J. Am. Chem. SOC.1992, 114, 3499-3505 /EtOAc) (lit. mp 113 oC);2''HNMR 6 9.04(d, 1 H,J = 2.4 Hz), 8.36 (dd, 1 H,J = 2.4, 8.7 Hz),7.57 (d, 1 H,J = 8.7 Hz), 3.08 (q, 2 H,J = 7.3 Hz),1.46 (t, 3 H,J = 7.3 Hz); I3C NMR 6 147.29,144.59,143.60, 126.98,126.75, 121.65,26.71, 12.44;IR (film) 3098,2986, 1589,1516, 1451, 1344, 1308, 1246, 154, 1098, 1053,921,832,734 cm-l; GC-MS m / e (%) 228
3499
ministered by the American Chemical Society, the Herman Frasch Foundation, the National Science Foundation, and SociEtE Nationale Elf Aquitaine. Registry No. la, 81788-92-3;lb, 139631-56-4;IC, 139631-57-5;Id, 139631-59-7;2a, 64251-85-0;Zb, 33945-99-2;2c, 139631-58-6;M , 76832-59-2;3, 139631-66-6;3 4 139631-71-3;3b, 139631-72-4;4, 6863-32-7; 5, 139631-60-0;6,68846-57-1;7,139631-61-1; 8, 13963162-2;9,139631-63-3;10,74379-74-1; 11, 139631-65-5; 12, 139631-68-8; 13, 139631-69-9; 14, 139631-70-2; 15,7343-55-7;18, 139631-64-4; 19, sulfene, 917-73-7;bromocyclopropane, 4333-56-6. 13963 1-67-7;
(M+, 59), 200 (16), 184 (30),183 (loo), 137 (16), 106
(20), 95 (28), 91 (17), 79 (28). 77 (16), 69 (38), 63 (52), 62 (16).
Acknowledgment. We thank Shu Hai Zhao and Mohan Thiruvazhi for experimental assistance and gratefully acknowledge support from the donors of the Petroleum Research Fund, ad-
Spin Coupling Constants in Aldoses Enriched with 1 3 C at the Terminal Hydroxymethyl Carbon: Effect of Coupling Pathway Structure on Jccin Carbohydrates 13C-13C
Jian Wu,t Paul B. Bondo,t Tapani Vuorinen,s and Anthony S.Serianni*,t Contribution from the Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana 46556, Omicron Biochemicals, Inc., 19882 Alou Lane, South Bend, Indiana 46637, and Laboratory of Wood Chemistry, Helsinki University of Technology, SF-02150 Espoo, Finland. Received July 31, 1991
Abstract: Eight aldohexoses (allo, altro, galacto, gluco, gulo, ido, manno, talo), four aldopentoses (arabino, lyxo, ribo, xylo), and two aldotetroses (erythro, threo) have been prepared with "C-enrichment (99 atom-% I3C) at the terminal hydroxymethyl (CH20H) carbon. High-resolution IH-decoupled ')C NMR spectra were obtained at 75 and 125 MHz in order to obtain one-bond (lJCc)and longer range (2Jcc, 3Jcc) l3C-I3C spin coupling constants involving the terminal carbons of the more abundant furanose and pyranose forms of these monosaccharides in 2H20. In some cases spectral interpretation was assisted by the use of one-dimensional INADEQUATE I3C spectra. The effect of aldopyranose and aldofuranose ring structure and conformation on the magnitudes of these couplings, especially 2Jccc and 'JccCc,was probed. Results show that 'JCC? is highly affected by the orientation of terminal hydroxyl substituents along the C-C-C coupling pathway and that 3Jcccc is not only affected by molecular dihedral angle (Le., Karplus relationships) but also by substituent geometry along the C-C-C-C coupling pathway.
need is particularly acute for .Icc, since it is well recognized that relatively subtle changes in structure along the coupling pathway may dramatically affect their magnitudese8 Seminal studies in non-carbohydrate systems conducted by Barfield and co-worke r ~ ~ -have " clearly shown that Jcc depends highly on pathway
Introduction In recent years, nuclear magnetic resonance (NMR) spectroscopy has emerged as a powerful tool to investigate the structures and conformational features of biologically-important molecules in solution. The development of multidimensional modes of data collection'v2 has played a dominant role in this regard, especially in studies of macromolecules such as proteins and nucleic acids. Implicit in these new methods is the fundamental assumption that an intelligent integration of different N M R parameters can lead to more reliable models of solution behavior. For example, the combined use of IH-lH spin couplings (JHH) and noes, measured from COSY and NOESY spectra, respectively, has been important in computer-aided three-dimensional structure determinations of protein^.^,^ Thus, studies aimed a t an improved understanding of how specific N M R parameters are affected by molecular structure and dynamics are critical to the development of NMR-based strategies to probe the solution properties of molecules. While numerous N M R studies of carbohydrates have used IH-IH spin couplings to assess molecular structure and conformation: interest in I3C-IH (JCH) and 13C-'3C (.Ic spin c) couplings is increasing, partly because modern N M R methods are available that permit their measurement without the need for %enrichmenL6v7 Thus, while the problem of measurement has been reduced, a real need exists for systematic investigations of the dependencies of these couplings on carbohydrate structure. This
(1)(a) Emst, R.R.; Bodenhausen, G.; Wokaun, A. Principles of Nuclear Magnetic Resonance in One and Two Dimensions; Oxford University Press: New York, 1987. (b) Pulse Methods in ID and 2D Liquid-Phase N M R Brey, W. S . , Ed.; Academic Press: New York, 1988. (2) (a) Vuister, G. W.; de Waard, P.; Boelens, R.; Vliegenthart, J. F. G.; Kaptein, R. J . Am. Chem. Soc. 1989, 1 1 1 , 772. (b) Fesik, S.W.; Gampe, R. T., Jr.; Zuiderweg, E. R. P. J . Am. Chem. SOC.1989,111,770.(c) Kay, L. E.; Clore, G. M.; Bax, A.; Gronenborn, A. M. Science 1990,249,411. (3)Wtithrich, K.NMR of Proteins and Nucleic Acids; John Wiley and Sons: New York, 1986. (4) (a) Bax, A. Ann. Rev. Biochem. 1989,58, 223. (b) Clore, G. M.; Gronenborn, A. M. Crit. Reu. Biochem. Mol. Biol. 1989,24, 479. (5)(a) For a general discussion of the use of JHH in the conformational analysis of cyclic compounds, see: Booth, H. Prog. NMR Spectrosc. 1969, 5 , 149. (b) Bentley, R.Ann. Rev. Biochem. 1972,41, 953. (6)(a) Bax, A,; Freeman, R. J . Am. Chem. SOC.1982,104, 1099. (b) Morat, C.; Taravel, F. R.; Vignon, M. R. Magn. Reson. Chem. 1988,26,264. (7) (a) Bax, A.; Freeman, R.;Kempsell, S. P. J . Am. Chem. SOC.1980, 102,4849.(b) Bax, A,; Freeman, R.; Frenkiel, T. A.; Levitt, M. H. J . Magn. Reson. 1980,43, 478. (8)Marshall, J. L. Carbon-Carbon and Carbon-Proton NMR Couplings: Applications to Organic Stereochemistry and Conformational Analysis; Verlag Chemie: Weinheim/Bergstrasse, Germany, 1983. (9) (a) Barfield, M.; Burfitt, I.; Doddrell, D. J. Am. Chem. SOC.1975,97, 2631. (b) Barfield, M.; Conn, S. A,; Marshall, J. L.; Miiller, D. E. J . Am. Chem. SOC.1976,98,6253. (10) Marshall, J. L.; Conn, S. A,; Barfield, M. Org. Magn. Reson. 1977, 9. 404.
t University of Notre Dame.
*OmicronBiochemicals, Inc. University of Technology.
1 Helsinki
0002-786319211514-3499%03.00/0 0 1992 American Chemical Society 1-
Wu et al.
3500 J . Am. Chem. SOC.,Vol. 114, No. 9, 1992
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PPM Figure 1. (A) The partial ‘H-decoupled ”C N M R spectrum (75 MHz)
of ~-[6-’~C]idose.(B-D) INADEQUATE I3C N M R spectra of L-[~-’)C]idose obtained with a 6 (B), 50 (C), and 100 ms (D) mixing time. Shorter mixing times enhance the detection of carbons directly bonded to the enriched site, while longer mixing times suppress the one-bond correlations and enhance the detection of coupled carbons further removed from the enriched site. Close inspection of data in (D) reveals small couplings to C3 of the a-furanose (-76.4 ppm) and to C4 of the a-pyranose (-71.7 ppm) that were not detected in the conventional I D spectrum in (A).
structure and that faulty interpretations can occur when these couplings are applied without a detailed knowledge of this dependency. In a previous study,’* we examined Jcc in the aldoses involving the C1 (anomeric) carbon. Results showed that the relative orientation of the hydroxyl substituents on the coupled carbons, and on carbons along the coupling pathway, significantly affects the coupling magnitude. For example, 2Jc1,c3 in aldohexopyranoses was found to be large (-4.3 Hz)in structures having 0 1 , C1, C2, C3, and 0 3 coplanar, and displacing either 0 1 or 0 3 out of this plane reduced this coupling to a small or zero value. 3Jc1,c6 was found to depend on the orientation of 0 1 with respect to the coupling pathway, with larger couplings observed when 0 1 , C1, 05,and C5 are coplanar.I2 While the orientation of 0 6 with respect to this coupling pathway is also likely to affect 3Jc1,c6in aldohexopyranoses, this effect could not be assessed due to the inherent rotational mobility of the exocyclic C5-C6 bond. The present investigation aims to extend this earlier work to Jcc involving the terminal hydroxymethyl carbons of the simple aldoses. (4-13C)Aldotetroses, (5-13C)aldopentoses, and (6-13C)aldohexoses have been prepared, high-resolution I3C NMR spectra at 125 MHz have been obtained and analyzed, and the I3C-l3C couplings between the terminal carbon and the remaining pyranosy1 and furanosyl ring carbons have been interpreted in terms of ring structure, configuration, and conformation.
(typically sine-bell or gaussian) to improve the detection of small couplings. Spectra were referenced externally to the C1 chemical shift of ~-~-(l-’~C)glucopyranose (97.4 ppm), and reported chemical shifts and I3C-”C spin couplings are accurate to f0.1 ppm and *O.l Hz, respectively. High-resolution ‘H N M R spectra (500 MHz) were collected on the same spectrometer at 21’ with sweep widths of 3400 Hz and 56K data points, giving a digital resolution of 0.061 Hz/pt. IH-IH spincoupling constants are accurate to f O . l Hz. One-dimensional INADEQUATE ”C N M R ~ p e c t r a ’ ~aJt~21° in * H 2 0 were obtained on a General Electric GN-300 FT-NMR spectrometer operating in the quadrature-phase mode. FIDs were processed with exponential multiplication (EM) to yield spectra with a digital resolution of 0.075 Hz/pt. Synthesis of (13C)-LabeledAldoses. Detailed procedures for the synthesis of the ”C-labeled aldoses used in this study have been published elsewhere,I3J4 and only a brief account of the methods is given here. ~-(6-”C)Glucosewas prepared from 1,2-0-isopropyIidene-a-~-xylopentodialdo-l,4-furanoseand K ’ T N as described by King-Morris et ~-(6-”C)Mannosewas obtained from ~ - ( 6 - ~ ~ C ) g h c obys emolybdatecatalyzed C2-epimeri~ation.’~ o-(5-I3C)Ribose and ~-(5-’~C)arabinose were prepared by treatment of D-(6-13C)ghcose with P b ( 0 A ~ ) ~to l ’ give ~-(4-’~C)erythrose, which was chain-extended by cyanohydrin r e d u c t i ~ n ’ ~ Jto~ give J ~ the target labeled pentoses. ~-(4-”C)Erythrose was epimerized with sodium molybdateI6 to yield, after chromatography, ~-(4-’~C)threose.~ 4 4 ”C)Threose was chain-extended by cyanohydrin reduction”J4J8 to give, after chromatography, ~-(5-”C)xyloseand D-(5-”C)lyXOSe. ~-(6-”C)Idose was prepared from 1,2-O-isopropylidene-a-~-xy~opentodialdc-l,4-furanoseand K”CN as described by King-Morris et all5 ~-(6-’~C)Gulose was obtained from ~-(6-’)C)idoseby C2-epimerization with sodium molybdate.I6 ~ - ( 6 - ~ ~ C ) A l land o s e ~ - ( 6 - ~ ~ C ) t a lwere o s e prepared as described by King-Morris et aLt5for the preparation of ~ - ( 6 - ~ ~ C ) g l u c by o s esubstituting 1,2-O-isopropylidene-cu-~-ribo-pentodialdo-1,4-furanose for 1,2Oisopropylidene-cu-D-xylo-pentodialdo-1,Cfuranose as the starting sugar in the cyanohydrin reduction reaction. Epimerization of purified ~ - ( 6 ’3C)alloseand ~ - ( 6 - ~ ~ C ) t a lwith o s e sodium molybdatet6 gave D-(6-I3C)altrose and ~-(6-~~C)galactose, respectively. The labeled aldoses were identified by ”C N M R spectroscopy using chemical shift data reported previously.I2
Results and Discussion A. Solution Properties of the Aldoses. Aqueous solutions of the c4
