Packaging and storage—
Preserve in tight, light-resistant containers. Store at 25
, excursions permitted between 15
and 30
.
Chromatographic purity—
Detection reagent—
Transfer 2.5 g of cadmium acetate to a 500-mL volumetric flask, add 10 mL of glacial acetic acid, dilute with alcohol to volume, and mix. Just prior to use, prepare a 0.2 in 100 solution of ninhydrin in the cadmium acetate solution for use as the Detection reagent.
Solvent mixture—
Prepare a solution of methanol and water (4:1), and mix.
Ammonium chloride reference solution—
Dissolve 60 mg of ammonium chloride in 10.0 mL of water, and mix.
Standard stock solution—
Dissolve
USP Labetalol Hydrochloride RS in
Solvent mixture, and mix to obtain a solution having a known concentration of 40 mg per mL.
Standard solution 1—
Quantitatively dilute a portion of the Standard stock solution with Solvent mixture to obtain a solution having a known concentration of 0.2 mg per mL.
Standard solution 2—
Quantitatively dilute a portion of the Standard solution 1 with Solvent mixture to obtain a solution having a known concentration of 0.1 mg per mL.
Test solution—
Dissolve 200 mg of Labetalol Hydrochloride in 5.0 mL of
Solvent mixture, and mix.
Procedure I—Apply separately 5-µL portions of the
Standard stock solution, Standard solution 1, Standard solution 2, and the
Test solution to a suitable thin-layer chromatographic plate (see
Chromatography 
621
) coated with a 0.25-mm layer of chromatographic silica gel mixture. Allow the spots to dry, and develop the chromatograms in a solvent system consisting of a mixture of dichloromethane, methanol, and ammonium hydroxide (15:5:1) until the solvent front has moved about three-fourths of the length of the plate. Remove the plate from the developing chamber, mark the solvent front, and allow the solvent to evaporate. Examine the plate under short-wavelength UV light: the
RF value of the principal spot from the
Test solution corresponds to that of the principal spot from the
Standard stock solution.
Spray the plate with
Detection reagent, heat the plate at 105
for 15 minutes, cool to room temperature, and examine the chromatogram: no individual secondary spot observed in the chromatogram of the
Test solution is greater in size or intensity than the principal spot observed in the chromatogram of
Standard solution 1 (0.5% each).
[NOTE—The spots appear as dark orange spots on a light orange to yellow background. A “negative image” spot (white) near the origin may be observed in the chromatogram of the
Test solution. This is due to the formation of ammonium chloride during the chromatographic procedure and may be ignored.
]
Procedure II—Apply separately 10-µL portions of the
Ammonium chloride reference solution, the
Standard stock solution, Standard solution 1, Standard solution 2, and the
Test solution to a suitable thin-layer chromatographic plate (see
Chromatography 621), coated with a 0.25-mm layer of chromatographic silica gel mixture. Allow the spots to dry, and develop the chromatograms in a solvent system consisting of a mixture of ethyl acetate, isopropyl alcohol, water, and ammonium hydroxide (25:15:8:2) until the solvent front has moved about three-fourths of the length of the plate. Remove the plate from the developing chamber, mark the solvent front, and allow the solvent to evaporate. Examine the plate under short-wavelength UV light: no individual secondary spot (other than that due to ammonium chloride) observed in the chromatogram of the
Test solution is greater in size or intensity than the principal spot observed in the chromatogram of
Standard solution 1 (0.5% each).
Total impurities—
The sum of the intensities of all secondary spots (other than those due to ammonium chloride) observed in the chromatograms of the Test solution from both Procedure I and Procedure II does not exceed 1.0%.
Diastereoisomer ratio—
1-Butaneboronic acid solution—
Dissolve 1-butaneboronic acid in pyridine, previously dried over a suitable molecular sieve, and mix to obtain a solution having a known concentration of 20 mg per mL.
System suitability solution—
Dissolve an accurately weighed quantity of
USP Labetalol Hydrochloride RS in
1-Butaneboronic acid solution, and dilute quantitatively and stepwise with
1-Butaneboronic acid solution to obtain a solution having a known concentration of about 1.4 mg of
USP Labetalol Hydrochloride RS per mL. Allow the solution to stand at room temperature for 20 minutes before using.
Test solution—
Transfer about 1 mg of Labetalol Hydrochloride to a 1-mL reaction vial, add 0.7 mL of 1-Butaneboronic acid solution, and mix until the labetalol hydrochloride is completely dissolved. Allow the solution to stand at room temperature for 20 minutes before using.
Chromatographic system (see Chromatography 621)—
The gas chromatograph is equipped with a flame-ionization detector and a 2-mm × 1.8-m glass column packed with 10% phase G3 on 100- to 120-mesh support S1AB. The column temperature is maintained at about 320
, and the injection port and the detector block temperatures are maintained at about 340
. Nitrogen is used as the carrier gas at the flow rate of about 30 mL per minute. Chromatograph the
System suitability solution, and record the peak responses as directed for
Procedure: the relative retention times are about 0.8 for the diastereoisomer B 1-butaneboronate derivative and 1.0 for the diastereoisomer A 1-butaneboronate derivative; the resolution,
R, between the diastereoisomer A 1-butaneboronate derivative and diastereoisomer B 1-butaneboronate derivative peaks is not less than 1.5; and the relative standard deviation of the ratios of the peak areas of the diastereoisomers for replicate injections is not more than 2.0%.
Procedure—
Inject about 2 µL of the
Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the diastereoisomer A content, in percentage, taken by the formula:
100rA / (rA + rB),
in which
rA is the peak area of the diastereoisomer A 1-butaneboronate derivative peak; and
rB is the peak area of the diastereoisomer B 1-butaneboronate derivative peak. The diastereoisomer A content is not less than 45.0% and not more than 55.0%.
Assay—
Mobile phase—
Prepare a suitable filtered and degassed mixture of 0.1
M monobasic sodium phosphate and methanol (65:35). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard preparation—
Dissolve an accurately weighed quantity of
USP Labetalol Hydrochloride RS in
Mobile phase to obtain a solution having a known concentration of about 0.4 mg per mL.
Assay preparation—
Transfer about 40 mg of Labetalol Hydrochloride, accurately weighed, to a 100-mL volumetric flask, dilute with Mobile phase to volume, and mix.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 230-nm detector and a 4.6-mm × 20-cm column that contains packing L1 and is maintained at 60 ± 1
. The flow rate is about 1.5 mL per minute. Chromatograph the
Standard preparation, and record the peak responses as directed for
Procedure: the column efficiency determined from the analyte peak is not less than 700 theoretical plates; the tailing factor for the analyte peak 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 5 µL) of the
Standard preparation and the
Assay preparation into the chromatograph, record the chromatograms, and measure the area responses for the major peaks. Calculate the quantity, in mg, of labetalol hydrochloride (C
19H
24N
2O
3·HCl) in the portion of Labetalol Hydrochloride taken by the formula:
100C(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Labetalol Hydrochloride RS in the
Standard preparation; and
rU and
rS are the peak area responses obtained from the
Assay preparation and the
Standard preparation, respectively.
;)
