*3.13. Acute Systemic Toxicity Assay*

All animal procedures were approved by the ethical committee of animal research in the Ocean University of China, and complied with the requirements of the National Act on the use of experimental animals (China). The acute systemic toxicity of tilapia collagen was investigated according to ISO 10993-11:2009. The collagen samples were cut into 1×1 cm sizes with 0.5-mm thickness and sterilized by 60Co before using. The leach liquor was prepared at a ratio of three cm2/mL collagen into saline between the total surface area of the materials for 72 h at 37 ◦C. Then, 20 Kunming (KM) mice (Licensed ID: SCXK 2014-0007) were divided into four groups randomly. Three experimental groups were intraperitoneally injected the leach liquor of ASC, PSC, and PC with a dose of 50 mL/kg respectively, and the blank control group was injected saline with a dose of 50 mL. The weight of mouse was weighed and recorded immediately after injection and at four hours, 24 h, 48 h, and 72 h after injection. The criteria that were used to assess acute systemic toxicity are shown in Table 2.


**Table 2.** The criteria used to assess acute systemic toxicity.
