Terminalia chebula Fruit

Terminalia chebula Fruit

Proposed For Comment Version 0.3

Terminalia chebula Fruit

 


DEFINITION

The article consists of the pericarp of dried mature fruit of Terminalia chebula Retz. (Family Combretaceae). It contains NLT 15% of hydrolyzable tannins calculated as the sum of chebulagic acid and chebulinic acid, on the dried basis.

 

SYNONYMS

Myrobalanifera citrina Houtt.

Myrobalanus chebula (Retz.) Gaertn.

Myrobalanus gangetica (Roxb.) Kostel.

Terminalia gangetica Roxb.

Terminalia reticulata Roth.

 

POTENTIAL CONFOUNDING MATERIALS

None known.

 

SELECTED COMMON NAMES

Chinese: 诃子, 訶梨勒

English: Myrobalan, black myrobalan, chebulic myrobalan, gali nut, Indian gall-nut, ink nut, yellow myrobalan

French: Badamier chébule, Myrobalan chébule

German: Chebulische Myrobalane, Rispiger Myrobalanenbaum

Hindi: हर्रा,  हरीतक

Japanese: ハリタキ, イ エロ-ターミナリア, ミ ロバランノキ

Russian: Харитаки

Spanish: Mirobalanos índicos

Thai: สมอ ไทย

 

CONSTITUENTS OF INTEREST

Hydrolyzable tannins: Chebulagic acid and chebulinic acid

Triterpene: Arjungenin

 

IDENTIFICATION

• A. Botanical Characteristics

Macroscopic: The surface of the dry fruit is somewhat wrinkled and shows five slightly thick but well-defined longitudinal ridges that are 2–3 mm wide and 2 mm thick. The surface color varies from light yellowish-brown to a nearly uniform brown with yellowish markings or patches. In some fruits, the basal portion is narrower and somewhat elongated or tapering.

Microscopic: The epidermis consists of a single layer, the cells are rectangular and tangentially elongated, and covered with a thick cuticle. Below the epidermis, the outer mesocarpic region consists of 6–10 layers of cells filled with abundant starch grains and tannin. In the mesocarpic region, sclereids are present, about 4–6 cells thick and arranged tangentially. The sclerified cells are thick-walled, lignified, and elongated. Below the sclerenchymatous layer occurs a layer of 20–25 mesocarpic cells which are parenchymatous in nature, oval to round in shape, and filled with starch grains and tannin. In the mesocarpic region, vascular strands occur at a certain place and consist only of spiral elements. The mesocarpic cells contain abundant rosettes of calcium oxalate. Starch grains present in cells of the outer and inner mesocarpic regions are simple or compound, and oval to round.

• B.  HPTLC for Articles of Botanical Origin <203>

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

Standard solution B: 100 mg/mL of USP Terminalia chebula Fruit Dry Extract RS in methanol. Sonicate for 10 min, centrifuge or filter, and use the supernatant or the filtrate.

Sample solution: Sonicate about 1.0 g of Terminalia chebula Fruit, finely powdered, in 100 mL of methanol for 10 min, centrifuge or filter, and use the supernatant or the filtrate.

Chromatographic system

Adsorbent: Chromatographic silica gel F254 mixture with an average particle size of 5 µm

Application volume: 5 µL of Standard solution A, and 1 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 formate, toluene, formic acid, and water (30: 1.5: 4: 3)

Developing distance: 7 cm

Derivatization reagent: Natural products reagent (NP reagent): 1 g of diphenylboinic acid aminoethylester dissolved in 200 mL of ethyl acetate

Analysis

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

Apply the Samples as bands and dry in air. Develop in a saturated chamber (20 min with filter paper), remove the plate from the chamber, and dry in air. Examine under UV light at 254 nm. Heat the plate then treat the plate at 100° for 3 min, then treat with the Derivatization reagent. Examine under UV light at 365 nm.

System suitability: Under UV 254 nm prior to derivatization, the chromatogram of Standard solution B exhibits an intense band in the lower half similar in RF and color to the chebulagic acid band in the chromatogram of Standard solution A. Another intense band above the chebulagic acid band is the chebulinic acid band. Two additional bands appear above the chebulinic acid band, one near RF 0.4 and another near RF 0.7. Two additional bands appear below the chebulagic acid band. Under UV 365 nm after derivatization, about six intense blue fluorescence bands appear in the chromatogram. A pale blue band similar in RF and color to the chebulagic acid band also appears.

Acceptance criteria: Under UV 254 nm prior to derivatization, the chromatogram of Sample solution exhibits two intense bands in the lower half of the chromatogram with the lower band corresponding in RF and color to the chebulagic acid in the chromatogram of Standard solution A. One intense band above chebulagic acid band is chebulinic acid. About four additional bands appear, two broad bands above chebulinic acid, one near an RF of approximately 0.7, and one below chebulagic acid. Under UV 365 nm after derivatization, about four pale blue fluorescence bands appear in the lower half, and one blue fluorescence band appears with an RF of approximately 0.7. A greenish white band near RF 0.5, characteristic to Terminalia bellirica, should not be observed.

• C. HPLC

Analysis: Proceed as directed in the Assay for Content of Constituents of Hydrolyzable Tannins.

Acceptance criteria: The chromatogram of the Sample solution exhibits peaks at retention times corresponding to those for chebulagic acid, ellagic acid, and chebulinic acid in Standard solution B. The most prominent peak in the chromatogram corresponds to ellagic acid, followed by chebulagic acid, which appears before ellagic acid, and chebulinic acid appears after ellagic acid, with almost equal responses.

 

ASSAY

• Content of Constituents of Hydrolyzable Tannins

Solution A: Dissolve 140 mg of monobasic potassium phosphate in 900 mL of water, add 0.5 mL of o-phosphoric acid, dilute with water to 1 L, and mix.

Solution B: Acetonitrile

Mobile phase: See Table 1.

 

Table 1

Time
(min)

Solution A
(%)

Solution B
(%)

0

100

0

7

92

8

12

85

15

22

80

20

25

50

50

28

100

0

32

100

0

Standard solution A: 0.3 mg/mL each of USP Chebulagic Acid RS in methanol

Standard solution B: Dissolve 0.1 g of USP Terminalia chebula Fruit Dry Extract RS in 100 mL of boiling water, sonicate, and pass through a membrane filter of 0.45-μm or finer pore size.

Sample solution: Transfer 0.3 g of Terminalia chebula Fruit, finely powdered and accurately weighed, to a 100-mL volumetric flask. Add 70 mL of methanol and reflux at 80° for 30 min. Cool down and add 30 mL of water. Sonicate for 5 min and dilute with water to volume. Pass through a membrane filter of 0.45-µm pore size.

Chromatographic system

(See Chromatography <621>, System Suitability.)

Detector: UV 254 nm

Column: 4.6-mm x 25-cm; 5-µm packing L1 (similar to Phenomenex Luna C18)

Column temperature: 25°

Flow rate: 1.5 mL/min

Injection volume: 10 µL

System suitability

Samples: Standard solution A and Standard solution B

Suitability requirements

Resolution: NLT 2.0 between chebulagic acid and the peak after, Standard solution B

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

Relative standard deviation: NMT 2.0% for the chebulagic acid peak in repeated injections, Standard solution A

Chromatogram similarity: The chromatogram of Standard solution B is similar to the reference chromatogram provided with the lot of USP Terminalia chebula Fruit Dry Extract RS being used.

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 Terminalia chebula Fruit Dry Extract RS being used, identify the retention times of the peaks corresponding to chebulagic acid and chebulinic acid in the Sample solution. The approximate relative retention times of the peaks for chebulagic acid and chebulinic acid are 1.0 and 1.21, respectively.

Separately calculate the percentages of chebulagic acid and chebulinic acid in the portion of Terminalia chebula Fruit taken:

 

Result = (rU/rS) x CS x (V/W) x F x 100

 

rU     = peak area of the analyte from the Sample solution

rS     = peak area of chebulagic acid from Standard solution A

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

V      = volume of the Sample solution (mL)

W     = weight of Terminalia chebula Fruit taken to prepare the Sample solution (mg)

F      = conversion factor for the analytes (1 for chebulagic acid and 1.14 for chebulinic acid)

 

Add the percentages of chebulagic acid and chebulinic acid.

Acceptance criteria: NLT 15% on the dried basis

 

CONTAMINANTS

• Elemental Impurities—Procedures <233>

Acceptance criteria

Arsenic: NMT 2.0 µg/g

Cadmium: NMT 0.5 µg/g

Lead: NMT 5.0 µg/g

Mercury: NMT 0.2 µg/g

• Articles of Botanical Origin <561>, Pesticide Residues Analysis: 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 <561>, Methods of Analysis, Foreign Organic Matter: NMT 1.0%

• Articles of Botanical Origin <561>, Methods of Analysis, Alcohol-Soluble Extractives, Method 1: NLT 40.0%

• Articles of Botanical Origin <561>, Methods of Analysis, Water-Soluble Extractives, Method 2: NLT 60.0%

• Loss on Drying <731>

Sample: 1 g of Terminalia chebula Fruit, finely powdered

Analysis: Dry the Sample at 105° for 2 h.

Acceptance criteria: NMT 8%

• Articles of Botanical Origin <561>, Methods of Analysis, Total Ash: NMT 15%

• Articles of Botanical Origin <561>, Methods of Analysis, Acid-Insoluble Ash: NMT 5%

 

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(s) of the plant contained in the article.

• USP Reference Standards <11>

USP Chebulagic Acid RS

USP Terminalia chebula Fruit Dry Extract RS

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Other Versions

Proposed For Comment Version 0.2
Proposed For Development Version 0.1