Lagerstroemia speciosa Leaf Powder

Lagerstroemia speciosa Leaf Powder

Proposed For Comment Version 0.2

Lagerstroemia speciosa Leaf Powder


 

DEFINITION

The article consists of the dried leaves of Lagerstroemia speciosa (L.) Pers. (Family Lythraceae) reduced to a powder or very fine powder. It contains NLT 0.2% of corosolic acid, on the dried basis.

 

POTENTIAL CONFOUNDING MATERIALS

Terminalia cuneata Roth

 

CONSTITUENTS OF INTEREST

Pentacyclic triterpene acids: Corosolic acid, virgatic acid, and oleanolic acid

 

IDENTIFICATION

• A. Botanical Characteristics

Macroscopic: Grayish-green powder

Microscopic: Fragments of upper epidermis, polygonal cells, some contain calcium oxalate crystals, no stomata; fragments of lower epidermis cells, irregular shapes, with slightly wavy walls, showing anomocytic stomata; fragments of upper epidermal cells with underlying palisade cells; fragments of parenchyma cells, some contain prisms  of calcium oxalate, other cells contain cluster crystals of calcium oxalate; oil cells; fragments of lignified fibers; fragments of spiral vessels; fragments of pitted vessels associated with fibers

B. Thin-Layer Chromatography

Standard solution A: 0.2 mg/mL of USP Corosolic Acid RS in methanol

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

Sample solution: Sonicate about 0.2 g of Lagerstroemia speciosa Leaf Powder, in 10 mL of methanol for 10 min, centrifuge, and use supernatant.

Chromatographic system

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

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

Application volume: 6 µL each of Standard solution A and Standard solution B and 8 µL 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: Toluene, ethyl acetate, and glacial acetic acid (55:45:0.5)

Developing distance: 6 cm

Derivatization reagent: Anisaldehyde reagent – 85 mL of ice-cooled methanol mixed with 10 mL of glacial acetic acid, 5 mL of sulfuric acid, and 0.5 mL of p-anisaldehyde

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 an un-saturated chamber, remove the plate from the chamber, and dry. Treat with Derivatization reagent, and heat for 3 min at 100°. Examine under visible light.

System suitability: Under visible light, the chromatogram of Standard solution B exhibits in the lower half six characteristics bands in the following order of increasing RF: a blue band close to the start, due to asiatic acid; two minor blue bands; the most intense band, a violet band, with similar RF and color as the corosolic acid band in the chromatogram of Standard solution A; a minor brownish band; and a minor blue band due to virgatic acid. Standard solution B also exhibits two minor violet bands, that are separated, at about three-fourths of the chromatogram. The band with the lower RF is due to oleanolic acid.

Acceptance criteria: Under visible light, the chromatogram of the Sample solution exhibits the most intense band is a violet band corresponding in color and RF to the band due to corosolic acid in the chromatogram of Standard Solution A, and the following bands in the lower half of the chromatogram, corresponding to similar bands in Standard solution B: a minor blue band close to the start; and two minor bands, a brownish band and a blue band, at RF above the corosolic acid band. Sample solution also exhibits two minor violet bands, that are separated, at about three-fourths of the chromatogram.

C. HPLC

Analysis: Proceed as directed in the test for Content of Corosolic Acid.

Acceptance criteria: The chromatogram of the Sample solution exhibits an intense peak at a retention time corresponding to that of corosolic acid in Standard solution A. The Sample solution chromatogram exhibits, among other peaks, a peak of lesser intensity due to virgatic acid and a minor peak due to oleanolic acid.

 

ASSAY

• Content of Corosolic Acid

Solution A: 0.1% Phosphoric acid in water

Solution B: Acetonitrile

Mobile phase: A mixture of Solution A and Solution B (4:6)

Standard solution A: 0.1 mg/mL of USP Corosolic Acid RS in methanol

Standard solution B: 5.0 mg/mL USP Lagerstroemia speciosa Powdered Extract RS in methanol; sonicate if necessary. Before injection, pass through a membrane filter of 0.45-µm or finer pore size. Discard the first few mL of the filtrate.

Sample solution: Transfer about 5.0 g of Lagerstroemia speciosa Leaf Powder, accurately weighed, to a round-bottom flask. Add 75 mL of methanol and reflux for 15 min; set aside to settle and decant the supernatant.  Repeat the extraction three more times and combine the extract, flter, concentrate under reduced pressure, transfer to a 100-mL volumetric flask, adjust with methanol to volume, and mix.  Before injection, pass through a membrane filter of 0.45-µm or finer pore size. Discard the first few mL of the filtrate.

Chromatographic system

(See Chromatography <621>, System Suitability.)

Detector: UV 205 nm

Column: 4.6-mm × 25-cm; 5-µm packing L1 (similar to Hibar® 250-4,6; Lichrospher® 100, RP-18e; or Luna C18 100A)

Flow rate: 1.6 mL/min

Injection volume: 20 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 Lagerstroemia speciosa Powdered Extract RS being used.

Resolution: NLT 1.5 between the corosolic acid peak and the peak before, Standard solution B

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

Relative standard deviation: NMT 2.0%, determined from the corosolic acid peak in repeated injections, Standard solution A

Analysis

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

Using the chromatograms of Standard solution A, Standard solution B, and the reference chromatogram provided with the lot of USP Lagerstroemia speciosa Powdered Extract RS being used, identify the retention time of the peaks corresponding to corosolic acid, virgatic acid, and oleanolic acid in the Sample solution chromatogram. The approximate relative retention times of the different peaks for corosolic acid, virgatic acid, and oleanolic acid are 1.0, 1.1, and 3.2, respectively.

Calculate the percentage of corosolic acid in the portion of Lagerstroemia speciosa Leaf Powder taken:

 

Result = (rU/rS) × CS × (V/W) × 100

 

rU    = peak area of corosolic acid from the Sample solution

rS    = peak area of corosolic acid from Standard solution A

C  = concentration of corosolic acid in Standard solution A (mg/mL)

    = volume of the Sample solution (mL)

W    = weight of Lagerstroemia speciosa Leaf Powder taken to prepare the Sample solution (mg)

Acceptance criteria: NLT 0.2% corosolic acid, calculated on dried basis

 

CONTAMINANTS

• Elemental Impurities—Procedures <233>

Acceptance criteria

Arsenic: NMT 2.0 µg/g

Cadmium: NMT 1.0 µg/g

Lead: NMT 10.0 µg/g

Mercury: NMT 1.0 µg/g

• Articles of Botanical Origin,General Method for Pesticide Residues Analysis <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

• Articles of Botanical Origin, Alcohol-Soluble Extractives, Method 1 <561>: NLT 10.0%

• Articles of Botanical Origin, Water-Soluble Extractives, Method 1 <561>: NLT 18.0%

• Loss on Drying <731>

Analysis: Dry 2.0 g of Lagerstroemia speciosa Leaf Powder at 105° for 2 h.

Acceptance criteria: NMT 10.0%

• Articles of Botanical Origin, Total Ash <561>

Analysis: 2.0 g of Lagerstroemia speciosa Leaf Powder

Acceptance criteria: NMT 7.0%

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

Analysis: 4.0 g of Lagerstroemia speciosa Leaf Powder

Acceptance criteria: NMT 2.0%

 

ADDITIONAL REQUIREMENTS

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

• Labeling: The label states the Latin binomial and the part of the plant from which the article was derived.

• USP Reference Standards <11>

USP Corosolic Acid RS

USP Lagerstroemia speciosa Powdered Extract RS

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Other Versions

Final Authorized Version 1.0