A process for the preparation of 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid

Abstract

Process for preparing 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) of formula (I), (I),comprising oxidizing a diol of formula (IIa) in an oxidizing step, (IIa) and / or a hydroxymethyl ketone according to formula (IIb): (IIb).

Description

[0001] A Process for the Preparation of 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid

[0002] Technical Field of the Invention

[0003] The present invention relates to a process for the preparation of 2-oxo-4- (hydroxy(methyl)phosphinoyl)butyric acid (PPO). In particular, the present invention relates to the catalytic oxidation of a diol precursor to 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO).

[0004] Background of the Invention

[0005] The herbicide glufosinate is a non-selective, foliarly applied herbicide considered to be one of the safest herbicides from a toxicological or environmental standpoint. Current commercial chemical synthesis methods for glufosinate yield a racemic mixture of L- and D-glufosinate (Duke et al. 2010 Toxins 2:1943-1962). However, L-glufosinate (also known as phosphinothricin or (S)-2-amino-4-(hydroxy(methyl)phosphonoyl)butanoic acid) is much more potent than D-glufosinate (Ruhland et al. (2002) Environ. Biosafety Res. 1:29-37). Therefore, methods are needed to produce only or primarily the active, L-glufosinate form. Previously, cost effective methods to generate pure L-glufosinate, or a mixture of D- and L-glufosinate enriched for L-glufosinate, have not been available. Therefore, efficient reactions have been developed leading to L-glufosinate, such as proposed in WO 2017 / 151573 Al. This reaction route implements 2-oxo-4- (hydroxy(methyl)phosphinoyl)butyric acid (PPO) as intermediate species. Synthesis routes to PPO have been further investigated inter alia in EP 0 030 424 Bl.

[0006] WO 2024 / 051121 Al discloses a method for preparing a pesticide intermediate 4- (hydroxymethylphosphonyl)-2-carbonylbutyric acid (PPO). However, the method disclosed therein comprises the step of mixing acryloyl chloride, a first solvent, a polymerization inhibitor, a catalyst and potassium ferrocyanide. This reaction results in the formation of acryloyl cyanide, which is a highly toxic compound, which can be absorbed through skin, mouth, and inhalation. Hence, industrial application of such a method requires a secure safety protocol and measures to ensure that it is followed. This makes the overall process complicated, prone to error, expensive and still bears the risk of a hazard.

[0007] Summary of the invention

[0008] Hence, efficiency of the production process of PPO can always be improved. Improving the efficiency of the production process of PPO will also improve the overall efficiency of the production of glufosinate, in particular L-glufosinate. Hence, it is an object of the present invention to provide a process for the production of PPO, which is easy to carry out, has good yields, easy to work-up and the starting materials are easily accessible.

[0009] It has now been found out that above-mentioned object can be achieved by a process for preparing 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I), and / or PPO salts, more particularly PPO-sodium or PPO-ammonium: comprising oxidizing in an oxidating step a diol of formula (Ila): and / or a hydroxymethyl ketone according to formula (lib):

[0010] (lib).

[0011] It has been further surprisingly found that above-mentioned object can be achieved by the use of a mixture comprising 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I)

[0012] (I), and aditionally a diol according to formula (Ila): and / or a hydroxymethyl ketone according to formula (lib):

[0013] (lib). for preparing glufosinate and / or glufosinate salts, more particularly glufosinate, glufosinate- sodium or glufosinate-ammonium.

[0014] Finally, it has been surprisingly found that above-mentioned object can be achieved by a mixture comprising 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I)

[0015] (I), and aditionally a diol according to formula (Ila): and / or a hydroxymethyl ketone according to formula (lib):

[0016] (lib). for preparing glufosinate and / or glufosinate salts, more particularly glufosinate, glufosinate- sodium or glufosinate-ammonium.

[0017] Definitions

[0018] As used in this specification and in the appended claims, the singular forms of "a" and "an" also include the respective plurals unless the context clearly dictates otherwise. In the context of the present invention, the terms "about" and "approximately" denote an interval of accuracy that a person skilled in the art will understand to still ensure the technical effect of the feature in question. The term typically indicates a deviation from the indicated numerical value of ± 20 %, preferably ± 15 %, more preferably ± 10 %, and even more preferably ± 5 %. It is to be understood that the term "comprising" is not limiting. For the purposes of the present invention the term "consisting of" is considered to be a preferred embodiment of the term "comprising of". If hereinafter a group is defined to comprise at least a certain number of embodiments, this is meant to also encompass a group which preferably consists of these embodiments only. Furthermore, the terms "first", "second", "third" or "(a)", "(b)", "(c)", "(d)" etc. and the like in the description and in the claims, are used for distinguishing between similar elements and not necessarily for describing a sequential or chronological order. It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the invention described herein are capable of operation in other sequences than described or illustrated herein. In case the terms "first", "second", "third" or "(a)", "(b)", "(c)", "(d)", "i", "ii" etc. relate to steps of a method or use or assay there is no time or time interval coherence between the steps, i.e. the steps may be carried out simultaneously or there may be time intervals of seconds, minutes, hours, days, weeks, months or even years between such steps, unless otherwise indicated in the application as set forth herein above or below. It is to be understood that this invention is not limited to the particular methodology, protocols, reagents etc. described herein as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the present invention that will be limited only by the appended claims. Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art.

[0019] As used herein the term “does not comprise” or “free of” means in the context that the composition of the present invention is free of a specific compound or group of compounds, which may be combined under a collective term, that the composition does not comprise said compound or group of compounds in an amount of more than 0.8 % by weight, based on the total weight of the composition. Furthermore, it is preferred that the composition according to the present invention does not comprise said compounds or group of compounds in an amount of more than 0.5 % by weight, preferably the composition does not comprise said compounds or group of compounds at all.

[0020] The term " p / atinum-group meta / s" as used therein denotes the chemically closely related metals platinum, palladium, iridium, rhodium, ruthenium, and osmium, which in nature generally occur together. The use of platinum or palladium, especially platinum, is preferred.

[0021] Ther term "aqueous alkaline medium" as used herein denotes a reaction mixture that reacts alkaline, i.e. which has a pH value of larger than 7.

[0022] Detailed Description of the Invention

[0023] As set out above, the present invention is concerned with a process, a use, and a mixture, which will be described in the following in more detail. Process of the invention

[0024] The present invention is concerned with a process for preparing 2-oxo-4- (hydroxy(methyl)phosphinoyl)butyric acid (PPO) of formula (I) comprising oxidizing in an oxidating step (0) a diol of formula (Ila) and / or a hydroxymethyl ketone according to formula (lib):

[0025] (lib).

[0026] This process enables an easy access to PPO, as the diol can be produced e.g. by radical addition of methylphosphinic acid butyl ester to 3,4-dihydroxy butene or its acetylated derivative and follow up acidic deprotection.

[0027] The oxidizing step according to the process of the present invention can be carried out in the presence of a chemical catalyst (OC) or an enzyme (OE).

[0028] Process using an oxidation step in the presence of an enzyme (OE)

[0029] In the following the preferred embodiment of the present invention is described, in which the oxidating step (0) is carried out in the presence of an enzyme (OE).

[0030] Preferably, if the process according to the present invention is carried out using an enzyme, the enzyme present in the oxidating step (OE) is an Oxidoreductase ECI enzyme. More preferably, the Oxidoreductase ECI enzyme is selected from the group consisting of EC1.1, EC1.2 or EC1.10, more preferably EC1.1.1, EC1.1.3, EC1.2.1, EC1.2.3. Most preferably, the Oxidoreductases ECI enzyme is an enzyme according to SEQ ID NO:10. Preferably, not only one enzyme can be used, but combinations of Oxidoreductases ECI, preferably a combination of the classes EC1.1 and EC1.2.

[0031] If the process according to the present invention is carried out using an enzyme, the oxidating step (OE) the of process is preferably carried out at a temperature in the range of from 25 to 45 ° C, more preferably 30 to 42 ° C, and most preferably 35 to 40 ° C. Also preferably, if the process according to the present invention is carried out using an enzyme, the oxidating step (OE) of the process is carried out in an aqueous solution, preferably a buffered aqueous solution. Most preferably, the pH value of the buffered aqueous solution is in the range of from 8.8 to 7.2, most preferably the pH value of the buffered aqueous solution is 7. Preferably, the buffer aqueous solution is a solution of ammonia in water or isopropyl amine in water. Moreover, if the process according to the present invention is carried out using an enzyme, the oxidating step (OE) of the process is carried out for a period in the range of from 5 to 48 h.

[0032] Preferably, the oxidating step (OE) is carried out in an ex vivo environment. In case that the oxidating step of the process of the present invention is an oxidating step (OE) in the presence of an enzyme, preferably of an Oxidoreductase ECI enzyme, the whole process according to the present invention is carried out in an ex vivo environment.

[0033] The oxidating step (OE) of the process of the present invention is preferably carried out in the presence of a Catalase enzyme in addition to the Oxidoreductase ECI enzyme, preferably a Catalase enzyme according to CAS-Nr. 9001-05-2. It has been surprisingly found that the presence of the Catalase enzyme significantly improves the yields of the compound according to formula (I). Without wishing to be bound by theory it is believed that the improvement in yield is caused by the removal of intermediately formed hydrogen peroxide from the reaction mixture in the oxidating step.

[0034] Process using an oxidation step in the presence of a chemical catalyst (OC)

[0035] In the following the alternatively preferred embodiment is described, in which the oxidation step (O) is carried out in the presence of a chemical catalyst (OC).

[0036] Hence, if the process according to the present invention is carried out in the presence of a chemical catalyst, the chemical catalyst of the oxidating step (OC) is preferably a platinumgroup metal catalyst. Also preferably, the oxidizing step (OC) is carried out in an aqueous solution in the presence of a gas comprising molecular oxygen and an activator. More preferably, the oxidizing step (OC) is carried out in an aqueous solution of an alkali in the presence of a gas comprising molecular oxygen and an activator.

[0037] The platinum-group metal used as the catalyst in the oxidating step (OC) can be added to the reactants in a variety of forms, for example in elementary, i.e. metallic, form, for instance as so-called "black", in combination with other platinum-group metals or in the form of a compound, for example as an oxide or also in the form of some other compound. Preferably, the platinum-group metal used as the catalyst in the oxidating step (OC) is present in metallic form. Preferably, the platinum-group metal catalyst used as the catalyst in the oxidating step (OC) is selected from platinum and palladium.

[0038] The platinum-group metals used as the catalyst in the oxidating step (OC) can be applied to supports. Preferably, the support of the platinum-group metal catalyst as used in the oxidating step (OC) is selected from the group consisting of active charcoal, graphite, kieselguhr, silica gel, spinels, aluminum oxide, asbestos, calcium carbonate, magnesium carbonate, barium sulphate, or organic support material. Active charcoals have proved particularly suitable, for example, inexpensive pulverulent active charcoals, produced from wood, which are extensively used for decolorizing purposes. Hence, preferably, the platinum-group metal catalyst used in the oxidating step (OC) of the process according to the present invention is a supported platinum-group metal catalyst, more preferably a charcoal supported platinum-group metal catalyst.

[0039] The amount of the platinum-group metal comprised in the supported platinum-group metal catalyst as used in the oxidating step (OC) of the process according to the present invention can be less than 10 wt.-% with respect to the total weight of the platinum-metal group catalyst, preferably is in the range of from 0.1 to 5 wt.-%.

[0040] Furthermore, the platinum-group metal catalyst as used in the oxidating step (OC) of the process according to the present invention preferably comprises an activator. More preferably, the activator is selected from lead and / or a compound thereof and / or bismuth and / or a compound thereof.

[0041] The amounts in which the platinum-group metal catalysts are used in the oxidating step (OC) of the process according to the present invention depend on the desired rate of oxidation, the form of the catalyst, the nature and amount of the activator, and so on, and can in a specific case easily be determined by preliminary experiments.

[0042] Preferably, the amount of platinum-group metal used per mole compounds according to formulae (Ila) of (lib) in the oxidating step (OC) of the process according to the present invention is less than 1,000 mg. In most cases sufficiently high reaction rates are achieved with an amount of platinum-group metal of 20 to 400 mg per mole of compounds according to formulae (Ila) or (lib).

[0043] It has been found that tar formation is avoided when using the platinum-group metal catalyst in the oxidating step (OC) of the process according to the present invention. Hence, the platinum-group metal catalysts used in the oxidating step (OC) of the process according to the present invention can be used repeatedly. As a result of this reuse, the consumption of platinum-group metal catalyst per mole of compounds according to formulae (Ila) or (lib) can be reduced to 5 mg or less, before reprocessing of the platinum-group metal catalyst becomes necessary.

[0044] Preferably, in the oxidating step (OC) of the process according to the present invention, the activator is present in an amount of less than 0.1 mole or more with respect to the amount of compounds according to formulae (Ila) or (lib), more preferably of 5 x 10’5to 1 x 10’1mole, even more preferably 1 x 10’4to 1 x 10’2mole.

[0045] The activator as used in the oxidating step (OC) of the process according to the present invention preferably comprise the metals lead and / or bismuth as such, that is to say in the elementary form, and / or in the form of their compounds, for example as oxides, or as salts of hydracids, such as chlorides, bromides, iodides, sulfides, selenides and tellurides; or as salts of inorganic oxy-acids, such as nitrates, nitrites, phosphites, phosphates, sulfates, carbonates, perchlorates, antimonates, arsenates, selenites, selenates, and borates; or as salts of oxy-acids derived from transition metals, for example vanadates, niobates, tantalates, chromates, molybdates, tungstates, and permanganates; or as salts of organic aliphatic or aromatic acids, for example formates, acetates, propionates, benzoates, salicylates, lactates, mandelates, glyoxylates, arylglyoxylates and citrates; or as phenolates and the like. The activators may, in each particular case, be soluble, partially soluble or insoluble in the reaction mixture.

[0046] Hence, the activator used in the oxidating step (OC) of the process according to the present invention preferably comprises one or more compounds selected from the group consisting of lead in elemental form, bismuth in elemental form, lead in oxide form, bismuth in oxide form, lead in hydracid salt form, bismuth in hydracid salt form, lead in form of a salt of an inorganic oxyacid, bismuth in form of a salt of an inorganic oxyacid, lead in form of a salt of a transition-metal-comprising oxyacid, bismuth in form of a salt of a transition-metal- comprising oxyacid, lead in form of a salt of an organic aliphatic or aromatic acid, and bismuth in form of a salt of an organic aliphatic or aromatic acid, more preferably is selected from lead in elemental form and bismuth in elemental form, and most preferably is bismuth in elemental form.

[0047] In a preferred embodiment of the process according to the present invention, in the oxidating step (OC), combinations of these activators with one another and / or with other elements or compounds, not specified as an activator, can also be used. The activators as used in the oxidating step (OC) of the process according to the present invention may be present in various valency levels or in a mixture of valency levels. Furthermore, changes in valency may also occur during the reaction. If the activators have not already been added as oxides and / or hydroxides, it is possible that they become entirely or partially converted to these in the alkaline medium. After the reaction, the platinum-group metal catalyst can be filtered off together with the sparingly soluble activator and be reused in further oxidation reactions. Losses of platinum-group metal catalyst and / or activator, if these occur, must be made up.

[0048] The activator used in the oxidating step (OC) of the process according to the present invention can be added to the reactants as a solid, preferably in a finely divided form, or in the form of a solution. It is also possible to add the activator when preparing the platinum- group metal catalyst, or to impregnate the platinum-group metal catalyst with the activator. The activator can also serve as a support for the platinum metal. Preferably, the activator is incorporated into the platinum-group metal catalyst.

[0049] If the oxidating step (OC) of the process according to the present invention is carried out in the presence of a chemical catalyst, the oxidating step (OC) is preferably carried out in the presence of an aqueous alkaline medium.

[0050] The alkali of the aqueous alkaline medium of the oxidating step (OC) of the process according to the present invention can be added to a solution or suspension of the compounds according to formulae (Ila) or (lib) in water, or the compounds according to formulae (Ila) or (lib) can be dissolved or suspended in the aqueous alkaline solution. Preferably, the amount of alkali is chosen to provide 0.3 to 5, preferably 0.5 to 3, equivalents of alkali per mole of compounds according to formulae (Ila) or (lib) to be oxidized. The use of from 0.9 to 2 equivalents of alkali per mole of compounds according to formulae (Ila) or (lib) to be oxidized is particularly preferred.

[0051] Preferably, the alkali is selected from the list consisting of sodium hydroxide, potassium hydroxide, or carbonate, more preferably the alkali is selected from the list consisting of sodium hydroxide or potassium hydroxide, and most preferably the alkali is sodium hydroxide.

[0052] In the oxidating step (OC) of the process according to the present invention, the concentration of the organic compounds in the aqueous alkaline reaction solution is in general selected so that both the diol according to formula (Ila), the ketone according to formula (lib), and the PPO according to formula (I) formed are present in solution under the reaction conditions. Where appropriate, the diol according to formula (Ila) and / or the ketone according to formula (lib) should be added in portions to the oxidation mixture, preferably together with part of the alkali. Most preferably, the final concentration of organic compounds in the reaction mixture is in the range of from 5 to 30 wt.% with respect to the total weight of the oxidation mixture. Under the conditions according to the invention, an oxidation effect is observable at all temperatures at which a liquid phase is present. Accordingly, the possible reaction temperature of the oxidating step (OC) of the process according to the present invention ranges from the solidification point to the boiling point of the reaction mixture. Preferably, if the oxidating step is carried out in the presence of a chemical catalyst, the oxidating step (OC) is carried out at a temperature of from 10 to 100 ° C, preferably 25 to 85 ° C, and most preferably 45 to 65 ° C.

[0053] The aerating step in the oxidizing step (OC) of the process according to the present invention can be carried out by bubbling gas comprising molecular oxygen through the reaction mixture. Alternatively, the gas comprising molecular oxygen can be applied under a pressure of 0.5 to 10 atm.

[0054] Preferably, the gas comprising molecular oxygen as used in the oxidating step (OC) of the process according to the present invention can be pure oxygen or air.

[0055] The oxidating step of the process of the present invention, when being carried out in the presence of a chemical catalyst, preferably a platinum-group metal catalyst, is preferably carried out in the presence of a Catalase enzyme, preferably a Catalase enzyme according to CAS-Nr. 9001-05-2. It has been surprisingly found that the presence of the Catalase enzyme significantly improves the yields of the compound according to formula (I). Without wishing to be bound by theory it is believed that the improvement in yield is caused by the removal of intermediately formed hydrogen peroxide from the reaction mixture in the oxidating step.

[0056] If a Catalase enzyme is present in the reaction mixture in the oxidating step (OC) of the process of the present invention when being carried out in the presence of a chemical catalyst, preferable a platinum-group metal catalyst, the reaction temperature is preferably in the range of from 25 to 45 ° C, more preferably 30 to 42 ° C, and most preferably 35 to 40 ° C

[0057] Use of the Invention

[0058] Furthermore, the present invention is concerned with the use of a mixture comprising 2- oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I) and aditionally a diol according to formula (Ila) and / or a hydroxymethyl ketone according to formula (lib):

[0059] (lib). for preparing glufosinate and / or glufosinate salts, more particularly glufosinate, glufosinate- sodium or glufosinate-ammonium.

[0060] In particular, the PPO is preferably used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by a transaminase (TA) enzyme, by aminating the PPO to L-glufosinate by an L-amino acid dehydrogenase (LAAD) enzyme, or by chemical conversion. Preferably, the PPO is used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by a transaminase (TA) enzyme or by aminating the PPO to L-glufosinate by an L-amino acid dehydrogenase (LAAD) enzyme. If the PPO is used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by a transaminase (TA) enzyme, the aminating is preferably carried out in the presence of an amine group from one or more amine donors. If the PPO is used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by an L-amino acid dehydrogenase (LAAD) enzyme, the aminating is preferably carried out in the presence of an ammonia source.

[0061] Mixture of the invention

[0062] Finally, the present invention is concerned with a mixture comprising 2-oxo-4- (hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I)

[0063] (I), and aditionally a diol according to formula (Ila) and / or a hydroxymethyl ketone according to formula (lib):

[0064] (lib).

[0065] In a preferred embodiment, the present invention is concerned with a mixture comprising 2- oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I)

[0066] (i), and aditionally a diol according to formula (Ila) and a hydroxymethyl ketone according to formula (lib):

[0067] (lib).

[0068] It has been found that the mixture can be used to produce glufosinate preferably L- glufosinate, by aminating the PPO to L-glufosinate by a transaminase (TA) enzyme, by aminating the PPO to L-glufosinate by an L-amino acid dehydrogenase (LAAD) enzyme, or by chemical conversion. Preferably, the mixture can be used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by a transaminase (TA) enzyme or by aminating the PPO to L-glufosinate by an L-amino acid dehydrogenase (LAAD) enzyme. If the mixture is used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by a transaminase (TA) enzyme, the amination is preferably carried out in the presence of an amine group from one or more amine donors. If the mixture is used to produce glufosinate, preferably L-glufosinate, by aminating the PPO to L-glufosinate by an L-amino acid dehydrogenase (LAAD) enzyme, the amination is preferably carried out in the presence of an ammonia source.

[0069] Preferably, PPO is the predominant compound among the mixture according to the present invention. Preferably, the amount of PPO in the mixture according to the present invention is 80% or greater, 85% or greater, 90% or greater, or about 95% or greater, 97% or greater, 98% or greater based on the combined weight of PPO, diol according to formula (Ila) and ketone according to formula (lib).

[0070] Preferably, the amount of diol according to formula (Ila) in the mixture according to the present invention is 10% or less, 5% or less, 2.5% or less, or 1% or less based on the combined weight of PPO, and diol according to formula (Ila) and ketone according to formula (lib).

[0071] Preferably, the amount of ketone according to formula (lib) in the mixture according to the present invention is 10% or less, 5% or less, 2.5% or less, or 1% or less based on the combined weight of PPO, and diol according to formula (Ila) and ketone according to formula (lib).

[0072] These compositions can optionally occur as dried powders or dissolved in aqueous or nonaqueous carrier and additional chemical species can optionally be present. Optionally, the composition is prepared and used in an ex vivo environment.

[0073] Examples

[0074] Inventive Example IE1

[0075] 1 M NaOH was added to a solution of 2-hydroxy-4-(hydroxy(methyl)phosphinoyl)butanol (100 mg) in water (6 mL) to adjust the pH to 8. 500 mg Pt / Bi on carbon (water content 59%, 1.63% Pt, 0.41% Bi) were added. The reaction mixture was heated to 55 ° C and the pH was adjusted throughout the reaction to 8 by addition of aq. NaOH while aerating with oxygen. After 28 h NMR showed a conversion of 10% to the keto acid (PPO). PPO and intermediates were identified by both NMR and HPLC.

[0076] HPLC Method: The phosphorous containing compounds were separated by an Aminex HPX- 87 H, 300*7.8 mm column. Temperature 30 ° C, Flowrate 0.5 mL / min, Rl Detection, Eluent 5 mM sulfuric acid in water. HPLC retention times: 2-hydroxy-4- (hydroxy(methyl)phosphinoyl)butanol: 16.0 min, PPO 9.7 min,

[0077] Inventive Example IE2

[0078] 1 M NaOH was added to a solution of 2-hydroxy-4-(hydroxy(methyl)phosphinoyl)butanol (100 mg) in water (6 mL) to adjust the pH to 8. 500 mg Pt / Bi on carbon (water content 59%, 1.63% Pt, 0.41% Bi) were added. Catalase (10 mg, CAS: 9001-05-2) was added to the reaction mixture. The reaction mixture was heated to 40 ° C and aerated. The pH value was adjusted throughout the reaction to 8 by addition of aq. NaOH. After 26 h a conversion of 59% to the keto acid (PPO) was measured by NMR.

[0079] Inventive Example IE3 - Cloning of oxidoreductases

[0080] Enzymes were identified from public databases (NCBI, Uniprot). The DNA sequences corresponding to the enzymes were codon-optimized for expression in Escherichia coli (E. coli) and cloned (Twist Bioscience) in pDHE 19.2 vector (DE 19848129). The gene of interest lies under control of a rhamnose inducible promoter (rhaBAD). The synthesized plasmids were used to transform competent cells (Chung, C.T. et al., Proc Natl Acad Sci U S A, 1989, 86, 2172) of the E. coli strain TG10 (WO 2004 / 050877 Al). The E. coli strain TG10 is a rhaA- -derivate of E. coli TGI (DSMZ 6056) transformed with pHSG575 (Takeshita, S. et al., Gene, 1987, 61, 63) and pAgro4 (pBB541 in Tomoyasu, T. et al., Mol. Microbiol., 2001, 40, 397). GalOx (SEQ ID NO:9) may be cloned and expressed as described in Birmingham, W.R., et al., Nature Communications, 2021, 12, 4946) or WO 2016 / 150629 Al. The sequences as listed in Table 1 were cloned based on the above-mentioned protocols.

[0081] Table 1: Tested Enzymes

[0082] Inventive Example IE4 - Recombinant production of oxidases

[0083] E. coli TG10 transformed with the oxidase-containing plasmids were grown in 10 ml LB medium (Bertani, G., J Bacteriol, 1951, 62, 293) supplemented with 100 pg / ml ampicillin, 50 pg / ml spectinomycin, 20 pg / ml chloramphenicol and 12.5 pg / ml tetracyline at 37 ° C for 16 h (200 rpm). This preculture was used to inoculate 100 ml of LB medium supplemented with 100 pg / ml ampicillin, 50 pg / ml spectinomycin, 20 pg / ml chloramphenicol and 12.5 pg / ml tetracyline until the optical density reaches A600 nm of 0.6. To the cultures, 0.2 mM isopropyl-E-D-thiogalactopyranoside, and 0.5 g / L rhamnose were added and incubated further at 22 ° C for 16-18 h (200 rpm). The biomass was harvested by centrifugation at 4000*g for 15 min at 4 C and the pellets were frozen in -20 ° C until further use. Recombinant production of GalOx (SEQ ID NO:10) is described in Birmingham, W.R., et al., Nature Communications, 2021, 12, 4946) or WO 2016 / 150629 Al.

[0084] Inventive Example IE5 - Preparation of the biocatalyst

[0085] The frozen pellets were thawed in 0.1 M potassium phosphate buffer pH 7.8 (3 mL buffer / g pellet). The cell suspension was distributed in 2 ml vials and lyzed in Homogenisator (Precellys ®, VWR) for 2*30 sec cycles. The cell-free lysate was further centrifuged at 12000*g for 15 min at 4 ° C. The clarified supernatant (cell-free extract) was used for the activity assay.

[0086] Inventive Example IE6 - ABTS assay - Activity screening of oxidoreductase

[0087] Enzymatic activity was measured at 30 ° C by incubating the cell-free extract (12.5 vol%) in 200 pl of 0.1 M air-saturated potassium phosphate buffer pH 7.8 containing 5 g / L 2- hydroxy-4-(hydroxy(methyl)phosphinoyl)butanol, 0.2 g / L ABTS (2,2’-azino-bis(3- ethylbenzothiazoline-6-sulfonic acid)) and 0.5 g / L HRP (Horse radish peroxidase). The reaction was monitored by measuring the absorbance of ABTSox at 420 nm up to 2 h. In the presence of GalOx-Protein (SEQ ID NO:10) 2-hydroxy-4- (hydroxy(methyl)phosphinoyl)butanol was oxidized. In the presence of Aoxl-Protein (SEQ ID NO:4) 2-hydroxy-4-(hydroxy(methyl)phosphinoyl)butanol was oxidized. Whereas in the absence of protein no oxidation was observed.

[0088] Comparative Example CE1:

[0089] PPO

[0090] 65 mg of 2-oxo-3-butenoic acid was suspended in 1 mL of methylene chloride. The reaction mixture was heated to 30 ° C. Subsequently, 100 pL of dichloro(methyl)phosphane was added, and the mixture was stirred at 30 ° C for 5 hours, after which it was allowed to cool to room temperature. The reaction mixture was then stirred at room temperature for an additional 15 hours. Following this, 10 mL of deionized water was added, and the reaction mixture was stirred for 30 min. Then an additional 10 mL of methylene chloride was added. The phases were separated, and both phases were concentrated under reduced pressure until dry and analyzed by HPLC. Analysis of the HPLC traces revealed that the starting material was completely consumed, but no desired product was observed in either phase.

[0091] HPLC Method: Aminex HPX-87 H, 300*7.8 mm column. Temperature 30 ° C, Flowrate 0.5 mL / min, Rl Detection, Eluent 5 mM sulfuric acid in water. HPLC retention times: 2-oxo- 3-butenoic acid 17.3 min, PPO 9.7 min.

[0092] Sequences

[0093] SEQ I D NO:1 atgagcgatattactgtgacgaattgggcgggtaacatcacctatacggcgaaagaactgctgcgtccgcacagcctgg acgcgctgcgcgcattggttgcagacagcgcgcgtgtccgcgtgctgggtagcggccatagctttaacgaaattgcggagccgg gtgatggcggcgtgctgctgtccctggccggtttgccatccgttgtcgacgtcgatactgcggcccgtacggtgcgtgtcggtggc ggcgttcgctacgcagagctggctcgtgtcgttcacgcgcgtggtttagcgctgccgaacatggccagcctgccgcacatcagcg ttgcgggcagcgtggcgaccggtacgcatggcagcggtgttggtaatggctctctggcgagcgtagttagagaagtggaactggt gacggcagatggcagcaccgtcgttatcgctcgtggcgacgagcgttttggtggcgcggttacgtcgctgggtgccctgggtgttg tcaccagcttgaccctggacctggagccggcatacgaaatggaacaacatgtttttaccgaactgccgctggctggcctggatcc ggcgaccttcgaaacggttatggcagctgcgtactccgtgtctctgtttaccgattggcgcgcaccgggttttcgccaagtctggtt gaagcgtcgtaccgaccgccctctggacggcttcccgtatgcagcaccggcggcggagaaaatgcacccggtgccgggtatgc cagccgttaattgcactgagcaattcggcgtgccgggtccgtggcacgagcgcctgccgcatttccgtgccgagttcaccccgag cagcggtgctgaactccagtcagagtacctgatgccgcgtgagcacgcgctggcagcactgcacgcgatggacgcgatccgtg aaaccctggctcctgtgttgcagacctgtgagattcgcaccgtggctgccgacgcacagtggctgagcccggcttatggtcgtga taccgtagccgcacattttacgtgggtcgaggacaccgccgcggtgctgccggttgttcgccgcctggaagaagcactggtcccg ttcgcggcacgtccgcactggggtaaagtgttcacggtcccggcaggcgagttgcgtgcgctgtacccgcgtcttgcggatttcgg tgcactggccggtgccctggacccagcgggcaagtttaccaacgcgttcgtccgtggtgtgctggcgggt

[0094] SEQ I D NO:2

[0095] MSDITVTNWAGNITYTAKELLRPHSLDALRALVADSARVRVLGSGHSFN EIAEPGDGGVLL SLAGLPSVVDVDTAARTVRVGGGVRYAELARVVHARGLALPNMASLPH ISVAGSVATGTHGSGV GNGSLASVVREVELVTADGSTVVIARGDERFGGAVTSLGALGVVTSLTLDLEPAYEMEQHVFTEL PLAGLDPATFETVMAAAYSVSLFTDWRAPGFRQVWLKRRTDRPLDGFPYAAPAAEKM HPVPG MPAVNCTEQFGVPGPWHERLPHFRAEFTPSSGAELQSEYLMPREHALAALHAMDAI RETLAPV LQTCEI RTVAADAQWLSPAYGRDTVAAHFTWVEDTAAVLPVVRRLEEALVPFAARPHWGKVFTV PAGELRALYPRLADFGALAGALDPAGKFTNAFVRGVLAG

[0096] SEQ I D NO:3 atgatgggtcatccggaagaggttgatgtcattgtgtgtggcggcggtccggctggttgcgtagttgcgggccgtctggcg tacgcggaccccaccctcaaggtgatgctgatcgaaggcggggcgaataaccgcgatgacccctgggtttatcgtccgggaatt tatgtacgcaacatgcagcggaacggcatcaatgataaagcgacgttctacacagacaccatggcgtcgagttatttgcgcggt cggagaagcatagtcccgtgtgccaacatcctggggggtggtagctcaattaacttccagatgtacactcgggcgtcagcgagc gattgggatgatttcaagacggaagggtggacgtgtaaggacctgttgcctctgatgaagcgtctggaaaattaccagaaaccgt gcaataacgatacccacggctatgacggtccgattgctattagcaatggcggccagatcatgcctgtggcgcaggatttccttag agcagcacacgctatcggggttccatattccgatgatattcaggatctgaccaccgctcatggtgcggagatttgggccaaatat atcaatcgccataccggtcgtcgcagtgatgcagctactgcctatgtgcacagtgtgatggacgtgcaagataatttatttctgcgt tgcaacgcacgcgtgtcacgcgtcttgttcgatgataacaataaagccgtgggcgtcgcctatgttccaagccgtaatcgtaccc atggcggtaaattacatgagaccatagtaaaagcgcgtaaaatggttgttctgagctctggcaccctcggcacaccgcaaatcct agagcgctcgggagttggtaatggagaactgctgcgccaactcggtattaaaatcgtttcggatctgccaggcgtcggtgaacag tatcaggaccactacaccacgctgagcatctatcgcgtctccaacgaatctattaccaccgatgatttcctgcgaggggtgaaag acgtacagcgtgaactgtttaccgaatgggaagtaagcccggaaaaagcccgtttatcttcgaatgcaattgacgccggctttaa aattcgccctacggaggaggaactgaaagaaatgggcccggagttcaacgagctgtggaaccgttactttaaagataaacccg ataaaccggtgatgtttggctctattgtcgcgggcgcttacgcagaccacactcttctaccgccgggtaaatatattacgatgtttc agtatttagaatatccggcaagccgcggcaaaattcatatcaaatcccaaaacccatacgtggaaccgttctttgacagcgggtt tatgaacaataaggcggattttgccccgatccgctggagctacaagaaaaccagggaagtggcgcgacgcatggacgcatttc gtggagaattaacgtcccaccatccgcgttttcaccccgcaagtccggccgcttgcaaagacatcgatattgaaacagccaagc agatctacccggatggactgacggttggtatccatatgggctcgtggcatcagccttccgaaccgtacaaacatgataaggtgat cgaggacataccatacacggaggaagatgataaagcgattgacgattgggtcgcggaccacgtcgaaactacctggcatagtc tgggtacttgtgccatgaaaccgcgtgaacaaggtggagtagttgataaacgtcttaacgtgtatggtactcaaaatttgaaatgc gtggacctgtcaatctgcccggataacctgggcacaaacacgtattctagcgcgttgctcgtaggggaaaaaggggccgatttg attgcagaagagcttggcttaaaaattaaaaccccacacgcccctgttccacatgctccagtgcccacaggccgccctgcgacc caacaggtgcgcggatcctga

[0097] SEQ I D NO:4

[0098] MGH PEEVDVIVCGGGPAGCVVAGRLAYADPTLKVMLIEGGANNRDDPWVYRPGIYVRNM QRNGINDKATFYTDTMASSYLRGRRSIVPCAN ILGGGSSINFQMYTRASASDWDDFKTEGWTCK DLLPLMKRLENYQKPCNNDTHGYDGPIAISNGGQIM PVAQDFLRAAHAIGVPYSDDIQDLTTAH GAEIWAKYINRHTGRRSDAATAYVHSVMDVQDN LFLRCNARVSRVLFDDN NKAVGVAYVPSRN RTHGGKLH ETIVKARKMVVLSSGTLGTPQILERSGVGNGELLRQLGIKIVSDLPGVGEQYQDHYT TLSIYRVSN ESITTDDFLRGVKDVQRELFTEWEVSPEKARLSSNAIDAGFKI RPTEEELKEMGPEF NELWNRYFKDKPDKPVMFGSIVAGAYADHTLLPPGKYITMFQYLEYPASRGKIHIKSQNPYVEP FFDSGFMNN KADFAPI RWSYKKTREVARRMDAFRGELTSHHPRFHPASPAACKDI DIETAKQIY PDGLTVGIH MGSWHQPSEPYKH DKVI EDI PYTEEDDKAIDDWVADHVETTWHSLGTCAMKPRE QGGVVDKRLNVYGTQNLKCVDLSICPDNLGTNTYSSALLVGEKGADLIAEELGLKIKTPHAPVPH APVPTGRPATQQVR

[0099] SEQ I D NO:5 atgtcaacctctagctctgacccgtttttcaattttgccaaatccagcttccgctctgcggctgcacagaaagcgagtgcat ctagtctgccgccgttgcccgggccggataaaaaagttccggggatggatattaaatatgatgtggttattgtcggatctggtccg attggttgcacgtacgctcgggagcttgttggtgcgggctataaagtcgctatgtttgatataggcgagatcgactcgggcctgaa aatcggcgcgcataaaaaaaacaccgtggagtaccagaaaaacattgataaattcgttaacgtaattcagggccagctcatgtc cgtttcggtgcctgtgaacaccctggtcgtggacactctcagcccgacgtcgtggcaggccagcacgttctttgtacgcaacggct caaacccagagcaggacccactgcgtaatctgtcaggccaagcggtaactcgcgtggtcggaggcatgagcacccactggac atgcgcaacgccccgcttcgatcgggaacagcgtccgctccttgtcaaagacgacgctgatgccgacgacgcggaatgggatc gtttgtacaccaaagccgaatcatattttcaaaccggtacggatcaatttaaagagtccatccgtcataatctggttctgaataaac tgaccgaggaatacaaaggtcagcgtgattttcagcagatcccgctggctgcgacccgccgctcgcctacttttgtcgaatggag ttccgcgaatacggtgtttgatttacaaaatcgtccgaacacggacgctccggaagaaagatttaacttattcccggccgtcgcct gtgagagggtggtacgcaatgcgttgaatagcgaaatcgaatccttacacatccatgacctaattagcggggaccgattcgaaa ttaaggccgatgtttacgtcttgaccgccggtgcggttcataatacccaattactggttaacagtggttttgggcagctgggccgcc ccaatcctgcgaacccgcctgaactgttaccgagcctcggtagctatattacggaacagtctcttgtgttctgtcagacggtgatg agcacagaattgattgattcggtgaagtcggatatgaccattcgtgggacacctggcgagctaacttatagtgtcacgtatacgcc cggcgcgagtaccaacaaacacccggattggtggaacgaaaaagttaagaaccacatgatgcaacatcaggaagatccgttg cccataccgtttgaagacccagaaccacaggtaaccaccctgttccaacctagccatccgtggcacacccaaatccaccgtgat gccttctcatacggtgcggtgcagcagtcgattgattcccggctgattgtagactggcgtttctttggccgtactgaaccgaaagag gagaacaaactgtggttctcggataagattactgacgcctacaatatgccgcagccgacctttgattttcgctttccggcaggccg tactagcaaagaagcggaagatatgatgacagatatgtgcgttatgtccgccaaaatcggtggatttctgccaggaagcctgcc gcagtttatggagccaggcctggtgctgcacctgggtggtacccatcgaatgggctttgatgaaaaggaagataattgttgcgtg aacactgacagccgcgtgtttgggtttaaaaatctgttccttggcggttgcggtaacatcccaaccgcatacggagcgaacccta cgctgacagcaatgtcattagcgatcaagtcctgtgaatatatcaagcaaaatttcaccccaagtccgttcacatctgaagcaca a

[0100] SEQ I D NO:6

[0101] MSTSSSDPFFN FAKSSFRSAAAQKASASSLPPLPGPDKKVPGMDI KYDVVIVGSGPIGCTY ARELVGAGYKVAMFDIGEI DSGLKIGAHKKNTVEYQKNIDKFVNVIQGQLMSVSVPVNTLVVDTL SPTSWQASTFFVRNGSNPEQDPLRNLSGQAVTRVVGGMSTHWTCATPRFDREQRPLLVKDDA DADDAEWDRLYTKAESYFQTGTDQFKESI RH NLVLN KLTEEYKGQRDFQQIPLAATRRSPTFVE WSSANTVFDLQNRPNTDAPEERFNLFPAVACERVVRNALNSEIESLHIHDLISGDRFEI KADVYV LTAGAVHNTQLLVNSGFGQLGRPNPAN PPELLPSLGSYITEQSLVFCQTVMSTELI DSVKSDMTI RGTPGELTYSVTYTPGASTNKHPDWWNEKVKNHMMQHQEDPLPIPFEDPEPQVTTLFQPSHP WHTQI HRDAFSYGAVQQSIDSRLIVDWRFFGRTEPKEEN KLWFSDKITDAYNMPQPTFDFRFPA GRTSKEAEDMMTDMCVMSAKIGGFLPGSLPQFMEPGLVLHLGGTHRMGFDEKEDNCCVNTD SRVFGFKNLFLGGCGNI PTAYGAN PTLTAMSLAIKSCEYI KQNFTPSPFTSEAQ

[0102] SEQ I D NO:7 atgagcacgtcgagcagtgatccgttctataatttcgcgaaaacgagctttaaatcagcggcggctcagaaagcctcag ccacaagtctgccgcctctccccggccccgaccagaaagtaccgggtatggatattaaatatgacgttgttattgtcggcagcgg cccgatcggttgcacctacgcgcgcgaactggtggaagcgggttataaagtggcgatgttcgacattggcgaaattgactcagg gcttaagatcggtgcgcataagaagaacaccgtcgagtatcaaaaaaatattgacaaatttgtcaatgttattcagggccaattg atgtctgtgtctgtaccggtaaataaactggttgtggataccttaagtccgaccagttggcaagcgtccacgtttttcgttcgcaatg gctcaaacccggagcaggacccgcttcggaatctgagcggccaggcagttactcgcgtggtgggtggcatgtctacgcactgga cgtgtgcaaccccgcggtttgatagggagcagcgtccactgctggtcaaagatgatccggatgccgatgatgcgatttgggacc agctgtatacgaaagcagaaagctattttaagaccgggacagatcagtttaacgaatctatacgtcataaccttgttttgaacaaa ttagctgaagaatataaaggtcagcgtacctttcagcagattccattggcggccacccgtcgcaacccgacttttgtggaatggtc gagcgccaatactgtgtttgatctgcaaaaccgcccgaatatcgacgcacccgaagaacgcttcaatctttttcctgcggtggcgt gtgagagagtaatgcgtaacgcctcaaataccgctattgaatcactgcatatccgggatctgatttccggtgatcgctttgctatcc aggcagatgtctatgtgctgaccgcgggagcggttcacaacacccaattactggttaattccggttttgggaagctgggccgtccg gacccggccaaccccccagaattactgcctttcttaggctcctacattacagagcagtccctggtcttttgccagaccgtaatgtct accgaactgatcgattcggtgaagtcggacatgacgattatcggtaaccctggcgaactgggctactcggtgtcttacatgccgg gtgcttcgactaacaaacacccagattggtggaatgagaaagtccagaaccacatgatgcaacaccaagaagacccgttacct attccatttgaagatccagaaccgcaagtgaccacgttgtttcagccgagccatccttggcatacacagatccatcgagatgcctt tagctacggtgccgtacagcaaagcattgatagccgtctgatagtcgattggcgctttttcggccgtaccgagcccaaagaggag aataaattgtggttcagtgacaaaattacggacgcctacaacatgccgcagccgactttcgacttccgcttcccagcaggacgta ccagtcaagaagctgaagatatgatgacggacatgtgcgttatgtccgcgaaaatcggtggctttctgccggggtccctgccgca atttatggaaccggggctagtcctccacctgggaggaactcatcgcatggggttcgacgaacaggaagataactgttgcgtggat acagatagccgtgtgttcggtttcaacaatctcttcctcggtggatgtggcaacatccccactgcatacggcgccaacccaacctt gacggcaatgagtctggcaatcaaatcgtgcgagtatatcaaaaaaaattttacccctagcccgtttaccccagctcaatga

[0103] SEQ I D NO:8

[0104] MSTSSSDPFYNFAKTSFKSAAAQKASATSLPPLPGPDQKVPGMDI KYDVVIVGSGPIGCTY ARELVEAGYKVAM FDIGEIDSGLKIGAHKKNTVEYQKNIDKFVNVIQGQLMSVSVPVN KLVVDTL SPTSWQASTFFVRNGSNPEQDPLRNLSGQAVTRVVGGMSTHWTCATPRFDREQRPLLVKDDP DADDAIWDQLYTKAESYFKTGTDQFN ESIRHNLVLNKLAEEYKGQRTFQQI PLAATRRNPTFVE WSSANTVFDLQNRPNI DAPEERFNLFPAVACERVMRNASNTAIESLHIRDLISGDRFAIQADVYVL TAGAVH NTQLLVNSGFGKLGRPDPANPPELLPFLGSYITEQSLVFCQTVMSTELIDSVKSDMTII GN PGELGYSVSYMPGASTN KH PDWWN EKVQNH MMQHQEDPLPIPFEDPEPQVTTLFQPSHP WHTQI HRDAFSYGAVQQSIDSRLIVDWRFFGRTEPKEEN KLWFSDKITDAYNMPQPTFDFRFPA GRTSQEAEDMMTDMCVMSAKIGGFLPGSLPQFMEPGLVLHLGGTHRMGFDEQEDNCCVDTD SRVFGFNN LFLGGCGN IPTAYGANPTLTAMSLAIKSCEYI KKNFTPSPFTPAQ

[0105] SEQ I D NO:9 atggcatctgcacctattggtagcgccattcctcgcaacaactgggccgtcacttgcgacagtgcacagtcgggaaatg aatgcaacaaggccattgatggcaacaaggataccttttggcacacattctatggcgccaacggggatccaaagccccctcac acatacacgattgacatgaagacaactcagaacgtcaacggcttgtctgtgctgcctcgacaggatggtaaccaaaacggctgg atcggtcgccatgaggtttatctaagctcagatggcacaaactggggcagccctgttgcgtcaggtagttggttcgccgactctac tacaaaatactccaactttgaaactcgccctgctcgctatgttcgtcttgtcgctatcactgaagcgaatggccagccctggacta gcattgcagagatcaacgtcttccaagctagttcttacacagccccccagcctggtcttggacgctggggtccgactattgactta ccgattgttcctgcggctgcagcaattgaaccgacatcgggacgagtccttatgtggtcttcatatcgcaatgatgcatttgaagg atcccctggtggtatcactttgacgtcttcctgggatccatccactggtattgtttccgaccgcactgtgacagtcaccaagcatgat atgttctgccctggtatctccatggatggtaacggtcagatcgtagtcacaggtggcaacgatgccaagaagaccagtttgtatga ttcatctagcgatagctggatcccgggacctgacatgcaagtggctcgtgggtatcagtcatcagctaccatgtcagacggtcgtg tttttaccattggaggctccttcagcggtggcgtatttgagaagaatggcgaagtctatagcccatcttcaaagacatggacgtcc ctacccaatgccaaggtcaacccaatgttgacggctgacaagcaaggattgcttttttcagacaaccacgcgtggctctttggatg gaagaagggttcggtgttccaagcgggacctagcacagccatgaactggtactataccagtggaagtggtgatgtgaagtcagc cggaaaacgccagtctaaccgtggtgtagcccctgatgccatgtgcggaaacgctgtcatgtacgacgccgttaaaggaaagat cctgacctttggcggctccccagattatacagactctgacgccacaaccaacgcccacatcatcaccctcggtgaacccggaac atctcccaacactgtctttgctagcaatgggttgtactttgcccgaacgtttcacacctctgttgttcttccagacggaagcacgttta ttacaggaggccaacgacgtggaattccgttcgaggattcaaccccggtatttacacctgagatctacgtccctgaacaagaca ctttctacaagcagaaccccaactccattgttcgcgcctaccatagcatttcccttttgttacctgatggcagggtatttaacggtgg tggtggtctttgtggcgattgtaccacgaatcatttcgacgcgcaaatctttacgccaaactatctttacgatagcaacggcaatct cgcgacacgtcccaagattaccagaacctctacacagagcgtcaaggtcggtggcagaattacaatctcgacggattcttcgat tagcaaggcgtcgttgattcgctatggtacagcgacacacacggttaatactgaccagcgccgcattcccctgactctgacaaac aatggaggaaatagctattctttccaagttcctagcgactctggtgttgctttgcctggctactggatgttgttcgtgatgaactcggc cggtgttcctagtgtggcttcgacgattcgcgttactcag

[0106] SEQ ID NO:10

[0107] MASAPIGSAISRNNWAVTCDSAQSGNECNKAI DGNKDTFWHTFYGANGDPKPPHTYTIDM KTT QNVNGLSM LPRQDGNQNGWIGRH EVYLSSDGTNWGSPVASGSWFADSTTKYSN FETRPARYV RLVAITEANGQPWTSIAEINVFQASSYTAPQPGLGRWGPTIDLPIVPAAAAIEPTSGRVLMWSSYR NDAFGGSPGGITLTSSWDPSTGIVSDRTVTVTKHDMFCPGISMDGNGQIVVTGGN DAKKTSLY DSSSDSWIPGPDMQVARGYQSSATMSDGRVFTIGGSWSGGVFEKNGEVYSPSSKTWTSLPNA KVN PMLTADKQGLYRSDNHAWLFGWKKGSVFQAGPSTAMNWYYTSGSGDVKSAGKRQSNRG VAPDAMCGNAVMYDAVKGKILTFGGSPDYQDSDATTNAH IITLGEPGTSPNTVFASNGLYFART FHTSVVLPDGSTFITGGQRRGIPFEDSTPVFTPEIYVPEQDTFYKQNPNSIVRVYHSISLLLPDGR VFNGGGGLCGDCTTN HFDAQIFTPNYLYNSNGN LATRPKITRTSTQSVKVGGRITISTDSSISKAS LI RYGTATHTVNTDQRRIPLTLTNNGGNSYSFQVPSDSGVALPGYWMLFVMNSAGVPSVASTIR VTQ

Claims

Claims1. Process for preparing 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) of formula (I)comprising oxidizing in an oxidizing step a diol of formula (Ila):(Ha), and / or a hydroxymethyl ketone according to formula (lib):(lib).

2. The process according to claim 1, wherein the oxidizing step is carried out in the presence of a chemical catalyst or an enzyme.

3. The process according to claim 2, wherein the oxidizing step is carried out in the presence of an enzyme, wherein the enzyme is an Oxidoreductases ECI, preferably an Oxidoreductase ECI selected from the group consisting of EC1.1, EC1.2, and EC1.10, more preferably selected from the group consisting of EC1.1.1, EC1.1.3, EC1.2.1, and EC1.2.3.

4. The process according to claim 1, wherein the oxidizing step is carried out in the presence of a chemical catalyst, wherein the chemical catalyst is a platinum-group metal catalyst, wherein the oxidizing step is further carried out in the presence of a gas comprising molecular oxygen and an activator.

5. The process according to claim 4 wherein the activator comprises one or more compounds selected from the group consisting of lead in elemental form, bismuth in elemental form, lead in oxide form, bismuth in oxide form, lead in hydracid salt form, bismuth in hydracid salt form, lead in form of a salt of an inorganic oxyacid, bismuth in form of a salt of an inorganic oxyacid, lead in form of a salt of a transition-metal- comprising oxyacid, bismuth in form of a salt of a transition-metal-comprising oxyacid, lead in form of a salt of an organic aliphatic or aromatic acid, and bismuth in form of asalt of an organic aliphatic or aromatic acid, preferably is selected from lead in elemental form and bismuth in elemental form, most preferably is bismuth in elemental form.

6. The process according to any of the preceding claims 4 to 5, wherein the platinumgroup metal catalyst is selected from platinum and palladium.

7. The process according to any of the preceding claims 4 to 6, wherein the activator is incorporated into the platinum-group metal catalyst.

8. The process according to any of the preceding claims 4 to 7, wherein the platinumgroup metal catalyst is a supported catalyst, preferably a charcoal-supported catalyst.

9. The process according to any of the preceding claims 4 to 8, wherein the process is carried out at a temperature of from 10 to 100 ° C, preferably 25 to 85 ° C, and most preferably 45 to 65 ° C.

10. The process according to any of the preceding claims 1 to 8, wherein the oxidizing step is carried out in the presence of a Catalase enzyme.

11. process according to claim 10, wherein the process is carried out at a temperature of from 25 to 45 ° C, preferably 30 to 42 ° C, and most preferably 35 to 40 ° C.

12. The process according to any of the preceding claims 4 to 11, wherein the process is carried out with the gas comprising molecular oxygen under a pressure of 0.5-10 atmospheres.

13. The use of a mixture comprising 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I):and aditionally a diol according to formula (Ila):and / or a hydroxymethyl ketone according to formula (lib):for preparing glufosinate and / or glufosinate salts, more particularly glufosinate, glufosinate-sodium or glufosinate-ammonium.

14. A mixture comprising 2-oxo-4-(hydroxy(methyl)phosphinoyl)butyric acid (PPO) according to formula (I):(I), and aditionally a diol according to formula (Ila):(Ha), and / or a hydroxymethyl ketone according to formula (lib):(lib).

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