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
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