Timolol Maleate

Edition: BP 2025 (Ph. Eur. 11.6 update)

(Ph. Eur. monograph 0572)

Timolol Maleate

C₁₇H₂₈N₄O₇S 432.5 26921-17-5

Action and use

Beta-adrenoceptor antagonist.

For Timolol Eye Drops only: treatment of glaucoma.

Preparations

Dorzolamide and Timolol Eye Drops

Timolol Eye Drops

Timolol Tablets

Ph Eur

DEFINITION

(2S)-1-[(1,1-Dimethylethyl)amino]-3-[[4-(morpholin-4-yl)-1,2,5-thiadiazol-3-yl]oxy]propan-2-ol (Z)-butenedioate.

Content

98.5 per cent to 101.0 per cent (dried substance).

CHARACTERS

Appearance

White or almost white, crystalline powder or colourless crystals.

Solubility

Soluble in water and in ethanol (96 per cent).

mp

About 199 °C, with decomposition.

IDENTIFICATION

First identification: A, B.

Second identification: A, C, D.

A. Specific optical rotation (2.2.7): -6.2 to -5.7.

Dissolve 1.000 g in 1 M hydrochloric acid and dilute to 10.0 mL with the same acid.

B. Infrared absorption spectrophotometry (2.2.24).

Comparison timolol maleate CRS.

C. Thin-layer chromatography (2.2.27).

Test solution Dissolve 5 mg of the substance to be examined in methanol R and dilute to 5 mL with the same solvent.

Reference solution Dissolve 5 mg of timolol maleate CRS in methanol R and dilute to 5 mL with the same solvent.

Plate TLC silica gel GF254 plate R.

Mobile phase concentrated ammonia R, methanol R, methylene chloride R (1:20:80 V/V/V).

Application 10 μL.

Development Over 2/3 of the plate.

Drying In air.

Detection Expose to iodine vapour for 2 h.

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

D. Triturate 0.1 g with a mixture of 1 mL of dilute sodium hydroxide solution R and 3 mL of water R. Shake with 3 quantities, each of 5 mL, of ether R. To 0.1 mL of the aqueous layer add a solution containing 10 mg of resorcinol R in 3 mL of sulfuric acid R. Heat on a water-bath for 15 min; no violet-red colour develops. Neutralise the remainder of the aqueous layer with dilute sulfuric acid R and add 1 mL of bromine water R. Heat on a water-bath for 15 min, then heat to boiling and cool. To 0.2 mL of this solution add a solution containing 10 mg of resorcinol R in 3 mL of sulfuric acid R. Heat on a water-bath for 15 min; a violet-red colour develops. Add 0.2 mL of a 100 g/L solution of potassium bromide R and heat for 5 min on a water-bath; the colour becomes violet-blue.

TESTS

Solution S

Dissolve 0.5 g in carbon dioxide-free water R and dilute to 25 mL with the same solvent.

Appearance of solution

Solution S is clear (2.2.1) and not more intensely coloured than reference solution B8 (2.2.2, Method II).

pH (2.2.3)

3.8 to 4.3 for solution S.

Enantiomeric purity

Liquid chromatography (2.2.29). Carry out the test protected from actinic light.

Solvent mixture methylene chloride R, 2-propanol R (10:30 V/V).

Test solution Dissolve 30.0 mg of the substance to be examined in the solvent mixture and dilute to 10.0 mL with the solvent mixture.

Reference solution (a) Dissolve 30 mg of timolol maleate CRS in the solvent mixture and dilute to 10 mL with the solvent mixture.

Reference solution (b) Dissolve 3 mg of (R)-timolol CRS (impurity A) in the solvent mixture and dilute to 10.0 mL with the solvent mixture. Dilute 1.0 mL of the solution to 10.0 mL with the solvent mixture.

Reference solution (c) Dilute 1 mL of reference solution (a) to 100 mL with the solvent mixture. Mix 1 mL of this solution with 1 mL of reference solution (b).

Reference solution (d) Dilute 1.0 mL of the test solution to 100.0 mL with the solvent mixture.

Column:

– size: l = 0.25 m, Ø = 4.6 mm;

– stationary phase: cellulose derivative of silica gel for chiral separation R (5 μm).

Mobile phase diethylamine R, 2-propanol R, hexane R (2:40:960 V/V/V).

Flow rate 1 mL/min.

Detection Spectrophotometer at 297 nm.

Injection 5 μL.

Elution order Impurity A is eluted first.

System suitability:

– resolution: minimum 4.0 between the peaks due to impurity A and the (S)-enantiomer in the chromatogram obtained with reference solution (c);

– the retention times of the principal peaks due to the (S)-enantiomer in the chromatograms obtained with the test solution and reference solution (a) are identical.

Limit:

– impurity A: not more than the area of the principal peak in the chromatogram obtained with reference solution (d) (1.0 per cent).

Liquid chromatography (2.2.29).

Test solution Dissolve 0.100 g of the substance to be examined in mobile phase A and dilute to 20 mL with mobile phase A.

Reference solution (a) Dilute 1.0 mL of the test solution to 100.0 mL with mobile phase A. Dilute 1.0 mL of this solution to 10.0 mL with mobile phase A.

Reference solution (b) Dissolve 3 mg of timolol impurity F CRS in methanol R and dilute to 20.0 mL with the same solvent. Dilute 1.0 mL of the solution to 10.0 mL with mobile phase A (solution A). Dissolve the contents of a vial of timolol for system suitability CRS (containing impurities B, C and D) in 1.0 mL of solution A.

Reference solution (c) Dissolve 2 mg of the substance to be examined and 20 mg of maleic acid R in 10 mL of acetonitrile R. Evaporate 1 mL of the solution to dryness under a stream of nitrogen R in an amber glass vial. Heat the open vial at 105 °C for 1 h. Reconstitute the residue with 1.0 mL of mobile phase A (in situ preparation of impurity E).

Column:

– size: l = 0.150 m, Ø = 3.9 mm;

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

Mobile phase:

– mobile phase A: mixture of equal volumes of methanol R and a 4.32 g/L solution of sodium octanesulfonate R previously adjusted to pH 3.0 with glacial acetic acid R;

– mobile phase B: methanol R;

Time (min)	Mobile phase A (per cent V/V)	Mobile phase B (per cent V/V)
0 – 10	97.5	2.5
10 – 11	97.5 → 70	2.5 → 30
11 – 20	70	30

Flow rate 1.2 mL/min.

Detection Spectrophotometer at 295 nm.

Injection 20 μL.

Identification of impurities Use the chromatogram supplied with timolol for system suitability CRS and the chromatogram obtained with reference solution (b) to identify the peaks due to impurities B, C, D and F; use the chromatogram obtained with reference solution (c) to identify the peak due to impurity E.

Relative retention With reference to timolol (retention time = about 7.5 min): maleic acid = about 0.1;

impurity D = about 0.3; impurity E = about 0.4; impurity B = about 0.7; impurity F = about 0.8; impurity C = about 2.1.

System suitability Reference solution (b):

– resolution: minimum 1.5 between the peaks due to impurities B and F.

Limits:

– correction factor: for the calculation of content, multiply the peak area of impurity D by 0.6;

– impurities B, C, D, E, F: for each impurity, not more than twice the area of the principal peak in the chromatogram obtained with reference solution (a) (0.2 per cent);

– 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 4 times the area of the principal peak in the chromatogram obtained with reference solution (a) (0.4 per cent);

– disregard limit: 0.5 times the area of the principal peak in the chromatogram obtained with reference solution (a) (0.05 per cent); disregard any peak due to maleic acid.

Loss on drying (2.2.32)

Maximum 0.5 per cent, determined on 1.000 g by drying in an oven at 105 °C.

Sulfated ash (2.4.14)

Maximum 0.1 per cent, determined on 1.0 g.

ASSAY

Dissolve 0.350 g in 60 mL of anhydrous acetic acid R. Titrate with 0.1 M perchloric acid, determining the end-point potentiometrically (2.2.20).

1 mL of 0.1 M perchloric acid is equivalent to 43.25 mg of C₁₇H₂₈N₄O₇S.

STORAGE

Protected from light.

IMPURITIES

Specified impurities A, B, C, D, E, F.

Other detectable impurities (the following substances would, if present at a sufficient level, be detected by one or other of the tests in the monograph. They are limited by the general acceptance criterion for other/unspecified impurities and/or by the general monograph Substances for pharmaceutical use (2034). It is therefore not necessary to identify these impurities for demonstration of compliance. See also 5.10. Control of impurities in substances for pharmaceutical use) G, H, I, J.

Timolol Maleate