Vitamin A Tablets
DEFINITION
Vitamin A Tablets contain retinyl acetate or retinyl palmitate in an amount equivalent to NLT 95.0% and NMT 120.0% of the labeled amount of vitamin A, as retinol (C20H30O).
IDENTIFICATION
[Note—Use low-actinic glassware. ]
• A. Procedure
Sample solution:
A solution containing the equivalent of 6 µg/mL of retinol from powdered Tablets in methylene chloride
Analysis:
To 1 mL of Sample solution add 10 mL of antimony trichloride TS.
Acceptance criteria:
A transient blue color appears at once.
• B. Thin-Layer Chromatographic Identification Test 
201
201
Standard solution:
A solution containing the equivalent of 0.5 mg/mL of retinol from USP Retinyl Acetate RS or USP Retinyl Palmitate RS in methylene chloride
Sample solution:
To a portion of finely powdered Tablets, equivalent to 5 mg of retinol, add 15 mL of water, sonicate, and shake vigorously for 2 min. Extract with 10 mL of methylene chloride by shaking for 2 min. Centrifuge, and use the lower layer of methylene chloride extract.
Application volume:
10 µL, 8-mm band
Developing solvent system:
Cyclohexane and ether (4:1)
Spray reagent:
Phosphomolybdic acid TS
Analysis:
Proceed as directed in the chapter, using the Developing solvent system. Locate the spots on the plate using the Spray reagent.
Acceptance criteria:
The RF value of the principal spot from the Sample solution corresponds to that from the Standard solution.
• C.
The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.
ASSAY
• Procedure
[Note—Use low-actinic glassware. ]
Mobile phase:
n-Hexane
Standard solution 1:
A solution containing the equivalent of 15 µg/mL of retinol from USP Retinyl Acetate RS in n-hexane
Standard solution 2:
A solution containing the equivalent of 15 µg/mL of retinol from USP Retinyl Palmitate RS in n-hexane
System suitability solution:
Mix equal volumes of Standard solution 1 and Standard solution 2.
Sample solution:
Finely powder NLT 20 Tablets. To a portion of the powder, equivalent to 5 Tablets, add 15 mL of water, and sonicate for 5 min. Add 15 mL of n-hexane, and shake for 15 min on a wrist-action shaker in a water bath maintained at 60
. Add 10 mL of dimethyl sulfoxide, and shake for an additional period of 30 min on a wrist-action shaker in a water bath maintained at 60. [Note—Set up the wrist-action shaker to ensure that the contents of the container are mixed vigorously and thoroughly. ] Centrifuge, and transfer the hexane layer to a 100-mL volumetric flask. Repeat the extraction with three additional 15-mL portions of n-hexane by thoroughly shaking each for 5 min. Dilute the extracts in the volumetric flask with n-hexane to volume. Further dilute this solution with n-hexane to obtain a solution with a concentration equivalent to 15 µg/mL of retinol.
. Add 10 mL of dimethyl sulfoxide, and shake for an additional period of 30 min on a wrist-action shaker in a water bath maintained at 60. [Note—Set up the wrist-action shaker to ensure that the contents of the container are mixed vigorously and thoroughly. ] Centrifuge, and transfer the hexane layer to a 100-mL volumetric flask. Repeat the extraction with three additional 15-mL portions of n-hexane by thoroughly shaking each for 5 min. Dilute the extracts in the volumetric flask with n-hexane to volume. Further dilute this solution with n-hexane to obtain a solution with a concentration equivalent to 15 µg/mL of retinol.
Chromatographic system
Mode:
LC
Detector:
UV 325 nm
Column:
4.6-mm × 15-cm; 3-µm packing L8
Flow rate:
1 mL/min
Injection size:
40 µL
System suitability
Sample:
System suitability solution
Suitability requirements
Resolution:
NLT 10 between all-trans-retinyl palmitate and all-trans-retinyl acetate
Relative standard deviation:
NMT 3.0%
Analysis
Samples:
Standard solution 1 or Standard solution 2 and Sample solution
Calculate the percentage of the labeled amount of vitamin A, as retinol (C20H30O), in the portion of Tablets taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak area of the all-trans-retinyl ester from the Sample solution |
| rS | = | = peak area of the all-trans-retinyl ester from the appropriate Standard solution |
| CS | = | = concentration of retinol in the appropriate Standard solution (µg/mL) |
| CU | = | = nominal concentration of vitamin A, as retinol, in the Sample solution (µg/mL) |
Acceptance criteria:
95.0%–120.0% of the labeled amount of vitamin A, as retinol (C20H30O)
PERFORMANCE TESTS
• Dissolution 711
711
[Note—Perform this test under light conditions that minimize photo degradation. ]
Medium:
1% (w/v) sodium ascorbate and 1% (w/v) octoxynol 9 in 0.05 M phosphate buffer pH 6.8; 900 mL
Apparatus 2:
75 rpm
Time:
45 min
Standard solution:
Dissolve a suitable amount of USP Retinyl Acetate RS or USP Retinyl Palmitate RS in isopropyl alcohol, and dilute with Medium to obtain the concentration similar to that expected in the Sample solution. [Note—The amount of isopropyl alcohol should be 5%–10%. ]
Sample solution:
Withdraw a portion of the solution under test, pass through a suitable filter of 0.45-µm pore size, and use the pooled sample as the test specimen.
Solution A:
Methanol and water (90:10)
Solution B:
Methanol and isopropyl alcohol (55:45)
Mobile phase:
See the gradient table below.
| Time (min) |
Solution A (%) |
Solution B (%) |
|---|---|---|
| 0 | 100 | 0 |
| 8 | 0 | 100 |
| 13 | 0 | 100 |
| 13.1 | 100 | 0 |
| 15 | 100 | 0 |
Chromatographic system
Mode:
LC
Detector:
UV 325 nm
Column:
4.6-mm × 10-cm; 3-µm packing L1
Flow rate:
1.0 mL/min
Injection size:
50 µL
System suitability
Sample:
Standard solution
Suitability requirements
Tailing factor:
NMT 1.5 for retinyl acetate and NMT 2.0 for retinyl palmitate
Relative standard deviation:
NMT 2.0%
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of the labeled amount of retinyl acetate or retinyl palmitate dissolved:
Result = (rU/rS) × (CS × V/L) × 100
| rU | = | = peak area of the all-trans-retinyl ester from the Sample solution |
| rS | = | = peak area of the all-trans-retinyl ester from the appropriate Standard solution |
| CS | = | = concentration of retinol in the appropriate Standard solution (µg/mL) |
| V | = | = volume of Medium, 900 mL |
| L | = | = label claim of vitamin A, as retinol (µg/Tablet) |
Tolerances:
NLT 75% (Q) of the labeled amount of retinyl acetate or retinyl palmitate is dissolved.
• Uniformity of Dosage Units 905:
Meet the requirements
905:
Meet the requirements
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in tight containers, and store at room temperature. Protect Tablets from light.
• Labeling:
Label it to indicate the chemical form of vitamin A present, and to indicate the vitamin A activity in terms of the equivalent amount of retinol. Vitamin A activity may be stated also in USP units. USP units of activity for vitamins, where such exist or formerly existed, are equivalent to the corresponding international units, where such formerly existed. International units (IU) for vitamins also have been discontinued; however, the use of IU on the labels of vitamin products continues. Where articles are labeled in terms of Units in addition to the required labeling, the relationship of the USP Units or IU to mass is as follows. One USP Vitamin A Unit = 0.3 µg of all-trans-retinol (vitamin A alcohol) or 0.344 µg of all-trans-retinyl acetate (vitamin A acetate) or 0.55 µg of all-trans-retinyl palmitate (vitamin A palmitate), and 1 µg of retinol (3.3 USP Vitamin A Units) = 1 retinol equivalent (RE).
Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Natalia Davydova
Scientific Liaison 1-301-816-8328 |
(DS2010) Monographs - Dietary Supplements |
| 711 |
Margareth R.C. Marques, Ph.D.
Senior Scientific Liaison 1-301-816-8106 |
(GCDF2010) General Chapters - Dosage Forms |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 5031
Pharmacopeial Forum: Volume No. 36(5) Page 1219