HK1208198B - Branched fatty acids as liquid cation exchangers - Google Patents

Description

The present invention relates to a process for removing an organic compound from an aqueous solution and a reaction mixture comprising an aqueous solution containing an organic compound.

A fundamental problem in the biotechnological production of fine chemicals from renewable raw materials which are otherwise synthesized from fossil fuels is the conversion of the product once obtained, which is typically in a high-volume aqueous phase, into an organic phase, which is carried out on the one hand to concentrate a finished intermediate or final product and to allow synthetic processing in organic solution in subsequent reaction steps, if necessary, and on the other to improve the yield of the reaction in the aqueous phase by removing the desired product or to allow the reaction to proceed in a technically reasonable manner.

The distribution of a compound in a two-phase equilibrium system comprising an aqueous hydrophilic phase and an organic hydrophobic phase that do not mix is largely dependent on the physicochemical properties of the compound concerned. Whereas compounds with a high content of or consisting exclusively of unsubstituted hydrocarbons predominantly enrich in the hydrophobic phase, compounds with a high content of polar groups such as heteroatomic functionalities and particularly compounds with charges predominantly or practically occur exclusively in the aqueous phase, making it difficult to convert to an organic phase.

The distribution of a compound in the two-phase system mentioned above after equilibrium is often determined by means of distribution coefficients, e.g. the Nernst equation. A special coefficient of distribution is K_ow, also known as the p-value, which characterizes the equilibrium of distribution of a compound between an octanol and an aqueous phase:

Examples of industrially highly demanded organic compounds that are positively charged in aqueous solution at physiological pH are 12-aminolauric acid (ALS) and its derivatives, in particular the methyl ester (ALSME). ALS is an important starting product in the production of polymers. ALS is traditionally produced from fossil raw materials in a low-yield process using laurinlactam, which is produced by trimeration of butadiene, then hydration to form cyclododecan, then oxidation to cyclododecan, transformation with hydroxylaurin and then Beck-rearrangement.

The state of the art teaches the extraction of positively charged organic compounds by contacting an aqueous reaction mixture including a metabolic cell with an organic phase including an organic solvent. For example, DE10200710060705 describes the extraction of the product ALSME by shaking with acetic acid ethyl ester from an aqueous reaction mixture. Asano et al. (2008) reveal the extraction of ALS with toluene from an aqueous reaction solution including an ALS synthesizing enzyme (Asano, Y., Fukuta, y., Yoshida, Y., and Komeda, H. (2008): The Screening, Characterization, and Use of ω-Laurohydroxyacetyl A: Synthesis of Biochemical Enzymes: New Amino Acid, 12-81-2150, Acid, 214 (2008).

US 4 855 053 describes a method for the extraction of organic compounds such as amino acids, amines, lactames and phenols from aqueous solutions using branched and unbranched carbonic acids.

The present invention is therefore intended to provide a method for the removal of positively charged organic matter as described in claim 1 and a corresponding reaction mixture as described in claim 11.

These and other tasks are solved by the subject matter of the attached independent claims, embodiments of which are derived from the subclaims.

In a first aspect, the problem underlying the invention is solved by a process for removing an organic compound from an aqueous solution as claimed in claim 1.

In a second embodiment of the first aspect, which is also an embodiment of the first embodiment, the problem is solved by a method where the ratio of liquid cation exchanger to organic compound in step (b) is at least 1.

In a third embodiment of the first aspect, which is also an embodiment of the first to second embodiments, the problem is solved by a method with a volume ratio of organic solution to aqueous solution of 1:10 to 10:1.

In a fourth embodiment of the first aspect, which is also an embodiment of the first to third embodiments, the problem is solved by a process in which the liquid cation exchanger is a branched fatty acid of the formula (H)₃C)_{2 and}CH-(CH_{2 and}(b)_n-COOH or an unprotonated form thereof and n is at least 4, preferably at least 8 and preferably 14

In a fifth embodiment of the first aspect, which is also an embodiment of the first to fourth embodiments, the problem is solved by a process where the aqueous solution continues to contain a metabolic cell.

In a sixth embodiment of the first aspect, which is also an embodiment of the first to fifth embodiments, the problem is solved by a process in which the organic solution also contains at least one organic solvent, preferably a fatty acid and/or a fatty acid ester.

In a seventh embodiment of the first aspect, which is also an embodiment of the first to sixth embodiments, the problem is solved by a process in which the hydrophobic organic solution contains the liquid hydrophobic cation exchanger in a volume proportion of 20 to 80%, preferably 25 to 75%.

In an eighth embodiment of the first aspect, which is also an embodiment of the first to seventh embodiments of the first aspect, the organic compound is a diamine selected from the group comprising butandiamine, 1,5-pentandiamine, 1,6-hexandiamine, 1,8-octandiamine, 1,14-tetradecandiamine, 1,18-octadecandiamine, 2-methyl-1,5-diaminopentane, 2,2-dimethyl-1,5-diaminopentane, 4,4'-diaminodicyclohexylmethane, 3,3'-dimethyl-4,4'-diaminodicyclohexylmethane, 3,3',5,5'-tetramethyl-4,4'-diaminodicyclohexylmethane, 2.2.4- or 2.4.4-trimethylhexandiamine, 1.4-diaminethylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxy

In a ninth embodiment of the first aspect, which is also an embodiment of the first to eighth embodiments, the temperature at step (b) is between 28 and 70 °C, preferably between 30 and 37 °C.

In a second aspect, the problem underlying the invention is solved by a reaction mixture as claimed in claim 11. Other Other Other^{2 and}R³H^OtherThe following shall be reported:^{1 and}(I) Other Other or

In a first embodiment of the second aspect, the problem is solved by a reaction mixture, whereby the liquid hydrophobic cation exchanger is a branched fatty acid of the formula (H₃C)_{2 and}CH-(CH_{2 and}(b)_n-COOH or an unprotonated form thereof and n is at least 4, preferably at least 8 and preferably 14

In a second embodiment of the second aspect, which is also an embodiment of the first embodiment of the second aspect, the aqueous solution continues to contain a metabolic cell.

In a third embodiment of the second aspect, which is also an embodiment of the first to second embodiments of the second aspect, the organic compound is a diamine selected from the group comprising butandiamine, 1,5-pentandiamine, 1,6-hexandiamine, 1,8-octandiamine, 1,14-tetradecandiamine, 1,18-octadecandiamine, 2-methyl-1,5-diaminopentane, 2,2-dimethyl-1,5-diaminopentane, 4,4'-diaminodicyclohexylmethane, 3,3'-dimethyl-4,4'-diaminodicyclohexylmethane, 3,3',5,5'-tetramethyl-4,4'-diaminodicyclohexylmethane, 2.2.4- or 2.4.4-trimethylhexamine, 1.4-diaminethylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylenedioxylen

The inventors of the present invention have found that the efficiency of removing a positively charged organic compound from an aqueous solution to a hydrophobic organic solution can be increased surprisingly when this organic solution contains a liquid cation exchanger, which is a saturated alkane acid with at least one alkyl substituent. Without being bound by any theory, the inventors of the present invention assume that the negative charge or negative charges of the liquid cation exchanger interact ionically with the positive charge or positive charges of the organic compound and that this interaction leads to at least one positive charge masking which increases the masking in the organic phase of the solution.

Err1:Expecting ',' delimiter: line 1 column 465 (char 464)₃- (CH)_{2 and}(b)_n-COOH or an unprotonated form thereof, in which at least one hydrogen atom from the alkyl chain is replaced by an alkyl substituent of the formula -CH_{2 and}(b)_m-H is substituted, where n and m may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 respectively and independently of each other. In the most preferred embodiment, the saturated alkane acid with at least one alkyl substituent is a compound from the group comprising isostaric acid, isopalmitic acid, isomyristinic acid, phytanic acid, 2-hexyldecylic acid, 2-butyldecylic acid, 15-methylhexadecylic acid, 13-methyltetrahydrocetacetic acid. In a particularly preferred embodiment, the exchanged carbon atoms in the alkyl are substituted, especially with the latter.

In a preferred embodiment, the liquid cation exchanger is a branched fatty acid that contains exclusively non-cyclic substituents, i.e. the molecule is linear, i.e. it does not contain ring-shaped structures.

Like all compounds in this notification, a saturated alkane acid with at least one alkyl substituent in a preferred embodiment includes equally unprotonated, partially and fully protonated forms of the compound.

The method of the invention is suitable for the removal of structurally diverse organic compounds with a positive charge and a positive or neutral total charge from an aqueous solution. Other Other Other^{2 and}R³H⁺-A- NO⁴R⁵H⁺(ii) the following Other Other where A is an alkyl group with at least three, preferably at least six, preferably at least eight carbon atoms, preferably unsubstituted and straight-chained. A may be a branched or unbranched, linear or cyclic alkane or an aromatic or heteroaromatic substituent or substituent with at least two of the substituents specified in formula (II). In a particularly preferred embodiment, the organic compound is a compound of formula II, and A is an alkyl chain of formula - (CH2) where n is 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 26, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 40 and preferably in addition to R2^{2 and}= R³= R⁴= R⁵= H. and where R^{2 and}, R³, R⁴, and R⁵The preferred embodiment is aminolauric acid or an ester thereof, preferably the methyl ester.

Err1:Expecting ',' delimiter: line 1 column 101 (char 100)

The aqueous solution from which the organic compound is removed is preferably a buffer solution containing water or an aqueous culture medium containing water or even more preferably water than the predominant solvent. For example, the solvent content of water in the aqueous solution is more than 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, 99 per cent by volume or 100 per cent by volume. The professional is aware of numerous aqueous buffer culture media suitable for the preservation or culture of cells, especially biotechnologically important cells. These include equally complete media such as LB media, minimal media such as M9 media, minimally complex excipients or combinations of the above-mentioned selective media such as peptone, andErr1:Expecting ',' delimiter: line 1 column 261 (char 260)In a preferred embodiment, the pH of the aqueous culture medium is kept in step (b) for at least 0,5 h, preferably at least 2 h, preferably at least 6 h, preferably at least 12 h, in the pH range 4 to 9, preferably between 4,5 and 8,5, preferably between 6,2 and 7,2. The professional is familiar with the way in which the pH of an aqueous solution, especially an aqueous solution containing metabolic cells and the media necessary for their culture, can be adjusted and regulated by the addition of acid or base, such as sulphuric acid or ammonia water.In another preferred embodiment, the temperature is between 0 and 45 °C, preferably between 15 and 40 °C, preferably between 20 and 37 °C.

Err1:Expecting ',' delimiter: line 1 column 130 (char 129)

Err1:Expecting ',' delimiter: line 1 column 101 (char 100)Err1:Expecting ',' delimiter: line 1 column 291 (char 290)i.e. total neutral total charge.

Err1:Expecting ',' delimiter: line 1 column 110 (char 109)

Err1:Expecting ',' delimiter: line 1 column 103 (char 102)

Err1:Expecting ',' delimiter: line 1 column 261 (char 260)_{2 and}Err1:Expecting ',' delimiter: line 1 column 259 (char 258)

The contact between aqueous and organic solution shall be carried out under appropriate conditions and in particular over a period sufficient to allow the organic compound to pass sufficiently from the aqueous phase to the organic phase, ideally even to bring it into equilibrium. This period and conditions may be determined by the practitioner in the course of routine experimentation. In a preferred embodiment, step (b) shall take at least 0,25, 0,5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 18, 24, 36 or 48 hours or longer. Additional information on possible ways of carrying out the invention is described in EP11154707.

The temperature at step (b) depends not only on the properties of the liquid cation exchanger but, in particular, in the case where contact between the aqueous and organic solution occurs during the aqueous phase of the reaction, also on the temperature requirements of any reactions occurring in the aqueous phase. In particular, in the case where a metabolic cell is catalytically active in the aqueous phase, the temperature must be suitable for maintaining this activity. In a preferred embodiment, the temperature at step (b) is 0 to 100 °C, preferably 20 to 80 °C, 28 to 70 °C, 30 to 37 °C, 35 to 40 °C.

The pH of step (b) must also take into account the requirements of any concurrent reactions, the transition of the dissolved organic compound to the liquid ion exchanger phase, the stability of conduits, products, intermediates or agents.

In order to convert the organic compound from the aqueous phase to the organic one as completely as possible, a sufficient amount of the liquid hydrophobic cation exchanger is required. In a preferred embodiment of the present invention, the ratio of the substance to the liquid cation exchanger and organic compound is at least 1, i.e. at least one molecule of liquid hydrophobic cation exchanger is used per molecule of the organic compound. In an even more preferred embodiment, the ratio is greater than 2, 3, 5, 10, 15, or 20, respectively, 1.5 to 3.

The volume ratio of the organic solution to the aqueous solution, together with the cation exchanger/organic compound ratio, is important for an efficient process. In a particular embodiment, it is 100:1 to 1:100, preferably 20:1 to 1:20, and preferably 10:1 to 1:10, 4:1 to 1:4, 3:1 to 1:3 or most preferably 1:2 to 2:1.

In addition to the liquid hydrophobic cation exchanger, the hydrophobic organic phase may still contain a hydrophobic solvent. This may be used to increase the uptake capacity of a liquid hydrophobic cation exchanger in the hydrophobic phase and prevent undesirable behavior such as flocculation. In a preferred embodiment, the solvent is an educt of a reaction occurring in the aqueous solution, most strongly preferred a substrate catalyzed reaction occurring in the aqueous solution. In a preferred embodiment, it is an enzyme ester of a fatty acid. In a particularly preferred embodiment, it is a fluorescent ester of a methylated acid, which is used as a hydrophobic or hydrophobic mixing agent, and can also be used as a hydrophobic or hydrophobic exchanger.

The percentage of solvent, if present, in the hydrophobic organic phase is 1 to 99% by volume (vol.%) in a preferred embodiment, 10 to 90%, preferably 20 to 80%, preferably 25 to 75% by volume.

Err1:Expecting ',' delimiter: line 1 column 709 (char 708)Err1:Expecting ',' delimiter: line 1 column 773 (char 772)Err1:Expecting ',' delimiter: line 1 column 293 (char 292)

The method can be used to first oxidize and then aminate fatty acids or their esters, for example by an enzyme system as described in international patent application WO 2009/077461. The metabolic cell is a cell that has a recombinant alkane hydroxylase and a transaminase, preferably at least one enzyme from the group consisting of alcohol hydroxylase, alanine hydroxylase and lactamhydrolase.

Err1:Expecting ',' delimiter: line 1 column 673 (char 672)Err1:Expecting ',' delimiter: line 1 column 360 (char 359)Err1:Expecting ',' delimiter: line 1 column 246 (char 245)

Err1:Expecting ',' delimiter: line 1 column 103 (char 102)

Err1:Expecting ',' delimiter: line 1 column 103 (char 102)

Err1:Expecting ',' delimiter: line 1 column 103 (char 102)⁺For example, the alanine hydrogenases from Bacillus subtilis (database code L20916), Rhizobium leguminosarum (database code CP001622), Vibrio proteolytikus (database code AF070716), Mycobacterium tuberculosis (database code X63069), Enterobacter aerogenes (database code AB013821) may be used, and the following substances may be used:

Err1:Expecting ',' delimiter: line 1 column 673 (char 672)Err1:Expecting ',' delimiter: line 1 column 361 (char 360)Err1:Expecting ',' delimiter: line 1 column 246 (char 245)

Err1:Expecting ',' delimiter: line 1 column 247 (char 246)Err1:Expecting ',' delimiter: line 1 column 346 (char 345)Err1:Expecting ',' delimiter: line 1 column 107 (char 106)

For optimal electron donation to CYP153 family cytochrome P450 monooxygenase from the reducing agent, preferably NADH, it is preferable to use the monooxygenase together with its functionally interacting ferredoxin reductase and its functionally interacting ferredoxin, which may be isolated or, when used with a metabolic cell, co-expressed polypeptides or polypeptides N- or C-terminally fused with CYP153 family cytochrome P450 monooxygenase. Whether a ferredoxin reductase or a ferredoxin with a given P450 monooxygenase from the CYP153 family is a functionally interacting polypeptide can be easily determined by the presence of a specialist and three substrate oxide reductase agents.Alternatively, the enzyme test described by Scheps, D., Malca, H., Hoffmann, B., Nestl, B. M., and Hauer, B. (2011) Org. Biomol. Chem., 9, 6727 may be used, which shows a significant increase in reaction rate in the case of functionally interacting polypeptides. In a particularly preferred embodiment, the cytochrome P450 monooxygenase of the CYP153 family, ferredoxin and ferredoxin reductase are derived from the same organism. In a particularly preferred embodiment, the ferredoxin reductase variant Alvorax borkumensis2 (databank YP_196923) or one of its variants derived from Alvorax borkumensis2 (databank YP_19692 or SK_19692 or a variant thereof derived from Alvorax borkumensis2 or from Alvorax cytochrome PK_19692 or from SK_19692 or from Alvorax borkumensis2 (databank SK_19692) or a variant thereof derived from Alvorax borkumensis2 (databank YP_19692 or a variant thereof derived from Alvorax borkumensis2 or SK_19393 or from Alvorax borkumensis2 or from Alvorax borkumensis2 or from Alvorax borkumensis2 or from SK_19692 or from Alvorax borkumensis2 or from Alvorax borkumensis2 or from Alvorax borkumensis2 or from SK_19392 or from Alvorax borkumensis2 or from Alvorax borkumensis2 or from Alvorax borkumensis2 or from Alvorax borkum.

Err1:Expecting ',' delimiter: line 1 column 650 (char 649)which has a homology, here used synonymously with identity, of 70, 75, 80, 85, 90, 92, 94, 96, 98, 99%, or more with respect to the corresponding original wild-type nucleic acid or amino acid sequence, preferably deleted or replaced with amino acids other than those forming the catalytically active centre or essential for structure or folding, or with the latter merely conservatively replaced, such as glutamate instead of aspartate or leucine instead of valine. The state of the art describes algorithms that can be used to calculate the extent of two-sequence homology, e.g. Arthur Lesk (2008), Introduction to bioinformatics,Err1:Expecting ',' delimiter: line 1 column 672 (char 671)preferred 2,_M- and/or k_CatErr1:Expecting ',' delimiter: line 1 column 209 (char 208)

Err1:Expecting ',' delimiter: line 1 column 129 (char 128)Err1:Expecting ',' delimiter: line 1 column 840 (char 839)Hybridisation takes place in a preferred embodiment under strict conditions, i.e. only hybrids are produced where the probe and target sequence, i.e. the polynucleotides treated with the probe, are at least 70% identical. It is known that the severity of hybridisation, including the washing steps, is influenced or determined by variations in buffer composition, temperature and salt concentration. The hybridisation reaction is generally carried out at relatively low stress compared to the washing steps (Hybaid Hybridisation Guide, UK, Teddington, 1996). For the corresponding reaction, for example, a 5C buffer hybridisation reaction can be carried out at a caffer temperature.The test chemical is used to determine the concentration of polynucleotides in the test medium, and the concentration of polynucleotides in the test medium is used to determine the concentration of polynucleotides in the test medium. The test chemical is used to determine the concentration of polynucleotides in the test medium.The concentration of salt may be reduced to a concentration corresponding to 0,2 x SSC or 0,1 x SSC, if necessary. By gradually increasing the hybridisation temperature in steps of approximately 1-2°C from 50°C to 68°C, polynucleotide fragments may be isolated, which, for example, are, as a result of increasing preference, at least 70% or at least 80% or at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the sequence identity of the introduced nucleic acid.Err1:Expecting ',' delimiter: line 1 column 292 (char 291)

The second step is to separate the hydrophobic organic solution from the aqueous culture medium. This is an easy process for the professional to perform, simply by stopping the vessel and then decanting a phase of the culture, due to the inherent ability of this system to form two phases. Alternatively, a separation funnel can be used. In the case of sufficiently different boiling points, it is possible to reduce the boiling phase at lower temperatures, which will usually be the organic phase, by applying a pressure. Small amounts of water remaining in the organic phase can be removed using inorganic dry agents such as calcium hydrogels, calcium chloride, sulphur dioxide, sodium hydroxide or sodium nitrate.

Further guidance on the implementation of the invention can be found in PCT/EP2011/071491, where the fatty acids described therein are to be replaced by those according to the invention.

Claims (15)

1. Method for removing an organic compound from an aqueous solution, comprising the steps

   a) providing the aqueous solution which contains the organic compound, and a hydrophobic organic solution, wherein the latter comprises a liquid hydrophobic cation exchanger,

   b) contacting the aqueous solution and the hydrophobic organic solution, and

   c) separating off the hydrophobic organic solution from the aqueous solution, wherein the liquid hydrophobic cation exchanger is a saturated alkanoic acid having at least one alkyl substituent, which comprises at least 12 carbon atoms, wherein the organic compound is a compound of the formula         NR²R³H⁺-A- NR⁴R⁵H⁺     (II),

   wherein A is an alkylene group having at least three, preferably at least six, particularly preferably eight, carbon atoms, which is preferably unsubstituted and straight-chain,

   and wherein R², R³ and R⁴, and R⁵, in each case and independently of one another, are selected from the group consisting of hydrogen, methyl, ethyl and propyl; wherein the pH in the aqueous solution in step b) is 6 to 8, preferably 6.2 to 7.2.
2. Method according to Claim 1, wherein the molar ratio of liquid cation exchanger to organic compound in step b) is at least 1.
3. Method according to any one of Claims 1 to 2, wherein the volumetric ratio of organic solution to aqueous solution is 1:10 to 10:1.
4. Method according to any one of Claims 1 to 3, wherein the liquid cation exchanger is a branched-chain fatty acid of the formula (H₃C)₂CH-(CH₂)_n-COOH or an unprotonated form thereof and n is at least 4, preferably at least 8, and most preferably is 14.
5. Method according to any one of Claims 1 to 4, wherein the liquid cation exchanger is a branched-chain fatty acid which exclusively comprises substituents that are other than cyclic substituents.
6. Method according to any one of Claims 1 to 5, wherein the aqueous solution additionally comprises a metabolically active cell.
7. Method according to any one of Claims 1 to 6, wherein the organic solution additionally contains at least one organic solvent, preferably a fatty acid and/or a fatty acid ester.
8. Method according to Claim 7, wherein the hydrophobic organic solution contains the liquid hydrophobic cation exchanger in a volumetric fraction of 20 to 80%, preferably 25 to 75%.
9. Method according to any one of Claims 1 to 8, wherein the organic compound is a diamine from the group consisting of 1,4-butanediamine, 1,5-pentanediamine, 1,6-hexanediamine, 1,8-octanediamine, 1,14-tetradecanediamine, 1,18-octadecanediamine, 2-methyl-1,5-diaminopentane, 2,2-dimethyl-1,5-diaminopentane, 4,4'-diaminodicyclohexylmethane, 3,3'-dimethyl-4,4'-diaminodicyclohexylmethane, 3,3',5,5'-tetramethyl-4,4'-diaminodicyclohexylmethane, 2,2,4- or 2,4,4-trimethylhexamethylenediamine, 1,4-diaminocyclohexane, 4,4'-diaminodicyclohexylpropane and isophoronediamine.
10. Method according to any one of Claims 1 to 9, wherein the temperature in step b) is between 28 and 70°C, preferably between 30 and 37°C.
11. Reaction mixture comprising an aqueous solution which contains an organic compound and a hydrophobic organic solution, wherein the hydrophobic organic solution comprises a liquid hydrophobic cation exchanger, wherein the liquid hydrophobic cation exchanger is a saturated alkanoic acid having at least one alkyl substituent, which comprises at least 12 carbon atoms, wherein the organic compound is a compound of the formula         NR²R³H⁺-A- NR⁴R⁵H⁺     (II),

    wherein A is an alkylene group having at least three, preferably at least six, particularly preferably eight, carbon atoms which is preferably unsubstituted and straight-chain,

    and wherein R², R³, R⁴, and R⁵ , in each case and independently of one another, are selected from the group consisting of methyl, ethyl, propyl and butyl, and

    wherein the pH in the aqueous solution is 6 to 8, preferably 6.2 to 7.2.
12. Reaction mixture according Claim 11, wherein the liquid hydrophobic cation exchanger is a branched-chain fatty acid of the formula (H₃C)₂CH-(CH₂)_n-COOH or an unprotonated form thereof and n is at least 4, preferably at least 8, and most preferably 14.
13. Reaction mixture according to any one of Claims 11 to 12, wherein the liquid cation exchanger is a branched-chain fatty acid which exclusively comprises substituents that are other than cyclic substituents.
14. Reaction mixture according to any one of Claims 11 to 13, wherein the aqueous solution additionally comprises a metabolically active cell.
15. Reaction mixture according to one of Claims 11 to 14, wherein the organic compound is a diamine from the group consisting of butanediamine, 1,5-pentanediamine, 1,6-hexanediamine, 1,8-octanediamine, 1,14-tetradecanediamine, 1,18-octadecanediamine, 2-methyl-1,5-diaminopentane, 2,2 dimethyl-1,5-diaminopentane, 4,4'-diaminodicyclohexylmethane, 3,3'-dimethyl-4,4'-diaminodicyclohexylmethane, 3,3',5,5'-tetramethyl-4,4'-diaminodicyclohexylmethane, 2,2,4- or 2,4,4-trimethylhexamethylenediamine, 1,4-diaminocyclohexane, 4,4'-diaminodicyclohexylpropane and isophoronediamine.