Chemo-enzymatic total synthesis of 3-epiaustraline, australine, and 7-epialexine

Article abstract of DOI:10.1021/jo000933d

Sequential enzymatic aldol reaction and bis-reductive amination leads to the total syntheses of tetrahydroxylated pyrrolizidine alkaloids, 3-epiaustaline (14), australine (1), and 7-epialexine (11). This approach allows for their rapid construction without the need for protecting group manipulation of the hydroxyl functionality. In addition, an improved procedure for the asymmetric epoxidation of divinyl carbinol (3) was described, and the product was used in a concise synthesis of the required triol 7 and ent-7.

Full text of DOI:10.1021/jo000933d

8
264
J . Org. Chem. 2000, 65, 8264-8268
Ch em o-En zym a tic Tota l Syn th esis of 3-Ep ia u str a lin e, Au str a lin e,
a n d 7-Ep ia lexin e
Alex Romero and Chi-Huey Wong*
Department of Chemistry and the Skaggs Institute for Chemical Biology, The Scripps Research Institute,
1
0550 North Torrey Pines Road, La J olla, California 92037
wong@scripps.edu
Received J une 20, 2000
Sequential enzymatic aldol reaction and bis-reductive amination leads to the total syntheses of
tetrahydroxylated pyrrolizidine alkaloids, 3-epiaustaline (14), australine (1), and 7-epialexine (11).
This approach allows for their rapid construction without the need for protecting group manipulation
of the hydroxyl functionality. In addition, an improved procedure for the asymmetric epoxidation
of divinyl carbinol (3) was described, and the product was used in a concise synthesis of the required
triol 7 and ent-7.
Naturally occurring tetrahydroxlated pyrrolizidines
such as australine (1) and alexine (2) (Figure 1), and their
stereoisomers, have been isolated from the genera
Castanospermum and Alexa.1 Interest in their total
synthesis has arisen due to their ability to inhibit
2
glucosidases, as well as for their antiviral and retroviral
3
activities. Although there are several methodologies for
F igu r e 1.
Sch em e 1. Retr osyn th etic An a lysis of
constructing simple pyrrolizidines, no direct de novo
synthetic methods have been developed until recently for
4
constructing complex molecules such as 1 and 2. Many
Au str ia lin es a n d Alexin es
of the earlier synthetic methods that have been reported
entailed routes employing carbohydrates as starting
5
materials, all of which, including the most recent ap-
proaches, require protecting group strategies.
6
a,b
Synthetic efforts toward iminocyclitols by us
and
others6 have demonstrated the utility of a sequential
enzymatic aldol and reductive amination reaction se-
quence for the stereoselective construction of hydroxlated
piperidines and pyrrolidines. In our continuing efforts to
c,d
*
To whom correspondence should be addressed. Tel: (858) 784-
487. Fax: (858) 784-2409.
1) Alexine isolation: (a) Nash, R. J .; Fellows, L. E.; Dring, J . V.;
Fleet, G. W. J .; Derome, A. E.; Hamor, T. A.; Scofield, A. M.; Watkin,
D.J . Tetrahedron Lett. 1988, 29, 2487. Australine isolation: (b)
Molyneux, R. J .; Benson, M.; Wong, R. Y.; Tropea, J . E.; Elbein, A. D.
J . Nat. Prod. 1988, 51, 1198.
2
(
expand the scope of enzymes in organic synthesis, we now
illustrate the first chemo-enzymatic approach to the
synthesis of bicyclic iminocyclitols, 3-epiaustraline, aus-
traline, and 7-epialexine, that bypasses the need for
protecting group manipulation of the hydroxyl function-
ality. The retrosynthesis is outlined in Scheme 1. The
highly functionalized pyrrolizidine skeleton was envi-
sioned to arise from an intramolecular bis-reductive
amination reaction of a nonprotected tetrahydroxylated
keto-aldehyde, which sets the stereocenter corresponding
(
2) (a) Elbein, A. D.; Molyneux, R. J . Chemical and Biological
Perspectives; Wiley: New York, 1986; Vol. 5. (b) Elbein, A. D. Annu.
Rev. Biochem. 1987, 56, 497. (c) Elbein, A. D. FASEB 1991, 5, 3055.
(
d) Nishimura, Y. In Studies in Natural Products Chemistry, Vol. 10;
Elsevier: Amsterdam, 1992. Fellows, L. E.; Nash, R. J . Sci. Prog.
Oxford) 1990, 74, 245.
3) (a) Elbein, A. D.; Tropea, J . E.; Molyneux, R. J . US Pat. Appl.
(
(
US 289, 907; Chem Abstr. 1990, 113, P91444p. (b) Fellows, L. E.; Nash,
R.J . PCT Int. Appl. WO GB Appl. 89/7951; Chem. Abstr. 1991, 114,
43777.
7
to C-3. This keto-aldehyde would be generated from a
1
(
4) (a) Denmark, S. E.; Hurd, A. R. J . Org. Chem. 2000, 65, 2875-
stereospecific enzymatic aldol reaction between dihy-
droxyacetone phoshpate (DHAP, available in four steps)
2
2
7
886. (b) Denmark, S. E.; Herbert, B. J . Org. Chem. 2000, 65, 2887-
896. (c) Denmark, S. E.; Herbert, B. J . Am. Chem. Soc. 1998, 120,
357-7358. (d) Denmark, S. E.; Martinborough, E. A. J . Am. Chem.
8
Soc. 1999, 121, 3046-3056. (e) White, J . D.; Hrnciar, P.; Yokochi, A.
(6) (a) Gijsen, H.; Qiao, L.; Fitz, W.; Wong, C.-H. Chem. Rev. 1996,
96, 443-473 and references therein. (b) Look, G. C.; Fotsch, C.; Wong,
C.-H. Acc. Chem. Res. 1993, 26, 182-190. (c) Hung, R.; Straub, J . A.;
Whitesides, G. M. J . Org. Chem. 1991, 56, 3849-3855. (d) Straub, A.;
Effenberger, F.; Fischer, P. J . Org. Chem. 1990, 55, 3926-3932.
(7) For a related example of a bis-reductive amination of dicarbonyl
sugars for the construction of monocyclic iminocyclitols, see: Baxter,
E. W.; Reitz, A. B. J . Org. Chem. 1994, 59, 3175-3185.
F. T. J . Am. Chem. Soc. 1998, 120, 7359-7360.
(5) (a) Pearson, W. H.; Hines, J . V. Tetrahedron Lett. 1991, 32, 5513.
(
b) Ikota, N. Tetrahedron Lett. 1992, 33, 2553. (c) Choi, S.; Bruce, I.;
Fairbanks, A. J .; Fleet, G. W.; J ones, A. H.; Nash, R. J .; Fellows, L. E.
Tetrahedron Lett. 1991, 32, 5517. (d) For a review of the reported
syntheses of hydroxylated pyrrolizidine alkaloids, see: Giovanni, C.;
Zanardi, F.; Rassu, G.; Pinna, L. Advances in the Stereoselective
Synthesis of Hydroxylated Pyrrolizidines. Org. Prep. Proced. Int. 1996,
(8) J ung, S,-H.; J eong, J .-H.; Miller, P.; Wong, C.-H. J . Org. Chem.
1994, 59, 7182-7184.
2
8, 641-682.
1
0.1021/jo000933d CCC: $19.00 © 2000 American Chemical Society
Published on Web 11/05/2000

Total Synthesis of 3-Epiaustraline
J . Org. Chem., Vol. 65, No. 24, 2000 8265
Sch em e 2a
Sch em e 3a
a
Conditions: (a) EtOH, 65%.
mixture of diastereomers.14 The major isomer was sepa-
rated and submitted to sodium periodate cleavage fol-
lowed by aldol addition reaction with DHAP mediated
by fructose-1,6-diphosphate aldolase (FDPA, obtained
from rabbit muscle). Enzymatic cleavage of the phosphate
group with acid phosphatase (Pase) led to 8 in 25%
yield.15
The NMR spectrum of 8 showed an ∼7:1 mixture of
1
the pyranose 9 and ketone 8. COSY and H NMR analysis
of the peracetate 10 established the cis orientation
between substituents at C-4 and C-5 (J H4-H5 ) 1.8 Hz)
and thus established formation of syn product 7 as the
major stereoisomer resulting from the allyl indium ad-
dition reaction.
a
Conditions: (a) (-)-DIPT, cumene hydroperoxide, CH3Cl3, -45
°
C, 86%, >99% ee; (b) 0.5 N NaOH, 98%; (c) 30% NH4OH, rt, then
ethylformate, EtOH, 90 °C, 95%; (d) O3, MeOH, -78 °C, then In,
allyl bromide, H2O, rt, 56% (after separation of diastereomers);
(
e) NaIO4, H2O, then DHAP, FDPA, then Pase, 25%; (f) O3,
MeOH-H2O, -78 °C, then H2 Pd/C, then HCl and H2 Pd/C, 66%.
DIPT ) diisopropyl tartrate, DHAP ) dihydroxyacetone phos-
phate, FDPA ) fructose-1,6-diphosphate aldolase, Pase ) acid
phosphate.
A key feature in the successful enzymatic aldol reaction
was the mild generation of a sensitive R-N-formyl alde-
hyde through the use of NaIO
4
in water.16 In addition,
the choice of the N-formyl protecting group was instru-
mental, since the enzymatic aldol reaction between
DHAP and the analogous N-Cbz-protected aldehyde
failed to give any desired aldol product.
and an appropriate aldehyde to contruct the stereocenters
corresponding to C-1 and C-2 of australines and alexines.9
Since the final two sequences do not require protection
of the hyroxyl groups, we planned a synthesis of the
required aldehyde that would be amenable to this non-
protecting group strategy.
The synthesis begins with Sharpless asymmetric ep-
oxidation of divinylcarbinol (3) to furnished epoxide 4 in
excellent yield with high enantiomeric purity (86%, >99%
ee)10 (Scheme 2). In our attempts to repeat and optimize
The final bis-reductive amination sequence was ac-
complished by ozonolysis of 9, followed by a reductive
workup of the ozonide with Pd/C and hydrogen. Cleavage
of the formamide with HCl in the presence of Pd/C and
hydrogen resulted in concomitant reduction of the inter-
mediate iminium ion affording 7-epialexine (11) (66%)
17,18
as a 8:1 mixture of stereoisomers.
Concentration of
the known literature procedure for the epoxidation of this
the reaction mixture was required after the ozonolysis
step in order to avoid the N-methylated side product 12,
arising from competitive intermolecular reductive ami-
nation with formaldehyde.
substrate,1
1a-c
we discovered that substituting cumene
hydroperoxide1 for tert-butyl hydroperoxide produced
1c
better results. Base-induced rearrangement of epoxide
4
followed by regioselective nucleophilic opening with
In a related example (Scheme 3), while studying the
reactivity of highly functionalized iminium ions, we
observed that in the absence of a hydride source, protic
solvents (i.e., MeOH and EtOH) serve as nucleophiles to
give the mixed N,O-ketals in moderate yields. When
1
1d
ammonia
and protection of the resulting amine with
1
2
ethyl formate gave the formamide 6. Attempts at
selective ring opening of the epoxide 5 using either NaN
or TMSN failed, since only a mixture of regioisomeric
3
3
azides was obtained. Ozonolysis of 6 afforded a mixture
of hemiacetals that were directly subjected to allyl
bromide and indium,13 which generated triol 7 as a 3:1
(13) Paquette, L. A.; Bennett, G. D.; Isaac, M. B.; Chatriwalla, A.
J . Org. Chem. 1998, 63, 1836-1845.
(
14) The use of other allylating reagents and conditions (i.e.,
(
9) All four possible stereoisomers can be obtained depending on the
allyltributylstannane) gave lower selectivity.
choice of aldolase enzyme, see ref 6a and: Zannetti, M. T.; Knorst, M;
(15) We detected ∼10-15% of a third stereoisomer, tentatively
assigned as the product arising from epimerization of the starting
aldehyde.
Fessner, W.-D. Chem. Eur. J . 1999, 5, 1882-1890.
(10) For comparison of optical rotation with known literature values,
see ref 11a-c.
(16) Other oxidative or reducing stategies to generate this aldehyde
would most likely lead to facile epimerization. For a review on optically
active N-protected R-amino aldehydes, see: J urczak, J .; Golebiowski,
A. Chem. Rev. 1989, 89, 149-164.
(
11) (a) J ager, V.; Schroter, D.; Koppenhoefer, B. Tetrahedron 1991,
4
7, 2195-2210. (b) Schreiber, S. L.; Schreiber, T. S.; Smith, D. B. J .
Am. Chem. Soc. 1987, 109, 1525-1529. (c) Katsuki, T.; Martin, V. S.
Organic Reactions; Paquette, L. A., Ed.; Wiley: New York, 1996; Vol.
(17) The major stereoisomer was assigned as 7-epialexine upon
comparison of physical and spectral data to literature values, see refs
5a and 18.
4
7
8, pp 1-300. (d) J ager, V.; Stahl, U.; Hummer, W. Synthesis 1991,
76-782.
(
12) Other formylating conditions (i.e., acetic anhydride and formic
(18) Fleet, G. W.; Haraldsson, M.; Nash, R. J .; Fellows, L. E.
Tetrahedron Lett. 1988, 29, 5441-5444.
acid) gave low yields.

8
266 J . Org. Chem., Vol. 65, No. 24, 2000
Romero and Wong
Sch em e 4a
divinyl carbinol 3 was described leading to a concise
synthesis of the required triol 7 and ent-7.
Exp er im en ta l Section
Gen er a l Meth od s. The reagents and solvents were reagent
grade and used as supplied. Fructose-1,6-diphosphate aldolase
(rabbit muscle) and acid phosphatase (sweet potato) were
purchased from Sigma. Solvent evaporation was performed
under reduced pressure using a rotary evaporator, followed
by evacuation (<0.1 mmHg) to constant sample weight. Silica
gel 60 (230-240 mesh) or reverse phase silica gel (RP-18, 40-
6
3 mesh)was used in chromatograhpy. Standard conditions
using Ac O and pyridine were employed for the acetylation
reactions.
2
Mod ified P r oced u r e for th e P r ep a r a tion of (2S,3R)-
,2-Ep oxy-4-p en ten -3-ol (4). A mixture of crushed 4 Å
1
molecular sieves (4.0 g) and CH
2 2
Cl (120 mL) was cooled to
-35 °C, and titanium tetraisopropoxide (3.5 mL, 11.9 mmol)
and (R,R)-(-)-diisopropyl tartrate (3.3 mL, 15.5 mmol) were
added by syringe. After the mixture was stirred at -35 °C for
30 min, divinylcarbinol (10 g, 119 mmol) was added by addition
funnel, followed by cumene hydroperoxide (35 mL, 238 mmol).
The reaction mixture was stirred at -35 °C for 36 h. Aqueuous
a
Conditions: (a) NaIO4, H2O, DHAP, FDPA, then Pase, 30%;
(
b) O3, MeOH-H2O, -78 °C, then H2 Pd/C, then HCl and H2 Pd/
C, 70%; (c) O3, MeOH-H2O, -78 °C, then HCl, rt, then NaOAc,
NaCNBH3, AcOH, 52%. DHAP ) dihydroxyacetone phosphate,
FDPA ) fructose-1,6-diphosphate aldolase, Pase ) acid phos-
phatase.
saturated Na
2
SO
4
(10 mL) was added, and the mixture was
diluted with Et
2
O (100 mL). After the mixture was stirred at
ambient temperature for 3 h, the resulting slurry was filtered
through a pad of Celite, and the resulting yellow solution was
concentrated. Excess cumene alcohol and cumene hydroper-
oxide were removed by silica gel chromatography (Hex/EtOAc,
employing anhydrous MeOH and shorter reaction times
in the reductive amination leading to 11, similar mixed
N,O-ketals products were detected. This observation may
provide a new route to iminocyclitols containing a leaving
group as inhibitors of glycosidase.
ent-7 was prepared using similar methods starting
from ent-4 and submitted to sodium periodate cleavage
and enzymatic aldol reaction as described for 7 (Scheme
4
:1 then 100% Et
2
O). Kugelrohr distillation (120 °C, 30 mm/
25
Hg) provided 10.2 g (86%) of 4 as a colorless oil: [a]
D
-53° (c
)); H NMR (500
) δ 5.83 (ddd, J ) 16.9, 10.6, 6.2 Hz, 1H), 5.38 (br
d, J ) 17.2 Hz, 1H), 5.25 (d, J ) 10.6 Hz, 1H), 4.33 (br s, 1H),
11a
25
1
0
.73, CHCl
3
) (lit. [R]
D
-49° (c 0.73, CHCl
3
MHz, CDCl
3
3
1
4
.09 (dd, J ) 6.6, 2.9 Hz, 1H), 2.80-2.74 (m, 2H), 2.05 (br, s,
H); 13C NMR (125 MHz, CDCl
) δ 135.4, 117.6, 70.1, 53.9,
3
3.5; LRMS (ESI) m/z 99 (M - H).
4
). Pyranose 13 was obtained as a 1:1 mixture of
1
5
(2R,3R)-2,3-E p oxy-4-p en t en ol (5). Epoxide 4 (4.0 g, 40
anomeric stereoisomers in 30% yield. Ozonolysis of this
mixture and intramolecular bis-reductive amination
produced 3-epiaustraline (14) in 70% yield as a single
isomer.2
The demonstrated facial selectivity of the reductions
leading to 7-epialexine (11) and 3-epiaustraline (14) was
mmol) was added to 40 mL of 0.5 N NaOH and stirred for 20
min at room temperature. Extraction with CH Cl
(3 × 100
mL), drying of the combined organic layers (Na SO ), and
concentration gave 3.9 g (98%) of 5 as a colorless oil: [R]
2
2
2
4
0,21
25
D
1a
22
1
+50° (c 2.0, CHCl
3
) (lit.1 [R]
D
3
+56° (c 2.5, CHCl )); H NMR
(
500 MHz, CDCl
3
) δ 5.61-5.54 (m, 1H), 5.49 (dd, J ) 17.3, 1.5
Hz, 1H), 5.29 (dd, J ) 10.3, 1.5 Hz, 1H), 3.91 (dd, J ) 12.9,
consistent with related reductions of iminium ions,
leading to simple cis-pyrrolizidines.22 Surprisingly, when
1.8 Hz, 1H), 3.65 (dd, J ) 12.5, 3.3 Hz, 1H), 3.39 (dd, J ) 7.7,
2
.2 Hz, 1H), 3.07-3.05 (m, 1H), 2.53 (br s, 1H); 13C NMR (125
the reductive amination of 13 was carried out with
NaCNBH , australine (1) was produced as the major
3
MHz, CDCl
9 (M - H).
2S,3S)-3-N-F or m yla m in e-4-p en ten e-1,2-d iol (6). Ep-
3
) δ 134.6, 120.0, 61.2, 60.1, 55.9; LRMS (ESI) m/z
9
2
0
stereoisomer (8:1) in 52% yield. It is possible that the
inversion in facial selectivity arises from a hydroxyl-
directed iminium ion reduction, where the reducing agent
is a soluble hydride source.7
In summary, a short chemoenzymatic method utilizing
a stereospecific aldol reaction coupled with selective bis-
reductive amination have been applied toward the total
synthesis of tetrahydroxylated pyrrolizidines, 3-epiau-
straline, australine, and 7-epialexine. This strategy can
be potentially be used to access other stereoisomers and
analogues related to the hydroxylated pyrrolizidine class
of alkaloids. Additionally, an improved epoxidation of
(
oxide 5 (3.9 g, 39 mmol) was added to a 30% solution of
ammonium hydroxide (50 mL) at room temperature. After 24
h, the solution was concentrated to give 4.5 g (quantitative)
2
5
11d
22
of pure amino diol: [R]
D
-17° (c 1.0, MeOH) (lit. [R]
c 1.4, MeOH)); H NMR (500 MHz, CDCl ) δ 5.94 (ddd, J )
7.3, 10.3, 7.3 Hz, 1H), 5.20 (dd, J ) 17.3, 10.3 Hz, 2H), 3.65-
D
-19°
1
(
3
1
3
13
.61 (m, 1H), 3.55-3.52 (m, 2H), 3.41-3.38 (m, 1H); C NMR
) δ 137.7, 115.4, 74.3, 63.4, 56.2; LRMS (ESI)
m/z 118 (M + H).
(125 MHz, CDCl
3
EtOH (10 mL) was added to a mixture of amino diol (4.5 g)
and ethyl formate (32 mL, 400 mmol). The homogeneous
solution was heated at 90 °C for 18 h. To cleave the formate
esters, the solution was concentrated, and MeOH (100 mL)
was added. This solution was heated to 80 °C for 12 h. The
solution was concentrated, and resulting oil was subjected to
(
(
19) Romero, A.; Wong, C.-H. Unpublished results.
20) 3-Epiaustraline and australine were assigned based upon
comparison of physical and spectral data to literature values, see ref
d.
(
separated (isomerically enriched to 75%) and submitted to ozonolysis
and reductive amination to give 14 as a single isomer, thus providing
evidence that pyranose 13 is a mixture of anomeric stereoisomers.
silica gel chromatograhy (CH
2 2
Cl /MeOH, 7:1) to afford 5.4 g
2
5
4
(95%) of formamide 6 as a colorless oil: [R]
D
-15° (c 1.0,
OD) δ 8.10 (s, 1H), 5.85 (ddd,
J ) 17.1, 10.5, 6.6 Hz, 1H), 5.19-5.13 (m, 2H), 4.49 (br t, 1H),
.59-3.56 (m, 1H), 3.50-3.42 (m, 2H); 13C NMR (125 MHz,
CD OD) δ 163.3, 135.0, 117.9, 74.5, 64.4, 53.9; LRMS (ESI)
m/z 146 (M + H).
21) Each of the two pyranose 13 stereoisomers were partially
1
MeOH); H NMR (500 MHz, CD
3
3
(
22) Hesse, M.; J anowitz, A.; Vavrecka, M. Tetrahedron Lett. 1991,
3
3
2, 5543-5546.

Total Synthesis of 3-Epiaustraline
J . Org. Chem., Vol. 65, No. 24, 2000 8267
(
2S,3S,4R)-3-N-F or m yla m in e-6-h ep ten e-1,2,4-tr iol (7).
A solution of 6 (2.7 g, 19 mmol) and MeOH (70 mL) was cooled
to -78 °C, and a gentle stream of O was bubbled into the
Hydrogen chloride (0.04 mL, 4 M in dioxane, 0.18 mmol)
was added to a mixture of aldehyde (prepared above), pal-
3
ladium on carbon (10%) (5 mg), and MeOH-H
2
O (4 mL, 6:1)
solution until a faint blue color appeared. The ozonide was
quenched with dimethyl sulfide (2.7 mL, 37 mmol) at -78 °C,
and the reaction was allowed to warm to room temperature.
The solution was concentrated, and the yellow oil was used in
the next step without purification.
Indium powder (2.1 g, 19 mmol) and allyl bromide (3.2 mL,
7 mmol) were added to a solution of aldehyde (prepared
above) and H O (40 mL). The mixture was stirred vigorously
for 12 h, at which point 1 N NaOH was added to bring the
reaction mixture to pH 6.5. Celite (1.0 g) was added, and the
resulting slurry was filtered and concentrated to produce 3.5
g (98%) of 7 as 3:1 mixture of diastereomers. Purification by
under an atmosphere of H (balloon). After 72 h, the mixture
2
was filtered and concentrated to yield 33 mg of crude product.
This residue was loaded onto Dowex 50W-X2 ion-exchange
resin (200 mg) in H
NH OH. After concentration, the yellow residue was submitted
to reversed-phase silica gel chromatography (100% H O) to
afford 21 mg (70%) of 3-epiaustraline (14) as a single isomer
2 2
O and eluted with H O followed by 2 N
4
2
3
2
5
4d
25
(
1
(
white solid): [R]
D
+5.9° (c 1.0, MeOH) (lit. [R]
D
+6.2° (c
2
1
.0, MeOH)); H NMR (600 MHz, D O) δ 4.39 (br t, 1H), 4.27
t, J ) 2.6 Hz, 1H), 4.08 (br t, 1H), 3.91 (dd, J ) 12.1, 5.3 Hz,
2
1
1
3
H), 3.82 (dd, J ) 12.3, 7.4 Hz, 1H), 3.54 (dd, J ) 4.6, 2.9 Hz,
H), 3.43-3.40 (m, 1H), 3.18 (ddd, J ) 12.3, 10.0, 6.2 Hz, 1H)
.02-2.99 (m, 1H), 1.95-1.86 (m, 2H); 13C NMR (150 MHz,
2 2
silica gel chromatography (CH Cl /MeOH, 7:1) afforded 2.0 g
2
5
1
D
2
O) δ 77.3, 75.1, 73.1, 68.9, 63.5, 55.7, 44.9, 33.9; HRMS
MALDI-FTMS) m/z 190.1071 (M + H, 190.1074 calcd for
N).
Au str a lin e (1). A 1:1 mixture of anomeric isomers 13 (25
mg, 0.10 mmol) and MeOH (2 mL) was cooled to -78 °C, and
a gentle stream of O was bubbled into the solution until a
faint blue color appeared. The ozonide was quenched with
palladium on carbon (10%) (5 mg) under an atmosphere of H
of 7: [R]
D
3
+57° (c 1.0, MeOH); H NMR (600 MHz, CD OD)
(
δ 8.12 (s, 1H), 5.78-5.75 (m, 1H), 5.02-4.96 (m, 2H), 4.06-
8 16 4
C H O
4
.04 (m, 1H), 3.84 (d, J ) 8.4 Hz, 1H), 3.65-3.63 (m, 1H), 3.57
(
2
dd, J ) 11.7, 3.4 Hz, 1H), 3.45 (dd, J ) 12.8, 5.9 Hz, 1H),
1
3
.34-2.32 (m, 2H); C NMR (150 MHz, CD
17.8, 72.3, 69.2, 64.6, 53.0, 39.8; HRMS (FAB) m/z 190.1081
N).
P yr a n ose (13). Sodium periodate (1.4 g, 6.4 mmol) was
added to a solution of triol ent-7 (1.0 g, 5.3 mmol) and H
13 mL) at room temperature. After 30 min, BaCl (647 mg,
.7 mmol) was added, and the resulting precipitate was filtered
3
OD) δ164.5, 135.8,
1
3
(
8 16 4
M + H, 190.1079 calcd for C H O
2
(
balloon) at -78 °C, and the reaction was allowed to warm to
2
O
room temperature. The solution was concentrated and the
colorless oil was used in the next step without purification.
(
2
2
through a pad of Celite. Dowex 50W-X8 acidic resin was added
to the filtrant until the solution became acidic (pH 1.1). The
resin was filtered, and the pH of the solution was adjusted to
Hydrogen chloride (0.08 mL, 4 M in dioxane, 0.3 mmol) was
added to a solution of aldehyde (prepared above) and MeOH-
2
H O (4 mL, 6:1). After 72 h, sodium acetate (25 mg, 0.3 mmol)
6
.7 with 2 N NaOH solution. Dihydroxyacetone phosphate
was added followed by sodium cyanoborohydride (8 mg, 0.12
mmol) and acetic acid (0.03 mL, 0.50 mmol). After the mixture
was stirred for 36 h at room temperature, 1 N HCl was added
to bring the solution to pH 1.1. The mixture was concentrated,
and the residue was loaded onto Dowex 50W-X2 ion-exchange
8
2
(DHAP) (17 mL, 0.37 M in H O, 6.4 mmol) was added, and
the pH was readjusted to 6.7 with 2 N NaOH. Fructose-1,6-
diphosphate-aldolase (500 units, from rabbit muscle) was
added, and the mixture was stirred slowly at room tempera-
ture. After 72 h, the pH of the mixture was adjusted to 4.7,
acid phosphatase (sweet potato) (250 units) was added, and
the mixture was incubated at 38 °C for 12 h. The reaction
mixture was neutralized to pH 7.0 with 2 N NaOH and
concentrated. MeOH (100 mL) was added, and the mixture
was filtered though a pad of Celite and concentrated. Silica
gel chromatography (CH Cl /MeOH, 6:1) gave 390 mg (30%)
2 2
of 13 as a 1:1 mixture of anomeric isomers. Characterization
data for the triacetate of each anomeric isomer is provided.
Data for the first triacetate off the silica gel column (1:1, CH
resin (100 mg) in H
NH OH. After concentration, the yellow residue was submitted
to reversed-phase silica gel chromatography (100% H O) to
afford 10 mg (52%) of australine (1) as a 8:1 stereoisomeric
mixture. Data for australine (1): 1H NMR (600 MHz, D
O) δ
2 2
O and eluted with H O followed by 2 N
4
2
2
4.27 (dt, J ) 4.0, 2.2 Hz, 1H), 4.14 (t, J ) 7.9 Hz, 1H), 3.80
(dd, J ) 9.4, 8.1 Hz, 1H), 3.70 (dd, J ) 11.8, 3.5 Hz, 1H), 3.51
(dd, J ) 11.6, 6.6 Hz, 1H), 3.09 (dd, J ) 7.5, 4.3 Hz, 1H), 3.05
(ddd, J ) 9.9, 7.5, 2.2 Hz, 1H), 2.64-2.61 (m, 2H), 1.95-1.92
(m, 1H), 1.87-1.82 (m, 1H); 13C NMR (150 MHz, D
O) δ 78.0,
72.2, 69.8, 69.7, 68.6, 61.9, 51.0, 34.4; HRMS(MALDI-FTMS)
m/z 190.1073 (M + H, 190.1074 calcd for C N).
2
-
2
1
2 3
Cl /EtOAc): H NMR (600 MHz, CDCl ) δ 8.33 (s, 1H), 5.89
(
5
5
d, J ) 9.5 Hz, 1H), 5.66 (ddd, J ) 17.2, 10.3, 7.3 Hz, 1H),
8 16 4
H O
.26 (dd, J ) 10.6, 4.4 Hz, 1H), 5.13 (d, J ) 10.6 Hz), 5.10-
.07 (m, 2H), 4.68 (ddd, J ) 4.7, 2.2, 1.5 Hz, 1H), 4.34-4.31
P yr a n ose (9). Following the general procedure for the
preparation of 13, triol 7 (690 mg, 3.65 mmol) was submitted
to periodate cleavage and aldol condensation to afford 225 mg
of 9 (25%) after silica gel chromatography (CH Cl /MeOH, 7:1).
(m, 1H), 4.24 (d, J ) 11.7 Hz, 1H), 3.89 (d, J ) 11.7 Hz, 1H),
2
3
1
2
3
.34-2.31 (m, 1H), 2.21-2.17 (m, 1H), 2.11 (s, 3H), 2.09 (s,
2
2
H), 1.98 (s, 3H); 13C NMR (150 MHz, CDCl
70.1, 161.5, 132.3, 118.7, 95.8, 69.7, 68.9, 67.8, 66.4, 48.4, 34.8,
) δ 170.8, 170.3,
3
2 2
Silica gel TLC Rf 0.6 (CH Cl /MeOH, 7:1). Existing as a
1
mixture of pyranose and ketone (7:1). Data for pyranose 9: H
+
0.8, 20.7; HRMS (MALDI-FTMS) m/z 396.1267 (M + Na ,
NMR (600 MHz, D O) δ 8.05 (s, 1H), 5.75-5.73 (m, 1H), 5.07-
2
96.1265 calcd for C16
H
23
O
9
NNa).
5.02 (m, 2H), 4.41-4.38 (ddd, J ) 9.0, 7.2, 2.2 Hz, 1H), 3.94
(br s, 1H), 3.86 (app t, 1H), 3.69 (br d, 1H), 3.65 (d, J ) 11.6
Data for second triacetate: 1H NMR (600 MHz, CDCl
3
) δ
1
3
Hz, 1H), 3.41 (d, J ) 11.6, 1H), 2.24-2.16 (m, 2H); C NMR
8
.18 (s, 1H), 5.82-5.77 (m, 1H), 5.74 (d, J ) 9.5 Hz, 1H), 5.34
(
6
2
2
150 MHz, D O) δ 169.9, 132.8, 116.9, 97.2, 68.9, 66.0, 64.8,
(
(
app t, 1H), 5.11-5.05 (m, 3H), 4.21 (d, J ) 11.7 Hz, 1H), 4.16
app q, J ) 10.3 Hz, 1H), 3.97 (d, J ) 11.7 Hz, 1H), 3.89 (ddd,
+
3.6, 47.5, 33.8; HRMS (FAB) m/z 270.0956 (M + Na ,
70.0954 calcd for C
P en ta a ceta te (10): H NMR (600 MHz, CDCl
9
H
19
O
5
NNa).
J ) 10.3, 8.1, 2.2 Hz, 1H), 3.73 (br s, 1H), 2.39-2.36 (m, 1H),
.29-2.26 (m, 1H), 2.10 (s, 3H), 2.06 (s, 3H), 2.02 (s, 3H); 13
NMR (150 MHz, CDCl
C
1
2
3
) δ 8.53 (s,
3
) δ 171.4, 171.3, 169.6, 160.9, 133.4,
1H), 5.75-5.70 (m, 1H), 5.66 (d, J ) 8.8 Hz, 1H), 5.56 (app t,
J ) 8.4 Hz, 1H), 5.04 (dd, J ) 16.4 Hz, 1H), 5.01 (d, J ) 9.9
Hz, 1H), 4.97 (ddd, J ) 6.2, 4.4, 1.8 Hz, 1H), 4.90 (dd, J ) 8.4,
1.1 Hz, 1H), 4.82 (d, J ) 11.7 Hz, 1H), 4.51 (d, J ) 11.7 Hz,
1
2
17.7, 95.2, 70.96, 70.93, 70.8, 66.5, 51.9, 35.4, 20.7, 20.69,
+
0.62; HRMS (MALDI-FTMS) m/z 396.1267 (M + Na , 396.1265
calcd. for C16
23 9
H O NNa).
1
3
1
7
H), 2.16-1.13 (m, 2H), 2.114 (s, 3H), 2.110 (s, 3H), 2.10 (s,
3
-Ep ia u str a lin e (14). A 1:1 mixture of anomeric isomers
3 (40 mg, 0.16 mmol) and MeOH (3 mL) was cooled to -78
C, and a gentle stream of O was bubbled into the solution
H), 2.09 (s, 3H), 2.04 (s, 3H); 13C NMR (150 MHz, CD
OD) δ
1
°
3
70.3, 169.7, 169.4, 169.1, 168.5, 161.9, 133.2, 118.3, 89.9, 74.9,
3
3.2, 71.1, 66.4, 56.5, 37.0, 21.4, 21.4, 20.7, 20.5; HRMS (FAB)
until a faint blue color appeared. The ozonide was quenched
with palladium on carbon (10%) (5 mg) under an atmosphere
+
m/z 480.1478 (M + Na , 480.1476 calcd for C20
27
H O11NNa).
of H
2
(balloon) at -78 °C, and the reaction was allowed to
7-Ep ia lexin e (11). Following the general procedure for the
preparation of 3-epiaustraline (14), 9 (30 mg, 0.12 mmol) was
submitted to ozonolysis followed by reductive amination to
give, after reverse phase silica gel chromatography (100%
warm to room temperature. The solution was concentrated,
and the colorless oil was used in the next step without
purification.

8
268 J . Org. Chem., Vol. 65, No. 24, 2000
O), 15 mg of 7-epialexine (11) (66%) as a 8:1 mixture of
Romero and Wong
H
2
Ack n ow led gm en t. This research was supported by
2
5
18
25
stereoisomers: [R]
D
-7.2° (c 0.45, MeOH) (lit. [R]
D
-10.6°
the NIH (GM 4415).
1
(c 0.56, H
2
O)); H NMR (600 MHz, D O) δ 4.52-4.50 (m, 1H),
2
4
3
1
.25 (t, J ) 8.3 Hz, 1H), 3.94-3.92 (m, 2H), 3.91-3.89 (m, 1H),
Su p p or tin g In for m a tion Ava ila ble: Spectral data for the
triacetate of 13. COSY data for 10 and NMR data for
compounds 7, 10, 11, 13, 14, and 1. This material is available
free of charge via the Internet at http://pubs.acs.org.
.64-3.62 (m, 1H), 3.30-3.26 (m, 1H), 3.12-3.06 (m, 2H),
.95-1.91 (m, 1H), 1.89-1.85 (m, 1H); 13C NMR (150 MHz,
D O) δ 76.2, 73.9, 70.9, 65.6, 62.8, 57.9, 45.3, 32.8; HRMS
2
(
C
MALDI-FTMS) m/z 190.1078 (M + H, 190.1074 calcd for
8
H
16
O
4
N).
J O000933D