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Y. Ma et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3450–3452
7
8
9
10
Scheme 3.
10 penetrated the cell membranes much more quickly
(Fig. 3 and Table 2) and therefore has greater potential
as an intracellular iron probe. Studies are currently in
progress with these probes in both hepatocytes and
lymphocytes.
Figure 3. Fluorescence intensity of 10 lM 6 (kex 324 nm; kem 464 nm)
and 6 lM 10 (kex 383 nm; kem 447 nm) recorded during incubation at
20 °C in the presence of suspension of washed human erythrocyte
ghosts resealed in the presence of 100 lM iron(II) and 1 mM ascorbic
acid. The suspension medium A also contained 25 mM MOBS and
1 mM ascorbic acid, and medium B contained 25 mM MOBS and
1 mM ascorbic acid plus 0.1 mM DTPA.
Acknowledgments
The work was financially supported from MRC
(G9901131). We are grateful to Universities UK for
ORS awards.
sealed ghosts to chelate iron but also the iron leaked
from the ghosts to combine with the probe in the super-
natant. The two processes are consistent with a fit of the
respective data to a double exponential decay function,
as shown in Table 2. However, in the presence of DTPA
(Fig. 3-6B), the effluxed iron was preferentially bound to
the high-affinity ligand, DTPA, and led to a single-expo-
nential decay function, representing permeation of the
fluorescent chelator into the resealed ghosts (Fig. 3
and Table 2).
References and notes
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Anal. Biochem. 2002, 304, 1.
Previous experiments on human erythrocyte ghosts indi-
cate that enhanced lipophilicity of iron fluorescent
probes led to enhanced membrane penetration.10 To in-
crease the lipophilicity of fluorescent probe 6, more
hydrophobic diamine derivatives were reacted with the
acidic pyranone 3 to produce the corresponding bicyclic
lactam derivatives 7, 8, 9 and 10 (Scheme 3). ClogP val-
ues (calculated logP values; the logarithm of partition
coefficients were calculated using Chemoffice 6.0 from
Cambridgesoft Corporation, Cambridge, UK) for the
molecules are À1.16, À1.28, À0.85 and À0.40, respec-
tively. Compared with 6, the most hydrophobic probe
12. Petrat, F.; de Groot, H.; Sustmann, R.; Rauen, U. Biol.
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Table 2. Permeability parameters for 6 and 10 across human eryth-
rocyte ghost membranes
ÀDTPA
+DTPA
14. Dobbin, P. S.; Hider, R. C.; Hall, A. D.; Taylor, P. D.;
Sarpong, P.; Porter, J. B.; Xiao, G. Y.; van der Helm, D.
J. Med. Chem. 1993, 36, 2448.
15. Hare, L. E.; Lu, M. C.; Sullivan, C. B.; Sullivan, P. T.;
Counsell, R. E. J. Med. Chem. 1974, 17, 1.
16. Spectroscopic data for 6: MS: 181 (M+1), 1H NMR
(360 MHz, DMSO-d6): d (ppm) 3.64–3.68 (m, 2H), 4.57 (t,
J = 5.8 Hz, 2H), 7.89 (s, 1H), 8.41 (s, 1H), 9.07 (br s, 1H);
Anal. Calcd for C8H11ClN2O4: C, 40.95; H, 4.73; N, 11.94.
Found: C, 41.62; H, 4.45; N, 11.99
6
A1 (a.u.)
t1 (s)
127
108
65
117
127
—
A2 (a.u.)
t2 (s)
283
—
10
A1 (a.u.)
t1 (s)
127
15
252
24
A2 (a.u.)
t2 (s)
14
207
—
—
17. Bajorath, J. Nat. Rev. Drug. Discov. 2002, 1, 882.