Iron reagent— Dissolve 1.5 g of ferric chloride in 30 mL of water, add 3 mL of perchloric acid and 15 mL of nitric acid, and heat the solution until dense, white fumes are produced. Cool, add 40 mL of water and 10 mL of nitric acid, and dilute with perchloric acid to 100 mL. [note—This concentrate may be stored and used for several months.] Transfer 4.0 mL of this concentrate to a 100-mL volumetric flask, add 36 mL of alcohol, dilute with water to volume, and mix.

Procedure— Place a portion of finely powdered Tablets, equivalent to about 250 mg of testolactone, in a 100-mL volumetric flask, add 85 mL of chloroform, and shake by mechanical means for 30 minutes. Add chloroform to volume, mix, and centrifuge the solution. Transfer 2 mL of the supernatant to a small test tube, and evaporate with the aid of a gentle current of air to dryness. Add 0.1 mL of Hydroxylamine reagent, heat in a water bath at about 60 for 1 minute, then add 1 mL of Iron reagent: a rose-violet color is produced.

Test specimen— Place a portion of finely powdered Tablets, equivalent to about 100 mg of testolactone, in a glass-stoppered, 35-mL centrifuge tube. Add 20 mL of chloroform, insert the stopper, and shake by mechanical means for 10 minutes. Centrifuge the mixture, and filter the supernatant into a 50-mL beaker. Evaporate on a steam bath to dryness, and dry the residue in vacuum at 100 for 3 hours.

1.667C(AU / AS)