JPH0324455A - Column filler for liquid chromatography - Google Patents

Abstract

PURPOSE:To elongate the life of a column by using silica for base material, coating the surface with organic silicone having a bridge structure, and providing a layer having a polyamine layer as the outermost layer. CONSTITUTION:At first, the surface of silica which is to become base material is covered with an organic silicone layer having a bridge structure, and the dissolution of the matrix of silica is protected. Then, the reaction active group of the protecting layer is utilized, and polyamine is made to react. An amino group is introduced into the surface of the protecting layer. Here, the bridged organic silicone layer having siloxane bond is used. As the bridged organic silicone layer having syloxane bond, epoxysylane compound or its hydrolysis condensation product is used. Thus, the silica in the base material is protected with the bridged organic silicone layer. Since there is the polyamine layer on the surface of the protecting layer, the dissolution of the silica itself is suppressed, and a column filler having the long column life can be obtained.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention relates to liquid chromatography (hereinafter referred to as LC).
This article relates to column packing materials used in sugar analysis. More specifically, the present invention relates to an LC column packing material for sugar analysis that uses silica as a base material and has an excellent column life by coating the surface with a specific coating material.

[Prior art] As is well known, in LC, a separation column filled with a packing material is placed in a constant temperature bath, a liquid used as a mobile phase is forced into the column using high pressure, and the sample to be measured is separated by the column. It has a mechanism to detect objects using an ultraviolet absorption detector, a refractive index detector, etc.

Conventional LC columns for sugar analysis are broadly classified into the following four types.

■ Analysis using NH2 type silica-based packing material (normal phase mode).

■ Analysis using a sulfonic acid type ion exchanger (ligand exchange mode, + gel flow).

■ Analysis using an anion exchanger (method using a borate buffer and utilizing complex formation).

■ Analysis using immobilized lectin and boronic acid-binding packing material (affinity chromatography).

[Problems to be Solved by the Invention] Although the normal phase mode method (ii) above has versatility, it has the drawback that the column life is extremely short. This is mainly due to the following points.

A. The silica matrix is dissolved by the NH2 group.

That is, the silica of the base material is dissolved by the alkalinity of the amino group.

B, there is reaction inhibition due to side reactions during the introduction of NH2 groups. That is, as shown in the following reaction formula, some NH2 groups are chemically bonded to the silica surface, while others are not chemically bonded.

[Silica surface] -0H+ (CH30) 3 Si (
CH2 ) 3 NH20H OH [Silica surface] -08+ (CH30)3 Si(CH
2)3NH2-{CH2)3N82
Formula [II. ] Here, formula [I] is the desired reaction, and formula [1] is a side reaction. -OH of formula [I1]...
-N}12- is adsorption by hydrogen bond, not chemical bond. Therefore, there is a problem that the bond is unstable and easily released. If the amine compound is liberated, it may cause measurement errors or dissolve the silicate base material as described above.

In order to solve the problems of the conventional sugar analysis described above, the present invention uses silica as a base material, coats the surface with organic silicone having a crosslinked structure, and provides a layer having a polyamine layer as the outermost layer. This provides an LC column packing material for sugar analysis that has a long column life.

[Means for Solving the Problems] To achieve the above object, the present invention provides a column packing material for liquid chromatography using silica as a base material, in which the surface of the silica of the base material is bonded by siloxane bonds. This column packing material for liquid chromatography is coated with a crosslinked organic silicone, and has a polyamine chemically bonded to an organic group of the crosslinked organic silicone.

The main points of the present invention are as follows.

■ The surface of the silica substrate is first coated with a crosslinked organic silicone layer to protect the silica matrix from dissolving.

(2) Utilizing the reactive groups possessed by the protective layer, the polyamine is reacted to introduce amino groups onto the surface of the protective layer.

In the above, a crosslinked organic silicone layer having a siloxane bond is used as the protective layer. The crosslinked organic silicone layer having a siloxane bond may be of any type, but it is preferable to use an epoxysilane compound or a hydrolyzed condensate thereof, more preferably γ-glycidoxytrimethoxysilane (GMS), or These include its hydrolyzed condensates.

The reaction formula when GMS is used will be explained below.

For convenience of explanation, the reaction of two molecules is shown.

[Silica table i 1-OH+ (Ctobo)3S
i (C}L! )3 0CH2 CHCH2\
/ (GMS) 0 0 0H ...Formula [B] Reaction (1) of the above reaction formula [A] is carried out using a hydrolysis catalyst such as triethylamine, potassium fluoride, or acetic acid in a solvent such as toluene. Hydrolysis is carried out in the presence of water. In reaction (2) of the above reaction formula [B], an epoxy group and a polyamine are reacted to open the epoxy group and simultaneously bond a primary or secondary amino group in the polyamine. Any polyamine may be used as this polyamine. As the ring-opening catalyst for the epoxy group, the following can be used.

A. Metal acetylacetone salts such as aluminum acetylacetone, salts such as aluminum alcoholate salts.

B. Various amine compounds.

C. Acid anhydride.

D. Imidazole compound.

E. Various metal salt compounds.

The column packing material of the present invention is preferably used in an acetonitrile/water system as a solution and as a column for sugar analysis as an analytical sample.

[Function] The silica base material is protected by a crosslinked organic silicone layer (protective layer), and since a polyamine layer exists on the surface of this protective layer, the dissolution of the silica itself is suppressed, and the NH2 column has a long service life. A similar LC column packing material for sugar analysis is obtained.

[Example 1] Hereinafter, one embodiment of the present invention will be described in detail using the drawings.

After vacuum drying the silica gel (Log) at 120°C for 6 hours, 7ml of GMS, dry toluene, 1.0mA'H
201 Add some catalyst (triethylamine, potassium fluoride, etc.) and heat to reflux for 24 hours.

At this time, as shown in the above reaction formula [A], the alkoxy moiety of GMS is hydrolyzed by the action of the existing H20 and the catalyst to produce silanol, which results in (1) silanol groups on the silica surface, and (2) silylation. A synthesis reaction occurs when the silanol groups of the agent and heat are applied.

In other words, GMS molecules bond to the silica surface while polymerizing, resulting in the formation of epoxy groups (
A hydrophilic polymer layer with reactive active groups is formed on the silica surface.

After heating and refluxing, wash with toluene, acetone, and ether.
Dry at 70°C for 20br (vacuum drying).

Then, 7 ml of polyethyleneimine (Mw=600)
Silica was added to a solution of 70 ml of DMF and reacted at room temperature for 24 hours.

At this time, as shown in reaction formula [B] above, a reaction occurs between the epoxy group and the primary or secondary amino group in the polyamine, and the polyamine layer is chemically bonded onto the hydrophilic polymer layer. .

A slurry of the obtained packing material is packed into a 4Mφ×150mm column according to a conventional method.

FIG. 1 shows a schematic diagram of the silica surface obtained by the present invention. The silica surface is eventually coated with two components.

FIG. 2 shows an example in which silica having an average pore diameter of 120 μm and an average particle diameter of 3 μm is used.

The conditions are as follows.

Column = 4 lungs φ x 150 mark column Mobile phase: Acetonitrile/H2 0 = 6 5/3
5 Detector: Refractive index detector temperature = 40°C The sample is α-, β-γ-cyclodextrin and N
It can be seen that the separation is as good as with the H2 type column.

The column was stable, and similar chromatograms were obtained even after 1000 injections.

[Effects of the Invention] In order to solve the problems of the prior art described above, the present invention uses silica as a base material, coats the surface with organic silicone having a crosslinked structure, and has a layer having a polyamine layer as the outermost layer. By providing this, it was possible to provide an LC column packing material for sugar analysis that has a long column life. In other words, since protection was provided by a crosslinked organic silicone layer, the basic problem of silica dissolution could be solved.

[Brief explanation of drawings]

FIG. 1 is a model diagram of the silica surface of the column packing material of the present invention. FIG. 2 is an example of analysis of sugar (cyclodextrin) using the column packing material of the present invention.

Claims (1)

[Claims] (1) In a column packing material for liquid chromatography using silica as a base material, the surface of the silica base material is coated with a crosslinked organic silicone bonded by a siloxane bond, and a polyamine is attached to the organic group of the crosslinked organic silicone. A column packing material for liquid chromatography characterized by being chemically bonded.