Rhodiola rosea Root and Rhizome Powder

Rhodiola rosea Root and Rhizome Powder

Proposed For Comment Version 0.2

Rhodiola rosea Root and Rhizome Powder


 

DEFINITION

The article consists of the dried roots and rhizomes of Rhodiola rosea L. (Family Crassulaceae) reduced to a powder or very fine powder.  It contains NLT 0.3% of the phenylpropenoid glycosides rosarin, rosavin, and rosin calculated as rosavin, and NLT 0.08% of salidroside on the dried basis.

 

Potential Confounding Materials

Related Rhodiola species including R. kirilowii, R. yunnanensis, R. crenulata, R. sacra, and R. sachalinensis

 

Constituents of Interest

Phenylpropenoid glycosides: Rosarin, rosavin, and rosin

Salidroside

 

IDENTIFICATION

• A. Botanic Characteristics

Macroscopic: Pinkish-brown or light brown in color

Microscopic: It shows fragments of cork cells, subrectangular or polygonal, containing yellowish-brown pigments, 10–150 µm in diameter; parenchymatous cells of cortex, subsquare or polygonal, containing reddish-brown pigments; stone cells, subrounded, subtriangular, subrectangular or irregular shape, some elongated, mostly 14–70 µm in diameter, up to 270 µm in length; fibers mostly in bundles, long fusiform in shape, ends oblique-sharp or blunt-round, with oblique or criss-cross striations, 10–60 µm in diameter; and reticulate and pitted vessels, up to 120 µm in diameter.

• B. Thin-Layer Chromatography

Standard solution A: 1.0 mg/mL of USP Rosavin RS in methanol

Standard solution B: 50 mg/mL of USP Rhodiola rosea Powdered Extract RS in methanol. Sonicate for 10 min, centrifuge, and use the supernatant.

Sample solution: Sonicate about 0.5 g of Rhodiola rosea Root and Rhizome Powder in 5 mL of methanol for 10 min, centrifuge, and use the supernatant.

Chromatographic system

(See Chromatography <621>, Thin-Layer Chromatography.

Adsorbent: Chromatographic silica gel with an average particle size of 5 µm (HPTLC plates)

Application volume: 3 µL of Standard solution A and 5 µL each of Standard solution B and Sample solution, as 8-mm bands

Relative humidity: Condition the plate to a relative humidity of about 33% using a suitable device.

Temperature: 25°

Developing solvent system: Ethyl acetate, methanol, water, and formic acid (77:13:10:2)

Developing distance: 6 cm

Derivatization reagent: Dissolve 1 g of diphenylamine in acetone, add 1 mL of aniline, and mix. Carefully add 7.5 mL of phosphoric acid, and mix.

Analysis 

Samples: Standard solution A, Standard solution B, and Sample solution

Apply the Samples as bands to a suitable HPTLC plate and dry in air. Develop the chromatograms in a saturated chamber, remove the plate from the chamber, and dry. Treat with Derivatization reagent, heat for 5 min at 120°, and examine under visible light.

System suitability: The chromatogram of Standard solution B exhibits, in the lower half, three gray bands and two brownish bands, one above and the other below the gray bands. The most intense band in the chromatogram is the brownish band with an RF below the gray bands; the most intense gray band is the lower band at an RF corresponding to the band due to rosavin in the chromatogram of Standard solution A; the upper gray band due to rosarin is less intense.

Acceptance criteria: The chromatogram of the Sample solution exhibits a gray band corresponding to the band due to rosavin in the chromatogram of Standard solution A. It exhibits the following bands corresponding to similar bands in the chromatogram of Standard solution B: two additional gray bands and two brownish bands, one above and the other below the gray bands; the most intense band in the chromatogram is the brownish band with an RF below the gray bands; the most intense gray band is the lower band due to rosavin.

C. HPLC

Analysis: Proceed as directed in the Assay for Content of Phenylpropenoid Glycosides and Salidroside.

Acceptance criteria: The chromatogram of the Sample solution exhibits peaks at the retention times corresponding to the peaks due to salidroside, tyrosol, rosarin, rosavin, rosin, and rosiridin in the chromatogram of Standard solution B. The ratio of the content of phenylpropenoid glycosides (rosarin, rosavin, and rosin) to the content of salidroside is about 3:1.

 

ASSAY

• Content of Phenylpropenoid Glycosides and Salidroside

Solution A: Water

Solution B: Acetonitrile

Mobile phase: See Table 1.

 

Table 1

Time
(min)

Solution A
(%)

Solution B
(%)

0

94

6

6

83

17

7

80.3

19.7

9

80.3

19.7

10

0

100

12

94

6

17

94

6

 

Solvent: 75% Methanol in water

Standard solution A: 1.0 mg/mL of USP Rosavin RS in methanol

Standard solution B: 4.0 mg/mL of USP Rhodiola rosea Powdered Extract RS in methanol. Sonicate to dissolve, if necessary. Before injection, pass through a membrane filter of 0.45-µm or finer pore size.

Standard solution C: 1.0 mg/mL of USP Salidroside RS in methanol

Sample solution: Transfer about 5.0 g of Rhodiola rosea Root and Rhizome Powder, accurately weighed, to a 25-mL flask. Add 7 mL of Solvent, sonicate at 37° for 15 min, and filter into a 10-mL volumetric flask. Wash the residue on the filter paper twice, using 1 mL of Solvent each time. Add the washings to the volumetric flask, dilute with Solvent, and mix. Before injection, pass through a membrane filter of 0.45-µm or finer pore size, discarding the first few mL of the filtrate.

Chromatographic system 

(See Chromatography <621>, System Suitability.)

Mode: LC

Detector: UV 205 nm

Column: 3.0-mm × 10-cm; 2.5-µm packing L1 (similar to Luna C18-HST)

Column temperature: 40±1°

Flow rate: 1.0 mL/min

Injection volume: 1 µL

System suitability 

Samples: Standard solution A and Standard solution B

Suitability requirements 

Chromatogram similarity: The chromatogram from Standard solution B is similar to the reference chromatogram provided with the lot of USP Rhodiola rosea Powdered Extract RS being used.

Resolution: NLT 1.5 between the rosarin and rosavin peaks, Standard solution B

Tailing factor: NMT 2.0 for the rosavin peak, Standard solution A

Relative standard deviation: NMT 2% determined from the rosavin peak in repeated injections, Standard solution A

Analysis 

Samples: Standard solution A, Standard solution B, Standard solution C, and Sample solution

Using the chromatograms of Standard solution A, Standard solution B, Standard solution C, and the reference chromatogram provided with the lot of USP Rhodiola rosea Powdered Extract RS being used, identify the retention time of the peaks corresponding to salidroside, tyrosol, rosarin, rosavin, rosin, and rosiridin in the Sample solution.

Separately calculate the percentages of rosarin, rosavin, and rosin as rosavin:

 

Result = (rU/rS) × CS × (V/W) × 100

 

rU    = peak area of the relevant analyte in the Sample solution

rS    = peak area of rosavin in Standard solution A

CS   = concentration of rosavin in Standard solution A (mg/mL)

    = volume of the Sample solution (mL)

W    = weight of Rhodiola rosea Root and Rhizome Powder taken to prepare the Sample solution (mg)

Calculate the percentage of phenylpropenoid glycosides as the sum of the percentages of rosarin, rosavin, and rosin.

Calculate the percentage of salidroside:

 

Result = (rU/rS) × CS × (V/W) × 100

 

rU     = peak area of salidroside in the Sample solution

rS     = peak area of salidroside in Standard solution C

CS    = concentration of salidroside in Standard solution C (mg/mL)

     = volume of the Sample solution (mL)

W     = weight of Rhodiola rosea Root and Rhizome Powder taken to prepare the Sample solution (mg)

Acceptance criteria

Phenylpropenoid glycosides: NLT 0.3% on the dried basis

Salidroside:  NLT 0.08% on the dried basis

 

CONTAMINANTS

Elemental Impurities—Procedures <233>

Acceptance criteria

Arsenic: NMT 2.0 µg/g

Cadmium: NMT 1.0 µg/g

Lead: NMT 5.0 µg/g

Mercury: NMT 1.0 µg/g

Articles of Botanical Origin, General Method for Pesticide Residues Analysis <561>: Meets the requirements

Articles of Botanical Origin, Test for Aflatoxins <561>: Meets the requirements

Microbial Enumeration Tests <61>: The total aerobic bacterial count does not exceed 105 cfu/g, the total combined molds and yeasts count does not exceed 103 cfu/g, and the bile-tolerant Gram-negative bacteria does not exceed 103 cfu/g.

Tests for Specified Microorganisms <62>: Meets the requirements of the tests for the absence of Salmonella species and Escherichia coli

 

SPECIFIC TESTS

Loss on Drying <731>

Analysis: Dry 1.0 g of Rhodiola rosea Root and Rhizome Powder at 105° for 2 h.

Acceptance criteria: NMT 12.0%

Articles of Botanical Origin, Total Ash <561>

Analysis: 2.0 g of Rhodiola rosea Root and Rhizome Powder

Acceptance criteria: NMT 12.0%

Articles of Botanical Origin, Acid-Insoluble Ash <561>

Analysis: 2.0 g of Rhodiola rosea Root and Rhizome Powder

Acceptance criteria: NMT 3.0%

 

ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in well-closed containers, protected from light and moisture. Store at room temperature.

Labeling: The label states the Latin binomial and the parts of the plant from which the article was obtained.

USP Reference Standards <11>

USP Aflatoxins RS

USP Rhodiola rosea Powdered Extract RS

USP Rosavin RS

USP Salidroside RS

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