Raw Material Test Method - Ambroxol HCl

Title: Raw Material Test Method

Material : AMBROXOL HYDROCHLORIDE BP

Generic Name                        :           AMBROXOL HYDROCHLORIDE

Empirical Formula               :           C13H19Br2ClN2O

Molecular weight      :           414.6

CAS No.                     :           [23828-92-4] 

01.              Appearance:

Reference: BP

Acceptance criteria: White or yellowish crystalline powder.

Procedure:

Spread approximately 3.0 g of sample over a white paper and examine visually. Check the appearance for color, nature and any visible foreign particles. Record its appearance, extraneous matter, if any. The sample should be white or yellowish crystalline powder.

02.              Solubility:

Reference: BP

Acceptance criteria: sparingly soluble in water, soluble in methanol, practically insoluble in methylene chloride.

Procedure:

Sparingly soluble in water:

Take about 0.1 gm of the sample in a clean and dry test tube. To each of these add 10 ml of water and mix. The solution should be clear without any suspended particles.

Soluble in methanol:

Take about 1.0 gm of the sample in a clean and dry test tube. To each of these add 30 ml of methanol and mix. The solution should be clear without any suspended particles.

Practically insoluble in methylene chloride.:

Take about 10 mg of the sample in a 200 ml dry and clean volumetric flask. To it add 100 ml of methylene chloride and shake. Gradually add methylene chloride to it and shake. The dispersion should not be clear (suspended particles will be observed).

03.              Identification:

A.    UV-Vis Spectrum:

Reference: BP

Acceptance criteria: The solution shows two absorption maxima at 245 nm and 310 nm. The ratio of the absorbance measured at 245 nm to that measured at 310 nm is 3.2 to 3.4

Procedure:

Dissolve 20.0 mg in 0.05 M sulphuric acid and dilute to 100.0 ml with the same acid. Dilute 2.0 ml of the solution to 10.0 ml with 0.05 M sulphuric acid. Examined between 200 nm and 350 nm.

Spectral range. 200-350 run.

Absorption maxima At 245 run and 310 run.

Absorbance ratio. A245/A310 = 3.2 to 3.4.

B.     IR Spectrum:

Reference: BP

Acceptance criteria: The IR spectrum of the sample must be concordant with that of the standard.

Instrument:

Shimadzu

Acceptance criteria:

IR Spectrum of the test sample must be concordant with that of Ambroxol hydrochloride RS/Working standard. The correlation should be at least 0.950.

Procedure:

Take about 1 mg of Ambroxol hydrochloride (sample) and 300 mg of previously dried KBr. Mix uniformly by mortar & pestle. Transfer the mixture into a disc making die and spread the powder uniformly with help of punch. Transfer the die set to the disc making press and compress to form a thin disc. Remove the disc from the die set. Transfer to sample holder and keep the sample holder in the instrument sample compartment.

Scan the sample over the range from 4000 cm-1 to 400 cm-1. Make sure that the transmittance of the standard and sample spectrum at the wavelength of 2000 cm-1 is on or above the 60 % mark.

IR Spectrum of the test sample shall exhibit maximum only at the same wave number as that of Ambroxol hydrochloride working standard.

             C.         By TLC:

Reference: BP

Acceptance criteria:

             The principal spot in the chromatogram obtained with the test solution is similar in position and size to the principal spot in the chromatogram obtained with the reference solution.

Procedure:

Test solution:

Dissolve 50 mg of the substance to be examined in methanol R and dilute to 5 ml with the same solvent.

Reference solution.

Dissolve 50 mg of ambroxol hydrochloride CRS in methanol R and dilute to 5 ml with the same solvent.

Plate: TLC silica gel F254 plate R.     

Mobile phase: concentrated ammonia R, 1-propanol R,ethyl acetate R, hexane R (1:10:20:70 V/V/V/V).

Application: 10 µl.

Development: over 2/3 of the plate.

Drying: in air.

Detection: examine in ultraviolet light at 254 nm

D.Chloride Test:

Reference: BP

Acceptance criteria:

With silver nitrate TS, solutions of chlorides yield a white, curdy precipitate that is insoluble in nitric acid but is soluble in a slight excess of 6 N ammonium hydroxide

Preparation of Silver Nitrate TS:

Dissolve about 1.75 g of silver nitrate in 100 mL of water.

Preparation of Silver Nitrate TS:

Dilute 35 mL of Ammonia Water, Stronger with water to make 100 ml.

Procedure:

Dissolve 25 mg in 2.5 ml of water R, mix with 1.0 ml of dilute ammonia R1 and allow to stand for 5 min. Filter and acidify the filtrate with dilute nitric acid R. Add one drop of diluted nitric acid and 0.5 mL of silver nitrate TS to the filtrate: a white, curdy precipitate is formed. Centrifuge the mixture without delay and decant the supernatant layer. Wash the precipitate with three 1-mL portions of nitric acid solution (1 in 100), and discard the washings.

Add ammonia TS dropwise to this precipitate. It dissolves readily. When a monograph specifies that an article responds to the test for dry chlorides, mix the solid to be tested with an equal weight of manganese dioxide, moisten with sulfuric acid, and gently heat the mixture: chlorine, which is recognizable by the production of a blue color with moistened starch iodide paper, is evolved.

Solution S. Dissolve 0.75 g in methanol R and dilute to 15 ml with the same solvent.

04.              Appearance of solution:

Solution S is clear (2.2.1) and not more intensely coloured than reference solution Y6 (2.2.2, Method II).

05.  pH (2.2.3): 4.5 to 6.0.

Dissolve 0.2 g in carbon dioxide-free water R and dilute to 20 ml with the same solvent.

06.  Related substances. Liquid chromatography (2.2.29).

Prepare the solutions immediately before use.

Test solution. Dissolve 50.0 mg of the substance to be examined in water R and dilute to 50.0 ml with the same solvent.

Reference solution (a). Dilute 1.0 ml of the test solution to 100.0 ml with water R. Dilute 1.0 ml of this solution to 10.0 ml with the mobile phase.

Reference solution (b). Dissolve 5 mg of the substance to be examined in 0.2 ml of methanol R and add 0.04 ml of a mixture of 1 volume of formaldehyde solution R and 99 volumes of water R. Heat at 60 °C for 5 min. Evaporate to dryness under a current of nitrogen. Dissolve the residue in 5 ml of water R and dilute to 20 ml with the mobile phase.

Column: — size: l = 0.25 m, Ø = 4.0 mm,        

Stationary phase: octadecylsilyl silica gel for chromatography R (5 µm).

Mobile phase: a mixture of equal volumes of acetonitrile R and a solution prepared as follows: dissolve 1.32 g of ammonium phosphate R in 900 ml of water R, adjust to pH 7.0 with phosphoric acid R and dilute to 1000 ml with water R.

Flow rate: 1 ml/min.

Detection: spectrophotometer at 248 nm.

Injection: 20 µl.

Sensitivity: reference solution (a).

Run time: 3 times the retention time of Ambroxol.

Identification of impurities: Use the chromatogram obtained with referance solution (b). To identify the peak due to impurity B

Relative retention: With reference to ambroxol ( retention time = about 9 min): impurity B = about 6.

System suitability: reference solution (b).

 — resolution: minimum of 4.0 between the peaks due to impurity B and ambroxol

Limits:

Unspecified impurities: For each impurity not more than the area of the principal peak in the chromatogram obtained with reference solution (a) (0.10 per cent),

Total: not more than 3 times the area of the principal peak in the chromatogram obtained with reference solution (a) (0.3 per cent),

Disregard limit: 0.1 times the area of the principal peak in the chromatogram obtained with reference solution (a). (0.05 per cent)

07.  Loss on drying (2.2.32) :

Procedure:

Dry the weighing bottle at 105°C until constant weight and weigh (W1) in gm. Weigh accurate 1.0 gm of sample (W2) in the weighing bottle. Dry the sample at 105°C for 1 hour. Cool the sample in desiccator and weigh (W3) in gm. Calculate the loss on drying (LOD) in percentage by following formula.

LOD (% w/w) =  

Acceptance criteria: Maximum 0.5%.

08.  Sulphated ash (2.4.14):

Procedure:

Ignite a silica crucible 600 ± 50° for 30 minutes; allow cooling in a desiccator over silica gel and weigh (W1) in gm. Weigh 1 gm of sample (W2) and transfer into the crucible. Moisten the test sample with a small amount of sulphuric acid (usually 1 ml) and heat gently at as low a temperature as practicable until the sample is thoroughly charred. After cooling, moisten the residue with a small amount of sulphuric acid, heat gently until white fumes are no longer evolved and ignite at 600° ± 50° until the residue is completely incinerated. Ensure that flames are not produced at any time during the procedure. Allow the crucible to cool in a desiccator over silica gel, weigh it again and weigh (W3) of the residue.

If the weight of residue so obtained exceeds the prescribed limit, repeat the moistening with sulphuric acid and ignition, as previously, for 30 min periods until 2 consecutive weighings do not differ by more than 0.5 mg or until the percentage of residue complies with the prescribed limit.

Calculate the ash in percentage by following formula:

Sulfated ash (% w/w) =    

 Where,

W1      =          Weight of empty ignited crucible in gm.

W2      =          Weight of test sample before ignition in gm.

W3      =          Weight of crucible after ignition with the residue sample in gm

Acceptance criteria: Maximum 0.1%.

09. ASSAY

Dissolve 0.300 g in 70 ml of ethanol (96 per cent) R and add 5 ml of 0.01 M hydrochloric acid. Carry out a potentiometric titration (2.2.20), using 0.1 M sodium hydroxide. Read the volume added between the two points of inflexion.

1 ml of 0.1 M sodium hydroxide is equivalent to 41.46 mg of C13H19Br2ClN2O.

Or Alternative UV Method:

Sample solution:

Weigh accurately about 50 mg of test sample and transfer it into a 100 ml volumetric flask and dissolve in 10 ml of 0.1ml HCl. Dilute it with 0.1ml HCl upto the mark and mix well. Transfer 2 ml of this solution to a 100 ml volumetric flask and dilute it with 0.1ml HCl upto the mark, mix well.

Standard solution:

Weigh accurately about 50 mg of CRS Ambroxol hydrochloride and transfer it into a 100 ml volumetric flask and dissolve in 10 ml of 0.1ml HCl. Dilute it with 0.1ml HCl upto the mark and mix well. Transfer 2 ml of this solution to a 100 ml volumetric flask and dilute it with 0.1ml HCl upto the mark, mix well.

Measurement:

Determine the absorbance of sample and standard solution in 1-cm cell at 245 nm using 0.1N Hydrochloric acid as blank.

Calculate the quantity of Ambroxol hydrochloride in percentage in the sample taken by the formula:

Content Ambroxol hydrochloride of (% w/w as it is) =

Where,

A sam               =          Absorbance from sample solution

A std                 =          Absorbance from standard solution

Wstd                 =          Weight of Ambroxol Hcl Working standard taken to prepare the standard solution                                  

Wsam                     =          Weight of test sample taken to prepare the sample solution

Pstd                        =          Potency of CRS/Working standard in percentage

Content of Ambroxol hydrochloride (% w/w on dried basis)     =

 A = Assay (% w/w as it is)

Acceptance Criteria: 99.0% to 101% (On Dried Basis)