JPH0196009A - Method for stabilizing against hydrolysis of aminoalkyl silica by crosslinking together with transforming amino groups into tertiary forms - Google Patents

Abstract

PURPOSE:To extend the life of separation column, by using column packing obtd. by crosslinking between amino groups by reacting aminoalkyl bound silica with halomethyl compd., together with transforming them into tertiary amino groups. CONSTITUTION:The aminoalkyl silica of formula I or II (where, R is hydrocarbon residue having 2-10 carbon atoms) is prepd. The halomethyl compd. of formula III or IV (where, X is halogen atom R' is epoxy residue of CnH2n+1O, ether residue of CnH2n+1O or alkylphenyl group of formula V, R'' is hydrocarbon residue of CnH2n, carbonyl group CO, S atom, phenylene group of formula VI or alcohol residue of formula VII) is prepd. The halomethyl compd. is reacted with the above-mentioned aminoalkyl silica to crosslink between the amino groups, together with transforming the amino groups into tertiary forms.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a novel method for improving packing materials for separation columns used in liquid chromatography.

[Prior art] Aminoalkyl-bonded silica is a column packing material for normal phase chromatography, but it is also used in mobile phase systems that utilize the ion exchange properties of amino groups, and in mixtures of water and organic solvents such as in the separation of sugars. It is used in a wide range of fields, including mobile phase systems. However, when a mobile phase containing water, particularly a mobile phase with a pH of 8 or higher, is used, the aminoalkylsilyl group is hydrolyzed and eliminated, resulting in a corresponding decrease in separation ability. Alternatively, if a carbonyl compound, acid amide, acid chloride, thionyl chloride, halomethyl compound, etc. is present in the sample, it will react with the amino group hydrogen of the packing material and change to the corresponding substituent, resulting in separation. This brings about a change in Noh. Since the aminoalkyl-bonded silica conventionally used contains primary to secondary amino groups, it has an extremely short life as a filler and has the drawback of requiring frequent replacement of the filler.

[Problems to be solved by the invention] Primary to secondary amino groups react extremely easily with carbonyl compounds, acid anhydrides, acid chlorides, dionyl chloride compounds, halomethyl compounds, etc., resulting in relatively stable substitutions. compounds such as Schiff bases, acid amides, and tertiary amino compounds. Acylation of amino groups with acid chlorides or acid anhydrides has already been carried out, but since the product is an acid amide, there are drawbacks such as the inability to obtain the separation ability inherent in amino groups. Thionyl chloride compounds have similar problems. Compared to these, the behalomethyl compound is a tertiary amino compound, but the separation ability inherent in the amino base is hardly impaired, and moreover, the amino groups can be crosslinked, making it an extremely stable bonding group against hydrolysis. The present invention applies this principle.

[Means for solving problems] E S r RN
II or 35iRN11 (C111C111N1
(). ~,. The aminoalkyl-bonded silica represented by -CIltCIItNIIT can be
1. Reacts with a halomethyl compound represented by [l″CIl!X, III IIIIII
1ll-N-C11*-R
By modifying to aminoalkyl-bonded silica with a crosslinked structure represented by a general formula such as '-CII*4-C11t-R'-C1lt-N-', the conventional separation ability unique to amino groups is impaired. Improves hydrolytic stability without oxidation.

[Function] The present invention reacts conventional primary to secondary aminoalkyl-bonded silica with a halomethyl compound to modify it into aminoalkyl-bonded silica in which amino groups are crosslinked, thereby creating a filling with good hydrolytic stability. This is a method for preparing a drug.

Further, the separation characteristics can be changed by selecting the halomethyl compound to be reacted. The details will be explained below.

Aminopropyl-bonded silica 5i-CIItClltCIItN, which has been conventionally used as a normal phase packing material
IIt undergoes hydrolysis in a water-containing mobile phase of 9118 or higher to eliminate the aminopropylsilyl group. In contrast, the tertiary cross-linked aminopropyl-bonded silica of the present invention has good hydrolytic stability, so that the bonding group does not detach.

An example is shown in FIG.

The product of the present invention, which has been developed by Kashinaka, is a tertiary cross-linked aminopropyl-bonded silica obtained by reacting conventional aminopropyl-bonded silica with 1,3-dibromopropane [bis(bromomethyl)methane], and is clearly stable. .

In this example, each silica was placed in a flask and the pH was adjusted to 1.
0.0 buffer solution was added, the temperature was maintained at 50° C. with stirring, and after hydrolysis for a predetermined period of time, the silica in the flask was collected, thoroughly washed and dried, and its nitrogen content was measured using a conventional method. The nitrogen retention rate on the vertical axis is the nitrogen content before hydrolysis.
It is shown as 0.

In addition, after 60 minutes of hydrolysis, the silica was washed and dried, and
Examples of chromatograms in which monosaccharides were separated using a mixed solvent of acetonitrile and water as a mobile phase packed in a x4.6 mm stainless steel column are shown in FIGS. 2 and 3, respectively. As is clear from this, the separation ability of the product of the present invention is good, whereas the separation ability of the conventional product after hydrolysis is also lower than that of the untreated product.

The above results were exactly the same even when the type of halomethyl compound was changed, and the products of the present invention which were tritreated and crosslinked were extremely stable against hydrolysis, and several examples thereof are shown in Table 1. Conventional product A in the table
is aminopropyl-bonded silica, and B is N-2-aminoethyl-3-aminopropyl-bonded silica. In addition, the hydrolytic stability of each filler was 4.6 mm to 150.
Fill the column with 1 ml of pl+9 buffer per minute.
The retention time of adenosine diphosphate was measured, and the ratio before and after the fluid flow was expressed as the retention capacity.

Table 1 Comparison of hydrolytic stability (retention ability) between conventional product and product of the present invention Conventional product A 582
Conventional product B 703 l is tertiaryized and cross-linked with bis(chloromethyl)ether.
Tertiary crosslinking of 9641 with 1,2-dibromoethane 985! α
9962 was tertiary-crosslinked with α°-dichloro-p-xylene, and 9972 was tertiary-crosslinked with 1,2-dibromoethane.
Tertiary cross-linked product with 3-dibromopropane Tertiary cross-linked product of 9982 with chloromethyloxirane 99 As is clear from Table 1, when a buffer solution of pl + 9 is passed for 7 consecutive days, all conventional products lose hydration. Column deterioration occurs due to decomposition, and retention capacity also decreases.

In contrast, the product of the present invention has 3
It can be seen that it is extremely stable even after graded crosslinking. This is a very limited example, and XCl1. R″-CII,
All halomethyl compounds represented by X or XCIIJ' exhibit the same stability.

Thus, it has been found that an excellent filler that is stable against hydrolysis can be obtained by tertiating and crosslinking the conventionally used 1-2 treated aminoalkyl-bonded silica with a halomethyl compound. Next, a more detailed explanation will be given in Examples. However, it is not limited to this.

Example ! Three 50 g of spherical silica gels with an average particle size of 5 μm were placed in a Lof flask, and 250 ml of toluene and 50 g of aminopropyldimethylmethoxysilane were sequentially added to the flask.
The reaction was heated under reflux for 0 hours. After the reaction was completed, it was filtered and washed in a conventional manner and dried at 60° C. under reduced pressure overnight. Put three 5g of this aminopropyl-bonded silica into a Lof flask and add 20g of xylene.
m1. Predetermined amounts of catalyst and 1,3-dibromopropane were added one after another, and the mixture was heated under reflux for 5 hours under a nitrogen stream to react. After the reaction is complete, filter, xylene, ethanol, acetone,
After washing sequentially with water and finally with acetone, 80℃ under reduced pressure.
It was dried overnight. Approximately 100 m of the obtained tertiary crosslinked silica
g was placed in a test tube, 0.5 ml of ninhydrin test solution was added, and the mixture was heated in a hot bath to examine the degree of tertiaryization of amino groups.

Table 2 shows the results with respect to the amount of 1,3-dibromopropane charged. In the table, the symbol ○ indicates that tertiaryization is complete and is negative to the ninhydrin reagent, △ indicates that it is weakly colored and contains a small amount of unreacted primary amine, and the symbol X indicates that it is deeply colored and contains a considerable amount of primary amine. The remaining items are shown below.

Table 2 Degree of tertiary crosslinking of aminopropyl-bonded silica 1.3-dibromo Catalyst Propane charge (triethylamine) 0 (g) O (g) X
l 0.5 X2
1.0 △3
1.5 05 2.5
010 5.0
050 25.0 0 again,
Figures 1 to 3 show the hydrolytic stability of aminopropyl-bonded silica and tertiary crosslinked silica with 5 g of 1,3-dibromopropane.

Example 2 Three 50 g of spherical silica gels with an average particle size of 5 μm were placed in a flask, and 250 ml of toluene was added. 55 g of N4-aminoethyl-3-aminopropyltrimethoxysilane was successively added, and the mixture was heated under reflux for 10 hours to react. After the reaction was completed, it was filtered and washed in a conventional manner and dried at 60° C. under reduced pressure overnight. N-2-aminoethyl-3 thus prepared
Three 5 g pieces of -aminopropyl-bonded silica were placed in a flask, 20 ml of xylene and 5 g of 1.2-dibromoethane were added to each flask, and the mixture was heated under reflux for 5 hours under a nitrogen stream to react. After the reaction was completed, it was washed and dried in the same manner as in Example 1.
A graded crosslinked product was prepared. Under the same conditions, 1.3
- A tertiary crosslinking reaction was carried out with dibromopropane, chloromethyloxirane, and a 1:1 mixture thereof.

The hydrolytic stability of the silica thus prepared is shown in Table 1.
Of these, 2, 6, and 7.8, and the tertiary crosslinked ones are stable.

The above Example 1 corresponds to the general formula, and Example 2 corresponds to Ni1s -N-C
1lt-R″-CIlt-N- or N11°CIlt CI[.

■ 1l CIlt + X-C11t-R' + X-CI
lt-11'-CIlt-X -C11゜I CIL C11t CIl, CIl.

Cl1x C11t corresponds to each.

[Effect] As a result of tertiaryizing conventional aminoalkyl-bonded silica with a halomethyl compound and at the same time cross-linking the amino groups, it is possible to eliminate carbonyl compounds and other reactive impurities in the sample and during hydrolysis without impairing the original separation properties. It has the great effect of extending column life, which is the most important aspect of liquid chromatography.

[Brief explanation of the drawing]

Figure 1 shows a comparison of the effects of the present invention with conventional products. Figure 2 shows a comparison of the separation ability of the product of the present invention before and after hydrolysis. Figure 3 compares the separation performance of the conventional product before and after hydrolysis. % Kyoda Hito Tokyo Chemical Industry Co., Ltd. 0 :'to 60 90 12
0 150 180 210 Hydrolysis time (minutes) Figure 1 Comparison of hydrolysis stability Retention time (minutes) A. Retention time before hydrolysis (minutes) B. After hydrolysis treatment Figure 2 Separation ability of the product of the present invention

Claims (1)

[Claims] ≡SiRNH_2 or ≡SiRNH(CH_2CH_2
NH)_0_~_1_0-CH_2CH_2NH_2, XCH_2R
A novel method for hydrolytically stabilizing aminoalkyl-bonded silica, which is characterized by reacting with a halomethyl compound represented by ' or Where, R″ means a phenyl group, etc. Also, R″ is a hydrocarbon residue of C_nH_2_n, a carbonyl group of CO, an S atom, ▲ formula,
There are chemical formulas, tables, etc. ▼ Phenylene group, ▲ Numerical formulas, chemical formulas, tables, etc. ▼ means alcohol residues, etc. n is an integer from 1 to 10.