CN111351892A

CN111351892A - Radix ophiopogonis detection method

Info

Publication number: CN111351892A
Application number: CN201811570788.4A
Authority: CN
Inventors: 孙秋亚; 江廷峰; 李东慧
Original assignee: Hebei Wanbang Folon Pharmaceutical Co ltd
Current assignee: Hebei Wanbang Folon Pharmaceutical Co ltd
Priority date: 2018-12-21
Filing date: 2018-12-21
Publication date: 2020-06-30

Abstract

The invention relates to the technical field of traditional Chinese medicines, and particularly relates to a detection method of radix ophiopogonis. The ruscogenin is detected by adopting a thin-layer chromatography identification method, so that the technical problem of the radix ophiopogonis identification method is solved. The method has strong durability, and can monitor the quality of the radix ophiopogonis products more effectively.

Description

Radix ophiopogonis detection method

Technical Field

The invention relates to the technical field of traditional Chinese medicines. In particular to a detection method for identifying the traditional Chinese medicine radix ophiopogonis by adopting thin-layer chromatography.

Background

Ophiopogon japonicus is recorded in the section of 'Chinese pharmacopoeia' 2015 edition, only a reference medicinal material is used as a reference under the identification item of the Ophiopogon japonicus in the original quality standard, and silica gel GF is adopted₂₅₄When the plate is unfolded for detection and observed under an ultraviolet lamp at 254nm, the spots are occasionally unclear and are difficult to compare and judge. The ruscogenin reference substance is added in the method as a reference, the silica gel G plate is adopted for unfolding, the spots are clear and identifiable, the quality of the medicinal materials can be better controlled, and the method removes the use of trichloromethane, so that the method is more environment-friendly.

Disclosure of Invention

The invention aims to solve the technical problem of designing a method for identifying and detecting the radix ophiopogonis medicinal material, so that the product quality is effectively controlled.

The invention provides a detection method for identifying a radix ophiopogonis medicinal material.

Drawings

The effect of the invention can be illustrated by fig. 1-5, and the detection method is stable and reliable.

FIG. 1 is a thin layer chromatogram for identification of Ophiopogon japonicus;

FIG. 2 is a graph of a homemade layout in a durability test;

FIG. 3 is a graph of a typical commercial plate in a durability test;

FIG. 4 is a graph of a durability test at 10-20 ℃;

FIG. 5 is a 25-30 ℃ chart in a durability test.

Detailed Description

The invention can be realized by the following scheme:

1. instrument, reagent and test sample information

(1) Instrument for measuring the position of a moving object

An electronic balance: model HK-JC-220AB

A water bath kettle: type HH-6

A drying oven: model 202-2

(2) Reagent and reagent

Methanol, hydrochloric acid, toluene, glacial acetic acid, and 10% sulfuric acid ethanol solution.

(3) Reference medicinal material

Radix ophiopogonis: the batches 121013, 201310, 181122-02, 121013, 201310, 181122-03,

121013-.

Ruscogenin: lot No. 111909-201204-160215-04, China food and drug inspection research institute.

(4) Test sample

Radix ophiopogonis: the lot numbers 2017-.

(5) Method of producing a composite material

Sample amount of spotting: 3-5 mu l

Thin-layer plate: silica gel G thin layer plate

Developing solvent toluene-methanol-glacial acetic acid (80: 5: 0.25)

Color developing agent: 10% sulfuric acid ethanol solution

Color development conditions are as follows: baking at 110 ℃ for 5-7 minutes

2. Sample assay

① preparation of control solution

Taking 3g of radix Ophiopogonis as reference material, adding 50ml of methanol, heating and refluxing for 2h, standing, taking supernatant, evaporating to dryness, adding 40ml of water into residue for dissolving, adding 1ml of hydrochloric acid, slightly boiling, heating and refluxing for 1h, cooling, transferring into a separating funnel, extracting with water saturated n-butanol for 3 times, 40ml each time, combining extractive solutions, washing with water for 2 times, 50ml each time, discarding water washing solution, recovering n-butanol to dryness, adding 2ml of methanol into residue for dissolving, and taking the residue as reference material solution.

② preparation of control solutions

Precisely weighing ruscogenin 5mg, adding methanol 5ml to obtain 1mg solution per 1ml, and making into reference solution.

③ preparation of test solution

Taking 3g of radix ophiopogonis, adding 50ml of methanol, heating and refluxing for 2h, standing, taking supernatant, evaporating to dryness, adding 40ml of water into residue for dissolving, adding 1ml of hydrochloric acid, slightly boiling, heating and refluxing for 1h, cooling, transferring into a separating funnel, extracting with water-saturated n-butanol for 3 times, 40ml each time, combining extracting solutions, washing with water for 2 times, 50ml each time, discarding water washing solution, recovering n-butanol to dryness, adding 2ml of methanol into residue for dissolving, and taking the residue as a control solution.

④ method of measurement

Performing a thin-layer chromatography test according to general rule 0502, sucking 3-5 mu l of each of a control medicinal material solution, a control solution and a sample solution, respectively dropping the control medicinal material solution, the control solution and the sample solution on the same silica gel G thin-layer plate, presaturating the silica gel G thin-layer plate for 10-15 min by using toluene-methanol-glacial acetic acid (80: 5: 0.25) as a developing agent, developing, taking out, drying, spraying a 10% sulfuric acid ethanol solution, and drying the silica gel G thin-layer plate for 5-7 min at 110 ℃ until spots are clearly developed.

⑤ test standards

Spots of the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution and the reference medicinal material.

⑥ test results

The dwarf lilyturf tuber medicinal materials (the batch numbers are 2017-217, 2018-440 and 2018-167) show spots with the same color on the positions corresponding to the color spectrum of the reference substance and the color spectrum of the reference substance. The chromatographic results are shown in FIG. 1.

3. Durability test

(1) Influence of different brands of silica gel G thin layer plates on test

① silica gel G thin layer plates of different brands, including commercially available silica gel G thin layer plate and self-made silica gel G thin layer plate

Commercially available silica gel G thin layer plates: activation was carried out for 30 minutes at 110 ℃ just before use.

The preparation method of the self-made silica gel G thin-layer plate comprises the following steps: grinding and mixing 1 part of stationary phase (silica gel G) and 3 parts of adhesive (0.25% sodium carboxymethylcellulose aqueous solution) in the same direction in a mortar, removing bubbles on the surface, pouring into a coater, smoothly moving the coater on a glass plate for coating (the thickness is 0.2-0.3 mm), taking down the coated thin-layer glass plate, placing the glass plate on a horizontal table, airing at room temperature, and drying at 110 ℃ for 30 minutes, namely, in a drying box with a drying agent for later use.

② test method

And (3) respectively dropping the prepared reference substance, the reference medicinal material solution and three batches of sample solutions on the same common commercially available silica gel G plate and a self-made silica gel G plate, presaturating for 10-15 min by taking toluene-methanol-glacial acetic acid (80: 5: 0.25) as a developing agent, developing, taking out, drying in the air, spraying with 10% sulfuric acid ethanol solution, and drying at 110 ℃ for 5-7 min until spots are clearly developed.

③ test standards

The spots of the sample, the reference substance and the reference medicinal material should be clearly distinguished, and the spots with the same color should be displayed on the corresponding positions of the chromatogram of the sample and the chromatogram of the reference medicinal material.

④ test results

The spots of the sample, the reference substance and the reference medicinal material should be clearly distinguished, and the spots with the same color should be displayed on the corresponding positions of the chromatogram of the sample and the chromatogram of the reference medicinal material. The results of the self-made plate spectrum are shown in figure 2, and the results of the common commercial plate are shown in figure 3.

(2) Effect of different temperatures on the test

① test method

Respectively dropping the prepared reference substance, reference medicinal material solution and three batches of sample solutions on the same silica gel G thin-layer plate, simultaneously preparing 2 thin-layer plates, and respectively developing at 10-20 deg.C and 25-30 deg.C with toluene-methanol-glacial acetic acid (80: 5: 0.25) as developing agent.

② test standards

At the temperature of 10-20 ℃ and 25-30 ℃, spots of the test sample, the reference sample and the reference drug are clear and distinguishable, and spots with the same color are displayed on the corresponding positions of the chromatogram of the test sample and the chromatogram of the reference drug.

The spectra of the 2 thin-layer plates should be consistent.

③ test results

The spectra of the 2 thin-layer plates are consistent. The graph results at 10-20 ℃ are shown in figure 4, and the graph results at 25-30 ℃ are shown in figure 5.

Claims

1. The detection method of radix ophiopogonis is characterized by comprising the following steps of:

preparing a contrast medicinal solution, namely taking 3g of radix ophiopogonis contrast medicinal material, adding 50ml of methanol, heating and refluxing for 2h, standing, taking supernatant to be dried by distillation, adding 40ml of water into residues for dissolving, adding 1ml of hydrochloric acid, slightly boiling, heating and refluxing for 1h, cooling, transferring into a separating funnel, extracting for 3 times with water-saturated n-butanol, 40ml each time, combining extracting solutions, washing for 2 times with water, 50ml each time, discarding a water washing solution, recovering the n-butanol to be dry, and adding 2ml of methanol into residues for dissolving to serve as the contrast medicinal solution;

preparation of control solution A solution containing ruscogenin 5mg, methanol 5ml, and 1mg per 1ml is prepared by weighing ruscogenin 5mg, and making into control solution;

preparing a test solution by taking 3g of radix ophiopogonis medicinal material, adding 50ml of methanol, heating and refluxing for 2h, standing, taking supernatant to be dried by distillation, adding 40ml of water into residue to dissolve, adding 1ml of hydrochloric acid, slightly boiling, heating and refluxing for 1h, cooling, transferring into a separating funnel, extracting with water-saturated n-butanol for 3 times, 40ml each time, combining extracting solutions, washing with water for 2 times, 50ml each time, discarding a water washing solution, recovering the n-butanol to be dry, adding 2ml of methanol into residue to dissolve, and taking the residue as a control medicinal material solution;

the determination method is tested according to thin-layer chromatography of general 0502 in Chinese pharmacopoeia, 3-5 mul of a control medicinal material solution, a control solution and a test solution are respectively sucked and respectively spotted on the same silica gel G thin-layer plate, toluene-methanol-glacial acetic acid (80: 5: 0.25) is used as a developing agent, pre-saturation is carried out for 10-15 min, the solution is developed, taken out, dried in the air, sprayed with 10% sulfuric acid ethanol solution, and dried at 110 ℃ for 5-7 min until spots are clearly developed.