CN103140588B - 包含异黄酮-苷元、雌马酚和露纳素(lunasin)的基于发酵大豆的混合物,其制备方法及其在食品、医药和化妆品领域的相关应用 - Google Patents
包含异黄酮-苷元、雌马酚和露纳素(lunasin)的基于发酵大豆的混合物,其制备方法及其在食品、医药和化妆品领域的相关应用 Download PDFInfo
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Abstract
本发明涉及一种混合物，该混合物除包含雌马酚和露纳素(lunasin)之外，还包含异黄酮-苷元，例如大豆苷元(daidzein)、染料木素(genistein)和黄豆黄素(glycitein)，所述混合物基于用乳酸菌发酵的大豆，所述乳酸菌分离自食物基质；还涉及所述混合物用于保护人肠细胞和皮肤及其类似物的应用，其中，所述保护尤其涉及防止炎性状态和保护屏障功能，及其治疗毛发脱落的应用。
Description
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本发明涉及一种基于已发酵大豆的混合物、所述混合物的制备方法以及所述混合物在食品、医药和化妆品领域的应用，其中，所述混合物包含异黄酮-苷元(isoflavones-aglicones)、雌马酚和露纳素。具体而言，本发明涉及一种混合物，该混合物除包含雌马酚和露纳素(lunasin)之外，还包含异黄酮-苷元，如大豆苷元(daidzein)、染料木素(genistein)和黄豆黄素(glycitein)，所述混合物基于已用乳酸菌发酵的大豆，所述乳酸菌分离自食物基质；本发明还涉及所述混合物用于保护皮肤及其附件和保护人肠细胞的应用，其中，所述保护尤其涉及防止炎性状态和保护屏障功能，及其用于治疗脱发的应用。
异黄酮是天然存在于各种植物(特别是大豆)中的双酚类化合物(Tsangalis等，2002.Enzymatic transformation of isoflavone phytoestrogens in soymilk byβ-glucosidase producing bifidobacteria(“用产生β-葡萄糖苷酶的双歧杆菌对大豆乳中异黄酮植物激素进行酶转化”).Food Res.Int.Sci.67:3104-3113)。大豆源异黄酮和基于大豆的食物制品属于四种类型的化合物：苷元、丙二酰基-、乙酰基-和β-葡萄糖苷-共轭物(Tsangalis等，2002.Enzymatic transformation ofisoflavone phytoestrogens in soymilk byβ-glucosidase producing bifidobacteria.(“用产生β-葡萄糖苷酶的双歧杆菌对大豆乳中异黄酮植物激素进行酶转化”)Food Res.Int.Sci.67:3104-3113)。多于90％大豆异黄酮总浓度的大豆异黄酮以β-葡萄糖苷衍生物的形式出现。由于β-葡萄糖苷衍生物具有强疏水性特征和高分子量，其不被人体吸收。因此，为使该化合物可被生物利用继而被代谢，必须将其水解至苷元。在通过肠道期间，肠β-葡萄糖苷酶和细菌β-葡萄糖苷酶的活力会使水解成为苷元(例如大豆苷元、染料木素和黄豆黄素)的作用发生。游离形式的所述苷元具有类似于雌激素的结构，并在人体内模拟雌二醇的功能(Setchell和Cassidi,1999.Dietary isoflavones:biological effects and relevance tohuman health.(“食物异黄酮：生物效应以及和人体健康的关系”)J.Nutr.131:758-767)。通常，异黄酮苷元的使用与激素病理学风险降低有关(Kruzer 2000.Hormon effects of soy isoflavones:studies in premenopausal and postmenopausalwomen(“大豆异黄酮的激素效应：在绝经前后的妇女中的研究”).J.Nutr.130:660-661)，也与骨质疏松症、更年期、癌症和心血管病理引起的死亡的较低发生率有关(Nagata等.1998.Decreased serum total cholesterol concentration isassociated with high intake of soy products in Japanase men and women(“日本男女中血清总胆固醇浓度降低与大量摄取大豆制品相关”))J.Nutr.128:209-213)。
雌马酚是一种非甾体类化合物雌激素，属于异黄酮类。对人类而言，雌马酚的主要来源是大豆衍生物，是最富含大豆苷元和苷元型大豆苷元的代表(其直接前体)(Axelson等，1984.Soya a dietary source of the non-steroidal oestrogenequol in man and animals(“大豆，人和动物的非甾体雌激素雌马酚的膳食来源”).J.Endocrinol.102:49-56)。与其它异黄酮-苷元不同，雌马酚是唯一具有核心手性中心的异黄酮-苷元，该手性中心由于杂环中缺乏双键所致(Setchell等，2002.The clinical importance of the metabolite equol a clue to the effectiveness of soyand its isoflavones(“代谢物雌马酚的临床重要性，大豆及其异黄酮有效性的线索”).Am.Soc.Nutr.Sci.125:3577-3583)。通常，雌马酚容易通过结肠壁吸收，但怠于代谢(Setchell等，2002.The clinical importance of the metabolite equol aclue to the effectiveness of soy and its isoflavones(“代谢物雌马酚的临床重要性，大豆及其异黄酮有效性的线索”).Am.Soc.Nutr.Sci.125:3577-3583)。与其大豆苷元前体相比，雌马酚显示出一组有趣的性质：高雌激素活性(Muthyala等，2004.Equol,a natural estrogenic metabolita from soy isoflavones:convenientpreparation and resolution of R-and S-equols and their differing binding andbiological activity through estrogen receptors alpha and beta(“雌马酚，源自大豆异黄酮的天然雌激素代谢物：方便的制备和解析R-和S-雌马酚，以及它们对α-和β-雌激素受体的不同结合活性和生物活性”).Bioorg.Med.Chem.12:1559-1567)、抗氧化剂活性(Mitchell等，1998.Antioxidant efficacy ofphytoestrogens in chemical and biological model systems(“在化学和生物模型系统中植物雌激素的抗氧化剂功效”).Arch.Biochem.Biophys.360:142-148)以及抗雄激素活性(Lund等，2004.Equol is a novel anti-androgen that inhibits prostategrowth and hormone feedback(“雌马酚，一种抑制前列腺生长和激素反馈的新型抗雄激素物质”).Bio.Reprod.70:1188-1195)。
大豆异黄酮在人体内的代谢有广泛记载(Axelson等，1984.Soya a dietarysource of the non-steroidal oestrogen equol in man and animals(“大豆，人和动物的非甾体雌激素雌马酚的膳食来源”).J.Endocrinol.102:49-56；Bannwart等，1984.Identification of o-desmethylangolensin,a metabolita of daidzein and ofmatairesinol,one likely plant precursor of the animal lignan enterolactone in humanurine(“鉴定邻-脱甲基安哥拉紫檀素，一种大豆苷元和罗汉松脂素的代谢物，与人尿液中动物木脂素肠内酯类似的植物前体”).Finn.Chem.Lett.5:120-125)。在通过肠道期间，将糖苷型异黄酮代谢为苷元并将苷元代谢为雌马酚的能力仅限于30～50％的西方国家人口(Frankefeld等，2005.High concordance ofdaidzein-metabilizing phenotypes in individuals measured 1to 3years apart(“在1-3年期间检测个体中大豆苷元代谢表型的高度一致性”).Brit.J.Nutr.94:873-876)。可根据两个主要策略来提高大豆衍生的异黄酮的生物利用度：在使用前富集苷元和雌马酚，或者通过摄取适合于原位合成所述化合物的有活性的活细菌来调节肠内微生物群(Tsangalis等，2004.Development o fan isoflavoneaglycone-enriched soymilk using soy germ,soy protein isolate and bifidobacteria(“采用大豆胚芽、大豆蛋白分离物和双歧杆菌制备富含大豆异黄酮苷元的大豆乳的开发”).Food Res.Intern.37:301-312)。各种研究都考虑了在大豆乳的发酵过程中使用双歧杆菌和乳酸菌来将糖苷型异黄酮转化为苷元和/或雌马酚(Chun等，2007.Conversion of isoflavone glucoside to aglycones in soymilk byfermentation with lactic acid bacteria(“通过乳酸菌发酵在大豆乳中将异黄酮葡萄糖苷转化为苷元”).J.Food Sci.72:39-44；Donkor和Shah 2008.Production ofβ-glucosidase and hydrolysis of isoflavone phytoestrogens by Lactobacillusacidophilus,Bifidobacterium lactis and Lactobacillus casei in soymilk(“采用嗜酸乳杆菌(Lactobacillus acidophilus)、双歧乳酸杆菌(Bifidobacterium lactis)和干酪乳杆菌(Lactobacillus casei)在大豆乳中生产β-葡萄糖苷酶和水解异黄酮植物雌激素”).J.Food Sci.73:15-20；Pham和Shah 2007.Biotransformation of isoflavoneglycosides by Bifidobacterium animalis in soymilk supplemented with skim milkpowder.(“采用动物双歧杆菌(Bifidobacterium animalis)在添加了脱脂奶粉的大豆乳中生物转化异黄酮葡萄糖苷”)J.Food Sci.72:316-324；Tsangalis等，2002；Tsangalis等，2004；Wei等，2007.Using Lactobacillus and Bifidobacterium toproduct the isoflavone algycones in fermented soymilk(“采用乳杆菌和双歧杆菌在发酵的大豆乳中产生异黄酮苷元”).Int.J.Food Microbiol.117:120-124)。但是这些研究中明显存在一定局限性：(i)已考虑的细菌生物型/种类的数量很有限；(ii)用于发酵步骤的细菌仅来源于人的粪便材料；(iii)用于发酵的底物数量很有限，其中无一考虑涉及使用生物大豆粉(soya biological flour)；(iv)制备仅基于异黄酮/苷元或雌马酚，并且就雌马酚的生产而言，其最大浓度是0.521mg/100ml(Tsangalis等，2002.Enzymatic transformation of isoflavonephytoestrogens in soymilk byβ-glucosidase producing bifidobacteria(“用产生β-葡萄糖苷酶的双歧杆菌在大豆乳中酶促转化异黄酮植物雌激素”).Food Res.Int.Sci.67:3104-3113)；(v)没有研究测试制品的生物有效性，特别是在皮肤保护方面；以及(vi)没有研究配制出包括异黄酮-苷元、雌马酚和露纳素的制剂。
露纳素是一种生物活性肽(43个氨基酸残基，分子量约为5400Da)，其在大豆中被首次鉴定(Galve等，2001.Chemopreventive property of a soybean peptide(lunasin)that binds to deacetylated histones and inhibits acetylation(“结合去乙酰组蛋白并且抑制乙酰化的大豆肽(露纳素，lunasin)的化学防护特性”).CancerRes.61:7473–7478)，并接着在大麦中(Jeong等，2002.Barley lunasin suppressesras-induced colony formation and inhibits core histone acetylation in mammaliancells(“大麦露纳素在哺乳动物细胞中抑制ras-诱导的克隆形成并且抑制核心组蛋白的乙酰化”).J.Agric.Food Chem.50:5903-5908)、小麦中(Jeong等，2007.Thecancer preventive peptide lunasin from wheat inhibits core histone acetylation(“源自小麦的抗癌肽露纳素抑制核心组蛋白的乙酰化”).Cancer Lett.255:42–48)和苋属植物中(Silva-Sánchez等，2008.Bioactive peptides in amaranth(Amaranthushypochondriacus)seed(“苋属植物千穗谷(Amaranthus hypochondriacus)种子中的生物活性肽”).J.Agric.Food Chem.56:1233–1240)也被找到。大豆中，露纳素的浓度根据栽培品种、培养土壤气候环境和收获后的谷物所受的技术加工而变化。在Loda栽培品种中发现了非常高浓度的露纳素(约11mg/g)，而在其它大豆品种中(例如，Imari品种)，露纳素的含量不超过5～6mg/g(Wang等.2008.Analysis of soybean protein derived peptides and the effect of cultivar,environmental conditions,and processing of lunasin concentration in soybean andsoy products(“大豆蛋白衍生肽的分析以及栽培品种、环境条件和露纳素加工对于大豆和大豆制品中露纳素浓度的影响”).J.AOAC Intern.91:936-944)。露纳素在多肽链的C端含有9个天冬氨酸残基。这种组成有利于提高对低乙酰化的染色质区域(肽可结合于该区域)亲和性，从而抑制乙酰化-脱乙酰动力学，因而在癌发生中起到肿瘤抑制物的作用。也有报道露纳素可对癌发生现象发挥保护活性，归因于其对由ras基因诱导的细胞增殖的抑制和对H3组蛋白乙酰化作用的抑制(Jeong等，2003.Characterization of lunasin isolated from soybean(“从大豆中分离的露纳素的表征”).J Agric Food Chem.51:7901-7906)。从文献数据来看，很明显，至今没有研究考虑过在乳酸菌发酵过程中从大豆衍生物中富集露纳素。
基于如上所报道的考虑，某些因素似乎起到了具有显著革新意义的作用：(i)使用基于大豆的底物，可能是生物来源；(ii)使用分离自食物基质而非粪便来源的乳酸菌；(iii)优化适合于促成含更大量的功能性分子，如异黄酮-苷元、雌马酚和露纳素，的制剂配制的生物技术方法；(iv)通过体外和离体的分析，证实所得制剂的功效，诸如对皮肤和人肠细胞的保护效果，尤其涉及抑制炎性状态及保持屏障功能。
目前，本发明作者开发出了一种新方法，该方法适合于提供基于已发酵大豆的混合物，该混合物富集了异黄酮、苷元、雌马酚和露纳素。本发明所述的混合物通过使用特定乳酸菌混合物发酵大豆而获得，所述乳酸菌仅来源于食物基质，而非粪便来源。由于本发明所述的混合物富含异黄酮-苷元、雌马酚和露纳素，特别是露纳素，其在皮肤和人肠细胞的保护方面具有特殊功效，其中，所述保护尤其涉及防止炎性状态及保持屏障功能。
目前，本发明作者考虑了以下事项：(i)根据β-葡萄糖苷酶对对硝基苯基-β-D-吡喃葡萄糖苷(pNPG)的活性，对103种仅分离自食物基质的乳酸菌进行筛选，选出4种乳酸菌作为混合起始物用于发酵基于大豆粉的底物，其中，所述4种乳酸菌的名称如下：植物乳杆菌(L.plantarum)DPPMA24W(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23756)和DPPMASL33(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23755)、发酵乳杆菌(L.fermentum)DPPMA114(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23757)和鼠李糖乳杆菌(L.rhamnosus)DPPMAAZ1(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23758)；(ii)使用基于大豆的14种不同底物，选出最理想的是基于生物大豆粉，使用混合起始物发酵的制品；(iii)通过对生物大豆粉底物发酵方法的优化得到的制剂配方包含1.45mg/100ml(57.0μM)大豆苷元、3.9mg/100ml(140.3μM)染料木素、0.58mg/100ml(20.4μM)黄豆黄素、0.9mg/100ml(37.3μM)雌马酚，和8.4mg/100ml露纳素；(iv)基于上述功能性化合物的制品显示对皮肤表皮的保护效果、对炎性状态的积极抑制效果和肠细胞屏障功能。
如本发明所述的乳酸菌属于乳(酸)杆菌(Lactobacillus)种，并且分离自意大利中部与南部用于面包制作的天然酵母和来自普利亚区地区的佩科里诺干酪(Pecorino)型的陈年“凝乳拉伸型干酪(pasta filata)”乳酪。分离自所述食物基质的乳酸菌所通常显示的代谢和环境适应特性与在人类和动物胃肠道寄居的微生物差异不大。选择并使用了植物乳杆菌(L.plantarum)DPPMA24W(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23756)、植物乳杆菌(L.plantarum)DPPMASL33(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23755)、发酵乳杆菌(L.fermentum)DPPMA114(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23757)，和鼠李糖乳杆菌(L.rhamnosus)DPPMAAZ1(2010年7月8日保藏于德国微生物菌种保藏中心(DSMZ)，DSM号23758)。
已经标准化并最佳化的生物技术方法涉及：通过所述四种乳酸菌在基于不同大豆粉的底物(优选生物来源)上，于30～37℃发酵48～96小时。在发酵过程结束时，可通过离心从培养肉汤移除细胞或不移除细胞，并利用干燥或冷冻干燥使上清脱水。
用于发酵基于生物大豆的制品的生物技术方法描述如下。
作为发酵各种大豆乳制品的结果，合成得到3.9～57.0μM大豆苷元、7.8～140.3μM染料木素、6.7～20.4μM黄豆黄素、7.6～37.3μM雌马酚和约8.4mg/100ml露纳素。上述浓度的上限是指生物大豆粉来源的大豆乳的发酵。根据一种可能的配方，显示来自生物大豆的发酵产物的应用适合于：(i)保护表皮并增强其屏障功能；(ii)抑制经干扰素-γ(IFN-γ)和脂多糖(LPS)诱导后的Caco-2/TC7细胞的炎性状态；(iii)刺激屏障功能，以跨上皮电阻(Transepithelial ElectricResistance(TEER))测试证明；和(iv)抑制白细胞介素-8(IL-8)的合成。
采用微生物、色谱技术和测试对体外和离体细胞培养物的分析所作的补充分析证明了：如本发明所述，使用由分离自食物基质且未用于先前研究的乳酸菌种类组成的混合起始物发酵生物大豆粉，提供(i)苷元、雌马酚和露纳素的伴随性合成，这在之前的研究中从未被发现，和(ii)抗炎性状态的保护功效，增强表皮和人肠细胞的屏障功能。
因此，本发明的特殊目标是一种通过大豆发酵制备基于已发酵大豆的混合物的方法，其中，所述混合物包括异黄酮-苷元、雌马酚和露纳素，所述大豆发酵使用下述四种乳酸菌混合物：植物乳杆菌(Lactobacillus.plantarum)DSM23755、植物乳杆菌(Lactobacillus.plantarum)DSM 23756、发酵乳杆菌(Lactobacillus.fermentum)DSM 23757和鼠李糖乳杆菌(Lactobacillus.rhamnosus)DSM 23758。根据本发明所述方法获得的混合物是富含异黄酮-苷元、露纳素、雌马酚的混合物，即，与使用不同于本发明所用那些乳酸菌的乳酸菌的方法所获的已知混合物相比，本发明所述的混合物含有更大百分比的异黄酮-苷元、露纳素和雌马酚。
本发明所述的方法包括或由以下步骤组成：a)培养增殖所述四种乳酸菌，植物乳杆菌(Lactobacillus plantarum)DSM 23755、植物乳杆菌(Lactobacillusplantarum)DSM 23756、发酵乳杆菌(Lactobacillus.fermentum)DSM 23757和鼠李糖乳杆菌(Lactobacillus.rhamnosus)DSM 23758；
b)将所述乳酸菌的水性悬液接种于基于大豆的底物；优选乳酸菌水性悬液以占总底物体积1～4％的量接种于所述底物，所述水性悬液中的各生物型细胞密度约为log 9.0cfu/ml；
c)在30～37℃，优选30℃，孵育48～96小时，优选96小时。
适用的基于大豆的底物是，例如，大豆粉，优选生物大豆粉，大豆乳和如本发明所述的其它市售配方。
本发明所述方法还可包括步骤d)离心肉汤培养物以分离细胞和乳酸菌。具体来说，所述肉汤培养物的离心可在4℃以10,000×g进行15min。
本发明所述方法还可包括步骤e)通过干燥或冷冻干燥使步骤d)所获的上清脱水。根据一个实施方式，省略了步骤d)的配方可包含活性的活乳酸菌。组合物的制备可涉及在脱水步骤结束时添加合适的赋形剂进行配制，以获得依情况而定的适合于口服或局部使用形式的制品。
本发明的另一个目的是一种混合物，该混合物包含异黄酮-苷元、雌马酚和露纳素，其基于已发酵的大豆，所述大豆可获自不含移除乳酸菌的步骤d)的上述方法。因此，所述混合物包含如本发明上述的乳酸菌。如上所述，如本发明所述的混合物是富含异黄酮-苷元、雌马酚和露纳素的混合物，即，与使用不同于本发明所用那些乳酸菌的乳酸菌的方法所获的已知混合物相比，本发明所述的混合物含有更高百分比的异黄酮-苷元、雌马酚和露纳素。
本发明还涉及一种药物或化妆品组合物，该组合物包含上述混合物和一种或多种药学上或化妆品中可接受的赋形剂和/或佐剂，或由这些物质组成。
根据另一个实施方式，本发明所述的混合物可作为食品补足剂(foodintegrator)使用。例如，所述混合物也可用于传统食品，例如烘焙或意大利面制品。
此外，本发明所述的混合物或组合物可用于治疗皮肤或肠的紊乱或疾病。具体而言，所述混合物或组合物可用于抗皮肤屏障功能改变，例如，用于预防或治疗皮肤敏感、皮肤干燥、牛皮癣、特应性皮炎、皮脂溢性皮炎、皮屑、刺激性皮肤病、湿疹皮肤病、接触皮肤病、溃疡、痤疮、皮肤老化。此外，可将本发明所述的混合物或组合物用于肠屏障功能改变的情况，例如用于治疗或预防腹腔疾病、食物不耐症、克罗恩病(Crohn’s disease)。
本发明所述的混合物或组合物可用于化妆品领域，例如用于治疗毛发脱落，或用于医药领域，用于治疗脱毛症或休止期脱发)。
具体而言，本发明所述的混合物或组合物可通过局部方式给药，例如以乳霜、洗液、糊剂、油膏、凝胶、溶液、乳剂、悬液形式，或者全身给药，例如通过口服方式，例如以小瓶、咀嚼片、丸、小袋等形式。
当然，根据本发明包括移除乳酸菌的步骤d)的方法所得的混合物，或者包含相同成分的药物或化妆品组合物也可有利地用于上述指征，因为与使用不同于本发明所用那些乳酸菌的乳酸菌的方法所获的已知混合物相比，该混合物或组合物含有更高百分比的异黄酮-苷元、雌马酚和露纳素。
此外，本发明的一个目标是以下四种乳酸菌的混合物：植物乳杆菌(Lactobacillus plantarum)DSM 23755、植物乳杆菌(Lactobacillus plantarum)DSM 23756、发酵乳杆菌(Lactobacillus fermentum)DSM 23757和鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758。最后，本发明的一个目标是植物乳杆菌(Lactobacillus plantarum)DSM 23755或植物乳杆菌(Lactobacillus plantarum)DSM 23756或发酵乳杆菌(Lactobacillus fermentum)DSM 23757或鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758。
现在根据本发明的优选实施方式，配合具体相关的所附图片，对本发明进行示例性的描述，而不用于限制本发明。
图1显示属于不同种的103种乳酸菌生物型的β-葡萄糖苷酶在pNPG合成底物上的活性。所有用于该分析的乳酸菌都以完全创新的方式提前分离，并仅分离自食物基质。乳酸菌生物型由编码显示，以表明其与菌种之间的对应性，请参考本文(实施例1)的方法说明。数据是三次三个重复样品实验数据的平均值。由箱线图报告统计学的详细说明。
图2A显示通过混合起始物进行的乳酸化进程，所述混合起始物在来源于14种不同粉末的大豆乳的存在下选出。图2B显示包括混合起始物的乳酸菌生物型的细胞密度，所述混合起始物在来源于14种不同粉末的大豆乳的存在下选出。数据是三次三个重复样品实验数据的平均值。由箱线图报告统计学的详细说明。
图3显示露纳素在使用所选混合起始物的大豆乳发酵过程中的合成(mg/100ml)，所述大豆乳获自生物大豆粉(OFS)。数据是三次三个重复样品实验数据的平均值。
图4显示在已发酵的生物大豆乳(OFS)和PBS中暴露0小时和24小时后，重建表皮的跨上皮电阻(TEER)(欧姆×cm2)，所述生物大豆乳使用所选混合起始物发酵。数据是三次三个重复样品实验数据的平均值。
图5显示来自Caco-2/TC7细胞的一氧化氮(NO)释放(μM)。所述细胞经大豆乳和用作标准样品的化合物(10μM)(雌马酚、大豆苷元、染料木素和黄豆黄素)预处理24小时，所用大豆乳获自未经发酵或用所选混合起始物发酵的生物大豆粉，稀释到雌马酚终浓度为10μM并无菌过滤。接着，细胞由干扰素-γ(IFN-γ)(1000U/ml)和脂多糖(LPS)(100ng/ml)刺激24小时。将含有DMSO(1％，v/v)或甲醇(0.5％，v/v)的DMEM培养基用作阴性对照。数据是三次三个重复样品实验数据的平均值。星号表示与阴性对照相比的有意义差异(P<0.01)。
图6显示孵育24、48和72小时后Caco-2/TC7细胞的跨上皮电阻(TEER)(欧姆×cm2)。在如下几种情况下进行孵育：存在干扰素-γ(IFN-γ)(1000U/ml)(-■-)；存在IFN-γ和获自生物大豆粉的大豆乳，其中，未经发酵大豆乳(-▲-)或用所选混合起始物发酵、稀释到雌马酚终浓度(10μM)并无菌过滤的大豆乳(-×-)。DMEM培养基用作阴性对照(-◆-)。数据是三次三个重复样品实验数据的平均值。星号表示与阴性对照相比的有意义差异(P<0.01)。
图7显示来自Caco-2/TC7细胞的白细胞介素-8(IL-8)的释放(pg/ml)，其中Caco-2/TC7细胞由白细胞介素-1β(IL-1β)(2ng/ml)刺激24小时，并接着用大豆乳和用作标准样品的化合物(10μM)(雌马酚、大豆苷元、染料木素和黄豆黄素)处理(24小时)，所用大豆乳获自未经发酵或用所选混合起始物发酵的生物大豆粉，稀释到雌马酚终浓度为10μM并无菌过滤。将包含DMSO(1％，v/v)或甲醇(0.5％，v/v)的DMEM培养基用作阴性对照。数据是三次三个重复样品实验数据的平均值。星号表示与阴性对照相比的有意义差异(P<0.01)。
图8显示包含露纳素和不含露纳素的生物质对细胞增殖的影响。
图9显示包含露纳素和不含露纳素的生物质对Bcl-2和Bax蛋白表达的影响。
实施例1：分离自食物基质的103种乳酸菌生物型的β-葡萄糖苷酶活性
本研究使用了103种乳酸菌生物型，所述乳酸菌属于以下种：消化乳杆菌(Lactobacillus alimentarius)(10N，2B，5A)、短乳杆菌(Lactobacillus brevis)(5Z，DPPMA869)、干酪乳杆菌(Lactobacillus casei)(LC10)、干酪乳杆菌干酪亚种(Lactobacillus casei subsp.Casei)(2047，2756，2763，2766)、干酪乳杆菌假植物亚种(Lactobacillus casei subsp.Pseudoplantarum)(2742，2745，2749，2750)、弯曲乳杆菌(Lactobacillus curvatus)(13H5，14H10，1Hd，2042，2081，2768，2770，2771，2775，SAL23，SAL35)、德氏乳杆菌保加利亚亚种(Lactobacillusdelbrueckii subsp.Bulgaricus)(11842，B15Z)、发酵乳杆菌(Lactobacillusfermentum)(DPPMA114，D13)、戈氏乳杆菌(Lactobacillus gasseri)(B30W)、瑞士乳杆菌(Lactobacillus helveticus)(15009，B26W，PR4)、希氏乳杆菌(Lactobacillushilgardii)(51B)、类食品乳杆菌(Lactobacillus paralimentarius)(15α，15β，16R，8D，DPPMA238)、副干酪乳杆菌(Lactobacillus paracasei)(12H8，1Hb，B14F5，B18S，B25F3，PF6，B61F5)、帕卡乳杆菌(Lactobacillus pacarbuchneri)(B10F5，B48F3，B48F5，B9F5t，BF1，BF2)、类植物乳杆菌(Lactobacillus paraplantarum)(4DE，DPPMA667)、戊糖乳杆菌(Lactobacillus pentosus)(8CF，12H5，12H6)、植物乳杆菌(Lactobacillus plantarum)(14H4，17N，19A，1A7，2A，30，3DM，4H1，4H10，DB200，DPPMASL33，DPPMA24W)、鼠李糖乳杆菌(Lactobacillusrhamnosus)(11，19，DPPMAAZ1，DPPMAAZ21)、沙克乳杆菌(Lactobacillussakei)(9I，SAL1，SAL18)、罗斯乳杆菌(Lactobacillus rossiae)(10A，15R，3D，5C1，5α，CF51，CI35，CR20，E18)、旧金山乳杆菌(Lactobacillussanfranciscensis)(16α，A17，BB12，DE9，E19，H10)、乳酸乳球菌乳酸亚种(Lactococcus lactis subsp.Lactis)(10γ)、戊糖片球菌(Pediococcuspentosaceus)(C16F5，C25F3，C30F5t，C6F5，C7F3，C9F5t，C29F5)和食窦魏斯氏菌(Weissella cibaria)(10XA16，3XA4，5S，5XF12)。所有生物型属于巴里大学植物保护和应用微生物学系的收藏和培养(the Collezione di Colture delDipartimento di Protezione delle Piante e Microbiologia Applicata dell’Universitàdegli Studi di Bari)，早前分离自食物基质(用于面包制作和乳酪的天然酵母)。乳酸菌生物型在MRS培养基(奥克欧德公司(Oxoid)，英国贝辛斯托克)中于30℃或37℃增殖24小时。
细胞培养24小时，通过离心(于4℃以10,000×g离心15min)收集，用磷酸盐缓冲液(50mM，pH 7.0)洗涤两次，并以log 9.0cfu/ml的细胞密度在水中重悬，用于β-葡萄糖苷酶活性分析。β-葡萄糖苷酶活性通过从对硝基苯酚-β-D-葡萄糖苷(pNPG)底物(西格玛奥德里奇化学公司(Sigma Aldrich Chemical Corporate)，美国密苏里州圣路易斯市)释放的对硝基苯酚量化。分析使用了含有900μlpNPG(终浓度)的磷酸盐缓冲液(0.5M,pH 7.5)和100μl细胞悬液。在40℃孵育该混合物，并通过在95℃热处理5min阻断该反应。在410nm检测吸光度。将一个β-葡萄糖苷酶活性单位(U)定义为，在分析条件下，1min释放1nmol对硝基苯酚所需的酶量(De Angelis等，2005.Purification and characterization of anintracellular family 3β-glucosidase from Lactobacillus sanfranciscensis CB1(“从旧金山乳杆菌CB1纯化和鉴定细胞内β-葡萄糖苷酶家族3”).Ital.J.Food Sci.17:131-142.)。
实施例2：大豆乳的制备和发酵
在大豆乳制作中使用了生物大豆(有机农业大豆，OFS)(ECNS公司(ECorNaturaSi)，意大利维罗纳市)、大豆蛋白分离物(SPI)(Supro Soja 80阿普托尼亚公司(Aptonia)，法国阿斯克新城)和不同的市售大豆粉制品(嘉吉调质溶液大豆蛋白(Cargill Texturizing Solutions Soy Protein)，比利时根特市)。OFS经洗涤并静置在蒸馏水中过夜。将潮湿并肿胀的大豆手工去皮，并用温水(约90℃)以1:10的比例稀释，用PBI国际均质器(PBI International homogeniser)(意大利米兰市)匀浆。该匀浆步骤以10,000×g进行2min，然后暂停1min，再以14,000×g处理2min。离心悬液(于4℃，7,000×g，10min)，无菌过滤大豆乳通过0.22μm孔径的滤器(密理博公司(Millipore Corporation)，贝德福德市)。其pH是6.2。SPI用蒸馏水(40℃)以0.4:10的比例稀释，并在搅拌(120rpm)下于约55℃热处理30min。在室温冷却后，用5M的NaOH调整pH至6.7(Tsangalis等，2002)。在高压灭菌器内于121℃进行灭菌15min。按照Chun等人描述的方法(Chun等，2007.Conversion of isoflavone glucoside to aglycones in soymilk by fermentation withlactic acid bacteria(“通过乳酸菌发酵将大豆乳中的异黄酮葡萄糖苷转化为苷元”).J.Food Sci.72:39-44)，将所述市售大豆粉制品用蒸馏水(40℃)以1:10的比例稀释。pH值约为6.3。在高压灭菌器内于121℃灭菌15min。
将基于β-葡萄糖苷酶活性挑选出的4种乳酸菌的混合细胞悬液接种(1～4％,v/v)于不同类型的大豆乳。4种乳酸菌生物型的每种的初始细胞密度是log 7.0cfu/ml。发酵在搅拌下(120rpm)于30℃进行96小时。将大豆乳冷冻干燥，重悬于DMEM培养基中并无菌过滤，用于人肠细胞测试。
实施例3：监测乳酸菌，测定异黄酮-苷元、雌马酚和露纳素
在各大豆乳类型的发酵过程中，使用RAPD-PCR技术对作为混合起始物的乳酸菌进行监测(植物乳杆菌(Lactobacillus plantarum)DSM 23755对应于DPPMASL33、植物乳杆菌(Lactobacillus plantarum)DSM 23756对应于DPPMA24W、发酵乳杆菌(Lactobacillus fermentum)DSM 23757对应于DPPMA114和鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758对应于DPPMAAZ1)。使用任选序列(P75’AGCAGCGTGG 3’(SEQ ID No:1)，和M13,5’GAGGGTGGCGGTTCT 3’(SEQ ID NO:2))的两个引物(英杰公司(Invitrogen)，意大利米兰市)，随机扩增质粒和细菌DNA染色体的不同区域用于分型(De Angelis等，2001.Characterization of non-starter lactic acid bacteria(NSLAB)from ewes’Italian cheeses based on phenotypic,genotypic and cell-wall protein analyses(“基于表型、基因型和细胞壁蛋白测试来鉴定来自母羊意大利奶酪的非起始物乳酸菌(NSLAB)”).Appl.Environ.Microbiol.67:2011-2020；Rossetti e Giraffa,2005.Rapid identification of dairy lactic acid bacteria by M13-generated,RAPD-PCRfingerprint databases(“用M13产生的RAPD-PCR指纹数据库快速鉴定乳制品乳酸菌”).J.Microbiol.Met.63:135-144).
根据Otieno和Shah描述的方法，从发酵的大豆乳样品提取异黄酮-苷元和雌马酚(Otieno和Shah,2007.A comparison of changes in the transformation ofisoflavones in soymilk using varying concentrations of exogenous andprebiotic-derived endogenousβ-glucosidases(“用不同浓度的外源和益生菌来源的内源β-葡萄糖苷酶对于大豆乳中异黄酮转化的影响比较”).J.Appl.Microbiol.103:601-612)。根据Maubach等人描述的方法进行HPLC色谱分析，以测定化合物(Maubach等，2003.Quantitation of soy-derived phytoestrogens in human breasttissue and biological fluids by high-performance liquid chromatography(“用高效液相色谱量化人乳腺组织和生物液中大豆来源的植物雌激素”).J.Chromatogr.784:137-144)。
通过HPLC色谱法，使用设有C18 Xterra Waters柱和214nm UV检测器的AKTA纯化系统(GE医疗保健(GE Healthcare))，测定发酵前和发酵中所获的大豆乳(来源于生物大豆粉)中的露纳素，用由5％ACN+0.05％TFA(洗脱液A)和ACN+0.05％TFA(洗脱液B)组成的混合物溶剂洗脱(Wang等，2008.Analysis ofsoybean protein derived peptides and the effect of cultivar,environmentalconditions,and processing of lunasin concentration in soybean and soy products(“大豆蛋白衍生肽分析以及培养品种、环境条件和处理过程对于大豆和大豆制品中的露纳素浓度的影响”).J.AOAC Intern.91:936-944)。使用合成的露纳素(EZB公司(EZBiolab)，美国印第安那卡梅尔市)作为标准样品。
实施例4：重建表皮测试和TEER(跨上皮电阻)测量
人重建表皮(重建的人表皮)由多层人表皮的正常角质化细胞构成。它是将人角质化细胞在化学限定培养基(MCDM 153)(不添加胎牛血清)中，在惰性多孔聚碳酸酯支持物上，于气液界面培养17天后所得的完全分化的表皮。在该生长阶段，形态分析显示有活力的多层化表皮和由多于十层紧密细胞层组成的角质层。根据过去描述过的方法使用人重建表皮(Di Cagno等，2009.Synthesis ofγ-amino butyric acid(GABA)by Lactobacillus plantarumDSMZ19463:functional grape must beverage and dermatological application(“用植物乳杆菌(Lactobacillus plantarum)DSMZ19463合成γ-氨基丁酸(GABA)：功能性未发酵葡萄汁饮料和皮肤科应用”).Appl Biotechnol Microbiol DOI:10.1007/s00253-009-23704)。
用Millicell-ERS电压电阻表(密理博公司(Millipore)，马萨诸塞州贝德福德市)实行TEER测量。测量以欧姆×cm2表示。
实施例5：Caco-2/TC7细胞测试
在Dulbecco(DMEM)培养基中培养人源Caco-2细胞(克隆TC7)，所述培养基中添加胎牛血清(10％)、非必需氨基酸(1％)、庆大霉素/链霉素(50μg/ml)、谷氨酰胺(2mM)和4-(2-羟乙基)-1-哌嗪基-乙磺酸(1％)(Di Cagno等，2010.Quorumsensing in sourdough Lactobacillus plantarum DC400:induction of plantaricin A(PlnA)under co-cultivation with other lactic acid bacteria and effect of PlnA onbacterial and Caco-2cells(“酵原植物乳杆菌DC400的群体感应：在与其它乳酸菌共培养的条件下诱导植物乳杆菌素A(PlnA)以及PlnA对细菌和Cao-2细胞的作用”).Proteomics付印中)。通过中性红(Neutral Red)染料摄取测试来测定细胞活性(Balls等，1987.纽约学术出版社Goldber A.M.编的“Approaches tovalidation alternative methods in toxicology”(《毒理学中可选的验证方法研究》)中的第45-58页)。在用不同制品处理24～72小时后，用PBS缓冲液洗涤细胞，再用中性红溶液(33mg/l)于37℃孵育4小时。然后，再次用PBS缓冲液洗涤细胞，并用裂解溶液(包含1％醋酸的50％乙醇水溶液)处理。用Novapath酶标仪(伯乐公司(Biorad)，加利福尼亚赫尔克里士市)进行读板(Di Cagno等，2010)。
通过测量氧化产物(即，硝酸盐和亚硝酸盐)来测定从Caco-2/TC7细胞释放的一氧化氮(NO)。在用不同制品孵育后，将细胞培养物上清与等体积的Griess试剂(1％，p/v，对氨基苯磺酸的0.5M HCl溶液和0.1％，p/v，N-1-萘基乙二胺盐酸盐)混合。在室温孵育30min后，在540nm检测吸光度，参考用亚硝酸钠制作的标准曲线来测定亚硝酸盐浓度。
为了检测TEER，将Caco-2/TC7细胞接种于(7.5×104个细胞/ml)有聚乙烯滤器(0.4μm孔径)的24孔微板容器中。处理前，在37℃孵育细胞21天。用不同制品进行处理18、24和48小时。然后通过TEER测量来测定细胞层的完整性。
为检测释放的白细胞介素-8(IL-8)，先用白细胞介素-1β孵育Caco-2/TC7细胞(24小时)，然后用不同制品进一步刺激24小时。通过ELISA分析(BenderMedSystems)测定促炎性因子IL-8的合成。根据试剂盒的说明书，使用标准曲线进行定量。
结果
(1)基于β-葡萄糖苷酶活性选择乳酸菌
预先测试分离自食物基质的103种乳酸菌生物型在pNPG合成底物上的β-葡萄糖苷酶活性。活性在0至202.3U之间变化(图1)。主要属于以下种的四十八个生物型没有表现β-葡萄糖苷酶活性：消化乳杆菌(L.alimentarius)、短乳杆菌(L.brevis)、干酪乳杆菌(L.casei)、德氏乳杆菌保加利亚亚种(L.delbrueckii subsp.Bulgaricus)、瑞士乳杆菌(L.helveticus)、希氏乳杆菌(L.hilgardiii)、类食品乳杆菌(L.paralimentarius)、类植物乳杆菌(L.paraplantarum)、戊糖乳杆菌(L.pentosus)、旧金山乳杆菌(L.sanfranciscensis)、乳酸乳球菌乳酸亚种(Lc.lactissubsp.Lactis)、帕卡乳杆菌(L.parabuchneri)和食窦魏斯氏菌(W.cibaria)。活性平均值是3U，25°和75°数据百分位的相应值是0U和45.5U。属于乳酸菌不同种的二十五个生物型显示高于55U的β-葡萄糖苷酶活性。植物乳杆菌(L.plantarum)DPPMA24W、发酵乳杆菌(L.fermentum)DPPMA114、鼠李糖乳杆菌(L.rhamnosus)DPPMAAZ1和植物乳杆菌(L.plantarum)DPPMASL33显示较高的活性(分别为202.35±7.08、163.15±6.52、146.60±5.84和144.34±7.19U)。这些生物型的β-葡萄糖苷酶活性值在箱线图加误差棒范围之外。基于这些结果，挑出这四种乳酸菌用于混合起始物的配制，所述混合起始物将用于发酵不同的大豆乳类型。
(2)大豆乳的发酵和功能化合物的合成
表1报告了用于制备大豆乳的不同大豆粉类型的化学组成、蛋白质分散性和粒径。表1显示14种大豆粉的化学组成、蛋白质分散性和粒径，所述大豆粉通过所选的混合起始物用于生产功能性化合物，所述混合起始物包括：植物乳杆菌(Lactobacillus plantarum)DSM 23755(对应于DPPMASL33)、植物乳杆菌(Lactobacillus plantarum)DSM 23756(对应于DPPMA24W)、发酵乳杆菌(Lactobacillus fermentum)DSM 23757(对应于DPPMA114)和鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758(对应于DPPMAAZ1)。
使用所选混合起始物发酵十四种大豆乳类型，所述混合起始物包括植物乳杆菌(L.plantarum)DPPMA24W和DPPMASL33、发酵乳杆菌(L.fermentum)DPPMA114和鼠李糖乳杆菌(L.rhamnosus)DPPMAAZ1。所有底物都经历了乳酸化过程(图2A)。发酵96小时后，ΔpH值从0.59±0.06至1.19±0.09变化，所述ΔpH值分别获自Provasoy 68288和低脂大豆粉。ΔpH平均值是0.93，并且25°和75°数据百分位相应值的范围是0.79和1.01。发酵后，所有大豆乳类型的pH值在范围5.1～5.3内。
在所有大豆乳类型的发酵过程中，乳酸菌都有生长(图2B)。Δlog cfu/ml的值从0.99±0.29至1.61±0.30变化，所述值分别获自来自全脂大豆粉和低脂大豆粉的大豆乳类型。细胞密度增长的平均值是1.31Δlog cfu/ml，对应于细胞密度绝对值8.31cfu/ml。对应于25°和75°的数据百分位值的范围是1.21和1.43。通过24～36小时的孵育时间完成乳酸菌的生长。通过RAPD-PCR分型测定，用于混合起始物的所有四种乳酸菌生物型在不同大豆乳类型上生长的细胞密度相似。
不同大豆乳类型的共轭异黄酮的初始浓度从142.3±12.5至171.5±10.8mM变化，从102.2±8.6至123.0±11.3mM变化，以及从10.5±1.1至18.0±0.9mM变化(分别对应大豆黄酮苷、染料木苷和黄豆黄苷)。相反，在不同大豆乳类型中观察到了低浓度(0～7.8±0.5μM)的苷元(表2)。表2显示在使用所选混合起始物的大豆乳(获自14种不同粉末)发酵过程中，异黄酮-苷元(大豆苷元、染料木素和黄豆黄素)和雌马酚的浓度(μM)。数据是三次三个重复样品实验数据的平均值。
除获自全脂大豆粉的大豆乳以外，在所有大豆乳类型的孵育过程中，苷元浓度都有所增加。在孵育96小时后，在OFS大豆乳中观察到了大豆苷元的最高浓度(57.0±4.0μM，对应于1.45mg/100ml)，接下来是Prolia 68238(50.7±2.1μM)和Prolia 68237(46.4±1.7μM)。在上述三种大豆乳类型中也有染料木素的最高终浓度(140.3±9.4-3.9mg/100ml、102.9±6.4和94.0±5.3μM，分别对应于OFS、Prolia 68238和68237)。在所有大豆乳类型中，与其它苷元相比，黄豆黄素的浓度较低。黄豆黄素的最高浓度在Prolia 68237和68238以及OFS大豆乳类型中(分别是23.9±2.4、22.5±1.3和20.4±1.0μM-0.58mg/100ml)。就从生物大豆粉(OFS)生产的大豆乳而言，共轭异黄酮转化为相应苷元的转化率是0.72、0.85和0.98(对应于大豆黄酮苷转化为大豆苷元，染料木苷转化为染料木素，以及黄豆黄苷转化为黄豆黄素)。虽然苷元浓度在孵育过程中有所增加，但24小时后所有三种共轭异黄酮的水解速率都在范围1.0～0.95内。
孵育前，在所有大豆乳类型中都没有检测到雌马酚的存在(表2)。在发酵过程中，各种大豆乳类型都不能合成雌马酚。孵育96小时后，在Prolia 68238和68237大豆乳类型中检测到的雌马酚最高浓度(分别是20.0±1.1和18.5±0.9μM)，尤其是在OFS大豆乳中(37.3±1.5μM，对应于0.9mg/100ml)。由于同时合成大豆苷元，所以无法检测其转化为雌马酚的转化率。许多研究考虑使用分离自人粪便材料的潜力益生菌，以使大豆乳富集异黄酮-苷元(Chun等，2007.Conversion of isoflavoneglucoside to aglycones in soymilk by fermentation with lactic acid bacteria(“通过乳酸菌发酵将大豆乳中的异黄酮葡萄糖苷转化为苷元”).J.Food Sci.72:39-44；Donkor和Shah 2008.Production ofβ-glucosidase and hydrolysis of isoflavonephytoestrogens by Lactobacillus acidophilus,Bifidobacterium lactis and Lactobacilluscasei in soymilk(“嗜酸乳杆菌(Lactobacillus acidophilus)、乳双歧杆菌(Bifidobacterium lactis)和干酪乳杆菌(Lactobacillus casei)在大豆乳中产生β-葡萄糖苷酶和水解异黄酮植物雌激素”).J.Food Sci.73:15-20；Pham和Shah 2007.Biotransformation of isoflavone glycosides by Bifidobacterium animalis in soymilksupplemented with skim milk powder(“在添加了脱脂奶粉的大豆乳中用动物双歧杆菌(Bifidobacterium animalis)生物转化异黄酮葡萄糖苷”).J.Food Sci.72:316-324；Tsangalis等，2002；Tsangalis等，2004；Wei等，2007.Using Lactobacillusand Bifidobacterium to product the isoflavone algycones in fermented soymilk(“使用乳杆菌和双歧杆菌在发酵大豆乳中产生异黄酮苷元”).Int.J.FoodMicrobiol.117:120-124)。所用的微生物全部是双歧杆菌或属于不同种的各种乳酸菌。本发明选择了之前从未使用过的四种新生物型用于合成异黄酮-苷元和雌马酚，所述生物型对应于植物乳杆菌(L.Plantarum)DPPMA24W和DPPMASL33、发酵乳杆菌(L.Fermentum)DPPMA114和鼠李糖乳杆菌(L.Rhamniosus)DPPMAAZ1。仅有有限数量的研究也考虑了在大豆乳的发酵过程中合成雌马酚。在用双歧杆菌发酵的大豆乳中合成了雌马酚(Tsangalis等，2002.Enzymatictransformation of isoflavone phytoestrogens in soymilk byβ-glucosidase producingbifidobacteria(“产β-葡萄糖苷酶的双歧杆菌在大豆乳中酶促转化异黄酮植物雌激素”).Food Res.Int.Sci.67:3104-3113)。在发酵24小时后，相比于使用假长双歧杆菌(Bifidobacterium pseudolongum)和长双歧杆菌(Bifidobacterium longum)生物型所获的雌马酚产量0.338和0.433mg/100ml，用动物双歧杆菌(Bifidobacteriumanimalis)合成了雌马酚最高浓度(0.521mg/100ml)。用本研究所选混合起始物发酵的OFS大豆乳含有更高浓度的雌马酚，即37.3μM，对应于0.9mg/100ml。
基于上述报道的结果，认为由生物大豆粉(OFS)生产的大豆乳是用于合成异黄酮-苷元和雌马酚的最佳底物。基于我们所知，之前没有研究考虑使用获自生物培养大豆粉的大豆乳来合成异黄酮-苷元和雌马酚。
因此，用HPLC方法检测了露纳素的浓度(Wang等，2008.Analysis of soybeanprotein derived peptides and the effect of cultivar,environmental conditions,andprocessing of lunasin concentration in soybean and soy products(“大豆蛋白源肽的分析和培养品种、环境条件和加工对于大豆和大豆制品中露纳素浓度的影响”).J.AOAC Intern.91:936-9444)。孵育前，露纳素的浓度约为3.2mg/100ml(图3)。在发酵过程中，所选混合起始物促使露纳素持续增加，在孵育96小时结束时，其浓度约为8.4mg/100ml。基于我们所知，之前没有研究考虑在由使用乳酸菌发酵的大豆乳组成的同一制剂中，相伴合成异黄酮-苷元、雌马酚和露纳素。这种生物活性肽(露纳素)的生理学效应在文献中有广泛引证(Jeong等，2003.Characterization oflunasin isolated from soybean(“分离自大豆的露纳素的表征”).J Agric Food Chem.51:7901-7906；Jeong等.2007.The cancer preventive peptide lunasin from wheatinhibits core histone acetylation(“源自小麦的抗癌肽露纳素抑制核心组蛋白的乙酰化”).Cancer Lett.255:42–48)。
基于上述结果，将已发酵的OFS大豆乳用于皮肤保护和关于人肠细胞的分析。
(3)重建表皮测试和TEER(跨上皮电阻)测量
将获自生物大豆粉并用所选混合起始物发酵的OFS大豆乳以10μM的雌马酚终浓度用于处理人重建表皮(根据模型)。该模型已经广泛实验并受科学界认可(Di Cagno等，2009.Synthesis ofγ-amino butyric acid(GABA)byLactobacillus plantarum DSMZ19463:functional grape must beverage anddermatological application(“用植物乳杆菌(Lactobacillus plantarum)DSMZ19463合成γ-氨基丁酸(GABA)：功能性未发酵葡萄汁饮料和皮肤科应用”).Appl BiotechnolMicrobiol DOI:10.1007/s00253-009-23704)。处理24小时后，进行TEER检测。该类分析已受国际科学界广泛认可，以角质层的完整性作为参考，用于评价组织的腐蚀能力和屏障功能。具体而言，凭藉该检测，可得到关于角质层水平存在的致密层状结构的信息，该信息与紧密连接的完整性和表皮厚度相关。这些因子作为一个整体来界定有效的屏障功能。图4显示在已发酵OFS大豆乳的存在下，TEER值呈现显著增长(P<0.05)，这证明了该分子在皮肤水平的保护性作用。使用化学合成雌马酚和露纳素的混合物得到了相同结果。
根据目前所知，这是首个关于包含异黄酮-苷元、雌马酚和露纳素的基于大豆乳制品的应用例，该应用例证明其具有刺激皮肤屏障功能的作用。
(4)Caco-2/TC7细胞测试
为测试由生物大豆粉(OFS)生产的大豆乳中所含异黄酮-苷元的免疫调节性质，使用中性红(NR)摄取测试，首先评价了浓度在5～100μM的标准化合物(雌马酚、大豆苷元、染料木素和黄豆黄素)对Caco-2/TC7细胞的细胞毒性。染料木素、黄豆黄素和雌马酚显示出与甲醇和DMSO(阴性对照)相似的表现，并没有对细胞增殖产生显著影响。处理72小时后，浓度高于100μM的大豆苷元显著地抑制了细胞增殖(P<0.03)。
用10μM浓度的OFS已发酵大豆乳(稀释至雌马酚终浓度为10μM)或未经发酵的大豆乳预先处理Caco-2/TC7细胞24小时。这些化合物或制品没有表现出诱导释放NO，而显示与阴性对照(即甲醇和DMSO)相似的表现(图5)。接着，每24小时，用INF-γ(1000U/ml)和LPS(100ng/ml)刺激Caco-2/TC7细胞。用阴性对照、大豆甙元或未经发酵的OFS大豆乳做保护性处理，该处理明显提高了NO的释放(P<0.05)，从而模拟了Caco-2/TC7细胞的炎性状态。相反，使用雌马酚或已发酵的OFS大豆乳的处理明显抑制了NO的释放(分别是P<0.002和P<0.007)。采用染料木素和黄豆黄素处理也观察到了NO释放受到显著抑制(P<0.05)。由于预先已证明用于测试的10μM浓度的异黄酮-苷元无毒，所以Caco-2/TC7细胞的死亡并不一定干扰NO释放。
在本研究的培养条件下，Caco-2/TC7细胞发展了肠上皮细胞的形态及功能特性，包括紧密的细胞连接，其完整性通过TEER测定测量。预先在标准化合物(10～100μM)、已发酵OFS大豆乳(稀释至雌马酚浓度为10μM)或未经发酵的OFS大豆乳的存在下测定TEER。除了雌马酚，在72小时的孵育过程中没有观察到100μM浓度的化合物(1000U/ml)对TEER的影响。用INF-γ(1000U/ml)处理Caco-2/TC7细胞促使了TEER值的显著降低(P<0.003)(图6)。当Caco-2/TC7细胞既受INF-γ刺激，又经已发酵OFS大豆乳处理时，INF-γ的负效应显著减弱(P<0.007)。在未经发酵的OFS大豆乳存在下，观察到可忽略不计的效应。染料木素、黄豆黄素、尤其是雌马酚显示与已发酵的OFS大豆乳相似的趋势。大豆苷元没有干扰由INF-γ引起的负效应。
白细胞介素-8(IL-8)是C-X-C趋化因子家族中的一员，并在嗜中性细胞的活化中起到重要作用，因而引发炎性反应。当Caco-2/TC7细胞受到白细胞介素-1β(2ng/ml)处理时，观察到IL-8合成的显著增加(P<0.001)(图7)。当Caco-2/TC7细胞受到白细胞介素-1β刺激，同时也受到雌马酚和大豆苷元处理时，观察到IL-8合成的显著减少(P<0.005)。使用已发酵的OFS大豆乳的处理观察到对IL-8合成的最高抑制(P<0.001)。相反，使用染料木素、黄豆黄素或已发酵OFS大豆乳的处理没有造成IL-8合成的减少(P<0.10)。
上述报告的结果清楚显示，由已发酵OFS大豆乳引起的人肠细胞的屏障功能刺激效应以及抗炎性，主要归因于雌马酚和一些异黄酮-苷元的存在。露纳素也可能引起附加效应。
(5)用于合成大豆苷元、染料木素、黄豆黄素、雌马酚和露纳素的生物技术方法和其在皮肤病学领域中应用的开发
如上文其它部分所述，开发了用于合成异黄酮-苷元(大豆苷元、染料木素和黄豆黄素)、雌马酚和露纳素的生物技术方法和该方法用于皮肤病学领域中的应用。所述方法包括：
a)在MRS培养基中以纯培养方式培养植物乳杆菌(L.plantarum)DPPMA24W和DPPMASL33、发酵乳杆菌(L.fermentum)DPPMA114和鼠李糖乳杆菌(L.rhamnosus)DPPMAAZ1；
b)收集、洗涤并接种细胞悬液于不同大豆乳类型，优选从按农学生物方法培养且实验室去皮的大豆粉制得的无菌大豆乳；
c)用所选混合起始物发酵大豆乳48～96小时，优选96小时，于30～37℃发酵，优选30℃；
d)离心分离细胞。依照不同的方法，所述制品还可包含乳酸菌细胞；
e)通过干燥或冷冻干燥方法使所述制品脱水；
f)通过添加合适的赋形剂制备组合物，以获得适于口服或局部给药使用的形式(视情况而定)。
实施例6：体外评估对比包含露纳素的生物质与不含露纳素的生物质对毛发生长的刺激作用。
体外研究比较包含露纳素的生物质(BL)与不含露纳素的生物质(B)作为毛发生长的促进剂。
材料与方法
在包含了2mM L-谷氨酰胺、1×抗真菌和抗生素溶液(1000u g/ml硫酸链霉素、1000单位/ml青霉素G和2.5μg/ml两性霉素B)和10％胎牛血清的培养基(达尔伯克(氏)改良伊格尔(氏)培养基(Dulbecco's modified Eagle's medium，DMEM))中培养毛乳头细胞(Derma papilla cells，DPCs)。在细胞融合时，用不含血清的DMEM培养细胞24小时，然后用不同浓度的包含或不含露纳素的生物质处理细胞。
用MTT方法测定细胞增殖(Mosmann，1983)。接种DPCs于96孔板(104个细胞/孔)，孵育24小时后加入待分析物质。用ELISA酶标仪于570nm测量吸光度。
进一步对Bcl2进行蛋白质印迹(western blot)实验。用包含Tris-HCl(50mM，pH7.4)、EDTA 2mM、亮抑蛋白酶肽(leuptin)100μg/ml和NaCl 100mM的缓冲液提取蛋白质。
通过SDS-PAGE将50ug蛋白质上样并分离。1:500稀释抗Bcl-2、Bax和肌动蛋白(actin)的单克隆抗体，用ECL系统检测抗原-抗体复合物，并用图像测密计(伯乐公司(Bio-Rad)GS-700)分析结果。
结果与讨论
在测试浓度范围内(0.01-0.5μM)，包含露纳素的生物质以剂量依赖的方式诱导了DPCs的体外增殖提高(p<0.05)(图1)。
与不含露纳素的生物质不同，包含露纳素的生物质具有引起Bcl-2蛋白表达增加和Bax蛋白表达减少的效应(图2)。
这些数据表明，包含露纳素的生物质通过对DPCs的增殖和抗凋亡效应刺激毛发生长，并可因此延长毛发生长初期阶段。
Claims (27)
1.用于制备基于已发酵大豆的混合物的方法，所述基于已发酵大豆的混合物包含异黄酮-苷元、雌马酚和露纳素lunasin，所述方法使用下述四种乳酸菌的混合物发酵大豆：植物乳杆菌(Lactobacillus plantarum)DSM 23755、植物乳杆菌(Lactobacillus plantarum)DSM 23756、发酵乳杆菌(Lactobacillus fermentum)DSM 23757和鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758。
2.如权利要求1所述的方法，其特征在于，所述方法包括以下步骤或由以下步骤组成：
a)培养增殖所述四种乳酸菌：植物乳杆菌(Lactobacillus plantarum)DSM23755、植物乳杆菌(Lactobacillus plantarum)DSM 23756、发酵乳杆菌(Lactobacillus fermentum)DSM 23757和鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758；
b)将所述乳酸菌的水性悬液接种于基于大豆的底物；
c)于30～37℃，孵育48～96小时。
3.如权利要求2所述的方法，其特征在于，所述步骤c)的孵育在30℃进行。
4.如权利要求2所述的方法，其特征在于，所述步骤c)的孵育进行96小时。
5.如权利要求2所述的方法，其特征在于，所述乳酸菌的水性悬液以1～4％总底物体积的量接种于所述底物，且在所述水性悬液中，各菌株的细胞密度为log 9.0 cfu/ml。
6.如权利要求2所述的方法，其特征在于，基于大豆的底物选自大豆粉和大豆乳。
7.如权利要求6所述的方法，其特征在于，所述大豆粉为生物大豆粉。
8.如权利要求2-7中任一项所述的方法，其还包括步骤d)离心培养物以除去乳酸菌细胞，其中所述培养物为肉汤培养物。
9.如权利要求8所述的方法，其特征在于，所述肉汤培养物的离心在4℃以10,000×g进行15 min。
10.如权利要求2-7中任一项所述的方法，其还包括步骤e)通过干燥使步骤c)中所得的培养物脱水。
11.如权利要求10所述的方法，其特征在于，所述干燥是冷冻干燥。
12.如权利要求8所述的方法，其特征在于，其还包括步骤e)通过干燥使步骤d)中所得的上清脱水。
13.如权利要求12所述的方法，其特征在于，所述干燥是冷冻干燥。
14.基于已发酵大豆的混合物，所述混合物包含异黄酮-苷元、雌马酚和露纳素，所述混合物通过如权利要求1～13中任一项所述的方法获得。
15.药物组合物，所述组合物包含如权利要求14所述的混合物以及一种或多种药学上可接受的赋形剂和/或佐剂，或者由如权利要求14所述的混合物以及一种或多种药学上可接受的赋形剂和/或佐剂组成。
16.化妆品组合物，所述组合物包含如权利要求14所述的混合物以及一种或多种化妆品中可接受的赋形剂和/或佐剂，或者由如权利要求14所述的混合物以及一种或多种化妆品中可接受的赋形剂和/或佐剂组成。
17.权利要求14所述的混合物以所述混合物本身或联合一种或多种赋形剂和/或佐剂在作为食品补足剂中的应用。
18.一种食品补足剂，其包含权利要求14所述的混合物本身或其与一种或多种赋形剂和/或佐剂的联合。
19.如权利要求14所述的混合物或如权利要求15所述的药物组合物在制备用于治疗皮肤或肠壁的紊乱或疾病的组合物中的应用。
20.如权利要求14所述的混合物或如权利要求15所述的药物组合物或如权利要求16所述的化妆品组合物的应用，其用于化妆品应用。
21.如权利要求20所述应用的应用，所述化妆品应用是用于治疗毛发脱落。
22.如权利要求14所述的混合物或如权利要求15所述的药物组合物或如权利要求16所述的化妆品组合物在制备用于治疗脱毛症或休止期脱发的组合物中的应用。
23.以下四种乳酸菌的混合物：植物乳杆菌(Lactobacillus plantarum)DSM23755、植物乳杆菌(Lactobacillus plantarum)DSM 23756、发酵乳杆菌(Lactobacillus fermentum)DSM 23757和鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758。
24.乳酸菌植物乳杆菌(Lactobacillus plantarum)DSM 23755。
25.乳酸菌植物乳杆菌(Lactobacillus plantarum)DSM 23756。
26.乳酸菌发酵乳杆菌(Lactobacillus fermentum)DSM 23757。
27.乳酸菌鼠李糖乳杆菌(Lactobacillus rhamnosus)DSM 23758。
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| ITRM2010A000378 | 2010-07-12 | ||
| ITRM2010A000378A IT1405780B1 (it) | 2010-07-12 | 2010-07-12 | Miscela arricchita di isoflavoni-agliconi, equolo e lunasina a base di soia fermentata, procedimento per la sua preparazione e relativi usi in campo alimentare, medico e cosmetico. |
| PCT/IT2011/000240 WO2012007978A2 (en) | 2010-07-12 | 2011-07-11 | Fermented soya based mixture comprising isoflavones- aglicones, equol and lunasil, process for the preparation and uses thereof in food, medical and cosmetic fields. |
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| CN103140588B true CN103140588B (zh) | 2015-09-30 |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| JP6322580B2 (ja) * | 2012-12-11 | 2018-05-09 | キッコーマン株式会社 | 豆乳発酵エキスおよび胚軸発酵エキス |
| WO2014090259A1 (en) * | 2012-12-12 | 2014-06-19 | Herrens Mark Aps | Product comprising red clover extract and methods for producing the same |
| CN103933548A (zh) * | 2014-04-17 | 2014-07-23 | 中国药科大学 | 露那辛多肽在防治白内障药物中的应用 |
| CN108138124A (zh) | 2015-08-31 | 2018-06-08 | 科.汉森有限公司 | 具有抗真菌活性的发酵乳杆菌细菌 |
| KR102394638B1 (ko) * | 2015-09-30 | 2022-05-09 | (주)아모레퍼시픽 | 유산균 유래의 세포외 소낭을 포함하는 탈모 방지 또는 육모 촉진용 조성물 |
| KR102132827B1 (ko) * | 2015-10-21 | 2020-07-22 | 주식회사 단정바이오 | 락토바실러스 람노서스 비타피원 균주, 이를 이용한 칡 또는 대두 배아 발효물의 제조방법 및 이를 이용한 칡 또는 대두 배아 발효물 |
| CN105596275B (zh) * | 2016-01-06 | 2018-07-27 | 名臣健康用品股份有限公司 | 一种含有双菌发酵豆奶提取物的化妆品基质及应用 |
| JP6626869B2 (ja) * | 2016-10-03 | 2019-12-25 | 株式会社バイオジェノミクス | 善玉菌生産物質の製造方法及び食品 |
| CN106520625B (zh) * | 2016-12-05 | 2019-09-10 | 厦门和美科盛生物技术有限公司 | 一种共生乳酸菌发酵液、制备方法及其用途 |
| CN106399208B (zh) * | 2016-12-05 | 2019-09-10 | 厦门和美科盛生物技术有限公司 | 一种适合食品加工应用的乳酸菌发酵液、制备方法及用途 |
| KR101840376B1 (ko) * | 2017-10-20 | 2018-03-20 | (주)코엔바이오 | 탈모 예방, 발모 촉진 또는 성기능 개선능이 있는 락토바실러스 퍼멘텀 균주 및 이를 포함하는 조성물 |
| CN108338946A (zh) * | 2018-03-12 | 2018-07-31 | 黑龙江军门现代农业发展有限公司 | 一种含有丝肽的美容面膜及其制备方法 |
| JP2018186823A (ja) * | 2018-07-13 | 2018-11-29 | 株式会社ダイセル | イソフラバノン類の製造方法 |
| CN109223603B (zh) * | 2018-09-21 | 2021-01-29 | 黑龙江省中医药科学院 | 治疗脱发组合物的制备方法 |
| CN109402000A (zh) * | 2018-10-31 | 2019-03-01 | 南京师范大学 | 一株产β-葡萄糖苷酶乳酸菌及其筛选方法和富含活性黄酮苷元酸奶的制备方法 |
| KR20240023456A (ko) * | 2022-08-11 | 2024-02-22 | 코스맥스 주식회사 | 천연 비타민을 포함하는 복합발효물의 탈모 방지 및 발모 촉진용 조성물 |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2006093408A2 (en) * | 2005-03-01 | 2006-09-08 | Csm Nederland B.V. | Bacteria that naturally overproduce folate |
| CN1944634A (zh) * | 2006-03-08 | 2007-04-11 | 沈阳农业大学 | 水解大豆异黄酮糖苷酶工程菌株、其构建方法及其用途 |
Family Cites Families (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5026553A (en) * | 1989-08-07 | 1991-06-25 | Dale E. Swinney | Swinney's hair growth formula |
| KR100454760B1 (ko) * | 1997-01-22 | 2005-01-13 | 후지 세이유 가부시키가이샤 | 발효두유(豆乳)제조방법 |
| WO1998042200A1 (fr) * | 1997-03-20 | 1998-10-01 | Jean James Garreau | Produits alimentaires a base de lait de soja et leur procede de fabrication |
| ES2262240T3 (es) * | 1997-07-05 | 2006-11-16 | Societe Des Produits Nestle S.A. | Postre congelado con bacterias lacticas y fibras fermentables. |
| AUPP112497A0 (en) * | 1997-12-24 | 1998-01-22 | Novogen Research Pty Ltd | Compositions and method for protecting skin from UV induced immunosupression and skin damage |
| JP4163276B2 (ja) * | 1998-02-05 | 2008-10-08 | 独立行政法人理化学研究所 | 機能性組成物 |
| BR9914476B1 (pt) * | 1998-10-09 | 2010-11-30 | composição cosmética. | |
| AUPQ008299A0 (en) * | 1999-04-30 | 1999-05-27 | G.J. Consultants Pty Ltd | Isoflavone metabolites |
| FR2831395B1 (fr) * | 2001-10-25 | 2004-06-11 | Triballat Laiteries | Procede de fabrication de produits fermentes a base de soja, produits fermentes obtenus par ce procede et ferments correspondants |
| US20060251750A1 (en) * | 2002-09-30 | 2006-11-09 | Tabor Aaron T | Soy formulations and their use in skin care |
| AU2005211165B8 (en) * | 2004-01-28 | 2011-07-14 | Nestec S.A. | Nutritional composition for improving skin condition and preventing skin diseases |
| ES2316146T3 (es) * | 2006-07-07 | 2009-04-01 | Alpro Nv | Metodo para preparar un analogo de producto lacteo. |
-
2010
- 2010-07-12 IT ITRM2010A000378A patent/IT1405780B1/it active
-
2011
- 2011-07-11 RS RS20170403A patent/RS55900B1/sr unknown
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2006093408A2 (en) * | 2005-03-01 | 2006-09-08 | Csm Nederland B.V. | Bacteria that naturally overproduce folate |
| CN101203599A (zh) * | 2005-03-01 | 2008-06-18 | 荷兰Csm有限公司 | 天然高产叶酸菌株 |
| CN1944634A (zh) * | 2006-03-08 | 2007-04-11 | 沈阳农业大学 | 水解大豆异黄酮糖苷酶工程菌株、其构建方法及其用途 |
Non-Patent Citations (1)
| Title |
|---|
| 产胞外β-葡萄糖苷酶乳酸菌的筛选及其酶学性质的初步研究;万振堂 等;《食品与发酵工业》;20090430;第35卷(第4期);第28－32、37页 * |
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| Publication number | Publication date |
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| CN103140588A (zh) | 2013-06-05 |
| US20130231291A1 (en) | 2013-09-05 |
| LT2593570T (lt) | 2017-06-26 |
| HRP20170603T1 (hr) | 2017-06-30 |
| CA2803322C (en) | 2020-02-25 |
| SI2593570T1 (sl) | 2017-08-31 |
| WO2012007978A2 (en) | 2012-01-19 |
| EP2593570A2 (en) | 2013-05-22 |
| HUE032690T2 (en) | 2017-10-30 |
| JP2013538187A (ja) | 2013-10-10 |
| RU2013105731A (ru) | 2014-08-20 |
| ITRM20100378A1 (it) | 2012-01-13 |
| MX2013000353A (es) | 2013-03-22 |
| EP2593570B1 (en) | 2017-03-01 |
| JP5951604B2 (ja) | 2016-07-13 |
| RS55900B1 (sr) | 2017-09-29 |
| CA2803322A1 (en) | 2012-01-19 |
| BR112013000778A2 (pt) | 2016-06-07 |
| IT1405780B1 (it) | 2014-01-24 |
| WO2012007978A3 (en) | 2012-05-18 |
| RU2564576C2 (ru) | 2015-10-10 |
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