Temperature-induced hemoglobin dynamics: oxygenation-dependent evolution of hemoglobin spectral response in the near-infrared

Significance: Monitoring thermally induced optical changes can guide thermal therapies, but these signals depend strongly on hemoglobin oxygenation, leading to different spectral responses under ex vivo and in vivo conditions. Clarifying these effects is essential for interpreting absorption measurements during treatment. Aim: We characterized near-infrared absorption spectra of hemoglobin during heating under controlled oxygenated and deoxygenated conditions and related the signatures to hemoglobin chemistry. Approach: Using time-domain diffuse optical spectroscopy, we measured 680- to 1110-nm absorption spectra of blood phantoms containing oxyhemoglobin (oxyHb) or deoxyhemoglobin (deoxyHb) during controlled heating. Multi-concentration measurements yielded extinction coefficients for native and thermally denatured species. Results: Heating oxyHb caused a broad absorption rise consistent with autoxidation of ferrous (Fe2+) heme to ferric (Fe3+) forms, similar to methemoglobin and hemichromes. Heated deoxyHb showed distinct peaks at 774 and 874 nm that disappeared upon re-oxygenation, after which spectra converged to treated oxyHb. The oxygen-dependent kinetics of the 774/874 nm bands support a ferrous-denatured intermediate that converts to ferric hemichromes when oxygen becomes available. Conclusions: Thermal denaturation of hemoglobin follows distinct oxygenation-dependent spectroscopic pathways. The extracted extinction coefficients provide essential inputs for optical models of laser-based ablation and improve diffuse optical monitoring of thermal therapies.