INTRODUCTION:

The principal objective of any pharmaceutical plant is to manufacture products of requisite attribute and quality consistently, at the lowest possible cost.¹ Although validation studies have been conducted in the pharmaceutical industry for a long time, there is an ever increasing interest in validation owing to their industry’s greater emphasis in recent years on quality assurance program and is fundamental to an efficient production operation. The concept of validation has expanded through the years to embrace a wide range of activities from analytical methods used for the quality control of drug substances and drug products to computerized systems for clinical trials, labeling or process control, Validation is founded on, but not prescribed by regulatory requirements and is best viewed as an important and integral part of cGMP.^2,3

The validation concept is firstly evolved in USA in 1978. Validation is documented evidence which gives the high degree of assurance, that a specific process will consistently produce a product meeting its predetermined specifications and quality attributes. The word validation simply means assessment of validity or action of proving effectiveness. Validation is a team effort where it involves people from various disciplines of the plant. To enhance the effectiveness and safety of the drug product after approval, many regulatory agencies such as the United States Food and Drug Administration (FDA) also require that the drug product be tested for its identity, strength, quality, purity and stability before it can be released for use. For this reason, pharmaceutical validation and process controls are important in spite of the problems that may be encountered. Validation mainly based on, FDA regulations describing current good manufacturing practice (cGMP) for finished pharmaceuticals are provided in 21 CF parts 210 and 211. The cGMP regulations require that manufacturing processes be designed and controlled to assure that in process materials and the finished product meet predetermined quality requirements and do so consistently and reliably. Validation is therefore one element of quality assurance associated with a particular process, as the process differs so widely, there is no universal approach to validation and regulatory bodies such as FDA and EC who have developed general non-mandatory guide lines. Then word validation simply means, ‘assessment of validity’ or action of proving effectiveness. According to European community for medicinal products, validation is ‘action of proving’, in accordance with the principles of GMP that any procedures, process, requirement, material, activity or system actually leads to expected results.^4,5

MATERIALS AND METHOD:

Materials:

Isoniazid USP/BP/Ph. Eur Active Ingredient Amsal chem., Microcrystalline cellulose NF Diluent RANQ Remedies, Pregelatinized Starch NF Binder Colorcon, Colloidal Silicon Dioxide NF Glident Wacker chemie, colour Ponceau 4R Supra LD Binder Prepartion (colourant) Roha Dye chem., Saccharin Sodium USP Binder Prepartion (Sweetener) Suzhou fine, Purified Water IP/BP/USP/LD Binder Prepartion (solvent) Lupin Ltd. Aurangabad, Rifampin Part USP Active Ingredient Lupin Tarapur, Microcrystalline Cellulose NF Diluent RANQ Remedies, Crospovidone NF Disintegrant ISP Chemical, Colloidal Silicon Dioxide NF Binder Wacker chemie, Pregelatinized Starch NF Disintegrant And Binder Colorcon, Ascorbic Acid USP Preservative DSM, Purified Water IP/BP/USP Solvent Lupin Ltd., Analysis report of all the raw material was checked and only approved raw material was used.

Methods:^6-12

Average Weight:

Select 20 Tablets randomly from the pooled sample. Weigh 20 tablets individually and calculate the average weight. Report the value in mg.

Average weight = Total Weight of 20 tablets/ 20

Hardness:

Select 10 tablets from collective sample and measured the hardness of tablet using calibrated hardness tester like Monsanto hardness tester.

Friability:

The friability of tablet was determined using Roche Friabilator. Preweighed Rifampin and Isoniazid tablets were transferred into friabilator. The friabilator was operated at 25rpm for 4 minutes i.e. 100 revolutions. Tablets are dropped from a distance of 6 inches with each revolution. The tablets were deducted and weighed again. The percentage friability was calculated by,

F = W initial-W final/W final *100

Thickness:

Tablet thickness was measure by calibrated Vernier calliper scale and is measure in mm.

Weight Variation:

Twenty tablets was randomly selected from each formulation and weighed individually to check for weight variation. The following percentage deviation in weight variation according to IP was allowed.

Content Uniformity:

20 tablets were crushed and powder equivalent to 50mg of Rifampin and Isoniazid and dissolve in 100ml of water stirred and filtered. 1ml of the solution was diluted to 10ml with water and the absorbance was taken at 475 nm with the help of HPLC. The amount of drug present in each formulation was determined by HPLC.

In Vitro Disintegration Test:

Disintegration test was performed on 6 tablets. Place one tablet in each of the six tubes of the basket, add disc to each tube and operate the apparatus using water at 37 ± 2⁰C as the immersion fluid. At the end of 30 minutes, lift the basket from the fluid and observed the tablets. All the tablets should disintegrate. Record the disintegration time in minute and seconds. If 1 or 2 tablets are fail to disintegrate, repeat the test on 12 additional tablets not less than 16 of the total of 18 tablets should disintegrate.

In Vitro Dissolution Study:

In vitro dissolution study was performed in 900ml 0.1 N HCl as medium using USP Type I (Basket) apparatus at 100rpm for 30 minutes (37±0.5⁰C). Aliquots of the dissolution medium (40ml) were withdrawn at specific time intervals and replaced immediately with equal volume of fresh medium. The samples were filtered through Distek, 0.45μ cellulose filter paper and analyzed for drug content by measuring the absorbance at 475nm.

Dissolution Medium Preparation:

8.5ml of HCl added in the 1000ml of water.

Standard preparation: (For INH)

Weigh accurately about 66mg of Isoniazid working standard in to 100ml volumetric flask and dissolve in 50 ml of water and make up to the volume with water.

Standard preparation: (For Rifa.)

Weigh accurately about 33mg of Rifampin working standard in to 200ml volumetric flask and dissolve in 25 ml of solution and make up to the volume with water.

0.05 M Potassium Dihydrogen Orthophosphate (buffer):

Accurately weighed and dissolved about 6.8gm of Potassium Dihydrogen Orthophosphate in 1000ml of water.

Mobile Phase Preparation:

Separately measured 990ml of 0.05M Potassium Dihydrogen Orthophosphate buffer in 10ml of acetonitrile, checked the pH and adjusted to 4.0 ± 0.05 ml with 2% orthophosphoric acid. The Buffer filtered through 0.45μ filter paper and degassed accordingly.

Chromatographic Conditions:

Column: μ Bondapack C18, 300 × 3.9mm, 10μ or equivalent

Flow rate: 1.0ml/min

Wavelength: 254nm

Injection Volume: 20μl

Run time: 15 min

Procedure:

Filter the standard and test preparation through 0.45μ membrane filter. Injected standard in 5 replicate injections and test solution in single and record chromatograms. Calculate the system suitability by using formula.

% Rifampin and Isoniazid Dissolve

AT × WS × 10 × 900 × 25 × P × 100

% (Dissolution) = ----------------------------------------------

AS × 200 × 25 × 1 × 10 × 100 × 150

Where,

AT = Absorbance of Rifampin and Isoniazid in test solution

AS = Absorbance of Rifampin and Isoniazid in standard solution

WS = weight of standard (mg)

P = Potency of standard for Rifampin and Isoniazid

100 = Percentage factor

Drug concentration was calculated, and % release was calculated.

Assay:

Buffer Preparation (pH-6.8):

Weighed 2.8gm of anhydrous disodium hydrogen orthophosphate in 2000ml of purified water, checked pH and adjusted by Orthophosphoric acid.

Mobile phase preparation:

Mobile Phase A:

Separately measured 960ml of above buffer in 40ml acetonitrile, mixed, filtered through 0.45μ filter paper and degas suitably.

Mobile Phase B:

Separately measured 450 ml of above buffer in 550 ml acetonitrile, mixed, filtered through 0.45 μ filter paper and degas suitably.

Diluent:

accurately weighed and dissolved 2.8gm of anhydrous disodium hydrogen orthophosphate in 2000ml of purified water, checked Initial pH and adjusted to 6.8 ± 0.05 with Orthophosphoric acid.

Standard preparation:

Weighed accurately 80mg Rifampin WS and 40mg Isoniazid WS in individual 500ml volumetric flask, dissolved in 100ml methanol, sonicated 1 min and added about 375ml individually Diluent, further sonicate 10 min, makeup the final volume.

Test Preparation:

Finely powder not less than 20 Tab. and weigh 189gm of powder in 500ml volumetric flask. Added about 100ml of methanol to dissolve Rifampin, sonicate 1 min, add 375ml of diluent and sonicate 10 min and make up the final volume.

Procedure:

Filter standard and test preparation through 0.45μ Membrane filter. Inject volume in 5 replicate injections and test solution in duplicate and record chromatogram. Calculate the system suitability by using formula.

Formula for calculation of assay of Rifampin in tablet:

AT × WS × 500 × W × P × 100

% Rifampin= -----------------------------------------------

AS × 500 × WT × 75 × 100

Formula for calculation of assay of Isoniazid in tablet:

AT × WS × 500 × W × P × 100

% Isoniazide= -----------------------------------------------

AS × 500 × WT × 75 × 100

Where,

AT = Avg. area of test solution of Rifampin and Isoniazid peaks respectively.

AS = Avg. area of standard solution for Rifampin and Isoniazid peaks respectively.

WS = Weight of standard (mg) Rifampin and Isoniazid peaks respectively.

P = Potency of standard for Rifampin and Isoniazid

100 = Percentage factor

Drug concentration was calculated, and % release was calculated.

Loss on Drying:

Took clean capillary stopperd bottle, dried at 600C for 30 minutes and cool in the desiccator.

1. Weight of dried empty bottle. (W1)

2. Weight of bottle with 100mg sample. (W2)

Dried the bottle and sample at 600C and at pressure of 5 mm for 3 hours and cool in the desiccator.

3. Weight bottle and 100 mg sample after drying. (W3)

Calculate the LOD by using following formula:

W2-W3

% LOD = -------------- × 100

W2-W1

RESULT AND DISCUSSION:

Blend uniformity after Dry Mixing Stage (08 Minutes):

Blend uniformity done for checking the flow property of powder blend. Also done for checking the uniform mixing of all excipient and lubricants. Samples collected from RMG from different locations and uniformity of blend checked. The BU of isoniazid at dry mixing stage for three process performance qualification batches was found in the range of 98.1-100.0%, 99.9-101.95% and 102.2-105.4% respectively and complies as per acceptance criteria. Mean of the Isoniazid at dry mixing stage for three process performance qualification batches was found 99.4%, 100.9% and 103.3% respectively.

The BU of Rifampin at dry mixing stage for three process performance qualification batches was found in the range of 96.0-102.1 %, 99.6-102.1% and 93.6-101.7% respectively and complies as per acceptance criteria. Mean of the Isoniazid at dry mixing stage for three process performance qualification batches was found 100.5%, 101.4% and 100.1% respectively.

Fig. No. 01: Blend Uniformity at Dry mixing stage (INH)

Drying:

% LOD of dried granules comprising of top, middle, bottom layers of FBD bowl of Isoniazid part, for three performance qualification batches was found in the range of 1.30- 1.73% w/w, 1.29-1.53% w/w and 1.27-1.49% w/w respectively and complies as per limit.

% LOD of dried granules comprising of top, middle, bottom layers of FBD bowl of Rifampin part, for three performance qualification batches was found in the range of 1.30-1.73% w/w, 1.29-1.53% w/w and 1.27-1.49% w/w respectively and complies as per

limit.

Fig No. 02: Blend Uniformity at Dry mixing stage (Rifa)

Fig. no. 03: Loss on Drying of Dried Granules (INH)

Fig. No. 04: Loss on Drying of Dried Granules (Rifa Part)

Lubrication:

Blend uniformity of Lubricated blend (5 minutes):

Blend uniformity done for checking the flow property of powder blend. Also done for checking the uniform mixing of all excipient and lubricants. Samples collected from octagonal blender from different locations and uniformity of blend checked.

The blend uniformity of INH at lubrication stage for three process performance qualification batches was found in the range of 96.2-103.7, 96.0-106.5, 96.6-100.2 % respectively and complies as per acceptance criteria.

Fig. no. 05: Blend uniformity of Lubricated blend (INH)

Dissolution of Compressed Tablets:

The rate of % dissolution rate of rifampin and isoniazid at initial, middle and end stage of compression of three consecutive batches were found within acceptance criteria.

Fig.no. 06: Dissolution of compressed tablets (INH Part)

Fig.no. 07: Dissolution of compressed tablets (Rifa Part)

Evaluation:

The three process performance qualification batches of Rifampin and Isoniazid tablets were fabricated using the raw materials procured from the approved vendors. These raw materials were used after approval of QC laboratory stating compliance to the respective raw material specifications in all aspects. Samples were withdrawn from blend uniformity analysis at dry mixing stage. The observed results of BU of Isoniazid for three batches and Rifampin were complying with specified acceptance criteria. Samples were withdrawn for BU and composite blend analysis at lubrication stage. The observed result of BU of Isoniazid and Rifampin for three batches was complying with specified acceptance criteria. Samples were withdrawn at lubrication stage for analysis of physical characteristics of blend. Observe results for information only. Samples were withdrawn from lubricated blend unloaded in IPC for BU of first batch. The observed BU of Rifampin and Isoniazid of blend unloaded in IPC for first batch were complying with specified acceptance criteria. The in process analytical results of three batches were found similar and there is no variability observed within the batch and also no variability observed within the manufactured process qualification batch. During tablet compression, in-process checks were concluded and all the in-process results of description, group weight, hardness, thickness, DT, friability were found to comply with the acceptance criteria. Samples were collected from 40 locations spread across entire batch run during compression for determining the uniformity of dosage units. The observed results uniformity of dosage units (Isoniazid and Rifampin) were found to comply as per acceptance criteria. Sample withdrawn from initial, middle and end of tablet compression operation for determining dissolution. The results of dissolution at initial, middle and end of compression were found within acceptance criteria. After coating samples were also collected for finished product analysis and found to comply in accordance with the acceptance criteria. The results of uniformity of dosage units and dissolution were found to comply with specific acceptance criteria.

CONCLUSION:

Rifampin 150 mg and Isoniazid 75mg tablets were manufactured with a set of operational parameters and manufacturing instructions and process validation study was performed as per USFDA guideline. The results of various stages of the process like dry mixing, wet granulation, drying, lubrication, compression and coating were found satisfactory. The results of finished product analysis were also found within acceptance criteria. Therefore, it can be concluded that process validation of solid oral dosage form of Rifampin 150 mg and Isoniazid 75mg tablets in the expansion area was done successfully.

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Received on 18.09.2020 Modified on 20.11.2020

Asian Journal of Pharmacy and Technology. 2021; 11(2):105-110.

DOI: 10.52711/2231-5713.2021.00017