Characterization of the tissue light penetration in prostate photodynamic therapy (PDT) is important to plan the arrangement and weighting of light sources so that sufficient light fluence is delivered to the treatment volume. The optical properties (absorption [μ_a], transport scattering [μ_s^′] and effective attenuation [μ_eff] coefficients) of 13 patients with locally recurrent prostate cancer were measured in situ using interstitial isotropic detectors. Measurements were made at 732 nm before and after motexafin lutetium (MLu)–mediated PDT in four quadrants. Optical properties were derived by applying the diffusion theory to the fluence rates measured at several distances (0.5–5 cm) from a point source. μ_a and μ_s^′ varied between 0.07 and 1.62 cm⁻¹ (mean 0.37 ± 0.24 cm⁻¹) and 1.1 and 44 cm⁻¹ (mean 14 ± 11 cm⁻¹), respectively. μ_a was proportional to the concentration of MLu measured by an ex vivo fluorescence assay. We have observed, on average, a reduction of the MLu concentration after PDT, presumably due to the PDT consumption of MLu. μ_eff varied between 0.91 and 6.7 cm⁻¹ (mean 2.9 ± 0.7 cm⁻¹), corresponding to an optical penetration depth (δ = 1/μ_eff) of 0.1–1.1 cm (mean 0.4 ± 0.1 cm). The mean penetration depth at 732 nm in human prostate is at least two times smaller than that found in normal canine prostates, which can be explained by a four times increase of the mean value of μ_s^′ in human prostates. The mean light fluence rate per unit source strength at 0.5 cm from a point source was 1.5 ± 1.1 cm⁻², excluding situations when bleeding occurs. The total number of measurements was N = 121 for all mean quantities listed above. This study showed significant inter- and intraprostatic differences in the optical properties, suggesting that a real-time dosimetry measurement and feedback system for monitoring light fluences during treatment should be considered for future PDT studies.