Medium: water; 900 mL.

Apparatus 2: 50 rpm.

Time: 15 minutes.

Buffer solution and Mobile phase— Prepare as directed in the Assay.

0.05% 9-Fluorenylmethyl chloroformate solution— Transfer 100 mg of 9-fluorenylmethyl chloroformate to a 200-mL volumetric flask, dilute with acetonitrile to volume, and mix. Prepare this solution fresh.

Borate buffer— Dissolve 6.2 g of boric acid in approximately 950 mL of water, adjust with 1 N sodium hydroxide to a pH of 9.0, and dilute with water to 1 L.

Diluent— Transfer 176.4 g of sodium citrate dihydrate to a 1000-mL volumetric flask, dissolve in and dilute with Dissolution Medium to volume, and mix.

Standard stock solution— Dissolve an accurately weighed quantity of USP Alendronate Sodium RS in Dissolution Medium, and dilute quantitatively and stepwise with the same solvent to obtain a solution having a known concentration corresponding to the concentration that would be obtained by dissolving 1 Tablet in 900 mL of the same Medium. Calculate the concentration, C, in mg per mL, of anhydrous alendronate sodium in this solution.

Standard solution— Transfer 5.0 mL of the Standard stock solution to a 50-mL polypropylene screw-cap centrifuge tube containing 1.0 mL of Diluent and 5.0 mL of Borate buffer, and mix for about 3 minutes. Add 4.0 mL of 0.05% 9-Fluorenylmethyl chloroformate solution, and agitate for about 30 seconds. Allow the solution to stand at room temperature for 25 minutes. Add 25 mL of methylene chloride, and agitate for about 40 seconds. Centrifuge the mixture for 5 minutes. Use a portion of the clear upper aqueous layer.

Reagent blank— Using 5 mL of water, proceed as directed for Standard solution, beginning with “to a 50-mL polypropylene screw-cap centrifuge tube.”

Test solution— After 15 minutes, withdraw a portion of the solution under test, and centrifuge immediately. Using 5.0 mL of the clear supernatant, proceed as directed for Standard solution, beginning with “to a 50-mL polypropylene screw-cap centrifuge tube.”

Chromatographic system (see Chromatography 621)— Proceed as directed in the Assay. Chromatograph the Standard solution, and record the peak responses as directed for Procedure: the capacity factor, k¢, is not less than 2.0; and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 50 µL) of the Standard solution, the Test solution, and the Reagent blank into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of alendronic acid (C4H13NO7P2) dissolved by the formula: in which C is defined under the Standard stock solution; and rU and rS are the peak areas obtained from the Test solution and the Standard solution, respectively. [NOTE—827.1 is the molecular weight conversion factor (C4H13NO7P2 / C4H12NNaO7P2) multiplied by the volume of the Medium (900 mL).]

Tolerances— Not less than 80% (Q) of the labeled amount of alendronic acid (C4H13NO7P2) is dissolved in 15 minutes. Tablets labeled for weekly dosing: not less than 75% (Q) of the labeled amount of alendronic acid (C4H13NO7P2) is dissolved in 15 minutes.

(Official January 1, 2007)

Diluent— Transfer 29.4 g of sodium citrate dihydrate to a 1000-mL volumetric flask, dissolve in and dilute with water to volume, and mix.

Buffer solution— Transfer 14.7 g of sodium citrate dihydrate and 7.05 g of anhydrous dibasic sodium phosphate to a 1000-mL volumetric flask, dissolve in about 900 mL of water, adjust with phosphoric acid to a pH of 8.0, dilute with water to volume, and mix.

0.1% 9-Fluorenylmethyl chloroformate solution— Transfer 250 mg of 9-fluorenylmethyl chloroformate to a 250-mL volumetric flask, dilute with acetonitrile to volume, and mix. Prepare this solution fresh just prior to use.

Borate solution— Transfer 38.1 g of sodium borate to a 1000-mL volumetric flask, dissolve in and dilute with water to volume, and mix.

Mobile phase— Prepare a filtered and degassed mixture of Buffer solution, acetonitrile, and methanol (75:20:5). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard stock preparation— Prepare a solution of USP Alendronate Sodium RS in diluent containing 0.03 mg of anhydrous alendronate sodium per mL.

Standard preparation— Transfer 5.0 mL of the Standard stock preparation to a 50-mL polypropylene screw-cap centrifuge tube containing 5 mL of Borate solution, and mix for about 3 minutes. Add 4 mL of 0.1% 9-Fluorenylmethyl chloroformate solution, and agitate for about 30 seconds. Allow the solution to stand at room temperature for 25 minutes. Add 25 mL of methylene chloride, and agitate for about 40 seconds. Centrifuge the mixture for 10 minutes. Use the clear upper aqueous layer.

Assay stock preparation— Transfer not fewer than 10 Tablets to a 1000-mL volumetric flask. Add 500 mL of Diluent, shake by mechanical means for 30 minutes, and sonicate for 5 minutes. Dilute with Diluent to volume, mix, and centrifuge a portion of this solution. Quantitatively dilute a portion of the clear supernatant to a concentration in a range of 0.02 to 0.03 mg per mL.

Assay preparation— Using 5.0 mL of the Assay stock preparation, proceed as directed for Standard preparation, beginning with “to a 50-mL polypropylene screw-cap centrifuge tube.”

Reagent blank— Using 5 mL of Diluent, proceed as directed for Standard preparation, beginning with “to a 50-mL polypropylene screw-cap centrifuge tube.”

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 266-nm detector and a 4.1-mm × 25-cm column that contains packing L21. The column is maintained at a constant temperature of about 35. The flow rate is about 1 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the capacity factor, k¢, is not less than 2.0; and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 50 µL) of the Standard preparation, the Assay preparation, and the Reagent blank into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of alendronic acid (C4H13NO7P2) in the portion of Tablets taken by the formula: in which D is the dilution factor for the Assay stock preparation; C is the concentration, in mg per mL, of anhydrous USP Alendronate Sodium RS in the Standard stock preparation; and rU and rS are the peak areas obtained from the Assay preparation and the Standard preparation, respectively. [NOTE—0.919 is the molecular weight conversion factor (C4H13NO7P2 / C4H12NNaO7P2).]