Method for the estimation of Ketosteroids content

Method for the estimation of Ketosteroids content

Glasswares and Apparatus Required:

R.B. flask 250mL, separating funnel 250mL, beaker(100 mL)

Reagents And Solvents Required:

1.    Diethyl ether L.R. Grade

2.    Distilled water

3.    Sodium sulphate anhydrous

Procedure:

Weigh accurately about 5.0gm of sample extract powder & transfer into a 250mL R.B. flask. Add about 50mL of 80%v/v methanol & reflux on water bath for one hour. Cool & filter. Repeat the process in the same manner once again. Wash the residue in the R.B. flask with 80% v/v methanol (2x20mL).  Combine the filtrate & washings. Evaporate the solutions to 25mL.  Transfer this solution into a separating funnel. Add 15mL of water & extract the final solution with 25mLx3 times with solvent ether. Collect the ethereal layer & discard the aqueous phase. Ethereal layer is washed with (2x20mL) of 5% w/w sodium hydroxide solution. Discard the aqueous phase. Further wash the ethereal layer slowly with 20mL x 5 times or more to make it alkaline free. Ether layer is filtered through anhydrous sodium sulphate, washing the Na₂SO₄ layer by 10mLx 2 times with ether. Combine the filtrate & washings. Evaporate on water bath & dry at 80 ⁰c  to constant weight.

     

Calculation:

Percentage of Ketosteroids content (w/w) is calculated as:

                                                      wt. of Residue in gm. X    100

           % of Ketosteroids   =      -----------------------------------------------

                                                           wt. of sample in gm

   

For raw material : 10gm sample (20 # ) is to be taken in the case of raw material and the quantity of  methanol (80%) shall be increased accordingly. Rest of the method is same as above.