11_chapter 4.pdf - Shodhganga

C h a p t e r 4 Ma t e r i a l s a n d Me t h o d s

66 Formulation & Evaluation of Floating DDS for some se lected drugs

4.1 Materials

4.2 Instruments and Equipments

4.3 Analytical Methods

4.4 Formulations

4.4.1 Formulation of Non-Effervescent floating tablets

4.4.2 Formulation of Effervescent floating tablets

4.4.3 Formulation of Hollow microspheres

4.5 Evaluation



4.1 MATERIALS

Sl. No. Drugs SOURCE

1 Verapamil Hydrochloride Glenmark Pharmaceutical Ltd.,

Mumbai, India

2 Rosiglitazone maleate Matrix Lab, Hyderabad, India

3 Losartan Potassium Suven life sciences, Hyderabad, India

Polymers and Chemicals

4 Polyvinyl pyrrolidone (PVP) SRL, Mumbai, India

5 Chitosan Sigma Aldrich, USA

6 Hydrochloric acid Reachem lab, Chennai, India

7 Accurel®

MP 1000 Membrana, Obernburg, Germany

8 Karaya gum Sigma Aldrich, USA

9 Lactose Loba Chemie, Mumbai, India

10 Sodium bicarbonate Loba Chemie, Mumbai, India

11 Ethyl cellulose 7cps Loba chemie, Mumbai, India

12 Polyethylene oxide Aldrich, Mumbai

13 Hydroxy propyl methyl cellulose Ranbaxy, Baddi, India

14 Eudragit L-100 Evonik, Mumbai, India

15 Potassium chloride Lobachemie, Mumbai, India

14 Tween 80 Lobachemie, Mumbai, India

15 Dichloromethane Lobachemie, Mumbai, India

16 Ethanol Lobachemie, Mumbai, India

17 Barium sulphate Lobachemie,, Mumbai, India



4.2 INSTRUMENTS AND EQUIPMENTS

Sl. No. NAME OF THE

EQUIPMENT

MODEL/ MANUFACTURER

1 Digital balance Shinko Sansui, Japan

2 Hot air oven Tempo, India

3 Magnetic stirrer Remi equipments, India

4 Tablet punching machine Rimek, Minipress- 1(model-1674),

Karnavati, India

5 Micrometer screw gauge Mitutoyo, Japan

6 Dissolution apparatus (8 basket) Electrolab, India

7 UV-Visible spectrophotometer Shimadzu-1800, Japan

8 FT-IR spectrophotometer Shimadzu-8400 S, Japan

9 KBr Press Techno search instruments, India

10 Scanning electron microscopy

(SEM)

Joel SEM analysis Instrument, Model

JSM 840A, Japan

11 Differential scanning

calorimetry (DSC)

Shimadzu DSC-60, Japan

12 Tablet hardness tester Inweka, IHT 100, Ahmedabad , India

13 Digital pH meter Elico-LI120pH(type003),Hyderabad

14 X- Ray Machine Bharat Electronics Ltd. Pune, India

15 Accu- Chek Glucometer Roche Diagnostics India Pvt Ltd,

India



REAGENTS104

0.1 N Hydrochloric acid (HCl)

8.5 ml of concentrated hydrochloric acid solution was diluted with distilled

water upto 1000 ml to give 0.1 N HCl.

0.2 M Potassium chloride

14.911 g of potassium chloride was dissolved in 1000 ml of distilled water.

0.2 M Potassium dihydrogen phosphate

Accurately weighed 27.218 g of potassium dihydrogen orthophosphate was

dissolved in 1000 ml of distilled water.

Hydrochloric acid buffer (pH 1.2)

50.0 ml of 0.2 M potassium chloride was placed in a 200 ml volumetric flask,

to this 85.0 ml of 0.2 M hydrochloric acid was added and then made up to the volume

with water.



4.3 ANALYTICAL METHODS

4.3.1. Verapamil hydrochloride: The method described by Florey K was followed.81

Stock solution: Verapamil hydrochloride in pH 1.2 hydrochloric acid (HCl) buffer

(100 g/ml).

Scanning: From the stock solution, a suitable concentration (10 g/ml) was prepared

with pH 1.2 Hydrochloric acid buffer solution and UV scan was taken

between 200-400 nm. The spectrum is given in figure 4.01. The absorption

maxima of 278 nm was selected and utilized for further studies.

Figure 4.01: UV-Spectra of Verapamil hydrochloride in pH 1.2 hydrochloric

acid buffer



Standard Plot: From the stock solution, 10, 20, 30, 40, 50, and 60 g/ml

solutions of Verapamil hydrochloride were prepared in pH 1.2 hydrochloric acid

buffer solution. The absorbance was measured at 278 nm and a graph of concentration

versus absorbance was plotted. Standard plot data of Verapamil hydrochloride in pH

1.2 hydrochloric acid buffer solution is reported in table 4.01 and graph in figure 4.02.

Table 4.01: Standard plot data for Verapamil hydrochloride in pH 1.2

hydrochloric acid buffer

*Standard deviation, n = 3

Figure 4.02: Standard plot for Verapamil hydrochloride in pH 1.2 hydrochloric

acid buffer

Concentration

(µg/ml)

Absorbance at 278 nm

(mean ± SD*)

10 0.124 ± 0.000

20 0.233 ± 0.070

30 0.361 ± 0.038

40 0.480 ± 0.102

50 0.590 ± 0.007

60 0.715 ± 0.001



4.3.2 Rosiglitazone Maleate: Method described by Shiva SK et al. was followed.105

Stock Solution: Rosiglitazone maleate in pH 1.2 hydrochloric acid buffer solution

(100 g/ml).

Scanning: From the stock solution, a suitable concentration (10 g/ml) was prepared

with pH 1.2 Hydrochloric acid buffer solution and UV scan was taken

between the wavelengths of 200-400 nm. The spectrum is given in figure

4.03. The absorption maxima of 228 nm was selected and utilized for

further studies.

Figure 4.03: UV-Spectra of Rosiglitazone maleate in pH 1.2 hydrochloric acid

buffer



Standard Plot: From the stock solution, 4, 8, 12, 16 and 20 g/ml solutions of

Rosiglitazone maleate were prepared in pH 1.2 hydrochloric acid buffer solution. The

absorbance was measured at 228 nm and a graph of concentration versus absorbance

was plotted. Standard plot data of Rosiglitazone maleate in pH 1.2 hydrochloric

acidbuffer solution is reported in table 4.02 and graph in figure 4.04.

Table 4.02: Standard plot data for Rosiglitazone maleate in pH 1.2 hydrochloric

acid buffer

*Standard deviation, n = 3

Figure 4.04: Standard plot for Rosiglitazone maleate in pH 1.2

hydrochloric acid buffer

Concentration

(µg/ml)


(mean ± SD*)

4 0.169 ± 0.0316

8 0.234 ± 0.0110

12 0.488 ± 0.0870

16 0.641 ± 0.0095

20 0.803 ± 0.0063



4.3.3 Losartan potassium: Method described by Kalyani et al106

was followed.

Standard solution: Accurately weighed 50 mg of Losartan potassium was dissolved

in 100 ml of pH 1.2 hydrochloric acid buffer to get a solution containing 500 μg/ml of

drug.

Scanning: From the standard solution, a solution was prepared to give a

concentration of 8 μg/ml in pH 1.2 hydrochloric acid buffer and UV scan was taken

between the wavelengths of 200-400 nm. The spectrum is reported in the figure 4.05.

The absorption maxima of 248 nm was selected and utilized for further studies.

Figure 4.05: UV-Spectra of Losartan potassium in pH 1.2 hydrochloric acid

buffer



Standard Plot: From the standard solution, a stock solution was prepared to

give a concentration of 50 μg/ml in pH 1.2 Hydrochloric acid buffer. Aliquots of 0.4,

0.6, 0.8, 1.0, 1.2 and 1.4 ml of stock solution was pipetted out into 10 ml volumetric

flasks. The volume was made up to the mark with pH 1.2 hydrochloric acid buffer.

These dilutions gave 2, 3, 4, 5, 6 & 7 μg/ml concentration of losartan potassium

respectively. The absorbances of prepared solutions of losartan potassium in pH 1.2

hydrochloric acid buffer were measured at 248 nm spectrophotometrically against pH

1.2 Hydrochloric acid buffer as blank. Standard plot data of losartan potassium in pH

1.2 Hydrochloric acid buffer is reported in table 4.03 and graph in figure 4.06.

Table 4.03: Standard plot data for Losartan potassium in pH 1.2 hydrochloric

acid buffer

*Standard deviation, n=3

Figure 4.06: Standard plot of Losartan potassium in pH 1.2 hydrochloric acid

buffer

Concentration

(μg/ml)


(Mean ± S.D*)

2 0.214 ± 0.0020

3 0.287 ± 0.0010

4 0.419 ± 0.0015

5 0.526 ± 0.0010

6 0.626 ± 0.0026

7 0.734 ± 0.0200



4.4. FORMULATIONS

4.4.1 Formulation of Non-Effervescent floating tablets

Floating matrix tablets were prepared by direct compression method. All the

ingredients were blended together to get homogenous mixture. Accurel® MP1000 as

low density polypropylene foam powder, karaya gum as release retardant, chitosan as

swellable polymer, lactose as diluent and magnesium stearate as lubricant were used.

Powder mass was compressed into tablets using a 10 station rotary tablet punching

press with 12 mm punch and die set. Each tablet contained 50 mg of losartan

potassium. Composition of each tablet is given in table 4.04.

Table 4.04: Formulation chart of non-effervescent floating Losartan potassium

tablets

Ingredients (mg) G-I G-II G-III G-IV G-V G-VI G-VII G-VIII G-IX

Losartan

Potassium

50 50 50 50 50 50 50 50 50

Accurel®MP1000 150 150 150 150 150 150 150 150 150

Karaya Gum 40 50 60 40 50 60 40 50 60

Chitosam 30 40 50 40 50 30 50 30 40

Lactose 76.5 56.5 36.5 66.5 46.5 56.5 56.5 66.5 46.5

Magnesium

Stearate

3.5 3.5 3.5 3.5 3.5 3.5 3.5 3.5 3.5

Total Weight 350 350 350 350 350 350 350 350 350



4.4.2 Formulation of Effervescent floating tablets

The floating tablets of verapamil hydrochloride were prepared by direct

compression technique. For each tablet formulation, drug, HPMC-K15M, karaya

gum, sodium bicarbonate, and diluents were blended homogeneously for 10 min

followed by addition of magnesium stearate. The total weight of each tablet was

300 mg. The amount of karaya gum used was in the range of 40–90 mg, whereas

HPMC was used in the range of 20-40 mg. The powder mixture was further mixed for

5 min in a mortar. The resultant mixture was compressed into tablets using a Rimek

rotary tablet machine. Thirteen formulations were prepared by changing the amount

of the ingredients as shown in table 4.05.

Table 4.05: Formulation chart of effervescent floating Verapamil hydrochloride

tablets

Ingredients

(mg)

F1 F2 F3 F4 F5 F6 F7 F8 F9 F10 F11 F12 F13

Verapamil

Hydrochloride 120 120 120 120 120 120 120 120 120 120 120 120 120

Karaya Gum 40 40 40 40 70 70 70 70 70 90 90 90 90

HPMC K15 M 20 40 30 30 20 40 20 40 30 20 40 30 30

Sodium

Bicarbonate

20 20 10 30 10 10 30 30 20 20 20 10 40

PVP K30 15 15 15 15 15 15 15 15 15 15 15 15 15

Magnesium

Stearate 5 5 5 5 5 5 5 5 5 5 5 5 5

Lactose 70 50 70 50 60 40 40 20 40 30 10 30 00

Total weight 300 300 300 300 300 300 300 300 300 300 300 300 300



4.4.3 Formulation of Hollow microspheres71,107, 108

Floating microspheres with a central hollow cavity were prepared by using a

modified Quasi-emulsion diffusion technique. Weighed quantities of Rosiglitazone

maleate (RSM), ethyl cellulose, polyethylene oxide and hydroxy propylmethyl

cellulose (HPMC K15M) were dissolved in a mixture of ethanol and dichloromethane

(1:1 solvent ratio) at room temperature in a magnetic stirrer at 50 rpm for 50 min.

This solvent was poured drop wise into 100 ml distilled water containing 2 ml of

Tween 80 maintained at a temperature of 50 ± 2 °C. The resultant solution was stirred

with a pitched-blade-type impeller type agitator at 1100 rpm for 3 h to allow the

volatile solvent to evaporate. This resulted in the formation of microspheres. Different

ratios of polymers were used to prepare the microspheres. Eleven formulations were

prepared by changing the amount of ingredients as shown in table 4.06 and

diagrammatic representation of preparation of hollow microspheres is shown in the

figure 4.07.

Table 4.06: Formulation chart of Rosiglitazone maleate hollow microspheres

INGREDIENTS F1 F2 F3 F4 F5 F6 F7 F8 F9 F10 F11

Rosiglitazone maleate

(gm) 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1

Ethyl cellulose (gm) - - 2 1 1 1 2 1 2 1 -

Polyethylene oxide

(gm) - 1 1 2 - - - - - - -

HPMC K15M (gm) - - - - 1 2 1 - - - -

Eudragit S100 (gm) - - - - - - - 1 1 2 1

Solvent ratio * (ml) 1:1 1:1 1:1 1:1 1:1 1:1 1:1 1:1 1:1 1:1 1:1

Tween 80(ml) 2 2 2 2 2 2 2 2 2 2 2

* Ethanol and dichloromethane of 30 ml each.



Figure 4.07: Diagrammatic representation of preparation of hollow

microspheres by Quassi-emulsion technique



4.5 EVALUATION

All the formulations were evaluated for the following parameters

Non-Effervescent floating tablets and Effervescent floating tablets

Weight variation

Friability

Hardness

Diameter & thickness

Uniformity of drug content

Fourier transform infrared spectroscopy (FT-IR)

Differential scanning calorimetry (DSC)

In vitro buoyancy studies

Water uptake studies

In vitro drug release studies

Mathematical model fitting of obtained drug release data

In vivo studies

Stability studies



Floating Hollow microspheres

Percentage yield

Drug loading and Entrapment efficiency

Fourier transform infrared spectroscopy (FT-IR)

Differential scanning calorimetry (DSC)

Scanning electron microscopy

Sphericity of the microspheres

Micromeritic properties

In vitro buoyancy of microspheres

In vivo floating behaviour

In vitro drug release studies

Mathematical model fitting of obtained drug release data

In vivo drug release studies

Stability studies



4.5.01 Technological characteristics of floating tablets109

4.5.01.1 Weight variation test

20 tablets from each formulation were randomly picked up and weighed

individually and the average weight was calculated. The individual weights were then

compared with the average weight. For the tablets of average weight 350 mg, the %

deviation allowed is ± 5 %.

4.5.01.2 Friability

Ten tablets were weighed and placed in a Roche friabilator and rotated at

25 rpm for 4 min. The tablets were taken out, dedusted, and reweighed. The

percentage friability of the tablets was calculated using the equation:

% F = {1-(Wt/W)} ×100

Where, % F is percentage friability, W is the initial weight of tablet and Wt is the final

weight of tablets after revolutions.

Compressed tablets with a loss of less than 1 % are generally considered acceptable.

4.5.01.3 Hardness

The hardness of core tablets was measured using Inweka hardness tester. A

total of five tablets from each formulation were taken for the study and the average of

the three is reported. It is expressed in kg.

4.5.01.4 Thickness and diameter

Thickness and diameter of the tablets were determined by using Mitutoyo

micrometer screw gauge. The average of five tablets from each formulation was

taken. It is expressed in millimeter.



4.5.01.5 Uniformity of drug content

Drug content uniformity was determined by randomly selecting 5 tablets were

powdered. The quantity equivalent to single dose of the drug was dissolved in HCl

buffer solution, pH 1.2 for 5 hours with occasional shaking and diluted to 100 ml with

buffer. After filtration to remove insoluble residue, 1 ml of the filtrate was diluted to

10 ml with the buffer. The absorbance was measured at the required λmax using a UV

visible spectrophotometer. The experiments were carried out in triplicate for all

formulations and average values were recorded.

The drug content was calculated using the following equation:

% Drug content = conc. (μg/ml) × Dilution factor × 100/ 50

4.5.02 Drug-excipient compatibility studies

4.5.02.1 Fourier transform infra red spectroscopy (FT-IR)

In order to evaluate the integrity and compatibility of the drug in the

formulation, drug-excipient interaction studies were performed. Pure drug and

optimized formulations were analyzed by Fourier transform infra-red (FTIR)

spectroscopy. FTIR spectra of pure drug and its formulations were obtained by a

FT-IR Shimadzu 8400S (Japan) spectrophotometer using the KBr pellet method. The

samples were scanned from 400 to 4,000 cm−1

wave number.

4.5.02.2 Differential scanning calorimetry (DSC)

Differential scanning calorimetry was performed on pure sample of drug and

its formulation. Calorimetric measurements were made with empty cell (high purity

alpha alumina discs) as the reference. The dynamic scans were taken in nitrogen



atmosphere at the heating rate of 10 °C min-1

. The energy was measured as Joules per

kilocalorie.

4.5.03 In vitro floating studies110

The in vitro buoyancy was characterized by floating lag time and total floating

time. The test was performed using a USP dissolution apparatus type-II (basket) using

900 ml of 0.1 N HCl buffer solution at 100 rpm at 37 ± 0.5°C. The time required for

the formulation to rise to the surface of the dissolution medium and the duration for

which the formulation constantly floated on the dissolution medium were noted as

floating lag time and total time, respectively.

4.5.04 Water uptake studies111

The swelling of the polymers was measured by their ability to absorb water

and swell. The water uptake study of the tablet was done using a USP dissolution

apparatus type-II (basket) in 900 ml of pH 1.2 Hydrochloric acid buffer at 100 rpm.

The medium was maintained at 37 ± 0.5°C throughout the study. At regular time

intervals, the tablets were withdrawn, blotted to remove excess water, and weighed.

Swelling characteristics of the tablets were expressed in terms of water uptake (WU)

as:

WU (%) =Weight of Swollen tablet- Initial weight of tablet X 100

Initial weight of tablet

4.5.05 Percentage drug entrapment efficiency112

Floating microspheres equivalent to 4 mg of drug was dissolved in 10 ml

ethanol. The samples were assayed for drug content using UV spectrophotometer at

228 nm after suitable dilution. No interference was found due to the other components

of floating microspheres at 228 nm. The percentage drug entrapment efficiency and

yield were calculated as follows



% Drug entrapment efficiency = Calculated drug concentration × 100

Theoretical drug concentration

4.5.06 Yield of floating microspheres

The yield was determined by weighing the microspheres and then the

percentage yield was calculated with respect to the weight of the input materials, i.e.,

weight of rosiglitazone maleate and polymers used. The formula for calculation of

percentage yield is as follows

% Yield= Total weight of floating microspheres × 100

Total weight of drug and polymer

4.5.07 Scanning electron microscopy (SEM)

The surface morphology of the microspheres was examined by scanning

electron microscopy (SEM; JSM-5200, Jeol, Tokyo, Japan) operated at 15 KV on

samples, gold-sputtered for 120 s at 10 mA, under argon at low pressure.

4.5.08 Sphericity of the microsphere112

The sphericity of the prepared microspheres can be confirmed using a camera

lucida by taking the tracings of the microspheres on a black paper. The tracings help

to calculate the circulatory factor and confirm the sphericity of microspheres if the

obtained values are nearer to 1.

To determine the sphericity, the tracings of prepared microspheres

(magnification 45x) were taken on a black paper using camera lucida, (Model-Prism

type, Rolex, India). Circulatory factor (S) was calculated using

S= P2/12.5xA

Where A is area (cm2) and, P is the perimeter of the circular tracing.



4.5.09 Micromeritic properties of microsphere113, 114

The microspheres were characterized by their micromeritic properties, such as

particle size, bulk density, compressibility index and angle of repose (values useful in

prediction of flowability).

4.5.09.1 Particle size

The particle size of the microspheres was measured using an optical

microscopic method and the mean particle size was calculated by measuring 425

particles with the help of a calibrated ocular micrometer with stage micrometer.

4.5.09.2 Angle of repose

The flow characteristics of microspheres are measured by angle of repose.

Improper flow of microspheres is due to frictional forces between the microspheres.

These frictional forces are quantified by an angle of repose. Angle of repose is the

maximum angle possible between the surface of a pile of the microspheres and the

horizontal plane. Relationship between angle of repose and powder flow is given in

table 4.07. Fixed funnel method was employed. A funnel that was secured with its tip

at a given height above the graph paper was placed on a flat horizontal surface.

Microspheres were carefully poured through the funnel until the apex of the conical

pile just touches the tip of the funnel. The radius and height of the pile were then

determined. The angle of repose ( ) for samples were calculated using the formula

Tan θ = Height

Radius



Table 4.07: Relationship between angle of repose and powder flow

Angle of repose ( ) Flowability

<25 Excellent

25-30 Good

30-40 Passable

>40 Very poor

4.5.09.3 Tapped bulk density

The tapping method was used to determine the tapped density of the

microspheres using tapped density testing apparatus (Electrolab tapped density tester

ETD-1020) and percent compressibility index as follows

Tapped density = Mass of microspheres

Volume of microspheres after tapping

4.5.09.4 Compressibility (Carr’s) index

Carr‟s index is a dimensionless quantity, which proved to be useful to the

same degree as the angle of repose values for predicting the flow behavior. Apparent

bulk density was determined by pouring the samples in bulk into a graduated cylinder.

Tapped density was determined by placing a graduated cylinder containing a known

mass of powder on a mechanical Electrolab tap density tester. Samples were tapped

until no further reduction in volume of the sample was observed. Relationship

between powder flowability & % compressibility is shown in table 4.08. Carr‟s index

is calculated using the formula

% Compressibility index = 1- V/Vo × 100

Where Vo and V are the volumes of the sample before and after the standard tapping.



Table 4.08: Relationship between powder flowability & % compressibility

% Compressibility range Flow description

5-15 Excellent (free flowing granules)

12-16 Good (free flowing powder granules)

18-21 Fair (powdered granules)

23-28 Poor (very fluid powders)

28-35 Poor (fluid cohesive forces)

35-38 Very Poor

>40 Extremely poor

4.5.10 Floating Characteristics

4.5.10.1 In vitro buoyancy of microspheres71

The floatation study was carried out to ascertain the floating behaviour of the

microspheres prepared with various polymer combinations. Floating behaviour of

hollow microspheres was studied using a USP dissolution test apparatus II by

spreading the microspheres (100 mg) on 900 ml of 0.1 N HCl containing 0.02 % v/v

tween 80 as surfactant. The medium was agitated with a paddle rotating at 100 rpm

and maintained at 37° ± 0.5 °C for 12 h. Both the floating and the settled portions of

microspheres were collected separately. The microspheres were dried and weighed.

The percentage of floating microspheres was calculated using the following equation

% Floating capability = Weight of floating hollow microspheres × 100

Initial weight of hollow microspheres



4.5.10.2 In vivo floating behavior115

Barium sulphate loaded microspheres were prepared by adopting the

procedure as described earlier, except for using barium sulphate instead of drug.

Healthy rabbit weighing approximately 2.3 Kg was used to assess in vivo floating

behaviour. Ethical clearance for the handling of experimental animals was obtained

from the institutional animal ethical committee (IAEC) of JSS College of Pharmacy,

Mysore constituted for the purpose. The animal was fasted for 12 h and the first X-ray

photographed to ensure absence of radio opaque material in the stomach. The rabbit

were made to swallow barium sulphate loaded microspheres with 30 ml of water.

During the experiment, rabbits were not allowed to eat but water was provided. At

predetermined time intervals, the radiograph of abdomen was taken using an X-ray

machine.

4.5.11 In vitro drug release study108, 116,117

The release rate of drug from formulations was determined using USP

dissolution testing apparatus II (basket type). The dissolution test was performed

using 900 ml of 0.1 N HCl, at 37± 0.5 oC and 50 to 100 rpm. Aliquots (5mL) were

withdrawn at regular intervals for 12 h, sample was replaced by its equivalent volume

of fresh dissolution medium to maintain the sink condition. The samples were

analyzed spectrophometrically at wavelength corresponding to absorption maxima of

the drugs. The release kinetics was fitted into various models using PCP dissolution

v2.08 software.

4.5.12 Mechanism of drug release, 118,119,120,121

The different mathematical models may be applied for describing the kinetics

of the drug release process from dosage forms the most suited being the one which

best fits to the experimental results. The best models describe drug release from



pharmaceutical dosage form resulting from a simple phenomenon, or when this

phenomenon, by being the rate-limiting step, conditions all the process occurring in

the system. The kinetics of release from formulations were determined by finding the

best fit of the release data to zero order, first order, matrix(Higuchi), Hixson-Crowell,

and Korsmeyer- Peppas plots. Higuchi developed several theoretical models to study

release of high and low water soluble drugs incorporated in the semi-solid and/or

solid matrices. According to this model, drug release was described as a square root of

time-dependent diffusion process based on Fick‟s law. This relation can be used to

describe drug dissolution from several types of modified release pharmaceutical

dosage forms.

Qt =K H = √t

where KH is Higuchi‟s rate constant, and Qt is the amount of drug released at

time t. If a plot of square root of time versus cumulative amount of drug release yields

a straight line, and the slope is 1 or more than 1, then the particular dosage form is

considered to follow Higuchi kinetics of drug release. In some experimental situations

the release mechanism deviates from the Fick‟s equation, following an anomalous

behavior (Non-Fickian release). In these cases a more generic equation can be used.

Korsmeyer et al. developed a simple, semi-empirical, relating exponentially the drug

release to the lapsed time.

Qt/ Qα =Ktn

where Qt/Qα is the fraction of drug released at time t; K is the constant

comprising a structural and geometric characteristics of the tablets; and n, the release

exponent, is a parameter that depends on the release mechanism and is thus used to

characterize it.



Peppas used this n value in order to characterize different release

mechanisms. If the n value is 0.5 or less, the release mechanism follows Fickian

diffusion, and higher values (0.5 < n < 1) for mass transfer follow a non-Fickian

model (anomalous transport).

Hixson-Crowell recognized that area of the particle is proportional to the cubic

root of its volume, and derived an equation as follows

Wo1/3

- Wt

1/3 =Ks t

where Wo is the initial amount of drug, Wt is the remaining amount of drug in dosage

form at time t, and KS is a constant incorporating the surface volume relation.

4.5.13 In vivo evaluation38, 122

In vivo evaluation studies of the optimized microsphere formulation and pure

drug were carried out on normal healthy male albino rats selected with average body

weight of about 300-350 gm. They were housed individually in polypropylene cages,

maintained under standard conditions (12 h light and 12-h dark cycle; 27±2°C;

50±10% relative humidity); the animals were fed with standard rat pellet diet and

water with glucose. Ethical clearance for the handling of experimental animals was

obtained from the institutional animal ethical committee (IAEC) constituted for the

purpose. Non-insulin dependent diabetes mellitus (NIDDM) was induced in animals

which were fasted overnight by a single intraperitoneal injection of alloxan at the dose

of 120 mg/kg for all group animals except the group I animals, which served as

control. The blood glucose level was determined after 72 h of alloxan administration

using Glucometer. The animals with blood glucose level more than 187 mg/dl were

chosen for the experiment. All the animals showed hyperglycemia after 72 h of

alloxan administration. Only the rats found with permanent NIDDM were used for

in vivo evaluation studies. For the control (group I & II), the animals were kept fasting



overnight and water with glucose adlibitum. For group III and group IV, pure drug

and hollow microspheres were administered orally with oral gauss in the morning

following overnight fasting. No food and liquid except water with glucose were given

to the animals during the experiment. After collection of zero-hour blood sample, F3

was administered orally through oral gauss. Blood sample was collected from the tail

vein of the rat at every 1 h interval. Plasma glucose levels were determined using one

touch ACCU-Chek Active®. The number of animals required for the in vivo

evaluation studies is given the table 4.09.

Table 4.09: Number of animals required for the in vivo evaluation studies

Group

number

Treatment No. of animals

1 Positive control (normal control) 6

2 Negative control (diabetic control rat administered) 6

3 Pure drug (Rosiglitazone maleate of 4-mg/kg) 6

4 Optimized formulation ( F3 of 4-mg/kg) 6

Total 24



Figure 4.08: Administration of alloxan by I.P route

Figure 4.09: Administration of microspheres by oral gauze in suspension

form



Figure 4.10: Blood collection from rat tail vein

In vivo studies were carried out to monitor the gastric retention property of

floating tablet formulations. Barium sulfate loaded formulations were used as X- ray

markers. The study was conducted after obtaining approval from the Institutional

animal ethics committee of JSS College of Pharmacy, Mysore. Albino rabbits (2.5 kg)

were used in the study. Before the test, the rabbits were fasted overnight and the

formulations were administered orally to the rabbits with water. X- Ray pictures were

taken at different time intervals after the administration of the formulations.



4.5.14 Stability studies123

Stability testing of drug products begins as a part of drug discovery and ends

with the demise of the compound or commercial product. FDA and ICH specifies the

guidelines for stability testing of new drug products, as a technical requirement for the

registration of pharmaceuticals for human use. The objective of stability testing is to

investigate the effect of environmental factors on changes in product quality with time

so as to establish its shelf life and recommend its storage conditions.

Drug decomposition or degradation occurs during storage, because of

chemical alteration of the active ingredients or due to product instability, leading to

lower concentration of the drug in the dosage form, hence the stability of

pharmaceutical preparation needs to be evaluated. The objective of stability studies is

to predict the shelf life of a product by accelerating the rate of decomposition,

preferably by increasing the temperature and relative humidity (RH) conditions.

A drug formulation is said to be stable if it fulfills the following requirements:

It contains at least 90% of the stated active ingredient

It contains effective concentration of the added preservatives, if any

It does not exhibit discoloration or precipitation, nor develops foul odour

It does not develop irritation or toxicity

Formulations were packed in a screw capped bottle and studies were carried out

for 12 months by keeping at

25± 2°C and 60 ± 5% RH

30 ± 2 °C and 65 ± 5% RH

and for 6 months for accelerated storage condition at

40 ± 2°C and 75 ± 5% RH



Samples were withdrawn on 0, 3, 6 and 12 months for long term storage

condition and 0, 3 and 6 months for accelerated storage condition and checked for

changes in physical appearance and drug content as per ICH Q1A (R2) guidelines.

Graphs were plotted using Sigmaplot 12.0 to determine the statistical significance.

Results obtained in the methods and the conclusions arrived from them are

provided in the following chapters.

Documents

11_chapter 4.pdf - Shodhganga