Phytonadione
(fye toe'' na dye' one).
C31H46O2 450.70 1,4-Naphthalenedione, 2-methyl-3-(3,7,11,15-tetramethyl-2-hexadecenyl)-, [R-[R*,R*-(E)]]-; Phylloquinone [84-80-0]. DEFINITION
Phytonadione is a mixture of E- and Z-isomers containing NLT 97.0% and NMT 103.0% of phytonadione isomers (C31H46O2). It contains NMT 21.0% of the Z-isomer.
IDENTIFICATION
• B. Ultraviolet Absorption 197U
Analytical wavelength:
248 nm
Sample solution:
10 µg/mL in n-hexane
Acceptance criteria:
Meets the requirements. Absorptivities do not differ by more than 3.0%.
ASSAY
• Procedure
[NoteProtect solutions containing phytonadione from exposure to light. ]
Mobile phase:
n-Hexane and n-amyl alcohol (2000: 1.5)
Internal standard solution:
2.5 mg/mL of cholesteryl benzoate in Mobile phase
Standard solution:
Prepare a 96-µg/mL solution of USP Phytonadione RS in Mobile phase. Pipet 10 mL of this solution and 7 mL of Internal standard solution into a 25-mL volumetric flask, and dilute with Mobile phase to volume.
Sample solution:
Prepare as directed for the Standard solution, using Phytonadione instead of USP Phytonadione RS.
Chromatographic system
Mode:
LC
Detector:
UV 254 nm
Column:
4.6-mm × 25-cm; packing L3
Flow rate:
1 mL/min
Injection size:
50 µL
System suitability
Sample:
Standard solution
[NoteThe relative retention times for the internal standard, (Z)-phytonadione, and (E)-phytonadione are 0.7, 0.9, and 1.0, respectively. ]
Suitability requirements
Resolution:
NLT 1.5 between (Z)-phytonadione and (E)-phytonadione
Relative standard deviation:
NMT 2.0% for the ratios of the sum of peak areas of (Z)-phytonadione and (E)-phytonadione to the peak area of the internal standard
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of phytonadione (C31H46O2) in the portion of Phytonadione taken:
Result = (RU/RS) × (CS/CU) × 100
Acceptance criteria:
97.0%103.0%
OTHER COMPONENTS
• Z-Isomer Content
[NoteProtect solutions containing phytonadione from exposure to light. ]
Mobile phase, Internal standard solution, Sample solution, Chromatographic system, and Analysis:
Proceed as directed in the Assay, except calculate the percentage of Z-isomer in the portion of Phytonadione taken:
Result = [rZ/(rZ + rE)] × 100
Acceptance criteria:
NMT 21.0%
IMPURITIES
• Limit of Menadione
Sample:
20 mg of Phytonadione
Analysis:
Mix the Sample with 0.5 mL of a mixture of 6 N ammonium hydroxide and alcohol (1:1). Add 1 drop of ethyl cyanoacetate, and shake gently.
Acceptance criteria:
No purple or blue color is produced.
SPECIFIC TESTS
• Refractive Index 831:
1.5231.526
• Reaction:
A 50-mg/mL solution of Phytonadione in dehydrated alcohol is neutral to litmus.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in tight, light-resistant containers.
Auxiliary Information
Please check for your question in the FAQs before contacting USP.
USP35NF30 Page 4318
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