Medium: Phosphoric acid solution (1 in 1000); 250 mL, in a tall-form beaker.

Apparatus 7— Proceed as directed in the chapter, using the transdermal system holder–cylinder (see Figure 7b). Center the Transdermal System onto a dry, unused 10-cm × 10-cm piece of Cuprophan dialysis membrane with the adhesive side against the membrane, taking care to eliminate air bubbles between the membrane and the release surface. Attach the membrane to the cylinder using two Parker O-rings, such that one of the borders of the transdermal system is aligned to the groove and it is wrapped around the cylinder. The filled beakers are weighed and pre-equilibrated to 32.0 ± 0.3, prior to immersing the test sample. Reciprocate at a frequency of about 30 cycles per minute with an amplitude of 2.0 ± 0.1 cm. At the end of each time interval, transfer the test sample to a fresh beaker containing the appropriate volume of Medium, weighed and pre-equilibrated to 32.0 ± 0.3. At the end of each release interval, allow the beakers to cool to room temperature, make up for evaporative losses by adding water to obtain the original weight, and mix. This solution is the final Test solution.

Times: 2, 12, and 24 hours.

Mobile phase— Transfer 0.2 mL of N,N-dimethyloctylamine to a 1-L volumetric flask, add 220 mL of acetonitrile, and mix. Add 300 mL of water, 0.2 mL of glacial acetic acid, 0.20 g of anhydrous sodium acetate, and 0.55 g of sodium 1-dodecanesulfonate, and dilute with water to volume. Mix for 1 hour until clear. Filter and degas. Make adjustments if necessary (see System Suitability under Chromatography 621). (Equilibration of the column may take as long as 3 hours.)

Standard solution— Dissolve an accurately weighed quantity of USP Nicotine Bitartrate Dihydrate RS in Medium, and dilute quantitatively, and stepwise if necessary, with Medium to obtain a solution having a known concentration of about 0.142 mg of nicotine bitartrate per mL (or 0.046 mg nicotine as free base per mL). [NOTE—About 80 mL of this solution is required in order to prepare the System suitability solution.]

System suitability solution— Transfer 8 mg (free base) of nicotine to a 100-mL volumetric flask, and dissolve in 10 mL of acetonitrile. Add 5 mL of 30 percent hydrogen peroxide, and allow 15 minutes to react. Dilute with Medium to volume, and mix. Transfer 20 mL of this solution to a 100-mL volumetric flask, dilute with Standard solution to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 15-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 resolution, R, between nicotine and any degradation peaks is not less than 1.1; the tailing factor is not more than 2.0; and the relative standard deviation for replicate injections is not more than 1.5%.

Procedure— Separately inject equal volumes (about 50 µL) of filtered portions of the Standard solution and the solution under test into the chromatograph, record the chromatograms, and measure the responses for the major peaks.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified below, conforms to Acceptance Table 4.

Phosphate buffer— Dissolve 40.0 g of sodium chloride, 1.0 g of potassium chloride, 8.66 g of dibasic sodium phosphate, and 1.0 g of monobasic potassium phosphate in 5 L of water.

Medium: Phosphate buffer; 500 mL.

Apparatus 6: 50 rpm, double-sided tape being used to attach the Transdermal System to the cylinder.

Times: 6 and 24 hours.

Mobile phase— Proceed as directed in the Assay.

System suitability solution— Transfer 1.0 mL of the System suitability solution, prepared as directed in the Assay, to a 100-mL volumetric flask, dilute with Medium to volume, and mix.

Standard solution— Pipet 6.0 mL of the Standard preparation, prepared as directed in the Assay, into a 50-mL volumetric flask, dilute with Medium to volume, and mix. Dilute quantitatively and stepwise with Medium to obtain an appropriate final concentration.

Test solution— At each of the test times, withdraw a 2-mL aliquot of the solution under test. [NOTE—Replace the aliquots withdrawn for analysis with fresh portions of Medium.]

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 260-nm detector and a 4.6-mm × 12.5-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the Standard solution used for the 6-hour interval, and record the peak responses as directed for Procedure: the resolution, R, between 4,4¢-dipyridyl dihydrochloride and nicotine is not less than 5.0; 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 100 µL) of the filtered portion of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to Acceptance Table 4.

Medium: water; 900 mL.

Apparatus 5: 50 rpm, the stainless steel disk assembly being replaced with a 5-cm watch glass for an 11-mg Transdermal System and an 8-cm watch glass for a 22-mg Transdermal System.

Times: 1, 2, and 4 hours.

Standard solution— Prepare a solution of USP Nicotine Bitartrate RS in water having a known concentration of nicotine similar to that of the solution under test.

Procedure— Determine the amount of C10H14N2 released by employing UV absorption at the wavelength of maximum absorbance at about 259 nm, in comparison with the Standard solution, using water as the blank.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to the following Acceptance Table.

Medium: 0.025 N hydrochloric acid; 600 mL.

Apparatus 5: 50 rpm, a convex screen being used to hold the Transdermal System in position during testing.

Times: 4 and 16 hours.

Standard solution and Procedure— Proceed as directed for Procedure in Test 3.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to Acceptance Table 4.

Phosphate buffer, Medium, and Apparatus— Proceed as directed under Test 2.

Times: 3, 6, and 24 hours.

Mobile phase— Proceed as directed in the Assay.

System suitability solution, Standard solution, Test solution, and Chromatographic system— Proceed as directed under Test 2.

Procedure— Proceed as directed under Test 2 except to inject about 30 µL.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to Acceptance Table 5.

Medium: Phosphoric acid solution (1 in 1000); 250 mL, in a tall-form beaker.

Apparatus 7— Proceed as directed in the chapter, using the transdermal system holder–cylinder (see Figure 4b). Center the Transdermal System onto a dry, unused 10-cm × 10-cm piece of Cuprophan dialysis membrane with the adhesive side against the membrane, taking care to eliminate air bubbles between the membrane and the release surface. Attach the membrane to the cylinder using two Parker O-rings, such that one of the borders of the transdermal system is aligned to the groove and it is wrapped around the cylinder. The filled beakers are weighed and pre-equilibrated to 32.0 ± 0.3, prior to immersing the test sample. Reciprocate at a frequency of about 30 cycles per minute with an amplitude of 2.0 ± 0.1 cm. At the end of each time interval, transfer the test sample to a fresh beaker containing the appropriate volume of Medium, weighed and pre-equilibrated to 32.0 ± 0.3. At the end of each release interval, allow the beakers to cool to room temperature, make up for evaporative losses by adding water to obtain the original weight, and mix. This solution is the final Test solution.

Times: 2, 12, and 24 hours.

Mobile phase— Transfer 0.2 mL of N,N-dimethyloctylamine to a 1-L volumetric flask, add 220 mL of acetonitrile, and mix. Add 300 mL of water, 0.2 mL of glacial acetic acid, 0.20 g of anhydrous sodium acetate, and 0.55 g of sodium 1-dodecanesulfonate, and dilute with water to volume. Mix for 1 hour until clear. Filter and degas. Make adjustments if necessary (see System Suitability under Chromatography 621). [Note—Equilibration of the column may take as long as 3 hours.]

Standard solution— Dissolve an accurately weighed quantity of USP Nicotine Bitartrate Dihydrate RS in Medium, and dilute quantitatively, and stepwise if necessary, with Medium to obtain a solution having a known concentration of about 0.142 mg of nicotine bitartrate per mL (or 0.046 mg nicotine as free base per mL). [NOTE—About 80 mL of this solution is required in order to prepare the System suitability solution.]

System suitability solution— Transfer 8 mg (free base) of nicotine to a 100-mL volumetric flask, and dissolve in 10 mL of acetonitrile. Add 5 mL of 30 percent hydrogen peroxide, and allow 15 minutes to react. Dilute with Medium to volume, and mix. Transfer 20 mL of this solution to a 100-mL volumetric flask, dilute with Standard solution to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 15-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 resolution, R, between nicotine and any degradation peaks is not less than 1.1; the tailing factor is not more than 2.0; and the relative standard deviation for replicate injections is not more than 1.5%.

Procedure— Separately inject equal volumes (about 50 µL) of filtered portions of the Standard solution and the solution under test into the chromatograph, record the chromatograms, and measure the responses for the major peaks.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified below, conforms to Acceptance Table 1.

Phosphate buffer— Dissolve 40.0 g of sodium chloride, 1.0 g of potassium chloride, 8.66 g of dibasic sodium phosphate, and 1.0 g of monobasic potassium phosphate in 5 L of water.

Medium: Phosphate buffer; 500 mL.

Apparatus 6: 50 rpm, double-sided tape being used to attach the Transdermal System to the cylinder.

Times: 6 and 24 hours.

Mobile phase— Proceed as directed in the Assay.

System suitability solution— Transfer 1.0 mL of the System suitability solution, prepared as directed in the Assay, to a 100-mL volumetric flask, dilute with Medium to volume, and mix.

Standard solution— Pipet 6.0 mL of the Standard preparation, prepared as directed in the Assay, into a 50-mL volumetric flask, dilute with Medium to volume, and mix. Dilute quantitatively and stepwise with Medium to obtain an appropriate final concentration.

Test solution— At each of the test times, withdraw a 2-mL aliquot of the solution under test. [NOTE—Replace the aliquots withdrawn for analysis with fresh portions of Medium.]

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 260-nm detector and a 4.6-mm × 12.5-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the Standard solution used for the 6-hour interval, and record the peak responses as directed for Procedure: the resolution, R, between 4,4¢-dipyridyl dihydrochloride and nicotine is not less than 5.0; 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 100 µL) of the filtered portion of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to Acceptance Table 1.

Medium: water; 900 mL.

Apparatus 5: 50 rpm, the stainless steel disk assembly being replaced with a 5-cm watch glass for an 11-mg Transdermal System and an 8-cm watch glass for a 22-mg Transdermal System.

Times: 1, 2, and 4 hours.

Standard solution— Prepare a solution of USP Nicotine Bitartrate RS in water having a known concentration of nicotine similar to that of the solution under test.

Procedure— Determine the amount of C10H14N2 released by employing UV absorption at the wavelength of maximum absorbance at about 259 nm, in comparison with the Standard solution, using water as the blank.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to the following Acceptance Table.

Medium: 0.025 N hydrochloric acid; 600 mL.

Apparatus 5: 50 rpm, a convex screen being used to hold the Transdermal System in position during testing.

Times: 4 and 16 hours.

Standard solution and Procedure— Proceed as directed under Test 3.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to Acceptance Table 1.

Phosphate buffer, Medium, and Apparatus— Proceed as directed under Test 2.

Times: 3, 6, and 24 hours.

Mobile phase— Proceed as directed in the Assay.

System suitability solution, Standard solution, Test solution, and Chromatographic system— Proceed as directed under Test 2.

Procedure— Proceed as directed under Test 2 except to inject about 30 µL.

Tolerances— The amount of C10H14N2 released, as a percentage of the labeled amount of the dose absorbed in vivo, at the times specified, conforms to Acceptance Table 1.

(162.23/462.41)(100C/N)(rU / rS),