Leflunomide
C12H9F3N2O2 270.21
4-Isoxazolecarboxamide, 5-methyl-N-[4-(trifluoromethyl)phenyl]-;
,,-Trifluoro-5-methyl-4-isoxazolecarboxy-p-toluidide
[75706-12-6].
(le floo' noe mide).
+'&img=../../images/v35300/cas-75706-12-6.gif&casNumber=75706-12-6&usp=35&nf=30&chemicalStructureImage=true&ID='+parent.myID,600,500);)
C12H9F3N2O2 270.21
4-Isoxazolecarboxamide, 5-methyl-N-[4-(trifluoromethyl)phenyl]-;
,,-Trifluoro-5-methyl-4-isoxazolecarboxy-p-toluidide
[75706-12-6].DEFINITION
Leflunomide contains NLT 98.0% and NMT 102.0% of C12H9F3N2O2, calculated on the dried basis.
IDENTIFICATION
• A. Infrared Absorption 
197K
197K
Sample:
Dry the substance for 10 min at 130
.
.
• B.
The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.
ASSAY
• Procedure
Mobile phase:
Acetonitrile, triethylamine, and water (70:1:130). Adjust with phosphoric acid to a pH of 4.
Standard solution:
0.5 mg/mL of USP Leflunomide RS in acetonitrile and Mobile phase (1:9). [Note—First dissolve in acetonitrile. Protect solutions from light. ]
System suitability solution:
0.5 mg/mL of USP Leflunomide RS, 0.15 mg/mL of USP Leflunomide Related Compound B RS, and 0.05 mg/mL of USP Leflunomide Related Compound C RS in Mobile phase. [Note—Dissolve the Reference Standards in acetonitrile, and dilute with Mobile phase. ]
Sample solution:
0.5 mg/mL of Leflunomide in acetonitrile and Mobile phase (1:9). [Note—First dissolve in acetonitrile. Protect solutions from light. ]
Chromatographic system
Mode:
LC
Detector:
UV 210 nm
Column:
4-mm × 12.5-cm; packing L1
Flow rate:
1 mL/min
Injection size:
20 µL
System suitability
Sample:
System suitability solution
[Note—The relative retention times for leflunomide related compound B and leflunomide related compound C are 0.2 and 0.9, respectively. ]
Suitability requirements
Resolution:
NLT 1.0 between the leflunomide and leflunomide related compound C peaks
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of C12H9F3N2O2 in the portion of Leflunomide 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 Leflunomide RS in the Standard solution (mg/mL) |
| CU | = | = nominal concentration of Leflunomide in the Sample solution (mg/mL) |
Acceptance criteria:
98.0%–102.0% on the dried basis
IMPURITIES
Organic Impurities
• Procedure 1: Limit of Leflunomide Related Compound A
Mobile phase, System suitability solution, and Chromatographic system:
Proceed as directed in the Assay.
Standard stock solution:
0.125 mg/mL of USP Leflunomide Related Compound A RS, in acetonitrile and Mobile phase (1:19)
Standard solution:
0.5 µg/mL of USP Leflunomide Related Compound A RS, from the Standard stock solution in Mobile phase
Sample solution:
2.5 mg/mL of Leflunomide, in acetonitrile and Mobile phase (1:9)
Injection size:
20 µL
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of leflunomide related compound A in the portion of Leflunomide taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak area of leflunomide related compound A from the Sample solution |
| rS | = | = peak area of leflunomide related compound A from the Standard solution |
| CS | = | = concentration of USP Leflunomide Related Compound A RS in the Standard solution (mg/mL) |
| CU | = | = concentration of Leflunomide in the Sample solution (mg/mL) |
Acceptance criteria:
NMT 0.02 %
• Procedure 2
Mobile phase, Sample solution, System suitability solution, and Chromatographic system:
Proceed as directed in the Assay.
Standard solution:
0.5 µg/mL of USP Leflunomide RS, from the Standard solution in Mobile phase
Sensitivity solution:
0.25 µg/mL of Leflunomide, from the Standard solution in Mobile phase
System suitability
Samples:
System suitability solution and Sensitivity solution
Resolution:
NLT 1.0 between leflunomide and leflunomide related compound C
Signal-to-noise ratio:
NLT 10, Sensitivity solution
Analysis
Samples:
Standard solution and Sample solution
[Note—Disregard any peak with an area less than the leflunomide peak from the System suitability solution. Continue the elution for two times the retention time of the leflunomide peak. ]
Calculate the percentage of each related compound and any unknown impurity (see Impurity Table 1) in the portion of Leflunomide taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak area for each impurity from the Sample solution |
| rS | = | = peak area of leflunomide from the Standard solution |
| CS | = | = concentration of USP Leflunomide RS in the Standard solution (mg/mL) |
| CU | = | = concentration of Leflunomide in the Sample solution (mg/mL) |
Impurity Table 1
| Name | Relative Retention Time |
Relative Response Factor |
Acceptance Criteria, NMT (%) |
|---|---|---|---|
| 5-Methylisoxazole-carboxylic acid | 0.05 | 1.0 | 0.1 |
| Leflunomide related compound B | 0.22 | 1.0 | 0.3 |
| N-(2¢-Trifluoromethylphenyl)-5-methylisoxazole-4-Carboxamide | 0.29 | 1.0 | 0.1 |
| 2-Cyano-acetic acid-(4¢-trifluoromethyl)-anilide | 0.36 | 1.0 | 0.1 |
| Leflunomide related compound C | 0.94 | 1.0 | 0.1 |
| Any other individual impurity | — | — | 0.1 |
| Total impurities, excluding leflunomide related compound B and leflunomide related compound C | — | — | 0.2 |
| Total impurities | — | — | 0.4 |
SPECIFIC TESTS
• Melting Range or Temperature 741:
164–168
741:
164–168
• Loss on Drying 731:
Dry a sample in a vacuum over diphosphorus pentoxide at 60 for 4 h: it loses NMT 0.5% of its weight.
731:
Dry a sample in a vacuum over diphosphorus pentoxide at 60 for 4 h: it loses NMT 0.5% of its weight.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in a well-closed container. Store at a temperature not exceeding 30.
.
• USP Reference Standards 11
11
USP Leflunomide RS 
') + '&img=../../images/v35300/cas-75706-12-6.gif',600,500);)
USP Leflunomide Related Compound A RS
USP Leflunomide Related Compound B RS
USP Leflunomide Related Compound C RS
Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Clydewyn M. Anthony, Ph.D.
Senior Scientific Liaison 1-301-816-8139 |
(SM22010) Monographs - Small Molecules 2 |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 3645
Pharmacopeial Forum: Volume No. 35(5) Page 1158