Chromatography column

Separated chromatography

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This entry is made by Compilation and application of scientific encyclopedia entries of "Science Popularization China" to examine.

The chromatographic column is Gel chromatography The main body in technology is usually glass tube or Plexiglass tube 。 According to the composition of the sample mixture in the stationary phase and mobile phase partition coefficient Different, thus separating Chromatography Law.

Chinese name: Chromatography column

Foreign name: Chromatography column
Category: Separated chromatography

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Specifications

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The diameter of the chromatographic column does not affect the resolution. With a large amount of samples, the diameter of the column can be increased. Generally, the gel column with a diameter greater than 2 cm should be prepared, but the samples should be evenly distributed on the gel column bed when adding samples. In addition, the larger the diameter, the larger the volume of eluant and the greater the dilution of sample. The degree of separation depends on the column height. In order to separate different components, the gel column bed must have an appropriate height. The degree of separation is related to the square root of the column height. However, because the soft gel column is too high, it is usually not more than 1m. Used for family separation short column Generally, the gel column is 20-30cm long, the column height is 5:1-10:1 compared with the diameter, and the gel bed volume is 4-10 times of the sample solution volume. Fractionation The line between column height and diameter is 20:1-100:1. The commonly used gel columns are 50 × 25cm and 10 × 25cm. Under the filter plate of chromatographic column Dead volume It should be as small as possible. If the dead volume under the palm filter plate is large, the possibility of re mixing between the separated components is large. As a result, the elution peak shape will be affected, and the trailing image will appear, reducing Resolution 。 During precise separation, the dead volume shall not exceed 1/1000 of the total bed volume^[1] 。

Operation method

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Extraction and separation by medium pressure column chromatography Oridonin For example:

1. Adsorbent treatment and column filling

A certain amount Thin layer chromatography silica gel load enamel Place the tray in the oven at 105 。 C After 2H activation, take it out and place it at room temperature to install the column. The column shall be filled with dry method to fill the whole column without leaving Dead volume After sealing, use plunger metering solvent pump as per acetone one Petroleum ether (5:95) Pump the solvent in proportion until the solvent flows out.

2. Sample loading

Apply proper amount of 400G ethanol extract of Rabdosia rubescens acetone Dissolve, pump into the middle pressure column head, and then pump into Petroleum ether 1000RID。

3. Elution

In proportion Petroleum ether one acetone by Eluate , 60ML/MIN flow rate gradient elution The operating pressure of the column is 2~3KG/C. Collect as per each 500ML, TLC detection Developing agent Acetone chloroform (2:8), Chromogenic agent 5% Vanillin concentrated sulfuric acid Ethanol solution

When the elution is complete, merge the same streams and recover the solvent. take Oridonin The streams are combined and evaporated to dryness. Add proper amount of methanol and heat dissolve it, then place it naturally to precipitate crystals, which is oridonin.

4. Regeneration and balance of chromatographic column

After TLC detection, all the active ingredients flow out acetone 5000ML regeneration, acetone I Petroleum ether (5:95) After the balance, the sample can be loaded again and used repeatedly (the number of uses varies with different raw materials, and generally can be used continuously for more than 2-6 months).

5. Recrystallization

Oridonin After complete crystallization and precipitation, Decompression filtration And then wash with a small amount of methanol（ Washing liquid Keep it for the next hot dissolved oridonin). Lying oridonin with appropriate amount of methanol, it is placed and crystallized again. After the crystallization is complete, it is filtered and a small amount of oridonin is used Ether Wash and dry the crystals at room temperature Vacuum drying oven Medium, 35 degrees vacuum drying to Constant weight 。 Sealed in a bottle dryer Save in.

6. Thin layer chromatography

Take appropriate amount Oridonin Reference substance And the oridonin sample obtained by the above method dissolved in methanol Silicone G Thin-layer plate Upper, with chloroform I acetone (8:2) is Developing agent Carry out development, 5% vanillin concentrated sulfuric acid Color of ethanol solution at 105. C Bake until the spots are clear. It shows that under this development system, oridonin A sample and control sample have the same RF value And showed the same blue-green spots. There were no impurity spots in the prepared oridonin sample.^[1]

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Registration No	GSYJX (Jin) Zi 2009 No. 1400388	Name of manufacturer (Chinese)
Product performance, structure and composition	Main composition of the product: chromatography column is composed of chromatography column, filler and sealing plug. The column is Stainless steel material ； The filler is about 0.6mL of hydrophilic polymer formed by isobutene ester synthesis; Sealing bolt is made of polyether ether ketone resin（ PEEK ）Composition.	Scope of application	This product is used for the treatment of different glycosylated hemoglobin Perform separation.
Registered Agent	Chimico Pharmaceutical Technology Development (Beijing) Co., Ltd	After sales service organization	Aikelai International Trading (Shanghai) Co., Ltd
Approval date	2009.02.26	Expiration date	2013.02.25
Name of manufacturer (English)	ARKRAY Factory, Inc.	Address of manufacturer (Chinese)	1480 Koji, Konan-cho, Koka, Shiga, JAPAN
Production site	1480 Koji, Konan-cho, Koka, Shiga, JAPAN	Country of production (Chinese)	Japan
Product name (Chinese)	Chromatography column	Product name (English)	COLUMN UNIT
Specification and model	HS-VII	Product standard	Registration standard for imported products YZB/JAP 0057-2009 Chromatographic Column

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