Scholarly article on topic 'In vitro and in vivo evaluation of gastroretentive floating drug delivery system of ofloxacin'

In vitro and in vivo evaluation of gastroretentive floating drug delivery system of ofloxacin Academic research paper on "Chemical sciences"

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{Ofloxacin / "Gastroretentive drug delivery system" / "Box–Behnken design" / Bioavailability / Floating}

Abstract of research paper on Chemical sciences, author of scientific article — Rajani Shakya, Panna Thapa, Ranendra N. Saha

Abstract This study aimed to develop hydrophilic matrix based controlled release gastroretentive drug delivery system of ofloxacin and conducting its in vitro and in vivo evaluations. Effervescent floating gastroretentive drug delivery system of ofloxacin was prepared utilizing Box–Behnken statistical design with 3 factors, 3 levels and 15 experimental trials. Formulation optimization was done by setting targets on selected responses. In vivo studies were carried out for the optimized formulation with 12 healthy human volunteers and obtained pharmacokinetic parameters were compared with the marketed once daily formulation, “Zanocin OD”. Optimized formulation showed satisfactory controlled in vitro drug release for more than 12 h with excellent buoyancy properties (floating lag time <1 min, floating duration >16 h). Optimized and marketed formulations were found to have similar in vitro release profile (f2 = 79.22) and also were found to be bioequivalent. Serum ofloxacin concentration was well maintained above its reported minimum inhibitory concentrations for most of the pathogens for sufficiently longer duration. C max and AUC values of optimized formulation were found to be significantly higher than of marketed product despite their bioequivalence. Bettertherapeutic effect can be expected since ofloxacin exhibits concentration dependent killing. Hence, gastroretention can be a promising approach to enhance bioavailability of ofloxacin with narrow absorption window in upper GIT.

Academic research paper on topic "In vitro and in vivo evaluation of gastroretentive floating drug delivery system of ofloxacin"

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Original Research Paper

In vitro and in vivo evaluation of gastroretentive floating drug delivery system of ofloxacin

Rajani Shakya a'*, Panna Thapa a, Ranendra N. Saha b

a Department of Pharmacy, Kathmandu University, P.O. Box: 6250, Dhulikhel, Kavre, Nepal b Birla Institute of Technology and Science, Pilani, Rajasthan, India



Article history: Received 2 April 2013 Received in revised form 7 May 2013

Accepted 13 June 2013

Keywords: Ofloxacin

Gastroretentive drug delivery system

Box—Behnken design



This study aimed to develop hydrophilic matrixbased controlled release gastroretentive drug delivery system of ofloxacin and conducting its in vitro and in vivo evaluations. Effervescent floating gastroretentive drug delivery system of ofloxacin was prepared utilizing Box— Behnken statistical design with 3 factors, 3 levels and 15 experimental trials. Formulation optimization was done by setting targets on selected responses. In vivo studies were carried out for the optimized formulation with 12 healthy human volunteers and obtained pharmacokinetic parameters were compared with the marketed once daily formulation, "Zanocin OD". Optimized formulation showed satisfactory controlled in vitro drug release for more than 12 h with excellent buoyancy properties (floating lag time <1 min, floating duration >16 h). Optimized and marketed formulations were found to have similar in vitro release profile (f2 = 79.22) and also were found to be bioequivalent. Serum ofloxacin concentration was well maintained above its reported minimum inhibitory concentrations for most of the pathogens for sufficiently longer duration. Cmax and AUC values of optimized formulation were found to be significantly higher than of marketed product despite their bioequivalence. Bettertherapeutic effect can be expected since ofloxacin exhibits concentration dependent killing. Hence, gastroretention can be a promising approach to enhance bioavailability of ofloxacin with narrow absorption window in upper GIT.

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The importance of controlled drug delivery systems that release drug over an extended period of time has long been recognized in the pharmaceutical field. Application of such controlled release technology to oral drug delivery system

however has been limited because the actual time for effective drug delivery is restricted by gastrointestinal transit time. Gastric retention devices are designed to prolong the gastric residence time of oral controlled release dosage forms. They thus result in increased contact time for drugs that act locally, increased absorption of drugs that have absorption windows in

* Corresponding author. Tel.: +977 11 661399, +977 9841299482 (mobile). E-mail address: (R. Shakya).

Peer review under responsibility of Shenyang Pharmaceutical University

1818-0876/$ — see front matter © 2013 Shenyang Pharmaceutical University. Production and hosting by Elsevier B.V. All rights reserved. http://dx.doi.Org/10.1016/j.ajps.2013.07.025

upper part of gastrointestinal tract (GIT), and better absorption for drugs less soluble in the intestinal fluid [1]. Several approaches have been developed to achieve extended gastric residence time of the oral drug delivery systems such as bio-adhesive system, swelling and expanding systems, floating systems and delayed gastric emptying devices. Amongst these methods, floating drug delivery system is preferred one that offers a simple and practical approach to achieve Gastro-retention [2]. Floating dosage forms have a bulk density lower than that of gastric fluids and therefore remain buoyant on the stomach contents to prolong the gastric retention time [3-7].

Ofloxacin is widely used fluoroquinolone and has been reported as one among the five most frequently prescribed antibiotics in Nepal [8]. It is a broad spectrum antibiotic effective against wide range of Gram-negative and Grampositive microorganisms. Biological half life of this drug is from 5 to 6 h due to which frequent administration is required. To avoid the drawbacks of frequent administrations such as plasma level fluctuations and patient non-compliance it is desirable to have a controlled release dosage forms of oflox-acin. It has been reported that bioavailability of ofloxacin is strongly dependent on the local physiology of GIT. It is readily soluble in the acidic environment of the stomach and thus is preferentially absorbed from the upper part of GIT [9]. In the alkaline environment of intestine, precipitation of the drug occurs decreasing its absorption.

This study was conducted with an aim to develop floating gastroretentive tablet formulation incorporating 400 mg ofloxacin into hydrophilic polymeric matrix which would release the drug in stomach and upper part of GIT in a controlled manner. Since ofloxacin has site-specific absorption from these regions, gastroretention of the dosage form will improve its oral bioavailability [10-12].

2. Materials and methods

2.1. Materials

Ofloxacin (Batch no. A5/206), hydroxypropyl methyl cellulose (HPMC) K100M (Batch no. HP121406 MC) and crosspovidone (Batch no. YPVPP09319040) were obtained from Nepal Pharmaceutical Laboratories Pvt. Ltd., Birgunj, Nepal as gift samples. Sodium bicarbonate (NaHCO3), citric acid, polyvinyl pyrrolidone (PVP K-30), magnesium stearate, lactose and iso-propyl alcohol were purchased from local suppliers. Marketed product, "Zanocin OD", (manufactured by Ranbaxy, India; Batch no. 2033597), used as a reference, was purchased from the local retail pharmacy.

2.2. Box-Behnken statistical design

One of the widely used response surface designs, a Box-Behnken statistical design with 3 factors, 3 levels, and 15 runs with triplicate center points was employed for the formulation of floating gastroretentive tablets of ofloxacin. Formulation design, optimization and other investigations were done using Statgraphics Centurion XV software from Stat-Point Technologies, USA, version 15.2.06. The independent variables or the factors were the amount of HPMC K100M

(X1), crosspovidone (X2) and NaHCO3 (X3). Levels of these factors were set in the formulation design on the basis of the results of preliminary study and are coded as -1, 0, and +1 (Table 1). The responses selected were the cumulative percentage drug release at 2 h (Y1), 8 h (Y2) and 12 h (Y3), floating lag time (Y4) and the total floating time (Y5). Formulation optimization was done by setting targets for these response variables.

2.3. Preparation of floating gastroretentive tablets

Tablets were prepared by conventional wet granulation method using HPMC K100M as a release retardant, cross-povidone as a swelling agents and NaHCO3 as gas generating agent. Citric acid was also incorporated in the formulation to provide sufficiently acidic medium for NaHCO3 to react and maintain buoyancy. The compositions of designed 15 formulations are listed in Table 2. All ingredients (except gas generating agents and magnesium stearate) were passed through sieve no. 60 and mixed in a polybag for 10 min and granulated using PVP K30 (in isopropyl alcohol). The wet mass was passed through sieve number 14 and dried in hot air oven at 50 °C for 1.5 h. Dried granules were mixed with remaining ingredients and compressed using 16-station rotary tablet press (Rimek Minipress-I, India) using 13 mm flat punch in order to obtain controlled release floating gastroretentive tablets containing 400 mg of ofloxacin.

2.4. In vitro analysis

2.4.1. In vitro drug release study

Drug release from the tablets was studied using USP dissolution apparatus (Electrolab TDT-081, India), type I (basket method). A tablet was placed inside a basket and immersed in a dissolution vessel (n = 6) containing 900 ml of 0.1 N HCl (pH 1.2), used as dissolution media at 37 ± 0.5 °C and stirred at a speed of 100 rpm. The amount of drug released after 0.5,1, 2,3, 4, 6, 8, 10, 12, 14 and 16 h was determined using UV-visible spectrophotometer (Shimadzu Corporation, 1601, Japan) at 294 nm. Release profiles of the designed formulations were compared with that of the marketed formulation, "Zanocin OD". Similarity and difference factors were calculated using appropriate formulas.

2.4.2. In vitro buoyancy study

The floating property of the tablets was visually determined in triplicate. The floating lag time and the total floating time were determined in the USP dissolution apparatus containing 0.1 N HCl (pH 1.2, maintained at 37 ± 0.5 °C). The time required

Table 1 - Independent variables and their levels in Box-Behnken design.

Level (mg per tablet)

Low Middle High

Xj (amount of HPMC K100M) X2 (amount of crosspovidone) X3 (amount of NaHCO3) -1 (40) -1 (60) -1 (60) 0 (70) +1 (100) 0 (105) +1 (150) 0 (75) +1 (90)

Table 2 - The composition of 15 formulations as per Box-Behnken design.

Ingredients (mg) F1 F2 F3 F4 F5 F6 F7 F8 F9 F10 F11 F12 F13 F14 F15

Ofloxacin 400 400 400 400 400 400 400 400 400 400 400 400 400 400 400

HPMC K100M 40 70 100 70 100 100 70 100 70 40 40 70 70 40 70

Crosspovidone 60 105 105 150 105 150 105 60 150 105 105 60 105 150 60

NaHCO3 75 75 60 60 90 75 75 75 90 60 90 90 75 75 60

Citric acid 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30

PVP K-30 40 40 40 40 40 40 40 40 40 40 40 40 40 40 40

Mag. Stearate 15 15 15 15 15 15 15 15 15 15 15 15 15 15 15

Lactose 150 75 60 45 30 0 75 90 15 120 90 105 75 60 135

Total 810 810 810 810 810 810 810 810 810 810 810 810 810 810 810

for the tablet to rise to the surface of the medium and float was considered as floating lag time. The duration of time the dosage form remained constantly on the surface of medium was considered as the total floating time.

2.4.3. Drug release kinetics

In vitro release data were subjected to model fitting analysis to know the order of drug release by treating the data according to zero order, first order and Higuchi's release kinetic equations. Since these kinetic models are generally unable to explain the drug release mechanism from polymeric matrices that undergo swelling and/or erosion during dissolution process, the release data were further fitted into Ritger—Peppas empirical equation [7,13,14]. According to this equation,

Here, Mt/MN is the fractional drug release at time t; K is the release rate constant and n is the diffusional exponent indicative of the release mechanism.

2.4.4. Compatibility of ofloxacin with excipients To investigate the chemical interaction, Fourier Transformed Infra Red (FTIR) analysis of admixture of ofloxacin and the excipients used in the formulation were carried out over the range of 400-4000 cm-1 using FTIR spectrometer (Shimadzu, IR Prestige 21, se Japan). The spectra produced by the pure drug alone and in combination with excipients were compared to confirm the interaction.

In vivo evaluation

In order to ascertain the pharmacokinetic property and expected clinical efficacy, in vivo evaluation of the optimized formulation containing 400 mg ofloxacin was carried out. Open label, randomized, two treatments, two periods, two sequences, single dose, two ways crossover comparative bioavailability study was conducted using healthy human subjects. Prior ethical approval was taken from the Institutional Review Committee (IRC) of Kathmandu University School of Medical Sciences/Dhulikhel Hospital (IRC KUSMS), for carrying out this study (Protocol approval number: 34/11). Pharmacokinetic data of optimized formulation was compared with in vivo data of marketed extended release product of ofloxacin 400 mg (Zanocin OD, Batch no. 1760135).

Twelve healthy, non-smoking, adult Nepalese male volunteers were enrolled in the study. Written informed consent was

obtained from each subject after adequate explanation of the objectives, methods and potential hazards of the study. All volunteers gave medical screening examination before dosing to establish their fitness to participate in the study. Enrolled volunteers were randomly divided into two groups. One group administered optimized formulation and another group administered marketed formulation, "Zanocin OD" on the first study day under fasting condition. After a washout period of 7 days, on the second study day, volunteers exchanged formulations. In both study days, 5 ml of pre-dose blood sample was collected 15 min before drug administration. Later blood sample was collected at 0.5,1, 2,3,4, 5, 6, 7, 8,10,12,16 and 24 h of drug administration. Blood samples were collected in vacutainer tubes without anticoagulant. It was allowed to clot at room temperature for 20 min followed by centrifuging for 15 min at 5000 rpm. Serum was transferred into a separate serum container and was promptly frozen at -40 °C until assay.

2.6. Drug analysis

2.6.1. Sample preparation

To 180 ml of thawed human serum sample, 20 ml of ciproflox-acin (internal standard) was added and vortex mixed for 30 s. 600 ml of methanol was added to the tube. After mixing for another 1 min the tube was centrifuged at 5000 rpm for 5 min using a cooling centrifuge (Remi, India). 20 ml of filtered supernatant was injected into the HPLC column for analysis.

2.6.2. Chromatographic conditions

Serum samples were analyzed for ofloxacin with a validated bioanalytical method using HPLC (Shimadzu, Japan) comprising of a pump (LC-20AD), autosampler (SIL-20AD), photo-diode array detector (SPD-M20A) and a column oven (CTO-10 ASVP). The data collection and integration was accomplished using LC solutions software.

Separation was performed on a reverse phase CAPCELL PAK C18, MG type column (250 mm length, 4.6 mm internal diameter, 5 mm particle size) from Shiseido Fine Chemicals. Mobile phase consisting of acetonitrile and 0.0625% triethyl-amine in water (12.5: 87.5, pH adjusted to 2.5 with ortho-phosphoric acid) was delivered at a flow rate of 1.5 ml/min. The system was operated at wavelength of 294 nm and temperature of column oven was set at 40 °C.

The peaks of ofloxacin and ciprofloxacin were resolved with good symmetry. The retention time for ofloxacin and ciprofloxacin was 8.2 ± 0.8 and 9.3 ± 0.76 min respectively. The

typical assay run time was 12 min. The bioanalytical method was validated as per ICH guidelines.

The serum drug concentration-time data was subjected to non-compartmental analysis using a pharmacokinetic software, WinNonlin®, standard edition, version 5.2.1 (Pharsight Corporation, USA), to obtain various pharmacokinetic parameters. Relative bioavailability was determined as the ratio of AUC0-n of optimized formulation to the AUC0-N of marketed product.

3. Results and discussion

3.1. Compatibility of ofloxacin with excipients

FTIR study revealed absence of potential chemical interaction between the drug and excipients. The infrared spectrum of ofloxacin alone showed characteristic peaks at 3427, 3043, 2968, 2785, 1716, 1620, 1550, 1458, 1056 cm"1 [15]. All of these typical peaks of ofloxacin remained unaffected in the spectrum of ofloxacin in combination with HPMC K100M and also in presence of other excipients used in the formulation.

3.2. In vitro buoyancy

Most of the formulations were found to have good buoyancy properties (Table 3). They floated immediately upon immersion in to the media and remained floated for 16 h. NaHCO3 and citric acid were employed as gas forming agents dispersed in the matrix and this formulation was found to be appropriate for achieving desired buoyancy characteristics. Addition of NaHCO3 was found essential to ensure rapid floating. Furthermore, since the pH of the stomach is elevated under fed condition (~3.5), citric acid was also incorporated in the formulation to provide sufficiently acidic medium for NaHCO3 to react. This will allow the system to float independent of the pH of the medium. Upon immersion, NaHCO3 starts reaction immediately with the acidic

dissolution media and added citric acid. This reaction generates sufficient amount of CO2 which get entrapped and protected within the gel layer formed by hydration of HPMC K100M. This leads to decreased density of the tablet (reported as 1.004-1.010 g/cm3), as a result of which the tablet becomes buoyant [2,3,5,11,16-18]. Buoyancy property is further facilitated by relatively good acid solubility of ofloxacin which causes faster penetration of dissolution media into the matrix. This in turn causes quicker initiation of reaction resulting in faster generation of CO2 making the tablets more buoyant.

It has been reported in several literatures that the amount of NaHCO3 is directly related to the floating lag time [3,11,19-21]. As the amount of NaHCO3 increases, floating lag time decreases due to the generation of larger amount of effervescence. However in this study, quantity of HPMC K100M was found to have more prominent effect on floating characteristics instead. Formulation containing higher amount of HPMC K100M had better buoyancy property. Decreasing its amount decreased floating duration and increased floating lag time [20,22]. This effect is more noticeable in formulations containing lesser amount of NaHCO3. When floating characteristics of the fifteen formulations were compared, a longer floating lag time and a shorter duration of floating were observed in formulations F1, F10, F11 and F14 containing least (40 mg per tablet) amount of HPMC K100M (Table 3). The matrix formed with lower concentration of this polymer seems not to be capable enough to hold the bubbles and float. Also there was quicker loss of tablet shape integrity in these formulations which have further worsened their buoyancy property.

3.3. In vitro drug release

In vitro drug release and buoyancy study revealed that values of responses for these 15 formulations varied markedly indicating strong relationship between responses and the factors (Table 3). The range of response Yi was from 28.84% in F12 to 61.64% in F10. Response Y2 ranged from 53.98% in F12 to a

Table 3 - Observed responses in designed fifteen formulations.

Formulations Responsesa

Yi (%) Y2 (%) Y3 (%) Y4 (s) Ys(h)

F1 50.62 ± 7.20 81.09 ± 7.81 96.00 ± 1.89 1.00 ± 00 1.00 ± 00

F2 34.89 ± 2.07 70.79 ± 5.49 85.61 ± 4.71 25.00 ± 5 16.00 ± 00

F 3 34.29 ± 7.38 70.12 ± 6.33 80.97 ± 2.04 1.00 ± 00 16.00 ± 00

F 4 39.93 ± 1.14 77.75 ± 3.97 92.34 ± 1.53 1.00 ± 00 16.00 ± 00

F 5 31.26 ± 4.03 65.11 ± 5.43 83.37 ± 5.79 1.00 ± 00 16.00 ± 00

F 6 29.91 ± 3.97 55.08 ± 4.46 68.99 ± 5.47 1.00 ± 00 16.00 ± 00

F 7 33.14 ± 3.64 69.95 ± 5.27 84.57 ± 4.87 1.00 ± 00 16.00 ± 00

F 8 39.51 ± 3.76 74.35 ± 6.62 79.57 ± 5.34 54.00 ± 7 16.00 ± 00

F 9 34.71 ± 2.48 69.11 ± 2.63 81.44 ± 3.27 1.00 ± 00 16.00 ± 00

F10 61.64 ± 3.51 96.70 ± 4.14 100.00 ± 00 95.00 ± 5 1.00 ± 0.15

F 11 48.89 ± 4.17 69.28 ± 4.96 82.97 ± 6.28 156.00 ± 10 0.50 ± 0.10

F 12 28.84 ± 4.13 53.98 ± 4.59 69.61 ± 4.95 1.00 ± 00 7.00 ± 0.27

F 13 37.16 ± 6.66 75.10 ± 4.38 90.63 ± 0.47 1.00 ± 00 16.00 ± 00

F 14 41.64 ± 2.77 75.15 ± 6.59 98.36 ± 1.88 22.00 ± 15 4.00 ± 0.23

F 15 31.28 ± 4.62 70.48 ± 1.92 88.31 ± 0.87 78.00 ± 12 4.00 ± 0.25

a Data is expressed as mean ± standard deviation.

maximum of 81.09% in F1. Similarly another response Y3 was in the range of 68.99 in F6 to 100% in F10.

Majority of the formulations extended the drug release for 16 h. Drug release from the hydrophillic matrix tablet is known to be a complex interaction between diffusion, swelling, and erosion mechanisms. These processes are controlled by the hydration of HPMC, which forms the gel barrier through which the drug diffuses [23]. Higher polymer concentration increase the diffusion path length for the drug due to forming greater amount of gel which retard drug release from the formulation. The growth of erosion front, diffusion front, and swelling front decrease with the increase in polymer proportion because of the formation of a stronger gel layer, which make the entry of medium into the matrix difficult [24-27].

In the present formulation design, amount of HPMC K100M varied from 40 to 100 mg per tablet (5-12% of total tablet weight). Formulations F1, F10, F11 and F14, containing low concentration of HPMC K100M, gave high release rate of 50.62%, 61.64%, 48.89% and 41.64% respectively at the end of 2 h. Also these formulations completed drug release within a short time. When quantity of HPMC K100M was increased to 100 mg per tablet in formulations F3, F5, F6 and F8, release at the end of 2 h decreased down to 34.29%, 31.26%, 29.91% and 39.51% respectively. These formulations successfully extended drug release for sufficiently longer duration. At the end of 16 h drug release from formulations F3, F5, F6 and F8 were found to be 93.08, 93.02, 80.01 and 90.54% respectively.

It has been observed that high release occurred in all designed formulations at the end of 2 h. Marketed product also gave a high release rate of 37.08% at the end of 2 h. Such type of release could be due to the reason that the gel layer, which controls drug release needs sometime to become effective. Till the gel barrier is being formed, high rate of erosion occurs resulting in high initial drug release [23,27,28]. After some time release rate slows down as polymer hydration occurs.

In vitro release data were fitted in to different release kinetic models and it was observed that regression coefficient was highest for the Higuchi model (except for F10 & F11). The n values for all formulations, except F1, F10, F11 and F14, were found to be lying within the range of 0.45-0.89, indicating anomalous or non-Fickian type of diffusion to be the predetermining mechanism of drug release.

Mathematical relationships for the measured responses and the independent variables or factors were generated with the help of software Statgraphics Centurion XV and are shown in Equations (1)-(5). These equations represent the quantitative effect of variables (X1, X2, X3) and their interactions on the response. Coefficient with more than one factor term and those with higher order terms represent interaction terms and quadratic relationship respectively. A positive sign represents synergistic effect, while a negative sign indicates antagonistic effect or an inverse relationship between the factors and response [29]. Correlation coefficient (r2) for the equations indicates the percentage variability in model fitting for that particular variable. The adjusted r2 value is more suitable for comparing models with different number of independent variables which can be obtained by including only statistically significant (P < 0.05) coefficients in the equation. In this study final equation was considered with maximum adjusted r2 value. The adjusted r2 value for responses Ya, Y2, Y3 and Y5

were found to be 79.97, 80.51, 84.73 and 88.27% respectively, which indicate good fit. When ANOVA was performed at 95% confidence interval to estimate the significance of the model, factors X1 (HPMC) and X3 (NaHCO3) were identified as critical influencing parameters for selected responses Y^ Y2 and Y3. Another response, Y5, i.e. the total floating time was found to be affected significantly by the amount of HPMC (X1) and CP (X2) whilst no significant effect of formulation variables was observed on Y4, i.e. floating lag time.

Y1 = 111.946 - 1.518X! - 0.195X3 + 0.009 X2 (1)

r2 = 86.41%, r2 (adjusted) = 79.97%

Y2 = 169.080 - 1.456Xa + 0.587X2 - 1.351X3 + 0.004X1

- 0002XjX2 + 0.012XjX3 - 0.002X2 (2)

r2 = 87.43%, r2(adjusted) = 80.51%

Y3 = 173.766 - 1.130Xa + 0.908X2 - 1.775X3 + 0.005X1

- 0008XJX2 + 0.014XjX3 - 0.003X2 + 0.004X2X3 (3)

r2 = 86.39%, r2(adjusted) = 84.73%

Y4 = 932.302 - 5.271Xa - 19.122X3 + 0.031X1 + 0.127X2 (4)

r2 = 37.53%, r2(adjusted) = 20.50%

0.002X2 - 0.014X3

r2 = 92.46%, r2(adjusted) = 88.27%

3.4. Formulation optimization and evaluation

For formulation optimization, target values for the responses (Y1 - Y5) were set on the basis of in vitro drug release and buoyancy study of marketed product, "Zanocin OD" (Table 4). Composition of the optimized formulation given by the software using this technique termed as "multiple response optimization" is shown in Table 5. This combination maximized the desirability over the indicated region to 0.93 (Fig. 1).

Optimized formulation had good physicochemical properties and was found to fulfill the requirement of an optimum

Table 4 — Multiple response optimization.

Response (Range observed in Fl — F15) Target value

% Cumulative Low High

release at

at the end of,

(i)2h 29.91 ± 3.97 61.64 ± 3.51 37.08

(ii) 8 h 53.98 ± 4.59 96.70 ± 4.14 77.99

(iii) 12 h 68.99 ± 5.47 100.00 ± 00 92.03

Floating lag time (s) 1.00 ± 00 156.00 ± 10 Minimize

Total floating 0.50 ± 0.10 16.00 ± 00 Maximize

duration (h)

Table 5 — Composition of optimized formulation.

Composition Amount per % per

tablet (mg) tablet weight

Ofloxacin 400.00 49.38

HPMC K100M 66.02 8.15

Crosspovidone 107.88 13.32

NaHCO3 67.17 8.29

Citric acid 30.00 3.70

PVP K30 40.00 4.94

Magnesium Stearate 15.00 1.85

Lactose 83.93 10.36

Total 810.00 100.00

Crosspovidone = 105.0



™ 0'2~of

™ 0.6-C1.7 " 0'g-0.®

60 70 80


Fig. 1 - Counter plot showing effect of HPMC K100M and NaHCO3 on desirability factor.

controlled release formulation because of better regulation of drug release at the end of 2, 8 and 12 h. The predicted release profile, given by the software was found to be quite close to the profile obtained experimentally which indicates the validity of the developed model (Table 6). Floating lag time of optimized formulation was 2.56 s and it remained floated for more than 16 h in vitro. Thus, besides controlling drug release for an extended duration, the formulation seemed to have an excellent floating potential which is prerequisite for prolong residence of the dosage form in the stomach.

The respective release profiles of the marketed and optimized formulation superimposed over each other which indicates analogy of their release performances (Fig. 2). Comparison of release profile of optimized and marketed formulations gave similarity factor (f2) of 79.22 and difference

Table 6 - Response variables and release kinetics of marketed product and optimized formulation (observed and predicted).

Formulation Response variables Higuchi n

Yi Y2 Y3 Y4 Y5 KH r2 (%) (%) (%) (s) (h)

Marketed Optimized

(predicted) Optimized (observed)

37.08 77.99 92.02 2.00 >16 27.10 0.98 0.55 37.08 77.06 93.82 2.72 17.36 23.45 0.99 0.52

39.77 76.09

2.56 >16 26.01 0.99 0.52

-•— M arketed formulation -A— Optimized formulation

Time (h)

Fig. 2 - Comparative release profile of marketed and optimized formulation.

factor (f1) of 3.84. Table 6 shows the comparison of the response variables and other release parameters of these formulations. Percentage cumulative drug release at 2 (Y1), 8 (Y2) and 12 h (Y3) from optimized formulation were found to be quite close to that of marketed product. Buoyancy parameters like floating lag time (Y4) and total floating duration (Y5) of these formulations were also lying in close proximity to each other (Table 6). Drug release from both optimized and marketed formulation followed Higuchi's release kinetics and the values of n were found to be 0.52 and 0.55 respectively suggesting the release mechanism to be anomalous transport.

Pharmacokinetic evaluation

The mean serum concentration versus time profiles of the optimized and marketed formulations were similar and

c3 Ü a

-♦— Optimized formulation Marketed formulation

Time (h)

Fig. 3 - Overlain mean serum ofloxacin concentration-time profiles after oral single dose administration of optimized and marketed formulation in 12 healthy human volunteers.

Table 7 - Pharmacokinetic profiles of optimized and marketed formulation.

Parameters Marketed Optimized P value

formulation formulation

Cmax (mg/ml) 3.47 ± 0.70 3.94 ± 0.39 0.04

AUC0—24 (mg h/ml) 36.85 ± 4.77 41.80 ± 4.83 0.03

AUC0—„ (mg h/ml) 43.33 ± 8.82 47.94 ± 6.41 0.06

tmax(h) 5.67 ± 1.97 5.75 ± 1.48 —

t *(h) 7.36 ± 2.57 7.20 ± 1.52 —

Kel 0.10 ± 0.32 0.10 ± 0.02 —

nearly superimposable (Fig. 3). Pharmacokinetic parameters of these formulations are presented in Table 7.

90% confidence interval of the ratio of mean AUC0-24, AUC0-n and Cmax of the optimized to marketed formulation were found to be in the range of 94.27%-103.85%; 99.97%-110.11% and 98.59-111.86% respectively. These values are within the bioequivalence accepted range of 80%-125% [30,31]. Thus, the two formulations can be regarded bio-equivalent and hence interchangeable.


Controlled release floating gastroretentive tablet dosage form of ofloxacin was successfully developed using Box-Behnken statistical design. Serum drug level monitoring in human subjects for 24 h showed extended drug release for sufficiently longer duration making its once daily administration sufficient. Serum ofloxacin concentration was maintained well above the reported minimum inhibitory concentrations for most of the pathogens for longer duration. The optimized formulation, when compared to the conventional immediate release preparation, seems to be promising for improving bioavailability of ofloxacin for enhancing its therapeutic efficacy along with improving patient convenience due to less frequent dosing requirement.


The authors would like to express their hearty gratitude to Nepal Pharmaceutical Lab Pvt. Ltd for supplying ofloxacin raw material and excipients. We are also thankful to Mr. Rajan Shrestha, Department of Pharmacy, Kathmandu University for the help provided during tablet preparation.


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