Add the following:
Powdered Malabar-Nut-Tree, Leaf Extract
DEFINITION
Powdered Malabar-Nut-Tree, Leaf Extract is prepared from Malabar-Nut-Tree, Leaf using suitable solvents such as water, methanol, or a mixture of these solvents. The ratio of plant material to extract is between 8:1 and 5:1. It contains NLT 90.0% and NMT 110.0% of the labeled amount of vasicine. It may contain suitable added substances as carriers.
IDENTIFICATION
• A. Thin-Layer Chromatographic Identification Test
Standard solution A:
0.5 mg/mL of USP Vasicine RS in methanol. Sonicate to dissolve if necessary.
Standard solution B:
50 mg/mL of USP Powdered Malabar-Nut-Tree, Leaf Extract RS in methanol. Sonicate for about 15 min, centrifuge, and use the supernatant.
Sample solution:
50 mg/mL of Powdered Malabar-Nut-Tree, Leaf Extract in methanol. Sonicate for about 15 min, centrifuge, and use the supernatant.
Adsorbent:
Chromatographic silica gel with an average particle size of 10–15 µm (TLC plates)
Application volume:
10 µL, as 4-mm bands
Developing solvent system:
A mixture of ethyl acetate, methanol, and ammonia (8: 2: 0.2)
Analysis
Samples:
Standard solution A, Standard solution B, and Sample solution
Apply the Samples as bands to a suitable thin-layer chromatographic plate (see Chromatography 
621
, Thin-Layer Chromatography). Use a saturated chamber. Develop the chromatograms until the solvent front has moved up about three-fourths of the plate. Remove the plate from the chamber, dry, and examine under UV at 254 nm.
621, Thin-Layer Chromatography). Use a saturated chamber. Develop the chromatograms until the solvent front has moved up about three-fourths of the plate. Remove the plate from the chamber, dry, and examine under UV at 254 nm.
Acceptance criteria:
The chromatogram of the Sample solution exhibits a quenching zone at an RF value of approximately 0.35 for vasicine, corresponding to a zone in the chromatogram of Standard solution A. The chromatogram of the Sample solution also exhibits an additional quenching zone at an RF value of approximately 0.53 for vasicinone, corresponding to a similar zone in the chromatogram of Standard solution B. Other minor zones may be observed in the Sample solution and Standard solution B chromatograms.
• B. HPLC:
The chromatogram of the Sample solution from the test for Content of Vasicine shows a main peak at a retention time corresponding to that of vasicine in the chromatogram of Standard solution A. Identify other peaks in the Sample solution by comparison with the chromatogram of Standard solution B and the reference chromatogram provided with the lot of USP Powdered Malabar-Nut-Tree, Leaf Extract RS being used. The Sample solution shows an additional peak corresponding to vasicinone.
COMPOSITION
• Content of Vasicine
Buffer solution:
Dissolve 1.36 g of anhydrous potassium dihydrogen phosphate in 900 mL of HPLC grade water. Add 2.0 mL of phosphoric acid. Dilute with water to 1000 mL, and filter.
Mobile phase:
Buffer solution, acetonitrile, and tetrahydrofuran (92:5:3)
Standard solution A:
0.1 mg/mL of USP Vasicine RS in methanol. Sonicate to dissolve if necessary.
Standard solution B:
5.0 mg/mL of USP Powdered Malabar-Nut-Tree, Leaf Extract RS in methanol. Sonicate for about 15 min. Before injection, pass through a membrane filter of 0.45-µm or finer pore size.
Sample solution:
5.0 mg/mL of Powdered Extract in methanol. Sonicate for about 15 min. Before injection, pass through a membrane filter of 0.45-µm or finer pore size, and discard the first part of the filtrate.
Chromatographic system
Mode:
LC
Detector:
UV 280 nm
Column:
4.6-mm × 25-cm; 5-µm packing L10
Flow rate:
1.0 mL/min
Injection size:
20 µL
System suitability
Samples:
Standard solution A and Standard solution B
[Note—The approximate relative retention times for vasicine and vasicinone peaks are 1.00 and 1.23, respectively. ]
Suitability requirements
Chromatogram similarity:
The chromatogram from Standard solution B is similar to the reference chromatogram provided with the lot of USP Powdered Malabar-Nut-Tree, Leaf Extract RS being used.
Resolution:
NLT 2.0 between the vasicine and vasicinone peaks, Standard solution B
Relative standard deviation:
NMT 2.0% determined from the vasicine peak in repeated injections, Standard solution A
Analysis
Samples:
Standard solution A, Standard solution B, and Sample solution
Using the chromatogram of Standard solution A, Standard solution B, and the reference chromatogram provided with the lot of USP Powdered Malabar-Nut-Tree, Leaf Extract RS being used, identify the retention times of the peaks corresponding to vasicine and vasicinone.
Calculate the percentage of vasicine in the portion of Powdered Extract taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response of vasicine from the Sample solution |
| rS | = | = peak response of vasicine from Standard solution A |
| CS | = | = concentration of USP Vasicine RS in Standard solution A (mg/mL) |
| CU | = | = concentration of Powdered Malabar-Nut-Tree, Leaf Extract in the Sample solution (mg/mL) |
Acceptance criteria:
90.0%–110.0% of the labeled amount of vasicine
CONTAMINANTS
• Heavy Metals, Method III 231:
NMT 20 ppm
231:
NMT 20 ppm
• Articles of Botanical Origin, General Method for Pesticide Residues Analysis 561:
Meets the requirements
561:
Meets the requirements
• Microbial Enumeration Tests 2021:
The total aerobic bacterial count does not exceed 104 cfu/g, and the total combined molds and yeasts count does not exceed 103 cfu/g.
2021:
The total aerobic bacterial count does not exceed 104 cfu/g, and the total combined molds and yeasts count does not exceed 103 cfu/g.
• Absence of Specified Microorganisms 2022:
Meets the requirements of the tests for absence of Salmonella species and Escherichia coli
2022:
Meets the requirements of the tests for absence of Salmonella species and Escherichia coli
SPECIFIC TESTS
• Loss on Drying 731:
Dry 1.0 g of Powdered Extract at 105
for 3 h: it loses NMT 5.0% of its weight.
731:
Dry 1.0 g of Powdered Extract at 105 for 3 h: it loses NMT 5.0% of its weight.
• Other Requirements:
It meets the requirements of the test for Residual Solvents in Botanical Extracts 565.
565.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in well-closed containers, protected from light and moisture, and store at controlled room temperature.
• Labeling:
The label states the Latin binomial and, following the official name, the part of the plant from which the article was derived. It meets other labeling requirements in Botanical Extracts 565.
565.
USP35
Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Maged H. Sharaf, Ph.D.
Principal Scientific Liaison 1-301-816-8318 |
(DS2010) Monographs - Dietary Supplements |
| 2021 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| 2022 |
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 1379
Pharmacopeial Forum: Volume No. 36(6) Page 1616