AAcadcadeemmicic SciScieenncceess

International Journal of Pharmacy and Pharmaceutical Sciences

ISSN- 0975-1491 Vol 4, Issue 3, 2012

Re s e arch A r t i c l eIN VITRO EVALUATION OF FLOATING MATRIX TABLETS OF AMOXICILLIN AND

METRONIDAZOLE FOR THE ERADICATION OF HELICOBACTER PYLORI

LAILA H. EMARA1, AYA R. ABDOU1, AHMED A. ELASHMAWY*1, RANIA M. BADR1,NADIA M. MURSI2

1Industrial Pharmacy Laboratory, Medical and Pharmaceutical Chemistry Department, Division ofPharmaceutical Industries, National

Research Centre, ElTahrir Street, Dokki, Giza 12622, Egypt. 2Department of Pharmaceutics, Faculty ofPharmacy, Cairo University, Cairo,

11562, Egypt. Email:as h m a w y a @yah o o . c o m

Received: 20 Mar 2012, Revised and Accepted:05 May 2012

ABSTRACTThe aim of this study was to develop floating tablets for amoxicillin trihydrate (AmoxTH) and metronidazole, for thetreatment of Helicobacter pylori (H.pylori) mediated peptic ulcer. Tablets were developed based on gasformation techniqueto prolong gastricresidence time with the desired in vitro release profile. Poly(ethylene oxide) (PEO) andhydroxypropylmethylcellulose (HPMC) of various grades were used as swellable polymers, while NaHCO3 and CaCO3 wereused as the gas generating agents. Tablets were evaluated for physical parameters, foating characteristics and in vitrorelease rate studies using a well controlled openloop system of the fowthrough cell apparatus. Results showed that theshortest floating lag time values recorded were 0.40 and 1.00 min, corresponding to AmoxTH tablets containing 150mg HPMCK4M and metronidazole tablets containing 240 mg PEO of MW 8,000,000, respectively. The relation betweenfoating lag time and MW of PEO was directly proportional with AmoxTH tablets, and inversely proportional with metronidazoletablets. Tablets of both drugs were able to remain foating, without disintegration, for more than 6.0 hours in 0.1N HCl (pH1.2), except in case of AmoxTH/PEO of MW 100,000 and metronidazole/PEO of MW 100,000, 300,000 and 900,000. Thehighest percentage of AmoxTH released (82.1%) was obtained from the doublelayer tablet prepared with PEO of MW8,000,000, in the gas generating layer and PEO of MW 900,000 in the drugreleasing layer. While, the highest percentage ofmetronidazole released (75.3%) was obtained from the singlelayer foating tablet containing PEO of MW 100,000. Under thesame storage conditions, AmoxTH doublelayer tablet was found to show similar release profiles before and after storage,while metronidazole showed higher release rate profile.Keywords: Gas formation technique, Amoxicillin trihydrate, Metronidazole, Flowthrough cell apparatus,Floating tablets, Storage.

INTRODUCTIONDrug release from new drug delivery systems can besustained for up to 24 h for many drugs using currentrelease technologies. However, the real challenge in thedevelopment of oral controlled release dosage forms isto prolong the residence time of dosage forms in thestomach or upper gastrointestinal (GI) tract until the drugis completely released1. Rapid GI transit could result inincomplete drug release from the drug delivery system inthe absorption zone leading to diminished efficacy of theadministered dose2. Several approaches are currentlyused to retain the dosage form in stomach. These includedifferent systems e.g.: foating3,4, bioadhesive5, swellingand expanding systems6,7 as well as other delayed gastricemptying devices8,9. The principle of floating preparationsoffers a simple and practical approach to achieveincreased residence time for the dosage form in stomachand sustained drug release10,11.H.pylori is one of the major causative agents of pepticulcer12. According to the guidelines presented by theEuropean Helicobacter Study Group (EHSG)1317, ulcerpatients who are H.pylori positive require treatment withantimicrobial agents, in addition to antisecretory drugs.Although many antibacterial agents have very lowminimum inhibitory concentration (MIC) against H.pylori invitro18, no single agent is effective in the eradication ofthe infection in vivo when administered alone19. The MIC50values (concentrations resulting in 50% inhibition) ofamoxicillin and metronidazole, for instance, are as low as0.008 g/ml and 2 g/ml, respectively18. In addition, singleantibiotic therapy is strongly discouraged to prevent thedevelopment of resistant strains20. There could be one orseveral reasons for the failure of singleantibiotic therapy

against H.pylori. Firstly, the organism resides in themucus gel close to the acidic environment of thegastric fuid. Many antibacterial agents, such as penicillinand erythromycin, degrade rapidly in acidic medium.Secondly, the drug must diffuse into the mucus layerand the bacterial glycocalyx to furnish concentrationssufficient for antibacterial activity. For eradication ofH.pylori in the stomach the concentrations of antibacterialagents reaching the site of infection from tablets orcapsules might not be bactericidal against organismslocated in the mucus layer and protected by theglycocalyx. Lastly, the contact time of antibacterial drugswith the organism needs to be

sufficiently long for successful eradication of H.pylorifrom the gastric mucosa21, which can be achievedthrough a gastroretentive dosage form.Amoxicillin is the 4hydroxy analogue of ampicillin, and isused in a similar variety of susceptible infections. It isgiven as part of treatment regimens to eradicate H.pyloriinfection in patients with peptic ulcer disease22.Metronidazole is a 5nitroimidazole derivative with activityagainst anaerobic bacteria and protozoa. It is also used toeradicate H.pylori in peptic ulcer disease, in combinationwith antimicrobials, and either bismuth compounds orproton pump inhibitors23.Therefore, certain amoxicillin or metronidazole foatingsystems were developed for better eradication ofH.pylori, including: amoxicillin foating minimatrices24,gellan based amoxicillin foating in situ gelling systems25,amoxicillin foating tablets2628, amoxicillin floatingalginate beads29, foating metronidazole tablets3035 andbeads3639.Poly(ethylene oxide) (PEO) is among various hydrophilicpolymers that, in the presence of water, form a hydrogelthat could control the release of the active moiety

either by swelling or by swelling/erosion. PEOs havebeen proposed as alternatives to cellulose or otherethylene glycol derivatives in the production of controlledrelease drug delivery systems40. The rate and kinetics ofdrug release from hydrophilic matrix is dependant onvarious factors such as types of polymer, solubility ofdrug, polymer content, particle size of drug and polymeras well as types and amounts of excipients used in theformulation41,42.Hydroxypropylmethylcellulose (HPMC), a semisyntheticcellulose derivative, is widely used as a matrix in oralcontrolled release tablet formulations43,44. Its widespreaduse is mainly due to its generally regarded as safe(GRAS) status and biodegradable nature. Furthermore, itis compatible with numerous drugs, accommodates highlevels of drug loading and can be easily incorporated toform matrix tablets by direct blending or granulation44,45.The availability of a wide range of viscosity grades alsoallows the formulator to modify the release of drugsfrom HPMC matrix tablets according to the therapeuticneed. Drug release from HPMC matrix tabletinvolves complex mechanisms46. In contact with water,HPMC swells

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to form a gel, which serves as a barrier to drugdiffusion. Drug release from the HPMCdrug matrixinvolves solvent penetration into the dry matrix,gelatinization of the polymer, dissolution of the drug anddiffusion of the solubilized drug through the gel layer.Concomitantly, outer layers of the tablet become fullyhydrated and dissolve, a process generally referred to aserosion47.The objective of this study was to develop floating tabletsof amoxicillin and metronidazole for treatment of pepticulcers caused by Helicobacter pylori. PEO and HPMC wereused as swellable polymers for their foating and drugrelease retardant properties48. An effervescent mixture,for tablet floating, was used to release carbon dioxide inpresence of acidic medium. Tablets were prepared bydirect compression as a low cost method with no drugexposure to harsh conditions such as hightemperature49. The effect of different formulationvariables on the tablet floating characteristics as well asthe in vitro release rate profile was investigated.MATERIALS AND METHODS MaterialsAmoxicillin Trihydrate (AmoxTH) and metronidazole weregifted from EIPICO (Egypt). Poly(ethylene oxide) (PEO)[molecular weights (MW) 100,000, 300,000, 900,000,4,000,000, and 8,000,000] andhydroxypropylmethylcellulose (HPMC): K4M, K15M andK100M, were purchased from Aldrich (Germany). Sodiumbicarbonate (NaHCO3) was from Kahira Pharm. (Egypt).Calcium carbonate (CaCO3) and lactose were from BDHLaboratory Supplies (England). Magnesium stearate wasobtained from Peter Greven (Nederland, Germany) andmethanol (HPLC grade) was from Prolabo (France).Distilled water was used for all experiments (Milli RO plus10, Millipore, USA). All other reagents were of analyticalgrade.TabletPreparationFormulation of AmoxTH (375 mg/tablet) or metronidazole(250 mg/tablet) in swellable effervescent floating tabletswas carried out as single and double layer tablets.These tablets were formulated with the use of swellablepolymers (PEO or HPMC) and an effervescent mixturecomposed of sodium bicarbonate and calcium carbonatein the ratio of (1:2)35, respectively. The final weight of thetablet was adjusted to 0.7 gm by adding lactose as filler.

All ingredients (for each formula) in their specified ratios(Tables 1,2 and 3), were sieved through 710 m sieve (meshnumber 25)except for magnesium stearate which was sieved through425 m sieve (mesh number 40). Blending of allingredients was carried out simultaneously usingpolyethylene bag50, after which tablets were preparedfrom different blends by direct compression at 1.5tonscompression force (Single Punch Press Tablet Machine,Stokes Merrill Model 5117A, USA). For such formulae, around die (13 mm internal diameter) with flatfacedpunches were employed to give round flatsurface tablets.Characterization of the preparedtabletsPhysicalparametersAutoTest4 (automatic tablettesting system, Dr.Schleuniger Pharmatron, Germany) was used fordetermination of thickness, diameter, weight, andhardness of the prepared tablets (mean of twentytablets for each formula was calculated).ContentUniformityTwenty tablets of each formula were weighed, grinded,and the weight equivalent to one tablet was transferredquantitatively into100 ml glassstoppered volumetric fask. The volume wasthen completed to the mark with either 0.1 N HCl (pH 1.2)or dilute HCl (1in 100) for AmoxTH24 and metronidazole51 tablets,respectively. The volumetric flasks were shaken using"temperaturecontrolled shaking waterbath (LabLine,USA)" for 30 min in 37C water bath. The solution wasthen filtered, and the absorbance was measuredspectrophotometrically at the predetermined max at 272and 277 forAmoxTH and metronidazole,respectively24,51.Determination of Floating Lag Time and FloatingDuration of the Prepared TabletsThe time required for the tablets to emerge on thedissolution medium surface (foating lag time) andthe time the tablets remained foating on thedissolution medium surface (foating duration) wereinspected visually in 1L jacketed jar connected toJulabo circulator (F10VC, Germany), filled with 400 ml 0.1N HCl, pH1.252, at 370.5C53. The results were registered as anaverage of three repetitions.

TabletCode

Table 1: Composition of AmoxTH foating tablets (375 mg/tablet)

Tablet Composition (mg) aPEO Molecular Weight HPMC Grade Calcium

Sodium

8,000,000

4,000,000

900,000

300,000

100,000

K4M K15M K100M Carbonate BicarbonateA1 240 0 0 0 0 0 0 0 40 20

A2 0 240 0 0 0 0 0 0 40 20A3 0 0 240 0 0 0 0 0 40 20A4 0 0 0 240 0 0 0 0 40 20A5 0 0 0 0 240 0 0 0 40 20A6 0 0 0 0 0 150 0 0 40 20A7 0 0 0 0 0 225 0 0 40 20A8 0 0 0 0 0 0 150 0 40 20A9 0 0 0 0 0 0 0 150 40 20

a 1% magnesium stearate was used as lubricant.

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Table 2: Composition of metronidazole foating tablets

(250 mg/tablet) Tablet Code Tablet Composition (mg) a P EO Molecu lar We igh t Calcium Carbonate Sodium Bicarbonate

8,000,000

4,000,000

900,000

300,000

100,000M1 240 0 0 0 0 40 20

M2 0 240 0 0 0 40 20M3 0 0 240 0 0 40 20M4 0 0 0 240 0 40 20M5 0 0 0 0 240 40 20M6 120 0 120 0 0 40 20a 1% magnesium stearate was used as lubricant.

Table 3: Composition of the double layer foating tablets

Layer Function Total Weight ofEach Layer (mg)

Composition a (mg)

1 Swellable gas generating layer 200 PEO of MW 8,000,000 1402 Swellable / sustainable Drug containing

layer500 Calcium carbonate 40

Sodium bicarbonate 20PEO of MW 900,000 75AmoxTH b 375Lactose 50Or 75PEO of MW 900,000 250Metronidazole c 175Lactose

a 1% magnesium stearate was used as lubricant; b Tablet A.d.; c Tablet M.d.

In vitro releasestudiesThese studies were carried out using the openloopsystem of the flowthrough cell, USP Apparatus 4, which iscomposed of Dissotest CE6 equipped with a CY 750piston pump (Sotax, Switzerland). Each tablet wasplaced into the large dissolution cell (22.6 mm diameter)according to the cell design shown in Figure (1), as thisdesign allowed for foating observations while studyingthe release profile. Built in filtration system (0.7 mWhatmann GF/F and GF/D glass microfiber filters, andglass wool) was used throughout the study. Thedissolution medium was 0.1N HCL (pH 1.2), which wasfiltered (on 0.45 m filter), degassed, and then pumpedat a laminar flow rate of 8.0 0.2 ml/min.Temperature of the

dissolution medium was kept constant at 37 0.5 C. Atpredetermined time intervals, volume fractions werecollected and then analyzed spectrophotometrically forAmoxTH or metronidazole content by measuring theabsorbance at the corresponding max (272 nm forAmoxTH and 277 nm for metronidazole) against 0.1NHCl (pH 1.2) as blank. Each formula was tested intriplicate for up to 6.0 h and the mean value wascalculated.The release kinetics was computed by fitting the releaserate data to zeroorder, firstorder, secondorder, Higuchi,and HixsonCrowell cuberoot models. The criteria forselecting the most appropriate model were based onthe best goodness of fit and the smallest sum of squaredresiduals (SSR)5457.

Fig. 1: Dissolution cell design used for the prepared floating tablets: laminar flow with free tablet position (glass beads filling the entry cone)

Effect of storage on drugreleaseThe double layer foating tablet for each drug (A.d andM.d, cf. Table3) was stored at ambient conditions for 12 months, inamber tightly closed glass containers away from directlight. After 12 months the tablets were reevaluated fortheir appearance, floating properties,and in vitro release profile. The similarity factor (2), asproposed byMoore and Flanner58 was calculated from the meanrelease data and used to evaluate the effect of storage

on the release profile. 2 is defined as:

2 = 50 x log {[1+ ( 1/n ) t=1n ( Rt Tt ) 2 ]0.5 x 100}

Where, n is number of data time points collected duringthe in vitro release test, Rt and Tt are the cumulativerelease percentages released at the selected (n) time

point of the fresh and stored tablets, respectively.The (2) value is a measure of the similarity between twodissolution curves and its value ranges from 0 and100. A high (2) value indicates high similaritybetween two release rate profiles. FDA

suggests that two dissolution profiles are considered similar if the similarity factor 2 is between 50 and 10059,60.RESULTS ANDDISCUSSIONPhysical parameters of foatingtabletsWeightvariationAll the prepared foating tablets showed acceptable weightvariation range. The average tablet weight ranged from668.0 to 734.0 mg for AmoxTH formulations and from666.6 to 729.6 mg for metronidazole formulations. Notmore than two tablets deviated from the averageweight by more than 5.0% (percentage deviation foruncoated and filmcoated tablets weighing more than 250mg), and none deviated by more than twice thatpercentage61.Thickness anddiameterThe prepared tablets showed good uniformity of thickness and diameter. The values of tablet thickness were in the range of 4.70 5.35 mm for AmoxTH formulations and from 4.31 to 4.83 mm for metronidazole formulations. The average diameter ranged from

12.66 to 12.76 mm for AmoxTH formulations andfrom 12.65 to12.69 mm for metronidazoleformulations.HardnessAmoxTH formulae average tablet hardness wasbetween 9.90 and12.30 kP, while that of metronidazole formulae wasbetween 4.80 and 6.70 kP.ContentuniformityThe drug concentration was not less than 92% and didnot exceed100% of the labeled claim. This result indicated that allthe prepared formulations complied with the limits ofpharmacopoeia for content uniformity, i.e., the averagepercentage of drug content of all formulae was foundto be within the range of 85% and 115% of the labelclaim62.In vitroevaluationTables (4 and 5) summarize the results of floating lagtime, floating duration as well as release characteristicsfor the prepared AmoxTH and metronidazole tablets.

Table 4: In vitro release data mean and foating behavior of AmoxTH floating tablets (375 mg/tablet)

Tablet Code Floating Lag Time (min)

Floating Duration (min)

Release% Release ModelA1 4.2 >360 12.89 ZeroorderA2 2.3 >360 15.03 ZeroorderA3 1.6 >360 30.76 ZeroorderA4 1.4 >360 38.71 ZeroorderA5 a 53.40 Hixson and Crowell CubeRootA6 0.40 >360 23.87 FirstorderA7 0.88 >360 18.19 FirstorderA8 2 >360 26.95 ZeroorderA9 5.25 >360 18.87 Zeroordera Failed to float and disintegrated.

Table 5: In vitro release data mean and floating behavior of metronidazole foating tablets (250 mg/tablet)

Tablet Code Floating Lag Time (min)

Floating Duration (min)

Release % Release ModelM1 1 > 360 14.93 FirstorderM2 3.5 > 360 14.88 FirstorderM3 8 120 33.86 ZeroorderM4 a 49.59 ZeroorderM5 a 75.29 Hixson and Crowell CubeRootM6 7.5 > 360 16.09 SecondOrdera Failed to float and disintegrated.

Floating lag time and foatingdurationWhen designing gastric floating drug delivery system, thetwo mechanisms most frequently used are low density oftablet ingredients and the use of gas generatingagents63. A lowdensity drug delivery system could beachieved by using PEO and/or HPMC in a polymericdelivery system. Upon contact with water, a hydrogellayer will be formed to act as a gel boundary for thedelivery system. The other mechanism of foatation, gasgeneration, was also incorporated into these tablets. Thiswas achieved by incorporating sodium bicarbonate andcalcium carbonate into the delivery system. When waterpenetrates into the tablet, generation of carbon dioxideoccurs, which becomes trapped in the polymeric systemand helps the foatation of the delivery system63.Single layertabletsFigure (2) shows the foating lag time values of AmoxTHand metronidazole tablets based on PEO. In case ofAmoxTH/PEO based tablets, there was a direct relationship

between PEO molecular weight and foating lag timevalues, where decreasing PEO molecular weight from8,000,000 to 300,000 (Tablets A1A4) decreased foatinglag time from 4.2 to 1.4 min, while with further decreaseof PEO molecular weight (i.e. 100,000), tablets (A5) failedto float (cf. table 4).On the other hand, in case of metronidazole tablets, therelation between the PEO molecular weight and foatinglag time was inversely proportional (cf. Figure 2). Also,tablets prepared with lower MW PEO

of 300,000 (M4) & 100,000 (M5) failed to foat. Theseresults might be due to the different physicochemicalproperties of the two drugs.It is worthy to mention that the tablets (A5, M4 and M5)disintegrated rather than swelled and hence failed to foat.This means that all the produced carbon dioxide escapedinto the test medium rather than being entrapped withinthe tablet. These results might be considered if it will berequired to prepare a combined floating dosage form ofthese two drugs for improved patient compliance.Metronidazole tablets containing (1:1) blend of PEO of MW900,000 and 8000,0000 was prepared (M6). It was foundthat the floating lag time was governed by the lower PEOmolecular weight (cf. Table 5). As the floating lag time

values were 8.0 min versus 7.5 min for tablet made withpure PEO of MW 900,000 (M3) and the 1:1 PEO blend(M6), respectively, while foating lag time of PEO of MW8,000,000 (M1) was 1.0 min (cf. Table 5).In case of AmoxTH/HPMC based tablets (cf. Table 4), itwas found that an increase in HPMC (K4M) from 150mg/tablet (A6) to 225 mg/tablet (A7), increased thefloating lag time by more than 50% (i.e. from 0.40 minto 0.88 min). Therefore, 150 mg/tablet HPMC wasselected to study the effect of different HPMC viscositygrades on the floating lag time. Figure 3, shows thatthere was a direct relation between increasing HPMCviscosity and foating lag time. The floating lag timevalues were 0.40, 2.00 and 5.25 min for tablets A6 (K4M),A8 (K15M) and A9 (K100M), respectively.

Fig. 2: Chart of floating lag time values of AmoxTH and metronidazole tablets

Fig. 3: Chart of floating lag time values of AmoxTH tablets based on different HPMC grades

Double layertabletsA double layer tablet for each drug was prepared tostudy this system and its impact on both floatingand drug release properties. The gasgenerating layerwas prepared with PEO of MW8,000,000, calcium carbonate and sodium bicarbonate.The drug releasing layer, containing either AmoxTH ormetronidazole, was prepared with PEO of MW 900,000and lactose (c.f. table 3).

The results revealed that the floating lag time values,which corresponds to PEO of MW 8,000,000, wereincreased in case of the double layer tablets compared tothe single layer tablets for both drugs. It was found thatthe floating lag time values were extended to be 15.0min versus 4.2 min in case of AmoxTH double (A.d) andsingle layer (A1) tablets, respectively. While in caseof metronidazole, the foating lag time values were 20.0versus 1.0 min for the double (M.d) and single layer (M1)tablets, respectively (c.f. Table 4 and 5). Theprolongation of foating lag time values,

observed in double layer tablets of both drugs, might bedue to the less amount of PEO of MW 8000,0000 used inthe double layer tablet which was 140 mg compared to240 mg in single layer tablet (cf. Tables 13). However,these foating lag time values were still less than thegastric emptying time64.The foating duration was more than 6.0 h for thedouble layer tablets of both drugs. This study could beextended to lessen the floating lag time to preventpremature emptying from the stomach.It was interesting to point out that all tablets floated inless than 0.50 min in the dissolution cell during the invitro flowthrough cell dissolution study (dynamic test),compared to the previously described floating behaviorstudy52 (static test). Also, this observation had beenpointed out by Rosa et al.52 where sotalol HCl tabletsfloated after 27.7 min in the beaker method understati c conditions and after 10.3 min during the dissolutiontest using USP basket method.

In vitro drugreleaseA specific in vitro release method for evaluation of gastroretentive AmoxTH formulations in acidic test medium wasadopted based on a previous study using an openloopsystem of the flowthrough cell65, which provides acontinuous fow of fresh dissolution medium during thestudy (i.e. perfect sink condition). This method wasdesigned to overcome the errors in the calculation of theamount of AmoxTH released without the interference ofAmoxTH degradation product. The study suggested thatfor evaluation of gastroretentive AmoxTH products, thebeaker method as well as USP paddle and basket are notrecommended if the analysis of AmoxTH will be carriedout by UV/spectrophotometer. The openloop system ofthe flowthrough cell was safely applied to evaluate suchproducts, where AmoxTH was accurately analyzed byUV/spectrophotometric. On the other hand, the USP paddleand basket methods are not recommended if the analysisof AmoxTH will be carried out by UV/spectrophotometer,where in such cases samples should be analyzed byHPLC65.

Single layertabletsPEO basedtabletsFigures (4 and 5) show the drug release rate profiles ofAmoxTH and metronidazole from tablets containing PEOof different molecular weight, respectively. The drugrelease rate studies revealed that PEO of the highestmolecular weight (MW 8,000,000) gave the lowest drugrelease percentage after 6.0 h (12.89% and14.93% for AmoxTH and metronidazole tablets,respectively) which might be due to its low erosionproperty. The highest drug release was obtained from thelower molecular weight (PEO MW 100,000) which gave50.94% and 75.29% for AmoxTH and metronidazoletablets, respectively. Therefore, it is obvious that therelease rate from PEO matrices was inversely proportionalto PEOs molecular weight. Similar results were reportedin another study by Ali et al.48, on Celecoxid floatingformulations containing PEO of MW 2,000,000 and4,000,000.

Fig. 4: AmoxTH release rate profles from the single layer foating tablets containing PEO of different molecular weights

Fig. 5: Metronidazole release rate profles from the single layer floating tablets containing PEO of different molecular weights

In a trial to obtain a reasonable release rate withminimum foating lag time, tablets containing (1:1) blendof PEO of MW 900,000 and8000,0000 were prepared (M6) (cf. table 2). Resultsrevealed thatthe release rate was governed by the highmolecular weight polymer, where metronidazole releasepercentages after 6.0 h were14.93% versus 16.09% for tablets made with purePEO of MW8000,000 (M1) and the 1:1 PEO blend (M6), respectively(cf. Figure5). On the other hand, the foating lag time wasgoverned by the lower molecular weight polymer asshown in Table 5. Therefore, this might be a challenge forthe formulator scientist.Also, metronidazole release rate could be tuned byfurther changes in the polymer blend ratio. In a studycarriedout by Korner el al.66, hydrophilic matrix tabletswere made by mixing PEO of MW 100,000 and 2,000,000in different ratios (10:90, 30:70, 50:50, 70:30 and90:10). They reported that the polymer (PEO) dissolutionrate decreased with increasing the fraction of the longpolymer (2,000,000) in the tablet.HPMC/AmoxTH basedtabletsFigure 6 shows AmoxTH release rate profiles from A6, A7,A8 and A9 tablets containing 150 (HPMC K4M), 225 mg(HPMC K4M), 150 mg

(HPMC K15M) and 150 mg (HPMC K100M),respectively. It was found that when the polymer contentincreased from 150 mg (A6) to225 mg (A7), the percentage of AmoxTH released after6.0 h decreased slightly from 23.87 to 18.19%. This mightbe due to an increase in tortuosity and length of thediffusion path through the matrix67. Tablets A6 and A8containing 150 mg HPMC K4M and K15M, respectively,showed almost the same release rate profiles (23.87%and 26.95% released after 6.0 h, respectively), while thetablet containing K100M (A9) showed the least AmoxTHpercentage released after 6.0 h (18.87%). This mightbe due to the fact that HPMC K4M has relatively low gelstrength, least entanglement and smallest diffusion pathlength compared to HPMC K100M68. In addition, asdescribed by Siepmann and Peppas69, the overall drugrelease mechanism of HPMC matrices is sequentiallygoverned as follows: (i) At the beginning, steep waterconcentration gradients are formed at the polymer/waterinterface resulting in water imbibition into the matrix. (ii)Due to the imbibition of water, HPMC swells resulting indramatic changes of polymer and drug concentrationsand increasing dimensions of the system. (iii) Uponcontact with water, the drug dissolves and diffuses out ofthe device due to concentration gradients. (iv) Withincreasing water content, the diffusion coefficient of thedrug increases substantially69.

Fig. 6: AmoxTH release rate profles from the single layer floating tablets containing HPMC of different grades

Double layertablets

In vitro releasestudy

Figure (7) shows the photograph of the double layer tabletin the flow through cell. Figure 7 (B) shows that the twolayers swelled as a result of fluid ingress, and as a resultof the gradual and continuous generation of gas in thefirst layer, the tablet gradually foated. The drug releaserate was controlled by polymer swelling, matrix erosion,drug diffusion and the gel thickness dynamics69. Thus bothfoating and drug release could be optimizedindependently.

Figure (8) shows the release rate profiles for AmoxTH andmetronidazole double layer tablets against theircorresponding single layer tablets containing PEO of MW900,000. The results revealed that the percentages ofdrug released after 6.0 h were

82.13% and 57.64% for AmoxTH (A.d) and metronidazole(M.d) double layer tablets, respectively. While in caseof single layer tablets containing PEO of MW 900,000, thepercentages of drug released after 6.0 h were 30.76% and33.86% for AmoxTH (A3) and metronidazole (M3),respectively. It is worthy to mention that the higherrelease percentages of both drugs in case of the doublelayer tablets might be due to the effect of drug/polymerratio that have been increased in case of the double layertablets (cf. Tables 13).

Effect of storage on therelease rateFigures (9 and 10) show the release rate profiles ofdouble lay er floating tablets of AmoxTH (A.d) andmetronidazole (M.d) before and after storage forone year at room temperature. After storage,foating lag time and duration as well as the physicalappearance of AmoxTH and metronidazole tablets werenot affected.AmoxTH tablets exhibited similar release rateprofiles after 1 year storage (2 values > 50), whichmight indicate good stability. On the other hand,metronidazole tablets showed a pronounced increase

in the drug release rate after 1 year storage.The similarity factor 2 value for the metronidazoletablets was 25.0, which was out of the FDA limit ofacceptance 59. Therefore, metronidazole tablets(M.d) was not considered stable under thesestorage conditions as indicated by the 2 value ofthe release rate data.This different behavior observed for AmoxTH andmetronidazole tablets may be due to the variationbetween type and degree of drugpolymer interaction,changes of degree of crystallinity or amorphous phasesof the drug and drug molecular weight in each case,which could lead to the instability of the stored tablets.

Fig. 7: Photographs of AmoxTH double layer tablet (A.d) in the flowthrough cell.Key: A: Cell filling, gasgeneration and swelling of the tablet; B: Magnified photograph of the swelled tablet; C: Floated tablet

Fig. 8: Release rate profiles for PEO of MW 900,000 based foating tablets (double versus single layer). Key: A: AmoxTH tablets; B: Metronidazole tablets

Fig. 9: AmoxTH release rate profles from the double layer foating tablets before and after one yearstorage at ambient conditions

Fig. 10: Metronidazole release rate profles from the double layer foating tablets before and after one yearstorage at ambient conditions

Other researchers had similar results of instability ofmetronidazole/PEO formulae of MW 1,000,000 and7,000,000 by storage70. In case of lower molecular weightpolymer (1,000,000), a significant increase inmetronidazole release (f2 < 50) was observed afterstoring the samples under stress conditions. The reasonbehind this phenomenon was reported to be the result ofstructural changes of PEO, which lead to strongerpolymerpolymer interaction, resulting in the decrease ofthe strength of the secondary bonds formed between thepolymer chains and the active ingredient molecules. Onthe other hand, no such changes was seen in the case ofthe higher molecular weight form, although earlierstudies by these group of researchers confirmedstructural alterations similar to those of the low molecularweight polymer70. This suggested that not only themodified physical properties of the polymer matrixdetermine the behavior of the dosage form in the courseof storage but also the characteristics of the molecules.Also, the authors reported that, in the case oftheophylline, drug release of high molecular weightmatrices increased to an even greater extent70. Inaddition, it was reported that metronidazole release ishigher than

that of theophylline, which can partly be attributed to thegreater density of metronidazole molecules in thepolymer matrix. This could lead to the higherconcentration gradient of metronidazole, resulting in thefaster diffusion of this drug. Another contributing factorwas found to be that the interaction between theophyllineand PEO was stronger, the energy gain of the Hbondbetween these two molecules was greater than in case ofmetronidazole70.

Kinetic study of drugrelease data

By applying the linear regression method and subjectingthe release rate data to different release kinetics andmechanisms (zeroorder, firstorder, secondorder,Higuchi diffusion and HixsonCrowell), most of AmoxTHtablets were found to follow mainly the zeroorder releasemodel, except for the tablets containing HPMC:150 mg/tablet and 225 mg/tablet K4M (A6 and A7,respectively) which followed firstorder model. Meanwhile,tablets (A5) which contained PEO of MW 100,000 andAmoxTH double layer tablets (A.d) followed Hixson and

Crowell cuberoot model (cf. Table 4).

While for Metronidazole floating tablets, results revealedthat tablets (M1) and (M2) containing PEO of MW8,000,000 and 4,000,000, respectively, followed the firstorder release model, while tablets (M3) and (M4)containing PEO of MW 900,000 and 300,000 respectively,followed the zeroorder release model. Tablets (M5)containing PEO of MW 100,000 followed Hixson andCrowell Cube Root model. Tablets (M6) that contained(1:1) blend PEO of MW8,000,000 and 900,000, as well as metronidazoledouble layer tablets (M.d) followed the secondorderrelease model (cf. Table 5).CONCLUSIONSThis study demonstrates the feasibility of prolonging thegastric residence time of antiH.pylori drugs via oraladministration of the proposed floating tablets.Furthermore, sustained release of the model drugs(metronidazole and AmoxTH) from such foatingtablets can be achieved over a period of at least 6.0 h.These dosage forms need further in vitro optimization, toshorten the foating lag time as well as a clinical trialinvolving patients suffering from peptic ulcer.This stomach targeted dosage form could maintain theminimum inhibitory concentration for sufficient time toallow for local eradication and thereby achieve betterefficiency of therapy with improved patient compliance,reduced costs and minimized side effects caused byimmediate release dosage forms.REFERENCES1. Baumgartner S, Kristi J, Vrecer F, VodopivecP, Zorko B.

Optimization of floating matrix tablets and evaluation of their gastric residence time. Int J Pharm2000; 195: 125135.

2. Iannuccelli V, Coppi G, Bernabei MT,Cameroni R. Air compartment multipleunit systemfor prolonged gastric residence. Part I. Formulationstudy. Int J Pharm 1998; 174:4754.

3. Menon A, Ritschel WA, Sakr A. Development andevaluation of a monolithic floating dosage form forfurosemide. J Pharm Sci1994; 83: 239245.

4. Whitehead L, Fell JT, Collett JH, Sharma HL, SmithAM. Floating dosage forms: an in vivo studydemonstrating prolonged gastric retention. J ContRel 1998; 55: 312.

5. Santus G, Lazzarini G, Bottoni G. An invitroin vivo investigation of oral bioadhesivecontrolled release furosemide formulations. Eur JPharm Biopharm 1997; 44: 3952.

6. Deshpande AA, Rhodes CT, Shah NH, Malick AW.Controlled release drug delivery systems forprolonged gastric residence: an overview. Drug DevInd Pharm 1996; 22: 531540.

7. Deshpande AA, Shah NH, Rhodes CT, Malick W.Development of a novel controlledrelease system forgastric retention. Pharm Res 1997; 14: 815819.

8. Singh B, Kim K. Floating drug delivery systems: anapproach to oral controlled drug delivery via gastricretention. J Cont Rel2000; 63: 235259.

9. Chawla G, Bansal A. A means to address regionalvariability in intestinal drug absorption. PharmTechnol 2003; 27: 5068.

10. Vijayakumar A, Senthilnathan B, RavichandiranV. A review

article on different types of floating drug delivery systems. Int J Pharm Pharm Sci 2012; 4(1): 4550.

11. Sahu AK, Verma A, Singh SK. Preparation ofhydrophilic swelling controlledrelease floatingmatrix tablets containingHPMC and chitosan. Int J Pharm Pharm Sci 2012; 4(1): 406411.

12. Pandit V, Suresh S, Joshi H. Peptic ulcer and itsmanagement. J Pharm Res 2008; 1(2): 245252.

13. Malfertheiner P, Megraud F, O'Morain C, Bazzoli F,ElOmar E, Graham D, et al. Current concepts in themanagement of Helicobacter pylori infection: theMaastricht III Consensus Report. Gut. 2007; 56(6):772781.

14. Fischbach L, Evans EL. Metaanalysis: the effect ofantibiotic resistance status on the efficacy of tripleand quadruple first line therapies for Helicobacterpylori. Aliment Pharmacol Ther2007; 26(3): 343357.

15. Stenstrom B, Mendis A, Marshall B. Helicobacterpylorithe latest in diagnosis and treatment. AustFam Physician 2008;37(8): 608612.

16. Graham DY, Shiotani A. New concepts of resistancein the treatment of Helicobacter pylori infections. NatClin Pract Gastroenterol Hepatol 2008; 5(6): 321331.

17. Qasim A, O'Morain CA, O'Connor HJ. Helicobacterpylori eradication: role of individual therapyconstituents and therapy duration. Fundam ClinPharmacol 2009; 23(1): 4352.

18. Ateshkadi A, Lam NP, Johnson CA. Helicobacterpylori and peptic ulcer disease. Clin Pharm 1993; 12:3448.

19. Murray DM. Clinical relevance of infection byHelicobacter pylori. Clin Microbiol Newsletter 1993;15: 3337.

20. Hunt, R. H. Helicobacter pylori eradication: a criticalappraisal and current concerns. Scand. J. Gastroentrol1995; 30: 7376.

21. Shah S, Qaqish R, Patel V, Amiji M. Evaluation ofthe factors influencing stomachspecific delivery ofantibacterial agents for Helicobacter pyloriinfection. J Pharm Pharmacol 1999; 51:667672.

22. Sweetman SC, editor, Martindale The CompleteDrug Reference, Pharmaceutical Press, London, 34thedition, 2005. p. 900902.

23. De Boer WA, Tytgat GN. Regular review:treatment of

Helicobacter pylori infection. BMJ 2000;320(7226): 3134.

24. Badhan AC, Mashru RC, Shah PP, Thakkar AR,Dobaria NB.

Development and evaluation of sustained releasegastroretentive minimatrices for effectivetreatment of H. pylori infection. AAPSPharmSciTech 2009; 10(2): 459467.

25. Rajinikanth PS, Balasubramaniam J, Mishra B.Development and evaluation of a novel floating insitu gelling system of amoxicillin for eradication ofHelicobacter pylori. Int J Pharm2007; 335(12): 114122.

26. Yellanki SK, Sayed JA, Goranti S.Hydrodynamically balanced bilayer tablets ofglycerol monooleate coated amoxicillin trihydrate: Anovel approach to prolong the local action by gastricretention. Int Res J Pharm Sci 2010; 1(1): 3841.

27. Tokumura T, Machida Y. Preparation of amoxicillinintragastric buoyant sustainedrelease tablets andthe dissolution characteristics. J Cont Rel 2006;110(3): 581586.

28. Hilton AK, Deasy PB. Invitro and invivo evaluationof an oral sustainedrelease foating dosage formof amoxicillin trihydrate. Int J Pharm 1992; 86(1): 7988.

29. Whitehead L, Collett JH, Fell JT. Amoxycillin releasefrom a foating dosage form based on alginates.Int J Pharm 2000;210(12): 4549.

30. Asnaashari S, Khoei NS, Zarrintan MH, Adibkia K,Javadzadeh Y.

Preparation and evaluation of novel metronidazolesustained release and floating matrix tablets. PharmDev Technol 2011;16(4): 400407.

31. LaraHernndez B, HernndezLen A, VillafuerteRobles L.

Effect of stearic acid on the properties ofmetronidazole/methocel K4M foating matrices.Brazilian J Pharm Sci 2009; 45: 497505.32. GutirrezSnchez PE, HernndezLen A,

VillafuerteRobles L.Effect of sodium bicarbonate on the properties ofmetronidazole foating matrix tablets. Drug DevInd Pharm2008; 34(2): 171180.33. CedilloRamrez E, VillafuerteRobles L,

HernndezLen A.Effect of added pharmatose DCL11 on the sustainedrelease of metronidazole from methocel K4M andcarbopol 971P NF foating matrices. Drug Dev IndPharm 2006; 32(8): 955965.

34. Javadzadeh Y, Hamedeyazdan S, Adibkia K, KiafarF, Zarrintan

MH, BarzegarJalali M. Evaluation of drug releasekinetics and physicochemical characteristics ofmetronidazole foating beads based on calciumsilicate and gasforming agents. Pharm Dev Technol2009; 15(4): 329338.

35. Yang L, Eshraghi J, Fassihi R. A newintragastric delivery system for the treatment ofHelicobacter pylori associated gastric ulcer: in vitroevaluation. J Cont Rel 1999; 57(3): 215222.

36. Sriamornsak P, Asavapichayont P, Nunthanid J,Luangtana Anan M, Limmatvapirat S, PiriyaprasarthS. Waxincorporated emulsion gel beads of calciumpectinate for intragastric foating drug delivery. AAPSPharmSciTech 2008; 9(2): 571576.

37. Sriamornsak P, Sungthongjeen S,Puttipipatkhachorn S. Use of pectin as a carrier forintragastric floating drug delivery: Carbonate saltcontained beads. Carbohydr Polym 2007; 67(3):436445.

38. Ishak RA, Awad GA, Mortada ND, Nour SA.Preparation, in vitro and in vivo evaluation ofstomachspecific metronidazole loaded alginatebeads as local antiHelicobacter pylori therapy. J ContRel 2007; 119(2): 207214.

39. Murata Y, Sasaki N, Miyamoto E, Kawashima S. Useof floating alginate gel beads for stomachspecificdrug delivery. Eur J Pharm Biopharm 2000; 50(2):221226.

40. Jeong B, Kim SW, Bae YH. Thermosensitive solgelreversible hydrogels. Adv Drug Deliv Rev 2002; 54:3751.

41. Levina M, RajabiSiahboomi AR. The influence of excipients on

drug release from hydroxypropyl methylcellulosematrices. J Pharm Sci 2004; 93: 27462754.

42. Bravo SA, Lamas MC, Salomon CJ. Swellablematrices for the controlled release of diclofenacsodium: formulation and in vitro studies. Pharm DevTechnol. 2004; 9: 7583.

43. Dahl TC, Calderwood T, Bormeth A, Trimble K, Piepmeier E.

Infuence of physicochemical properties ofhydroxypropyl methylcellulose on naproxen releasefrom sustainedrelease matrix tablets. J Cont Rel1990; 14(1): 110.

44. Li CL, Martini LG, Ford JL, Roberts M. The use ofhypromellose in oral drug delivery. J Pharm Pharmacol2005; 57(5): 533546.

45. Sung KC, Nixon PR, Skoug JW, Ju TR, Gao P, Topp EM,, et al. Effect of formulation variables on drug andpolymer release from HPMCbased matrix tablets. Int JPharm 1996; 142(1): 5360.

46. Alderman DA. A review of cellulose ethers inhydrophilic matrices for oral controlledreleasedosage forms. Int J Pharm Technol Prod Manuf 1984;5: 19.

47. Ghimire M, Hodges L, Band J, O'Mahony B, McInnesFJ, Mullen AB, et al. In vitro and in vivo erosionprofiles of hydroxypropylmethylcellulose (HPMC)matrix tablets. J Cont Rel 2010; 147(1): 7075.

48. Ali J, Arora S, Ahuja A, Babbar AK, Sharma RK, Khar RK.

Formulation and development of floating capsules ofcelecoxib:in vitro and in vivo evaluation. AAPS PharmSciTech2007; 8(4):312319.

49. BaniJaber AK, Alkawareek MY, AlGousous JJ, Abu Helwa AY.

Floating and sustainedrelease characteristics ofeffervescent tablets prepared with a mixed matrix ofEudragit L10055 and Eudragit E PO. Chem PharmBull (Tokyo) 2011; 59(2):155160.

50. Nama M, Gonugunta CS, Reddy Veerareddy P.Formulation and evaluation of gastroretentive dosageforms of clarithromycin. AAPS PharmSciTech 2008;9(1): 231237.

51. United States Pharmacopeia and NationalFormulary USP 30 NF 25; The United StatesPharmacopeial Convention, Inc.: Rockville, MD, 2007.p. 2655.

52. JimnezCastellanos MR, Zia H, Rhodes CT, Design and testing

in vitro of a bioadhesive and floating drug deliverysystem for oral application. Int J Pharm 1994; 105(1):6570.

53. Tadros MI. Controlledrelease effervescentfoating matrix tablets of ciprofoxacinhydrochloride: Development,optimization and in vitroin vivo evaluation in healthyhuman volunteers. Eur J Pharm Biopharm 2010; 74:332339.

54. Ostle B. Statistics In Research: The Iowa StateUniversity Press, Ames, Iowa, USA. 1960. p. 64.

55. Parab PV, Oh CK, Ritschel WA. Sustainedrelease from precirol (glycerol palmitostearate)matrix. Effect of mannitol and hydroxypropylmethylcellulose on the release of theophylline. DrugDev Ind Pharm 1986; 12(8): 13091327.

56. Philip AK, Pathak K. Osmotic fow throughasymmetric membrane: a means for controlleddelivery of drugs with varying solubility. AAPSPharmSciTech 2006; 7(3): E1E11.

57. Sood A, Panchagnula R. Drug release evaluation of diltiazem CR

preparations. Int J Pharm 1998; 175(1): 95107.58. Moore JW, Flanner HH. Mathematical comparison

of dissolution profiles. Pharm Tech 1996; 20: 6474.59. United States Food and Drug Administration. Guidance for

industry, dissolution testing of immediate release solid oral dosage forms; 1997.

60. Shah VP, Tsong Y, Sathe P, Liu JP. In vitrodissolution profile comparison statistics andanalysis of the similarity factor, f2.Pharm Res 1998; 15(6): 889896.

61. British pharmacopoeia. London, The Pharmaceutical Press;

1998. p. 242.62. United States Pharmacopeia and National

Formulary USP 27 NF 22; The United StatesPharmacopeial Convention, Inc.: Rockville, MD, 2004.p. 454458.

63. Li S, Lin S, Chien YW, Daggy BP, Mirchandani HL.Statistical optimization of gastric floating system fororal controlled delivery of calcium. AAPSPharmSciTech 2001; 2(1): 1122.

64. Singh BN, Kim KH. Floating drug delivery systems:an approach to oral controlled drug delivery viagastric retention. J Cont Rel2000; 63(3): 235259.

65. Emara LH, Abdou AR, ElAshmawy AA, MursiNM. In vitro release evaluation of gastroretentiveamoxicillin preparations for treatment of HelicobacterPylori. The 38th Annual Meeting& Exposition of the Controlled Release Society,NationalHarbor, Maryland, USA; 2011: July 30 August 3.

66. Krner A, Larsson A, Piculell L, Wittgren B. Tuningthe polymer release from hydrophilic matrix tabletsby mixing short and long matrix polymers. J PharmSci. 2005; 94(4): 759769.

67. Nagarwal RC, Ridhurkar DN, Pandit JK. In vitrorelease kinetics and bioavailability ofgastroretentive cinnarizine hydrochloride tablet.AAPS PharmSciTech 2010; 11(1): 294303.

68. Someshwar K, Chithaluru K, Ramarao T, Kumar KK.

Formulation and evaluation of effervescent floating tablets of tizanidine hydrochloride. Acta Pharm 2011;61: 217226.

69. Siepmann J, Peppas NA. Modeling of drug releasefrom delivery systems based on hydroxypropylmethylcellulose (HPMC), AdvDrug Del Rev 2001; 48: 139157.

70. Kiss D, Suvegh K, Zelko R. The effect of storage andactive ingredient properties on the drug releaseprofile of poly(ethylene oxide) matrix tablets.Carbohydr Polym 2008;74(4): 930933.