Test solution: 5 mg per mL, in water.

Diluent— Transfer 2 mL of lead-free nitric acid into a 1000-mL volumetric flask, dilute with water to volume, and mix.

Standard stock solution— Dissolve 79.9 mg of lead nitrate in 100 mL of Diluent in a 500-mL volumetric flask, and dilute with Diluent to volume. Transfer 10.0 mL of the resulting solution into a 100-mL volumetric flask, and dilute with Diluent to volume. Each mL of the Standard stock solution contains the equivalent of 10 µg of lead.

Standard solutions— Dilute aliquots of the Standard stock solution quantitatively, and stepwise if necessary, with Diluent to obtain solutions having concentrations of 0.03 and 0.015 µg per mL.

Test solution— Transfer 160 mg of Neotame, accurately weighed, to a 10-mL volumetric flask. Dissolve in and dilute with Diluent to volume.

Blank— Use the Diluent as the blank.

Procedure— Separately inject equal volumes (about 15 µL) of the Standard solutions, the Blank, and the Test solution into a suitable graphite furnace atomic absorption spectrophotometer set at 283.3 nm and equipped with an autosampler, pyrolytically coated graphite tubes, a solid pyrolytic graphite platform, and an adequate means of background correction. Use a hollow-cathode lamp as the source, use argon as the purge gas, and breathing-quality air as the alternate gas. Optimize the instrument program as recommended by the manufacturer for lead, using a char temperature of 500 and an atomization temperature of 2000. Correct the area responses of all Test solutions and Standard solutions for the Blank area response. Generate the appropriate lead calibration algorithm, and determine the lead concentration in the Test solution, in µg per mL. Calculate the percentage of lead in the portion of Neotame taken by the formula: in which C is the blank-corrected lead concentration in the Test solution, in µg per mL; and W is the weight, in µg, of Neotame taken to prepare the Test solution: not more than 0.0002% is found.

Mobile phase— Proceed as directed in the Assay.

Related compound A standard solution— Dissolve an accurately weighed quantity of USP Neotame Related Compound A RS in Mobile phase to obtain a solution having a known concentration of about 0.03 mg per mL.

Detector sensitivity solution— Transfer 2 mL of the Related compound A standard solution to a 50-mL volumetric flask, dilute with Mobile phase to volume, and mix.

Standard solution— Prepare as directed for Standard preparation in the Assay.

Test solution— Transfer about 100 mg of Neotame, accurately weighed, to a 50-mL volumetric flask, dissolve in and dilute with Mobile phase to volume, and mix. [NOTE—This solution is stable for up to 32 hours when stored at a temperature of 0 to 10.]

Chromatographic system (see Chromatography 621)— Use the same system as directed in the Assay except to chromatograph the Related compound A standard solution and the Detector sensitivity solution, and record the peak responses as directed for Procedure: the signal-to-noise ratio for the Detector sensitivity solution is not less than 10; and the relative standard deviation for replicate injections of the Related compound A standard solution is not more than 5.0%.

Procedure— Separately inject equal volumes (about 25 µL) of the Related compound A standard solution, the Standard solution, and the Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the percentage of related compound A in the portion of Neotame taken by the formula: in which CS is the concentration, in mg per mL, of USP Neotame Related Compound A RS in the Related compound A standard solution; CU is the concentration, in mg per mL, of Neotame in the Test solution; and rU and rS are related compound A peak responses obtained from the the Test solution and Related compound A standard solution, respectively: not more than 1.5% is found. Calculate the percentage of other impurites in the portion of Neotame taken by the formula: in which CS is the concentration, in mg per mL, of USP Neotame RS in the Standard solution; and CU is the concentration, in mg per mL, of Neotame in the Test solution; Ai is the sum of the responses of all impurity peaks (except that of related compound A and the solvent peak, if observed) in the Test solution; and AS is the response of the neotame peak in the Standard solution: not more than 2.0% is found.

(Official January 1, 2007)

Mobile phase— Dissolve 3.0 g of sodium 1-heptanesulfonate in 740 mL of water in a suitable 1000-mL vessel, add 3.8 mL of triethylamine, and mix. Adjust the resulting solution with phosphoric acid to a pH of 3.5, and dilute with water to 750 mL. Add 250 mL of acetonitrile, adjust with phosphoric acid to an apparent pH of 3.7, mix, filter, and degas. Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve an accurately weighed quantity of USP Neotame RS in Mobile phase, and dilute quantitatively, and stepwise if necessary, with Mobile phase to obtain a solution having a known concentration of about 1.0 mg per mL.

Assay preparation— Transfer about 50 mg of Neotame, accurately weighed, to a 25-mL volumetric flask, dissolve in and dilute with Mobile phase to volume, and mix. Pipet 5 mL of the resulting solution into a 10-mL volumetric flask, dilute with Mobile phase to volume, and mix. [NOTE—This solution is stable for up to 32 hours when stored at a temperature of 0 to 10].

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 210-nm detector and a 4.6-mm × 10-cm column that contains packing L1. The flow rate is about 1.5 mL per minute. The column temperature is maintained at 45. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the tailing factor is not more than 2.0; and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 25 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of C20H30N2O5 in the portion of Neotame taken by the formula: in which C is the concentration, in mg per mL, of USP Neotame RS in the Standard preparation; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.NF24