Powdered Cat's Claw
DEFINITION
Powdered Cat's Claw is Cat's Claw reduced to a powder or very fine powder. It contains NLT 0.3% of pentacyclic oxindole alkaloids as isopteropodine, calculated on the dried basis, as the sum of speciophylline, uncarine F, mitraphylline, isomitraphylline, pteropodine, and isopteropodine.
IDENTIFICATION
• A. Thin-Layer Chromatographic Identification Test
Standard solution:
100 mg of USP Powdered Cat's Claw Extract RS in 2 mL of methanol. Sonicate for 5 min, shaking occasionally, heat in a water bath at 60
for 15 min, cool, and centrifuge.
for 15 min, cool, and centrifuge.
Sample solution:
5 g of Powdered Cat's Claw in 10 mL of methanol. Sonicate for 5 min, shaking occasionally. Heat the mixture in a water bath at 60 for 15 min, cool, and filter.
for 15 min, cool, and filter.
Adsorbent:
Chromatographic silica gel mixture with an average particle size of 10–15 µm (TLC plates)
Application volume:
20 µL, as bands that are 1 cm in length
Developing solvent system:
Ethyl acetate and hexane (95:5)
Spray reagent A:
Dissolve 0.85 g of basic bismuth nitrate in 10 mL of glacial acetic acid and 40 mL of water by heating. Filter if necessary (Solution A). Dissolve 8 g of potassium iodide in 30 mL of water (Solution B). Mix Solution A and Solution B (1:1) to obtain a stock solution. Dilute 1 mL of the stock solution with 2 mL of glacial acetic acid and 10 mL of water. [Note—Use freshly mixed Solution A and Solution B. ]
Spray reagent B:
Use a 10% solution of sodium nitrite in water.
Analysis
Samples:
Standard solution and Sample solution
Develop the chromatogram to a length of NLT 12 cm, and dry the plate in a current of air.
Acceptance criteria:
Examine the plate under short UV light. The Sample solution chromatogram shows multiple zones that correspond in RF values to those observed from the Standard solution chromatogram. Other zones of varying intensities may be observed in the Sample solution. Spray the plate with Spray reagent A followed by Spray reagent B, and examine the plate under daylight. The Sample solution chromatogram shows multiple orange-brown zones that correspond in color and RF values to those observed in the Standard solution chromatogram. Other colored zones of varying intensities may be observed in the Sample solution.
• B.
The Sample solution exhibits peaks for speciophylline, uncarine F, mitraphylline, isomitraphylline, pteropodine, and isopteropodine at retention times that correspond to those in Standard solution A, as obtained in the test for Content of Pentacyclic Oxindole Alkaloids and Limit of Tetracyclic Oxindoles.
COMPOSITION
• Content of Pentacyclic Oxindole Alkaloids and Limit of Tetracyclic Oxindoles
Standard solution A:
Dissolve an accurately weighed quantity of USP Powdered Cat's Claw Extract RS in methanol, shaking for 1 min. Dilute with methanol to obtain a solution having a known concentration of about 0.5 mg of the labeled amount of total oxindole alkaloids per mL. Pass through a filter of 0.45-µm or finer pore size.
Standard solution B:
0.1 mg/mL of USP Isopteropodine RS in methanol. Pass through a nylon filter of 0.45-µm or finer pore size.
Sample solution:
Accurately weigh approximately 750 mg of Powdered Cat's Claw, and place in a 10-mL centrifuge tube. Sonicate with 2.5 mL of methanol for 10 min. Centrifuge, and transfer this solution to a 10-mL volumetric flask. Repeat the extraction three additional times combining the extracts in the 10-mL volumetric flask, and dilute with methanol to volume. Transfer about 3 mL of the solution to a test tube containing 300 mg of polyamide powder, and shake for 1 min. Pass through a nylon filter of 0.45-µm or finer pore size, discarding the first part of the filtrate.
Solution A:
Prepare a filtered and degassed 10 mM pH 7.0 phosphate buffer by mixing 6 mL of 1 N sodium hydroxide, 10 mL of 1 M monobasic potassium phosphate, and sufficient water to make 1000 mL. Adjust to a pH of 7.0 ± 0.1 by adding more of either solution.
Solution B:
Acetonitrile
Solution C:
Methanol and glacial acetic acid (99:1)
Mobile phase:
See Table 1.
Table 1
| Time (min) |
Solution A (%) |
Solution B (%) |
Solution C (%) |
|---|---|---|---|
| 0 | 65 | 35 | 0 |
| 17 | 65 | 35 | 0 |
| 25 | 50 | 50 | 0 |
| 30 | 50 | 50 | 0 |
| 31 | 0 | 0 | 100 |
| 36 | 0 | 0 | 100 |
| 39 | 65 | 35 | 0 |
| 49 | 65 | 35 | 0 |
Chromatographic system
Mode:
LC
Detector:
UV 245 nm
Column:
4.6-mm × 10-cm; end-capped 3-µm packing L1
Flow rate:
0.75 mL/min
Injection size:
10 µL
System suitability
Samples:
Standard solution A and Standard solution B
Suitability requirements
Chromatogram similarity:
The chromatogram obtained using Standard solution A is similar to the Reference Chromatogram provided with the USP Powdered Cat's Claw Extract RS.
Tailing factor:
NMT 2.0 for the isopteropodine peak, Standard solution B
Relative standard deviation:
NMT 2.0% for the isopteropodine peak in repeated injections, Standard solution B
Analysis
Samples:
Standard solution A, Standard solution B, and Sample solution
Measure the areas of the analyte peaks. Identify the retention times of the peaks corresponding to speciophylline, uncarine F, mitraphylline, isomitraphylline, rhynchophylline, isorhynchophylline, pteropodine, and isopteropodine by comparison of the chromatogram of Standard solution A with the Reference Chromatogram provided with the lot of the USP Powdered Cat's Claw Extract RS used.
Separately calculate the percentages of speciophylline, uncarine F, mitraphylline, isomitraphylline, rhynchophylline, isorhynchophylline, pteropodine, and isopteropodine, as isopteropodine, in the portion of Powdered Cat's Claw taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response for each relevant alkaloid from the Sample solution |
| rS | = | = peak response for isopteropodine from Standard solution B |
| CS | = | = concentration of USP Isopteropodine RS in Standard solution B (mg/mL) |
| CU | = | = concentration of Powdered Cat's Claw in the Sample solution (mg/mL) |
Calculate the content of pentacyclic oxindole alkaloids by adding the percentages of speciophylline, uncarine F, mitraphylline, isomitraphylline, pteropodine, and isopteropodine.
Calculate the content of tetracyclic oxindole alkaloids by adding the individual percentages of rhynchophylline and isorhynchophylline.
Acceptance criteria
Pentacyclic oxindole alkaloids:
NLT 0.3%
Tetracyclic oxindole alkaloids:
NMT 0.05%
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:
It meets the requirements of the tests for absence of Salmonella species and Escherichia coli. The total aerobic microbial count does not exceed 105/g; the total combined molds and yeasts count does not exceed 103/g; and the bile-tolerant Gram-negative bacteria do not exceed 103/g.
2021:
It meets the requirements of the tests for absence of Salmonella species and Escherichia coli. The total aerobic microbial count does not exceed 105/g; the total combined molds and yeasts count does not exceed 103/g; and the bile-tolerant Gram-negative bacteria do not exceed 103/g.
SPECIFIC TESTS
• Botanic Characteristics:
Presence of fragments of cork and suberized cells, with cell walls evenly thickened; presence of phelloderm sclereids; fragments of fibers crossed by vascular rays are darkened due to the presence of sand-like calcium oxalate microcrystals; solitary or two- to three-compound starch grains up to 15 µm in diameter; absence of styloids, typically present in U. guianensis
• Loss on Drying 731:
Dry at 105 for 2 h: it loses NMT 7.0% of its weight.
731:
Dry at 105 for 2 h: it loses NMT 7.0% of its weight.
• Articles of Botanical Origin, Total Ash 561:
NMT 8.0%
561:
NMT 8.0%
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in tight, light-resistant containers, and store at 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.
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 |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 1226
Pharmacopeial Forum: Volume No. 32(4) Page 1124