Assay—
Transfer to a suitable separator an accurately measured volume of Injection, equivalent to about 20 mg of pyridostigmine bromide. Add 10 mL of 1 N hydrochloric acid, and mix. Extract with four 20-mL portions and one 15-mL portion of ethyl ether, and discard the extracts. Transfer the aqueous layer to a 100-mL volumetric flask, using about 25 mL of water to aid the transfer. Place the flask on a steam bath and warm, with the aid of a stream of nitrogen, to evaporate any residual ether, then cool the flask to room temperature, dilute with water to volume, and mix. Dilute 20.0 mL of the resulting solution with 0.1 N hydrochloric acid to 100.0 mL, and mix. Dissolve an accurately weighed quantity of
USP Pyridostigmine Bromide RS in 0.1 N hydrochloric acid to obtain a Standard solution having a known concentration of about 40 µg per mL. Concomitantly determine the absorbances of both solutions in 1-cm cells at the wavelength of maximum absorbance at about 269 nm, with a suitable spectrophotometer, using 0.1 N hydrochloric acid as the blank. Calculate the quantity, in mg, of C
9H
13BrN
2O
2 in each mL of the Injection taken by the formula:
(0.5C / V)(AU / AS),
in which
C is the concentration, in µg per mL, of
USP Pyridostigmine Bromide RS in the Standard solution,
V is the volume, in mL, of Injection taken, and
AU and
AS are the absorbances of the solution from the Injection and the Standard solution, respectively.
191
.