Abstract: OBJECTIVE To prepare a low toxicity nattokinase(NK) injection. METHODS By adding the γ-polyglutamic acid(γ-PGA) to the NK solution, the NK-γ-PGA complex injection was prepared. Taking survival time, survival state and the tail state of the surviving mouse at 48 h after a single tail intravenous injection as toxicity evaluation indexes, the acute toxicity test was used to study the effects of pH and γ-PGA concentration on the complexation of NK and γ-PGA separately. The in vitro thrombolytic test was used to analyze the reason of the toxicity reduction of NK-γ-PGA complex injection. RESULTS When the NK-γ-PGA complex injection(NK 10 kU·mL^-1, γ-PGA 0.924 mg·mL^-1, pH 6.0) was administrated by a single tail intravenous injection at a dose of 120 kU·kg^-1, the death rate of mice was 0, and the mouse tail had no swelling and necrosis, which indicated that when the pH was 6.0 and the concentration of γ-PGA was 0.924 mg·mL^-1, the complexation of NK and γ-PGA was the best. The results of in vitro thrombolytic test showed that the addition of γ-PGA did not reduce the activity of the NK, but had a sustained-release effect to reduce drug toxicity. CONCLUSION The NK-γ-PGA complex injection(NK 10 kU·mL^-1, γ-PGA 0.924 mg·mL^-1, pH 6.0) prepared in this study achieved the goal of reducing NK injection toxicity without changing NK activity.