For Sulfamethoxazole content:
To about 4 g of the oral suspension add 30 ml of 0.1M Sodium hydroxide, shake and extract with four 50 ml quantities of chloroform, washing each extract with the same two 10 ml quantities of 0.1M Sodium hydroxide. Reserve the combined chloroform extracts for the assay for trimethoprim. Dilute the combined aqueous solution and washings to 250 ml with water, filter and dilute 5 ml of the filtrate to 200 ml with water (solution A).
Note: Carry out the following procedure protected from light.
Using 2 ml of solution A. Add 0.5 ml of 4M hydrochloric acid and 1 ml of a 0.1% w/v solution of sodium nitrite and allow to stand for 2 minutes. Add 1 ml of a 0.5% w/v solution of ammonium sulphamate and allow to stand for 3 minutes. Add 1 ml of a 0.1% w/v solution of N-(1-naphthyl) ethylenediamine dihydrochloride and allow to stand for 10 minutes. Dilute the resulting solution to 25 ml with water and measure the absorbance at 538 nm, using in the reference cell a solution prepared in the same manner but using 2 ml of water in place of solution A. Dissolve about 0.25 g of Sulfamethoxazole BPCRS or Sulfamethoxazole WS in 50 ml of 0.1M Sodium hydroxide and dilute to 250 ml with water. Dilute 5 ml of the resulting solution to 200 ml with water (solution B).
Repeat the procedure using 2 ml of solution B and beginning at the words ‘Add 0.5 ml of …’
Calculate the content of C10H11N3O3S from the values of the absorbances obtained using the declared content of Sulfamethoxazole (C10H11N3O3S) in Sulfamethoxazole BPCRS or Sulfamethoxazole WS. Determine the weight per ml of the oral suspension and calculate the content of Sulfamethoxazole (C10H11N3O3S), weight in volume.
For Trimethoprim content:
Extract the chloroform solution reserved in the assay for Sulfamethoxazole with four 50 ml quantities of 1M Acetic acid. Wash the combined extracts with 5 ml of chloroform and dilute the aqueous extracts to 250 ml with 1M Acetic acid. To 10 ml of this solution add 10 ml of 1M Acetic acid and sufficient water to produce 100 ml and measure the absorbance of the resulting solution at the maximum at 271 nm.
Calculation: Calculate the content of Trimethoprim (C14H18N4O3)taking 204 as the value of A (1%, 1 cm) at the maximum at 271 nm. Using the weight per ml of the oral suspension, calculate the content of Trimethoprim (C14H18N4O3), weight in volume.
Test absorbance 1000 250 100
————————x————- x ——————– x ———— x 5 x Weight per ml.
204 100 test wt. (in mg) 10