CN116270987A - 具有美白、抗氧化、祛黄褐斑功能的中药组合物及其制备方法 - Google Patents
具有美白、抗氧化、祛黄褐斑功能的中药组合物及其制备方法 Download PDFInfo
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Abstract
本发明涉及一种具有美白、抗氧化、祛黄褐斑功能的中药组合物及其制备方法,本发明将大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥等生产工艺制成大米肽粉更易被人体吸收,将中药材通过粉碎、榨汁、二次发酵、膜过滤、真空冷冻干燥、低温超微粉碎技术处理制得含SOD中药发酵粉,将这些先进技术创造性地组合在一起,摸索出相应的操作步骤及工艺参数,应用于具有美白、抗氧化、祛黄褐斑功能的中药组合物制备方法中,尽可能地保留药物、食材的主要功效成分,在服用量显著下降的同时也提升了功效及生物利用度。
Description
技术领域
本发明涉及功能性组合物技术领域,具体涉及一种美白、抗氧化、祛黄褐斑功能的中药组合物及其制备方法。
背景技术
皮肤黑化的主要因素是黑色素产生、转运及沉着。黑色素是由位于表皮基底层的黑素细胞产生的,黑素细胞约占基底细胞的10%。黑素产生后,黑色素细胞借助树枝状突起向基底细胞和棘层细胞输送黑素颗粒。黑色素产生后,转移至角质层沉积下,皮肤“变黑”,而这些也会随着角质层的代谢而脱落。而酪氨酸是形成黑色素非常重要的原料,酪氨酸在黑色素细胞内通过酪氨酸酶及多种氧化酶的催化生成黑色素。产生的黑色素一部分进入毛细血管,最终被肾脏代谢;另一部分则转移至角质层沉积,导致皮肤“变黑”。内外因素会共同作用,影响黑素细胞功能与皮肤色度。外因有紫外线、污染等,内因有遗传、激素、炎症水平等。因此,防止紫外线照射以及保持良好的身体状态对于美白是非常重要的。结合黑色素生成途径及皮肤“变黑”的原因,抑制黑色素的生产及转运是美白的主要途径。
美白原料都是围绕黑色素产生的机理以及信号转导通路的调控发挥作用的。但不同原料作用机理不同,给皮肤带来的影响也是相差很大的,如敏感肌和皮肤屏障较弱的人群就要慎重选择含酸类的美白产品。
常见的美白作用机理有以下几种:抑制黑色素生成路径中各环节所需的酶(如酪氨酸酶)活性及表达;抗氧化剂可以还原黑色素生成环节中的氧化产物,抑制多巴的氧化产物多巴醌的产生,从而抑制黑色素的产生,达到美白效果;抑制已经生成的黑色素向角质层转运,使其无法因顺利到达角质层而引起的视觉上的“变黑”;黑素细胞的代谢受到角质形成细胞、成纤维细胞等释放的多种调控因子的影响,黑素合成相关的信号转导通路较为复杂,多种信号转导因子都会促进黑色素的合成,调控阻碍信号转导可以减少黑色素的合成;加快皮肤新陈代谢及角质层脱落;其他间接的美白方式,如防晒措施可以避免因紫外线照射引发的黑色素生成而达到间接美白的效果。另外,生活中养成健康的饮食习惯和作息安排,保持良好的身体状态,也可使皮肤在视觉上透亮光泽。
近年来,过量自由基所产生的氧化性损害及其在人体疾病所扮演的角色,日益受到重视。自由基是一种具有高度活性的粒子,能氧化许多重要的生物分子,如蛋白质、脂质、脱氧核糖核酸(DNA)等。而这些主要生物分子的大量被氧化,已发现与多类的疾病有关,包括肾脏病、动脉硬化、缺血性心脏病、糖尿病、发炎性疾病、癌症、神经病、高血压、败血症、急性外伤、眼睛疾病(如白内障)、肺部疾病、血液病等,也与器官移植和药物毒性有关。所以在人体内,过量自由基也就成为许多疾病包括慢性病的共同起源。氧在体内进行正常的代谢过程中会形成不稳定的自由基,超氧阴离子就是自由基的一种。由于其不稳定,易从其他分子中夺取电子,形成新的稳定物质,同时又可能产生了一个新的自由基。当其从细胞膜上夺取电子,且体内的抗氧化剂耗尽时,过量产生的自由基严重损害与接触的任何胶原蛋白及其它结缔组织,引起细胞膜的过氧化伤害,释放导致炎症的介质。这些介质能够抑制免疫系统,干扰人体的DNA密码,造成多种危害生命的疾病。而OPC遇到自由基时,它能将电子“馈赠”给自由基,而自身又不会因被氧化形成新的自由基,从而可切断形成自由基的反应链,通过这种中和作用清除自由基。衰老是一个非常复杂的多因素过程,以细胞功能生理生化方面的损害为特征,与自由基生成增加紧密相关。
多酚是在植物性食物中发现的天然化合物,可作为抗氧化剂并可能降低疾病风险。多酚类物质包括类黄酮、酚酸、多酚酰胺、白藜芦醇、姜黄素和木脂素等。潜在的健康益处包括降低患心脏病、糖尿病、癌症、炎症和认知能力下降的风险。多酚在人体内充当抗氧化剂的角色,这意味着它们有助于保护身体并中和对细胞造成损害的自由基。自由基是由正常细胞过程和外部因素(如辐射、空气污染、吸烟和化学暴露)产生的高活性氧分子。如果没有抗氧化剂来中和自由基,细胞就会受伤,从而增加罹患糖尿病、心脏病、恶性肿瘤等健康问题的风险。
黄褐斑是一种色素代谢障碍性疾病,目前病因尚不清楚。研究认为,需氧生物在新陈代谢过程中,一些氧化反应都伴随着自由基形成。如黄嘌呤氧化酶催化黄嘌呤或次黄嘌呤氧化生成尿酸,同时可形成超氧阴离子、自由基等许多酶的氧化反应。空气中氧化性污染物如O2、NO2、NO等也能在体内启动自由基反应,光线照射(特别是紫外线)也能使H2O生成OH-等。自由基是带有未成对电子的分子,由于未成对电子趋于成对,所以反应极为活泼。机体内一面不断产生自由基,而另一面又有一套清除自由基的机制,使其产生与清除达到平衡,使机体免受损伤。随年龄增长,自由基水平上升,而清除自由基的酶活力下降。机体内产生过多自由基,导致许多新陈代谢过程的破坏,引起多种疾病并加速机体老化。尤其是体内氧自由基(亦称活性氧),当其攻击机体内细胞内蛋白质、核酸和脂类时,使之发生交联、变性、沉积,形成比原有分子大许多倍的聚合物-脂褐素,进而形成黄褐斑,使皮肤改变。临床上治疗黄褐斑主要通过药物治疗,黄褐斑是多见于中青年女性面部对称性的色素沉着性皮肤病,其原因包括紫外线照射、化妆品、妊娠、内分泌紊乱、口服避孕药、过度疲劳、种族及遗传等。黄褐斑的治疗通常包括系统药物治疗和外用药物治疗。常见的系统治疗药物为氨甲环酸片、维生素C片、维生素E软胶囊。常见的外用治疗药物为氢醌乳膏、壬二酸乳膏、复方熊果苷乳膏、维A酸乳膏。但是“是药三分毒”,况且每个人对于药物的适应程度不一样,有可能会产生严重的副作用而使人们无法接受。
研究表明老年动物机体内SOD、GSH-PX活性显著下降,MDA水平明显升高,这和文献报道结果一致,由于机体内抗氧化能力减弱,自由基水平的升高,致使机体内氧化反应加速,反应活泼的氧自由基攻击机体细胞的蛋白质核酸和脂类时,使细胞功能丧失,生物分子交联变性,形成比原有分子大许多倍的聚合物—脂褐素,进而形成黄褐斑,使皮肤颜色改变。研究认为随年龄增长机体氧化与抗氧化反应失衡是引起黄褐斑形成发展的原因之一。
我国人口基数大,爱美人士特别普遍,且皮肤不好,过氧化体质或有黄褐斑的人群基数特别庞大,迫切需要开发出能够真正实现美白、抗氧化、祛黄褐斑功能且副作用较小的产品。因此充分利用中华传统中药、药食同源食材、新食品原料、天然食物等宝贵资源,系统、完善地研究、利用及开发出具有美白、抗氧化、祛黄褐斑功能的产品,普及推广为大众享用,对提高我国人民健康水平具有极其深远的现实意义,同时也会带来巨大的社会效益和经济效益。
在现有技术中,申请号CN201711055559.4、名为含有何首乌干细胞提取物的抗衰老护肤品及其制备方法的中国发明专利公开了何首乌干细胞的制备方法以得到所述何首乌干细胞提取物。何首乌具有较好的抗衰老功效,这是因为何首乌干细胞提取物可以快速激活休眠成纤维细胞,促进细胞分裂、增殖,提升细胞DNA损伤修复能力,促进细胞自我修复更新,分泌胶原物质,调节胶原纤维,增强皮肤弹性,具有强抗氧化性,清除自由基,缓解色素色斑沉着,同时赋予肌肤充足水分和超强锁水能力,让皮肤富有弹性、紧致光滑。但据《何首乌主要成分大黄素、大黄酸和二苯乙烯苷对肝细胞、肝癌细胞的影响》,何首乌的大黄素、大黄酸等成分,具有肝毒性,会引发不良反应,在高浓度、长时间作用下有细胞毒作用。因此,需要寻找一种能够替代何首乌主要成分的产品。
发明内容
本发明所要解决的第一个技术问题是针对现有技术的现状,提供一种具有美白、抗氧化、祛黄褐斑功能的中药组合物。
本发明所要解决的第二个技术问题是针对现有技术而提供上述具有美白、抗氧化、祛黄褐斑功能的中药组合物的制备方法。
本发明解决第一个上述技术问题所采用的技术方案为:该具有美白、抗氧化、祛黄褐斑功能的中药组合物,其特征在于:按重量计,采用以下原料组分制备而成:
本发明组合物配方的原料中,大米肽粉是一种食源性低聚肽,其是以大米蛋白为原料,经过生物酶解制备而得小分子低聚肽,源于大米蛋白但优于大米蛋白,富含多种人体必需氨基酸,具有抑制黑色素生成、美白护肤、减少皮肤皱纹、抵抗衰老、抗氧化、清除体内自由基以及辅助降血压的作用;
含SOD中药发酵粉源自中药材莳萝、曼陀茄根、酢浆草、荜澄茄经过酵母菌和乳酸菌发酵而得的中药发酵粉,富含SOD、花青素、植物多酚等功效成分,具有抗氧化、美白、祛黄褐斑等多重功效;
异叶青兰为多年生草本植物的干燥全草,味甘性寒,有助于提高耐缺氧能力,可降低由缺氧引起的红细胞体积增大,而且还可以降低红细胞的数量,从而降低血液的粘滞性,改善血循环。镇咳祛痰,抑菌消炎,清泻肝热,也可用于牙龈肿痛,内出血及口腔溃疡;
柠檬马鞭草又名橙香木,是马鞭草科柠檬马鞭草的干燥叶片,具有松弛精神紧张、回复元气、能利尿减肥,刺激肝胆、治疗支气管炎、鼻塞、咽喉疼痛。特别值得一提的是其富含多酚成分及天然植物SOD成分,柠檬马鞭草多酚及天然植物SOD具备超强的美白能力及抗氧化能力,通过抑制黑色素细胞中酪氨酸酶mRNA的表达,以及对黑色素形成过程中多个基因控制点的抑制实现美白的功效,还具有抵抗紫外线辐射,清除或抑制自由基生成,抑制脂质过氧化、调节抗氧化相关酶活性等机制发挥抗氧化作用;
蜜蜂花是野芝麻亚科多年生草本植物干燥全草,味苦涩性平;清热解毒,用于风湿麻木,皮肤瘙痒,疮疹,崩漏等症,其类黄酮含量较高,能充分发挥其抗氧化的功效;
海葱,别名虎眼万年青,风信子科海葱的干燥叶及鳞茎,舒筋活血,对于跌打扭伤、肌肉过度疲劳有良好的恢复作用,特别注意的是其多酚含量很高,具有强大的美白作用,可以深入皮肤内部并保持高活性,可消除自由基抗氧化,有效抑制黑色素生成过程中多种酶的活性,特别是抑制酪氨酸酶活性,同时还具有防止皮肤粗糙和抗炎、抗菌的功效,是皮肤美白和抗衰老非常有效的活性物;
黑果腺肋花楸果萃取物源自新食品原料黑果腺肋花楸果,是黑色野樱莓的成熟果实,蔷薇科腺肋花楸属灌木浆果,具有美白、提高免疫力、抗氧化的功效;
关山樱花萃取物源自新食品原料关山樱花,关山樱花是蔷薇科李亚科樱属关山樱的花,所含的樱花苷和樱花素,是樱花两大具有卓越抗氧活性的黄酮素,能极大促进糖分代谢,具有抑制糖化作用,还能抑制酪氨酸酶的活性,进而抑制黑色素具有美白效果。另外,通过防止糖化,促进了胶原蛋白和骨胶原的产生,达到柔软弹性肌肤、抑制皮肤炎症的作用;
线叶金雀花萃取物源自新食品原料线叶金雀花,线叶金雀花是南非西开普敦地区生长的一种草本针状灌木,富含抗氧化类黄酮,能起到清除人体自由基,调节人体机能,预防和控制多种疾病、防止细胞DNA受损,延缓衰老的功效;
桑白皮提取物源自中药桑白皮,桑白皮为桑科植物桑的干燥根皮,味甘性寒,归肺经,功能为泻肺平喘,利水消肿,用于肺热喘咳,水肿胀满尿少,面目肌肤浮肿,另外桑白皮能修复受损的细胞,淡化瘢痕,治疗皮肤损,还有营养皮肤、美白的特别作用;
半枝莲提取物源自中药半枝莲,半枝莲为唇形科植物半枝莲的干燥全草,味辛、苦、性寒,归肺、肝、肾经;功能为清热解毒,化瘀利尿,用于疔疮肿毒,咽喉肿痛,跌扑伤痛,水肿,黄疸,蛇虫咬伤;
菝葜提取物源自百合科植物中药菝葜的干燥根茎,味甘、微苦、涩,性平,归肝、肾经;功能为利湿去浊,祛风除痹,解毒散瘀,用于小便淋浊,带下量多,风湿痹痛,疔疮痈肿,能清热燥湿,泻火解毒;
菟丝子提取物源自中药菟丝子,菟丝子为旋花科植物南方菟丝子或菟丝子的干燥成熟种子,味辛、甘,性平,归肝、肾、脾经,功能为补益肝肾,固精缩尿,安胎、明目、止泻,更能消风祛斑,用于肝肾不足,腰膝酸软,阳痿遗精,遗尿尿频,肾虚胎漏,胎动不安,目昏耳鸣,脾肾虚泻。
优选地,所述大米肽粉,由大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥处理而获得,上述步骤获得的大米肽粉中分子量小于1000D的低聚肽成分占91.42%~92.56%;
SOD中药发酵粉中SOD含量达45000IU/g,多酚含量达2.0~5.0%,原花青素含量达15~25%;
异叶青兰、柠檬马鞭草、蜜蜂花、海葱制成的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚中的多酚含量均达75%~82%;
黑果腺肋花楸果萃取物中的原花青素达10.5%~12.3%、关山樱花萃取物中的樱花苷和樱花素分别达2.5%~2.9%、1.2%~1.6%、线叶金雀花萃取物中的总黄酮含量达25%~35%;
桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物,其功效成分分别为45%~50%的粗多糖,5.0%~6.0%的野黄芩苷,52%~56%的粗多糖和1.8%~2.4%的金丝桃苷。
为解决第二个技术问题,本发明提供一种制备如上述的具有美白、抗氧化、祛黄褐斑功能的中药组合物的方法,其特征在于:包括以下步骤:
(1)将桑白皮、半枝莲、菝葜、菟丝子通过超临界CO2萃取技术+真空冷冻干燥+低温超微粉碎技术处理制成桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物;
(2)将黑果腺肋花楸果、关山樱花、线叶金雀花经过水蒸气蒸馏、提取、浓缩、真空冷冻干燥、粉碎、沸腾制粒工艺处理制得黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物;
(3)将大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥等生产工艺制成大米肽粉;
(4)将中药材莳萝、曼陀茄根、酢浆草、荜澄茄通过粉碎、榨汁、二次复合发酵、膜过滤、真空冷冻干燥、低温超微粉碎技术处理制得含SOD中药发酵粉;
(5)将异叶青兰、柠檬马鞭草、蜜蜂花、海葱经过超临界CO2萃取、膜分离、离子交换及真空冷冻干燥技术处理制得纯度高的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚;
将上述步骤制成的桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物、黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物、大米肽粉、含SOD中药发酵粉以及异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚分别称量后,投入三维混合机中混合15~20min至完全均匀,混合频率15~20Hz,过60~100目筛网,即得本发明中药组合物。
进一步地,所述步骤(3)中大米肽粉可由以下制备方法制得:
(a)制粒:大米蛋白采用沸腾制粒,溶剂为水,溶剂添加量为10~15%,制粒温度为140~155℃,制成16~20目之间的大米蛋白颗粒;
(b)真空爆破:将所述大米肽粉颗粒在1.8~2.5Mpa饱和蒸气压下维持120~140s后,在15~30ms内释放压力,并迅速抽真空至真空度-60~-80kpa之间,保持80~120s,释放真空即得大米蛋白真空爆破组织;
(c)调浆:将大米蛋白真空爆破组织粉碎过80~100目筛,用纯净水配制成大米蛋白溶液,将大米蛋白溶液混合均匀,使之均匀分散在水溶液中。将混合均匀大米蛋白粉加入适当比例的纯净水,得到其分散液;调节大米蛋白质粉的分散液的pH至6~8;
(d)酶解:加热至50℃,保持其温度在50±5℃,在大米蛋白分散液中加入地衣芽孢杆菌蛋白酶和羧肽酶,均匀搅拌酶解1~3h,其中地衣芽孢杆菌蛋白酶加入量为底物含量的0.2%~0.4%,羧肽酶加入量为底物含量的0.05%~0.1%,酶解结束后用高温瞬时灭酶法进行灭酶;
(e)离心过滤:灭酶后的酶解液体经过离心机13000~16000rpm离心分离,去除杂质,收集清液,启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为25~50pm,滤膜孔径为2~4μm,操作压力为0.05~0.12MPa,确保滤液澄清透明,去除滤渣;
(f)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离氨基酸;
(g)真空浓缩:将离子交换后的溶液进行真空减压浓缩,浓缩温度60~80℃,真空度-0.085~-0.095Mpa,得低聚肽浓缩液,相对密度控制在1.05~1.09之间;
(h)喷雾干燥:将上述低聚肽浓缩液,进行喷雾干燥,工艺参数为喷口温度175~190℃,出口温度90~95℃,上料速度2.5~3.0L/min,得所需符合质量要求的大米肽粉。
大米肽粉氨基酸分布以及分子量分布数据如表1、表2所示。
表1大米肽粉氨基酸分布表
色氨酸 | 0.54 | 异亮氨酸 | 2.74 |
胱氨酸 | 0.18 | 亮氨酸 | 4.97 |
天门冬氨酸 | 7.74 | 酪氨酸 | 1.7 |
苏氨酸 | 3.24 | 苯丙氨酸 | 2.64 |
丝氨酸 | 4.26 | 赖氨酸 | 3.52 |
谷氨酸 | 15.4 | 组氨酸 | 1.58 |
甘氨酸 | 4.38 | 精氨酸 | 7.64 |
丙氨酸 | 4.71 | 脯氨酸 | 2.92 |
缬氨酸 | 3.8 | 蛋氨酸 | 0.95 |
表2大米肽粉分子量分布表
进一步地,所述步骤(2)中黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物可由以下制备方法制得:
(a)水蒸气蒸馏:将黑果腺肋花楸果或关山樱花或线叶金雀花经过破碎处理,过4~6目筛网;破碎组织经过水蒸气蒸馏2~4h,收集挥发油及药渣;
(b)提取浓缩:将药渣加药渣重8~10倍水,提取1~2次,1~2h/次,200目滤过。真空浓缩至相对密度为1.05~1.08的提取浓缩液;
(c)真空冷冻干燥:将上述提取浓缩液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-28~-32℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干萃取物;
(d)粉碎:将上述冻干萃取物粉碎,过80~100目筛;
(e)沸腾制粒:将上述挥发油和冻干粉末使用小型沸腾制粒的低温喷雾、沸腾制粒,喷雾温度为30~45℃,充分制粒,沸腾干燥制成挥发油包埋粉,即得黑果腺肋花楸果萃取物或关山樱花萃取物或线叶金雀花萃取物。
进一步地,所述步骤(5)中异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚可由以下制备方法制得:
(a)粉碎:将异叶青兰或柠檬马鞭草或蜜蜂花或海葱中药饮片粉碎成颗粒,颗粒过6~8目筛;
(b)超临界CO2萃取:将上述颗粒用CO2超临界萃取法提取,夹带剂为68%~75%(V/V)食用酒精,夹带剂添加量为投料量的5%~6%,萃取温度28~32℃,萃取压力29~31Mpa,CO2流量15~18kg/h,萃取时间70~90min,分离压力6-8MPa,分离温度28-32℃,得到萃取液;
(c)膜分离:启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为60~75μm,滤膜孔径为6~8μm,操作压力为0.15~0.25MPa,确保滤液澄清透明,去除滤渣;
(d)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离杂质;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-32~-35℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干物;冻干物粉碎,过80~100目筛即得异叶青兰多酚或柠檬马鞭草多酚或蜜蜂花多酚或海葱多酚。
进一步地,所述步骤(4)中含SOD中药发酵粉可由以下制备方法制得:
(a)粉碎:将中药材莳萝、曼陀茄根、酢浆草、荜澄茄以1~3:2~4:1~2:0.5~1.0的比例混合破碎,过30~60目筛;
(b)榨汁:将上述组织破碎物加入1.5~3倍的纯化水充分浸泡1~3h,温度保持在45~55℃,浸泡浓浆经过大型滚筒式榨汁机榨汁处理,离心去杂,即得澄清离心液,离心转速为16000~18000r/min;
(c)二次复合发酵:将离心液中加入乳酸克鲁维酵母,酵母添加量为离心液重量的0.15~0.25%,发酵温度为28~35℃,发酵时间为4~6h,发酵结束后继续加入植物乳杆菌,其菌株号为CGMCC NO.1258和鼠李糖乳杆菌其菌株号为R0011,两者乳杆菌的添加比例为1~2:0.5~1.2,两种乳杆菌的总添加量为离心液重量的0.3~0.35%,发酵温度为36~45℃,发酵时间为3~4h,发酵结束后过400~500目筛网;
(d)膜过滤:启动膜过滤设备,上述发酵清液经过微孔滤膜过滤,滤膜厚度为35~45μm,滤膜孔径为2~8μm,操作压力为0.05~0.12MPa,确保滤液澄清透明,去除滤渣;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-38~-45℃,预冻速率1.2~1.4℃/min,预冻终点温度-80℃,干燥室压力25~32pa,加热板温度35~40℃,干燥时间为14~16h,得冻干物;
(f)低温超微粉碎:将上述冻干物经过低温超微粉碎机处理,过400~500目筛,粉碎温度为18~25℃,即得含SOD中药发酵粉。
特别值得一提的是,我们选用的乳酸克鲁维酵母筛选自青藏高原日喀则卓木拉日雪山海拔3000米以上自然生长的天山雪莲,其发酵产生SOD的能力特别突出,属于稀缺菌种资源。上述中药材由于采用了粉碎、榨汁、二次复合发酵、膜过滤、真空冷冻干燥、低温超微粉碎技术处理,制成的含SOD中药发酵粉中SOD含量高达45000IU/g,多酚含量高达2.0~5.0%,原花青素含量高达15~25%。
与现有技术相比,本发明的优点在于:
(1)本发明针对传统醇提有溶剂残留,水提法提取杂质较多且成分易氧化的状况,将桑白皮、半枝莲、菝葜、菟丝子通过超临界CO2萃取技术+真空冷冻干燥+低温超微粉碎技术处理制成中药提取物,保留并富集了中药中主要的活性成分;本发明针对易挥发活性成分在提取时易损失的情况,将黑果腺肋花楸果、关山樱花、线叶金雀花经过水蒸气蒸馏、提取、浓缩、冻干、粉碎、沸腾制粒工艺处理制得萃取物;异叶青兰、柠檬马鞭草、蜜蜂花、海葱经过超临界CO2萃取+膜分离+离子交换+真空冷冻干燥工艺处理制成高纯度的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚强化其抗氧化、美白、祛黄褐斑的效果;本发明将大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥等生产工艺制成大米肽粉更易被人体吸收,将中药材通过粉碎、榨汁、二次发酵、膜过滤、真空冷冻干燥、低温超微粉碎技术处理制得含SOD中药发酵粉,将这些先进技术创造性地组合在一起,摸索出相应的操作步骤及工艺参数,应用于具有美白、抗氧化、祛黄褐斑功能的中药组合物制备方法中,尽可能地保留药物、食材的主要功效成分,在服用量显著下降的同时也提升了功效及生物利用度;
(2)本发明完全遵循中药养生的健康理念,充分利用中华传统中药、药食同源食材、新食品原料、天然食物等宝贵资源,开发出具有美白、抗氧化、祛黄褐斑功能且不良反应或副作用较小的产品。多种多酚原料结合中药提取物、含SOD中药发酵粉与大米肽,按照本发明技术方案复配,可以协同增效,充分实现其美白、抗氧化、祛黄褐斑的功能。即:本发明中药组合物能在清除体内毒素、祛除氧自由基的同时实现皮肤肤色的变白和黄褐斑斑点的淡化消失。
附图说明
图1为本发明实施例2中人群试食实验整体变白人数数据统计的结果图;
图2为本发明实施例2中人群试食实验平均变白程度数据统计的结果图;
图3为本发明实施例3中该款中药组合物清除DPPH自由基能力的结果图;
图4为本发明实施例3中该款中药组合物清除超氧阴离子自由基能力的结果图;
图5为本发明实施例3中该款中药组合物和VC对羟基自由基的清除效果的结果图;
图6为本发明实施例3中VC和该款中药组合物总还原力测定的结果图。
具体实施方式
以下结合实施例对本发明作进一步详细描述。
按重量组分计,本发明实施例1~3的配方如下:
原辅料 | 方案1 | 方案2 | 方案3 |
大米肽粉 | 24 | 40 | 24 |
含SOD中药发酵粉 | 28 | 11 | 25 |
异叶青兰多酚 | 1 | 0.5 | 1 |
柠檬马鞭草多酚 | 5 | 3 | 5 |
蜜蜂花多酚 | 0.5 | 1 | 1 |
海葱多酚 | 1.5 | 1 | 1.2 |
黑果腺肋花楸果萃取物 | 13.5 | 5 | 9.8 |
关山樱花萃取物 | 9 | 4 | 9 |
线叶金雀花萃取物 | 4 | 7.5 | 9 |
桑白皮提取物 | 2 | 5 | 3 |
半枝莲提取物 | 3 | 8 | 5 |
菝葜提取物 | 5 | 10 | 5 |
菟丝子提取物 | 3.5 | 4 | 2 |
合计 | 100 | 100 | 100 |
所述大米肽粉,由大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥处理而获得,上述步骤获得的大米肽粉中分子量小于1000D的低聚肽成分占91.42%~92.56%;
SOD中药发酵粉中SOD含量达45000IU/g,多酚含量达2.0~5.0%,原花青素含量达15~25%;
异叶青兰、柠檬马鞭草、蜜蜂花、海葱制成的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚中的多酚含量均达75%~82%;
黑果腺肋花楸果萃取物中的原花青素达10.5%~12.3%、关山樱花萃取物中的樱花苷和樱花素分别达2.5%~2.9%、1.2%~1.6%、线叶金雀花萃取物中的总黄酮含量达25%~35%;
桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物,其功效成分分别为45%~50%的粗多糖,5.0%~6.0%的野黄芩苷,52%~56%的粗多糖和1.8%~2.4%的金丝桃苷。
在上述各实施例中,具有美白、抗氧化、祛黄褐斑功能的中药组合物的制备方法为:
(1)将桑白皮、半枝莲、菝葜、菟丝子通过超临界CO2萃取技术+真空冷冻干燥+低温超微粉碎技术处理制成桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物;
(2)将黑果腺肋花楸果、关山樱花、线叶金雀花经过水蒸气蒸馏、提取、浓缩、真空冷冻干燥、粉碎、沸腾制粒工艺处理制得黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物;
(3)将大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥等生产工艺制成大米肽粉;
(4)将中药材莳萝、曼陀茄根、酢浆草、荜澄茄通过粉碎、榨汁、二次复合发酵、膜过滤、真空冷冻干燥、低温超微粉碎技术处理制得含SOD中药发酵粉;
(5)将异叶青兰、柠檬马鞭草、蜜蜂花、海葱经过超临界CO2萃取、膜分离、离子交换及真空冷冻干燥技术处理制得纯度高的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚;
具体地,
所述步骤(1)中异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚可由以下制备方法制得:
(a)粉碎:将异叶青兰或柠檬马鞭草或蜜蜂花或海葱中药饮片粉碎成颗粒,颗粒过6~8目筛;
(b)超临界CO2萃取:将上述颗粒用CO2超临界萃取法提取,夹带剂为68%~75%(V/V)食用酒精,夹带剂添加量为投料量的5%~6%,萃取温度28~32℃,萃取压力29~31Mpa,CO2流量15~18kg/h,萃取时间70~90min,分离压力6-8MPa,分离温度28-32℃,得到萃取液;
(c)膜分离:启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为60~75μm,滤膜孔径为6~8μm,操作压力为0.15~0.25MPa,确保滤液澄清透明,去除滤渣;
(d)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离杂质;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-32~-35℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干物;冻干物粉碎,过80~100目筛即得异叶青兰多酚或柠檬马鞭草多酚或蜜蜂花多酚或海葱多酚。
所述步骤(2)中黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物可由以下制备方法制得:
(a)水蒸气蒸馏:将黑果腺肋花楸果或关山樱花或线叶金雀花经过破碎处理,过4~6目筛网;破碎组织经过水蒸气蒸馏2~4h,收集挥发油及药渣;
(b)提取浓缩:将药渣加药渣重8~10倍水,提取1~2次,1~2h/次,200目滤过。真空浓缩至相对密度为1.05~1.08的提取浓缩液;
(c)真空冷冻干燥:将上述提取浓缩液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-28~-32℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干萃取物;
(d)粉碎:将上述冻干萃取物粉碎,过80~100目筛;
(e)沸腾制粒:将上述挥发油和冻干粉末使用小型沸腾制粒的低温喷雾、沸腾制粒,喷雾温度为30~45℃,充分制粒,沸腾干燥制成挥发油包埋粉,即得黑果腺肋花楸果萃取物或关山樱花萃取物或线叶金雀花萃取物。
所述步骤(3)中大米肽粉可由以下制备方法制得:
(a)制粒:大米蛋白采用沸腾制粒,溶剂为水,溶剂添加量为10~15%,制粒温度为140~155℃,制成16~20目之间的大米蛋白颗粒;
(b)真空爆破:将所述大米肽粉颗粒在1.8~2.5Mpa饱和蒸气压下维持120~140s后,在15~30ms内释放压力,并迅速抽真空至真空度-60~-80kpa之间,保持80~120s,释放真空即得大米蛋白真空爆破组织;
(c)调浆:将大米蛋白真空爆破组织粉碎过80~100目筛,用纯净水配制成大米蛋白溶液,将大米蛋白溶液混合均匀,使之均匀分散在水溶液中。将混合均匀大米蛋白粉加入适当比例的纯净水,得到其分散液;调节大米蛋白质粉的分散液的pH至6~8;
(d)酶解:加热至50℃,保持其温度在50±5℃,在大米蛋白分散液中加入地衣芽孢杆菌蛋白酶和羧肽酶,均匀搅拌酶解1~3h,其中地衣芽孢杆菌蛋白酶加入量为底物含量的0.2%~0.4%,羧肽酶加入量为底物含量的0.05%~0.1%,酶解结束后用高温瞬时灭酶法进行灭酶;
(e)离心过滤:灭酶后的酶解液体经过离心机13000~16000rpm离心分离,去除杂质,收集清液,启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为25~50μm,滤膜孔径为2~4μm,操作压力为0.05~0.12MPa,确保滤液澄清透明,去除滤渣;
(f)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离氨基酸;
(g)真空浓缩:将离子交换后的溶液进行真空减压浓缩,浓缩温度60~80℃,真空度-0.085~-0.095Mpa,得低聚肽浓缩液,相对密度控制在1.05~1.09之间;
(h)喷雾干燥:将上述低聚肽浓缩液,进行喷雾干燥,工艺参数为喷口温度175~190℃,出口温度90~95℃,上料速度2.5~3.0L/min,得所需符合质量要求的大米肽粉。
所述步骤(4)中含SOD中药发酵粉可由以下制备方法制得:
(a)粉碎:将中药材莳萝、曼陀茄根、酢浆草、荜澄茄以1~3:2~4:1~2:0.5~1.0的比例混合破碎,过30~60目筛;
(b)榨汁:将上述组织破碎物加入1.5~3倍的纯化水充分浸泡1~3h,温度保持在45~55℃,浸泡浓浆经过大型滚筒式榨汁机榨汁处理,离心去杂,即得澄清离心液,离心转速为16000~18000r/min;
(c)二次复合发酵:将离心液中加入乳酸克鲁维酵母,酵母添加量为离心液重量的0.15~0.25%,发酵温度为28~35℃,发酵时间为4~6h,发酵结束后继续加入植物乳杆菌,其菌株号为CGMCC NO.1258和鼠李糖乳杆菌其菌株号为R0011,两者乳杆菌的添加比例为1~2:0.5~1.2,两种乳杆菌的总添加量为离心液重量的0.3~0.35%,发酵温度为36~45℃,发酵时间为3~4h,发酵结束后过400~500目筛网;
(d)膜过滤:启动膜过滤设备,上述发酵清液经过微孔滤膜过滤,滤膜厚度为35~45μm,滤膜孔径为2~8μm,操作压力为0.05~0.12MPa,确保滤液澄清透明,去除滤渣;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-38~-45℃,预冻速率1.2~1.4℃/min,预冻终点温度-80℃,干燥室压力25~32pa,加热板温度35~40℃,干燥时间为14~16h,得冻干物;
(f)低温超微粉碎:将上述冻干物经过低温超微粉碎机处理,过400~500目筛,粉碎温度为18~25℃,即得含SOD中药发酵粉。
所述步骤(5)中异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚可由以下制备方法制得:
(a)粉碎:将异叶青兰或柠檬马鞭草或蜜蜂花或海葱中药饮片粉碎成颗粒,颗粒过6~8目筛;
(b)超临界CO2萃取:将上述颗粒用CO2超临界萃取法提取,夹带剂为68%~75%(V/V)食用酒精,夹带剂添加量为投料量的5%~6%,萃取温度28~32℃,萃取压力29~31Mpa,CO2流量15~18kg/h,萃取时间70~90min,分离压力6-8MPa,分离温度28-32℃,得到萃取液;
(c)膜分离:启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为60~75μm,滤膜孔径为6~8μm,操作压力为0.15~0.25MPa,确保滤液澄清透明,去除滤渣;
(d)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离杂质;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-32~-35℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干物;冻干物粉碎,过80~100目筛即得异叶青兰多酚或柠檬马鞭草多酚或蜜蜂花多酚或海葱多酚。
按照实施例1~3中的配方将复合中药由上述步骤制成的桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物、黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物、大米肽粉、含SOD中药发酵粉以及异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚分别称量后,投入三维混合机中混合15~20min至完全均匀,混合频率15~20Hz,过60~100目筛网,即得本发明中药组合物。
实验例1:祛黄褐斑功能人体试食试验
1、试用样品:使用实施例方案1中的中药组合物作为试食样品,人口服推荐剂量为每日2次,每次4g,温水送服,总计每日服药量为8g。
2、受试者纳入标准:
符合体检符合下列条件的自愿受试者。
2.1面部淡褐色至深褐色界限清楚的斑片,通常对称性分布,无炎性表现及鳞屑。
2.2无其它明显自觉症状。
2.3主要发生在青春期后,女性多发。
2.4病情有一定的季节性,夏重冬轻。
2.5无明显内分泌疾病,并排除其它疾病引起的色素沉着。
3、受试者排除标准:
3.1年龄在18岁以下或65岁以上者,妊娠或哺乳期妇女,过敏体质及对本中药提取物成分过敏者。
3.2合并有心血管、脑血管、肝、肾和造血系统等严重疾病及内分泌疾病,精神病患者。
3.3嗜酒者或吸烟者。
3.4短期内服用与受试功能有关的物品,影响到对结果的判断者。
3.5未按规定服用受试样品,无法判定功效或资料不全影响疗效或安全性判断者。
4、试验设计及分组要求:
采用自身和组间两种对照设计。按受试者的黄褐斑颜色、面积情况随机分为试食组、阳性对照组和阴性对照组,尽可能考虑影响结果的主要因素如户外活动情况、性别、年龄等,进行均衡性检验,以保证组间的可比性。每组受试者不少于40例。
5、受试样品的剂量和食用方法
试食组按推荐服用方法、服用量服用受试产品,对照组可服用安慰剂或采用空白对照。受试样品给予时间30天,必要时可以延长至45天。受试者在试验期间停止使用其他口服及外用有关养颜祛斑的用品。试验期间不改变原来的饮食习惯,正常饮食。
6、观察指标
6.1安全性指标
6.1.1一般状况:包括精神、睡眠、饮食、大小便、血压等;
6.1.2血、尿、便常规检查;
6.1.3肝、肾功能检查,胸透、心电图、腹部B超检查(在试验开始前检查以便排查排除情况)。
6.2功效性指标
6.2.1颜面部黄褐斑面积大小检测:用标尺测量受试前后整个颜面部黄褐斑面积(mm2)
6.2.2颜面部黄褐斑面积深浅检测:按照中国科学院地理研究所设计研制,测绘出版社1992年出版的《实用标准色卡》(第一版)中棕色(Y+M+BK即黄+品红+黑的叠色)色卡为黄褐斑深浅的判断标准:Ⅰ度(15、20、5),Ⅱ度(30、40、10),Ⅲ度(40、60、15)
7、数据处理和结果判定
对试食前后黄褐斑颜色积分和面积变化进行统计,同时计算有效率。
8、试验情况
8.1、一般情况
初始试验人群试食组40例,阴性对照组40例,阳性对照组40例,试食前后,受试者精神、睡眠、饮食、大小便、血压等状态正常,血、尿、便常规检查;肝、肾功能检查,胸透、心电图、腹部B超检查正常,无异常现象发生。例数、性别、人数、年龄、病程见表4所示。
表4.试食前自愿者一般资料对比
8.2、试验方法
试食组服用中药组合物实施例方案1的样品人口服推荐剂量为每日2次,每次4g,温水送服,总计每日服药量为8g。阳性对照组服用氨甲环酸胶囊,每日6粒,每日服药量为1500mg;阴性对照组服用相同剂量的安慰剂,观察时间为30天。30天后,通过诊断观察黄褐斑的个数、面积大小、色度变化情况,统计诊断数据进行综合分析。
疗效评价标准:治愈(黄褐斑几乎完全消失,面积减少大于90%,且不产生新黄褐斑),显效(黄褐斑颜色下降Ⅱ度,面积减少大于40%,且不产生新黄褐斑),有效(黄褐斑颜色下降Ⅰ度,面积减少大于10%,且不产生新黄褐斑),无效(黄褐斑颜色及面积无明显变化甚至加重),总有效率=治愈率+显效率+有效率。
8.3、试验结果
试食后三个组别自愿者服用效果数据对比如表5所示:
表5.试食后自愿者服用效果对比
由表5结果可知,试食组治愈0例,显效25例,有效13例,总有效率为95.0%,试食组总有效率显著高于阴性对照组(P<0.01),也高于阳性对照组。三组患者均无明显不良反应。
9、总结
结果判定:试食组按照规定的方法及服用量服用受试药物以后,临床症状大有好转,试食组黄褐斑面积显著减少,颜色积分明显下降,自身前后比较及与对照组比较,差别均有显著性,且不产生新的黄褐斑,总体有效率高于阴性对照组及阳性对照组,可判定本发明药物组合物具有祛黄褐斑的作用,提示在祛黄褐斑方面确有其独特功效。
实验例2:美白功能人体试食试验
1、试用样品:实验组20人使用实施例方案1中的中药组合物作为试食样品,人口服推荐剂量为每日2次,每次4g,温水送服,总计每日服药量为8g。空白组10人不吃试食样品改用相同剂量的安慰剂替换,和试食组一样坚持定时VISIA-CR检测。
2、体验者要求
2.1、体验者在产品体验期间尽可能避免辛辣、烟酒、油腻等刺激性食物的摄入;
2.2、体验者需保持良好作息,尽量在十一点之前就寝,减少熬夜,保持良好心情。体验者在产品体验期间尽量避免阳光暴晒;
2.3、产品体验期间,须暂停其他类美白产品的使用;
2.4、体验期间体验者若有服用激素类药物,须向组织者报备;
2.5、体验期间可适量增加运动,促进血液循环,但尽量减少户外运动或做好防护,注重日常防晒,减少外出,避免阳光直接照射。
2.6、试用者无明显内分泌疾病,并排除其他疾病引起色素沉着。
3、测试要求
3.1、体验者在九个测量日需保持面部,不化妆,呈素颜状态。测试前请用清水清洗,安静等待10分钟后,开始测量(不可用餐巾纸擦拭面部)。
3.2、每次测试时,测试者须保持自然放松状态、同一姿势、同一面部表情。
3.3、九个测量日须穿深色系上衣,避免衣领浅色或白色反光,导致分析误差。
3.4、分析区域尽量避免笑纹、反光区域、黑痣、头发、胎记、白斑、抓痕、疤痕。
4、项目方案
本试用方案暂定总时长为8周,需按照本中药组合物的食用方法以及用量,进行试用,每周需配合进行一次脸部皮肤状态测试。
食用方法:早晚各一次,每次4g,饭后一小时内食用;
测试时间:项目启动后,先测量0周数据,后每周一的上午,分别对优享官进行相关测试,利用VISIA-CR采集面部图像并分析相关指标。
5、验证仪器设备与测试要求
设备:VISIA-CR(面部图像分析仪),采集面部图像并分析指标,以VISIA-CR前中后期结果(斑点、棕色斑)对比分析及面部特定部位白皙度变化(LAB值)为结果判定。采用自身服用前后对照设计,采用VISIA-CR(面部图像分析仪)对试用者的前额和左右侧面部局部皮肤的颜色,提取L*,a*,b*值,取样时避免高亮反光、黑痣、泛红区域。并通过L*,b*值计算ITA(individual topology angle)值,计算方法为:
6、结果分析
如下图1、图2结果所示,食用本中药组合物1个月后试验组有54%的人变白了,2月后有72%的人变白了,空白组无明显变白。图2显示试食组1个月后平均变白程度为3.9%,2个月后平均变白程度为4.5%。而空白组1~2个月后变白程度分别为1.1%和0.4%。空白组白皙度的增长可能是由于日照时长及强度的减少。且试食组体验人群除了面部肤色变白以外,斑点、紫质及皱纹等指标均有所改善,证实本发明中药组合物有良好的美白效果。
实验例3:抗氧化功能体外抗氧化试验研究
1实验方法
1.1样品制备
选用中药组合物实施例方案1的样品,充分溶解于1mL纯化水中制成待测样品。
1.2清除DPPH自由基能力的测定
取2mL 0.2mmol·L-1的DPPH溶液,加入1mL待测样品及1mL H2O混匀,常温反应30min,在517nm处测定样品吸光值Ai。空白组用H2O代替样品,其余操作同样品组,测得空白组吸光值A0。维生素C(VC)作为阳性对照。其清除率计算公式为:
1.3清除超氧阴离子自由基能力的测定
取pH8.2的Tris-HCl缓冲溶液4.5mL,加入4.2mL H2O,混匀后于25℃恒温水浴中保温20min,同时将样品和25mmol邻苯三酚溶液一起放入25℃水浴中预热。保温20min后加入1mL待测样品及0.4mL邻苯三酚溶液,迅速混匀后于25℃下反应5min,之后加入1mL8 mmol·L-1HCl终止反应,于320nm处测得样品吸光值Ai。空白组用H2O代替样品,其余操作同样品组,测得空白组吸光值A0。VC作阳性对照,其清除率计算公式为:
1.4清除羟基自由基能力的测定
取10mmol·L-1水杨酸钠-乙醇溶液、10mmol·L-1-1FeSO4溶液、待测样品各1mL,混匀后37℃恒温水浴反应30min,之后加入1mL8.8 mmol·L-1H2O2溶液终止反应,于510nm处测得样品吸光值Ai。空白组用H2O代替样品,其余操作同样品组,测得空白组吸光值A0,Ai0为不加H2O2的本底吸光值。VC作为阳性对照,其清除率计算公式为:
1.5总还原力的测定
取2.5mL0.2 mol·L-1的磷酸盐缓冲溶液,2.5mL1%的K3Fe(CN)6溶液和2mL待测样品,混匀后50℃水浴保温30min,之后加入2.5mLH2O和0.5mL0.1%的FeCl3溶液,混匀,于700nm处测得样品吸光值Ai。空白组用H2O代替样品,其余操作同样品组,测得空白组吸光值A0。VC作为阳性对照,其总还原力计算公式为:
总还原力=Ai-A0
2体外抗氧化试验研究结果
2.1该款中药组合物清除DPPH自由基能力的研究
该款中药组合物和VC对DPPH自由基的清除效果见图3,由图可见两者的清除率与其浓度均表现出明显的正比关系。实验范围内测得该款中药组合物的IC50值为520.83μg·mL-1,VC的IC50值为92.59μg·mL-1。结果表明该款中药组合物对DPPH自由基的清除效果较好,但低于同浓度下的VC。
2.2该款中药组合物清除超氧阴离子自由基能力的研究
该款中药组合物和VC对超氧阴离子自由基的清除效果见图4,由图可见两者的清除效果均与浓度呈正比关系。VC的IC50值为909.09μg·mL-1,而该款中药组合物的IC50值为2884.62μg·mL-1,其表明该款中药组合物对超氧阴离子自由基有一定的清除能力但其清除效果要低于同浓度下的VC。
2.3该款中药组合物清除羟基自由基能力的研究
该款中药组合物和VC对羟基自由基的清除效果如图5所示,由图可见两者的清除率均与其浓度存在明显的量效关系。VC的IC50值为208.33μg·mL-1,而该款中药组合物的IC50值为2040.82μg·mL-1,其表明该款中药组合物对羟基自由基具有一定的清除能力但效果低于同浓度下的VC。
2.4该款中药组合物总还原力的研究
该款中药组合物和VC对Fe3+的总还原力结果如图6所示,由图可知,在一定浓度范围内,VC和该款中药组合物的总还原力随浓度增大而增强,具有明显的量效关系,但同浓度下该款中药组合物的总还原力要低于VC。
本实验通过体外活性评价模型实验结果的分析,总结出此款中药组合物的体外抗氧化活性低于同浓度的VC,其对DPPH自由基和羟基自由基具有较好的清除能力,对超氧阴离子自由基的清除能力较弱,对Fe3+有一定的还原能力。证实此款中药组合物具有较好的体外抗氧化活性,其活性具有明显的量效关系,说明它具有较好的抗氧化作用。
实验例4:对照组试验研究
表6
如表6所示,上述六个方案中,其中方案A为实施例方案一配方,作为对照组基准配方设置,样品浓度均为2000μg/mL。通过分析以上六款配方清除DPPH自由基的能力、清除超氧阴离子自由基能力、清除羟基自由基能力以及该款中药组合物总还原力的研究数据,如下表7的数据所示,从而反映出该基础配方在缺失2~3味不同原料后总体抗氧化效果之间存在的差异。
表7
如表7所示,本实验数据可知,通过体外活性评价模型实验结果的分析,总结出此款中药组合物在缺失2~3味不同原料后总体的体外抗氧化活性四项指标均要显著低于同浓度的实施例方案一配方(方案A),证实此款中药组合物中多种植物多酚成分(如异叶青兰多酚、柠檬马鞭草多酚、海葱多酚)、植物萃取物(关山樱花萃取物、线叶金雀花萃取物)、中药提取物(如桑白皮提取物、桑白皮提取物、菟丝子提取物)各成分之间存在较为理想的协同增效的关系,各组分以本申请权利要求书1中描述的特定的配方配伍后,能促进药物之间扬长避短、相互促进、相互提升、协同增效,大幅提升其抗氧化的功能效果,从而提升了药效。
以上所述仅为本发明的优选实施例,并非因此限制本发明的专利范围,凡是利用本发明权利要求书内容所作的等效结构或流程变换,或在不脱离本发明原理的前提下做出的若干改进和润饰,或直接或间接运用在其他相关的技术领域,均同理包括在本发明的专利保护范围内。
Claims (8)
2.根据权利要求1所述的具有美白、抗氧化、祛黄褐斑功能的中药组合物,其特征在于:所述大米肽粉,由大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥处理而获得,上述步骤获得的大米肽粉中分子量小于1000D的低聚肽成分占91.42%~92.56%
SOD中药发酵粉中SOD含量达45000IU/g,多酚含量达2.0~5.0%,原花青素含量达15~25%;
异叶青兰、柠檬马鞭草、蜜蜂花、海葱制成的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚中的多酚含量均达75%~82%;
黑果腺肋花楸果萃取物中的原花青素达10.5%~12.3%、关山樱花萃取物中的樱花苷和樱花素分别达2.5%~2.9%、1.2%~1.6%、线叶金雀花萃取物中的总黄酮含量达25%~35%;
桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物,其功效成分分别为45%~50%的粗多糖,5.0%~6.0%的野黄芩苷,52%~56%的粗多糖和1.8%~2.4%的金丝桃苷。
3.一种制备如权利要求1或2所述的具有美白、抗氧化、祛黄褐斑功能的中药组合物的方法,其特征在于:包括以下步骤:
(1)将桑白皮、半枝莲、菝葜、菟丝子通过超临界CO2萃取技术+真空冷冻干燥+低温超微粉碎技术处理制成桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物;
(2)将黑果腺肋花楸果、关山樱花、线叶金雀花经过水蒸气蒸馏、提取、浓缩、真空冷冻干燥、粉碎、沸腾制粒工艺处理制得黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物;
(3)将大米蛋白经制粒、真空爆破、调浆、酶解、离心过滤、离子交换、真空浓缩、喷雾干燥等生产工艺制成大米肽粉;
(4)将中药材莳萝、曼陀茄根、酢浆草、荜澄茄通过粉碎、榨汁、二次复合发酵、膜过滤、真空冷冻干燥、低温超微粉碎技术处理制得含SOD中药发酵粉;
(5)将异叶青兰、柠檬马鞭草、蜜蜂花、海葱经过超临界CO2萃取、膜分离、离子交换及真空冷冻干燥技术处理制得纯度高的异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚;
将上述步骤制成的桑白皮提取物、半枝莲提取物、菝葜提取物、菟丝子提取物、黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物、大米肽粉、含SOD中药发酵粉以及异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚分别称量后,投入三维混合机中混合15~20min至完全均匀,混合频率15~20Hz,过60~100目筛网,即得本发明中药组合物。
4.根据权利要求3所述的制备方法,其特征在于:所述步骤(3)中大米肽粉可由以下制备方法制得:
(a)制粒:大米蛋白采用沸腾制粒,溶剂为水,溶剂添加量为10~15%,制粒温度为140~155℃,制成16~20目之间的大米蛋白颗粒;
(b)真空爆破:将所述大米肽粉颗粒在1.8~2.5Mpa饱和蒸气压下维持120~140s后,在15~30ms内释放压力,并迅速抽真空至真空度-60~-80kpa之间,保持80~120s,释放真空即得大米蛋白真空爆破组织;
(c)调浆:将大米蛋白真空爆破组织粉碎过80~100目筛,用纯净水配制成大米蛋白溶液,将大米蛋白溶液混合均匀,使之均匀分散在水溶液中。将混合均匀大米蛋白粉加入适当比例的纯净水,得到其分散液;调节大米蛋白质粉的分散液的pH至6~8;
(d)酶解:加热至50℃,保持其温度在50±5℃,在大米蛋白分散液中加入地衣芽孢杆菌蛋白酶和羧肽酶,均匀搅拌酶解1~3h,其中地衣芽孢杆菌蛋白酶加入量为底物含量的0.2%~0.4%,羧肽酶加入量为底物含量的0.05%~0.1%,酶解结束后用高温瞬时灭酶法进行灭酶;
(e)离心过滤:灭酶后的酶解液体经过离心机13000~16000rpm离心分离,去除杂质,收集清液,启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为25~50μm,滤膜孔径为2~4μm,操作压力为0.05~0.12MPa,确保滤液澄清透明,去除滤渣;
(f)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离氨基酸;
(g)真空浓缩:将离子交换后的溶液进行真空减压浓缩,浓缩温度60~80℃,真空度-0.085~-0.095Mpa,得低聚肽浓缩液,相对密度控制在1.05~1.09之间;
(h)喷雾干燥:将上述低聚肽浓缩液,进行喷雾干燥,工艺参数为喷口温度175~190℃,出口温度90~95℃,上料速度2.5~3.0L/min,得所需符合质量要求的大米肽粉。
5.根据权利要求3所述的制备方法,其特征在于:所述步骤(2)中黑果腺肋花楸果萃取物、关山樱花萃取物、线叶金雀花萃取物可由以下制备方法制得:
(a)水蒸气蒸馏:将黑果腺肋花楸果或关山樱花或线叶金雀花经过破碎处理,过4~6目筛网;破碎组织经过水蒸气蒸馏2~4h,收集挥发油及药渣;
(b)提取浓缩:将药渣加药渣重8~10倍水,提取1~2次,1~2h/次,200目滤过。真空浓缩至相对密度为1.05~1.08的提取浓缩液;
(c)真空冷冻干燥:将上述提取浓缩液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-28~-32℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干萃取物;
(d)粉碎:将上述冻干萃取物粉碎,过80~100目筛;
(e)沸腾制粒:将上述挥发油和冻干粉末使用小型沸腾制粒的低温喷雾、沸腾制粒,喷雾温度为30~45℃,充分制粒,沸腾干燥制成挥发油包埋粉,即得黑果腺肋花楸果萃取物或关山樱花萃取物或线叶金雀花萃取物。
6.根据权利要求3所述的制备方法,其特征在于:所述步骤(5)中异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚可由以下制备方法制得:
(a)粉碎:将异叶青兰或柠檬马鞭草或蜜蜂花或海葱中药饮片粉碎成颗粒,颗粒过6~8目筛;
(b)超临界CO2萃取:将上述颗粒用CO2超临界萃取法提取,夹带剂为68%~75%(V/V)食用酒精,夹带剂添加量为投料量的5%~6%,萃取温度28~32℃,萃取压力29~31Mpa,CO2流量15~18kg/h,萃取时间70~90min,分离压力6-8MPa,分离温度28-32℃,得到萃取液;
(c)膜分离:启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为60~75μm,滤膜孔径为6~8μm,操作压力为0.15~0.25MPa,确保滤液澄清透明,去除滤渣;
(d)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离杂质;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-32~-35℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干物;冻干物粉碎,过80~100目筛即得异叶青兰多酚或柠檬马鞭草多酚或蜜蜂花多酚或海葱多酚。
7.根据权利要求3所述的制备方法,其特征在于:所述步骤(4)中含SOD中药发酵粉可由以下制备方法制得:
(a)粉碎:将中药材莳萝、曼陀茄根、酢浆草、荜澄茄以1~3:2~4:1~2:0.5~1.0的比例混合破碎,过30~60目筛;
(b)榨汁:将上述组织破碎物加入1.5~3倍的纯化水充分浸泡1~3h,温度保持在45~55℃,浸泡浓浆经过大型滚筒式榨汁机榨汁处理,离心去杂,即得澄清离心液,离心转速为16000~18000r/min;
(c)二次复合发酵:将离心液中加入乳酸克鲁维酵母,酵母添加量为离心液重量的0.15~0.25%,发酵温度为28~35℃,发酵时间为4~6h,发酵结束后继续加入植物乳杆菌,其菌株号为CGMCC NO.1258和鼠李糖乳杆菌其菌株号为R0011,两者乳杆菌的添加比例为1~2:0.5~1.2,两种乳杆菌的总添加量为离心液重量的0.3~0.35%,发酵温度为36~45℃,发酵时间为3~4h,发酵结束后过400~500目筛网;
(d)膜过滤:启动膜过滤设备,上述发酵清液经过微孔滤膜过滤,滤膜厚度为35~45μm,滤膜孔径为2~8μm,操作压力为0.05~0.12MPa,确保滤液澄清透明,去除滤渣;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-38~-45℃,预冻速率1.2~1.4℃/min,预冻终点温度-80℃,干燥室压力25~32pa,加热板温度35~40℃,干燥时间为14~16h,得冻干物;
(f)低温超微粉碎:将上述冻干物经过低温超微粉碎机处理,过400~500目筛,粉碎温度为18~25℃,即得含SOD中药发酵粉。
8.根据权利要求3所述的制备方法,其特征在于:所述步骤(1)中异叶青兰多酚、柠檬马鞭草多酚、蜜蜂花多酚、海葱多酚可由以下制备方法制得:
(a)粉碎:将异叶青兰或柠檬马鞭草或蜜蜂花或海葱中药饮片粉碎成颗粒,颗粒过6~8目筛;
(b)超临界CO2萃取:将上述颗粒用CO2超临界萃取法提取,夹带剂为68%~75%(V/V)食用酒精,夹带剂添加量为投料量的5%~6%,萃取温度28~32℃,萃取压力29~31Mpa,CO2流量15~18kg/h,萃取时间70~90min,分离压力6-8MPa,分离温度28-32℃,得到萃取液;
(c)膜分离:启动膜过滤设备,清液经过微孔滤膜过滤,滤膜厚度为60~75μm,滤膜孔径为6~8μm,操作压力为0.15~0.25MPa,确保滤液澄清透明,去除滤渣;
(d)离子交换:将过滤后的滤液经过离子交换柱及活性炭柱,进行脱盐脱色处理,纳滤去除游离杂质;
(e)真空冷冻干燥:将上述得到的滤液投入真空冷冻干燥设备中进行干燥,参数设定为:预冻初始温度-32~-35℃,预冻速率1.6~1.8℃/min,预冻终点温度-65℃,干燥室压力35~42pa,加热板温度45~50℃,干燥时间为18~22h,得冻干物;冻干物粉碎,过80~100目筛即得异叶青兰多酚或柠檬马鞭草多酚或蜜蜂花多酚或海葱多酚。
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CN117018755B (zh) * | 2023-10-10 | 2023-12-22 | 北京逯博士行为医学科技研究院有限公司 | 一种具有抗癌组分的混合物制备方法 |
CN117694549A (zh) * | 2024-02-05 | 2024-03-15 | 北京衡美金叶营养健康科技有限公司 | 一种保湿祛斑美白的组合物、制备方法及应用 |
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