Dexamethasone Sodium Phosphate
(dex'' a meth' a sone soe' dee um fos' fate).
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C22H28FNa2O8P 516.41

Pregna-1,4-diene-3,20-dione, 9-fluoro-11,17-dihydroxy-16-methyl-21-(phosphonooxy)-, disodium salt, (11,16)-.
9-Fluoro-11,17,21-trihydroxy-16-methylpregna-1,4-diene-3,20-dione 21-(dihydrogen phosphate) disodium salt [2392-39-4].
» Dexamethasone Sodium Phosphate contains not less than 97.0 percent and not more than 102.0 percent of C22H28FNa2O8P, calculated on the water-free and alcohol-free basis.
Packaging and storage— Preserve in tight containers.
USP Reference standards 11—
USP Dexamethasone RS Click to View Structure
USP Dexamethasone Phosphate RS Click to View Structure
Identification—
A: pH 9 Buffer with magnesium—Mix 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. [note—Sonication 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.
System suitability solution— Prepare a solution in Mobile phase containing in each mL 0.05 mg of USP Dexamethasone Phosphate RS and 0.02 mg of USP Dexamethasone RS.
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 (C22H29FO5) 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).
Mobile phase— Use variable mixtures of Solution A and Solution B as directed for Chromatographic system.
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
0–3.5 90 10 isocratic
3.5–23.5 90®60 10®40 linear gradient
23.5–34.5 60®5 40®95 linear gradient
34.5–59.5 5 95 isocratic
59.5–60 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.
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 C2H5OH 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 C2H5OH 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
0–3.5 90 10 isocratic
3.5–24 90®60 10®40 linear gradient
24–35 60®5 40®95 linear gradient
35–60 5 95 isocratic
60–60.1 5®90 95®10 linear gradient
60.1–65 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 C22H28FNa2O8P 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.
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Monograph Domenick Vicchio, Ph.D.
Senior Scientific Liaison
1-301-998-6828		 (SM42010) Monographs - Small Molecules 4
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USP35–NF30 Page 2842