phases were collected and the solvent was removed under vacuum.
The product was dried and 1.06 g (72% yield) of the product were
obtained.
selectin-coated nanoparticles to a selectin ligand, immobilized on
a sensor chip, was tested, and the resulting signal was henceforth
referred to as 100% binding. Pre-incubation of the selectin-coated
particles with varying concentrations of a selectin inhibitor (here a
gold colloid with a ligand shell) decreased the binding signal. The
calculated IC50 values (half inhibitory concentration) is the molar
concentration of the inhibitor needed to reduce the binding signal
to 50% of the initial value. Further details regarding the binding
assay can be found in the ESI† and in Ref. 22.
1H NMR (CDCl3, 400 MHz): d = 1.35–1.45 (m, 2H), 1.57 (quin,
2H), 1.63 (quin, 2H), 2.30 (s, 3H) 2.33 (t, 2H), 2.85 (t, 2H).
Synthesis of N-hydroxysuccinimide-6-acetylsulfanylhexanoate
An amount of 1.06 g (5.58 mmol) of 6-acetylsulfanylhexanoic
acid was added to a solution of 708 mg (6.16 mmol) of N-
hydroxysuccinimide in 100 ml of CH2Cl2. A solution of 1.27 g
(6.16 mmol) of dicyclohexylcarbodiimide (DCC) in 5 ml of
CH2Cl2 was added dropwise and after a few minutes a white solid
precipitated. The reaction mixture was stirred for 1 h. The solvent
was removed under vacuum and the crude product was dissolved
in little CH2Cl2 and filtered. The solvent was evaporated and 1.55 g
of a brownish oil were obtained (98% yield).
Cytotoxicity assay
R
The Promega CellTiter 96ꢀ AQueous Cell Proliferation Assay was
applied. L-selectin expressing T-cells (Jurkat) cells were seeded in
96-well plated at a density of 5000 cells/well. The Au-colloids NP-
1-sulf, NP-10-sulf and NP-16-sulf were added at concentrations of
0.003 nM, 0.03 nM, 0.3 nM, 3 nM and 30 nM. In comparison to
the untreated control cell proliferation was detected after 3 days
incubation by measuring the conversion of the tetrazolium com-
pound 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-
2-(4-sulfophenyl)-2H-tetrazolium (MTS) and an electron coupling
reagent, phenazine ethosulfat (PES) into a water-soluble formazan
that absorbs light at 490 nm by cellular dehydrogenases. The assay
was performed in triplicates at each concentration. The untreated
control was set to 100% proliferation.
1H NMR (CDCl3, 400 MHz): d = 1.46 (m, 2H), 1.61 (quin, 2H),
1.74 (quin, 2H), 2.30 (s, 3H), 2.59 (t, 2H), 2.82 (s, 4H), 2.86 (t, 2H).
Synthesis of [5-(2-hydroxy-1-hydroxymethyl-ethylcarbamoyl)-
pentyl]-S-thioacetate
An amount of 765 mg (8.41 mmol) of serinol 10 was added to 20 ml
of a solution of 1.62 g (5.64 mmol) of N-hydroxysuccinimide-6-
acetylsulfanylhexanoate in DMF. After a few minutes the solution
became cloudy and the mixture was stirred for 30 min. The solvent
was removed under vacuum and the residue was suspended in
Acknowledgements
◦
50 ml of water. The suspension was stored over night at 4–8 C.
The authors gratefully acknowledge financial support from
the Deutsche Forschungsgemeinschaft within the Sonder-
forschungsbereich 765, the Fonds der Chemischen Industrie (PhD
fellowship for F. P.) and the Alexander von Humboldt Foundation
(Fellowship for S. Y.).
The white precipitate was filtered off, washed with water, dried
under high vacuum and purified by chromatography (CH2Cl2). A
white solid was obtained (448 mg, 30% yield).
1H NMR (CDCl3, 400 MHz): d = 1.34 (quin, 2H), 1.55 (quin,
2H), 1.62 (quin, 2H), 2.19 (t, 2H), 2.29 (s, 3H), 2.82 (t, 2H), 3.72
(ddd, 4H), 3.92 (quin, 1H).
Notes and References
Synthesis of 17-sulf
1 M. P. Bevilacqua and R. M. Nelson, J. Clin. Invest., 1993, 91, 379.
2 D. Vestweber and J. E. Blanks, Physiol. Rev., 1999, 79, 181.
3 K. Ley, D. C. Bullard, M. L. Arbones, R. Bosse, D. Vestweber, T. F.
Tedder and A. L. Beaudet, J. Exp. Med., 1995, 181, 669.
4 W. S. Somers, J. Tang, G. D. Shaw and R. T. Camphausen, Cell, 2000,
103, 467.
5 A. Leppa¨nen, T. Yago, V. I. Otto, R. P. McEver and R. D. Cummings,
J. Biol. Chem., 2003, 278, 26391.
6 F. Li, P. P. Wilkins, S. Crawley, J. Weinstein, R. D. Cummings and R.
P. McEver, J. Biol. Chem., 1996, 271, 3255.
[5-(2-Hydroxy-1-hydroxymethyl-ethyl-carbamoyl)pentyl]-S-thio-
acetate (52 mg, 0.20 mmol) was dissolved in 2 ml of DMF
and 123 mg (0.80 mmol) of SO3·DMF in 1 ml of DMF were
added dropwise. The solution was stirred for 1 h. 3.2 ml of 1 M
NaOH were added and the reaction mixture was stirred for 5 h.
The completeness of the reaction was monitored by TLC. The
product was not isolated but directly reacted with the Au/citrate
nanoparticles.
7 R. M. Nelson, O. Cecconi, W. G. Roberts, A. Aruffo, R. J. Linhardt
and M. P. Bevilacqua, Blood, 1993, 82, 3253.
8 A. Koenig, K. Norgard-Sumnicht, R. Linhardt and A. Varki, J. Clin.
Invest., 1998, 101, 877.
9 M. Mammen, S.-K. Choi and G. M. Whitesides, Angew. Chem., 1998,
110, 2908, Angew. Chem., Int. Ed., 1998, 37, 2754.
10 M.-C. Bowman, T. E. Ballard, C. J. Ackerson, T. N. Tran, D. L.
Feldheim, D. M. Margolis and C. Melander, J. Am. Chem. Soc., 2008,
130, 6896.
11 D. A. Giljohann, D. S. Seferos, W. L. Daniel, M. D. Massich, P. C. Patel
and C. A. Mirkin, Angew. Chem., 2010, 122, 3352, Angew. Chem. Int.
Ed., 2010, 49, 3280.
12 (a) M. De, P. S. Ghosh and V. M. Rotello, Adv. Mater., 2008, 20, 4225;
(b) C. Kim, S. S. Agasti, Z. Zhu, L. Isaacs and V. M. Rotello, Nat.
Chem., 2010, 2, 962.
Synthesis of NP-17-sulf
A solution of 0.20 mmol 17-sulf in 6 ml of DMF/NaOH was
added to 50 ml of a 2.6 nM aqueous solution of Au/citrate and
the mixture was stirred for 72 h. Then the particles were dialysed
against 600 ml of water for three times. After concentration of the
solution to 10 ml a 12.7 nM solution of NP-17-sulf was obtained.
1H NMR (D2O, 400 MHz): d = 1.37 (m), 1.59 (m), 1.66 (m), 2.26
(t), 2.71 (m), 4.09 (m), 4.38 (m).
13 C. M. Niemeyer, Angew. Chem., 2001, 113, 4254, Angew. Chem., Int.
Ed., 2001, 40, 4128.
SPR based competitive selectin-ligand binding assay22
14 K. Riehemann, S. W. Schneider, T. A. Luger, B. Godin, M. Ferrari and
H. Fuchs, Angew. Chem., 2009, 121, 886, Angew. Chem., Int. Ed., 2009,
48, 872.
Selectin-ligand binding was analyzed by surface plasmon reso-
nance (SPR). First, the binding (detected as resonance units) of
This journal is
The Royal Society of Chemistry 2011
Org. Biomol. Chem., 2011, 9, 7448–7456 | 7455
©