Total synthesis of acinetoferrin

Article abstract of DOI:10.1021/jo9717144

The total synthesis of a novel siderophore, acinetoferrin, is described. The key transformation involves the tandem oxidation and acylation of the N₃-amino group of N₁-BOC propane diamine prior to coupling with the external carboxyls of citric acid. A 'one-pot-two-step' reaction converted a primary amine (RNH₂) into a O-benzoyl hydroxamate (i.e., RN(OOCPh)COR') in good yield (68%). These studies demonstrated the utility of the O-benzoyl protecting group in the synthesis of α,β-unsaturated hydroxamic acids.

Full text of DOI:10.1021/jo9717144

J . Org. Chem. 1998, 63, 1491-1495
1491
Tota l Syn th esis of Acin etofer r in
Queenie Xianghong Wang and Otto Phanstiel IV*
Center for Diagnostics and Drug Development, Department of Chemistry, University of Central Florida,
Orlando, Florida 32816-2366
Received September 15, 1997
The total synthesis of a novel siderophore, acinetoferrin, is described. The key transformation
involves the tandem oxidation and acylation of the N3-amino group of N1-BOC propane diamine
prior to coupling with the external carboxyls of citric acid. A “one-pot-two-step” reaction converted
a primary amine (RNH2) into a O-benzoyl hydroxamate (i.e., RN(OOCPh)COR′) in good yield (68%).
These studies demonstrated the utility of the O-benzoyl protecting group in the synthesis of R,â-
unsaturated hydroxamic acids.
In tr od u ction
interest as structurally modified derivatives of 1-3 may
be used as suitable oral vectors, while limiting the
proliferation of pathogenic bacterial strains, which grow
via citrate siderophores.11
Iron is an essential micronutrient required by mam-
mals and most other terrestrial lifeforms. Accessing iron
is problematic as it exists mainly as insoluble ferric
hydroxide polymers in the biosphere. Microorganisms
have evolved a group of low molecular weight, virtually
ferric ion specific chelating agents (i.e., siderophores) to
solubilize and transport iron within their environment.
Interestingly, the ability of these ligands to transport
ferric ion has found clinical use in the treatment of iron-
overload diseases.1
In this “across-kingdom” application, the siderophores
utilized for iron internalization in microorganisms are
found to promote the clearance of iron from humans.
Indeed, thalassemia patients have been treated success-
fully with deferrioxamine B (DFO) isolated from Strep-
tomyces pilosus for the past 35 years.1-6 However, a
major difficulty in iron chelation therapy is patient
compliance with the regimen involving triweekly infu-
sions of DFO. Since patients would be more receptive
to an orally administered drug, research has now focused
on the development of orally active iron chelators.
A number of siderophores contain a citric acid compo-
nent,7 as seen in schizokinen (1)8 isolated from Bacillus
megaterium, aerobactin (2)9 isolated from Aerobacter
strains, and nannochelin A (3)10 isolated from Nanno-
cystis exedens (Figure 1). This class of siderophores is of
Recently, acinetoferrin 4 was isolated from Acineto-
bacter haemolyticus bacteria by Okujo et al.12 This novel
chelator was shown to consist of a rather unusual trans-
2-octenoylhydroxamic acid appended to a citric acid
framework.12 Acinetoferrin represents an interesting
synthetic target because of its unusual 2(E)-octenoylhy-
droxamic acid moiety and because previous synthetic
strategies using O-benzyl-protected hydroxamates are
not readily applicable.13-16 Removal of such an O-benzyl
group from an acinetoferrin precursor via hydrogenolysis
would compromise the octenoylhydroxamate functionality
by a competing hydrogenation process. Attempts at
alternative reduction methods (EtSH, BF3‚Et2O) by Pat-
tenden et al. with a tris-O-benzyl-protected nannochelin
A precursor gave only a 30% yield of 3.17 Recognizing
the structural similarity of acinetoferrin and nannochelin
A, we hoped to apply our previous method18 for the
synthesis of R,â-unsaturated hydroxamic acids to the
synthesis of acinetoferrin.
Resu lts a n d Discu ssion
Acinetoferrin consists of two N-(3-aminopropyl)-N-
(hydroxy)-2(E)-octenamide fragments attached via the
primary amino group to the terminal carboxyl groups of
citric acid by amide bonds (Figure 1).12 Our strategy
involved the construction of a N-(3-aminopropyl)-N-
(hydroxy)-2(E)-octenamide precursor via a tandem oxida-
tion-acylation of the primary amino group of a protected
(1) Bergeron, R. J .; McManis, J . S. In The Development of iron
Chelators for Clinical Use; Bergeron, R. J ., Brittenham, G. M., Eds.;
CRC Press: Boca Raton, FL, 1994; pp 237-273.
(2) Aksoy, M.; Birdwood, G. F. B. Hypertransfusion and Iron
Chelation in Thalassemia; Hans Huber Publishers: Berne, 1985, 80.
(3) Graziano, J . H.; Markenson, A.; Miller, D. R.; Chang, A.; Bestack,
M.; Meyers, P.; Pisciotto, P.; Rifkind, A. J . Pediatr. 1978, 92, 648-
652.
(4) Pippard, M. J .; Callender, S. T. Br. J . Haemat. 1983, 54, 503-
507.
(5) Peter, H. H. Proteins of Iron Storage and Transport; Spik, G.,
Montreuil, J ., Gichton, R. R., Mazwier, J ., Eds.; Elsevier: Amsterdam,
1985, p 293.
(6) Warne, P., Ed. Ciba Geigy J . 1991, 4, 32-35.
(7) Neilands, J . B.; Ratledge, C. CRC Handbook of Microbiology, 2nd
ed., Vol. IV.; Laskin, A. L., Lechevalier, H. A., Eds.; CRC Press, Boca
Raton, FL, 1982; pp 565-574.
(8) Milewska, M. J .; Chimiak, A.; Glowacki, Z. J . Prakt. Chem. 1987,
329, No. 3, 447-456.
(9) Maurer, P. J .; Miller, M. J . J . Am. Chem. Soc. 1982, 104, 3096-
3101.
(11) Neilands, J . B. Siderophores as Biological Deferration Agents
and the Regulation of their Synthesis by Iron, ref 1, p 165.
(12) Okujo, N.; Sakakibara, Y.; Yoshida, T.; Yamamoto, S. Biometals
1994, 7, 170-176.
(13) Bergeron, R. J .; Wiegand, J .; McManis, J . S.; Perumal, P. T. J .
Med. Chem. 1991, 34, 3182-3187.
(14) Bergeron, R. J .; McManis, J . S. Tetrahedron 1989, 45, No. 16,
4939-4944.
(15) Bergeron, R. J .; McManis, J . S.; Perumal, P. T.; Algee, S. E. J .
Org. Chem. 1991, 56, 5560-5563.
(16) Bergeron, R. J .; McManis, J . S. CRC Handbook of Microbial
Iron Chelates 1991, 271-307.
(17) Mulqueen, G. C.; Pattenden G.; Whiting, D. A. Tetrahedron
1993, 40, 9137-9142.
(10) Kunze, B.; Trowitzsch-Kienast, W.; Hofle, G.; Reichenbach, H.
J . Antibiot. 1992, 45, No. 2, 147-150.
(18) Bergeron, R. J .; Phanstiel IV, O. J . Org. Chem. 1992, 57, 7140-
7143.
S0022-3263(97)01714-3 CCC: $15.00 © 1998 American Chemical Society
Published on Web 02/04/1998

1492 J . Org. Chem., Vol. 63, No. 5, 1998
Wang and Phanstiel
F igu r e 1. Iron chelators predicated upon a citric acid component.
propanediamine derivative prior to coupling with the
external carboxyls of citric acid.
phasic medium (CH2Cl2/aqueous buffer solution) favored
the formation of the benzoyloxy amine 7 at the expense
of the benzamide 8.21 Indeed, a careful study of the
benzoyl peroxide mediated oxidation of 6 revealed a
dramatic dependence on the pH of the aqueous phase and
the molar ratio of the amine and BPO. To date, the
highest yield of 7 (72%) was obtained with equal volumes
of a pH 10.5 carbonate buffer solution and 0.1 M amine
6 in CH2Cl2 using 2 equiv of benzoyl peroxide (versus the
amine) at room temperature.
While the selective coupling of amino substrates with
the terminal carboxyls of citric acid can be achieved
utilizing either 2-substituted-1,3-bis-activated esters of
citric acid or anhydromethylene citric acid,8,22 the re-
ported imide formation associated with the latter ap-
proach prompted the use of the former coupling meth-
odology. The selective activation of the 1,3 carboxyls of
citric acid was accomplished by the formation of the bis-
activated ester: 2-tert-butyl-1,3-di-N-(hydroxy)succinim-
idyl citrate, 10.8
Synthesis of tert-butylacinetoferrin (11) required the
coupling of the functionalized propylamine fragment to
the activated citric acid framework. Previous experience
revealed that higher yields were obtained in the conden-
sation step if the benzoyl group was removed prior to
amide formation.18 Therefore, 9 was treated with 10%
NH3/MeOH solution at -23 °C to give the “free” hydrox-
amate 12 (89%). Treatment of 12 with trifluoroacetic
acid (TFA) at 0 °C gave the N-(3-aminopropyl)-N-(hy-
droxy)-2(E)-octenamide, TFA salt (13). Condensation of
13 with a solution of the bis N-hydroxysuccinimide (NHS)
ester 108 and triethylamine (TEA) in 1,4-dioxane gave
tert-butylacinetoferrin (11) in 86% yield. Finally, treat-
In 1990 Milewska et al. reported the conversion of the
free Nδ-amine of NR-(benzyloxycarbonyl)-L-ornithine tert-
butyl ester to an O-benzoyl-protected hydroxamate using
benzoyl peroxide (BPO) followed by acetyl chloride.19 As
one strategy to access acinetoferrin involved the oxidation
and acylation of a propanediamine derivative, this tan-
dem sequence seemed a worthwhile approach. Indeed,
this transformation was employed in our earlier synthesis
of nannochelin A,18 albeit in 25% yield.
As shown in Scheme 1, the reaction of 1,3-propanedi-
amine (5) and di-tert-butyl dicarbonate afforded 3-(tert-
butoxycarbonylamino)propylamine (6) in 81% yield. Com-
pound 6 was dissolved in a biphasic carbonate buffer (pH
) 10.5) and reacted with 2 equiv of BPO dissolved in CH2-
Cl2 at room temperature. This oxidation step generated
the desired benzoyloxy amine 7 and benzamide 8. The
mixture was then acylated with trans-2-octenoyl chloride
to give the desired N-(3-(tert-butoxycarbonylamino)pro-
pyl)-N-(benzoyloxy)-2(E)-octenamide (9) in 68% isolated
yield. This is a significant increase in yield, when
compared to our earlier efforts with 3, and was realized
only after a careful study of the initial oxidation step.
The mono-oxidation of primary amines is difficult at
best as the products are often over-oxidized by the
reaction conditions.20 Early studies with 6 using benzoyl
peroxide as the oxidant gave the desired benzoyloxy
amine 7 and identified two competing pathways: N-
oxidation and N-acylation.21 We discovered that a bi-
(19) Milewska, M. J .; Chimiak, A. Synthesis 1990, 233-234.
(20) Gragerov, I. P.; Levit, A. F. J . Gen. Chem. USSR 1960, 30,
3690-3695.
(21) Wang, Q. X.; King, J .; Phanstiel IV, O. J . Org. Chem. 1997, 62,
8104-8108.
(22) Lee, B. H.; Miller, M. J . J . Am. Chem. Soc. 1982, 104, 3096-
3101.

Total Synthesis of Acinetoferrin
J . Org. Chem., Vol. 63, No. 5, 1998 1493
Sch em e 1. Syn th esis of Acin etofer r in (4)a
F igu r e 2. E and Z rotamers of a typical hydroxamic acid.
predominating. The preference for Z isomers in these
solvents has been observed by Brown and co-workers
with other hydroxamic acids.23
During our scale-up of 4, we observed a facile cycliza-
tion in the presence of TFA to give the imide 14. This
phenomenon was temperature dependent. Workup at
room temperature gave 4, whereas elevated temperatures
1
during TFA removal gave 14. The H NMR spectrum of
14 (in CD3OD) is shown in Figure 3. A sample of 4 in
CD3OD was 79% converted to 14 upon standing at room
temperature for 9 months. Similar cyclizations were
recently observed with another citric acid based chelator,
rhizoferrin,25 and other amide containing systems.26-28
In summary, the synthesis provided both proof of
structure of the natural product and a method, which
should allow facile access to a variety of acinetoferrin
homologues with different amino fragments. The mild,
yet selective, method of converting a primary amine into
a protected hydroxamate should be of immense value to
medicinal chemists interested in synthesizing function-
alized hydroxamic acids.
Exp er im en ta l Section
Ma ter ia ls a n d Meth od s. Lipophilic Sephadex LH-20 was
obtained from the Sigma Chemical Company. Silica gel 60
(70-230) mesh was purchased from EM Science, Darmstadt,
Germany. Solvents were freshly distilled prior to use. Re-
agents were purchased either from the ACROS Chemical
Company or the Aldrich Chemical Co. and were used without
further purification. trans-2-Octenoic acid was purchased from
Pfaltz and Bauer, Inc. All solutions are expressed in volume
% unless otherwise indicated. Ammonia-containing solutions
were prepared by measuring out concentrated aqueous NH4-
OH in the listed volume %.
a
Reagents: (a) di-tert-butyl dicarbonate, 10% TEA/MeOH; (b)
benzoyl peroxide (2 equiv)/CH2Cl2, pH ) 10.5 aqueous sodium
carbonate buffer, rt; (c) trans-2-octenoyl chloride, CH2Cl2; (d) 10%
NH3/MeOH; (e) TFA, 0 °C; (f) TEA, 10, dry dioxane, 15 °C.
Acin etofer r in (4). TFA (2 mL) was added dropwise to 11
(34 mg, 5.3 µmol) at 0 °C. After the addition was complete,
the stirred solution was allowed to warm to room temperature.
TLC (7% EtOH/CHCl3) showed that no starting material
remained after 1 h. The volatiles were removed under high
vacuum and gave a yellow oil, which was eluted on LH-20
Sephadex (3.5 g) with 10% EtOH/toluene to give acinetoferrin
(4) (30 mg, 97%). 4: Rf ) 0.31 (benzene:acetic acid:water (125:
72:3, matched lit. value),12 Rf ) 0.38 in 25% MeOH/CHCl3; 1H
NMR (500 MHz) (CD3OD) δ 6.84 (dt, 2H, olefinic), 6.61 (d, 2H,
olefinic), two triplets at 3.70 and 3.65 (2.1:1 integration ratio
respectively, total integration for both signals 4H, CH2NO),
3.20 (m, 4H, CH2NCO), 2.71 (d, 2H, CH2), 2.64 (d, 2H, CH2),
2.24 (dt, 4H, CH2CdC), 1.82 (m, 4H, CH2) 1.48 (m, 4H, CH2),
1.34 (m, 8H, CH2), 0.90 (t, 6H, CH3); 1H NMR (600 MHz)
(DMSO-d6) δ 9.83 (s, 1H, COOH), 7.96 (s, 2H), 6.70 (dt, 2H,
olefinic), 6.51 (d, 2H, olefinic), two triplets at 3.55 and 3.48
(2.3:1 integration ratio respectively, total integration for both
signals 4H, CH2NO), 3.02 (m, 4H, CH2NCO), 2.50 (dd, 4H,
ment of 11 with TFA gave acinetoferrin, (4) in 97% yield
(see Scheme 1).
The Rf values of 4 in several solvent systems (as
determined by TLC) were identical with those reported
for the natural product.12 Although we were unable to
obtain an authentic sample, the mass spectrum and the
proton NMR spectrum of 4 in CD3OD of the synthetic
material were in complete agreement with the published
spectra.12 In addition, 2-D NMR experiments confirmed
1
the C-H assignments made by Okujo.12 The H NMR
spectrum of 4 (in CD3OD) revealed both Z (Peak at δ 3.70)
and E (peak at δ 3.65) rotamers of acinetoferrin (4) (see
also ref 12), whereas in DMSO-d6 these isomers are
detected at δ 3.55 and 3.48, respectively. These signals
were assigned to the methylene group adjacent to the
hydroxamic acid nitrogen in structure 4 (e.g., CH2N(OH)-
COR).12 The rotamers are generated from the hindered
rotation about the hydroxamic acid carbon-nitrogen
bond (Figure 2) and do not pertain to the E-alkene
moiety. The E and Z rotamers were assigned by data
obtained from earlier model systems23,24 and were ob-
served in both CD3OD and DMSO-d6 with the Z isomer
(24) Bergeron, R. J .; McManis, J . S.; Phanstiel IV, O.; Vinson, J . R.
T. J . Org. Chem. 1995, 60, 109-114.
(25) Bergeron, R. J .; Xin, M.; Smith, R. E.; Wollenweber, M.;
McManis, J . S.; Ludin, C.; Abboud, K. A. Tetrahedron 1997, 53, 427-
434.
(26) Lee, B. H.; Miller, M. J . J . Org. Chem. 1983, 48, 24-31.
(27) Nau, C. A.; Brown, E. B.; Bailey, J . R. J . Am. Chem. Soc. 1925,
47, 2596-2606.
(28) Kisfaludy, L.; Schon, I.; Renyei, M.; Gorog, S. J . Am. Chem.
Soc. 1975, 97, 5588-5589.
(23) Brown, D. A.; Glass, W. K.; Mageswaran, R.; Mohammed, S.
A. Magn. Reson. Chem. 1991, 29, 40-45.

1494 J . Org. Chem., Vol. 63, No. 5, 1998
Wang and Phanstiel
F igu r e 3. 1H NMR spectrum of 14 in CD3OD.
CH2), 2.19 (dt, 4H, CH2CdC), 1.63 (m, 4H, CH2) 1.41 (m, 4H,
CH2), 1.28 (m, 8H, CH2), 0.86 (t, 6H, CH3); high-resolution
mass spectrum (FAB), theory for (C28H49N4O9) M + 1 )
585.3500, found M + 1 ) 585.3507.
twice with additional CH2Cl2 (2 × 100 mL). The organic layers
were combined, dried over anhydrous Na2SO4, filtered, and
concentrated to give the crude product. The crude amide 9
was subjected to flash column chromatography, eluting with
30% ethyl acetate/hexane, to give the N-(3-(tert-butoxycarbo-
nylamino)propyl)-N-(benzoyloxy)-2(E)-octenamide 9 (8.58 g:
68%) as a colorless oil. 9: Rf ) 0.33 in 30% ethyl acetate/
hexane; 1H NMR (CDCl3) δ 8.12 (d, 2H, aromatic), 7.71 (t, 1H,
aromatic H), 7.54 (t, 2H, aromatic H), 7.03 (dt, 1H, olefinic),
6.05 (d, 1H, olefinic), 5.20 (broad, 1H, NH), 3.92 (t, 2H, CH2-
NO), 3.24 (m, 2H, CH2), 2.15 (m, 2H, CH2CdC), 1.82 (m, 2H,
CH2), 1.64 (m, 2H, CH2), 1.53-1.15 (m, 13H, tert-butyl and 2
CH2), 0.83 (t, 3H, CH3). Anal. Calcd for C23H34N2O5: C, 66.01;
H, 8.19; N, 6.69. Found: C, 65.83; H, 8.17; N, 6.61.
3-(ter t-Bu toxyca r bon yla m in o)p r op yla m in e (6). 1,3-
Diaminopropane (5) (11.16 g, 150 mmol) was dissolved in 350
mL of a 10% TEA/MeOH solution. A solution of di-tert-butyl
dicarbonate (10.9 g, 50 mmol) and MeOH (20 mL) was added
to this mixture with vigorous stirring. The mixture was
refluxed for 2 h. The tert-butoxycarbonylation was complete
as evidenced by thin-layer chromatography (TLC) using 4%
MeOH/CHCl3. The mixture was concentrated and subjected
to flash column chromatography to give the mono-BOC amine
1
6 (7.06 g, 81%); Rf ) 0.24 (4% NH3/MeOH); H NMR (CDCl3)
δ 4.95 (br s, 1H, NH), 3.21 (m, 2H, CH2NBOC), 2.72 (t, 2H,
CH2N), 1.58 (m, 2H, CH2), 1.40 (s, 9H, tert-butyl).
2-ter t-Bu tyl-1,3-di-N-(h ydr oxy)su ccin im idyl Citr ate (10).
The bis NHS ester 10 was prepared by a previous method (see
ref 8 and an improved purification method in ref 18).
N-(3-(ter t-Bu toxycar bon ylam in o)pr opyl)ben zam ide (8).
The byproduct, 8, was also isolated (0.67 g, 8%) during the
Acin etofer r in ter t-Bu tyl Ester (11). Triethylamine (3.5
g, 34.7 mmol) was added dropwise to a mixture of 10 (1.25 g,
2.83 mmol) and 13 (1.82 g, 6.2 mmol) in 60 mL of dry dioxane
at 15 °C under nitrogen. The solution was allowed to warm
to room temperature and stirred overnight. The volatiles were
removed under high vacuum and the residue was chromato-
graphed using pretreated silica gel. Before use the silica gel
was made “iron free” by washing with methanol:acetone:10M
HCl (45:45:10), followed by a 10 wt % Na2CO3 solution, and
then rinsed with deionized water until pH 7 and air-dried.
Eluting the column with 7.5% MeOH/CHCl3 gave the tert-butyl
ester of acinetoferrin (11) (1.56 g, 86%). 11: Rf ) 0.27 in 7%
MeOH/CHCl3; 1H NMR (600 MHz) (CDCl3) δ 9.38 (broad s,
1H), 7.46 (s, 2H), 6.85 (m, 2H, olefinic), 6.60 (d, 2H, olefinic),
two multiplets at 3.74 and 3.68 (1:1 integration ratio respec-
tively, total integration 4H, CH2), 3.22 (broad s, 4H, CH2), 2.63
(dd, 4H, CH2), 2.19 (dt, 4H, CH2), 1.84 (m, 4H, CH2), 1.46 (s,
9H, tert-butyl), 1.44 (m, 4H, CH2), 1.29 (m, 8H, CH2), 0.88 (t,
6H, CH3); high-resolution mass spectrum (FAB): theory for
(C32H56N4O9) M + 1 ) 641.4126, found M + 1 ) 641.4143.
Anal. Calcd for C32H56N4O9: C, 59.98; H, 8.81; N, 8.74.
Found: C, 59.69; H, 8.69; N, 8.51.
N-(3-(ter t-Bu toxyca r bon yla m in o)p r op yl)-N-(h yd r oxy)-
2(E)-octen a m id e (12). A 10% NH3/MeOH solution (30 mL)
was added dropwise to 9 (3.7 g, 8.85 mmol) under nitrogen at
-23 °C using a dry ice/CCl4 bath. The reaction was monitored
by TLC (30% ethyl acetate/hexane). After 3 h, the reaction
was complete. The mixture was concentrated to give a crude
light yellow oil, which was subjected to flash column chroma-
tography on “pretreated” silica gel (see procedure for 11). The
product was eluted with 40% ethyl acetate/hexane to give 12
as a light yellow solid (2.47 g, 89%). Rf ) 0.29 in 45% ethyl
acetate/hexane. 12: 1H NMR (CDCl3) δ 6.83 (m, 1H), 6.57 (m,
1H), 5.15 (broad, 1H), 3.67 (t, 2H), 3.09 (m, 2H), 2.13 (m, 2H),
1
synthesis of 9. 8: Rf ) 0.29 in 40% ethyl acetate/hexane; H
NMR (CDCl3) δ 7.85 (d, 2H, aromatic H), 7.44 (m, 3H, aromatic
H), 4.92 (broad s, 1H, NH), 3.51 (m, 2H, CH2), 3.23 (m, 2H,
CH2), 1.72 (m, 2H, CH2), 1.43 (s, 9H, tert-butyl); high-resolution
mass spectrum (FAB): theory for (C15H23N2O3) M + 1 )
279.1709, found M + 1 ) 279.1703.
N-(3-(ter t-Bu t oxyca r b on yla m in o)p r op yl)-N-(b en zoy-
loxy)-2(E)-octen a m id e (9). A solution of benzoyl peroxide
(14.52 g, 60 mmol) and 225 mL of CH2Cl2 was added dropwise
at room temperature to a vigorously stirred mixture of 6 (5.22
g, 30 mmol) and 300 mL of a carbonate buffer solution (at pH
10.5). The buffer solution was prepared by combining 222 mL
of 0.75 N aqueous NaHCO3 and 78 mL of 1.5 N aqueous NaOH.
The starting material was consumed after 4 h as shown by
TLC (4% NH3/MeOH). Column chromatography (40% ethyl
acetate/hexane) was used to separate N-(benzoyloxy)-3-(tert-
butoxycarbonylamino)propylamine (7) and the benzamide 8
(e.g., Rf ) 0.43 and 0.29, respectively). A characterization of
7 is included for completeness. 7: 1H NMR (CDCl3) δ 8.03 (d,
2H, aromatic H), 7.58 (m, 1H, aromatic H), 7.47 (m, 2H,
aromatic H), 4.81 (broad m, 1H, NH), 3.22 (m, 4H, CH2), 1.82
(m, 2H, CH2), 1.41 (s, 9H, tert-butyl); high-resolution mass
spectrum (FAB), theory for (C15H23N2O4) M + 1 ) 295.1658,
found M + 1 ) 295.1645. However, further studies showed
that the next step could be accomplished without isolation of
7. After the first reaction was complete, a solution of trans-
2-octenoyl chloride (4.81 g, 30 mmol) in 30 mL of CH2Cl2 was
added dropwise over 15 min. Note: the acid chloride was
generated from trans-2-octenoic acid using refluxing thionyl
chloride and was distilled under vacuum prior to use. The
disappearance of 7 was monitored by TLC (40% ethyl acetate/
hexane). After the acylation was complete, the organic layer
was separated and the remaining water layer was washed

Total Synthesis of Acinetoferrin
J . Org. Chem., Vol. 63, No. 5, 1998 1495
1.75 (m, 2H), 1.53-1.15 (m, 15H, tert-butyl and CH2), 0.83 (t,
168.7, 171.6, 177.0, 180.6; high-resolution mass spectrum
(FAB), theory for (C28H47N4O8) M + 1 ) 568.3347, found M +
1 ) 568.3347. Anal. Calcd for C28H47N4O8: C, 59.24; H, 8.34;
N, 9.87. Found: C, 59.14; H, 8.16; N, 9.93.
3H); high-resolution mass spectrum (FAB), theory for (C16H31
-
N2O4) M + 1 ) 315.2283, found M + 1 ) 315.2273.
N-(3-Am in op r op yl)-N-(h yd r oxy)-2(E)-octen a m id e, Tr i-
flu or oa cetic Acid Sa lt (13). TFA (50 mL) was added
dropwise over 1 min to 12 (1.97 g, 6.27 mmol) at 0 °C under
nitrogen. The ice bath was removed and the solution stirred
for 5 min. TLC (45% ethyl acetate/hexane) showed that the
reaction was complete. The volatiles were removed under
reduced pressure to give the desired salt 13 (2.06 g). The solid
residue was consumed in the synthesis of tert-butylacineto-
ferrin (11).
Ack n ow led gm en t. The authors thank the generous
support of this work by the University of Central Florida
Department of Chemistry. In addition, the 600 and 500
MHz NMR spectra were obtained by Dr. Barry Sch-
weitzer (and Kerry Keertikar) at the Walt Disney
Memorial Cancer Institute in Orlando, FL, and Brian
Killday at the Harbor Branch Oceanographic Institute
in Ft. Pierce, FL, respectively. All mass spectral data
were provided by Dr. David Powell and Maria del Pilar
at the University of Florida. We also appreciate the
financial support from the National Science Foundation
in the purchase of our 200 MHz NMR instrument
(Grant CHE-8608881).
Im id e of Acin etofer r in (14). Compound 14 was generated
during the removal of TFA under heat and reduced pressure.
14: Rf ) 0.77 in 50% acetone/CHCl3, Rf ) 0.64 in benzene:
acetic acid:water (125:72:3), and Rf ) 0.46 in 15% MeOH/
1
CHCl3; H NMR (600 MHz) (CD3OD) δ 6.84 (m, 2H), 6.62 (d,
2H), 3.69 (m, 4H), 3.57 (t, 2H), 3.17 (m, 2H), 3.05 (d, 1H), 2.93
(d, 1H), 2.77 (d, 1H), 2.63 (d, 1H), 2.24 (m, 4H), 1.96 (m, 2H),
1.80 (m, 2H), 1.49 (m, 4H), 1.36 (m, 8H), 0.93 (t, 6H); 13C NMR
(600 MHz) (CD3OD) δ 14.5, 23.7, 26.2, 27.8, 29.3, 32.7, 33.6,
37.6, 37.7, 43.2, 43.3, 47.0, 47.1, 73.8, 120.5, 120.6, 148.4, 148.5,
J O9717144