Change to read:

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.

Test solution: 10 mg per mL, in water.

Change to read:

0.01 M Ammonium acetate buffer solution— Prepare as directed in the Assay.

Diluent— Adjust the pH of 0.01 M Ammonium acetate buffer solution with sodium hydroxide to 9, and mix. Prepare a degassed mixture of 0.01 M Ammonium acetate buffer solution and acetonitrile (19:1).

Solution A— Prepare a filtered and degassed mixture of 0.01 M Ammonium acetate buffer solution and acetonitrile (19:1).

Solution B— Prepare a filtered and degassed mixture of 0.01 M Ammonium acetate buffer solution and acetonitrile (3:1).

Standard stock solution A— Dissolve an accurately weighed quantity of USP Didanosine Related Compound A RS in Diluent, and dilute quantitatively, and stepwise if necessary, with Diluent to obtain a solution having a known concentration of about 0.05 mg per mL.

Standard stock solution B— Dissolve an accurately weighed quantity of USP Didanosine RS in Diluent, and dilute quantitatively, and stepwise if necessary, with Diluent to obtain a solution having a known concentration of about 0.025 mg per mL.

Standard stock solution C— Dissolve an accurately weighed quantity of USP Didanosine 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 0.025 mg per mL.USP32

Standard solution— Transfer 5.0 mL of Standard stock solution A, 3.0 mL of Standard stock solution B, and 3.0 mL of Standard stock solution CUSP32 to a 50-mL volumetric flask, dilute with Diluent to volume, and mix.

System suitability solution— Dissolve an accurately weighed quantity of USP Didanosine System Suitability Mixture RS in Diluent, and dilute quantitatively, and stepwise if necessary, with Diluent to obtain a solution having a known didanosineUSP32 concentration of about 0.5 mg per mL.

Test solution— Transfer about 50 mg of Didanosine, accurately weighed, to a 100-mL volumetric flask, dissolve in and dilute with Diluent to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 25-cm column that contains 5-µm packing L1. The flow rate is about 2.0 mL per minute. The chromatograph is programmed as follows. Chromatograph the Standard solution, and record the peak responses as directed for Procedure: the relative standard deviation for replicate injections, determined on the didanosine related compound A peak, is not more than 2.0%. [Note—For information purposes only, didanosine elutes between 6 and 7.5 minutes; the relative retention times are about 1.0 for didanosine, 0.28 for didanosine related compound A, and 2.11 for didanosine related compound B.USP32] Chromatograph the System suitability solution, and record the peak responses as directed for Procedure: the resolution, R, between didanosine and dideoxydidehydroinosine is not less than 3.0; and the column efficiency, determined on the dideoxydidehydroinosine peak, is not less than 6000 theoretical plates.

Procedure— Separately inject equal volumes (about 10 µL) of the Standard solution and the Test solution into the chromatograph, record the chromatograms for 30 minutes, and measure all the peak responses. Calculate the percentage of didanosine related compound A in the portion of Didanosine taken by the formula: in which CS is the concentration, in mg per mL, of USP Didanosine Related Compound A RS in the Standard solution; CU is the concentration, in mg per mL, of the Test solution; rU is the peak response of didanosine related compound A in the Test solution; and rS is the peak response of USP Didanosine Related Compound A RS in the Standard solution. Calculate the percentage of any specified impurity and any other individual impurities in the portion of Didanosine taken by the formula: in which CS is the concentration, in mg per mL, of USP Didanosine RS in the Standard solution; CU is the concentration, in mg per mL, of the Test solution; rU is the peak response of each impurity in the Test solution; and rS is the peak response of USP Didanosine RS in the Standard solution. The impurity limits meet the requirements specified in Table 1.

0.01M Ammonium acetate buffer solution— Dissolve 1.54 g of ammonium acetate in a 2000-mL volumetric flask, dilute with water to volume, and mix.

Mobile phase— Prepare a filtered and degassed mixture of 0.01 M Ammonium acetate buffer solution and acetonitrile (21:1). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve an accurately weighed quantity of USP Didanosine RS in water to obtain a solution containing 0.1 mg per mL.

Assay preparation— Transfer an accurately weighed quantity of 50 mg of Didanosine to a 500-mL volumetric flask. Dissolve in and dilute with water to volume. Mix the sample for 1 hour to dissolve completely before use.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 25-cm column that contains packing L1. The flow rate is about 2.0 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the column efficiency is not less than 6000 theoretical plates; the tailing factor is not more than 2.5; and the relative standard deviation for replicate injections is not more than 2.0%. [note—For information purposes only, the retention time of didanosine is between 7 and 11 minutes.]

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 responses for the major peaks. Calculate the percentage of C10H12N4O3 in the portion of Didanosine taken by the formula: in which C is the concentration, in mg per mL, of USP Didanosine RS in the Standard preparation; W is the weight, in mg, of Didanosine taken for the Assay preparation; 100 is the conversion factor to percentage; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.