Microsoft Word 曲敏二校 3318.doc

Size: px

Start display at page:

Download "Microsoft Word 曲敏二校 3318.doc"

桐毕
7 years ago
Views:

1 5 10 Vol. 5 No Journal of Food Safety and Quality Oct., 2014 SOD 曲敏 *, 秦丽楠, 刘羽佳, 范宏臣, 朱姝, 王金凤 (, ) 摘要 : 目的 (NBT) (SOD) 方法 SOD SOD, 结果 SOD, NBT , 2.6, 6, NBT, SOD, NBT 1.30~1.47, NBT, 结论 NBT SOD SOD 关键词 : ; ; ; The comparison of two methods of testing superoxide dismutase activity QU Min *, QIN Li-Nan, LIU Yu-Jia, FAN Hong-Chen, ZHU Shu, WANG Jin-Feng (Key Laboratory of Food Science and Engineering, Harbin University of Commerce, Harbin , China) ABSTRACT: Objective To compare the pyrogallol autoxidation and nitroblue tetrazolium (NBT) photoreduction methods for determination of superoxide dismutase (SOD) enzyme activity. Methods The paper based on the different methods to determine the SOD activity of standard, alfalfa and tomato, respectively, also compare the accuracy and sensitivity. Results The results showed that pyrogallol autoxidation and NBT photoreduction method of enzyme activity coefficient of variation were 0.84, 2.03, and the sample quantity which was 2.6 times of the former, dilution ratio was 6 times of the former, and the accuracy of pyrogallol autoxidation was higher than NBT photoreduction method but the sensitivity was poor, in detecting the SOD activity of standard. However, Pyrogallol autoxidation activity coefficient of variation was measured by NBT photoreduction reaction between 1.30 to 1.47 times the activities was measured by the coefficient of variation, the NBT photoreduction method has preferable stability and reproducibility than pyrogallol autoxidation in determining the SOD activity. Conclusion NBT photoreduction is an ideal testing method to determine the SOD activity of standard and plants. KEY WORDS: superoxide dismutase; pyrogallol autoxidation; NBT photoreduction method 基金项目 : (C ) ( ) (LBH-Q13098) Fund: Supported by Natural Science Foundation of Heilongjiang Province of China (C ), Science and Technology Research Project of Heilongjiang Province Department of Education ( ) and Postdoctoral Scientific Research Start-up Fund Project of Heilongjiang Province (LBH-Q13098) * 通讯作者 :,, qumin777@126.com *Corresponding author: QU Min, Professor, Harbin University of Commerce, Tongda Street 138, Daoli District, Harbin , China. qumin777@126.com

2 10, : SOD 引言 (superoxide dismutase, SOD),, (O 2- ), [1], [2] SOD 1997,, SOD SOD, [3] SOD, SOD -O 2 -, [4] C (NBT) -NBT [5-7],, (NBT) SOD, SOD SOD, SOD SOD, SOD 2 材料与仪器 2.1 材料与试剂 :, ;, : SOD, ;, ;, ; Tris EDTA-2Na, 2.2 仪器与设备 HWS 24, ; 722, ; TGL-16, ; DHP-9162, ; PHS-3 C ph, 3 实验方法 3.1 样品的处理苜蓿 SOD 的制备, 80, 10 g, 50 mmol/l ph 7.5, 4 24 h, r/min 20 min, SOD :, min, 8000 r/min 20 min, [8] : 0.5 mol/l HCl ph 3.6, 4 1 h,, : ( 1000 KDa), 1000 ml, 4 24 h( :, ) 番茄 SOD 的制备 [9], g, 2 50 mmol/l ph 7.5, : : -20, 15 min, 4 2 h, 8000 r/min 20 min,,,, 4, 8000 r/min 20 min,, 邻苯三酚自氧化法 [10,11],, : 25, 4.5 ml 50 mmol/l, ph 8.2 Tris-HCl 10 μl 50 mmol/l,,, Tris-HCl, 325 nm 30 s A, 6, OD/min : 25, 4.5 ml 50 mmol/l, ph 8.2 Tris-HCl, 20 min, 10 μl 50 mmol/l,,, Tris-HCl

3 3320 5, 325 nm 30 s A, 6, 50 1 / =(A 0 -A S )/A S 100 SOD (U/ml)=(A 0 -A S )/(A S 50 ) 4.5/V N : A 0 ; A S ; V ; N 3.3 NBT 光还原法 [12], 3 ml 130 mmol/l ; 750 μmol/l NBT; 20 μmol/l ; 100 μmol/l EDTA-2 Na; 50 mmol/l (ph 7.5), (, ), 4000 lx 20 min, NBT 560 nm,, 50% 1 4 结果与分析 4.1 两种方法测定 SOD 标准品酶活的比较两种测试方法的精确性 1 mg SOD 1 ml 50 mmol/l, ph 7.5 SOD, NBT 3 1 2, (SD=28.95) (CV=0.84) NBT (SD=74.28) (CV=2.03), NBT, 1 2,,, 测试方法的灵敏度 NBT SOD, NBT, 50,, 3, NBT 2.6 ;, NBT 6, NBT Table 1 表 1 邻苯三酚自氧化法测定 SOD 标准品数据结果 The results of pure SOD measurement data of the assay pyrogallol autoxidation SOD 1 SOD 2 SOD 3 (A 325 /min) (U/mL) (U/mL) SD CV 0.84 Table 2 表 2 NBT 光还原法测 SOD 标准品数据结果 The results of pure SOD measurement data of the assay NBT photoreduction SOD 1 SOD 2 SOD 3 50% (A 560 /min) (U/mL) (U/mL) SD CV 2.03

4 10, : SOD 3321 表 3 两种方法测定 SOD 标准品的比较 Table 3 Comparison of two methods of SOD measurement standards NBT /ml 两种方法测定植物 SOD 酶活的效果比较, SOD SOD,, SOD, NBT, [13] SOD, 325 nm, 50 1, 50, SOD,,,, NBT SOD, 4 SOD,, 24%, 35%, SOD 42% 50%, Table 4 表 4 NBT 法测苜蓿 SOD 酶活结果 The results of Alfalfa SOD enzyme activity test sensitivity with the assay NBT photoreduction 3 SOD (U/mL) SD CV Table 5 表 5 邻苯三酚自氧化法测番茄 SOD 酶活结果 The results of tomato SOD enzyme activity results with the assay pyrogallol autoxidation 3 SOD (U/mL) SD CV Table 6 表 6 NBT 法测番茄 SOD 酶活结果 The results of tomato SOD enzyme activity results with the assay NBT photoreduction 3 SOD (U/mL) SD CV

5 SOD, SOD 4, NBT SOD, NBT,,, SOD, 5 6, NBT SOD,,, NBT 1.30~1.47,, NBT,,, SOD,, SOD,,,,, NBT [14],,, ;,,, SOD [15], Fenton, MSOD Lewis, SOD, SOD NBT ; 5 6,, NBT, NBT 1 2,, 5 6, SOD,, SOD,, 5 结论 NBT, SOD SOD, SOD, NBT,,,, NBT SOD( SOD SOD ),, SOD,, SOD, NBT,, NBT,, NBT, SOD SOD, NBT, 参考文献 [1],,. SOD [J]., 2002, 18(4): Xu J, Zhao LH, Song GW. Chemiluminescence method for determination of SOD activity of interference factors and eliminating methods [J]. J Anal Sci, 2002, 18(4): [2],,,. SOD [J]., 2010, 8(2): Dou JH, Yu SX, Fan SL, et al. SOD and plant stress resistance [J]. Mol Plant Breeding, 2010, 8(2): [3] Wang,SY Shao B, Liu ST, et al. Purification and characterization

6 10, : SOD 3323 of Cu, Zn-superoxide dismutase from black soybean [J]. Food Res Int, 2012, 47(2): [4],,,. [J]., 1983, (5): Shao CB, Luo GH, Wang AG, et al. Several kinds of detecting superoxide dismutase (sod) reaction of comparison [J]. Plant Phy Commun, 1983, (5): [5],,,. SOD [J]., 2011, 32(6): Xu D, Zhao J, Huang HC, et al. Determination of SOD specific activity in animal and plant tissues by improved xanthine oxidase method[j]. Food Sci, 2011, 32(6): [6],,. [J]., 2002, 12(1): Xu YB, Zhao SJ, Guo Y. The commentary of some measure methods for superoxide dismutase [J]. Guangdong Pharm, 2002, 12(1): [7],,. [J]., 2008, 29(12): Jang Y, Mao L, Cai YQ. Advance of study on extraction and purification techniques and activity assay methods of superoxide dismutase in fruits and vegetables [J]. Food Sci, 2008, 29(12): [8]. [D]. :, He Y. Blueberry leaves superoxide dismutase extraction and purification and application[d]. Changchun: Jilin Agricultural University, [9] 珺,,. [J]., 2012, 40(7): Li J, Ma LT, Gao SL. Optimization of the extraction process of superoxide dismutase from tomatoes[j]. J Anhui Agric Sci, 2012, 40(7): [10]. SOD [J]., 2005, (10): Sun YJ. Garlic extraction of SOD[J]. Chem Biol Eng, 2005, (10): [11],. SOD [J]., 2000, 10(6): 673. Li YL, Zhang Y. The development of pyrogallol autoxidation method of SOD activity[j]. Chin J Health Laboratory Technol, 2000, 10(6): 673. [12]. SOD [D]. :, Song XQ. Isolation and Purification of SOD from Maize[D]. Baoding: Hebei Agricultural University, [13],. NBT SOD ( )[J]., 2000, 14(2): Zhang WJ, Ge C. NBT reduction method, the development of three light oxidation method was developed for the determination of SOD enzyme activity comparison (brief) [J]. J Hebei Vocational Tech Teachers Coll, 2000, 14(2): [14],,,. [J]., 2008, 36(4): Yang L, Chen GH, Cui JC, et al. Fluorescence dynamic research method to determine the activity of superoxide dismutase[j]. Chin J Anal Chem, 2008, 36(4): [15],,,. SOD?[J]., 2003, 37(1): Liao ZR, Chen WX, Ai CC, et al. The development of three phenol autoxidation method suitable for determination of SOD activity? [J]. J cent Chin Norm Univ, 2003, 37(1): 作者简介 ( 责任编辑 : 白洪健 ) 曲敏, 教授, 主要研究方向为新型植物蛋白及食品添加剂 qumin777@126.com

<4D6963726F736F667420576F7264202D203631372D3632312032303133303430333030312DBACEC0F25FD0A3B6D4B8E55F2DB6FED0A32D2D2DC8A5B5F4CDBCD6D0B5C4BBD8B3B5B7FBBAC52E646F63>

<4D6963726F736F667420576F7264202D203631372D3632312032303133303430333030312DBACEC0F25FD0A3B6D4B8E55F2DB6FED0A32D2D2DC8A5B5F4CDBCD6D0B5C4BBD8B3B5B7FBBAC52E646F63> 第 4 卷第 2 期食品安全质量检测学报 Vol. 4 No. 2 2013 年 4 月 Journal of Food Safety and Quality Apr., 2013 何莉 *, 姜笑寒 ( 广东省食品药品职业技术学校, 广州 510663) 摘要 : 本文通过科学地运用 html5+jsp+sql 技术, 建