Mental health is a significant public concern. Given that individuals spend extended periods indoors, understanding the psychophysiological health effects of indoor materials on well-being is crucial. This study compared the effects of wooden and resin plaster rooms on psychophysiological well-being. Well-being was assessed using heart rate variability measurements and an affective well-being questionnaire. Subjective perceptions were also evaluated using a semantic differential questionnaire. The results indicated higher heart rate variability in the wooden room compared to the reference room. Participants breathed about one breath per minute more slowly in the wooden room, with a negative correlation between heart rate variability and respiratory rate. Positive affect was elevated, and negative affect was reduced in the wooden room, which was also perceived more favorably in sensory evaluations. The findings suggest that wooden interiors are preferred over artificial materials, enhancing both physiological and psychological well-being.