Mercaptopurine Tablets
DEFINITION
Mercaptopurine Tablets contain NLT 93.0% and NMT 110.0% of the labeled amount of mercaptopurine (C5H4N4S·H2O).
IDENTIFICATION
Change to read:
•  A. The UV absorption spectrum exhibits a maximum at 325 ± 2 nm, and the ratio A255/A325 does not exceed 0.09.
Sample:  5 µg/mL of mercaptopurine in a mixture of methanol and water (1:1), from the Sample solution in the AssayUSP35
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
Solution B:  Methanol and Solution A (5:95)
Solution C:  Methanol and Solution A (30:70)
Mobile phase:  Solution B and Solution C (80:20)
Diluent:  Methanol and water (1:1)
Standard solution:  0.25 mg/mL of USP Mercaptopurine RS in a 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.
Sample stock solution:  0.5 mg/mL of mercaptopurine in Diluent from NLT 5 Tablets. Place the Tablets into a suitable volumetric flask, add methanol equivalent to 50% of the final volume, and shake mechanically for a minimum of 30 min. Dilute with water to volume. Pass through a PVDF filter of 0.45-µm pore size, and discard the first 3 mL of filtrate.
Sample solution:  0.25 mg/mL of mercaptopurine in Diluent from the Sample stock solution
Chromatographic system 
Mode:  LC
Detector:  UV 260 nm
Column:  4.6-mm × 10-cm; 3-µm packing L1
Column temperature:  30
Flow rate:  1.0 mL/min
Injection size:  10 µL
System suitability 
Sample:  Standard solution
Suitability requirements 
Tailing factor:  NMT 2.0
Relative standard deviation:  NMT 2.0%
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of mercaptopurine (C5H4N4S·H2O) in the portion of Tablets taken:
Result = (rU/rS) × (CS/CU) × (Mr1/Mr2) × 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== nominal concentration of mercaptopurine in the Sample solution (mg/mL)
Mr1== molecular weight of mercaptopurine, 170.19
Mr2== molecular weight of anhydrous mercaptopurine, 152.18
USP35
Acceptance criteria:  93.0%–110.0%
PERFORMANCE TESTS
Change to read:
•  Dissolution 711
Test 1 
Medium:  0.1 N hydrochloric acid; 900 mL
Apparatus 2:  50 rpm
Time:  60 min
Mobile phase:  0.1% acetic acid in water
Standard solution:  USP Mercaptopurine RS in Medium
Sample solution:  Pass a portion of the solution under test through a suitable filter. Dilute with Medium to a concentration that is similar to the Standard solution, if necessary.
Chromatographic system 
Mode:  LC
Detector:  UV 230 nm
Column:  3.9-mm × 15-cm; packing L1
Flow rate:  2.5 mL/min
Injection size:  10 µL
System suitability 
Sample:  Standard solution
[Note—The retention time for mercaptopurine is NLT 4 min. ]
Suitability requirements 
Relative standard deviation:  NMT 2.0%
Analysis 
Samples:  Standard solution and Sample solution
Tolerances:  NLT 80% (Q) of the labeled amount of C5H4N4H2OUSP35 is dissolved.
Test 2:  If the product complies with this test, the labeling indicates that it meets USP Dissolution Test 2.
Medium, Apparatus 2, Chromatographic system, and Analysis:  Proceed as directed for Test 1.
Time:  120 min
Tolerances  NLT 80% (Q) of the labeled amount of C5H4N4S·H2O is dissolved.
•  Uniformity of Dosage Units 905: Meets the requirements
IMPURITIES
Add the following:
Organic Impurities 
•  Procedure
Solution A:  0.1% (v/v) formic acid in water
Solution B:  Methanol and Solution A (2:98)
Solution C:  Methanol and Solution A (1:1)
Mobile phase:  See Table 1.
Table 1
Time
(min)
Solution B
(%)
Solution C
(%)
0 100 0
8 100 0
20 0 100
25 0 100
27 100 0
30 100 0
Standard stock solution:  0.06 mg/mL of USP Mercaptopurine RS in Solution A. [Note—Use methanol equivalent to 2.5% of the final volume to help dissolve. ]
Standard solution:  1.2 µg/mL of USP Mercaptopurine RS in Solution B from the Standard stock solution
Sensitivity solution:  0.06 µg/mL of USP Mercaptopurine RS in Solution B from the Standard solution
Sample stock solution:  0.5 mg/mL of mercaptopurine in a mixture of methanol and Solution A (1:9) from NLT 5 Tablets. Place the Tablets into a suitable volumetric flask, add methanol equivalent to 10% of the final volume, and shake mechanically for a minimum of 30 min. Dilute with water to volume. Pass through a PVDF filter of 0.45-µm pore size, and discard the first 3 mL of filtrate.
Sample solution:  0.12 mg/mL of mercaptopurine in Solution A. Transfer 6.0 mL of the Sample stock solution into a 25-mL volumetric flask, and dilute with Solution A to volume. Pass through a PVDF filter of 0.45-µm pore size, and discard the first 5 mL of filtrate. [Note—Inject the Sample solution within 1 h of preparation. ]
Chromatographic system 
Mode:  LC
Detector:  UV 260 nm
Column:  4.6-mm × 10-cm; 3-µm packing L1
Temperature 
Column:  30
Sample:  4
Flow rate:  1.0 mL/min
Injection size:  50 µL
System suitability 
Samples:  Standard solution and Sensitivity solution
Suitability requirements 
Tailing factor:  NMT 2.0, Standard solution
Signal-to-noise ratio:  NLT 10, Sensitivity solution
Relative standard deviation:  NMT 2.0%, Standard solution
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of each impurity in the portion of Tablets 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== nominal concentration of mercaptopurine in the Sample solution (mg/mL)
F== relative response factor for each individual impurity (see Table 2)
Acceptance criteria 
Individual impurities:  See Table 2. [Note—Disregard any impurity peak less than 0.05%. ]
Table 2
Name Relative
Retention
Time
Relative
Response
Factor
Acceptance
Criteria,
NMT (%)
Didanosine related compound Aa 0.54 6.3 0.3
Mercaptopurine 1.00
Mercaptopurine disulfideb 2.90 4.4 0.4
Any unspecified
impurity
1.0 0.2
Total impurities 0.6
a  Hypoxanthine.
b  1,2-Di(9H-purin-6-yl)disulfane.
USP35
ADDITIONAL REQUIREMENTS
•  Packaging and Storage: Preserve in well-closed containers.
•  Labeling: When more than one Dissolution test is given, the labeling states the Dissolution test used only if Test 1 is not used.
•  USP Reference Standards 11
USP Mercaptopurine RS Click to View Structure
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
711 Margareth R.C. Marques, Ph.D.
Senior Scientific Liaison
1-301-816-8106
(GCDF2010) General Chapters - Dosage Forms
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USP35–NF30 Page 3811
Pharmacopeial Forum: Volume No. 36(6) Page 1554