Betamethasone Oral Solution
» Betamethasone Oral Solution contains not less than 90.0 percent and not more than 115.0 percent of the labeled amount of betamethasone (C22H29FO5).
Packaging and storage— Store at controlled room temperature, protected from light. Preserve in a tight container.
USP Reference standards 11—
USP Betamethasone RS Click to View Structure
Identification—
A:
Test solution— Place a volume of Oral Solution equivalent to about 1 mg of betamethasone in a centrifuge tube. Add 15 mL of 0.1 N hydrochloric acid and 20 mL of ethyl acetate. Shake the tube for about 1 minute. Centrifuge to separate the phases. Transfer the upper phase (ethyl acetate) to a suitable container. Evaporate it to dryness on a steam bath under a gentle stream of nitrogen.
Allow to cool to room temperature. Dissolve the residue in about 0.5 mL of chloroform and methanol (1:1), using a vortex mixer. Transfer the solution to a 2-mL volumetric flask with small portions of a mixture of chloroform and methanol (1:1), dilute with the mixture of chloroform and methanol (1:1) to volume, and mix.
Evaporate 1 mL of the resulting solution on a steam bath just to dryness, and dissolve the residue in 0.5 mL of alcohol.
Standard solution— Prepare a solution of USP Betamethasone RS in alcohol having a concentration of about 1 mg betamethasone per mL.
Developing solvent system: a mixture of chloroform and diethylamine (2:1).
Procedure— Proceed as directed in the chapter. Locate the spots by lightly spraying with dilute sulfuric acid (1 in 2); heat on a hot plate or under a lamp until spots appear.
B: The retention time of the major peak in the chromatogram of the Assay preparation corresponds to that in the chromatogram of the Standard preparation, as obtained in the Assay.
Microbial limits 61— The total aerobic microbial count does not exceed 100 cfu per mL, and the total combined molds and yeasts count does not exceed 10 cfu per mL. It meets the requirements of the test for absence of Escherichia coli.
pH 791: between 2.8 and 3.6.
Deliverable volume 698— For Oral Solution packaged in multiple-unit containers: meets the requirements.
Related compounds— [note—Protect all standard and test solutions from light. ]
Diluent, Buffer, Solution A, Solution B, Mobile phase, Standard stock preparation, System suitability preparation, and Assay preparation —Prepare as directed in the Assay.
Standard solution— Quantitatively dilute with Diluent an aliquot of the Standard stock preparation, as prepared in the Assay, to obtain a solution having a known concentration of about 0.48 µg per mL.
Quantitative limit solution— Quantitatively dilute with Diluent an aliquot of the Standard solution to obtain a solution having a known concentration of about 0.024 µg per mL.
Test solution— Prepare as directed for the Assay preparation.
Chromatographic system (see Chromatography 621)— Proceed as directed in the Assay. Chromatograph the System suitability preparation, and record the peak responses as directed for Procedure: the relative retention times are about 1.0 for betamethasone and 1.2 for beclomethasone; the resolution, R, between betamethasone and beclomethasone is not less than 4.0; and the relative standard deviation of the betamethasone peak for replicate injections of the Quantitative limit solution is not more than 10%.
Procedure— Separately inject equal volumes (about 50 µL) of the Standard solution, the Quantitative limit solution, and the Test solution into the chromatograph, record the chromatograms, and measure the peak area responses for the betamethasone related compounds listed in Table 1 and for betamethasone. Calculate the percentage of each related compound in the portion of Oral solution taken by the formula:
100(CS /CT)(ri / rS)
in which CS is the concentration, in mg per mL, of betamethasone in the Standard solution; CT is the concentration, in mg per mL, of betamethasone in the Test solution, based on the label claim; ri is the peak area response for each related compound obtained from the Test solution; and rS is the peak area response of the betamethasone peak obtained from the Standard solution. The limits are as specified in Table 1.
Table 1
Related Compound Relative
Retention
Time		 Limit
(%)
Betamethasone 1.0 —
9,11-Expoxy-17,21-dihydroxy-16-methylpregna-1,4 diene-3,20-dione 1.25 1.3
17,21-Dihydroxy-16-methylpregna-1,4,11-triene-3,20-dione 1.33 0.7
Assay— [note—Protect all standard and assay preparations from light. ]
Diluent— Prepare a mixture of water and dehydrated alcohol (3:2).
Buffer— Accurately weigh 6.8 g of monobasic potassium phosphate, and transfer to a 1-L flask. Add 1 L of water, and mix until completely dissolved. Adjust with phosphoric acid to a pH of 2.9, and mix.
Solution A— Prepare a filtered and degassed mixture of Buffer and acetonitrile (75:25).
Solution B— Prepare a filtered and degassed mixture of Buffer and acetonitrile (55:45).
Mobile phase— Use variable mixtures of Solution A and Solution B as directed for Chromatographic system. Make adjustments if necessary (see System Suitability under Chromatography 621).
Standard stock preparation— Dissolve an accurately weighed quantity of USP Betamethasone RS in dehydrated alcohol, and sonicate until completely dissolved to obtain a solution having a known concentration of about 0.3 mg per mL. Quantitatively dilute with water an aliquot of the solution to obtain a solution having a known concentration of about 0.12 mg per mL.
System suitability preparation— Dissolve an accurately weighed quantity of beclomethasone in dehydrated alcohol, and sonicate until completely dissolved to obtain a solution having a concentration of about 0.3 mg per mL. Quantitatively dilute with water an aliquot to obtain a solution having a known concentration of about 0.12 mg per mL. Pipet 10.0 mL of this solution and 10.0 mL of the Standard stock preparation into a 25-mL volumetric flask, and dilute with Diluent to volume.
Standard preparation— Quantitatively dilute with Diluent an aliquot of the Standard stock preparation to obtain a solution having a known concentration of about 0.048 mg per mL.
Assay preparation— Transfer an accurately measured volume of Oral Solution, equivalent to about 1.2 mg of betamethasone, to a 25-mL volumetric flask, dilute with Diluent to volume, and mix.
Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 15-cm column that contains 4-µm packing L1. The flow rate is about 1.5 mL per minute. The chromatograph is programmed as follows.
Time
(minutes)		 Solution A
(%)		 Solution B
(%) 		Elution
0–25.0 100®0 0®100 linear gradient
25.0–25.1 0®100 100®0 linear gradient
25.1–35.0 100 0 re-equilibration
Chromatograph the System suitability preparation, and record the peak responses as directed for Procedure: the relative retention times are about 1.0 for betamethasone and 1.2 for beclomethasone; the resolution, R, between betamethasone and beclomethasone is not less than 4.0; the tailing factor of the betamethasone peak is not more than 1.5; and the relative standard deviation for replicate injections of the Standard preparation is not more than 2.0%.
Procedure— Separately inject equal volumes (about 50 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the peak area responses for the betamethasone peaks. Calculate the content of betamethasone (C22H29FO5) as a percentage of the labeled content of betamethasone in the Oral Solution taken by the formula:
100(CS / CU)(rU / rS)
in which CS is the concentration, in mg per mL, of betamethasone in the Standard preparation; CU is the concentration, in mg per mL, of betamethasone in the Assay preparation, based on the label claim; and rU and rS are the peak area responses of the betamethasone peak obtained from the Assay preparation and the Standard preparation, respectively.
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