A: Infrared Absorption 197K.

B: The 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.

Diluent— Dilute 100 mL of 1 N hydrochloric acid with dehydrated alcohol to 1000 mL.

Test preparation— Prepare a solution of Tolcapone having a concentration of 0.01 mg per mL in Diluent.

Procedure— Proceed as directed under Spectrophotometry and Light-Scattering 851, and measure the absorbance: the maximum is between 265.2 and 269.3, and the absorptivity is between 75.29 and 79.93.

Alcohol stock solution— Transfer 6.3 µL of absolute alcohol, using a microsyringe, to a 50-mL volumetric flask containing dimethylformamide, and mix.

Methylene chloride stock solution— Transfer 3.8 µL of methylene chloride, using a microsyringe, to a 50-mL volumetric flask containing dimethylformamide, and mix.

Standard solution— Transfer 10.0 mL of Alcohol stock solution and 1.0 mL of Methylene chloride stock solution to a 50-mL volumetric flask. Dilute with dimethylformamide to volume, and mix.

Test solution— Transfer about 200 mg of Tolcapone, accurately weighed, to a 10-mL volumetric flask, add 7 mL of dimethylformamide, and sonicate to dissolve. Dilute with dimethylformamide to volume, and mix.

Chromatographic system (see Chromatography 621)— The gas chromatograph is equipped with a flame-ionization detector, a 0.53-mm × 30-m fused silica column coated with 3.0-µm G43 stationary phase, and a 0.53-mm × 5-m fused silica column coated with 3.0-µm G3 stationary phase. The carrier gas is helium, flowing at a rate of 5 mL per minute. The column temperature is maintained at 35. The temperatures of the injection port and the detector are maintained at 120 and 260, respectively. Chromatograph the Standard solution, and record the peak responses as directed for Procedure: the relative retention time is about 0.7 for alcohol and 1.0 for methylene chloride; and the relative standard deviation for replicate injections is not more than 10.0%.

Procedure— Separately inject equal volumes (about 1 µL) of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the peak responses. Calculate the percentages (w/w) of alcohol and methylene chloride in the portion of Tolcapone taken by the formula: in which C is the concentration, in µL per mL, of each solvent in the Standard solution; D is the density, in mg per µL, of each solvent at 20; W is the weight, in mg, of Tolcapone taken to prepare the Test solution; and rU and rS are the peak areas of the appropriate analyte obtained from the Test solution and the Standard solution, respectively: not more than 0.25% of alcohol is found; and not more than 0.01% of methylene chloride is found. [NOTE—Condition the column at 220 for 15 minutes after each injection.]

TEST 1—

TEST 2—

(Official January 1, 2007)

Diluent— Prepare a mixture of methanol and acetonitrile (24:15).

System suitability solution— Dissolve an accurately weighed quantity of USP Tolcapone Related Compound A RS, USP Tolcapone RS, and USP Tolcapone Related Compound B RS in Diluent; and dilute quantitatively, and stepwise if necessary, with Diluent to obtain a solution having a known concentration of about 5 µg per mL, 5 µg per mL, and 10 µg per mL, respectively. Transfer 2.0 mL of this solution to a 100-mL volumetric flask, add about 63 mL of Diluent, dilute with water to volume, and mix.

Mobile phase— Prepare a filtered and degassed mixture of methanol, 0.05 M monobasic potassium phosphate having a pH of 2.0 ± 0.1, and acetonitrile (8:7:5). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve an accurately weighed quantity of USP Tolcapone RS in Diluent; and dilute quantitatively, and stepwise if necessary, with Diluent to obtain a solution having a known concentration of about 1.0 mg per mL. Transfer 5.0 mL of this solution to a 50-mL volumetric flask, add about 27.5 mL of Diluent, dilute with water to volume, and mix.

Assay preparation— Transfer about 50 mg of Tolcapone, accurately weighed, to a 50-mL volumetric flask, dissolve in and dilute with Diluent to volume, and mix. Transfer 5.0 mL of this solution to a 50-mL volumetric flask, add about 27.5 mL of Diluent, dilute with water to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 230-nm detector and a 4.0-mm × 25-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the System suitability solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.6 for tolcapone related compound A, 1.0 for tolcapone, and about 1.4 for tolcapone related compound B; and the resolution, R, between tolcapone related compound B and tolcapone is not less than 4.0. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the relative standard deviation for replicate injections is not more than 1.0%.

Procedure— Separately inject equal volumes (about 20 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the areas for the major tolcapone peaks. Calculate the quantity, in mg, of C14H11NO5 in the portion of Tolcapone taken by the formula: in which C is the concentration, in mg per mL, of USP Tolcapone RS in the Standard preparation; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.