Lutein
C40H56O2 568.87
-
-Carotene-3,3¢-diol (3R,3¢R,6¢R)
[127-40-2].
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C40H56O2 568.87
--Carotene-3,3¢-diol (3R,3¢R,6¢R)
[127-40-2].DEFINITION
Lutein is the purified fraction obtained from saponification of the oleoresin of Tagetes erecta L. It contains NLT 80.0% of total carotenoids calculated as lutein (C40H56O2). It contains NLT 74.0% of lutein and NMT 8.5% of zeaxanthin, both calculated as lutein (C40H56O2) on the anhydrous basis.
IDENTIFICATION
• A. Ultraviolet-Visible Absorption 
197U
197U
Wavelength range:
300–700 nm
Sample solution:
Prepare as directed for the Sample solution in the test for Content of Total Carotenoids.
Acceptance criteria:
It meets the requirements in the chapter. The absorbance ratio A446/A474 is 1.09–1.14.
• B.
The retention time for the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the test for Content of Lutein.
COMPOSITION
• Content of Total Carotenoids
[Note—Use low-actinic glassware. ]
Diluent:
Hexanes, acetone, toluene, and dehydrated alcohol (10:7:7:6)
Sample stock solution:
0.3 mg/mL of Lutein in Diluent
Sample solution:
3.0 µg/mL of Lutein in dehydrated alcohol from dilution of Sample stock solution
Instrumental conditions
Mode:
UV-Vis
Analytical wavelength:
446 nm
Blank:
Dehydrated alcohol
Analysis
Sample:
Sample solution
Calculate the percentage of total carotenoids (T) as lutein (C40H56O2) in the portion of Lutein taken:
Result = A/(F × C)
| A | = | = absorbance of the Sample solution |
| F | = | = coefficient of extinction (E1%) of lutein in alcohol (100 mL·g 1·cm1), 2550 |
| C | = | = concentration of lutein in the Sample solution (g/mL) |
Acceptance criteria:
NLT 80.0%
• Content of Lutein
Mobile phase:
Hexane and ethyl acetate (3:1)
Standard solution:
150 µg/mL of USP Lutein RS in Mobile phase
Sample solution:
Transfer 1 mL of the Sample stock solution from the test for Content of Total Carotenoids, and evaporate under a stream of nitrogen to dryness. Add 1 mL of Mobile phase, and sonicate to dissolve.
Chromatographic system
Mode:
LC
Detector:
UV-Vis at 446 nm
Column:
4.6-mm × 25-cm; 5-µm packing L3
Flow rate:
1.5 mL/min
Injection size:
10 µL
System suitability
Sample:
Standard solution
[Note—The relative retention times for lutein and zeaxanthin are about 1.0 and 1.05, respectively. ]
Suitability requirements
Resolution:
NLT 1.0 between lutein and zeaxanthin
Tailing factor:
NMT 2.0
Relative standard deviation:
NMT 2.0%
Analysis
Sample:
Sample solution
Calculate the percentage of the lutein peak as the total detected area in the portion of Lutein taken:
Result = (rU/rT) × 100
| rU | = | = peak response of lutein |
| rT | = | = sum of the responses of all the peaks |
Acceptance criteria:
NLT 85%
Calculate the percentage of lutein in the portion of Lutein taken:
Result = (rU/rT) × T
| rU | = | = individual peak response |
| rT | = | = sum of the responses of all the peaks |
| T | = | = percentage of total carotenoids as determined in the test for Content of Total Carotenoids |
Acceptance criteria:
NLT 74.0% of lutein on the anhydrous basis
• Zeaxanthin and Other Related Compounds
[Note—Use low-actinic glassware. ]
Mobile phase, Standard solution, Sample solution, and Chromatographic system:
Proceed as directed in Content of Lutein.
Analysis
Sample:
Sample solution
Calculate the percentage of zeaxanthin as the total detected area in the portion of Lutein taken:
Result = (rU/rT) × 100
| rU | = | = peak response of zeaxanthin |
| rT | = | = sum of the responses of all the peaks |
Acceptance criteria:
NMT 9.0%
Calculate the percentage of zeaxanthin in the portion of Lutein taken:
Result = (rU/rT) × T
| rU | = | = peak response of zeaxanthin |
| rT | = | = sum of the responses of all the peaks |
| T | = | = percentage of total carotenoids as determined in the test for Content of Total Carotenoids |
Calculate the percentage of other related compounds in the portion of Lutein taken:
Result = (rU/rT) × 100
| rU | = | = individual peak response of any other peak in the chromatogram (excluding zeaxanthin and lutein) |
| rT | = | = sum of the responses of all the peaks |
Acceptance criteria:
NMT 8.5% of zeaxanthin; NMT 1.0% of any other single related compound on the anhydrous basis
IMPURITIES
• Residue on Ignition 281:
NMT 2.0%
281:
NMT 2.0%
• Lead 251:
NMT 1 ppm
251:
NMT 1 ppm
• Heavy Metals, Method II 231:
NMT 5 ppm
231:
NMT 5 ppm
SPECIFIC TESTS
• Water Determination, Method I 921:
NMT 1.0%
921:
NMT 1.0%
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in hermetically sealed, light- and oxygen-resistant containers. Store in a cool place.
• USP Reference Standards 11
11
USP Lutein RS
Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Huy T. Dinh, M.S.
Scientific Liaison 1-301-816-8594 |
(DS2010) Monographs - Dietary Supplements |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 1368
Pharmacopeial Forum: Volume No. 33(2) Page 255