Monographs: Pharmaceutical substances: Digitoxinum - Digitoxin

Molecular formula. C₄₁H₆₄O₁₃

Relative molecular mass. 765.0

Graphic formula.

Chemical name.

3β-[(O-2,6-Dideoxy-β-D-ribo-hexopyranosyl-(1→4)-O-2,6-dideoxy-β-D-ribo-hexopyranosyl-(1→4)-2,6-dideoxy-β-D-ribo-hexopyranosyl)-oxy]-14-hydroxy-5β-card-20(22)-enolide; CAS Reg. No. 71-63-6.

Description. A white or almost white, microcrystalline powder; odourless.

Solubility. Practically insoluble in water; slightly soluble in ethanol (~750 g/l) TS.

Category. Cardiotonic.

Storage. Digitoxin should be kept in a well-closed container, protected from light.

Additional information. CAUTION: Digitoxin is extremely poisonous and should be handled with care.

Requirements

Definition. Digitoxin contains not less than 95.0% and not more than 105.0% of C₄₁H₆₄O₁₃, calculated with reference to the dried substance.

Identity tests

• Either tests A, B and D or tests B, C and D may be applied.

A. Carry out the examination as described under 1.7 Spectrophotometry in the infrared region. The infrared absorption spectrum is concordant with the spectrum obtained from digitoxin RS or with the reference spectrum of digitoxin.

B. Carry out the test as described under 1.14.1 Thin-layer chromatography, using kieselguhr R1 as the coating substance and a mixture of 10 volumes of formamide R and 90 volumes of acetone R to impregnate the plate, dipping it about 5 mm beneath the surface of the liquid. After the solvent has reached a height of at least 15 cm, remove the plate from the chromatographic chamber and allow to stand for at least 5 minutes. Use the impregnated plate within 2 hours, carrying out the chromatography in the same direction as the impregnation. As the mobile phase, use a mixture of 50 volumes of xylene R, 50 volumes of ethylmethylketone R and 4 volumes of formamide R. Apply separately to the plate 3 μl of each of 2 solutions (A) of the test substance, and (B) of digitoxin RS, each prepared by dissolving 50 mg in a mixture of equal volumes of chloroform R and methanol R to produce 10 ml and then diluting 1 ml to 5 ml with methanol R. Develop the plate for a distance of 12 cm. After removing the plate from the chromatographic chamber, allow it to dry at 115 °C for 20 minutes, cool, spray with a mixture of 15 volumes of a solution of 25 g of trichloroacetic acid R in 100 ml of ethanol (~750 g/l) TS and 1 volume of a freshly prepared 30 mg/ml solution of tosylchloramide sodium R, and then heat the plate at 115°C for 5 minutes. Allow to cool, and examine the chromatogram in daylight and in ultraviolet light (365 nm). The principal spot obtained with solution A corresponds in position, appearance, and intensity with that obtained with solution B.

C. Dissolve 1 mg in 1 ml of ethanol (~750 g/l) TS by heating gently. Cool the solution and add 1 ml of dinitrobenzene/ethanol TS and 1 ml of potassium hydroxide (1 mol/l) VS; a violet colour develops and then fades.

D. Dissolve 1 mg in 2 ml of a solution prepared by mixing 0.5 ml of ferric chloride (25 g/l) TS and 100 ml of glacial acetic acid R; cautiously add 1 ml of sulfuric acid (~1760 g/l) TS to form a lower layer; a brown ring, but no red colour, is produced at the junction of the two liquids, and after some time the acetic acid layer acquires a blue colour (distinction from allied glycosides).

Specific optical rotation. Use a 10 mg/ml solution in chloroform R and calculate with reference to the dried substance;

Sulfated ash. Not more than 1.0 mg/g.

Loss on drying. Dry to constant weight at 105°C; it loses not more than 20 mg/g.

Gitoxin. Dissolve about 5 mg, accurately weighed, in 1 ml of methanol R and dilute to 25 ml with a mixture of equal volumes of hydrochloric acid (~250 g/l) TS and glycerol R. Allow to stand for 1 hour. The absorbance of a 1-cm layer of this solution at 352 nm, when measured against a solvent cell containing a mixture of equal volumes of hydrochloric acid (~250 g/l) TS and glycerol R, is not more than 0.28 (preferably use 2-cm cells for the measurement and calculate the absorbance of a 1-cm layer); the gitoxin content is about 50 mg/g.

Assay. Dissolve about 0.05 g, accurately weighed, in sufficient methanol R to produce 25 ml; dilute 5.0 ml of this solution to 100 ml with methanol R. Place 5.0 ml of the dilute solution to be tested in a 25-ml volumetric flask, add 15 ml of alkaline trinitrophenol TS, and dilute to 25 ml with methanol R. Set aside for 30 minutes, protected from light, and measure the absorbance in a 1-cm layer at the maximum at about 490 nm against a solvent cell containing a solution prepared by diluting 15 ml of alkaline trinitrophenol TS to 25 ml with methanol R. Calculate the amount of C₄₁H₆₄O₁₃ in the substance being tested by comparison with digitoxin RS, similarly and concurrently examined.

Additional requirements for Digitoxin for parenteral use

Complies with the monograph for "Parenteral preparations".

Bacterial endotoxins. Carry out the test as described under 3.4 Test for bacterial endotoxins; contains not more than 111.0 IU of endotoxin RS per mg.