专业英语翻译葡萄籽提取物对面包的抗氧化活性和质量属性的影响

1、The effects of grape seed extract fortification on the antioxidant activity and quality attributes of bread葡萄籽提取物对面包的抗氧化活性和质量属性的影响Abstract: The antioxidant activity change of breads added with grape seed extract (GSE) was investigated. The results showed that bread with the addition of GSE had stron

2、ger antioxidant activity than that of blank bread, and increasing the level of GSE addition further enhanced the antioxidant capacity of the bread. However, thermal processing caused antioxidant activity of GSE added to bread to decrease by around 3040%. We also studied the effect of GSE on the form

3、ation of detrimental Ne-(carboxymethyl) lysine (CML), a famous advanced glycation endproduct in bread. According to the results, GSE could reduce CML in bread and acted in a dose-dependent manner. Meanwhile, except for an acceptable colour change, adding GSE to bread had only little effect on the qu

4、ality attributes of the bread. Altogether, our findings indicate that GSE-fortified bread is promising to be developed as a functional food with relatively lower CML-related health risks, yet a high antioxidant activity.Keywords: Antioxidant activity Grape seed extract Bread Advanced glycation endpr

5、oducts摘要：对加入葡萄籽提取物（GSE）对面包的抗氧化活性的变化进行了研究。结果表明，加入GSE的面包具有比空白面包（空白对照）更强的抗氧化活性，并且增加GSE的添加量会进一步提高面包的抗氧化能力。然而，热加工造成添加GSE的面包的抗氧化活性降低大约3040%。我们还研究了添加GSE对形成有害氖-（羧甲基）赖氨酸（CML）的影响，我们还研究了GSE对有害氖（羧甲基）赖氨酸（CML）的形成的影响，是面包中著名的晚期糖基化终末产物。根据研究结果，GSE能减少面包中CML的形成，并且呈现剂量依赖性。同时，除了具有可接受的颜色变化，添加GSE对面包质量属性的影响不大。总之，我们的研究结果表明，G

6、SE强化面包有望被开发作为功能性食品，有相对较低的CML含量和较低的健康风险，但具有较高的抗氧化活性。关键词：抗氧化活性；葡萄籽提取物；面包；晚期糖基化终末产物1. IntroductionPhenolic compounds are widely distributed in foods, such as fruits (Robards, Prenzler, Tucker, Swatsitang, & Glover, 1999), vegetables (Bonoli, Gallina Toschi, & Lercker, 2005) and cereals (Dykes &

7、amp; Rooney, 2007; Liyana-Pathirana & Shahidi, 2006). As naturally occurring antioxidants, phenolic compounds have been reported to possess diverse beneficial bioactivities, including anti-allergic, antiviral, anti-inflammatory and anti-mutagenic properties (Yao et al., 2004). Meanwhile, a large

8、 number of in vitro and animal studies have also suggested that phenolic compounds may be effective in protecting against cancer, and cardiovascular diseases. The protective effects might be mediated through their action as antioxidants to prevent oxidative damage induced by reactive oxygen species

9、to some important biomolecules (like DNA, lipids andproteins) under pathological conditions (Hollman, 2001; Yao et al.,2004). Recently, phenolic antioxidants have been viewed as an important class of food ingredients either as food additives or as novel ingredients to introduce extra health benefits

10、 to various food products. Considering the fact that heat treatment is a widespread processing method in the food industry, a salient question is whether these thermal processes would lead to significant alterations in the antioxidant capacities of phenolic additives. It has been reported that total

11、 antioxidant activities of tomatoes and carrots were enhanced with thermal processing (Dewanto, Wu, Adom, & Liu, 2002; Patras, Brunton, Da Pieve, Butler, & Downey, 2009) while antioxidant capacities of soybeans were lowered with similar processing (Xu & Chang, 2008).1.前言酚类化合物广泛分布于食品中，如水果

12、、蔬菜、谷物等。作为天然存在的抗氧化剂，酚类化合物已被报道具有多样有益的生物活性，包括抗过敏，抗病毒，抗炎和抗诱变性等。同时，大量的体外和动物研究还表明，酚类化合物可以有效地防止癌症和心血管疾病。其保护功能可能是通过它们作为抗氧化剂的作用来实现的，防止活性氧的氧化损伤从而引起一些重要生物分子（如DNA、脂质和蛋白质）处于病理状态。最近，酚类抗氧化剂已被视为食品成分的一类重要的或者作为食品添加剂或作为新成分引入到各种食品中有额外的健康益处。考虑到事实情况，热处理是在食品工业中的普遍的处理方法，一个突出的问题就是在这些热加工过程中是否会导致酚类添加剂抗氧化能力的显著改变。据报道，在热加工过程中番茄

13、和胡萝卜的抗氧化活性会增强，而大豆的抗氧化能力会降低。 So far, little work has been focused on evaluating the relationship between thermal processing of food and changes in antioxidant capacities of phenolic additives. For this consideration, we investigated the influence of thermal processing on antioxidant capacity of grape

14、 seed extract (GSE) in the present work. As a well-known nutraceutical product, GSE is an abundant source of catechins and proanthocyanidins with a strong antioxidant and free radical scavenging activity (Liang, Wang, Simon, & Ho, 2004; Wu, Wang, & Simon, 2005). Moreover, it shows other biol

15、ogical effects as well, such as inhibition of platelet aggregation, anti-inflammation and anti-ulcer activity (Saito, Hosoyama, Ariga, Kataoka, & Yamaji, 1998; Vitseva, Varghese, Chakrabarti, Folts, & Freedman, 2005). In this study, different amounts of GES were mixed with bread ingredients

16、before starting the bread-making program on bread makers. Comparing the antioxidant activities of the GSE-fortified bread with those of standard GSE solutions would enable estimation of the extent to which the thermal process might affect the antioxidant of GSE. Textural analysis, colour measurement

17、 and sensory evaluation were also conducted to investigate whether addition of GSE will affect quality attributes of bread or not.迄今为止，很少有研究工作集中于评价热加工处理与酚类添加剂抗氧化能力变化之间的关系。出于这种考虑，在目前工作中我们研究了热加工对葡萄籽提取物（GSE）抗氧化能力的影响。作为一种众所周知的营养保健品，GSE是儿茶素和花青素的丰富来源，具有很强的抗氧化活性和清除自由基的能力。此外，它也同样显示了多种生物学效应，例如抑制血小板聚集，抗炎，抗溃疡活

18、性等。在本研究中，在面包机开始面包制作程序之前，在面包配料中加入不同量的GSE。比较GSE强化面包与标准GSE溶液的抗氧化活性，能够估计热加工过程影响GSE抗氧化活性的程度。进行结构分析，颜色测量和感官评价来研究添加GSE是否会影响面包的质量属性。 In addition, the effect of GSE on the formation of Ne-(carboxymethyl)lysine (CML) in bread was studied. As a well-characterised detrimental advanced glycation endproduct (AGE)

19、 (Sebekova & Somoza, 2007; Tessier & Niquet, 2007), CML was widely found in a range of foods including bread (Charissou, Ait-Ameur, & BirlouezAragon, 2007; Hartkopf & Erbersdobler, 1994) and its content in crust was reported to be much higher than in crumb (Assar, Moloney, Lima, Mage

20、e, & Ames, 2009). Currently it is viewed as a potential toxicant in food. Moreover, it has become a biomarker associated with oxidative stress, atherosclerosis and diabetes in humans (Nerlich & Schleicher, 1999; Schleicher, Wagner, & Nerlich, 1997). Natural antioxidants like bean extract

21、s (Madhujith, Amarowicz, & Shahidi, 2004), peanut skin and some flavonoids have been proven to possess strong inhibitory effects on the formation of AGEs in vitro, which are mainly attributed to their potent antioxidant activities (Lou, Yuan, Yamazaki, Sasaki, & Oka, 2001; Peng et al., 2007;

22、 Wu & Yen, 2005). In addition, capability of certain phenolics such as tea catechins and proanthocyanidins to scavenge reactive carbonyl species (such as glyoxal, methylglyoxal) in glycation process has also been proposed to contribute to inhibition of AGE formation (Lo et al., 2006; Peng et al.

23、, 2008). Therefore, it is of great interest to investigate whether GSE fortification could reduce CML content in bread. In this regard, influence of different levels of GSE addition on CML content of bread was examined. 此外，研究了GSE对面包中氖-（羧甲基）赖氨酸（CML）形成的影响。作为一个特征明显的有害的晚期糖基化终末产物（AGE），CML广泛存在于各种食品中包括面包，并

24、且其含量在面包皮中比在面包心中高得多。目前，它被看作是食品的潜在毒物。而且，它已成为与人类氧化应激，动脉粥样硬化和糖尿病相关联的生物标志物。在体外实验中，天然抗氧化剂像豆子提取物，花生皮和一些黄酮类化合物已被证明对AGE的形成有很强的抑制作用，这主要归功于其强大的抗氧化活性。此外，某些酚类物质如茶叶中的儿茶素和原花青素清除活性羰基物质（如乙二醛，甲基乙二醛）的能力，也已经被提出在糖化过程中有助于抑制AGE形成。因此，研究GSE强化能否降低面包中的CML含量是非常有意义的。就这一点而言，研究了不同水平的GSE添加量对面包中CML含量的影响。2. Materials and methods2.1.

25、 ChemicalsIngredients for bread-making were bought from a local supermarket in Hong Kong. Grape seed extract with 95% proanthocyandins including catechin and epicatechin was a gift from Shenzhen BannerBio Inc. (Shenzhen, PR China). The compound Ne-(carboxy methyl) (CML) was purchased from NeoMPS (St

26、rasbourg, France). Ortho-phthalaldehyde (OPA), 2-mercaptoethanol, sodium borohydride, boric acid, sodium tetraborate decahydrate, sodium hydroxide, hydrochloric acid,2,2 0-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS), potassium peroxodisulphate and trolox were all obtained from SigmaAldr

27、ich Company (St. Louis, MO, USA). All analytical and HPLC grade solvents used were obtained from BDH Laboratory Supplies (Poole, UK).2.材料与方法2.1.化学药品面包制作的材料购买于香港当地的超市。葡萄籽提取物有95原花青素包括儿茶素和表儿茶素，由深圳Banner生物公司提供。化合物氖-（羧甲基）（CML）购买自NeoMPS。正-邻苯二醛（OPA），2-巯基乙醇，硼氢化钠，硼酸，四硼酸钠十水合物，氢氧化钠，盐酸，2,2-连-二（3-乙基苯并噻唑-6-磺酸）（AB

28、TS），过氧化钾和生育酚均全部购自Sigma-Aldrich公司。所有分析和HPLC级溶剂由BDH实验室提供。2.2. Preparation of breadBread was made using a bread maker (Moulinex, Ecully Cedex, France) bought from a local supermarket. The program 2 for fast basic white bread (500 g/each) was chosen for bread-making.Briefly, this program adopts the foll

29、owing sequential process: first kneading (5 min), rest (5 min), second kneading (20 min), first rising (15 min), third kneading (10 s), second rising (8 min and 50 s), fourth kneading (10 s), third rising (29 min and 50 s) and baking (43 min). The recipe for bread includes canola oil (3.5 tea spoon)

30、, water (190 mL), salt (1 tea spoon, about 5.7 g), sugar (2.5 tea spoon, about 8.88 g), milk powder (1.5 table spoon, about 10.28 g), white bread flour (350 g) and flaked dried yeast (1 tea spoon, about 2.85 g). GSE with different levels (300 mg, 600 mg and 1 g) was added into breads (500 g/each), r

31、espectively. All breads were made in three batches. Bread crusts were carefully sliced off and ground for later determination of CML.2.2. 面包的准备面包是用从当地超市购买的面包机制作的。选择程序2为快速基本白面包（500克/个）制作。简单地说，这个程序采用以下顺序加工：首先揉制（5分钟），静置（5分钟），第二揉制（20分），第一次上升（15分钟），第三次揉制（10秒），第二次上升（8分钟50秒），第四次揉制（10秒），第三次上升（29分钟50秒），然后烘烤（

32、43分钟）。面包配方包括低芥酸菜子油（3.5茶匙），水（190ml），盐（1茶匙，约5.7克），糖（2.5茶匙，约8.88克），奶粉（1.5汤匙，约10.28克），白面包粉（350克）和片状干酵母（1茶匙，约为28.5克）。GSE以不同水平（300毫克，600毫克和1克）分别加入到面包（500克/个）中。所有的面包都平行制作三组。将面包皮仔细切掉以便随后测定CML含量。2.3. Antioxidant activity measurementGround bread powders were vortex-dispersed (5 s), respectively, into water, 3

33、0%, 50% and 70% ethanol with a final concentration of 100 mg/mL. The samples were extracted by sonication for 30 min and then centrifuged (3233g) for 20 min. The supernatants were used for subsequent determination of antioxidant capacity. Meanwhile, 0.06, 0.12 and 0.2 mg/mL GSE solutions were prepar

34、ed as controls corresponding to different concentrations of GSE added to bread. Total antioxidant capacity was determined using the trolox equivalent antioxidant capacity (TEAC) assay according to the literature (Cheng, Chen, & Wang, 2007) with minor modifications. In brief, 7 mM ABTS salt solut

35、ion was reacted with 2.45 mM potassium peroxodisulphate solution and the reaction mixture was allowed to stand in the dark for 16 h at room temperature (25C) and was used in 2 days. The resultant radical solution was diluted with deionized water to an absorbance of 0.7 ± 0.05 at 734 nm. Fifty m

36、icrolitres of bread sample solution or standard (different concentrations of trolox) and 50L of water were added to 1.8 mL of diluted ABTS+ solution and absorbance was taken at 734 nm on a UV-1206 Spectrophotometer (Shimadzu, Kyoto, Japan) after 6-min incubation. Results were expressed as TEAC value

37、s (nmol trolox/mg bread sample). Triplicate analyses were performed.2.3.抗氧化活性测定研磨过得面包粉呈涡流分散（5秒），分别地加入水、30、50和70的乙醇，最终浓度为100mg/ml。将样品超声处理提取30分钟，然后离心（3233g）20分钟。上清液用于后续测定抗氧化能力。同时，制备0.06、0.12和0.2mg/ml的 GSE溶液作为相应不同浓度的GSE加入到面包中。总抗氧化能力的测定根据文献稍作修改，使用Trolox（生育酚）当量抗氧化能力（TEAC）法测定。简言之，7mM ABTS盐溶液与2.45mM过氧化钾溶液反

38、应，然后将反应混合物于室温下（25）避光静置（在黑暗中）16个小时，在两天之内使用。所得自由基溶液用去离子水稀释至在734nm处吸光度为0.7±0.05。五十微升的面包样品溶液或标准的（不同浓度的生育酚）和50L水加入到1.8ml稀释的ABTS溶液中，静置6min后，使用UV-1206分光光度计（岛津，京都，日本）在734nm处测定吸光度。结果以TEAC值表示（nmol生育酚/mg面包样品）。测定平行进行三次。2.4. Textural analysisAfter cooling for 1 h, breads were cut into slices of 25 mm thickn

39、ess with a bread knife. The central two slices were used to perform textural analysis on a TA-XT2 texture analyzer (Stable Micro System, Surrey, UK) equipped with a cylindrical probe of 20 mm in diameter. Based on texture profile analysis (TPA), hardness was calculated. Textural analysis was perform

40、ed in the mode of Measure Force in Compression”, while trigger type was set as auto-5 g at the speed of 2 mm/s followed by a 10 mm compression distance.2.5. Measurement of colour changesColour changes of bread slices with or without the addition of different concentrations of GSE was measured with a

41、 tristimulus reflectance colourimeter (Minolta CR-400 Chroma Meter, Konica Minolta, NJ, USA), and readings were expressed as L* , a*and b* values, where L * indicates whiteness (value 100) or blackness (value 0),a* indicates red (positive value) or green (negative value), and b* indicates yellow (po

42、sitive value) or blue (negative value). 2.6. Sensory evaluation Ten untrained panelists engaged in the sensory evaluation based on scoring various bread samples according to five attributes: sweetness, porosity, astringency, stickness and colour. In terms of corresponding intensity, each attribute w

43、as ranked as low, medium and high grades gaining 1, 2, 3 score, respectively. Sensory evaluation was conducted on three batches of bread. The mean scores and corresponding standard deviation values for each attribute were calculated for comparison.2.4.织构分析冷却1小时后，用面包刀将面包切成25mm的厚度。采用装配有直径20毫米圆柱形探针的TA-

44、XT2质构仪（稳定微系统，萨里，英国）对中间两片面包进行织构分析。根据纹理剖面分析（TPA），计算硬度。织构分析在 “测量压缩力量”模式上进行，触发器类型被设定为自动5g，以2mm /秒的速度，接着通过10mm的压缩距离。2.5. 颜色变化测定采用三色反射色度计（美能达CR-400色度仪，柯尼卡美能达，NJ，美国）测定加入或不加入不同浓度GSE后的面包片的颜色变化，并且读数表示为L *，a *和b*值，其中L*表示白度（值100）或黑度（值0），a*表示红色（正值）或绿色（负值），b*表示黄色（正值）或蓝色（负值）。2.6.感官评定10名未经训练的小组成员参与感官评定，根据他们对不同面包样品的

45、打分来评定，根据五种特性：甜度，孔隙率，收敛性（涩味），粘性和颜色。依照相对强度，每种特性分为低、中和高三个等级，分别获得1、2、3分。感官评价对三组面包进行。计算每种特性的平均得分和相应的标准偏差值用于比较。2.7. Determination of CML in bread crustHPLC analysis was adopted to determine CML content in bread crust with reference to the reported method (Drusch, Faist, & Erbersdobler, 1999) with some

46、 modifications. The sample preparation procedures include defatting, reduction, hydrolysis and chemical derivatization.2.7.1. DefattingBread crusts (1 g) were defatted using two successive extractions in 9 mL of a chloroform/methanol (2:1, v/v) solution followed by centrifugation (3233g at room temp

47、erature) for 20 min. Then, the defatted bread crusts were dried completely at 50C.2.7.2. ReductionDefatted bread crusts (100 mg) were reduced with 4 mL sodium borate buffer (0.2 M, pH 9.4) and 2 mL sodium borohydride (1 M in 0.1 M NaOH) and then incubated for 4 h at room temperature2.7.3. Hydrolysis

48、Subsequently, hydrochloric acid (HCl) was added to the reduced samples to achieve a final concentration of 6 M HCl. In order to avoid any oxidative process, samples were degassed with a stream of nitrogen for 5 min. Then samples were hydrolysed at 110C for 20 h. After hydrolysis, samples were dried

49、by rotary evaporation and then mixed with 10 mL of water before filtration. Two millilitres of the filtrates were concentrated by rotary evaporation and re-dissolved in sodium borate buffer (0.2 M, pH 9.4) for later derivatization.2.7.4. DerivatizationOPA (10 mg) was dissolved in 2 mL of methanol to

50、 obtain an OPA stock solution. The derivatization reagent was composed of 1 mL of OPA stock solution, 8lL of 2-mercaptoethanol and 3.992 mL borate buffer (consisting of 0.2 M boric acid and 0.2 M NaOH, pH 9.9) (Hanczko & Molnar-Perl, 2003). The reagent was prepared at least 90 min before use for

51、 derivatization and was disposed after 2 days. Hydrolysates (200lL) were mixed with 100lL of derivatization reagent and incubated for 3 min before subjecting to HPLC analysis.2.7.5. HPLC analysisAnalytical HPLC was carried out using a Waters 2695 Separation Module equipped with a Waters 2475 Multikf

52、luorescence detector. A pre-packed ODS-A column (1504.6 mm, 5lm, YMC Co. Ltd., Kyoto, Japan) was selected for HPLC analysis and detection was at 340 nm (excitation) and 455 nm (emission). The flow rate was 1.0 mL/min and the injection volume was 10lL. The mobile phases were: (solvent A) sodium aceta

53、te buffer (pH 6.7, 20 mM) acetonitrile (90/10, v/v) and (solvent B) acetonitrile. The elution started with 5% B and hold for 9 min, and then it was linear gradient to 70% B in 5 min and kept at 70% B till 17 min. The gradient was subsequently set back to 95% B within 1 min and the post running time

54、was 10 min. Peaks for CML-derivatives in bread samples were confirmed by comparison with an authentic compound. CML contents of the bread samples were calculated based on the peak areas of the corresponding CML derivatives.2.7. 测定面包皮的CML值根据文献报告的方法稍作修改，采用HPLC分析法测定面包皮的CML含量。样品制备程序包括脱脂、还原、水解和化学衍生化。2.7.

55、1脱脂取面包皮（1克）利用9ml氯仿/甲醇（2:1，v/v）溶液连续两次萃取脱脂，接着离心分离（室温3233g）20分钟。然后将脱脂面包皮在50下完全干燥。2.7.2还原 脱脂面包皮（100毫克）分别用4mL硼酸钠缓冲液（0.2M，pH值9.4）和2ml硼氢化钠（1M，在0.1M NaOH中）还原，然后在室温下培养4小时。2.7.3水解随后，还原样品中加入盐酸（HCl）以达到最终浓度6 M HCl。为了避免任何的氧化过程，将样品用氮气气流脱气5分钟。然后将样品在110下水解20小时。水解后，将样品通过旋转蒸发干燥，然后加入10ml水后过滤。取2ml滤液通过旋转蒸发浓缩，并且重新溶解在硼酸钠缓冲

56、液（0.2M，pH值9.4）中以便随后的衍生化。2.7.4衍生化取OPA（10mg）溶解于2ml甲醇中，得到OPA储备液。衍生化试剂是由1ml OPA储备液，8l 2-巯基乙醇和3.992ml硼酸盐缓冲液（由0.2M硼酸和0.2M NaOH组成，pH值9.9）组成。试剂至少在衍生前90min制备，并且可放置2天。水解产物（200l）与100l衍生化试剂混合，静置3分钟后进行HPLC分析。2.7.5HPLC分析HPLC分析是采用配备有Waters2475多荧光检测器的Waters2695分离模块进行。选择一种预包装的ODS-A柱（150 4.6mm，5m，YMC有限公司，京都，日本）用于HPLC

57、分析，检测是在340nm（激发）和455nm（发射）下进行。流速为1.0ml/分钟，注射体积为10L。流动相为：（溶剂A）乙酸钠缓冲液（pH6.7，20mM）-乙腈（90/10，v/v）和（溶剂B）乙腈。洗脱开始用5的B并保持9分钟，然后在5min内以线性梯度升至70的B，并且在70B保持17分钟。随后梯度被设置回至95B 1分钟内，然后运行10分钟。面包样品的CML衍生物的峰与真实化合物的峰进行对比。根据相对应的CML衍生物的峰面积来计算面包样品中的CML含量。 2.8. Statistical analysisStatistical analyses were performed usin

58、g the SPSS statistical package (SPSS Inc., Chicago, IL). Paired samples Ttest was applied to determine whether a particular treatment of the sample would result in a significant difference compared with the corresponding control.P< 0.05 was selected as the level decision for significant differenc

59、es.2.8.统计分许统计学分析使用SPSS软件进行。配对样本t检验用于测定特殊处理的样品是否具有显著性差异。选择水平P<0.05认为具有显著差异。3. Results and discussionThermal treatment is among the most popular ways of food processing. During heating, a complex array of chemical reactions takes place, which plays a pivotal role in determining the quality attributes (sensory characteristics, nutritional value and safety) of processed foods. While some compounds are destroyed during food processing, many more new compounds might be introduced into the food system. Some of these c