A: To 2 mL of a solution (1 in 50) add 2 mL of sulfuric acid, cool, and superimpose 2 mL of ferrous sulfate TS: a brown ring is produced at the junction of the two liquids.

B: Dissolve about 5 mg in a mixture of 1 mL of lead acetate TS and 1 mL of 2.5 N sodium hydroxide: a yellow color is produced. Heat the mixture for several minutes on a steam bath: the color changes to brown, and, on standing, a precipitate of lead sulfide separates.

C: A solution of it responds to Identification test A under Thiamine Hydrochloride Injection.

D: Dissolve about 5 mg in 5 mL of 0.5 N sodium hydroxide, and proceed as directed in Identification test B under Thiamine Hydrochloride Injection, beginning with “then add 0.5 mL of potassium ferricyanide TS”: the specified reaction is observed.

Solution A , Solution B, and Mobile phase—Prepare as directed in the Assay under Thiamine Hydrochloride.

Test solution— Dissolve quantitatively an accurately weighed quantity of Thiamine Mononitrate in Mobile phase to obtain a solution having a concentration of about 1.0 mg per mL.

Chromatographic system— Proceed as directed in the test for Chromatographic purity under Thiamine Hydrochloride.

Procedure— Proceed as directed in the test for Chromatographic purity under Thiamine Hydrochloride: the total of the responses of all secondary peaks is not greater than 1.0% of the total of the responses of all of the peaks.

(Official January 1, 2007)

Solution A , Solution B, Mobile phase, Internal standard solution, Standard preparation, and Chromatographic system—Proceed as directed in the Assay under Thiamine Hydrochloride.

Assay preparation— Transfer an accurately weighed quantity of about 200 mg of Thiamine Mononitrate to a 100-mL volumetric flask, dissolve in Mobile phase, dilute with Mobile phase to volume, and mix. Transfer 10.0 mL of this solution to a 50-mL volumetric flask, add 5.0 mL of Internal standard solution, dilute with Mobile phase to volume, and mix.

Procedure— Separately inject equal volumes (about 10 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the areas of the major peaks. Calculate the quantity, in mg, of C12H17N5O4S in the Thiamine Mononitrate taken by the formula: in which 327.36 and 337.27 are the molecular weights of thiamine mononitrate and thiamine hydrochloride, respectively, C is the concentration, in µg per mL, of USP Thiamine Hydrochloride RS in the Standard preparation, and RU and RS are the ratios of the peak areas of thiamine to methylbenzoate obtained from the Assay preparation and the Standard preparation, respectively.