Mercaptopurine
C5H4N4S·H2O 170.19
C5H4N4S 152.18
6H-Purine-6-thione, 1,7-dihydro-, monohydrate;
Purine-6-thiol monohydrate
[6112-76-1].
Anhydrous[50-44-2].
Infrared Absorption 
197K
USP35
The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.USP35
Solution A:
0.77 g/L of ammonium acetate in water
Inorganic Impurities
Organic Impurities
USP35
Preserve in tight containers, protected from light. Store at room temperature.USP35
11
USP Didanosine Related Compound A RS 
Hypoxanthine.
C5H4N4O 136.11
USP35
(mer kap'' toe pure' een).
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C5H4N4S·H2O 170.19
C5H4N4S 152.18
6H-Purine-6-thione, 1,7-dihydro-, monohydrate;
Purine-6-thiol monohydrate
[6112-76-1].Anhydrous
[50-44-2].DEFINITION
Mercaptopurine contains NLT 97.0% and NMT 102.0% of C5H4N4S, calculated on the anhydrous basis.
IDENTIFICATION
Change to read:
• A. Infrared Absorption 197KUSP35
Change to read:
• B.
The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.USP35
ASSAY
Change to read:
• Procedure
Solution A:
0.77 g/L of ammonium acetate in water
Mobile phase:
Methanol and Solution A (25:75)
Standard stock solution:
0.2 mg/mL of USP Mercaptopurine RS in the mixture of methanol and water (1:1). Transfer USP Mercaptopurine RS into a suitable volumetric flask, and add methanol equivalent to 50% of the final volume. Shake mechanically to dissolve, and dilute with water to volume.
Standard solution:
0.02 mg/mL of USP Mercaptopurine RS in Mobile phase from the Standard stock solution
Sample stock solution:
Transfer 25 mg of Mercaptopurine into a 50-mL volumetric flask. Add 25 mL of methanol, shake mechanically for at least 45 min, and dilute with water to volume. Transfer 20 mL of this solution into a 25-mL volumetric flask, and dilute with Mobile phase to volume.
Sample solution:
0.02 mg/mL of Mercaptopurine in Mobile phase from the Sample stock solution
Chromatographic system
Mode:
LC
Detector:
UV 325 nm
Column:
4.6-mm × 15-cm; 5-µm packing L68
Flow rate:
1.0 mL/min
Injection size:
20 µL
System suitability
Sample:
Standard solution
Suitability requirements
Tailing factor:
NMT 2.0
Relative standard deviation:
NMT 2.0%
Analysis
USP35
Samples:
Standard solution and Sample solution
Calculate the percentage of mercaptopurine (C5H4N4S) in the portion of Mercaptopurine taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response from the Sample solution |
| rS | = | = peak response from the Standard solution |
| CS | = | = concentration of USP Mercaptopurine RS in the Standard solution (mg/mL) |
| CU | = | = concentration of Mercaptopurine in the Sample solution (mg/mL) |
USP35
Acceptance criteria:
97.0%–102.0% on the anhydrous basis
IMPURITIES
Add the following:
Inorganic Impurities
• Residue on Ignition 281:
NMT 0.1%USP35
281:
NMT 0.1%USP35
Add the following:
Organic Impurities
• Procedure
Solution A and Mobile phase:
Prepare as directed in the Assay.
Standard stock solution A:
Use the Standard stock solution from the Assay.
Standard stock solution B:
0.026 mg/mL of USP Didanosine Related Compound A RS in a mixture of methanol and water (1:1). Transfer USP Didanosine Related Compound A RS into a suitable volumetric flask, and add methanol equivalent to 50% of the final volume. Shake mechanically to dissolve, and dilute with water to volume.
System suitability solution:
4.0 µg/mL of USP Mercaptopurine RS and 0.1 µg/mL of USP Didanosine Related Compound A RS in Mobile phase from Standard stock solution A and Standard stock solution B
Standard solution:
4.0 µg/mL of USP Mercaptopurine RS in Mobile phase from Standard stock solution A
Sensitivity solution:
0.2 µg/mL of USP Mercaptopurine RS in Mobile phase from the Standard solution
Sample solution:
Transfer 20 mg of Mercaptopurine into a 50-mL volumetric flask. Add 12.5 mL of methanol, shake mechanically to dissolve, and dilute with water to volume. [Note—Inject the Sample solution within 1 h of preparation. ]
Chromatographic system
Mode:
LC
Detector:
UV 260 nm
Column:
4.6-mm × 15-cm; 5-µm packing L68
Flow rate:
1.0 mL/min
Injection size:
50 µL
Run time:
NLT 7 times the retention time of the mercaptopurine peak
System suitability
Samples:
System suitability solution, Standard solution, and Sensitivity solution
Suitability requirements
Resolution:
NLT 3.0 between the mercaptopurine and didanosine related compound A peaks, System suitability solution
Tailing factor:
NMT 2.0, Standard solution
Signal-to-noise ratio:
NLT 10, Sensitivity solution
Relative standard deviation:
NMT 2.0% for the mercaptopurine peak, System suitability solution
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of each impurity in the portion of Mercaptopurine taken:
Result = (rU/rS) × (CS/CU) × (1/F) × 100
| rU | = | = peak response of each impurity from the Sample solution |
| rS | = | = peak response of mercaptopurine from the Standard solution |
| CS | = | = concentration of USP Mercaptopurine RS in the Standard solution (mg/mL) |
| CU | = | = concentration of Mercaptopurine in the Sample solution (mg/mL) |
| F | = | = relative response factor for each individual impurity (see Table 1) |
Acceptance criteria:
See Table 1. [Note—Disregard any impurity peak less than 0.05%. ]
Table 1
| Name | Relative Retention Time |
Relative Response Factor |
Acceptance Criteria, NMT (%) |
|---|---|---|---|
| Didanosine related compound Aa |
0.80 | 7.1 | 0.15 |
| Mercaptopurine | 1.00 | — | — |
| Mercaptopurine disulfideb | 5.14 | 2.9 | 0.15 |
| Any unspecified impurity |
— | 1.0 | 0.10 |
| Total impurities | — | — | 0.5 |
|
a
Hypoxanthine.
b
1,2-Di(9H-purin-6-yl)disulfane.
|
|||
USP35SPECIFIC TESTS
• Phosphorus
Standard phosphate solution:
43.96 µg/mL of dried monobasic potassium phosphate (equivalent to 10 µg of P)
Standard solution:
Add 2 mL of Standard phosphate solution to a 25-mL volumetric flask, add 1 mL of 15 N sulfuric acid, 0.5 mL of nitric acid, 0.75 mL of ammonium molybdate TS, and 1 mL of aminonaphtholsulfonic acid TS, then dilute with water to volume, and mix. Allow to stand for 5 min.
Sample solution:
Digest 200 mg with 2 mL of 15 N sulfuric acid in a large test tube, periodically adding nitric acid, dropwise and with caution. Continue heating until practically all of the liquid has evaporated and the residue is colorless. Transfer the residue, with the aid of small portions of water, to a 25-mL volumetric flask. Add 1 mL of 15 N sulfuric acid, 0.5 mL of nitric acid, 0.75 mL of ammonium molybdate TS, and 1 mL of aminonaphtholsulfonic acid TS, then dilute with water to volume. Allow to stand for 5 min.
Blank:
Add 2 mL of 15 N sulfuric acid to a large test tube, periodically adding nitric acid, dropwise and with caution. Continue heating until practically all of the liquid has evaporated and the residue is colorless. Transfer the residue, with the aid of small portions of water, to a 25-mL volumetric flask, add 1 mL of 15 N sulfuric acid, 0.5 mL of nitric acid, 0.75 mL of ammonium molybdate TS, and 1 mL of aminonaphtholsulfonic acid TS, then dilute with water to volume. Allow to stand for 5 min.
Instrumental conditions
Mode:
UV-Vis
Analytical wavelength:
750 nm
Analysis
Samples:
Standard solution, Sample solution, and Blank
Acceptance criteria:
The absorbance of the Sample solution is NMT that of the Standard solution (NMT 100 ppm).
• Water Determination, Method I 921:
NMT 12.0%
921:
NMT 12.0%
ADDITIONAL REQUIREMENTS
Change to read:
• Packaging and Storage:
Preserve in tight containers, protected from light. Store at room temperature.USP35
Change to read:
• USP Reference Standards 11
USP Didanosine Related Compound A RS ') + '&img=../../images/v35300/c11rs2106.gif',600,500);)
Hypoxanthine.
C5H4N4O 136.11
USP35
USP Mercaptopurine RS
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Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Feiwen Mao, M.S.
Senior Scientific Liaison 1-301-816-8320 |
(SM32010) Monographs - Small Molecules 3 |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 3810
Pharmacopeial Forum: Volume No. 36(6) Page 1552