Process For Manufacturing a Composite Sorbent Material For Chromatographical Separation of Biopolymers

Abstract

The present invention relates to a sorbent material for separation and purification of biopolymers, particularly nucleic acids, having a solid support substantially modified with a copolymer coating comprising aromatic monomers and crosslinking compounds and unsaturated esters or ethers preferably attached to the support via a vinylchlorsilane. The use of these materials for separation of nucleic acids, particularly a one-step isolation of DNA from lysates of different biological sources, is an object of the invention as well as a chromatographic column or cartridge at least partially filled with the sorbent material of the invention, a membrane-like device comprising the sorbent material of the invention, and a kit comprising the sorbent material of the invention in bulk or packed in chromatographic devices as well as other devices necessary for performing sample preparations.

1 . A polymer obtainable by a process of polymerizing at least three components [A],[B], and [M] wherein
 A is at least one aromatic or aliphatic compound having at least one polymerizable unsaturated moiety,   B is at least one cross-linkable aromatic or aliphatic compound, and   M is an organic non-saturated polymerizable compound different from [A] having hetero atoms in the C—C chain or in side chains wherein the process comprises the steps of   admixing the components [A], [B], and [M] sequentially or non-sequentially,   polymerizing the resulting mixture composition   removing unreacted material and   recovering and drying the composite material.   
     
     
         2 . The polymer of  claim 1  having the following structure
   {[A] x —[B] y —[M] z } p      wherein and x, y, and z are independent of each other and an integer of 1-100 and p is a number between 2 and 5000.   
     
     
         3 . A monomer having the structure [A] x —[B] y —[M] z  wherein
 A is at least one aromatic or aliphatic compound having at least one polymerizable unsaturated moiety,   B is at least one cross-linkable aromatic or aliphatic compound, and   M is an organic non-saturated polymerizable compound different from [A] having hetero atoms in the C—C chain or in side chains   and x, y, and z are independent of each other an integer of 1-100.   
     
     
         4 . A chromatographic material comprising essentially a polymer of  claim 1 . 
     
     
         5 . A composite sorbent material having a support which is at least partially covered by a polymer coating according to  claim 1 . 
     
     
         6 . A composite sorbent material having a support which is at least partially covered by the polymer coating according to  claim 1  having the general formula 
       
         
           
           
               
               
           
         
       
       obtainable by a process comprising the steps of
 modifying the support with a vinylchlorsilane 
 followed by addition of 
 a) at least one unsubstituted or substituted styrene or vinylnaphthalene [A] of general formula 
 
       
         
           
           
               
               
           
         
         b) at least one cross-linkable unsubstituted or substituted (1,4 or 1,2)-divinylbenzene [B] of general formula 
       
       
         
           
           
               
               
           
         
         c) optionally a non-saturated carboxylic acid or alkyl, aryl or hydroxylalkyl ester thereof (M) of general formula 
       
       
         
           
           
               
               
           
         
          respectively a non-saturated alcohol or ether of general formula 
       
       
         
           
           
               
               
           
         
         polymerizing the copolymer composition 
         removing unreacted material and 
         recovering and drying the composite material. 
       
     
     
         7 . The sorbent material according to  claim 5  wherein the support is a porous inorganic material selected from the group comprising inorganic metal oxides, such as oxides of aluminum, titanium, zirconium, silicon oxides, iron oxides, controlled pore glass (CPG), diatomaceous earth and combinations thereof. 
     
     
         8 . The sorbent material according to  claim 7 , wherein the porous inorganic metal oxides show a bidisperse distribution of the pore sizes, with mean pore diameters in the range of 2-15 nm for the smaller pore size and 20-100 nm for the larger pore size. 
     
     
         9 . The sorbent material of  claim 5 , wherein the support has an average pore size of 100-2000 Å, and a specific surface of 20-300m 2 /g. 
     
     
         10 . The sorbent material of  claim 5 , wherein the polymer coating has a thickness of about 10 to 250 Angström and micropores of less than 50 Å accessable to water, salts, and low molecular weight substances, the polymer coating being non-adsorptive towards nucleic acids and adsorptive towards proteins. 
     
     
         11 . The sorbent material of  claim 5 , wherein the mass ratio of copolymer components [A], [B], and [M] complies with the formula [A]:[B]:[M]=1:0.03-0.30:0.01-0.2. 
     
     
         12 . A chromatographic column, capillary or cartridge at least partially filled with the composite sorbent material according to  claim 5 . 
     
     
         13 . A membrane-like article comprising the composite sorbent according to  claim 5 , wherein the composite sorbent is embedded in a porous organic or inorganic matrix. 
     
     
         14 . A kit comprising a composite sorbent according to  claim 5 , the composite sorbent being in bulk or packed in chromatographic columns or cartridges or a membrane-like article. 
     
     
         15 . The sorbent material of  claim 5 , for performing fast sample preparations or chromatographic separations of nucleic acids. 
     
     
         16 . The sorbent material of  claim 5 , for separation of nucleic acids and/or proteins, wherein nucleic acids are passing said material without retention while proteins, salts and other low molecular weight substances are retained. 
     
     
         17 . The sorbent material of  claim 16 , for separation of DNA in one step. 
     
     
         18 . A process for manufacturing a composite sorbent material for chromatographic separation of biopolymers according to general formula 
       
         
           
           
               
               
           
         
       
       comprising the steps of:
 modifying the support with a dialkylchlorsilane having an ethylenically unsaturated moiety such as vinylchlorsilane 
 followed by addition of 
 a) at least one unsubstituted or substituted styrene or vinylnaphthalene [A] of general formula 
 
       
         
           
           
               
               
           
         
         b) at least one cross-linkable unsubstituted or substituted (1,4 or 1,2)-divinyl benzene [B] of general formula 
       
       
         
           
           
               
               
           
         
         c) optionally a non-saturated carboxylic acid or alkyl, aryl or hydroxylalkyl ester thereof (M) of general formula 
       
       
         
           
           
               
               
           
         
          respectively a non-saturated alcohol or ether of general formula 
       
       
         
           
           
               
               
           
         
         polymerizing the copolymer composition 
         removing unreacted material and 
         recovering and drying the composite material. 
       
     
     
         19 . The process according to  claim 18 , wherein the mass ratio of copolymer components [A], [B], and [M] complies with
   A:[B]:[M]=1:0.03-0.30:0.01-0.2.   
     
     
         20 . The process according to  claim 18 , characterized in that the step of polymerizing the copolymer composition is conducted by polymerization in organic solvents or by emulsion polymerization in water with an emulgator and a polymerization initiator. 
     
     
         21 . The process according to  claim 20 , wherein the mass ratio of water or organic solvent copolymer is 5/1-20/1. 
     
     
         22 . A method for purifying and/or isolating nucleic acids using the sorbent material of  claim 5 . 
     
     
         23 . The sorbent material of  claim 9 , wherein the support has an average pore size of 300-1000 Å. 
     
     
         24 . The sorbent material of  claim 9 , wherein the support has a specific surface of 20-100 m 2 /g. 
     
     
         25 . The sorbent material of  claim 10 , wherein the polymer coating has a thickness of about 10 to 100 Å. 
     
     
         26 . The kit of  claim 14 , further comprising filter materials, reagents and/or buffers or other devices or chemicals for performing sample preparations or chromatographic separations of biopolymers.