Methyltestosterone

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

Methyltestosterone contains NLT 97.0% and NMT 103.0% of methyltestosterone (C₂₀H₃₀O₂), calculated on the dried basis.

2 IDENTIFICATION

Change to read:

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: ▲197A or▲ (USP 1-May-2024) 197K

B. The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.

3 ASSAY

Change to read:

Procedure

▲[Note—Protect solutions containing methyltestosterone from light.]▲ (USP 1-May-2024)

Mobile phase: Acetonitrile and water (55:45)

Standard stock solution: 0.25 mg/mL of USP Methyltestosterone RS in methanol

Standard solution: 0.02 mg/mL of USP Methyltestosterone RS from the Standard stock solution in Mobile phase

System suitability stock solution: 0.25 mg/mL of USP Testosterone RS in methanol

System suitability solution: ▲0.02 mg/mL each of USP Methyltestosterone RS and USP Testosterone RS in Mobile phase prepared as follows.▲ (USP 1-May-2024)

Dilute 4 mL of the System suitability stock solution with the Standard solution to 50 mL.

Sample stock solution: 0.50 mg/mL of Methyltestosterone in methanol

Sample solution: 0.02 mg/mL of Methyltestosterone from the Sample stock solution in Mobile phase

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 241 nm

Column: 4-mm × 25-cm; ▲5-µm▲ (USP 1-May-2024) packing L1

Flow rate: 1 mL/min

Injection volume: 50 µL

System suitability

Samples: Standard solution and System suitability solution

[Note—The relative retention times for testosterone and methyltestosterone are about 0.8 and 1.0, respectively.]

Suitability requirements

Resolution: NLT 2.0 between testosterone and methyltestosterone, System suitability solution

▲ (USP 1-May-2024)

Tailing factor: NMT 2.7, Standard solution

Relative standard deviation: NMT ▲1.10%,▲ (USP 1-May-2024) Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of methyltestosterone (C₂₀H₃₀O₂) in the portion of Methyltestosterone taken:

Result = (rᵤ / rₛ) × (Cₛ / Cᵤ) × 100

rᵤ = peak response of methyltestosterone from the Sample solution

rₛ = peak response of methyltestosterone from the Standard solution

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

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

Acceptance criteria: 97.0%–103.0% on the dried basis

4 IMPURITIES

Change to read:

Organic Impurities

▲[Note—Protect solutions containing methyltestosterone from light.]▲ (USP 1-May-2024)

Solution A: Methanol and water (55:45)

Solution B: Methanol

Mobile phase: See Table 1.

Table 1

System suitability solution: ▲0.5 mg/mL of USP Methyltestosterone RS and 0.005 mg/mL of USP Testosterone RS in methanol

Sensitivity solution: 0.25 µg/mL of USP Methyltestosterone RS in methanol

Standard solution: 0.5 µg/mL of USP Methyltestosterone RS in methanol▲ (USP 1-May-2024)

Sample solution: 0.5 mg/mL of Methyltestosterone in methanol

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 254 nm

Column: 4.6-mm × 25-cm; 5-µm packing L1

▲Temperatures

Autosampler: 5°

Column: 30°▲ (USP 1-May-2024)

Flow rate: 1 mL/min

Injection volume: ▲10 µL▲ (USP 1-May-2024)

System suitability

Samples: ▲ (USP 1-May-2024) System suitability solution, ▲Sensitivity solution, and Standard solution

[Note—The relative retention times in Table 2 are provided as information that could aid in peak assignment.]

Table 2

ᵃ Androst-4-en-3,17-dione.

ᵇ 17α-Hydroxy-17-methylandrost-4-en-3-one.

▲ (USP 1-May-2024)

Suitability requirements

▲Resolution: NLT 1.5 between testosterone and methyltestosterone, System suitability solution▲ (USP 1-May-2024)

Relative standard deviation: NMT ▲5.0%, Standard solution▲ (USP 1-May-2024)

Signal-to-noise ratio: NLT ▲10, Sensitivity solution▲ (USP 1-May-2024)

Analysis

Samples: ▲Standard solution and▲ (USP 1-May-2024) Sample solution

▲Calculate the percentage of any specified and unspecified impurity in the portion of Methyltestosterone taken:

Result = (rᵤ / rₛ) × (Cₛ / Cᵤ) × (1 / F) × 100

rᵤ = peak response of any specified or unspecified impurity from the Sample solution

rₛ = peak response of Methyltestosterone from the Standard solution

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

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

F = relative response factor (see Table 3)

▲ (USP 1-May-2024)

Acceptance criteria: ▲See Table 3. The reporting threshold is 0.05%.

Table 3

▲ (USP 1-May-2024)

5 SPECIFIC TESTS

Optical Rotation 〈781S〉, Procedures, Specific Rotation

Sample solution: 10 mg/mL of Methyltestosterone in alcohol

Acceptance criteria: +79° to +85°

Loss on Drying 〈731〉

Analysis: Dry at 105° for 4 h.

Acceptance criteria: NMT 2.0%

6 ADDITIONAL REQUIREMENTS

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

USP Reference Standards 〈11〉

USP Methyltestosterone RS

USP Testosterone RS