Car☐yverdazyls: Water-soluble organic free radicals

JOURNAL

OF MAGNETIC

RESONANCE

99, 387-390 ( 1992)

Carboxyverdazyls: Water-Soluble Organic Free Radicals ARMIN J. MAYR, MICHAEL P. EASTMAN, CYNTHIA J. HARTZELL, DING DONG, AND CARLEE MCCLELLAN Department of Chemistry, Northern Arizona University, Flagstag Arizona 8601 l-5698 Received October 3 1, 199 1; revised January 2 1, I992

The advent of magnetic resonance imaging created a need for water-soluble paramagnetic materials as contrast agents. Complexes of Gd(III), Mn(II), and Fe( III) are commonly used for this purpose, since they have large magnetic moments and relax nuclear spins through electron-nuclear dipole interactions ( 1) . However, in vivo dissociation of these complexes may cause medical problems due to the toxicity of the metal ions and free ligand. The use of water-soluble nitroxide radicals in MRI has been explored to probe their potential for special applications (2). We report the synthesis and properties of new water-soluble verdazyls. These compounds are stable S = 1 organic radicals based on the tetrazine framework and may prove to be useful MRI contrast agents. The synthesis of carboxy-substituted verdazyls follows published procedures (3). Diazotization of aldehyde-phenylhydrazones 1 leads to deep red formazans 2, which, on condensation with formaldehyde, produce the dark green carboxyverdazyls 3a-3c. The carboxyverdazyls dissolve in dilute aqueous base to green radical anion (3’) solutions, whereas with acids the purple, water-soluble, diamagnetic cations 3” are formed (see Scheme 1) . A mechanism for the pH-dependent redox process involving 1,3,5-triphenylverdazyl has been proposed (3, 4). The uv/vis spectrum of 3a in water at pH 10 is very similar to the spectrum of triphenylverdazyl in nonpolar solvents (3) with respect to X,,, and t. A bathochromic shift of the absorption at approximately 7 10 nm is observed at verdazyl concentrations > 10e4 M and may indicate micellular aggregation of the surfactant-like verdazyl anions 3’ (see Fig. 1) . This shift is reversed in the presence of p- and y-cyclodextrin, suggesting a host-guest interaction of the phenyl substituents of the verdazyl with the cyclodexbins, thus disrupting the aggregates. EPR spectra of aqueous verdazyl solutions show nine lines from equivalent interaction of the unpaired electron with four nitrogen atoms (aN = 6 G). The similarity of these spectra with those of verdazyls in nonaqueous solutions suggests little direct interaction of water molecules with the radical centered at the tetrazine ring. As the verdazyl concentration increases to > 10e3 M, the emergence of a single line component is observed (Fig. 2a). This behavior is consistent with aggregated radicals undergoing Heisenberg spin exchange. Dispersion of the radicals by @-and y-cyclodextrin causes the single line component to disappear (Fig. 2b). 13C NMR provides detailed information on the structure of the cyclodextrin-nitroxide host-guest complex (5); the same experiments were carried out with the ver387

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388

NOTES

3 a R=COOH, R’=H 3 b R=H, R=COOH 3 c R=R’=COOH

SCHEME 1

720

1% [3al

FIG. 1. Plot of A,,,,, at approximately 7 10 nm, vs log [3a]

389

NOTES 9 a

1 1 Oi

0

1

2

3

4 FIELD (mT)

5

6

1

2

3

4 FIELD (ml)

5

6

i

71

04 0

7

FIG. 2. (a) x-band EPR spectrum of a 10e3 M solution of 3a in water at T = 297 K; (b) with 10m3Mycyclodextrin, conditions as in (a).

C6

c4

C3C2C5

Cl

A

l’~~‘1’~“l1~‘~1’~~‘l~~‘~~~‘~‘l’~~~~~~’~~’~~’~”’~~‘~~‘l~‘~~~~”’l~‘120

110

FIG.

100

90

80

70

60 PPM

3. (A) “C NMR spectrum of P-cyclodextrin ( IO-* M) in D,O; (B) with 10m4M3a.

390

NOTES

0

I

2

3

4

5

radical concentration,

6 mMol

FIG. 4. Plot of 1/ T, of water protons (5% H20 in DzO) vs concentration of 3a (filled diamonds) and DTBN (open diamonds). T, relaxation times were measured at 200 MHz and T = 303 K.

dazyls. Figure 3 shows the 13C NMR spectrum of y-cyclodextrin in the presence of 3a ( 10P4 M) in D20. Significant line broadening of the C3, C5, and, to some extent, C6 peaks has occurred, in agreement with the results of nitroxides interacting with ycyclodextrin (5). However, unlike with the nitroxides, changes in T2 were not accompanied by changes in T, . We have determined the effect of the water-soluble verdazyls on the relaxivity (Tl-‘Jobs of water protons at 200 MHz, using 5% H20/Dz0 mixtures and compared the result with that of di-tert-butylnitroxide (DTBN). Figure 4 shows the plot of for the verdazyl3a and DTBN. The slopes of ( Ti ’ jobs vs free radical concentration the straight lines ( TI-l),,,ag are a measure of the magnetic relaxation of the water protons induced by the paramagnetic species. The plot is linear for DTBN over a wide concentration range and apparently nonlinear for 3a; this effect may be attributed to verdazyl aggregation. At a radical concentration of 6.5 X lop4 M, the ( T, -‘)Obs value for DTBN (0.19 s-l) is similar to that for 3a (0.15 SK’), implying an outer sphere relaxation mechanism in both cases. ACKNOWLEDGMENTS This research was supported by MBRS-NIH and NSF-REU grants to Northern Arizona University, Northern Arizona University’s Organized Research Fund, and the Petroleum Research Fund (C.J.H.). Helpful discussions with J. R. Brainard of the Los Alamos National Laboratory are acknowledged. REFERENCES 1.

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Mu,q