A: Transfer to a centrifuge tube a quantity of powdered Tablets, equivalent to about 500 mg of metyrapone, add 10 mL of 1 N sodium hydroxide, and mix. Extract with 10 mL of chloroform, centrifuge, and filter: the IR absorption spectrum of this solution, determined in a 0.5-mm cell against chloroform, exhibits maxima only at the same wavelengths as that of a similar solution of USP Metyrapone RS.

B: Transfer to a centrifuge tube 1 mL of the filtrate obtained in Identification test A, add 20 mL of chloroform, and extract with 30 mL of 1 N sulfuric acid, centrifuging and filtering the sulfuric acid layer through a pledget of cotton. Dilute 1 mL of this solution with 1 N sulfuric acid to 100 mL: the UV absorption spectrum of the resulting solution exhibits maxima and minima at the same wavelengths as that of a similar solution of USP Metyrapone RS, concomitantly measured.

Medium: 0.1 N hydrochloric acid; 900 mL.

Apparatus 1: 100 rpm.

Time: 45 minutes.

Procedure— Determine the amount of C14H14N2O dissolved from UV absorbances at the wavelength of maximum absorbance at about 259 nm of filtered portions of the solution under test, suitably diluted with 0.1 N hydrochloric acid, in comparison with a Standard solution having a known concentration of USP Metyrapone RS in the same medium.

Tolerances— Not less than 60% (Q) of the labeled amount of C14H14N2O is dissolved in 45 minutes.

(Official January 1, 2007)

2 ,4-Dinitrophenylhydrazine in methanol—Shake by mechanical means about 1 g of 2,4-dinitrophenylhydrazine with 75 mL of methanol in a 100-mL flask for about 15 minutes, and filter through paper. Prepare fresh daily.

2 ,4-Dinitrophenylhydrazine hydrochloride—Mix 2 mL of hydrochloric acid with 23 mL of 2,4-Dinitrophenylhydrazine in methanol.

Potassium hydroxide in methanol— Dissolve 5 g of potassium hydroxide in 100 mL of methanol, and filter.

Standard preparation— Dissolve an accurately weighed quantity of USP Metyrapone RS in chloroform, and dilute quantitatively and stepwise with chloroform to obtain a solution having a known concentration of about 100 µg per mL.

Assay preparation— Weigh and finely powder not less than 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 25 mg of metyrapone, with the aid of 10 mL of 1 N sodium hydroxide, to a centrifuge tube. Shake gently, and extract with three 15-mL portions of chloroform. Centrifuge each extract, filtering through a pledget of cotton, previously washed with chloroform, into a 50-mL volumetric flask. Add chloroform to volume, and mix. Pipet 5 mL of this solution into a 25-mL volumetric flask, add chloroform to volume, and mix.

Procedure— Pipet 3 mL each of the Standard preparation, the Assay preparation, and chloroform to provide a blank, into separate 50-mL volumetric flasks. To each flask add 1 mL of 2,4-Dinitrophenylhydrazine hydrochloride, and shake lightly. Evaporate the solutions on a steam bath to near dryness. Wash down the sides of the flask with 1 mL of chloroform and methanol (1:1), and again evaporate the solutions to near dryness. Heat the flasks in an oven maintained at 110 to 120 for 30 minutes. Remove the flasks, pipet 10 mL of Potassium hydroxide in methanol into each flask, and heat again in the boiling water bath for 1 minute. Allow to cool to room temperature, insert the stoppers, shake by mechanical means for 5 minutes, add methanol to volume, and mix. Concomitantly determine the absorbances of the solutions in 1-cm cells relative to the blank, at the wavelength of maximum absorbance at about 450 nm, with a suitable spectrophotometer. Calculate the quantity, in mg, of metyrapone (C14H14N2O) in the portion of Tablets taken by the formula: in which C is the concentration, in mg per mL, of USP Metyrapone RS in the Standard preparation, and AU and AS are the absorbances of the solutions from the Assay preparation and the Standard preparation, respectively.