The antioxidative and antibacterial properties of low-molecular-weight secondary metabolite subfractions (ex-LMS) from cultures of Pycnoporus sanguineus cultivated under different temperature conditions (25°C [ex-LMSa] and 30°C [ex-LMSb]) were assessed. The antioxidative properties were studied using chemiluminometric measurement, an ABTS assay, and a DPPH reduction rate assay with Trolox and ascorbic acid as the control. The values noted for the ex-LMSb were significantly higher than those for ex-LMSa: 97%, 52%, and 31% for chemiluminometric measurement, the ABTS assay, and the DPPH assay, respectively, at a concentration of 50 µg/mL. Half-maximal effective concentrations reached 4.17 µg/mL for chemiluminometric measurement, 47.25 µg/mL for the ABTS assay, and 51.46 µg/mL for DPPH assay. Toxicity tests against Vibrio fischeri yielded 99.8% for ex-LMSa and 99.85% for ex-LMSb. Antibacterial activity toward Staphylococcus aureus was observed in the ex-LMSb fractions (inhibition zone, 23.5 mm; minimum inhibitory concentration, 0.12 mg/mL). Scanning electron microscopy images exhibited severe disruption of the bacterial cells treated with ex-LMSb compared with the control. The results obtained suggest that the extracellular fluid isolated from P. sanguineus-submerged cultures might be a good source of antioxidative and antibacterial compounds. In addition, the increase in the culture temperature evidently enhanced the bioactive properties of the preparation.