Measurements of short distances between trityl spin labels with CW EPR, DQC and PELDOR

Trityl based spin labels are emerging as a complement to nitroxides in nanometer distance measurements using EPR methods. The narrow spectral width of the trityl radicals prompts us to ask the question at which distance between these spin centers, the pseudo-secular part of the dipolar coupling and spin density delocalization have to be taken into account. For this, two trityl-trityl and one trityl-nitroxide model compounds were synthesized with well-defined interspin distances. Continuous wave (CW) EPR, double quantum coherence (DQC) and pulsed electron-electron double resonance (PELDOR) spectra were acquired from these compounds at commercial X-band frequencies. The data analysis shows that two of the compounds, with distances of up to 25 Å, fall into the strong coupling regime and that precise distances can only be obtained if both the spin density delocalization and the pseudo-secular part of the dipolar coupling are included in the analysis.