SEMMELWEIS UNIVERSITY
DOCTORAL SCHOOL OF PHARMACEUTICAL AND PHARMACOLOGYCAL SCIENCES
Chemical Deglucosylation of Secologanin and Vincoside
Derivatives
Ph.D. thesis
László Károlyházy
Under the supervision of Professor László F. Szabó
Semmelweis University
Department of Organic Chemistry
Director: Professor Péter Mátyus
Previous director: Professor László F. Szabó
Budapest, 2002.
Chemical Deglucosylation of Secologanin and Vincoside derivatives
Ph. D. Thesis
László Károlyházy
Under the supervision of Professor László F. Szabó, Ph. D.
Semmelweis University, Department of Organic Chemistry, 2002
Summary
The indole alkaloids are formed in the plants by the Mannich-type condensation of the monoterpenoid
glucoside secologanin and tryptamine. The first product of this coupling reaction catalyzed by strictosidine
synthase is strictosidine (with 3S configuration). Under enzyme-free conditions, both strictosidine and its 3R
epimer vincoside are formed in a 1:1 ratio. Strictosidine is the precursor of more than 2200 indole and related
alkaloids. The key reaction of their formation the deglucosylation. The removal of the glucosidic subunit by the
enzyme b-glucosidase is well known in the literature. However, the deglucosylation by aqueous acid or base was
used only sporadically. The main subject of this work is to study the chemical (non-enzymatic) acid-catalyzed
hydrolytic and the aminolytic deglucosylation.
1. Epimer-free strictosidine was prepared from secologanin with tryptamine in the presence of the
strictosidine synthase presented by A. I. Scott. The enzyme proved to be highly substrate-specific and
completely stereoselective. In strictosidine, the S configuration of C-3 was confirmed, and the main
conformations established.
2.1. The secologanin having the C-7 atom as the part of a formyl group gave lactams with primary
amines, and lactones with secondary or by catalytic effect of tertiary amines, without deglucosylation.
Secologanin derivatives protected by acetalisation or coupled with tryptamine at C-7 were deglucosylized by
primary amine with simultaneous incorporation of the amino subunit. The aglucone obtained from benzyl
vincoside could be transformed by treatment of acetic acid into a new, fused pentacyclic ringsystem with
participation of N-1.
2.2. The secologanin ethylene acetal was fragmented with a secondary amine into a C3 unit which proved
to be methyl 3-piperidinoacrylate and a C7 unit, which could not be identified. In aqueous acid, secologanin as
well as its ethylene acetal were fragmented likewise into a C3 unit which could not be identified, and a C7 unit,
which proved to be benzaldehyde.
3.1 In the aminolytic deglucosylation of benzylvincoside and benzyl-oxostrictosidine, both fragments
could be identified. The C3 unit proved to be again methyl 3-piperidinoacrylate, and the C7 fragment was found
as a second N-benzyl group in the appropriate (oxo)tryptamine derivatives.
3.2. In aqueous acid, the vincoside derivatives protected on neither, either, or both nitrogen atoms
afforded tetra- or pentacyclic aglucones, whose possible structures and formations were studied by graph
analysis.
4. The compounds obtained by our methods were compared with those obtained by enzymatic analysis or
as alkaloids from natural sources.
The structures (including, in most cases, also the stereochemistry) of the prepared compounds were
proved by chemical correlations and nuclear magnetic resonance (NMR) spectroscopy.
2
Introduction and program
The large family of the indole and related alkaloids containing more than 2200 individual compounds is
constructed in a Mannich-type reaction from two building blocks, secologanin and tryptamine,. In this coupling
reaction, a new center of chirality (Mannich center, C-3) is built up with complete stereoselectivity under the
catalytic effect of the enzyme strictosidine synthase. The further alkaloids are formed by subsequent
transformations and in large variety, first of all in the plant families Apocynaceae, Loganiaceae és Rubiaceae.
Many of them have characteristic biological effects, and had important influence on the elaboration of potent
drugs, as well as on the development of the methods applied in the synthesis and structure elucidation of organic
compounds.
Since two decades, the chemical background of the biogenetic processes of these alkaloids have been
thoroughly studied in the Department of Organic Chemistry of the Semmelweis University under the direction of
Prof. László F. Szabó. The investigations were focused on the stereoselective coupling reaction and on the
deglucosylation and its consequences.
Secologanin and the primary products formed from it are glucosides, however, the majority of the
alkaloids are aglucones. For understanding of their biogenesis, it was necessary to investigate the
deglucosylation, as the great synthetic potential of secologanin and tryptamine could be developped only after
the removal of the glucosyl group. In the plant celles, the deglucosylation is catalyzed by a glucosydase enzyme.
The same method is frequently used in the chemistry of the indole alkaloids under cell-free conditions. However,
its applicability is limited by the enzyme specifity. Chemical, i. e. non-enzymatic deglucosylation was used only
sporadically in this field. Therefore, the aim of our work was to elaborate chemical methods for the
deglucosylation of the secologanin derivatives. In the topics of simple secologanin derivatives, some result were
already obtained in our Department, and they should have been extended to the coupled products.
As an introductory step, it was necessary to confirm the configuration of C-3 (Mannich center) in the first
product of the biosynthesis. In the coupling reaction of secologanin with tryptamine, two epimers, strictosidine
and vincoside may be formed. While in the vincoside series, this configuration was unequivocally proved by X
ray diffraction analysis to be R, in the strictosidine series, there were only indirect indication for the S
configuration of it.
Therefore, the program of the dissertation work was as follows:
1. Preparation of epimer-free strictosidine and confirmation of the configuration at the Mannich center.
2. Further investigation of the acid-catalyzed hydrolytic and aminolytic deglucosylations in simple
secologanin derivatives.
3. Application of the methods mentioned above in the secologanin derivtives coupled with tryptamine.
4. Interpretation of the results from the point of view of the biogenesis of the indole alkaloids.
In order to achieve these purposes, it was necessary to prepare the appropriate educts and products of the
deglucosylations, and to prove their structure (including their stereostructure) by chemical correlations and
nuclear magnetic resonance (NMR) spectrscopy. It was expected, that the execution of this program can present
an at least partial insight into the chemical background of the biogenesis of the indole alkaloids.
3
Scientific results
1.1 Strictosidine synthase enzyme was prepared from the plasmide and according to the
prescription presented by A. I. Scott.
1.2. Preparation of strictosidine (77)* from secologanin and tryptamine with the catalysis of
strictosidine synthase. The process runned with 98% chemical yield and with complete
stereoselectivity (Figure 1).
10
10
9
11
8
12
13
6
1
N
4
3
2
NH
18
H
H
12
13
7
5
20
15
22
16
1
N
19
14
H3CO
6
8
5
H
H
9
11
7
O HO
21
2´
O
1´
O
3´
3
2
H
H
OH
5´
16
14
17
R = acetyl
21
20
19
OH
O
O
22
15
OH
6´
N
H
4´
17
4
O
O RO
18
H
H
18
O
H
H
H
19
20
HR
4
N
H H
H
H
H
H
7
13
9
12
10
H
1
N
H
16
17
3O
2
8
HS
H
H
H
H
H
H
O O
H
H
H
4'
O
H
5'
6'
H
H
H
H
11
8
H 13 H
1
H
H
H
4
3
N
1'
O
H
O
22
16
14
14
H
H
17
O
15
21
H 20
H
H
4'
O
5'
R
H
H
H
H
6'
O
R
H
R = acetyl
H
11
H
O
5
2
H
H
O
2'
7
N
3' H
O
H
6
R
O
H
H
H
12
H
O
OR
9
3'
H
H
O
O
2'
1'
21
15
14
5
6
H
O
OR
H
10
H
H
OR
O
19
H
H 18
H
O',O',O',O'-tetraacetyl-18,19-dihydrostrictosamide
strictosidine
Figure 1. Three-dimensional structure of strictosidine and O’,O’,O’,O’-tetraacetyl-18,19-dihydrostrictosamide.
1.3. The 3S configuration of the Mannich center was confirmed, and the main conformations of
strictosidine established by its chemical correlation with O´,O´,O´,O´-tetraacetyl-18,19-dihydrostrictosamide (91) and by detailed NMR investigations (3S, 15S, 20S, 21S, S11PP). By using the
graph and the expected coupling constant pattern around C-14 shown in Figure 2, it was possible to
select the single stereostructure obtained in the enzymatic coupling reaction out of the possible 648
stereoisomers.
*
The numbering of compounds is the same used in thesis.
4
H-3 H-15
H-14R
H-14S
H14R
az R sorban:
X = N4
Y = C2
H15
H14S
ap sc
3
sc ap H
P
3-14
M
3-14
C16
16
C
H
C
C20
X
Y sc sc
sc ap
H14S
14R
H
H3
S31
R32
P
14-15
20
C
H14S
H15
15
H
C16
C
P
3-14
Y
C20
S22(N)
R11(N)
P
14-15
M
14-15
Y
H14S
H14R
sc ap
3
sc sc H
P
S32 14-15
R31
C20
C
C20
H15
C16
H
Y
H
C20
S13
R23
H14R
sc sc
H3 ap sc
H15
P
3-14
15
S23
R13
M
3-14
Y
X
H14R
sc sc
sc sc
P
3-14
14S
C16
16
Y
H14S
ap sc
sc sc H3
H14R
sc ap
H3
ap sc
16
H14R
X
M
3-14
X
X
C20
C16
H15
S12(N)
P
R21 14-15
P
3-14
X
M
14-15
Y
H14S
ap ap 3
sc sc H
P
14-15
S21
R12(P)
M
14-15
H14R
X
H14R
sc sc
H3 ap ap
20
P
3-14
P
14-15
C16
Y
H15
15
C20
S11(P)
R22(P)
H14S
X
az S sorban
X = C2
Y = N4
Y
X
H14S
H3
S33
R33
Figure 2. Conformations around C-14.
1.4. According to our investigations, the strictosidine synthase has a very high substrate
specifity and a complete stereoselectivity. It was established, that the enzyme accepts, over
secologanin (1) and tryptamine (2), only 8,10-dihydrosecologanin (14) as substrate, out of several ones
investigated by us.
2.1. The acid-catalyzed hydrolytic removal of glucosyl unit of secologanin (1) and its ethylene
acetal (20) gave benzaldehyde (166) as single product. Its formation was interpreted by fragmentation
of the aglucone into a C3 and C7 unit followed by aromatization of the C7 fragment, the carbon atoms
5
of which are at the appropriate oxidation level. Unfortunately, the hypthetic C3 fragment could not be
isolated (Figure 3).
10
O
H
6
H
9
11
10
O
7
H
H
Oglc
O
4
3
O
X
4
3
N
2´
O
3´
H
9
11
4´
O
Oglc
O
4
2
H
6
1
10
8
7
O
163: X=NCH3, Y=a-OH
168: X=NCH3, Y=b-OH
170: X=O, Y=a-OH
5´
6´
H
3
O
20 : 8,10-vinyl
165: 8,10-ethyl
11
6
5
3
O
1 : 8,10-vinyl
14: 8,10-ethyl
H3CO
7
1
H3CO
2
Og
Oglc Y
O
10
8
8
1
5
H3CO
O
8
7
9
1
5
H3CO
NR
11
4
3
O
171: R=CH3, 8,10-vinyl
172: R=propil, 8,10-vinyl
173: R=propil, 8,10-ethyl
glc = b-D-glucopyranosyl
174
Figure 3. Simple secologanin derivatives.
2.2. Deglucosylation with amine involved its simultaneous incorporation into the aglucones.
However, in the case of secologanin (1) having C-7 as a free formyl group, the primary amine
associated to C-7, and gave as final product the lactam epimeric pair of 7-hydroxy-N-methylbakankozine (7a-OH: 7b-OH = 2:1) (163, 168) without deglucosylation. The b-hydroxy isomer (168)
could be isomerized into the thermodynamically more stable a-isomer.
2.3. As with secondary amine, the lactamization is already not possible, 7-hydroxy-sweroside
was formed. Only the 7a-hydroxy epimer could be isolated (Figure 3.).
2.4. When the formyl group of secologanin was protected as ethylene acetal (20), the amine
associated to C-3. With primary amine, after deglucosylation, with O®N isoster exchange and
elimination of water, the original dihydropyran ring was transformed into a dihydropyridine ring (171,
172 és 173) both in the secologanin- and the 8,10-dihydrosecologanin-ethylene acetal (20 és 165).
Also the secondary amine piperidine associated at C-3, however, in this case the deglucosylation was
followed by fragmentation into C3 and C7 units, of which only the C3 unit could be isolated as methyl
3-piperidinoacrylate (174) (Figure 3.).
2.5. It seems, that the deglucosylations carried out by aqueous acid and secondary amine are
complementary processes, as the two isolated fragments C3 (methyl 3-piperidinoacrylate) and C7
(benzaldehyde) were formed from the different carbon atoms of the same C10 secologanin aglucone
subunit.
3.1. Our investigations were also extended to such derivatives of secologanin which were
prepared in the coupling reaction with tryptamine (2) and oxotryptamine (5) (Figure 4.)
6
5
7
H
N
3
N
H
Bn
18
H
14
O
20
14
H
O
16
O
20
19
17
H
18
176
Bn = benzyl; glc = b-D-glucopyranosyl
18
19
14
H3CO
O
15
H
20
Oglc
21
H
22
O
16
21
17
O
113: 18,19-vinyl, R=H
115: 18,19-ethyl, R=H
117: 18,19-vinyl, R=CH3
181: 18,19-ethyl, R=CH3
Oglc
81 : 18,19-vinyl
120: 18,19-ethyl
O
H
R
17
Bn
N
3
N
16
15
21
H
H3CO
H
Oglc
22
N
3
N
H
19
Figure 4. The educt glucosides.
3.2. The aminolytic deglucosylation of benzylvincoside (113) and benzyl-oxovincoside (176)
was immediately followed by fragmentation. Like to the simpler secologanin derivatives, in both
compounds, the amine associated to the b-carbon atom of the a,b-unsaturated ester system (C-17), and
after deglucosylation, the intermediate was cleaved into a C3 fragment and a tryptamine derivative
containing the C7 fragment of the secologanin subunit. In these cases, both fragments could be isolated
(Figure 5.).
Bn
N
N
N
3
2
H
H
O
189
20
15
19
15
14
18
3
N
H
54
N
H
H3CO
Bn
N
18
19
21
H
4
O
H
20
16
Bn
18
19
H
H
Oglü
H
20
O
H
N
N
H
H3CO
N
17
N
O
193
190
O
16
17
Oglü
21
Bn
21
20
15
19
14
18
N
H
3
N
H
O
138
Bn
3
N
H
O 14
H
Bn
O
21
15
22
H
20
H
H CO
3
N
18
CH3
19
O
16
17
O
182
Figure 5. Fragmentation of the coupled compounds.
The C3 fragment proved again to be methyl 3-piperidinoacrylate (174). However, C7 fragment
was found as a second benzyl group in N,N-dibenzyltryptamine (54) and N,N-dibenzyl-oxotryptamine
(189), respectively. For the formation of this benzyl group, the C7 fragment had to be reduced, which
was interpreted by an internal redox system. The reduction was investigated in detail in the
fragmentation of (113). It was found, that parallel to the reduction of the C7 fragment, the C-16–C-17
7
single bond of the primary adduct benzyl-17-piperidino-16,17-dihydrovincoside (190) was oxidized
into a double bond by which benzyl-17-piperidinovincoside (193) was formed. This product, although
not in completely pure form, could be isolated. The process was completed by a reducing, and stopped
by an oxidating agent.
3.3. In the acid-catalyzed hydrolytic deglucosylation of benzyl-oxostrictosidine (176), benzyloxostrictosidine-aglukone (182) was isolated as a main product, which could be formed, after opening
of the dihydropyran ring, by isomerization and O-17®C-19 cyclization. This aglucone type
corresponds to the most stable one (39) (Figure 7.) of the N-methyl bakankosine, as it was already
established previously in our Department. However, the formation of benzyl-oxotryptamine (138) and
benzaldehyde (166) as side products were also observed. These fragments were probably formed in the
retro-Mannich reaction and subsequent fragmentation.
4. During the aminolytic deglucosylation of benzylvincoside (113) by primary amine, like to
simpler derivatives of secologanin, fragmentation was not observed. Also in this case, the
dihydropyran ring was transformed into a dihydropyridine ring (197) (Figure 6.). By treatment of this
product with glacial acetic acid, the pentacyclic compound having a seven-membered ring (198) was
obtained by N-1®C-19 and N-17®C-21 cyclization. Its complete structure including (also the
stereochemistry) was established (3R, 15S, 19S, R210N) by NMR spectroscopy and using the
information included into the graph of Figure 2. By catalytic hydrogenation of compound 198, the
benzyl group could be removed (199) and the seven-membered ring partially cleaved (200), but a
rearrangement from N-1 to N-4 was not observed.
N
H H
H3CO
3
Bn
N
18
19
14
15
20
N
16
17
O
197
H
19
21
H
22
18
20
15
H
16
N
Bn
18
17
22
H
19
OCH3
20
15
H3C
17
H3C
198
H
16
N
18
14
21
O
NH
3
N
14
21
CH3
N
3
N
OCH3
22
14
19
20
15
21
H
16
N
O
NH
3
N
H H
17
OCH3
22
O
H3C
199
200
Figure 6. Deglucosylation of a coupled product by amine without fragmentation.
5.1. Acid-catalytic hydrolytic deglucosylation of vincoside derivatives was studied in detail. In
these processes, derivatives of benzyl-vincoside and its 18,19-dihydroanalogue protected on either,
neither or both N atoms were used as educts in which N-1 was protected by methylation, and N-4 by
benzylation. The stereochemistry of the compounds was determined as described previously. and the
results are given in parentheses. No fragmentation was observed. In the different cyclizations, four
nucleophilic and four electrophilic sites may take part, which allow two types of cyclizations both in
four variations (Figure 7.). The first type involves only the secologanin subunit and results in oxa- or
carbacyclization, in the second type, further connection is built between one of the N atoms of the
tryptamine subunit and a site in the secologanin subunit, and results in azacyclization. In these
8
processes, two series of aglucones may be formed depending on the cyclization to N-1 (aglucone type
A) or N-4 (aglucone type B). The pentacyclic aglucones are formed by combination of the
oxa/carbacyclization and the azacyclization.
in single-aglucons
10
8
H
9
O
5
1
O-1®C-3 (2)
3
H
4
11
C-10®C-3 (10)
O-3®C-8 (11 and 12)
O-3®C-1 (1 and 9)
O
O
9
oxa/carbacyclizations
5
4
11
O
1
H
8
10
3
H
O
O
in alkaloid-aglucons
1
3
N
H
R1
4
NR4
H
H
20
H3CO
22
O
21
17
H
O
O
4
1
R1=H or Me
R4=H or Bn
18
19
3
N
H
R1
NR4
20
oxa(carba)cylizations
O-21®C-17 (3)
O-17®C-19 (7)
C-18®C-17 (9)
O-17®C-21 (5 )
H3CO
O
21
H
22
O
18
19
17
H
O
azacyclizations
3
N
R1 H
O
21
A
H
18
4 NR4
H
19
O
22
H
17
OCH3
N(1 or 4)®C-17 (2)
N(1 or 4)®C-21 (4 )
N(1 or 4)®C-22 (6 )
N(1 or 4)®C-19 (8)
1
3
N
R1
H
4
NR4
O
H
H
21
18
B
H3CO
H
22
19
O
17
O
O
Figure 7. Possible cyclizations in the alkaloide aglucones.
5.2. When both N atoms were unprotected, the deglucosylation was preceded by lactamization,
consequently only the deglucosylation of vincosamide (81) and its dihydro derivative (120) could be
studied. It was established, that in these reactions as well as in the deglucosylation of glucosides
protected at both N atoms (117 amd 181) compounds 202, 206, 204, 208 were formed according to
cyclizations O-17®C-21 and O-17®C-19, respectively (Figure 8.). These compounds have the same
structure in their secologanin subunit as had N- methylbakankosine and its 8,10;dihydro derivative.
Previously, it was already demonstrated in our Department, that the aglucones 39 and 29 were the
most stable ones under the circumstances applied during deglucosylation. The aglucones have the
following stereochemical characterizations: 202: 3R, 15S, 19S:19R=3:4, 20S, R12NN; 204: 3R, 15S,
19S:19R = 3:1, 20S, R11NP; 206: 3R, 15S, 20S, 21R, R12NN; 208: 3R, 15S, 20S, 21R, R11NP.
5.3. In the hydrolytic deglucosylation of the glucosides unprotected at N-1, and protected at N-4
(113 and 115), the first cyclization resulted in the same changes in the secologanin subunit of the
aglucones as described in point 5.2. However, they were followed by an azacyclization between N-1
and C-21 (Figure 8.). The glucoside 113 with a vinyl group afforded 203 having a new fused, the
9
educt 115 with ethyl group gave 207 having a new bridged, seven-membered ring in the pentacyclic
system. In debenzylation (203´ and 207´) of the aglucones, no rearrangement from N-1 to N-4 was
observed. Their stereochemical characteristics are as follows: 203’: 3R, 15S, 19S, 20S, 21R, R21PN,
207’: 3R, 15S, 20S, 21S, R21dPN.
H
1
N
3
H
4
N
H
1
O
22
N
H
4
O
H
O
19
O
H
N
3
H
4
N
N
H
17
+ H3COH
O
H
N
H
N
Bn
4
N
1
3
O
17
205
Bn
H
1
H
N
H
H3CO
OH
O
207
N
H
H3C
+ H2O
O
O
4
21
3
H
OH
H3CO
17
OCH3
206
19
21
H
OH
H
O
+ H2O
17
21
H
H3CO
O
H3C
O
21
H3C
N
17
4
3
21
H
H
3
204
19R:19S = 1:3
H
1
O
22
4
N
O
203
202
19R:19S = 4:3
1
H3CO
OCH3
17
H
21
H
1
O
19
O
19
H
3
H3C
21
Bn
4
N
1
N
Bn
3
17
21
H
N
17
O
208
209
O
H
H
1
OH
N
H
O
N
H3CCOO
O
4
H
NH
H
21
H
H
O
29
N
20
H
19
O
4
21
O
17
17
H
NH
1
N
OCH3
OCH3
H
O
O
39
O
203´
207´
Figure 8. Products of the acid-catalyzed hydrolytic deglucosylations.
5.4. The educt glucosides unprotected at N-4 and protected at N-1 could not be prepared,
because of their spontaneous lactamization. Therefore, the acid-catalyzed rearrangement was studied
in situ in the derivatives obtained by debenzylation with catalytic hydrogenation of the appropriate
benzyl aglucones 204 and 208. Under the catalytic hydrogenation carried out in aqueous acid, as a
result of izomerizations and N-4®C-21 cyclization, the pentacyclic 205 and tetracyclic 209 aglucones,
respectively, were isolated. Their stereochemical characteristics are: 205: 3R, 15S, 19S, 20R, R31NN,
209: 3R, 15S, 20?, R130N.
5.5. In order to interprete the results of deglucosylation, graphs (Figures 9 and 10) were
constructed.
10
N-1®C-19
O-17®C-21
N-1®C-19
O-21®C-17
H
1
N
N-1®C-17
O-21®C-17
19
H
1
17
H
N
+ H2O
H
1
N
O
* *
17
HO
O
HO
H
*
*
O
17
O
HO
H
17
H
18
O
H
N
HO
17
*
O
+ H3COH
H O
HO
18
19
HO
–
21
*
VE
17
*
18
24
*
H
1
N
18
17
9
18 19
–
H
NH
+ H2O
23
N-1®C-17
C-18®C-17
NH
*
*
21
O
H
1
H
1
N
O
HO
21
19
H
1
6(H)
21
H
17
O
19
N
CO2 Me
*
* *
18
*
–
*
19
18
H
21
7
17
H
O-17®C-19
17
H
1
N
*
HO
H
H
18 17*
*
CO2 Me
19
*
*
O
22
OH
N-1®C-21
C-18®C-17
NH
*
203
H
*
CO2 Me
17
17
O
NH
*
H
O
O
NH
*
H
19
CO2 Me
N-1®C-19
O-17®C-19
+ H3COH
H O
O
H
1
N
21
*
+ CH4
22
NH
*
19
O
CO2 Me
8
H
1
N
H
*
NH
*
N-1®C-17
O-17®C-19
22
H
17(H)
204
N-1®C-19
CO2 Me *
*
H
*
H
NH
*
VE
H
18
H O
O
17
H
1
N
H
MeO2C 22
*
* *
NH
*
CO2 Me
HO
N-1®C-22
H
N
VE
*
O
21
19
17
H
H
NH
O
H
22
1(H)
O
H
H
NH
*+ H3COH
22
VE
CO2 Me
17
N
N-1®C-19
C-18®C-17
N
O
H O
*
O
21
H
1
*
C-18®C-17
CO2 Me
O
H
H
N
H
MeO2C 22
H
*
H
*
19
H
NH
** *
18
H
17
H
5(H)
NH
H
1
CO2 Me
22
O
H
N-1®C-17
O-17®C-21
VE
*
21
H
HO
H
N
H
O
11
H
CO2 Me
16(H)
O-17®C-21
H
1
N-1®C-17
H
+ H 2O
207
VE
*
208
H
*
*
17
NH
*
17
19
N-1®C-22
C-18®C-17
17
3(H)
21
25
O
H
1
N
H
NH
N
O
22
–
CO2Me
22
21
H
CO2 Me
17
H
1
H
2(H)
19
VE
N-1®C-21
H 18
*
N
*
N-1®C-21
O-17®C-21
–
4(H)
NH
*
*
MeO2C 22*
NH
18
NH
NH
H
*
19
** 15(H)
21
OH
H
*
*
17
3
VE
O
OH
O-21®C-17
21
H
H
*
19
NH
*+ H COH
22
14
*
*
O
H
1
N
H
21
1
21
HO
18
1
*
*
18
H
N
+ H3COH
H
22
17
H
1
N
CO2 Me
H
–
NH
H
H
O
NH
10(H)
1
N
O
1
*
22
21
13
12(H)
11(H)
H
1
N
HO
CO2 Me
N-1®C-22
O-17®C-21
17
CO2 Me
N-1®C-22
O-21®C-17
H
*
17*
VE
*
H
NH
*
*
**
H
*
21
H
O
19
OH
*
21
N-1®C-21
O-21®C-17
21
*
CO2 Me
O
NH
*
H
1
N
NH
*
**
19
20
N-1®C-22
O-17®C-19
21
N-1®C-21
O-17®C-19
Figure 9. Possible cyclizations to N-1. Aglucone types of the series A.
The reaction matrices shown in the graphs are (with one exception) under thermodynamic
control. If N-1 is protected, and N-4 is free, the aglucone formed in the reaction mixture (204 and 208)
will be immediately stabilized by a second reaction (205 and 209, respectively) before having been set
in of the thermodynamic control. If N-4 is protected, and N-1 is free (113 and 115), the most stable
aglucone (203 and 207, respectively) will be formed suppressing the lactamization, which is
disfavored because of the non-planarity of the amide group. If both N atoms are free (81 and 120), the
lactamization to N-4 precedes the deglucosylation and affords the most stable aglucones (202 and
206). This structure is the more stable subunit under the circumstances of the deglucosylation, too. If
both N atoms are protected, the deglucosylation will be stopped at the tetracyclic level (205 and 209).
The secologanin subunit of all vincoside aglucone types are on the most stable level, which is formed
by O-17®C-19 cyclization in the “natural” series, and O-17®C-21 cyclization in the dihydro series.
By graph analysis it was possible to select the isolated aglucones as most favored structures, and to
find the shortest ways of their formations out of the large number of possibilities.
11
N-4®C-19
O-21®C-17
N-4®C-19
O-17®C-21
4
N-4®C-17
O-21®C-17
N
19
H
*
*
H
4N
*
H
H
19
*
4
*
H
N-4®C-21
O-21®C-17
4
O
*
+ H3COH
*
H
4
O
OH
17
10(H)
*H*
*
H
*H
**
H
18
H
H
*
19
O
22
H
O
H
*
*17
H
17
21
*H
H
H
*
*
22
H3CO
OCH3
H
O
*H
1(H)
H
O
21
VE
H
O
H3CO 22
4 N 19
H
O
O
18
H
H3CO
*
**
17
H
24
**
9
H
4 NH
18
H
–
H
*
OCH3
+ H2O
*
4
H
23
O
H
8
O
–
O
21
H
*
N
H
O
–
O
H
22
H3CO
18
N 21
OH
*H * H
* * 19
O
17
17
O
H
O
*
N 19 H
O
O
*
17
+ CH4 H
H
205
OH
4
H
H3CO
H
* *
H 3CO
7
4
4
O
* 19
* *
N-4®C-17
O17®C-19
17
OH
21
OCH3
O
19
O
*
N 17 H
H
O-17®C-19
N-4®C-19
18
N-4®C-17
C-18®C-17
H
*
22
O
N 17 H
O
*H*
*
H3CO
4
N 19
4
*
H
18
17
18
OH
21
O
H 18
**
21
H
17(H)
O
H
O
21
H3CO 22
O
N-4®C-19
C-18®C-17
*H
17
4
H
N-4®C-22
204
OH
C-18®C-17
OH
H
O
*
* *
17
6(H)
H
H
*
19
18
H
H
H
*
+ H3COH
17 H
OCH3
O
VE
O
N 22
*H
H
O
21
17
H
O
H
H
*
5(H)
N
*
H
4
19
H
4
VE
19
VE
25
N-4®C-17
O-17®C-21
O-17®C-21
O
4 NH
O
16(H)
OH
O
17
3(H)
17
+ H2O
H
21
4 NH
H
18
H
**
O
N-4®C-17
17
H
H3CO 22
–
O
H3CO
19
*
H
208
H
21
*
18
NH
N
*
VE
4
OH
2(H)
OH
4
O
–
N-4®C-21
O-17®C-21
VE
N-4®C-21
O
VE
21
OH
H
4(H)
19
N 17 OH
VE
N
*
O
–
18
19
O-21®C-17
4
* *
15(H)
H
*
22
18
NH
H3CO 22
21
O
N-4®C-22
C-18®C-17
*
H
O
+ H3COH
17
206
OH
21
209
H3CO
O
N
*H *
H
–
O
H
14
O
H
H
21
N 22
+ H3COH
O
N 22
*
H
17
13 OH
* 17
* * OH
H
4
O
H3CO
4
11(H)
*H
H
O
21
O
N-4®C-22
O-21®C-17
H
N-4®C-22
O-17®C-21
OH
21
H
21 H
H3CO
4
*
* * O
17
H3CO
O
12(H)
N
VE
H
OCH3
+ H2O
*
H
N 17 H
H
* *
21
*
+ H3COH
N 22
H
19
O
202
O
N-4®C-19
O-17®C-19
17
* * O
19
H
O
20
N-4®C-22
O-17®C-19
21
N-4®C-21
C-18®C-17
N-4®C-21
O-17®C-19
Figure 10. Possible cyclizations to N-4. Aglucone types of the series B.
6.1. The comparison of the results of the deglucosylation by acid-catalyzed hydrolysis with
those obtained by enzymatic deglucosylation described in the literature indicated the characteristic
differences in the thermodynamic versus kinetic control.
6.2. In the subgroup of structures involving cyclization to N-1, the comparison of the aglucones
obtained in our experiments with those of alkaloids isolated from natural sources suggested the
conclusion that N-1 cyclization may take place only, if the cyclization to N-4 is hindered because of
some structural reasons. The graph of N-1 cyclizations (Figure 9.) contained (at least partially) all
aglucone types, which are involved in the natural alkaloids, and helped to prognose the most
reasonable routs of their formation.
Publications
Articles
12
1. Károlyházy L.; Patthy Á.; Podányi B.; Szabó L. F.: Rekombináns sztriktozidin szintáz enzim
előállítása és szubsztrátspecifitásának vizsgálata. Gyógyszerészet, 1995, 39, 447-448.
2. Károlyházy L.; Patthy Á.; Szabó L. F.; Podányi B.: Aromatizációval járó fragmentációk a
szekologanin és indolalkaloidok kémiájában. Gyógyszerészet, 1996, 40, 481-482.
3. Patthy-Lukáts, Á.; Károlyházy, L.; Szabó, L. F.; Podányi, B.: First and Detailed Stereochemical
Analysis of Strictosidine. J. Nat. Prod., 1997, 60, 69-75.
4. Károlyházy L.; Patthy Á.; Szabó L. F.: Vinkozid aglükonok képződése és régiószelektív
gyűrűzáródása. Gyógyszerészet, 1998, 42, 539-543.
5. Károlyházy, L.; Patthy-Lukáts, Á.; Szabó, L. F.: Chemistry of Secologanin. Part 5. Graph
Analysis of the Acidic Deglycosylation of Vincoside Derivatives. J. Phys. Org. Chem., 1998, 11,
622-631.
6. Károlyházy, L.; Patthy-Lukáts, Á.; Szabó, L. F.; Podányi, B.: Chemistry of Secologanin. Part 7.
Chemical Fragmentation of Secologanin: Biomimetic Transition from the Aliphatic into the
Aromatic Skeleton. Tetrahedron Letters, 2000, 41, 1575-1578.
7. Károlyházy, L.; Patthy-Lukáts, Á.; Podányi, B.; Szabó, L. F.: Chemistry of Secologanin. Part 12.
Preparation of Aglycones Derived from Vincoside. (ready for publication).
Lectures and poster presentations
1. Károlyházy L.; Patthy Á.; Szabó L. F.; Podányi B.: Acetálformában védett szekologanin kémiai
reakciói. IX. Gyógyszerészkongresszus Budapest, 1993. szeptember 19-22.
2. Károlyházy L.; Patthy Á.; Podányi B.; Szabó L. F.; Scott A. I.; Fodor I.: Sztriktozidin szintáz
enzim termeltetése és szubsztrátspecifitás vizsgálata. III. Semmelweis Tudományos Fórum
Budapest, 1994. április 28. (Poster)
3. Károlyházy L.: Rekombináns sztriktozidin szintáz enzim előállítása és szubsztrátspecifitásának
vizsgálata. II. Clauder Ottó Emlékverseny Budapest, 1994. augusztus 24.
4. Károlyházy L.; Patthy Á.; Szabó L. F.; Podányi B.: Aromatizációval járó fragmentációk a
szekologanin és indolalkaloidok kémiájában. Pharma Kollokvium Budapest, 1995. április 5.
5. Podányi, B.; Károlyházy, L.; Patthy, Á.; Szabó, L. F.: Determination of the Conformation and
Configuration of Strictosidine. 50 Years of NMR Budapest, 1995. április 27-28.
6. Károlyházy L.; Patthy Á.; Podányi B.; Szabó L. F.: Deglükozilezést kísérő fragmentációk és
ciklizációk a szekologanin és indolalkaloidok kémiájában. MTA Alkaloidkémiai Munkabizottság
tudományos ülése Balatonfüred, 1995. május 3-4.
7. Károlyházy L.; Patthy Á.; Podányi B.; Szabó L. F.: A szekologanin származékok aromatizációval
járó fragmentációja. Vegyészkonferencia Debrecen, 1995. augusztus 27-28. (Poster)
8. Károlyházy L.; Patthy Á.; Podányi B.; Szabó L. F.: Vinkozid aglükonok újabb gyűrűzáró reakciói.
MTA Alkaloidkémiai Munkabizottság tudományos ülése Balatonfüred, 1996. április 25.
13
9. Patthy Á.; Károlyházy L.; Podányi B.; Morvai M.; Szabó L. F.: Régiószelektívitás vinkozid származékok savas deglükozilezésénél. X. Gyógyszerészkongresszus Budapest, 1996. május 15-17.
10. Károlyházy L.: Vinkozid aglükonok aromatizációval járó fragmentációja és régiószelektív
gyűrűzáródása. III. Clauder Ottó Emlékverseny Budapest, 1996. szeptember 26-28.
11. Károlyházy, L.; Patthy, Á.; Szabó, L. F.; Podányi, B.: Application of Graph Analysis in the
Bioorganic Reactivity of Alkaloidtype Secologanin Derivatives. Sixth European Symposium on
Organic Reactivity Louvain-la Neuve (Belgium), 1997. július 24-29. (Poster)
12. Károlyházy L.; Patthy Á.; Szabó L. F.: Gráfanalízis alkalmazása a triptaminnal kapcsolt
szekologanin származékokból képződő aglükonok esetében. Vegyészkonferencia Siófok, 1997.
Szeptember 1-3. (Poster)
13. Károlyházy, L.; Patthy, Á.; Szabó, L. F.; Podányi, B.: Application of Graph Analysis in the
Bioorganic Reactivity of Alkaloidtype Secologanin Derivatives. VI. Semmelweis Tudományos
Fórum Budapest, 1997. november 5-7. (Poster)
14. Károlyházy L.; Patthy Á.; Szabó L. F.: Indolalkaloid képződés kulcslépésének vizsgálata enzim
jelenlétében és távollétében. MTA Bioorganikus Kémiai Munkabizottság tudományos ülése Pécs,
1998. május 11-12.
15. Szabó L. F.; Patthy Á.; Károlyházy L.; Podányi B.: Terpenoid eredetű vegyületek biomimetikus
típusú aromatizációja. MTA Bioorganikus Kémiai Munkabizottság tudományos ülése Visegrád,
1999. szeptember 9-10.
14