## STP FOR AMITRIPTYLINE TABLETS

1.0) Description: Visual

2.0)  Average Weight: Check weight of 20 tablets at randomly and calculate the average weight by formula :

wt of 20 tablets (gm) x 1000 / 20. found avg. wt in mg

3.0)  Uniformity of weight:

Weigh 20 tablets selected at random and calculate the average weight. Not more than two of the individual weights deviate from the average weight by more                                              than  the percentage shown in table.

Tablets were weighed individually and the percentage of deviation of its weight from the average weight was determined for each tablet.

Formula for calculate the percentage of deviation

= (experimental weight – theoretical weight)    x 100

Theoretical weight

 Average weight of tablets Percentage deviation More than 80mg but Less than 250mg 7.5% 250mg or More 5%

4.0)      Identification Test:

1. Disperse a quantity of the powdered tablets containing 5mg of Amitriptyline HCl with 20ml of methanol and filter. To 1ml of the filtrate, add 1ml of 0 2.5 per cent w/v solution of sodium bicarbonate, 1ml of a 2 per cent w/v solution of sodium periodate and 1ml of a 0.3 per cent w/v solution of potassium permanganate, allow to stand for 15 minutes, acidify with dilute sulphuric acid and extract wth10.0ml of 2,2,4-trimethylpentane. When examined in the range 230nm to 360nm, resulting solution shows an absorption maximum only at 265nm.
2. Triturate a quantity of the powdered tablets containing 0.1gm of amitriptylinre HCl with 10ml of chloroform, filter and evaporate the filtrate to a low volume. Add ether until a turbidity is produced and allow standing. To about 50mg of the precipitate dissolved in 3ml of water add 1 drop of a 2.5 per cent w/v solution of quinhydrone in methanol; no red colour is produced within 15 minutes (distinction from nortriptyline).
3. The precipetate obtained in test B gives reaction (A) of chlorides.

5.0)      Hardness:

The standard method used for tablet hardness testing is compression testing. The tablet is placed between two jaws that crush the tablet. The machine measures the force applied to the tablet and detects when it fractures. Although compressive force is applied to the tablet, along the diameter of the tablet at right angles to the applied force.

6.0)      Disintegration Time:

Unless otherwise stated in the individual monograph, introduce one tablet into each tube and add a disc to each tube. The assembly is suspended in the liquid medium in a suitable vessel, preferably a 1-litre beaker. The volume of liquid is such that the wire mesh at its highest point is at least 15mm below the surface of the liquid, and at its lower point is at least 25mm above the bottom of the beaker. At no time should the top of the basket-rack assembly become submerged. There is a thermostatic arrangement for heating the liquid and maintaining the temperature at 37±2°.

If 1 or 2 tablets fail to disintegrate, repeat the test on 12 additional tablets; not less than 16 of the total of 18 tablets tested disintegrate.

If the tablets adhere to the disc and the preparation under examination fails to comply, repeat the test omitting the disc. The preparation complies with the test if all the tablets in the repeat test disintegrate.

7.0)      Dimension of Tablet:

Length, breadth and thickness are determined by vernier in mm

8.0)      Dissolution:

Medium: 900ml of 0.1 M Hydrochloric acid

Time: 45 minutes

Speed: 100 rpm

Withdraw a suitable volume of the medium and filter. Measure the absorbance of the filtered solution, suitably diluted with the medium if necessary, at the                  maximum at about 239nm. Calculate the content of Amitriptyline in the medium from the absorbance obtained from a solution of known concentration of                     amitriptyline HCl reference standard in the same medium.

Limit: NLT 80% of the stated amount of Amitriptyline HCl.

9.0)      Leak test:

The apparatus is used to test for the integrity of packed strips, blisters and Alu-Alu Blister pack containing tablets. Ensure apparatus bath is filled with purified water upto mark indicated and add 0.5% crystal violet solution in water. Samples are placed into the desiccators and the lid is placed in position. The pump starts to produce a vacuum 15inHg inside the desiccators and the vacuum is held for 1 minute. The sample remains at the required vacuum level for given time interval buzzer will sound after time is over and will cut off the vacuum pump. As the package is immersed in a colored dye solution the venting of the desiccators will allow any holes to be penetrated by the dye and the contents of the flexible packaging will also be stained with the same coloring material.

Examine all the strips for any leakage by opening the pockets manually. If anyone pocket shows evidence of leakage, reject the sample, stop the Blister / Strip machine and immediately take corrective action.

10.0)    Related substance:

Determined by thin layer chromatography, protected from light, coating the plate with silica gel G.

Mobile phase: A mixture of 85 volumes of cyclohexane, 15 volumes of ethyl acetate and 3 volumes of diethylamine.

Test solution: Extract a quantity of the powdered tablets containing 20mg of Amitriptyline HCl with 5ml of a mixture of 9 volumes of ethanol (95 per cent) and 1 volumes of 2 M HCl centrifuge and use the supernatant liquid, evaporated to drness and dissolve in 10ml of chloroform.

Reference solution (a): A 0.001 per cent w/v solution of dibenzosuberone reference standard in chloroform.

Reference solution (b): A 0.004 per cent w/v solution of cyclobenzaprine HCl referenec in chloroform.

Apply to the plate 10µl of each solution. Allow the mobile phase to rise 14cm in an unlined tank. Dry the plate in air until the odour of the solvent is no longer detectable, spray with a freshly prepared mixture of 4 volumes of formaldehyde solution and 96 volumes of sulphuric acid, heat at 105° for 10 minutes and examine under ultraviolet light at 365nm. Any spots in the chromatogram obtained with the test solution corresponding to dibenzosuberone and cyclobenzaprine HCl are not more intense then the spots in the chromatograms obtained with reference solutions (a) and (b) respectively and any other secondary spot is not more intense than the spot in the chromatogram obtained with reference solution (b).

11.0)    Uniformity of content:

(For tablets containing 10mg or less)

Determine by liquid chromatography

Test solution: Powder one tablet, shake with 2.5ml of 0.1 M HCl until completely disintegrated, add 5ml of methanol, shake for 30 minutes, dilute the suspension to 10ml with methanol, centrifuge and use the clear supernatant liquid.

Reference solution: Dissolve 25.0mg of amitriptyline HCl reference standard in 10ml of methanol and dilute to 25.0ml with methanol (50 per cent).

Chromatographic system:

• A stainless steel column 20cmX4.6mm packed with octadecylsilane bonded to porous silica or ceramic microparticles (10µm),
• Mobile phase: 0.03 M sodium hexanesulphonate in a mixture of equal volumes of acetonitrile and water, adjusted to pH 4.5 with G.A.A.
• Flow rate: 2ml per minute,
• Spectrophotometer set at 239nm,
• Injection volume: 20µl

Inject the reference solution and the test solution.

Calculate the content of amitriptyline HCl in tablet.

12.0)    Assay:

Determined by liquid chromatography.

Test solution: When tablets are film-coated, shake 20 tablets with 50ml of 0.1 M HCl until completely disintegrated, add 100ml of methanol, shake for 30 minutes, dilute the suspension to 200.0ml with methanol, shake for 30 minutes, dilute the suspension to 200.0ml with methanol, centrifuge and dilute a volume of the supernatant liquid equivalent to 25mg of Amitriptyline HCl to 100.0ml with methanol (50 per cent).

When tablets are sugar-coated, weigh and powder 20 tablets. Weigh a quantity of the powder containing 50mg of Amitriptyline HCl, shake with 50ml of 0.1 M Hcl for 30 minutes, add 100ml of methanol, shake for 30 minutes, dilute the mixture to 200.0ml with water, centrifuge and use the supernatant liquid.

Reference solution: Dissolve 50mg of amitriptyline HCl reference standard in 10ml of methanol and dilute to 200.0ml with methanol (50 per cent).

Chromatographic system:

• A stainless steel column 20cmX4.6mm packed with octadecylsilane bonded to porous silica or ceramic microparticles (10µm),
• Mobile phase: 0.03 M sodium hexanesulphonate in a mixture of equal volumes of acetonitrile and water, adjusted to pH 4.5 with G.A.A.
• Flow rate: 2ml per minute,
• Spectrophotometer set at 239nm,
• Injection volume: 20µl

Alternative Method: By UV Spectrophotometer

Test Solution: Weigh and powder 20 tablets. Weight sample equivalent to 50mg of Amitriptyline HCl 100ml volumetric flask add 20ml 0.1M HCl into it and sonicate to dissolve the test sample. Make up to mark with 0.1 M HCl. Filter and dilute 1ml of this solution to 50ml in 0.1 M HCl.

Reference Solution: Weigh 50mg of reference standard of Amitriptyline HCl in 100ml of volumetric flask. Add 20ml 0.1 M HCl into it and sonicate to dissolve. Make up to mark with 0.1 M HCl. Dilute 1ml of this solution to 50ml with same solvent.

Check Absorbance of test as well as reference standard at about 239nm in UV Spectrophotometer.

Calculate the content of Amitriptyline HCl in the tablets.

Formula:

Area of Test      STD Wt.(mg)      1            100                 50           Potency

—————–X————–X——-X—————X——–X————–X Average wt.

Area of STD        100                  50      Test Wt.(mg)       1                100

Acceptance criteria: 90.0%-110.0%

12.0)    MICROBIOLOGICAL PURITY:

Perform the test according to requirements of IP,

Total aerobic Microbial count (TAMC): NMT 103 CFU/g

Total combined yeasts/Moulds count (TYMC): NMT 102 CFU/g

Pathogens: in 1gm drug.

Escherichia Coli – Should be absent

Pseudomonas aeroginosa – Should be absent

Salmonella – Should be absent

Staphylococcus aureus– Should be absent

Abbreviations:

Wt.: Weight

mg: Miligram

ml: Milileter

STD: Standard

inHg: Inch of Mercury

rpm: Rounds per minute

CFU: Colony forming unit

G.A.A: Glacial acetic acid

HCl: Hydrochloric acid