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1 酶解蓝圆鲹蛋白制备降血压肽的工艺研究舒冰, 孙建华, 伍善广, 赵钟兴, 廖丹葵 (, ) 摘要 : 以蓝圆鲹 (Decapterus maruadsi) 鱼肉蛋白为原料, 选用 6 种蛋白酶在各自适宜的条件下酶解制备降血压肽, 通过检测酶解产物对血管紧张素转化酶 (ACE) 的体外抑制活性, 筛选出木瓜蛋白酶为制备蓝圆鲹降血压肽的最佳用酶考察了不同水解度的酶解产物对 ACE 抑制效果, 当水解度为 13.7% 时, 产物对 ACE 的最高抑制活性可达 67.4% 在此基础上, 分别了考察酶解时间初始酶用量初始底物浓度酶解温度和 ph 值对酶解工艺的影响, 得到制备蓝圆鲹降血压肽的适宜酶解工艺为 : 初始酶用量为 7000 U/g, 初始底物浓度为 25 g/l, 酶解温度为 45, ph 为 7.0, 酶解 240 min 采用超滤技术对酶解产物活性组分进行分离富集, 发现产物中高活性部分均可富集于 10 kda 和 5kDa 渗透液中, 且高活性组分分子量主要集中在 5 kda 以下, 该组分对 ACE 的活性抑制率为 88.47% 关键词 : 酶解 ; 蓝圆鲹 (Decapterus maruadsi); 蛋白 ; 降血压肽中图分类号 : S98 文献标识码 : A 文章编号 : (2013) (angiotension converting enzyme, ACE), I, [1], ACE, ACE 1977 ACE, ACE, ACE [1], ACE, ACE,,,,,, [2] [3] [4] [5] [6] ACE 鲹,,,, [7,8] 鲹 (Decapterus maruadsi), 6 鲹,,, 鲹鲹 1 材料与方法 1.1 材料与仪器 (HHL) (ACE) Sigma ; ; 鲹 : ; : : ( ; ) : (1986-),,,, :, : , shubingice@126.com;,,,,, liaodk@gxu.edu.cn Marine Sciences / Vol. 37, No. 3 /

2 A-88, ; 4K15, Sigma ; Aglient 1100, ; Labscale, Millipore 1.2 实验方法鲹, 75,, 20, 15 min, ph, ph, ph, 10 min, r/min 10 min, (DH), ph-stat [9] Cushman, HPLC [10] 鲹, ph kda 5 kda, ACE 1 6 Fig. 1 The ACE inhibitory activity of six kinds of hydrolysates 2.2 木瓜蛋白酶酶解蓝圆鲹蛋白工艺优化 (DH) (IP) 鲹 ACE 2 2 实验结果与分析 2.1 不同酶解产物的活性 6 鲹, ACE 1 1 ACE, ACE, ACE, [11-12], 鲹 6 ACE,,, ACE, 鲹 Fig. 2 2 The diversification for DH and ACE inhibitory ratio 2, 鲹,, ACE, 13.7%, ACE, ACE,, ACE,, ACE, ACE [13], ACE, 46 / 2013 / 37 / 3

3 [14] ACE, 13.0% 16.0%, 240 min g/l, 55, ph 7.0, 4 h,, 3 4 Fig. 4 Effect of different initial substrate concentrations on DH 3 Fig. 3 Effect of different initial enzyme concentrations on DH 3,, DH,,,, U/g,,,, U/g U/g, 55, ph 7.0, 4 h,, 4 4, DH,,,, 30 g/l,, 25 g/l U/g, 25 g/l, ph 7.0, 4 h, 5 5 Fig. 5 Effect of different temperatures on DH,,,, [15] 5, 45, ph U/g, 25 g/l, 45, ph 6 ph,,,,,, ph,, Marine Sciences / Vol. 37, No. 3 /

4 [15] 6, ph, ph,,, ph 7.0 Fig. 6 6 ph Effect of different ph values on DH 2.3 超滤前后多肽回收率及抑制活性 22.4 mg/l 500 ml, ph 7.0, 10 kda 5 kda 50 ml, ACE, 1 表 1 超滤后不同部分滤液的 ACE 抑制活性 Tab. 1 ACE Inhibitory activity of different ultrafiltrating solutions ACE (%) (%) kda () 5 kda () 1, 10 kda 5 kda, 5 kda ACE 88.47%, 1.72,, ACE 2~12, 15 [16], 3kDa, 5 kda, [17,18], 5 kda, 3 结论 (1) 6 鲹 ACE, ACE 鲹, 13.0% 16.0% ACE, 13.7% ACE 67.4% (2) ph, : U/g, 25 g/l, 45, ph 7.0, 240 min, 16.0%, ACE (3), 10 kda 5 kda, 5 kda ACE 88.47%, 5 kda : [1] Sang-Hoon Lee, Zhong-Ji Qian, Se-Kwon Kim. A novel angiotensin I converting enzyme inhibitory peptide from tuna frame protein hydrolysate and its antihypertensive effect in spontaneously hypertensive rats[j]. Food Chemistry, 2010, 118: [2] Marta Miguel, María A Manso, Mercedes Salaices, et al. Vascular effects and antihypertensive properties of kappa-casein macropeptide[j]. International Dairy Journal, 2007, 17(12): [3] Vanessa Vermeirssen, John Van Camp, Willy Verstraete. Fractionation of angiotensin I converting enzyme inhibitory activity from pea and whey protein in vitro gastrointestinal digests [J]. Journal of the Science of Food and Agriculture, 2005, 85(3): [4] Wu Jianping, Ding Xiaolin. Hypertensive and physiological effect of angiotensin converting enzyme inhibitory peptides derived from soy protein on spontaneously hypertensive rats[j]. Journal of Agricultural and Food Chemistry, (1): [5] Rotimi E Aluko, Erasmus D Monu. Functional and bioactive properties of quinoa seed protein hydrolysates[j]. Journal of Food Science, 2003, 68(4): / 2013 / 37 / 3

5 [6] Ali Bougatef, Naima Nedjar-Arroume, Ahmed Barkia, et al. Angiotensin I-converting enzyme (ACE) inhibitory activities of sardinelle(sardinella aurita) by-products protein hydrolysates obtained by treatment with microbial and visceral fish serine proteases[j]. Food Chemistry, 2008, 111(2): [7],,,. 鲹 [J]., 2005, 26(2): [8]. [J]., 2010(06): [9] Marquez M C, Vazquez M A. Modeling of enzymatic protein hydrolysis[j].process Biochemistry, 1999, 35: [10],,,. [J]., 2008, 32(8): [11],,. [J]., 2008, 29 (11): [12],,. ACE [J]., 2010, 31(14): 1-4. [13],. ACE [J]., 2007, 22 (6): [14] Chiang W D,Tsou M J,C hu Y H. Angiotensin I converting enzyme inhibitor derived from soy protein hydrolysate and produced by us ing m em brane reactor[j]. Food Chem, 2006, 98: [15]. [M]. :, 2002, [16] Robert M C,R azaname A, M utter M, et al. Identification of angiotensin-i-converting enzyme inhibitory peptides derived from sodium caseina te hydro lysates produced by Lactobacillus he lveticus N CC 2765[J]. J Agr Food Chem, 2004, 52(23): [17],,. [J]., 2010, 36(9): [18],,,. [J]., 2010, 12(1): Study on technology of antihypertensive peptides production from Decapterus maruadsi protein with enzymatic hydrolysis SHU Bing, SUN Jian-hua, WU Shan-guang, ZHAO Zhong-xing, LIAO Dan-kui (Schoo1 of Chemistry and Chemical Engineering, Guangxi University, Nanning , China) Received: Feb.,20,2012 Key words: hydrolyze; Decapterus maruadsi; protein; antihypertensive peptides Abstract: The protein from Decapterus maruadsi was hydrolyzed by six different proteolytic enzymes in their optional conditions. The inhibition percent (IP) of Angiotensin I-converting enzyme (ACE) by the hydrolysates was examined with high performance liquid chromatography. Results showed the papain was the best enzyme for producing ACEI. When the degree of hydrolysis (DH) was 13.7%, the inhibition of ACE by the hydrolysate reached as high as 67.4%. Further on, the influence of hydrolysis time, initial enzyme concentrations ([E]), initial substrate concentrations ([S]), hydrolysis temperatures and ph values was investigated. Results showed that the optimal condition for producing ACEI was as follows: [E] at 7000 U/g Pro, [S] at 25 g/l, hydrolysis temperature at 45, ph at 7.0 and hydrolysis time in 240 minutes. Then ultrafiltration technology was used to separate and enrich the active part of hydrolysates. Results showed that the higher active part of the hydrolysates can be enriched in the penetrating fluid of both 10 and 5 KDa. It was also found that the higher active hydrolysates were mainly lower than 5 KDa with an ACE inhibition rate of 88.47%. ( 本文编辑 : 康亦兼 ) Marine Sciences / Vol. 37, No. 3 /

2014-目录6期.indd

2014-目录6期.indd 50 0, Vol.5, No.06 食品科学工艺技术猪股骨头胶原蛋白降血压肽的分离纯化刘小红, 李诚 *, 付刚, 苏赵 ( 四川农业大学食品学院, 四川雅安 650) 摘要 : 依次采用碱性蛋白酶木瓜蛋白酶和风味蛋白酶对猪股骨头胶原蛋白进行酶解,