• Digoxin is the one of the important cardiotonic glycosides obtained from the leaf of Digitalis lanata, Digitalis purpurea

• Family- Scrophulariaceae

• Digoxin chemically, it is a glycoside produces three molecule of digitoxose sugar and one molecule of digoxygenin during hydrolysis. 

• It shows positive inotropic and negative chronotropic activity. 

• Digoxin hydrolyzed at acidic pH but stable at pH 7. 

• It starts its action within 30 minutes to 2 hours, half life is 30-40 hours and major eliminated by kidney. 

• The melting point of the digoxin is 230-265°C. 

Properties of digoxin are: 

It occurs as clear to white coloured crystals or crystalline powder. 

It tastes bitter and has no odour. 

It starts melting at 235ºC. 

It is soluble in dilute alcohol, pyridine, or mixture of chloroform and alcohol. 

It is insoluble in ether, acetone, ethyl acetate, chloroform, and water.

It is slightly soluble in diluted alcohol, and very slightly soluble in 40% propylene glycol. 

On heating, it gets decomposed and emits acrid smoke and irritating fumes. 

Industrial Production Industrial production of digoxin involves the following steps: 

1. Initial Extraction and Removal of Phenolics by Precipitation 

2. The powdered drug (20ml) is extracted with 100ml of 70% ethanol and heated on a hot plate for 20 minutes. 

3. The extract is cooled and filtered or centrifuged to remove solid plant debris. 

4. 150ml of water and 20ml of the strong lead sub-acetate solution is added to the filtrate or supernatant. 

5. A precipitate of phenolic compounds is obtained in the form of insoluble lead complexes. 

6. The mixture is centrifuged, the supernatant is pipetted out, and 10% H2SO4 is added dropwise till no precipitate forms. 

7. This treatment removes the excess amount of lead ions in the form of insoluble lead sulphate. 

8. Solvent Partition to Remove Cardenolides from Aqueous Layer 

9. The mixture is centrifuged, the supernatant is pipetted out and further extracted with 50ml of chloroform four times. 

10. The chloroform extract is combined and washed with 20 ml of water to remove any residual lead ions.

11. The chloroform extract is separated and dried using anhydrous sodium sulphate. 

12. The obtained chloroform solution is filtered into a round bottom flask. 

13. 5ml aliquots of chloroform are evaporated to dryness in separate evaporating dishes. 

14. In the final step, two tests are performed on the residue; the first is Kedde test for cardenolides and the second is the iron (III) chloride test for phenolic compounds. 

15. To isolate digitoxin and digoxin, TLC is performed and the reference solution for digoxin is 5%w/v solution in ethanol. 

Industrial Production Industrial production of digoxin involves the following steps: 

Method II

1. The dried powdered leaf was extracted with petroleum ether under reduced pressure. 

2. Discard the extract and digested the marc with water at 0-4°C so that the polysaccharide may remove. 

3. Again the filtrate is discarded and the marc is extracted with alcohol and water. 

4. Under reduced pressure alcoholic extract is concentrated at 50°C. 

5. This concentrate solution is treated with lead acetate to remove the impurities. 

6. In the water-soluble glycoside portion (obtain after the treatment of lead acetate) maintain the pH around 6 and then wash with non polar solvent. 

7. Discard the organic layer and treat the aqueous layer with 0.5 % sodium sulphate to remove the lead sulphate. 

8. The aqueous layer is then treated with chloroform and afterwards with ethanol.

9. The Chloroform portion contains less polar glycoside while the ethanolic portion contains more polar glycoside. 

10. The ethanolic portion further treated and subjected to chromatography to separate the digoxin. 

Paper Chromatography Process 

1. The sample solution [2mg in 5ml of CHCl 3 and CH3OH (1:1)] and standard solution of the same concentration are compared by placing 10 ml aliquots of each as separate spots on a filter paper strip (10cm from one end). 

2. The Chromatogram is developed in a closed chamber according to the descending technique, in which the end of the strip which is nearest to the sample is kept in a trough with equilibrated solvent and the trough is located in the upper part of the chamber. Thus, the solvent runs down through the paper. 

3. A saturated amount is placed by placing a small amount of both phases of the equilibrated solvent in the bottom of the chromatographic chamber. 

4. The chromatogram is removed from the chamber and air-dried when the solvent reaches a point within 5cm of the bottom edge of the strip.

5. Location of glycosidic zones in the sample and standard is determined by spraying the strip with a 25% solution of trichloroacetic acid in methanol followed by heating at 100C for a minute. 

6. The presence of a greyish green spot indicates the presence of digoxin in the sample, having R1 value same as in standard solution: 

7. On paper chromatograms, digoxin is determined by using Baljet picric reagent, 

8. Both digoxin and isodigoxin can be determined by using Keller -Killiani reagent (ferric chloride with sulphuric and glacial acetic acids). 

9. TLC densitometry is used as an alternative method to HPLC and GLC for the standardisation of crude medicinal plant extracts of Digitalis lanata. 

Estimation: 

Dissolve digoxin (equivalent to 500 mg) in 5 ml solvent (mixture of chloroform: methanol 65:35) and add 20 ml of glacial acetic acid. 

Shake one hour continuously and filter the solution. 

Dilute 5 ml of this filtrate to 25 ml with glacial acetic acid which contain 0.005 percent W/V ferric chloride and 2 percent V/V sulphuric acid and allow to stand for 90 minutes. 

Measure the absorbance at 519 mµ and compare with this standard solution prepared in same way. 

TLC study

Stationary phase: Silica gel G. 

Mobile phase - Cyclohexane: Acetone: Acetic acid (49:49:2). 

Detecting reagent: 5 percent sulphuric acid. 

View under UV far light digoxin appears as blue colour, compare with standard 

Utilisation 

• Digoxin is used along with other medications for treating heart failure. 

• It is also used to treat certain types of irregular heartbeat (such as chronic atrial fibrillation). 

• Treatment of heart failure helps to regulate the irregular heartbeat and also decreases the risk of blood clots, thus reducing the chances of having a heart attack or stroke. 

• Digoxin is a cardiac glycoside that acts by affecting some minerals (sodium and potassium) of the heart cells. 

• This reduces the strain on the heart and also helps in maintaining a normal, steady, and strong heartbeat.