Acetohydroxamic Acid

This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition

Issued and maintained by the United States Pharmacopeial Convention (USP)

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C₂H₅NO₂ 75.07

N-Acetyl hydroxyacetamide;

Acetohydroxamic acid CAS RN®: 546-88-3; UNII: 4RZ82L2GY5.

1 DEFINITION

Acetohydroxamic Acid, dried over phosphorus pentoxide for 16 h, contains NLT 98.0% and NMT 101.0% of acetohydroxamic acid (C₂H₅NO₂).

2 IDENTIFICATION

Change to read:

 A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197K (CN 1-May-2020)

B.

Sample solution: 20 mg/mL in water

Analysis: To 10 mL of the Sample solution add 2 drops of potassium permanganate TS.

Acceptance criteria: The pink color of the permanganate disappears.

3 ASSAY

Procedure

Ferric chloride solution: 20 mg/mL of ferric chloride in 0.1 N hydrochloric acid

Standard solution: 500 μg/mL of USP Acetohydroxamic Acid RS in 0.1 N hydrochloric acid

Sample solution: 500 μg/mL of Acetohydroxamic Acid, previously dried, in 0.1 N hydrochloric acid

Blank: 0.1 N hydrochloric acid

Analysis

Samples: Standard solutions, Sample solution, and Blank

Transfer 10.0 mL each of the Standard solution, Sample solution, and Blank to separate 100-mL volumetric flasks. To each flask add 50 mL of 0.1 N hydrochloric acid and 10.0 mL of Ferric chloride solution, and dilute with 0.1 N hydrochloric acid to volume. Without delay, concomitantly determine the absorbances of the solutions at the wavelength of maximum absorbance at about 502 nm using the Blank to set the instrument.

Calculate the percentage of acetohydroxamic acid (C₂H₅NO₂) in the portion of Acetohydroxamic Acid taken:

Result = (A_u/A_s) × (C_s/C_u) × 100

A_u = absorbance of the Sample solution

A_s = absorbance of the Standard solution

C_s = concentration of USP Acetohydroxamic Acid RS in the Standard solution (μg/mL)

C_u = concentration of Acetohydroxamic Acid in the Sample solution (μg/mL)

Acceptance criteria: 98.0%–101.0% on the previously dried basis

4 IMPURITIES

Residue on Ignition 〈281〉: NMT 0.1%

Limit of Hydroxylamine

Buffer: 1.36 g/L of monobasic potassium phosphate in water, adjusted with 1 M potassium hydroxide to a pH of 7.4

Solution A: 1 mg/mL of pyridoxal 5-phosphate monohydrate in Buffer, prepared in a low-actinic flask fresh before use

Standard stock solution: 2.0 mg/mL of hydroxylamine hydrochloride in water

Standard solutions: Transfer 5.0, 10.0, and 15.0 mL of the Standard stock solution to separate 100-mL volumetric flasks, and dilute with water to volume.

Sample solution: Transfer 1500 mg of Acetohydroxamic Acid, previously dried, to a 100-mL beaker, and dissolve in a sufficient amount of water to cover the electrode of a calibrated pH meter (about 60 mL). While stirring, adjust with 0.05 M potassium hydroxide to a pH of 7.4.

Transfer the contents of the beaker, with the aid of small portions of water, to a 100-mL volumetric flask, and dilute with water to volume.

Blank: Water

Analysis

Samples: Standard solutions, Sample solution, and Blank

Transfer 2.0 mL of each Standard solution, the Sample solution, and Blank into separate 100-mL volumetric flasks. To each flask add 4.0mL of Solution A. After 8 min, accurately timed, dilute the contents of each flask with Buffer to volume.

Immediately determine the fluorescence intensities of the solutions from the Standard solutions and the Sample solution in a fluorometer at an excitation wavelength of 350 nm and an emission wavelength of 450 nm, setting the instrument to zero with the Blank. Determine the best-fit straight line from the fluorescence intensities of the three Standard solutions versus the hydroxylamine hydrochloride concentrations, in μg/mL. From the best-fit straight line, determine the concentration, in μg/mL, of hydroxylamine hydrochloride in the Sample solution.

Calculate the percentage of hydroxylamine in the portion of Acetohydroxamic Acid taken:

Result = (C_u/C) × (M_r1/M_r2) × 100

C_u= concentration of hydroxylamine hydrochloride in the Sample solution (mg/mL)

C = concentration of Acetohydroxamic Acid in the Sample solution (mg/mL)

M_r1 = molecular weight of hydroxylamine, 33.03

M_r2 = molecular weight of hydroxylamine hydrochloride, 69.50

Acceptance criteria: NMT 0.5%

5 SPECIFIC TESTS

Loss on Drying 〈731〉

Analysis: Dry a sample over phosphorus pentoxide for 16 h.

Acceptance criteria: NMT 1.0%

Completeness of Solution 〈641〉: A 1.0-g portion dissolves in 10 mL of water to yield a clear solution.•

Color of Solution

Sample solution: 200 mg/mL in water

Blank: Water

Instrumental conditions

Mode: UV-Vis

Analytical wavelength: Between 400 and 750 nm

Cell: 1 cm

Acceptance criteria: The absorbance is NMT 0.050.

6 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in tight containers, and store in a cool, dry place.

USP Reference Standards 〈11〉

USP Acetohydroxamic Acid RS