Identification—
A:
To a portion of Injection, equivalent to about 1 mg of digitoxin, add 10 mL of water, and extract with 10 mL of chloroform. Evaporate the chloroform extract on a steam bath with the aid of a current of air to dryness. Dissolve the residue in 2 mL of a solution prepared by mixing 0.3 mL of
ferric chloride TS and 50 mL of glacial acetic acid, and underlay with 2 mL of sulfuric acid: at the zone of contact of the two liquids a brown color, which gradually changes to light green, then to blue, is produced, and finally the entire acetic acid layer acquires a blue color.
B:
To a portion of Injection, equivalent to about 0.2 mg of digitoxin, add 10 mL of water, and extract with 10 mL of chloroform. Evaporate the chloroform extract on a steam bath with the aid of a current of air to dryness. Add 2 mL of a freshly prepared 1 in 100 solution of m-dinitrobenzene in alcohol, and allow to stand for 10 minutes with frequent shaking. Add 2 mL of a 1 in 200 solution of tetramethylammonium hydroxide in alcohol, and mix: a red-violet color develops slowly and then fades.
C:
The retention time of the major peak in the chromatogram of the
Assay preparation corresponds to that of the major peak in the chromatogram of the
Standard preparation as obtained in the
Assay.
Assay—
Mobile phase, Standard preparation, System suitability preparation, and Chromatographic system—
Prepare as directed in the
Assay under
Digitoxin.
Assay preparation—
Transfer an accurately measured volume of Injection, equivalent to about 1 mg of digitoxin, to a 25-mL volumetric flask. Dilute with Mobile phase to volume, and mix.
Procedure—
Proceed as directed for
Procedure in the
Assay under
Digitoxin. Calculate the quantity, in µg, of C
41H
64O
13 in each mL of the Injection taken by the formula:
25(C / V)(rU / rS),
in which
C is the concentration, in µg per mL, of
USP Digitoxin RS in the
Standard preparation,
V is the volume, in mL, of Injection taken, and
rU and
rS are the peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
85
—
It contains not more than 111.0 USP Endotoxin Units per mg of digitoxin.