Further Studies on the Assignment of Signals in 13CMagnetic Resonance Spectra of Aldoses and Derived Methyl Glycosidesl

PHILIP A. J. GORINAND MYTOSK MAZUREK

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Prairie Regional Laboratory, National Research Council of Canada, Saskatoon, Saskatchewan S7N OW9 Received October 9,1974

PHILIPA. J. GORIN and MYTOSK MAZUREK. Can. J. Chem. 53,1212(1975). The I3C signals of most of the more common sugars and their methyl glycosides have been assigned, based on the a-carbon and a-carbon effects that occur on deuterium substitution. For pyranoses having the same hydroxyl configuration, replacement of the CH,OH substituent at C-5 with CH,, CO,H, CO,Me, or H is accompanied by displacement of the C-4, C-5, and when applicable C-6 signals. The magnitudes of the displacements in the glucopyranose and mannopyranose series are almost identical and were of aid in assigning signals in the 13Cspectrum of methyl (methyl a-D-mannopyranuronosid)uronate.The displacements, however, differ from corresponding ones observed in the galactopyranose series. Some assignments are made for the I3C signals of structurally related methyl aldofuranosides.
A. J. GORIN et MYTOSK MAZUREK. Can. J. Chem. 53,1212(1975). PHILIP On a attribue les signaux en r.m.n. du 13C de la plupart des sucres usuels ainsi que de leurs glycosides de methyle; ces attributions ont Cte faites en se basant sur les effets que produisent les carbones a et des substitutions par du deuterium. Pour des pyrannoses ayant la m&me configuration des groupes hydroxyles, le remplacement d'un substituant C H 2 0 H en C-5 par CH3, C 0 2 H , C0,Me ou H s'accompagne par un dkplacement des signaux en '2-4, C-5 et lorsque c'est possible en C-6. L'amplitude de ces deplacements dans les series glucopyrannoses et mannopyrannoses sont presque identiques et a ete fort utile pour attribuer les signaux dans le spectre r.m.n. du 13C du (a-D-mannopyrannuronoside de m6thyle)uronate de mithyle. Les dCplacements different toutefois des deplacements correspondants observCs en serie galactopyrannose. On a fait quelques attributions pour des signaux de I3C dans des aldofurannosides structurale. [Traduit par le journal] de methyle qui leur sont relib d'une f a ~ o n

In a previous study (I), the carbon-13 magnetic resonance (I3c.m.r.) spectra of deuterated derivatives of ap-D-glucose, ap-D-mannose, and ap-D-galactose were interpreted and discussed. With the aid of two suitable deuterated derivatives of each aldose it was possible to make an unambiguous assignment of each signal, since the signals of the 13C nucleus attached to the deuteron (a-carbon) disappeared and those of the adjacent P-carbons were displaced upfield by 0.02-0.10 p.p.m. (1). Since some of the assignments carried out elsewhere have proved to be incorrect, the studies in this laboratory have been extended to most of the frequently encountered monosaccharides and certain derived methyl glycosides. The preparation of the deuterated sugars was rendered easier, in most instances, by synthetic routes previously devised for the undeuterated counterparts. As in the above study, which was relatively brief, two deuterated derivatives of each sugar were selected and their 13c.m.r. spectra ration'NRCC No. 14432.

alized by observing the expected a-carbon (2) and the P-carbon deuterium isotope effects (1) (Table 1). Although, for the most part, the signal of the' a-carbon disappears, in the case of 5 - 2derivatives ~ of pentoses, or the 6-2H derivatives of 6-deoxyhexoses and methyl 6-deoxyhexopyranosides, the a-13C signals appear as triplets at 0.4 p.p.m. upfield (3). The P-carbon effect appeared as an upfield shift of 0.04-0.09 p.p.m. on monodeuteration, 0.07-0.12 p.p.m. on dideuteration, and 0.17 p.p.m. for the C-3 signal The - ~ ~signals ,. of of p - ~ - a r a b i n o ~ ~ r a n o s e - 4 , 5 a- and P-anomers of reducing sugars were distinguished by following the mutarotation process by 13c.m.r. spectroscopy, and those of isomeric methyl glycosides, which were components of mixtures, were obtained by reference to appropriate standards. The assignments made by these approaches are summarized in Tables 2-6. The results obtained on the methyl glycosides can be used in interpreting the 13c.m.r. spectra of polysaccharides and other sugar-containing polymers (4). Also, the following discussion will show that

GORIN AND MAZUREK: N.M.R. STUDIES OF ALDOSES

1213

TABLE 1. Upfield displacement of 13C-C-'H and 13C-'H signals of sugars* compared with those of corresponding I3C-C-'H and 13C-'H resonances, respectively, in p.p.m. Sugar Upfield displacement in p.p.m.t
-

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aB-~-Rhamnose-6-'H Methyl a~-~-xylopyranoside-3-'H Methyl ap-~-xyIopyranoside-5-'H Methyl a(3-o-arabinopyranoside-5-'H Methyl a(3-~-arabinopyranoside-4,5-'H~ Methyl up-D-galactopyranoside-4-'H Methyl a~-~-galactopyranoside-6-'H~ Methyl a~-~-ribofuranoside-3-'H Methyl aD-~-ri bofuranoside-5-'H Methyl ae-D-arabinofuranoside-pH Methyl a!3-~-arabinofuranoside-4,5-'H, Methyl ae-D-galactofuranoside-4-'H Methyl a~-~-galactofuranoside-6-'H~ Methyl 6-deoxy-a!3-~-galactopyranoside-4-'H Methyl 6-deoxy-a~-~-galactopyranoside-6-'H Methyl 6-deoxy-a~-~-glucopyranoside-6-'H Methyl 6-deoxy-ae-D-gl~copyranoside-4-~H Methyl Methyl Methyl Methyl Methyl Methyl a-L-rhamnopyranoside-4-'H a-L-rhamnopyranoside-6-'H up-D-glucopyranosiduronicacid-2-'H US-D-glucopyranosiduronic acid-5-'H (methyl a(3-D-glucopyranosid)uronate-pH (methyl ap-D-glucopyranosid)uronate-5-'H

C,-4(0.07), Cp-4(0.07), C.-5(0.42; J 20 HZ), Cp-5(0.37; J 23 HZ) Cm-4(0. lo), C,-4(0.12) C.-3(0. lo), C,-3(0.17) C=-3(0.08), C,-3(0.06), Cm-5(0.06) CB-5(0 .06) Cm-5(0. 04), CB-5(0. 04), Cm-6(0. 50; J 20 Hz), CB-6(0.50, J 20 HZ) Cm-5(0. 05), Cp-5(0.06) Cm-3(0. 08), C,-3(0.07), Cm-5(0. 06), Cp-5(0 .06) Cm-3(0.06),Cp,-3(0.05), Ca-5(0.05), Cp-5(0. 06) C.-5(0.09), Cp-5(0.08), Cap-6(0.41,J 18 HZ) Ca-4(0.06), CB-4(0. 06)
-

Ca-3(0.08), C,-3(0.09) Cm-3(0. 08), C,-3(0.08), Cm-5(0. 06), C,-5(0.06) Ca-5(0.12), CD-5(0. 1 1)
-

Ca-4(0.04), C,-4(0.04) Cm-3(0. 04), C,-3(0 04) C.-3(0. 04), C,-3(0 05), Cm-5(0. 08), CB-5(0. 06) Ca-5(0. 12), C,-5(0 12) C.-3(0.08), C,-3(0 07), Cm-5(006), Cp-5(0.05) C.-5(0.04), C,-5(0 05) C.-5(0.04), C,-5(0. 04) Cm-3(0. 05), Co-3(0 06), Cm-5(006), C,-5(0.05) C.-3(0. 08), Cp-5(0. 06) C.-5(0.05), Cap-6(0. 32; J 0 68) C.-3(0.04), Cp-3(0.05) C.-4(0. 07), C,-4(0.07) C,-3(0 06), C,-3(0.06) C.-4(0.06), Co-4(0.06)

*a and i3 subscripts refer to C-1 configuration of sugar. tunless otherwise stated, the signals of carbons attached to deuterium disappear.

signal displacements observed on changing certain substituents in structurally related sugars are consistent, so that they may be of aid in interpreting the spectra of other sugars. The chemical shifts of signals of 13C nuclei of methyl a-D-glucopyranoside may be compared with those of respective nuclei in methyl a-Raldopyranosides having a C1 conformation and the same configuration at C-2, C-3, C-4, and C-5 (Table 2). On going from methyl a-D-glucopyranoside to methyl 6-deoxy-a-D-glucopyranoside, methyl a-D-glucopyranosiduronic acid,

methyl (methyl a-D-glucopyranosid)uronate, and methyl a-D-xylopyranoside, marked differences of shift were not observed for the respective C-1, C-2, C-3, and OCH,-1 signals. Replacement of CH'OH at C-5 with CH, causes (2-4 and C-5 signals to be displaced at + 6 p.p.m. (downfield) and -4 p.p.m., respectively.' These figures are + 1.9 p.p.m. and -0.8 p.p.m., respectively, when comparisons are made with the spectra of
'Shifts are measured relative to external Me,Si, which is always upfield from observed signals.

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TABLE 2. Assignment of signals in I3C spectra of configurationally related sugars. The figures of the chemical shifts are in p.p.m., as are those in parentheses, which represent the difference of the shift from that of the corresponding carbon in a-D-glucose and methyl a-D-glucopyranoside a-D-Glucose Methyl a-D-glucopyranoside

92.7 93.3 (+0.6) 93.1 (+0.4)

72.14 72.5 (+0.4) 72.9 (+0.8)

a-D-Glucose (I) 73.4 70.4 a-D-Xylose 73.9 70.4 (+0.5) (0)

72.10 62.1 (-10.0)

61.3

100.3 100.6 (+0.3)

72.5 72.3 (-0.2) 72.6 (+O.l) 71.9 (-0.6) 71.94 (-0.6)

74.2

Methyl a-D-glucopyranoside 70.6 72.7 61.7

$
56.2 56.0 (-0.2) 56.2 (0) 56.7 (f0.5) 54.2
! -

Methyl a-D-xylopyranoside 74.3 70.4 62.0 (+O.l) (-0.2) (-10.7) Methyl 6-deoxy-a-D-glucopyranoside 73.9 76.2 68.7 17.6 (-0.3) (+5.6) (-4.0) (-44.1) Methyl a-D-glucopyranosiduronicacid 73.8 72.5 71.9 ? (-0.4) (+1.9) (-0.8)

E
P
C

6-Deoxy-a-D-glucose 73.6 76.4 68.6 (+0.2) (+6.0) (-3.5)

6
VI

18.0 (-43.3)

r
w
m *

100.3 (0) 100.7 (+0.4) 100.8 (+0.5)

VI

Methyl (methyl a-D-g1ucopyranosid)uronate 73.7 72.4 71.87 ? 56.8 (-0.5) (+1.8) (-0.8) (+0.6)

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TABLE 3. Assignment of signals in I3C spectra of configurationally related sugars. The figures of the chemical shifts are in p.p.m., as are those in parentheses, which represent the difference of the shift from that of the corresponding carbon in a-D-glucose and methyl P-D-glucopyranoside 13-D-Glucose
C-1 96.5 C-2 74.8 C-3 C-4 C-5 76.6 C-6 61.5 C-1 104.3 105.1 (+0.8) 96.8 (+0.3) 75.6 (+0.8) C-2 74.2 74.0 (-0.2) 74.5 (+0.3) 73.8 (-0.4) 73.7 (-0.5) C-3 76.9 76.9 (0)

Methyl a-D-glucopyranoside
C-4 C-5 C-6 OCHS-1 58.3 58.3 (0) 58.3 (0) 58.5 (+0.2) 58.7 (+0.4) 56.5 OCH3

a-D-Glucose(I) 76.4 70.3

Methyl a-D-glucopyranoside 70.8 76.9 61.9 Methyl P-D-xylopyranoside 70.4 66.3 (-0.4) (-10.6)

6-Deoxy-P-D-glucose 76.6 76.1 73.0 (+0.2) (+5.8) (-3.6)

18.0 (-43.5)

104.3 (0) 104.3 (0) 104.6 (+0.3)

Methyl 6-deoxy-P-D-glucopyranoside 76.7 76.2 73.0 17.8 (-0.2) (+5.4) (-3.9) (-44.1) Methyl a-D-glucopyranosiduronicacid 76.5 72.3 75.6 ? (+1.5) (-1.3) (-0.4) Methyl (methyl a-D-g1ucopyranosid)uronate 76.3 72.4 75.7 ? (-0.6) (+1.6) (-1.2)

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TABLE 4 . Assignment of signals in I3Cspectra of configurationally related sugars. The figures of the chemical shifts are in p.p.m., as are those in parentheses, which represent the difference of the shift from that of the corresponding carbon in a- and p-D-mannose and methyl a-D-mannopyranoside a - and p-D-Mannose Methyl a-D-mannopyranoside

a-D-Mannose (I)

Methyl a-D-mannopyranoside

95.0 95.0 ( 0 ) 94.6 94.6 ( 0 )

71.7 71.9 (+0.2) 72.3 72.4 (+O.l)

71.3 7 1 .I (-0.2) 74.1 73.8 (-0.3)

68.0 73.3 (+5.3) 67.8 72.9 (+5.l)

73.4 69.4 (-4.0) 77.2 73.1 (-4.1)

62.1 18.0 (-44.1) 62.1 18.0 (-44.1)

101.9 101.9 ( 0 ) 102.3 (+0.4)

71.2 71.0 (-0.2)

71.8 71.3 (-0.5)

68.0 73.1 (+5.1)

73.7 69.4 (-4.3)

62.1 17.7 (-44.4)

55.9 55.8 (-0.1) 56.5 (+0.6) 54.1

3
3
C 0

a-L-Rhamnose

Methyl a-L-rhamnopyranoside

r
M --

p-D-Mannose (I) p-L-Rhamnose

Methyl (methyl a-D-rnannopyranosid)uronate* 70.4 71.1 69.2 72.9 ?

(-0.8)

(-0.7)

(+1.2)

(-0.8)

*Suggested assignments.

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TABLE 5. Assignment of signals in I3Cspectra of configurationally related sugars. The figures of the chemical shifts are in p.p.m., as are those in parentheses, which represent the difference of the shift from that of the corrzsponding carbon in a- and p-D-galactoseand methyl a- and methyl p-D-galactopyranoside a - and p-D-Galactose
C- 1 93.6 93.7 (+O.l) 93.3 (-0.3) 97.7 C-2 69.8 69.6 (-0.2) 69.2 (-0.6) 73.3 C-3 C-4 C-5 71.7 C-6 62.5 C- l 100.5 101.0 (+0.5) 16.7 (-45.8) 62.3 100.5 (0) 104.9 105.1 (+0.2) 97.3 (-0.4) 72.8 (-0.5)

Methyl a - and methyl p-D-galactopyranoside

C-2 69.4 69.4 (0) 69.0 (-0.4) 71.8 71.8 (0) 71.5 (-0.3) C-3 C-4 C-5 C-6 62.3 OCHa-1 56.3 56.3 (0) 56.3 (0) 58.3 58.1 (-0.2) 58.3 (0)

5 >
L
C % '

a-D-Galactose (I) 70.56 70.63

Methyl a-D-galactopyranoside 70.6 70.4 71.8 Methyl p-L-(or D-)-arabinopyranoside 69.92 69.96 63.8 (-0.7) (-0.4) (-8.0)

I! '3

~-L-(oT D-)-Arabinose 69.8 69.8 63.6 (-0.8) (-0.8) (-8.1)

k
m

I:

6- Deoxy-a-D-galactose 70.4 73.0 67.4 (-0.2) (+2.4) (-4.3)

p-D-Galactose (I) 74.2 70.1
76.3

Methyl 6-deoxy-a-D-galactopyranoside 70.6 72.9 67.5 16.5 (0) (+2.5) (-4.3) (-45.8) Methyl p-D-galactopyranoside 73.9 69.8 76.2 Methyl a-L-(or D-)-arabinopyranoside 73.4 69.4 67.3 (-0.5) (-0.4) (-8.9) Methyl 6-deoxy-p-D-galactopyranoside 74.1 72.4 71.9 16.5 (+0.2) (+2.6) (-4.3) (-45.6)
62.1

e
C
~1

m 1

m 9

> r

R
V1

'3

6- Deoxy-p-D-galactose 74.0 72.5 71.9 (-0.2) (+2.4) (-4.4)

b 3

16.7 (-45.6)

104.8 (-0.1)

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TABLE 6. Assignments of signals in 13c.m.r. spectra of various methyl aldofuranosides. Chemical shifts and the figures in parentheses, which represent the difference of the shift of a given signal from that of the corresponding methyl galactofuranoside are in p.p.m. Methyl a-D-galactofuranoside
C- 1 103.1 103.2 (+o.l) 109.0 C-2 77.4 77.5 (+O.l) 75.3 C-3 C-4 C-5 C-6 63.4 OCH3-1 56.1 56.3 (+0.2) 56.3 C- 1 109.2 109.3 (i-0.1) 104.2 C-2 81.9 81.9 (0) 72.1

Methyl 13-D-galactofuranoside
C-3 C-4 C-6 C-6 63.9 0CH3-1 56.1

Methyl a-D-galactofuranoside 75.5 82.3 73.7 Methyl p-D-arnbinofuranoside 75.7 83.1 64.2 (+0.2) (+0.8) (-9.5) Methyl p-D-ribofuranoside 71.9 83.9 63.9

Methyl p-D-galactofuranoside 77.8 84.0 72.0 Methyl a-D-arabinofuranoside 77.5 8,4. 9 62.4 (-0.3) (+0.9) (-9.6) Methyl a-D-ribojiiranoside 70.8 85.5 62.2

GORIN AND MAZUREK:

N.M.R. STUDIES

OF ALDOSES

1219

C 0 2 H or C02CH3derivatives. When the spectra of methyl a-D-glucopyranoside and methyl a-Dxylopyranoside, and a-D-glucose and a-D-xylose are compared, respectively, only the shifts of C-5 signals are markedly different (- 1 1 and - 10 p.p.m., respectively). The displacements of the C-4 and C-5 signals recorded for the methyl P-D-aldopyranosides are similar to those recorded above in the a-series, as are those which are obtained when the spectra of the p-anomers of glucose, 6-deoxyglucose, and xylose are compared (Table 3). A similar approach can be used for comparison of the chemical shifts of the C-4 and C-5 signals of a- and p-D-mannose with those of aand P-L-rhamnose on one hand and for those of methyl a-D-mannopyranoside and methyl a-Lrhamnopyranoside on the other (Table 4). Again the C-4 and C-5 signal displacements are +.5 p.p.m. and - 4 p.p.m., respectively, without appreciable effects on those of C-1, C-2, and C-3. Since the result of replacing a C H 2 0 H group with a CH, group is the same in the glucopyranose and mannopyranose series, little difference would be expected between these two series when a C H 2 0 H group is replaced with a C 0 2 CH, group. Such a relationship allows a tentative assignment of signals in the 13c.m.r. spectrum of methyl (methyl a-D-mannopyranosid)uronate, whose deuterated derivatives would be difficult to prepare. Using the C H 2 0 H to C02Me displacements in the glucopyranose series, which are: C-2 (-0.6), C-3 (-0.5), C-4 (+1.8), and C-5 (-0.8), the nearest fit obtained by comparison of signal displacements of methyl a-D-mannopyranoside and the methyl ester are C-2 (-0.8), C-3 (-0.7), C-4 ( + 1.2), and C-5 (-0.8) (see Table 4). Comparisons can be made between the shifts of 13C signals of various sugars conformationally and configurationally related to galactose (Table 5). These are between: (i) a-D-galactose and 6-deoxy-a-D-galactose, (ii) the corresponding P-derivatives, (iii) methyl a-D-galactopyranoside and methyl 6-deoxy-a-D-galactopyranoside, and (io) the corresponding P-derivatives. In each comparison the C-5 and C-4 signals are dis+2.5 p.p.m., resplaced by -4 p.p.m. and pectively. The downfield displacement is less than +6 p.p.m. observed in the glucopyranose series, which contains sugars with equatorial rather than axial OH-4 groups. However, on

going from (i) a-D-galactose to P-L-arabinose, (ii) P-D-galactose to a-L-arabinose, (iii) methyl a-D-galactopyranoside to methyl P-L-arabinopyranoside, and (iv) methyl P-D-galactopyranoside to methyl a-L-arabinopyranoside, the C-5 signals are displaced by -9 to - 10 p.p.m. This is of the same order as is observed when the signals of glucose and xylose, or their corresponding methyl pyranosides are compared. The signals of 13Cspectra of some of the pyranose sugars described above and in a previous study (1) have been assigned previously by other methods. The results obtained by Perlin et al. (6), who correlated chemical shifts of 13C nuclei with electron densities based on molecular orbital data, agree in most cases. Assignments of 13C signals of ap-D-glucose, ap-D-mannose, P-Dgalactose, P-D-allose, ap-D-xylose, a-arabinose, methyl P-D-galactopyranoside, methyl P-D-glucopyranoside, and methyl ap-D-xylopyranoside were in agreement. In the cases of a-D-galactose, methyl a-D-glucopyranoside, methyl a-D-galactopyranoside, and methyl a-D-mannopyranoside, some were different. Differences were also observed for the C-2, C-3, and C-4 signals of P-arabinose but since they were resolved by only 0.2 p.p.m. and less, the discrepancy may be due to the use of H 2 0 as solvent in their study and D 2 0 in ours. The latter solvent was preferred since it appeared to suppress microbial activity and was suitable for 'H-',C decoupling studies. Less agreement was attained when the present data were compared with the results of Dorman and Roberts (7), who used a system of empirical constants based on steric or proximity effects to estimate chemical shift differences. Agreement was obtained when the assignments of 13C signals of P-D-glucose, ap-D-mannose, BD-galactose, a-D-allose, P-D-xylose, ap-L-rhamnose, p-fucose, and methyl P-D-glucopyranoside were compared. Their assignments of signals of a-D-glucose, a-D-galactose, ap-arabinose, P-Dallose, a-D-xylose, a-fucose, and methyl a-D-glucopyranoside differed. The disagreement that exists in the cases of a- and 0-arabinose can be explained partly by the differing interpretations of the 13c.m.r. spectrum. Dorman and Roberts (7) reported a signal for each anomer that could not be observed by Perlin et al. (6) or in the present study. Their discrepancy is likely due to the slowness of recording the spectra, relative to the speed of mutarotation.

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CAN. J. CHEM. VOL. 53, 1975

TABLE 7. Differences between the 13C chemical shifts of the anomers of D-glucose, and the respective ones of D-mannose and D-galactose, in p.p.m.

C- 1

C-2

C-3

C-4

C-5

C-6

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The signal displacements occurring on changing the nature of the substituent at C-5 are confined to C-4, C-5 (and C-6 in some cases). Thus, these effects are over much shorter range than those observed on going from an equatorial to an axial hydroxyl group, for example, P-D-glucose to P-D-galactoseor P-D-glucoseto p-D-mannose. As far as these longer range effects are concerned, it is evident that on going from P-Dglucose to p-D-mannose, with an axial OH-2 group, and to P-D-galactose with an axial OH-4 group, there is little correlation between signal displacements of carbons in the corresponding positions relative to the axial OH group (Table 7). It, therefore, does not seem profitable to pursue such an approach in predicting 13C chemical shifts of other sugars containing axial OH groups. A certain number of methyl furanosides were examined and their 13c.m.r.spectra rationalized. Not surprisingly, the C-1, C-2, C-3, C-4, and OCH, signals of methyl P-D-galactofuranoside correspond in their shifts to those of methyl P-L-arabinofuranoside and those of the corresponding P-hexoside to those of the a-pentoside (Table 6). Evidently the CH'OH and CHOHCH'OH substituents are similar in their shielding or deshielding effects and do not give rise to differences of conformation or populations of given conformations. A few of the deuterated compounds compiled in Table 1 were prepared by routes that have not been described previously. For example, methyl a-L-rhamnopyranoside was treated with phosgene in pyridine to give the 2,3-cyclic carbonate, which was then oxidized with phosphorus pentaoxide in dimethyl sulfoxide to the 2,3-cyclic

carbonate of methyl 6-deoxy-a-L-lyxohexopyranosid-4-ulose. Treatment with sodium borohydride provided a mixture containing methyl a-L-rhamnopyranoside which was identified by its 13c.m.r. spectrum and a larger proportion of another isomer, which although it was not isolated, is probably methyl 6-deoxy-ol-L-talopyranoside. The action of sodium borodeuteride on the ketone provided a mixture of the 4-'H derivatives of the methyl 6-deoxy-ol-L-hexopyranosides, which was used for assigning the 13C signals of methyl a-L-rhamnopyranoside. The mixture of aldoses prepared on hydrolysis contained ap-L-rhamnose-4-'H, which aided the assignment of ' signals of ap-L-rhamnose. D-Ribose-5-'H was prepared from I ,2-0isopropylidene-ol-D-allofuranose by successive periodate oxidation, sodium borodeuteride reduction and hydrolysis. ~-Ribose-3-'H was obtained from 1 ,2-0-isopropylidene-a-D-allofuranose-3-'H in a similar fashion, except that sodium borohydride was used as reducing agent.

Experimental
Carbon-1 3 magnetic resonance spectra were obtained using a Varian XL-100-15 spectrometer with Fourier transform from D 2 0 solutions (0.85 ml) of compound (20-100 mg) contained in a coaxial glass cylinder fitting snugly within a 12 mm diameter x 8 in. tube maintamed at 33". Larger amounts of solute were dissolved in D 2 0 (2 ml) contained in the 12 mm x 8 in. tube similar to that described above, except that it was fitted with a Teflon vortex plug. T o obtain a complete I3c.m.r. spectrum the spectral width was 5000 Hz, the acquisition time 0.4 s, and the pulse width 50 ps. In several experiments it was necessary t o improve the resolution and the spectral width was narrowed to 500 Hz, with the acquisition time 4 s and the pulse width 117 ps. Under these conditions the extent of the p-carbon deuterium isotope effect could

GORIN AND MAZUREK: N.M.R. STUDIES OF ALDOSES

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be measured by examination of a mixture of deuterated deuterated methyl ap-D-galactopyranosides by refluxing and undeuterated material. Frequently signals were in 3% methanolic hydrogen chloride for 3 h. superimposed when a 5000 H z scan was used and those S 2 H 2and 4,5-2H3 Derivatives of ap-D-Arabinopyracould be resolved by preparation of two o r three spectra nose, and Corresponding Deuterated Methyl with a 500 H z sweep width and suitable offsets. Signals ae-D-Arabinopyranosides and Methyl of CH, groups were recognized a s triplets in partially ap-DArabinofi~ranosides decoupled spectra. The chemical shifts are expressed as (3-~-Arabinopyranose-5-~H, was prepared from a-D6 c in p.p.m., relative to external Me4Si, whose shift rela- gluc0se-6-~H~ (I) by the method used by Isbell et at. (20) tive t o D 2 0 (lock signal) was obtained in a separate for the meoaration of ~-~-arabino~vranose-5-'~C from .experiment. a - ~ - g l u c o s e - 6C -~ .~ Some of the undeuterated chemicals used were ob~:D-~~ucose-5,6was 2~3 prepared by acid hydrolysis tained commercially and they are listed below with SUP- of ~ , ~ - 0 - i ~ ~ ~ ~ ~ ~ ~ ~ i (18) ~ ~ ~ ~ - ~ ~ l i e r s :a-D-xylose (Fisher Scientific Co., Fair Lawn, and converted to p-D-arabinopyranose-4,5-2H3 by the N.J.), a-L-arabinose (Nutritional Biochemicals Corp., same procedure, Cleveland, Ohio), a-L-rhamnose (Aldrich Chemical Co., ~ ~ t ~ of ~ of ~ thet arabinoses ~ t iin ~~0 ~ gave ~ Milwaukee, Wisc.), a-L-fucose, and a- and a-anomers of the ap-pyranosidic mixtures, methyl D-gluco~~ranoside and methyl D-galacto~~rano- Treatment of the anomeric mixture with refluxing 3% side, methyl a- and methyl 13-~-xylop~ranoside (Sigma methano]ic hydrogen chloride for 3 h gave a mixture of Chemical Co., St. Louis, Mo.), a - ~ - g l u c o s e - 2 - ~(Raylo H methyl ap-D-arabinopyrano~ideand methyl ae-D-arabinChemicals Ltd., Edmonton, Alta.), methyl e-L-arabino- 0f~ranosic-J~. In a parallel experiment using undeuterated pyranoside (K & K Labs Inc., Plainview, N.Y.). derivatives, the glycosidic mixture gave a 13c.m.r. specOther compounds were prepared according to Pro- trum with 23 of the possible 24 signals. Even when a cedures described in the appended references. These were: spectral width of 500 H~ was used the signal at 6 c 75.7 methyl a-~-rhamnopyran~side ( 8 ) , a - and P-anomers of from methyl a- and methyl p-D-arabinofuran~sidecould methyl 6-deoxy-L-galactopyranoside (9), methyl a-D-ara- not be resolved, binofuranoside (lo), methyl P-L-arabinofuranoside (1 l), 6-Deox~-ap-D-gatactose-6-2H, 6-Deoxy-ae-L-ga1acmethyl a - ~ - ~ ~ ~ b i ~ o p y r a n(12), o s i d methyl e p-D-galactoto~e-4-~H Derived , Methyl 6-Deoxy-aP-~-galactofuranoside and its minor a-anomer which are produced simultaneously (13), a- and of methyl ~ - g l u c ~ pyranosides, 6-Deoxy-ap-L-gtucose-4-2H~ and Methy16-Dcoxy-ap-L-g1uco~~ranoside-4-2H pyranosiduronic acid (14), methyl (methyl a-D-mannopyranosid)uronate (IS), and methyl 6-deoxy-a-~-~luco- Using a Illethod analogous to that of Schmid and Karrer (21), who reduced 1,2:3,4-di-0-isopropylidene-6pyranoside (16). The above glucopyranosiduronic acid derivatives were p-tolylsulfonyl-a-D-galactopyranose with lithium alumiconverted to their methyl esters by treatment for with num hydride to give 1,2:3,4-di-O-iso~rop~lidene-a-~fucopyranose, the 6-2H derivative was obtained by using cold 1% methanolic HCI. lithium aluminum deuteride. Heyns et at. (22), using Pt/H2 as reagent, reduced Preparation of Deuterated Sugars methyl 6-deoxy-a-L-xylohexopyranosid-4-ulose to give a ~ - ~ - X y l o ~ ~ r a n o s e -a~-~-Xylopyranose-3-~H, 5-~H, methyl 6-deoxy-a-L-galactopyranoside exclusively. Using Pt/D2 we obtained the 4-'H derivative, and when sodium and Derived Methyl ap-D-Xylopyranosides a-~-Xylose-5-~H was prepared from 1,2-0-isopropy- borodeuteride in water was used .as a reagent, some lidene-a-D-glucofuranose by successive oxidation with methyl 6-deo~~-a-~-glucopyranoside-4-~H was formed sodium periodate, reduction with sodium borodeuteride, also, as shown by the '3c.m.r. spectrum. Hydrolysis of each deuterated glycoside or glycosides and hydrolysis, as outlined by Isbell et at. (17) for the with 0.5 M H2S04 at 100" for 6 h gave the appropripreparation of a-D-xylose-5r3H. a - ~ - X y l o s e - S - ~ was H similarly prepared from 1,2-0- ate reducing sugars. Each deuterated glycoside was equiisopropylidene-a-D-glu~ofuranose-3-~H, obtained from librated with refluxing 3% methanolic hydrogen chloride partial acid hydrolysis of 1,2:5,6-di-0-isopropylidene-a- for 3 h to give the ae-glycosidic mixture. ~-glucofuranose-3-~H (18). ae-~-Rhamnose-6-~H, ap-~-~hamnose-4-~ Methyl H, Mutarotation of each of the deuterated X Y ~ O S ~ in S D20 a-D-Rhanmopyranoside-6-~H, and thy/ gave ae-D-xylopyranose. a-L-Rhamnopyrano~ide-4-~H Each of the ap-mixtures was treated with refluxing 3% Haskins et (23) hydrogenated methyl 2,3,4-tri-0methanolic hydrogen chloride to give methyl 4 - D - benzoy~-6-deoxy-6~iodo-a-~-ma~nopyran~sid to give xylopyranoside. methyl 2,3,4-tri-0-benzo~l-a-D-rhamnopyranosid using 4-2H and 6-2H, Derivatives of Methyl ap-D-ga/aCtoRaney nickel in the presence of diethylamine. They Profuranoside and Methyl ae-D-Galactopyranoside ceeded to prepare methyl a-D-rhamnopyranoside and The 4-2H and 6-'H derivatives of r-D-galactose (I) a-D-rhamnopyranose. The compounds were prepared as weredissolvedin0.5~methanolichydrogenchloridewhich 6-2H derivatives by substituting deuterium for hydrogen was left for 15 h at room temperature (19). The product in the reduction step. consisted mainly of methyl me-D-galactofuranoside T o methyl a - L - r h a m n ~ p ~ r a n o s i d (6.6 e g) in pyridine according to the l3c.m.r. spectrum. The signals of methyl (30 ml), phosgene (1.2 molar-equiv.) in toluene (20 mi) ae-D-galactofuranoside were assigned once the signals of was added with stirring. After 3 h, thesolution wasevapormethyl ap-D-galactopyranoside were accounted for. ated to a sirup, which was partitioned between chloroform The above furanosidic mixtures were converted to and water. The chloroform layer was washed with water
r ~ L

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14. S. A. BARKER, E. J. BOURNE and M. STACEY. Chem. Ind. (London) 970 (1951). 15. H. W. H. SCHMIDT. Tetrahedron Lett. 236 (1%7). 16. T. E. TIMELL, W. ENTERMAN, F. SPENCER and E. J. SOLTES. Can. J. Chem. 43,2296 (1965). H.L. FRUSH and J. D. MAYER. J. Res. 17. H. S. ISBELL, Natl. Bur. Standards. @A, 359(1960). 18. H. J. KOCHand A. S. PERLIN. Carbohyd. Res. 15,403 (1970). 19. P. A. LEVENE, A. L. RAYMOND and R. T. DILLON. J. Biol. Chem. 95,699(1932).

20. H. S. ISBELL, N. B. HOLTand H. L. FRUSH. J. Res. Natl. Bur. Standards 57,95 (1956). 21. H. SCHMID and P. KARRER. Helv. Chim. Acta. 32, 1371 (1949). and H . PAULSEN. Chem. 22. K. HEYNS,A. L. BARON Ber. 97,921 (1964). 23. W. T. HASKINS, R. M. HANN and C. S. HUDSON. J. Am. Chem. Soc. 68,628 (1946). 24. P. M. COLLINS. Tetrahedron 21,1809 (1965). 25. I. AUGESTAD and E. BERNER. Acta. Chem. Scand. 10, 911 (1956).