Method for estimation of Sennosides content

Method for estimation of Sennosides content

Reagents Required

1. Di Ethyl ether

2. Hydrochloric acid

3. Sodium Hydrogen Carbonate

4. Ferric chloride

5. Magnesium acetate

6. Distilled water

Glass wares, Apparatus & Instruments Required

1. Volumetric flask 100mL,500mL.

2. Evaporating dish

3. Pippette 10mL,20mL

4. Spectrophotometer

5. R.B. Flask

Reagents Preparation

1. Ferric chloride solution :-

  

    105 gm Ferric chloride hexahydrate in 1000 ml distilled water.

2. Magnesium acetate  solution:-

   Dissolve 0.5gm of Magnesium acetate  in  100mL methanol.

3. Dilute Hydrochloric Acid :-

    8.5 ml of  Hydrochloric acid  diluted to 100mL with wter. 

Procedure

Weigh accurately about 0.150 gm of the extract in  a 100mL volumetric flask, dissolve in water and dilute to 100.0mL with the water. Filter the solution, Discard the first 10mL of the filtrate. Transfer 20mL of the filtrate to a 150 ml seprating funnel. Add 0.1mL of dilute Hydrochloric acid and shake with three quanties, each of 15mL ether. Allow the layer to seprate and discard the ether layer.Add 0.10gm of sodium hydrogen carbonate to the aqueous layer and shake for three minutes. Centrifuge and trnasfer 10mL of the supernatant liquid to a 100mL round bottom flask having a ground-glass neck. Add 20 mL of Ferric chloride solution and mix. Heat for 20 minutes under a reflux condenser in a water bath with the water level above that of the liquid in the flask. Add 3 mL of Hydrochloric acid  and heat for a further 30 minute with frequent shaking, to dissolve the precipitate. Cool transfer the mixture to a seprating funel and shake with three quantities each of 25mL of ether. Combine the ether layer and wash with two quantities, each of 15mL of water.Transfer ether layer to a volumetric flask and dilute to 100 mL with ether. Everopate 10mL carefully to dryness and disslove the residue in 10mL of Magnesium acetate solution. Mesure the absorbance at 515nm using methanol as blank..        

Calculation:-

 Calculate the Percentage of Hydroxyanthracene glycosides calculated as sennoside B :-

                            Absorbance X 4.167

                % = ---------------------------------

                           Sample taken in gm