Vincristine Sulfate Injection
201
11
USP Vincristine Sulfate (Assay) RS
(RB 1-Jul-2011)
DEFINITION
Vincristine Sulfate Injection is a sterile solution of Vincristine Sulfate in Water for Injection. It contains NLT 90.0% and NMT 110.0% of the labeled amount of vincristine sulfate (C46H56N4O10·H2SO4).
[Caution—Handle Vincristine Sulfate Injection with great care because it is a potent cytotoxic agent.
]
IDENTIFICATION
Change to read:
• A. Thin-Layer Chromatographic Identification Test 201
Standard solution:
10 mg/mL of USP Vincristine Sulfate RS or USP Vincristine Sulfate (Assay) RS(RB 1-Jul-2011) in dichloromethane and methanol (3:1)
or USP Vincristine Sulfate (Assay) RS(RB 1-Jul-2011) in dichloromethane and methanol (3:1)
Sample solution:
Transfer a volume of Injection, equivalent to 2 mg of vincristine sulfate, to a small centrifuge tube. For each mL of solution add 1 drop of ammonium hydroxide. Add 0.2 mL of dichloromethane. Place the cap on the tube, shake it vigorously for NLT 1 min, and centrifuge for 1 min. Carefully withdraw the dichloromethane layer, and transfer to a small stoppered vial.
Chromatographic system
Mode:
TLC
Adsorbent:
0.25-mm layer of chromatographic silica gel mixture
Application volume:
20 µL
Developing solvent system:
Fresh ether, methanol, and methylamine solution (2 in 5) (19:2:1)
Spray reagent:
Dissolve 2.0 g of ceric ammonium sulfate in 100 mL of water with heating and stirring, and slowly add 100 mL of phosphoric acid. Filter if necessary.
Analysis
Samples:
Standard solution and Sample solution
Develop the chromatographic plate in a methanol prewash tank; for maximum sensitivity, dry it NMT 2 h before use. Score it about 15 cm above the points of application. Apply the Standard solution and the Sample solution about 2.5 cm from the lower edge of the plate, and dry thoroughly (a current of cool air may be used to help dry the spots). Place the plate in the nonequilibrated developing chamber that contains a paper liner around the back and sides and Developing solvent system to a depth of 2 cm. Remove the plate when the solvent moves to the scored line (about 80 min), and discard the solvent system. Dry the plate in a fume hood at room temperature, heat on a metal plate on a steam bath for 15 min, and spray the plate while still hot with Spray reagent. Continue heating the plate for 15 min to stabilize the spots.
Acceptance criteria:
The RF value and the color of the principal spot from the Sample solution correspond to those from the Standard solution.
• 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
Change to read:
• Procedure
Solution A:
Diethylamine and water (1:59). Adjust with phosphoric acid to a pH of 7.5.
Mobile phase:
Methanol and Solution A (7:3)
Standard solution:
1.2 mg/mL(RB 1-Jul-2011) of USP Vincristine Sulfate RS or USP Vincristine Sulfate (Assay) RS(RB 1-Jul-2011) in water
1.2 mg/mL(RB 1-Jul-2011) of USP Vincristine Sulfate RS or USP Vincristine Sulfate (Assay) RS(RB 1-Jul-2011) in water
System suitability solution:
1 mg/mL of (RB 1-Jul-2011) USP Vinblastine Sulfate RS in the Standard solution(RB 1-Jul-2011)
(RB 1-Jul-2011) USP Vinblastine Sulfate RS in the Standard solution(RB 1-Jul-2011)
Sample solution:
Nominally 1.2 mg/mL(RB 1-Jul-2011) from Injection in water
1.2 mg/mL(RB 1-Jul-2011) from Injection in water
Chromatographic system
Mode:
LC
Detector:
UV 297 nm
Columns
Precolumn:
Porous silica gel packing
Guard:
2- to 5-cm; packing L1
Analytical:
4.6-mm × 25-cm; packing L7
Flow rate:
1.5 mL/min
Injection size:
10 µL
System suitability
Samples:
Standard solution and System suitability solution
Suitability requirements
Resolution:
NLT 4.0 between vincristine sulfate and vinblastine sulfate, 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 vincristine sulfate (C46H56N4O10·H2SO4) 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 vincristine sulfate in the Standard solution (mg/mL) |
| CU | = | = nominal concentration of vincristine sulfate in the Sample solution (mg/mL) |
Acceptance criteria:
90.0%–110.0%
IMPURITIES
Change to read:
Organic Impurities
• Procedure
Standard solution, System suitability solution, and System suitability:
Proceed as directed in the Assay.
Solution A:
Diethylamine and water (3:197). Adjust with phosphoric acid to a pH of 7.5.
Solution B:
Methanol
Solution C:
Prepare a suitable dilution of any preservative present in the Injection, as identified in the labeling.
Mobile phase:
See Table 1.
Table 1
| Time (min) |
Solution A (%) |
Solution B (%) |
|---|---|---|
| 0 | 38 | 62 |
| 12 | 38 | 62 |
| 27 | 8 | 92 |
| 29 | 38 | 62 |
| 34 | 38 | 62 |
Sample solution A:
1 mg/mL of vincristine sulfate from Injection in water
Sample solution B:
0.04 mg/mL in water from Sample solution A
Chromatographic system:
Proceed as directed in the Assay, except use a Flow rate of 2 mL/min and an Injection size of 200 µL.
Analysis
Samples:
Solution C, Sample solution A, and Sample solution B
Inject Solution C to identify the peaks due to the preservatives. Disregard any peaks at these retention times for the calculations of any other individual impurity and total impurities.
Calculate the percentage of each impurity in the portion of Injection taken:
Result = [rUA/(SrUA + 25rUB)] × 100
| rUA | = | = peak response of each impurity appearing after the solvent peak from Sample solution A |
| rUB | = | = peak response of vincristine from Sample solution B |
Calculate the percentage of total impurities in the portion of Injection taken:
Result = [(SrUA/(SrUA + 25rUB)] × 100
The terms in the formula are as defined above.
Acceptance criteria:
See Table 2.
Table 2
| Name | Relative Retention Time |
Acceptance Criteria, NMT (%) |
|---|---|---|
| Vincristine | 1.0 | — |
| N-Desformylvincristine | 1.4 (RB 1-Jul-2011) |
3.0 |
| Any other individual impurity | — | 2.0 |
| Total impurities | — | 6.0 |
SPECIFIC TESTS
• Bacterial Endotoxins Test 85:
It contains NMT 62.5 USP Endotoxin Units/mg of vincristine sulfate.
85:
It contains NMT 62.5 USP Endotoxin Units/mg of vincristine sulfate.
• pH 791:
3.5–5.5
791:
3.5–5.5
• Sterility Tests 71:
Meets the requirements
71:
Meets the requirements
• Other Requirements:
It meets the requirements in Injections 1, Labeling.
1, Labeling.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in light-resistant glass containers, and store in a refrigerator.
• Labeling:
The label states: “For Intravenous Use Only—Fatal If Given By Other Routes.”
Where labeled as containing more than 2 mg, it must also be labeled as a Pharmacy Bulk Package (see Injections 1). The labeling directs that the drug be dispensed only in containers enclosed in an overwrap labeled as directed below. When packaged in a Pharmacy Bulk Package, it is exempt from the requirement in Injections 1 that the closure be penetrated only one time after constitution with a suitable sterile transfer device or dispensing set, when it contains a suitable substance or mixture of substances to prevent the growth of microorganisms.
1). The labeling directs that the drug be dispensed only in containers enclosed in an overwrap labeled as directed below. When packaged in a Pharmacy Bulk Package, it is exempt from the requirement in Injections 1 that the closure be penetrated only one time after constitution with a suitable sterile transfer device or dispensing set, when it contains a suitable substance or mixture of substances to prevent the growth of microorganisms.When dispensed, the container or syringe (holding the individual dose prepared for administration to the patient) must be enclosed in an overwrap bearing the statement: “Do Not Remove Covering Until Moment of Injection. For Intravenous Use Only—Fatal If Given By Other Routes.”
Change to read:
• USP Reference Standards 11
USP Endotoxin RS
USP Vinblastine Sulfate RS 
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[Note—No loss on drying determination is needed. ]
USP Vincristine Sulfate RS
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USP Vincristine Sulfate (Assay) RS
(RB 1-Jul-2011)
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 |
| 85 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| 71 |
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 5023
Pharmacopeial Forum: Volume No. 37(1)