Floating microspheres of Miglitol as gastro retentive drug delivery system: 32 full factorial design and in vitro evaluation

Background: The goal of this study was to develop a gastro retentive oating drug delivery system that would improve site specic activity, patient compliance and therapeutic ecacy. Methodology: Floating microspheres of Miglitol were formulated by double emulsion method using ethyl cellulose and eudragit E100 different weight ratio and PVA as an emulsier. It has been prepared with respect quantity of polymer concentration and stirring speed to evaluate for % buoyancy, drug entrapment eciency, particle size drug release rate. Result: The percent of buoyancy, drug entrapment eciency, particle size, and percentage yield were increased with increase the polymer mixture concentration. Among all formulation batches, F6 showed acceptable results drug entrapment eciency (86.57%) and buoyancy (94.25%). F10 formulation was prepared to check the predicted and actual factors and compared with optimized formulation F6. The drug release was increased as the polymer concentration was decrease. The kinetic model zero order had the highest regression coecient value, it was described as a sustained release dosage form. According to ICH guideline accelerated stability studies of F6 and F10 formulations were conducted for 90 days. After 90 days buoyancy and in vitro drug release was performed and the results were F6 and F10 buoyancy was found to be 88.21%, 87.22% and in vitro drug release was found to be 62.87%, 63.51%. Conclusion: The present study, showed compatibility of drug with polymers by FTIR in formulation. double The Miglitol present present showed compatibility of drug with polymers by FTIR in formulation. Floating microsphere of Miglitol was prepared by double emulsion technique. The F6 Miglitol oating microsphere was optimized formulation demonstrated with excellent drug entrapment performance (86.57%), good oating behaviour (94.25%), and the largest particle size (670µm). The F6 formulation showed the drug release prole in sustained manner more than 12hrs. Also F6 formulation has compared with the formulation F10 which was prepared to validate the predicted and actual values and it was found that F6 optimized formulation showed the better results. Formulated Miglitol microsphere were stable at 40ºC ± 2ºC and 75% ± 5% as per stability studies. From the present study, it can be concluded that the prepared oating microspheres remain for a longer period of time in simulated gastric uid and results in sustained release of drug, resulting in enhanced therapeutic effectiveness and safety with site specic drug delivery. Often, to improve patient compliance, reduce the dose frequency.

polysaccharides (such as starch, sucrose, and other sugar complexes) into monosaccharides including glucose. Since this effect slows the release of glucose, a high postprandial rise in blood glucose and serum insulin will be reduced. Miglitol has a short biological half-life (2hr) and is absorbed by the intestine. To have the greatest impact, it must be consumed with the main meal. It has a molecular weight of 207.2 and a Pka of 5.9 [6,8]. Miglitol is classi ed as a BCS class I compound, which means it is highly soluble and permeable [7]. Miglitol conventional dosage forms available in the market are tablet and lm coated tablets. As these dosage forms needs to be administered in multiple doses daily due to shorter half-life of drug, which ultimately reduces patient compliance.
To overcome these disadvantages some of research work has carried out. For instance; Miglitol Sustained release tablet [6], Miglitol Matrix tablet [5], Miglitol oral bioadhesive Controlled release tablet [22]. Compared to these developed formulations, the oating microspheres will improve the therapeutic e cacy with drug release in sustained manner. There is a need to establish a formulation that will keep the dosage form in the stomach for a longer period of time and release the drug slowly, resulting in enhanced therapeutic e cacy.
Microspheres are free owing solid spherical particles with diameters ranging from 1-1000 micrometers.
They consisting of proteins or synthetic polymers, which are biodegradable in nature. [11,25] A welldesigned dosage type controlled/sustained drug delivery system should be able to overcome the limitations of traditional therapy while also improving therapeutic e cacy. Each particle is essentially a drug mixture distributed in a polymer using a zero-order release procedure. The dissolution/disintegration of the matrix regulates drug release. Because of the size and shape of the microspheres, they have a ballbearing effect [12,24,27].
Several advantages have been developed to the gastric residence time (GRT) of dosage types, such as the oating drug delivery system (FDDS) or the hydrodynamically balanced system (HBS). Since it has a lower bulk density than gastric uids, it can oat in the gastric juice for an extended period of time without impacting the gastric-emptying rate. In the oating device, the drug is slowly released at a predetermined time, and the residual is excreted after the release is complete [10,13].
Multiple unit drug delivery systems, such as oating microspheres, are engineered as a sustained drug delivery to increase therapeutic e cacy and oral bioavailability and it has been gaining attention for the uniform distribution of these multiple-unit dosage forms in the stomach, which means better drug absorption and less local stomach discomfort [14,23]. As per review of literature oating drug delivery system has not been developed yet for this drug.
Thus, in the present study attempts were made to prepare, optimize and evaluate oating microspheres for effective therapeutic e cacy, sustained release of drug with reduced dose frequency and enhanced patient compliance.

Methods
Miglitol was obtained as a gift sample from Hetero Labs Ltd., Baddi, Himachal Pradesh. Ethyl cellulose was purchased from west coast lab, Mumbai. Eudragit E100 was purchased from Evonik pharma industries, Mumbai and Polyvinyl alcohol (PVA), Dichloromethane (DCM), and n-Hexane was purchased from Moly Chem, Mumbai. All other solvents and reagents were used as analytical grade in this analysis.
Factorial Design: The Design of Experiment (DOE) represented a maximum amount of information in a minimum number of runs. DOE is the easiest and most effective technique for controlling a critical parameter. Various preliminary trail formulations were carried out by varying concentrations of ethyl cellulose and eudragit E100 and also magnetic stirring speeds. On the basis of preliminary studies, the concentration of polymers and magnetic stirrer speed were selected to formulate the microspheres. The selected polymer concentration and stirrer speed were used to engender a 3 2 full factorial ( 2 factor and 3 levels) screening design, and 9 experiment runs were establish using JMP ® software (version 15). There are two independent variables and two dependent variables in this analysis. The concentration of ethyl cellulose (X 1 ) and concentration of eudragit E100 (X 2 ) were selected as independent variables (-1, 0, +1). The dependent variables were % buoyancy (Y 1 ) and drug entrapment e ciency (DEE) (Y 2 ). Response surface methodology (RSM) was used for the statistical analysis of effects of independent variables on dependent variables using Design-Expert® software (version-13) as shown in Table 1. Using this software screening design, different models were constituted, and the signi cance of the model was con rmed by statistical parameters. RSM two and three dimensional response counter plot was constituted to read the main effect and interaction between factors and runs [9]. The formulation 10 was prepared as to validate optimized formulation by using actual and predicted value coe cient obtained DOE.

Preparation of Microsphere:
Microspheres were prepared by double emulsion method using water in oil in water emulsion (W/O/W) with different polymer (Ethyl Cellulose & Eudragit E100) ratios [15]. Drug was dissolved in aqueous solvent (Distilled water) and polymer mixture (-1, 0, +1) was dissolved in organic solvent (DCM). Then the aqueous solution was mixed to the organic phase containing the polymer to obtain the primary emulsion (W/O). This primary emulsion was slowly injected (Syringe with 22G needle) into the 100ml of 2% PVA solution under magnetic stirrer (700 rpm, at 30ºC) to prepare the double emulsion (W/O/W). After preparing the double emulsion, 15ml of n-Hexane was added to the microsphere to harden it. The emulsion was held under the magnetic stirrer at the same speed for 5 hours in order to evaporate the organic solvent. The microspheres were then ltered using Whatman lter paper no 44, washed with n-Hexane, and allowed to dry overnight at room temperature [15,35]. The formulation master formula as given in Table 2.
Characterization oating of Microsphere: Drug Entrapment E ciency: The Miglitol Floating Microsphere of all the formulations were analysed by UV-Visible Spectrophotometer. The 100mg of all oating microspheres were diluted in 20 ml of DCM and with continuous stirring phosphate buffer pH 1.2 was added in divided quantise of 20ml three times to completely dissolve the microspheres. Then the obtain solution was transferred into the separating funnel. In separating funnel the aqueous phase was removed. Whatman lter paper was used to lter the separated aqueous solution.
Then the sample was analysed by UV-Visible Spectrophotometer at 220 nm [16]. Then the drug entrapment e ciency was calculated by the given formula [21].

Percentage of Buoyancy:
The USP type-2 paddle dissolution apparatus was used to achieve the oating behaviour. The 100 mg of all oating microsphere formulations were transferred into the dissolution basket. The 900 ml of phosphate buffer pH 1.2 was used as a oating medium with at 100 rpm up to 12hrs. After 12hrs the oating microspheres on the surface of oating media was ltered and kept for drying at room temperature overnight [16, 17,30]. After drying the microsphere buoyancy was calculated by the given formula.
Determination of Particle size: Particle size determination plays an important role in the drug release studies and particle oating property. The oating microsphere shape and particle size were evaluated by optical microscope. With a calibrated ocular micrometre, particle size was measured in the range of 200-700 [20].
Scanning Electron Microscopy: The dried oating microspheres morphological character of outer and internal surface was analysed by scanning electron microscope (SAIF Karnatak University Dharwad). The particle image was captured at 60X and 100 X magni cations by using electron microscope.
Percentage yield: The prepared all oating microspheres were collected and weighed. The weighed microsphere was divided by the total weight of initial non-volatile components which we were used for preparation of microspheres (Drug + Polymer) [18]. The provided formula was used to measure the percentage yield.
Lag time: The oating microspheres were weighed from various formulations and transferred to a beaker with a pH of 1.2 as the medium. The time taken by oating microsphere to rise to the surface was noted.
In vitro drug release study: The dried oating microspheres were evaluated by in vitro drug release pro le. The dissolution was conducted by using the 0.1 N HCl (pH 1.2) and the study was performed up to 12hrs. The dissolution apparatus USP type-2 paddle method was used to assess the drug release study for all formulations. The all different formulations of oating microsphere were weighed and transferred into the dissolution basket. The baskets contain 900ml of pH 1.2 water, which is same as the stomach pH. The rotational speed of the paddle was set to 100 rpm. Every periodic time interval 5ml solution was pipetted out from the dissolution basket and at the same time fresh 5ml dissolution medium was added. Then the sample was diluted with pH 1.2 and analyzed by UV-Visible Spectrophotometer at 220 nm [9].
Release kinetic study: The kinetic study was performed to determine the mechanism of drug release from oating microspheres. The drug release data were plotted into the different kinetic models, such as zero order, rst order, higuchi and korsemeyer-peppas kinetics [29,31].
Stability Studies: According to the ICH guidelines stability study was carried out for optimized formulations. Formulations of oating microspheres were held in stability chamber for 90 days with temperature of 40ºC ± 2ºC, and relative humidity of 75% ± 5% (19)   Characterization oating of Microsphere: Drug Entrapment E ciency: The drug entrapment e ciency of the all formulations of oating microsphere was found to be in the range of 86.57% -70.82 %. Drug entrapment e ciency was increasing with increase in Ethyl cellulose: Eudragit E100 polymer ratio. Because as the polymer concentration is increased, the polymer coats more drug particles, increasing the drug entrapment e ciency. The DEE percentage is showed in Table 3.
Percentage Buoyancy: The percentage of buoyancy of the oating microspheres in all formulations was found to be between 94.25% and 73.23%. The % buoyancy was increased due to increase in the polymer mixture concentration. The oating microsphere was oated in the simulated gastric juice for more than 12hrs. This means that the microsphere will be retained on the gastric juice for a longer period of time in order to enhance the gastric residence time of the dosage form. Table 3 displays the percentage of buoyancy.
Determination of Particle size: The particle size determination has been carried out for all formulations of oating microspheres, and was found to be in the range from 670.42 µm -410.45µm. It was observed that as the Ethyl cellulose: Eudragit E100 polymer ratio increased the particle size also increased. Because, the consistency of solution will increase with polymer ratios, resulting in increased particle size. Floating microsphere with mean particle size range from 500µm-1000µm, have been reported to possess higher oating ability [28]. Table 3 showed the measured particle size effects.
Scanning Electron Microscopy: The optimized oating microsphere (F6) surface morphology was explored by SEM. It has been examined at different magni cation of 60X and 100X. The images of microspheres were almost smooth and spherical shape and the small porous cavities were found on the surface of microspheres, which will aid in improving the oating property of microspheres. The F9 formulation of oating microspheres prepared higher ethyl cellulose and eudragit E100 ratios has shown rough surface in SEM images. This is attributed to increased polymer concentration increases particle size with rough surface. It's showed in Figure 2 Percentage yield: The percentage yield of all oating microsphere formulations was calculated. The calculated percentage yield ranged between 98.76% -91.656%. It indicates that the increased polymer concentration in microspheres leads to increased percentage yield. The calculated percentage yield is showed in Table 3.
Lag time: All nine formulations were evaluated for oating lag time, and the oating microsphere lag time was zero seconds for all formulations. Because, all the oating microsphere formulations possess oating property, when they are added to simulated gastric uid, microsphere will not go into the solvent and it will just oat on the simulated gastric uid. As a result, the oating lag time would be zero second.
In vitro drug release study: In vitro drug release study of oating microsphere was conducted for 12hrs using 0.1N HCl pH1.2 as a release medium by using USP type-2 dissolution apparatus. After 12 hours, the cumulative drug release of all 9 formulations showed that Formulation F9 had the highest cumulative drug release and Formulation F6 had the lowest cumulative drug release. The increased polymer concentration results in more amount of polymer coating around the drug that ultimately decreases the cumulative percent drug release. The cumulative percentage drug release was performed triplicates and the data was plotted in graph it has shown in Figure 3.
Release kinetic study: The data was obtained from in vitro drug release study and it has plotted in different kinetic models. The all formulations were showing the zero order kinetic model with maximum R 2 value. Zero order kinetic model is the best t model for sustained release dosage forms. As per the n-value of peppas model which was between 0.5 to 1, so mechanism was non-ckian transport [33]. The kinetic model tting release pro le of all formulation results are given in Table 4.
Stability Studies: According to ICH guidelines, all formulations were tested for stability for 30, 60 and 90 days. There was slight variation in shown in buoyancy and in-vitro drug of Miglitol oating microspheres on 90 th day at (40ºC ± 2ºC and 75% ± 5%). F6 formulations buoyancy was found to be 88.21% and in-vitro drug release was found to be 62.87%. F10 formulations buoyancy was found to be 87.22% and in vitro drug release was found to be 63.51%. There were no colour variations in the physical appearance of any of the Miglitol oating microsphere formulations, and there was no overlap on other particles.
Optimization of oating microsphere: Based on pilot study report, selected polymer concentration and stirring speed Miglitol oating microsphere was prepared by using JMP ® software 3 2 factorial designs (3 level 2 factor). Independent variables was polymer concentration A (Ethyl cellulose), B (Eudragit E100) 3 levels (-1, 0, +1) 500, 600, 700 and dependent variables was drug entrapment e ciency and percentage buoyancy. Miglitol oating microsphere was prepared by double emulsion method because of Miglitol is freely soluble in water to con rm with available literature and text books.
The optimization studies were performed in order to select the best polymer concentration of ethyl cellulose and eudragit E100 to obtain the maximum drug entrapment e ciency and buoyancy. As the polymer concentration (ethyl cellulose and eudragit e100) increase buoyancy and drug entrapment also increase simultaneously.
ANOVA was analysed with data, it was obtained that it follows quadratic model R 2 value for % of buoyancy was 0.9866 and drug entrapment was 0.9920.
The Equation for the buoyancy and drug entrapment e ciency 1 and 2 respectively, Design space is a multivariate mixture and interaction of independent factors and process factors that has been proved to enable quality assurance. To build design space and optimise all of the replies, a numerical optimization approach (desirability function) and a graphical optimization approach (overlay plot) were utilised.
Constraints on the dependent response and independent factors were used to obtain at the optimum formulation. The response, buoyancy, and drug entrapment e ciency restrictions were xed at range between 90-95 % and 95-98 %, respectively. As a result, because it falls in the yellow area of the overlay plot, with desirability equal to 1, Formulation F10 was regarded an optimum formulation.   Counter plot for DEE Figure 5 Counter plot for % buoyancy Figure 6 Counter plot for predication factor and overlay plot