Lonicera japonica Flower Dry Extract. Proposed For Development Version 0.1. Published on Herbal Medicines Compendium (

Transcription

1 Published on Herbal Medicines Compendium ( Lonicera japonica Flower Dry Extract Proposed For Development Version 0.1 Lonicera japonica Flower Dry Extract DEFINITION The article is prepared from the dried flower buds or in early opening flowers of Lonicera japonica Thunb. (Family Caprifoliaceae), collected before flowering in early summer, by extraction with water or hydroalcoholic mixtures. The ratio of starting crude plant material to Dry Extract is between 10:1 and 5:1. It contains NLT 90.0% and NMT 110.0% of the labeled amount of caffeoylquinic acids, calculated as the sum of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, 3,4-di-O-caffeoylquinic acid, 3,5-di-O -caffeoylquinic acid and 4,5-di-O-caffeoylquinic acid on the dried basis; and NLT 90.0% and NMT 110.0% of the labeled amount of iridoids, calculated as the sum of sweroside, secoxyloganin and centauroside on the dried basis. POTENTIAL CONFOUNDING MATERIALS Lonicera macranthoides Flower Lonicera hypoglauca Flower Lonicera confusa Flower Lonicera fulvotomentosa Flower CONSTITUENTS OF INTEREST Iridoids: Secoxyloganin, Sweroside, Centauroside Caffeoylquinic acids: Neochlorogenic acid, Chlorogenic acid, Cryptochlorogenic acid, 3,4-di-O-Caffeoylquinic acid, 3,5-di-O -Caffeoylquinic acid and 4,5-di-O-Caffeoylquinic acid Flavone: Luteolin-7-O-glucoside, Rutin IDENTIFICATION A. THIN-LAYER CHROMATOGRAPHY Standard solution A: 1.0 mg/ml of USP chlorogenic acid RS and 0.5 mg/ml USP luteolin-7-o-glucoside RS in methanol Standard solution B: 5.0 mg/ml of USP Lonicera japonica Flower Dry Extract RS in methanol, sonicate for 10 min and filter. Sample solution: 5.0 mg of Lonicera japonica Flower Dry Extract in 10 ml methanol, sonicate for 10 min and filter. (See Chromatography <621>, Thin-Layer Chromatography) Adsorbent: Use a chromatographic silica gel mixture with an average particle size of 5 µm (HPTLC plates) Application volume: 5 µl as 8-mm bands Relative Humidity: Condition the plate to a relative humidity of about 50% using a suitable device Temperature: 25º Developing solvent system: The upper layer solution of the mixture of ethyl acetate, formic acid and water (7:5:5) Developing distance: 6 cm Derivatization reagent: A solution of 3% aluminum chloride in ethanol Samples: Standard solution A, Standard solution B, and Sample solution Apply the samples as bands to a suitable high performance thin-layer chromatographic plate and dry in air. Develop the chromatograms in a saturated chamber, remove the plate from the chamber, and dry in air. Derivatize with Derivatization reagent, heat at 105º for 5 min, and examine immediately under UV light at 366 nm. System suitability: Under UV light at 366 nm, the chromatogram of Standard solution B exhibits four bands in the lower-third section; one blue fluorescent band corresponds in Rf to the band due to chlorogenic acid in the chromatogram of Standard solution A; one yellow band above chlorogenic acid corresponds in Rf to the band due to luteolin-7-o-glucoside in the chromatogram of Standard solution A; one faint blue fluorescent band is below chlorogenic acid; another yellow band below the faint blue fluorescent band is due to rutin.

2 Standard solution B exhibits three blue fluorescent bands in the middle-third section of the chromatogram; the blue fluorescent band with a highest Rf is due to 3,5-di-O-caffeoylquinic acid; the faint blue fluorescent band with a midle Rf is due to 4,5-di-O-caffeoylquinic acid; another faint blue fluorescent band is below 4,5-di-O-caffeoylquinic acid. Acceptance criteria: Under UV light at 366 nm, the Sample solution chromatogram exhibits an intense blue fluorescent band and a yellow band at Rfs corresponing to the bands due to chlorogenic acid and luteolin-7-o-glucoside in the chromatogram of Standard solution A, respectively. The Sample solution exhibits additional bands corresponding to similar bands in the chromatogram of Standard solution B, these includ one faint blue fluorescent band below chlorogenic acid, another yellow band below the faint blue fluorescent band in lower-third section; and three fluorescent bands in middle-third section. C. HPLC : Proceed as directed in the test for Content of Caffeoylquinic Acids. Acceptance criteria: The chromatogram of the Sample solution exhibits an intense peak with a retention time corresponding to chlorogenic acid in Standard solution A, the peaks related to caffeoylquinic acids for neochlorogenic acid, cryptochlorogenic acid, 3,4-di- O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid and 4,5-di-O-caffeoylquinic acid at retention times corresponding to the same caffeoylquinic acids in the Standard Solution B and the Reference Chromatogram provided with the lot of USP Lonicera japonica Flower Dry Extract RS being used. The content ratios for each compound versus chlorogenic acid are listed in Table 2. D. HPLC : Proceed as directed in the test for Content of Iridoids. Acceptance criteria: The chromatogram of Sample solution exhibits a peak with a retention time corresponding to secoxyloganin in Standard solution A, the peaks related to iridoids for sweroside and centauroside at retention times corresponding to the same iridoids in the Standard solution B and the Reference Chromatogram provided with the lot of USP Lonicera japonica Flower Dry Extract RS being used ASSAY CONTENT OF CAFFEOYLQUINIC ACIDS Solution A: 0.1% Phosphoric acid in water Solution B: Acetonitrile Mobile phase: See Table 1. Table 1 Time (min) A (%) B (%) [NOTE: Protect from light and proceed under low actinic light. The Standard solution and Sample solution are stable for 24 h at room temperature.] Standard solution A: 0.30 mg/ml of USP Chlorogenic Acid RS in methanol Standard solution B: 5.0 mg/ml of USP Lonicera japonica Flower Dry Extract RS in 75% methanol, sonicate for 15 min, and pass through a membrane filter of 0.22 μm or finer pore size. Sample solution: Accurately transfer about 50 mg of Lonicera japonia Flower Dry Extract into a suitable stoppered conical flask, accurately add 10 ml of 75% methanol. Weigh the filled flask with a precision of ± 0.1 mg and then sonicate for 30 min. After cooling to room temperature, adjust the weight to initial by adding methanol. Pass through a membrane filter of 0.22 μm or finer pore size, and discard the first portion of the filtrate. (See Chromatography <621>, System Suitability)

3 Mode: LC Detector: UV 327 nm Column: 4.6 mm 25 cm; 5-μm packing L1 (similar to Syncronis C18) Column temperature: 15 ± 1º Flow rate: 0.7 ml/min Injection volume: 2 µl System suitability Sample: Standard solution A and Standard solution B Suitability requirements Chromatogram similarity: The chromatogram of Standard solution B is similar to the reference chromatogram provided with the lot of USP Lonicera japonica Flower Dry Extact RS being used. Resolution: NLT 1.5 betwen the peak of chlorogenic acid and the peak of cryptochlorogenic acid, Standard solution B Tailing factor: NMT 2.0 for the peak of chlorogenic acid, Standard solution A Relative standard deviation: NMT 2.0% for chlorogenic acid, Standard solution A 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 Lonicera japonica Flower Dry Extract RS being used, identify the retention times of the peaks corresponding to neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, 3,4-di-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid and 4,5-di-O-caffeoylquinic acid. The approximate relative retention times (RRT), relative to chlorogenic acid, are provided in Table 2. (The RRTs are variable due to different column, mobile phase flow rate and so on.) Table 2 Analyte Approximate Relative Retention Time Conversion Factor Content Ratio neochlorogenic acid chlorogenic acid cryptochlorogenic acid ,4-di-O-caffeoylquinic acid ,5-di-O-caffeoylquinic acid ,5-di-O-caffeoylquinic acid Separately calculate the percentage of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, 3,4-di-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid, 4,5-di-O-caffeoylquinic acid in the portion of Lonicera japonica Flower Dry Extyract taken: Result = (r U /r S ) C S (V/W) F 100 r U = peak area of relevant analyte from the Sample solution r S = peak area of chlorogenic acid from the Standard solution A C S = concentration of USP Chlorogenic Acid RS in the Standard solution A (mg/ml) V = volume of the Sample solution (ml) W = weight of Lonicera japonica Flower Dry Extract taken to prepare the Sample solution (mg) F = conversion factor for analytes as provided in Table 2. Separately calculate the content of caffeoylquinic acids as the sum of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, 3,4-di-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid and 4,5-di-O-caffeoylquinic acid on the dried basis. Calculate the percentage of the labeled amount of caffeoylquinic acids in the Extract: Result = (P/L) 100 P = content of caffeoylquinic acids as determined above (%) L = Labeled amount of caffeoylquinic acids (%) Acceptance criteria: 90.0% % of the labeled amount of caffeoylquinic acids on the dried basis.

4 CONTENT OF IRIDOIDS Solution A, Solution B, Mobile phase, Standard solution B and Sample solution are prepared in the same way as in the test for Content of Caffeoylquinic Acids. Standard solution A: 0.05 mg/ml of USP Secoxyloganin RS in methanol (See Chromatography <621>, System Suitability) Mode: LC Detector: UV 240 nm Column: 4.6 mm 25 cm; 5-μm packing L1 (similar to Syncronis C18) Column temperature: 15 ± 1º Flow rate: 0.7 ml/min Injection volume: 2 µl System suitability Sample: Standard solution A and Standard solution B Suitability requirements Chromatogram similarity: The chromatogram of Standard solution B is similar to the reference chromatogram provided with the lot of USP Lonicera japonica Flower Dry Extact RS being used. Resolution: NLT 1.5 betwen the peak of secoxyloganin and the peak before it, Standard solution B Tailing factor: NMT 2.0 for the peak of secoxyloganin, Standard solution A Relative standard deviation: NMT 2.0% for secoxyloganin, Standard solution A 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 Lonicera japonica Flower Dry Extract RS being used, identify the peaks of sweroside, secoxyloganin, and centauroside. The approximate relative retention times for the peaks of sweroside, secoxyloganin, and centauroside are 0.85, 1.00 and 1.80, respectively Separately calculate the percentage of secoxyloganin, sweroside and centauroside in the portion of Lonicera japonica Flower Dry Extract taken: Result = (r U /r S ) C S (V/W) F 1100 r U = peak area of relevant analyte from the Sample solution r S = peak area of secoxyloganin from the Standard solution A C S = concentration of USP Secoxyloganin RS in the Standard solution A (mg/ml) V = volume of the Sample solution (ml) W = weight of Lonicera japonica Flower Dry Extract taken to prepare the Sample solution (mg) F = conversion factor for analytes (1.00 for secoxyloganin, 1.03 for sweroside, and 0.89 for centauroside) Calculate the content of iridoids as the sum of secoxyloganin, sweroside and centauroside on the dried basis. Calculate the percentage of the labeled amount of iridoids in the Extract: Result = (P/L) 100 P = content of iridoids as determined above (%) L = Labeled amount of iridoids (%) Acceptance criteria: 90.0% % of the labeled amount of iridoids on the dried basis. CONTAMINANTS ELEMENTAL IMPURITIES PROCEDURES <233> Acceptance criteria Arsenic: NMT 2.0 µg/g Cadmium: NMT 0.3 µg/g Lead: NMT 5.0 µg/g Mercury: NMT 0.2 µg/g Copper: NMT 20.0 µg/g

5 ARTICLES OF BOTANICAL ORIGIN, General Method for Pesticide Residues <561>: Meets the requirements. MICROBIAL ENUMERATION TESTS <2021>: The total aerobic bacterial count does not exceed 10 5 cfu/g, the total combined molds and yeasts count does not exceed 10 3 cfu/g, and the bile-tolerant Gram-negative bacteria does not exceed 10 3 cfu/g. TESTS FOR SPECIFIED MICROORGANISMS <2022>: Meets the requirements of the tests for absence of Salmonella species and Escherichia coli. ARTICLES OF BOTANICAL ORIGIN, Aflatoxins <561>: Meets the requirements. SPECIFIC TESTS Limit of Triterpenoid Saponins by TLC Standard solution: 1.0 mg/ml of USP macranthoidin B RS, and 1.0 mg/ml USP dipsacoside B RS in methanol. Sample solution: 5.0 mg of Lonicera japonica Flower Dry Extract in 10 ml methanol, sonicate for 10 min and filter. (See Chromatography <621>, Thin-Layer Chromatography) Adsorbent: Use a chromatographic silica gel mixture with an average particle size of 5 µm (HPTLC plates) Application volume: 3 µl as 8-mm bands Relative Humidity: Condition the plate to a relative humidity of about 50% using a suitable device Temperature: 25º Developing solvent system: The upper layer solution of a mixture of n-butanol, formic acid and water (4:1:5) Developing distance: 6 cm Derivatization reagent: A solution of 10% sulphuric acid in ethanol Samples: Standard solution and Sample solution Apply the samples as bands to a suitable high performance thin-layer chromatographic plate and dry in air. Develop the chromatograms in a saturated chamber, remove the plate from the chamber, and dry in air. Derivatize with Derivatization reagent, heat at 105º for 5 min, and examine immediately under UV light at 366 nm. System suitability: Under UV light at 366 nm, the chromatogram of Standard solution exhibits two clearly separated bands in the lower-third section, the band with a higher Rf is due to dipsacoside, the band with a lower Rf is due to macranthoidin B. Acceptance criteria: Under UV light at 366 nm, the Sample solution chromatogram do not exhibits bands at Rfs and colour corresponing to the bands due to dipsacoside and macranthoidin B in the chromatogram of Standard solution; the Sample solution chromatogram do not exhibits bands with similar color as the bands of dipsacoside and macranthoidin B, and with Rfs between the bands of dipsacoside and macranthoidin B. LOSS ON DRYING <731>: : Dry 1 g of Lonicera japonica Flower Dry Extract at 105º for 2 h. Acceptance criteria: NMT 5.0% ADDITIONAL REQUIREMENTS PACKAGING AND STORAGE: Preserve in well-closed containers, store at a cool and dry place, protect from light, moisture and moth. LABELING: The label states the Latin binomial and the part(s) of the plant from which the article was prepared. It meets other labeling requirements under <565> Botanical Extracts. USP REFERENCE STANDARDS <11> USP Lonicera japonica Flower Dry Extract RS USP Secoxyloganin RS USP Chlorogenic Acid RS USP Luteolin-7-O-Glucoside RS USP Dipsacoside RS USP Macranthoidin B RS Source URL (modified on 2014/06/13-2:53pm):