 |
Description  |
|
|
BACKGROUND OF THE INVENTION
The present invention relates to new sialic acid derivatives, in
particular, gangliosides or intermediates for producing gangliosides.
In general, glycolipids of mammal cells are the substances which are formed
by glycoside linkage between various combinations of various sugars such
as glucose, galactose, N-acetylglucosamine, N-acetylgalactosamine, fucose
and sialic acid, and a lipid structure called ceramide which in turn is
formed through an amide linkage between fatty acid and long-chain
aminoalcohol known to as sphingosine. And, these glycolipids belong to a
so-called sphingoglycolipid. Among these glycolipids, those which have
sialic acid are specifically called gangliosides.
Generally, such compounds locally exist in the outer molecular layer of the
bi-layer structure of cell membrane. The current study has proved that
these compounds play important roles in cells, such as identification of
cells, reception and response to information, receptor function,
differentiation, and proliferation, malignant alteration and behavior of
cells, and so forth.
It is, however, extremely difficult to isolate oligosaccharides containing
sialic acid from a living body. Therefore, fine synthesis of those
oligosaccharides is indispensable for clarification of correlation between
the accurate biological information and molecular structures of the
saccharides.
SUMMARY OF THE INVENTION
Accordingly, an object of the present invention is to provide sialic acid
derivatives useful as an intermediate for the synthesis of gangliosides,
as well as a method of producing such substances.
The present invention is concerned with sialic acid derivatives having the
following general formula (I):
##STR5##
wherein, R.sup.1 represents hydrogen, an acetyl group, a trityl group,
##STR6##
(R.sup.5 represents hydrogen or an acetyl group and R.sup.6 represents
hydrogen, sodium or a methyl group)
R.sup.2 represents hydrogen or an acetyl group,
one of R.sup.3 and R.sup.4 represents chlorine, --OAc, --OH,
##STR7##
(R.sup.7 represents a hydrogen atom or an acetyl group, R.sup.8 represents
a hydrogen atom, an acetyl group or a benzyl group, R.sup.9 represents a
hydrogen atom, an acetyl group, a benzyl group
##STR8##
wherein R.sup.10 represents a hydrogen atom or a benzoyl group), while
the other represents --COOR.sup.11 wherein R.sup.11 represents hydrogen,
sodium, or a methyl group.
DETAILED DESCRIPTION OF THE INVENTION
The present invention will be explained in detail below by referring to the
following process diagrams.
First, explanation will be made by referring to Process diagram 1.
##STR9##
In the first step, the compound (1) is dissolved in acetylchloride and the
solution is saturated with hydrogen chloride gas while being cooled by
ice, followed by stirring at 3 to 24 hours at room temperature, thus
obtaining the compound (2). Then, the compound (2) is dissolved in allyl
alcohol and silver salicylate is added to the solution. The solution is
then stirred for 5 to 24 hours at room temperature, to obtain the compound
(3).
The compound (3) is then dissolved in methanol and sodium methoxide is
added to the solution. The resultant solution is then stirred for 2 to 24
hours at room temperature, to obtain the compound (4).
Then, the compound (4) is dissolved in pyridine, to which tritylchloride is
added. Then, the resultant solution is stirred for 3 to 24 hours at room
temperature, to obtain the compound (5).
The compound (5) is dissolved in pyridine and acetic anhydride is added to
the solution, followed by stirring the solution for 2 to 24 hours at room
temperature, to obtain the compound (6).
Then, 90% aqueous solution of acetic acid is added to the compound (6) and
the thus obtained mixture is stirred for 2 to 24 hours at a temperature
ranging between room temperature and 80.degree. C., to obtain the compound
(7).
The compound (7) and the compound (2) are dissolved in anhydrous
tetrahydrofuran, and the thus formed mixture is added to activated
molecular sieves 4A. Then, silver triflate (AgOTf) dissolved in anhydrous
tetrahydrofuran is added to the molecular sieve containing the compounds
(7) and (2), while the molecular sieves are stirred at a temperature
between -30.degree. and 30.degree. C. Then, after lapse of 0.5 to 6 hours,
the compound (2) dissolved in anhydrous tetrahydrofuran is added thereto,
and stirring is effected for 2 to 24 hours at a temperature between
-30.degree. and 30.degree. C., whereby the compounds (8) and (9) are
obtained.
Secondly, another process for producing the present compounds will be
explained by referring to Process diagram 2.
##STR10##
The compound (1) is subjected to deallylation, to obtain the compound (2),
which in turn is then chlorinated to convert the compound (2) into the
compound (3). Then, the compound (3) is reacted with benzyl lactose to
obtain the compounds (4) and (5). Then, the hydroxy groups of the
compounds (4) and (5) are acetylated, to obtain the compounds (6) and (7),
respectively. The compound (6) is then changed into the compound (8)
through debenzylation, and the compound (8) is further changed into the
compound (9) through acetylation. The compound (9) is then changed into
the compound (10) through hydroxylation, and trichloroacetonitrile is made
to react with the compound (10), whereby the compound (11) is obtained.
Then, benzoylceramide is made to react with the compound (11), to obtain
the compound (12). The compound (12) is subjected to hydrolysis whereby
the compound (13), ganglioside iso GD.sub.3, is obtained.
In this case, the compound (1) corresponds to the compound (9) shown in the
Process diagram 1, and the process therefor is shown in the diagram.
Specifically speaking, the compound (1) is dissolved in a mixture solvent
of ethanol:water:acetic acid=20:5:1. Then, 10% Pd-C is added to the
resultant solution, which is then stirred for 5 hours to 4 days at room
temperature. The reaction solution is filtrated by Celite and iodine is
added to the filtrated solution, followed by stirring for 15 to 60 minutes
at room temperature. The reaction solution is then diluted with water, and
chloroform is added to the diluted solution. After washing with water, the
solution is further washed with sodium and then with saturated brine. The
solution is then dehydrated with anhydrous magnesium sulfate and then the
solution medium is distilled off, to obtain the compound (2).
The compound (2) is dissolved in tetrahydrofuran, to which toluene and
Vilsmeier's reagent [J.C.S., Perkin I, 754-757 (1976)] are added. The
resultant mixture is stirred for 2 hours to one day at room temperature,
whereby the compound (3) is obtained from the reaction solution.
A tetrahydrofuran solution of benzyl lactose and a tetrahydrofuran solution
of the compound (3) are added to molecular sieves, and the mixture is
stirred for 15 to 60 minutes at room temperature. Then, while the mixture
is cooled by ice-methanol, a tetrahydrofuran solution of silver triflate
and a tetrahydrofuran solution of tin chloride are added to the mixture.
After lapse of 30 minutes to 2 hours, a tetrahydrofuran solution of the
compound (3) is added and the mixture is stirred for 3 hours to 2 days,
thus obtaining the compounds (4) and (5).
The compounds (4) and (5) are dissolved in pyridine and acetic anhydride,
respectively, and dimethylaminopyridine is added to the respective
solutions. These solutions are stirred at room temperature for about 2 to
24 hours, to obtain the compounds (6) and (7). Subsequently, the compound
(6) is dissolved in methanol, and is reduced with 10% Pd-C at room
temperature for 2 hours to 2 days, whereby the compound (8) is obtained.
The compound (8) is dissolved in pyridine acetic anhydride and
dimethylaminopyridine is added to the solution, followed by stirring for
about 3 to 24 hours, whereby the compound (9) is obtained.
The compound (9) is dissolved in N,N-dimethylformamide and hydrazinium
acetate is added to the solution. The mixture is then stirred at 5 to 40
minutes at a temperature ranging between room temperature and 80.degree.
C., whereby the compound (10) is obtained.
The compound (10) is dissolved in methylene chloride and, while the
solution is cooled by ice, trichloroacetonitrile and
1,8-diazabicyclo[5,4,0]-7-undecen are added to the solution. The mixture
is then stirred for 1 to 2 hours, whereby the compound (11) is obtained.
The compound (11) and a chloroform solution of benzoylceramide are added to
molecular sieves, and, after stirring for 10 to 30 minutes, boron
triflouride-ether complex is added to the mixture while the latter is
cooled by ice, followed by stirring for about 3 hours to one day, whereby
the compound (12) is obtained.
The compound (12) is dissolved in methanol and tetrahydrofuran and sodium
methoxide is added to the solution. The solution is then stirred for 2
hours to one day at room temperature, and the reaction solution is
subjected to distillation under reduced pressure Subsequently, methanol,
tetrahydrofuran and water are added to the residue, and stirring is
conducted at room temperature for 2 hours to one day. The reaction
solution is then neutralized with IRC-50, to obtain the compound (13).
Further, another process for producing the present compounds will be
explained by referring to Process diagram 3.
##STR11##
The compound (1) is subjected to deallylation to obtain the compound (2),
which in turn is then chlorinated to produce the compound (3). Meanwhile,
the compound (4) is obtained through pyridylsulfidization of the compound
(2). The compounds (5) and (6) are formed by causing benzyl lactose to
react with the compounds (3) and (4), respectively. Then, the compounds
(5) and (6) are acetylated to produce the compounds (8) and (7). The
compound (8) is debenzylated to produce the compound (9), which in turn is
acetylated to produce the compound (10). The compound (10) is deacetylated
to produce the compound (10). The compound (11) is then made to react with
Cl.sub.3 CCN so as to form the compound (12). The compound (12) is then
made to react with the compound having the following formula (15), to
produce the compound (13).
##STR12##
(Bz represents a benzoyl group)
The compound (13) is then subjected to deacetylation, debenzoylation and
saponification, whereby ganglioside iso GM.sub.3 expressed by the general
formula (14) is obtained.
This production method will be explained more detail hereinunder.
10% Pd-C is added to the compound (1) in EtOH-H.sub.2 O-AcOH (20:5:1) and
the mixture is stirred for 1 hour to 4 days at a temperature between room
temperature and 100.degree. C., followed by filtration by Celite. The
filtrated liquid is then condensed under reduced pressure. To the reaction
liquid, 80% THF of iodine is added. The mixture is then stirred for 15 to
60 minutes at room temperature and the reaction solution is diluted with
water. Then, the diluted solution is washed with water after addition of
chloroform. The solution is then washed by aqueous NaHSO.sub.3 solution
and then by saturated brine. The washed solution is then dehydrated with
MgSO.sub.4 and the solution medium is distilled off to produce the
deallylated compound (2).
Then, Vilsmeier's reagent (J.C.S. Perkin 1, 754-757 (1976)) is added and
the mixture is stirred at room temperature for 2 to 24 hours to obtain the
chlorinated compound (3).
THF solution of benzyl lactose and THF solution of the compound (3) are
added to molecular sieves. After stirring at room temperature for 15 to 60
minutes, THF solution of silver triflate (AgOTf) and THF solution of
SnCl.sub.2 are added to the mixture while the mixture is being cooled by
ice-MeOH. The mixture is then stirred for 2 to 48 hours and filtrated by
Celite. The filtrated liquid is then washed by aqueous solution of
saturated sodium bicarbonate, water and saturated brine. The washed
mixture is then dried by anhydrous MgSO.sub.4 and the mixture medium is
distilled off to form the compounds (5) and (6).
2,2'-Dipyridylsulfide and tributylphosphine are added to the compound (2)
in dichloromethane. The resultant mixture is stirred at room temperature
for 2 to 48 hours to obtain the compound (4). Subsequently, a
dichloroethane solution of benzyl lactose and a dichloroethane solution of
AgOTf are added to molecular sieves. While the mixture is cooled by
ice-MeOH, SnCl.sub.2 and a dichloroethane solution of the compound (4) are
added thereto. The mixture is then stirred for 1 to 24 hours at 30.degree.
to 60.degree. C. and is filtrated by Celite. The flitrated liquid is
washed by an aqueous solution of saturated sodium bicarbonate and
saturated brine, and the washed solution is distilled off after drying by
anhydrous MgSO.sub.4, whereby the compounds (5) and (6) are obtained.
Then, pyridine and acetic anhydride are added to the compound (6) so as to
dissolve the latter. Then, dimethyl aminopyridine is added to the
dissolved compound (6) and the mixture is stirred for 1 to 24 hours at
room temperature, to obtain the acetylated compound (7). A similar process
is carried out on the compound (5), to obtain the compound (8).
10% Pd-C is added to the compound (8) in methanol. The mixture is then
catalytically reduced at room temperature for about 2 to 24 hours so as to
obtain the compound (9).
The compound (9) is then dissolved by addition of pyridine and acetic
anhydride, and dimethyl aminopyridine is added to the dissolved compound
(9). The mixture is then stirred at room temperature, obtain the
acetylated compound (10).
The compound (10) is then dissolved in DMF, to which H.sub.2 N.NH.sub.2
AcOH is added. After the resultant solution is stirred for 5 to 60 minutes
at a temperature between room temperature and 80.degree. C., EtOAc is
added to the solution, which is then washed with saturated brine and dried
by anhydrous MgSO.sub.4, followed by distilling-off, to obtain the
compound (11).
The compound (11) is dissolved in methylene chloride, to which
trichloroacetonitrile and DBU (1,8-diazobicyclo [5,4,0 ] undeca-7-en) are
added. The resultant mixture is stirred for 1 to 4 hours, to obtain the
compound (12).
Chloroform solution of the compound (12) and the compound (15) having the
following formula (15) are added to molecular sieves.
##STR13##
(Bz represents a benzoyl group)
Then, BF.sub.3.Et.sub.2 O is added to the mixture while the latter is
cooled by ice-MeOH and the thus formed mixture is stirred for 1 to 24
hours. After filtration by Celite, the mixture is distilled off under
reduced pressure, to obtain the compound (13).
The compound (13) is dissolved in a mixture solvent of MeOH:THF of 1:1,
which is then stirred at room temperature for 2 to 24 hours. After
distilling-off of the medium of the reaction solution, MeOH, THF and
H.sub.2 O are added to the residual material, which is stirred at room
temperature for about 2 to 24 hours. The reaction liquid is then
neutralized by IRC-50, followed by filtration, and the filtrated liquid is
distilled off under reduced pressure, whereby the ganglioside isoGM.sub.3
(14) is obtained as the final product.
UTILITY OF THE INVENTION
Novel compounds of the present invention described above are useful as
tumor markers and cell differentiation markers having differentiation
inducibility, or as synthesis intermediates of such markers.
Examples of the present invention will be described hereinunder. In the
following description of Examples, the numbers of compounds are the same
as those appearing in respective process diagram.
EXAMPLE
A. Examples concerning the process shown in Process diagram 1
REFERENCE EXAMPLE 1
The compound (3) was prepared as follows, in accordance with the teaching
of Carbohydr. Res. 78: (1980), 190-194.
10 g (18.3 mmol) of the compound (1) was dissolved in 50 g of
acetylchloride and the solution was saturated with hydrogen chloride gas
while being cooled by ice, followed by stirring for 15 hours at room
temperature. After distilling off the medium of the reaction solution,
ether was added to the residual material and then distillation was
repeated. In consequence, caramel-like compound (2) was obtained in an
amount of 9.02 g (94%).
Then, 8.42 g (11.5 mmol) of the compound (2) was dissolved in 70 ml of
allyl alcohol, and 5.36 g (21.9 mmol) of silver salicylate was added to
the solution. The solution was then stirred for 15 hours at room
temperature. The resultant reaction solution was filtrated by suction and
then condensed under reduced pressure. To the condensed solution, ethyl
acetate was added and the ethyl acetate layer was washed by water, 5%
sodium thiosulfate, saturated sodium bicarbonate and saturated brine. The
washed solution was then dried by anhydrous magnesium sulfate, followed by
distillation-off of the solution medium under reduced pressure. The
reaction product was then refined by silica gel column [Wakogel (produced
by Wako Junyaku Kogyo Kabushiki Kaisha) C-300, 300 g,
chloroform:ethanol=4:1], to obtain the compound (3) in an amount of 8.05 g
(91%).
Rf=0.57 (chloroform:ethanol=4:1)
melting point 155.degree. to 175.degree. C.
EXAMPLE 1
29 g (54.6 mmol) of the compound (3) was dissolved in 200 ml of methanol
and 20 ml of sodium methoxide was added to the solution, followed by
stirring for 2 hours at room temperature. Then, Amberlist 15, ion exchange
resin (produced by Roam and Hearth Co., U.S.A.) was added to the reaction
solution so as to neutralize the solution, followed by filtration and
distillation of the solution medium under reduced pressure, whereby the
compound (4) was obtained in an amount of 18.9 g (85%).
Rf=0.05 (chloroform:ethanol=4:1).
Rf=0.25 (chloroform:methanol=5:1).
[.alpha.].sup.23 -3.0 (c=1.0 methanol).
Element analysis C.sub.15 H.sub.25 NO.sub.9 1/2 H.sub.2 O. Calculated
value: C, 48.40; H, 7.04; N, 3.76. Measured value: C, 48.51; H, 6.76; N,
3.7.
EXAMPLE 2
8.5 g (23 mmol) of the compound (4) was dissolved in 100 ml of pyridine and
9 g (31.5 mmol) of tritylchloride was added to the solution, followed by
stirring for 6 hours at 50.degree. C. Then, triethylamine was added to the
reaction solution and the solution medium was distilled off under reduced
pressure. To the resultant material, chloroform was added, and the
chloroform layer was washed by water. Then, the washed solution was dried
with anhydrous magnesium sulfate, followed by distilling-off of the
solution medium. The reaction product was then refined by a silica gel
column (Wakogel C-300, 300 g, toluene ethyl acetate=1 5), whereby the
compound (5) was obtained in an amount of 9.6 g (72.8%).
Rf=0.74 (chloroform:methanol=3:2).
Element analysis C.sub.19 H.sub.33 O.sub.9 N. Calculated value: C, 67.42;
H, 6.49; N, 2.3. Measured value: C, 67.51; H, 6.39; N, 2.23.
[.alpha.].sup.21 -5.2.degree. (chloroform c=1.01).
EXAMPLE 3
8.0 g (13.2 mmol) of the compound (5) was dissolved in 50 ml of pyridine,
to which 50 ml of acetic anhydride was added. The solution was then
stirred for 15 hours at room temperature. The reaction solution was then
subjected to azeotropic distillation with toluene for 5 times, followed by
distilling-off of the solution medium under reduced pressure The residue
was recrystallized, whereby the compound (6) was obtained in an amount of
9.47 g (97.5%).
Melting point 216.degree. to 218.degree. C.
Rf: 0.65 (toluene:ethyl acetate=1:5).
[.alpha.].sup.22 +0.2 (chloroform c=1.0).
Element analysis C.sub.40 H.sub.45 NO.sub.12. Calculated value: C, 65.65;
H, 6.20; N, 1.91. Measured value: C, 65.70; H, 6.21; N, 1.90.
EXAMPLE 4
120 ml of a 90% aqueous solution of acetic acid was added to 9.46 g (12.9
mmol) of the compound (6), and stirring was effected for 5 hours at
55.degree. C. The reaction solution was then condensed under reduced
pressure and ethyl acetate was added to the condensed solution. The ethyl
acetate layer was then washed by saturated sodium bicarbonate and water,
and dehydration was conducted with anhydrous magnesium sulfate, followed
by distilling-off of the solution medium under reduced pressure. The
reaction product was then refined by a silica gel column (Wakogel C-300,
600 g, ethyl acetate:methanol=10:0.2), whereby the compound (7) was
obtained in an amount of 3.29 g (52%).
Rf: 0.29 (ethylacetate:methanol=10:0.3).
[.alpha.].sup.22 -29.4.degree. (chloroform c=0.93).
Element analysis C.sub.21 H.sub.31 NO.sub.12. Calculated value: C, 51.53;
H, 6.38; N, 2.68. Measured value: C, 50.92; H, 6.34; N, 2.79.
EXAMPLE 5
To 2 ml of anhydrous tetrahydrofuran, there were added 300 mg (0.6 mmol) of
the compound (7), 398 mg (0.78 mmol) of the compound (2), and 1.5 g of
activated molecular sieves 4A. While the mixture was stirred at
-10.degree. C., 2 ml of anhydrous tetrahydrofuran containing 462 mg (1.8
mmol) of silver triflate was added to the mixture. After lapse of 2 hours,
2 ml of anhydrous tetrahydrofuran containing 214 mg (0.42 mmol) of the
compound (2), and the mixture was stirred for 3 hours at -10.degree. C.
The reaction solution was then filtrated through Celite and ethyl acetate
was added to the filtrated solution. The ethyl acetate layer was washed by
saturated sodium bicarbonate and then by water. Subsequently, dehydration
was conducted with anhydrous magnesium sulfate, followed by
distillation-off of the solution medium under reduced pressure. The
reaction product was then refined by a silica gel column (Wakogel C-300,
100 g, 5% methanol-ethylacetate, then C-300, 50 g, carbon
tetrachloride:acetone 1:1), whereby compounds (8) and (9) were obtained.
Property of Compound (8)
[.alpha.].sup.D -8.2 (chloroform c=1.02).
Element analysis C.sub.41 H.sub.58 O.sub.24 N.sub.2. Calculated value: C,
51.14; H, 6.07; N, 2.91. Measured value: C, 51.42; H, 6.91; N, 2.93.
NMR 400 MHz, CDCl.sub.3, ppm, TMS 1.902, 2.010, 2.017, 2.036, 2.095, 2.157,
2.184, (-COCH.sub.3 .times.9), 2.427, 1H, dd, J=4.88, 12.94, H-2beq,
2.607, 1H, dd, J=4.64, 12.70, H-2aeq, 3.785, 3H, s, OCH.sub.3, 3.794, 3H,
s, OCH.sub.3, 4.89, 1H, m, H-4a, 5.15, 1H, m, H-4b.
Property of Compound (9)
[.alpha.].sub.23.sup.D -17.8 (chloroform c=0.98).
Element analysis C.sub.41 H.sub.58 O.sub.24 N.sub.2. Calculated value: C,
51.14; H, 6.07; N, 2.91. Measured value: C, 50.68; H, 6.04; N, 2.8.
NMR 400 MHz, CDCl.sub.3, ppm, TMS 1.875, 1.903, 2.021, 2.028, 2.058, 2.111,
2.132, 2.144, 2.167, s, COCH.sub.3 .times.9, 1.964, 2H, t, J=7.81, H-3aax,
H-3bax, 2.570, 1H, dd, J=4.64, 12.94, H-3eq, 2.619, 1H, dd, J=4.15, 12.45,
H-3eq, 3.786, 3H, s, OCH.sub.3, 3.790, 3H, s, OCH.sub.3 4.862, 2H, m,
H-4a, H-4b.
B. Examples concerning the process shown in Process diagram 2
EXAMPLE 1
500 mg (0.52 mmol) of the compound (1) was dissolved in 20 ml of a mixed
solvent of ethanol, water and acetic acid at a ratio of
ethanol:water:acetic acid=20:5:1. Then, 250 mg of 10% Pd-C was added to
the mixture, which was stirred for 2 days at 60.degree. C. The reaction
solution was filtrated by Celite and the solution medium was distilled off
under reduced pressure. Then, 40 ml of 80% tetrahydrofuran (20% water) was
added to the residue, and 429 mg (1.8 mmol) of iodine was added thereto,
and stirred for 30 minutes at room temperature. The reaction solution was
then diluted with water and chloroform was added to the diluted solution.
The mixture was then washed with water, followed by washing with an
aqueous solution of sodium hydrogen sulfate and then by washing with
saturated brine. The washed solution was then dried with anhydrous
magnesium sulfate and then the solution medium was distilled. The residue
was refined by a silica gel column (Wakogel by Wako Junyaku Kabushiki
Kaisha: C-300, 50 g, chloroform:methanol=10:0.5), whereby the compound (2)
was obtained in an amount of 400 mg (83.5%).
Rf=0.23 (chloroform:methanol=10:0.5).
Element analysis C.sub.38 H.sub.54 N.sub.2 O.sub.24 +2H.sub.2 O. Calculated
value: C, 47.60; H, 6.09; N, 2.91. Measured value: C, 47.82; H, 5.69; N,
2.89.
EXAMPLE 2
380 mg of the compound (2) was dissolved in 10 ml of tetrahydrofuran, to
which 4 ml of toluene and 960 mg (7.5 mmol) of Vilsmeier's reagent were
added. The mixture was then stirred for 15 hours at room temperature. The
reaction solution was then refined by a silica gel column (Wakogel C-300,
20 g, chloroform:methanol=10:0.5), whereby the compound (3) was obtained
in an amount of 361 mg (94.5%).
Rf=0.25 (chloroform:methanol=10:0.5).
EXAMPLE 3
4 ml of tetrahydrofuran solution containing 845 mg (0.96 mmol) of
4,6-free-benzyl lactose and 1 ml of tetrahydrofuran solution containing
180 mg (0.19 mmol) of the compound (3) were added to 1.5 g of molecular
sieves. The mixture was then stirred for 30 minutes at room temperature.
Then, while the mixture was cooled with ice-methanol, 1 ml of
tetrahydrofuran solution containing 800 mg (3.1 mmol) of silver triflate
and 1 ml of tetrahydrofuran solution containing 200 mg (1.05 mmol) of tin
chloride were added to the mixture. After lapse of 2 hours, 1 ml of
tetrahydrofuran solution containing 181 mg (0.19 mmol) of the compound (3)
was added to the resultant solution, followed by stirring for 15 hours.
The reaction solution was then filtrated with Celite and the filtrated
solution was washed with a saturated aqueous solution of sodium
bicarbonate and then with saturated brine. The washed solution was then
dried by anhydrous magnesium sulfate, followed by distilling-off of the
solution medium. The residue was then refined by a silica gel column
(Wakogel C-300, 100 g, chloroform:methanol=10:0.5), whereby the compound
(4) and the compound (5) were obtained in amounts of 74.1 mg (10.4%) and
11.7 mg (1.6%), respectively.
Property of Compound (4 )
Rf=0.26 (chloroform:methanol=10:0.5).
[.beta.].sub.D.sup.20 -11.74 (c=0.855, chloroform).
Element analysis C.sub.92 H.sub.110 O.sub.34 N.sub.2 +2H.sub.2 O.
Calculated value: C, 61.75; H, 6.20; N, 1.57. Measured value: C, 62.11; H,
6.12; N, 1.55.
Property of Compound (5)
Rf=0.29 (chloroform:methanol=10:0.5).
[.beta.].sub.D.sup.20 -9.67 (c=0.335, chloroform).
EXAMPLE 4
13 mg (0.007 mmol) of the compound (5) was dissolved in a mixture of 3 ml
of pyridine with 3 ml of acetic anhydride. Then, 8 mg of dimethyl
aminopyridine was added to the solution, which was stirred for 15 hours at
room temperature. The medium of the reaction solution was then distilled
off under reduced pressure and the residue was refined by a silica gel
column (Wakogel C-300, 10 g, chloroform:methanol=10:0.5), whereby the
compound (7) was obtained in an amount of 11.2 mg (84%).
Rf=0.39 (chloroform:methanol=10:0.5.
Element analysis C.sub.94 H.sub.112 O.sub.35 N.sub.2 +H.sub.2 O. Calculated
value: C, 60.51; H, 6.26; N, 1.50. Measured value: C, 60.31; H, 5.79; N,
1.83.
EXAMPLE 5
93.8 mg (0.052 mmol) of the compound (4) was dissolved in a mixture of 4 ml
of pyridine with 4 ml of acetic anhydride. Then, 16 mg of dimethyl
aminopyridine was added to the solution, which was stirred for 15 hours at
room temperature. The medium of the reaction solution was then distilled
off under reduced pressure, and the residue was refined by a silica gel
column (Wakogel C-300, 20 g, chloroform:methanol=10:0.5), whereby the
compound (8) was obtained in an amount of 78.5 mg (82%).
Rf=0.27 (chloroform:methanol=10:0.5).
[.alpha.].sup.20 -7.20 (c=0.50, chloroform).
Element analysis C.sub.94 H.sub.112 O.sub.35 N.sub.2. Calculated value: C,
61.69; H, 6.17; N, 1.57. Measured value: C, 61.26; H, 6.41; N, 1.73.
EXAMPLE 6
74 mg (0.04 mmol) of the compound (6) was dissolved in 10 ml of methanol,
which was catalytically reduced for 15 hours at room temperature by 50 mg
of 10% Pd-C. The reaction solution was filtrated by Celite and the
solution medium was distilled off under reduced pressure, whereby the
compound (8) was obtained in an amount of 51 mg (98%).
Rf=0.54, 0.65 (butanol:ethanol:water=2:1:1).
Element analysis C.sub.50 H.sub.74 O.sub.34 N.sub.2. Calculated value: C,
48.15; H, 5.98; N, 2.25. Measured value: C, 48.04; H, 5.88; N, 2.86.
EXAMPLE 7
46 mg (0.037 mmol) of the compound (8) was dissolved in a mixture of 3 ml
of pyridine with 3 ml of acetic anhydride. Then, 20 mg of dimethyl
aminopyridine was added to the solution, which was stirred for 15 hours at
room temperature. The medium of the reaction solution was then distilled
off under reduced pressure, and the residue was refined by a silica gel
column (Wakogel C-300, 10 g, chloroform:methanol=10:0.5), whereby the
compound (9) was obtained in an amount of 55.3 mg (99%).
Rf=0.27 (chloroform:methanol=10:0.5).
Element analysis C.sub.62 H.sub.87040 N.sub.2. Calculated value: C, 49.64;
H, 5.85; N, 1.87. Measured value: C, 50.00; H, 5.55; N, 2.40.
EXAMPLE 8
48.5 mg (0.032 mmol) of the compound (9) was dissolved in dimethyl
formamide, to which 3.6 mg (0.039 mmol) of hydrazinium acetate was added,
followed by stirring for 15 minutes at 60.degree. C. After addition of
chloroform to the solution, the reaction solution was washed with water
and then with saturated brine. The washed solution was then dried by
anhydrous magnesium sulfate, followed by distilling-off the solution
medium under reduced pressure The residue was then refined by a silica gel
column (Wakogel C-300, 10 g, acetone:carbon tetrachloride=2:1), whereby
the compound (10) was obtained in an amount of 40 mg (85%).
Rf=0.36 (acetone carbon tetrachloride=2:1).
EXAMPLE 9
33 mg (0.023 mmol) of the compound (10) was dissolved in 1 ml of methylene
chloride and, while the solution is cooled by ice, 0.17 ml (1.7 mmol) of
trichloroacetonitrile and 6 .mu.l (0.043 mmol) of
1,8-diazabicyclo[5,4,0]-7-undecen were added thereto. The solution was
then stirred for 2 hours and the reaction solution was refined by a silica
gel column (Wakogel C-300, 10 g, acetone:carbon tetrachloride=2:1),
whereby the compound (11) was obtained in an amount of 35 mg (96%).
Rf=0.47 (acetone:carbon tetrachloride=2:1).
NMR 400 MHz, CDCl.sub.3, .delta., ppm, TMS 1.874, 1.902, 1.951, 2.025,
2.040, 2.066, 2.088, 2.095, 2.129, 2.133, 2.141, 2.159, 2.164, 2.181,
COCH.sub.3 .times.15, 2.553, 2H, m, H-3C, H-3d, 3.800, 3H, s, OCH.sub.3,
3.807, 3H, s, OCH.sub.3.
EXAMPLE 10
34 mg (0.021 mmol) of the compound (11) and 50 mg (0.066 mmol) of benzoyl
ceramide were dissolved in 2 ml of chloroform solution, which was added to
1 g of molecular sieves. Then, after stirring for 10 minutes, 15 .mu.l
(0.124 mmol) of boron tetrafluoride-diethylether complex was added to the
mixture, while the mixture is cooled by ice-methanol, and the mixture was
then stirred for 15 hours. The reaction solution was filtrated by Celite,
and then the solution medium was distilled off under reduced pressure.
After the distilling-off, the residue was refined by silica gel column
(Wakogel C-300, 20 g, ethyl acetate, then chloroform:methanol=10:0.5), and
HPTLC (developed at a ratio of acetone:carbon tetrachloride=1:1), whereby
3.6 mg (7.7%) of the compound (12) was obtained.
Rf=0.030 (cholorform:methanol=10:0.5).
[.alpha.].sub.D.sup.19 -7.33 (c=0.15 chloroform).
NMR 400 MHz, CDCl.sub.3, .delta., ppm, TMS 0.878, 6H, t, J=6.10, --CH.sub.3
.times.2, 1.252, s, --CH.sub.2 .times.32, 1.869, 1.899, 1.943, 1.964,
2.013, 2.022, 2.025, 2.064, 2.087, 2.125, 2.135, 2.145, 2.158,
--COCH.sub.3 .times.15, 2.56, 2H, m, H-3C, H-3d, 3.794, 6H, s, OCH.sub.3
.times.2, 5.454, 1H, dd, J=7.26, 13.22,
##STR14##
5.859, 1H, d, t, J=13.88, 6.94, --CH.dbd.CH--CH.sub.2 --, 7.443, 2H, t,
J=8.06, aromatic proton, 7.548, 1H, t, J=7.57, aromatic proton, 8.006, 2H,
d, J=7.08, aromatic proton.
EXAMPLE 11
3.6 mg (0.0016 mmol) of the compound (13) was dissolved in a mixture of 1
ml of methanol with 1 ml of tetrahydrofuran, to which 50 .mu.l of N-sodium
methoxide was added. The mixture thus formed was stirred for 15 hours at
room temperature. The reaction solution was then neutralized with IRC-50,
followed by filtration and distilling-off under reduced pressure. The
residue was refined with Sephadex LH-20
(chloroform:methanol:water=60:30:4.6), whereby the compound (13) was
obtained in an amount of 2.5 mg (96%). Decomposition point 260.degree. to
275.degree. C.
Rf=0.50 (butanol:ethanol:water=2:1:1).
NMR 400 MHz, d-6 DMSO/D.sub.2 O, 98:2 v/v (65.degree.), .delta., ppm, TMS
0.854, 6H, t, J=6.72, CH.sub.3 .times.2, 1.241, s, CH.sub.2 .times.32,
1.874, 3H, s, NHCOCH.sub.3, 1.882, 3H, s, NHCOCH.sub.3, 4.180, 1H, d,
J=7.56, H-1a, 4.232, 1H, d, J= 6.59, H-1b, 5.357, H, d, d, J=7.14, 14.5
Hz, --CH--CH.dbd.CH--, 5.548, 1H, d, t, J=7.14, 14.27, NHCO
--CH.dbd.CH--CH.sub.2
C. Examples concerning the process shown in Process diagram 3
Reference Example 1
2.5 g (4.7 mmol) of the compound (1) was dissolved in 60 ml of EtOH-H.sub.2
O-AcOH (20:5:1) and 2.5 g of 10% Pd-C was added to the solution. The
mixture was then stirred for 15 hours at 60.degree. C. The reaction
solution was filtrated by Celite and the solution medium was condensed
under reduced pressure. The residue was dissolved in 200 ml of 80% THF
(20% H.sub.2 O), to which 2.2 g (8.7 mmol) of iodine was added, followed
by stirring at room temperature for 30 minutes. Then, the reaction
solution was diluted by water, to which chloroform was added. Then, the
mixture was washed with water, further washed by aqueous solution of
NaHSO.sub.3 and then saturated brine. The washed solution was then dried
with MgSO.sub.4, and then the solution medium was distilled off. The
residual product was then refined by a silica gel column (Wakogel
C-300:300 g, chloroform:methanol=10:0.25), whereby the starting material
compound (2) was obtained in an amount of 1.74 g (yield 75%).
Properties of Compound (2)
Rf=0.55 (chloroform:methanol=10:1).
[.alpha.].sub.D.sup.21 -30.8 (c=1.02, CHCl.sub.3).
Reference Example 2
400 mg (0.81 mmol) of the compound (2) was dissolved in 12 ml of
toluene-THF (1:1) mixed solution, and 969 mg (7.57 mmol) of Vilsmeier's
reagent was added to the mixture. The mixture was then stirred at 15 hours
at room temperature. The reaction liquid was then refined by a silica gel
column (Wakogel C-300:20 g, chloroform:methanol=10:0.5), whereby the
compound (3) was obtained in an amount of 300 mg (yield 72.3%).
[Property of Compound (3)]
Rf=0.39 (chloroform:methanol=10:0.5).
Reference Example 3
10 ml of dichloroethane and 0.2 ml of DMF were added to 450 mg (0.92 mmol)
of the compound (2). While cooling the mixture with ice, 200 mg (1.68
mmol) of SOCl.sub.2 was added to the mixture, which was then stirred for
15 hours. The medium of the reaction solution was distilled off and then
azeotropic process by toluene was conducted, to produce the compound (3)
in an amount of 460 mg (yield 98%).
[Properties of Compound (3)]
Rf=0.39 (CHCl.sub.3 :MeOH=10:0.5).
[.alpha.].sub.D.sup.21 -63.degree. (c=1.0, CHCl.sub.3).
NMR (90 MHz, CDCl.sub.3, .delta. (ppm) TMS): .delta. 1.917, 2.059, 2.084,
2.089, 2.218, S, OCOCH.sub.3 .times.5, 2.80, 1H, dd, J=5.0, 13.0, H-3eq,
3.881, 3H, S, --OCH.sub.3, 5.20, 1H, m, H-4.
EXAMPLE 1
2 ml of THF solution of 820 mg (0.92 mmol) of Benzyl O-(2,
3-di-O-benzyl-.beta.-D-guluctopyranosyl)-(1.fwdarw.4)-2,3,6-tri-O-benzyl-.
beta.-D-glucopyranoside and 1 ml of THF solution of 150 mg (0.29 mmol) of
the compound (3) were added to 1.5 g of molecular sieves. The mixture was
stirred at room temperature for 1 hour. While cooled by ice-MeOH, 1 ml of
THF solution containing 800 mg (3.1 mmol) of AgOTf and 1 ml of THF
solution containing 200 mg (1.05 mmol) of SnCl.sub.2 to the mixture. After
lapse of 2 hours, 1 ml of THF solution was added to the mixture, which was
stirred for 15 hours. The reaction solution was filtrated by Celite and
was washed successively by an aqueous solution of saturated bicarbonate,
water and saturated brine, followed by drying by anhydrous MgSO.sub.4 and
distilling-off, whereby a reaction product in an amount of 284 mg (yield
26%) was obtained. The reaction product was then refined by a silica gel
column (Wakogel C-300:100 g, toluene:ethylacetate=1:2), whereby the
compound (5) and the compound (6) were obtained in amounts of 256 mg
(yield 23.5%) and 27 mg (yield 2.5%).
[Property of Compound (5)]
Rf=0.31 (toluene:ethylacetate=1:2).
[.alpha.].sub.D.sup.21 -53.3 (c=1.01, | | |
