Adenosine

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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C₁₀H₁₃N₅O₄       267.24

6-Amino-9-β-_D-ribofuranosyl-9H-purine;

9-β-_D-Ribofuranosyladenine     CAS RN^®: 58-61-7; UNII: K72T3FS567.

1 DEFINITION

Adenosine contains NLT 98.0% and NMT 102.0% of adenosine (C₁₀H₁₃N₅O₄), calculated on the dried basis.

2 IDENTIFICATION

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

B. The retention times of the major peaks of the Sample solution correspond to those of the Standard solution, as obtained in the Assay.

3 ASSAY

Change to read:

3.1 Procedure

Buffer: 6.8 g/L of potassium hydrogen sulfate and 3.4 g/L of tetrabutylammonium hydrogen sulfate in a solution prepared as follows. Transfer suitable quantities of potassium hydrogen sulfate and tetrabutylammonium (ERR 1-Oct-2024) hydrogen sulfate to an appropriate volumetric ask, and dissolve in 90% of the ask volume of water. Adjust with 2 N potassium hydroxide to a pH of 6.5, and dilute with water to volume.

Mobile phase: Buffer and water (60:40)

System suitability solution: 4 µg/mL each of USP Adenine RS and inosine in Mobile phase

Standard solution: 0.2 mg/mL of USP Adenosine RS in Mobile phase

Sample solution: 0.2 mg/mL of Adenosine in Mobile phase

3.2 Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 254 nm

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

Flow rate: 1.5 mL/min

Injection volume: 20 µL

Run time: NLT 1.5 times the retention time of the adenosine peak

3.3 System suitability

Samples: System suitability solution and Standard solution

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

Suitability requirements

Resolution: NLT 1.5 between adenine and inosine, System suitability solution

Tailing factor: NMT 2.0, Standard solution

Relative standard deviation: NMT 0.7%, Standard solution

3.4 Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of adenosine (C₁₀H₁₃N₅O₄) in the portion of Adenosine taken:

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

r_U = peak response from the Sample solution

r_S = peak response from the Standard solution  

C_S = concentration of USP Adenosine RS in the Standard solution (mg/mL)

C_U = concentration of Adenosine 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%

4.1 Organic Impurities

Buffer, Mobile phase, System suitability solution, and Chromatographic system: Proceed as directed in the Assay. Standard solution: 0.001 mg/mL of USP Adenosine RS in Mobile phase

Sample solution: 1 mg/mL of Adenosine in Mobile phase

4.2 System suitability

Samples: System suitability solution and Standard solution

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

4.3 Suitability requirements

Resolution: NLT 1.5 between adenine and inosine, System suitability solution

Relative standard deviation: NMT 5%, Standard solution

4.4 Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of each impurity in the portion of Adenosine taken:

Result = (r_U /r_S ) × (C_S /C_U) × (1/F) × 100

r_U = peak response from the Sample solution  

r_S = peak response from the Standard solution

C_S = concentration of USP Adenosine RS in the Standard solution (mg/mL)

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

F = relative response factor (see Table 1)

Acceptance criteria: See Table 1. Disregard peaks that are less than 0.05% of the adenosine peak.

Table 1

^a 1-β-d-Ribofuranosylpyrimidine-2,4(1H,3H)-dione.

^b 9-β-d-Ribofuranosylpurine-6(1H)-one.

^c 2-Amino-9-β-d-ribofuranosylpurine-6(1H)-one.

5 SPECIFIC TESTS

Optical Rotation, Specific Rotation 〈781S〉: −68° to −72°

Test solution: 20 mg/mL in sodium hydroxide solution (1 in 20), determined on a sample previously dried at 105° for 2 h

Loss on Drying 〈731〉

Analysis: Dry a sample at 105° for 2 h.

Acceptance criteria: NMT 0.5%

6 ADDITIONAL REQUIREMENTS

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

USP Reference Standards 〈11〉

USP Adenine RS

USP Adenosine RS