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1 一种新型功能性海带豆酱的制备工艺研究杨星星 1, 崔迎雪 1, 武凌宇 1, 王亚囡 1, 刘远平 2, 李钰金 2 1, 魏玉西 (1., ; 2., ) 摘要 : 为了探究一种新型纳豆菌发酵食品的最佳工艺, 利用纳豆菌发酵大豆和海带, 以发酵物中游离氨基酸态氮含量和纳豆激酶活性为评价指标, 通过单因素试验和正交试验确定最佳发酵条件结果表明, 发酵温度 42 发酵时间 54 h 接种量 1% 料水比(m/v)1.. 4 配料比( 大豆..海带 ) 时发酵产物中游离氨基酸态氮含量最高 (0.041%); 发酵温度 40 发酵时间 48 h 接种量 3% 料水比(m/V) 1.. 2, 配料比 ( 大豆..海带 ) 时发酵产物中纳豆激酶活性最高 ( 可达 IU/g) 关键词 : 海带 ; 大豆 ; 纳豆菌 ; 发酵 ; 游离氨基酸态氮 ; 纳豆激酶活性中图分类号 : TS254 文献标识码 : A 文章编号 : (2017) DOI: /hykx ,, 2000 [1-2] (Bacillus natto), 100, [3], [4],, [5-6], (Laminaria japonica Aresch), (Phaeophyta) (Laminarales) (Laminariaceae) (Laminaria),,, [7],,,,,,,,, [8],,,,,, 1 材料与方法 1.1 材料, ; ; (Bacillus natto), 1.2 仪器与设备 HC-2518 ; HH4 ; 721G ; 88-1 ; Sartorius PB-10 ph ( : ; : : ( )(2016ZDJS06A01) [Foundation: This study was financially supported by Shandong province key R & D projects (Key technologies)(no.2016zdjs06a01)] : (1994-),,,, :, : , @qq.com; (1964-),,,, :, : , yuxiw729@163.com; (1966-),,, :, r.lyj@163.com 48 / 2017 / 41 / 6

2 ) ; - ; ( ) 1.3 工艺流程 1.4 方法 1.4.1, 12 h ;,, 60, (free ammonia nitrogen, FAN) [9], X : ( V1 V0) c X (%) N m 20 /100, c: (mol/l); m: (g); V 1 : NaOH (ml); V 2 : NaOH (ml); N: (natto kinase, NK) Folin- [10-14], : V X ( A样 A对 ) K N T, A : OD 680nm ; A : OD 680nm ; K: 1 ( g); V: (ml); T: (min); N: h, Folin-,, g (, ) 2.5 g (, ), 22.5 ml, min, 1% 2% 3% 4% 5%, h ( ) 5 g 2.5 g, 15 ml(1.. 2) 22.5 ml(1.. 3) 30 ml(1.. 4) 37.5 ml(1.. 5), min, 2%, h g 2.5 g, 22.5 ml, min, 2%, h ( ) , 1 5 g 10 g (1.. 2); 2 5 g 15 g (1.. 3); 3 5 g 2.5 g (2.. 1); 4 5 g 1.67 g (3.. 1); 5 5 g 5g (1.. 1) 1.. 3, ml, min, 2%, h g 2.5 g, 22.5 ml, min, 2%, h 1.4.5, L 18 (3 7 ) 1.5 数据处理 SPSS 结果与讨论 2.1 单因素实验 2.1.1,,,,,,,,, Marine Sciences / Vol. 41, No. 6 /

3 [15-16],, 1 ;,,, [17],, 3 1 Fig. 1 Effect of inoculum quantity on free amino nitrogen concentration and nattokinase activity 1, 48 h, 2% (0.025%), 2%, ;, 2% ( IU) Fig. 2 Effect of solid-to-water ratio on free amino nitrogen concentration and nattokinase activity 2, 1.. 3, 0.038% IU;, 2.1.3,,, 3 Fig. 3 Effect of fermentation time on free amino nitrogen concentration and nattokinase activity 60 h, 48 h (0.036%),, 48 h ( IU) 2.1.4,,,,,, ;,,,,, 4 4, 3.. 1, (0.032%), (745.24IU) 2.1.5, [18], 5 50 / 2017 / 41 / 6

4 4 Fig. 4 Effect of mixing ratio on free amino nitrogen concentration and nattokinase activity 5 Fig. 5 Effect of fermentation temperature on free amino nitrogen concentration and nattokinase activity 5,,, 42, 0.032% IU 42,, 2.2 正交试验结果, R, C>B>A>D>E,,, k 1 k 2 k 3,, A 3 B 2 C 1 D 1 E 3, : h 3% 1.. 2, 3.. 1, IU/g 表 1 因素与水平排列表 Tab. 1 Factors and levels A (%) B (h) C ( ) D (m/v) E (m/m) 表 2 以纳豆激酶活力为评价指标的正交试验方案及结果 Tab. 2 Orthogonal array design with experimental values of nattokinase activity as evaluation item A (%) B (h) C ( ) D (m/v) E (m/m) (IU) Marine Sciences / Vol. 41, No. 6 /

5 A (%) B (h) C ( ) D (m/v) E (m/m) (IU) k k k R , R, C>A>E>B=D,, ; k 1 k 2 k 3, A 1 B 3 C 1 D 3 E 2, : h 1% , 0.041%,,, ;, 表 3 以游离氨基酸态氮含量为评价指标的正交试验方案及结果 Tab. 3 Orthogonal array design with experimental values of free amino nitrogen concentration as evaluation item A (%) B (h) C ( ) D (m/v) E (m/m) (g/100g) k k k R / 2017 / 41 / 6

6 3 结论,,, : : h 3% 1.. 2, 3.. 1,, IU/g; : h 1% ,, 0.041%,, : [1] Sumi H. Antibacterial activity of Bacillus natto growth inhibition against Escherichia coli O-157 [J]. Bioindustry, 1997, 14: [2] Sumi H. Accumulation of vitamin K (menaquione 7) in plasma after indigestion of natto and natto bacilli (B. subtillis natto)[j]. Food Science and Technology Research, 1999, 5(1): [3] Garrity G M.Bergey s Manual of Systematic Bacteriology[M]. 2nd ed.vol 1. New York: John Wiley & Sons, Inc., [4] Esaki H, Onozaki H, Osawa T. Antioxidative activity of fermented soybean products [M]. Washington D C: American Chemical Society, 1994: [5]. [J]., 2014, 39(9): Wang Lina. Optimization of liquid fermentation conditions of Bacilus nato [J].China Condiment, 2014, 39(9): [6] Murooka Y, Yamshita M. Traditional healthful fermented products of Japan[J]. J Ind Microbiol, 2008, 35(8): [7],,,. [J]., 2016, 37(12): Yao Haiqin, Wang Feijiu, Liu Fuli, et al. Chemical analysis and nutritional assessment of new varieties of Saccharina japonica[j]. Food Science, 2016, 37(12): [8],,. [J]., 2016, 02: 60-63, 69. Qiu Zhe, Sun Yuechun, Wu Haige.Nutritional components analysis of degraded Laminaria japonica by sodium alginate lyase[j]. Journal of Heilongjiang Bayi Agricultural University, 2016, 02: 60-63, 69. [9],. [M]. :, 2002: Wang Yonghua, Zhang Shuihua. Food Analysis[M]. Beijing: China Light Industry Press, 2002: [10],,,. [J]. ( ), 2011, 35(6): Zhang Anping, Li Huirong, Lv Wei, et al. Nattokinaes liquid fermentation condition optimizes[j]. Journal of Hebei Normal University (Natural Science Edition), 2011, 35(6): [11], [J],, 2013, 25(5): Hu Jing, Detection of nattokinase activity [J].Strait Pharmaceutical Journal, 2013, 25(5): [12] Fujita M, Nomura K, Hong K, et al. Purification and characterization of a strong fibrinolytic enzyme (nattokinase) in the vegetable cheese natto, a popular soybean fermented food in Japan [J]. Biochem Biophy Res Cbmmun, 1993, 197(3): [13],,,. [J]., 2010, 28(1): Chen Xiaofei, Zhou Fuzhong, Chen Guocan, et al. Characterization of nattokinase[j]. Henan Science, 2010, 28(1): [14],,,. ph [J]., 2009, 01: Zhao Zhengtao, Li Quanyang, Zhao Hongling, et al. Research on the characteristic of caseins at different ph[j]. Journal of Dairy Science and Technology, 2009, 01: [15],,,. [J]., 2015, 36(9): Wang Wenxiu, Yu Jia, Li Shan, et al. Effects of different conditions on free amino nitrogen content and nattokinase activity in Bacillus natto-fermented clam[j]. Food Science, 2015, 36(9): [16] Kubo Y, Inaka T, Hachiya T, et al. Development of a rifampicin-resistant Bacillus subtilis strain for natto fermentation showing enhanced exoenzyme production[j]. Journal of Bioscience and Bioengineering, 2013, 115(6): [17]. [D]. :, Wang gang. The solid ferment seperation and application of nattokinase[d]. Changchun: Jilin Agriculture Marine Sciences / Vol. 41, No. 6 /

7 University, [18],,,. [J]., 2016, 35(1): Huang Ting, Liu Liangzhong, Cao Yuxiang, et al. Optimization of solid-state fermentation technology of natto[j]. China Brewing, 2016, 35(1): Study on the optimal fermentation technology for a new type of bean sauce with kelp using Bacillus natto YANG Xing-xing 1, CUI Ying-xue 1, WU Ling-yu 1, WANG Ya-nan 1, LIU Yuan-ping 2, LI Yu-jin 2, WEI Yu-xi 1 (1. College of Life Sciences, Qingdao University, Qingdao , China; 2. Rongcheng Taixiang Food Co. Ltd., Rongcheng , China) Received: Jan. 10, 2017 Key words: kelp; soybean sauce; Bacillus natto; fermentation; free amino nitrogen; nattokinase activity Abstract: This study investigated the optimal fermentation technology for a new type of food using Bacillus natto. [Method] After fermentation of kelp and soybean by Bacillus natto, the concentration of free amino nitrogen and nattokinase activity were determined, and then the optimum conditions were confirmed by single factor analysis and orthogonal test. [Result] The maximum concentration of free amino nitrogen reached 0.041% when the fermentation was performed at 44 for 54 h with an inoculation amount of 1%, a solid-to-water ratio of (m/v), and a mixing ratio of Meanwhile, the maximum activity of nattokinase reached IU/g after 48 h of fermentation at 40 with an inoculation amount of 3%, a solid-to-water ratio of (m/v), and a mixing ratio of ( 本文编辑 : 康亦兼 ) 54 / 2017 / 41 / 6

H 2 SO ml ml 1. 0 ml C 4. 0 ml - 30 min 490 nm 0 ~ 100 μg /ml Zhao = VρN 100% 1 m V ml ρ g

H 2 SO ml ml 1. 0 ml C 4. 0 ml - 30 min 490 nm 0 ~ 100 μg /ml Zhao = VρN 100% 1 m V ml ρ g 16 6 2018 11 Chinese Journal of Bioprocess Engineering Vol. 16 No. 6 Nov. 2018 doi 10. 3969 /j. issn. 1672-3678. 2018. 06. 004 1 2 1 1 1. 330200 2. 330006 4 Box-Behnken 1 25 g /ml 121 92 min 2 6. 284%