Scheme 1. Reagents and conditions: (a) HCl (32% water), HCHO (37% in water), 0 °C → rt, 8 h; (b) N-substituted piperazine, (Me₂CH)₂NEt, CHCl₃, rt, 1 day; (c) propargylamine, (Me₂CH)₂NEt, CHCl₃, rt, 1 day.

Scheme 2. Reagents and conditions: (a) Boc₂O, MeOH, 0 °C → rt, 1 day; (b) propargyl bromide, (Me₂CH)₂NEt, 0 °C → rt, 1 day, (c) F₃CCOOH, 0 °C → rt, overnight; (d) 5-chloromethyl-8-quinoline 2, CHCl₃, (Me₂CH)₂NEt, rt, 1 day.

Scheme 3. Reagents and conditions: (a) propargyl bromide, NaHCO₃, DMSO, rt; (b) TrCl, DMF, rt; (c) 5-chloromethyl-8-quinoline 2, CHCl₃, (Me₂CH)₂NEt, rt; (d) CF₃COOH, rt.

Figure 1. Spectrophotometric detection of Fe(III) complexation with HLA20: (a) spectrum of 0.8 mM compound 9 (HLA20) in Tris buffer pH 7.4; (b) spectrum of 0.8 mM compound 9 (HLA20) in Tris buffer pH 7.4 after the addition of 0.15 mM Fe₂(SO₄)₃.

Figure 2. Spectrophotometric detection of the formation of Fe(III)–9 (HLA20) complex from Cu(II)–9 (HLA20) complex: (a) spectrum of 0.6 mM compound 9 (HLA20) in Tris buffer pH 7.4; (b) spectrum of (a) after the addition of 0.2 mM Cu(SO₄)₂. (c) spectrum of (b) after the addition of 0.1 mM Fe₂(SO₄)₃.

Figure 3. Change in absorption at λ_max = 590 nm with [Fe(III)]/[HLA20] molar ratio in Tris buffer (pH 7.4).

Figure 4. Positive ion electrospray ionization mass spectrum of a solution of iron(III) sulfate and compound 9 (HLA20).

Figure 5. Dequenching of CA–Fe complexes by chelators. Iron precomplexed calcein (CA–Fe) solutions of 1 μM in Hepes 20 mM buffered saline (pH 7.4 = HBS) were incubated with various concentrations of chelators at rt for 1 h. The graph shows the original fluorescence data given in arbitrary units (a.u.) against chelator concentration.

Figure 6. Dequenching of CA/iron complexes in K562 cells by extracellularly added the test chelators. CA-loaded K562 cells were treated with various chelators (5 μM concentration) at the time indicated (arrow). Scale, arbitrary fluorescence units (a.u.) against time (s).

Figure 7. Relative permeability of iron chelators in K562 cells. Values were mean ± SEM (n = 6), given relative to those obtained with SIH, for which 100 represents an apparent rate constant of 0.0069 s⁻¹ (equivalent to a t_1/2 of ingress of 145 s for a 5 μM chelator solution).

Figure 8. Effects of apomorphine and iron chelators 2, 3, and 9 and desferal on 6-OHDA-toxicity in differentiated P19 cells. Differentiated P19 cells were treated for 6 h with 50 μM 6-hydroxydopamine (6OHDA) in full medium containing either 5 μM of indicated chelators or 1 μM apomorphine. The cell viability was measured with Alamar blue assay 4 h later. % Protection = the % activity relative to the system treated with no 6OHDA. Data are means ± SEM, n = 3. p < 0.05, compared with control.

Figure 9. Typical EPR spectra of DMPO-OH. Derived from hydroxyl radicals generated by photolysis of a 0.6% H₂O₂ pH 7.4 PBS buffer containing 0.1 M DMPO, with different irradiation time: A, B, and C: 40 and 46 min; D 40 min; E 46 min. (A) Control. (B) 0.1 mM 9 (HLA20). (C) 0.5 mM 9 (HLA20). (D) 1.0 mM 9 (HLA20). (E) 1.0 mM 9 (HLA20).

Figure 10. Concentration–response curves for inhibition at room temperature and pH = 7.4 (PBS buffer containing 0.1 M DMPO) of EPR DMPO-OH peaks by HLA20.

Inhibitory properties on lipid peroxidation (LPO) and in vitro monoamine oxidase (MAO) of the new chelators