Alanine

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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C₃H₇NO₂   89.09

L-alanine     CAS RN®: 56-41-7; UNII: OF5P57N2ZX.

1 DEFINITION

Alanine contains NLT 98.5% and NMT 101.5% of L-alanine (C₃H₇NO₂), calculated on the dried basis.

2 IDENTIFICATION

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197K

3 ASSAY

3.1 Procedure

Sample: 80 mg of Alanine Titrimetric system

(See Titrimetry 〈541〉.) Mode: Direct titration

Titrant: 0.1 N perchloric acid VS

Endpoint detection: Potentiometric

Blank: 3 mL of formic acid in 50 mL of glacial acetic acid

Analysis: Dissolve the Sample in a mixture of 3 mL of formic acid and 50 mL of glacial acetic acid, and titrate with Titrant. Calculate the

percentage of l-alanine (C₃H₇NO₂) in the portion of Alanine taken:

Result = [(V_S − V_B ) × N × F × 100]/W

V_S = Titrant volume consumed by the Sample (mL)

V_B = Titrant volume consumed by the Blank (mL)

N = actual normality of the Titrant (mEq/mL)

F = equivalency factor, 89.09 mg/mEq

W = Sample weight (mg)

Acceptance criteria: 98.5%–101.5% on the dried basis

4 IMPURITIES

Residue on Ignition 〈281〉: NMT 0.15%

Chloride and Sulfate, Chloride〈221〉

Standard solution: 0.70 mL of 0.020 N hydrochloric acid

Sample: 1.0 g of Alanine

Acceptance criteria: NMT 0.05%

Chloride and Sulfate, Sulfate〈221〉

Standard solution: 0.30 mL of 0.020 N sulfuric acid

Sample: 1.0 g of Alanine

Acceptance criteria: NMT 0.03%

Change to read:

Iron 〈241〉, Procedures, Procedure 1 (CN 1-Jun-2023) : NMT 30 ppm

Related Compounds

Mobile phase: 0.008 N sulfuric acid solution

System suitability solution: A mixture of 0.05 mg/mL of USP Fumaric Acid RS, 0.05 mg/mL of USP Maleic Acid RS, and 3 mg/mL of USP Malic Acid RS in water

Maleic acid standard solution: 0.05 mg/mL of USP Maleic Acid RS in water

Malic acid standard solution: 3 mg/mL of USP Malic Acid RS in water

Fumaric acid standard solution: 0.05 mg/mL of USP Fumaric Acid RS in water

Sample solution: 100 mg/mL of Alanine in water

4.1 Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 214 nm

Column: 7.8-mm × 30-cm; 9-µm packing L17

Column temperature: 30°

Flow rate: 0.6 mL/min

Injection volume: 10 µL

4.2 System suitability

Sample: System suitability solution

[Note—See Table 1 for the relative retention times.]

Suitability requirements

Resolution: NLT 1.5 between maleic acid and malic acid

Relative standard deviation: NMT 5.0% for each of fumaric acid, maleic acid, and malic acid

4.3 Analysis

Samples: Standard solutions and Sample solution

Calculate the percentage of each specied acid in the portion of Alanine taken:

Result = (r_U /r_S ) × (C_S /C_U) × 100

r_U = peak response of maleic acid, malic acid, or fumaric acid from the Sample solution

r_S = peak response of maleic acid, malic acid, or fumaric acid from the corresponding Standard solution  

C_S = concentration of USP Maleic Acid RS, USP Malic Acid RS, or USP Fumaric Acid RS in the corresponding Standard solution (mg/mL)

C_U = concentration of Alanine in the Sample solution (mg/mL)  

Calculate the percentage of any unspecied impurity in the portion of Alanine taken:

Result = (r_U /r_S ) × (C_S /C_U) × 100

r_U = peak response of any unspecied impurity from the Sample solution

r_S = peak response of fumaric acid from the Fumaric acid standard solution  

C_S = concentration of USP Fumaric Acid RS in the Fumaric acid standard solution (mg/mL)

C_U = concentration of Alanine in the Sample solution (mg/mL)

Acceptance criteria: See Table 1.

Table 1

5 SPECIFIC TESTS

Optical Rotation, Specific Rotation〈781S〉

Sample solution: 100 mg/mL in 6 N hydrochloric acid

Acceptance criteria: +13.7° to +15.1°

pH 〈791〉: 5.5–7.0, in a solution (1 in 20)

Loss on Drying 〈731〉

Analysis: Dry at 105° for 3 h.

Acceptance criteria: NMT 0.2%

6 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in tight containers, and store at controlled room temperature.

USP Reference Standards 〈11〉

USP l-Alanine RS

USP Fumaric Acid RS

USP Maleic Acid RS

USP Malic Acid RS