INTRODUCTION:

Oral administration of drugs is the most common and preferred route for delivery of therapeutic agents. The popularity of the oral route is due to patient compliance, ease of administration, accurate dosing, cost-effective manufacturing methods, and generally improved shelf-life of the product. The rationale for development of controlled release formulation of a drug is to enhance its therapeutic benefits, minimizing its side effects. Much advancement have come about by the simultaneous convergence of many factors, including the discovery of novel polymers, formulation optimization better understanding of physiological and pathological constraints, prohibitive cost of developing new drug entities and the introduction of biotechnology and biopharmaceutics in the drug product design^1-3.

Recently numerous hydrophilic polymers have been investigated and are currently used in the design of complex controlled release systems. Among desirable features, the polymers should posse’s inherent physicochemical characteristics which provide for the attainment of high gel state viscosity upon swelling, ability to maintain constant gel layer integrity over a prolonged period and hence low erosion rate, and complete dissolution of polymer upon exhaustion of drug release. The most widely used polymers are the nonionic polyethylene oxide (PEO), hydroxypropyl methyl cellulose (HPMC), Hydroxypropyl cellulose (HPC) types. The cationic chitosan types and anionic alginate types, the attainment of high gel state viscosity, maintenance of constant gel layer, in a monolithic sense for linear drug release over a prolonged period of time is not easily achievable and still remains a challenge^4-5.

The drug such as Labetalol HCl was selected taking in to consideration of their physicochemical, biopharmaceutical properties and rationale of clinical efficacy.

Labetalol hydrochloride is a selective α-and nonselective β- adrenergic blocking agent. It is used in management of hypertension, alone or in combination with other classes of antihypertensive agents. It is one of several preferred initial therapies in hypertensive patients with heart failure, post- MI, high coronary disease risk, or Diabetes mellitus. It can be used as monotherapy for initial management of uncomplicated hypertension. It is also effective in controlling blood pressure in pregnant women with moderate to severe hypertension and severe pregnancy- induced hypertension. Labetalol hydrochloride has a dosage of, 100 mg twice daily initially. Labetalol hydrochloride is rapidly and almost completely absorbed from the GI tract following oral administration. It undergoes extensive first-pass metabolism in the liver and/or GI mucosa. Absolute bioavailability is about 25%. The half life of drug is short (2 to 4 hr).

Based on the above physical, chemical, biopharmaceutical, properties and clinical relevance, Labetalol HCl was selected as drug candidate for developing matrix tablets as controlled release systems.

Therefore to improve bioavailability, to reduce frequency of administration, to avoid the adverse effect of drug, which was caused by conventional dosage form, to maintain plasma levels of drug within a safe and effective range and to improve patient compliance, in this study an attempt has been made to formulate controlled release matrix tablets of Labetalol hydrochloride using various hydrophilic and hydrophobic polymers and the effect of various hydrophilic and hydrophobic polymers on drug release and other parameters were studied to optimize the final formulation.

MATERIALS AND METHODS:

Materials

Labetalol HCl was obtained as gift sample from Celon Labs Ltd. Hyderabad, India. All other materials, excipients, solvents and reagents were either analytical or Pharmacopoeial grade and they were procured from S.D. fine Chemicals Mumbai.

Methods

Drug- polymer interaction studies

Fourier Transform Infra-Red (FT-IR) spectral analysis

Fourier–Transform Infrared (FT–IR) spectrums of pure Atorvastatin calcium and combination of drug and excipients were obtained by a Fourier-Transform Infrared spectrophotometer, (FTIR-8300, Shimadzu, Japan) using the KBr disk method (2 mg sample in 200 mg KBr). The scanning range was 400 to 4000 cm^-1 and the resolution was 1cm^-1. This spectral analysis was employed to check the compatibility of drugs with the excipients used.

Differential Scanning Calorimetry (DSC) analysis

DSC analysis was performed using Shimadzu DSC-60, Shimadzu Limited Japan. A 1:1 ratio of drug and excipient was weighed into aluminium crucible. And sample was analyzed by heating at a scanning rate of 20⁰C over a temperature range 40-430⁰C under nitrogen environment.

Formulation of Labetalol matrix Tablets

The matrix tablets containing Labetalol was prepared by a direct compression technique. POLYOX WSR 301, WSR 303, Eudragit L 100, Eudragit S 100, ethyl cellulose, sodium alginate, xanthan gum, karaya gum and guar gum were used as polymers for controlling the drug release. The composition of various matrix tablet formulations were given in tables 8a and 8b. The Controlled release tablet formulations consisted of a drug and polymer were prepared in different ratios. The dose of the drug was maintained constantly while the proportion of polymers was varied for various matrix tablets.

Method of Preparation:

The formulations prepared are shown in tables 8a and 8b together with their compositions. The drug, polymer/s, and diluent were screened through # 40 and preblended using a lab scale double cone blender. The lubricant was added and the blend was mixed again prior to compression. The tablet blends were directly compressed by using ten station tabletting machine (Cemach machinery Co. Pvt, Ltd Ahmadabad, India) using a flat faced 6 mm punches (table 1 and 2).

Evaluation of Labetalol powder blend:

powder blend prepared by direct compression method were evaluated for various properties like bulk density, tapped density, compressibility index, Hausner ratio, flow properties (angle of repose) by using standard procedures. All studies were carried out in triplicate (n=3) and average values are reported with respective standard deviation.

Angle of repose:

The angle of repose of powder blend was determined by the funnel method. The accurately weight powder blend were taken in the funnel. The height of the funnel was adjusted in such a way the tip of the funnel just touched the apex of the powder blend. The powder blends were allowed to flow through the funnel freely on to the surface. The diameter of the powder blend cone was measured and angle of repose was calculated using the following equation.

tan θ = h / r,θ = tan^-1 (h / r)

Where, h = height of the powder cone.

r = radius of the powder cone.

Bulk density:

Both loose bulk density (D_b) and tapped bulk density (D_t) was determined. A quantity of 2 gm of powder blend from each formula, previously shaken to break any agglomerates formed, was introduced in to 10 ml measuring cylinder. After that the initial volume was noted and the cylinder was allowed to fall under its own weight on to a hard surface from the height of 2.5 cm at 2 second intervals. Tapping was continued until no further change in volume was noted. D_b and D_t were calculated using as the following equations.

D_b = Weight of the powder blend /Untapped Volume of the packing

D_t =Weight of the powder blend/Tapped Volume of the packing

Table 1: Composition of different batches of Labetalol matrix tablets

Ingredients Mg/tablets)	FORMULATIONS
Ingredients Mg/tablets)	F1	F2	F3	F4	F5	F6	F7	F8	F9
Labetalol	100	100	100	100	100	100	100	100	100
Polyox-WSR 303	60	120	180	60	120	180	60	120	180
Lactose	132.5	72.5	12.5	--	--	--	--	--	--
Starch 1500	--	--	--	132.5	72.5	12.5	--	--	--
DCP	--	--	--	--	--	--	132.5	72.5	12.5
MCC	--	--	--	--	--	--	--	--	--
Ethyl cellulose	--	--	--	--	--	--	--	--	--
Eudragit L100	--	--	--	--	--	--	--	--	--
Eudragit S 100	--	--	--	--	--	--	--	--	--
Magnesium stearate	1.5	1.5	1.5	1.5	1.5	1.5	1.5	1.5	1.5
aspartame	3	3	3	3	3	3	3	3	3
Orange flavour	3	3	3	3	3	3	3	3	3
Total weight	300	300	300	300	300	300	300	300	300

Table 1:Continued.....

Ingredients Mg/tablets)	FORMULATIONS
Ingredients Mg/tablets)	F10	F11	F12	F13	F14	F15	F16	F17	F18
Labetalol	100	100	100	100	100	100	100	100	100
Polyox-WSR 303	60	120	180	90	90	90	90	90	90
Lactose	--	--	--	--	--	--	--	--	--
Starch 1500	--	--	--	--	--	--	---	--	--
DCP	--	--	--	--	--	--	--	--	--
MCC	132.5	72.5	12.5	42.5	72.5	42.5	12.5	42.5	12.5
Ethyl cellulose	--	--	--	60	90	--	--	--	--
Eudragit L100	--	--	--	--	--	60	90	--	--
Eudragit S 100	--	--	--	--	--	--	--	60	90
Magnesium stearate	1.5	1.5	1.5	1.5	1.5	1.5	1.5	1.5	1.5
aspartame	3	3	3	3	3	3	3	3	3
Orange flavour	3	3	3	3	3	3	3	3	3
Total weight	300	300	300	300	300	300	300	300	300

#all the batches contained 0.5% w/w magnesium stearate; MCC-Microcrystalline cellulose; DCP – Dicalcium phosphate

Table 2: Composition of different batches of Labetalol matrix tablets

Ingredients Mg/tablets)	FORMULATIONS
Ingredients Mg/tablets)	F19	F20	F21	F22	F23	F24	F25
Labetalol	100	100	100	100	100	100	100
Polyox-WSR 303	90	90	90	90	90	90	90
Polyox-WSR 301	--	--	--	--	--	--	--
Karaya gum	60	90	--	--	--	--	--
Sodium alginate	---	--	60	90	--	--	--
MCC	42.5	12.5	42.5	12.5	42.5	12.5	42.5
Xanthan gum	--	--	--	--	60	90	--
Guar gum	--	--	--	--	--	--	60
Magnesium stearate	1.5	1.5	1.5	1.5	1.5	1.5	1.5
aspartame	3	3	3	3	3	3	3
Orange flavour	3	3	3	3	3	3	3
Total weight	300	300	300	300	300	300	300

Table 2: Continued....

Ingredients Mg/tablets)	FORMULATIONS
Ingredients Mg/tablets)	F26	F27	F28	F29	F30	F31	F32
Labetalol	100	100	100	100	100	100	100
Polyox-WSR 303	90	--	--	--	--	--	--
Polyox-WSR 301	--	60	120	180	60	120	180
Karaya gum	--	--	--	--	--	--	--
Sodium alginate	--	--	--	--	--	--	--
MCC	12.5	132.5	72.5	12.5	132.5	72.5	12.5
Xanthan gum	--	--	--	--	--	--	--
Guar gum	90	--	--	--	--	--	--
Magnesium stearate	1.5	1.5	1.5	1.5	1.5	1.5	1.5
aspartame	3	3	3	3	3	3	3
Orange flavour	3	3	3	3	3	3	3
Total weight	300	300	300	300	300	300	300

# All the batches contained 0.5% w/w magnesium stearate; MCC- Microcrystalline cellulose; DCP – Dicalcium phosphate

Compressibility index

The Compressibility Index of the powder blend was determined by Carr’s (compressibility) index. It is a simple test to evaluate the D_t and D_b of a powder blend and the rate at which it packed down. The formula for Carr’s Index is as below:

Where, D_t is the tapped density of the powder, D_b is the bulk density of the powder

Hausner’s ratio

Hausner’s ratio is an index of ease of powder flow; it is calculated by following formula.

Hausner ratio = D_t/D_b

Evaluation of Labetalol matrix tablets

The prepared tablets were evaluated for general appearance, content uniformity, hardness, friability, weight variation, thickness, diameter, disintegration time and swelling index, in vitro dissolution profile using methods specified in Indian Pharmacopoeia. The following evaluation tests were carried out on formulated tablets which includes;

i) General appearance:

The morphological characterization which includes size, shape, colour, presence or absence of odour, taste surface texture of the tablets was determined.

ii) Thickness and diameter:

Five tablets were picked from each formulation randomly and thickness and diameter was measured individually. It is expressed in mm and standard deviation was also calculated. The tablet thickness and diameter was measured using vernier calliper.

iii) Hardness:

Hardness indicates the ability of a tablet to withstand mechanical shocks while handling. The hardness of the tablets was determined using Monsanto hardness tester. It is expressed in kg/cm². Five tablets were randomly picked and hardness of the same tablets from each formulation was determined. The mean and standard deviation values were also calculated.

iv) Friability test:

Friability test is performed to assess the effect of friction and shocks, which may often cause tablet to chip, cap or break. Roche Friabilator was used for the purpose. Pre-weighed sample of ten tablets were placed in the Friabilator, which was then operated at 25 rpm for 4 minutes or ran upto 100 revolutions. After 100 revolutions the tablets were dusted and reweighed. Compressed tablets should not lose more than 1% of their weight.

The % friability was then calculated by the following formula:

Percentage friability =

(Initial weight - Final weight /Initial weight) × 100

v) Weight variation:

20 tablets were selected randomly from each formulation and weighed individually to check for weight variation. The US Pharmacopoeia allows a little variation in the weight of a tablet.

In all the formulations the tablet weight was 300 mg; hence 7.5% weight variation was allowed.

vi) Drug content:

Five tablets were weighed individually and powdered. The powder equivalent to average weight of tablets was weighed and drug was extracted in Phosphate buffer pH 6.8, the drug content was determined measuring the absorbance at 302 nm after suitable dilution using a Shimadzu UV- Vis double beam spectrophotometer 1601.

vii) In vitro disintegration time:

In vitro disintegration time was performed by apparatus specified in USP at 50 rpm. Phosphate buffer pH 6.8, 900 ml was used as disintegration medium, and the temperature of which was maintained at 37±2°C and the time in second taken for complete disintegration of the tablet with no palpable mass remaining in the apparatus was measured in seconds.

viii) Swelling index:

The extent of swelling was measured in terms of % weight gain by the tablet. The swelling behaviour of all formulation was studied. One tablet from each formulation was kept in a Petri dish containing pH 6.8 phosphate buffers. At the end of 0.5,1,2,3, 4,5,6,7,8 and 12hrs tablets were withdrawn, soaked on tissue paper and weighed and then percentage weight gain by the tablet was calculated by formula;

ix) In vitro drug release studies:

The following procedure was employed throughout the study to determine the in vitro dissolution rate for all the formulations.

Drug release study was carried out by using USP dissolution rate test apparatus-II (Electrolab, Mumbai, India). The study was conducted at 37°C and 50 rpm in 900 ml pH 6.8-phosphate buffer and studied for drug release up to 24 h. Five ml of sample was withdrawn at different time intervals, filtered and the drug content was estimated at 302 nm after suitable dilution.

x) Data Analysis:

To examine the drug release kinetics and mechanism, the cumulative release data of optimized formulation was fitted to models representing zero order (Q v/s t), first order [Log(Q₀-Q) v/s t], Higuchi’s square root of time (Q v/s t_1/2) and Peppas double log plot (log Q v/s log t) respectively, where Q is the cumulative percentage of drug released at time t and (Q₀-Q) is the cumulative percentage of drug remaining after time t.

xi) Scanning Electron Microscopy

The optimized formulation (F24) was selected for Scanning Electron Microscopy (SEM) analysis. The tablet surface morphology was studied at zero time and 22^nd hour of dissolution. The morphological characters of these 2 scans were compared to hypothesize the mechanism of drug release and swelling.

xii) Stability Studies:

Stability of a drug has been defined as the ability of a particular formulation, in a specific container, to remain within its physical, chemical, therapeutic and toxicological specifications.

In the present study, stability studies were carried out at 40⁰C±2⁰C/75%±5% RH for a period of 90 days for the selected formulations. The formulations were then evaluated for changes in the physicochemical properties, swelling study and in vitro drug release.

RESULTS AND DISCUSSION:

Drug-Excipients Compatibility Studies:

Fourier transform infrared (FTIR) analysis

Physical mixture of Labetalol and formulative ingredients were subjected for IR spectroscopic analysis to ascertain whether there was any interaction between drug and excipients used. The IR spectras showed similar characteristic peaks at their respective wavelengths with minor differences. The similarity in the peaks indicated the compatibility of drug with formulation excipients. IR spectra of the physical mixture of drug with formulative ingredients were depicted in figure 1 to 2.

Table 3: Pre compression evaluation of Labetalol powder blend

Formu lations	Bulk density (gm/cm³)*	Tapped density (gm/cm³)*	Carr’s Index (%)*	Hausner ratio (H_R)*	Angle of repose(q)*	flowability	Drug Content (%)
F1	0.673±0.010	0.776±0.029	13.265±1.672	1.152±0.02	25.19	Good	94.32
F2	0.589±0.023	0.666±0.031	11.607±1.262	1.131±0.025	23.89	Good	97.42
F3	0.628±0.031	0.702±0.028	10.476±1.623	1.117±0.027	26.68	Good	103.10
F4	0.661±0.028	0.758±0.039	13.065±1.213	1.149±0.031	26.53	Good	98.20
F5	0.611±0.033	0.717±0.027	12.962±1.278	1.148±0.008	21.28	Good	96.75
F6	0.634±0.007	0.634±0.018	11.538±1.291	1.130±0.023	23.77	Good	99.38
F7	0.568±0.025	0.653±0.016	10.344±2.328	1.115±0.032	25.62	Good	97.04
F8	0.584±0.027	0.640±0.026	10.619±1.259	1.118±0.039	24.94	Good	101.16
F9	0.573±0.031	0.694±0.023	10.434±1.906	1.157±0.029	27.61	Good	98.30
F10	0.603±0.008	0.660±0.013	13.636±2.018	1.150±0.011	24.50	Good	98.50
F11	0.573±0.023	0.680±0.036	11.818±0.775	1.134±0.028	24.61	Good	97.05
F12	0.667±0.032	0.715±0.031	11.447±1.243	1.29±0.031	23.51	Good	98.4
F13	0.633±0.039	0.649±0.035	11.498±2.332	1.118±0.029	20.24	Good	97.25
F14	0.574±0.027	0.717±0.014	12.447±1.259	1.126±0.039	21.09	Good	96.42
F15	0.628±0.029	0.648±0.019	11.316±2.329	1.122±0.027	22.36	Good	98.29
F16	0.574±0.009	0.718±0.009	12.212±1.837	1.134±0.029	20.42	Good	99.85
F17	0.584±0.011	0.711±0.017	14.051±2.985	1.136±0.033	22.13	Good	98.44
F18	0.627±0.034	0.714±0.029	12.220±1.916	1.112±0.031	22.42	Good	99.74
F19	0.628±0.015	0.702±0.036	11.538±1.213	1.117±0.023	22.20	Good	98.40
F20	0.634±0.17	0.709±0.028	12.264±1.105	1.139±0.028	24.02	Good	96.83
F21	0.622±0.012	0.648±0.019	10.476±1.958	1.130±0.010	23.47	Good	97.09
F22	0.603±0.008	0.660±0.013	13.636±2.018	1.150±0.011	23.32	Good	97.57
F23	0.573±0.023	0.680±0.036	11.818±0.775	1.134±0.028	22.70	Good	96.15
F24	0.611±0.033	0.717±0.027	12.962±1.278	1.148±0.008	22.62	Good	95.07
F25	0.634±0.007	0.634±0.018	11.538±1.291	1.130±0.023	22.15	Good	94.49
F26	0.574±0.027	0.717±0.014	12.447±1.259	1.126±0.039	23.25	Good	95.42
F27	0.584±0.011	0.711±0.017	14.051±2.985	1.136±0.033	22.13	Good	98.44
F28	0.667±0.032	0.715±0.031	11.447±1.243	1.29±0.031	23.51	Good	98.4
F29	0.634±0.007	0.634±0.018	11.538±1.291	1.130±0.023	23.77	Good	99.38
F30	0.589±0.023	0.666±0.031	11.607±1.262	1.131±0.025	23.89	Good	97.42
F31	0.573±0.023	0.680±0.036	11.818±0.775	1.134±0.028	22.70	Good	96.15
F32	0.661±0.028	0.758±0.039	13.065±1.213	1.149±0.031	26.53	Good	98.20

*All values are expressed as mean ± SD, n=3

The results of angle of repose and compressibility index (%) ranged from 20.24 to 27.61 and 10.344 to 14.051 respectively. The results of LBD and TBD ranged from 0.573 to 0.673 and 0.634 to 0.776 respectively. The drug content in a weighed amount of powder blend of all formulations ranged from 94.32 to 103.10%. Hausner’s ratio ranges from 1.112 to 1.157. Results of angle of repose (<30) indicate good flow properties of the powder blend. This was further supported by lower compressibility index values. Generally, compressibility index values up to 15% result in good to excellent flow properties. The drug content in the weighed amount of powder blend of all formulations was found to be uniform. All these results indicate that the powder blend prepared from different batches possessed good flow properties, compressibility, and drug content. The results of evaluation of labetalol powder blend are shown in table 3.

Post-compressional parameters:

Controlled release formulations for Labetalol were prepared by direct compression method using Cemach 10 station mini press. The direct compression process used for the preparation of matrix tablets was found to be ideal and is easy to reproduce. Polymers such as poly(ethylene oxides) [Polyox WSR 301 and Polyox WSR 303], ethyl cellulose, Polymethacrylates [Eudragit S 100 and Eudragit L 100], xanthan gum, guar gum, karaya gum and sodium alginate were used in the preparation of matrix tablets as controlled release polymers and exhibited good flow properties. These polymers were found to be ideal for the preparation of controlled release matrix tablets. Thirty two matrix tablet formulations were prepared with Labetalol by employing various polymers at different concentrations. As PEO’s are hydrophilic, the involvement of water or moisture makes the wet granulation process highly problematic. Therefore a dry process that produces acceptable powder characteristics and does not intervene with drug release characteristics is desirable. Hence the dry process such as direct compression technique was employed in the present investigation for the preparation of controlled release matrix tablets.

All the tablet formulations were evaluated for parameters such as shape, colour, thickness, hardness, friability, weight variation, drug content, in vitro disintegration time, in vitro dissolution studies, swelling study, model fitting of release profile and stability studies.

a) General appearance:

All the matrix tablets from each batch were found to be flat, white in color, circular in shape and having good physical appearance. There was no change in the color and odour of the tablets from all the batches.

b) Thickness and Diameter:

Thickness and diameter of formulated matrix tablets was ranged from 4.12±0.08 to 5.28±0.01mm and 7.01±0.01 to 7.05±0.03mm respectively. The values are almost uniform in all formulations.

c) Hardness:

The hardness of the tablets of all batches ranged from 6.5 ± 0.1 to 6.6 ± 0.2kg/cm ². It can be observed from the results that the hardness of all batches of matrix tablets was found to be uniform.

d) Friability:

Tablet hardness is not an absolute indicator of strength. Another measure of tablet strength is friability. Conventional compressed tablets that lose less than 1% of their weight are generally considered acceptable. In the present study, the percentage friability for all the formulations was below 1% indicating that the friability is within the prescribed limits.

e) Weight variation:

In a weight variation test, the United State Pharmacopoeial limit for the percentage deviation for the tablets of more than 324 mg is ±5%. The average percentage deviation of all tablet formulations was found to be within the above limit, it was found to be form 300±2.0 mg to 301±2.0 mg. and hence all formulations passed the test for uniformity of weight as per official requirements.

f) Drug content:

The content uniformity test was performed for all the formulations and drug content in the formulated tablets was ranged from 98.8±0.14 to 100.0±0.20. The results indicated that drug content was found to be uniform among different batches of the tablets.

g) In vitro disintegration study

All the tablets were found to be non–disintegrating in water and aqueous fluids of acidic (1.2) and alkaline (6.8) pH. As the tablets formulated with various polymers were non-disintegrating with acidic and alkaline fluids, they are considered suitable for oral controlled release. The results of evaluation of labetalol matrix tablets are presented in table 4.

Swelling Behavior of Labetalol matrix tablets

Since the rate of swelling is related and may affect the mechanism and kinetics of drug release, the penetration of the dissolution medium and swelling of tablets were determined. The extent of swelling was measured in terms of percentage weight gain by the tablets.

The swelling behavior of selected matrix tablet formulations (formulations containing natural gums) was studied. The swelling behavior indicates the rate at which the matrix tablet absorbs water from dissolution media and swells. The water uptake and swelling started slowly and continued for 12 hours. Constant and prolonged release of drug will occur in such situation because of increase in diffusion path length due to swelling of the matrix. The formulation containing xanthan gum and guar gum exhibited a high degree of swelling when compared with the formulations containing karaya gum and sodium alginate. Hence the drug release was extended for prolonged time for the formulation containing xanthan and guar gum. The tablets appeared swollen from the beginning and a viscous gel layer was formed when they came into the contact with the dissolution medium. The swelling behaviour of selected batches of Labetalol matrix tablets is presented in figure 5.

Table 4: Post compression evaluation of Labetalol matrix tablets

Formulation code	Diameter (mm)*	Thickness (mm)*	Hardness (kg/cm²)*	Friability (%)**	Weight variation (mg)***	Drug content (%)*	DT (min)	Appear-ance
F1	7.02±0.02	4.12±0. 08	6.6 ± 0.1	0.19	300 ± 2.0	98.9 ± 0.05	ND*	++
F2	7.01±0.02	4.27±0.01	6.5± 0.2	0.16	301 ± 2.0	99.3 ± 0.07	ND*	+++
F3	7.03±0.02	4.42±0.06	6.5 ± 0.3	0.16	300 ± 2.0	99.4 ± 0.02	ND*	+
F4	7.01±0.01	4.67±0.05	6.5 ± 0.2	0.15	300 ± 2.0	98.8 ± 0.14	ND*	++
F5	7.01±0.03	4.46±0.01	6.5 ± 0.2	0.18	301 ± 2.0	100.0 ±0.02	ND*	+
F6	7.01±0.04	5.19±0.06	6.6 ± 0.2	0.16	300 ± 2.0	100.0 ±0.01	ND*	+++
F7	7.02±0.01	4.37±0.08	6.5 ± 0.3	0.18	301 ± 2.0	99.3 ± 0.11	ND*	++
F8	7.03±0.02	4.36±0.01	6.5 ± 0.3	0.14	300 ± 2.0	99.3 ± 0.10	ND*	+
F9	7.04±0.01	5.01±0.02	6.5 ± 0.2	0.14	300 ± 2.0	99.1 ± 0.14	ND*	++
F10	7.05±0.03	4.97±0.03	6.5 ± 0.1	0.13	301 ± 2.0	99.4 ± 0.13	ND*	+
F11	7.02±0.01	5.15±0.02	6.5 ±0.2	0.12	301 ± 2.0	99.4 ± 0.13	ND*	+++
F12	7.01±0.02	5.28±0.01	6.5 ± 0.3	0.16	300 ± 2.0	100.0 ±0.02	ND*	++
F13	7.01±0.05	4.20±0.04	6.5 ± 0.2	0.17	300 ± 2.0	99.7 ± 0.11	ND*	+++
F14	7.02±0.06	4.26±0.03	6.5 ± 0.1	0.12	300 ± 2.0	100.0 ±0.01	ND*	+
F15	7.03±0.03	4.36±0.06	6.5 ± 0.3	0.15	300 ± 2.0	99.4 ± 0..20	ND*	++
F16	7.01±0.02	4.39±0.08	6.5 ± 0.2	0.16	300 ± 2.0	98.8 ± 0.24	ND*	++
F17	7.02±0.01	4.47±0.01	6.5 ± 0.4	0.10	300 ± 2.0	100.0 ±0.10	ND*	+++
F18	7.03±0.03	4.49±0.02	6.5 ± 0.1	0.15	300 ± 2.0	98.9 ± 0.31	ND*	+++
F19	7.04±0.02	4.37±0.03	6.5 ± 0.2	0.14	300 ± 2.0	100.0 ±0.11	ND*	++
F20	7.05±0.01	4.36±0.06	6.5 ± 0.2	0.10	300 ± 2.0	99.1 ± 0.15	ND*	+++
F21	7.02±0.01	4.85±0.07	6.5 ± 0.1	0.18	300 ± 2.0	99.4 ± 0.16	ND*	++
F22	7.01±0.03	4.12±0.08	6.5 ±0.2	0.16	300 ± 2.0	99.7± 0.11	ND*	+++
F23	7.03±0.02	4.27±0.08	6.5 ± 0.3	0.12	300 ± 2.0	100.0 ±0.10	ND*	++
F24	7.01±0.02	4.42±0.06	6.5 ± 0.4	0.15	300 ± 2.0	99.4 ± 0.24	ND*	+++
F25	7.01±0.03	4.67±0.00	6.5 ± 0.5	0.10	300 ± 2.0	100.0 ±0.20	ND*	++
F26	7.01±0.02	4.42±0.01	6.5 ± 0.1	0.15	300 ± 2.0	98.9 ±0.26	ND*	+++
F27	7.05±0.03	4.97±0.03	6.5 ± 0.1	0.13	301 ± 2.0	99.4 ± 0.13	ND*	+
F28	7.02±0.01	5.15±0.02	6.5 ±0.2	0.12	301 ± 2.0	99.4 ± 0.13	ND*	+++
F29	7.02±0.01	4.37±0.08	6.5 ± 0.3	0.18	301 ± 2.0	99.3 ± 0.11	ND*	++
F30	7.03±0.02	4.36±0.01	6.5 ± 0.3	0.14	300 ± 2.0	99.3 ± 0.10	ND*	+
F31	7.01±0.03	4.12±0.08	6.5 ±0.2	0.16	300 ± 2.0	99.7± 0.11	ND*	+++
F32	7.03±0.02	4.27±0.08	6.5 ± 0.3	0.12	300 ± 2.0	100.0 ±0.10	ND*	++

*All values are expressed as mean ± SE, n=5; **all values are expressed as mean ± SE, n=10; ***all values are expressed as mean ± SE, n=20; += Average; ++= good, +++= excellent, DT: disintegration time, ND: Non disintegrating.

Figure 5: Swelling behaviour of selected batches of Labetalol matrix tablets containing different concentrations of natural polymers

In vitro Drug Release Study

The in vitro drug release characteristics were studied in 900 ml of phosphate buffer pH 6.8 for a period of 24 hours using USP XXIII dissolution apparatus 2 (paddle type at 50 rpm).

Labetalol HCl release from the matrix tablets contained POLYOX WSR 301, POLYOX WSR 303 were performed in pH 6.8 phosphate medium for 24 hrs. From the In vitro dissolution studies it was also observed that the high molecular weight poly (ethylene oxide) i.e. POLYOX WSR 303, effectively controlled the release rate of the drugs for an extended period of time than the low molecular weight POLYOX WSR 301. This was due to their high molecular weight poly (ethylene oxides) such as POLYOX WSR 303 has slow swelling rates, where the release rate of the drug will be slow when compared with the low molecular weight poly(ethylene oxides).

The effect of hydrophilic and hydrophobic diluents on the drug release was studied. The diluents selected were lactose, microcrystalline cellulose, dicalcium phosphate, and starch 1500. The drug release from the matrix tablets containing lactose as diluent was rapid when compared to the other diluents such as microcrystalline cellulose, DCP and starch 1500. This was due to hydrophilic nature of the lactose than other diluents, which resulted in faster penetration of dissolution media i.e. easier penetration of dissolution medium into the tablet matrix, which lead to weaker gel strength, higher erosion of gel layer and therefore faster drug release from the matrix. Insoluble but weakly swellable fillers such as MCC and DCP remained within the gel structure and resulted in the slow release rate of the drugs. Hence MCC and DCP as insoluble diluents provided the slower rate of drug release. These excipients have minimum swelling property which had contributed to the swollen matrix for PEO and retarded the penetration of dissolution medium. Reports indicated that MCC has strong tablet binding properties which decrease the tablet porosity. This nature of MCC is also responsible for the extended drug release from the matrix tablets. The formulations containing starch 1500 which is slightly soluble filler, the drug release from the matrix tablets were extended up to 16 hrs. This was due to the diluent retarded the easier penetration of dissolution medium into matrix and thus prevented the polymer matrix erosion. The swelling nature of starch 1500 upon exposure to dissolution medium resulted in the formation of a gel layer which controlled the release rate of Labetalol from matrix tablets. The matrix tablet formulations F3, F6, F9 and F12 extended the drug release up to 18 hrs. The influence of binary polymeric systems on the drug release was evaluated. The matrix tablets were formulated using combinations of both hydrophilic and hydrophobic polymers. For this hydrophilic PEO and hydrophobic eudragits and ethyl cellulose were used in the formulation of matrix tablets. Combinations of synthetic and natural polymers were also used in preparing the matrix tablets to study their influence on controlled release of drug. Combinations of PEO with various natural gums like xanthan gum, karaya gum, guar gum and sodium alginate were used in the matrix tablet formulations.

The drug release from the matrix tablets containing PEO and eudragits (F15-18) showed linear drug release over a period of 18 hrs. The initial burst release of the drug was not observed in these formulations. This was due to the presence of insoluble eudragit in the matrix which retarded the faster drug diffusion due to the formation of a rigid matrix. The gel structures formed around the matrix tablets were rigid when compared with the formulations containing PEO and natural gums. The ‘n’ value obtained from the peppas plots for these formulations were in the range of 0.5 to 0.89. These values indicated that the drug release is by both diffusion and matrix relaxation mechanisms for these matrix tablets containing Labetalol.

The drug release from the matrix tablets containing PEO and ethyl cellulose (F13 and F14) was extended over a period of 20 hrs. The presence of hydrophobic ethyl cellulose in the matrix prevented the rapid gel formation. Since ethyl cellulose is a hydrophobic polymer it cannot swell similar to that of PEO and thus retarded the rapid drug diffusion from the matrix. The ‘n’ value obtained from the peppas plots for these formulations were in the range of 0.5 to 0.89. These values indicated that the drug release is by both diffusion and matrix erosion mechanisms for these matrix tablets containing Labetalol.

The drug release form the tablets containing PEO and xanthan gum (Formulations F23 and F24) showed controlled release from 16 to 22 hrs than the formulations containing combination of PEO and various gums like guar gum, karaya gum and sodium alginate. This is due to high degree of swelling and slow erosion due to polymer relaxation for xanthan gum than other gums which has been observed from the swelling index studies. The drug release from the tablets containing PEO, karaya gum and guar gum (Formulations F19, F20, F25 and F26) was extended up to 18 hrs. It was also observed that increase in the concentration of gums, the drug release was extended. This is due the hydrophilic nature of the poly (ethylene oxide) and gums. These tablets showed greater water uptake which resulted in the formation of highly viscous gel layer around the tablet. The formed gel layer resulted in the longer diffusional path length, there by retarding the drug diffusion.

A different drug release profile was observed from the matrix tablets containing PEO and sodium alginate (Formulations F21 and F22). Initial burst release of the drug was not observed which was also observed for the formulations containing eudragits. This was due to the hydrophilic polymer sodium alginate is an anionic linear polysaccharide which is insoluble at acidic pH. But the sodium alginate retarded the matrix tablet to swell. They remain insoluble and retarded the rapid drug diffusion. And hence the initial burst release of the drug was not observed.

It was also observed that an increasing in the PEO content, the drug release rate from the matrix tablets was decreased. The correlation coefficient values calculated for the first order plots for the matrix tablet formulations were linear, which indicated the drug release from the matrix tablet formulations followed first order kinetics (figure 6-11).

Formulation F24 showed extended drug release over a period of 22 hrs and other formulations showed lower drug release. Based on the swelling index studies and in vitro drug release data formulation F24 was considered optimized batch.

Figure 6: Comparative in vitro release profile from different batches of Labetalol matrix tablet (F1-F6)

Figure7: Comparative in vitro release profile from different batches of Labetalol matrix tablet (F7-F12)

Figure 8: Comparative in vitro release profile from different batches of Labetalol matrix tablet (F13-F18)

Figure 9: Comparative in vitro release profile from different batches of Labetalol matrix tablet (F19-22)

Figure 10: Comparative in vitro release profile from different batches of Labetalol matrix tablet (F23-26)

Figure 11: Comparative in vitro release profile from different batches of Labetalol matrix tablet (F27-32)

Scanning Electron Microscopy

The surface morphology of optimized formulation (F24) at zero time and at 22^nd hour of dissolution study was observed. SEM photographs before dissolution it showed intact surface without any perforations, channels, or troughs. After dissolution, the solvent front enters the matrix and moves slowly toward the center of the tablet. The drug diffuses out of the matrix after it comes in contact with dissolution medium. The images of the tablet showed the presence of both gelling structures and pores on the surface. Thus, the presence of both pores and gelling structure indicates the combination of diffusion and erosion mechanism in the release of labetalol from the matrix tablet of batch F24. The SEM photographs of labetalol matrix tablet (F24) were shown in figure 12.

At zero time of dissolution study

At 22^nd hour of dissolution study

Figure 12: SEM photomicrographs of optimized batch of labetalol matrix tablet (F24)

Mechanism of drug release:

To determine the mechanism of drug release kinetics from optimized formulation F24, the dissolution data were treated according to Higuchi (cumulative percentage of drug released vs. square root of time), Korsmeyer-Peppas model (log cumulative percentage of drug released vs. log time) equations and Hixson-Crowell model (cube root % drug remaining vs time) along with zero order (cumulative amount of drug released vs. time) pattern. The data were processed for regression analysis using MS EXCEL statistical function. It can be observed from the results that the release rate data of optimized formulation of labetalol matrix tablets F24 formulated using xanthan gum as the matrix did not follow a zero-order release pattern. By using Higuchi’s kinetics or square-root kinetics this would explain why drug diffuses at a comparatively slower rate as the distance of diffusion increases. In our experiments, the in-vitro release profiles of drug from optimized formulation F24 could be best expressed by Higuchi’s equation, as the plots showed high linearity (R²= 0.9908).

To confirm the diffusion mechanism, the data were fit into Korsmeyer-Peppas model. The optimized formulation F24 showed high linearity (R²= 0.9907, with slope (n) values 0.6661, this (n) value indicating that coupling of diffusion and erosion mechanism so called anomalous non-Fickian diffusion and may indicate that the drug release is controlled by more than one mechanism, which indicate that formulation F24 release the drug by diffusion coupled with erosion mechanism. The result of modelling and drug release kinetics of optimized labetalol matrix tablet Batch F24 were shown in table 5.

Stability studies:

The formulation which showed good in vitro performance (F24) was subjected to accelerated stability studies. These studies were carried out by investigating the effect of temperature on the physical properties of the tablets and on drug release from the matrix as per ICH guidelines. Formulation F24 was subjected to accelerated stability studies.

The results indicated that there was no visible and physical changes observed in the matrix tablets after storage. It was also observed that there was no significant change in drug release from the matrix tablets. The slow and controlled drug release characteristics of the matrix tablets remained unaltered. Thus the drug release characteristics of controlled release matrix tablets designed were found to be quite stable (figure 13).

Figure 13: Comparison of in vitro release profile of optimized formulation of Labetalol matrix tablet (F24) after stability study

CONCLUSION:

The following conclusions were drawn from the results:

· Labetalol was freely water soluble drugs found to be suitable for formulating as controlled release matrix tablets with POLYOX WSR 301 and POLYOX WSR 303 by direct compression process.

· The direct compression process employed for the preparation of matrix tablets were found suitable with the drugs and all the polymers used.

· The physical parameters evaluated for the matrix tablet formulations such as weight uniformity, hardness, friability and drug content were uniform and were within the IP limits.

· Weight uniformity of matrix tablet formulations were uniform in all the cases and were maintained with in I.P specified limits.

· Hardness of all the matrix tablet formulations was found to be within the range of 6.5 ± 0.5 Kg/cm². Friability loss was negligible, less than 0.20 % for all the matrix formulations.

· Drug content was evaluated for all the matrix tablet formulations and found to be uniform. Drug content for the matrix tablet formulations was found to be within the specified range for labetalol extended release tablets USP.

· FTIR spectral studies of selected formulations of labetalol exhibited no major interactions between the drug, polymer and diluents.

· The matrix tablets containing POLYOX WSR 301 extended the release of labetalol up to 12 hrs. The drug release from these matrix tablets followed anomalous transport mechanism. As the concentration of the polymers increased the drug release was extended. Formulations F28, F29, F31 and F32, were found to extend the linear drug release upto 12 hrs.

· The drug release from the matrix tablets containing POLYOX WSR 303 was extended up to 22 hrs. Drug release from the tablets depended up on the polymer concentration, polymer combinations and type of diluents used. Drug release form these matrix tablets followed anomalous transport mechanism. Formulations F6, F9, F12, F14, F18, F20, F22, F24, F26 were found to extend the linear drug release upto 22 hrs.

· Diluents such as microcrystalline cellulose, dicalcium phosphate, starch 1500 have high influence on extending the drug release over a prolonged period of time. The order of delay in drug release in presence of diluents in the matrix tablet formulations were Starch 1500 > DCP > MCC > Lactose.

· Binary polymeric systems used for the preparation of the matrix tablets for labetalol had influence on extending the drug release. Among the binary polymeric systems used for the preparation of matrix tablets, combination of poly (ethylene oxide) (Polyox WSR 303) and ethyl cellulose, eudragits, xanthan gum, guar gum, karaya gum extended the drug release for a period of 18 to 22 hrs. Formulations F12, F14, F20, F24 and F26 showed prolonged release for a period of 18-22 hrs.

· Log percentage drug undissolved versus time plots for first order release rate constant of all the prepared matrix tablets were found to be linear with R²values of 0.91-0.97.

· Amount of drug release versus square root of time plots for all the matrix tablet formulations were linear which indicated that the drug release from the matrix tablet formulations is by diffusion process.

· The ‘n’ value obtained from the peppas plots for the matrix tablet formulations were in the range of 0.5 to 0.89. These values indicated that the drug release is by both diffusion and matrix relaxation mechanisms for the matrix tablets containing Labetalol.

· No significant changes were observed in the physical characteristics and in the drug release profiles of selected matrix tablet formulations of Labetalol after storing them at accelerated storage conditions.

ACKNOWLEDGEMENTS:

The authors are thankful to Principal and Management of Karavali College of Pharmacy, Mangalore for providing all the facilities and support for this research project. The authors are also thankful to Celon Labs Ltd. Hyderabad, India for generous gift samples of Labetalol HCl.

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3. Ritchel WA. Biopharmaceutics and pharmacokinetic aspects in the design of controlled release per-oral drug delivery system. Drug Dev Ind Pharm 1989; 15: 1073-103.

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Received on 04.04.2016 Accepted on 25.04.2016

Asian J. Pharm. Res. 2016; 6(2): 107-120

DOI: 10.5958/2231-5691.2016.00018.6

Formulation	First order		Zero order		Higuchi’s		Peppa’s
F24	r²	K	r²	K	n	r²	n	r²
F24	0.991	0.087	0.780	4.41	27.37	0.993	0.67	0.991