First synthesis of biopterin α-D-glucoside

Article abstract of DOI:10.3987/COM-08-S(F)60

A novel glycosyl donor, 4,6-di-O-acetyl-2,3-di-O-(4-methoxybenzyl)-α- D-glucopyranosy bromide (15) was efficiently prepared from D-glucose in 8 steps. The first synthesis of 2'-O-(α-D-glucopyranosyl)biopterin (2) was achieved by treatment of the key precursor, N²-(N,N- dimethylaminomethylene)-1'-O-(4-methoxybenzyl)-3-[2-(4-nitrophenyl)ethyl] biopterin (6) with 15 in the presence of silver triflate and tetramethylurea, followed by removal of the protecting groups.

Full text of DOI:10.3987/COM-08-S(F)60

HETEROCYCLES, Vol. 77, No. 2, 2009
747
HETEROCYCLES, Vol. 77, No. 2, 2009, pp. 747 - 753. © The Japan Institute of Heterocyclic Chemistry
Received, 25th July, 2008, Accepted, 5th September, 2008, Published online, 8th September, 2008
DOI: 10.3987/COM-08-S(F)60
FIRST SYNTHESIS OF BIOPTERIN α-D-GLUCOSIDE
Tadashi Hanaya,* Hiroki Baba, and Hiroshi Yamamoto†
Department of Chemistry, Faculty of Science, Okayama University, Tsushima-
naka, Okayama 700-8530, Japan. E-mail: hanaya@cc.okayama-u.ac.jp
†
School of Pharmacy, Shujitsu University, Nishigawara, Okayama 703-8516,
Japan
Abstract –A novel glycosyl donor, 4,6-di-O-acetyl-2,3-di-O-(4-methoxy-
benzyl)-α-D-glucopyranosy bromide (15) was efficiently prepared from
D-glucose in 8 steps. The first synthesis of 2’-O-(α-D-glucopyranosyl)biopterin
(2) was achieved by treatment of the key precursor, N2-(N,N-dimethylamino-
methylene)-1’-O-(4-methoxybenzyl)-3-[2-(4-nitrophenyl)ethyl]biopterin (6) with
15 in the presence of silver triflate and tetramethylurea, followed by removal of
the protecting groups.
Some pterins having a hydroxyalkyl side-chain at C–6, a representative example being biopterin (1), have
been found as glycosidic forms in certain prokaryotes; for example, 2’-O-(α-D-glucopyranosyl)biopterin
(2)1-4 isolated from cyanobacteria and limipterin [2’-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-
biopterin] (3)5 isolated from a green sulfur photosynthetic bacterium. Glycosides of other pterins such
as ciliapterin (L-threo-biopterin),6 neopterin,7 and 6-hydroxymethylpterin8 were also isolated from
cyanobacteria, anaerobic photosynthetic bacteria, and chemoautotrophic archaebacteria. Although
biopterin α-D-glucoside (2) is the most noteworthy among these pterin glycosides because of its abundant
occurrence in various kinds of cyanobacteria, Anacystis nidulans,1 Oscillatoria sp.,2 Synechococcus sp.,3
and Spirulina platensis,4 there has been no report for synthesis of 2 since its first discovery in 1958.
O
N
OH
O
N
OH
O
N
OH
3
6
2'
N
N
N
N
N
3'
HN
HN
H2N
HN
H2N
1'
OH
O
O
7
H2N
N
2
OH
HO
HO
1 (biopterin)
OH
O
OH
O
HO
OH
NHAc
2
3
This paper is dedicated to Professor Emeritus Keiichiro Fukumoto on the occasion of his 75th birthday.

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HETEROCYCLES, Vol. 77, No. 2, 2009
The physiological function of the parent pterins has been studied in detail: e.g., 1 exhibits enzyme
cofactor activity in aromatic amino acid hydroxylation9 and nitric oxide synthesis10 as the form of its
tetrahydro derivative. By contrast, the functional roles of pterin glycosides have remained obscure,
although some inhibitory activities against tyrosinase11 and photostabilization of photosynthetic
pigments12 were reported for 2. Despite a considerable interest from the viewpoint of their biological
activities and functions as well as structural proof of hitherto reported natural products, attempts at
preparation of pterin glycosides have scarcely been made so far, except for our synthetic studies on
limipterin (3) and ciliapterin glycosides.13-15 In a previous paper,14 we developed an efficient synthetic
protocol for the pterin 2’-O-glycosides by way of the key intermediate, N2-(N,N-dimethylamino-
methylene)-1’-O-(4-methoxybenzyl)-3-[2-(4-nitrophenyl)ethyl]biopterin (6) derived from L-rhamnose (or
D-xylose) via 4 and 5 (Scheme 1). Glycosylation of 6 with tetra-O-benzoyl-α-D-glucopyranosyl
bromide (7) in the presence of silver triflate afforded the 2’-O-(β-D-glucopyranosyl)biopterin derivative
(8). In the present study, we therefore have undertaken to prepare an efficient glycosyl donor leading
the preponderant production of pterin α-glycosides. We now describe the first synthesis of the
representative, natural pterin glycoside, 2’-O-(α-D-glucopyranosyl)biopterin (2).
O
NH2
HN
O
N
OPMB
OH
O
OPMB
H2N
N
NH2
N
N
L-rhamnose
D-xylose
OH
HN
H2N
H3C
NaHCO3
MeOH-H2O
O
4
5
O
OPMB
BzO
O
NPE
N
N
N
1) Me2NCH(OMe)2
DMF
2) Ac2O, Py
O
N
OPMB
OH
OBz
N
BzO
Br
NPE
N
N
N
O
7
N
N
N
BzO
BzO
3) NPEOH, PPh3
DEAD, THF
N
CF3SO3Ag
O
OBz
TMU, CH2Cl2
4) NaOMe, MeOH
75%
6
BzO
OBz
NPE: p-nitrophenylethyl
PMB: p-methoxybenzyl
8
Scheme 1
The stereoselective formation of the β-glycoside (8) from 6 was mainly caused by participation of the
2-O-benzoyl group of the glycosyl donor (7) through the formation of an acyloxonium ion intermediate.16
In order to avoid such a neighboring group participation, we sought to introduce an ether substituent for
protection of 2-OH of a glycosyl donor. Taking into consideration the available combination of
protecting groups employed for the synthetic pathway, p-methoxybenzyl (PMB) and acetyl groups were
respectively chosen for protection of 2,3-OH and 4,6-OH of the glycosyl moiety.
Penta-O-acetyl-β-D-glucopyranose (9),17 derived from D-glucose, served as the starting material for
preparation of methyl 4,6-di-O-acetyl-2,3-di-O-PMB-1-thio-β-D-glucopyranose (14) and its

HETEROCYCLES, Vol. 77, No. 2, 2009
749
α-D-glucopyranosyl bromide derivative (15), the potential glycosyl donors for the pterin α-glycosides
(Scheme 2). Treatment of 9 with thiourea and boron trifluoride etherate, followed by the action of
methyl iodide and triethylamine, afforded the methyl 1-thio-β-D-glucopyranose derivative (10).18
Methanolysis of 10 in the presence of sodium methoxide and the subsequent acetalization with
2,2-dimethoxypropane in the presence of p-toluenesulfonic acid provided the 4,6-O-isopropylidene
derivative (11). 19
Treatment of 11 with p-methoxybenzyl chloride and sodium hydride in DMF gave
the 2,3-di-O-PMB derivative (12). Hydrolysis of 12 in 70% acetic acid afforded methyl 2,3-di-O-PMB-
1-thio-β-D-glucopyranoside (13), which was then treated with acetic anhydride and pyridine to give the
desired 4,6-di-O-acetyl derivative (14).20 The transformation of the thioglycoside (14) to the
D-glucopyranosyl bromide (15)21 was achieved by reaction with bromine in dichloromethane in the
presence of 2,6-lutidine.
SC(NH2)2, BF3-OEt2
MeCN, reflux, 20 min,
AcO
AcO
AcO
AcO
O
1) NaOMe, MeOH
rt, 2 h
OAc
SMe
SMe
O
PMBCl
O
OAc
O
OAc
OH
2) Me2C(OMe)2
TsOH, DMF
rt, 12 h
then MeI, TEA, rt, 2 h
89%
NaH, Bu4NI
DMF, rt, 10 h
75%
O
OAc
OAc
10
OH
9
11
90% (2 steps)
O
HO
AcO
AcO
Br2
2,6-lutidine
SMe
SMe
SMe
Ac2O, Py
O
OPMB
O
OPMB
70% AcOH aq.
O
OPMB
O
OPMB
CH2Cl2, rt, 2 h
90%
MeOH, rt, 10 h
99%
O
HO
rt, 12 h
98%
AcO
AcO
Br
OPMB
12
OPMB
13
OPMB
14
PMBO
15
Scheme 2
Glycosylation of the 1’-O-PMB-biopterin derivative (6) with glycosyl donors (14, 15) was examined
under various conditions in the presence of activators (Scheme 3). Treatment of 6 with the thioglycoside
(14) in dichloromethane at room temperature in the presence of methyl triflate22 or
N-iodosuccinimide-silver triflate23 as activators resulted in the formation of unidentified, decomposed
compounds instead of the desired glycoside.24 While glycosylation of 6 with 4.0 mol equiv. of the
glycosyl bromide (15) in dichloromethane in the presence of tetrabutylammonium bromide and
N-ethyldiisopropylamine25 did not proceed, the same reaction in the presence of silver triflate (2.0 mol
equiv.) and tetramethylurea (TMU)26 (1.0 mol equiv.) afforded an inseparable anomeric mixture (85:15)
of the 2’-O-(α-D-glucopyranosyl)biopterin derivative (16a) and its β-anomer (16b) in 56% yield, along
with the recovery of 6 (38%). The α-anomeric structure of 16a was derived from its J1,2 value (3.5 Hz)
of 1H-NMR, while the larger J1,2 value (8.0 Hz) confirmed the β-form of 16a.
Separation of these isomers was achieved by removal of PMB groups and the subsequent acetylation.
Thus, the mixture of 16a,b was treated with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) in
dichloromethane, followed by acetylation with acetic anhydride in pyridine, afforded the

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HETEROCYCLES, Vol. 77, No. 2, 2009
2’-O-(2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl)biopterin derivative (17a) in 43% (total yield from 6)
and its β-anomer (17b) in 8%.27
O
N
OPMB
O
O
N
OPMB
NPE
N
NPE
N
N
N
N
N
N
O
N
OPMB
OH
O
15
N
N
N
NPE
N
N
N
+
N
AcO
OPMB
OAc
CF3SO3Ag, TMU
CH2Cl2, rt, 3 h
O
OPMB
N
PMBO
O
56%
AcO
6
(38% recovery of 6)
OAc
(85:15)
OPMB
16a
16b
1) DDQ, CH2Cl2, H2O, rt, 2 h
2) Ac2O, Py, rt, 12 h
91%
O
OAc
O
O
N
OAc
O
OH
NPE
N
NPE
N
N
N
N
N
N
N
HN
H2N
1) NH3 aq., MeOH
rt, 12 h
O
O
N
N
N
N
N
N
+
2) DBU, DMF
rt, 12 h
OAc
AcO
OH
HO
OAc
OAc
O
OAc
OH
AcO
O
90%
AcO
O
OAc
OH
2
17a (43% from 6)
17b (8%)
Scheme 3
Removal of the protecting groups of 17a was carried out according to the following steps: treatment with
aqueous ammonia (to cleave the N,N-dimethylaminomethylene and acetyl groups) and then with DBU28
(to cleave NPE group) furnished the desired 2’-O-(α-D-glucopyranosyl)biopterin (2) in 90% overall yield.
The precise parameters obtained on 1H- and 13C-NMR spectra for 2 are listed in Table 1; the spectral data
of the synthetic compound (2) were found to be essentially identical with those reported for natural
product.4,11
The present work thus demonstrates the first synthesis of biopterin α-D-glucoside (2) by use of the key
intermediate 1’-O-PMB-biopterin derivative (6) and the novel glycosyl donor (15) to preferentially
provide an α-glycoside. Extension of this work including improvement of selectivity and yield for
glycosylation as well as applications of these findings in synthesizing other natural pterin α-glycosides is
in progress.
ACKNOWLEDGEMENTS
We are grateful to the SC-NMR Laboratory of Okayama University for the NMR measurements and to
Okayama Foundation for Science and Technology (to T. H.) which partially supported this work.

HETEROCYCLES, Vol. 77, No. 2, 2009
751
1
13
Table 1. 600 MHz H- and 151 MHz C-NMR Spectral Parameters [chemical shifts (δ) and coupling
constants (Hz)] for biopterin α-D-glucoside (2) in D2O
——————————————————————————————————————————————
Pterin moiety
Glycosyl moiety
———————————————
—————————————————————————
Com-
pound
H-7
H-1’
H-2’
H3-3’
H-1
H-2
H-3
H-4
H-5
H2-6
(J1’,2’
)
(J2’,3’
)
(J1,2)
(J2,3)
(J3,4)
(J4,5)
(J5,6)
——————————————————————————————————————————————
Synthetic a 8.79 4.83
4.07
(6.1)
4.06
(5.9)
1.30
5.00
(3.9)
4.99
(3.3)
3.41
3.32
3.22
2.40
3.43
(7.1)
(10.0) (9.0)
(10.0) (3.6)
Natural b
8.79 4.82
(7.3)
1.29
3.40
(9.2)
3.30
(9.2)
3.21
(9.2)
2.38
(3.4)
c
——————————————————————————————————————————————
Pterin moiety
Glycosyl moiety
———————————————————————————
——————————————
C-2
C-4
C-4a C-6
C-7
C-8a C-1’ C-2’ C-3’ C-1 C-2 C-3 C-4 C-5 C-6
——————————————————————————————————————————————
Synthetic d 155.7 165.4 128.3 152.2 149.6 155.2 75.6 75.5 14.9 95.8 71.6 73.4 69.5 72.5 60.5
Natural e 155.7 165.4 128.6 152.8 150.1 155.4 76.0 76.0 15.2 96.3 72.1 73.9 70.1 72.9 61.2
——————————————————————————————————————————————
a The solvent peak (δ 4.79) was used as an internal standard.
b Ref. 4 (at 270 MHz). The internal or external standard is not shown.
c Not reported.
d 1,4-Dioxane (δ 67.2) was used as an internal standard.
e Ref. 11 (at 100 MHz). The internal or external standard is not shown.
REFERENCES AND NOTES
1. H. S. Forrest, C. Van Baalen, and J. Myers, Arch. Biochem. Biophys., 1958, 78, 95.
2. T. Matsunaga, J. G. Burgess, N. Yamada, K. Komatsu, S. Yoshida, and Y. Wachi, Appl. Microbiol.
Biotechnol., 1993, 39, 250.
3. Y. K. Choi, Y. K. Hwang, Y. H. Kang, and Y. S. Park, Pteridines, 2001, 12,121.
4. Y. Noguchi, A. Ishii, A. Matsushima, D. Haishi, K. Yasumuro, T. Moriguchi, T. Wada, Y. Kodera,
M. Hiroto, H. Nishihara, M. Sekine, and Y. Inada, Mar. Biotechnol., 1999, 1, 207.
5. K. W. Cha, W. Pfleiderer, and J. J. Yim, Helv. Chim. Acta, 1995, 78, 600.
6. (a) M. Ikawa, J. J. Sasner, J. F. Haney, and T. L. Foxall, Phytochemistry, 1995, 38, 1229. (b) S.-H.
Cho, J.-U. Na, H. Youn, C.-S. Hwang, C.-H. Lee, and S.-O. Kang, Biochim. Biophys. Acta, 1998,
1379, 53.
7. (a) A. Suzuki and M. Goto, J. Biochem., 1968, 63, 798. (b) X. Lin and R. H. White, J. Bacteriol.,

752
HETEROCYCLES, Vol. 77, No. 2, 2009
1988, 170, 1396.
8. H. W. Lee, C. H. Oh, A. Geyer, W. Pfleiderer, and Y. S. Park, Biochim. Biophys. Acta, 1999, 1410,
61.
9. (a) S. Kaufman and E. E. Kaufman, ‘Folates and Pterins,’ ed. by R. Blakley and S. J. Benkovic, J.
Wiley & Sons, New York, 1985, Vol. 2, pp. 251–352. (b) S. Kaufman and D. B. Fisher, ‘Molecular
Mechanisms of Oxygen Activation,’ ed. by O. Hayaishi, Academic Press, New York, 1974, pp.
285–369. (c) G. R. Guroff, J. Biol. Chem., 1969, 244, 142.
10. (a) N. S. Kwon, C. F. Nathan, D. J. Stuehr, J. Biol. Chem., 1989, 264, 20496. (b) M. A. Marletta,
Cell, 1994, 78, 927.
11. Y. Wachi, S. Yoshida, K. Komatsu, and T. Matsunaga, Jpn. Patent, 05,286,989, 1993 (Chem. Abstr.,
1994, 120, 161782t).
12. T. Saito, H. Ishikawa, Y. Hada, K. Fukui, Y. Kodera, A. Matsushima, and Y. Inada, Dyes Pigments,
2003, 56, 203.
13. (a) T. Hanaya, K. Soranaka, K. Harada, H. Yamaguchi, R. Suzuki, Y. Endo, H. Yamamoto, and W.
Pfleiderer, Heterocycles, 2006, 67, 299. (b) T. Hanaya, H. Baba, H. Toyota, and H. Yamamoto,
Tetrahedron, 2008, 64, 2090.
14. T. Hanaya, H. Toyota, and H. Yamamoto, Synlett, 2006, 2075.
15. (a) T. Hanaya, H. Baba, and H. Yamamoto, Carbohydr. Res., 2007, 342, 2159. (b) T. Hanaya, H.
Baba, M. Kanemoto, and H. Yamamoto, Heterocycles, 2008, 76, 635.
16. G. Wulff and G. Röhle, Angew. Chem., Int. Ed. Engl., 1974, 13, 157, and references cited therin.
17. M. L. Wolfrom and A. Thompson, Methods Carbohydr. Chem., 1963, 2, 211.
18. F. M. Ibatullin, K. A. Shabalin, J. V. Jänis, and A. G. Shavva, Tetrahedron Lett., 2003, 44, 7961; use
of larger amounts of methyl iodide (10 mol equiv.) and TEA (12 mol equiv.) than those reported in
this lit. led to a much improved yield (89%) of 10 from 9 (lit., 57% by use of 1.1 equiv. of methyl
iodide and 3.1 equiv. of TEA).
1
19. All newly isolated compounds hereafter in this paper gave satisfactory analytical and H-NMR
(mostly at 600 MHz) data, which will be presented in a full paper in the near future.
1
20. H-NMR for 14 (500 MHz, CDCl3) δ = 1.94, 2.06 (3H each, 2s, AcO-4,6), 2.22 (3H, s, MeS-1), 3.48
(1H, dd, J1,2 = 9.8, J2,3 = 8.8 Hz, H-2), 3.54 (1H, ddd, J4,5 = 10.1, J5,6a = 5.2, J5,6b = 2.4 Hz, H-5), 3.61
(1H, t, J3,4 = 9.5 Hz, H-3), 3.80 (6H, s, MeO), 4.08 (1H, dd, J6a,6b = 12.2 Hz, Hb-6), 4.20 (1H, dd,
2
Ha-6), 4.35 (1H, d, H-1), 4.59, 4.77 (2H each, 2d, J = 11.0 Hz, CH2O-2 or 3), 4.67, 4.81 (2H each,
2d, 2J = 10.1 Hz, CH2O-2 or 3), 5.02 (1H, t, H-4), 6.855, 6.86, 7.18, 7.30 (2H each, 4 d, Jo,m = 8.8 Hz,
C6H4).
1
21. H-NMR for 15 (500 MHz, CDCl3) δ = 1.95, 2.05 (3H each, 2s, AcO-4,6), 3.53 (1H, dd, J2,3 = 9.2,
J1,2 = 3.9 Hz, H-2), 3.80, 3.81 (3H each, 2s, MeO), 3.92 (1H, t, J3,4 = 9.5 Hz, H-3), 4.02 (1H, dd,
J6a,6b = 12.5, J5,6b = 2.1 Hz, Hb-6), 4.14 (1H, ddd, J4,5 = 10.4, J5,6a = 4.6, Hz, H-5), 4.26 (1H, dd, Ha-6),
4.58, 4.80 (2H each, 2d, 2J = 11.3 Hz, CH2O-2 or 3), 4.62, 4.67 (2H each, 2d, 2J = 11.6 Hz, CH2O-2
or 3), 5.05 (1H, dd, H-4), 6.26 (1H, d, H-1), 6.86, 6.88, 7.19, 7.29 (2H each, 4 d, Jo,m = 8.6 Hz, C6H4).
22. H. Lönn, Carbohydr. Res., 1985, 139, 105.

HETEROCYCLES, Vol. 77, No. 2, 2009
753
23. H. Yin, F. W. D’Souza, and T. L. Lowary, J. Org. Chem., 2002, 67, 892.
24. In model experiments using thioglycoside (14) with 2-propanol in CH2Cl2, the corresponding
glycosides were obtained in good yields: 95% (α:β = 60:40) by use of MeOTf; 91% (56:44) by use of
NIS-AgOTf.
25. R. U. Lemieux, K. B. Hendriks, R. V. Stick, and K. James, J. Am. Chem. Soc., 1975, 97, 4056.
26. S. Hanessian and J. Banoub, Methods Carbohydr. Chem., 1980, 8, 247.
1
27. H-NMR for 17a (600 MHz, CDCl3) δ 1.18 (3H, d, J2’, 3’ = 6.4 Hz, H3-3’), 1.98, 1.99, 1.995, 2.08,
3
2.19 (3H each, 5s, AcO-1’, AcO-2,3,4,6*), 3.16 [2H, t, J = 7.8 Hz, CH2CH2-N(3)], 3.19, 3.23 (3H
each, 2s, Me2N), 3.83 (1H, ddd, J4,5 = 10.3, J5,6a = 4.2, J5,6b = 2.2 Hz, H-5*), 3.94 (1H, dd, J6a,6b
=
12.5 Hz, Hb-6*), 4.19 (1H, dd, Ha-6*), 4.45 (1H, qd, J1’,2’ = 4.6 Hz, H-2’), 4.58, 4.60 [1H each, 2dt, 2J
= 12.0 Hz, CH2-N(3)], 4.80 (1H, dd, J2,3 = 10.3, J1,2 = 3.9 Hz, H-2*), 5.00 (1H, dd, J3,4 = 9.5 Hz,
H-4*), 5.22 (1H, d, H-1*), 5.33 (1H, dd, H-3*), 6.06 (1H, d, H-1’), 7.42, 8.14 (2H each, 2d, Jo,m = 8.8
*
Hz, C6H4), 8.86 (1H, s, CH=N-2), 8.90 (1H, s, H-7), for glycosyl moiety. 1H-NMR for 17b (600
MHz, CDCl3) δ 1.31 (3H, d, J2’, 3’ = 6.4 Hz, H3-3’), 1.88, 1.95, 2.01, 2.09, 2.13 (3H each, 5s, AcO-1’,
3
AcO-2,3,4,6*), 3.18 [2H, t, J = 7.6 Hz, CH2CH2-N(3)], 3.19, 3.235 (3H each, 2s, Me2N), 3.70 (1H,
ddd, J4,5 = 10.0, J5,6a = 5.1, J5,6b = 2.4 Hz, H-5*), 4.13 (1H, dd, J6a,6b = 12.2 Hz, Hb-6*), 4.23 (1H, dd,
2
Ha-6*), 4.54 (1H, qd, J1’,2’ = 5.9 Hz, H-2’), 4.61, 4.63 [1H each, 2dt, J = 12.4 Hz, CH2-N(3)], 4.66
(1H, d, J1,2 = 8.1 Hz, H-1*), 4.87 (1H, dd, J2,3 = 9.5 Hz, H-2*), 5.01 (1H, dd, J3,4 = 9.5 Hz, H-4*), 5.06
(1H, dd, H-3*), 5.94 (1H, d, H-1’), 7.43, 8.15 (2H each, 2d, Jo,m = 8.5 Hz, C6H4), 8.75 (1H, s, H-7),
8.84 (1H, s, CH=N-2), *for glycosyl moiety.
28. (a) T. Hanaya, K. Torigoe, K. Soranaka, H. Yamamoto, Q. Yao, and W. Pfleiderer, Pteridines, 1995,
6, 1. (b) F. Himmelsbach, B. S. Schulz, T. Trichtinger, R. Charubala, and W. Pfleiderer,
Tetrahedron, 1984, 40, 59.