Cladribine

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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C₁₀H₁₂ClN₅O₃ 285.69 

Adenosine, 2-chloro-2′-deoxy-; 

2-Chloro-2′-deoxyadenosine CAS RN®: 4291-63-8; UNII: 47M74X9YT5. 

1 DEFINITION 

Cladribine contains NLT 98.0% and NMT 102.0% of cladribine (C₁₀H₁₂ClN₅O₃), calculated on the anhydrous basis. 

2 IDENTIFICATION 

Change to read: 

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

Procedure 

Buffer: Dissolve 9.96 g of triethylamine phosphate in 500 mL of water. Add another 500 mL of water, and adjust with potassium hydroxide to a pH of 6.1. 

Diluent: Methanol and water (10:90) 

Mobile phase: Methanol and Buffer (22:78) 

System suitability solution: 0.02 mg/mL each of USP Cladribine RS and USP Cladribine Related Compound A RS in Diluent Standard solution: 0.5 mg/mL of USP Cladribine RS in Diluent 

Sample solution: 0.5 mg/mL of Cladribine in Diluent 

Chromatographic system 

(See Chromatography 〈621〉, System Suitability.) 

Mode: LC 

Detector: UV 265 nm 

Column: 4.6-mm × 15-cm; packing L1 

Flow rate: 1.0 mL/min 

Injection volume: 10 µL 

System suitability 

Samples: System suitability solution and Standard solution 

Suitability requirements 

Resolution: NLT 1.5 between the cladribine and cladribine related compound A peaks, System suitability solution 

Tailing factor: NMT 2.0 for the cladribine peak, System suitability solution 

Relative standard deviation: NMT 2.0%, Standard solution 

Analysis 

Samples: Standard solution and Sample solution 

Calculate the percentage of cladribine (C₁₀H₁₂ClN₅O₃) in the portion of Cladribine 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 Cladribine RS in the Standard solution (mg/mL) 

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

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

4 IMPURITIES 

Residue on Ignition 〈281〉: NMT 0.1% 

Organic Impurities 

Buffer, Diluent, Mobile phase, System suitability solution, Sample solution, and Chromatographic system: Proceed as directed in the Assay. System suitability 

Sample: System suitability solution 

Suitability requirements 

Resolution: NLT 1.5 between cladribine and cladribine related compound A 

Tailing factor: NMT 2.0 for the cladribine peak 

Analysis 

Sample: Sample solution 

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

Result = (r_U/r_T)  × 100 

r_U = peak response of each individual impurity  

r_T = sum of the responses of all the peaks

Acceptance criteria: See Table 1. 

Table 1 

5 SPECIFIC TESTS 

Optical Rotation, Specific Rotation〈781S〉 

Sample solution: 10 mg/mL in dimethylformamide 

Acceptance criteria: −17.0° to −21.0° 

Water Determination, Method I 〈921〉: NMT 4.0% 

ADDITIONAL REQUIREMENTS 

Packaging and Storage: Preserve in well-closed containers, and protect from light. Store between 2° and 8°. 

USP Reference Standards 〈11〉 

USP Cladribine RS 

USP Cladribine Related Compound A RS 

2-Methoxy-2’-deoxyadenosine. 

C₁₁H₁₅N₅O₄ 281.27