Nattokinase activity (NK), biogenic amine content and sensory properties of natto are of great significance to consumers, which are affected by strains and fermentation methods. In this study, changes in the pH, biogenic amine and free amino nitrogen (FAN) contents, NK and protease activities, and sensory characteristics of natto prepared using Bacillus subtilis GUTU09 combined with different strains (Lactobacillus, Bifidobacterium and Mucor) and fermentation methods were investigated. The combination of two strains showed the best fermentation performance among all samples. The NK and protease activity and FAN content in double-strain fermentation increased by 10.33 FU/g, 88.78 U/g, and 2.34 g/kg, respectively, compared with those in single-strain fermentation. Sensory evaluation demonstrated that mixed fermentation primarily affected the sensory acceptance. This method also reduced the contents of various biogenic amines in natto compared with single-strain fermentation. Tyramine, cadaverine, spermine, and spermidine were significantly reduced, whereas histamine was slightly increased. The total biogenic amines decreased from 390.76 mg/kg to a minimum of 16.16 mg/kg. Some Mucor strains also reduced the contents of various biogenic amines. In the dual-bacteria fermentation of Mucor and GUTU09, co-fermentation has advantages over stage-fermentation, with higher NK and protease activity and higher sensory scores. Correlation analysis showed that the formation and accumulation of some biogenic amines in natto prepared using different microbial combinations were related to NK activity and pH. All these results showed that the quality of natto was improved by mixed fermentation and suitable fermentation methods, which laid a foundation for its potential industrial application.

纳豆激酶活性（NK）、生物胺含量和纳豆的感官特性对消费者具有重要意义，它们受菌株和发酵方法的影响。本研究中，采用枯草芽孢杆菌GUTU09 与不同菌株（乳杆菌、双歧杆菌和毛霉) 和发酵方法进行了研究。两种菌株的组合在所有样品中表现出最好的发酵性能。与单菌株发酵相比，双菌株发酵NK、蛋白酶活性和FAN含量分别增加10.33 FU/g、88.78 U/g和2.34 g/kg。感官评价表明混合发酵主要影响感官接受度。与单菌株发酵相比，该方法还降低了纳豆中各种生物胺的含量。酪胺、尸胺、精胺和亚精胺显着减少，而组胺略有增加。总生物胺从 390.76 mg/kg 降至最低 16.16 mg/kg。一些毛霉菌株还降低了各种生物胺的含量。在毛霉和GUTU09双菌发酵中，共发酵优于阶段发酵，NK和蛋白酶活性更高，感官评分更高。相关性分析表明，使用不同微生物组合制备的纳豆中某些生物胺的形成和积累与NK活性和pH值有关。这些结果表明，混合发酵和适宜的发酵方法提高了纳豆的品质，为其潜在的工业应用奠定了基础。