3664 J. Org. Chem. Vol. 39, No. 25, 1974
Niedballa and Vorbruggen
A General Synthesis of N-Glycosides. III.192 A Simple Synthesis of Pyrimidine Disaccharide Nucleosides U. Niedballa and H. Vorbruggen" Research Laboratories of Schering A.G., Department Allgemeine Synthetische Chemie, 1 Berlin 65, Muedlerstrasse 170-1 72, Germany Received March 29,1474 Silylated uracils as well as 6-azauracils react smoothly with peracetylated reducing disaccharides in the presence of SnC14 to give the corresponding disaccharide nucleosides. Preliminary experiments show that tri- and tetrasaccharides react analogously.
Wolfrom and coworkers3 have described the synthesis of acetylated purine disaccharide nucleosides via the chloromercuric salts, which gave the pure crystalline 9P-disaccharide purine nucleosides in -10-30% overall yields. More recently Rao and Lerner4 prepared analogously Ip-cellobiosyl- and 1/3-lactosylbenzimidazole, but, to our knowledge, no disaccharide pyrimidine nucleosides have as yet been prepared starting from disaccharides prior to our own work. Thus we wondered whether the readily available peracetylated oligosaccharides with a reducing end group, especially acetylated disaccharides like octaacetylcellobiose, -lactose, and -maltose, would not react with silylated pyrimidines and SnC14 to give the corresponding pyrimidine disaccharide nucleosides. We expected a smooth reaction since it is well known5 that Friedel-Crafts catalysts like SnC14 or TiC14 cleave the 1-0-acetyl bond in these acetylated oligosaccharides in preference to the glycosidic linkage between the sugar residues to give a C-1 cation. However, in our initial reactions of crystalline disaccharide octaacetates with silylated pyrimidines and SnC14 the yields of acetylated pyrimidine disaccharide nucleosides varied widely and several other products were formed. A detailed investigation of the reaction of octaacetylcelbbiose (1) with silylated 6-azauracil (2) revealed that commercial as well as our own samples of 1 always contained up to 0.5% ethanol or other alcohols from recrystallization. Since alcohols and acetic acid react with SnCl, or Tic14 to give HC1,2 the disaccharide bond in 1 or in the already formed disaccharide nucleoside 3 was cleaved. Thus not only 3 but also the 6-azauracil N-glucosides 4 and 5 were formed by the reaction of 2 with the cations obtained by the indicated types of glycosidic bond cleavage of 1 and 3 (Scheme I). The nucleoside 4 was identified by tlc comparison with an authentic sample. Furthermore both acetylated nucleosides 4 and 5 gave on saponification the known free 2-(p-
D-glucopyranosyl)-2,3,4,5-tetrahydro-1,2,4-triazine-3,5dione (8),lthus making the structure of 5 highly probable. To avoid formation of HCI the acetylated cellobiose was therefore pulverized and dried a t elevated temperature in high vacuum or by azeotropic distillation. Repetition of the reaction of dried 1 with 2 and SnC14 in boiling 1,2-dichloroethane now gave the crystalline acetylated disaccharide nucleoside 3 in 80% yield as the only product. Reaction of 2 with hepta-0- acetylcellobiosyl chlorides in the presence of SnC14 in 1,2-dichloroetbane gave beside 3 the N3- as well as the Nl,N3-bis(disaccharide) nucleoside.6 In most subsequent experiments the carefully dried peracetylated disaccharides were treated with silylated pyrimidines and SnC14 in boiling 1,2-dichloroethane yielding 60-80% of the c~rrespondingperacetylated disaccharide nucleosides, which often either crystallized spontaneously or after purification over a column of alumina. Methanolysis with sodium methylate in methanol gave in high yields the free disaccharide nucleosides. The structures of the acetylated disaccharide nucleosides are supported by their mass spectra as exemplified by the fragmentation of the maltose nucleoside 6 (Scheme 11). Further obvious fragments observed were mle 744 (M H), 685 (M - CH3COOH), 684 (744 - CH&OOH), 625 (M - 2 CH&OOH), 624 (744 - 2CH3COOH). An additional proof for the structure of the free disaccharide nucleosides was provided when the free disaccharide nucleoside 7a with an a-glucosidic bond was incubated with maltase (a-glucosidase) in aqueous buffer solution to give the glucoside 8 in practically quantitative yield, which was identical with an authentic sample (Scheme 111). Since peracetylated tri- and tetrasaccharides are not commercially available we prepared and reacted a mixture of peracetylated cellotriose (9) and cellotetraose (10) with silylated 6-azauracil (2) and obtained a mixture of the acetylated oligosaccharide nucleosides 11 and 12 in -70% yield, which were separated by preparative tlc, and the
Scheme I
cleavage
3
OAc 1
3
4
5
J. Org. Chem., L701.39, No. 25, 1974 3665
A General Synthesis of N-Glycosides. I11 Scheme
Scheme I11
II
0
0
-
HOCH,
maltase
pH 6 buffer, 40 hr, 25'
OH
OH 7a
+ glucose
OH 8
m/e 444
AcOCH,
AcOCH,
I
Scheme IV
I
@J$+
AcO
OAc
OAc
m/e 618 ACOCH,
0
OAc
OAc
9,n-1 10,n=2
'
6Ac
.m/e 127
m/e 331
structures were assigned by nmr ratio of H-5 and H-1' to acetyl CH3 (Scheme IV). In Tables I and I1 the formation of acetylated and free disaccharide nucleosides is summarized. Besides the already mentioned octaacetylcellobiose (l), octaacetylmaltose (13) and octaacetyllactose (14) were treated with 3 3 bis(trimethylsilyloxy)-1,2,4-triazine (2), 6-methyl-3,5S,O-bis(trimethylsilyl)-1,2,4-triazine (15), 6-methyl-3,5bis(trimethylsilyloxy)-1,2,4-triazine (16), 5-ethyl-2,4-bis(trimethylsily1oxy)pyrimidine (17), 2,4-bis(trimethylsily1oxy)pyrimidine (18), and 2,4-O,N-bis(trimethylsilyl)-2hydroxy-4-aminopyrimidine(28). Experimental Section For instruments and the purification of solvents compare part I of this series.2 Tlc systems follow: system A [toluene-acetic acid-Hz0 (5:5:1)];7 system B (ethyl acetate); system C [ethyl acetate-hexane (2:1)]; system D In-butyl acetate-methyl glycol-HzO (4:1:2)]; and system E [l-propanol-aqueous NH3 (7:3)]. For the column chromatography neutral alumina (Woelm) and standard silica gel Merck (Darmstadt) were used.
OAc
OAc 11, n =1 12. n = 2
Preparation of the Acetylated Disaccharides. The acetylated disaccharides were prepared according to standard methods and dried either by heating 1-2 hr to 80-100° mm) or by azeotropic distillation with 1,a-ethylene chloride: Octaacetyl-a-cellobiose,8 octaacetyl-P-maltose," octaacetyl-P-lactose10 were obtained. Preparation of Acetylated Oligosaccharides. The acetolysis of cellulose was performed according to Hess and Dziengel.11 The crude product was dried in U ~ C U Oa t 50" for 2 days and extracted with hot methanol several times. The methanolic solution was concentrated in uucuo. On stirring over night crystals of octaacetyla-cellobiose (I, mp 222-223') had formed, which were filtered.
3666 J. Org. Chem. Vol. 39, No. 25, 1974
Niedballa and Vorbriiggen Table I
Protected nucleoside Silylated
Peracetyled
~
1
3
Analysis, % c a l c d (found)
-
_
I
_
I
pyrimidine disaccharide Y i e l d , % a
2
_
80
Mp,*
[a120DC
O c
- 59.0
241- 242
E-i,d
Formula ( m o l w t )
5.88
C29H37"3019
(731.64)
2
13
15
1
7
72
1%
95
130-132
- 31.5
5.88
cZ RH3?"3019
- 17.1
6.61
C30H39N3018S
(731.64) (761.73)
16
1
20
63
150-153
- 48.0
5.73
C30H39N3019
15
13
21
70
181-182
- 68.9
6.60
C30H383018S
(745.66) ('761.73)
- 35.7
5.86
80
- 29.6
5.91
cZ 9H3TN30i9
66
-3.1
6.61
C30H3 S N 3 0 i $
10
13
22
63
2
14
23
15
14
24
125- 126
C30H39N3019
(745.66) (73 1.64) (761.73)
16
14
25
69
1'7
1
26
78
5.80
I8
13
21
71
5.85
- 31.5
5.75
C30H39N30i9
(745.66) C32H42"2019
(758.71) 19
3 0% 8N20
(730.81) 28
14
29
81
6.15
9.1
C3CH39N3018
(729.67) a
C
H
N
47.61 (47.50) 47.61 (47.40) 47.31 (47.02) 48.32 (48.02) 47.31 (47.02) 48.32 (47.97) 47.61 (47.67) 47.31 (47.34) 48.32 (48.07) 50.66 (50.39) 49.31 (49.06) 49.38 (49.19
5.10 (5.31) 5.10 (5.48) 5.16 (5.46) 5.27 (5.40) 5.16 (5.50) 5.27 (5.49) 5.10 (5.38) 5.16 (5.39) 5.27 (5.33) 5.58 (5.76) 5.34 (5.39) 5.39 (5. 55)
5.74 (6.03) 5.74 (5.93) 5.51 (5.56) 5.64 (5.63) 5.51 (5.61) 5.64 (5.51) 5.74 (5.56) 5.51 (5.72) 5.64 (5.65) 3.69 (3.61) 3.86 (3.75) 5.76 (5.61)
5
4.21 (4.20)
4.21 (4.11)
4.21 (4.17)
According to standard procedure. b From ethanol. c c 1, CHC13. d CDC13, parts per million (d, J = 9 Hz).
Table I1 1 _ _
FreP nucleoside
Analysis, 74 c a l c d ( f o u n d )
Protected
-
nucleoside Y i e l d , gff
3
3a
95
"c
[ O]ZODC
bI..1t d
202-205
- 39.0
5.63
Mp,b
Formula ( m o l w t ) Ci5H23W3012*
HZo
(455.50)
7
7%
75
188-190
- 47.3
5.66
Ci5H23N3012'
"2O
(455.50) 19
19a
74
2 74-2 75"
- 18.8
6.65
C16H25N3Q11S
20
2053
82
260e
- 33.3f
5.65
C16H25N3012
21
2 la
65
207-209
- 60.8
6.63
CIgH2rjN30ijS
(467.47) (451.40) (499.62) 22
22a
79
174- 175
-55.v
5.65
C16H25N3012
23
23a
72
292-294
- 24.3
5.68
C15H23N30i2
24
24a
83
218-221
- 10.0
7.30g
C16H25N301iS
25
2 5a
78
2 56-2 58"
- 20. 8f
5.65
CiBH25N3012
26
26a
C18H28N2012
27
2"la
C16H24N20i2
(451.40) (437.38) (467.47) (451.40)
CH3OH
C
H
N
39.55 (39.74) 39.55 (39.84) 41.11 (41.02) 42.57 (42.83) 40.87 (40.60) 42.57 (42.43) 41.19 (41.17) 41.11 (40.84) 42.57 (42.84) 46.55
5. 53 (5.84) 5.53 (5.76) 5.39 (5.48) 5.59 (5.82) 5.88 (5.93) 5.59 (5.71) 5.30 (5.44) 5.39 (5. 56) 5.59 (5.84) 6.08
44.03
5.54
6.42
44.14 (43.86)
5.79 (5.93)
9.65 (9.49)
5
9.22 (8.96) 9.22 (8.97) 8.99 6.86 (8. 81) (6.68) 9.31 (9.07) 8.45 6.59 (8.35) (6.49 9.31 (9.19) 9.61 (9.47) 8.99 6.86 (8.84) (6.72) 9.31 (9.18) 6.03
(464.42) (436.37) 29
29a
90
11. of
5.70
Ci6H25N301i
(435.40) a According to standard procedure. b Prom ethanol-Hz0. tion. f c, 0.4 (ethanol-HzO. 3:2). g In pyridine-&.
c
c, 0.4 (ethanol-HzO, 3:l). d DzO, parts per million (d, J = 9 Hz).e Decomposi-
J . Org. Chem., Vol. 39, No. 25, 1974 3667
A General Synthesis of N-Glycosides. I11
Enzymatic Cleavage. 2-(~-D-Maltosyl)-2,3,4,5-tetrahydroThe residual oil gave on methanolysis12 the free oligosaccharides. 1,2,4-triazine-3,5-dione (7a, 60 mg, 0.37 mmol) was dissolved in acThe methanolic solution was neutralized with HzS04 and evapoetate buffer, pH 6 (10 ml, 0.1 N ) . After addition of a-glucosidase rated in uucuo to dryness. The aqueous extract13 was reacetylated (2 mg, Boehringer-Mannheim, suspension in 3.2 m ammonium sulaccording to Hess and Dziengelll with acetic anhydride-pyridine fate solution, pH -6, 1 ml) the reaction was incubated a t 25" and to give the acetates, which were separated by column chrmatograthe enzymatic cleavage was followed by tlc. After 40 hr all starting phy on silica gel eluting with mixtures of ether-acetone. material had disappeared and a new product had formed, which The purity of the oligosaccharide fractions was checked by tlc was identical in polarity with 2-(/3-~-glucopyranosy1)-2,3,4,5-tet(system A). The spots were detected by spraying with HzSOq and rahydro-1,2,4-triazine-3,5-dione (8). The solution was filtered over heating to l l O o for 5 min. For the silyl Hilbert-Johnson reaction a a column of Dowex 50 E ' and the column was washed with methachromatography fraction was used, which according to tlc (system nol-HzQ. Filtrate and washings were combined and evaporated in A) consisted of -60% hendecaacetylcellotriose (9), tetradecaacetyluucuo to dryness. Preparative tlc on silica gel (system E) gave a cellotetraose (IO), and some octaacetylcellobiose ( I ) . fluorescent band which was eluted with methanol-HzO and crysPreparation of the Heterocycles. The 1,2,4-triazine derivatallized from ethanol to give 35.9 mg (95.1%) of 8 as white needles, tives were prepared according to Gut14 and silylated according to mp 211-212° (lit.2 210--212O). standard methods.2 Reaction of a Mixture o f Kendecaacetylcellotriose (9) and A. General Procedure for the Preparation of Disaccharide Tetradeeaacetylcellotetraose (10) with 2. To a mixture of 9, 10, Nucleosides. To a solution of peracetylat,ed disaccharide (15 and some 1 (2.0 g) and 2 (8 mmol) in L,2-dichloroethane (100 rnl), mmol) and of the persilylated pyrimidine (16.5 mmol) in 1,2-diSnC14 (0.8 ml, 6.8 mmol) was added. After 2 hr of reflux and workchloroethane (150 ml) SnC14 (1.26 ml, 10.8 mmol) in 1,2-dichloup (2.1 g colorless foam) an aliquot of the mixture of 11 and 12 (1.6 roethane (20 ml) was added. The reaction mixture was either reg) was subjected to preparative tlc (silica gel, system D) to give the fluxed for 2 hr under exclusion of moisture (preparation of 3, 7, nucleosides I1 and 12. 14-20, 29) or kept for 4 hr a t 60' (preparation of 21, 22). The 2-(Deca-O- acetyl-~-D-ceilotriosyl)-2,3,4,5-tetrahydro-l,2~~cooled solution was diluted with methylene chloride (200 ml) and triazine-3,5-dione (11):nmr (CDC13) 6 7.39 (s, 1,H-5), 5.84 (d, 1, J poured into ice-saturated NaHC03 solution (200 ml). The organic = 9 Hz, H-l'), 2.2-1.9 (m, 30, OAc). layer was separated and the aqueous solution was washed with 2-(Trideca-O-acetyl-~-D-cellotetraosyl)-2,3,4,5-tetrahydromethylene chloride (100 ml). The combined organic solution was 1,2,4-triazine-3,5-dione (12): nmr (CDC13) 6 7.39 (s, 1, H-5), 5.83 filtered through sand-Celite, dried over NazS04, and evaporated (d, 1,J = 9 Hz,H.-l'), 2.2-1.8 (m, 39, QAc). in U U C U O . The residue was chromatographed on neutral alumina (activity 111) and eluted with ethyl acetate-methanol. B. General Procedure for the Methanolysis of Acetylated A c ~ ~ ~ w ~ e We d ~arer indebted n ~ ~ ~to. Dr. G.-A Hoyer Disaccharide Nucleosides. Acetylated disaccharide nucleoside and Dr. D. Rosenberg for the nmr and mass spectra and Di(4.4 mmol) was suspended in dry methanol (20 ml) and 1 N NaOCH3 solution (8.8 mmol) was added whereupon the subplorn-Ing. G. Huber for the analyses. stance dissolved and after 3Q-40 min a white precipitate was formed. After stirring the reaction solution overnight a t room temperaRegistry No.-I, 5346-90-7; 2, 17331-61-2; 3, 38909-08-9; 38, ture the alkaline solution was neutralized by addition of wet 38909-17-0; 7, 38909-10-3; ?a, 38909-16-9; 9, 31873-41-3; 10, 52523-12-3; 11, 39020-67-2; 12, 52523-13-4; 13, 20880-60-8; 14, Dowex 50 H+, whereupon the precipitate dissolved. The resin was 52554-32-2; 15, 52571-10-5; 16, 17331-64-5; 17, 31167-05-2; 18, filtered and washed with methanol and water. After evaporation of 10457-14-4; 19, 52523-14-5; 19a, 52523-15-6; 20, 38909-07-8; 20a, the combined filtrates the residue was crystallized from ethanol or a mixture of ethanol-water. 38909-15-8; 21, 38909-09-0; 21a, 38909-18-1; 22, 38909-06-7; 22a, 2-(Hepta-0- acetyl-/3-D-cellobiosyl)-2,3,4,5-tetrahydro-l,2,- 38909-14-7; 23, 38909-11-4; 23a, 38909-19-2; 24, 38909-12-5; 24a, 4-triazine-3,5-dione (3). On reaction of impure 1 (containing al38909-20-5; 25, 52,523-16-7; 25a, 52523-17-8; 26, 38909-23-8; 26a, cohol) the crude reaction mixture showed on tlc evaluation (system 38909-24-9; 27, 52523-18-9; 27a, 52523-19-0; 28, 18037-10-0; 29, B) besides 3 a number of less polar products, one of which was 52523-20-3; 29a, 52523-21-4. identical with an authentic sample of 4. A mixture of 4 and t h e slightly slower moving 5 was isolated by preparative tlc (system B) and saponified with methanolic ammonia to give a product which gave only a single spot on tlc (system E) identical with an authen-(1) Synthesis of Nucleosides No. 1 1 . t,ic sample of 8. (2)Part I: J. Org. Chem., 39, 3654 (1974).Part II: J. Org. Chem., 39, 3660 Reaction of Hepta-0 -acetylcellobiosyl Chloride with 2. To 11974). a chilled solution of hepta-0- acetylcellobiosyl chloride (2.5 g, 3.8 (3)iaj M.' L. Woifrom, P. McWain, F. Shafizadeh, and A. Thompson, ,J. mmol) and 2 (4.5 mmol) in dry 1,2-dichloroethane (30 mi), SnC14 Amer. Chem. Soc., 81, 6080 (1959);(b) M. L. Wolfrom, P. McWain, and (0.1 ml, 0.86 mmol) in 1,2-dichloroethane (5 ml) was added. After 4 A. Thompso, ibid., 82,4353 (1960). (4) R. Rao and L. M. Lerner, J. Ory. Chem., 87,3741 (1972). days a t 22' and work-up the crude product contained according to (5) E. Pacsu, Chem. Ber., til, 1508 (1928);I. F . Szabb, II Farkas, R. Bogtlc examination (silica gel, system C) a t least two products. The nar, and H.Gross, Acta Chim. Acad. Sci. Hung., 64,67 (1970). nucleosides were separated by fractional precipitation from etha(6)The formation of: Ni,N~-bisnucleosdesin reactions with acetylated 1nol. On dissolving the crude reaction product in a small volume of halo sugars and silylated 5-methyl-6-aza~iraciihas been observed by R. hot ethanol a precipitate of pure 3 was obtained. Further fractional H. Hall, J. Amer. Chem. Soc., 80, 1145 (1958). (7)G. Quinkeri, B. Wegemund, F. Hbmburg: and b. Cimboilek, Chem. Ber., precipitation gave the Nq- as well as the Nz,N4-bisglycoside. 97,958 (1964).For the use of two-phase tic systems, compare t i . VorYield of 3: 1.68 g (60.5%); , , ,A (CH3OH) 263 nm, , , ,A (CH30H brijggen and P. Strehlke, bid.. 108, 3039 (1973). 4-NaQH) (0.1 N ) 262 nm. (8)L. Gattermann and H. Wieland, "Die Praxis des organischen Chemik4-(Hepta-0- acetyl-~-D-cellobiosyl)-2,3,4,5-tetrahyd~o-l,2,4ers," Waiter de Gruyter and Co., Berlin, 1962,p 345. , , ,A (CH30H) 259 nm, A,, triazine-3,s-dione: yield, 0.25 g (8.9%); (9)W. Koenigs and E. Knorr, Chem. Ber., 34, 4343 (laoi). (IO) M. Schmoeger, Chem. Ber., 25, 2452 (1892). (CH3OH 4-NaOH) (0.1N) 305 nm. (11)K . Hess and K. Dziengel, Chem. Ber., 68, 1594 (1935). 2,4-Bis(hepta-0- acetyl-/3-~-cellobiosyl)-2,3,4,5-tetrahydro1,2,4-triazine-3,5-dione: yield, 0.148 g (5.3%); , , ,A (CH30I-I) 263 ;: 234 (1932), nm, Amax (CH30H -I-NaOH) (0.1 N ) 262 nm; nmr (CDC13) 6 7.35 (b) J. Gut, (14)(a)J. Gut, Co//ecf.Czech. Chem. Commun., 23, 1588 (1958); (s, 1, H - 8 , 5.83 (m, 2, H-l'N4, H-l'N2), 2.2-1.8 (m, 42, OAc). Advan. Heterocycl. Chem., 1, 203 (1963).
[iii ~
~
~
~
~
~
~
