WO2005009981A1 - Reactifs en phase solide pouvant se diviser et se lier de maniere multifonctionnelle, et procede pour leur production - Google Patents

Reactifs en phase solide pouvant se diviser et se lier de maniere multifonctionnelle, et procede pour leur production Download PDF

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Publication number
WO2005009981A1
WO2005009981A1 PCT/DE2004/001684 DE2004001684W WO2005009981A1 WO 2005009981 A1 WO2005009981 A1 WO 2005009981A1 DE 2004001684 W DE2004001684 W DE 2004001684W WO 2005009981 A1 WO2005009981 A1 WO 2005009981A1
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Prior art keywords
carrier material
general formulas
linker
polymeric surface
functionalized
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PCT/DE2004/001684
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German (de)
English (en)
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Thomas RÜHL
Klaus Burger
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Universität Leipzig
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/107—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by chemical modification of precursor peptides
    • C07K1/1072—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by chemical modification of precursor peptides by covalent attachment of residues or functional groups
    • C07K1/1077—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by chemical modification of precursor peptides by covalent attachment of residues or functional groups by covalent attachment of residues other than amino acids or peptide residues, e.g. sugars, polyols, fatty acids
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D263/00—Heterocyclic compounds containing 1,3-oxazole or hydrogenated 1,3-oxazole rings
    • C07D263/02—Heterocyclic compounds containing 1,3-oxazole or hydrogenated 1,3-oxazole rings not condensed with other rings
    • C07D263/08—Heterocyclic compounds containing 1,3-oxazole or hydrogenated 1,3-oxazole rings not condensed with other rings having one double bond between ring members or between a ring member and a non-ring member
    • C07D263/16—Heterocyclic compounds containing 1,3-oxazole or hydrogenated 1,3-oxazole rings not condensed with other rings having one double bond between ring members or between a ring member and a non-ring member with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D263/18—Oxygen atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D317/00—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms
    • C07D317/08—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3
    • C07D317/10—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 not condensed with other rings
    • C07D317/32—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 not condensed with other rings with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D317/34—Oxygen atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D327/00—Heterocyclic compounds containing rings having oxygen and sulfur atoms as the only ring hetero atoms
    • C07D327/02—Heterocyclic compounds containing rings having oxygen and sulfur atoms as the only ring hetero atoms one oxygen atom and one sulfur atom
    • C07D327/04—Five-membered rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/04—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers
    • C07K1/042—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers characterised by the nature of the carrier
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/04—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers
    • C07K1/047—Simultaneous synthesis of different peptide species; Peptide libraries
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/13—Labelling of peptides
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B80/00—Linkers or spacers specially adapted for combinatorial chemistry or libraries, e.g. traceless linkers or safety-catch linkers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00277—Apparatus
    • B01J2219/00452—Means for the recovery of reactants or products
    • B01J2219/00454—Means for the recovery of reactants or products by chemical cleavage from the solid support
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00583—Features relative to the processes being carried out
    • B01J2219/00603—Making arrays on substantially continuous surfaces
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00583—Features relative to the processes being carried out
    • B01J2219/00603—Making arrays on substantially continuous surfaces
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • B01J2219/0061—The surface being organic
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00583—Features relative to the processes being carried out
    • B01J2219/00603—Making arrays on substantially continuous surfaces
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • B01J2219/00612—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports the surface being inorganic
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00583—Features relative to the processes being carried out
    • B01J2219/00603—Making arrays on substantially continuous surfaces
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • B01J2219/00623—Immobilisation or binding
    • B01J2219/00626—Covalent
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00583—Features relative to the processes being carried out
    • B01J2219/00603—Making arrays on substantially continuous surfaces
    • B01J2219/00639—Making arrays on substantially continuous surfaces the compounds being trapped in or bound to a porous medium
    • B01J2219/00641—Making arrays on substantially continuous surfaces the compounds being trapped in or bound to a porous medium the porous medium being continuous, e.g. porous oxide substrates
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B50/00—Methods of creating libraries, e.g. combinatorial synthesis
    • C40B50/14—Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support

Definitions

  • the invention relates to surface-functionalized carrier materials, each having a polymer surface and at least one linker compound covalently bonded to it.
  • Such materials can be used for the covalent immobilization of biomolecules, in particular amino acids, peptides or proteins or molecules with amino or carboxy groups, for creating substance libraries in combinatorial chemistry, for the synthesis of amino acids, peptides, proteins or molecules with at least one peptide structural unit
  • Solid phases can be used in peptide chemistry and for the production of affinity labeling derivatives.
  • the invention further describes a method for producing the surface-functionalized carrier materials.
  • the further chain structure includes a condensation of the next amino acid to the amino group (the TV terminus) of the already immobilized amino acid or - in the case of further synthesis - the immobilized peptide.
  • the synthesis from the C to the IV Therminus of the peptide takes place.
  • the Boc strategy derived from the Merrifield concept (R. Arshady et al., J. Chem. Soc. Pekin Trans. I 1981, 529-537) and the Fmoc strategy (LA Carpino, GY Hau, J Org. Chem. 1972, 37, 3404-3409), in which the amino group of the amino acids to be linked is protected by certain protective groups, the peptide is ultimately bound to the solid phase via the carboxy function of the first amino acid.
  • chloroformic acid ester units immobilized on the surface of polystyrene are used as the starting point for peptide synthesis (RL Letsinger, MJ Komet, J. Am. Chem. Soc. 1963, 85, 2149-2144; R Matsueda et al., J. Am. Chem. Soc. 1975, 97, 2573-2575).
  • the synthesis is carried out here by intravenous linkage of the amino acids protected as tert-butyl ester in the order given by the target sequence in the direction of the C-terminus of the peptide.
  • a disadvantage of this prior art is that, depending on the desired synthesis strategy, differently functionalized support materials have to be used, since the chemical properties of the respective surface functions of the support material only permit an IV-terminal or a C-terminal linkage of the amino acid.
  • Surface-functionalized carrier materials are known which allow the user to selectively bind a C-terninal or an IV-T ⁇ ninal binding of an amino acid or another molecule with corresponding functional groups (Sh. N. Khattab; A. El-Faha; AM El-Massry; EME Mansour; MM Abd. El-Rahman Letters in Peptide Science 2001, 7, 331-345; M. Lebl et al., US 5635598, Jun., 1997; P. Wessig et al., WO 02/051917 AI, July 2002).
  • the production of the known multifunctional, linkable and cleavable solid phase reagents has the disadvantage that additional steps for the protection and / or activation of the surface-functionalized carrier material have to be applied
  • the surface functionalized carrier materials which allow the user either a C-terminal or a H-terminal linkage of an amino acid or another molecule with corresponding functional groups, have the disadvantage that their production activates the e ⁇ ien and protects the other C-
  • the term and the protection of the IV terminus are required and additional steps have to be taken to split off the respective protecting group at the C and IV terminus and to activate the C terminus. Quantitative activation of the C-terminus can be difficult.
  • the coupling step of the linker to the solid phase is designed to be side-selective, so that the carboxy group available for subsequent derivatization is protected is.
  • the IV terminal is protected in order not to react undesirably with the activated C-terminus during the coupling step to the solid phase.
  • the object of the invention is to provide easy-to-handle solid phase supports with at least one linker compound covalently bonded to them.
  • the carrier should covalently immobilize molecules with amino or carboxy groups, especially amino acids, peptides or proteins, and thus z. B. simplify the production of substance libraries in combinatorial chemistry.
  • a method for producing the surface-functionalized carrier material is also to be provided.
  • the object is achieved by a surface-functionalized carrier material having a polymeric surface and at least one linker compound covalently bonded to it according to the general formulas (1) to (16) or (1 ') to (16'),
  • n 1 to 12
  • R 1 and R 2 independently of one another are H or an alkyl group
  • L is a Spacer means that links the linker compound to the polymeric surface.
  • the surface-functionalized carrier material according to the general formulas (1) to (16) according to the invention therefore consists of: 1. a polymeric surface (P) or a polymeric surface with a spacer (L) which links the linker compound to the polymeric surface 2. and one Hexafluoroacetone (F 3 CC-CF 3 ) protected linker compound.
  • the linker compound is protected by hexafluorothioacetone (F 3 CC-CF 3 ).
  • linker compounds consist of hexafluoroacetone or hexafluorothioacetone-protected and carboxyl-activated ⁇ -amino-dicarboxylic acids of the general formula (43),
  • n has the meaning 1 to 12 and R 1 and R 2 independently of one another denote H or an alkyl group, ⁇ -hydroxy-dicarboxylic acids of the general formula (44),
  • n 1 to 12 and Ri is H or an alkyl group
  • n 1 to 12 and Ri is H or an alkyl group, iV-substituted Glycm derivatives of the general formula (46),
  • n has the meaning 1 to 12.
  • the carboxy group is activated towards nucleophiles and is available in a reactive form.
  • hexafluoroacetone or hexafluorothioacetone as a protective group advantageously makes it possible to combine protection and activation on the one hand and derivatization and deblocking on the other hand in one step, thereby saving synthesis steps.
  • the surface-functionalized support material according to the general formulas (1) to (16) or (1 ') to (16') can advantageously be water or molecules with hydroxyl, thiol or amino group at the C-terminal or molecules with carboxy-, sulfonyl - or either phosphoryl groups. O-, S-, or iV-terminal can be bound covalently.
  • the binding of molecules with carboxy, sulfonyl or phosphoryl groups in activated form or by a coupling reagent to the surface-functionalized support material according to the general formulas (1) to (16) or (! ') to (16 5 ) is made possible by splitting off the hexafluoroacetone or hexafluorothioacetone group by reaction with the hydroxyl, thiol or amino group.
  • the surface-functionalized support material according to the general formulas (1) to (16) or (1 ') to (16') can thus advantageously be used for the covalent immobilization of photolabile victim groups, for the covalent immobilization of biomolecules, in particular amino acids, peptides or proteins or other molecules with amino or hydroxy, thiol (C-terminal immobilization) and / or carboxy groups (O-, S- or iV-terminal immobilization) can be used.
  • the surface-functionalized carrier material according to the general formulas (1) to (16) or (1 ') to (16') is therefore advantageously suitable for creating substance libraries in combinatorial chemistry, for the synthesis of amino acids, peptides, proteins or Molecules with at least one peptide structural unit on solid phases in peptide chemistry and for obtaining affinity labeling derivatives (J. Jenssen, K. Sewald, N. Sewald, Bioconjugate. Chem. 2004, 15, 594-600).
  • the surface-functionalized carrier material according to the general formulas (1) to (16) or (1 ') to (16') is advantageously suitable for use in photoaffinity labeling and its subsequent steps.
  • the carrier material according to the invention serves as a carrier for a photosensitive group and a molecule with affine properties to form a partner molecule.
  • the partner molecule is enriched, covalently bound by a photochemical reaction and broken down chemically or enzymatically.
  • the polymer surface (P) is then removed via a predetermined breaking point by a cleavage reaction.
  • the cleaning processes involved are simple here and the time required is considerably reduced compared to the methods known in solution.
  • the carboxy function of the surface-functionalized carrier material according to the general formulas (1) to (16) or (1 ') to (16') integrated in the lactone ring is capable of amide, thioester or ester formation.
  • the amino, thiol or hydroxy function deprotected in the course of the amide, thioester or ester formation is thus immediately available for derivatization.
  • Complex cleaning steps such as flash Column chromatography (FSC), which are essential for synthesis in solution, is gone.
  • the quantitative removal of the hexafluoroacetone or hexafluorodioacetone hydrate is possible by simply washing with large amounts of water (advantageously dilute citric acid).
  • the amine used and the acylation reagent used in the subsequent step are removed by simple washing with appropriate solvents, so the two reagents can be used in excess in the two reaction steps (3-5 equivalents), which considerably increases the yield.
  • the surface-functionalized support material (33) according to the invention (Scheme 1) with high occupancy of linker compounds protected from hexafluoroacetone or hexafluorothioacetone can advantageously be used for a three-stage synthesis of (50) (Scheme 2).
  • ring opening e.g. by aminolysis
  • electrophiles e.g. acylation, sulfonation or phosphoryation
  • an oxidation e.g. formation of disulfides
  • the ring opening is preferably carried out with amines, even under aqueous conditions.
  • the spacer L is preferably an amino acid, a peptide or another compound which contains at least one amino or hydroxy function for the attachment of the linker compound and at least one carboxy function for the immobilization to the solid phase.
  • the carrier material preferably consists entirely or on its surface of an organic polymer.
  • the polymeric surface (P) preferably contains hydroxyl groups (for example Wang resins) which enable the linker compound or the spacer L to be bound.
  • the esters, amide or urethane bonds formed by the binding of the linker compound or the spacer L can advantageously be cleaved by simple methods known per se, after derivatization has taken place.
  • the organic polymer is preferably selected from polypropylene, polyethylene, polysulfone, polystyrene, polyvinyl chloride, polyacrylonitrile, cellulose, amylose, agarose, polyamide, polyimide, polytetrafluoroethylene, polyvinylidene difluoride, polyester, polycarbonate, polyacrylate, polyacrylamide or derivatives, copolymers or blends of these substances.
  • the organic polymer is particularly preferably a Wang resin, a PEGA resin (2-acrylamidoprop-l-yl- (2-aminoprop-l-yl) polyethylene glycolsoo and dimethylaciylamide cross-linked with bis 2-acrylamidoprop-l-yl polyethylene glycolsoo) or a Rink Amide resin.
  • the carrier material is an inorganic and / or mineral material, preferably a glass, a silicate, a ceramic material or a metal or a composite of at least one inorganic and / or mineral material and at least one organic polymer.
  • the carrier material is preferably in the form of a membrane, a film, a plate, a microtiter plate, a reaction vessel, a slide, a fiber, a hollow fiber, a nonwoven, a tissue, a powder, a granulate or particles and is porous or non-porous , It is preferably a powder.
  • a membrane In the form of a membrane, it has a symmetrical or asymmetrical pore structure. It preferably has pores with a diameter of 1 nm to 10 ⁇ m.
  • the invention further relates to a process for producing the surface-functionalized carrier material according to the invention with a polymeric surface and at least one linker compound covalently bonded to it according to the general formulas (1) to (16) or (1 ') to (16').
  • n, R 1 and R 2 have the above meaning and L is a spacer which is intended to link the linker compounds to the polymer surface, or the linker compounds according to the general formulas (17 ') to (32')
  • hexafluoroacetone has long been known as a protective group for ⁇ -amino, ⁇ -hydroxy, ⁇ -mercapto-dicarboxylic acids and iV-substituted glycine derivatives
  • the lactone ring is activated by the two trifluoromethyl substituents. So far, the ring opening in solution systems with amines, hydroxy compounds or water. When the ring opening of hexafluoroacetone-protected ⁇ -amino-dicarboxylic acids with excess strong nucleophiles at room temperature in solution, an elimination rank reaction disadvantageously dominates. Due to the often stable half-aminal or half-thioacetal intermediate stage, a large amount of water is required for quantitative elimination, which makes additional washing and extraction steps necessary, which are expensive to carry out in solution systems, before working up by flash column chromatography.
  • the carrier material according to the general formulas (1) to (16) and (1 ') to (16') according to the invention enables rapid workup after simple removal of the hexafluoroacetone or hexafluorothioacetone by simply washing the carrier material.
  • hexafluoroacetone protective grapple technology to solid-phase systems has so far failed because hexafluoroacetone is a very toxic and aggressive gas, the hydrates of which are also regularly used as solvents for H-containing polymers.
  • the polymer is in the hexafluoroacetone and hexafluoroacetone hydrate present in the solvent.
  • the dissolved polymer makes it impossible to isolate the product by filtration.
  • the surface-functionalized support material according to the general formulas (1) to (16) or (1 ') to (16') is therefore, in order to avoid dissolving the polymeric surface, not by direct reaction of the linker compound bound to the support material with hexafluoroacetone or hexafluorothioacetone.
  • linker compounds are reacted in excess with linkable functional groups of solid-phase reagents (e.g. Wang resin).
  • the reaction is preferably carried out with catalysis (e.g. by adding DMAP - dimethylaminopyridine) or without catalysis.
  • linker compounds (occupancy reagent) (e.g. 1.5 equivalents), the number of runs, the temperature range (e.g. 0 ° C), the solvent (e.g. dry pyridine, chloroform) and the base (e.g. pyridine, NaHCO 3 ) or propene oxide are correct according to the substances used.
  • the substances obtained are cleaned by methods known per se.
  • the surface-functionalized carrier material obtained according to the general formulas (1) to (16) or (1 ') to (16') is preferably characterized by IR spectroscopy and the occupancy (amount of substance of the linker compound bound to the polymer divided by the mass of substance ) determined by elemental analysis.
  • the surface-functionalized carrier material obtained according to the general formulas (1) to (16) or (1 ') to (16') is characterized in the IR spectrum by the signals of the carbonyl groups of the linker compound which is linked to the polymeric surface ,
  • the occupancy of the substances obtained can be determined by elemental analysis (e.g. fluorine, nitrogen and / or sulfur elemental analysis).
  • the linker compounds according to the general formulas (17) to (20) are prepared by directly reacting the corresponding compounds according to the general formulas (43) to (46) with hexafluoroacetone in solution.
  • linker compounds of the general formulas (17 ') to (20') are correspondingly prepared by direct reaction of the corresponding compounds of the general formulas (43) to (46) with hexafluorothioacetone in solution.
  • the linker compounds according to general formulas (21) to (24) are prepared from the compounds according to general formulas (17) to (20) by known methods by converting the carboxy group into an isocyanate group in solution.
  • the linker compounds according to the general formulas (21 ') to (24') are correspondingly prepared from the compounds according to the general formulas (17 ') to (20').
  • the linker compounds according to general formulas (25) to (28) are prepared from the compounds according to general formulas (17) to (20) by coupling the free carboxy group with a linker in solution to form an ester or amide bond.
  • the linker compounds of the general formulas (25 ') to (28') are correspondingly prepared from the compounds of the general formulas (17 ') to (20').
  • the linker compounds according to general formulas (29) to (32) are prepared from the compounds according to general formulas (21) to (24) by reaction of the isocyanate function with a linker in solution to form a urea or urethane bond.
  • the linker compounds of the general formulas (29 ') to (32') are correspondingly prepared from the compounds of the general formulas (21 ') to (24').
  • the ⁇ -amino, ⁇ -hydroxy or ⁇ -thiol group and selectively the ⁇ -carboxy function are protected in one step.
  • the advantages of the invention can be seen in particular in the following: Since the protection of the amino, hydroxy or thiol function and the activation of the carboxy function in the If new processes are combined into one synthesis step, the application of the new process means that there is no need for synthesis steps. Since the Linke ⁇ * compounds are bound to the resin as an activated species, it is guaranteed that every molecule anchored with the resin is also activated. This cannot be guaranteed in the case of subsequent activation that only takes place on the resin. Since a washing process takes place after each reaction step, the dissolved reagents can be used in large excess. This leads to optimal yields. The reaction sequence can be automated.
  • the carboxy group of the compounds according to the general formulas (17) to (20) or (17 ') to (20') and (25) to (32) or (25 ') to (32') is activated by processes known per se, preferably by reaction with thionyl chloride or phosphorus pentachloride in an acid chloride, by reaction with DAST (diethylaminosulfur trifluoride) in an acid fluoride or by a coupling reagent, for example TBTU (O- (1 H-bezotriazol-1-yl) -H, N, iV, N'-tetramethyluronium tetrafluoroborate).
  • TBTU O- (1 H-bezotriazol-1-yl
  • the compounds according to the general formulas (25) to (32) or (25 ') to (32') are preferably selected from the compounds according to the general formulas (17) to (24) or (17 ') to (24' ) obtained by attaching the spacer L.
  • the spacer L is linked to the linker compound according to the general formulas (17) to (20) or (17 ') to (20') by acylation to give the ester or amide and the linkage of the spacer L to the linker compound according to general formulas (21) to (24) or (21 ') to (24') by addition to urethane or urea.
  • the spacer L is preferably an amino acid, a peptide or another compound with at least one amino or hydroxy function for the functionalization of the compounds according to the general formulas (17) to (24) or (17 ') to (24 ') and a carboxy function for immobilization on the polymer surface.
  • the linking of the linker compounds of the general formulas (17) to (20) or (17 ") to (20 ') and the general formulas (25) to (32) or (25') to (32 ') with the polymer Surface is preferably carried out by acylation to give the ester or amide and that of the linker compounds of the general formulas (21) to (24) or the general formulas (21 ') to (24') is preferably carried out by addition to the urethane or urea.
  • a trapping reagent for hydrogen chloride preferably NaHCO 3 or pyridine or propene oxide, is preferably used.
  • linker compounds according to the general formulas (17) to (32) or (17 ') to (32') are either used once.
  • the process is preferably carried out several times, preferably twice to three times, in order to achieve a quantitative conversion.
  • the surface-functionalized carrier material according to the invention is preferably produced in a temperature range between -100 ° C. and + 100 ° C., preferably -50 ° C. to 50 ° C., particularly preferably at room temperature.
  • the reaction is carried out in a solvent or solvent mixture.
  • Chloroform and / or dichloromethane is preferably used as the solvent for base-sensitive substances and pyridine is preferably used for base-insensitive substances.
  • washing liquid Water, ethyl acetate or another organic solvent or a solvent mixture is preferably used as the washing liquid.
  • the invention also relates to the use of the carrier material according to the invention for the synthesis of amino acids, peptides, proteins or molecules with at least one peptide structure unit, a first amino acid to be used for the synthesis being covalently bound to the carrier material according to the invention and chain extension by successively attaching further amino acids and / or a chemical modification takes place.
  • the first amino acid used for the synthesis is preferably bound to the support material by a peptide bond between the amino group of the amino acid and the acid function (activated by hexafluoroacetone or hexafluorothioacetone) of the linker compound.
  • the further attachment of amino acids can advantageously take place N-, O- or S-terminal to the amino, hydroxy or thiol group of the linker compound or C-terminal to the carboxy group of the attached amino acid.
  • the N-, O- or S- or C-terminal binding of the amino acid to the linker compound is controlled by blocking the amino group or the carboxy group of the amino acid and / or the amino, hydroxyl or thiol group the corresponding linker compound with chemical protecting groups.
  • N-, O- or S-terminal connection is to be made via a coupling reagent (e.g. TBTU)
  • a coupling reagent e.g. TBTU
  • Activation of the carboxy function by a coupling reagent e.g. TBTU
  • Carboxyfunction of the amino acid to be linked to be protected In addition, the N, O or S terminus of the linker connection must be protected.
  • amino protective group is the Boc protective group (tert-butoxycarbonyl)
  • the methyl ester represents a protected carboxy function.
  • thiol or alcohol protective group is the Bzl protective group (benzyl
  • the ER spectra were recorded on the FTIR TM Genisis Series from ATI Mattson.
  • the elemental analyzes were measured on the VarioEL V2.6 from Elementar Analysensysteme GmbH.
  • Occupancy (IV elementary analysis): 92% of the max. Occupancy (0.6 mmol / g)
  • Example 3 1 g (0.6 mmol) of Wang resin (Acros, 1% cross-linked with DVB, 0.5-0.6 mmol / g, 200-400 mesh) are placed in 15 ml of dry chloroform. After a swelling time of 30 min, 271 mg (0.9 mmol) of (R) - (5-oxo-2,2-bis-trifluomomethyl- [1,3] dioxolan-4-yl) acetyl chloride in 1 ml of dry chloroform are stirred added. After 40 min, 76 mg (0.9 mmol) NaHCO 3 is added. The reaction mixture is stirred for 25 h. The product is then filtered off, washed with ice water and ethyl acetate and dried in an oil pump vacuum.
  • Wang resin Acros, 1% cross-linked with DVB, 0.5-0.6 mmol / g, 200-400 mesh
  • Occupancy (_E elementary analysis): 63% of the max. Occupancy (0.6 mmol / g)
  • Example 4 1 g (0.8 mmol) of Wang resin ( ⁇ ovabiochem, 1% cross-linked with DNB, 0.5-1J mmol / g, 100-200 mesh) are placed in 10 ml of dry chloroform. After a swelling time of 30 min, 389 mg (1.2 mmol) of racemic (5-oxo-2,2-bis-h ⁇ fluoromethyl- [1,3] oxathiolan-4-yl) acetyl chloride in 1 ml of dry chlorofomi are added with stirring. After 10 min 65 mg (0.8 mmol) of dry pyridine in 1 ml of dry chlorofomi is added dropwise. The reaction mixture is stirred for 44 h. The product is then filtered off, washed with ethyl acetate and dried in an oil pump vacuum.
  • Wang resin ⁇ ovabiochem, 1% cross-linked with DNB, 0.5-1J mmol / g, 100-200 mesh
  • Another rule for obtaining the substance is as follows.
  • Example 5 1,352 g (0.5 mmol) of HMPA-PEGA resin (Novabiochem, 0.2-0.4 mmol / g) dried in an oil pump vacuum are placed in 20 ml of dry chloroform. After a swelling time of 30 min, 257 mg (0.8 mmol) of racemic (5-oxo-2,2-bis-hifluoromethyl- [1,3] oxathiolan-4-yl) acetyl chloride in 1 ml of dry chlorofomi are added with stirring. After 10 minutes, 43 mg (0.5 mmol) of dry pyridine in 1 ml of dry chloroform is added dropwise. The reaction mixture is stirred for 50 h. The product is then filtered off, washed with ethyl acetate and dried in an oil pump vacuum.
  • racemic 5-oxo-2,2-bis-hifluoromethyl- [1,3] oxathiolan-4-yl
  • acetyl chloride in 1
  • Example 6 1 g (0.6 mmol) of Wang resin (Acros, 1% cross-linked with DNB, 0.5-0.6 mmol / g, 200-400 mesh) is placed in 10 ml of dry pyridine. After 5 minutes of treatment in an ultrasonic bath and a swelling time of 30 minutes, the mixture is cooled to below 0 ° C. with dry ice and 322 with stirring mg (1.2 mmol) 3-isocyanatomethyl-2,2-bis (ti'ifluormethyl) -l, 3-oxazolidin-5-one added. The reaction mixture is treated for 5 min with ice water cooling in an ultrasonic bath, stirred for 2 h at 4 ° C. (ice water) and overnight at room temperature. The product is then filtered off, washed with ethyl acetate and dried in an oil pump vacuum.
  • Wang resin Acros, 1% cross-linked with DNB, 0.5-0.6 mmol / g, 200-400 mesh
  • Occupancy (N-elementary analysis): 93% of the max. Occupancy (0.6 mmol / g)
  • Example 8 500 mg (0.3 mmol) of Wang resin (Acros, 1% cross-linked with DNB, 0.5-0.6 mmol / g, 200-400 mesh) is placed in 10 ml of dry chloroform. After a swelling time of 30 min, 133 mg (0.5 mmol) of racemic 4-isocyanatomethyl-2,2-bis-trifluomomethyl-[1,3] oxathiolan-5-one in 1 ml of dry chloroform are added with stirring. The reaction mixture is stirred for 8 days at room temperature. The product is then filtered off, washed with ethyl acetate and dried in an oil pump vacuum.
  • Wang resin Acros, 1% cross-linked with DNB, 0.5-0.6 mmol / g, 200-400 mesh
  • Occupancy (S elementary analysis): 25% of the max. Occupancy (0.6 mmol / g)
  • Example 9 0.507 g (0.4 mmol) of Rinkamid-4-methylbenzhyd ⁇ ylamine polymer resin (Acros, 1% cross-linked with DNB, 0.4-0.8 mmol / g, 200-400 mesh) are placed in 5 ml of dry chloroform. After a swelling time of 30 min, 146 mg (0.5 mmol) of (R) - (5-oxo-2,2-bis-trifluoromethyl- [1,3] dioxolan-4-yl) acetyl chloride in 85 mg propene oxide are added with stirring , The The reaction mixture is stirred overnight. The product is then filtered off, washed with ethyl acetate and dried in an oil pump vacuum.
  • Rinkamid-4-methylbenzhyd ⁇ ylamine polymer resin Acros, 1% cross-linked with DNB, 0.4-0.8 mmol / g, 200-400 mesh
  • Occupancy 50% of the max. Occupancy (0.8 mmol / g)
  • Occupancy (S elementary analysis): 71% of the max. Occupancy (0.8 mmol / g)
  • Example 12 Synthesis on the surface-functionalized support material (33) according to the invention (Scheme 2)
  • Scheme 2 shows reagents and conditions for the synthesis: i) means DMAP, Py, RT (room temperature), ii) means H-Lys (Z) -OMe x HCl, DMAP, Py, RT. iii) stands for DMAP, Py, RT. iv) means TFA, H 2 O, RT.
  • the yield of reaction (33) ⁇ (50) is 45%.
  • Scheme 3 shows the reagents and conditions: i) means 2 HFA, DMF, RT, where the
  • Yield is 90%, ii) means SOCl 2 , RT, the yield being questioned 100%, iii) means trimethylsilyl azide, toluene, 0 ⁇ 80 ° C., the yield being 40%.
  • Hexafluoroacetone is slowly introduced into a slurry of 4.97 g (263 mmol) (51) and 20 ml of dry DMF (ultrasound) with vigorous stirring. After the reaction has ended (hexafluoroacetone backflow in a dry ice cooler, bubble counter, 19 F-NMR, clear solution), the solution is concentrated in an oil pump vacuum, mixed with ice water (ultrasound) and freeze-dried. The residue is taken up in chloroform, filtered, concentrated in vacuo, mixed with ice water (ultrasound) and freeze-dried. No further cleaning steps are necessary. Yield: 30.36 g (90%), colorless crystals, mp 79-80 ° C.

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  • Chemical Kinetics & Catalysis (AREA)
  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
  • Peptides Or Proteins (AREA)
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Abstract

L'invention concerne des matériaux supports fonctionnalisés en surface, présentant chacun une surface polymère et au moins un composé de liaison lié par covalence à cette dernière. L'invention concerne également la production et l'utilisation desdits matériaux. Dans les matériaux supports selon l'invention, un groupe a-amino-, a-thiol ou a-hydroxy et un groupe carboxy sont protégés par de l'hexafluoroacétone et, parallèlement, des composés de liaison activés par des groupes carboxy sont liés aux réactifs en phase solide, qui possèdent des fonctions hydroxy ou amine (par exemple des résines Wang) par l'intermédiaire de ponts ester, amine ou uréthane. Ces matériaux peuvent être utilisés pour l'immobilisation covalente de biomolécules, la création de bibliothèques de substances en chimie combinatoire, pour la synthèse d'acides aminés, de peptides, de protéines ou de molécules comportant au moins une unité de structure peptidique sur des phases solides en chimie des peptides et pour l'obtention de dérivés de marquage d'affinité.
PCT/DE2004/001684 2003-07-23 2004-07-22 Reactifs en phase solide pouvant se diviser et se lier de maniere multifonctionnelle, et procede pour leur production WO2005009981A1 (fr)

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Cited By (2)

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JP2019073621A (ja) * 2017-10-16 2019-05-16 国立大学法人金沢大学 炭素繊維強化プラスチックの製造方法、炭素繊維強化プラスチック及びセルロース系樹脂
CN114455775A (zh) * 2022-01-05 2022-05-10 江苏合普环保科技有限公司 一种醛类生产中高盐废水的生物工程菌处理方法

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WO1996026213A1 (fr) * 1995-02-21 1996-08-29 Degussa Aktiengesellschaft Procede de preparation de l-aspartyl-d-alanine-n-(thietane-3-yl)-amides

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WO1989008643A1 (fr) * 1988-03-11 1989-09-21 Bioresearch, Inc. Amino acides-n-carboxyanhydrides avec protection urethane
WO1996026213A1 (fr) * 1995-02-21 1996-08-29 Degussa Aktiengesellschaft Procede de preparation de l-aspartyl-d-alanine-n-(thietane-3-yl)-amides

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BURGER K ET AL: "A new efficient approach to N-linked glycopeptoids", TETRAHEDRON LETTERS, ELSEVIER SCIENCE PUBLISHERS, AMSTERDAM, NL, vol. 42, no. 17, 23 April 2001 (2001-04-23), pages 3061 - 3063, XP004233341, ISSN: 0040-4039 *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2019073621A (ja) * 2017-10-16 2019-05-16 国立大学法人金沢大学 炭素繊維強化プラスチックの製造方法、炭素繊維強化プラスチック及びセルロース系樹脂
JP7038994B2 (ja) 2017-10-16 2022-03-22 国立大学法人金沢大学 炭素繊維強化プラスチックの製造方法、炭素繊維強化プラスチック及びセルロース系樹脂
CN114455775A (zh) * 2022-01-05 2022-05-10 江苏合普环保科技有限公司 一种醛类生产中高盐废水的生物工程菌处理方法

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