Pyridoxine Hydrochloride

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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1 DEFINITION

Pyridoxine Hydrochloride contains NLT 98.0% and NMT 102.0% of pyridoxine hydrochloride (C₈H₁₁NO₃ · HCl), calculated on the dried basis.

2 IDENTIFICATION

Change to read:

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

B. Identification Tests—General, Chloride 〈191〉: Meets the requirements

3 ASSAY

Procedure

Mobile phase: Mix 10 mL of glacial acetic acid, 0.6 g of sodium 1-hexanesulfonate, and 700 mL of water in a 1000-mL volumetric flask. Adjust with glacial acetic acid or 1 N sodium hydroxide to a pH of 3.0. Add 235 mL of methanol, and dilute with water to volume.

Internal standard solution: 5 mg/mL of p-hydroxybenzoic acid in Mobile phase

Standard solution: Prepare a 0.5-mg/mL solution of USP Pyridoxine Hydrochloride RS in Mobile phase. Transfer 10.0 mL of this solution and 1.0 mL of Internal standard solution to a 100-mL volumetric flask, and dilute with Mobile phase to volume.

Sample solution: Prepare a 0.5-mg/mL solution of Pyridoxine Hydrochloride in Mobile phase. Transfer 10.0 mL of this solution and 1.0 mL of Internal standard solution to a 100-mL volumetric flask, and dilute with Mobile phase to volume.

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 280 nm

Column: 4.6-mm × 25-cm; packing L1

Flow rate: 1.5 mL/min

Injection size: 20 µL

System suitability

Sample: Standard solution

[Note—The relative retention times for pyridoxine and p-hydroxybenzoic acid are about 0.7 and 1.0, respectively.]

Suitability requirements

Resolution: NLT 2.5 between pyridoxine and p-hydroxybenzoic acid

Relative standard deviation: NMT 3.0% for the ratios of the pyridoxine peak area response to the internal standard peak area response

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of pyridoxine hydrochloride (C₈H₁₁NO₃ · HCl) in the portion of Pyridoxine Hydrochloride taken:

Result = (Rᵤ/Rₛ) × (Cₛ/Cᵤ) × 100

Rᵤ = internal standard ratio (peak response of pyridoxine/peak response of the internal standard) from the Sample solution

Rₛ = internal standard ratio (peak response of pyridoxine/peak response of the internal standard) from the Standard solution

Cₛ = concentration of USP Pyridoxine Hydrochloride RS in the Standard solution (mg/mL)

Cᵤ = concentration of Pyridoxine Hydrochloride in the Sample solution (mg/mL)

Acceptance criteria: 98.0%–102.0% on the dried basis

4 IMPURITIES

Residue on Ignition 〈281〉: NMT 0.1%

5 SPECIFIC TESTS

Content of Chloride

Sample: 500 mg of Pyridoxine Hydrochloride

Blank: 50 mL of methanol

Titrimetric system

(See Titrimetry 〈541〉.)

Mode: Direct titration

Titrant: 0.1 N silver nitrate VS

Endpoint detection: Visual

Analysis: Dissolve the Sample in 50 mL of methanol. Add 5 mL of glacial acetic acid and 2–3 drops of eosin Y TS. Titrate with the Titrant. Perform a Blank determination.

Calculate the percentage of chloride (Cl) in the Sample taken:

Result = {[(V − Vᵦ) × N × F]/W} × 100

V = Titrant volume consumed by the Sample (mL)

Vᵦ = Titrant volume consumed by the Blank (mL)

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

F = equivalency factor, 35.45 mg/mEq

W = Sample weight (mg)

Acceptance criteria: 16.9%–17.6% on the dried basis

Loss on Drying 〈731〉: Dry a sample in vacuum over silica gel for 4 h: it loses NMT 0.5% of its weight.

6 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in tight, light-resistant containers.

USP Reference Standards 〈11〉

USP Pyridoxine Hydrochloride RS