Alkaline ascorbate reagent— Mix 100 mL of alcohol, 80 mL of sodium hydroxide solution (1 in 5), and 8 mL of ascorbic acid solution (1 in 50). Prepare fresh on the day of use.

Standard preparation— Place about 18 mg of USP Epinephrine Bitartrate RS, accurately weighed, in a 100-mL volumetric flask, add sodium bisulfite solution (1 in 1000) to volume, and mix. Dilute this solution quantitatively with water to obtain a solution having a known concentration of about 10 µg of epinephrine per mL.

Procedure— Pipet a volume of Injection, equivalent to about 10 µg of epinephrine, into a 50-mL beaker. Into another 50-mL beaker pipet 1 mL of Standard preparation. Treat the contents of each beaker as follows. Add 10 mL of dilute hydrochloric acid (1 in 120), and heat gently to reduce the volume of solution to about 5 mL. Allow to cool to room temperature, then add 5 mL of sodium acetate solution (1 in 5), followed by 0.5 mL of potassium ferricyanide solution (1 in 400), and mix. At 2 minutes, accurately timed, after the last addition, add 20 mL of Alkaline ascorbate reagent, transfer the contents to a corresponding 50-mL volumetric flask with the aid of water, add water to volume, and mix. At 15 to 20 minutes after the addition of the Alkaline ascorbate reagent, determine the fluorescences of each solution and of a reagent blank, with a suitable fluorometer, with an excitation wavelength setting of 420 nm and a fluorescence wavelength setting of 520 nm. Calculate the quantity, in µg, of C9H13NO3 in each mL of Injection taken by the formula: in which C is the concentration of epinephrine, in µg per mL, of USP Epinephrine Bitartrate RS in the Standard preparation, V is the volume, in mL, of Injection taken, and IU, IS, and B are the fluorescence readings of the solutions from the Injection, the Standard preparation, and the reagent blank, respectively.

(Official January 1, 2007)

Standard preparation— Transfer to a 125-mL separator about 50 mg, accurately weighed, of USP Procaine Hydrochloride RS, and dilute with water to 20 mL.

Assay preparation— Transfer to a 125-mL separator an accurately measured volume of Injection, equivalent to about 50 mg of procaine hydrochloride, and dilute with water to 20 mL.

Procedure— To the Standard preparation and also to the Assay preparation add 5 mL of 6 N ammonium hydroxide, then treat each as follows. Extract with five 25-mL portions of chloroform, and filter the combined extracts through about 1 g of anhydrous sodium sulfate supported on a pledget of glass wool. Receive the filtrate in a 200-mL volumetric flask, add chloroform to volume, and mix. Transfer 3.0 mL of this solution to a 100-mL volumetric flask, add chloroform to volume, and mix. Concomitantly determine the absorbances of both solutions at the wavelength of maximum absorbance at about 280 nm, with a suitable spectrophotometer, using chloroform as the blank. Calculate the quantity, in mg, of C13H20N2O2·HCl in each mL of the Injection taken by the formula: in which W is the weight, in mg, of USP Procaine Hydrochloride RS used, V is the volume, in mL, of Injection taken, and AU and AS are the absorbances of the solutions obtained from the Assay preparation and the Standard preparation, respectively.