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ligand modification influence the wavelength of activation of platinum(IV) diazido complexes. This is in agreement with previous DFT calculations performed on complexes 56 and 57 which exhibited bands at longer wavelengths compared to complex 40.41 _{The trapping of the} _N

3 radicals from photo-irradiated

complexes 40, 44 and 57-59 suggested the release of the azide ligands in azidyl (N3) radical form is a common photo-decomposition pathway for platinum(IV)

diazido complexes.

3.4.4.2 Gamma ray irradiation

Cis-platin was first reported to be a radio-sensitiser by Zák and Drobník.68 Second and third generation platinum anticancer complexes are also efficient radio- sensitisers.69,70 However, this appears to be the first report on the activation of complex 40 with a gamma-ray irradiation source. Initial gamma-ray irradiation of complex 40 in the presence of spin trap, DMPO, did not generate EPR spectra suitable for full characterisation or qualitative analysis. It is most probably a decay product from the initial trapping of the N3 radicals by DMPO.

Employing the phosphorus spin trap, DEPMPO was more efficient in allowing the resultant EPR spectra to be interpreted in terms of N3 radical trapping. The

detection of the DEPMPO-N3 spin adduct suggests gamma-ray irradiation is

effective in photo-activating complex 40. Two probable activation pathways of complex 40 with the 137Cs gamma-ray irradiation source have potential to occur. Firstly, the 137_{Cs gamma-ray irradiation source has potential to ionise the solvent,}

140 H2O, leading to the generation of aqueous free radicals, hydrogen ion and low

amounts of H2 and H2O2 (Equation 3.3). Previous studies by Khan reported on the

one-electron reduction of trans-[PtIV(NH3)4(OH)2]2+ by the aquated electron

generated from pulse radiolysis, leading to the formation of a trans- [PtIII(NH3)4(OH)2]+ species. This platinum(III) species underwent additional

reaction with the hydroxyl radicals generated from the pulse radiolysis of water, forming trans-[Pt(NH3)4(H2O)2]3+.71 Additionally, free ammonia was also

detected in these reactions.

In this work, gamma-ray irradiations were performed in water, therefore ionisation of the solvent as shown in Equation 3.3 is possible. Through the hydrated electron, a one-electron reduction of the PtIV metal centre of complex 40 could occur, generating a platinum(III) intermediate. Although it is not fully understood, it is proposed the reactive nature of this platinum(III) transient species may result in the loss of the azide ligand in azidyl radical form, which is subsequently trapped by DEPMPO. Alternatively, the 137_{Cs, parent nuclide spontaneously decays via}

beta decay to the daughter nuclide, 137Ba and emits a β- particle, an antineutrino (ῡe) and Qβ- energy (Equation 3.4). The β- particle corresponds to an electron (e-),

where its presence satisfies the energy conservation.72

Therefore, the reduction of the PtIV metal centre may also be initiated from this electron in a similar pathway as described above. One-electron reduction of cis- platin has also been reported with the hydrated electron, forming a platinum(I) type species.73 Free ammonia from trans-[PtIV(NH3)4(OH)2]2+ was also detected in the

141 study by Khan et al.. Therefore in this work, the lack of cytotoxicity in DLD-1 cells from gamma-ray irradiated complex 40 may be due to the loss of coordinated pyridine. The loss of carrier ligands from platinum anticancer complexes has been associated with reduced cytotoxicity.74-76

Interestingly, the detection of the DEPMPO-N3 spin adduct from complex 40 was

obtained using clinically relevant gamma-ray irradiation doses. To date, the maximum dosage applied to patients is 45 Gy, given in aliquots of 1.8 – 2 Gy over a period of 5-8 weeks.37 However, despite the detection of the DEPMPO-N3 spin

adduct, it should be noted that the 137Cs gamma-ray irradiation source emitting a photon with energy of ca. 662 keV is more destructive towards complex 40, compared to visible light with a power outage of ca. 64 mW cm-2_{. Therefore, the}

detection of the DEPMPO-N3 spin adduct has potential to be due to complete

molecular fragmentation of complex 40 irradiated with 137Cs gamma-ray irradiation source. Additional UV-visible and NMR spectroscopy studies of complex 40 irradiated with gamma-rays are required to fully comprehend these preliminary results and elucidate the mechanism of action.

3.5 Conclusion

In this Chapter, photo-activation of complex 40 with blue light gave rise to the release of the azide ligand in azidyl (N3) radical form, which was detected by spin

trapping EPR spectroscopy using a variety of nitrone spin traps namely: DMPO, 4-POBN and DEPMPO. Varying the spin trap led to differences in efficiency and life-times of the resultant spin adducts. The phosphorus spin trap DEPMPO appeared to exhibit the fastest rate towards N3 radicals. In contrast, the spin trap,

142 4-POBN displayed the lowest level of N3 radical trapping, this was attributed to

steric hindrance due to the presence of the 4-pyridyl and t-butyl groups present in 4-POBN.

The first generation cyclic nitrone spin trap, DMPO generated the quartet of triplets spectrum, indicative of DMPO-N3 spin adduct formation. However, with a

known rate slower than N3 radical dimerisation, a portion of formed N3 radicals

are believed to undergo radical dimerisation. The distribution of spin density on the atoms involved in the hyperfine coupling was correlated with the lifetime of the spin adducts under investigation.

Interestingly, Brownian motion of the formed N3 radicals appears to account for

the observed increase in the amount of spin adduct formed upon changing the solvent from PBS/H2O to PBS/D2O. In contrast, the reduction in the DMPO-N3

spin adduct formed in RPMI-1640 was attributed to the various components in cell culture medium which have potential to behave as azidyl radical quenchers. Furthermore, the trapping of N3 radicals from related photo-activatable

platinum(IV) diazido complexes, trans,trans,trans-[Pt(N3)2(OH)2(MA)(py)] (44,

MA = methylamine), trans,trans,trans-[Pt(N3)2(OH)(SAD)(py)2] (56, SAD =

succinate), trans,trans,trans-[Pt(N3)2(OH)(ethyl-methyl-SAD)(py)2] (57) and

trans,trans,trans-[Pt(N3)2(OH)(N-MI)(py)2] (58, N-MI = N-methylisatoate)

suggested the release of the azide ligand(s) as a common photo-decomposition pathway of platinum(IV) diazido complexes. Finally, it was shown that complex

40 could be activated by gamma-rays from a 137Cs gamma radiation source at doses