The spectrum from Complex I was always weak, and it was only observed at X-band frequencies; its intensity was too low to produce a spectrum at S-band frequencies.

However, it dominated the weak EPR spectra obtained with both the EGCG and GA in the slightly acidic pH range. The contributions from both Complexes II and III increased with increasing pH above pH 7, and above pH 12, only complex III was detected in the solutions which contained more than 2-fold excess of the poyphenols. At this high pH, the spectrum of a Cu(II) glycerol complex was observed from solutions with lower polyphenol concentrations. Thus Complex III might correspond to mixed polyphenol/glycerol complexes of Cu(II), but the formation of a complex between Cu(II) and EGCG with a similar spectrum to that of Complex III in Fig. 3d was observed using pure H2O as the solvent (i.e. without glycerol). All of the spectra from the Raf activation Cu(II) complexes are complicated by the presence of appreciable linewidth anisotropy; their analysis to produce estimates of rotational correlation Fulvestrant times is described below (after consideration of the frozen solution spectra). Representative frozen solution spectra from the Cu(II)/EGCG reaction at X-band and S-band frequencies are shown in Fig. 6 and Fig. 7

for a Cu:EGCG ratio of 1:5. The g// region in Fig. 6 is expanded to provide better clarity, since this represents the part of the spectrum where different complexes (indicated by stick

diagrams) can be discerned. The full range of X-band spectra is available as supplementary information (Figures S5-8). Very similar results were observed with the Cu:GA system and these are also available as supplementary information (Figures S9–12). The spectrum in Fig. 6a corresponds to the uncomplexed [Cu(H2O)6]2 + ion, and that in Fig. 6b belongs primarily to Complex I. Increasing the pH gave results that correspond to mixtures of all three complexes in different ratios (Fig. 6c and d) and at very high pH, Complex III was the major species detected (Fig. 6e). The “pepper” function in the Easyspin software package was used to simulate the spectra of the three mononuclear Cu-EGCG Lck complexes (Fig. 8), and their parameters are summarised in Table 1 along with the corresponding values derived by simulation of the Cu(II)/GA spectra. The various Cu-complexes are distinguished by a progressive shift in g// to lower values and A// to higher values from the uncomplexed ion through Complex I to Complex III (Table 1). Table 1 also includes the parameters for the Cu-glycerol complex, which can be formed at very high pH. However, since its g- and A-values differ significantly from those of Complex III, it can be concluded that polyphenol complexes dominate the spectra in high pH solutions with a Cu:EGCG ratio of 1:5.