Fluconazole Injection
[Note—On the basis of the synthetic route, perform either (a) Procedure 1 and Procedure 2, or (b) Procedure 3, or (c) Procedure 4. Procedure 3 is recommended if bistriazole ketone and epoxyfluconazole (see Table 4) are potential impurities. Procedure 4 is recommended if fluconazole bromohydrine and epoxyfluconazole (see Table 5) are potential impurities. ]
USP35
USP35
DEFINITION
Fluconazole Injection is a sterile solution of Fluconazole in a suitable vehicle. It contains NLT 90.0% and NMT 110.0% of the labeled amount of fluconazole (C13H12F2N6O).
IDENTIFICATION
• A.
The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.
ASSAY
Change to read:
• Procedure
Buffer:
0.82 g/LUSP35 of sodium acetate in water. Adjust with 1 N acetic acid solution to a pH of 5.0.
0.82 g/LUSP35 of sodium acetate in water. Adjust with 1 N acetic acid solution to a pH of 5.0.
Diluent:
Methanol and Buffer (20:80)
Solution A:
Methanol and Buffer (5:95)
Solution B:
Acetonitrile and methanol (60:40)
Mobile phase:
See Table 1.
Table 1
| Time (min) |
Solution A (%) |
Solution B (%) |
|---|---|---|
| 0 | 80 | 20 |
| 9 | 80 | 20 |
| 15 | 15 | 85 |
| 18 | 80 | 20 |
| 25 | 80 | 20 |
System suitability solution:
0.04 mg/mL each of benzyl alcohol and USP Fluconazole RS in Diluent
Standard solution:
0.2 mg/mL of USP Fluconazole RS in Diluent
Sample solution:
Equivalent to 0.2 mg/mL of fluconazole from Injection in Diluent
Chromatographic system
Mode:
LC
Detector:
UV 261 nm
Column:
4.0-mm × 10-cm; 3-µm packing L1
Flow rate:
1 mL/min
Injection size:
100 µL
System suitability
Samples:
System suitability solution and Standard solution
[Note—The relative retention times for benzyl alcohol and fluconazole are about 0.8 and 1.0, respectively, from the System suitability solution. ]
Suitability requirements
Resolution:
NLT 1.8 between benzyl alcohol and fluconazole, System suitability solution
Column efficiency:
NLT 4000 theoretical plates for the fluconazole peak, System suitability solution
Tailing factor:
NMT 1.5 for the fluconazole peak, System suitability solution
Relative standard deviation:
NMT 2.0%, Standard solution
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of the labeled amount of fluconazole (C13H12F2N6O) in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response from the Sample solution |
| rS | = | = peak response from the Standard solution |
| CS | = | = concentration of the Standard solution (mg/mL) |
| CU | = | = nominal concentration of fluconazole in the Sample solution (mg/mL) |
Acceptance criteria:
90.0%–110.0%
IMPURITIES
Change to read:
Organic Impurities
[Note—On the basis of the synthetic route, perform either (a) Procedure 1 and Procedure 2, or (b) Procedure 3, or (c) Procedure 4. Procedure 3 is recommended if bistriazole ketone and epoxyfluconazole (see Table 4) are potential impurities. Procedure 4 is recommended if fluconazole bromohydrine and epoxyfluconazole (see Table 5) are potential impurities. ]
USP35
• Procedure 1: For Nonpolar Impurities
Buffer and Diluent:
Prepare as directed in the Assay.
Solution A:
Methanol and Buffer (5:95)
Solution B:
Acetonitrile and methanol (60:40)
Mobile phase:
See Table 2.
Table 2
| Time (min) |
Solution A (%) |
Solution B (%) |
|---|---|---|
| 0 | 77 | 23 |
| 5 | 77 | 23 |
| 30 | 40 | 60 |
| 43 | 77 | 23 |
| 50 | 77 | 23 |
System suitability solution:
2.4 and 20 µg/mL, respectively of 1,4-benzoquinone and USP Fluconazole RS in Diluent
Standard solution:
2 µg/mL of USP Fluconazole RS in Diluent
Diluted standard solution:
0.2 µg/mL of USP Fluconazole RS in Diluent from the Standard solution
Sample solution:
Equivalent to 1.0 mg/mL of fluconazole from Injection in Diluent
Chromatographic system
Mode:
LC
Detector:
UV 261 nm
Column:
4.0-mm × 10-cm; 3-µm packing L1
Flow rate:
1 mL/min
Injection size:
100 µL
System suitability
Samples:
System suitability solution, Standard solution, and Diluted standard solution
[Note—The relative retention times for 1,4-benzoquinone and fluconazole are about 0.5 and 1.0, respectively, System suitability solution. ]
Suitability requirements
Resolution:
NLT 5.0 between 1,4-benzoquinone and fluconazole, System suitability solution
Column efficiency:
NLT 2500 theoretical plates for the fluconazole peak, System suitability solution
Tailing factor:
NMT 1.5 for the fluconazole peak, System suitability solution
Relative standard deviation:
NMT 5.0%, Standard solution
Ratio of average peak area:
Between 8.0 and 12.0 for the ratio of the fluconazole peak from the Standard solution to that of the Diluted standard solution
Analysis
Samples:
Standard solution and Sample solution
[Note—For the following calculations, do not include peaks eluting before fluconazole and do not include impurities at relative retention times of 2.00–2.12 and 3.14–3.26. The disregarded impurities at the specified relative retention times are process impurities monitored in the drug substance. Furthermore, disregard any peak due to an excipient or any peak less than 0.02%. This test is for determination of the late-eluting peaks, and hence the early-eluting peaks are not quantitated using this procedure. ]
Calculate the percentage of the largest unknown nonpolar impurity in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response of the largest impurity from the Sample solution |
| rS | = | = peak response from the Standard solution |
| CS | = | = concentration of fluconazole in the Standard solution (mg/mL) |
| CU | = | = concentration of fluconazole, based on the label claim and the extent of dilution, in the Sample solution (mg/mL) |
Calculate the percentage of unknown nonpolar impurities in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = sum of the peak areas of the unknown peaks from the Sample solution |
| rS | = | = peak response from the Standard solution |
| CS | = | = concentration of the Standard solution (mg/mL) |
| CU | = | = concentration of fluconazole, based on the label claim and the extent of dilution, in the Sample solution (mg/mL) |
Acceptance criteria
Largest unknown nonpolar impurity:
NMT 0.1%
Total unknown nonpolar impurities:
NMT 0.5%
• Procedure 2: For Polar Impurities
Buffer, Diluent, Mobile phase, System suitability solution, and Chromatographic system:
Proceed as directed in the Assay.
Standard solution:
2 µg/mL of USP Fluconazole RS in Diluent
Diluted standard solution:
0.2 µg/mL of USP Fluconazole RS in Diluent from the Standard solution
Sample solution:
Equivalent to 0.2 mg/mL of fluconazole from Injection in Diluent
System suitability
Samples:
System suitability solution, Standard solution, and Diluted standard solution
[Note—The relative retention times for benzyl alcohol and fluconazole are about 0.8 and 1.0, respectively, System suitability solution. ]
Suitability requirements
Resolution:
NLT 1.8 between benzyl alcohol and fluconazole, System suitability solution
Column efficiency:
NLT 4000 theoretical plates for the fluconazole peak, System suitability solution
Tailing factor:
NMT 1.5 for the fluconazole peak, System suitability solution
Relative standard deviation:
NMT 5.0%, Standard solution
Ratio of average peak area:
Between 8.0 and 12.0 for the ratio of the fluconazole peak from the Standard solution to that of the Diluted standard solution
Analysis
Samples:
Standard solution and Sample solution
[Note—The relative retention times of known related compounds versus fluconazole are included in Table 3. ]
Calculate the percentage of the single largest unknown polar impurity in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response of the largest impurity from the Sample solution |
| rS | = | = peak response from the Standard solution |
| CS | = | = concentration of fluconazole in the Standard solution (mg/mL) |
| CU | = | = concentration of fluconazole, based on the label claim and the extent of dilution, in the Sample solution (mg/mL) |
Calculate the quantity, as a percentage, of the unknown polar impurities in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = total peak area response of all the unknown peaks from the Sample solution |
| rS | = | = peak response from the Standard solution |
| CS | = | = concentration of fluconazole in the Standard solution (mg/mL) |
| CU | = | = concentration of fluconazole, based on the label claim and the extent of dilution, in the Sample solution (mg/mL) |
Acceptance criteria
[Note—Disregard aminofluconazole quaternary salt, fluconazole isomer, and fluconazole diol impurities from Table 3 in the following calculation because these are process impurities that are monitored in the drug substance. ] USP35
Largest unknown polar impurity:
NMT 0.1%
Total unknown polar impurities:
NMT 0.5%
Total unknown polar and nonpolar impurities:
NMT 1.0% (sum of results from Procedure 1 and Procedure 2)
[Note—Disregard any peak due to an excipient or any peak less than 0.03%. ]
Table 3
| Name | Relative Retention Time |
|
|---|---|---|
| Hydroxymethylfurfural (if dextrose is present)a | 0.22–0.28 | |
| Aminofluconazole quaternary saltb | 0.30–0.36 | |
| Unidentified impurity (if dextrose is present)c | 0.37–0.43 | |
| Fluconazole isomerd | 0.47–0.59 | |
| Fluconazole diole | 0.68–0.74 | |
| Cyclohexanonef | 0.77–0.83 | |
| Fluconazole | 1.0 | |
|
a
5-Hydroxymethylfurfural.
b
4-Amino-1-(2-(2,4-difluorophenyl)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl)-4H-1,2,4-triazolium bromide.
c
Dextrose-related compound.
d
2-(2,4-Difluorophenyl)-1-(1H-1,2,4-triazol-1-yl)-3-(4H-1,2,4-triazol-4-yl)propan-2-ol.
e
2-(2,4-Difluorophenyl)-3-(1H-1,2,4-triazol-1-yl)propane-1,2-diol.
f
A process impurity associated with drug product packaged in bags.
|
||
USP35
• Procedure 3
Buffer:
0.63 g/LUSP35 of ammonium formate in water
0.63 g/LUSP35 of ammonium formate in water
Mobile phase:
Acetonitrile and Buffer (14:86)
Standard solution:
2 mg/mL of USP Fluconazole RS in Buffer. [Note—Use approximately 14% of the total volume as acetonitrile, and sonicate if necessary to facilitate dissolution. ]
Sensitivity solution:
1 µg/mL of USP Fluconazole RS in Mobile phase from the Standard solution
Sample solution:
Equivalent to 2 mg/mL of fluconazole in 0.9% aqueous solution of sodium chlorideUSP35
Equivalent to 2 mg/mL of fluconazole in 0.9% aqueous solution of sodium chlorideUSP35
Chromatographic system
Mode:
LC
Detector:
UV 260 nm
Column:
4.6-mm × 15-cm; 3.5-µm packing L1
Column temperature:
30
Flow rate:
1 mL/min
Injection size:
20 µL
System suitability
Samples:
Standard solution and Sensitivity solution
[Note—The retention time for fluconazole is between 12 and 14 min. ]
Suitability requirements
Signal-to-noise ratio:
NLT 10, Sensitivity solution
Column efficiency:
NLT 3000 theoretical plates, Standard solution
Tailing factor:
NMT 2.0, Standard solution
Relative standard deviation:
NMT 2%, Standard solution
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of each impurity in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response of each impurity from the Sample solution |
| rS | = | = peak response of fluconazole from the Standard solution |
| CS | = | = concentration of fluconazole in the Standard solution (mg/mL) |
| CU | = | = concentration of fluconazole, based on the label claim, in the Sample solution (mg/mL) |
Acceptance criteria
Individual impurities:
See Table 4.
Total impurities:
NMT 0.5%
Table 4
| Name | Relative Retention Time |
Acceptance Criteria, NMT (%) |
|
|---|---|---|---|
| Bistriazole ketonea | 0.13 | 0.2 | |
| Fluconazole isomerb | 0.5 | 0.2 | |
| Fluconazole | 1.0 | — | |
| Epoxyfluconazolec | 2.6 | 0.2 | |
| Any other individual impurity | — | 0.2 | |
|
a
1,3-Bis(1H-1,2,4-triazol-1-yl)propan-2-one.
b
2-(2,4-Difluorophenyl)-1-(1H-1,2,4-triazol-1-yl)-3-(4H-1,2,4-triazol-4-yl)propan-2-ol.
c
1-[2-(2,4-Difluorophenyl)-2,3-epoxypropyl]-1H-1-2,4-triazole.
|
|||
Procedure 4
Buffer:
13.4 g/L of dibasic sodium phosphate heptahydrate in water. Adust with phosphoric acid to a pH of 7.0.
Mobile phase:
Acetonitrile and Buffer (26:74)
Standard solution:
2 mg/mL of USP Fluconazole RS in Mobile phase
Sample solution:
Equivalent to 2 mg/mL of fluconazole in 0.9% aqueous solution of sodium chloride
Chromatographic system
Mode:
LC
Detector:
UV 260 nm
Column:
4.6-mm × 25-cm; 5-µm packing L7
Flow rate:
0.5 mL/min
Injection size:
50 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 each impurity in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response of each impurity from the Sample solution |
| rS | = | = peak response of fluconazole from the Standard solution |
| CS | = | = concentration of fluconazole in the Standard solution (mg/mL) |
| CU | = | = concentration of fluconazole, based on the label claim, in the Sample solution (mg/mL) |
Table 5
| Name | Relative Retention Time |
Relative Response Factor |
Acceptance Criteria, NMT (%) |
|
|---|---|---|---|---|
| Aminofluconazole quaternary salta | 0.57 | 0.74 | 0.1 | |
| Fluconazole isomerb | 0.68 | 0.93 | 0.1 | |
| Fluconazole diolc | 0.91 | 1.3 | 0.1 | |
| Fluconazole | 1.0 | 1.0 | — | |
| Fluconazole bromohydrined | 2.58 | 1.1 | 0.1 | |
| Epoxyfluconazolee | 2.59 | 0.90 | 0.1 | |
| Any other individual impurity | — | 1.0 | 0.1 | |
|
a
4-Amino-1-(2-(2,4-difluorophenyl)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl)-4H-1,2,4-triazol-1-ium bromide.
b
2-(2,4-Difluorophenyl)-1-(1H-1,2,4-triazol-1-yl)-3-(4H-1,2,4-triazol-4-yl)propan-2-ol.
c
2-(2,4-Difluorophenyl)-3-(1H-1,2,4-triazol-1-yl)propane-1,2-diol.
d
1-Bromo-2-(2,4-difluorophenyl)-3-(1H-1,2,4-triazol-1-yl)propan-2-ol.
e
1-[2-(2,4-Difluorophenyl)-2,3-epoxypropyl]-1H-1-2,4-triazole.
|
||||
USP35SPECIFIC TESTS
• Sterility Tests 
71
:
Meets the requirements
71:
Meets the requirements
• Bacterial Endotoxins Test 85:
NMT 0.416 USP Endotoxin Unit/mg of fluconazole
85:
NMT 0.416 USP Endotoxin Unit/mg of fluconazole
• Other Requirements:
Meets the requirements under Injections 1
1
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Store at controlled room temperature.
• Labeling:
Label to indicate the vehicle used. If a test for Organic Impurities other than Procedure 1 and Procedure 2 is used, then the labeling states with which Organic Impurities test the article complies.
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Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Behnam Davani, Ph.D., M.B.A.
Senior Scientific Liaison 1-301-816-8394 |
(SM12010) Monographs - Small Molecules 1 |
| 71 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| 85 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 3205
Pharmacopeial Forum: Volume No. 36(6) Page 1530