Betamethasone

(Ph Eur monograph 0312)

C22H29FO5       392.5      378-44-9

 

Ph Eur


 

 

Definition

 

Betamethasone contains not less than 97.0 per cent and not more than the equivalent of 103.0 per cent of 9-fluoro-11b,17,21-trihydroxy-16b-methylpregna-1,4-diene-3,20-dione, calculated with reference to the dried substance.

 

Characters

 

A white or almost white, crystalline powder, practically insoluble in water, sparingly soluble in ethanol, very slightly soluble in methylene chloride.

 

Identification

 

First identification

 

B, C.

 

Second identification

 

A, C, D, E.

 

A.    Dissolve 10.0 mg in ethanol  R and dilute to 100.0 ml with the same solvent. Place 2.0 ml of the solution in a stoppered tube, add 10.0 ml of phenylhydrazine-sulphuric acid solution R, mix and heat in a water-bath at 60C for 20 min. Cool immediately. The absorbance (2.2.25) of the solution measured at 419 nm is not greater than 0.10.

 

B.    Examine by infrared absorption spectrophotometry (2.2.24), comparing with the spectrum obtained with betamethasone CRS. If the spectra obtained in the solid state with the substance to be examined and the reference substance show differences, dissolve the substance to be examined and the reference substance separately in the smallest necessary quantity of methylene chloride R and evaporate to dryness on a water-bath. Using the residues, record the spectra again.

 

C.    Examine by thin-layer chromatography (2.2.27), using as the coating substance a suitable silica gel with a fluorescent indicator having an optimal intensity at 254 nm.

 

Test solution

 

Dissolve 10 mg of the substance to be examined in a mixture of 1 volume of methanol  R and 9 volumes of methylene chloride R and dilute to 10 ml with the same mixture of solvents.

 

Reference solution (a)

 

Dissolve 20 mg of betamethasone CRS in a mixture of 1 volume of methanol  R and 9 volumes of methylene chloride R and dilute to 20 ml with the same mixture of solvents.

 

Reference solution (b)

 

Dissolve 10 mg of dexamethasone CRS in reference solution (a) and dilute to 10 ml with the same solution.

 

Apply separately to the plate 5 ml of each solution. Develop over a path of 15 cm using a mixture of 5 volumes of butanol  R saturated with water  R, 10 volumes of toluene R and 85 volumes of ether  R. Allow the plate to dry in air and examine in ultraviolet light at 254 nm. 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 reference solution (a). Spray with alcoholic solution of sulphuric acid R. Heat at 120C for 10 min or until the spots appear. Allow to cool. Examine the chromatograms in daylight and in ultraviolet light at 365 nm. The principal spot in the chromatogram obtained with the test solution is similar in position, colour in daylight, fluorescence in ultraviolet light at 365 nm and size to the principal spot in the chromatogram obtained with reference solution (a). The test is not valid unless the chromatogram obtained with reference solution (b) shows two spots which may however not be completely separated.

 

D.    Mix about 5 mg with 45 mg of heavy magnesium oxide R and ignite in a crucible until an almost white residue is obtained (usually less than 5 min). Allow to cool, add 1 ml of water  R, 0.05 ml of phenolphthalein solution R1 and about 1 ml of dilute hydrochloric acid R to render the solution colourless. Filter. Add 1.0 ml of the filtrate to a freshly prepared mixture of 0.1 ml of alizarin S solution R and 0.1 ml of zirconyl nitrate solution R. Mix, allow to stand for 5 min and compare the colour of the solution with that of a blank prepared in the same manner. The test solution is yellow and the blank is red.

 

E.    Add about 2 mg to 2 ml of sulphuric acid R and shake to dissolve. Within 5 min, a deep reddish-brown colour develops. Add the solution to 10 ml of water  R and mix. The colour is discharged and a clear  solution remains.

 

Tests

 

Specific optical rotation (2.2.7)

 

Dissolve 0.125 g in methanol  R and dilute to 25.0 ml with the same solvent. The specific optical rotation is +118 to +126, calculated with reference to the dried substance.

 

Related substances

 

Examine by liquid chromatography (2.2.29).

 

Test solution

 

Dissolve 25.0 mg of the substance to be examined in a mixture of equal volumes of acetonitrile R and methanol  R and dilute to 10.0 ml with the same solvent.

 

Reference solution (a)

 

Dissolve 2 mg of betamethasone CRS and 2 mg of methylprednisolone CRS in mobile phase A and dilute to 100.0 ml with the same mobile phase.

 

Reference solution (b)

 

Dilute 1.0 ml of the test solution to 100.0 ml with mobile phase A.

 

The chromatographic procedure may be carried out using:

 

a stainless steel column 0.25 m long and 4.6 mm in internal diameter packed with octadecylsilyl silica gel for chromatography R (5 mm),

 

as mobile phase at a flow rate of 2.5 ml/min, a linear-gradient programme using the following conditions:

 

Mobile phase A In a 1000 ml volumetric flask mix 250 ml of acetonitrile R with 700 ml of water  R and allow to equilibrate; adjust the volume to 1000 ml with water  R and mix again,

 

Mobile phase B Acetonitrile R,

 

as detector a spectrophotometer set at 254 nm,

 

maintaining the temperature of the column at 45C.

 

Equilibrate the column with mobile phase B at a flow rate of 2.5 ml/min for at least 30 min and then with mobile phase A for 5 min. For subsequent chromatograms, use the conditions described from 40 min to 46 min.

 

Adjust the sensitivity of the system so that the height of the principal peak in the chromatogram obtained with 20 ml of reference solution (b) is not less than 50 per cent of the full scale of the recorder.

 

Inject 20 ml of reference solution (a). When the chromatograms are recorded in the conditions described above, the retention times are: methylprednisolone, about 11.5 minutes and betamethasone, about 12.5 minutes. The test is not valid unless the resolution between the peaks corresponding to methylprednisolone and betamethasone is at least 1.5; if necessary, adjust the concentration of acetonitrile in mobile phase A.

 

Inject separately 20 ml of the mixture of equal volumes of acetonitrile R and methanol  R as a blank, 20 ml of the test solution and 20 ml of reference solution (b). In the chromatogram obtained with the test solution: the area of any peak, apart from the principal peak, is not greater than the area of the principal peak in the chromatogram obtained with reference solution (b) (1.0 per cent) and not more than one such peak has an area greater than half the area of the principal peak in the chromatogram obtained with reference solution (b) (0.5 per cent); the sum of the areas of all the peaks, apart from the principal peak, is not greater than twice the area of the principal peak in the chromatogram obtained with reference solution (b) (2.0 per cent). Disregard any peak due to the blank and any peak with an area less than 0.05 times the area of the principal peak in the chromatogram obtained with reference solution (b).

 

Loss on drying (2.2.32)

 

Not more than 0.5 per cent, determined on 0.500 g by drying in an oven at 100C to 105C.

 

Assay

 

Dissolve 0.100 g in alcohol  R and dilute to 100.0 ml with the same solvent. Dilute 2.0 ml of the solution to 100.0 ml with alcohol  R. Measure the absorbance (2.2.25) at the maximum at 238.5 nm.

 

Calculate the content of C22H29FO5 taking the specific absorbance to be 395.

 

Storage

 

Store protected from light.

 

IMPURITIES

 

A.    dexamethasone,

 

B.    21-chloro-9-fluoro-11b,17-dihydroxy-16b-methylpregna-1,4-diene-3,20-dione,

 

C.    17,21-dihydroxy-16b-methylpregna-1,4,9(11)-triene-3,20-dione,

 

D.    9-fluoro-11b,17-dihydroxy-16b-methyl-3,20-dioxopregna-1,4-dien-21-yl ethoxycarboxylate,

 

E.    9,11b-epoxy-17,21-dihydroxy-16b-methyl-9b-pregna-1,4-diene-3,20-dione,

 

F.    17,21-dihydroxy-16b-methylpregna-1,4,11-triene-3,20-dione,

 

G.    11a,17,21-trihydroxy-16b-methylpregna-1,4-diene-3,20-dione,

 

H.    14-fluoro-11b,17,21-trihydroxy-16b-methyl-8a,9b,14b-pregna-1,4-diene-3,20-dione,

 

I.     8-fluoro-11b,17,21-trihydroxy-16b-methyl-8a,9b-pregna-1,4-diene-3,20-dione,

 

J.     17,21-dihydroxy-16b-methylpregna-1,4-diene-3,20-dione.

 

 

 


Ph Eur

 

Action and use

 

Corticosteroid.

 

Preparation

 

Betamethasone Tablets