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Chapter.

Methodology

5. METHODOLOGY
5.1 Characterization of API 1. Official monograph: In house 2. Description: A white to off-white powder 3. Identification: By HPLC: The retention time of the principal peak in the sample preparation for assay should corresponds with the retention time of the principal peak in the standard preparation for assay1. 4. Related substances: Unknown Impurities: Not more than 0.1% Total Impurities: Not more than 0.5% 5. Assay by HPLC: 99.87% (98% - 102%w/w) 6. Loss of drying: 0.67% (Not more than 1.0%) 7. Heavy metals: Not more than 0.001% w/w 8. Sulphated ash: Not more than 0.2% w/w 9. pH: Observed 4.0 Specification: 3.5 5.0 10. Melting point: 142~146c 11. Half-life:13 hour 5.2 Selection and justification of excipients2 Diluents: In view of the low or medium dose of drug it is essential to add bulking agents or diluents to increase the weight of the tablet. Dibasic calcium phosphate was used as diluents because of its good compaction property and good flow property of the coarse grade material and also it is insoluble in water. So used as diluents in SR preparation. Matrix-forming polymers: HPMC which is most widely used matrix-forming polymer in these days because of its excellent compatibility, multifunctional property. It is available in different grades depending upon its attached group. It enables control time release of active ingredient by pH independent swelling and permeability.

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Chapter.5

Methodology

Lubricants: Magnesium Stearate, talc &colloidal silicon dioxide as lubricating agent. Table 5.1 Ingredients and their function
Sr. NO 1. 2. 3. AED LACTOSE MONOHYDRETE HPMC K4M API Diluent Thickening agent; viscosity-increasing agent. 3. 4. 5. 6. HPMC K100M HPMC K15M IPA Microcrystalline cellulose(MCC) 7. 8. 8. Colloidal silicon dioxide TALC Magnesium stearate Glidant (0.25-3%) Glidant Lubricant (0.25-4%) Matrix-forming polymer Matrix-forming polymer Binder solution Diluent Ingredients Function

5.2 Preformulation study Preformulation can be defined as investigation of physical and chemical properties of drug substance alone and when combined with excipients. Preformulation studies are the first step in the rational development of dosage form of a drug substance. The objectives of preformulation studies are to develop a portfolio of information about the drug substance, so that this information is useful to develop formulation. Preformulation investigations are designed to identify those physicochemical properties and excipients that may influence the formulation design, method of manufacture, and pharmacokinetic-biopharmaceutical properties of the resulting product. Followings studies performed for in the preformulation study. 3-7 5.3 Physical appearance:5.3.1 Description Colour: - A white to off-white powder

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Chapter.5
Table 5.2 Effect of angle of repose () on flow property Angle of repose () < 20 20-30 30-34 >35 5.3.2 Solubility: Freely soluble in water, 0.1N HCl and pH 4.5 Acetate buffer. Soluble in pH 6.8 phosphate buffer and pH 7.5 phosphate buffer . Type of flow Excellent Good Passable Very poor

Methodology

5.3.3 Bulk density: a) Loose bulk density: Weigh accurately 25 g of drug (M), which was previously passed through 20 # sieve and transferred in 100 ml graduated cylinder. Carefully level the powder without compacting, and read the unsettled apparent volume (V0). Calculate the apparent bulk density in gm/ml by the following formula

Bulk density = Weight of powder / Bulk volume..


b) Tapped bulk density: Weigh accurately 25 g of drug, which was previously passed through 20 # sieve and transfer in 100 ml graduated cylinder. Then mechanically tap the cylinder containing the sample by raising the cylinder and allowing it to drop under its own weight using mechanically tapped density tester that provides a fixed drop of 14 2 mm at a nominal rate of 300 drops per minute. Tap the cylinder for 500 times initially and measure the tapped volume (V1) to the nearest graduated units, repeat the tapping an additional 750 times and measure the tapped volume (V2) to the nearest graduated units. If the difference between the c) two volumes is less than 2% then final the volume (V2).Calculate the tapped bulk density in gm/ml by the following formula:

Tapped density = Weight of powder / Tapped volume

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Chapter.5
5.3.4 Carrs index

Methodology

The Compressibility Index of the powder blend was determined by Carrs compressibility index. It is a simple test to evaluate the BD and TD of a powder and the rate at which it packed down. The formula for Carrs Index is as below:

Carrs index (%) = [(TD BD)*100] / TD..

5.3.5 Hausners ratio The Hausners ratio is a number that is correlated to the flowability of a powder or granular material.

Hausners ratio = TD / BD..

Table 5.2 Effect of carrs index and hausners ratio on flow property Carrs index (%) < 10 1115 1620 2125 2631 3237 >38 Flow character Excellent Good Fair Passable Poor Very poor Very, very poor Hausners ratio 1.001.11 1.121.18 1.191.25 1.261.34 1.351.45 1.461.59 >1.60

5.3.6 Angle of repose The angle of repose of API powder 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 blend was allowed to flow through the funnel freely on to the surface.

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Chapter.5

Methodology

The diameter of the powder cone was measured and angle of repose was calculated using the following equation.

Where, h and r are the height and radius of the powder cone respectively. 5.3.7 Particle size analysis: ( By malvern master seizer, dry method) For many active substances, particle size has an impact on powder flow; content uniformity and drug dissolution. In order to assure consistent product quality, the particle size of the API has been characterized. From the results obtained, the limits will be derived which will be routinely applied by the API manufacturer during analysis of drug. Particle size of drug was determined by Malvern particle size analyser. D (0.10) = 2.39 , D (0.50) = 7.37 , D (0.90) = 19.46 .

5.3.8: Assay Standard solution Parameters for Preparation of Standard Solution Column: Inertsil ODS 3r (2504.6) Colum Id: AD3/Lc-074/10 Wavelength: 220 nm Flow rate: 1.0ml/min Injection volume: 10l Column temp. : 45C Diluent: Water: CAN

First prepare buffer solution by 2.7gm KHPO4in 2 litre purified water. Then prepare mobile phase by addition of ACN with the ratio of 40:60.Then prepare diluent with addition of water: methanol (40:60). Finally 50 mg API is added in to the 25 ml diluent. The max of API in the above media was determined by scanning a suitable dilution of the stock. From the stock

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Chapter.5

Methodology

solution, various dilutions were made to obtain solutions of 1, 5, 10, 15 and 20 _g/ml, and absorbance was measured for each dilution. 5.3.9 Drug excipients compatibility study: API and excipients were been thoroughly mixed in predetermined ratio given in below table and passed through the 40# sieve. The blend was to be filled in transparent glass vials and were closed with gray coloured rubber stoppers and further sealed with aluminium seal and charged in to stress condition at above condition. Similarly API should also be kept at all condition as for the samples. Samples were withdrawn for analysis within two day of sampling date as per the compatibility study plan. Physical observation should be done at every week up to 1 month and DSC studies were carried out to determine the compatibility of excipients with the drug.8

5.3.10InVitro Release study of theoretical and Hypothetical Release profile


Dissolution parameter: Medium: 0.1N Hydrochloric acid Volume: 900ml Apparatus: IP (Paddle) RPM: 50 rpm Time point: 1, 2, 4, 6, 8, 10, 12 hrs. Temperature: 37C 0.5C Table 5.3InVitro Release study of Innovator and Hypothetical Release profile Dissolution Time (hrs.) 1 2 900ml, USP - II (Paddle) Apparatus, 50 RPM Theoretical % Drug Release 23.9 36 PH6.8 Phosphate buffer 4 8 12 53.5 89.6 97.6 50-55% 85-90% NLT 90% Page 56 Hypothetical % Drug Release 0.1N HCL 15-25% 30-35%

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Chapter.5

Methodology

5.3.11 Drug- Excipient Compatibility Study

Figure 5.5 Drug- Excipient Compatibility Study


Ratio of Dry Ingredients mix Drug: Initial 1 week 2 week 3 week 4 week

No: 1
API

Excipi ent

-----

White crystalline powder

No change

No change

No change

No change

LACTOSE MONOHY DRETE

1:1

White crystalline powder

no change

no change

no change

No change

HPMC K4M

1:1

White crystalline powder

No change

No change

No change

No change

HPMC K100M

1:1

White crystalline powder

No change

No change

No change

No change

HPMC K15M

1:1

White crystalline powder

No change

No change

No change

No change

Microcrys talline cellulose( MCC)

1:1

White crystalline powder

No change

No change

No change

No change

Colloidal silicon dioxide

1:1

White crystalline powder

No change

No change

No change

No change

TALC

1:1

White crystalline powder

No change

No change

No change

No change

Magnesiu m stearate

1:1

White crystalline powder

No change

No change

No change

No change

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Chapter.5
5.4. Formulation of trials:
5.6.FORMULATION OF TRIALSF001 TO F003

Methodology

SR NO 1 2

INGREDIENT

F-1

F-2

F-3

QUANTITY OF DRUG LACTOSE MONOHYDRATE

300 177

300 177

300 169.5

3 4 5 6 7 8

HPMC K4M HPMC K15M HPMC K100M IPA MCC COLLOIDAL SILICONE DIOXIDE

40

40

50 Q.S 56.5 6

Q.S 59 6

Q.S 59 6

9 10 11

TALC MG STEARATE TOTAL

12 6 600

12 6 600

12 6 600

5.4.1 (A) Method of preparation of Extended release tablets:


Method: API SR Tablets were prepared by Wet granulation technique. For F/001 to F/007:

Step 1: Sifting: API was passed through 16 #sieve, and pass HPMC K4M, HPMC K15M, LACTOSE
MONOHYDRETE through 40#seive

Step 2: Wet Granulation: by ISOPROPAYL ALCOHOL

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Chapter.5
Granulation:

Methodology

Table 5.8 Wet granulation parameters BINDER Time Impeller speed Dry mixing Binder addition Extra IPA addition Kneading 2minute 150RPM 1500RPM 40ml 2 minute 150RPM Off 150ml 2minute 150RPM Off 10minute 150RPM Off Chopper speed

Step 3: Drying: Granules were dried in Fluidized bed dryer. Inlet temperature: 45C Outlet temperature: 35 C LOD: 0.62 to 1.03 % Step 4: Sifting: Granules were passed through 20 # by manually. And retained 20# grenules passed through co-mill 1905 sieve Step 5: calculation Calculation of extra granular part Step 6: shifting MCC (Microcrystalline cellulose), colloidal silicone dioxide and TALC were passed through 40 # and Magnesium stearate through 60 #.

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Chapter.5
Step 5: Blending and Lubrication:

Methodology

Blending time for MCC, TALC & Colloidal silicon dioxide was 10 minutes and lubricated with magnesium stearate for 3 minutes. Step 6: Compression: The prepared blend was compressed (16 8.0 mm oval shape standard shape with concave,) using 16 station tablet compression machine (Cadmach, Ahmedabad, India).

5.4.1 (B) Evaluation of powder blend: Evaluation parameters of powder blend were done as in Preformulation study.

5.4.1 (C) Evaluation of tablets: 1. Appearance: Twenty tablets of each formulation were taken to check any discoloration or surface ruffness in the tablet formulation. 2. Weight variation test: To study weight variation twenty tablets of the formulation were weighed using a Mettler Toledo electronic balance and the test was performed according to the official method.9 3. Hardness: The hardness of five tablets was determined using the Dr.schleunizer type hardness tester and the average values were calculated. 4. Thickness and diameter: The Thickness and Diameter of the tables was determined by using Digital vernier calipers. Five tablets were used, and average values were calculated. 5. Friability: The friability of twenty tablets was measured by Roche friabilator for 4min at 25rpm for 100 revolutions. Accurately weigh twenty tablets placed into Roche friabilator for 100 revolutions than dedust the tablets and weigh.

% Friability = (W0 - W)* 100 / W..


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Chapter.5
5.5. Formulation of trialsF004 TO F008:-

Methodology

Table 5.9 Formulation of trialsF004 TO F008 Sr no 1 2 QUANTITY OF DRUG LACTOSE MONOHYDRATE 3 4 5 5 6 HPMC K4M HPMC K15M HPMC K100M IPA MCC(MICROCRYSTE LLAIYINE CELLULOSE) 7 8 9
COLLOIDAL SILICONE DIOXIDE

INGREDIENT

F/004

F/005

F/006

F/007

300 162

300 162

300 162

300 162

60 Q.S 54

10 50 Q.S 54

20 40 Q.S 54

30 30 Q.S 54

6 12 6

6 12 6

6 12 6

6 12 6

TALC MGNESIUM STEARATE

10

TOTAL

600

600

600

600

5.5.1 (A) Method of preparation of Extended release tablets:


Method: API SR Tablets were prepared by Wet granulation technique. For F/001 to F/007: Step 1: Sifting: API was passed through 16 # sieve, 0and pass HPMC K4M, HPMC K15M, LACTOSE MONOHYDRETE through 40#seive

Step 2: Wet Granulation: by ISOPROPAYL ALCOHOL

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Chapter.5
Granulation:
Table 5.10 Wet granulation parameters BINDER Time Impeller speed Dry mixing Binder addition Extra IPA addition Kneading 2minute 150RPM 1500RPM 40ml 2 minute 150RPM 1500RPM 150ml 2minute 150RPM Off 10minute 150RPM Off hopper speed

Methodology

Step 3: Drying: Granules were dried in Fluidized bed dryer. Inlet temperature: 45C Outlet temperature: 35 C LOD: 0.62 to 1.03 % Step 4: Sifting: Granules were passed through 20 # by manually. And retained 20# grenules passed through co-mill 1905 sieve Step 5: calculation Calculation of extra granular part Step 6: shifting MCC (Microcrystalline cellulose), colloidal silicone dioxide and TALC were passed through 40 # and Magnesium stearate through 60 #. Step 5: Blending and Lubrication: Blending time for MCC, TALC & Colloidal silicon dioxide was 10 minutes and lubricated with magnesium stearate for 3 minutes.

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Chapter.5
Step 6: Compression:

Methodology

The prepared blend was compressed (16 8.0 mm oval shape standard shape with concave,) using 16 station tablet compression machine (Cadmach, Ahmedabad, India).

5.5.1 (B) Evaluation of powder blend: Evaluation parameters of powder blend were done as in Preformulation study.

5.5.1 (C) Evaluation of tablets: 1. Appearance: Twenty tablets of each formulation were taken to check any discoloration or surface ruffness in the tablet formulation. 2. Weight variation test: To study weight variation twenty tablets of the formulation were weighed using a Mettler Toledo electronic balance and the test was performed according to the official method.9 3. Hardness: The hardness of five tablets was determined using the Dr.schleunizer type hardness tester and the average values were calculated. 4. Thickness and diameter: The Thickness and Diameter of the tables was determined by using Digital vernier calipers. Five tablets were used, and average values were calculated. 5. Friability: The friability of twenty tablets was measured by Roche friabilator for 4min at 25rpm for 100 revolutions. Accurately weigh twenty tablets placed into Roche friabilator for 100 revolutions than dedust the tablets and weigh.

% Friability = (W0 - W)* 100 / W

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Chapter.5
5.5.2 (D).4 Selection of packaging material:

Methodology

Tablets will be packed in a blister formed by the following primary packaging components. Base foil: PVC/PVDC blister pack. Lidding Material: 0.025 mm aluminum foil
Justification: PVC/PVDC Blister pack provides complete protection against light, water vapour, gases etc.

5.6Conclusion From the result, concluded that batch taken in combination with HPMC K4M&HPMC K100M (30+30%) in has good sustained property. Batch F007 was charged for Accelerated stability study. 5.7. Stability studies of optimized tablets The ICH Guidelines have established that long term stability testing should be done at 25C2C / 60%5% RH; stress testing should be done at 40C2C / 75%5% RH for 6 months. If significant change occurs at these stress conditions, then the formulation should be tested at an intermediate condition at 30C2C /75%5% RH. Table 6.7 shows different temperatures and period of stability testing. Table 5.11 ICH guidelines for stability study Minimum time period Study Storage condition covered by data at submission 12 months 6 months 6 months

Long term Intermediate Accelerated

25C2C / 60%5% RH 30C2C / 65%5% RH 40C2C / 75%5% RH

The stability studies of the optimized tablets were carried out at 40C temperature and 75 % relative humidity (accelerated stability) in stability chamber for three months .Tablets were withdrawn at 1, 2, 3 months intervals and evaluated for disintegration time, hardness, drug content and in vitro release.

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