Packaging and storage
Preserve in tight containers.
Identification
A:
pH 9 Buffer with magnesiumMix 3.1 g of boric acid and 500 mL of water in a 1-L volumetric flask, add 21 mL of 1 N sodium hydroxide and 10 mL of 0.1 M magnesium chloride, dilute with water to volume, and mix.
Alkaline phosphatase solution
Transfer 95 ± 5 mg of alkaline phosphatase enzyme to a 50-mL volumetric flask, dissolve by adding pH 9 Buffer with magnesium to volume, and mix. Prepare this solution fresh daily.
Standard solution
Weigh 15 mg of
USP Dexamethasone RS into a 5-mL volumetric flask. Dissolve in and dilute with ethyl acetate to volume.
[NOTESonication may be required to ensure dissolution.
]
Test solution
Weigh 20 mg of Dexamethasone Sodium Phosphate into a 15-mL centrifuge tube. Add 5.0 mL of Alkaline phosphatase solution , shake vigorously, and allow to stand for 30 minutes. Add 5.0 mL of ethyl acetate, shake vigorously, centrifuge, and use the upper, ethyl acetate layer.
Procedure
Apply 10-µL portions of the
Test solution and the
Standard solution to a thin-layer chromatographic plate (see
Chromatography 621) coated with a 0.25-mm layer of chromatographic silica gel mixture. Develop the chromatogram in a mobile phase consisting of a mixture of chloroform, methanol, and water (180:15:1) to a distance of three-fourths of the length of the plate. Air-dry the plate and observe under short-wavelength UV light: the
RF value of the principal spot obtained from the
Test solution corresponds to that obtained from the
Standard solution.
B:
The residue from the ignition of it meets the requirements of the tests for
Phosphate 191 and for
Sodium 191.
Specific rotation 781S:
between +74
and +82
, calculated on the water-free and alcohol-free basis.
Test solution:
10 mg per mL, in water.
pH 791:
between 7.5 and 10.5, in a solution (1 in 100).
Water, Method I 921
Determine the water content. The sum of the percentages of water content, and alcohol content, determined as directed in the test for
Alcohol, does not exceed 16.0%.
Limit of phosphate ions
Standard phosphate solution
Dissolve 143.3 mg of dried monobasic potassium phosphate, KH2PO4, in water to make 1000.0 mL. This solution contains the equivalent of 0.10 mg of phosphate (PO4) in each mL.
Phosphate reagent A
Dissolve 5 g of ammonium molybdate in 1 N sulfuric acid to make 100 mL.
Phosphate reagent B
Dissolve 350 mg of p-methylaminophenol sulfate in 50 mL of water, add 20 g of sodium bisulfite, mix to dissolve, and dilute with water to 100 mL.
Procedure
Dissolve about 50 mg of Dexamethasone Sodium Phosphate, accurately weighed, in a mixture of 10 mL of water and 5 mL of 2 N sulfuric acid contained in a 25-mL volumetric flask, by warming if necessary. Add 1 mL each of Phosphate reagent A and Phosphate reagent B, dilute with water to 25 mL, mix, and allow to stand at room temperature for 30 minutes. Similarly and concomitantly, prepare a standard solution, using 5.0 mL of Standard phosphate solution instead of the 50 mg of the substance under test. Concomitantly determine the absorbances of both solutions in 1-cm cells at 730 nm, with a suitable spectrophotometer, using water as the blank. The absorbance of the test solution is not more than that of the standard solution. The limit is 1.0% of phosphate (PO4).
Limit of free dexamethasone
Mobile phase
Prepare a solution containing 7.5 mL of triethylamine in 1 L of water. Adjust by the addition of phosphoric acid to a pH of 5.4. Prepare a filtered and degassed mixture of 74 parts of the resulting solution with 26 parts of methanol. Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard solution
Dissolve an accurately weighed quantity of
USP Dexamethasone Phosphate RS in
Mobile phase to obtain a solution containing about 0.5 mg per mL. Prepare a second solution by dissolving an accurately weighed quantity of
USP Dexamethasone RS in a mixture of methanol and water (1:1) to obtain a solution containing about 50 µg per mL. Transfer 10.0 mL of the first solution and 1.0 mL of the second solution to a 100-mL volumetric flask. Dilute with
Mobile phase to volume, and mix to obtain a solution having known concentrations of 50 µg of
USP Dexamethasone Phosphate RS per mL and 0.5 µg of
USP Dexamethasone RS per mL.
Test solution
Transfer about 50 mg of Dexamethasone Sodium Phosphate, accurately weighed, to a 100-mL volumetric flask, dissolve in and dilute with Mobile phase to volume, and mix. Further dilute 5.0 mL of this solution with Mobile phase to 50.0 mL.
Chromatographic system
(see
Chromatography 621)The liquid chromatograph is equipped with a 254-nm detector and a 4.5-mm × 25-cm column that contains 5-µm packing L11. The flow rate is about 1.2 mL per minute. Chromatograph the
Standard solution and the
System suitability solution, record the peak responses as directed for
Procedure, and determine the chromatographic characteristics from chromatograms obtained from the
System Suitability: the column efficiency determined from the analyte peak is not less than 900 theoretical plates; the tailing factor for the analyte peak is not more than 1.6; the resolution,
R, between dexamethasone phosphate and dexamethasone is not less than 1.8; and the relative standard deviation for replicate injections is not more than 1.0%.
Procedure
Separately inject equal volumes (about 20 µL) of the
Standard solution and the
Test solution into the chromatograph, record the chromatograms, and measure the responses for the dexamethasone peaks. Calculate the quantity, in µg, of dexamethasone (C
22H
29FO
5) in the portion of Dexamethasone Sodium Phosphate taken by the formula:
1000C(rU / rS),
in which
C is the concentration, in µg per mL, of
USP Dexamethasone RS in the
Standard solution; and
rU and
rS are the peak responses obtained from the
Test solution and the
Standard solution, respectively: not more than 1.0% is found.
Chromatographic purity
Acetate buffer
Dissolve 7 g of ammonium acetate in 1 L of water, adjust with glacial acetic acid to a pH of 4.0, and mix.
Solution A
Prepare a filtered and degassed mixture of methanol, water, and
Acetate buffer (7:7:6). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Solution B
Prepare a filtered and degassed mixture of methanol and
Acetate buffer (7:3). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Test solution
Transfer about 25 mg of Dexamethasone Sodium Phosphate, accurately weighed, to a 25-mL volumetic flask, dissolve in and dilute with Solution A 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 packing L7. The flow rate is about 1 mL per minute. The column temperature is maintained at 40
. The chromatograph is programmed as follows.
Time (minutes) |
Solution A
(%) |
Solution B
(%) |
Elution |
0 |
90 |
10 |
equilibration |
03.5 |
90 |
10 |
isocratic |
3.523.5 |
90®60 |
10®40 |
linear gradient |
23.534.5 |
60®5 |
40®95 |
linear gradient |
34.559.5 |
5 |
95 |
isocratic |
59.560 |
5®90 |
95®10 |
linear gradient |
Chromatograph the
Test solution, and record the peak responses as directed for
Procedure: the resolution between the major peak and the nearest impurity is not less than 1.0; and the relative standard deviation for replicate injections is not more than 4.0%.
Procedure
Separately inject equal volumes (about 15 µL) of the
Test solution into the chromatograph, record the chromatogram, and measure the peak responses. Calculate the percentage of each impurity in the portion of Dexamethasone Sodium Phosphate taken by the formula:
100(ri / rs),
in which
ri is the peak response for each impurity; and
rs is the sum of the responses of all peaks: not more than 1.0% of any individual impurity is found, and not more than 2.0% of total impurities is found.
Organic volatile impurities, Method IV 467:
meets the requirements.
Alcohol content, Method II 611
Proceed as directed in the chapter except to use column packing S8 and to use the following modifications.
Internal standard solution
Pipet 1 mL of isopropyl alcohol into a 100-mL volumetric flask, add water to volume, and mix.
Standard stock solution
Prepare a solution of alcohol in water (1 in 50). Determine the specific gravity at 25
(see
Specific 841), and obtain the percentage of C
2H
5OH by reference to the
Alcoholometric Table in the section
Reference Tables.
Standard solution
Into a 10-mL volumetric flask pipet 4 mL of Standard stock solution and 5 mL of Internal standard solution, add water to volume, and mix. Inject 2 µL of this solution into the gas chromatograph.
Test solution
Transfer about 500 mg of Dexamethasone Sodium Phosphate, accurately weighed, into a 10-mL volumetric flask. Pipet 5 mL of Internal standard solution into the flask, and mix to dissolve. Add water to volume, and mix. Inject 2 µL of this solution into the gas chromatograph.
Calculation
Calculate the percentage of alcohol in the Dexamethasone Sodium Phosphate taken by the formula:
4(S / W)(Z / Y),
in which
S is the percentage of alcohol in the
Standard stock solution; W is the weight, in g, of Dexamethasone Sodium Phosphate used in the
Test solution; and
Y and
Z are the ratios of the alcohol peak heights to the internal standard peak heights for the
Standard solution and the
Test solution, respectively. The content of C
2H
5OH is not more than 8.0%.
Assay
Buffer solution
Dissolve 7.0 g of ammonium acetate in 1 L of water, adjust with glacial acetic acid to a pH of 4.00 ± 0.05, and mix.
Solution A
Prepare a filtered and degassed mixture of methanol, water, and Buffer solution (350:350:300).
Solution B
Prepare a filtered and degassed mixture of methanol and Buffer solution (700:300).
Mobile phase
Use variable mixtures of
Solution A and
Solution B as directed for
Chromatographic system. Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard preparation
Dissolve an accurately weighed quantity of
USP Dexamethasone Phosphate RS in
Solution A to obtain a solution having a known concentration of about 0.92 mg per mL.
Assay preparation
Dissolve an accurately weighed quantity of Dexamethasone Sodium Phosphate in Solution A, and mix to obtain a solution having a concentration of about 1.0 mg per mL.
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 L7. The column temperature is maintained at about 40
. The flow rate is about 1.0 mL per minute. The chromatograph is programmed as follows.
Time (minutes) |
Solution A
(%) |
Solution B
(%) |
Elution |
0 |
90 |
10 |
equilibration |
03.5 |
90 |
10 |
isocratic |
3.524 |
90®60 |
10®40 |
linear gradient |
2435 |
60®5 |
40®95 |
linear gradient |
3560 |
5 |
95 |
isocratic |
6060.1 |
5®90 |
95®10 |
linear gradient |
60.165 |
90 |
10 |
isocratic |
Chromatograph the
Standard preparation, and record the peak responses as directed for
Procedure: the relative standard deviation is not more than 2.0%. Chromatograph the
Assay preparation, and record the peak responses as directed for
Procedure: the resolution,
R, between dexamethasone phosphate and the nearest impurity eluting after it is not less than 1.0.
Procedure
Separately inject equal volumes (about 15 µL) of the
Standard preparation and the
Assay preparation into the chromatograph, record the chromatograms, and measure the areas for the major peaks. Calculate the quantity, in mg, of C
22H
28FNa
2O
8P in the portion of Dexamethasone Sodium Phosphate taken by the formula:
(516.41 / 472.45)C(rU / rS),
in which 516.41 and 472.45 are the molecular weights of dexamethasone sodium phosphate and dexamethasone phosphate, respectively;
C is the concentration, in mg per mL, of
USP Dexamethasone Phosphate RS in the
Standard preparation; and
rU and
rS are the peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.