DD228461B1 - METHOD FOR THE PRODUCTION OF ANION EXCHANGERS - Google Patents

Abstract

Process for the preparation of chemically elutable, mechanically stable anion exchangers. From WP 245 785 known shaped or non-formed macromolecular masses with chemically active packing such. B. cyanuric chloride according to the invention with compounds of the general formula

Description

X - fl - _N _

in the X NH ₂ -, OH or SH groups

R alkyl groups having 1-10 C atoms, aromatics, photolytic groups such as

substituted o-nitrobenzyl or substituted methyl-phenancylester, heterocycler R ¹ , R ² , R ³ alkyl groups with 1-4 C-atoms or H-atoms Y halogen, hydroxyl groups

are, implemented in a conventional manner.

2. The method according to item 1, characterized in that compounds of the general formula I as an intermediate by reaction of compounds of general formula II

3 NH ₃ -RY II

with tertiary amines of the general formula III

-R ¹

NR ² III

-R ³

in which R, R ¹ , R ² , R ³ , Y have the abovementioned meaning, are formed.

Field of application of the invention

The invention relates to a process for the preparation of chemically elutable, mechanically stable anion exchangers based on macromolecular compositions. The anion exchangers are used for analytical and preparative tasks in molecular biology, biochemistry and genetic engineering.

Characteristic of the known state of the art

Surface carriers with anion exchange properties are known. At present there are two surface carriers with anion exchange properties based on cellulose paper and a plastic membrane on the market. The first case is a DEAE paper (manufacturer Whatman [England]), which is obtained by first derivatizing cellulose according to US Pat. No. 3,823,133 with diethylaminoethyl chloride and then using a suitable dispersing agent to form a sheet Cellulose paper is further processed. Although this anion exchange surface carrier has a high max. Binding capacity of about 300-400 μg / cm ² for nucleic acids, its too low mechanical stability characteristics such as abrasion, tearing, runoff of whole layers, the reason why it can hardly be used for biochemical investigations. The second case is a DEAE plastic membrane (NA-45, manufactured by Schleicher & Schüll [USA]), which has high mechanical stability but whose binding capacity is too low at 10-20 pg / cm.

Both carriers share the substituent, the diethylaminoethyl group. However, carrier materials having this substituent as the ionic bonding group are nucleic acids (oligonucleotides, DNA) chemically, e.g. As by piperidine, not elutable.

The aim of the invention is to produce inexpensively mechanically stable and chemically elutable anion exchangers which can be used in particular in molecular biology and biochemistry.

Explanation of the essence of the invention

The invention has for its object to produce novel anion exchangers on a macromolecular basis, which have a high binding capacity and which are chemically elutable.

It is known from WP 245785 known shaped or non-formed macromolecular masses with chemically active packing,

From a macromolecular compound in a concentration of 5-80VoI .-%

From a macromolecular compound with 4,6-dihalo-1,3,5-triazine groupings in a concentration of 1-80 vol.%

From 2,4,6-trihalo-1,3,5-triazine in a concentration of 0.5-50% by volume

From halides in a concentration of up to 20% by volume

- Where appropriate, from buffering substances up to 5 vol .-%, according to the invention with compounds of general formula I.

- Λ-

RNR -K ³

implemented in a conventional manner, wherein

X is NH ₂ , OH or SH groups

R alkyl groups having 1-10 C atoms, aromatics, photolytic groups such as substituted o-Nitrobenzylester or

Methylphenancylester as well as heterocycles R ¹ , R ² , R ³ alkyl groups with 1-4 C-atoms or H-atoms Y halides or hydroxyl groups.

are. Instead of directly reacting the shaped and non-formed macromolecular masses with chemically active packing with compounds of general formula I can advantageously compounds of formula I intermediary by reaction of compounds of general formula II

YR-NH ₃ ⁺ Y "II

with tertiary amines of the general formula III

-R ¹ NR ² III

be generated.

In the first case, shaped macromolecular masses can be reacted with chemically active packing materials such as mechanically stable cellulose papers, nonwovens and woven fabrics, thus obtaining mechanically stable anion exchange surface supports with high binding capacities for nucleic acids of up to 120 pg / cm ² .

In the second case, non-formed macromolecular masses with chemically active packing such. B. spherical, crystalline and fibrous cellulose reacted and obtained in this way anion exchangers, which can be used either in this form or processed with liquid dispersants to form shaped anion exchange surface carriers.

If compounds of the general formula I or compounds of the general formula II are used in conjunction with tertiary amines for reacting the shaped and non-formed macromolecular masses with chemically active fillers, where R = alkyl groups having 1-10 C atoms or aromatics, this results Anion exchangers which are chemically elutable with primary, secondary and tertiary amines. This chemical elution is of particular importance for the development of new solid-phase sequencing methods of nucleic acid fragments or for the isolation of DNA fragments from agarose or polyacrylamide gels. If compounds of the general formula I are used for reacting the shaped and non-formed macromolecular masses with chemically active packing, where R = substituted o-nitrobenzyl or substituted methylphenancyl esters, anion exchangers are obtained which are photolytically, i. H. are elutable using electromagnetic radiation.

All the above-mentioned ion exchangers are additionally with ionic agents, ie with high ionic strengths of the salts such as NH ₄ HCO ₃ , (NH ₄ I ₂ CO ₃ , Et ₃ NHCO ₃ , Et ₃ NHAc, NH ₄ Ac, K ₂ HPO ₄ , NaCl, among others eluted

 The term "chemically elutable" thus includes in the context of the present invention:

1. elution by chemical compounds, their mixtures or solutions in solvents including water, optionally with heating,

2. elution with ions in solutions of solvents including water,

3. Elution by cleavage of bonds by means of radiation including light

4. Elution by thermal treatment in bulk or in the presence of solvents including water.

The anion exchangers according to the invention have a high binding capacity and, in particular in the surface form, have excellent mechanical stability. For this reason, the use of shaped anion exchangers is particularly indicated in molecular biological and biochemical research. Compared with conventional anion exchangers, the exchangers according to the invention have, in addition to the ionic elution, further elution possibilities with chemical agents or electromagnetic radiation.

embodiments example 1

Cellulose paper (Whatman 540) is activated according to WP 245785 with cyanuric chloride. The surface-treated in this way paper (5 χ 7cm) is (are stored welded foil _r under N fumigation at -20 ⁰ C) from its special packaging removed and placed in a glass reaction chamber in which 2 mmol (450mg) of triethyl-1 -amino-ethyl-ammonium bromide in 7ml acetonitrile were submitted. The reaction chamber is shaken for about 12 hours at room temperature. After completion of the reaction, the paper is washed several times with water and dried between blotting paper. Instead of the solvent acetonitrile and aqueous buffer systems can be used.

Example 2

400mg crystalline, underivatized cellulose is activated according to WP 245785 with cyanuric chloride and then reacted in a 25 ml round bottom flask with 2 mmol (450 mg) triethyl-1-amino-ethyl-ammonium bromide in 7 ml acetonitrile for 12 h at room temperature. After completion of the reaction, the cellulose is washed on a frit several times with water and dried as in Example 1.

Example 3

Cellulose paper (Whatman 540) is surface-activated as in Example 1 with cyanuric chloride and then placed in a glass reaction chamber, were presented in the 2mmol (410 mg) of 1-bromo-2-amino-ethane hydrobromide in 7 ml of acetonitrile. The reaction is then started with the addition of 5 ml of triethylamine. The reaction chamber is sealed with Parafilm® and shaken at room temperature for about 12 hours After completion of the reaction, the paper is washed as in Example 1 and dried.

Example 4

400mg of crystalline underivatized cellulose e.g. Whatman is activated as in Example 2 with cyanuric chloride and then mixed in a 25 ml round bottom flask with 2 mmol (410 mg) of i-bromo-2-amino-ethane hydrobromide in 7 ml of acetonitrile. The reaction is started by addition of 5 ml of triethylamine and after 12 h at room temperature by washing the cellulose and drying, as described in Example 2, terminated.

Example 5

Surface-activated Whatman 540 paper (5 × 7 cm) from Example 1 is reacted in a reaction chamber with 2 mmol (574 mg) of p-aminobenzyl-triethyl-ammonium bromide in 7 ml of acetonitrile for 12 h at room temperature and worked up further as in Example 1.

Example 6

400 mg of crystalline Whatman cellulose surface-activated according to Example 2 is reacted with 2 mmol (574 mg) of p-aminobenzyl-triethylammonium bromide in 7 ml of acetonitrile, as described in Example 2, and worked up.

Example 7

Surface-activated Whatman 540 paper (5χ7 cm) from example 1 is mixed with 2 mmol (540 mg) of p-aminobenzylbromide hydrobromide in 7 ml of acetonitrile in a reaction vessel and the reaction started with 5 ml of triethylamine. After 12 h at room temperature, the paper is washed as in Example 1 and dried.

Example 8

400mg of crystalline, according to Example 2 surface-activated Whatman cellulose is reacted analogously to Example 4mitmmol (540 mg) p-aminobenzyl bromide hydrobromide in acetonitrile and triethylamine, washed after completion of the reaction and dried.

Example 9

Surface-activated Whatman 540 paper (5 χ 7 cm) from Example 1 is reacted with 2 mmol of glycine (o-nitro-p-methyl-triethylammonium bromide) benzyl ester in 7 ml of acetonitrile for 12 h at room temperature in a reaction chamber and worked up as in Example 1 on.

Example 10

Semi-synthetic paper of the type Hekosyn® (polyamide and cellulose) is surface-activated as in Example 1 with cyanuric chloride and then placed in a glass reaction chamber, were presented in the 2mmol (410 mg) 1-bromo-2-amino-ethane hydrobromide in 7 ml of acetonitrile. The reaction is then started with the addition of 5 ml of triethylamine. The reaction chamber is sealed with parafilm and shaken for about 12 hours at room temperature. After completion of the reaction, the paper is washed as in Example 1 and dried.

Claims (2)

1. A process for the preparation of chemically elutable, mechanically stable anion exchangers based on macromolecular masses with chemically active packing consisting of A macromolecular compound in a concentration of 5-80% by volume A macromolecular compound with 4,6-dihalo-1,3,5-triazine groupings in a concentration of 1-80% by volume 2,4,6-trihalo-1,3,5-triazine in a concentration of 0.5-50% by volume - halides in a concentration up to 20VoI .-% Optionally from buffering substances up to 5% by volume, characterized in that the masses with compounds of general formula I.