Easy access to water-soluble fullerene derivatives via 1,3-dipolar cycloadditions of azomethine ylides to C60

Full text of DOI:10.1021/jo961522t

9070
J . Org. Chem. 1996, 61, 9070-9072
Sch em e 1
Ea sy Access to Wa ter -Solu ble F u ller en e
Der iva tives via 1,3-Dip ola r Cycloa d d ition s
of Azom eth in e Ylid es to C60
Tatiana Da Ros and Maurizio Prato*
Dipartimento di Scienze Farmaceutiche, Universita` di
Trieste, Piazzale Europa 1, 34127 Trieste, Italy
Fabiola Novello and Michele Maggini
Centro Meccanismi di Reazioni Organiche del CNR,
Dipartimento di Chimica Organica, Universita` di Padova,
Via Marzolo 1, 35131 Padova, Italy
Elena Banfi
Dipartimento di Scienze Biomediche, Via A. Fleming 22,
34127 Trieste, Italy
Sch em e 2
Received August 6, 1996
One of the most promising fields of applications of the
fullerenes is based on their biological properties. Very
exciting findings, pioneered by Wudl,1 Nakamura,2 and
co-workers, have stimulated much interest and generated
further successful investigations.3-7 From a practical
point of view, a fundamental issue that must be ad-
dressed when dealing with fullerenes is the solubilization
of the candidate molecule or ion in the polar media
necessary for biological tests.7 Fullerenes, in fact, are
not soluble in water or in polar organic solvents. They
must be chemically modified, and a solubilizing append-
age must be covalently attached.8,9 In this paper we
report a general method for the solubilization of fullerenes
and a preliminary evaluation of their biological properties
against some microorganisms.
nicotine in order to check the influence of the new form
of carbon on the toxicity of nicotine.
With a suitable functionalization method in hand,10 we
were attracted by the idea that a very interesting
molecule, which we may call fulleronicotine for its
resemblance to natural nicotine, could be obtained in a
single step from commercially available reactants (Scheme
1).
Fulleronicotine (1) is a fullerene homologue of natural
nicotine, the well-known alkaloid abundantly found in
tobacco leaves. In fulleronicotine, the 3,4 bond of the
pyrrolidine ring is fused to a 6,6 ring junction of C60. As
nicotine has a range of biological activities and is used
as a pharmacological tool in a number of applications,11
it seemed interesting to prepare a fullerene-substituted
However, when we turned to the biological evaluation
of 1, we had to cope with its very low solubility in water-
miscible solvents. Attempts to solubilize 1 in water by
known methodologies12 failed to give reproducible results.
To obtain a soluble form of fulleronicotine, it was
necessary to start with an N-functionalized glycine. A
triethylene glycol chain was chosen as the solubilizing
appendage owing to its ready availability and high
solubilizing power.
N-Substituted glycine 2 was obtained by condensation
of glycine benzyl ester with aldehyde 3c under reductive
conditions (NaBH3CN), followed by hydrogenolysis of the
benzyl ester. Glycine 2, a very useful reactant for
preparing water-soluble derivatives, readily condensed
with a number of aldehydes and C60 to give the novel C60-
fused functionalized pyrrolidines 4a -c (Scheme 2, yields
34-39%, see Experimental Section).
All compounds 4a -c exhibit moderate solubility in a
9:1 ratio of water-DMSO. It was found that the best
procedure for preparing the aqueous solutions was to
dissolve the compounds in DMSO and to dilute 1:9 with
pure water. In this binary solvent an upper limit
concentration was obtained for 4c (ca. 3 × 10-5 M),
whereas the solubility for 4a ,b was slightly lower (ca.
1.5 × 10-5 M). A UV-vis absorption spectrum of 4a in
H2O-DMSO (9:1, 1 × 10-5 M) is reported in Figure 1. It
shows that the main absorptions of the fullerene core are
still evident, although, in analogy to other fullerene
(1) Sijbesma, R.; Srdanov, G.; Wudl, F.; Castoro, J . A.; Wilkins, C.;
Friedman, S. H.; DeCamp, D. L.; Kenyon, G. L. J . Am. Chem. Soc.
1993, 115, 6510.
(2) Tokuyama, H.; Yamago, S.; Nakamura, E.; Shiraki, T.; Sugiura,
Y. J . Am. Chem. Soc. 1993, 115, 7918.
(3) Boutorine, A.; Tokuyama, H.; Takasugi, M.; Isobe, H.; Nakamura,
E.; He´le`ne, C. Angew. Chem., Int. Ed. Engl. 1994, 33, 2462.
(4) Toniolo, C.; Bianco, A.; Maggini, M.; Scorrano, G.; Prato, M.;
Marastoni, M.; Tomatis, R.; Spisani, S.; Palu`, G.; Blair, E. D. J . Med.
Chem. 1994, 37, 4558.
(5) An, Y.-Z.; Chen, C.-H. B.; Anderson, J . L.; Sigman, D. S.; Foote,
C. S.; Rubin, Y. Tetrahedron 1996, 52, 5179.
(6) Nakamura, E.; Tokuyama, H.; Yamago, S.; Shiraki, T.; Sugiura,
Y. Bull. Chem. Soc. J pn. 1996, 69, 2143.
(7) For a recent review, see: J ensen, A. W.; Wilson, S. R.; Schuster,
D. I. Bioorg. Med. Chem. 1996, 4, 767.
(8) Hirsch, A. The Chemistry of the Fullerenes; Thieme: Stuttgart,
Germany, 1994.
(9) Diederich, F.; Thilgen, C. Science 1996, 271, 317.
(10) Maggini, M.; Scorrano, G.; Prato, M. J . Am. Chem. Soc. 1993,
115, 9798.
(11) The Pharmacological Basis of Therapeutics; Hardman, J . G.,
Goodman Gilman, A., Limbird, L. E., Eds.; McGraw-Hill: New York,
1996; 9th ed., p 192.
(12) Yamakoshi, Y. N.; Yagami, T.; Fukuhara, K.; Sueyoshi, S.;
Miyata, N. J . Chem. Soc., Chem. Commun. 1994, 517.
S0022-3263(96)01522-8 CCC: $12.00 © 1996 American Chemical Society

Notes
J . Org. Chem., Vol. 61, No. 25, 1996 9071
Syn th esis of N-(3,6,9-Tr ioxa d ecyl)glycin e (2). To a solu-
tion of 265 mg (1.63 mmol) of aldehyde 3c and 552 mg (1.63
mmol) of glycine benzyl ester 4-toluenesulfonate in 12 mL of
methanol-acetic acid (99:1) was added 127 mg (2.02 mmol) of
sodium cyanoborohydride in 30 min. The solution was stirred
at rt for 2.5 h and then was poured into 30 mL of saturated
NaHCO3 solution. The product was extracted with ethyl acetate,
the organic phase was washed with brine and dried with
anhydrous NaSO4, and the solvent was removed in vacuo. Since
the reaction mixture consisted of an unseparable mixture of
glycine benzyl ester, the desired N-(3,6,9-trioxadecyl)glycine (2)
(ethyl acetate-methanol (9:1), Rf ) 0.40), and the dialkylated
N,N-bis(3,6,9-trioxadecyl)glycine, the residue was dissolved in
dioxane and treated with 342 mg (2.00 mmol) of benzyl chloro-
formate. The N-Cbz-N-(3,6,9-trioxadecyl)glycine benzyl ester
was then purified by column chromatography (ethyl acetate-
petroleum ether (1:1), Rf ) 0.36). Yield: 355 mg (49% from
aldehyde 3c). 1H-NMR (200 MHz): δ 7.35 (s, 5H), 7.29 (m, 5H),
5.25 (s, 2H), 5.18 (m, 2H), 4.21 and 4.16 (two s, 2H), 3.55 (m, 12
H), 3.35 (s, 3H). 13C-NMR (50 MHz): δ 169.7, 156.2, 155.8,
136.3, 136.4, 128.5, 128.4, 128.3, 128.3, 128.1, 127.9, 127.8, 127.7,
71.8, 70.4, 70.3, 67.5, 67.3, 66.7, 66.6, 58.9, 50.2, 48.5, 47.9. EI-
MS: m/z 311 (M - COOCH2Ph)+, 267, 149, 91, 68 (100). IR
(film, cm-1): 2975, 2940, 1750, 1715, 1700. N-Cbz-N-(3,6,9-
trioxadecyl)glycine (750 mg) was dissolved in methanol (40 mL),
and 50 mg of Pd/C was added. The suspension was stirred at
rt under H2 atmosphere for 18 h and then was filtered on Celite,
and the solvent evaporated. Amino acid 2 was obtained in
quantitative yield and recrystallized from methanol-ethyl ether.
Mp: >250 °C. 1H-NMR (200 MHz, CDCl3): δ 3.78 (broad t, 2H),
3.51 (m, 12H), 3.30 (s, 3H), 3.18 (broad t, 2H). 13C-NMR (50
MHz, CDCl3): δ 170.5, 71.8, 70.4, 70.4, 70.3, 66.5, 58.9, 49.9,
46.7. IR (cm-1): 3350, 1650, 1111.
Gen er a l P r oced u r e for th e Syn th esis of F u ller en e
Der iva tives 1 a n d 4a -c. To a solution of 100 mg (0.14 mmol)
of C60 in 50 mL of toluene were added 0.70 mmol of aldehyde
and 0.14 mmol of N-substituted glycine. The mixture was
heated to reflux for 2 h, brought to rt, poured on top of a silica
gel column, eluted with toluene, and then eluted with toluene-
acetate (9:1). The compounds were dissolved in dichloromethane
and precipitated by addition of methanol.
Der iva tive 1: 39% (toluene-ethyl acetate (8:2), Rf ) 0.24).
1H-NMR (250 MHz): δ 9.00 (m, 1H), 8.60 (m, 1H), 8.16 (m, 1H),
7.38 (dd, J ) 6.0 Hz, J ) 7.5 Hz, 1H), 5.02 (d, J ) 9.5 Hz, 1H),
4.99 (s, 1H), 4.32 (d, J ) 9.4 Hz, 1H), 2.83 (s, 3H). 13C NMR
(62.5 MHz): δ 155.5, 153.2, 152.2, 151.8, 150.3, 149.6, 147.0,
146.0, 145.9, 145.8, 145.4, 145.2, 145.1, 145.0, 144.9, 144.3, 142.9,
142.8, 142.4, 142.3, 142.0, 141.7, 141.6, 140.0, 139.9, 137.0, 136.2,
136.1, 135.8, 123.3, 80.8, 69.8, 68.6, 39.7. UV-vis (cyclohex-
ane): λmax nm 430, 317, 256, 212. MALDI m/z: 855 (M + H)+,
720 (C60)+. Anal. Calcd for C68H10N2: C, 95.54; H, 1.18; N, 3.28.
Found: C, 93.71; H, 0.97; N, 3.19.
F igu r e 1. UV-vis absorption spectrum for a solution of 4a
in H2O-DMSO (9:1, 1 × 10-5 M).
derivatives dissolved in polar solvents, bands are broader
and less structured, especially in the low-energy region.13
In preliminary biological tests, 4c was found to be
active against a variety of microorganisms. Different
species of bacteria and different fungal strains were killed
in a slightly modified agar diffusion test: two strains,
clinical isolates CA1 and Z11, of Candida albicans, a
fastidious pathogenic eukariote; strain ATCC 6633 of
Bacillus subtilis, a spore-forming, Gram-positive bacte-
rium; strain AB1153 of Escherichia coli, a Gram-negative
enteric bacterium; a clinical isolate, strain 261/6 of
Mycobacterium avium, an acid fast, emerging pathogen
resistant to most antimicrobial drugs. In the latter case,
70% inhibition was observed with a concentration of 26
µg/mL, whereas complete inhibition was achieved with
concentrations 10 times higher.
The nature of the activity of the fullerene derivatives
4a -c is yet to be determined. However, the rather wide
range of biological activity exhibited makes these new
water-soluble fullerene derivatives interesting candidates
for further investigations. Work along these lines is
underway.
Exp er im en ta l Section
Ma ter ia ls a n d Meth od s. Details regarding instrumentation
used in this paper have been described elsewhere.14 All other
reagents were used as purchased from Fluka and Aldrich. All
solvents were distilled prior to use.
Der iva tive 4a : 37% (toluene-ethyl acetate (1:1), Rf ) 0.12).
1H-NMR (200 MHz): δ 9.01 (m, 1H), 8.58 (dd, J ) 4.8 Hz, J )
1.6 Hz, 1H), 8.19 (broad d, J ) 8.1 Hz, 1H), 7.37 (dd, J ) 8.1
Hz, J ) 4.8 Hz, 1H), 5.21 (d, J ) 9.8 Hz, 1H), 5.21 (s, 1H) 4.33
(d, J ) 9.8 Hz, 1H), 3.90-4.15 (m, 2H), 3.75 (m, 6H), 3.45 (m,
6H), 2.92 (m, 1H). 13C-NMR (50 MHz, CS2-CDCl3): δ 156.2,
153.8, 152.7, 152.3, 150.7, 150.0, 147.3, 146.3, 146.3, 16.2, 146.2,
146.2, 146.1, 146.1, 145.9, 145.7, 145.7, 145.6, 145.5, 145.5, 145.4,
145.4, 145.3, 145.3, 145.2, 144.7, 144.5, 144.4, 144.3, 143.1, 143.0,
142.7, 142.6, 142.5, 142.2, 142.1, 142.1, 142.0, 141.9, 141.8, 141.7,
141.6, 140.2, 140.1, 139.5, 137.3, 137.0, 136.3, 136.0, 135.6, 133.1,
123.6, 79.7, 76.0, 72.0, 70.8, 70.7, 70.1, 69.2, 67.6, 59.1, 52.1.
UV-vis (cyclohexane): λmax nm 701, 430, 323, 309, 255, 212.
MALDI m/z: 987 (M + H+). Anal. Calcd for C74H22N2O3: C,
90.05; H, 2.25; N, 2.84. Found: C, 88.85; H, 2.06; N, 2.76.
Der iva tive 4b: 38% (toluene-ethyl acetate (1:1), Rf ) 0.08).
1H-NMR (250 MHz): δ 8.68 (m, 2H), 7.81 (m, 2H), 5.23 (d, J )
7.8 Hz, 1H), 5.21 (s, 1H), 4.35 (d, J ) 7.8 Hz, 1H), 4.01 (2m,
2H), 3.78 (m, 6H), 3.57 (m, 2H), 3.38 (m, 4H), 2.99-2.90 (m, 1H).
13C-NMR (62.5 MHz): δ 155.9, 153.7, 152.4, 152.0, 149.5, 147.4,
147.3, 147.3, 146.3, 146.3, 146.2, 146.2, 146.2, 146.1, 146.1, 145.9,
145.9, 145.6, 145.6, 145.5, 145.4, 145.4, 145.3, 145.3, 145.3, 145.2,
145.2, 144.7, 144.5, 144.4, 144.3, 143.1, 143.0, 142.7, 142.6, 142.5,
142.2, 142.1, 142.1, 142.0, 141.9, 141.8, 141.7, 141.6, 141.5, 140.2,
139.9, 139.4, 137.1, 136.3, 136.1, 135.5, 124.5, 80.9, 75.5, 71.9,
70.7, 70.6, 70.0, 69.3, 67.4, 59.0, 52.1. UV-vis (cyclohexane):
Syn th esis of 3,6,9-Tr ioxa d eca n a ld eh yd e (3c). To a solu-
tion of oxalyl chloride (3 mL) in dichloromethane (75 mL) under
nitrogen and cooled in a dry ice-acetone bath were carefully
added 5 mL of dimethyl sulfoxide in 15 mL of CH2Cl2. The
solution was stirred for 10 min, and then a solution of 5 mL
(29.7 mmol) of triethylene glycol monomethyl ether in 30 mL of
CH2Cl2 was added dropwise. The mixture was stirred for 15
min, and then triethylamine (20 mL) was added dropwise over
a period of 20 min. The reaction mixture was left for 30 min at
-78 °C and then allowed to reach rt. After a standard workup
with a saturated aqueous NaCl solution, the crude was purified
by flash chromatography (eluant: ethyl acetate-methanol (9:
1), Rf ) 0.54) affording 2.7 g (56%) of pure aldehyde 3c. Bulb-
to-bulb distillation at 90 °C (0.01 Torr) gave a clear homogeneous
oil, which gave a single peak in GC-MS analysis. 1H-NMR (250
MHz, C6D6): δ (ppm) 9.74 (t, J ) 0.7 Hz, 1H), 4.16 (d, J ) 0.7
Hz, 2H), 3.72 (m, 2H), 3.64 (m, 2H), 3.56 (m, 2H), 3.38 (s, 3H).
13C-NMR (62.5 MHz, C6D6): δ (ppm) 200.1, 76.8, 72.2, 71.3, 71.0,
70.8, 58.6. EI-HRMS: m/z 162 (M+, <0.3%, exact mass not
measurable), 133.0899 ([M - CHO]+, 3%; C6H13O3 requires
133.0861).
(13) Diederich, F.; J onas, U.; Gramlich, V.; Herrmann, A.; Ringsdorf,
H.; Thilgen, C. Helv. Chim. Acta 1993, 76, 2445.
(14) Bianco, A.; Maggini, M.; Scorrano, G.; Toniolo, C.; Marconi, G.;
Villani, C.; Prato, M. J . Am. Chem. Soc. 1996, 118, 4072.

9072 J . Org. Chem., Vol. 61, No. 25, 1996
Additions and Corrections
λmax nm 700, 430, 320, 309, 255, 212. MALDI m/z: 987 (M +
H+). Anal. Calcd for C74H22N2O3: C, 90.05; H, 2.25; N, 2.84.
Found: C, 89.68; H, 1.99; N, 2.80.
ane): λmax nm 702, 430, 321, 306, 255, 213. MALDI m/z: 1041
(M+). Anal. Calcd for C75H31NO6: C, 86.44; H, 3.00; N, 1.34.
Found: C, 85.73; H, 2.70; N, 1.30.
Der iva tive 4c: 34% (toluene-methanol (9:1), Rf ) 0.22). 1H-
NMR (200 MHz): δ 4.97 (d, J ) 9.8 Hz, 1H), 4.64 (m, 1H), 4.38
(s, 1H), 4.36 (m, 1H), 4.26 (d, J ) 9.8 Hz, 1H), 4.04 (t, J ) 5.5
Hz, 2H), 3.66 (m, 18H), 3.36 (s, 3H), 3.35 (s, 3H). 13C-NMR (50
MHz) δ 156.1, 154.8, 154.2, 152.6, 147.2, 147.2, 147.1, 146.6,
146.3, 146.2, 146.2, 146.1, 146.0, 145.9, 145.8, 145.7, 145.6, 145.5,
145.4, 145.3, 145.2, 144.7, 145.3, 144.6, 144.4, 143.2, 143.0, 142.7,
142.6, 142.2, 142.1, 142.1, 142.0, 141.7, 141.6, 140.2, 140.1, 139.6,
139.3, 137.3, 136.4, 135.9, 135.5, 74.8, 73.5, 72.2, 72.0, 71.9, 70.9,
70.8, 70.6, 70.6, 70.3, 70.1, 67.7, 59.1, 52.3. UV-vis (cyclohex-
Su p p or tin g In for m a tion Ava ila ble: 1H and/or 13C NMR
spectra of compounds 2, 3c, and 4a -c (11 pages). This
material is contained in libraries on microfiche, immediately
follows this article in the microfilm version of the journal, and
can be ordered from the ACS; see any current masthead page
for ordering information.
J O961522T
Additions and Corrections
Vol. 40, 1975
Chloro sulfide 3 was prepared according to W. Adam
and M. Heil: Adam, W.; Heil, M. J . Am. Chem. Soc. 1991,
113, 1730. The chlorination with CCl4 in a basic medium
follows the principle given in the following: Meyers, C.
Y.; Matthews, W. S.; Ho, L. L.; Kolb, V. M.; Parady, T.
E. In Catalysis in Organic Synthesis; Smith, G. V., Ed.;
Academic Press: New York, 1977.
For a precedent to this type of radical probe, see:
Makosza, M.; Kwast, A. Bull. Soc. Chim. Belg. 1994, 103,
445.
Alber t P a d w a ,* An d r ew Au , Geor ge A. Lee, a n d
Willia m Ow en s. Photochemical Ring-Opening Reactions of
Substituted Chromenes and Isochromenes.
Page 1148. The NMR spectrum of trans-2-methoxy-
3-hydroxy-2-phenylindan (33) should read as follows:
NMR (CDCl3) δ 3.10 (s, 3H), 3.90 (broad s, 2H), 4.93
(s, 1H), and 6.8-7.8 (m, 9H). The integration pattern
previously reported was inadvertently switched.
J O964025L
J O964020O
J osep h A. Leon etti, Tim Gr oss, a n d R. Da n iel Little* .
Cycloaddition-Fragmentation as a Route to Bicyclic Ring Sys-
tems. Use of the Intermolecular Diyl Trapping Reaction.
Vol. 61, 1996
J a m es A. Ma r sh a ll* a n d Kevin W. Hin k le. Stereoselective
SE2′ Additions of Enantioenriched Allylic Tin and Indium
Reagents to Protected Threose and Erythrose Aldehydes:
General Strategy for the Stereocontrolled Synthesis of Precursors
to the Eight Diastereomeric Hexoses and Their Enantiomers.
Page 1787. We are pleased to acknowledge the as-
sistance of Ms. J anette Gunther with various aspects of
this research.
A
J O964016C
Page 107. The correct structures for transition states
7.2 and 7.3 are shown below.
Syed A. Abba s, M. Bila yet Hossa in , Dick va n d er Helm ,
F r a n cis J . Sch m itz,* Ma u r een La n ey, Ron n el Ca bu sla y,
a n d R a n d a ll C. Sch a t zm a n . Alkaloids from the Tunicate
Polycarpa aurata from Chuuk Atoll.
Page 2709. The correct spelling is of the first author’s
name is Syed A. Abbas.
J O964019P
J O9640231
An d r ew G. H. Wee* a n d J . Slobod ia n . Metal-Catalyzed
Reaction of Indoline Diazoamides Possessing
Substituent: Site-Selectivity, Diastereoselectivity, and Chemose-
lectivity.
a C-2 CH2X
J .-M. Ma tta lia , M. Ch a n on , a n d C. J . M. Stir lin g*. A New
Radical Clock for Testing the Possibility of Electron Transfer
from Carbanions.
Page 2899. References 15a,b cited for 12a in the
supplementary material section should read 7a,b. Refer-
ence 7a should be cited as follows: Cliff, G. R.; J ones,
G.; Woollard, J . McK. Tetrahedron Lett. 1973, 2401.
Under Gen er a l in the Experimental Section, “... refer-
ence 5” should read “... reference 1”.
Page 1153. The kinetic relation is k ) kH([5]/
[1])[Bu3SnH]0, where kH ) 3.6 × 106 M-1 s-1 (80 °C).
Page 1154. R ed u ct ion s of Ch lor ocyclop r op yl-
m eth yl P h en yl Su lfon e 4 w ith Bu 3Sn H. The deter-
mined ratios are 5/1. In refluxing cyclohexane this ratio
is 92/8 when [Bu3SnH]0 ) 0.26 M (5 equiv) and 0.27 M
(6.6 equiv).
J O9640276