REVIEW

Physiological Function of Lycopene and Its Application in Swine and Chicken Production

  • MEI Huadi , 1 ,
  • LI Yuanfei 1 ,
  • MA Xianyong 1, 2 ,
  • YU Miao , 1, 2, *
Expand
  • 1 Guangdong Engineering and Technology Research Center for Quality and Safety Control and Evaluation of Livestock and Poultry Meat, Guangdong Key Laboratory of Livestock and Poultry Breeding and Nutrition, State Key Laboratory of Livestock and Poultry Breeding, South China Key Laboratory of Animal Nutrition and Feed, Ministry of Agriculture, Institute of Animal Science, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
  • 2 Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming 525000, China
* associate professor, E-mail:

Received date: 2022-07-19

  Online published: 2023-02-13

Abstract

Lycopene is a kind of natural carotenoid, which has various physiological functions such as anti-oxidation, anti-inflammation, immunomodulatory and lipid-lowering, and has natural, environment-friendly and non-toxic characteristics. Lycopene has shown beneficial functions, such as improved performance, increased intestinal health and promoted meat quality in swine and chicken production research. Therefore, lycopene as a new type of feed additive, has broad development and application prospects. This paper summarized the physical and chemical properties, physiological functions, action mechanism and application of lycopene in swine and chicken production, providing theoretical references for the application of lycopene as a green feed additive in swine and chicken production.

Cite this article

MEI Huadi , LI Yuanfei , MA Xianyong , YU Miao . Physiological Function of Lycopene and Its Application in Swine and Chicken Production[J]. Chinese Journal of Animal Nutrition, 2023 , 35(2) : 727 -737 . DOI: 10.12418/CJAN2023.070

番茄红素是一种天然、无公害、无毒副作用的类胡萝卜素,主要存在于红色或橙色的水果和蔬菜中,如番茄、粉红葡萄柚、玫瑰果、木瓜等[1];它还存在于一些非红色或非橙色植物中,如芦笋和欧芹[2]。番茄红素凭借其来源天然,且具有多种生理功能,如抗氧化[3]、抗炎[4]、免疫调节[5]和调节机体代谢[6]等,在猪和鸡生产中的应用逐渐增多。在猪和鸡生产应用中表明,番茄红素具有改善其生产性能、肠道健康、肉品质和调节机体代谢等作用,因而具有良好的应用前景。因此,本文通过综述近些年国内外学者对番茄红素的研究,总结其生理功能、作用机制及其在猪和鸡生产中的应用,以期为番茄红素的饲料开发利用以及猪和鸡的生态健康养殖提供参考。

1 番茄红素的理化性质

番茄红素是脂肪烃类胡萝卜素,由8个异戊二烯单元组装而成,含有40个碳原子的碳链,其中包含11个共轭双键和2个非共轭双键,相对分子质量为536.85,熔点为173.5 ℃左右,以细长针状结晶体形式存在,为深红色粉末状,它还具有明显的亲脂性,使其易溶于有机溶剂,如氯仿、己烷、苯等,难溶或不溶于乙醇、甲醇和水,对光、温度、酸、催化剂等较为敏感[7]。在自然界中,番茄红素主要以全反式异构体的形式出现,通常称为全-E-番茄红素。在光、温度或化学反应的影响下,番茄红素可以从全反式异构体转化为单顺式或多顺式异构体[8]。因此,在西红柿中发现的大约90%的番茄红素以全反式异构体的形式存在,而在加工产品中多是以顺式异构体的形式存在[9]

2 番茄红素的生理功能及作用机制

2.1 抗氧化功能

正常生理条件下,自由基的产生和清除之间处于一种动态平衡,当出现氧化应激时,这种平衡就会被破坏,继而导致DNA、脂肪和蛋白质的损伤,最终使机体受到损害[10]。番茄红素由于其具有多不饱和结构,一方面可直接清除自由基,另一方面还可通过调控酶促抗氧化防御系统和酶促氧化损伤系统来间接清除自由基,从而避免机体受到氧化损伤。

2.1.1 直接清除自由基

番茄红素是自然界中一种极强的抗氧化剂,其清除单线态氧效率分别是维生素E的100倍和β-胡萝卜素的2倍[11]。番茄红素也可通过加成反应清除羟基自由基[12]。在体外和细胞内的研究发现,番茄红素还可以有效地与过氧亚硝酸盐反应,发挥过氧亚硝酸盐清除剂的作用[13-14]。另有研究表明,番茄红素还具有清除二氧化氮、硫酰及硫酰自由基的功能[15]。在极性或非极性条件下,番茄红素清除过氧化氢(H2O2)自由基的速度都要高于β-胡萝卜素[16]

2.1.2 调节抗氧化酶和氧化酶活性

番茄红素可通过调控酶促抗氧化防御系统和酶促氧化损伤系统,从而缓解机体的氧化损伤。在北京鸭雏鸭上的研究发现,番茄红素可通过提高肝脏中谷胱甘肽S-转移酶(glutathione S-transferase,GST)和过氧化氢酶(catalase,CAT)活性,同时提高肝脏总抗氧化能力(total antioxidant capacity,T-AOC),降低肝脏丙二醛(malondialdehyde,MDA)含量,进而缓解由黄曲霉毒素对肝脏诱导的氧化损伤[17]。Xia等[18]在高脂饮食小鼠模型中研究显示,番茄红素可通过增加血清T-AOC和超氧化物歧化酶(superoxide dismutase,SOD)活性,从而发挥抗氧化功能。番茄红素还可抑制由炎症引起的氧化应激。Li等[19]在葡聚糖硫酸钠诱导的溃疡性结肠炎小鼠模型中的研究表明,番茄红素可通过提高小鼠结肠中SOD、CAT、谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)活性和谷胱甘肽(glutathione,GSH)含量,并降低结肠过氧化物酶活性和MDA含量,继而发挥抗氧化作用,缓解炎症诱导的氧化应激水平。番茄红素还可通过抑制还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶(reduced nicotinamide adenine dinucleotide phosphate oxidase,NOX)活性,来减少活性氧(reactive oxygen species,ROS)生成,进而改善由乙醇和脂肪酸棕榈油酸引起的腺泡细胞氧化损伤[20]
核因子E2相关因子2(nuclear factor erythroid 2-related factor 2,Nrf2)是抗氧化系统中重要的转录因子,对激活抗氧化相关基因,如CATSODGSTGSH-Px至关重要[21-22]。Kelch样ECH关联蛋白1(kelch-like ECH-associated protein 1,Keap1)是一种Nrf2结合蛋白,作为Nrf2的负调节因子发挥作用,在正常情况下可防止Nrf2易位至细胞核[23]。但在氧化应激情况下,Keap1蛋白可迅速与Nrf2解偶联,促使Nrf2移位入核,并与细胞核内抗氧化反应元件(antioxidant response element,ARE)序列结合,进而增强抗氧化基因转录[24]。而番茄红素可通过上调ARENrf2表达,从而刺激Ⅱ相解毒抗氧化酶的产生,减少ROS对细胞的损害[25]。研究发现,番茄红素可直接或间接通过激活蛋白激酶B(protein kinase B,Akt/PKB)信号通路来诱导Nrf2基因转录,进而上调NAD(P)H:醌氧化还原酶1(NAD(P)H quinone reductase 1,NQO1)和血红素氧化酶-1(heme oxygenase-1,HO-1)的蛋白和mRNA表达水平,同时提高SOD、CAT和GSH-Px活性,最终缓解砷对小鼠肾脏造成的DNA损伤或抑制H2O2诱导的心肌细胞氧化损伤[26-27]。番茄红素还可通过促进磷酯酰肌醇-3-激酶(phosphatidylinositol 3-kinase,PI3K)/Akt/Nrf2信号转导,同时增加B细胞淋巴瘤/白血病-2(B-cell lymphoma/leukaemia-2,Bcl-2)的蛋白表达水平,继而保护M146L细胞免受氧化应激和凋亡[28]
综上所述,番茄红素一方面可直接与自由基结合,中断其链式反应;另一方面可通过激活Nrf2抗氧化信号通路,进而提高相关抗氧化酶的mRNA表达水平并提高其活性;同时降低氧化酶的活性,起到减少自由基生成并加快自由基清除的作用,最终维持细胞或机体的氧化平衡。

2.2 降脂功能

番茄红素可通过多种作用途径来减弱脂质的合成,同时加快脂质转运和动员,进而达到降脂效果。研究表明,番茄红素一方面可通过抑制信号转导及转录激活因子3(signal transducer and activator of transcription 3,STAT3)或激活腺苷酸激活蛋白激酶(adenosine 5’-monophosphate-activated protein kinase,AMPK)信号通路,来限制固醇调节元件结合蛋白-1(sterol regulatory element-binding protein-1,SREBP-1)、脂肪酸合酶(fatty acid synthase,FAS)和乙酰辅酶A羧化酶1(acetyl CoA carboxylase 1,ACC1)的基因表达,从而减少脂质的合成,最终降低小鼠肝脏或肉鸡血清总胆固醇(total cholesterol,TC)、甘油三酯(triglyceride,TG)和低密度脂蛋白(low density lipoprotein,LDL)含量[29-30];另一方面还可通过增强肉碱棕榈酰转移酶1、乙酰辅酶A氧化酶和过氧化物酶体增殖物激活受体(peroxisome proliferators-activated receptors,PPARα的mRNA表达水平,加强脂质的氧化分解,进而降低小鼠血清TG和非酯化脂肪酸含量[31]。还有研究发现,番茄红素可通过下调3-羟基-3-甲基戊二酰辅酶A还原酶(HMG-CoA reductase,HMGR)活性和前蛋白转化酶枯草杆菌蛋白酶/kexin 9型基因转录,来降低大鼠血浆高密度脂蛋白(high density lipoprotein,HDL)、TC和TG含量[32]。在巨噬细胞中,番茄红素还可通过抑制HMGR和LDL受体活性,从而抑制胆固醇的合成并减少LDL的细胞内吞,最终发挥降脂功能[33]。在种鸡中的研究显示,番茄红素可提高其肝脏PPARγ共激活因子1α(PPARγ coactlvator 1α,PGC1α)、PPARα、维甲酸X受体α(retinoid X receptor α,RXRα)和视黄酸受体α(retinotic acid receptor α,RARα)的基因表达,降低肝脏脂肪酸结合蛋白(fatty acids binding protein,FABP)1和FABP10的基因表达;提高空肠PPARγRXRαRXRγ以及十二指肠RARα的基因表达,降低空肠脂肪酸转运蛋白4的基因表达,这也是番茄红素调节脂肪代谢的重要机制之一[34]

2.3 抗癌功能

体内外的研究均发现,番茄红素具有较好的抗癌作用,可通过调节生长因子信号传导、细胞周期停滞和/或细胞凋亡诱导、转移和血管生成以及调节抗炎和抗氧化活性等,进而抑制前列腺癌、卵巢癌、乳腺癌、肝癌、肺癌等癌细胞的增殖[35-44]。大量证据表明,番茄摄入量和血液番茄红素含量与患前列腺癌的风险呈负相关[35]。另有研究发现,番茄红素可通过抑制正常大鼠前列腺组织中雄激素信号传导,并降低胰岛素样生长因子Ⅰ的mRNA表达水平,进而预防前列腺癌[36];番茄红素也可通过抑制人前列腺癌LNCaP细胞甲羟戊酸途径和损害Ras膜定位及其细胞信号传导,来促使细胞周期停滞在G0/G1期,并诱导细胞凋亡,继而治疗前列腺癌[37]。研究显示,番茄红素还可干扰卵巢癌的发生和发展,它通过上调Nrf2或下调核因子-κB(nuclear factor κB,NF-κB)和STAT3信号,从而在卵巢细胞中发挥抗炎和抗氧化功效,防止蛋鸡自发性卵巢癌的形成[38];还可通过下调整合素(integrin,ITG)β1、ITGα5、基质金属蛋白酶(matrix metalloproteinase,MMP)-9、黏着斑激酶、整合素连接激酶和上皮间质转化标记物的基因表达以及丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)活化,抑制小鼠卵巢原发性癌细胞和转移癌细胞增殖,并降低卵巢癌生物标记物CA125的水平,进而发挥其抗转移和抗增殖作用[39]。研究表明,番茄红素可分别通过抑制肺α7烟碱乙酰胆碱受体或NF-κB和细胞色素P450E1的蛋白表达水平,进而抑制烟草致癌物诱发的雪貂肺癌或肝癌细胞增殖[40]。番茄红素还可以抑制人肝癌SK-Hep-1细胞的侵袭、迁移和黏附,其机制可能是通过降低NOX的mRNA和蛋白表达水平以及MMP-9和MMP-2活性[41]。还有研究表明,番茄红素可通过上调p53和B细胞淋巴瘤/白血病-2相关X蛋白(B-cell lymphoma/leukaemia-2-associated X protein,Bax)的基因表达,来抑制人乳腺癌MCF-7细胞的增殖,并促进其细胞凋亡,最终起到抗癌作用[42]。除上述途径外,番茄红素还通过抑制人脐静脉内皮细胞迁移和血管内皮生长因子的活性,继而减少血管生成,有效预防癌症[43-44]

2.4 抗炎及免疫调节功能

番茄红素可通过上调和下调多种信号通路,限制炎症介质相关因子的转录和产生,进而改善炎症。在小鼠巨噬细胞RAW264.7中的研究显示,番茄红素可通过增强HO-1基因转录,抑制NF-κB和c-Jun氨基末端激酶信号通路的激活,来降低炎症酶环氧合酶-2(cyclooxygenase-2,COX-2)和一氧化氮合酶2的mRNA表达水平,同时降低肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素(interleukin,IL)-6和IL-1β的mRNA表达水平,从而减少炎症介质的产生,最终抑制脂多糖(lipopolysaccharide,LPS)诱导的细胞炎症[45-46]。在小鼠软骨细胞中的研究发现,番茄红素可通过激活Nrf2/HO-1信号通路,继而抑制IL-1β引起的NF-κB和STAT3途径的激活,限制TNF-αIL-6、COX-2和iNOS的基因转录,并降低TNF-α、IL-6和前列腺素含量,最终发挥抗炎作用[47]。番茄红素还可通过抑制MAPK和NF-κB的信号转导,同时刺激PPARγ的基因转录,继而减少IL-1β、IL-6、IL-8和TNF-α的分泌,最终改善氧化甾醇诱导的THP-1巨噬细胞炎症[48]。高迁移率族蛋白框1(high mobility group box 1,HMGB1)是一种非组蛋白DNA结合蛋白,由坏死细胞和暴露在促炎信号下的免疫细胞产生,通过吸引和激活炎症细胞和介质,并与晚期糖基化终末产物受体和Toll样受体结合,进而产生重要的促炎作用[49]。据报道,番茄红素可通过降低细胞黏附分子的基因表达,继而抑制HMGB1诱导的单核细胞黏附和迁移,减少LPS诱导的HMGB1释放、HMGB1介导的TNF-α和分泌型磷脂酶A2分泌,最终发挥抗炎功能[50]。有研究表明,番茄红素还可通过促进Nrf2/HO-1通路的激活,同时增强肝巨噬细胞自噬,来抑制核苷酸结合寡聚化结构域样受体蛋白3(nucleotide-binding oligomerization domain-like receptor protein 3,NLRP3)的活化,进而降低TNF-αIL-1β的mRNA表达水平,减轻小鼠急性肝损伤和改善细胞炎症[51]
番茄红素除了调节炎症介质外,还可增强机体细胞免疫和体液免疫。番茄红素具有保护淋巴细胞免受氧化损伤[52]、促进T淋巴细胞的正常分化、提高CD4+/CD8+比值[53]、增强自然杀伤(natural killer,NK)细胞活性等作用[54]。在卵巢癌大鼠模型中的研究报道,番茄红素可有效提高大鼠的免疫功能,其机制可能是通过促进T淋巴细胞转化,增加CD4+/CD8+比值,同时降低炎症细胞因子水平[55]。据报道,健康男性每天摄入330 mL番茄汁(37.0 mg/d番茄红素)会促进淋巴细胞增殖,并增强NK细胞的细胞毒性,从而发挥免疫调节功能[56]。还有研究发现,番茄红素可以调节小鼠T淋巴细胞干扰素-γ(interferon-γ,IFN-γ)和IL-2产生的潜力或间接激活T细胞[57]。据报道,补充富含番茄红素的饮食可增加小鼠脾B淋巴细胞的产生和血清免疫球蛋白(immunoglobulin,Ig)G含量,进而改善小鼠免疫功能[58]
综上可知,番茄红素具有较好的抗炎和免疫调节活性,一方面通过调节炎症相关因子的产生与释放;另一方面通过促进B、T淋巴细胞的产生、刺激T细胞的正常分化和增强自然杀伤(NK)细胞的活性等增强免疫力。

3 番茄红素在猪和鸡生产中的应用

番茄红素作为一种新型绿色添加剂和膳食补充剂,在猪和鸡生产中具有提高生产性能、改善肠道健康和产品品质等功能,具有较好的应用潜力。

3.1 番茄红素在猪生产中的应用

3.1.1 番茄红素对猪生产性能的影响

Sun等[59]研究发现,饲粮中添加50 mg/kg的番茄红素可提高母猪生产性能,提高母猪产活仔猪数,降低产死仔数,同时提高仔猪初生窝重和断奶窝重,这可能是由于番茄红素降低胎盘ROS、H2O2、IL-1β、IL-8和TNF-α含量及其基因表达,提高胎盘GSH-Px活性、分泌型免疫球蛋白A(secretory immunoglobulin A,SIgA)、IgG和IgM含量以及T-AOC,提高初乳中IgA、IgG和乳糖含量,从而改善了母猪的胎盘健康和乳汁成分,进而提高了母猪的生产性能。在“长白×约克夏×杜洛克”三元杂交育肥猪上的研究表明,饲粮中添加20 mg/kg的番茄红素4周后,育肥猪的体重(body weight,BW)和平均日增重(average daily gain,ADG)得到了提高[60]。然而也有研究发现,在“皮特兰×长白×大白”三元杂交育肥猪饲粮中添加12.5、25.0、37.5和50.0 mg/kg的番茄红素4周后,不影响其生产性能[61]。这可能是由于猪品种、饲养管理等因素的不同而产生不同的研究结果。由此可见,虽然番茄红素可提高母猪的生产性能,但是目前还需要进一步的试验来验证番茄红素对猪生产性能的积极效果。番茄红素对猪生产性能的影响见表1
表1 番茄红素对猪生产性能的影响

Table 1 Effects of lycopene on performance of swine

试验动物
Experimental
animals
饲养时间
Feeding
period
添加量
Addition
amount
对生产性能的影响(与对照组相比)
Effect on performance (compared with
control group)
参考文献
Reference
母猪
Sows
妊娠至
仔猪断奶
50 mg/kg 总产仔数、产活仔猪数、初生窝重、断奶窝重和
断奶仔猪数分别提高7.76%、12.37%、11.50%、
11.58%和11.63%,产死仔数降低66.67%
[59]
育肥猪
Fattening pigs
28 d 20 mg/kg 体重和平均日增重提高 [60]
育肥猪
Fattening pigs
28 d 12.5、25.0、37.5
和50 mg/kg
对生产性能无负面影响 [61]

3.1.2 番茄红素对猪的其他作用效果

饲粮中添加番茄红素还可具有改善肉品质、肠道健康及增强机体免疫力等作用。在“杜洛克×长白×约克夏”三元育肥猪中的研究表明,饲粮中添加200 mg/kg番茄红素可通过提高空肠绒隐比、CAT活性、封闭蛋白-1(Claudin-1)的mRNA和蛋白表达水平,降低十二指肠IL-1β、NF-κB、TNF-α和MDA含量,进而维护肠道结构及其屏障功能和改善肠道免疫反应,最终保持其肠道健康[62]。番茄红素可提高猪背最长肌的肌肉红度、肌内脂肪和粗蛋白质含量,降低肌肉亮度和黄度,进而改善育肥猪屠宰后猪肉的颜色、营养价值和多汁性;通过核因子E2相关因子1(nuclear factor erythroid 2-related factor 1,Nrf1)/钙/钙调蛋白依赖性蛋白激酶β轴激活AMPK/沉默调节蛋白1/PGC-1α信号通路,诱导背最长肌肌肉纤维从快肌纤维转化为慢肌纤维,进而提高其肉品质;还可通过增强背最长肌、血清和肝脏中抗氧化酶活性或基因表达,从而改善机体抗氧化功能[63]。在“皮特兰×长白×大白”三元育肥猪中的研究发现,随着饲粮中番茄红素水平(12.5、25.0、37.5和50 mg/kg)的增加,血浆白蛋白和血清抗牛血清白蛋白含量以及血液淋巴细胞数量呈线性增加[64-65];贮存期间猪腰最长肌的MDA含量呈线性降低,并能有效防止腰最长肌的脂肪氧化,降低猪腰最长肌解冻过程中的水分流失[66];降低血清TC和LDL含量以及LDL/HDL[61];进而增强机体免疫功能,改善肌肉氧化稳定性和肌肉品质,调节脂质代谢。在猪胚胎发育的体外培养中,0.1 μmol/L番茄红素可通过调节氧化应激和线粒体依赖的细胞凋亡来促进猪的胚胎发育[67]。还有研究表明,一定浓度的番茄红素可提高猪精子冷冻稀释液解冻后的精子质量,其中最佳添加水平为2.0 μmol/L[68]
综上所述,饲粮中添加一定浓度番茄红素可能通过改善母猪乳汁质量以及胎盘、肠道和机体健康,进而提高猪生产性能,但大多集中在育肥猪中,因此应进一步探究番茄红素在猪其他生长阶段的作用效果,并确定其在猪饲粮中的适宜添加量,以便番茄红素可更好的应用在猪生产中。

3.2 番茄红素在鸡生产中的应用

3.2.1 番茄红素对鸡生产性能的影响

研究显示,爱拔益加肉鸡饲粮中添加100 mg/kg番茄红素,可提高肉鸡BW和采食量(feed intake,FI),降低F/G[30]。在罗斯肉鸡饲粮中添加100 mg/kg番茄红素,可提高体增重(body weight gain,BWG),降低F/G[69]。番茄红素还对饲粮中添加黄曲霉毒素B1(aflatoxin B1,AFB1)的肉鸡生长具有改善作用,番茄红素(100、200和400 mg/kg)可使降低的ADG恢复50%~70%[70],推测可能与番茄红素提高了肠道消化酶活性以及改善肠道氧化还原平衡、肠道免疫和肠黏膜结构及屏障功能有关[71]。在海兰褐蛋鸡中的研究显示,饲喂添加10和20 mg/kg番茄红素饲粮,产蛋重和产蛋率均降低[72]。另有研究报道,罗曼白蛋鸡饲粮中补充20 mg/kg番茄红素,可提高产蛋重,而产蛋率并未降低[73]。以上结果的差异可能是由于不同蛋鸡品种引起的。同时,种鸡饲粮中添加40 mg/kg番茄红素,不仅可提高雏鸡初生BW,而且还可提高整个试验期间种鸡的鸡蛋受精率和孵化率,其机制可能是番茄红素提高了雏鸡十二指肠、空肠和回肠的绒毛高度及免疫器官指数和种鸡的抗氧化能力[74-75]。番茄红素对鸡生产性能的影响见表2
表2 番茄红素对鸡生产性能的影响

Table 2 Effects of lycopene on performance of chickens

试验动物
Experimental
animals
饲养阶段
Feeding
stage
添加量
Addition
amount
对生产性能的影响(与对照组相比)
Effect on performance (compared with
control group)
参考文献
Reference
爱拔益加肉鸡
AA broilers
1~42日龄 100、200和
400 mg/kg
100 mg/kg添加组体重和采食
量提高,料重比降低
[30]
爱拔益加肉鸡
AA broilers
1~42日龄 100、200和
400 mg/kg
100、200和400 mg/kg添加组平均日增重提高,
200和400 mg/kg添加组料重比降低
[70]
罗斯肉鸡
Ross broilers
21~84日龄 50、100和
200 mg/kg
100和200 mg/kg添加组体增重
提高,料重比降低
[69]
海兰褐蛋鸡
Hy-Line brown laying hens
38~42周龄 10和
20 mg/kg
10和20 mg添加组产蛋重分别降低3.95%和
3.49%,产蛋率分别降低2.47%和4.16%
[72]
罗曼白蛋鸡
Roman white laying hens
20~32周龄 20 mg/kg 平均蛋重提高3.86% [73]
兴华种鸡
Xinghua breeding chickens
26~31周龄 40 mg/kg 雏鸡初生体重提高1.38% [74]
兴华种鸡
Xinghua breeding chickens
26~31周龄 40 mg/kg 鸡蛋受精率和孵化率分别提高
3.80%和4.69%
[75]

3.2.2 番茄红素对鸡的其他作用效果

饲粮中添加番茄红素可有效改善鸡的肉蛋品质、机体代谢等。在爱拔益加肉鸡中的研究表明,100 mg/kg番茄红素可降低腹脂重和腹脂率,分别降低了21.58%和22.63%,同时还降低血清TG、TC和LDL含量,其机制可能是通过提高肝脏AMPK-α基因表达,降低肝脏SREBP-1、FASACC基因表达,从而减少脂质合成,加快脂质分解[30]。在对罗斯肉鸡的研究中发现,饲粮添加100和200 mg/kg番茄红素可提高血清葡萄糖和HDL含量以及血清CAT和GSH-Px活性,降低血清谷丙转氨酶和碱性磷酸酶活性[69];饲粮添加12 mg/kg番茄红素可提高腿肌多不饱和脂肪酸和α-生育酚含量,降低腿肌多不饱和脂肪酸过氧化指数[76];添加75 mg/kg的番茄红素还可降低腿肌胆固醇和MDA含量[77];这些研究表明,饲粮中添加一定浓度的番茄红素可调节肉鸡糖脂代谢,降低脂质沉积,提高机体抗氧化能力,增强肌肉氧化稳定性和改善肉品质,并对肝脏具有一定的积极影响。
在罗曼白蛋鸡研究中,饲粮添加20 mg/kg番茄红素可提高蛋黄颜色、蛋黄重、蛋重和蛋黄比例,还可通过降低肠道甾醇转运蛋白、微体三酰甘油转运蛋白、胆固醇酰基转移酶2、肝脏SREBP1、三磷酸腺苷柠檬酸裂解酶和肝X受体α的基因表达,增加肝脏ATP结合转运体G亚家族成员5/8蛋白基因表达,进而降低蛋黄胆固醇含量,最终改善鸡蛋品质[73]。给处于产蛋高峰期的海兰W36蛋鸡饲粮中添加20 mg/kg番茄红素,90 d后发现可降低蛋黄亚麻油酸和花生烯酸含量以及ω3/ω6多不饱和脂肪酸比值,饲粮添加40 mg/kg番茄红素还会降低蛋黄总多不饱和脂肪酸含量[78],这说明番茄红素具有调节蛋黄脂质分布的能力。饲粮中添加10 mg/kg番茄红素可通过增强肝脏PPARγSREBP-2、FABP4和脂蛋白脂酶基因表达,降低肝脏葡萄糖转运蛋白2/8基因表达,提高肝脏IL-6基因表达,进而调节白来航蛋鸡机体糖代谢、脂代谢和免疫反应[79]
由此可见,番茄红素在鸡生产中取得了良好进展,可通过多种途径有效改善机体健康状况,并不同程度提高肉鸡生长性能和种鸡繁殖性能,但对蛋鸡产蛋性能的积极效果有待考证,还需要加大番茄红素在蛋鸡上的应用,探究其应用效果。

4 小结

在无抗养殖背景下,番茄红素因其具有天然、无公害、无毒副等特点,同时具有抗炎、抗氧化、免疫调节等生理功能,使它成为一种新型的绿色饲料添加剂,逐渐应用于猪和鸡生产中。在鸡生产方面,饲粮中添加一定浓度的番茄红素,在不同品种、不同生理状况和生长阶段下均可促进其生长、改善肠道健康等,还可提高种鸡繁殖性能,但对蛋鸡产蛋性能的积极效果有待考证。在猪生产方面,饲粮中一定浓度番茄红素可提高母猪繁殖性能,改善母猪乳汁成分和胎盘健康,还可改善育肥猪肉品质、肠道和机体健康等,但在猪其他生长阶段的研究比较缺乏,且对猪生长性能的效果也有待进一步研究。
综上所述,番茄红素在猪和鸡生产中的研究还相对有限,不够系统和深入,在其他畜禽动物上的研究也比较缺乏,在不同种类的畜禽饲粮中的适宜添加量也不明确,且其作用机制和靶点也有待深入探究。因此,未来可加强其在猪不同生理状况和生长阶段下的研究以及蛋鸡生产中的应用研究,探究其适宜添加量,揭示其具体作用机制,这对促进番茄红素的饲料开发以及畜禽生态健康养殖的良性循环发展具有重大意义。
[1]
SAINI R K, A BEKHIT A E D, ROOHINEJAD S, et al. Chemical stability of lycopene in processed products:a review of the effects of processing methods and modern preservation strategies[J]. Journal of Agricultural and Food Chemistry, 2020, 68(3):712-726.

DOI

[2]
HEDAYATI N, NAEINI M B, NEZAMI A, et al. Protective effect of lycopene against chemical and natural toxins:a review[J]. BioFactors, 2019, 45(1):5-23.

DOI

[3]
CAO L, ZHAO J, MA L, et al. Lycopene attenuates zearalenone-induced oxidative damage of piglet sertoli cells through the nuclear factor erythroid-2 related factor 2 signaling pathway[J]. Ecotoxicology and Environmental Safety, 2021, 225:112737.

DOI

[4]
XU Z G, LIU C, WANG R, et al. A combination of lycopene and human amniotic epithelial cells can ameliorate cognitive deficits and suppress neuroinflammatory signaling by choroid plexus in Alzheimer’s disease rat[J]. The Journal of Nutritional Biochemistry, 2021, 88:108558.

DOI

[5]
FALLAH R, KIANI A, KHALDARI M. Supplementing lycopene combined with corn improves circulating IgG concentration in pregnant ewes and their lambs[J]. Tropical Animal Health and Production, 2021, 53(3):360.

DOI PMID

[6]
WANG J, GENG T H, ZOU Q H, et al. Lycopene prevents lipid accumulation in hepatocytes by stimulating PPARα and improving mitochondrial function[J]. Journal of Functional Foods, 2020, 67:103857.

DOI

[7]
SHI J, MAZZA G, LE MAGUER M, Lycopene from tomatoes[M]//SHI J,MAZZA G,LE MAGUER M.Functional foods:biochemical and processing aspects. Boca Raton: CRC Press, 2002:135-167.

[8]
SHI J, LE MAGUER M, BRYAN M, et al. Kinetics of lycopene degradation in tomato puree by heat and light irradiation[J]. Journal of Food Process Engineering, 2003, 25(6):485-498.

DOI

[9]
XIANQUAN S, SHI J, KAKUDA Y, et al. Stability of lycopene during food processing and storage[J]. Journal of Medicinal Food, 2005, 8(4):413-422.

DOI PMID

[10]
HALLIWELL B. Biochemistry of oxidative stress[J]. Biochemical Society Transactions, 2007, 35(5):1147-1150.

DOI

[11]
SIES H, STAHL W. Vitamins E and C,beta-carotene,and other carotenoids as antioxidants[J]. The American Journal of Clinical Nutrition, 1995, 62(6 Suppl):1315S-1321S.

DOI

[12]
PRASAD A K, MISHRA P C. Modeling the mechanism of action of lycopene as a hydroxyl radical scavenger[J]. Journal of Molecular Modeling, 2014, 20(5):2233.

DOI PMID

[13]
PANASENKO O M, SHAROV V S, BRIVIBA K, et al. Interaction of peroxynitrite with carotenoids in human low density lipoproteins[J]. Archives of Biochemistry and Biophysics, 2000, 373(1):302-305.

PMID

[14]
MUZANDU K, ISHIZUKA M, SAKAMOTO K Q, et al. Effect of lycopene and β-carotene on peroxynitrite-mediated cellular modifications[J]. Toxicology and Applied Pharmacology, 2006, 215(3):330-340.

DOI

[15]
MORTENSEN A, SKIBSTED L H. Relative stability of carotenoid radical cations and homologue tocopheroxyl radicals.A real time kinetic study of antioxidant hierarchy[J]. FEBS Letters, 1997, 417(3):261-266.

DOI

[16]
GALANO A, FRANCISCO-MARQUEZ M. Reactions of OOH radical with β-carotene,lycopene,and torulene:hydrogen atom transfer and adduct formation mechanisms[J]. The Journal of Physical Chemistry B, 2009, 113(32):11338-11345.

DOI

[17]
EL-SHESHTAWY S M, EL-ZOGHBY A F, SHAWKY N A, et al. Aflatoxicosis in pekin duckling and the effects of treatments with lycopene and silymarin[J]. Veterinary World, 2021, 14(3):788-793.

DOI PMID

[18]
XIA B K, ZHU R Y, ZHANG H, et al. Lycopene improves bone quality and regulates AGE/RAGE/NF-кB signaling pathway in high-fat diet-induced obese mice[J]. Oxidative Medicine and Cellular Longevity, 2022, 2022:3697067.

DOI PMID

[19]
LI Y P, PAN X, YIN M Y, et al. Preventive effect of lycopene in dextran sulfate sodium-induced ulcerative colitis mice through the regulation of TLR4/TRIF/NF-κB signaling pathway and tight junctions[J]. Journal of Agricultural and Food Chemistry, 2021, 69(45):13500-13509.

DOI PMID

[20]
LEE J, LIM J W, KIM H. Lycopene inhibits oxidative stress-mediated inflammatory responses in ethanol/palmitoleic acid-stimulated pancreatic acinar AR42J cells[J]. International Journal of Molecular Sciences, 2021, 22(4):2101.

DOI

[21]
CHEN S R, ZOU L Y, LI L, et al. The protective effect of glycyrrhetinic acid on carbon tetrachloride-induced chronic liver fibrosis in mice via upregulation of Nrf2[J]. PLoS One, 2013, 8(1):e53662.

DOI

[22]
XU M, CHEN X L, HUANG Z Q, et al. Effects of dietary grape seed proanthocyanidin extract supplementation on meat quality,muscle fiber characteristics and antioxidant capacity of finishing pigs[J]. Food Chemistry, 2022, 367:130781.

DOI

[23]
LIANG M C, WANG Z X, LI H, et al. L-arginine induces antioxidant response to prevent oxidative stress via stimulation of glutathione synthesis and activation of Nrf2 pathway[J]. Food and Chemical Toxicology, 2018, 115:315-328.

DOI PMID

[24]
DE HAAN J B. Nrf2 activators as attractive therapeutics for diabetic nephropathy[J]. Diabetes, 2011, 60(11):2683-2684.

DOI PMID

[25]
PALOZZA P, CATALANO A, SIMONE R, et al. Lycopene as a guardian of redox signalling[J]. Acta Biochimica Polonica, 2012, 59(1):21-25.

PMID

[26]
RAMADAN S S, ALMEER R, ALBASHER G, et al. Lycopene mitigates arsenic-induced nephrotoxicity with activation of the Nrf2 pathway in mice[J]. Toxin Reviews, 2022, 41(2):446-456.

DOI

[27]
ZHENG S L, DENG Z Y, CHEN F, et al. Synergistic antioxidant effects of petunidin and lycopene in H9c2 cells submitted to hydrogen peroxide:role of Akt/Nrf2 pathway[J]. Journal of Food Science, 2020, 85(6):1752-1763.

DOI

[28]
FANG Y C, OU S S, WU T, et al. Lycopene alleviates oxidative stress via the PI3K/Akt/Nrf2pathway in a cell model of Alzheimer’s disease[J]. PeerJ, 2020, 8:e9308.

DOI

[29]
ZENG Z H, HE W, JIA Z, et al. Lycopene improves insulin sensitivity through inhibition of STAT3/Srebp-1c-mediated lipid accumulation and inflammation in mice fed a high-fat diet[J]. Experimental and Clinical Endocrinology & Diabetes, 2017, 125(9):610-617.

[30]
WAN X L, YANG Z F, JI H R, et al. Effects of lycopene on abdominal fat deposition,serum lipids levels and hepatic lipid metabolism-related enzymes in broiler chickens[J]. Animal Bioscience, 2021, 34(3):385-392.

DOI

[31]
FENNI S, HAMMOU H, ASTIER J, et al. Lycopene and tomato powder supplementation similarly inhibit high-fat diet induced obesity,inflammatory response,and associated metabolic disorders[J]. Molecular Nutrition & Food Research, 2017, 61(9):1601083.

[32]
SULTAN ALVI S, ANSARI I A, KHAN I, et al. Potential role of lycopene in targeting proprotein convertase subtilisin/kexin type-9 to combat hypercholesterolemia[J]. Free Radical Biology and Medicine, 2017, 108:394-403.

DOI PMID

[33]
FUHRMAN B, ELIS A, AVIRAM M. Hypocholesterolemic effect of lycopene and β-carotene is related to suppression of cholesterol synthesis and augmentation of LDL receptor activity in macrophages[J]. Biochemical and Biophysical Research Communications, 1997, 233(3):658-662.

DOI

[34]
TIAN H C, LIU G B, GUO Y Q, et al. Lycopene supplementation regulates the gene expression profile and fat metabolism of breeding hens[J]. Journal of Animal Physiology and Animal Nutrition, 2020, 104(3):936-945.

DOI PMID

[35]
ROWLES J L, RANARD K M, APPLEGATE C C, et al. Processed and raw tomato consumption and risk of prostate cancer:a systematic review and dose-response meta-analysis[J]. Prostate Cancer and Prostatic Diseases, 2018, 21(3):319-336.

DOI

[36]
HERZOG A, SILER U, SPITZER V, et al. Lycopene reduced gene expression of steroid targets and inflammatory markers in normal rat prostate[J]. The FASEB Journal, 2005, 19(2):272-274.

[37]
PALOZZA P, COLANGELO M, SIMONE R, et al. Lycopene induces cell growth inhibition by altering mevalonate pathway and Ras signaling in cancer cell lines[J]. Carcinogenesis, 2010, 31(10):1813-1821.

DOI PMID

[38]
SAHIN K, YENICE E, TUZCU M, et al. Lycopene protects against spontaneous ovarian cancer formation in laying hens[J]. Journal of Cancer Prevention, 2018, 23(1):25-36.

DOI PMID

[39]
HOLZAPFEL N P, SHOKOOHMAND A, WAGNER F, et al. Lycopene reduces ovarian tumor growth and intraperitoneal metastatic load[J]. American Journal of Cancer Research, 2017, 7(6):1322-1336.

PMID

[40]
AIZAWA K, LIU C, TANG S Y, et al. Tobacco carcinogen induces both lung cancer and non-alcoholic steatohepatitis and hepatocellular carcinomas in ferrets which can be attenuated by lycopene supplementation[J]. International Journal of Cancer, 2016, 139(5):1171-1181.

DOI PMID

[41]
JHOU B Y, SONG T Y, LEE I, et al. Lycopene inhibits metastasis of human liver adenocarcinoma SK-Hep-1 cells by downregulation of NADPH oxidase 4 protein expression[J]. Journal of Agricultural and Food Chemistry, 2017, 65(32):6893-6903.

DOI

[42]
PENG S J, LI J, ZHOU Y, et al. In vitro effects and mechanisms of lycopene in MCF-7 human breast cancer cells[J]. Genetics and Molecular Research, 2017, 16(2):13.

[43]
ELGASS S, COOPER A, CHOPRA M. Lycopene inhibits angiogenesis in human umbilical vein endothelial cells and rat aortic rings[J]. British Journal of Nutrition, 2012, 108(3):431-439.

DOI

[44]
ŞAHIN M, ŞAHIN E, GÜMÜŞLÜ S. Effects of lycopene and apigenin on human umbilical vein endothelial cells in vitro under angiogenic stimulation[J]. Acta Histochemica, 2012, 114(2):94-100.

DOI

[45]
KAWATA A, MURAKAMI Y, SUZUKI S, et al. Anti-inflammatory activity of β-carotene,lycopene and tri-n-butylborane,a scavenger of reactive oxygen species[J]. In Vivo, 2018, 32(2):255-264.

[46]
MARCOTORCHINO J, ROMIER B, GOURANTON E, et al. Lycopene attenuates LPS-induced TNF-α secretion in macrophages and inflammatory markers in adipocytes exposed to macrophage-conditioned media[J]. Molecular Nutrition & Food Research, 2012, 56(5):725-732.

[47]
ZHAN J D, YAN Z J, KONG X J, et al. Lycopene inhibits IL-1β-induced inflammation in mouse chondrocytes and mediates murine osteoarthritis[J]. Journal of Cellular and Molecular Medicine, 2021, 25(7):3573-3584.

DOI

[48]
PALOZZA P, SIMONE R, CATALANO A, et al. Lycopene prevention of oxysterol-induced proinflammatory cytokine cascade in human macrophages:inhibition of NF-κB nuclear binding and increase in PPARγ expression[J]. The Journal of Nutritional Biochemistry, 2011, 22(3):259-268.

DOI

[49]
MOZOS I, STOIAN D, CARABA A, et al. Lycopene and vascular health[J]. Frontiers in Pharmacology, 2018, 9:521.

DOI PMID

[50]
LEE W, KU S K, BAE J W, et al. Inhibitory effects of lycopene on HMGB1-mediated pro-inflammatory responses in both cellular and animal models[J]. Food and Chemical Toxicology, 2012, 50(6):1826-1833.

DOI PMID

[51]
XUE R, QIU J N, WEI S, et al. Lycopene alleviates hepatic ischemia reperfusion injury via the Nrf2/HO-1 pathway mediated NLRP3 inflammasome inhibition in Kupffer cells[J]. Annals of Translational Medicine, 2021, 9(8):631.

DOI PMID

[52]
RISO P, PINDER A, SANTANGELO A, et al. Does tomato consumption effectively increase the resistance of lymphocyte DNA to oxidative damage?[J]. The American Journal of Clinical Nutrition, 1999, 69(4):712-718.

DOI

[53]
KOBAYASHI T, IIJIMA K, MITAMURA T, et al. Effects of lycopene,a carotenoid,on intrathymic T cell differentiation and peripheral CD4/CD8 ratio in a high mammary tumor strain of SHN retired mice[J]. Anti-Cancer Drugs, 1996, 7(2):195-198.

DOI

[54]
WATZL B, BUB A, BLOCKHAUS M, et al. Prolonged tomato juice consumption has no effect on cell-mediated immunity of well-nourished elderly men and women[J]. The Journal of Nutrition, 2000, 130(7):1719-1723.

DOI

[55]
柴静波, 李萍, 裴志萍. 番茄红素对荷卵巢癌大鼠免疫功能的影响及其机制研究[J]. 实用肿瘤学杂志, 2017, 31(1):13-17.

CHAI J B, LI P, PEI Z P. The effects and mechanism of lycopene on immune function of ovarian cancer rats[J]. Practical Oncology Journal, 2017, 31(1):13-17. (in Chinese)

[56]
WATZL B, BUB A, BRIVIBA K, et al. Supplementation of a low-carotenoid diet with tomato or carrot juice modulates immune functions in healthy men[J]. Annals of Nutrition and Metabolism, 2003, 47(6):255-261.

DOI PMID

[57]
YAMAGUCHI M, HASEGAWA I, YAHAGI N, et al. Carotenoids modulate cytokine production in Peyer’s patch cells ex vivo[J]. Journal of Agricultural and Food Chemistry, 2010, 58(15):8566-8572.

DOI

[58]
GARCIA A L, RÜHL R, HERZ U, et al. Retinoid- and carotenoid-enriched diets influence the ontogenesis of the immune system in mice[J]. Immunology, 2003, 110(2):180-187.

PMID

[59]
SUN S S, MENG Q W, BAI Y S, et al. Lycopene improves maternal reproductive performance by modulating milk composition and placental antioxidative and immune status[J]. Food & Function, 2021, 12(24):12448-12467.

[60]
AN B K, KIM D H, JOO W D, et al. Effects of lycopene and tomato paste on oxidative stability and fatty acid composition of fresh belly meat in finishing pigs[J]. Italian Journal of Animal Science, 2019, 18(1):630-635.

DOI

[61]
FACHINELLO M R, GASPARINO E, PARTYKA A V S, et al. Dietary lycopene alters the expression of antioxidant enzymes and modulates the blood lipid profile of pigs[J]. Animal Production Science, 2020, 60(6):806-814.

DOI

[62]
LIU A M, CHEN X L, HUANG Z Q, et al. Effects of dietary lycopene supplementation on intestinal morphology,antioxidant capability and inflammatory response in finishing pigs[J]. Animal Biotechnology, 2022, 33(3):563-570.

DOI

[63]
WEN W X, CHEN X L, HUANG Z Q, et al. Dietary lycopene supplementation improves meat quality,antioxidant capacity and skeletal muscle fiber type transformation in finishing pigs[J]. Animal Nutrition, 2022, 8:256-264.

DOI

[64]
FACHINELLO M R, FERNANDES N L M, DE SOUTO E R, et al. Lycopene affects the immune responses of finishing pigs[J]. Italian Journal of Animal Science, 2018, 17(3):666-674.

DOI

[65]
FACHINELLO M R, PARTYKA A V S, KHATLAB A D S, et al. 706 Lycopene affects hepatic gene expression of the main antioxidant enzymes in gilts[J]. Journal of Animal Science, 2017, 95(Suppl.4):344.

[66]
FACHINELLO M R, GASPARINO E, MONTEIRO A N T R, et al. Effects of dietary lycopene on the protection against oxidation of muscle and hepatic tissue in finishing pigs[J]. Asian-Australasian Journal of Animal Sciences, 2020, 33(9):1477-1486.

DOI PMID

[67]
KANG H G, LEE S, JEONG P S, et al. Lycopene improves in vitro development of porcine embryos by reducing oxidative stress and apoptosis[J]. Antioxidants, 2021, 10(2):230.

DOI

[68]
王捷. 番茄红素、芝麻酚和黄芩素对猪精子冷冻保存效果的影响[D]. 硕士学位论文. 杨凌: 西北农林科技大学, 2017.

WANG J. Effects of lycopene,sesamol and baicalein on cryopreservation of boar spematozoa[D]. Master’s Thesis. Yangling: Northwest A&F University, 2017. (in Chinese)

[69]
MEZBANI A, KAVAN B P, KIANI A, et al. Effect of dietary lycopene supplementation on growth performance,blood parameters and antioxidant enzymes status in broiler chickens[J]. Livestock Research for Rural Development, 2019, 31(1):12.

[70]
SARKER M T, WANG Z Y, YANG H M, et al. Evaluation of the protective effect of lycopene on growth performance,intestinal morphology,and digestive enzyme activities of aflatoxin B1 challenged broilers[J]. Animal Science Journal, 2021, 92(1):e13540.

[71]
SARKER M T, WAN X L, YANG H M, et al. Dietary lycopene supplementation could alleviate aflatoxin B1 induced intestinal damage through improving immune function and anti-oxidant capacity in broilers[J]. Animals, 2021, 11(11):3165.

DOI

[72]
AN B K, CHOO W D, KANG C W, et al. Effects of dietary lycopene or tomato paste on laying performance and serum lipids in laying hens and on malondialdehyde content in egg yolk upon storage[J]. The Journal of Poultry Science, 2019, 56(1):52-57.

DOI

[73]
ORHAN C, KUCUK O, SAHIN N, et al. Lycopene supplementation does not change productive performance but lowers egg yolk cholesterol and gene expression of some cholesterol-related proteins in laying hens[J]. British Poultry Science, 2021, 62(2):227-234.

DOI PMID

[74]
SUN B, CHEN C, WANG W, et al. Effects of lycopene supplementation in both maternal and offspring diets on growth performance,antioxidant capacity and biochemical parameters in chicks[J]. Journal of Animal Physiology and Animal Nutrition, 2015, 99(1):42-49.

DOI

[75]
SUN B, MA J, ZHANG J, et al. Lycopene regulates production performance,antioxidant capacity,and biochemical parameters in breeding hens[J]. Czech Journal of Animal Science, 2014, 59(10):471-479.

DOI

[76]
ROZBICKA-WIECZOREK A J, WIESYK E, BRZÓSKA F, et al. Fatty acid profile and oxidative stress of thigh muscles in chickens fed the ration enriched in lycopene,selenium compounds or fish oil[J]. Annals of Animal Science, 2014, 14(3):595-609.

DOI

[77]
ENGLMAIEROVÁ M, BUBANCOVÁ I, VÍT T, et al. The effect of lycopene and vitamin E on the growth performance,quality and oxidative stability of chicken leg meat[J]. Czech Journal of Animal Science, 2011, 56(12):536-544.

DOI

[78]
SHEVCHENKO L V, DAVYDOVYCH V A, USHKALOV V O, et al. The effect of astaxanthin and lycopene on the content of fatty acids in chicken egg yolks[J]. Regulatory Mechanisms in Biosystems, 2020, 11(4):568-571.

DOI

[79]
JANG I S, MOON Y S. Effects of lycopene on the expression of lipid metabolism,glucose transport and pro-inflammatory related genes in chickens[J]. Korean Journal of Poultry Science, 2015, 42(3):231-238.

DOI

Outlines

/