SSRT and fatigue life tests of SUS301 austenitic stainless steel were performed to examine the effect of hydrogen on the mechanical properties. Ni content of SUS301, 6.00–8.00 mass%, is lower than that of SUS304 in JIS standard for austenitic stainless steels. In the case of SSRT tests, specimens with and without hydrogen charging were tested in laboratory air at room temperature (R.T.), −45 °C, and −80 °C. The 0.2% offset yield strength (Ys) of the hydrogen charged specimens was less than 300 MPa in the tested temperature range. The tensile strength (Ts) and total elongation (El) of hydrogen charged specimens decreased remarkably. With decreasing testing temperature, fracture surface facet of the hydrogen charged specimens became dominant. Therefore, the effect of hydrogen on the tensile properties of SUS301 is supposed to be large. Specimens with and without hydrogen charging were fatigued in laboratory air at R.T., and specimens without hydrogen charging were fatigued in 100 MPa hydrogen gas atmosphere at R.T. Number of cycles (Nf) at finite fatigue life region of the hydrogen charged specimens and of the specimens tested in hydrogen gas were two orders shorter than that of the specimens tested in air. However, the finite fatigue life region of the hydrogen charged specimens and the specimens tested in hydrogen gas showed a different profile. Additionally, ferrite equivalents of all fatigue tested specimens and fatigued fracture surface morphology suggested the fatigue fracture mechanism between the hydrogen charged specimens tested in air and the non-charged specimens tested in 100 MPa hydrogen gas seems to be different. Therefore, further investigations are required to clear this difference.