摘要:
J. CHEM. SOC. PERKIN TRANS. I 1993 Synthesis of the Oligosaccharide Segment of a Novel Phenolic Glycolipid Antigen from Mycobacterium haemophilum M. K. Gurjar" and K. Revathi Reddy Indian Institute of Chemical Technology, Hyderabad 500 007, India The synthesis of the trisaccharide segment from Mycobacterium haemophilum phenolic glucolipid has been described. The synthesis of the disaccharide derivative 12 containing a free hydroxy group at C-2' was first examined by using 2-0-acetyl-4-0-benzyl-3-0-methyl-L-rhamnopyranosyltric hloro- acetimidate 9 as a glycosyl donor and 10as an aglycone followed by Zemplen deacetylation. The same glycosyl donor 9 was utilised for second coupling reaction with 12 to give the requisite trisaccharide 3 isolated after deacetylation, methylation and debenzylation.The highdegrees of a-selectivity in both the coupling reactions were substantiated by using 'H NMR, 13C NMR and partially decoupled 13C NMR spectral studies. The isolation of species-specific antigens from Mycobacterium (M.)leprae and M. tuberculosishave renewed significant interest in mycobacterium glycolipids. ' Recently it has also been observed that M. kansasii' and M. avium serotype 4 are found in the majority of patients who are HIV-1 positive. The structure and antigenicity of the phenolic glycolipid 1 from M. 0I OMe 1 /O-? MeOHo-tJ=7OH Me0 OMe 2 R = CHz(CH2)1&HCH2CH(CH2)&HCHCH2CHgI1 I1 OMe OMe Me OMe haemophilum that distinguished it from other mycobacteria has recently been de~cribed.~ The oligosaccharide segment was established as 2,3-di-O-Me-~-Rhap-(a1 -+2)-3-0-Me-~-Rhap-(a1 +4)-2,3-di-O-Me-~-Rhap-(al--+ which showed' novel structural similarities with that of the glycosyl residue: 3'6-di-0- Me-D-Glcp-(P1 +4)-2,3-di-O-Me-~-Rhap-(a1-+2)-3-0-Me-L-Rhap-(al+ of M.leprae phenolic glycolipid 2. For example, the glycolipids obtained from both M. haemophilum and M. leprae contained 2-linked 3-0-Me-a-~-Rhap and 4-linked 2,3- di-0-Me-L-Rhap but attached in a reverse sequence. It is also suggested that this relationship may prove extremely useful in explaining the nonavailability and pathogenicity associated with M. leprae. Development of an efficient strategy for the oligosaccharide preparation of a mycobacterium genus is a major objective of our laboratory.' Although a plethora of synthetic approaches for 0-glycosylation have been described with variable degrees of anomeric selectivity, we used Schmidt's trichloroacetimidate approach 'because of its simplicity and reliability.In addition, synthesis of the antigenic oligosaccharide was basic to an understanding of the structure-activity relationship necessary for the development of a synthetic antigen required for serodiagnosis and vaccination.' We described herein our synthetic studies on the oligosaccharide obtained from M. haemophilum. Results and Discussion The first stage in a route to the trisaccharide 3 involved preparation of the disaccharide derivative 12 containing a free 2'-hydroxy group.The glycosyl donor, namely, 2-0-acetyl-4-0- benzyl-3-0-methyl-~-rhamnopyranosyltrichloroacetimidate 9 was used for the two successive glycosylations and the presence of acetyl group at 0-2 was expected to provide a high degree of a-selectivity. Methyl 4-0-benzyl-3-0-methyl-a-~-rhamnopyranoside6was synthesised by a modified route in which the methyl 4-0-benzyl- a-L-rhamnopyranoside 4 was first transformed into the corresponding dibutyltin acetal derivative 5 by using dibutyltin oxide in refluxing benzene for 5 h followed by treatment with methyl iodide in the same solvent at reflux temperature for an additional 5 h to afford 6 (90%).lo Hydrolysis of the 0-glycoside bond by heating of the compound with 1.5 mol dm-3 sulfuric acid and dioxane at 100°C for 3 h and then conventional acetylation with acetic anhydride, pyridine and dimethyl- aminopyridine (catalytic) at room temperature gave the diacetate 7' obtained as a mixture of a-and P-anomers (7 :3; 65%). With a view to removing the acetyl group at the anomeric position, 7 was treated with freshly prepared tributyltin ethoxide in refluxing 1,2-dichloroethane for 3 h to give 8(93%).Subsequently, 8 was converted ' into the trichloroacetimidate derivative 9 by employing trichloroacetonitrile, 1'8-diazabi-cyclo[5.4.0]undec-7-ene in methylene dichloride at room temperature for 30min. The coupling reaction between 9 and the methyl 2,3-di-0- methyl-a-L-rhamnopyranoside' 10 in the presence of a catalytic amount of boron trifluoride-diethyl ether and 4 A molecular sieves in methylene dichloride at 0 "C afforded the disaccharide 11 (80%).The 'H NMR spectrum of 11 revealed anomeric protons at 6 4.72 (d, J1,21.2 Hz) and 5.12 (d, J1,,2, 1270 J. CHEM. SOC. PERKIN TRANS. I 1993 OR OAc 7 R=Ac "I8 R=Hc0 R = C(=NH)CC13 OM8 OMe OMe viii OMe HC?+ OM8 OMe 10 Scheme 1 Reagents and conditions: i, Bu,SnO, C6H6, heat, 5 h; ii, MeI, Bu,NI, C6H,, heat, 5 h; 111, 1.5 mol dm-3 H,SO,, dioxane, heat, 3 h, then Ac,O, Py,DMAP, 24 h; iv, Bu,SnOEt, CICH,CH,CI, heat, 3 h; v, DBU, CCI,CN, CH,CI,, 30 min; vi, 10, BF,.OEt,, CH,CI,, 0 O, 30 min; vii, NAOMe, MeOH, 2 h; viii, 9, BF,-OEt,, CH,CI,, 0 O, 30 min; ix, NaH, MeI, THF, 3 h; x, 10% Pd-C, H,, MeOH, 1 atm, 20 h.1.5 Hz), whilst the remaining protons had chemical shifts as expected. The partially decoupled "C NMR spectrum of 11 provided satisfactory stereochemical information since the observed coupling constants (JC-l,H-l167.0 and JC-l',H-l'173.0 Hz) were greater than 165 Hz, an indication l4 of a-configurations at both anomeric centres. Zemplen deacetylation of 11 gave the requisite disaccharide 12 containing a 2'-OH group free for the next coupling reaction. Compound 12 was also hydrogenolysed in the presence of 10% palladium on charcoal in methanol at normal temperature and pressure to give 13 which constituted the inner segment of the oligosaccharide framework of M. haemophilum. The structure of 13 was confirmed by 'H NMR and partially decoupled '3C NMR spectral studies.The second coupling reaction of 12 with 9 under the same conditions, as described earlier, provided the trisaccharide 14 (70%). In the 'H NMR spectrum of 14, three distinct singlets due to 1-H, 1'-H and 1"-H were located at 6 4.66, 5.04 and 5.13. Resonances characteristic of methoxy, benzyloxy and acetoxy groups appeared in the region expected of the assigned structure. However, further proof for 14 was gleaned from the 13C NMR spectrum in which chemical shifts at S 98.3 (C-1, Jc-1,l-H165.0 HZ), 98.5 (C-1, Jc-1*,1*-H171.4 Hz) and 100.4 (C-1 ", ,e-H 171.5 Hz) confirmed aconfigurations. Removal of the acetyl group in 14 under Zemplen conditions gave 15 whose structure was also scrutinised by 'H NMR, 13C NMR and partially decoupled '3CNMR spectral studies.The methylation of 15 with sodium hydride-methyl iodide in dry tetrahydro- furan at room temperature gave 16 which was finally de- benzylated by hydrogenation over palladium-on-charcoal in methanol at normal temperature and pressure to give the crystalline trisaccharide 3 in whose NMR spectral analysis, resonances due to anomeric protons were located at 6 4.66, 5.08 and 5.16 whilst anomeric carbons appeared at 6 98.1 (C- 1, Jc-l.l.H 167.0 Hz) and 100.7 173.0 Hz), 98.4 (C-1', Jc-ls,ls-H (C-I", Jc-l,.,l,,-H174.0 Hz). In addition the chemical ionisation mass spectrum showed a molecular ion peak at m/z 540 (M+). Experimental General Methods.-NMR spectra were recorded on a Varian Gemini 200 MHz spectrophotometer with tetramethylsilane as internal indicator.Chemical ionisation mass spectra were recorded on a VG MICROMASS 7070H instrument with acetone as ionisation molecule. Optical rotations were deter- mined on a JASCO DIP 360 digital polarimeter and are expressed in units of lo-' deg cm2 g-'. Methyl 4-0-Benzyl-3-O-methyl-a-~-rhamnopyranoside6.-A solution of methyl 4-O-benzyl-a-~-rhamnopyranoside4 (3.0 g, 11.2 mmol) dibutyltin oxide (3.34 g, 13.4 mmol) and benzene (100 cm3) was heated under reflux with azeotropic removal of water for 5 h. The solution was cooled to room temperature and methyl iodide (2.13 g, 15.0 mmol) and tetrabutylammonium iodide (1.5 g) were added to it. The reaction mixture was then further heated under reflux for 5 h, cooled to room temperature, J.CHEM. SOC. PERKlN TRANS. 1 1993 washed with water, dried and concentrated. The residue was chromatographed on silica gel by eluting with ethyl acetate- light petroleum (1 :4), to afford pure 6 (2.85 g, 90%), isolated as a syrup: [a]D -81.1 (c 0.6, chloroform) {lit.,” CarlD-83 (chloroform)}; S(CDCl,, 200 MHz) 1.21 (3 H, d, J5.66.5, 6-, 6’-, 6”-H), 3.36,3.48 (6 H, 2 s, 2 x OMe), 3.36 (1 H, t, J3,4= J4,5 90,4-H), 3.50(1 H,dd, J2.34.0, J3,49.0,3-H),4.04(1 H,dd, J1.2 1 .O, J2.3 4.0,2-H), 4.62,4.86 (2 H, 2d, J 12.0, PhCH2), 4.68 (I H, br s, I-H) and 7.3 (5 H, m, Ph). 1271 isolated as a syrup: [a]D -73.8 (c 1.41, chloroform); G(CDC13) 1.26 (3 H, d, J5,6 6.5, 5-CH3), 1.30 (3 H, J5*.6,6.5, 5’- CH,), 2.40 (1 H, br s, OH), 3.38, 3.46, 3.50, 3.52 (12 H, 4 s, 4 x OMe),4.62,4.88(2H,2d,J11.6,PhCH2),4.72(1H,d,J1.2 2.0, 1-H), 5.10 (1 H, d, J,.,,.2.1, 1‘-H) and 7.35 (5 H, m, Ph); 6 97.6 (C-I, JC-l,l-H 173.0).165.0) and 98.7 (C-1’, Jc-l,,13H Methyl 2,3-Di-0-methyl-4-0-(3-O-methyl-a-~-rhamnopyran-osy1)-a-L-rhamnopyranoside13.-A solution of 12 (0.10 g, 0.219 mmol), 10% palladium-on-charcoal (20 mg) in methanol (5 cm3) was stirred under an atmosphere of hydrogen at normal 1,2-Di-O-acetyl-4-O-ben~yl-3-O-methyl-a,~-~-rhamnop~~ran-ose 7.-A mixture of compound 6 (2.85 g, 10.1 mmol), dioxane (10 cm3) and 1.5 mol dm-’ sulfuric acid (10 cm3) was heated on a boiling water-bath for 3 h after which it was neutralised with solid barium carbonate.The solid was filtered off and washed with methanol and the combined filtrate and washings were concentrated, traces of moisture being removed by co-distillation with toluene. Acetic anhydride (3 cm3), pyridine (10 cm3) and 4.4-dimethylaminopyridine(30 mg) were added to the residue and the mixture stored for 24 h. Ice- water (5 cm3) was added to the reaction mixture which was then extracted with ethyl acetate. The ethyl acetate layer was washed with 1 mol dm-, HCI, water, aqueous sodium carbonate and water and then dried. The residue obtained after concentration was purified by column chromatography on silica gel with ethyl acetate-light petroleum (1 :9) to give 7(2.3 g, 65%) (a :P = 7 :3) isolated as a syrup: G(CDCl,, 200 MHz) 1.30 (d, J5,66.5, 5a- CH,), 1.34 (d, J5,66.5, SP-CH,), 2.12, 2.18 (2 S, a-CH3C02), 2.10, 2.22 (2 s, P-CH3C02), 3.42 (s, P-OMe), 3.44 (s, a-OMe), 4.62, 4.88 (2 H, 2d, J 12.0, PhCH2), 5.26 (m, 2a-H), 5.50 (m, 2P-H), 5.68 (s, IP-H), 5.94 (d, J 1.0, la-H) and 7.3 (5 H, m, Ph).Methyl 2,3-Di-O-methyl-4-0-(2-O-acetyl-4-0-benzyl-3-0-methyl-a-L-rhamnopyranosy1)-a-L-rhamnopyranoside 11.-A solution of compound 7 (2.3 g, 6.51 mmol), tributyltin ethoxide (2.5 cm3) and l72-dich1oroethane (20 cm3) was heated under reflux for 3 h and then concentrated. The residue was purified on silica gel by eluting with ethyl acetate-light petroleum (2: 3) to give 8 (1.95 g) which was treated with 1,8-diazabicyclo- C5.4.01undec-7-ene (0.9 cm3) and trichloroacetonitrile (1.5 cm3) in methylene dichloride (20 cm3) at room temperature for 30 min.The solution was passed through a small column of silica gel and eluted with methylene dichloride to give 9 (2.75 g, 93%). Compound 9 (2.75 g, 6.0 mmol) was added to a stirred solution of methyl 2,3-di-O-methyl-a-~-rhamnopyranoside 10 (0.83 g, 40 mmol), 4 A molecular sieves (2.0 g) in dichloromethane (30 cm3) and the mixture cooled to 0°C. Boron trifluoridediethyl ether (50 mm3) was added to the mixture which, after 30 min, was quenched with triethylamine (0.1 cm3). The solution was filtered, washed with water, dried and concentrated. The residue was chromatographed on silica gel by eluting with ethyl acetate-light petroleum (1 :4), to give 11 (1.6 g, 80%), isolated as a crystalline solid, m.p.125-127 “C; [a]D -53.2 (c 1.2, chloroform) (Found: C, 60.0; H, 7.5. Cak. for C25H38010: C, 60.2; H, 7.6%); GH(CDC13) 1.29 (6 H, d, J 6.5, 5,5’-CH3), 2.12 (3 H, S, CH,CO2), 3.37, 3.44, 3.46, 3.50 (12 H, 4 s, 4 x OMe), 4.61, 4.91 (2 H, 2d, J 11.7, PhCH,), 4.72 (1 H, d, J 1.2 I .2, I -H), 5.I2 (1 H, d, J1,,2, 1.5, I ’-H), 5.40 (I H, dd, J28.3, 4.0, 2‘-H) and 7.3 (5 H, m, Ph); 6, 98.1 (C-I, J,-,,,-, 167.0) and 99.4(C-1’, Jc-l,.l-H173.0). pressure and room temperature for I2 h after which the catalyst was filtered off and the filtrate concentrated. The residue was purified by column chromatography on silica gel by using ethyl acetate-hexane (9:l) to give 13 (0.075 g, 93%), isolated as a syrup; [a]D -65 (c 1.o, chloroform) (Found: C, 52.15; H, 8.0.Calc. for C16H3009: C, 52.45; H, 8.2%); G,(CDCl,) 1.28 (6 H, d, J6.0, 5,5‘-CH,), 3.34, 3.42, (6 H, 2s, 2 x OMe), 3.48 (6 H, s, 2 x OMe),4.04(1 H, m, 2-H), 4.66(1 H, d, J1,21.0, 1-H), 5.14 (1 H, d, J1,,2, 165.0) and 101.11.0, l’-H);& 98.0 (C-I, Jc-l.l-H (C-l’, Jc_1*.1,-H 170.2). Methyl 2,3-Di-O-methyl-4-0-[4-O-benzyl-3-O-methyl-2-0-(2-O-acetyl-4-O-benzyl-3-O-methyl-a-~-rhamnopyranosyl)-a-~-rhamnopyranosyl]-a-L-rhamnopyranoside14.-Boron trifluor-ide-diethyl ether (30 mm3) was added to a stirred solution of 12 (1.O g, 2.19 mmol), 9 (I .5 g, 3.3 mmol) and 4 A molecular sieves (2.0 g) in dichloromethane (20 cm3) at 0 “C. After 30 min the reaction mixture was worked-up and the residue chromato- graphed on silica gel with ethyl acetate-light petroleum (1 :3) as eluent to afford 14 (1.15 g, 70%), isolated as a syrup; [alD -57.3 (C 2.48, chloroform): d~(cDC13) 1.20, 1.26, 1.30 (9 H, 3d, J6.4, 5,5’,5”-CH3), 2.15 (3 H, S, CH,CO2), 3.32, 3.38, 3.43, 3.46, 3.48 (15 H, 5s, 5 x OMe), 4.58,4.60,4.83,4.86 (4 H, 4d, J 11.5, 2 x PhCH,), 4.66 (I H, S, I-H), 5.04 (1 H, S, 1‘-H), 5.13 (1 H, s, 1”-H), 5.40 (1 H, m, 2”-H), 7.3 (10 H, m, 2 x Ph); S, 98.3 (C-I, Jc-l,l-H 165.0), 98.5 (C-l‘, Jc-le,lr.H171.4) and 100.4 (C-1”,Jc-l,,,1 r3-H 17 1.5).Methyl 2,3-Di-O-methyl-4-0-[4-O-benzyl-3-0-methyl-2-0-(4-O-benzyl-3-O-methyl-a-~-rhamnopyranosyl)-a-~-rhamnopyr-anosyll-a-L-rhamnopyranoside15.-Compound 14 (0.70 g, 0.94 mmol) was dissolved in methanol (10 cm3) and sodium (40 mg) was added to the solution.After 2 h at room temperature, the reaction mixture was neutralised with Amberlite IR 120 (H) resin, filtered and concentrated. The residue was purified on a column of silica gel with ethyl acetate-light petroleum (2 :3) as eluent to give 15 (0.62 g, 9473, isolated as a solid, m.p. 51-53 “C; [a]D -64.4 (c 1.5, chloroform) (Found: C, 63.3; H, 7.45. Cak. for C37H54013: C, 62.9; H, 7.6%); &(CDCI,) 1.3 (9 H, m, 5-, 5’-, 5”-CH,), 3.37, 3.42, 3.49, 3.51, 3.54 (15 H, 5 s, 5 x OMe), 4.60,4.62,4.87,4.90 (4 H, 4 d, J 1 I .7,2 x PhCH,), 4.70 (d, J1,2 1.0, I-H), 5.14 (1 H, s, 1’-H), 5.19 (d, J1.,e2,,1.0, 1”-H) and 7.3 (10 H, m, 2 x Ph); 6, 98.5 (GI, JC-l,l-H 166.0), 100.5 (C-1’ + C- 1 ”,J 169.0).Methyl 2,3-Di-0-rnethyl-4-0-[4-O-benzyl-3-O-methyl-2-0-(4-0-benzy f-2,3-di-O-methyl-a-~-rhamnopyranosyl)-a-~-rham-nopyranosyo-a-L-rhamnopyranoside 16.-Methyl iodide (0.3 cm3) was added to a solution of compound 15 (0.40 g, 0.56 mmol) sodium hydride (0.15 g) and dry tetrahydrofuran (5cm3) and the Methyl 2,3-Di-O-methyl-4-0-(4-O-benzyl-3-O-methyl-a-~-mixture stirred for 3 h at room temperature. Methanol (1 cm3) rhumnopyranosj4)-a-L-rhamnopyrunoside12.-Sodium (0.1 g) was added to a solution of 11 (1.6 g, 3.2 mmol) in methanol (20 cm3) and the mixture stirred at room temperature for 2 h. It was then deionised with Amberlite IR 120 (H) resin, filtered and concentrated. The residue was purified on silica gel by using ethyl acetate-light petroleum (2:3) to give 12 (1.39 g, 95%), was then added to it to decompose sodium hydride after which it was concentrated and partitioned between water-ethyl acetate (I :1;60cm3). The ethyl acetate layer was dried and concentrated and the residue chromatographed on silica gel with ethyl acetate-light petroleum (1 :2) as eluent to give 16 (0.38 g, 9379, isolated as a syrup: [aID -78.5 (c 0.6, chloroform): dH(CDC1,) 1272 J.CHEM. SOC. PERKIN TRANS. I 1993 1.24, 1.27, 1.30 (9 H, 3 d, J6.0,5-, 5f-,5”-CH3), 3.35, 3.40,3.48, Part A, eds. G. B. Kubica and G. L. Wayne, Marcel Dekker, New York, 1984,467.3.50 (12 H, 4 s, 4 x OMe), 3.56 (6 H, s, 2 x OMe), 4.59, 4.61, 2 G. T. Valainis, I. M. Cardona and D. L. Greer, J.Acquired Immune 4.85,4.89(4H,4d,J10.6,PhCH2),4.78(1H,d,J1,21.5,l-H),Defic. Syndr., 1991,4, 5 16.5.12 (2 H, d, J 1.2, 1‘-H, 1”-H) and 7.3 (10 H, m, 2 x Ph). 3 M. K. Gurjar and G. Viswanadham, Tetrahedron Lett., 1991, 32, 6 194. 4 S. B. Gurdyal, M. McNeil, E. M. David, P. Francoise, M. Ridell and Methyl 2,3- Di-O-methyl-4-0-[3-O-methyl-2-0-(2,3-di-O-methyl-a-~-rhamnopyranosyl)-a-~-rhamnopyranosyl]-a-~-rham-P. J. Brennan, Biochemistry, 1991,30, 7772. 5 M. K. Gurjar and K. Revathi Reddy, Carbohydr. Rex, 1992,226,233;nopyranoside 3.-A solution of compound 16 (0.30 g, 0.41 mmol), 10% palladium-on-charcoal (30 mg) in methanol (5 cm3) was stirred under an atmosphere of hydrogen at normal pressure and room temperature for 20 h. The residue was purified by column chromatography on silica gel by using ethyl acetate-methanol (30:l) to give crystalline solid 3 (0.195 g, 8679, m.p.158-159 “C; [aID -97.3 (c 1.0, chloroform) (Found: C, 52.9; H, 8.0. Calc. for C24H440,3: C, 53.3; H, 8.1%); d~(cDCl3) 1.26, 1.28, 1.30 (9 H, 3 d, J 6.5, 5-, 5f-, 5”-CH3), 3.36,3.44(6H,2s,2 x OMe),3.48(12H,brs,4 x OMe),4.66 (1 H,s, 1-H),5.08(1 H,s, 1’-H)and5.16(1 H,s, l”-H);6,98.1 (C-1, Jc-l.l-H 167.0) and 100.7 (C- 173.0), 98.4 (C-lf, Jc-13,1,-H l”, Jc l,e.l,,.H 174.0);m/z (CIMS) 540 (M+). References 1 P. J. Brennan, Microbial Lipids, 1988, vol. 1, Academic Press, London, p. 203; P. J. Brennan, in The Mycobacteria. A Source Book, M. K. Gurjar, M. K. and A. S. Mainkar, Tetrahedron, 1992,48,6729; M. K. Gurjar and P. S. Mainkar, Carbohydr. Rex, 1993,293, 297. 6 H. Paulsen, Angew. Chem., Int. Ed. Engl., 1982,21, 155. 7 R. R. Schmidt, Angew. Chem., Int. Ed. Engl., 1986,25,212. 8 D. Chatterjee, S. N. Cho, C. Stewart, J. T. Douglas, T. Fujiwara and P. J. Brennan, Carbohydr. Res., 1988, 183, 241. 9 V. Pozsgay, Carbohydr. Res., 1969, 10,466. 10 V. Pozsgay, Carbohydr. Rex, 1979,69,284. 11 T. Fujiwara and S. Izumi, Agric. Biol. Chem., 1987,51,2539. 12 M. K. Gurjar and U. K. Saha, Tetrahedron Lett., 1992,33,4979. 13 K. Bock, T. Hvidt, J. Marino-Albernas and V. Verez-Bencomo, Carbohydr. Rex, 1990,200,33. 14 V. Pozsgay and A. Neszmely, Carbohydr. Rex, 1980, 80, 196; R. Kasai, M. Okihara, J. Asakawa, K. Mizuyani and D. Tanaka, Tetrahedron, I979,35, 1427. Paper 3/0009 1E Received 6th January 1993 Accepted 25th February 1993
ISSN:1472-7781
DOI:10.1039/P19930001269
出版商:RSC
年代:1993
数据来源: RSC