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Novel class of difluorovinylphosphonate analogues of PEP |
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Journal of the Chemical Society, Perkin Transactions 1,
Volume 1,
Issue 8,
1997,
Page 1249-1254
Aparecida M. Kawamoto,
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摘要:
J. Chem. Soc. Perkin Trans. 1 1997 1249 Novel class of difluorovinylphosphonate analogues of PEP Aparecida M. Kawamoto*,a and Malcolm M. Campbell b a Chemistry Division Space Activity Institute São Jose dos Campos SP Brazil b School of Chemistry University of Bath UK A new class of difluorovinylphosphonate analogues of PEP 4,4-difluoro-4-(diethoxyphosphinoyl)- 2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 (E)- and (Z)-4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and 4,4-difluoro-4-phosphono- 2-methylenebutanoic acid 16 have been synthesized. Methyl 3,3-difluoro-3-(diethoxyphosphinoyl)-2- hydroxy-2-methylpropionate 21 and the corresponding acid 3,3-difluoro-3-phosphono-2-hydroxy-2- methylpropionic acid 23 have also been synthesized.These compounds are designed to act as potential inhibitors in the shikimic acid pathway. The unique physiological and physical properties of organo- fluorine compounds make them attractive for use as medicinal products herbicides and polymers. Although the replacement of a phosphate functional group with a phosphonate moiety in biologically important molecules constitutes an attractive strategy for the design of nonhydrolysable substrate analogues as inhibitors or alternate substrate analogues and for enzymes that process naturally occurring phosphates it has been proposed that the corresponding 1,1-difluoroalkylphosphonate should be a superior replacement since this surrogate should more accurately mimic the steric and polar character of the phosphate function.1 Nieschalk et al.2 described the synthesis of monofluoro- and difluoro-methylenephosphonate analogues of sn-glycerol-3- phosphate as substrates for glycerol-3-phosphate dehydrogenase.The synthesis of fluorophosphonate derivatives of N9-benzylguanine as potent slow-binding multisubstrate analogue inhibitors of purine nucleoside phosphorylase has been described by Halazy et al.3 Small peptides containing the non-hydrolysable phosphotyrosyl mimetic difluorophosphonomethylphenylalanine (F2Pmp) have been shown to be extremely potent proteintyrosine phosphatase inhibitors with the fluorines increasing inhibitory potency 1000-fold relative to the unfluorinated species. Bien Ye et al.4 reported the synthesis of one such inhibitor [difluoro(4-hydroxy-2-naphthyl)methyl]phosphonic acid which is prepared in 12 steps from commercially available 1,3- dihydroxynaphthalene.Phosphoenol pyruvate (PEP) plays an important role in the shikimic acid pathway for the formation of 5-enolpyruvylshikimic- 3-phosphate (5-EPS-3-P) 5 which is enzymatically synthesized by the nucleophilic attack of the 5-OH of the shikimate 3-phosphate (S3P) on the C-2 position of PEP with the elimination of phosphate 5 (Fig. 1). The reaction proceeds through a tetrahedral intermediate 3 which has previously been isolated and characterized by Anderson et al.6 Structural mimics of this intermediate are indeed potent EPSPS inhibitors.7 The tetrahedral intermediate 3 although stable under alkaline conditions is hydrolysed readily at neutral pH although the configuration of the ketal carbon has not been elucidated.5 It also decomposes under acidic conditions to form pyruvate and S3P.6 Replacement of the C]O]P group in the phosphoenol pyruvate by C]CF2]P is one strategy used to stabilize the ketal phosphate structure of the tetrahedral intermediate 3 and gives some stable analogues that could be potential inhibitors of EPSP synthase.There are a variety of PEP analogues that have been examined as alternate substrates and/or inhibitors of 5-EPS-3-P synthase. Walker and Jones 8 reported the first evidence that (Z)- 3-fluoro-PEP 6 functions as a pseudo substrate for 5-EPS-3-P synthase producing in one step the unexpected monofluoro analogue 7 which remains tightly bound at the enzyme site. Philion et al.9 described the synthesis of disodium salt 8 which is an isopolar and isosteric analogue of PEP. According to the authors this analogue was envisioned to be a potential Michael acceptor which could bind irreversibly to an enzyme active site for which PEP is a substrate.PEP has also been tested as an inhibitor of prolidase by Radzicka and Wolfenden.10 They described the action of derivatives of phosphoenol pyruvic acid—fluorinated chlorinated or brominated—as strong competitive inhibitors of prolidase which is an enzyme present in microorganism and mammalian tissues where it is believed to catalyse terminal degradation of exogenous proteins. In humans a deficiency of prolidase results in a complex clinical syndrome involving mental retardation. Fig. 1 Proposed mechanism of 5-EPS-3-P synthase PO O CO2 – OH H PO CO2 – PO O CO2 – OH CO2 - OP PO O CO2 – OH •• CO2 - •• 1 O CO2 – OH 3 2 H + CO2 – 4 5 Pi 2– O3PO O CO2 – OH CO2 – OPO3 2– CH2F – O2C OPO3 2– H F HO P CO2 – +Na CH2 O Na+ –O F F 6 7 8 1250 J.Chem. Soc. Perkin Trans. 1 1997 Scheme 1 (EtO)2P CF2Br O (EtO)2P CF2C O H CHCO2H (EtO)2P CF2CH2CHBrCO2H O (EtO)2P CF2C O H CHCO2H (HO)2P CF2CH2 O C( CH2)CO2H (HO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2H (EtO)2P CF2ZnBr O ClCH CHCO2H CH2 C(Br)CO2H DBU CH2Cl2/RT H2C C(CH2Br)CO2H TMSI THF/RT TMSI RT 40% H2C C(CH2Br)CO2Me 50% 12.5% 11 CuBr/THF/RT Cul/THF/RT CuBr/THF/RT CuBr/THF/RT 83.2% 49% 20% 33% 10 12 Activated Zn0 60 °C 99% 13 15 (EtO)3P 17 14 16 18 (EtO)2P CF2ZnBr O 9 90% CF2Br2 Et2O/RT 11 RT = room temperature This work describes the synthesis of a new class of di- fluorovinylphosphonate analogues of PEP 4,4-difluoro-4- (diethoxyphosphinoyl)-2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)butanoate 17 (E)-4,4- difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and (Z)- 4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 and 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 16.Compounds 14 16 and 17 are expected to act as effective inhibitors of EPSP synthase and compound 15 is expected to act as a potential inhibitor of prolidase. Results and discussion The proposed routes to the targets compounds 12–18 are outlined in Scheme 1. Reaction with zinc dust of diethyl bromodifluoromethylphosphonate 10 prepared from triethyl phosphite 9 and dibromodifluoromethane 11 gave the stable [(diethoxyphosphinoyl) difluoromethyl]zinc bromide 11 12 which by four different routes afforded compounds 12–18 respectively.Reaction of 11 with 2-bromoacrylic acid gave the novel intermediate 2-bromo-4,4-difluoro-4-(diethoxyphosphinoyl)- butanoic acid 13 which was treated with DBU in dichloromethane to yield compound 14. The E isomer was formed and this might be the result of elimination from Newman conformation A rather than B (Fig. 2) although there are mechanisms for the conversion of the Z isomer into the E isomer. In the gauche conformation the three groups [Br CO2H and (EtO)2P(O)CF2] are near each other resulting in a steric strain and less stable conformation. The NOE experiments confirmed the E configuration of compound 14. Thus irradiation of 2a-H (d 6.40) had no effect on the 3b-H resonance. Furthermore irradiation at the Fig. 2 CO2H Br H H H CF2P(O)(OEt)2 CO2H Br H H (EtO)2P(O)CF2 H anti gauche A B 3b-H resonance had no effect on the 2a-H resonance.Since the NOE effect is only noticeable over short distances generally 2–4 Å it is clear that the two protons (2a and 3b) are in the transposition. Reaction of 11 with cis-3-chloroacrylic acid afforded compound 12 in the same Z configuration as the starting material. This configuration was established by a comparative NMR analysis of compounds 12 and 14. The chemical shifts and coupling constants for both compounds are given in Table 1. The b vinylic hydrogen signal for compound 12 was shifted downfield to d 6.9; a smaller coupling constant for compound 12 indicated a Z configuration. The NOE experiments were used to confirm the Z configuration of the compound 12. Irradiation of 2-H (d 6.37) increased the integration for 3-H (42%) whilst irradiation of 3-H (d 6.01) increased the integration for 2-H (42%).The NOE effect observed in compound 12 indicated a short distance 2–4 Å between 2-H and 3-H which confirmed their cis disposition. A proposed mechanism for the formation of this product is illustrated in Fig. 3.13 Fig. 3 Proposed mechanism for the preparation of compound 12 C C H Cl H CO2H Cu0 C C H CuI H CO2H Cl C C H R H CO2H C C H Cu H CO2H R – Zn +Br R R = (EtO)2(O)PCF2 – CuII RCuI Cu0 – RI RZnBr Table 1 2a Vinylic proton 3b Vinylic proton Chemical Coupling Chemical Coupling Compound shift constant (Hz) shift constant (Hz) E-14 Z-12 6.40 6.37 15.8 12.9 6.92 6.01 15.9 12.8 J. Chem. Soc. Perkin Trans. 1 1997 1251 Compounds 15 and 17 were prepared by the reaction of compound 11 with methyl 2-(bromomethyl)acrylate and methyl 2-(bromomethyl)acrylic acid respectively.It was thought that the complete hydrolysis of compounds 15 and 17 with TMSI would yield the corresponding difluorodihydroxybutanoic acid 16 via the tris(trimethylsilyl) phosphonate ester as intermediate. However the methyl ester 17 was not hydrolysed in this fashion and yielded compound 18 instead. The presence of the OMe group was evident from the NMR spectra where it gave rise to a singlet at d 3.63 in the 1H NMR spectrum and a singlet at d 53.55 in the 13C NMR spectrum. The organic zinc compound 11 is relatively stable (days to months) at room temperature and reacts with a wide variety of electrophiles in the presence of a catalytic amount of cuprous bromide to give the corresponding difluoroalkylphosphonates in good yields as reported earlier.12,14 However the unexpected low yields in some of the target compounds may be a result of a hydrolysis side reaction of 11 with the starting acid compounds leading to the diethyl difluoromethylphosphonate 20 as a byproduct.When methyl acrylate was treated with compound 11 in order to verify the reactivity of the ester with the organic zinc reagent there was no reaction even when the mixture was heated overnight in refluxing THF. Compound 21 another target compound because of its potential use in the shikimate pathway is outlined in Scheme 2. Diethyl (difluoromethyl)phosphonate 20 prepared from diethyl phosphite 19 and chlorodifluoromethane,15 reacted with methyl pyruvate to yield compound 21 which was hydrolysed to compound 23 at 60 8C in the presence of solvent (THF ether DCM) over a period of 7 days.It was shown that the hydrolysis of the methyl ester was incomplete even after 3 weeks under reflux. The presence of the OMe group was confirmed by 1H NMR analysis in D2O which showed one singlet at approximately d 3.87. Another important reaction the dehydration of compound 21 to give the corresponding olefin 22 which is an analogue of PEP were unsuccessful. Thus the method of Hofmann et al.16 for dehydration of a secondary alcohol led only to decomposition of alcohol 21. Conversion of the hydroxy group of 21 into a triflate or acetate followed by elimination was investigated. Treatment of the alcohol with trifluoromethanesulfonic anhydride gave the triflate but it failed to undergo elimination.Although Posner et al.17 used Woelm alumina at room temperature to effect high-yield dehydrosulfonation of both secondary cyclic and acyclic alcohols and primary sulfonate esters attempted use of basic alumina (Brockman I) for the desired elimination resulted only in recovery of starting material after several days. Other unsuccessful attempts to dehydrate the compound 21 to give the corresponding olefin 22 were as follows dehydration by using DAST following the method described by Blackburn and Kent;18 reaction of the OH group of compound 21 with acetyl chloride or mesyl choride and then elimination with a strong base (LDA NaH or DBU); dehydration of com- Scheme 2 (EtO)2PH (EtO)2P CF2H O C CO2Me O Me (EtO)2P CF2C(Me)CO2Me O OH (EtO)2P CF2 O C( CH2)CO2Me THF BuLi diisopropylamine – 78 °C 7 h Na CHF2Cl THF 0 °C (HO)2P CF2C(Me)CO2H O 20 19 80% OH 22 23 21 dehydration hydrolysis 99% pound 21 using Martin Sulfurane dehydrating agent bis[a,abis( trifluoromethyl) benzenemethanolato]diphenylsulfur.Experimental Melting points were determined on a commercially available apparatus (Electrothermal melting point apparatus) or Büchi 510 and are uncorrected. Elemental microanalysis was carried out using a Carlo Erba 1106 Elemental Analyser. Infrared spectra were recorded in the range of 4000–600 cm21 using a Perkin-Elmer 1600 FT-IR spectrophotometer and peaks are reported (nmax) in wavenumbers (cm21). Spectra of liquid samples were taken as Nujol mulls or in chloroform solution as indicated. 1H NMR Spectra were recorded on a JEOL GX FT-270 (270 MHz) spectrometer although where indicated a JEOL GX FT-400 (400 MHz) spectrometer was used.13C NMR Spectra were recorded on a JEOL GX FT-270 spectrometer operating at 67.8 MHz and using 90 and 135 DEPT pulse sequences to aid multiplicity determination. Chemical shifts (d) are expressed in ppm downfield from internal tetramethylsilane (SiMe4). Mass spectra were recorded using a VG Analytical 7070 E instrument with a VG 2000 data system. Electron ionisation (EI) was produced using an ionising potential of 70 eV. Chemical ionisation (CI) was employed using isobutane as the reagent gas although where indicated ammonia was also used. All general reagents and solvents were purified and dried when required using the methods described in D. D. Perrin W. L. F. Armarego and D. R. Perrin Purification of Laboratory Chemicals Pergamon Press Oxford 1980.Diethyl bromodifluoromethylphosphonate 10 This compound [dH(CDCl3) 1.4 (6 H t) and 4.3–4.4 (4 H m)] was prepared by the reaction in diethyl ether of triethyl phosphite and dibromodifluoromethane at room temperature.11 [(Diethoxyphosphinoyl)difluoromethyl]zinc bromide 11 This compound [dH(CDCl3) 1.4 (6 H t) and 4.2–4.3 (4 H m)] was prepared by the reaction in dry THF of 10 with acidwashed zinc powder at 60 8C. Diethyl (difluoromethyl)phosphonate 20 This compound [dH(CDCl3) 1.4 (6 H t) 4.2–4.3 (4 H m) and 5.9 (1 H td)] was prepared by the reaction in THF of diethyl phosphite with chlorodifluoromethane at 0 8C.15 Methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 To the solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1800 mg 5.41 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by methyl 2-(bromomethyl)acrylate (1000 mg 5.6 mmol) added dropwise at room temperature.The mixture was stirred overnight after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (3 7) as the eluent to give the title compound (757 mg 49%); RF 0.48 (light petroleum–ethyl acetate 1 1) (Found C 42.1; H 6.10. C10H17F2O5P requires C 42.0; H 6.0%); nmax(liquid film)/cm21 3502 2988 1725 (C]] O) 1634 (C]] CH2) 1274 (P]] O) and 1042 (OCH3); dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 3.09–3.25 (td CF2CH2 JH,F 19.64 JH,P 4.76) 3.79 (s OCH3) 4.29 (m CH3CH2OP) 5.89 (s vinylic H) and 6.47 (s vinylic H); dC(CDCl3) 1.62 (d CH3CH2OP JC,P 5.5) 34.89 (td CF2CH2 JC,F 21.15 JC,P 16.53) 52.02 (s OCH3) 64.42 (d CH3CH2OP JC,P 7.3) 118.79 (td CF2 JC,F 261.5 JC,P 216.7) 131.15 (s C]] CH2) and 166.45 (s C]] O); m/z (EI) 286 (M1 34%) 255 1252 J.Chem. Soc. Perkin Trans. 1 1997 (M1 2 OMe 25) 199 (58) and 109 (100); m/z (CI) 287 (MH1 100%). 4,4-Difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 15 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1200 mg 3.4 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by 2-bromomethylacrylic acid (600 mg 3.64 mmol). The mixture was stirred overnight at room temperature after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3).The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform– methanol (98.5 1.5) as eluent to give the title compound (769 mg 83.2%); RF 0.40 (CHCl3–MeOH 9 1) (Found C 39.5; H 5.6. C9H15F2O5P requires C 39.7; H 5.6%); nmax(liquid film)/ cm21 3498 (CO2H) 1725 (C]] O) 1634 (C]] CH2) and 1269 (P]] O); dH(CDCl3) 1.39 (t CH3CH2OP J 7.08) 3.17 (td CF2CH2 JH,F 19.5 JH,P 4.88) 4.25–4.23 (m CH3CH2OP) 5.98 (s vinylic H) and 6.6 (s vinylic H); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.6 (td CF2CH2 JC,F 29.2 JC,P 16.5) 64.8 (d CH3CH2OP JC,P 7.7) 118.1 (td CF2 JC,F 261.1 JC,P 217.1) 133.0 (s C]] CH2) and 170.3 (s C]] O); m/z (CI) 273 (MH1 100%); m/z (EI) 272 (M1 2%) 227 (M1 2 CO2H 12) 201 [M1 2 C(CH2)CO2H 13] 199 (50) and 109 (100).4,4-Difluoro-4-(phosphono)-2-methylenebutanoic acid 16 Compound 15 (1200 mg 4.4 mmol) in dry THF (100 cm3) was stirred with TMSI (2100 mg 10.5 mmol) under N2 at room temperature for 6 h. The excess of silylating reagent and ethyl iodide were removed in vacuo to give the bis(trimethylsilyl)- phosphonate esters which were dissolved in diethyl ether (30 cm3) and then treated with water (20 cm3) to give the title compound 16 (475 mg 50%); this was purified by column chromatography (CHCl3–MeOH 90 10); RF 0.33 (chloroform– methanol 1 1) mp 72 8C; nmax(D2O)/cm21 3424 2527 1700 (C]] O) 1630 (C]] C) and 1209 (P]] O); dH(D2O) 3.14 (dt CF2CH2 JH,F 20.4 JH,P 2.47) 5.89 (s vinylic H) and 6.34 (s vinylic H); dC(D2O) 35.65 [q CF2CH2C(]] CH2) JC,P 21.5 JC,F 36.9] 122.7 (td CF2 JC,P 204.9 JC,F 271.8) 132.9 (s C]] CH2) and 171.8 (C]] O); m/z (2ve FAB) 215 (MH2 35%) 197 (20) 177 (12) and 159 (10).4,4-Difluoro-4-(diethoxyphosphinoyl)-2-bromobutanoic acid 13 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1304 mg 4.03 mmol) in dry THF (3 cm3) was added a catalytic amount of cuprous iodide followed by 2-bromoacrylic acid (700 mg 4.64 mmol) dissolved in dry THF (3 cm3) added dropwise at room temperature. The mixture was stirred for 4 days after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (450 mg 33%); RF 0.46 (CHCl3– MeOH–AcOH 90 8 2) (Found C 28.6; H 4.3.C8H14- BrF2O5P requires C 28.3; H 4.2%); nmax(liquid film)/cm21 3459 (CO2H) 3057 2981 1739 (C]] O) 1596 1243 (P]] O) and 1174; dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 2.59–2.83 (m CF2CHH) 3.09–3.34 (m CF2CHH) 4.25–4.36 (m CH3CH2OP) and 4.55 (dd CH2CHBr J2,3b 4.39 J2,3a 9.28); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.9 (s CF2CH2- CHBr) 39.3 (dd CF2CH2CHBr JC,F 36.35 JC,P 19.85) 65.5 (d CH3CH2OP JC,P 8.9) 118.7 (td CF2 JC,P 219.2) and 171.6 (s C]] O); dF(CDCl3) 2112.1 (dddd JF,F 301.7 JF,P 105.2 J3b,F 25.4 J3a,F 12.7 1 F) and 2113.2 (dddd JF,F 301.7 JF,P 105.7 J3b,F 25.5 J3a,F 11.6 1 F); dP(CDCl3) 5.08 (t 1H decoupled JP,F 104; m 1H coupled JP,3a = JP,3b 4.03); m/z (CI) 339 341 (MH1 98%) 321 323 (M1 2 OH 20) and 293 295 (M1 2 CO2H 7).(E)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 To a solution of the butanoic acid 13 (490 mg 1.44 mmol) in dry CH2Cl2 (6 cm3) was added dropwise DBU (439 mg 2.88 mmol) at 0 8C. The solution was allowed to warm to room temperature after which it was stirred overnight. The solution was then acidified to pH 2.0 with KHSO4 (0.5 M) washed with brine and extracted with CH2Cl2. After work-up the product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the product as an amber liquid (43.6 mg 12.5%); RF 0.55 (CHCl3– MeOH–AcOH 90 8 2); nmax(liquid film)/cm21 3423 2917 (CO2H) 1722 (C]] O) 1641 (C]] C) and 1443 1260 (P]] O); dH(CDCl3) 1.38 (t CH3CH2OP J 7.15) 4.23–4.36 (m CH3CH2OP) 6.40 (dq CF2CH]] CH J2a,3b 15.8 J2a,F 5.31 J2a,P 2.57) 6.92 (dtd CF2CH]] CH J3b,2a 15.8 J3b,F 12.7 J3b,P 1.95) and 9.99 (br s CO2H); dC(CDCl3) 16.3 (d CH3CH2OP JC,P 5.5) 65.4 (d CH3CH2OP JC,P 6.6) 117.9 (td CF2 JC,F 260.0 JC,P 218.2) 127.9 (q CF2CH]] CH JC,P = JC,F 7.0) 136.3 (td CF2CH]] CH JC,P 13.2 JC,F 22.05) and 167.6 (s C]] O); m/z (CI) 259 (MH1 259.0547.C8H13O5F2P requires M 259.0547 100%) and 213 (M1 2 CO2H 3). (Z)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1191 mg 3.58 mmol) in dry THF (6 cm3) was added under N2 a catalytic amount of cuprous bromide followed by cis-3-chloroacrylic acid (382 mg 3.59 mmol).The mixture was stirred for 24 h at room temperature after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (924 mg 20%); RF 0.35 (CHCl3–MeOH–AcOH 90 8 2) (Found C 37.2; H 5.4. C8H13F2O5P requires C 37.2; H 5.1%); nmax(liquid film)/cm21 3417 2989 (CO2H) 2571 1731 (C]] O) 1657 (C]] C) 1620 1479 1396 and 1254 (P]] O); dH(CDCl3) 1.41 (t CH3CH2OP J 7.05) 4.29–4.40 (m CH3CH2OP) 6.01 (dtd CF2CH]] CH J3b,2a 12.8 J3b,F 12.7 J3b,P 1.94) and 6.37 (dq CF2CH]] CH J2a,3b 12.9 J2a,F 2.47 J2a,P 2.47); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 66.2 (d CH3CH2OP JC,P 6.6) 116.1 (td CF2 JC,F 261.7 JC,P 214.9) 126.9 (td CF2CH]] CH JC,P 13.6 JC,F 23.9) 129.9 (q CF2CH]] CH JC,P = JC,F 7.2) and 166.1 (s C]] O); m/z (CI) 259 (MH1 259.0547.C8H13O5F2P requires M 259.0547 100%) 241 (M1 2 OH 35) and 213 (M1 2 CO2H 20). Methyl [3,3-difluoro-3-(diethoxyphosphinoyl)-2-hydroxy-2- methyl]propionate 21 A solution of butyllithium (2.91 cm3 4.66 mmol) in hexane was added at 0 8C to a stirred solution of diisopropylamine (472 mg 4.66 mmol) in dry THF (10 cm3) and the mixture was stirred for 30 min. It was then cooled to 278 8C and treated with a solution of diethyl difluoromethylphosphonate (761 mg 4.05 mmol) in dry THF (10 cm3) pre-cooled to 278 8C added slowly. The mixture was then stirred for 1 h at 278 8C.Methyl pyruvate (623 mg 6.1 mmol) in dry THF (10 cm3) pre-cooled to 278 8C was added dropwise to the mixture which was then stirred at 278 8C for 6 h slowly warmed to room temperature and then stirred for an additional 2 h. The reaction mixture was then poured into dry diethyl ether (50 cm3) and washed with saturated aqueous NH4Cl (3 × 10 cm3). The organic layer was then dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (1 1) as eluent to give a colourless oil; RF 0.26 (light petroleum–ethyl acetate 1 1) (Found C 37.2; H 6.1. C9H17F2O6 requires C 37.2; J. Chem. Soc. Perkin Trans. 1 1997 1253 H 5.9%); nmax(liquid film)/cm21 3474 2990 1747 (C]] O) 1657 1265 (P]] O) 1168 1022 (OCH3); dH(CDCl3) 1.38 (t CH3CH2OP JH,H 7.1) 1.62 [t CF2C(OH)CH3 JH,F 1.47] 3.87 (s OCH3) 4.01 (s OH) and 4.29 (m CH3CH2OP); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 4.4) 19.1 (s CCH3) 53.5 (s OCH3) 64.8 (d CH3CH2OP JC,P 6.6) 117.7 (td CF2 JC,F 274.3 JC,P 207.1) and 171.9 (s C]] O); dF(CDCl3) 2115.1 (dd JF,F 306.9 JF,P 98.9 1 F) and 2118.4 (dd JF,F 306.9 JF,P 102.3 1 F); dP(CDCl3) 5.1 (t 1H decoupled JP,F 101.1; m 1H coupled JP,H 7.74); m/z (EI) 290 (M1 2%) 231 (M1 2 CO2Me 68) 187 (95) and 175 (100); m/z (CI) 291 (MH1 100%).3,3-Difluoro-3-phosphono-2-hydroxy-2-methylpropionic acid 23 The ester 21 (1006 mg 0.3466 mmol) was stirred with TMSI in excess (30 cm3) without solvent at room temperature for 2 days and then heated to 60 8C for 5 days. The excess of silylating reagent and ethyl iodide were removed under reduced pressure to give the trisilylated ester.This was dissolved in diethyl ether (50 cm3) and hydrolysed with water (3 × 20 cm3) to give a viscous brown product (76 mg 100%); nmax(D2O)/cm21 3416 2518 1724 (C]] O) 1451 1209 (P]] O) and 1084; dH(D2O) 1.44 (s CH3); dC(D2O) 19.0 (s CH3) 118.4 (td CF2 JC,F 269.9 JC,P 191.4) and 173.8 (s C]] O); m/z (1ve FAB) 221 (MH1 100%) 175 (M1 2 CO2H 65) 149 (20) and 91 (60); m/z (2ve FAB) 219 (MH2 218.9861. C4H7F2O6P requires MH2 218.9870 100%). Acknowledgements We thank CNPq Conselho Nacional de Desenvolvimento Cientifico e Tecnologico for the financial support of this work. References 1 S. F. Martin D. W. Dean and A. S. Wagman Tetrahedron Lett. 1992 33 1839. 2 J. Nieschalk A. S. Batsanov D. O’Hagan and J.A. K. Howard Tetrahedron 1996 52 165. 3 S. Halazy A. Ehrard A. Eggenspiller V. Berges-Gross and C. Danzin Tetrahedron 1996 52 177. 4 Bien Ye and T. R. Burke Jr. Tetrahedron 1996 52 9963. 5 M. M. Campbell M. Sainsbury and P. A. Searle Synthesis 1993 179. 6 K. S. Anderson J. A. Sikorski and K. A. Johnson Biochemistry 1988 27 1604. 7 P. D. Pansegrau K. S. Anderson T. Widlanski J. E. Ream R. D. Sammons J. A. Sikorski and J. R. Knowles Tetrahedron Lett. 1991 32 2589. 8 M. C. Walker and C. R. Jones J. Am. Chem. Soc. 1992 14 7601. 9 D. P. Philion and D. G. Cleary J. Org. Chem. 1992 57 2763. 10 A. Radzicka and R. Wolfenden Biochemistry 1991 30 4160. 11 D. J. Burton and R. M. Flynn J. Fluorine Chem. 1977 10 329. 12 D. J. Burton T. Ishiara and M. Murata Chem. Lett. 1982 755. 13 A. C.Cope and W. G. Dauben Org. React. 1992 41 135. 14 D. J. Burton and L. G. Sprague J. Org. Chem. 1989 54 613. 15 D. E. Bergstrom and P. W. Shum J. Org. Chem. 1988 53 3953. 16 R. V. Hofmann R. D. Bishop P. M. Fitch and R. Hardenstein J. Org. Chem. 1980 45 919. 17 G. H. Posner G. M. Gunia and K. A. Babiak J. Org. Chem. 1977 42 3173. 18 G. M. Blackburn and D. E. Kent J. Chem. Soc. Perkin Trans. 1 1986 913. Paper 6/03094G Received 2nd May 1996 Accepted 15th November 1996 © Copyright 1997 by the Royal Society of Chemistry J. Chem. Soc. Perkin Trans. 1 1997 1249 Novel class of difluorovinylphosphonate analogues of PEP Aparecida M. Kawamoto*,a and Malcolm M. Campbell b a Chemistry Division Space Activity Institute São Jose dos Campos SP Brazil b School of Chemistry University of Bath UK A new class of difluorovinylphosphonate analogues of PEP 4,4-difluoro-4-(diethoxyphosphinoyl)- 2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 (E)- and (Z)-4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and 4,4-difluoro-4-phosphono- 2-methylenebutanoic acid 16 have been synthesized.Methyl 3,3-difluoro-3-(diethoxyphosphinoyl)-2- hydroxy-2-methylpropionate 21 and the corresponding acid 3,3-difluoro-3-phosphono-2-hydroxy-2- methylpropionic acid 23 have also been synthesized. These compounds are designed to act as potential inhibitors in the shikimic acid pathway. The unique physiological and physical properties of organo- fluorine compounds make them attractive for use as medicinal products herbicides and polymers. Although the replacement of a phosphate functional group with a phosphonate moiety in biologically important molecules constitutes an attractive strategy for the design of nonhydrolysable substrate analogues as inhibitors or alternate substrate analogues and for enzymes that process naturally occurring phosphates it has been proposed that the corresponding 1,1-difluoroalkylphosphonate should be a superior replacement since this surrogate should more accurately mimic the steric and polar character of the phosphate function.1 Nieschalk et al.2 described the synthesis of monofluoro- and difluoro-methylenephosphonate analogues of sn-glycerol-3- phosphate as substrates for glycerol-3-phosphate dehydrogenase.The synthesis of fluorophosphonate derivatives of N9-benzylguanine as potent slow-binding multisubstrate analogue inhibitors of purine nucleoside phosphorylase has been described by Halazy et al.3 Small peptides containing the non-hydrolysable phosphotyrosyl mimetic difluorophosphonomethylphenylalanine (F2Pmp) have been shown to be extremely potent proteintyrosine phosphatase inhibitors with the fluorines increasing inhibitory potency 1000-fold relative to the unfluorinated species.Bien Ye et al.4 reported the synthesis of one such inhibitor [difluoro(4-hydroxy-2-naphthyl)methyl]phosphonic acid which is prepared in 12 steps from commercially available 1,3- dihydroxynaphthalene. Phosphoenol pyruvate (PEP) plays an important role in the shikimic acid pathway for the formation of 5-enolpyruvylshikimic- 3-phosphate (5-EPS-3-P) 5 which is enzymatically synthesized by the nucleophilic attack of the 5-OH of the shikimate 3-phosphate (S3P) on the C-2 position of PEP with the elimination of phosphate 5 (Fig.1). The reaction proceeds through a tetrahedral intermediate 3 which has previously been isolated and characterized by Anderson et al.6 Structural mimics of this intermediate are indeed potent EPSPS inhibitors.7 The tetrahedral intermediate 3 although stable under alkaline conditions is hydrolysed readily at neutral pH although the configuration of the ketal carbon has not been elucidated.5 It also decomposes under acidic conditions to form pyruvate and S3P.6 Replacement of the C]O]P group in the phosphoenol pyruvate by C]CF2]P is one strategy used to stabilize the ketal phosphate structure of the tetrahedral intermediate 3 and gives some stable analogues that could be potential inhibitors of EPSP synthase.There are a variety of PEP analogues that have been examined as alternate substrates and/or inhibitors of 5-EPS-3-P synthase. Walker and Jones 8 reported the first evidence that (Z)- 3-fluoro-PEP 6 functions as a pseudo substrate for 5-EPS-3-P synthase producing in one step the unexpected monofluoro analogue 7 which remains tightly bound at the enzyme site. Philion et al.9 described the synthesis of disodium salt 8 which is an isopolar and isosteric analogue of PEP. According to the authors this analogue was envisioned to be a potential Michael acceptor which could bind irreversibly to an enzyme active site for which PEP is a substrate. PEP has also been tested as an inhibitor of prolidase by Radzicka and Wolfenden.10 They described the action of derivatives of phosphoenol pyruvic acid—fluorinated chlorinated or brominated—as strong competitive inhibitors of prolidase which is an enzyme present in microorganism and mammalian tissues where it is believed to catalyse terminal degradation of exogenous proteins.In humans a deficiency of prolidase results in a complex clinical syndrome involving mental retardation. Fig. 1 Proposed mechanism of 5-EPS-3-P synthase PO O CO2 – OH H PO CO2 – PO O CO2 – OH CO2 - OP PO O CO2 – OH •• CO2 - •• 1 O CO2 – OH 3 2 H + CO2 – 4 5 Pi 2– O3PO O CO2 – OH CO2 – OPO3 2– CH2F – O2C OPO3 2– H F HO P CO2 – +Na CH2 O Na+ –O F F 6 7 8 1250 J. Chem. Soc. Perkin Trans. 1 1997 Scheme 1 (EtO)2P CF2Br O (EtO)2P CF2C O H CHCO2H (EtO)2P CF2CH2CHBrCO2H O (EtO)2P CF2C O H CHCO2H (HO)2P CF2CH2 O C( CH2)CO2H (HO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2H (EtO)2P CF2ZnBr O ClCH CHCO2H CH2 C(Br)CO2H DBU CH2Cl2/RT H2C C(CH2Br)CO2H TMSI THF/RT TMSI RT 40% H2C C(CH2Br)CO2Me 50% 12.5% 11 CuBr/THF/RT Cul/THF/RT CuBr/THF/RT CuBr/THF/RT 83.2% 49% 20% 33% 10 12 Activated Zn0 60 °C 99% 13 15 (EtO)3P 17 14 16 18 (EtO)2P CF2ZnBr O 9 90% CF2Br2 Et2O/RT 11 RT = room temperature This work describes the synthesis of a new class of di- fluorovinylphosphonate analogues of PEP 4,4-difluoro-4- (diethoxyphosphinoyl)-2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)butanoate 17 (E)-4,4- difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and (Z)- 4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 and 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 16.Compounds 14 16 and 17 are expected to act as effective inhibitors of EPSP synthase and compound 15 is expected to act as a potential inhibitor of prolidase. Results and discussion The proposed routes to the targets compounds 12–18 are outlined in Scheme 1. Reaction with zinc dust of diethyl bromodifluoromethylphosphonate 10 prepared from triethyl phosphite 9 and dibromodifluoromethane 11 gave the stable [(diethoxyphosphinoyl) difluoromethyl]zinc bromide 11 12 which by four different routes afforded compounds 12–18 respectively. Reaction of 11 with 2-bromoacrylic acid gave the novel intermediate 2-bromo-4,4-difluoro-4-(diethoxyphosphinoyl)- butanoic acid 13 which was treated with DBU in dichloromethane to yield compound 14. The E isomer was formed and this might be the result of elimination from Newman conformation A rather than B (Fig.2) although there are mechanisms for the conversion of the Z isomer into the E isomer. In the gauche conformation the three groups [Br CO2H and (EtO)2P(O)CF2] are near each other resulting in a steric strain and less stable conformation. The NOE experiments confirmed the E configuration of compound 14. Thus irradiation of 2a-H (d 6.40) had no effect on the 3b-H resonance. Furthermore irradiation at the Fig. 2 CO2H Br H H H CF2P(O)(OEt)2 CO2H Br H H (EtO)2P(O)CF2 H anti gauche A B 3b-H resonance had no effect on the 2a-H resonance. Since the NOE effect is only noticeable over short distances generally 2–4 Å it is clear that the two protons (2a and 3b) are in the transposition.Reaction of 11 with cis-3-chloroacrylic acid afforded compound 12 in the same Z configuration as the starting material. This configuration was established by a comparative NMR analysis of compounds 12 and 14. The chemical shifts and coupling constants for both compounds are given in Table 1. The b vinylic hydrogen signal for compound 12 was shifted downfield to d 6.9; a smaller coupling constant for compound 12 indicated a Z configuration. The NOE experiments were used to confirm the Z configuration of the compound 12. Irradiation of 2-H (d 6.37) increased the integration for 3-H (42%) whilst irradiation of 3-H (d 6.01) increased the integration for 2-H (42%). The NOE effect observed in compound 12 indicated a short distance 2–4 Å between 2-H and 3-H which confirmed their cis disposition.A proposed mechanism for the formation of this product is illustrated in Fig. 3.13 Fig. 3 Proposed mechanism for the preparation of compound 12 C C H Cl H CO2H Cu0 C C H CuI H CO2H Cl C C H R H CO2H C C H Cu H CO2H R – Zn +Br R R = (EtO)2(O)PCF2 – CuII RCuI Cu0 – RI RZnBr Table 1 2a Vinylic proton 3b Vinylic proton Chemical Coupling Chemical Coupling Compound shift constant (Hz) shift constant (Hz) E-14 Z-12 6.40 6.37 15.8 12.9 6.92 6.01 15.9 12.8 J. Chem. Soc. Perkin Trans. 1 1997 1251 Compounds 15 and 17 were prepared by the reaction of compound 11 with methyl 2-(bromomethyl)acrylate and methyl 2-(bromomethyl)acrylic acid respectively. It was thought that the complete hydrolysis of compounds 15 and 17 with TMSI would yield the corresponding difluorodihydroxybutanoic acid 16 via the tris(trimethylsilyl) phosphonate ester as intermediate.However the methyl ester 17 was not hydrolysed in this fashion and yielded compound 18 instead. The presence of the OMe group was evident from the NMR spectra where it gave rise to a singlet at d 3.63 in the 1H NMR spectrum and a singlet at d 53.55 in the 13C NMR spectrum. The organic zinc compound 11 is relatively stable (days to months) at room temperature and reacts with a wide variety of electrophiles in the presence of a catalytic amount of cuprous bromide to give the corresponding difluoroalkylphosphonates in good yields as reported earlier.12,14 However the unexpected low yields in some of the target compounds may be a result of a hydrolysis side reaction of 11 with the starting acid compounds leading to the diethyl difluoromethylphosphonate 20 as a byproduct.When methyl acrylate was treated with compound 11 in order to verify the reactivity of the ester with the organic zinc reagent there was no reaction even when the mixture was heated overnight in refluxing THF. Compound 21 another target compound because of its potential use in the shikimate pathway is outlined in Scheme 2. Diethyl (difluoromethyl)phosphonate 20 prepared from diethyl phosphite 19 and chlorodifluoromethane,15 reacted with methyl pyruvate to yield compound 21 which was hydrolysed to compound 23 at 60 8C in the presence of solvent (THF ether DCM) over a period of 7 days. It was shown that the hydrolysis of the methyl ester was incomplete even after 3 weeks under reflux. The presence of the OMe group was confirmed by 1H NMR analysis in D2O which showed one singlet at approximately d 3.87.Another important reaction the dehydration of compound 21 to give the corresponding olefin 22 which is an analogue of PEP were unsuccessful. Thus the method of Hofmann et al.16 for dehydration of a secondary alcohol led only to decomposition of alcohol 21. Conversion of the hydroxy group of 21 into a triflate or acetate followed by elimination was investigated. Treatment of the alcohol with trifluoromethanesulfonic anhydride gave the triflate but it failed to undergo elimination. Although Posner et al.17 used Woelm alumina at room temperature to effect high-yield dehydrosulfonation of both secondary cyclic and acyclic alcohols and primary sulfonate esters attempted use of basic alumina (Brockman I) for the desired elimination resulted only in recovery of starting material after several days.Other unsuccessful attempts to dehydrate the compound 21 to give the corresponding olefin 22 were as follows dehydration by using DAST following the method described by Blackburn and Kent;18 reaction of the OH group of compound 21 with acetyl chloride or mesyl choride and then elimination with a strong base (LDA NaH or DBU); dehydration of com- Scheme 2 (EtO)2PH (EtO)2P CF2H O C CO2Me O Me (EtO)2P CF2C(Me)CO2Me O OH (EtO)2P CF2 O C( CH2)CO2Me THF BuLi diisopropylamine – 78 °C 7 h Na CHF2Cl THF 0 °C (HO)2P CF2C(Me)CO2H O 20 19 80% OH 22 23 21 dehydration hydrolysis 99% pound 21 using Martin Sulfurane dehydrating agent bis[a,abis( trifluoromethyl) benzenemethanolato]diphenylsulfur. Experimental Melting points were determined on a commercially available apparatus (Electrothermal melting point apparatus) or Büchi 510 and are uncorrected.Elemental microanalysis was carried out using a Carlo Erba 1106 Elemental Analyser. Infrared spectra were recorded in the range of 4000–600 cm21 using a Perkin-Elmer 1600 FT-IR spectrophotometer and peaks are reported (nmax) in wavenumbers (cm21). Spectra of liquid samples were taken as Nujol mulls or in chloroform solution as indicated. 1H NMR Spectra were recorded on a JEOL GX FT-270 (270 MHz) spectrometer although where indicated a JEOL GX FT-400 (400 MHz) spectrometer was used. 13C NMR Spectra were recorded on a JEOL GX FT-270 spectrometer operating at 67.8 MHz and using 90 and 135 DEPT pulse sequences to aid multiplicity determination. Chemical shifts (d) are expressed in ppm downfield from internal tetramethylsilane (SiMe4).Mass spectra were recorded using a VG Analytical 7070 E instrument with a VG 2000 data system. Electron ionisation (EI) was produced using an ionising potential of 70 eV. Chemical ionisation (CI) was employed using isobutane as the reagent gas although where indicated ammonia was also used. All general reagents and solvents were purified and dried when required using the methods described in D. D. Perrin W. L. F. Armarego and D. R. Perrin Purification of Laboratory Chemicals Pergamon Press Oxford 1980. Diethyl bromodifluoromethylphosphonate 10 This compound [dH(CDCl3) 1.4 (6 H t) and 4.3–4.4 (4 H m)] was prepared by the reaction in diethyl ether of triethyl phosphite and dibromodifluoromethane at room temperature.11 [(Diethoxyphosphinoyl)difluoromethyl]zinc bromide 11 This compound [dH(CDCl3) 1.4 (6 H t) and 4.2–4.3 (4 H m)] was prepared by the reaction in dry THF of 10 with acidwashed zinc powder at 60 8C.Diethyl (difluoromethyl)phosphonate 20 This compound [dH(CDCl3) 1.4 (6 H t) 4.2–4.3 (4 H m) and 5.9 (1 H td)] was prepared by the reaction in THF of diethyl phosphite with chlorodifluoromethane at 0 8C.15 Methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 To the solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1800 mg 5.41 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by methyl 2-(bromomethyl)acrylate (1000 mg 5.6 mmol) added dropwise at room temperature. The mixture was stirred overnight after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3).The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (3 7) as the eluent to give the title compound (757 mg 49%); RF 0.48 (light petroleum–ethyl acetate 1 1) (Found C 42.1; H 6.10. C10H17F2O5P requires C 42.0; H 6.0%); nmax(liquid film)/cm21 3502 2988 1725 (C]] O) 1634 (C]] CH2) 1274 (P]] O) and 1042 (OCH3); dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 3.09–3.25 (td CF2CH2 JH,F 19.64 JH,P 4.76) 3.79 (s OCH3) 4.29 (m CH3CH2OP) 5.89 (s vinylic H) and 6.47 (s vinylic H); dC(CDCl3) 1.62 (d CH3CH2OP JC,P 5.5) 34.89 (td CF2CH2 JC,F 21.15 JC,P 16.53) 52.02 (s OCH3) 64.42 (d CH3CH2OP JC,P 7.3) 118.79 (td CF2 JC,F 261.5 JC,P 216.7) 131.15 (s C]] CH2) and 166.45 (s C]] O); m/z (EI) 286 (M1 34%) 255 1252 J.Chem. Soc. Perkin Trans. 1 1997 (M1 2 OMe 25) 199 (58) and 109 (100); m/z (CI) 287 (MH1 100%). 4,4-Difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 15 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1200 mg 3.4 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by 2-bromomethylacrylic acid (600 mg 3.64 mmol). The mixture was stirred overnight at room temperature after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform– methanol (98.5 1.5) as eluent to give the title compound (769 mg 83.2%); RF 0.40 (CHCl3–MeOH 9 1) (Found C 39.5; H 5.6.C9H15F2O5P requires C 39.7; H 5.6%); nmax(liquid film)/ cm21 3498 (CO2H) 1725 (C]] O) 1634 (C]] CH2) and 1269 (P]] O); dH(CDCl3) 1.39 (t CH3CH2OP J 7.08) 3.17 (td CF2CH2 JH,F 19.5 JH,P 4.88) 4.25–4.23 (m CH3CH2OP) 5.98 (s vinylic H) and 6.6 (s vinylic H); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.6 (td CF2CH2 JC,F 29.2 JC,P 16.5) 64.8 (d CH3CH2OP JC,P 7.7) 118.1 (td CF2 JC,F 261.1 JC,P 217.1) 133.0 (s C]] CH2) and 170.3 (s C]] O); m/z (CI) 273 (MH1 100%); m/z (EI) 272 (M1 2%) 227 (M1 2 CO2H 12) 201 [M1 2 C(CH2)CO2H 13] 199 (50) and 109 (100). 4,4-Difluoro-4-(phosphono)-2-methylenebutanoic acid 16 Compound 15 (1200 mg 4.4 mmol) in dry THF (100 cm3) was stirred with TMSI (2100 mg 10.5 mmol) under N2 at room temperature for 6 h.The excess of silylating reagent and ethyl iodide were removed in vacuo to give the bis(trimethylsilyl)- phosphonate esters which were dissolved in diethyl ether (30 cm3) and then treated with water (20 cm3) to give the title compound 16 (475 mg 50%); this was purified by column chromatography (CHCl3–MeOH 90 10); RF 0.33 (chloroform– methanol 1 1) mp 72 8C; nmax(D2O)/cm21 3424 2527 1700 (C]] O) 1630 (C]] C) and 1209 (P]] O); dH(D2O) 3.14 (dt CF2CH2 JH,F 20.4 JH,P 2.47) 5.89 (s vinylic H) and 6.34 (s vinylic H); dC(D2O) 35.65 [q CF2CH2C(]] CH2) JC,P 21.5 JC,F 36.9] 122.7 (td CF2 JC,P 204.9 JC,F 271.8) 132.9 (s C]] CH2) and 171.8 (C]] O); m/z (2ve FAB) 215 (MH2 35%) 197 (20) 177 (12) and 159 (10). 4,4-Difluoro-4-(diethoxyphosphinoyl)-2-bromobutanoic acid 13 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1304 mg 4.03 mmol) in dry THF (3 cm3) was added a catalytic amount of cuprous iodide followed by 2-bromoacrylic acid (700 mg 4.64 mmol) dissolved in dry THF (3 cm3) added dropwise at room temperature.The mixture was stirred for 4 days after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (450 mg 33%); RF 0.46 (CHCl3– MeOH–AcOH 90 8 2) (Found C 28.6; H 4.3. C8H14- BrF2O5P requires C 28.3; H 4.2%); nmax(liquid film)/cm21 3459 (CO2H) 3057 2981 1739 (C]] O) 1596 1243 (P]] O) and 1174; dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 2.59–2.83 (m CF2CHH) 3.09–3.34 (m CF2CHH) 4.25–4.36 (m CH3CH2OP) and 4.55 (dd CH2CHBr J2,3b 4.39 J2,3a 9.28); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.9 (s CF2CH2- CHBr) 39.3 (dd CF2CH2CHBr JC,F 36.35 JC,P 19.85) 65.5 (d CH3CH2OP JC,P 8.9) 118.7 (td CF2 JC,P 219.2) and 171.6 (s C]] O); dF(CDCl3) 2112.1 (dddd JF,F 301.7 JF,P 105.2 J3b,F 25.4 J3a,F 12.7 1 F) and 2113.2 (dddd JF,F 301.7 JF,P 105.7 J3b,F 25.5 J3a,F 11.6 1 F); dP(CDCl3) 5.08 (t 1H decoupled JP,F 104; m 1H coupled JP,3a = JP,3b 4.03); m/z (CI) 339 341 (MH1 98%) 321 323 (M1 2 OH 20) and 293 295 (M1 2 CO2H 7).(E)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 To a solution of the butanoic acid 13 (490 mg 1.44 mmol) in dry CH2Cl2 (6 cm3) was added dropwise DBU (439 mg 2.88 mmol) at 0 8C.The solution was allowed to warm to room temperature after which it was stirred overnight. The solution was then acidified to pH 2.0 with KHSO4 (0.5 M) washed with brine and extracted with CH2Cl2. After work-up the product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the product as an amber liquid (43.6 mg 12.5%); RF 0.55 (CHCl3– MeOH–AcOH 90 8 2); nmax(liquid film)/cm21 3423 2917 (CO2H) 1722 (C]] O) 1641 (C]] C) and 1443 1260 (P]] O); dH(CDCl3) 1.38 (t CH3CH2OP J 7.15) 4.23–4.36 (m CH3CH2OP) 6.40 (dq CF2CH]] CH J2a,3b 15.8 J2a,F 5.31 J2a,P 2.57) 6.92 (dtd CF2CH]] CH J3b,2a 15.8 J3b,F 12.7 J3b,P 1.95) and 9.99 (br s CO2H); dC(CDCl3) 16.3 (d CH3CH2OP JC,P 5.5) 65.4 (d CH3CH2OP JC,P 6.6) 117.9 (td CF2 JC,F 260.0 JC,P 218.2) 127.9 (q CF2CH]] CH JC,P = JC,F 7.0) 136.3 (td CF2CH]] CH JC,P 13.2 JC,F 22.05) and 167.6 (s C]] O); m/z (CI) 259 (MH1 259.0547.C8H13O5F2P requires M 259.0547 100%) and 213 (M1 2 CO2H 3). (Z)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1191 mg 3.58 mmol) in dry THF (6 cm3) was added under N2 a catalytic amount of cuprous bromide followed by cis-3-chloroacrylic acid (382 mg 3.59 mmol). The mixture was stirred for 24 h at room temperature after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo.The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (924 mg 20%); RF 0.35 (CHCl3–MeOH–AcOH 90 8 2) (Found C 37.2; H 5.4. C8H13F2O5P requires C 37.2; H 5.1%); nmax(liquid film)/cm21 3417 2989 (CO2H) 2571 1731 (C]] O) 1657 (C]] C) 1620 1479 1396 and 1254 (P]] O); dH(CDCl3) 1.41 (t CH3CH2OP J 7.05) 4.29–4.40 (m CH3CH2OP) 6.01 (dtd CF2CH]] CH J3b,2a 12.8 J3b,F 12.7 J3b,P 1.94) and 6.37 (dq CF2CH]] CH J2a,3b 12.9 J2a,F 2.47 J2a,P 2.47); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 66.2 (d CH3CH2OP JC,P 6.6) 116.1 (td CF2 JC,F 261.7 JC,P 214.9) 126.9 (td CF2CH]] CH JC,P 13.6 JC,F 23.9) 129.9 (q CF2CH]] CH JC,P = JC,F 7.2) and 166.1 (s C]] O); m/z (CI) 259 (MH1 259.0547. C8H13O5F2P requires M 259.0547 100%) 241 (M1 2 OH 35) and 213 (M1 2 CO2H 20).Methyl [3,3-difluoro-3-(diethoxyphosphinoyl)-2-hydroxy-2- methyl]propionate 21 A solution of butyllithium (2.91 cm3 4.66 mmol) in hexane was added at 0 8C to a stirred solution of diisopropylamine (472 mg 4.66 mmol) in dry THF (10 cm3) and the mixture was stirred for 30 min. It was then cooled to 278 8C and treated with a solution of diethyl difluoromethylphosphonate (761 mg 4.05 mmol) in dry THF (10 cm3) pre-cooled to 278 8C added slowly. The mixture was then stirred for 1 h at 278 8C. Methyl pyruvate (623 mg 6.1 mmol) in dry THF (10 cm3) pre-cooled to 278 8C was added dropwise to the mixture which was then stirred at 278 8C for 6 h slowly warmed to room temperature and then stirred for an additional 2 h. The reaction mixture was then poured into dry diethyl ether (50 cm3) and washed with saturated aqueous NH4Cl (3 × 10 cm3).The organic layer was then dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (1 1) as eluent to give a colourless oil; RF 0.26 (light petroleum–ethyl acetate 1 1) (Found C 37.2; H 6.1. C9H17F2O6 requires C 37.2; J. Chem. Soc. Perkin Trans. 1 1997 1253 H 5.9%); nmax(liquid film)/cm21 3474 2990 1747 (C]] O) 1657 1265 (P]] O) 1168 1022 (OCH3); dH(CDCl3) 1.38 (t CH3CH2OP JH,H 7.1) 1.62 [t CF2C(OH)CH3 JH,F 1.47] 3.87 (s OCH3) 4.01 (s OH) and 4.29 (m CH3CH2OP); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 4.4) 19.1 (s CCH3) 53.5 (s OCH3) 64.8 (d CH3CH2OP JC,P 6.6) 117.7 (td CF2 JC,F 274.3 JC,P 207.1) and 171.9 (s C]] O); dF(CDCl3) 2115.1 (dd JF,F 306.9 JF,P 98.9 1 F) and 2118.4 (dd JF,F 306.9 JF,P 102.3 1 F); dP(CDCl3) 5.1 (t 1H decoupled JP,F 101.1; m 1H coupled JP,H 7.74); m/z (EI) 290 (M1 2%) 231 (M1 2 CO2Me 68) 187 (95) and 175 (100); m/z (CI) 291 (MH1 100%).3,3-Difluoro-3-phosphono-2-hydroxy-2-methylpropionic acid 23 The ester 21 (1006 mg 0.3466 mmol) was stirred with TMSI in excess (30 cm3) without solvent at room temperature for 2 days and then heated to 60 8C for 5 days. The excess of silylating reagent and ethyl iodide were removed under reduced pressure to give the trisilylated ester. This was dissolved in diethyl ether (50 cm3) and hydrolysed with water (3 × 20 cm3) to give a viscous brown product (76 mg 100%); nmax(D2O)/cm21 3416 2518 1724 (C]] O) 1451 1209 (P]] O) and 1084; dH(D2O) 1.44 (s CH3); dC(D2O) 19.0 (s CH3) 118.4 (td CF2 JC,F 269.9 JC,P 191.4) and 173.8 (s C]] O); m/z (1ve FAB) 221 (MH1 100%) 175 (M1 2 CO2H 65) 149 (20) and 91 (60); m/z (2ve FAB) 219 (MH2 218.9861.C4H7F2O6P requires MH2 218.9870 100%). Acknowledgements We thank CNPq Conselho Nacional de Desenvolvimento Cientifico e Tecnologico for the financial support of this work. References 1 S. F. Martin D. W. Dean and A. S. Wagman Tetrahedron Lett. 1992 33 1839. 2 J. Nieschalk A. S. Batsanov D. O’Hagan and J. A. K. Howard Tetrahedron 1996 52 165. 3 S. Halazy A. Ehrard A. Eggenspiller V. Berges-Gross and C. Danzin Tetrahedron 1996 52 177. 4 Bien Ye and T. R. Burke Jr. Tetrahedron 1996 52 9963. 5 M. M. Campbell M. Sainsbury and P. A. Searle Synthesis 1993 179.6 K. S. Anderson J. A. Sikorski and K. A. Johnson Biochemistry 1988 27 1604. 7 P. D. Pansegrau K. S. Anderson T. Widlanski J. E. Ream R. D. Sammons J. A. Sikorski and J. R. Knowles Tetrahedron Lett. 1991 32 2589. 8 M. C. Walker and C. R. Jones J. Am. Chem. Soc. 1992 14 7601. 9 D. P. Philion and D. G. Cleary J. Org. Chem. 1992 57 2763. 10 A. Radzicka and R. Wolfenden Biochemistry 1991 30 4160. 11 D. J. Burton and R. M. Flynn J. Fluorine Chem. 1977 10 329. 12 D. J. Burton T. Ishiara and M. Murata Chem. Lett. 1982 755. 13 A. C. Cope and W. G. Dauben Org. React. 1992 41 135. 14 D. J. Burton and L. G. Sprague J. Org. Chem. 1989 54 613. 15 D. E. Bergstrom and P. W. Shum J. Org. Chem. 1988 53 3953. 16 R. V. Hofmann R. D. Bishop P. M. Fitch and R. Hardenstein J. Org. Chem. 1980 45 919.17 G. H. Posner G. M. Gunia and K. A. Babiak J. Org. Chem. 1977 42 3173. 18 G. M. Blackburn and D. E. Kent J. Chem. Soc. Perkin Trans. 1 1986 913. Paper 6/03094G Received 2nd May 1996 Accepted 15th November 1996 © Copyright 1997 by the Royal Society of Chemistry J. Chem. Soc. Perkin Trans. 1 1997 1249 Novel class of difluorovinylphosphonate analogues of PEP Aparecida M. Kawamoto*,a and Malcolm M. Campbell b a Chemistry Division Space Activity Institute São Jose dos Campos SP Brazil b School of Chemistry University of Bath UK A new class of difluorovinylphosphonate analogues of PEP 4,4-difluoro-4-(diethoxyphosphinoyl)- 2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 (E)- and (Z)-4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and 4,4-difluoro-4-phosphono- 2-methylenebutanoic acid 16 have been synthesized.Methyl 3,3-difluoro-3-(diethoxyphosphinoyl)-2- hydroxy-2-methylpropionate 21 and the corresponding acid 3,3-difluoro-3-phosphono-2-hydroxy-2- methylpropionic acid 23 have also been synthesized. These compounds are designed to act as potential inhibitors in the shikimic acid pathway. The unique physiological and physical properties of organo- fluorine compounds make them attractive for use as medicinal products herbicides and polymers. Although the replacement of a phosphate functional group with a phosphonate moiety in biologically important molecules constitutes an attractive strategy for the design of nonhydrolysable substrate analogues as inhibitors or alternate substrate analogues and for enzymes that process naturally occurring phosphates it has been proposed that the corresponding 1,1-difluoroalkylphosphonate should be a superior replacement since this surrogate should more accurately mimic the steric and polar character of the phosphate function.1 Nieschalk et al.2 described the synthesis of monofluoro- and difluoro-methylenephosphonate analogues of sn-glycerol-3- phosphate as substrates for glycerol-3-phosphate dehydrogenase.The synthesis of fluorophosphonate derivatives of N9-benzylguanine as potent slow-binding multisubstrate analogue inhibitors of purine nucleoside phosphorylase has been described by Halazy et al.3 Small peptides containing the non-hydrolysable phosphotyrosyl mimetic difluorophosphonomethylphenylalanine (F2Pmp) have been shown to be extremely potent proteintyrosine phosphatase inhibitors with the fluorines increasing inhibitory potency 1000-fold relative to the unfluorinated species.Bien Ye et al.4 reported the synthesis of one such inhibitor [difluoro(4-hydroxy-2-naphthyl)methyl]phosphonic acid which is prepared in 12 steps from commercially available 1,3- dihydroxynaphthalene. Phosphoenol pyruvate (PEP) plays an important role in the shikimic acid pathway for the formation of 5-enolpyruvylshikimic- 3-phosphate (5-EPS-3-P) 5 which is enzymatically synthesized by the nucleophilic attack of the 5-OH of the shikimate 3-phosphate (S3P) on the C-2 position of PEP with the elimination of phosphate 5 (Fig. 1). The reaction proceeds through a tetrahedral intermediate 3 which has previously been isolated and characterized by Anderson et al.6 Structural mimics of this intermediate are indeed potent EPSPS inhibitors.7 The tetrahedral intermediate 3 although stable under alkaline conditions is hydrolysed readily at neutral pH although the configuration of the ketal carbon has not been elucidated.5 It also decomposes under acidic conditions to form pyruvate and S3P.6 Replacement of the C]O]P group in the phosphoenol pyruvate by C]CF2]P is one strategy used to stabilize the ketal phosphate structure of the tetrahedral intermediate 3 and gives some stable analogues that could be potential inhibitors of EPSP synthase.There are a variety of PEP analogues that have been examined as alternate substrates and/or inhibitors of 5-EPS-3-P synthase. Walker and Jones 8 reported the first evidence that (Z)- 3-fluoro-PEP 6 functions as a pseudo substrate for 5-EPS-3-P synthase producing in one step the unexpected monofluoro analogue 7 which remains tightly bound at the enzyme site.Philion et al.9 described the synthesis of disodium salt 8 which is an isopolar and isosteric analogue of PEP. According to the authors this analogue was envisioned to be a potential Michael acceptor which could bind irreversibly to an enzyme active site for which PEP is a substrate. PEP has also been tested as an inhibitor of prolidase by Radzicka and Wolfenden.10 They described the action of derivatives of phosphoenol pyruvic acid—fluorinated chlorinated or brominated—as strong competitive inhibitors of prolidase which is an enzyme present in microorganism and mammalian tissues where it is believed to catalyse terminal degradation of exogenous proteins.In humans a deficiency of prolidase results in a complex clinical syndrome involving mental retardation. Fig. 1 Proposed mechanism of 5-EPS-3-P synthase PO O CO2 – OH H PO CO2 – PO O CO2 – OH CO2 - OP PO O CO2 – OH •• CO2 - •• 1 O CO2 – OH 3 2 H + CO2 – 4 5 Pi 2– O3PO O CO2 – OH CO2 – OPO3 2– CH2F – O2C OPO3 2– H F HO P CO2 – +Na CH2 O Na+ –O F F 6 7 8 1250 J. Chem. Soc. Perkin Trans. 1 1997 Scheme 1 (EtO)2P CF2Br O (EtO)2P CF2C O H CHCO2H (EtO)2P CF2CH2CHBrCO2H O (EtO)2P CF2C O H CHCO2H (HO)2P CF2CH2 O C( CH2)CO2H (HO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2H (EtO)2P CF2ZnBr O ClCH CHCO2H CH2 C(Br)CO2H DBU CH2Cl2/RT H2C C(CH2Br)CO2H TMSI THF/RT TMSI RT 40% H2C C(CH2Br)CO2Me 50% 12.5% 11 CuBr/THF/RT Cul/THF/RT CuBr/THF/RT CuBr/THF/RT 83.2% 49% 20% 33% 10 12 Activated Zn0 60 °C 99% 13 15 (EtO)3P 17 14 16 18 (EtO)2P CF2ZnBr O 9 90% CF2Br2 Et2O/RT 11 RT = room temperature This work describes the synthesis of a new class of di- fluorovinylphosphonate analogues of PEP 4,4-difluoro-4- (diethoxyphosphinoyl)-2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)butanoate 17 (E)-4,4- difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and (Z)- 4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 and 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 16.Compounds 14 16 and 17 are expected to act as effective inhibitors of EPSP synthase and compound 15 is expected to act as a potential inhibitor of prolidase. Results and discussion The proposed routes to the targets compounds 12–18 are outlined in Scheme 1.Reaction with zinc dust of diethyl bromodifluoromethylphosphonate 10 prepared from triethyl phosphite 9 and dibromodifluoromethane 11 gave the stable [(diethoxyphosphinoyl) difluoromethyl]zinc bromide 11 12 which by four different routes afforded compounds 12–18 respectively. Reaction of 11 with 2-bromoacrylic acid gave the novel intermediate 2-bromo-4,4-difluoro-4-(diethoxyphosphinoyl)- butanoic acid 13 which was treated with DBU in dichloromethane to yield compound 14. The E isomer was formed and this might be the result of elimination from Newman conformation A rather than B (Fig. 2) although there are mechanisms for the conversion of the Z isomer into the E isomer. In the gauche conformation the three groups [Br CO2H and (EtO)2P(O)CF2] are near each other resulting in a steric strain and less stable conformation.The NOE experiments confirmed the E configuration of compound 14. Thus irradiation of 2a-H (d 6.40) had no effect on the 3b-H resonance. Furthermore irradiation at the Fig. 2 CO2H Br H H H CF2P(O)(OEt)2 CO2H Br H H (EtO)2P(O)CF2 H anti gauche A B 3b-H resonance had no effect on the 2a-H resonance. Since the NOE effect is only noticeable over short distances generally 2–4 Å it is clear that the two protons (2a and 3b) are in the transposition. Reaction of 11 with cis-3-chloroacrylic acid afforded compound 12 in the same Z configuration as the starting material. This configuration was established by a comparative NMR analysis of compounds 12 and 14. The chemical shifts and coupling constants for both compounds are given in Table 1.The b vinylic hydrogen signal for compound 12 was shifted downfield to d 6.9; a smaller coupling constant for compound 12 indicated a Z configuration. The NOE experiments were used to confirm the Z configuration of the compound 12. Irradiation of 2-H (d 6.37) increased the integration for 3-H (42%) whilst irradiation of 3-H (d 6.01) increased the integration for 2-H (42%). The NOE effect observed in compound 12 indicated a short distance 2–4 Å between 2-H and 3-H which confirmed their cis disposition. A proposed mechanism for the formation of this product is illustrated in Fig. 3.13 Fig. 3 Proposed mechanism for the preparation of compound 12 C C H Cl H CO2H Cu0 C C H CuI H CO2H Cl C C H R H CO2H C C H Cu H CO2H R – Zn +Br R R = (EtO)2(O)PCF2 – CuII RCuI Cu0 – RI RZnBr Table 1 2a Vinylic proton 3b Vinylic proton Chemical Coupling Chemical Coupling Compound shift constant (Hz) shift constant (Hz) E-14 Z-12 6.40 6.37 15.8 12.9 6.92 6.01 15.9 12.8 J.Chem. Soc. Perkin Trans. 1 1997 1251 Compounds 15 and 17 were prepared by the reaction of compound 11 with methyl 2-(bromomethyl)acrylate and methyl 2-(bromomethyl)acrylic acid respectively. It was thought that the complete hydrolysis of compounds 15 and 17 with TMSI would yield the corresponding difluorodihydroxybutanoic acid 16 via the tris(trimethylsilyl) phosphonate ester as intermediate. However the methyl ester 17 was not hydrolysed in this fashion and yielded compound 18 instead. The presence of the OMe group was evident from the NMR spectra where it gave rise to a singlet at d 3.63 in the 1H NMR spectrum and a singlet at d 53.55 in the 13C NMR spectrum.The organic zinc compound 11 is relatively stable (days to months) at room temperature and reacts with a wide variety of electrophiles in the presence of a catalytic amount of cuprous bromide to give the corresponding difluoroalkylphosphonates in good yields as reported earlier.12,14 However the unexpected low yields in some of the target compounds may be a result of a hydrolysis side reaction of 11 with the starting acid compounds leading to the diethyl difluoromethylphosphonate 20 as a byproduct. When methyl acrylate was treated with compound 11 in order to verify the reactivity of the ester with the organic zinc reagent there was no reaction even when the mixture was heated overnight in refluxing THF.Compound 21 another target compound because of its potential use in the shikimate pathway is outlined in Scheme 2. Diethyl (difluoromethyl)phosphonate 20 prepared from diethyl phosphite 19 and chlorodifluoromethane,15 reacted with methyl pyruvate to yield compound 21 which was hydrolysed to compound 23 at 60 8C in the presence of solvent (THF ether DCM) over a period of 7 days. It was shown that the hydrolysis of the methyl ester was incomplete even after 3 weeks under reflux. The presence of the OMe group was confirmed by 1H NMR analysis in D2O which showed one singlet at approximately d 3.87. Another important reaction the dehydration of compound 21 to give the corresponding olefin 22 which is an analogue of PEP were unsuccessful. Thus the method of Hofmann et al.16 for dehydration of a secondary alcohol led only to decomposition of alcohol 21.Conversion of the hydroxy group of 21 into a triflate or acetate followed by elimination was investigated. Treatment of the alcohol with trifluoromethanesulfonic anhydride gave the triflate but it failed to undergo elimination. Although Posner et al.17 used Woelm alumina at room temperature to effect high-yield dehydrosulfonation of both secondary cyclic and acyclic alcohols and primary sulfonate esters attempted use of basic alumina (Brockman I) for the desired elimination resulted only in recovery of starting material after several days. Other unsuccessful attempts to dehydrate the compound 21 to give the corresponding olefin 22 were as follows dehydration by using DAST following the method described by Blackburn and Kent;18 reaction of the OH group of compound 21 with acetyl chloride or mesyl choride and then elimination with a strong base (LDA NaH or DBU); dehydration of com- Scheme 2 (EtO)2PH (EtO)2P CF2H O C CO2Me O Me (EtO)2P CF2C(Me)CO2Me O OH (EtO)2P CF2 O C( CH2)CO2Me THF BuLi diisopropylamine – 78 °C 7 h Na CHF2Cl THF 0 °C (HO)2P CF2C(Me)CO2H O 20 19 80% OH 22 23 21 dehydration hydrolysis 99% pound 21 using Martin Sulfurane dehydrating agent bis[a,abis( trifluoromethyl) benzenemethanolato]diphenylsulfur.Experimental Melting points were determined on a commercially available apparatus (Electrothermal melting point apparatus) or Büchi 510 and are uncorrected. Elemental microanalysis was carried out using a Carlo Erba 1106 Elemental Analyser. Infrared spectra were recorded in the range of 4000–600 cm21 using a Perkin-Elmer 1600 FT-IR spectrophotometer and peaks are reported (nmax) in wavenumbers (cm21).Spectra of liquid samples were taken as Nujol mulls or in chloroform solution as indicated. 1H NMR Spectra were recorded on a JEOL GX FT-270 (270 MHz) spectrometer although where indicated a JEOL GX FT-400 (400 MHz) spectrometer was used. 13C NMR Spectra were recorded on a JEOL GX FT-270 spectrometer operating at 67.8 MHz and using 90 and 135 DEPT pulse sequences to aid multiplicity determination. Chemical shifts (d) are expressed in ppm downfield from internal tetramethylsilane (SiMe4). Mass spectra were recorded using a VG Analytical 7070 E instrument with a VG 2000 data system. Electron ionisation (EI) was produced using an ionising potential of 70 eV.Chemical ionisation (CI) was employed using isobutane as the reagent gas although where indicated ammonia was also used. All general reagents and solvents were purified and dried when required using the methods described in D. D. Perrin W. L. F. Armarego and D. R. Perrin Purification of Laboratory Chemicals Pergamon Press Oxford 1980. Diethyl bromodifluoromethylphosphonate 10 This compound [dH(CDCl3) 1.4 (6 H t) and 4.3–4.4 (4 H m)] was prepared by the reaction in diethyl ether of triethyl phosphite and dibromodifluoromethane at room temperature.11 [(Diethoxyphosphinoyl)difluoromethyl]zinc bromide 11 This compound [dH(CDCl3) 1.4 (6 H t) and 4.2–4.3 (4 H m)] was prepared by the reaction in dry THF of 10 with acidwashed zinc powder at 60 8C. Diethyl (difluoromethyl)phosphonate 20 This compound [dH(CDCl3) 1.4 (6 H t) 4.2–4.3 (4 H m) and 5.9 (1 H td)] was prepared by the reaction in THF of diethyl phosphite with chlorodifluoromethane at 0 8C.15 Methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 To the solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1800 mg 5.41 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by methyl 2-(bromomethyl)acrylate (1000 mg 5.6 mmol) added dropwise at room temperature.The mixture was stirred overnight after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (3 7) as the eluent to give the title compound (757 mg 49%); RF 0.48 (light petroleum–ethyl acetate 1 1) (Found C 42.1; H 6.10.C10H17F2O5P requires C 42.0; H 6.0%); nmax(liquid film)/cm21 3502 2988 1725 (C]] O) 1634 (C]] CH2) 1274 (P]] O) and 1042 (OCH3); dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 3.09–3.25 (td CF2CH2 JH,F 19.64 JH,P 4.76) 3.79 (s OCH3) 4.29 (m CH3CH2OP) 5.89 (s vinylic H) and 6.47 (s vinylic H); dC(CDCl3) 1.62 (d CH3CH2OP JC,P 5.5) 34.89 (td CF2CH2 JC,F 21.15 JC,P 16.53) 52.02 (s OCH3) 64.42 (d CH3CH2OP JC,P 7.3) 118.79 (td CF2 JC,F 261.5 JC,P 216.7) 131.15 (s C]] CH2) and 166.45 (s C]] O); m/z (EI) 286 (M1 34%) 255 1252 J. Chem. Soc. Perkin Trans. 1 1997 (M1 2 OMe 25) 199 (58) and 109 (100); m/z (CI) 287 (MH1 100%). 4,4-Difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 15 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1200 mg 3.4 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by 2-bromomethylacrylic acid (600 mg 3.64 mmol).The mixture was stirred overnight at room temperature after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform– methanol (98.5 1.5) as eluent to give the title compound (769 mg 83.2%); RF 0.40 (CHCl3–MeOH 9 1) (Found C 39.5; H 5.6. C9H15F2O5P requires C 39.7; H 5.6%); nmax(liquid film)/ cm21 3498 (CO2H) 1725 (C]] O) 1634 (C]] CH2) and 1269 (P]] O); dH(CDCl3) 1.39 (t CH3CH2OP J 7.08) 3.17 (td CF2CH2 JH,F 19.5 JH,P 4.88) 4.25–4.23 (m CH3CH2OP) 5.98 (s vinylic H) and 6.6 (s vinylic H); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.6 (td CF2CH2 JC,F 29.2 JC,P 16.5) 64.8 (d CH3CH2OP JC,P 7.7) 118.1 (td CF2 JC,F 261.1 JC,P 217.1) 133.0 (s C]] CH2) and 170.3 (s C]] O); m/z (CI) 273 (MH1 100%); m/z (EI) 272 (M1 2%) 227 (M1 2 CO2H 12) 201 [M1 2 C(CH2)CO2H 13] 199 (50) and 109 (100).4,4-Difluoro-4-(phosphono)-2-methylenebutanoic acid 16 Compound 15 (1200 mg 4.4 mmol) in dry THF (100 cm3) was stirred with TMSI (2100 mg 10.5 mmol) under N2 at room temperature for 6 h. The excess of silylating reagent and ethyl iodide were removed in vacuo to give the bis(trimethylsilyl)- phosphonate esters which were dissolved in diethyl ether (30 cm3) and then treated with water (20 cm3) to give the title compound 16 (475 mg 50%); this was purified by column chromatography (CHCl3–MeOH 90 10); RF 0.33 (chloroform– methanol 1 1) mp 72 8C; nmax(D2O)/cm21 3424 2527 1700 (C]] O) 1630 (C]] C) and 1209 (P]] O); dH(D2O) 3.14 (dt CF2CH2 JH,F 20.4 JH,P 2.47) 5.89 (s vinylic H) and 6.34 (s vinylic H); dC(D2O) 35.65 [q CF2CH2C(]] CH2) JC,P 21.5 JC,F 36.9] 122.7 (td CF2 JC,P 204.9 JC,F 271.8) 132.9 (s C]] CH2) and 171.8 (C]] O); m/z (2ve FAB) 215 (MH2 35%) 197 (20) 177 (12) and 159 (10).4,4-Difluoro-4-(diethoxyphosphinoyl)-2-bromobutanoic acid 13 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1304 mg 4.03 mmol) in dry THF (3 cm3) was added a catalytic amount of cuprous iodide followed by 2-bromoacrylic acid (700 mg 4.64 mmol) dissolved in dry THF (3 cm3) added dropwise at room temperature.The mixture was stirred for 4 days after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (450 mg 33%); RF 0.46 (CHCl3– MeOH–AcOH 90 8 2) (Found C 28.6; H 4.3. C8H14- BrF2O5P requires C 28.3; H 4.2%); nmax(liquid film)/cm21 3459 (CO2H) 3057 2981 1739 (C]] O) 1596 1243 (P]] O) and 1174; dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 2.59–2.83 (m CF2CHH) 3.09–3.34 (m CF2CHH) 4.25–4.36 (m CH3CH2OP) and 4.55 (dd CH2CHBr J2,3b 4.39 J2,3a 9.28); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.9 (s CF2CH2- CHBr) 39.3 (dd CF2CH2CHBr JC,F 36.35 JC,P 19.85) 65.5 (d CH3CH2OP JC,P 8.9) 118.7 (td CF2 JC,P 219.2) and 171.6 (s C]] O); dF(CDCl3) 2112.1 (dddd JF,F 301.7 JF,P 105.2 J3b,F 25.4 J3a,F 12.7 1 F) and 2113.2 (dddd JF,F 301.7 JF,P 105.7 J3b,F 25.5 J3a,F 11.6 1 F); dP(CDCl3) 5.08 (t 1H decoupled JP,F 104; m 1H coupled JP,3a = JP,3b 4.03); m/z (CI) 339 341 (MH1 98%) 321 323 (M1 2 OH 20) and 293 295 (M1 2 CO2H 7).(E)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 To a solution of the butanoic acid 13 (490 mg 1.44 mmol) in dry CH2Cl2 (6 cm3) was added dropwise DBU (439 mg 2.88 mmol) at 0 8C. The solution was allowed to warm to room temperature after which it was stirred overnight. The solution was then acidified to pH 2.0 with KHSO4 (0.5 M) washed with brine and extracted with CH2Cl2.After work-up the product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the product as an amber liquid (43.6 mg 12.5%); RF 0.55 (CHCl3– MeOH–AcOH 90 8 2); nmax(liquid film)/cm21 3423 2917 (CO2H) 1722 (C]] O) 1641 (C]] C) and 1443 1260 (P]] O); dH(CDCl3) 1.38 (t CH3CH2OP J 7.15) 4.23–4.36 (m CH3CH2OP) 6.40 (dq CF2CH]] CH J2a,3b 15.8 J2a,F 5.31 J2a,P 2.57) 6.92 (dtd CF2CH]] CH J3b,2a 15.8 J3b,F 12.7 J3b,P 1.95) and 9.99 (br s CO2H); dC(CDCl3) 16.3 (d CH3CH2OP JC,P 5.5) 65.4 (d CH3CH2OP JC,P 6.6) 117.9 (td CF2 JC,F 260.0 JC,P 218.2) 127.9 (q CF2CH]] CH JC,P = JC,F 7.0) 136.3 (td CF2CH]] CH JC,P 13.2 JC,F 22.05) and 167.6 (s C]] O); m/z (CI) 259 (MH1 259.0547. C8H13O5F2P requires M 259.0547 100%) and 213 (M1 2 CO2H 3).(Z)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1191 mg 3.58 mmol) in dry THF (6 cm3) was added under N2 a catalytic amount of cuprous bromide followed by cis-3-chloroacrylic acid (382 mg 3.59 mmol). The mixture was stirred for 24 h at room temperature after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (924 mg 20%); RF 0.35 (CHCl3–MeOH–AcOH 90 8 2) (Found C 37.2; H 5.4. C8H13F2O5P requires C 37.2; H 5.1%); nmax(liquid film)/cm21 3417 2989 (CO2H) 2571 1731 (C]] O) 1657 (C]] C) 1620 1479 1396 and 1254 (P]] O); dH(CDCl3) 1.41 (t CH3CH2OP J 7.05) 4.29–4.40 (m CH3CH2OP) 6.01 (dtd CF2CH]] CH J3b,2a 12.8 J3b,F 12.7 J3b,P 1.94) and 6.37 (dq CF2CH]] CH J2a,3b 12.9 J2a,F 2.47 J2a,P 2.47); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 66.2 (d CH3CH2OP JC,P 6.6) 116.1 (td CF2 JC,F 261.7 JC,P 214.9) 126.9 (td CF2CH]] CH JC,P 13.6 JC,F 23.9) 129.9 (q CF2CH]] CH JC,P = JC,F 7.2) and 166.1 (s C]] O); m/z (CI) 259 (MH1 259.0547.C8H13O5F2P requires M 259.0547 100%) 241 (M1 2 OH 35) and 213 (M1 2 CO2H 20). Methyl [3,3-difluoro-3-(diethoxyphosphinoyl)-2-hydroxy-2- methyl]propionate 21 A solution of butyllithium (2.91 cm3 4.66 mmol) in hexane was added at 0 8C to a stirred solution of diisopropylamine (472 mg 4.66 mmol) in dry THF (10 cm3) and the mixture was stirred for 30 min.It was then cooled to 278 8C and treated with a solution of diethyl difluoromethylphosphonate (761 mg 4.05 mmol) in dry THF (10 cm3) pre-cooled to 278 8C added slowly. The mixture was then stirred for 1 h at 278 8C. Methyl pyruvate (623 mg 6.1 mmol) in dry THF (10 cm3) pre-cooled to 278 8C was added dropwise to the mixture which was then stirred at 278 8C for 6 h slowly warmed to room temperature and then stirred for an additional 2 h. The reaction mixture was then poured into dry diethyl ether (50 cm3) and washed with saturated aqueous NH4Cl (3 × 10 cm3). The organic layer was then dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (1 1) as eluent to give a colourless oil; RF 0.26 (light petroleum–ethyl acetate 1 1) (Found C 37.2; H 6.1.C9H17F2O6 requires C 37.2; J. Chem. Soc. Perkin Trans. 1 1997 1253 H 5.9%); nmax(liquid film)/cm21 3474 2990 1747 (C]] O) 1657 1265 (P]] O) 1168 1022 (OCH3); dH(CDCl3) 1.38 (t CH3CH2OP JH,H 7.1) 1.62 [t CF2C(OH)CH3 JH,F 1.47] 3.87 (s OCH3) 4.01 (s OH) and 4.29 (m CH3CH2OP); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 4.4) 19.1 (s CCH3) 53.5 (s OCH3) 64.8 (d CH3CH2OP JC,P 6.6) 117.7 (td CF2 JC,F 274.3 JC,P 207.1) and 171.9 (s C]] O); dF(CDCl3) 2115.1 (dd JF,F 306.9 JF,P 98.9 1 F) and 2118.4 (dd JF,F 306.9 JF,P 102.3 1 F); dP(CDCl3) 5.1 (t 1H decoupled JP,F 101.1; m 1H coupled JP,H 7.74); m/z (EI) 290 (M1 2%) 231 (M1 2 CO2Me 68) 187 (95) and 175 (100); m/z (CI) 291 (MH1 100%).3,3-Difluoro-3-phosphono-2-hydroxy-2-methylpropionic acid 23 The ester 21 (1006 mg 0.3466 mmol) was stirred with TMSI in excess (30 cm3) without solvent at room temperature for 2 days and then heated to 60 8C for 5 days. The excess of silylating reagent and ethyl iodide were removed under reduced pressure to give the trisilylated ester. This was dissolved in diethyl ether (50 cm3) and hydrolysed with water (3 × 20 cm3) to give a viscous brown product (76 mg 100%); nmax(D2O)/cm21 3416 2518 1724 (C]] O) 1451 1209 (P]] O) and 1084; dH(D2O) 1.44 (s CH3); dC(D2O) 19.0 (s CH3) 118.4 (td CF2 JC,F 269.9 JC,P 191.4) and 173.8 (s C]] O); m/z (1ve FAB) 221 (MH1 100%) 175 (M1 2 CO2H 65) 149 (20) and 91 (60); m/z (2ve FAB) 219 (MH2 218.9861. C4H7F2O6P requires MH2 218.9870 100%).Acknowledgements We thank CNPq Conselho Nacional de Desenvolvimento Cientifico e Tecnologico for the financial support of this work. References 1 S. F. Martin D. W. Dean and A. S. Wagman Tetrahedron Lett. 1992 33 1839. 2 J. Nieschalk A. S. Batsanov D. O’Hagan and J. A. K. Howard Tetrahedron 1996 52 165. 3 S. Halazy A. Ehrard A. Eggenspiller V. Berges-Gross and C. Danzin Tetrahedron 1996 52 177. 4 Bien Ye and T. R. Burke Jr. Tetrahedron 1996 52 9963. 5 M. M. Campbell M. Sainsbury and P. A. Searle Synthesis 1993 179. 6 K. S. Anderson J. A. Sikorski and K. A. Johnson Biochemistry 1988 27 1604. 7 P. D. Pansegrau K. S. Anderson T. Widlanski J. E. Ream R. D. Sammons J. A. Sikorski and J. R. Knowles Tetrahedron Lett. 1991 32 2589. 8 M. C. Walker and C. R. Jones J. Am.Chem. Soc. 1992 14 7601. 9 D. P. Philion and D. G. Cleary J. Org. Chem. 1992 57 2763. 10 A. Radzicka and R. Wolfenden Biochemistry 1991 30 4160. 11 D. J. Burton and R. M. Flynn J. Fluorine Chem. 1977 10 329. 12 D. J. Burton T. Ishiara and M. Murata Chem. Lett. 1982 755. 13 A. C. Cope and W. G. Dauben Org. React. 1992 41 135. 14 D. J. Burton and L. G. Sprague J. Org. Chem. 1989 54 613. 15 D. E. Bergstrom and P. W. Shum J. Org. Chem. 1988 53 3953. 16 R. V. Hofmann R. D. Bishop P. M. Fitch and R. Hardenstein J. Org. Chem. 1980 45 919. 17 G. H. Posner G. M. Gunia and K. A. Babiak J. Org. Chem. 1977 42 3173. 18 G. M. Blackburn and D. E. Kent J. Chem. Soc. Perkin Trans. 1 1986 913. Paper 6/03094G Received 2nd May 1996 Accepted 15th November 1996 © Copyright 1997 by the Royal Society of Chemistry J.Chem. Soc. Perkin Trans. 1 1997 1249 Novel class of difluorovinylphosphonate analogues of PEP Aparecida M. Kawamoto*,a and Malcolm M. Campbell b a Chemistry Division Space Activity Institute São Jose dos Campos SP Brazil b School of Chemistry University of Bath UK A new class of difluorovinylphosphonate analogues of PEP 4,4-difluoro-4-(diethoxyphosphinoyl)- 2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 (E)- and (Z)-4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and 4,4-difluoro-4-phosphono- 2-methylenebutanoic acid 16 have been synthesized. Methyl 3,3-difluoro-3-(diethoxyphosphinoyl)-2- hydroxy-2-methylpropionate 21 and the corresponding acid 3,3-difluoro-3-phosphono-2-hydroxy-2- methylpropionic acid 23 have also been synthesized.These compounds are designed to act as potential inhibitors in the shikimic acid pathway. The unique physiological and physical properties of organo- fluorine compounds make them attractive for use as medicinal products herbicides and polymers. Although the replacement of a phosphate functional group with a phosphonate moiety in biologically important molecules constitutes an attractive strategy for the design of nonhydrolysable substrate analogues as inhibitors or alternate substrate analogues and for enzymes that process naturally occurring phosphates it has been proposed that the corresponding 1,1-difluoroalkylphosphonate should be a superior replacement since this surrogate should more accurately mimic the steric and polar character of the phosphate function.1 Nieschalk et al.2 described the synthesis of monofluoro- and difluoro-methylenephosphonate analogues of sn-glycerol-3- phosphate as substrates for glycerol-3-phosphate dehydrogenase.The synthesis of fluorophosphonate derivatives of N9-benzylguanine as potent slow-binding multisubstrate analogue inhibitors of purine nucleoside phosphorylase has been described by Halazy et al.3 Small peptides containing the non-hydrolysable phosphotyrosyl mimetic difluorophosphonomethylphenylalanine (F2Pmp) have been shown to be extremely potent proteintyrosine phosphatase inhibitors with the fluorines increasing inhibitory potency 1000-fold relative to the unfluorinated species. Bien Ye et al.4 reported the synthesis of one such inhibitor [difluoro(4-hydroxy-2-naphthyl)methyl]phosphonic acid which is prepared in 12 steps from commercially available 1,3- dihydroxynaphthalene.Phosphoenol pyruvate (PEP) plays an important role in the shikimic acid pathway for the formation of 5-enolpyruvylshikimic- 3-phosphate (5-EPS-3-P) 5 which is enzymatically synthesized by the nucleophilic attack of the 5-OH of the shikimate 3-phosphate (S3P) on the C-2 position of PEP with the elimination of phosphate 5 (Fig. 1). The reaction proceeds through a tetrahedral intermediate 3 which has previously been isolated and characterized by Anderson et al.6 Structural mimics of this intermediate are indeed potent EPSPS inhibitors.7 The tetrahedral intermediate 3 although stable under alkaline conditions is hydrolysed readily at neutral pH although the configuration of the ketal carbon has not been elucidated.5 It also decomposes under acidic conditions to form pyruvate and S3P.6 Replacement of the C]O]P group in the phosphoenol pyruvate by C]CF2]P is one strategy used to stabilize the ketal phosphate structure of the tetrahedral intermediate 3 and gives some stable analogues that could be potential inhibitors of EPSP synthase.There are a variety of PEP analogues that have been examined as alternate substrates and/or inhibitors of 5-EPS-3-P synthase. Walker and Jones 8 reported the first evidence that (Z)- 3-fluoro-PEP 6 functions as a pseudo substrate for 5-EPS-3-P synthase producing in one step the unexpected monofluoro analogue 7 which remains tightly bound at the enzyme site. Philion et al.9 described the synthesis of disodium salt 8 which is an isopolar and isosteric analogue of PEP.According to the authors this analogue was envisioned to be a potential Michael acceptor which could bind irreversibly to an enzyme active site for which PEP is a substrate. PEP has also been tested as an inhibitor of prolidase by Radzicka and Wolfenden.10 They described the action of derivatives of phosphoenol pyruvic acid—fluorinated chlorinated or brominated—as strong competitive inhibitors of prolidase which is an enzyme present in microorganism and mammalian tissues where it is believed to catalyse terminal degradation of exogenous proteins. In humans a deficiency of prolidase results in a complex clinical syndrome involving mental retardation. Fig. 1 Proposed mechanism of 5-EPS-3-P synthase PO O CO2 – OH H PO CO2 – PO O CO2 – OH CO2 - OP PO O CO2 – OH •• CO2 - •• 1 O CO2 – OH 3 2 H + CO2 – 4 5 Pi 2– O3PO O CO2 – OH CO2 – OPO3 2– CH2F – O2C OPO3 2– H F HO P CO2 – +Na CH2 O Na+ –O F F 6 7 8 1250 J.Chem. Soc. Perkin Trans. 1 1997 Scheme 1 (EtO)2P CF2Br O (EtO)2P CF2C O H CHCO2H (EtO)2P CF2CH2CHBrCO2H O (EtO)2P CF2C O H CHCO2H (HO)2P CF2CH2 O C( CH2)CO2H (HO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2Me (EtO)2P CF2CH2 O C( CH2)CO2H (EtO)2P CF2ZnBr O ClCH CHCO2H CH2 C(Br)CO2H DBU CH2Cl2/RT H2C C(CH2Br)CO2H TMSI THF/RT TMSI RT 40% H2C C(CH2Br)CO2Me 50% 12.5% 11 CuBr/THF/RT Cul/THF/RT CuBr/THF/RT CuBr/THF/RT 83.2% 49% 20% 33% 10 12 Activated Zn0 60 °C 99% 13 15 (EtO)3P 17 14 16 18 (EtO)2P CF2ZnBr O 9 90% CF2Br2 Et2O/RT 11 RT = room temperature This work describes the synthesis of a new class of di- fluorovinylphosphonate analogues of PEP 4,4-difluoro-4- (diethoxyphosphinoyl)-2-methylenebutanoic acid 15 methyl 4,4-difluoro-4-(diethoxyphosphinoyl)butanoate 17 (E)-4,4- difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 and (Z)- 4,4-difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 and 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 16.Compounds 14 16 and 17 are expected to act as effective inhibitors of EPSP synthase and compound 15 is expected to act as a potential inhibitor of prolidase. Results and discussion The proposed routes to the targets compounds 12–18 are outlined in Scheme 1. Reaction with zinc dust of diethyl bromodifluoromethylphosphonate 10 prepared from triethyl phosphite 9 and dibromodifluoromethane 11 gave the stable [(diethoxyphosphinoyl) difluoromethyl]zinc bromide 11 12 which by four different routes afforded compounds 12–18 respectively.Reaction of 11 with 2-bromoacrylic acid gave the novel intermediate 2-bromo-4,4-difluoro-4-(diethoxyphosphinoyl)- butanoic acid 13 which was treated with DBU in dichloromethane to yield compound 14. The E isomer was formed and this might be the result of elimination from Newman conformation A rather than B (Fig. 2) although there are mechanisms for the conversion of the Z isomer into the E isomer. In the gauche conformation the three groups [Br CO2H and (EtO)2P(O)CF2] are near each other resulting in a steric strain and less stable conformation. The NOE experiments confirmed the E configuration of compound 14. Thus irradiation of 2a-H (d 6.40) had no effect on the 3b-H resonance. Furthermore irradiation at the Fig.2 CO2H Br H H H CF2P(O)(OEt)2 CO2H Br H H (EtO)2P(O)CF2 H anti gauche A B 3b-H resonance had no effect on the 2a-H resonance. Since the NOE effect is only noticeable over short distances generally 2–4 Å it is clear that the two protons (2a and 3b) are in the transposition. Reaction of 11 with cis-3-chloroacrylic acid afforded compound 12 in the same Z configuration as the starting material. This configuration was established by a comparative NMR analysis of compounds 12 and 14. The chemical shifts and coupling constants for both compounds are given in Table 1. The b vinylic hydrogen signal for compound 12 was shifted downfield to d 6.9; a smaller coupling constant for compound 12 indicated a Z configuration. The NOE experiments were used to confirm the Z configuration of the compound 12.Irradiation of 2-H (d 6.37) increased the integration for 3-H (42%) whilst irradiation of 3-H (d 6.01) increased the integration for 2-H (42%). The NOE effect observed in compound 12 indicated a short distance 2–4 Å between 2-H and 3-H which confirmed their cis disposition. A proposed mechanism for the formation of this product is illustrated in Fig. 3.13 Fig. 3 Proposed mechanism for the preparation of compound 12 C C H Cl H CO2H Cu0 C C H CuI H CO2H Cl C C H R H CO2H C C H Cu H CO2H R – Zn +Br R R = (EtO)2(O)PCF2 – CuII RCuI Cu0 – RI RZnBr Table 1 2a Vinylic proton 3b Vinylic proton Chemical Coupling Chemical Coupling Compound shift constant (Hz) shift constant (Hz) E-14 Z-12 6.40 6.37 15.8 12.9 6.92 6.01 15.9 12.8 J. Chem. Soc. Perkin Trans. 1 1997 1251 Compounds 15 and 17 were prepared by the reaction of compound 11 with methyl 2-(bromomethyl)acrylate and methyl 2-(bromomethyl)acrylic acid respectively.It was thought that the complete hydrolysis of compounds 15 and 17 with TMSI would yield the corresponding difluorodihydroxybutanoic acid 16 via the tris(trimethylsilyl) phosphonate ester as intermediate. However the methyl ester 17 was not hydrolysed in this fashion and yielded compound 18 instead. The presence of the OMe group was evident from the NMR spectra where it gave rise to a singlet at d 3.63 in the 1H NMR spectrum and a singlet at d 53.55 in the 13C NMR spectrum. The organic zinc compound 11 is relatively stable (days to months) at room temperature and reacts with a wide variety of electrophiles in the presence of a catalytic amount of cuprous bromide to give the corresponding difluoroalkylphosphonates in good yields as reported earlier.12,14 However the unexpected low yields in some of the target compounds may be a result of a hydrolysis side reaction of 11 with the starting acid compounds leading to the diethyl difluoromethylphosphonate 20 as a byproduct.When methyl acrylate was treated with compound 11 in order to verify the reactivity of the ester with the organic zinc reagent there was no reaction even when the mixture was heated overnight in refluxing THF. Compound 21 another target compound because of its potential use in the shikimate pathway is outlined in Scheme 2. Diethyl (difluoromethyl)phosphonate 20 prepared from diethyl phosphite 19 and chlorodifluoromethane,15 reacted with methyl pyruvate to yield compound 21 which was hydrolysed to compound 23 at 60 8C in the presence of solvent (THF ether DCM) over a period of 7 days.It was shown that the hydrolysis of the methyl ester was incomplete even after 3 weeks under reflux. The presence of the OMe group was confirmed by 1H NMR analysis in D2O which showed one singlet at approximately d 3.87. Another important reaction the dehydration of compound 21 to give the corresponding olefin 22 which is an analogue of PEP were unsuccessful. Thus the method of Hofmann et al.16 for dehydration of a secondary alcohol led only to decomposition of alcohol 21. Conversion of the hydroxy group of 21 into a triflate or acetate followed by elimination was investigated. Treatment of the alcohol with trifluoromethanesulfonic anhydride gave the triflate but it failed to undergo elimination.Although Posner et al.17 used Woelm alumina at room temperature to effect high-yield dehydrosulfonation of both secondary cyclic and acyclic alcohols and primary sulfonate esters attempted use of basic alumina (Brockman I) for the desired elimination resulted only in recovery of starting material after several days. Other unsuccessful attempts to dehydrate the compound 21 to give the corresponding olefin 22 were as follows dehydration by using DAST following the method described by Blackburn and Kent;18 reaction of the OH group of compound 21 with acetyl chloride or mesyl choride and then elimination with a strong base (LDA NaH or DBU); dehydration of com- Scheme 2 (EtO)2PH (EtO)2P CF2H O C CO2Me O Me (EtO)2P CF2C(Me)CO2Me O OH (EtO)2P CF2 O C( CH2)CO2Me THF BuLi diisopropylamine – 78 °C 7 h Na CHF2Cl THF 0 °C (HO)2P CF2C(Me)CO2H O 20 19 80% OH 22 23 21 dehydration hydrolysis 99% pound 21 using Martin Sulfurane dehydrating agent bis[a,abis( trifluoromethyl) benzenemethanolato]diphenylsulfur.Experimental Melting points were determined on a commercially available apparatus (Electrothermal melting point apparatus) or Büchi 510 and are uncorrected. Elemental microanalysis was carried out using a Carlo Erba 1106 Elemental Analyser. Infrared spectra were recorded in the range of 4000–600 cm21 using a Perkin-Elmer 1600 FT-IR spectrophotometer and peaks are reported (nmax) in wavenumbers (cm21). Spectra of liquid samples were taken as Nujol mulls or in chloroform solution as indicated. 1H NMR Spectra were recorded on a JEOL GX FT-270 (270 MHz) spectrometer although where indicated a JEOL GX FT-400 (400 MHz) spectrometer was used.13C NMR Spectra were recorded on a JEOL GX FT-270 spectrometer operating at 67.8 MHz and using 90 and 135 DEPT pulse sequences to aid multiplicity determination. Chemical shifts (d) are expressed in ppm downfield from internal tetramethylsilane (SiMe4). Mass spectra were recorded using a VG Analytical 7070 E instrument with a VG 2000 data system. Electron ionisation (EI) was produced using an ionising potential of 70 eV. Chemical ionisation (CI) was employed using isobutane as the reagent gas although where indicated ammonia was also used. All general reagents and solvents were purified and dried when required using the methods described in D. D. Perrin W. L. F. Armarego and D.R. Perrin Purification of Laboratory Chemicals Pergamon Press Oxford 1980. Diethyl bromodifluoromethylphosphonate 10 This compound [dH(CDCl3) 1.4 (6 H t) and 4.3–4.4 (4 H m)] was prepared by the reaction in diethyl ether of triethyl phosphite and dibromodifluoromethane at room temperature.11 [(Diethoxyphosphinoyl)difluoromethyl]zinc bromide 11 This compound [dH(CDCl3) 1.4 (6 H t) and 4.2–4.3 (4 H m)] was prepared by the reaction in dry THF of 10 with acidwashed zinc powder at 60 8C. Diethyl (difluoromethyl)phosphonate 20 This compound [dH(CDCl3) 1.4 (6 H t) 4.2–4.3 (4 H m) and 5.9 (1 H td)] was prepared by the reaction in THF of diethyl phosphite with chlorodifluoromethane at 0 8C.15 Methyl 4,4-difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoate 17 To the solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1800 mg 5.41 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by methyl 2-(bromomethyl)acrylate (1000 mg 5.6 mmol) added dropwise at room temperature.The mixture was stirred overnight after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (3 7) as the eluent to give the title compound (757 mg 49%); RF 0.48 (light petroleum–ethyl acetate 1 1) (Found C 42.1; H 6.10. C10H17F2O5P requires C 42.0; H 6.0%); nmax(liquid film)/cm21 3502 2988 1725 (C]] O) 1634 (C]] CH2) 1274 (P]] O) and 1042 (OCH3); dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 3.09–3.25 (td CF2CH2 JH,F 19.64 JH,P 4.76) 3.79 (s OCH3) 4.29 (m CH3CH2OP) 5.89 (s vinylic H) and 6.47 (s vinylic H); dC(CDCl3) 1.62 (d CH3CH2OP JC,P 5.5) 34.89 (td CF2CH2 JC,F 21.15 JC,P 16.53) 52.02 (s OCH3) 64.42 (d CH3CH2OP JC,P 7.3) 118.79 (td CF2 JC,F 261.5 JC,P 216.7) 131.15 (s C]] CH2) and 166.45 (s C]] O); m/z (EI) 286 (M1 34%) 255 1252 J.Chem. Soc. Perkin Trans. 1 1997 (M1 2 OMe 25) 199 (58) and 109 (100); m/z (CI) 287 (MH1 100%). 4,4-Difluoro-4-(diethoxyphosphinoyl)-2-methylenebutanoic acid 15 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1200 mg 3.4 mmol) in dry THF (6 cm3) was added a catalytic amount of cuprous bromide followed by 2-bromomethylacrylic acid (600 mg 3.64 mmol). The mixture was stirred overnight at room temperature after which it was filtered poured into water (10 cm3) and extracted with diethyl ether (3 × 10 cm3).The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform– methanol (98.5 1.5) as eluent to give the title compound (769 mg 83.2%); RF 0.40 (CHCl3–MeOH 9 1) (Found C 39.5; H 5.6. C9H15F2O5P requires C 39.7; H 5.6%); nmax(liquid film)/ cm21 3498 (CO2H) 1725 (C]] O) 1634 (C]] CH2) and 1269 (P]] O); dH(CDCl3) 1.39 (t CH3CH2OP J 7.08) 3.17 (td CF2CH2 JH,F 19.5 JH,P 4.88) 4.25–4.23 (m CH3CH2OP) 5.98 (s vinylic H) and 6.6 (s vinylic H); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.6 (td CF2CH2 JC,F 29.2 JC,P 16.5) 64.8 (d CH3CH2OP JC,P 7.7) 118.1 (td CF2 JC,F 261.1 JC,P 217.1) 133.0 (s C]] CH2) and 170.3 (s C]] O); m/z (CI) 273 (MH1 100%); m/z (EI) 272 (M1 2%) 227 (M1 2 CO2H 12) 201 [M1 2 C(CH2)CO2H 13] 199 (50) and 109 (100).4,4-Difluoro-4-(phosphono)-2-methylenebutanoic acid 16 Compound 15 (1200 mg 4.4 mmol) in dry THF (100 cm3) was stirred with TMSI (2100 mg 10.5 mmol) under N2 at room temperature for 6 h. The excess of silylating reagent and ethyl iodide were removed in vacuo to give the bis(trimethylsilyl)- phosphonate esters which were dissolved in diethyl ether (30 cm3) and then treated with water (20 cm3) to give the title compound 16 (475 mg 50%); this was purified by column chromatography (CHCl3–MeOH 90 10); RF 0.33 (chloroform– methanol 1 1) mp 72 8C; nmax(D2O)/cm21 3424 2527 1700 (C]] O) 1630 (C]] C) and 1209 (P]] O); dH(D2O) 3.14 (dt CF2CH2 JH,F 20.4 JH,P 2.47) 5.89 (s vinylic H) and 6.34 (s vinylic H); dC(D2O) 35.65 [q CF2CH2C(]] CH2) JC,P 21.5 JC,F 36.9] 122.7 (td CF2 JC,P 204.9 JC,F 271.8) 132.9 (s C]] CH2) and 171.8 (C]] O); m/z (2ve FAB) 215 (MH2 35%) 197 (20) 177 (12) and 159 (10).4,4-Difluoro-4-(diethoxyphosphinoyl)-2-bromobutanoic acid 13 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1304 mg 4.03 mmol) in dry THF (3 cm3) was added a catalytic amount of cuprous iodide followed by 2-bromoacrylic acid (700 mg 4.64 mmol) dissolved in dry THF (3 cm3) added dropwise at room temperature. The mixture was stirred for 4 days after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo.The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (450 mg 33%); RF 0.46 (CHCl3– MeOH–AcOH 90 8 2) (Found C 28.6; H 4.3. C8H14- BrF2O5P requires C 28.3; H 4.2%); nmax(liquid film)/cm21 3459 (CO2H) 3057 2981 1739 (C]] O) 1596 1243 (P]] O) and 1174; dH(CDCl3) 1.39 (t CH3CH2OP J 7.05) 2.59–2.83 (m CF2CHH) 3.09–3.34 (m CF2CHH) 4.25–4.36 (m CH3CH2OP) and 4.55 (dd CH2CHBr J2,3b 4.39 J2,3a 9.28); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 34.9 (s CF2CH2- CHBr) 39.3 (dd CF2CH2CHBr JC,F 36.35 JC,P 19.85) 65.5 (d CH3CH2OP JC,P 8.9) 118.7 (td CF2 JC,P 219.2) and 171.6 (s C]] O); dF(CDCl3) 2112.1 (dddd JF,F 301.7 JF,P 105.2 J3b,F 25.4 J3a,F 12.7 1 F) and 2113.2 (dddd JF,F 301.7 JF,P 105.7 J3b,F 25.5 J3a,F 11.6 1 F); dP(CDCl3) 5.08 (t 1H decoupled JP,F 104; m 1H coupled JP,3a = JP,3b 4.03); m/z (CI) 339 341 (MH1 98%) 321 323 (M1 2 OH 20) and 293 295 (M1 2 CO2H 7).(E)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 14 To a solution of the butanoic acid 13 (490 mg 1.44 mmol) in dry CH2Cl2 (6 cm3) was added dropwise DBU (439 mg 2.88 mmol) at 0 8C. The solution was allowed to warm to room temperature after which it was stirred overnight. The solution was then acidified to pH 2.0 with KHSO4 (0.5 M) washed with brine and extracted with CH2Cl2. After work-up the product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the product as an amber liquid (43.6 mg 12.5%); RF 0.55 (CHCl3– MeOH–AcOH 90 8 2); nmax(liquid film)/cm21 3423 2917 (CO2H) 1722 (C]] O) 1641 (C]] C) and 1443 1260 (P]] O); dH(CDCl3) 1.38 (t CH3CH2OP J 7.15) 4.23–4.36 (m CH3CH2OP) 6.40 (dq CF2CH]] CH J2a,3b 15.8 J2a,F 5.31 J2a,P 2.57) 6.92 (dtd CF2CH]] CH J3b,2a 15.8 J3b,F 12.7 J3b,P 1.95) and 9.99 (br s CO2H); dC(CDCl3) 16.3 (d CH3CH2OP JC,P 5.5) 65.4 (d CH3CH2OP JC,P 6.6) 117.9 (td CF2 JC,F 260.0 JC,P 218.2) 127.9 (q CF2CH]] CH JC,P = JC,F 7.0) 136.3 (td CF2CH]] CH JC,P 13.2 JC,F 22.05) and 167.6 (s C]] O); m/z (CI) 259 (MH1 259.0547.C8H13O5F2P requires M 259.0547 100%) and 213 (M1 2 CO2H 3). (Z)-4,4-Difluoro-4-(diethoxyphosphinoyl)but-2-enoic acid 12 To a solution of [(diethoxyphosphinoyl)difluoromethyl]zinc bromide (1191 mg 3.58 mmol) in dry THF (6 cm3) was added under N2 a catalytic amount of cuprous bromide followed by cis-3-chloroacrylic acid (382 mg 3.59 mmol).The mixture was stirred for 24 h at room temperature after which it was filtered poured into brine (10 cm3) and extracted with diethyl ether (3 × 10 cm3). The combined extracts were dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with chloroform–methanol–acetic acid (95 4 1) as eluent to give the title compound (924 mg 20%); RF 0.35 (CHCl3–MeOH–AcOH 90 8 2) (Found C 37.2; H 5.4. C8H13F2O5P requires C 37.2; H 5.1%); nmax(liquid film)/cm21 3417 2989 (CO2H) 2571 1731 (C]] O) 1657 (C]] C) 1620 1479 1396 and 1254 (P]] O); dH(CDCl3) 1.41 (t CH3CH2OP J 7.05) 4.29–4.40 (m CH3CH2OP) 6.01 (dtd CF2CH]] CH J3b,2a 12.8 J3b,F 12.7 J3b,P 1.94) and 6.37 (dq CF2CH]] CH J2a,3b 12.9 J2a,F 2.47 J2a,P 2.47); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 5.5) 66.2 (d CH3CH2OP JC,P 6.6) 116.1 (td CF2 JC,F 261.7 JC,P 214.9) 126.9 (td CF2CH]] CH JC,P 13.6 JC,F 23.9) 129.9 (q CF2CH]] CH JC,P = JC,F 7.2) and 166.1 (s C]] O); m/z (CI) 259 (MH1 259.0547.C8H13O5F2P requires M 259.0547 100%) 241 (M1 2 OH 35) and 213 (M1 2 CO2H 20). Methyl [3,3-difluoro-3-(diethoxyphosphinoyl)-2-hydroxy-2- methyl]propionate 21 A solution of butyllithium (2.91 cm3 4.66 mmol) in hexane was added at 0 8C to a stirred solution of diisopropylamine (472 mg 4.66 mmol) in dry THF (10 cm3) and the mixture was stirred for 30 min. It was then cooled to 278 8C and treated with a solution of diethyl difluoromethylphosphonate (761 mg 4.05 mmol) in dry THF (10 cm3) pre-cooled to 278 8C added slowly. The mixture was then stirred for 1 h at 278 8C.Methyl pyruvate (623 mg 6.1 mmol) in dry THF (10 cm3) pre-cooled to 278 8C was added dropwise to the mixture which was then stirred at 278 8C for 6 h slowly warmed to room temperature and then stirred for an additional 2 h. The reaction mixture was then poured into dry diethyl ether (50 cm3) and washed with saturated aqueous NH4Cl (3 × 10 cm3). The organic layer was then dried (Na2SO4) and concentrated in vacuo. The product was purified using column chromatography on silica gel with ethyl acetate–light petroleum (bp 60–80 8C) (1 1) as eluent to give a colourless oil; RF 0.26 (light petroleum–ethyl acetate 1 1) (Found C 37.2; H 6.1. C9H17F2O6 requires C 37.2; J. Chem. Soc. Perkin Trans. 1 1997 1253 H 5.9%); nmax(liquid film)/cm21 3474 2990 1747 (C]] O) 1657 1265 (P]] O) 1168 1022 (OCH3); dH(CDCl3) 1.38 (t CH3CH2OP JH,H 7.1) 1.62 [t CF2C(OH)CH3 JH,F 1.47] 3.87 (s OCH3) 4.01 (s OH) and 4.29 (m CH3CH2OP); dC(CDCl3) 16.2 (d CH3CH2OP JC,P 4.4) 19.1 (s CCH3) 53.5 (s OCH3) 64.8 (d CH3CH2OP JC,P 6.6) 117.7 (td CF2 JC,F 274.3 JC,P 207.1) and 171.9 (s C]] O); dF(CDCl3) 2115.1 (dd JF,F 306.9 JF,P 98.9 1 F) and 2118.4 (dd JF,F 306.9 JF,P 102.3 1 F); dP(CDCl3) 5.1 (t 1H decoupled JP,F 101.1; m 1H coupled JP,H 7.74); m/z (EI) 290 (M1 2%) 231 (M1 2 CO2Me 68) 187 (95) and 175 (100); m/z (CI) 291 (MH1 100%).3,3-Difluoro-3-phosphono-2-hydroxy-2-methylpropionic acid 23 The ester 21 (1006 mg 0.3466 mmol) was stirred with TMSI in excess (30 cm3) without solvent at room temperature for 2 days and then heated to 60 8C for 5 days.The excess of silylating reagent and ethyl iodide were removed under reduced pressure to give the trisilylated ester. This was dissolved in diethyl ether (50 cm3) and hydrolysed with water (3 × 20 cm3) to give a viscous brown product (76 mg 100%); nmax(D2O)/cm21 3416 2518 1724 (C]] O) 1451 1209 (P]] O) and 1084; dH(D2O) 1.44 (s CH3); dC(D2O) 19.0 (s CH3) 118.4 (td CF2 JC,F 269.9 JC,P 191.4) and 173.8 (s C]] O); m/z (1ve FAB) 221 (MH1 100%) 175 (M1 2 CO2H 65) 149 (20) and 91 (60); m/z (2ve FAB) 219 (MH2 218.9861. C4H7F2O6P requires MH2 218.9870 100%). Acknowledgements We thank CNPq Conselho Nacional de Desenvolvimento Cientifico e Tecnologico for the financial support of this work. References 1 S. F. Martin D. W. Dean and A. S. Wagman Tetrahedron Lett. 1992 33 1839. 2 J. Nieschalk A.S. Batsanov D. O’Hagan and J. A. K. Howard Tetrahedron 1996 52 165. 3 S. Halazy A. Ehrard A. Eggenspiller V. Berges-Gross and C. Danzin Tetrahedron 1996 52 177. 4 Bien Ye and T. R. Burke Jr. Tetrahedron 1996 52 9963. 5 M. M. Campbell M. Sainsbury and P. A. Searle Synthesis 1993 179. 6 K. S. Anderson J. A. Sikorski and K. A. Johnson Biochemistry 1988 27 1604. 7 P. D. Pansegrau K. S. Anderson T. Widlanski J. E. Ream R. D. Sammons J. A. Sikorski and J. R. Knowles Tetrahedron Lett. 1991 32 2589. 8 M. C. Walker and C. R. Jones J. Am. Chem. Soc. 1992 14 7601. 9 D. P. Philion and D. G. Cleary J. Org. Chem. 1992 57 2763. 10 A. Radzicka and R. Wolfenden Biochemistry 1991 30 4160. 11 D. J. Burton and R. M. Flynn J. Fluorine Chem. 1977 10 329. 12 D. J. Burton T. Ishiara and M. Murata Chem.Lett. 1982 755. 13 A. C. Cope and W. G. Dauben Org. React. 1992 41 135. 14 D. J. Burton and L. G. Sprague J. Org. Chem. 1989 54 613. 15 D. E. Bergstrom and P. W. Shum J. Org. Chem. 1988 53 3953. 16 R. V. Hofmann R. D. Bishop P. M. Fitch and R. Hardenstein J. Org. Chem. 1980 45 919. 17 G. H. Posner G. M. Gunia and K. A. Babiak J. Org. Chem. 1977 42 3173. 18 G. M. Blackburn and D. E. Kent J. Chem. Soc. Perkin Trans. 1 1986 913. Paper 6/03094G Received 2nd May 1996 Accepted 15th November 1996 © Copyright 1997 by the Royal Society of Chemistry
ISSN:1472-7781
DOI:10.1039/a603094g
出版商:RSC
年代:1997
数据来源: RSC
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