Alkaline borate solution— Dissolve 6.2 g of boric acid and 7.45 g of potassium chloride in 500 mL of water, add 210 mL of sodium hydroxide solution (1 in 25), and mix. Add water to make 2000 mL of solution, and mix.

Standard solution— [note—Prepare the Standard solution and the Test solution concomitantly.] Dissolve an accurately weighed quantity of USP Mephobarbital RS in Alkaline borate solution to obtain a solution having a concentration of about 10 mg per mL, and quantitatively dilute with water to obtain a solution having a known concentration of about 1.5 mg per mL.

Test solution— Transfer 1 Tablet to a glass-stoppered centrifuge tube, crush the tablet, and add 25.0 mL of Alkaline borate solution. Insert the stopper, shake for 10 minutes, and centrifuge until clear, filtering the supernatant, if necessary. Dilute a portion of the subsequent liquid quantitatively, and stepwise if necessary, with water to obtain a solution having a concentration of about 1.5 mg of mephobarbital per mL.

Procedure— Transfer 3.0 mL each of the Standard solution and the Test solution to separate 200-mL volumetric flasks, dilute each with a 1 in 3 solution of Alkaline borate solution in water to volume, and mix. Concomitantly determine the absorbances of the solutions in 1-cm cells at the wavelength of maximum absorbance at about 245 nm, with a suitable spectrophotometer, using a 1 in 3 solution of Alkaline borate solution in water as the blank. Calculate the quantity, in mg, of C13H14N2O3 in the Tablet taken by the formula: in which T is the labeled quantity, in mg, of mephobarbital in the Tablet; C is the concentration, in mg per mL, of USP Mephobarbital RS in the Standard solution; D is the concentration, in mg per mL, of mephobarbital in the Test solution, based upon the labeled quantity per Tablet and the extent of dilution; and AU and AS are the absorbances of the solutions from the Test solution and the Standard solution, respectively.