 0.5 °C at 100 rpm for 18 h. Periodically 5 ml of sample was withdrawn and replaced by dissolution media in order to maintain sink condition. The withdrawn samples were filtered through a Wattman filter paper 41 and the concentration of metoprolol tartrate was measured spectrophotometrically at 221.5 and 224 nm for acidic and basic media, respectively, after suitable dilutions (Patel et al. 2011). 2.2.3 In vivo study 2.2.3.1 Radiology (X-rays) The optimized batch was chosen for the preparation of the barium sulfate-loaded beads. The barium sulfate beads were prepared as of optimized batch but the drug was replaced with barium sulfate (100 mg). Healthy rabbit weighing approximately 2.3 kg was used to assess in vivo floating behavior. Ethical clearance for the handling of experimental animals was obtained (CPCSEA). The animal was fasted for 12 h and the first X-ray photographed to ensure the absence of radio opaque material in the stomach. The rabbits were made to swallow barium sulfate-loaded beads with 30 ml of water. During the experiment, rabbits were not allowed to eat but water was provided. At predetermined time intervals, the radiograph of abdomen was taken using an X-ray machine (Gangadharappa et al. 2011). 2.2.3.2 Stability study Stability study has become an integral part of formulation development. It generates information on which proposal for nature of drug or dosage and their recommended storage conditions are based. The accelerated stability study was conducted for selected formulation as per the ICH guidelines. Accelerated testing 40 ± 2 °C/75 ± 5 % RH for 6 months. As per ICH guidelines, beads of promising formulation were selected and subjected to accelerated stability study. Weighed quantity of the samples was kept in glass vials, sealed with rubber plugs and exposed to controlled temperature (40 ± 2 °C) and relative humidity (75 ± 5 %) for a period of 6 months in humidity control oven (Lab Control, Ajinkya IM 3500 Series, India). 2.2.3.3 Photo-stability study A light source that is designed to produce an artificial daylight fluorescent lamp combining visible and ultraviolet (UV) outputs, xenon, or metal halide lamp is used. D65 is the internationally recognized standard for outdoor daylight as defined in ISO 10977 (1993). ID65 is the equivalent indoor indirect daylight standard. For a light source emitting significant radiation below 320 nm, an appropriate filter may be fitted to eliminate such radiation. After 1, 2, 3 and 6 months, the samples were taken out and analyzed for drug entrapment
efficiency, buoyancy study, and in vitro release (Gadad et al. 2009; Amrutkar et al. 2012). 2.2.3.4 Release kinetics One of the most important and challenging areas in the drug delivery field is to predict the release of the active agent as a function of time using both simple and sophisticated mathematical models. The importance of such models lies in their utility during both the design stage of a pharmaceutical formulation and the experimental verification of a release mechanism. In order to identify a particular release mechanism, experimental data of statistical significance are compared to a solution of the theoretical model. To analyze the mechanism for the drug release and drug release rate kinetics of the dosage form, the data obtained was fitted into Zero order, First order, Higuchi matrix and Korsmeyer-Peppas. In this study by comparing the R2 values obtained, the best-fit model was selected. Drug release kinetics can be analyzed by various mathematical models, which are applied considering the amounts of drug released from 0 to 24 h. The following plots were made: Cumulative percentage drug release versus time (zero-order kinetic model); log cumulative percentage drug remaining versus time (first-order kinetic model); cumulative percentage drug release versus square root of time (Higuchi model); and log cumulative percentage drug remaining versus log time (Korsmeyer’s Peppas model) (Lin and Kawashima 1987).
3 Result and discussion 3.1 Pre-formulation study 3.1.1 Compatibility study The FT-IR spectra of pure metoprolol tartrate and the combination of metoprolol tartrate with the polymer show similar characteristic functional peak. The similarity in the peaks indicates the compatibility of metoprolol tartrate with polymers as shown in Fig. 1. 3.1.2 Formulation of floating calcium pectinate beads Based upon the results of preliminary study, FC formulation was selected for further studies because it showed highest entrapment efficiency of 70.89 %, produced floating beads buoyancy for 24 h and highest in vitro drug release of 96.04 %. Three formulations (FCE, FCF and FCG) of metoprolol tartrate beads were made. The formulations were prepared by varying polymer ratio and keeping the concentration of drug and other ingredients same as showed in table: the method used to prepare the calcium pectinate beads by
Eur J Drug Metab Pharmacokinet Fig. 1 FTIR spectra
dripping method using 21-gauge needle into the 3 % calcium chloride solution containing 10 % v/v acetic acid. The beads were formed due to the cross-linking of the pectin with divalent calcium ions of the calcium chloride solution. The reaction between sodium bicarbonate and acetic acid was occurred liberating carbon dioxide as gas bubbles which are responsible for floating of beads. 3.1.3 Characterization of beads 3.1.3.1 Particle size analysis and surface morphology The mean surface diameter of all three formulations was between 1.134 ± 0.018 and 1.198 ± 0.008 (mm),
tabulated in Table 3; this indicates that size of beads increased significantly with increase in viscosity there by subsequent increase in interfacial tension, resulting in the formation of larger particles. Surface morphology of beads batch no FCF was observed. It was shown that beads were spherical in shape, with slightly rougher surface/shrinkage and discrete as shown in Fig. 2. The surface topography reveals that the beads were highly porous because of rapid escape of the carbon dioxide during formulation. The cross section of beads from batch FCF showed a hollow core in the matrix, which may be because of the presence of gas generating agent. The thick matrix
Table 3 Characterization of floating beads Batch code
Particle size (mm)*
Drug entrapment efficiency (%)*
Swelling index (%)*
Bulk density (g/cm3)*
Porosity (%)
Floating ability in pH 1.2 (h)
FCE
1.134 ± 0.02
68.44 ± 3.81
1.51 ± 0.15
1.35 ± 0.05
28.94
[12
FCF FCG
1.176 ± 0.01 1.198 ± 0.01
76.84 ± 6.37 72.72 ± 4.23
1.95 ± 0.13 1.83 ± 0.13
1.13 ± 0.19 0.87 ± 0.13
32.73 34.08
[24 [24
* Values expressed are mean ± SD (n = 3)
Eur J Drug Metab Pharmacokinet
boundaries around the hollow core observed may be due to the coalescence of the gas bubbles formed in the wet beads. 3.1.3.2 Drug entrapment efficiency Drug entrapment in the beads includes drug entrapped within the polymer matrices. The values of total % entrapment efficiency of the drug were in the range of 53.78–76.84 % for dried beads as shown in Table 3. The actual drug content and drug entrapment efficiency was found to be more for the batch FCF with respect to other batch; this may be due to greater extent of crosslinking and thereby greater entrapment efficiency. Fig. 2 SEM photographs of metoprolol tartrate formulation beads
3.1.3.3 Bead porosity and bulk density The bulk density of the beads (Batch no FCG) was less as compared with the batch no FCE and FCF. As bulk density increases it was observed that size and porosity decreases (Table 3). 3.1.3.4 Determination of buoyancy of beads Only beads of batch no FD had a buoyancy lag time of 3 min. Batches FA, FD and FCE produced floating beads remained floating for more than 12 h and FC, FB, FCF and FCG for more than 24 h. The floating property of hollow/porous beads may be attributed to the low bulk density and the porosity of the beads, implying that the beads will have the propensity to exhibit an excellent buoyancy effect in vivo.
Eur J Drug Metab Pharmacokinet
Batches FA, FD and FCE floating beads remained floating for \24 h. This may be attributed because these batches could not maintain matrix integrity for more than 12 h. 3.1.3.5 Determination of swelling index The swelling behavior of the polymer is also an important factor controlling the release of the drugs from the bead systems. The extent of swelling of the formulated beads showed that the swelling was related to different polymer ratio with swelling being more significant for beads with increased gel formation this may be due greater extent of crosslinking between the polymers. The SI was found to be in the range of 1.51–1.95 % as shown in Table 3. 3.1.3.6 Drug release study The dissolution study of all the formulations of metoprolol tartrate beads was carried out in different media namely pH 1.2 and pH 6.8, 7.4 phosphate buffer. All these beads released 6.32–14.02 % of the drug in acidic medium irrespective of time. Batch FCE, FCF and FCG showed 14.02, 8.72 and 6.32 % drug release within 6 h in acidic medium, respectively. There was a slow release for 6 h. After 6 h there was burst release in phosphate buffer and the drug release observed for about 18 h. The drug release profile in phosphate buffer is shown in Fig. 3. The porous beads showed excellent lag time in drug release profile in acidic pH, this may be due to insolubility of pectin. At acidic pH, calcium pectinate and locust bean
Fig. 3 In vitro release profile for formulations
remained protonated into insoluble form with reduced swelling. The second phase of pulsed release in pH 6.8 and 7.4 can be attributed to rapid swelling and gel relaxation of calcium pectinate, locust bean at alkaline pH. 3.1.3.7 In vivo study The in vivo gastric residence of the batch FCF was studied by radiological study (X-ray) of radio-labeled beads using rabbit as animal model. In stomach, the insoluble beads were acted as indigestible food particle. Radiographic image 1–6 (Fig. 4) shows X-ray scans taken on the rabbit during radiological study. It can be interpreted from the images that the beads were clumped together intact and remained floating for 8 h of the study. 3.1.3.8 Stability study In view of potential utility of the formulation, stability study was carried out on batch FCF for 6 months according to ICH guidelines. Formulations were subjected to drug entrapment, floating behavior and in vitro release study after 30, 60, 90 and 180 days. On comparing the optimized formulation with initial data of % entrapment efficiency is 70.03 and cumulative % drug release is 88.79. Result showed (Table 4) that there were no significant changes observed in the appearance, drug entrapment efficiency, buoyancy study and in vitro release analysis of formulation. It confirms that formulation FCF was stable at a temperature of 40 ± 2 °C/75 ± 5 % and photo stable at the end of 180 days.
Eur J Drug Metab Pharmacokinet Fig. 4 Radiographic images taken on the rabbit during radiological studies
Table 4 Stability study Time (days)
Drug entrapment efficiency (%)*
Floating duration (h)
Percent cumulative drug release at the end of 24 h*
0
75.24 ± 6.33
[24
95.28 ± 0.81
30
74.55 ± 6.13
[24
95.54 ± 0.73
60 90
74.25 ± 6.06 73.71 ± 5.70
[24 [24
93.11 ± 1.48 91.83 ± 0.94
180
70.03 ± 5.40
[24
88.79 ± 0.90
* Values expressed are mean ± SD (n = 3)
3.1.3.9 Drug release kinetics Based on regression coefficient values (R2), all the formulations followed first-order drug release kinetics. From Peppas model, it was found that batch no FA, FB, FC, FCE, FCF and FCG showed anomalous transport kinetics, i.e., a combined mechanism of
pure diffusion and Case II transport and batch no FD showed non-Fickian diffusion (Table 5).
4 Conclusion The hollow beads containing metoprolol tartrate showed excellent buoyancy in acidic environment of stomach. The enhanced buoyancy of porous beads makes them excellent candidate for intragastric floating drug delivery, by slowing down the gastric emptying. The pulsatile drug delivery was characterized by rapid and complete drug release from the drug loaded porous beads due to the fast disintegration in the basic medium after a lag time in acidic environment. The release from porous beads was due to faster entry of the gastrointestinal fluid through the weak matrix of the bead in the buffer. Overall, the buoyant beads provided a lag phase while showing gastro
Eur J Drug Metab Pharmacokinet Table 5 Model fitting Batch no.
Zero order R
2
First order n
R
2
Higuchi model n
R
2
Korsmeyer-peppas model n
R2
n
FA
0.7968
4.433
0.9034
-0.040
0.7771
22.24
0.7356
1.164
FB
0.8572
4.624
0.9492
-0.051
0.8124
22.87
0.8049
1.143
FC
0.8299
4.856
0.9462
-0.061
0.8010
24.24
0.7853
1.139
FD
0.7431
4.810
0.9065
-0.055
0.7777
24.99
0.7542
0.987
FCE
0.7902
4.580
0.8987
-0.048
0.8033
23.46
0.8072
0.968
FCF
0.7865
5.083
0.9372
-0.067
0.7707
25.56
0.7585
1.186
FCG
0.8430
4.341
0.9428
-0.039
0.7958
21.42
0.7396
1.200
2
R regression coefficient, n slope
retention followed by a pulsatile drug release that would be beneficial for hypertension. Acknowledgments The authors are highly thankful to Dr. A. D. Taranalli, Principal KLEU’s college of pharmacy, Belgaum for providing all the facilities required for the project. Authors wish to thanks Low methoxy pectin (LMP), was obtained as generous gift sample from Krishna Pectins Pvt. Ltd, Jalgaon (India). Metoprolol tartrate was obtained as generous gift sample from Astrazeneca Pharmaceuticals Pvt Ltd, Bangalore, Karnataka, India. Xanthan gum, Sodium CMC, Guar gum, Locust bean, Gellan gum, Sodium bicarbonate from Hi-media Laboratories Pvt Ltd. Mumbai for providing drug and polymers as a gift sample.
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ORIGINAL PAPER
Development of hollow/porous floating beads of metoprolol for pulsatile drug delivery Sangmesh S. Taranalli • Panchaxari M. Dandagi Vinayak S. Mastiholimath
•
Received: 24 April 2013 / Accepted: 25 March 2014 Ó Springer International Publishing Switzerland 2014
Abstract The purpose of this work was to develop hollow calcium pectinate beads for floating pulsatile release of metoprolol tartrate intended for chronopharmacotherapy. Floating pulsatile concept was applied to increase the gastric residence of the dosage form having lag phase followed by a burst release. To overcome limitations of various approaches for imparting buoyancy, hollow/porous beads were prepared by simple process of acid-base reaction during ionotropic cross-linking using low methoxy pectin, xanthan gum, sodium carboxy methyl cellulose, guar gum, locust bean, gellan gum and calcium chloride as a cross-linking agent. Based on the preliminary studies optimized polymers were selected for formulation design with different polymers ratio concentrations. The obtained floating beads were studied for entrapment efficiency, buoyancy study, swelling index, surface morphology, in vitro release, stability studies and in vivo floating study. The floating beads obtained were porous, float up to 12–24 h. The radiological studies (X-rays) pointed out the capability of the system to release drug in lower parts of GIT after a programmed lag time for hypertension. The floating beads provided expected two-phase release pattern with initial lag time during floating in acidic medium followed by rapid pulse release in phosphate buffer. From the accelerated stability studies, it was observed that the formulations are quite stable. All formulations followed firstorder release kinetics by diffusion mechanism. This S. S. Taranalli (&) P. M. Dandagi Department of Pharmaceutics, KLEU’s College of Pharmacy, Belgaum 590010, Karnataka, India e-mail: [email protected] V. S. Mastiholimath Department of Quality Assurance, KLEU’s College of Pharmacy, Belgaum, India
approach suggested the use of hollow calcium pectinate microparticles as promising floating pulsatile drug delivery system for site- and time-specific release of drugs acting as per chronotherapy of diseases. Keywords Hollow beads Calcium pectinate beads Floating pulsatile drug delivery Metoprolol tartrate Chronotherapy Radiology
1 Introduction A delivery system with a release profile that is characterized by a time period of no release (lag time) followed by a rapid and complete drug release (pulse release) can be called as an ideal pulsatile drug delivery system. Pulsatile delivery system provides one or more rapid release pulses at predetermined lag times or at specific sites resulting in better absorption of the drug, and there by providing more effective plasma concentration time profile (Venkatesh 2005). Natural biodegradable polysaccharides like pectin, locust bean, guar gum, chitosan, sodium alginate and gellan gum have been used in controlled drug delivery. Various approaches to induce buoyancy in cross-linked beads have been used (Whitehead et al. 2000; Iannuccelli et al. 1998; Sriamornsak and Nunthanid 1999). The use of sodium bicarbonate as buoyancy imparting agent to produce floating beads is simplest among the various approaches and has been attempted successfully by many workers (Bussmer et al. 2003). Their floating property was based on the evolution of CO2 when in contact with acidic environment followed by the ability of polymer gel to entrap it, which decreases their density below one. These beads have been used to achieve prolonged gastric residence time, for sustained release/stomach-specific drug delivery providing
Eur J Drug Metab Pharmacokinet
an opportunity for both local and systemic drug action (Rajnikanth et al. 2007). In cardiovascular disease, capillary resistance and vascular reactivity are higher in the morning and decreases latter in the day. Platelet agreeability is increased and fibrinolytic activity decreased in the morning, leading to a state of relative hypercoagulability of the blood. Therefore, the frequencies of myocardial infarction (MI) and of sudden cardiac death are more prone from morning to noon. Ambulatory blood pressure measurements show a significant circadian variation to characterize blood pressure. Increased heart rate, blood pressure, imbalanced autonomic tone, and circulating level of catecholamines controlling the cardiac arrhythmias show important circadian variation and trigger the genesis of the circadian pattern of cardiac arrhythmias. Atrial arrhythmias appear to exhibit circadian pattern usually with a higher frequency in the daytime and lower frequency in the night time with the abnormal foci under the same long-term autonomic regulation as normal pacemaker tissue. According to study, ventricular tachyarrhythmias show late morning peak in the patients with MI sometime in the distant past morning peak and afternoon peak in patients with recent MI. Both pharmacokinetics and pharmacodynamics of some oral nitrates, calcium channel blocker and b-adrenoceptor antagonist medications have been shown to be influenced by the circadian time of their administration (Mandal et al. 2010). Metoprolol is a cardioselective b1-adrenergic blocking agent used for acute MI, heart failure, angina pectoris and mild to moderate hypertension. It may also be used for supraventricular and tachyarrhythmias and prophylaxis for migraine headaches. Metoprolol competes with adrenergic neurotransmitters such as catecholamines for binding at beta (1)-adrenergic receptors in the heart. Beta (1)-receptor blockade results in a decrease in heart rate, cardiac output, and blood pressure (Metoprolol tartrate (internet) 2012). All beta blockers are nearly equally effective in decreasing frequency and severity of attacks and in increasing exercise tolerance in classical Angina, but cardioselective agents. Long-term b-blocker therapy lowers risk of sudden cardiac death among ischemic heart disease patients. In angina pectoris, beta blockers are to be taken on a regular schedule not on as and when required basis (Tripathi 2008). It is moderately lipophilic drug belonging to class I of BCS classification having high solubility and high permeability. Since the half-life of metoprolol tartrate is 3–4 h, two multiple doses are needed to maintain a constant plasma concentration for a good therapeutic response and improved patient compliance (Metoprolol tartrate (internet) 2012). Although it is well absorbed in the gastrointestinal tract, its bioavailability is 40–60 % as a result of extensive first pass metabolism (Metoprolol (internet) 2012). Thus, there is a strong clinical need and market potential for a
dosage form that will deliver metoprolol tartrate in a controlled manner to a patient needing this therapy, thereby resulting in a better patient compliance. Multi-particulate or multiple unit systems offer various advantages over single-unit systems. These include no risk of dose dumping, flexibility of blending units with different release patterns, relative merits of bioavailability more consistent blood levels, reproducible and avoid all or none effect. The aim of the study was to design and characterize hollow/porous floating beads of metoprolol tartrate for pulsatile drug delivery for the treatment of hypertension.
2 Materials and method 2.1 Materials Low methoxy pectin (LMP) was obtained as generous gift sample from Krishna Pectins Pvt. Ltd., Jalgaon, Maharashtra, India. Metoprolol tartrate was obtained as generous gift sample from Astrazeneca Pharmaceuticals Pvt. Ltd., Bangalore (India). Xanthan gum, Sodium CMC, Guar gum Locust bean, Gellan gum and Sodium bicarbonate from Himedia Laboratories Pvt. Ltd., Mumbai, and Calcium Chloride from Loba Chem Pvt. Ltd., Mumbai. All other chemicals used were of analytical grade. 2.2 Method 2.2.1 Formulation of metoprolol tartrate beads 2.2.1.1 Preliminary studies for selection of polymer combination Four formulations were prepared using different polymers as shown in Table 1. Evaluation parameters such as drug entrapment efficiency, buoyancy study of the beads and in vitro drug release for the formulated beads were done and the promising batch was selected on the basis of above parameters for the further study. 2.2.1.2 Ionotropic gelation/bead formation A polymeric solution was prepared by dissolving various amounts of polymers in 15 ml of deionized water. Then, gas-forming agent, such as sodium bicarbonate and metoprolol tartrate (100 mg), was added into the solution (Table 2). The dispersion was sonicated for 30 min to break the lumps and to remove air bubbles. The resultant dispersion was dropped via a 21-gauge syringe needle into 3 % w/v calcium chloride (CaCl2) solution containing 10 % (v/v) acetic acid. The distance between the tip of the needle and the surface of the CaCl2 medium was about 6 cm. The beads formed were stirred at 150 ± 5 rpm using magnetic stirrer for 15 min for curing. The beads were separated by
Eur J Drug Metab Pharmacokinet Table 1 Preliminary studies for selection of polymer combination Batch no.
Drug (mg)
Low methoxy pectin (mg)
Xanthan gum (mg)
Sodium CMC cellulose (mg)
Guar gum (mg)
FA
100
300
50
100
–
FB
100
300
50
–
FC
100
300
50
–
FD
100
300
50
–
Locust bean (mg)
Gellan gum (mg)
Sodium bicarbonate (mg)
–
–
150
100
–
–
150
–
100
–
150
–
–
100
150
Table 2 Formulation code for metoprolol tartrate floating beads Batch no.
Metoprolol tartrate (mg)
Low methoxy Pectin (mg)
FCE
100
300
FCF FCG
100 100
300 300
Xanthan gum (mg)
Locust bean (mg)
Sodium bicarbonate (mg)
Deionized water (ml)
30
120
150
15
60 120
90 30
150 150
15 15
filtration, washed three times with distilled water and subsequently oven dried at 45 °C for 6 h (Dupuis et al. 2006). 2.2.2 Characterization of beads 2.2.2.1 Particle size analysis and surface morphology 100 beads were analyzed for their size distribution by optical microscopy. The mean diameter was determined by measuring the number of divisions covered by beads using ocular micrometer previously calibrated using stage micrometer. The surface Morphology was analyzed with a scanning electron microscope (JEOL JSM-6360 SEM) operated at an acceleration voltage of 5 kV (Pornsak et al. 2005; Gadad et al. 2009). 2.2.2.2 Determination of drug entrapment efficiency Accurately weighed quantities (50 mg) of beads from each batch were placed in 100 ml phosphate buffer, pH 7.4 and mechanically agitated on shaker at 200 rpm for 24 h. The resultant dispersions were filtered and analyzed spectroscopically at 224 nm. The percentage entrapment efficiency was calculated using following equation (Claire et al. 2011). Drug entrapment efficiency % ¼ ðActual drug content in the beads =theoretical drug contentÞ 100: 2.2.2.3 Bead porosity and bulk density The bead porosity was assessed using mercury porosimetry (Autoscan 60 Porosimeter, Quantachrome software, USA). The pressure was applied from 0 to 6,000 psi. The mercury intrusion data were recorded and plotted against pressure. Standard values for the contact angle and surface tension of mercury were used for calculations. The bulk densities of the beads
were also measured using same mercury porosimeter (Bulgarelli et al. 2002). 2.2.2.4 Buoyancy study Floating property of beads was evaluated using USP XXIII type II dissolution test apparatus (Electrolab TDT-06P, Mumbai, India) filled with 900 ml in pH 1.2 containing 0.02 % w/v Tween 80, using paddle at a rotation speed of 100 rpm. The temperature of medium was maintained at 37 ± 2 °C. 100 beads of each batch were placed in the media. Floating ability was observed visually (Huimin et al. 2012; Mastiholimath et al. 2008). 2.2.2.5 Determination of swelling index The swelling behavior of the beads was studied in pH 7.4 phosphate buffer solution. Approximately 100 mg of beads was taken in a dissolution basket and weighed; the baskets along with the beads were immersed in 7.4 phosphate buffer solution. The weight of the basket along with the beads was determined after 1 h, and then every hour. The swelling index (SI) of each formulation was calculated using the following equation (Sandolo et al. 2011; Patel et al. 2011): %SI ¼
W2 W1 100 W1
where, W1 is weight of the dry beads and basket and W2 is weight of the swollen beads and basket. 2.2.2.6 In vitro drug release The dissolution study of beads equivalent to 100 mg of metoprolol tartrate was performed using a USP XXIII type 2 dissolution test apparatus. The drug release study was performed in 900 ml for 6 h in pH 1.2. Then the dissolution medium was replaced with phosphate buffer of pH 6.8 for 3 h, the average small intestinal transit time is about 3 h. After 9 h, the dissolution medium was replaced with phosphate buffer
Eur J Drug Metab Pharmacokinet
of pH 7.4 maintained at 37.0 ± 0.5 °C at 100 rpm for 18 h. Periodically 5 ml of sample was withdrawn and replaced by dissolution media in order to maintain sink condition. The withdrawn samples were filtered through a Wattman filter paper 41 and the concentration of metoprolol tartrate was measured spectrophotometrically at 221.5 and 224 nm for acidic and basic media, respectively, after suitable dilutions (Patel et al. 2011). 2.2.3 In vivo study 2.2.3.1 Radiology (X-rays) The optimized batch was chosen for the preparation of the barium sulfate-loaded beads. The barium sulfate beads were prepared as of optimized batch but the drug was replaced with barium sulfate (100 mg). Healthy rabbit weighing approximately 2.3 kg was used to assess in vivo floating behavior. Ethical clearance for the handling of experimental animals was obtained (CPCSEA). The animal was fasted for 12 h and the first X-ray photographed to ensure the absence of radio opaque material in the stomach. The rabbits were made to swallow barium sulfate-loaded beads with 30 ml of water. During the experiment, rabbits were not allowed to eat but water was provided. At predetermined time intervals, the radiograph of abdomen was taken using an X-ray machine (Gangadharappa et al. 2011). 2.2.3.2 Stability study Stability study has become an integral part of formulation development. It generates information on which proposal for nature of drug or dosage and their recommended storage conditions are based. The accelerated stability study was conducted for selected formulation as per the ICH guidelines. Accelerated testing 40 ± 2 °C/75 ± 5 % RH for 6 months. As per ICH guidelines, beads of promising formulation were selected and subjected to accelerated stability study. Weighed quantity of the samples was kept in glass vials, sealed with rubber plugs and exposed to controlled temperature (40 ± 2 °C) and relative humidity (75 ± 5 %) for a period of 6 months in humidity control oven (Lab Control, Ajinkya IM 3500 Series, India). 2.2.3.3 Photo-stability study A light source that is designed to produce an artificial daylight fluorescent lamp combining visible and ultraviolet (UV) outputs, xenon, or metal halide lamp is used. D65 is the internationally recognized standard for outdoor daylight as defined in ISO 10977 (1993). ID65 is the equivalent indoor indirect daylight standard. For a light source emitting significant radiation below 320 nm, an appropriate filter may be fitted to eliminate such radiation. After 1, 2, 3 and 6 months, the samples were taken out and analyzed for drug entrapment
efficiency, buoyancy study, and in vitro release (Gadad et al. 2009; Amrutkar et al. 2012). 2.2.3.4 Release kinetics One of the most important and challenging areas in the drug delivery field is to predict the release of the active agent as a function of time using both simple and sophisticated mathematical models. The importance of such models lies in their utility during both the design stage of a pharmaceutical formulation and the experimental verification of a release mechanism. In order to identify a particular release mechanism, experimental data of statistical significance are compared to a solution of the theoretical model. To analyze the mechanism for the drug release and drug release rate kinetics of the dosage form, the data obtained was fitted into Zero order, First order, Higuchi matrix and Korsmeyer-Peppas. In this study by comparing the R2 values obtained, the best-fit model was selected. Drug release kinetics can be analyzed by various mathematical models, which are applied considering the amounts of drug released from 0 to 24 h. The following plots were made: Cumulative percentage drug release versus time (zero-order kinetic model); log cumulative percentage drug remaining versus time (first-order kinetic model); cumulative percentage drug release versus square root of time (Higuchi model); and log cumulative percentage drug remaining versus log time (Korsmeyer’s Peppas model) (Lin and Kawashima 1987).
3 Result and discussion 3.1 Pre-formulation study 3.1.1 Compatibility study The FT-IR spectra of pure metoprolol tartrate and the combination of metoprolol tartrate with the polymer show similar characteristic functional peak. The similarity in the peaks indicates the compatibility of metoprolol tartrate with polymers as shown in Fig. 1. 3.1.2 Formulation of floating calcium pectinate beads Based upon the results of preliminary study, FC formulation was selected for further studies because it showed highest entrapment efficiency of 70.89 %, produced floating beads buoyancy for 24 h and highest in vitro drug release of 96.04 %. Three formulations (FCE, FCF and FCG) of metoprolol tartrate beads were made. The formulations were prepared by varying polymer ratio and keeping the concentration of drug and other ingredients same as showed in table: the method used to prepare the calcium pectinate beads by
Eur J Drug Metab Pharmacokinet Fig. 1 FTIR spectra
dripping method using 21-gauge needle into the 3 % calcium chloride solution containing 10 % v/v acetic acid. The beads were formed due to the cross-linking of the pectin with divalent calcium ions of the calcium chloride solution. The reaction between sodium bicarbonate and acetic acid was occurred liberating carbon dioxide as gas bubbles which are responsible for floating of beads. 3.1.3 Characterization of beads 3.1.3.1 Particle size analysis and surface morphology The mean surface diameter of all three formulations was between 1.134 ± 0.018 and 1.198 ± 0.008 (mm),
tabulated in Table 3; this indicates that size of beads increased significantly with increase in viscosity there by subsequent increase in interfacial tension, resulting in the formation of larger particles. Surface morphology of beads batch no FCF was observed. It was shown that beads were spherical in shape, with slightly rougher surface/shrinkage and discrete as shown in Fig. 2. The surface topography reveals that the beads were highly porous because of rapid escape of the carbon dioxide during formulation. The cross section of beads from batch FCF showed a hollow core in the matrix, which may be because of the presence of gas generating agent. The thick matrix
Table 3 Characterization of floating beads Batch code
Particle size (mm)*
Drug entrapment efficiency (%)*
Swelling index (%)*
Bulk density (g/cm3)*
Porosity (%)
Floating ability in pH 1.2 (h)
FCE
1.134 ± 0.02
68.44 ± 3.81
1.51 ± 0.15
1.35 ± 0.05
28.94
[12
FCF FCG
1.176 ± 0.01 1.198 ± 0.01
76.84 ± 6.37 72.72 ± 4.23
1.95 ± 0.13 1.83 ± 0.13
1.13 ± 0.19 0.87 ± 0.13
32.73 34.08
[24 [24
* Values expressed are mean ± SD (n = 3)
Eur J Drug Metab Pharmacokinet
boundaries around the hollow core observed may be due to the coalescence of the gas bubbles formed in the wet beads. 3.1.3.2 Drug entrapment efficiency Drug entrapment in the beads includes drug entrapped within the polymer matrices. The values of total % entrapment efficiency of the drug were in the range of 53.78–76.84 % for dried beads as shown in Table 3. The actual drug content and drug entrapment efficiency was found to be more for the batch FCF with respect to other batch; this may be due to greater extent of crosslinking and thereby greater entrapment efficiency. Fig. 2 SEM photographs of metoprolol tartrate formulation beads
3.1.3.3 Bead porosity and bulk density The bulk density of the beads (Batch no FCG) was less as compared with the batch no FCE and FCF. As bulk density increases it was observed that size and porosity decreases (Table 3). 3.1.3.4 Determination of buoyancy of beads Only beads of batch no FD had a buoyancy lag time of 3 min. Batches FA, FD and FCE produced floating beads remained floating for more than 12 h and FC, FB, FCF and FCG for more than 24 h. The floating property of hollow/porous beads may be attributed to the low bulk density and the porosity of the beads, implying that the beads will have the propensity to exhibit an excellent buoyancy effect in vivo.
Eur J Drug Metab Pharmacokinet
Batches FA, FD and FCE floating beads remained floating for \24 h. This may be attributed because these batches could not maintain matrix integrity for more than 12 h. 3.1.3.5 Determination of swelling index The swelling behavior of the polymer is also an important factor controlling the release of the drugs from the bead systems. The extent of swelling of the formulated beads showed that the swelling was related to different polymer ratio with swelling being more significant for beads with increased gel formation this may be due greater extent of crosslinking between the polymers. The SI was found to be in the range of 1.51–1.95 % as shown in Table 3. 3.1.3.6 Drug release study The dissolution study of all the formulations of metoprolol tartrate beads was carried out in different media namely pH 1.2 and pH 6.8, 7.4 phosphate buffer. All these beads released 6.32–14.02 % of the drug in acidic medium irrespective of time. Batch FCE, FCF and FCG showed 14.02, 8.72 and 6.32 % drug release within 6 h in acidic medium, respectively. There was a slow release for 6 h. After 6 h there was burst release in phosphate buffer and the drug release observed for about 18 h. The drug release profile in phosphate buffer is shown in Fig. 3. The porous beads showed excellent lag time in drug release profile in acidic pH, this may be due to insolubility of pectin. At acidic pH, calcium pectinate and locust bean
Fig. 3 In vitro release profile for formulations
remained protonated into insoluble form with reduced swelling. The second phase of pulsed release in pH 6.8 and 7.4 can be attributed to rapid swelling and gel relaxation of calcium pectinate, locust bean at alkaline pH. 3.1.3.7 In vivo study The in vivo gastric residence of the batch FCF was studied by radiological study (X-ray) of radio-labeled beads using rabbit as animal model. In stomach, the insoluble beads were acted as indigestible food particle. Radiographic image 1–6 (Fig. 4) shows X-ray scans taken on the rabbit during radiological study. It can be interpreted from the images that the beads were clumped together intact and remained floating for 8 h of the study. 3.1.3.8 Stability study In view of potential utility of the formulation, stability study was carried out on batch FCF for 6 months according to ICH guidelines. Formulations were subjected to drug entrapment, floating behavior and in vitro release study after 30, 60, 90 and 180 days. On comparing the optimized formulation with initial data of % entrapment efficiency is 70.03 and cumulative % drug release is 88.79. Result showed (Table 4) that there were no significant changes observed in the appearance, drug entrapment efficiency, buoyancy study and in vitro release analysis of formulation. It confirms that formulation FCF was stable at a temperature of 40 ± 2 °C/75 ± 5 % and photo stable at the end of 180 days.
Eur J Drug Metab Pharmacokinet Fig. 4 Radiographic images taken on the rabbit during radiological studies
Table 4 Stability study Time (days)
Drug entrapment efficiency (%)*
Floating duration (h)
Percent cumulative drug release at the end of 24 h*
0
75.24 ± 6.33
[24
95.28 ± 0.81
30
74.55 ± 6.13
[24
95.54 ± 0.73
60 90
74.25 ± 6.06 73.71 ± 5.70
[24 [24
93.11 ± 1.48 91.83 ± 0.94
180
70.03 ± 5.40
[24
88.79 ± 0.90
* Values expressed are mean ± SD (n = 3)
3.1.3.9 Drug release kinetics Based on regression coefficient values (R2), all the formulations followed first-order drug release kinetics. From Peppas model, it was found that batch no FA, FB, FC, FCE, FCF and FCG showed anomalous transport kinetics, i.e., a combined mechanism of
pure diffusion and Case II transport and batch no FD showed non-Fickian diffusion (Table 5).
4 Conclusion The hollow beads containing metoprolol tartrate showed excellent buoyancy in acidic environment of stomach. The enhanced buoyancy of porous beads makes them excellent candidate for intragastric floating drug delivery, by slowing down the gastric emptying. The pulsatile drug delivery was characterized by rapid and complete drug release from the drug loaded porous beads due to the fast disintegration in the basic medium after a lag time in acidic environment. The release from porous beads was due to faster entry of the gastrointestinal fluid through the weak matrix of the bead in the buffer. Overall, the buoyant beads provided a lag phase while showing gastro
Eur J Drug Metab Pharmacokinet Table 5 Model fitting Batch no.
Zero order R
2
First order n
R
2
Higuchi model n
R
2
Korsmeyer-peppas model n
R2
n
FA
0.7968
4.433
0.9034
-0.040
0.7771
22.24
0.7356
1.164
FB
0.8572
4.624
0.9492
-0.051
0.8124
22.87
0.8049
1.143
FC
0.8299
4.856
0.9462
-0.061
0.8010
24.24
0.7853
1.139
FD
0.7431
4.810
0.9065
-0.055
0.7777
24.99
0.7542
0.987
FCE
0.7902
4.580
0.8987
-0.048
0.8033
23.46
0.8072
0.968
FCF
0.7865
5.083
0.9372
-0.067
0.7707
25.56
0.7585
1.186
FCG
0.8430
4.341
0.9428
-0.039
0.7958
21.42
0.7396
1.200
2
R regression coefficient, n slope
retention followed by a pulsatile drug release that would be beneficial for hypertension. Acknowledgments The authors are highly thankful to Dr. A. D. Taranalli, Principal KLEU’s college of pharmacy, Belgaum for providing all the facilities required for the project. Authors wish to thanks Low methoxy pectin (LMP), was obtained as generous gift sample from Krishna Pectins Pvt. Ltd, Jalgaon (India). Metoprolol tartrate was obtained as generous gift sample from Astrazeneca Pharmaceuticals Pvt Ltd, Bangalore, Karnataka, India. Xanthan gum, Sodium CMC, Guar gum, Locust bean, Gellan gum, Sodium bicarbonate from Hi-media Laboratories Pvt Ltd. Mumbai for providing drug and polymers as a gift sample.
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