Isoleucine
(eye'' soe loo' seen).
DEFINITION
Isoleucine contains NLT 98.5% and NMT 101.5% of l-isoleucine (C6H13NO2), calculated on the dried basis.
IDENTIFICATION
ASSAY
• Procedure
Sample:
130 mg of Isoleucine
Blank:
Mix 3 mL of formic acid and 50 mL of glacial acetic acid.
Titrimetric system
(See Titrimetry 541.)
Mode:
Direct titration
Titrant:
0.1 N perchloric acid VS
Endpoint detection:
Potentiometric
Analysis:
Dissolve the Sample in 3 mL of formic acid and 50 mL of glacial acetic acid. Titrate with the Titrant. Perform the Blank determination.
Calculate the percentage of isoleucine (C6H13NO2) in the Sample taken:
Result = {[(VS VB) × N × F]/W} × 100
Acceptance criteria:
98.5%101.5% on the dried basis
IMPURITIES
• Residue on Ignition 281:
NMT 0.3%
• Chloride and Sulfate, Chloride 221
Standard solution:
0.50 mL of 0.020 N hydrochloric acid
Sample:
0.73 g of Isoleucine
Acceptance criteria:
NMT 0.05%
• Chloride and Sulfate, Sulfate 221
Standard solution:
0.10 mL of 0.020 N sulfuric acid
Sample:
0.33 g of Isoleucine
Acceptance criteria:
NMT 0.03%
• Iron 241:
NMT 30 ppm
• Heavy Metals, Method I 231:
NMT 15 ppm
• Related Compounds
System suitability solution:
0.4 mg/mL each of USP l-Isoleucine RS and USP l-Valine RS in 0.1 N hydrochloric acid
Standard solution:
0.05 mg/mL of USP l-Isoleucine RS in 0.1 N hydrochloric acid. [NoteThis solution has a concentration equivalent to 0.5% of that of the Sample solution. ]
Sample solution:
10 mg/mL of Isoleucine in 0.1 N hydrochloric acid
Chromatographic system
Mode:
TLC
Adsorbent:
0.25-mm layer of chromatographic silica gel mixture
Application volume:
5 µL
Developing solvent system:
Butyl alcohol, glacial acetic acid, and water (3:1:1)
Spray reagent:
2 mg/mL of ninhydrin in a mixture of butyl alcohol and 2 N acetic acid (95:5)
System suitability
Suitability requirements:
The chromatogram of the System suitability solution exhibits two clearly separated spots.
Analysis
Samples:
System suitability solution, Standard solution, and Sample solution
After air-drying the plate, spray with Spray reagent, and heat between 100 and 105 for 15 min. Examine the plate under white light.
Acceptance criteria:
Any secondary spot of the Sample solution is not larger or more intense than the principal spot of the Standard solution.
Individual impurities:
NMT 0.5%
Total impurities:
NMT 2.0%
SPECIFIC TESTS
• Optical Rotation, Specific Rotation 781S
Sample solution:
40 mg/mL in 6 N hydrochloric acid
Acceptance criteria:
+38.9 to +41.8
• Loss on Drying 731:
Dry a sample at 105 for 3 h: it loses NMT 0.3% of its weight.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in well-closed containers.
Auxiliary Information
Please check for your question in the FAQs before contacting USP.
USP35NF30 Page 3573
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