The safety of pufferfish (Lagocephalus gloveri) liver oil and the contents of some nutritional components were examined to obtain important information on their use as high valued functional foods. Pufferfish liver oil was extracted by the hot-water method using $1\%$ NaOH solution to remove toxic compounds, and then purified using a general purifying method of fish oil. Any extraordinary clinical symptoms were not observed from all groups administrated with pufferfish liver oil throughout the test period. None of the rats died when administrated the highest concentration of 10 mL/kg of the pufferfish liver oil. Vitamin A content was 114.2 ppm, as a retinal equivalent in the oil extracted using hot-water, but the content was higher (169.3 ppm) in oil extracted using n-hexane. Vitamin D and E were not detected in ppm in oil extracted using hot-water. Vitamin D in the pufferfish liver oil extracted using n-hexane was also undetected, but vitamin E was at 32.5 ppm. Among the 18 minerals detected, the sodium content was the highest at 253.5 ppm, followed by 13.9 ppm ofpotassium, 1.5 ppm of calcium, 0.2 ppm of magnesium, and other trace minerals. The contents of EPA and DHA were $0.8\%;and;14.8\%$, respectively, in the pufferfish liver oil extracted using hot-water. Considering these results, there is potential that pufferfish liver oil could be used as a functional food.