A: The relative retention time of the major peak in the chromatogram of the Assay preparation corresponds to that in the chromatogram of the Standard preparation, as obtained in the Assay.

B: It meets the requirements of the tests for Sodium 191.

Internal standard solution— Dissolve a quantity of biphenyl in methylene chloride to obtain a solution containing 5 mg per mL.

Standard stock preparation— Prepare a solution of USP Valproic Acid RS in Internal standard solution having a concentration of about 8 mg per mL.

Standard preparation— Transfer 5.0 mL of the Standard stock preparation into a 50-mL volumetric flask, and dilute with methylene chloride to volume.

Assay preparation— Transfer an accurately measured volume of Injection, equivalent to 400 mg of valproic acid, into a suitable container; add about 20 mL of 5% (v/v) hydrochloric acid; shake by mechanical means for 2 minutes; add 50.0 mL of the Internal standard solution; and shake by mechanical means for 1 hour. Allow the phase to separate (approximately 1 hour). The bottom organic layer remains cloudy, and at times a slight emulsion may persist. If an emulsion forms, break it up by stirring it with a glass rod. Pipet 5 mL of the extract from the bottom organic layer into a 50-mL volumetric flask, and dilute with methylene chloride.

Chromatographic system (see Chromatography 621)— The gas chromatograph is equipped with a flame-ionization detector and a 2-mm × 1.8-m glass column packed with 10% phase G34 on 80- to 100-mesh support S1A. The column temperature is maintained at about 155, the injection port temperature is maintained at about 275, and the detector block temperature is maintained at about 300. Dry helium is used as the carrier gas, at a flow rate of about 20 mL per minute. Chromatograph the Standard preparation as directed for Procedure: the resolution, R, between the valproic acid and biphenyl peaks is not less than 3.0; and the relative standard deviation of the peak area ratios for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 2 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the peak areas for the valproic acid and biphenyl peaks. Calculate the quantity, in mg, of valproic acid in the volume of Injection taken by the formula: in which C is the concentration, in mg per mL, of USP Valproic Acid RS in the Standard preparation; RU and RS are the peak area ratios obtained from the Assay preparation and the Standard preparation, respectively; and D is the appropriate dilution factor used to prepare the Assay preparation.