A new convenient synthesis of 2-trifluoromethyl substituted aspartic acid and its isopeptides. Part 11

摘要

J. CHEM. SOC. PERKIN TRANS. t 1992 A New Convenient Synthesis of 2-Trif luoromethyl Substituted Aspartic Acid and its Isopeptides. Part 11 ‘ Norbert Sewald,” Jurgen Riede,b Peter Bissingerb and Klaus Burgera a Organisch- Chemisches tnstitut, Technische Universitat Miinchen, Lichtenbergstrasse 4, W-8046 Garching, Germany Anorganisch- Chemisches tnstitut, Technische Universitat Miinchen, Lichtenbergstrasse 4, W-8046 Garching, Germany The reaction of methyl 2-benzyloxycarbonylimino-3,3,3-trifluoropropionatewith acetyl chloride-triethyl- amine yields methyl 2- benzyloxy-6-oxo-4,5-dihydro-4-trifluoromethyl-l,3-oxazine-4-carboxylate,a 9-activated equivalent for 2-trifluoromethyl substituted aspartic acid. This reagent offers a versatile route to 9-derivatized 2-trifluoromethyl substituted aspartic esters via nucleophilic ring cleavage; with amino acid esters isoaspartyl peptides are formed.The diastereoisomeric dipeptides have been separated by flash chromatography; for benzyl N-benzyloxycarbonyl -R -2-trifIuoromet hyl -p-aspartyl-(a-methylester) -S-prolinate an X-ray analysis is presented. Benzyl N-benzyloxycarbonyl-S-2-trifluoro-methyl-9-aspartyl- (a-methylester) -S-prolinate exists as a mixture of cis-trans conformers in solution at room temperature. Peptides with isoaspartyl substructures like compound 1 play an important role in bi~chemistry.~.~ The linkage of aspartic acid to the next amino acid is formed with the f3-carboxy group. Aging of peptides containing aspartic acid and mismatches during peptide synthesis in organisms lead to an accumulation of those isoaspartyl peptides in viuo.In eucaryotic cells, the mismatched peptides and proteins can be recognized and reisomerized by a repair mechanism. The free a-carboxy group i of the isoaspartic moiety is methylated by the enzyme carboxy R20& CF,methylase. Loss of methanol leads to the intermediate ~formation of the succinimide 3 which is hydrolysed to give a HN~o~R mixture of aspartyl 4 and isoaspartyl peptides 1, the latter re- R’ ~0~~3 entering the isomerization cycle (Fig. 1). Iii IV t R202C&iR3 -iii ““2CpJOHHN Hooc~OHH2N x HCI XoR’ R1AO 6 Scheme 1 Reagenrs: i, CH,=CHOR3; ii, R30H, H+; iii, KMnO,; iv, H,C(C0,R3),; v, H30+ and decarboxylation processe~.~.~ The replacement of natural amino acids in peptides by non-natural amino acids is a widely used strategy for stabilization of the scissible peptide bond.3 There are only few reports in the literature on strategies for Fig. 1 the synthesis of 2-trifluoromethyl substituted aspartic acid 6 7-9 (Scheme 1). Most of them start from acyl imines 5 of 3,3,3-As correctly incorporated aspartyl residues are not trifluoropyruvates. methylated enzymatically, the net result of this reaction cycle is the conversion of isoaspartyl peptides to aspartyl pep tide^.^ Mismatched peptides not being recognized by the enzyme are degraded partially and excreted; significant amoilnts of isoaspartyl peptides can be isolated from urine. 7The introduction of a a-trifluoromethyl group into the 5 backbone of aspartic acid would facilitate further investigations Scheme 2 Recrgents: i, CH,=CH[CH,],MgX (n = 1-4); ii, KMnO, on the metabolism of isoaspartyl peptides in vitro by 19FNMR spectroscopy. Furthermore, x-trifluoromethyl substituted Recently we presented a versatile route to N-protected 2-tri- amino acids are known to be potent inhibitors of pyridoxal fluoromethyl substituted o-carboxylic x-amino acids 7 lo phosphate-dependent enzymes, which catalyse transamination (Scheme 2).268 J. CHEM. SOC. PERKIN TRANS. 1 1992 Results and Discussion The acyl imines of hexafluoroacetone 8 or 3,3,3-trifluoro-pyruvates 5 can be considered as 1,4-dipolar species with nucleophilic character at position 1 and strongly electrophilic character at position 4.During our investigation into the reaction behaviour of ketene and substituted ketene derivatives towards the acyl imines 8 of hexafluoroacetone (Scheme 3), originally reported by Gambaryan and Zeifman," we concluded that, under the reaction conditions applied by these authors ('ketene gener- ation' with acetyl chloride and base in situ), no free ketene should be involved. ' 8 9 8 10 Scheme 3 Reagents: i, R,C=C=O;ii, CH,COCI, NEt, On reaction of the acyl imines with acetyl chloride-triethyl- amine, an anionic intermediate is formed by nucleophilic attack of the acetyl chloride anion (Scheme 4).It cyclizes to 10 by a 6-em-trig process according to Baldwin's rules.' Stable substituted ketenes like diphenylketene or bis(trifluoromethy1)- ketene normally add to 4,4-bis(trifluoromethyl) substituted heterodienes like acyl imines 8 12.14-1 to yield six-membered cycloadducts 9 across the ketene CO-bond.11 10 Scheme 4 During the course of our research into the synthesis of trifluoromethyl substituted heterocycles and amino acids, we recognized that heterocycle 12, formed by reaction of the 2-acylimino-3,3,3-trifluoropropionates5a with acetyl chloride in the presence of triethylamine, is a P-activated derivative of 2-trifluoromethylaspartic acid. Position 6 is highly activated towards nucleophilic attack due to its anhydride-like structure (Scheme 5). $O,Me F3C, ,C 0,Me 5a 12 Scheme 5 Ring cleavage by hydrochloric acid (1 mol dmP3) at ambient temperature yields N-protected 2-trifluoromethylaspartyl X-methyl ester 13 (Scheme 6).Benzylamine cleaves the six-membered ring 12 at room temperature within minutes. Under the reaction conditions applied product 14a undergoes spontaneous cyclization to give F3C, ,C02Me HCI (1 rnol dm4) -Meo2c~OH ZH N Ph-0 12 13 Z-TFMAsp(OH)-OMe Scheme 6 succinimide 15 (Scheme 7) which is a trifluoromethyl substituted analogue of species 3 known from the biochemical studies mentioned above. Secondary amines like N-benzyl- aniline and 12 form derivatives of 2-trifluoromethylasparagine, a ring closure to give the succinimide now not being possible. F3C, /CO,Me R 14aR=H 14b R = Ph II -MeoH 15 Scheme 7 With esters of a-amino acids the 2-trifluoromethyl-P-aspartyl dipeptides 16, 18 and 19 are formed instantaneously at 0°C (Scheme 8).Cyclization to succinimides can be prevented by bulky residues in the amino acid ester backbone. Minor amounts of succinimide 17 can be isolated with less sterically hindered amino acids like alaninates. They have been characterized by NMR and GC-MS. The benzyloxycarbonyl group can be cleaved by hydrogen- ation according to standard procedures (lh20) (Scheme 9). The fully protected or, preferably, the N-deprotected diastereoisomeric dipeptides (4.8.20)are conveniently separated by flash chromatography. The strategy outlined offers a unique, preparatively simple access to dipeptides containing N-terminal 2-trifluoromethyl-P-aspartyl residues. Furthermore, the facile resolution of the diastereoisomers enables, after cleavage of the dipeptide, the preparation of enantiomerically pure 2-trifluoro- methyl substituted aspartic acid.This fact is especially im- portant as effective enantio- or diastereo-selective syntheses of 2-trifluoromethyl substituted amino acids are not reported in the literature. Synthetic routes to the optical isomers of 2-tri-fluoromethyl substituted amino acids generally rely on fluorin- ation of an optically active precursor l7 or resolution by chemical l8 or biochemical l9 means.20 The structures of the products described are confirmed by standard analytical techniques. The most important diagnostic criterion for the identification of the succinimide of type 17 is loss of methanol; in the 'H NMR spectra the signal corresponding to the methyl group of the ester function is no longer observed.One diastereoisomer of the dipeptide Z-TFMAsp-(S-Pro- 0Bzl)-OMe* 19a is obtained as a crystalline solid after * TFM amino acids = trifluoromethyl amino acids; e.g. 2-TFMAsp = 2-trifluoromethyl aspartic acid. J. CHEM. SOC. PERKIN TRANS. 1 1992 ZHN F3C C02Me H2;4c02BUf "c -16Z-TFMAsp(SAla-OBu')-OMeNloPhfiOAO + CH3 12 + H C02Bur."7*W3 17 F3C, ,C02Me 0"C Ph-0 y 2 Ph ZHN Ph 12 18 Z-TFMAsp(S-Phe4Me)-OMe 12 19 Z-TFMAsp(SPro-OBzl)-OMe Scheme 8 Ph 18 Z-TFMAsp(S-Phe-OMe)-OMeI" t Me02C CF30 H p02Me H2NuN5 l!i Ph 20 H-TFMAsp( S-Phe--OMe)-OMe Scheme 9 chromatographic resolution.It was characterized by X-ray (Table 1) analysis (Fig. 2).* The S-proline fragment provides a reference system for the absolute configuration at the chiral centre of the 2-trifluoro- methylisoaspartyl moiety, which can therefore be identified as having the R-configuration. In the crystalline state, the trans configuration at the peptide bond is favoured. The second diastereoisomer 19b of this dipeptide [Z-S-TFMAsp-(S-Pro- 0Bzl)-OMe] is a stable mixture of two conformers (19ba, 19bb) in solution. Obviously, rotation around the peptide bond between the two amino acids is hindered, as is often observed in proline derivatives2 For peptides composed of a-amino acids, the trans conformers are generally expected to be the more stable ones. In the presence of proline, the energy difference *Tables of bond lengths and bond angles, thermal parameters and hydrogen atom co-ordinates have been deposited with the Cambridge Crystallographic Data Centre (see Instructions for Authors, J.Chem. Soc., Perkin Trws. 1, 1992, Issue I). Fig. 2 between the cis and trans forms is thought to be quite low. In the case of the side chain of oxytocine (S-Bzl-Cys-Pro-Leu-Gly- NH,), the trans form predominates in ['H,]DMSO over the cis form in an approximate ratio of 3:2.,' The ratio of the conformers 19ba and 19bb as measured by 19F NMR depends on the solvent polarity (in CDCl, 1.3:l; in [*H,]methanol 1.9:1). The structurally relevant NMR signals of the conformer mixtures have been assigned using COSY and C,H-correlation techniques.The structural assignment is based on two-dimensional NOE measurements (Fig. 3). In one conformer (19bb), which is less populated, a spatial relation exists between the r-proton of Table 1 Fractional atomic coordinates for 19a Atom XIu Ylh ZI'C 0.263 82(7) 0.189 5(3) 0.136 88(6) 0.304 48(7) 0.028 6(2) 0.061 13(5) 0.332 69(8) -0.095 3(2) 0.144 33(6) 0.594 7( I) 0.156 3(3) 0.073 33(7) 0.377 5(1) 0.420 33) 0.079 44( 8) 0.687 60(9) 0.223 9(3) 0.164 19(6) 0.744 9( 1) 0.505 4(3) 0.121 77(8) 0.531 86(9) 0.428 O( 3) 0.115 74( 7) 0.408 76(9) 0.288 6(3) -0.009 76(6) 0.394 4( 1) 0.647 7(3) 0.007 47(6) 0.386 2( 1) 0.505 2(3) 0.190 50(6) 0.425 50(9) 0.171 4(3) 0.21 3 45(6) 0.801 l(1) 0.867 O(2) 0.137 2(4) 0.2 I7 3( 5) 0.2 18 06(9) 0.193 9(1) 0.931 9(2) -0.107 6(6) 0.217 4(1) 0.931 4(2) 0.094 4( 7) 0.193 8(1) 0.867 2(2) -0.188 9(6) 0.242 O( 1) 0.802 l(2) -0.067 3(5) 0.242 2( 1) 0.730 2(2) 0.267 3(6) 0.217 6(1) 0.701 4(1) 0.356 l(4) 0.119 8(1) 0.661 4(1) 0.290 8(4) 0.065 0( 1) 0.710 9(2) 0.149 4( 5) 0.026 4( 1) 0.690 6( 2) -0.078 7( 5) 0.043 9( 1) 0.605 7( 1) -0.068 7(4) 0.053 9(1) 0.533 O( 1) 0.463 5( 1) 0.240 9(4) 0.099 4(4) 0.096 85(9) 0.097 9( 1) 0.393 8( I) 0.231 9(4) 0.115 14(9) 0.323 4( 1) 0.088 O(4) 0.114 3(1) 0.395 9( 1) 0.435 4(4) 0.022 4 l(9) 0.402 2( 1) 0.482 2( 1) 0.502 4(2) 0.695 l(5) 0.738 4(4) 0.938 6(5) -0.054 2( 1) -0.071 57(9) -0.093 3(1) 0.574 6(2) 0.627 8(2) 0.975 8(5) 0.816 3(5) -0.113 2(1) -0.1 10 9(1) 0.609 3(2) 0.536 9(2) 0.401 8(1) 0.616 5(5) 0.578 l(5) 0.324 7(4) -0.088 2( 1) -0.068 6(1) 0.177 21(9) 0.431 4( 2) 0.237 O(7) 0.274 O( I ) Table 2 Characteristic I3C NMR shift values ofcompounds 19 (S) 19ba 19bb 19a (f rans) (cis) proline X-C 58.67 58.74 59.56 proline p-C 29.05 29.31 30.84 proline y-C 24.53 24.48 22.64 H-19bb Scheme 10 proline (6 4.64) and the more deshielded /I-proton (6 3.92) of TFMAsp, whereas in the other conformer (19ba),a relationship between one TFMAsp P-proton (6 4.28) and the geminal 6-protons (6 3.48/3.87) of the proline residue is found.Therefore, 19bb has cis and 19ba trms conformation (Scheme 10). In addition, the carbon NMR shift values support this assignment. According to the literature, the proline y-carbon J. CHEM. SOC. PERKIN TRANS. I 1992 t I #@ 0 I 4 3 2 6, Fig. 3 NOESY spectrum of 19b atom should be more shielded in the cis isomer. Furthermore, the shift differences between the proline p-and y-carbon atoms should be greater for the cis isomer (8-10 ppm) than for the truns isomer (5-6 ppmj.2' The observed data (Table 2) support the assignment. The values in Table 2 clearly indicate that the diastereoisomer 19a has a trunsoid conformation in CDCl, solution. Experimental For chromatography silica gel 60 (63-200 pm, Merck) and for flash chromatography silica gel 60 (3&63 pm, Riedel-de Haen) were used.Chloroform, dichloromethane and ethyl acetate were distilled over calcium chloride; diethyl ether and dioxane were predried over calcium chloride-potassium hydroxide and dried over sodium benzophenone ketyl under nitrogen. Melting points (not corrected) were determined using a Tottoli apparatus (Buchi SMP-20); elemental microanalyses were carried out with a Heraeus CHN-Elemental Analyzer. IR spectra were recorded using Perkin-Elmer 157 G or 257 spectrophotometers; 'H, 13C and 19F NMR spectra were recorded with a Bruker AM 360 spectrometer at 360,90 and 339 MHz, respectively. 19F NMR spectra were obtained using JEOL FX 90 Q (84 MHzj and Bruker AC 250 (235 MHz) spectrometer.As reference standard TMS was used for 'H and 13C NMR spectra (internal) and trifluoroacetic acid for '"F NMR spectra (external). All J-values are given in Hz. Mass spectra were recorded from electron ionization (EI, 70 eVj with a Varian MAT CH5 instrument. GC-MS analyses were carried out with a Carlo Erba 4160 gas chromatograph (column SE 30) and a Varian MAT M 112s mass spectrometer. Optical rotation values were measured using a Perkin-Elmer 241 MC polari- meter. The X-ray analysis was performed on a Enraf-Nonius- CAD4-diffractometer using Mo-KX radiation and a graphite monochromator. Mctiij*l 2-Bi~ri~j~lo.\-j~-4,5-cliiij~cl~~-4-tr~uoran~i~thj~-6-0.\-0-I ,3-o.\-arine-4-c.urho.~j~lut~12.-To a solution of acyl imine 5a 23 (14.5 g, 50 mmol) and freshly distilled acetyl chloride (39.3 g, 500 mmolj in absolute diethyl ether (250 cm3) at 0 "C, triethylamine (5.5 g, 55 mmolj was slowly added.The solution was stirred for 2 h at 0 C, then hydrolysed with ice water and extracted with diethyl ether. The organic phases were combined, dried over MgSO,, filtered, and evaporated to dryness. Filtration through silica gel (eluent, CHCI,) yielded 12 (1 1.9 g, 72"J as a pale J. CHEM. SOC. PERKIN TRANS. I 1992 yellow oil which partially crystallized on standing (Found: C, 50.70; H, 3.85; N, 4.35. Cl4H1,F,NO, requires C, 50.76; H, 3.65; N, 4.23%); vmax(film)/cm-' 1850 (CO) and 1760 (CH); 6,(360 MHz,CDC13)3.30(1 H,d,J17.4,5-H),3.40(1 H,d,J17.4,5-H), 3.69 (3 H, S, CO,Me), 5.19 (1 H, d, J 12.2.C6H5CH20), 5.28 (1 H, d, J 12.2, C,H,CH,O) and 7.34 (5 H, m, Ar-H); 6,(90 MHz, CDCI, and DEPT-135) 45.32 (C-5), 53.84 (CO,Me), 60.40 (9, 2J['3C'9F{ 'H)] 33.5, C-4), 69.05 (C6H5CH20), 122.70 (q, 1J['3C'9F('H}] 281.7, CF,), 147.43 (C-2), 159.98 (C-6), 164.44 (CO,Me), 128.38, 128.77, 128.87 (CHarom) and 134.51 (Carom); 6,(84 MHz, CDC1,) 5.2 (s); m/z 331 (M. 15%), 224 (2, M -C~HSCH~O),108 (46, C,HSCH,OH), 107 (77, C6H5CH20) and 91 (100, C,H,CH,). Hydrolysis of Methyl 2-Benzylo.xy-6-oxo-4-tr~uoromethyl-4,Sdihydro-1,3-0.xazin-4-carboxylate 12.--Compound 12 (1.7 g, 5 mmol) was stirred at room temperature in a solution of hydrochloric acid (1 mol drn-,; 5 cm3) in dioxane (20 cm3). The reaction progress was monitored by 19FNMR.After 3 h, the reaction mixture was extracted with chloroform; the organic layer was dried over MgS0, and evaporated to dryness in uacuo to yield 1-methyl hydrogen N-benzyloxycarbonyl-2-trifluoro-methylaspartate 13 (1.1 g, 62%) as an orange oil (Found: C, 48.25; H,4.20; N, 3.95. cl4Hl4F3No6 requires C, 48.15; H, 4.04; N, 4.01%); vma,(film)/cm-' 3400 (OH), 3360-3260 (NH) and 1745 (CO); 6,(360 MHz, CDCI,) 3.32 (1 H, br d, J 17.3, H,), 3.83 (3 H, s, CO,Me), 4.19 (1 H, br d, J 17.3, H6), 5.05 (1 H, d, J 12.3, C~HSCH~O), 5.10 (1 H, S, J 12.3, C,H,CH20), 6.31 (1 H, br s, NH), 7.31 (5 H, m, Ar-H) and 10.07 (1 H, br, C0,H); 6,(W MHz, CDCl,) 33.09 (br, CB), 54.52 (CO,Me), 63.10 (9, 2J['3C'9F{ 'H)] 29.5, C=), 67.39 (br, C,H,CH,O), 123.35 (4, 'J[13C'9F{ 'H}] 288.0, CF,), 154.30 (OCONH), 166.10 (CO,Me), 173.37 (C?), 128.09, 128.39, 128.62 and 135.71 (Carom); SF(84 MHz, CDCI,) 3.1 (s); m/z 349 (M, 479, 331 (1 , M -HZO), 224 (2, 331 -C,H,CH,O), 108 (IW, C,H,CH,OH), 107 (49, C,H,CH20) and 91 (99, C7H7).Reaction of 12 with Benzy1amine.-A solution of 12 (2.6 g, 7.8 mmol) in absolute dichloromethane (10 cm3) was treated with benzylamine (2 cm3, 18.3 mmol) and stirred at room temperature. The progress of the slightly exothermic reaction was monitored by 19FNMR. After 1 h, ice water (50 cm3) was added. The reaction mixture was acidified (pH 6) with hydrochloric acid (1 mol drn-,), extracted with dichloromethane (50 cm3), and the organic layer was dried over MgSO, and evaporated to dryness in uacuo.The remaining yellow oil crystallized slowly after purification by column chromatography over silica gel (eluent, CHCI,) to yield 1-benzyl-3-benzyloxy- carbonylamino-3-trifluoromethylpyrrolidine-2,5-dione15 (2.25 g, 71%), m.p. 96°C (Found: C, 59.15; H, 4.50; N, 6.95. C20H17F3N204 requires C, 59.11; H, 4.22; N, 6.89%); vmax(KBr)/ cm-' 3330 (NH), 1720 (CO) and 1705 (CO); 6,-(360 MHz, CDCI,) 3.16 (1 H, d, J 18.6,4-H), 3.31 (1 H, br, d, J 18.6, 4-H), 4.69 (2 H, br s, C,H,CH,N), 5.06 (2 H, br s, C,H,CH,O), 5.86 (1 H, s, NH) and 7.26-7.35 (10 H, m, Ar-H); 6,(90 MHz, CDCI,, and DEPT-135) 36.27 (br, C-4), 43.27 (C,H,CH,N), 62.15 (9, 2J['3C'9F{ 'H)] 29.9, C-3), 68.01 (C,H,CH,O), 123.1 1 (q, 1J['3C'9F{ 'H)] 285.2, CF,), 154.38 (OCONH), 169.71 (br, C-2), 171.56 (C-5), 128.1 1, 128.31, 128.35, 128.51, 128.64, 128.66 (CHarOm), 134.48, and 135.00 (Carom);6,(84 MHz, CDCI,) 1.8 (s); m/z 406 (M, l%), 315 (18, M -C,H,CH,), 298 (7, M -C,H,CH,OH), 255 (6, 298 -HNCO), 132 (1 5, C,H,CH=N-CO), 108 (22, C,H,CH,OH) and 9 1 (C,H&H,).Reuction qf'12 with N-Benzyluniline.-A solution of 12 (1.7 g, 5 mmol) and N-benzylaniline (0.9 g, 5 mmol) in absolute dichloromethane (20 cm3) was stirred at room temperature. The 27 1 reaction progress was monitored by '9FNMR. The solvent was removed in uucuo. The residue, a yellow oil, was purified by flash chromatography on silica geI (eluent, ethyl acetate-hexane, 1:10) yielding methyl N6-benzyl-Na-benzyloxycarbonyl-Na-phenyl-2-trifluoromethylasparaginate 14b (0.8 g, 31%) as a colourless oil (Found: C, 62.45; H, 4.85; N, 5.40.C27H25F3N205 requires C, 63.03; H, 4.90; N, 5.44%); v,,,(film)/cm-' 3390 (NH), 1750 (CO), 1730 (CO), 1650 (CO) and 1500 (N-CO); 6,(360 MHz, CDC1,) 3.07 (1 H, d, J 16.7, Ha), 3.91 (3 H, S, CO,Me), 3.96 (1 H, d, J 16.7, Ha), 4.74 (1 H, d, J 14.4, NCH,C,H,), 4.93 (1 H, d, J 14.4, NCH,C6H,), 5.16 (2 H, S, C,H,CH,O), 6.65 (1 H, br s, NH), 7.06 (2 H, m, Ar-H), 7.19 (2 H, m, Ar-H), 7.25 (3 H, m, Ar-H), 7.36 (4 H, m, Ar-H) and 7.42 (4 H, m, Ar-H); 6,(90 MHz, CDCI,) 33.92 (Ca), 53.27 (NCH,C,- H,), 54.41 (CO,Me), 63.81 (9, 2J['3C'9F { 'H)] 28.6, C,), 67.1 1 (C,H,CH,O), 123.93 (9, 'J[ 13C19F( 'H)] 288.0, CF,), 154.30 (OCONH), 167.12 (CO,Me), 167.65 (Cy), 127.70, 128.02, 128.39, 128.58, 128.59, 128.80, 128.81, 128.88, 130.00, 136.44, 137.26 and 141.30 (Carom); dF(84 MHz, CDCl,) 3.4 (s); m/z 514 (M, 2%), 406 (30, M -C6H,CH,OH), 347 (7, 406 -COZCH,), 224 (13, 406 -C~H~CHZNC~H,),183 (13, C6H5CH,NHC,H,), 108 (20, C,H,CH,OH), 107 (1 7, C6H5CH20) and 91 (100, C7H7).Synthesis of 2-TriJluoromethylisoaspurtyl Peptides.-To a solution of 12 (2.5 g, 7.5 mmol) in absolute diethyl ether (100 cm3) or absolute dichloromethane (100 cm3) at 0 "C was slowly added a solution of the corresponding a-amino acid ester (10 mmol) in diethyl ether (10 cm3) or dichloromethane (10 cm3), respectively. The reaction mixture was stirred at 0 "C until the 19FNMR spectrum of the solution indicated that the reaction had gone to completion.After evaporation of the solvent, polar impurities were removed by filtration through a 10 cm layer of silica gel (eluent, ethyl acetate-hexane, 2: 1). The remaining' colourless oil contained the two diastereoisomeric dipeptides and, in the case of alanine tert-butyl ester, minor amounts of non-polar impurities like the succinimide 17. The products were purified by flash chromatography on silica gel (50 cm column; eluent, ethyl acetate-hexane, 1:5); resolution of the diastereoisomers was thereby achieved. tert-Butyl N-Benzylo~~ycurbonyl-2-triJluoromethyl-~-uspart~?l-(a-methylester)-S- Alaninate [Z-TFMAsp(S-Ala-0Bu')-OMe] 16.-tert-Butyl alaninate (1.5 g, 10 mmol) was used to yield 16a (0.8 g, 23%), 16b (0.8 g, 23%) and the diastereoisorneric succinimides 17 (0.3 g, 8%) as colourless oils.Diastereoisomer 16a (Found: C, 52.55; H, 5.70; N, 5.80. CZ1H2,F3N207requires C, 52.94; H, 5.71; N, 5.88%); [z]L5 -5.3 (c 1.0 in CHCI,); vmax(film)/cm-' 3395 (NH), 1750 (CO), 1730 (CO), 1670 (CO) and 1510 (N-CO); 6,(360 MHz, CDCI,), 1.18 (3 H, d, J 7.0, AlaH,), 1.45(9H,s,AlaC02Bu'),3.21(1 H,d, J15.3,TFMAsp H,), 3.88 (1 H br, d, J 15.3, TFMAsp Ha), 3.90 (3 H, s, C0,Me) 4.36 (1 H, dq, J 7.6, 7.0, Ala Hs), 5.07 (2 H, br s, C6H,CH,0), 6.31 (lH,brd,J7.6,AIaNH),6.45(1H,brs,TFMAspNH)and 7.33 (5 H, m, Ar-H); 6,(90 MHz, CDCI,) 18.25 (Ala Ca), 27.94 (Bu'), 35.37 (br, TFMAsp C,), 48.72 (Ala Cm), 54.28 (CO,Me), 63.54 (9, 2J[13C19F{1H)] 29.5, TFMAsp CJ, 67.10 (C,H,CH,O), 82.18 (Oh'), 123.63 (4,'J['3C19F('H)] 288.2, CF,), 154.22 (OCONH), 166.54 (TFMAsp CO,Me/TFMAsp Cy), 171.66 (Ala CO,Bu'), 128.04, 128.28, 128.57 and 135.84 (Carom);&(84 MHz, CDCI,) 2.8 (s);m/z 477 (M + 1,O.lx), 476 (0.1, M), 420 (4, M -C4Hg), 403 (2, M + 1 -C4Hg -HZO), 375 (5, 403 -CO), 332 (13, M -C,H,,N02), 268 (I, M + 1 -C4H8 -CO -H2O -C,H,CH,O), 267 (2, M -C4H8 -CO -H2O -C,H,CH,O), 108 (5, C,H,CH,OH), 107 (6, C,H,CH,O), 91 (100, C,H,), 57 (1 3, C4H9) and 44 (26, (332).Diastereoisomer 16b (Found: C, 52.95; H, 5.25; N, 5.90. 272 C21H27F3N207requires C, 52.94; H, 5.71; N, 5.88%); calk5 -7.0 (c 1.0 in CHCI,); vmax(film)/cm-' 3390 (NH), 1750 (CO), 1730 (CO), 1670 (CO) and 1510 (N-CO); 6,(360 MHz, CDCl,) 1.33 (3 H, d, J 7.0, Ala H,), 1.44 (9 H, s, Ala CO,Bu'), 3.16 (1 H, d, J 14.1, TFMAsp H,), 3.90 (1 H, br d, J 14.1, TFMAsp H,), 3.91 (3 H, s, CO,CH,), 4.39 (1 H, dq, J9.0,7.0, Ala Ha), 5.08 (1 H, d, J 12.3, C,H,CH,O), 5.17 (1 H, d, J 12.3, C,H,CH,O), 6.39 (1 H, br s, TFMAsp NH), 6.48 (1 H, d, J9.0, Ala NH) and 7.35 (5 H, m, Ar-H); 6,-(90 MHz, CDCl,) 18.38 (Ala C,), 27.92 (Bu'), 35.51 (TFMAsp C,), 48.69 (Ala C,), 54.26 (C02Me), 63.65 (q, 2J['3C'9F('H)] 29.7, TFMAsp Ca), 67.30 (C,H,CH,O), 82.01 (CO~BU'), 123.63 (9, 'J['3C'9F('H}] 288.4, CF,), 154.41 (OCONH), 166.50 (TFMAsp CO,Me), 166.80 (TFMAsp Cy), 171.49 (Ala CO,Bu'), 128.09, 128.21, 128.51 and 135.84 (Carom); 6,(84 MHz, CDCl,) 2.9 (s); m/z 477 (M + 1, 0.1%) 476 (0.1, M), 420 (4, M -C4H8), 403 (2, M + 1 -C4HB -HZO), 375 (4, 403 -CO), 332 (12, M -C,H,,NO,), 268 (1, M + 1 -C4H8 -CO -H2O -C6H,CH,O), 267 (2, M -C4H8 -CO -H2O -C6H5-CHZO), 108 (4, C,H=,CH,OH), 107 (7, C,H,CH,O), 91 (100, C7H7), 57 (14, C4H,) and 44 (29, CO,).tert- Butyl 2-( 3- Benzyloxycarbonylamino-3 -triJuoromethyl-pyrrolidin-2,5-dion-1-yl)-propionate 17, diastereoisomeric mix- ture, C20H23F,N20,; SH(36O MHz, CDCl,) 1.42 (9 H, s, Bu'), 1.50 (3 H, d, J 7.2, 3-H), 3.12/3.15 (1 H, d/d, J 17.1/17.1, pyrrolidindione 4-H), 3.48 (1 H, br, d, J 17.1, pyrrolidindione 4- H), 4.72/4.76 (1 H, q/q, 1 H, J 7.2/7.2, 2-H), 5.10 (2 H, br s, C6H5CH20), 5.99/6.00 (1 H, s/s, NH) and 7.34 (5 H, m, Ar-H); 6,-(90 MHz, CDCl,), 13.73/13.93 (C-3), 27.71 (Bu'), 36.27/36.45 (pyrrolidindione C-4), 49.63/49.74 (C-2), 6 1.93/62.22 (q/q, ,J[ 13C19F( 'H}] 30.0/30.1, pyrrolidindione C-3), 67.91/68.06 (C,H,CH,O), 82.82/82.92 (OBU'), 123.18 (9, 1J['3C'9F( 'H}] 286.2,CF3), 154.05/154.57(OCONH), 167.23/167.35(pyrrolidin-dione C-2), 168.91/ 169.10 (pyrrolidindione C-5), 170.96 (C- 1), 128.37, 128.39, 128.61, 128.68, 128.70, 128.72, 135.12 and 135.32 (Carom); &(84 MHz, CDCI,) 0.7/0.9 (s); GC-MS: diastereoisomer 1: m/z 388 (M -C4HS, 473, 370 (5, 388 -HZO), 342 (2,370 -CO), 253 (5,388 -CO -C,H,CH,O),235 (3, 253 -H,O), 209 (2, 253 -CO,), 108 (14, C,H,CH,OH), 107 (30, C,H,CH,O), 91 (100, C7H7) and 57 (66, C,H,); diastereoisomer 2: m/z 388 (M -C,H,, 373, 370 (3, 388 -H,O), 342 (1, 370 -CO), 253 (3, 388 -CO -C~HSCH~O),235 (2, 253 -H,O), 209 (2, 253 -CO,), 108 (15, C6HsCH,OH), 107 (30, C,H,CH,O), 91 (100, C7H7) and 57 (72, C4H9).Methyl N-Benzyloxycarbonyl-2-trijluoromethyl-~-aspartyl-(x-methylester)-S-phenylalaninate [Z-TFMAsp(S- Phe-0Me)- OMe] lS.-MethyI S-phenylalaninate (1.9 g, 10 mmol) was used to yield the two diastereoisomeric dipeptides 18a (1.2 g, 32%) and 18b (1.2 g, 32%) as colourless oils. Diastereoisomer 18a (Found: C, 56.25; H, 4.90; N, 4.95. C2,H2,F3N207 requires C, 56.47; H, 4.94; N, 5.49%); +6.9 (c 0.5 in CHCI,); v,,,(film)/cm-' 3390 (NH), 1730 (CO), 1670 (CO) and 1500 (N-CO); 6,(360 MHz, CDCI,), 2.99 (1 H, dd, J 13.9, 6.6, Phe H,), 3.10 (1 H, dd, J 13.9, 5.8, Phe H,), 3.11 (1 H, d, J 15.2, TFMAsp H,),3.64(3 H,s,CO,Me),3.86(3 H,s,CO,Me),3.89 (1 H, br d, J 15.2, TFMAsp H,), 4.74 (1 H, m, Phe Ha), 4.95 (1 H, d, J 12.1, C6HsCH20), 5.01 (1 H, d, J 12.1, C,H,CH,O), 6.38 (1 H, s, TFMAsp NH), 6.51 (1 H, br d, J8.5, Phe NH) and 7.07- 7.34 (10 H, m, Ar-H); 6,-(90 MHz, CDCI,), 35.33 (TFMAsp C,), 37.51 (Phe C,), 52.27 (Phe CO,Me), 53.36 (Phe Ca), 54.31 (TFMAsp CO,Me), 63.62 (4, 2J['3C'9F(1H)] 29.6, TFMAsp c%), 67.26 (C,H,CH,O), 123.59 (q, 1J['3C'9F('H)] 288.0, CF,), 154.57 (OCONH), 166.38 (TFMAsp CO,Me), 167.29 (TFMAsp Cy), 17 1.32 (Phe CO,Me), 127.20, 128.08, 128.22, 128.49, 128.67, 129.15, 135.72 and 135.80 (Carom); 6,(84 MHz, CDCI,) 2.9 (s); WZ/Z 510 (M, 40/,), 419 (2, M -C7H7), 348 [5, Z-J.CHEM. SOC. PERKIN TRANS. 1 1992 TFMAsp(NH,)-OMe], 241 (3, 348 -C,H,CH,O), 162 (46, C,H,CH=CHCO,CH,) and 91 (100, C,H,).Diastereoisomer 18b (Found: C, 56.10; H, 4.80; N, 5.20. C2,H,,F,N2O7 requires C, 56.47; H, 4.94; N, 5.49%); [x];~+ 19.9(c 0.3 in CHCI,); v,,,(film)/cm-' 3390 (NH), 1730 (CO), 1670 (CO) and 1500 (N-CO); 6,(360 MHz, CDCl,) 2.86 (1 H, dd, J 13.8, 6.0, Phe H,), 3.02 (1 H, dd, J 13.8, 5.4, Phe H,), 3.22(1 H,d,J 15.3,TFMAsp H,),3.65 (3 H,s,CO,Me),3.88 (3 H, s, CO,Me), 3.90 (1 H, br d, J 15.3, TFMAsp HJ, 4.78 (1 H, ddd, J 7.9, 6.0, 5.4, Phe Ha), 5.01 (1 H, d, J 12.3, C,H,CH,O), 5.09 (1 H, d, J 12.3, C,H,CH,O), 6.22 (1 H, br, J7.9, Phe NH), 6.42 (1 H, s, TFMAsp NH), 7.08 (2 H, m, Ar-H) and 7.22-7.32 (8 H, m, Ar-H); 6,(90 MHz, CDCl,) 35.06 (TFMAsp C,), 37.92 (Phe C,), 52.28 (Phe CO,Me), 53.17 (Phe Ca), 54.33 (TFMAsp CO,Me), 63.42 (q, 2J['3C'9F(1H)] 29.4, TFMAsp Ca), 67.07 (C6H5CH20), 123.62 (9, 1J['3C'9F('H}] 287.9, CF,), 154.23 (OCONH), 166.45 (TFMAsp CO,Me), 166.80 (TFMAsp Cy), 171.41 (Phe CO,Me), 127.21, 127.90, 128.27, 128.57, 128.66, 129.25, 135.46 and 135.82 (Carom); 6,(84 MHz, CDCl,) 2.6 (s); m/z 510 (M, 573, 419 (2, M -C7H7), 348 [6, Z-TFMAsp- (NH,)-OMe], 241 (4, 348 -C,H,CH,O), 162 (44, C,H,CH= CHCO,CH,) and 91 (100, C7H7).Benzyl N-Benzyloxycarbonyl-R-2-trijluorometh~l-~-aspart~~l-(a-methylester)-S-prolinate [Z-R-TFMAsp(S-Pro-0Bzl)-OMe] 19.-Benzyl S-prolinate (1.5 g, 10 mmol) was used to yield the two diastereoisomeric dipeptides 19a (0.8 g, 20%) and 19b (0.8 g, 20%). After resolution by flash chromatography, the diastereo- isomer 19a was recrystallized from benzene-hexane (m.p.68 "C). Diastereoisomer 19a (Found: C, 58.22; H, 4.98; N, 5.24. C,,H,,F,N,O, requires C, 58.21; H, 5.07; N, 5.22%); [x]k5 -49.3 (c 1.0 in CHCI,); v,,,(KBr)/cm-' 3400 (NH), 1750 (CO), 1640 (CO) and 1500 (N-CO); 6,(360 MHz, CDCl,) 1.9Cb2.05 (4H,m,ProHC,/ProHY),3.22(1H,d,J16.2,TFMAspHB),3.58 (2 H, m, Pro H6), 3.81 (3 H, s, CO,Me), 4.19 (1 H, br d, J 16.2, TFMAsp HC,), 4.40 (1 H, dd, J8.0,3.0, Pro HC,), 5.02 (1 H, d, J 12.4, C,H,CH,O), 5.06 (I H, d, J 12.4, C,H,CH,O), 5.08 (1 H, d, J 12.4, C6H5CH20), 5.16 (1 H, d, J 12.4, C6H5CH20), 6.45 (1 H, br s, NH) and 7.25-7.35 (10 H, m, Ar-H); 6,(90 MHz, CDCI,, and DEPT-135) 24.53 (Pro Cy), 29.05 (Pro C,), 33.06 (TFMAsp C,), 46.97 (Pro C6), 54.20 (CO,CH,), 58.67 (Pro CJ, 63.19 (9, 'J[' ,C1 9F( 'H}] 29.0, TFMAsp CJ, 66.75 (2 x C,H,CH,O), 123.75 (q, 1J[13C'9F('H)] 288.0, CF,), 154.17 (OCONH), 166.29 (TFMAsp CO,Me), 166.65 (TFMAsp CJ, 171.69 (Pro CO,Bzl), 127.66, 127.92, 128.08, 128.22, 128.39, 128.46, 128.54, 135.73 and 136.18 (Carom); 6,(84 MHz, CDCI,) 2.5 (s); m/z 536 (M, 5%), 428 (1, M -C,H,CH,OH), 401 (7, M -CO,CH,C,H,), 332 (2, M -C12H,,N02), 293 (7, 401 -C,H,CH,OH), 204 (4, Cl2H1,NO2), 91 (83, C7H7) and 70 (100, C,H,N).A single crystal was sealed in a glass capillary under argon. Crystal data. C26H27F3N207r M = 536.5. Orthorhombic, space group P212121 (No. 19), n = 17.756(2),b = 6.171(l), c = 23.146(3) A, V = 2536.16 A3, Z = 4, D,= 1.405 g cm-,. Colourless needles, p(Mo-Kx) = 1.09 cm-'.Data collection and processing. CAD4 diffractometer, graphite-monochromated Mo-Kx radiation, i = 0.710 69 A, T = -55 "C, 4062 reflections measured, 3528 unique (Rint= 0.0203), giving 3284 with F, 2 20(F0),no absorption correction. Structure analysis nnd refinement. Direct methods (SHELXS- 86 24), anisotropic refinement with all non-hydrogen atoms; all hydrogen atoms found subsequently in difference Fourier maps and refined isotropically. The weighting scheme )I' = 1.1673 c2(F,) gave satisfactory agreement analyses; final R and R, values are 0.036 and 0.027. Benqd N-Ben~~~lo.~~~ccrrhon~~l-S-2-tri~uoro~~~tli~l-~-~ispci~t~~l-(x-met1i~~Iestlier)-S-prolinate [Z-S-TFMAsp(S-Pro-OBz1)-OMS] 19b (2 conformers; Found: C, 57.80; H, 4.80; N, 4.80.J. CHEM. SOC. PERKIN TRANS. i 1992 C26H27F3N207requires c, 58.21; H, 5.07; N, 5.227;); [xlf: -49.2 (c 1.O in CHCI,); v,,,(film)/cm-' 3390 (NH), 1750 (CO), 1730 (CO), 1650 (CO) and 1500 (N-CO); G,(conformer 19ba, 360 MHz, CDCI,) 1.96 (1 H, m, Pro H,), 2.00 (1 H, m, Pro H,), 2.00 (1 H, m, Pro H,), 2.17 (1 H, m, Pro Hb), 3.20 (1 H, d, J 15.6, TFMAsp H,), 3.48 (1 H, m, Pro H6), 3.87 (1 H, m, Pro H6), 3.87 (3H,s,CO2Me),4.28(lH,brd,J15.6,TFMAspHD),4.51(1 H, br dd, J8.9, 3.4, Pro Ha), 5.04 (1 H, d, J 12.3, C6H,CH,0), 5.06 (1 H, d, J 13.6, C6H5CH,0), 5.10 (1 H, d, J 12.3, C,H,CH,O), 5.13(1 H,d,J13.6,C6HsCH,0),6.40(1H,brs,NH)and7.25-7.38 (10 H, m, Ar-H); G,(conformer 19bb, 360 MHz, CDCI,) 1.70 (1 H, m, Pro Hy), 1.73 (1 H, m, Pro Hy), 1.98 (1 H, m, Pro H,), 2.10 (1 H, m, Pro H,), 3.06 (1 H, d, J 15.6, TFMAsp H,), 3.31 (1 H, m, Pro H6), 3.46 (1 H, m, Pro H6), 3.87 (3 H, s, C02Me),3.92(1H,brd,J15.6,TFMAspHB),4.64(1H,br,d,J 7.0, Pro Ha), 4.97 (1 H, d, J 12.8, C,H,CH,O), 5.08 (1 H, d, J 12.8, C,H,CH,O), 5.15 (1 H, d, J 13.6, C,HsCH,O), 5.21 (1 H, d, J 13.6, C6H5CH,0), 6.41 (1 H, br s, NH) and 7.25-7.38 (10 H, m, Ar-H); &(conformer 19ba, 90 MHz, CDCI,, and DEPT-135) 24.48 (Pro Cy), 29.31 (Pro Cb), 33.38 (br, TFMAsp C,), 47.16 (Pro C,), 54.22 (CO,Me), 58.74 (Pro Cu), 63.41 (q, 2J[13C19F{1H)] 29.0, TFMAsp C,), 66.62, 66.71 (2 x C,-H,CH,O), 123.80 (9, 'J['3C'9F{ 'H)] 288.0, CF,), 154.16 (OCONH), 166.66, 166.71 (TFMAsp CO,Me/C,), 171.01 (Pro C0,Bzl); &(conformer 19bb, 90 MHz, CDCI,) 22.64 (Pro C,), 30.84 (Pro C,), 33.52 (br, TFMAsp C,), 46.37 (Pro C6), 54.30 (CO,Me), 59.56 (Pro C,), 63.43 (4, 2J['3C19F{ 'H)] 29.1, TFMAsp Cu), 66.36, 67.12 (2 x C6H5CH,0), 123.70 (4, 1J['3C'9F(1H}] 228.0, CF,), 154.17 (OCONH), 166.33 (TFMAsp CO,Me), 166.68 (TFMAsp-C,), 171.74 (Pro CO~BZI),127.51, 127.88, 128.00, 128.06, 128.13, 128.24, 128.43, 128.44, 128.48, 128.53, 128.61, 128.73, 135.23, 135.87, 136.10and 136.35 (Carom of both conformers); 6,(84 MHz, CDCI,), 2.4 (s); 2.8 (s); ratio of integral values in CDCI, 1.9: 1, in [2H4]methanol 1.3: 1; m/z 536 (M, 273, 428 (1, M -C~HSCH~OH),401 (4, M -CO,CH,C6H,), 332 (3, M -C12H,,N02), 293 (6, 401 -C,HSCH,OH), 204 (7, C12H14N02),91 (100, C7H7) and 70 (92, C4H8N).NOESY experiment: 32 scans (preceded by 2 dummy scans) were recorded at 25 "C into 2K data blocks for each of the 256 t, values with a mixing time of 600 ms, a relaxation delay of 2 s and spectral widths of 3597.12 Hz. Phase senstivity was achieved by the TPPI method. The longitudinal relaxation time TI was determined by an inversion-recovery experiment. The mixing time was randomized in the range &3%. After zero-filling to 2K x 2K data were apodized with shifted square sine bell functions. After FT and phase correction, a baseline correction in both dimensions was applied. Hydrogenolytic Cleavage of the N- Protecting Group. Methyl 2-Trijluoromethyl-P-aspartyl-(x-methylester)-S-phenylalaninate [H- TFMAsp(S- Phe-0Me)-OMe] 20.-A solution of the diastereoisomeric mixture of the dipeptide Z-TFMAsp(L-Phe- 0Me)-OMe 18 (2.0 g, 4 mmol) in absolute methanol (20 cm3) was treated with hydrogen in presence of palladium on charcoal (50 mg, 10% Pd) until hydrogen was no longer consumed.The catalyst was filtered off, the solvent was evaporated in vacuo, and the two diastereoisomers 20a (0.7 g, 48%) and 20b (0.7 g, 48%) were resolved by flash chromatography on silica gel (50 cm column, eluent ethyl acetate-hexane, 1 :2); diastereoisomer 20a (Found: C, 50.70; H, 5.00; N, 7.45. CI6Hl9F3N20, requires C, 51.07; H, 5.09; N, 7.44%); [XI'," +38.9 (c 1.2 in CHCI,); vm,x(film)/cm-l 3380 (NH), 3340 (NH), 1745 (CO), 1670 (CO) and 1540 (N-CO); S,(360 MHz, CDCI,) 2.33 (2 H, br s, NH,), 2.59 (1 H, d, J 15.7, TFMAsp H,), 3.01 (1 H, dd, J 13.9, 6.7,PheH,),3.08(1 H,br,d,J15.7,TFMAspHP),3.1l(1 H,dd, J13.9,5.6, Phe H,),3.71 (3 H,s,CO,Me),3.73 (3 H,s,CO,Me), 4.83 (1 H, ddd, J 7.8, 6.7, 5.6, Phe Hu), 7.00 (1 H, br d, J 7.8, Phe 273 NH), 7.10 (2 H, m, Ar-H) and 7.24-7.32 (3 H, m, Ar-H); S,(90 MHz, CDCI,) 37.84 (Phe C,), 37.86 (TFMAsp C,), 52.41 (Phe, CO,Me), 53.31 (Phe C,), 53.66 (TFMAsp CO,Me), 62.94 (9, 'J[' ,CI9F( 'H)] 27.5, TFMAsp Ca), 124.22 (4, 1J['3C'9Ff 'HI] 285.4, CF,), 168.00 (TFMAsp C,), 168.94 (br, TFMAsp CO,Me), 171.89 (Phe CO,Me), 127.23, 128.69, 129.22 and 135.77 (Carom); 6,(84 MHz, CDCI,) -0.74 (s); m/z 376 (M, 11%), 317 (19, M -CO,CH,), 285 (5, M -C7H,), 180 (28, C5H,F30,), 170 [17, CF,C(CO,CH,)NH,CH,], 162 (100, C6H5CH=CHC0,CH,), 156 [l8, CF,C(CO,CH,)=NH,], 91 (15, C7H7) and 88 (36, C4H802).Diastereoisomer 20b (Found: C, 50.80; H, 5.10; N, 7.15. C16Hl,F,N,0, requires C, 51.07; H, 5.09; N, 7.44%); [xlf:+ 121.1 (c 1.7 in CHCI,); v,,,(film)/cm-' 3370 (NH), 3340 (NH), 1740 (CO), 1665 (CO) and 1535 (N-CO); 6,(360 MHz, CDCI,) 2.38 (2 H, br, s, NH,), 2.59 (1 H, d, J 15.7, TFMAsp H,), 3.03 (1 H, dd, J 14.0, 6.5, Phe H,), 3.07 (1 H, br d, J 15.7, TFMAsp H,), 3.13 (1 H, dd, J 14.0, 5.6, Phe H,), 3.71 (3 H, s, CO,Me), 3.80 (3 H, s, CO,Me), 4.81 (1 H, ddd, J 7.5, 6.5, 5.6, Phe Ha), 6.97 (1 H, br d, J7.5, Phe NH), 7.1 1 (2 H, m, Ar-H) and 7.26-7.30 (3 H, m, Ar-H); 6,(90 MHz, CDCI,) 37.65 (Phe C,), 37.81 (TFMAsp C,), 52.42 (Phe, CO,Me), 53.30 (Phe Ca), 53.73 (TFMAsp CO,Me), 62.95 (9, 2J['3C'9F('H)] 27.5, TFMAsp C,), 124.18 (9, 1J['3C'9F('H}] 285.4, CF,), 167.90 (TFMAsp C,), 168.78 (br, TFMAsp CO,Me), 171.68 (Phe CO,Me), 127.23, 128.66, 129.20 and 135.74 (Carom); 6,(84 MHz, CDCI,) -0.76 (s); m/z 376 (M, 11%), 317 (16, M -COZCH,), 285 (7, M -C7H7), 180 (23, C,H,F,O,), 170 [17, CF,C(C02CH,)NH,CH,], 162 (100, C,H,CH=CH-CO,CH,), 156 [18, CF,C(CO,CH,)=NH,], 91 (15, C,H7) and 88 (39, C4H802). 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