Near-infrared (NIR) spectroscopy was used to measure blood oxygen saturation (SO2) in vivo, in normal rat brain and in subcutaneously-implanted rat 9L gliosarcoma. Changes in cranial and tumor blood SO2 were measured during lethal pentobarbital overdose. After sacrifice, SO2 of cranial blood fell rapidly to a mean of 5.0% of the pre-sacrifice values, whereas SO2 of tumor blood stabilized at a mean of 72.4% of the pre-sacrifice values. This suggests that oxygen consumption by tumor is very low compared to brain. Cranial blood had a higher SO2 than tumor blood before sacrifice (p = 0.03), and a lower SO2 after sacrifice (p = 0.02). The magnitude of the change in SO2 after sacrifice was greater in normal brain than in tumor (p = 0.02), showing that brain tissue uses a greater proportion of the oxygen in ischemic blood than does tumor tissue. To determine the effect of carmustine (BCNU) treatment on tumor and cranial blood SO2, we compared BCNU-treated rats with sham-treated rats. Continuous NIR measurements before and immediately following treatment (ie. over 30-60 min) showed that tumor blood SO2 tended to increase after BCNU treatment, whereas SO2 tended to decrease following sham-treatment. The difference in SO2 between treated and control tumors was significant at 60 min (p = 0.02). Thus BCNU treatment can potentially result in immediate increases in tumor oxygenation. The increase in treated tumor blood SO2 occurred despite the fact that there was no change in cranial blood SO2 even at day 4 following treatment. Tumor blood SO2 was inversely correlated with tumor size (p = 0.001), confirming that blood is more poorly oxygenated in large tumors.