Identification—
Dissolve 60 mg of
USP Chloroprocaine Hydrochloride RS in 10 mL of water in a 60-mL separator, and in a second 60-mL separator mix a volume of Injection, equivalent to 60 mg of chloroprocaine hydrochloride, with sufficient water to obtain 10 mL of solution. Add 5 mL of dilute ammonium hydroxide (4 in 10) to each, mix, and immediately extract each with four 10-mL portions of chloroform, passing the extracts from the Reference Standard and the test specimen through cotton filters into separate 50-mL volumetric flasks. Dilute each with chloroform to volume, and mix. Add a mixture of chloroform and methanol (4:1) to a suitable chromatographic chamber arranged for thin-layer chromatography (see
Chromatography 
621
), cover the chamber, and allow the system to equilibrate for 15 minutes. Apply separately 10-µL portions of the chloroform solutions obtained from the Reference Standard and the test specimen to a suitable thin-layer chromatographic plate coated with a 0.25-mm layer of chromatographic silica gel mixture. Allow the spots to dry, and develop the chromatogram until the solvent front has moved about three-fourths of the length of the plate. Remove the plate from the developing chamber, mark the solvent front, and allow the solvent to evaporate. Locate the spots on the plate by examination under short-wavelength UV light: the
RF value of the principal spot obtained from the test solution corresponds to that obtained from the Standard solution (
RF about 0.40). After viewing, spray the chromatogram with
iodoplatinate TS: violet-blue colored spots, characteristic of tertiary nitrogen compounds, are visible.
Related compounds—
4-Amino-2-chlorobenzoic acid—
Using the chromatograms obtained as directed for the
Assay, calculate the percentage of 4-amino-2-chlorobenzoic acid (C
7H
6ClNO
2) in the chloroprocaine hydrochloride contained in the Injection taken by the formula:
10,000(C / Wa)(rU / rS),
in which
C is the concentration, in mg per mL, of 4-amino-2-chlorobenzoic acid in the
Standard preparation; Wa is the quantity, in mg, of chloroprocaine hydrochloride in the portion of Injection taken to prepare the
Assay preparation, determined as directed in the
Assay preparation; and
rU and
rS are the 4-amino-2-chlorobenzoic acid peak responses obtained from the
Assay preparation and the
Standard preparation, respectively: not more than 3.0% is found.
Assay—
Mobile phase, Standard 4-amino-2-chlorobenzoic acid solution, Standard preparation, System suitability solution, and Chromatographic system—
Proceed as directed in the
Assay under
Chloroprocaine Hydrochloride.
Assay preparation—
Transfer an accurately measured volume of Injection, equivalent to about 100 mg of chloroprocaine hydrochloride, to a 100-mL volumetric flask, add 40 mL of methanol, dilute with water to volume, and mix.
Procedure—
Proceed as directed in the
Assay under
Chloroprocaine Hydrochloride Calculate the quantity, in mg, of chloroprocaine hydrochloride (C
13H
19ClN
2O
2·HCl) in each mL of the Injection taken by the formula:
100(C / V)(rU / rS),
in which
V is the volume, in mL, of Injection taken, and the other terms are as defined therein.
