康 杰（美）

0 前言

在運動過程中，由于新陳代謝增加，氧利用率升高，從而導致高活性氧從線粒體滲漏[1]。除此之外，肌肉收縮本身也會激活磷脂酶A2，啟動一系列酶，從而引起活性物質(zhì)的增加[2]?；钚匝鯐淖兗毎Y構和功能，并導致肌肉損傷、免疫功能障礙和身體疲勞[3]。在過去的40年中，我們對運動所引起的氧化應激生物學意義的討論迅速增加?，F(xiàn)在我們認識到，雖然高水平的自由基會損傷細胞成分，但中低水平的氧化劑在細胞中發(fā)揮多種調(diào)節(jié)作用，如控制基因表達、調(diào)節(jié)細胞信號通路和調(diào)節(jié)骨骼肌力量輸出[4]，同時也可以刺激糖原再合成[5]、降低感染的風險[6]，甚至可以通過啟動和促進對訓練的適應性反應來提高運動成績[7-10]?；钚晕镔|(zhì)有害還是有益，取決于個體的運動持續(xù)時間、運動強度、身體屬性和營養(yǎng)狀況[11]。

無論是專業(yè)運動員還是運動愛好者，攝入抗氧化劑都是常見的做法。盡管并沒有證據(jù)證明其益處，但各種營養(yǎng)補充劑的市場依舊十分巨大[12]。事實上，抗氧化劑是專業(yè)運動員和運動愛好者最常使用的運動補充劑之一[13-14]。雖然這些產(chǎn)品已被吹捧為預防運動引起的氧化損傷和提高運動表現(xiàn)的手段，但對于其功效依舊缺乏證據(jù)。此外，一些研究表明抗氧化劑對受過訓練者的健康和運動表現(xiàn)有不良影響[15-16]。越來越多的證據(jù)表明，自由基在細胞中扮演重要的生理功能，并且抗氧化劑和自由基之間的平衡是獲得生理適應性的必要前提[17-20]。因此，我們有必要評估如何謹慎使用抗氧化劑，特別是在專業(yè)運動員中。

本綜述旨在提供研究證據(jù)，證明抗氧化劑在改善健康和運動表現(xiàn)方面的功效。文章開篇是有關活性物質(zhì)、抗氧化防御系統(tǒng)和運動誘導的氧化應激的概述。接下來是關于活性物質(zhì)在調(diào)節(jié)訓練導致的適應性中的作用以及抗氧化劑對運動表現(xiàn)影響的文獻綜述。文章最后還提供了有實際證據(jù)支撐的建議，從而幫助專業(yè)運動員或運動愛好者在補充抗氧化劑上作出明智的決定。

1 細胞內(nèi)主要活性物質(zhì)的產(chǎn)生

自由基一詞是指活性氧和氮類物質(zhì)，由于攜帶未配對的價電子而具有高活性。在動物肌肉纖維中，5種主要自由基具有生物學影響。第一種是過氧化物（O2-），在線粒體和細胞質(zhì)中形成，通過線粒體中電子傳遞鏈的少量氧分子提前釋放為O2-[21]。在黃嘌呤轉(zhuǎn)化為尿酸過程中，還原型輔酶Ⅰ，或黃嘌呤氧化酶（XO）也可在細胞外間隙形成O2-。XO主要存在于微血管內(nèi)皮細胞中，但也存在于白細胞中，劇烈運動后可能滲入肌纖維[22]。第二種是過氧化氫（H2O2），可以在XO轉(zhuǎn)化次黃嘌呤-黃嘌呤尿酸過程中釋放，也可以通過線粒體、胞質(zhì)溶膠和細胞外間隙中的過氧化物歧化酶（SOD）亞型由O2-形成[23-24]。第三種是羥基自由基（OH），通過O2-或H2O2與金屬離子如鐵或銅反應形成[24]。第四種是一氧化氮（NO），由L-精氨酸通過一氧化氮合酶（NOS）形成，主要是骨骼肌中的神經(jīng)元亞型（nNOS）與內(nèi)皮型 NOS（eNOS）[25-26]。 最后一種是過氧自由基過氧亞硝酸鹽（ONOO-），當O2-與NO反應時在胞質(zhì)溶膠中形成[27]。因為它們的起源是緊密相連的，運動時電子傳遞鏈和NOS的活化導致這5種自由基各自的數(shù)量增加。

底物消耗會導致谷胱甘肽還原酶活性下降，高溫會加速線粒體解偶聯(lián)。這兩者也可能促進運動過程中自由基的產(chǎn)生。此外，會導致酸中毒的無氧運動帶來的短暫缺氧，可能增加氧化應激反應[28]。最后，運動的機械應力本身，如外部沖擊，肌肉對骨骼的牽拉，肌肉的離心收縮和肌肉之間的摩擦等，也可以促進自由基的形成[29]。

2 活性物質(zhì)的利弊

細胞和細胞外空間暴露在來自外源和內(nèi)源的大量活性物質(zhì)中。外源性活性物質(zhì)來源包括氧氣、輻射、空氣污染物、異生素、藥物、酒精、重金屬、細菌、病毒、日光、食物和運動等。盡管如此，內(nèi)源性活性物質(zhì)來源更重要也更廣泛，因為在整段生命過程中它會持續(xù)產(chǎn)生。

作為正常代謝的一部分，所有需氧細胞都會產(chǎn)生活性物質(zhì)?；钚晕镔|(zhì)在疾病的發(fā)生、發(fā)展中發(fā)揮著重要作用[30]。活性氧和活性氮由于具有高活性，能夠使其他生物學層面上重要的分子發(fā)生變形，從而損傷細胞結構，阻礙細胞功能的實現(xiàn)。O2-、H2O2和OH能夠獲得不飽和脂肪酸中與雙鍵相鄰的質(zhì)子，如細胞膜中的質(zhì)子。于是這些脂肪酸形變的連鎖反應開始，形成脂質(zhì)過氧化物。這個過程稱為 “脂質(zhì)過氧化”，會導致細胞膜功能不良[24]。脂質(zhì)雙分子層的破壞改變了細胞膜的流動性和通透性，并可能導致膜結合蛋白活性降低[31]。NO可以氧化蛋白質(zhì)并改變其結構，從而損害其功能并影響基因轉(zhuǎn)錄[25,32-35]。同樣，OH、NO和ONOO-可以氧化核苷酸，損傷DNA，從而導致腫瘤的出現(xiàn)[36]。NO也被認為對肌纖維的收縮性有直接的抑制作用[37]。最后，氧化損傷也導致炎癥[38]和細胞凋亡[39]，并可能最終導致細胞功能下降。

雖然自由基一般只被看作是對細胞的威脅，但這種片面的想法也開始受到挑戰(zhàn)。越來越多的證據(jù)表明，自由基在調(diào)節(jié)肌肉適應過程中對氧化還原敏感的信號通路發(fā)揮重要作用[40]。最近幾項動物研究以及一些涉及運動員的研究提出了相關框架，涉及包括O2-、H2O2和NO在內(nèi)的活性物質(zhì)作為重要細胞信號的功能作用。有氧耐力訓練后，絲裂原活化蛋白激酶信號通路的活化可增強線粒體生物合成和毛細血管化（血管生成）、肌肉增殖和葡萄糖轉(zhuǎn)運能力[19,41]。人們已經(jīng)發(fā)現(xiàn)，這些對訓練的適應可能依賴于自由基引起的細胞氧化還原電位的改變[40]或O2-的短暫出現(xiàn)[17-18]，因為這些似乎刺激了該通路內(nèi)某些重要轉(zhuǎn)錄因子的上調(diào)。

另外，胞外間隙中由O2-形成的H2O2起到血管擴張劑的作用，可以優(yōu)化血流速度。一氧化氮合成酶在內(nèi)皮細胞中產(chǎn)生的NO也會帶來支持收縮肌的動脈血管舒張[42]，從而導致血流速度增加[43]。由此而引起的肌纖維微血管剪切應力的增加可刺激肌肉血管生成[44]。內(nèi)源性氧化劑防御，特別是O2-，也因活性氧的負反饋而上調(diào)[19,45]。

自由基也可能有急性積極作用。低濃度時，它們有助于維持肌肉力量輸出[40]。此外，在吞噬作用的氧爆過程中，巨噬細胞釋放 O2-、H2O2和 NO，作為清除受損或死亡細胞物質(zhì)的一部分，這有助于加快修復過程[46]。

3 抗氧化防御系統(tǒng)

為了對抗活性物質(zhì)，生命機體配備了高效的抗氧化防御系統(tǒng)。這些包括非酶、酶和膳食抗氧化劑。谷胱甘肽、尿酸、硫辛酸、膽紅素和輔酶Q10等都是非酶類抗氧化劑，這些抗氧化劑是內(nèi)源性的，通常是細胞代謝的副產(chǎn)物。主要的酶抗氧化劑是SOD，過氧化氫酶，谷胱甘肽過氧化物酶（GPX）和谷胱甘肽還原酶，而大多數(shù)已知的膳食抗氧化劑是生育酚（維生素 E）、抗壞血酸（維生素 C）和類胡蘿卜素（β-胡蘿卜素）。此外，各種多酚化合物近年來已被推廣為營養(yǎng)抗氧化劑。在涉及抗氧化劑的研究中，α-硫辛酸和藥物如N-乙酰半胱氨酸和別嘌呤醇也受到了評估。

在人體骨骼肌纖維中，幾種內(nèi)源性酶和底物共同作用從而清除自由基。SOD將O2-還原成H2O2。在細胞溶質(zhì)中，H2O2隨后可以通過GPX轉(zhuǎn)化為水，或通過硫氧還蛋白而形成過氧化物氧還酶。H2O2也可能通過過氧化氫酶轉(zhuǎn)化為水和分子氧[26]。二肽肌肽和絲氨酸也通過清除O2-和OH[47]起到抗氧化劑的作用。

不在人體內(nèi)合成的非酶促抗氧化劑，必須從外源獲得，包括維生素A（β-胡蘿卜素）、維生素C（抗壞血酸）和維生素 E（α-生育酚），這些維生素也被稱為膳食抗氧化劑。這些物質(zhì)能夠通過質(zhì)子捐贈清除各種自由基。維生素A屬于一組稱為類胡蘿卜素的紅色、橙色和黃色色素[48]。其他包括α-胡蘿卜素、β-隱黃質(zhì)、番茄紅素、葉黃素和玉米黃質(zhì)。β-胡蘿卜素是活性最強、最活躍的類胡蘿卜素，被食用后，它轉(zhuǎn)化為視黃醇，這是一種易于使用的維生素A。除了其維生素A原功能外，β-胡蘿卜素還被認為具有抗氧化特性[49]，并可能對免疫系統(tǒng)有積極影響[50]，且有抗癌作用[51]。維生素C是一種抗氧化劑，是人類一系列基本代謝反應（包括膠原合成）的輔助因子[52]。除了人類，這種水溶性維生素幾乎在所有生物中都是內(nèi)源性的?？箟难岬碾x子形式L-抗壞血酸是一種強還原劑，其氧化形式被酶和谷胱甘肽還原。維生素E是指包括生育酚和生育三烯酚的一組脂溶性化合物。α-生育酚是最具生物活性的形式，已被證明可以保護細胞免受脂質(zhì)過氧化[53-54]，并預防與氧化應激相關的慢性疾病[51,55]。其氧化形式可被其他抗氧化劑如維生素C、視黃醇、泛醇、谷胱甘肽、半胱氨酸和α-硫辛酸循環(huán)回活性形式[56]。

非酶促抗氧化劑還包括輔酶Q10、多酚、α-硫辛酸和N-乙酰半胱氨酸。輔酶Q10也稱為泛醌，是一種脂溶性維生素樣物質(zhì)，存在于大多數(shù)真核細胞中，主要存在于線粒體中[57]。它是電子傳遞鏈的一個組成部分，在細胞的能量產(chǎn)生中起著一定的作用。其還原形式泛醇，是體內(nèi)重要的抗氧化劑。多酚是一組水溶性植物源物質(zhì)，其特征是有一個以上的酚基[58]。對于已經(jīng)鑒定出的幾千種多酚，可以根據(jù)它們的結構和復雜性分成不同的類型，即黃酮類、木脂素類、芪類、香豆素類和單寧類。黃酮類化合物是最大的一類酚類化合物。水果和蔬菜中有豐富的多酚。例如，紅葡萄酒含有多種多酚類化合物，已被證明具有治療慢性疾病的藥理學特性[59-60]。α-硫辛酸是由辛酸衍生而來的有機硫化合物。它是4種線粒體酶復合物的重要輔助因子，因此，它是有氧代謝的關鍵。α-硫辛酸可能具有強大的抗氧化潛力，可以循環(huán)利用維生素E[61]；然而，它在組織中的積累是有限的。N-乙酰半胱氨酸是內(nèi)源性合成的抗氧化劑谷胱甘肽的副產(chǎn)物，它是一種半胱氨酸衍生物，在谷胱甘肽維持和代謝中發(fā)揮作用。鑒于其抗氧化特性，N-乙酰半胱氨酸已被用作營養(yǎng)補劑[43]。

內(nèi)源性和外源性抗氧化劑均可保護機體免受氧化應激的影響。更具體地說，酶和非酶抗氧化劑的協(xié)調(diào)網(wǎng)絡存在于胞內(nèi)和胞外，從而在自由基損傷蛋白質(zhì)、脂質(zhì)或DNA之前清除自由基。酶抗氧化劑是細胞蛋白質(zhì)，可以催化去除活性物質(zhì)從而防止氧化應激。非酶促抗氧化劑，如食物中所含的谷胱甘肽或抗氧化劑，可通過催化反應以外的方式消除自由基。為了最大程度免受自由基介導的損傷，酶促和非酶促抗氧化劑有策略地在整個細胞中將細胞區(qū)室化 （例如，細胞器vs.膜vs.胞質(zhì)溶膠）[4]?？寡趸瘎┖脱趸瘎醋杂苫┲g的平衡通常被稱為 “氧化還原平衡”，如圖1所示。氧化應激是抗氧化劑和氧化劑失衡的結果；這發(fā)生在自由基的產(chǎn)生超過抗氧化能力時。相反，當抗氧化能力大大超過自由基產(chǎn)生速率時，就會發(fā)生還原性應激。

圖1 自由基與抗氧化劑的關系Figure 1 Relationship between Radicals and Antioxidants

4 運動導致的氧化應激

在收縮過程中，骨骼肌是活性物質(zhì)的主要來源，也是其主要靶標之一[4]。運動使攝氧量（VO2）比靜息值高20倍[62]。在運動肌肉細胞的線粒體中，這意味著氧氣使用量增加了200倍[62]。20世紀70年代后期首次有了針對運動導致的氧化應激的描述，在運動的人的呼出氣體中[33]和運動的大鼠的組織[63]中發(fā)現(xiàn)脂質(zhì)過氧化產(chǎn)物的水平增加。1982年Davies等首次提供直接證據(jù)，證明高強度運動會顯著增加大鼠肌肉和肝臟自由基生成并造成線粒體膜損傷[64]。有人認為這可以同時對線粒體生物合成產(chǎn)生刺激作用。然而，大多數(shù)早期研究集中于氧化劑在肌肉中的破壞作用，并尋找抗氧化劑的潛在益處。

在過去的30年中，人們對運動相關的活性物質(zhì)的來源和后果的認識有了顯著提高。新出現(xiàn)的證據(jù)表明，肌肉收縮導致的自由基產(chǎn)生主要發(fā)生在肌肉的胞質(zhì)溶膠中，并且其數(shù)量多少受諸如環(huán)境條件、運動強度和持續(xù)時間等因素的影響[65-66]。具體而言，骨骼肌自由基的產(chǎn)生隨著運動強度和持續(xù)時間的增加而增加。此外，在炎熱的環(huán)境中和在高海拔地區(qū)（即約4 000 m）工作期間，收縮骨骼肌會產(chǎn)生更多的自由基[67-68]。因此，運動導致的肌肉自由基產(chǎn)生的幅度在很大程度上取決于運動條件。

雖然收縮骨骼肌會產(chǎn)生自由基，但運動并不總是會對骨骼肌造成氧化損傷。例如，低強度和短時間運動通常不會促進骨骼肌的氧化應激[4]。盡管如此，在中等強度到高強度下進行的長時間耐力運動往往會導致未經(jīng)訓練的個體的骨骼肌的氧化損傷。此外，重復性離心收縮，尤其是當運動員試圖適應新的運動強度時，會使骨骼肌承受相當大的壓力，可能會導致肌肉的氧化損傷[69-70]。損傷性運動也會誘發(fā)炎癥反應，進一步增加活性物質(zhì)的形成[71]。然而，這些研究往往缺乏關于受試者氧化還原狀態(tài)的信息，因此未能提供證據(jù)來證明活性物質(zhì)在肌肉損傷中所起到的作用。在訓練有素的耐力運動員的骨骼肌中，具有適應良好的內(nèi)源性抗氧化緩沖系統(tǒng)，可以抵抗運動引起的氧化應激[4]。因此，運動是否導致氧化應激取決于幾個因素，包括運動強度、持續(xù)時間以及個體的運動訓練狀態(tài)。

5 活性物質(zhì)和運動訓練適應

細胞適應自由基數(shù)量的增加，從而更能抵抗氧化應激的不利影響[72]。然而，必須強調(diào)的是，單次運動和定期運動的效果是完全不同的。定期運動能帶來許多有益的影響，身體適應氧化劑水平升高，而在單次劇烈運動時，適應性的變化是微不足道的。劇烈運動帶來的調(diào)整涉及增加血管舒張以增強血流和能量轉(zhuǎn)運，以及通過酶的變構活性發(fā)生動力學轉(zhuǎn)變，但這些可能不足以恢復氧化劑-抗氧化劑的動態(tài)平衡[20]。內(nèi)源性防御機制的長期刺激需要持續(xù)存在的生理刺激來維持一定程度的促氧化環(huán)境，并有效地使抗氧化系統(tǒng)超負荷[73]。在運動訓練，身體適應運動導致的氧化應激，并變得更能抵抗之后的氧化應激。這是通過多種不同的機制實現(xiàn)的，如上調(diào)氧化還原敏感基因表達和抗氧化酶水平[10,17]，增加酶活性[14,74]，刺激蛋白質(zhì)周轉(zhuǎn)[75]，改善DNA修復系統(tǒng)[76-77]，增加線粒體生物合成[8]和增加肌肉中熱休克蛋白的含量[78-79]。這些適應可以正面影響損傷后骨骼肌的重塑，并減少炎癥和細胞凋亡現(xiàn)象[20,80-81]。

中等水平的活性物質(zhì)似乎是各種生理過程所必需的，而過量的自由基產(chǎn)生會導致氧化損傷。這可以用激素的概念來描述，激素是一種劑量-反應關系，其中低劑量的物質(zhì)是刺激性的或有益的，高劑量是抑制性的或有毒性的[82]。為了適應活性物質(zhì)增加，線粒體的適應性反應也符合如此的毒物興奮效應，因此被稱為線粒體毒物興奮效應或線粒體效應[83]。活性物質(zhì)的激效作用，可能是定期運動對身體健康和運動表現(xiàn)有益的機制[82]。從活性物質(zhì)作為骨骼肌功能內(nèi)源性調(diào)節(jié)因子發(fā)揮的作用，就可見一斑?；钚晕镔|(zhì)是達到最佳收縮運動效果是必需的。肌肉肌絲，如肌球蛋白和肌鈣蛋白，以及肌質(zhì)網(wǎng)中的蛋白質(zhì)是對氧化還原敏感的，這使活性物質(zhì)具有改變肌肉收縮的能力[84]。

基于Reid關于氧化還原狀態(tài)對肌肉力量產(chǎn)生作用的模型，對活性物質(zhì)的反應可用鐘形曲線描述，如圖2所示[85-86]。在基線時，低水平的自由基對疲勞肌肉的收縮作用似乎不是很理想。來自Reid的研究數(shù)據(jù)表明，活性物質(zhì)的適度增加會導致肌肉力量增加，而抗氧化劑會消耗氧化劑水平并抑制力量。在較高的自由基濃度下，這種情況被逆轉(zhuǎn)，力量輸出因時間和劑量的增加而減少[85-87]。

圖2 自由基對肌肉性能的影響Figure 2 Effect of Free Radicals on Muscle Performance

6 抗氧化劑是否有益

運動員使用抗氧化劑，通常是為了防止運動引起的氧化應激的有害作用，加速肌肉功能的恢復，并提高運動能力[12-13,88-91]。目前，含有抗氧化劑的營養(yǎng)補劑無論在零售店還是從互聯(lián)網(wǎng)廠商都能廣泛購買。生產(chǎn)補充劑的公司提供的常見抗氧化劑包括維生素E、維生素C和β-胡蘿卜素。其他抗氧化產(chǎn)品包括葡萄提取物、白藜蘆醇、葉黃素、番茄紅素、硫辛酸、綠茶復合物等。研究表明，抗氧化劑的使用因地區(qū)和人群的不同而不同。盡管如此，世界各地的抗氧化劑使用率很高，一項研究報告稱，約62%的初級田徑運動員使用營養(yǎng)補劑，其中多種維生素和礦物質(zhì)是最受歡迎的[92]。

支持耐力運動員使用抗氧化劑的人認為，由于嚴格的運動訓練導致骨骼肌損傷性自由基產(chǎn)生增加，抗氧化劑對保護骨骼肌纖維免受氧化損傷至關重要。這一觀點得到了實驗證據(jù)的支持，其證明補充維生素C足以鈍化運動導致的自由基產(chǎn)生[93]。另一個支持使用抗氧化劑的論點是，許多耐力運動員的飲食缺乏抗氧化劑[94]。那些長期限制能量攝入、經(jīng)常從事體重控制或減肥性運動、刻意避免攝入某些食物類別或飲食不平衡的運動員，缺乏維生素的風險最大。只有少數(shù)膳食抗氧化劑有指定的推薦每日膳食供給量（RDA）。這些抗氧化劑食物的RDA包括：維生素C—男性為90 mg，女性為75 mg，維生素E—15 mg，硒—55 μg。因此，對于飲食中一種或多種抗氧化劑含量較少的運動員來說，補充抗氧化劑可能對他們有益，但是，建議在開始補充治療前咨詢營養(yǎng)師。

反對耐力運動員使用抗氧化劑的原因有以下幾點。首先，沒有證據(jù)表明運動導致的骨骼肌自由基對人體健康有害。定期運動可降低各種原因的死亡率，因此，運動導致的自由基增加似乎不太可能是不健康的[95]。此外，定期耐力運動訓練可促進肌纖維中酶促抗氧化劑的增加，從而改善內(nèi)源性保護，抵抗運動介導的氧化損傷[4]。因此，這種訓練導致的內(nèi)源性抗氧化劑的增加足以防止來自其他來源的氧化損傷。最后，如果耐力運動員保持營養(yǎng)均衡的等熱量飲食，他們應該不需要飲食以外的抗氧化劑。這些考慮因素已得到該領域和美國運動醫(yī)學學院專家的認可[96-97]。

也許對于耐力運動員使用抗氧化劑，最強烈的反對論點如下。首先，新的研究表明，抗氧化劑可以防止運動引起的骨骼肌適應[8,10]。令人信服的證據(jù)表明，運動導致的活性物質(zhì)的產(chǎn)生是促進包括抗氧化酶、線粒體蛋白和熱休克蛋白在內(nèi)的眾多骨骼肌蛋白表達的必需信號[4,65]。另一個反對運動員使用抗氧化劑的觀點是，目前的許多研究都不支持抗氧化劑補充對人體健康有益的觀點。例如，對68項隨機抗氧化劑試驗（共232 606名參與者）的薈萃分析得出結論，膳食補充β-胡蘿卜素、維生素A和維生素E不能改善健康狀況，并可能提高死亡率[98]。這份詳細報告的結論是，維生素C和硒對人類死亡率的作用尚不清楚，需要進一步研究才能提出建議。

7 研究證據(jù)——抗氧化劑作為運動增補劑

當調(diào)查抗氧化劑在運動表現(xiàn)中的作用時，結果普遍不一致，大多數(shù)研究稱其沒有益處。20世紀70年代初，Sharman等人的研究表明補充維生素E對青少年男性游泳運動員的耐力表現(xiàn)沒有有益的影響[99]。而且，與抗氧化劑組相比，安慰劑組在運動訓練中表現(xiàn)出更大的心肺功能改善作用，這可能是對于補充劑不利影響的首次報道。在隨后的研究中，維生素E在提高游泳運動員[100]、專業(yè)自行車運動員[101-103]、非耐力訓練男子[104]、大學運動員[105]和馬拉松運動員[106]的成績方面被證明無效。此外，在對從事有氧訓練的久坐老年人群的研究中，補充維生素E未能進一步提高受試者的身體機能指標[107]。輔酶Q10的補充劑對男性[108-110]的運動表現(xiàn)沒有任何明顯的影響，無論其年齡和訓練狀態(tài)如何。雖然有人假設多種抗氧化劑協(xié)同作用或許能更有效地對抗氧化應激，但事實上維生素E、維生素C、輔酶Q10和其他維生素和礦物質(zhì)的組合未能改善競技男子運動員[111]、自行車運動員[112-113]、鐵人三項運動員[114-115]、足球運動員[116-117]、抗阻訓練男子[118]、超耐力運動員[119]和中等訓練男子[120]的運動表現(xiàn)。

另一方面，已有多項研究顯示抗氧化劑對身體表現(xiàn)有積極但有限的影響。輔酶Q10可以提高專業(yè)越野滑雪運動員的最大攝氧量（VO2max）和有氧、無氧閾，從而增加運動能力和加快恢復速度[121]。同樣，在導致疲勞的運動試驗期間，無論是未經(jīng)訓練[122-123]還是受過訓練的個體[124-125]，補充輔酶Q10對運動表現(xiàn)、疲勞感覺和恢復都有有益的影響。維生素E補充劑也被證明對登山者在高海拔地區(qū)的表現(xiàn)[126]和雪橇狗的耐力表現(xiàn)[127]有益。在兩項早期研究中，補充維生素C可以提高未受過訓練的男性學生[128]和運動員[129]的運動能力。在Aguilo等人的一項研究中，補充維生素E、維生素C和β-胡蘿卜素組合后的男性運動員在最大運動試驗后表現(xiàn)出較低的血乳酸水平，運動訓練3個月后VO2max較對照組增加更多[130]。

已有多項調(diào)查顯示多酚具有增強運動表現(xiàn)的作用，包括槲皮素[31-34]、白藜蘆醇[135]，以及來自葡萄提取物[136]和甜菜根汁的多酚化合物[137-140]。新的證據(jù)表明，酚類化合物的抗氧化能力應該不是其保護作用的唯一機制，其保護作用也可以是它們與細胞信號級聯(lián)反應中的各種關鍵蛋白相互作用所介導的[141]。然而，這些發(fā)現(xiàn)遠未達成共識，很多研究的結果是相互矛盾。例如，槲皮素補充劑已被證明對久坐的人[142-143]或騎自行車的人[144]沒有增強運動表現(xiàn)的作用。另外還有研究發(fā)現(xiàn)，對于受電刺激等長收縮的小鼠，白藜蘆醇不能改善它們的肌力輸出和肌肉疲勞性[145]。有趣的是，在Marshall等人的一項研究中，維生素C被證明會減緩賽狗的速度[146]。

最近有人提出，在長時間亞極量運動時，N-乙酰半胱氨酸急性給藥可能會延緩人體肌肉疲勞。Medved等人研究了N-乙酰半胱氨酸對未經(jīng)訓練的男性的肌肉疲勞和運動表現(xiàn)的影響。雖然N-乙酰半胱氨酸在高強度間歇運動中表現(xiàn)出可以調(diào)節(jié)血液氧化還原狀態(tài)，但對于疲勞的出現(xiàn)時間沒有影響[147]。同一研究小組也觀察到，在一組混合訓練和未訓練但經(jīng)常運動的個體中，長時間運動時，N-乙酰半胱氨酸輸液對疲勞的出現(xiàn)時間沒有影響[148]。然而，在同一項研究中，抗氧化劑改善了血漿K+濃度的調(diào)節(jié)機制，并提出N-乙酰半胱氨酸的作用取決于個體的訓練狀態(tài)[148]。最后，據(jù)報道，長時間亞極量運動時N-乙酰半胱氨酸輸液可延后一組訓練有素的人疲勞的出現(xiàn)時間，這可能是通過增加肌肉半胱氨酸和谷胱甘肽的利用率而完成的[149]。攝入N-乙酰半胱氨酸帶來的副作用是會讓某些人覺得惡心。因此，對于那些在使用該補充劑時出現(xiàn)惡心癥狀的運動員，N-乙酰半胱氨酸可能不會改善他們的耐力表現(xiàn)。重要的是，補充N-乙酰半胱氨酸對于健康的長期影響仍然未知。

抗氧化劑在運動員中的普及導致了該領域出現(xiàn)大量小型研究。然而，這些研究在研究設計、運動方案、人群、補充方案和分析方法等方面差異很大，這使得這個問題仍然沒有定論。許多評估抗氧化劑對運動表現(xiàn)影響的研究質(zhì)量低，受試者人數(shù)少，其中一些研究沒有堅持高質(zhì)量的試驗要求（例如對照、雙盲和隨機化）。因此，將抗氧化劑作為一種有效的助劑，評估其功效時，需要謹慎。

8 研究證據(jù)——訓練中使用抗氧化劑面臨的干擾

最近，人們對高劑量外源性抗氧化劑如維生素C和維生素E在耐力訓練中的療效提出了疑問，一些研究表明這些抗氧化劑實際上可能會適得其反[18,20,41,111]。如前所述，人們一直認為活性氧對內(nèi)源性抗氧化系統(tǒng)的適應以及線粒體和血管生成起著重要的信號作用。當自由基生成被過度抑制時，這些信號可能因此被削弱或消除。

對與運動相關的氧化應激升高的一種反應是通過上調(diào)強大的抗氧化酶如SOD和GPX來加強氧化劑防御。然而，抗氧化劑可能會通過干擾自由基介導的信號來阻止這種適應[17,41]。尤其是運動員參與高強度訓練時，自由基的產(chǎn)生水平特別高。Knez等人的研究顯示，那些服用了抗氧化劑的運動員，在半鐵人三項或全鐵人三項運動后，其氧化損傷明顯大于未服用抗氧化劑的運動員，且能量生產(chǎn)系統(tǒng)在訓練過程中面臨同樣多的挑戰(zhàn)，會通過增加肌纖維的線粒體質(zhì)量、毛細血管密度以及改善底物的供應和利用，來增強運動能力[14]。在這里，動物和人類的對照研究也提供了強有力的證據(jù)，表明包括維生素C在內(nèi)的口服抗氧化劑可以干擾運動導致的信號傳導和隨后線粒體酶細胞色素C的表達，線粒體酶細胞色素C代表線粒體體積[8]，并可以改善胰島素敏感性[138]。此外，在參與耐力訓練計劃的個體中，急性補充維生素C（1g）和維生素E（600 IU）似乎可防止運動引起的血管舒張[150]，后者可鈍化血流帶來的血管生成刺激。如果eNOS的NO釋放被阻斷，血管生成也可以被阻止[44,151]。在 Gomez-Cabrera等人的研究中，對照組的人VO2max大約是每天攝入1 g維生素C的人的兩倍[8]。

另一個抗氧化劑干擾訓練的例子，出現(xiàn)在進行劇烈運動、不習慣的運動，特別是離心運動后的肌肉損傷時。維生素C和維生素E已被證明可延緩愈合和力量恢復，并在這種肌肉損傷性運動后增加氧化應激[36,152-154]。

總的來說，強勢的自由基清除行為實際上會通過抑制依賴自由基的適應信號來減少訓練刺激和有效性（圖3）。從希望通過訓練提高能力的運動員和教練員的角度來看，這些發(fā)現(xiàn)很有趣。難道很多人在不知不覺中通過普通的做法來抵消訓練效果，比如在耐力訓練后飲用富含抗氧化劑的恢復飲料或每天服用多種維生素。

圖3 目前關于運動訓練中補充抗氧化劑的共識Figure 3 Current Consensus on Antioxidant Supplementation during Exercise Training

9 實際意義和建議

關于補充劑的研究結果對營養(yǎng)學家、醫(yī)生、從業(yè)者、運動訓練師、教練和運動員以及普通人群都有重要意義。有證據(jù)表明高劑量抗氧化劑會排除運動訓練的健康促進作用并干擾自由基介導的生理適應，因此在使用抗氧化劑時要謹慎。以下建議是根據(jù)當前的研究證據(jù)制定的，可以指導那些想通過服用抗氧化劑以維持健康或增強運動表現(xiàn)的人。

（1）經(jīng)常運動的人需要優(yōu)化他們的營養(yǎng)結構，而不是使用補充劑。

（2）他們應通過食用多種水果、蔬菜、全谷物和堅果來獲得富含抗氧化劑的飲食結構。

（3）比起膠囊，全食中的抗氧化劑比例更優(yōu)，而且含有眾多的植物化學物質(zhì)，可以協(xié)同作用，從而優(yōu)化抗氧化劑的效果。

（4）抗氧化劑少量存在于食物中，因此，食用富含水果和蔬菜的飲食不太可能會導致抗氧化劑 “過量”。然而，如果通過膳食補充劑攝入大量抗氧化劑，便會有更大的中毒風險或者影響健康狀況。

（5）當個體有較高水平的氧化應激、能量攝入受限、從事大量減肥行為、飲食中去除了一個或多個食物群、飲食不平衡導致攝入微量營養(yǎng)素密度低時，可能需要補充抗氧化劑。

（6）在某些情況下，補充抗氧化劑可能是有利的，如過度訓練、肌肉損傷、比賽和高海拔訓練營，因為自由基的產(chǎn)生得到了強化，內(nèi)源性防御被削弱。

（7）普通膳食抗氧化劑（即維生素E和維生素C）已被證明不會改善運動表現(xiàn)或加速運動恢復。

（8）使用抗氧化劑N-乙酰半胱氨酸治療可改善亞極量運動時的人體運動表現(xiàn)。然而，N-乙酰半胱氨酸可能讓某些人出現(xiàn)惡心癥狀，補充N-乙酰半胱氨酸的長期影響仍然未知。

（9）在采用抗氧化劑方案之前，需要進行仔細的產(chǎn)品評估，該方案應該是有臨床監(jiān)督的，而且這只是短期的解決方案。

10 結論

運動促進肌肉中自由基的產(chǎn)生，長時間/劇烈運動會導致自由基產(chǎn)生與肌肉抗氧化劑之間的不平衡，從而導致氧化應激。為了防止自由基介導的損傷，肌肉細胞含有內(nèi)源性抗氧化劑來清除自由基。此外，從飲食中獲得的外源性抗氧化劑與內(nèi)源性抗氧化劑一起工作，形成一個細胞保護網(wǎng)絡來對抗自由基介導的氧化應激。運動員是否應該使用抗氧化劑仍然是一個重要且備受爭議的話題。目前，想向通過飲食攝入推薦營養(yǎng)素的運動員或經(jīng)常運動的人推薦抗氧化劑，可以依賴的科學證據(jù)很有限。事實上，高劑量的抗氧化劑可能會妨礙運動訓練對健康的促進作用，并干擾自由基介導的生理適應?？寡趸瘎┭a充劑通常不能改善運動成績，幾乎沒有證據(jù)證明它們在預防運動引起的肌肉損傷和增強恢復方面能起到作用。那些想增加抗氧化劑攝入量的人，應該考慮各種天然食品，而不是膠囊補充劑，并且應該意識到過量的抗氧化劑可能對健康和運動表現(xiàn)有害。

0 Introduction

During exercise,metabolism increases and oxygen utilization is elevated,leading to leakage of highly reac tive oxygen species from mitochondria[1].Aside from mitochondrial leakage,contraction itself activates phospholipase A2,initiating a cascade of enzymes and thereby increasing reactive species[2].Reactive oxygen species alter cell structure and function,and contribute to muscle damage,immune dysfunction,and fatigue[3].During the past four decades,our knowledge about the biological implications of exercise-induced oxidative stress has expanded rapidly.It is now appreciated that while high levels of free radicals can damage cellular components,low-to-moderate levels of oxidants play multiple regulatory roles in cells such as the control of gene expression,regulation of cell signaling pathways,and modulation of skeletal muscle force production[4].They can also be involved in stimulating glycogen re-synthesis[5],reducing susceptibility to the risk of infection[6],and they may even enhance athletic performance by initiating and promoting adaptive responses to training[7-10].The extent to which reactive species are damaging or helpful depends on the exercise duration,intensity,fitness attributes and nutritional status of the individual[11].

低溫配制工藝對虎杖膏中總二苯乙烯和總蒽醌含量的影響…………………………………………………… 王玉和等（21）：2911

Antioxidant supplementation is a common practice amongst both professional athletes and physically active individuals,and the market offering various nutrient supplements is immense despite the unclear evidence of their benefits[12].Indeed,antioxidants are among the most common sports supplements used by amateur and pro fessional athletes[13-14].Although these products have been touted as a means of preventing exercise-induced oxidative damage and enhancing performance,consistent evidence of their efficacy is lacking.Moreover,some studies suggest adverse effects of antioxidant supplementation on the health and performance of trained individuals[15-16].There is a growing body of evidence that the appearance of free radicals fulfils important physiological functions in cells,and that a balance between antioxidants and free radicals is necessary for desired physiological adaptations[17-20].Thus,it becomes necessary to evaluate the prudence of antioxidant supplementation,particularly among athletes.

This review is to provide research evidence with regard to the efficacy of using antioxidant supplementation in improving health and sports performance.The article begins with an overview of reactive species,antioxidant defense systems,and the exercise-induced oxidative stress.This is then followed by a review of literature concerning the role reactive species play in mediating training-induced adaptations and the effect of antioxidantsupplementation on exercise performance.The article also offers evidence-based recommendations that help athletes or those who are physically active in making a wise decision on antioxidant supplementation.

1 Production of Major Reactive Species in Cells

The term free radical refers to reactive oxygen and nitrogen species,which are highly reactive because of an unpaired valence electron.In animal muscle fibers,five main radicals have a biological impact.The first,superoxide (O2-),is formed in mitochondria and in the cytosol.A small amount of molecular oxygen passing through the electron transport chain in mitochondria is prematurely released as O2-[21]Superoxide can also be formed in the extracellular space by nicotinamide adenine dinucleotide phosphate hydrogen oxidase or by the enzyme xanthine oxidase(XO)during the conversion of xanthine to uric acid.XO is found mostly in microvascular endothelial cells,but is also present in leucocytes,which may infiltrate muscle fibers following strenuous exercise[22].The second,hydrogen peroxide(H2O2),can be released during the hypoxanthine→xanthine→uric acid conversion by XO,or it can be formed from O2-by superoxide dismutase(SOD)isoforms in mitochondria,cytosol,and the extracellular space[23-24].Third,the hy droxyl radical(·OH)is formed when O2-or H2O2reacts with metal ions such as iron or copper[24].The fourth radical,nitric oxide (NO·),is formed from L-arginine by nitric oxide synthase (NOS),mainly the neuronal isoform (nNOS)in skeletal muscle,but also endothelial NOS (eNOS)[25-26].Lastly,the peroxyl radical,peroxynitrite(ONOO-),is formed in the cytosol when O2-reacts with NO·[27].Because their origins are closely linked,increased activation of the electron transport chain and NOS during exercise leads to elevated production of each of these five radicals.

Substrate depletion,leading to a fall in glutathione reductase activity,and hyperthermia,which promotes mitochondrial uncoupling,may also contribute to free radical production during exercise.Furthermore,transient hypoxia during anaerobic exercise leading to acidosis may increase oxidative stress[28].Finally,mechanical stress of exercise,such as grinding,shearing,bending,and cutting,can itself increase free radical formation[29].

2 Negative and Beneficial Roles of Reactive Species

Cell s and extracellular spaces are exposed to a large variety of reactive species from both exogenous and endogenous sources.The exogenous sources include exposure to oxygen,radiation,air pollutants,xenobiotics,drugs,alcohol,heavy metals,bacteria,viruses,sunlight,food,and exercise.Nonetheless,exposure to endogenous sources is much more important and extensive because it is a continuous process during the life span.

Reactive species are generated by all aerobic cells as part of normal metabolism.The effect of reactive species plays an important role in the development of diseases[30].Because of their high reactivity,reactive oxygen species and reactive nitrogen species are able to deform other biologically important molecules,thus causing damage to cell structure and obstructing cell function.Superoxide,H2O2,and·OH are able to acquire the protons adjacent to double bonds in unsaturated fatty acids,such as those in cell membranes.This begins a chain reaction of deformation to these fatty acids forming lipid peroxides.This process,called“l(fā)ipid peroxidation”,results in poorly functioning cell membranes[24].The disruption of the lipid bilayer changes fluidity and permeability of the cell membrane and may lead to inactivity of membrane bound proteins[31].NO·can oxidize proteins and alter their structure,thereby impairing their function and affecting genetic transcription[25,32-35].Similarly,·OH,NO·,and ONOO-can oxidize nucleotides causing damage to DNA,which can lead to tumors[36].NO·has also been suggested to have a direct inhibitory effect on contractility in muscle fibers[37].Finally,oxidative damage also promotes inflammation[38]and apoptosis[39]and may eventually lead to decreased cellular functioning.

In addition,H2O2formed from O2-in the extracellular space acts as a vasodilator,which can optimize blood flow.NO·produced in endothelial cells by nitric oxide synthase also induces vasodilation in arteries that support the contracting muscle[42],leading to an increase in blood-flow velocity[43].The resulting increase in shear stress in the microvasculature of muscle fibers is an important stimulus for angiogenesis in muscle[44].Endoge nous oxidant-defense is also upregulated by negative feedback from reactive oxygen species,especially O2-[19,45].

Free radicals may also have acute positive effects.In low concentrations,they help maintain muscle force production[40].Furthermore,during the oxidative burst of phagocytosis,macrophages release O2-,H2O2,and NO·as part of the clearing out of damaged or dead cell material,which helps speed the repair process[46].

3 Antioxidant Defense Systems

To counter reactive species,the body is equipped with highly effective antioxidantdefense systems.These include non-enzymatic,enzymatic,and dietary antioxidants.Glutathione,uric acid,lipoic acid,bilirubin,and coenzyme Q10 are examples of non-enzymatic antioxidants that originate from endogenous sources and are often by-products of cellular metabolism.Principal enzymatic antioxidants are superoxide dismutase(SOD),catalase,glutathione peroxidase(GPX)and glutathione reductase,while most known examples of dietary antioxidants are tocopherols(vitamin E),ascorbic acid(vitamin C)and carotenoids(b-carotene).In addition,various polyphenolic com pounds have recently been promoted as nutrient antioxidants.α-Lipoic acid and pharmaceuticals such as N-acetylcysteine and allopurinol have also been evaluated in studies that involve antioxi-dant supplementation.

In human skeletal muscle fibers,several endogenous enzymes and substrates work together to scavenge free radicals.SOD reduces O2-to H2O2.In the cytosol,H2O2can thereafter be converted to water by glutathione peroxidase(GPX),which oxidizes glutathione(GSH),or one of several peroxiredoxins with the help of thioredoxin,or to water and molecular oxygen by catalase[26].The dipeptides carnosine and anserine also act as antioxidants by scavenging O2-and·OH[47].

Non-enzymatic antioxidants,which are not synthesized in humans,must be obtained exogenously,and include the vitamins A(b-carotene),C(ascorbic acid),and E(a-tocopherol),and these vitamins are also referred to as dietary antioxidants.These substances are able to scavenge various free radicals by proton donation.Vitamin A belongs to a group of red,orange and yellow pigments called carotenoids[48].Others include α-carotene,β-cryptoxanthin,lycopene,lutein,and zeaxanthin.β-Carotene is the most active carotenoid;after consumption it converts to retinol,a readily usable form of vitamin A.In addition to its provitamin A function,β-carotene is believed to have antioxidant properties[49]and may positively impact the immune system[50]and exhibit anticancerogenic effects[51].Vitamin C is an antioxidant and aco-factorin arangeof essential metabolic reactions in humans including collagen synthesis[52].This water-soluble vitamin is produced endogenously by almost all organisms except humans.L-ascorbate,an ion form of ascorbic acid,is a strong reducing agent and its oxidized form is reduced back by enzymes and glutathione.Vitamin E refers to a group of fat-soluble compounds that include tocopherols and tocotrienols.α-Tocopherol is the most biologically active form,and has been shown to protect the cells from lipid peroxidation[53-54]and to prevent chronic diseases associated with oxidative stress[51,55].Its oxidized form can be recycled back to the active form by other antioxidants,such as vitamin C,retinol,ubiquinol,glutathione,cysteine and a-lipoic acid[56].

Non-enzymatic antioxidants also include coenzyme Q10,polyphenols,α-Lipoic acid,and N-acetylcysteine.Coenzyme Q10,also known as ubiquinone,is a fat-soluble,vitamin-like substance,present in most eukaryotic cells,primarily in mitochondria[57].It is a component of the electron transport chain and plays a part in the energy production of a cell.Its reduced form,ubiquinol,acts as an important antioxidant in the body.Polyphenols are a group of water-soluble,plant-derived substances,characterized by the presence of more than one phenolic group[58].Several thousand polyphenols have been identified and they are divided into different groups according to their structure and complexity,i.e.,flavonoids,lignans,stilbenes,coumarins and tannins.Flavonoids are the largest group of phenolic compounds.Fruits and vegetables are a particularly rich source of polyphenols.For instance,red wine contains various polyphenolic compounds,which have been shown to possess pharmacological properties in the treatment of chronic diseases[59-60].α-Lipoic acid is an organosulfur compound derived from octanoic acid.It is an essential co-factor of the four mitochondrial enzyme complexes,therefore,is crucially involved in aerobic metabolism.α-Lipoic acid may have potent antioxidant potential and can recycle vitamin E[61];however,its accumulation in tissues is limited.N-acetylcysteine is a by-product of an endogenously synthesized antioxidant glutathione.It is a cysteine derivative and plays a role in glutathione maintenance and metabolism.Given its antioxidant property,N-acetylcysteine has been used as a nutritional supplement[43].

The body is protected against oxidative stress by both endogenous and exogenous antioxidants.More specifically,a coordinated network of enzymatic and non-enzymatic antioxidants exists in both the intracellular and extracellular locations to remove radicals before they damage proteins,lipids,or DNA.Enzymatic antioxidants are cellular proteins that catalytically remove reactive species to protect against oxidative stress.Non-enzymatic antioxidants,such as glutathione or antioxidants contained in food,can eliminate radicals by means other than a catalytic reaction.To provide optimal protection against radical-mediated damage,both enzymatic and non-enzymatic antioxidants are strategically compartmentalized(e.g.,organelles vs.membrane vs.cytosol)throughout the cell[4].The balance between antioxidants and oxidants (i.e.,radicals)is commonly referred to as “redox balance” and is illustrated in Figure 1.Oxidative stress results from an imbalance between antioxidants and oxidants;this occurs when radi-cal pro duction exceeds the antioxidant capacity.In contrast,reductive stress occurs when the antioxidant capacity greatly exceeds the rate of radical production.

4 Exercise-Induced Oxidative Stress

During contraction,skeletal muscle is a major source of reactive species,as well as one of the main targets[4].Exercise increases VO2by up to 20 times above resting values[62].In the mitochondria of exercising muscle cells,this translates to a 200-fold greater oxygen usage[62].Exercise induced oxidative stress was first described in the late 1970s when increased levels of lipid peroxidation products were found in the expired air of exercising humans[33]and the tissues of exercised rats[63].In 1982,Davies et al.[64]provided the first direct evidence that high-intensity exercise significantly increased radical production in the muscles and liver of rats and caused damage to mitochondrial membranes.It was suggested that this could,at the same time,deliver a stimulus to mitochondrial biogenesis.However,the majority of early studies focused on the damaging effects of oxidants in muscle and looked for the potential benefits of antioxidants.

Over the last 30 years,an understanding of the sources and consequences of exercise-related reactive species has advanced markedly.Emerging evidence indicates that contraction-induced radical production occurs primarily in the cytosol of the muscle and the magnitude of this production is influenced by factors,such as environmental conditions and the intensity and duration of exercise[65-66].Specifically,skeletal muscle radi cal production increases as a function of both the exercise intensity and duration. Moreover,contracting skeletal muscles produce more radicals during exercise in a hot environment and during work at high altitude(i.e.,～4 000 meters)[67-68].Therefore,the magnitude of exercise-induced muscle radical production can range widely depending upon the exercise conditions.

Although contracting skeletal muscles produce radicals,exercise bouts do not always result in oxidative damage to skeletal muscles.For example,low-intensity and short-duration exercise does not generally promote oxidative stress in skeletal muscles[4].Nonetheless,prolonged enduranceexerciseperformed atmoderateto-high intensities often results in oxidative damage to skeletal muscles of untrained individuals.In addition,repetitive eccentric con tractions,if unaccustomed in particular,place skeletal muscle under considerable stress that may cause muscle damage[69-70].Damaging exercise also induces an inflammatory response,which further increases formation of reactive species[71].However,these studies often lack the information about the subjects’redox status and therefore fail to provide evidence for the causal role of reactive species in muscle damage.Highly-trained endurance athletes have welladapted endogenous antioxidant buffer systems in their skeletal muscles that can resist exercise-induced oxida tive stress[4].Therefore,whether an exercise bout results in oxidative stress is dependent upon several factors,including the intensity and duration of exercise as well as the exercise training status of the individual.

5 Rea ctive Species and Exercise Training Adaptations

Cells adapt to increased free radicals production to become more resistant to the adverse effects of oxidative stress[72].It has to be emphasized,however,that the effects of a single bout of exercise and regular exercise are quite different.Regular physical activity brings about numerous beneficial effects and the body adapts to elevated oxidant levels,whilst with acute exercise,the adaptation is only marginal.Acute adjustment involves increased vasodilation to enhance blood flow and fuel transport and a kinetic shift via the allosteric activity of enzymes,which may not be sufficient to restore oxidant-antioxidant homeostasis[20].Long-term stimula tion of endogenous defense mechanisms requires the continuous presence of physiological stimuli that maintain a certain degree of pro-oxidative milieu,and effectively overload the antioxidant systems[73].With exer cise training,the body adapts to exercise-induced oxidative stress and becomes more resis tant to subsequent oxidative challenges.This is achieved through a number of different mechanisms,such as upregulation of redox-sensitive gene expression and antioxidant enzymes levels[10,17],an increase in enzyme activity[14,74],stimulation of protein turnover[75],improvement in DNA-repair systems[76-77],increased mitochondrial biogenesis[8],and increased muscle content of heat shock proteins[78-79].These adaptation can positively affects remodeling of skeletal muscle after injury and attenuate inflammation and apoptosis[20,80-81].

Moderate levels of reactive species appear necessary for various physiological processes,whereas an excessive radical production can cause oxidative damage.This may be described by the concept of hormesis,a dose-response relationship in which a low dose of a substance is stimulatory or beneficial and a high dose is inhibitory or toxic[82].The adaptive response of mito chondria to increased formation of reactive species is termed mitochondrial hormesis or mitohormesis[83].The hormetic action of reactive species could represent a mechanism underlying the health and performance benefits of regular physical activity[82].This can be seen in the role of reactive species as endogenous regulators of skeletal muscle function.Reactive species appear obligatory for optimal contractile activity.Muscle myofilaments,such as myosin and troponin,and proteins in the sarcoplasmic reticulum are redox-sensitive,which gives reactive species the ability to alter muscle contraction[84].

Based on Reid’s model for the role of redox state on muscle force production,responsesto reactive species can be described by a bell-shaped curve as shown in Figure 2[85-86].At baseline,low levels of free radicals appear to be suboptimal for the contraction of unfatigued muscle.The data from Reid’s studies suggest modest augmentation in reactive species causes muscle force to increase,while antioxidants deplete oxidant levels and depress force.At higher radical concentrations,this is reversed and force production decreases in a time-and dose-dependent manner[85-87].

6 Is Antioxidant Supplementation Beneficial?

It is common practice for athletes to use antioxidant supplements with the notion that they prevent the deleterious effects of exercise-induced oxidative stress,hasten recovery of muscle function,and improve performance[12-13,88-91].At the present,nutritional supplements containing antioxidants are widely available for purchase both in retail stores and from Internet vendors.Common antioxidants offered by supplement companies include vitamin E,vitamin C and β-carotene.Many other antioxidant products exist including grape extracts,resveratrol,lutein,lycopene,alpha lipoic acid,green tea complexes and numerous others.Studies re-veal that the incidence of antioxidant supplementation varies from country-to-country and across different segments of the population.Nonetheless,the use of antioxidant supplements is high around the world,as one study reported that～62%of junior track and field athletes use nutritional supplements,with multivitamins and minerals being the most popular[92].

Supporters of antioxidant supplementation for endurance athletes reason that because rigorous exercise training results in increased damaging radical production in skeletal muscles,antioxidant supplementation is essential to protect skeletal muscle fibers against oxidative damage.This notion is supported by experimental evidence demonstrating that vitamin C supplementation sufficiently blunts exercise-induced free radical production[93].Another argument used to support antioxidant supplementation is that many endurance athletes have diets that are deficient in antioxidants[94].Athletes who regularly restrict energy intake,have severe weight-loss practices,eliminate certain food groups or consume unbalanced diets are at the greatest risk for vitamin deficiency.Only a handful of dietary antioxidants have a designated Recommended Dietary Allowance(RDA).The RDAs for these include:vitamin C-90 mg for men and 75 mg for women,vitamin E-15 mg,and selenium-55 μg.Therefore,supplementation with an antioxidant could be beneficial for individuals who consume a diet low in one or more of these antioxidants;however,consultation with a dietitian before beginning a supplementation regimen is advised.

There are several arguments against antioxidant supplementation for endurance athletes.First,there is no evidence that exercise-induced radical production in skeletal muscle is harmful to human health.It is well established that regular exercise reduces all-cause mor tality and therefore,it seems unlikely that exercise-induced radical production is unhealthy[95].Further,regular endurance exercise training promotes increased enzymatic antioxidants in muscle fibers resulting in improved endogenous protection against exercise-mediated oxidative damage[4].Hence,this training-induced in crease in endogenous antioxidants may be adequate to protect against oxidative damage from other sources.Finally,if an endurance athlete maintains an isocaloric diet that is nutritionally well-balanced,it is likely that the individual does not need supplementary antioxidants above those consumed in the diet.These considerations have been acknowledged by experts in this field and American College of Sports Medicine[96-97].

Perhaps the strongest arguments against antioxidant supplementation for endurance athletes are the following.First,new studies reveal that antioxidant supplementation can prevent exercise-induced adaptations in skeletal muscle[8,10].Compelling evidence indicates that exercise-induced production of reactive species serves as a required signal to promote the expression of numerous skeletal muscle proteins including antioxidant enzymes,mitochondrial proteins,and heat shock proteins[4,65].Another argument against antioxidant supplementation in athletes is that much of the current research does not support the notion that antioxidant supplementation is beneficial to human health.For example,a meta-analysis of 68 randomized antioxidant supplement trials(total of 232 606 human participants)concluded that dietary supplementation with beta-carotene,vitamin A,and vitamin E does not improve health out comes and may increase mortality[98].This detailed report concluded that the roles of vitamin C and selenium on human mortality are unclear and require further study before a recommendation can be rendered.

7 Research Evidence-Antioxidant Supplements as Ergogenic Aids

There ha s been a general inconsistency of outcomes when investigating the role of antioxidant supplementation in exercise performance with the majority of the studies reporting no benefits.In the early 1970s,Sharman et al.[99]showed that supplementation with vitamin E had no beneficial effect on endurance perfor mance of adolescent male swimmers.Moreover,the placebo group demonstrated greater improvements of cardiorespiratory function with exercise training com pared with the antioxidant group,which may be the first report of the unfavorable effect of supplementation.In the studies that followed,vitamin E proved ineffective in improving performance in swimmers[100],profes sional cyclists[101-103],nonresistance-trained men[104],college athletes[105],and marathon runners[106].Furthermore,vitamin E supplements had no additive effect beyond that of aerobic training on indices of physical perfor mance in a group of older sedentary adults[107].Supplementation with coenzyme Q10 did not exhibit any significant effects on exercise performance of men[108-110])regardless of their age and training status.Despite the presumption that antioxidants work synergistically and may therefore be more efficient in combating oxidative stress,combinations of vitamins E,C,coenzyme Q10 and other vitamins and minerals failed to improve the exercise performance of competitive male runners[111],cyclists[112-113],triathletes[114-115],soccer players[116-117],resis tance-trained men[118],ultra-endurance runners[119],and moderately trained men[120].

On the other hand,there have been a number of studies showing positive but modest effects of antioxidant supplementation on physical performance.Coenzyme Q10 was associated with improved VO2maxand aerobic and anaerobic threshold of professional cross-country skiers that resulted in an increased exer cise capacity and a faster recovery rate[121].Similarly,supplementation with coenzyme Q10 indicated beneficial effects on performance,fatigue sensation,and recovery during fatigue-inducing exercise trials in both untrained volunteers[122-123]and trained individuals[124-125].Vitamin E supplementation was also shown to have a beneficial effect on the performance of climbers at high altitude[126]and endurance performance of sled dogs[127].In two early studies,supplementation with vitamin C was associated with an improved exercise capacity of untrained male students[128]and athletes[129].In a study by Aguilo et al.[130],male athletes supplemented with a combination of vitamin E,C and β-carotene exhibited lower blood lactate levels after a maximal exercise test and a greater increase in VO2maxafter 3 months of exercise training than the placebo group.

There have been a number of investigations showing the performance enhancing effects of polyphenols,including quercetin[31-34],resveratrol[135],and polyphenolic compounds from grape extract[136]and beetroot juice[137-140].Emerging evidence suggests that the antioxidant potential of phenolic compounds is unlikely to be the sole mechanism responsible fortheirprotective action,which could also be mediated by their interaction with various key proteins in the cell-signaling cascades[141].These findings,however,are far from reaching a consensus as there are studies showing conflicting results.Forexample,quercetin supplementation has been shown to have no ergogenic effects in sedentary individuals[142-143]or cyclists[144].It was also found that resveratrol did not improve muscle force output and muscle fatigability in mice subjected to electrically stimulated isometric contractions[145].Interestingly,in a study by Marshall et al.[146],vitamin C was shown to slow racing greyhounds.

More recently,it has been suggested that acute administration of N-acetylcysteine may delay human muscle fatigueduring prolonged submaximalexercise.Medved et al.[147]have studied the effect of N-acetylcysteine on muscle fatigue and performance in untrained men.Although N-acetylcysteine was shown to modulate blood redox status during high-intensity intermittent exercise,it did not affect time to fatigue.This same research group also observed no effect of NAC infusion on time to fatigue during prolonged exercise in a group of mixed trained and untrained but physically active in dividuals[148].In this same study,however,the antioxidant improved regulation of plasma K+concentration and it was suggested the ergogenic effect of N-acetylcysteine depends on an individual’s training status[148].Finally,N-acetylcysteine infusion during prolonged submaximal exercise was reported to augment time to fatigue in a group of well-trained individuals,possibly by increasing muscle cysteine and glutathione availability[149].A potential side effect is that the consumption of NAC can produce nausea in some individuals.Therefore,N-acetylcysteine supplementation may not improve endurance performance in those athletes who experience nausea when using this supplement.Importantly,the long-term health effects of supplementation with N-acetylcysteine remain unknown.

The popularity of antioxidant supplements with athletes has led to a plethora of small research studies in this area.However,these studies varied considerably in terms of research design,exercise protocol,population groups,supplementation regimen and analysis methods,which made this issue still remained inconclusive.Many of the studies evaluating the effects of antioxidants on exercise performance were of low quality with small subject numbers,and some of them did not adhere to all the accepted features of a high-quality trial(e.g.placebo-controlled,double-blind,and randomization).As such,caution is needed when evaluating the efficacy of using an antioxidant supplement as an ergogenic aid.

8 Research Evidence-Interferences of Antioxidant Supplementation with Training

Recently,questions have been raised about the efficacy of high doses of exogenous antioxidants such as vitamins C and E during endurance training,with several studies suggesting that these may actually be counterproductive[18,20,41,111].As mentioned earlier,it has been considered that reactive oxygen species play an important signaling role for adaptation of endogenous antioxidant systems and for mitochondrial genesis and angiogenesis.When radical appearance is overly suppressed,these signals may therefore be weakened or abolished.

One response to the elevated oxidative stress associated with exercise is increased oxidant defense via upregulation of powerful antioxidant enzymes like SOD and GPX.However,antioxidant supplementation may discourage such adaptations by interfering with the radical-mediated signal[17,41].This is especially the case when athletes are involved in high-intensity training during which radical production is particularly high.Knez et al.[14]reported significantly greater oxidative damage following half or full ironman triathlons in athletes who took antioxidant supplements than in those who did not.Similarly,the challenges faced by energy production systems during training stimulate enhance exercise capacity through increased mitochondrial mass and capillary density of muscle fibers and improved provision and utilization of substrate.Here,too,place bo-controlled studies with animals and humans have provided strong evidence that oral antioxidants including vitamin C can interfere with exercise-induced signaling and subsequent expression of the mitochondrial enzyme cytochrome c,which is representative of mito chondrial volume[8],and improvements to insulin sensitivity[138].Additionally,in humans involved in an endurance training program,acute supplementation of vitamins C(1 g)and E (600 IU)seemed to prevent exercise-induced va sodilation[150],which can blunt the blood flow-induced stimulus for angiogenesis.Angiogenesis can also be prevented if NO·release from eNOS is blocked[44,151].In the study by Gomez-Cabrera et al.[8],mean improve-ment in VO2max was about twice as great in humans who received a placebo than in those who received vitamin C at 1 g·day-1.

Another instance of antioxidant supplementation interfering with training is when muscle injury occurs,such as after intense,unaccustomed,and especially eccentric exercise.Vitamins C and E have been shown to delay healing and recovery of strength,and increase oxidative stress after such muscle-damaging exercise[36,152-154].

Collectively,it appears that over-dominant radical scavenging can actually reduce training stimuli and effectiveness by suppressing the radical-dependent signal for adaptation (Figure 3).Such findings are intriguing from the standpoint of athletes and coaches who wish to improve performance capacity through training.Could it be that many are unknowingly counteracting training effectiveness through ordinary practices such as consuming an antioxidant-rich recovery drink after an endurance training session or taking a daily multivitam in?

9 Practical Implications

The outcomes of supplementation studies have important implications for nutritionists,physicians,practitioners,athletic trainers,coaches,and athletes,as well as for the general population.Evidence that high doses of antioxidants preclude health-promoting effects of exercise training and interfere with radical-mediated physiological adaptations suggest caution in the use of antioxidant supplements.The following recommendations are developed based on the current research evidence and should help guide those who consider taking antioxidant supplements for maintaining health or enhancing performance:

（1）Physically active individuals need to optimize their nutrition rather than use supplements.

（2）Diets rich in antioxidants should be attained by consuming a variety of fruits,vegetables,whole grains,and nuts.

（3）Whole foods,rather than capsules,contain antioxidants presented in beneficial ratios and numerous phytochemicals that may act in synergy to optimize the effect of antioxidants.

（4）Antioxidants exist in small quantities in foods and therefore,there is limited risk of an antioxidant“overdose” by consuming a diet rich in fruits and vegetables.However,the ingestion of megadose of antioxidant via dietary supplements can increase the risk of toxicity and negative health consequences.

（5）Antioxidant supplementation may be warranted when individuals are exposed to high levels of oxidative stress,restrict their energy intake,use severe weight loss practices,eliminate one or more food groups from their diet,or consume unbalanced diets with low micronutrient density.

（6）There are certain circumstances in which an tioxidant supplementation is probably advantageous,such as overtraining,muscle injury,tournaments,competitions,and high-altitude training camps,since radical production is intensified and endogenous defense weakened.

（7）It has been demonstrated that supplementation with common dietary antioxidants(i.e.,vitamins E and C)does not improve exercise performance or accelerate recovery from exercise.

（8）Treatment with the antioxidant N-acetylcys teine has been shown to improve human exercise performance during submaximal exercise.However,N-acetylcysteine is associated with nausea in some individuals and the long-term effects of supplementation with N-acetylcysteine remains unknown.

（9）Careful product evaluation is required prior to adopting an antioxidant regimen,which should be clinically supervised and should only represent a short-term solution.

10 Conclusions

Exercise promotes radical production in the working muscles and prolonged/intense exercise can produce an imbalance between radical production and muscle antioxidants resulting in oxidative stress.To protect against the radical-mediated damage,muscle cells contain endogenous antioxidants to scavenge radi cals.Moreover,exogenous antioxidants obtained from the diet work with endogenous antioxidants to form a supportive network of cellular pro tection against radical-mediated oxidative stress.The question of whether or not athletes should use antioxidant supplements remains an important and highly debated topic.At present,there is limited scientific evidence to recommend antioxidant supplements to athletes or physically active individuals who consume the recommended nutrients through diet.In fact,high doses of antioxidants may preclude health-promoting effects of exercise training and interfere with radical-mediated physiological adaptations.Antioxidant supplements generally do not improve sports performance and there is little proof to support their role in prevention of exercise-induced muscle damage and enhance ment of recovery.Those who seek to augment their antioxidant intake should consider whole foods rather than capsules and should be aware of the fact that an overdose of antioxidants can be detrimental to health and performance.