Identification—
A:
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 for enalapril maleate.
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 for hydrochlorothiazide.
Dissolution 
711
—
Medium:
water; 900 mL.
Apparatus 2:
50 rpm.
Time:
30 minutes.
Determine the amount of C20H28N2O5·C4H4O4 dissolved, using filtered portions of the solution under test and following the Procedure for content uniformity of enalapril maleate, making any necessary volumetric adjustments, in comparison with a Standard solution of USP Enalapril Maleate RS having similar concentrations in the same Medium.
Determine the amount of C7H8ClN3O4S2 dissolved by employing UV absorption at the wavelength of maximum absorbance at about 320 nm in 1-cm cells, on filtered portions of the solution under test, suitably diluted with Medium, in comparison with a Standard solution having a known concentration of USP Hydrochlorothiazide RS dissolved in 20 mL of methanol and diluted with Medium.
Tolerances—
Not less than 80% (Q) of the labeled amount of enalapril maleate (C20H28N2O5·C4H4O4) and not less than 60% (Q) of the labeled amount of hydrochlorothiazide (C7H8ClN3O4S2) are dissolved in 30 minutes.
Uniformity of dosage units 905:
meet the requirements.
Procedure for content uniformity of enalapril maleate—
Buffer solution A—
Dissolve 136 g of monobasic potassium phosphate in 800 mL of water, adjust with phosphoric acid to a pH of 4.0, dilute with water to 1000 mL, and mix.
Buffer solution B—
Transfer 20.0 mL of Buffer solution A to a 1000-mL volumetric flask, dilute with water to volume, and mix.
Mobile phase—
Prepare a filtered and degassed mixture of water, acetonitrile, and
Buffer solution A (34:15:1). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard solution—
Dissolve an accurately weighed quantity of
USP Enalapril Maleate RS in
Buffer solution B to obtain a solution having a known concentration of about 100 µg per mL.
Test solution—
Transfer one finely powdered Tablet to a 50-mL volumetric flask, add about 30 mL of Buffer solution B, and sonicate for 15 minutes. Shake by mechanical means for 30 minutes, dilute with Buffer solution B to volume, sonicate for 30 minutes, mix, and filter, discarding the first portion of the filtrate. Dilute a portion of the filtrate with Buffer solution B quantitatively to obtain a solution containing about 100 µg per mL.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 215-nm detector and a 4.6-mm × 20-cm column containing 10-µm packing L7 and maintained at a temperature of 80
. The flow rate is about 2 mL per minute. Chromatograph the
Standard solution, and record the peak responses as directed for
Procedure: the capacity factor,
k¢, is not less than 2.5; the column efficiency determined from the analyte peak is not less than 1000 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 2.0%.
Procedure—
Separately inject equal volumes (about 50 µL) of the
Test solution and the
Standard solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of C
20H
28N
2O
5·C
4H
4O
4 in the Tablet taken by the formula:
(TC/D)(rU / rS),
in which
T is the labeled quantity, in mg, of enalapril maleate in the Tablet;
C is the concentration, in µg per mL, of
USP Enalapril Maleate RS in the
Standard solution; D is the concentration, in µg per mL, of enalapril maleate in the
Test solution; based upon the labeled quantity per Tablet and the extent of dilution; and
rU and
rS are the enalapril peak responses obtained from the
Test solution and the
Standard solution, respectively.
Procedure for content uniformity of hydrochlorothiazide—
Buffer solution A and Buffer solution B—
Prepare as directed under Procedure for content uniformity of enalapril maleate.
Standard solution—
Transfer about 50 mg of
USP Hydrochlorothiazide RS, accurately weighed, to a 200-mL volumetric flask. Add 20 mL of methanol to dissolve the material, dilute with
Buffer solution B to volume, and mix to obtain a stock solution. Transfer 5.0 mL of the stock solution to a 25-mL volumetric flask, dilute with
Buffer solution B to volume, and mix to obtain a solution having a known concentration of about 50 µg per mL.
Test solution—
Transfer 1 Tablet to a volumetric flask of a suitable size such that, when the hydrochlorothiazide is dissolved from the Tablet, a solution having a concentration of about 250 µg per mL is obtained. Add a volume of Buffer solution B equal to about half the capacity of the flask, and sonicate with occasional shaking for 15 minutes. Shake by mechanical means for 30 minutes, dilute with Buffer solution B to volume, sonicate for 30 minutes, mix, and filter, discarding the first portion of the filtrate. Transfer 5.0 mL of the clear filtrate to a 25-mL volumetric flask, dilute with Buffer solution B to volume, and mix.
Procedure—
Determine the absorbances of the
Standard solution and the
Test solution in 1-cm cells at the wavelength of maximum absorbance at about 320 nm and at 360 nm, with a suitable spectrophotometer, relative to
Buffer solution B as the blank. Calculate the quantity, in mg, of C
7H
8ClN
3O
4S
2 in the Tablet taken by the formula:
(TC / D)(A320 – A360)U /(A320 – A360)S,
in which
T is the labeled quantity, in mg, of hydrochlorothiazide in the Tablet;
C is the concentration, in µg per mL, of
USP Hydrochlorothiazide RS in the
Standard solution; D is the concentration, in µg per mL, of hydrochlorothiazide in the
Test solution, based upon the labeled quantity per Tablet and the extent of dilution; and the parenthetic expressions are the differences in absorbances of the two solutions at the wavelengths indicated by the subscripts for the
Test solution (
U) and the
Standard solution (
S), respectively.
Related compounds—
Buffer solution and Mobile phase—
Proceed as directed in the Assay for enalapril maleate.
Test solution—
Use the Assay preparation prepared as directed in the Assay for enalapril maleate.
Enalaprilat solution—
Transfer about 10 mg of
USP Enalaprilat RS, accurately weighed, to a 25-mL volumetric flask, dilute with water to volume, and mix.
Enalapril diketopiperazine solution—
Carefully place about 20 mg of
USP Enalapril Maleate RS in a 100-mL beaker to form a mound on the bottom of the beaker. Place the beaker on a hot plate at about one-half the maximum hot plate temperature setting. Heat for about 5 to 10 minutes until the solid is melted. Immediately remove the beaker from the hot plate, and allow to cool.
[NOTE—Avoid overheating to prevent heat-induced degradation, which would give rise to a brown color.
] To the cooled residue in the beaker add 50 mL of acetonitrile, and sonicate for a few minutes to dissolve. The solution typically contains, in each mL, between 0.2 mg and 0.4 mg of enalapril diketopiperazine.
Standard solution—
Transfer about 40 mg of
USP Enalapril Maleate RS, accurately weighed, to a 200-mL volumetric flask, and dissolve with about 50 mL of methanol. Pipet 1 mL each of
Enalaprilat solution and
Enalapril diketopiperazine solution into the volumetric flask, dilute with
Buffer solution to volume, and mix.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 215-nm detector and a 4.6-mm × 25-cm column that contains 5-µm packing L7. The column temperature is maintained at 65
, and the flow rate is about 1.5 mL per minute. Chromatograph the
Standard solution, and record the peak responses as directed for
Procedure: the relative retention times are about 0.3, 0.4, and 1.0 for enalaprilat, enalapril diketopiperazine, and enalapril, respectively; the resolution,
R, between any of the peaks is not less than 1.3; the column efficiency is not less than 700 theoretical plates for enalapril, 1500 for enalaprilat, and 1500 for enalapril diketopiperazine; the tailing factor is not more than 3.5; and the relative standard deviation for replicate injections is not more than 5.0% for enalaprilat and enalapril diketopiperazine and not more than 2.0% for enalapril.
Procedure—
Separately inject equal volumes (about 50 µL) of the
Standard solution and the
Test solution into the chromatograph, record the chromatograms, and measure the responses for the peaks. Calculate the quantity, in mg, of enalaprilat (enalapril related compound A) in the portion of Tablets taken by the formula:
0.2C(rU / rS),
in which
C is the concentration, in µg per mL, of enalaprilat in the
Standard solution; and
rU and
rS are the peak responses of enalaprilat obtained from the
Test solution and the
Standard solution, respectively.
Calculate the quantity, in mg, of enalapril diketopiperazine in the portion of Tablets taken by the formula:
0.2C(rU / rS),
in which C is the concentration, in µg per mL, of enalapril diketopiperazine in the Standard solution; and rU and rS are the peak responses of enalapril diketopiperazine obtained from the Test solution and the Standard solution, respectively: not more than 5.0% of total related compounds is found, calculated on the basis of the portion of Tablets taken as directed under Assay for enalapril maleate.
Assay for enalapril maleate—
Buffer solution—
Transfer 136 mg of monobasic potassium phosphate to a 1000-mL volumetric flask, add 800 mL of water, adjust with phosphoric acid to a pH of 2.0, dilute with water to volume, and mix.
Mobile phase—
Prepare a degassed and filtered solution of
Buffer solution and acetonitrile (6:4). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard preparation—
Transfer about 40 mg of
USP Enalapril Maleate RS, accurately weighed, to a 200-mL volumetric flask, add about 50 mL of methanol to dissolve, dilute with
Buffer solution to volume, and mix.
Assay preparation—
Weigh and finely powder not fewer than 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 40 mg of enalapril maleate, to a 200-mL volumetric flask, add about 50 mL of Buffer solution, and sonicate for 15 minutes. Add about 50 mL of methanol to the flask, sonicate for 15 minutes, dilute with Buffer solution to volume, mix, and filter.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 215-nm detector and a 4.6-mm × 25-cm column that contains 5-µm packing L7. The column temperature is maintained at 65
, and 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 is not less than 700 theoretical plates; the tailing factor is not more than 3.5; and the relative standard deviation for replicate injections is not more than 2.0%.
Procedure—
Separately inject equal volumes (about 50 µL) of the
Assay preparation and the
Standard preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of enalapril maleate (C
20H
28N
2O
5·C
4H
4O
4) in the portion of Tablets taken by the formula:
200C(rU / rS),
in which
C is the concentration, in µg per mL, of
USP Enalapril Maleate RS in the
Standard preparation; and
rU and
rS are the peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
Assay for hydrochlorothiazide—
Buffer solution—
Prepare as directed under Assay for enalapril maleate.
Mobile phase—
Prepare a filtered and degassed mixture of
Buffer solution and acetonitrile (9:1). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard preparation—
Transfer about 20 mg of
USP Hydrochlorothiazide RS, accurately weighed, to a 200-mL volumetric flask, add about 50 mL of methanol to dissolve, dilute with
Buffer solution to volume, and mix.
Assay preparation—
Prepare as directed for the Assay preparation under Assay for enalapril maleate, except to weigh a portion of the powdered Tablets equivalent to about 20 mg of hydrochlorothiazide.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 310-nm detector and a 4.6-mm × 20-cm column that contains 10-µm packing L7. The column temperature is maintained at 30
, and the flow rate is about 2.5 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; the column efficiency is not less than 1000 theoretical plates; 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 50 µL) of the
Assay preparation and the
Standard preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of hydrochlorothiazide (C
7H
8ClN
3O
4S
2) in the portion of Tablets taken by the formula:
200C(rU / rS),
in which
C is the concentration, in µg per mL, of
USP Hydrochlorothiazide RS in the
Standard preparation; and
rU and
rS are the peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
