CN114886808B - 龙胆发酵液的制备工艺及抗糖化用途 - Google Patents
龙胆发酵液的制备工艺及抗糖化用途 Download PDFInfo
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- CN114886808B CN114886808B CN202210520125.1A CN202210520125A CN114886808B CN 114886808 B CN114886808 B CN 114886808B CN 202210520125 A CN202210520125 A CN 202210520125A CN 114886808 B CN114886808 B CN 114886808B
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Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
- A61K8/9783—Angiosperms [Magnoliophyta]
- A61K8/9789—Magnoliopsida [dicotyledons]
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/005—Preparations for sensitive skin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/80—Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
- A61K2800/85—Products or compounds obtained by fermentation, e.g. yoghurt, beer, wine
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
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Abstract
本发明涉及化妆品技术领域,首次公开了龙胆发酵液的制备工艺及抗糖化用途。本发明以龙胆水提液作为发酵底物,以酿酒酵母和类干酪乳酸杆菌对龙胆水提液进行深层次的发酵加工,采用上述工艺制备的龙胆发酵液中活性成分总多酚、总蛋白、龙胆苦苷的含量明显高于龙胆水提液。本发明还提供了采用上述工艺制备的龙胆发酵液在抗糖化方面的功效,并证实了本发明的龙胆发酵液在较低浓度下的非酶糖基化抑制率远远高于龙胆水提液。
Description
技术领域
本发明涉及化妆品技术领域,特别涉及龙胆发酵液的制备工艺及抗糖化用途。
背景技术
近年来,糖基化终末产物(AGEs)主要作为糖尿病等疾病的标志物和羰基应激衰老理论中的关键物质进行研究。AGEs不仅会使皮肤弹性下降,色素堆积,产生皱纹这种外观性的变化,还会使皮肤屏障被破坏,皮肤相关细胞凋亡,发生炎症。此外,它具有不可逆性,不易被代谢,逐年累积下,皮肤从里至外发生深刻的变化。主流的抗糖成分主要是肌肽、玻色因、硫辛酸、葡萄籽提取物、黄酮类、多酚类、阿魏酸、革糖素等。其中,肌肽是经国际研究证实真正有效的抗糖成分,且安全无毒,肌肽能在糖侵蚀肌肤前,通过与糖反应的型态,代替体内蛋白质与糖反应,从而帮助保护蛋白质不被糖基化,达到抗糖化的作用。革糖素是经国际研究证实有效的抗糖成分,革糖素通过预防和清除两条通路达到抗糖功效,保护肌肤中的蛋白不易被糖化,帮助加速分解肌肤中的晚期糖化终末产物,有实验表明其与烟酰胺共同作用时,革糖素具有放大烟酰胺抑制黑色素转移能力的功效。化妆品技术领域的研发人员正寻找更多具有抗糖功效的成份。
龙胆(Gentiana scabra Bunge)是龙胆科、龙胆属多年生草本植物。龙胆的根及根茎入药,其味苦、性寒;归肝、胆经;具有清热燥湿、泻肝胆实火等作用。现有技术中通常采用常规的水提或醇提,获得龙胆提取物并加以利用。
发明内容
本发明的目的在于提供一种龙胆发酵液的制备工艺及抗糖化用途。
本发明的第一方面提供了一种龙胆发酵液的制备工艺,包括如下步骤:
(1)制备龙胆水提液:将龙胆粉碎过筛,加水浸泡后,进行超声辅助提取,过滤,取滤液,得到龙胆水提液;(2)菌种扩培:将酿酒酵母接种到YPD培养基,将类干酪乳酸菌接种到LB培养基,在摇床中培养;(3)龙胆发酵:将所述龙胆水提液接入培养后的菌种进行发酵,发酵温度为32℃,发酵时间为24小时,得到龙胆发酵液。
优选地,所述龙胆粉碎过筛后,按照料液比1:40(g/ml)的比例加水浸泡。
优选地,所述超声辅助提取的功率为550~570W,提取温度为70~80℃,提取45~60分钟。
优选地,所述扩培步骤中,培养至培养基的OD值为0.5~1.0,此时菌种处在对数期,即菌种已达到合适的接种浓度。
本发明的第二方面提供了以本发明第一方面的工艺制备的龙胆发酵液在化妆品或护肤品中的应用,其中,所述龙胆发酵液为化妆品和/或护肤品中的抗糖化功效成分。
本发明所提供的应用中,所述化妆品和/或护肤品施用于皮肤外表面。
优选地,本发明所提供的应用中,所述化妆品和/或护肤品选自眼霜、面霜、精华、乳液、面膜精华液中的一种或几种。
优选地,本发明所提供的应用中,所述龙胆发酵液作为添加剂分散于基质原料中,所述基质原料选自油质原料、粉质原料、胶质原料中的一种或几种。
优选地,本发明所提供的应用中,所述化妆品和/或护肤品还包括:表面活性剂、香精与香料、色素、保湿剂、防腐剂、抗氧剂、紫外线吸收剂、螯合剂、收敛剂、助渗剂、营养性添加剂中的一种或几种。
相对于现有技术而言,本发明首次提出以龙胆水提液作为发酵底物,以酿酒酵母和类干酪乳酸杆菌对龙胆水提液进行深层次的发酵加工,采用上述工艺制备的龙胆发酵液中活性成分总多酚、总蛋白、龙胆苦苷(单体化合物)的含量明显高于龙胆水提液。更重要的是,本发明还提供了采用上述工艺制备的龙胆发酵液在抗糖化方面的功效,并证实了本发明的龙胆发酵液在较低浓度下的非酶糖基化抑制率远远高于龙胆水提液。
具体实施方式
为使本发明的目的、技术方案和优点更加清楚,下面将对本发明的各实施方式进行详细的阐述。然而,本领域的普通技术人员可以理解,在本发明各实施方式中,为了使读者更好地理解本申请而提出了许多技术细节。但是,即使没有这些技术细节和基于以下各实施方式的种种变化和修改,也可以实现本申请所要求保护的技术方案。
本发明的部分实施方式提供了一种龙胆发酵液的制备工艺,包括如下步骤:(1)制备龙胆水提液:将龙胆粉碎过筛,加水浸泡后,进行超声辅助提取,过滤,取滤液,得到龙胆水提液;(2)菌种扩培:将酿酒酵母接种到YPD培养基,将类干酪乳酸菌接种到LB培养基,在摇床中培养;(3)龙胆发酵:将所述龙胆水提液接入培养后的菌种进行发酵,发酵温度为32℃,发酵时间为24小时,得到龙胆发酵液。
在本发明的部分实施方式中,所述龙胆粉碎过筛后,按照料液比1:40(g/ml)的比例加水浸泡。
在本发明的部分实施方式中,所述超声辅助提取的功率为550~570W,提取温度为70~80℃,提取45~60分钟。
在本发明的部分实施方式中,所述扩培步骤中,培养至培养基的OD值为0.5~1.0,此时菌种处在对数期,即菌种已达到合适的接种浓度。
本发明的第二方面提供了以本发明第一方面的工艺制备的龙胆发酵液在化妆品或护肤品中的应用,其中,所述龙胆发酵液为化妆品和/或护肤品中的抗糖化功效成分。
本发明的实施方式所提供的龙胆发酵液可应用于化妆品和/或护肤品中,所述化妆品和/或护肤品施用于皮肤外表面。
在本发明部分实施方式中,应用所述龙胆发酵液的化妆品和/或护肤品选自眼霜、面霜、精华、乳液、面膜精华液中的一种或几种。
本发明部分实施方式中,所述龙胆发酵液作为添加剂分散于基质原料中,所述基质原料选自油质原料、粉质原料、胶质原料中的一种或几种。
其中,所述油质原料可以是下述基质原料中的任意一种或几种组合:大豆油、橄榄油、杏仁油、蓖麻油、花生油、棉籽油、茶籽油、霍霍巴油、鳄梨油、椰子油、棕榈油、可可脂、米糠油、月见草油、小麦胚芽油、玉米胚芽油、棕榈蜡、木蜡、水貂油、海龟油、蛇油、蛋黄油、牛脂、马脂、猪脂、鹿脂、卵磷脂、羊毛脂、蜂蜡、鲸蜡、虫胶蜡、石蜡、凡士林、微晶蜡、地蜡、角鲨烷、羊毛脂蜡、羊毛脂醇、羊毛脂醇酯化物、乙酰化羊毛脂、乙酰化羊毛脂醇、羊毛脂酸、羊毛脂酸酯化物、聚氧乙烯羊毛脂、聚氧乙烯羊毛醇醚、聚氧丙烯羊毛醇醚、氢化羊毛脂、烷氧基化氢化羊毛脂、硅油(如:二甲基硅油、八甲基硅油、甲基苯基聚硅氧烷)、聚硅氧烷-聚氧烷基嵌段共聚物、甲基含氢硅油、Cn脂肪酸及其酯、Cn脂肪醇及其酯;其中,n≥10,并优选为10-24,更优选为12-20,如14、16、17、18。
所述Cn脂肪酸如:月桂酸、肉豆蔻酸、棕榈酸、硬脂酸、油酸、亚油酸、芥酸、癸酸。
所述Cn脂肪醇如:月桂醇、鲸蜡醇、硬脂醇、异硬脂醇、油醇、椰油醇。
所述Cn脂肪酸酯、Cn脂肪醇酯如:肉豆蔻酸异丙酯、肉豆蔻酸肉豆蔻酯、十四醇乳酸酯、十六醇乳酸酯、棕榈酸异丙酯、棕榈树异辛酯、硬脂酸丁酯、硬脂酸异辛酯、硬脂酸单甘油脂、硬脂酸聚乙二醇酯、油酸癸酯、辛酸甘油酯、癸酸甘油酯、甘油三油酸酯。
所述粉质原料可以是下述基质原料中的任意一种或几种组合:云母、碳酸镁、钛白粉、锌白粉、氢氧化钙、碳酸钙、高岭土、硬脂酸钙、硬脂酸锌、硬脂酸镁、滑石粉、二氧化硅、氢氧化铝、焦磷酸钙、碳酸氢钙、膨润土。
其中,所述油脂原料可以是植物油质原料、动物油质原料、矿物油质原料、合成油质原料、半合成油质原料中的任意一种或几种。如:淀粉、环糊精、黄原胶、角叉胶、瓜尔豆胶、***胶、黄芪胶、琼脂、虫胶、海藻酸钠、明胶、甲基纤维素、乙基纤维素、羧甲基纤维素钠、羟乙基纤维素、羟丙基纤维素、聚纤维素醚季铵盐、聚乙烯醇、聚乙烯吡咯烷酮、聚丙烯酸钠、聚氧乙烯、乙烯基吡咯烷酮/乙酸乙烯酯共聚物、卡波树脂、硅酸铝镁、C12-15醇苯甲酸酯。
其中,所述溶剂原料可以是下述基质原料中的任意一种或几种组合:水、乙醇、丙醇、异丙醇、异丁醇、乙二醇、1,2-丙二醇、环己烷、二氯二氟甲烷、四氟二氯乙烷、乙二醇单***、二甘醇单***、丙酮、甲基异丁基甲酮、醋酸乙酯、醋酸丁酯、乙酸戊酯、邻苯二甲酸二丁酯。
在本发明部分实施方式中,应用所述龙胆发酵液的提取物的化妆品和/或护肤品还包括:表面活性剂、香精与香料、色素、保湿剂、防腐剂、抗氧剂、紫外线吸收剂、螯合剂、收敛剂、助渗剂、营养性添加剂中的一种或几种。
其中,所述表面活性剂可以是:1)阴离子表面活性剂,如:C12-14脂肪醇硫酸铵、C12-14脂肪醇聚氧乙烯醚羧酸盐、十二烷基硫酸二乙醇胺盐、C12-14脂肪醇聚氧乙烯醚硫酸铵、十二烷基硫酸三乙醇胺、醇醚邻苯二甲酸单酯钠盐、酰化肽、月桂酸单乙醇酰胺硫酸钠、正辛醇聚氧丙烯醚琥珀酸单酯磺酸盐、十六醇琥珀酸单酯磺酸盐、磺基琥珀酸单酯二钠盐、琥珀酸单十八酯二钠盐、醇醚磺基琥珀酸单酯二钠盐、脂肪醇聚氧乙烯醚(钠)磺基琥珀酸单酯铵盐、油酰胺磺基琥珀酸单酯二钠盐、N-酰基谷氨酸钾盐、吡咯烷酮羧酸钠、聚丙烯酸钠、脂肪醇聚氧乙烯醚磷酸酯、脂肪醇聚氧乙烯醚磷酸单酯及其盐、脂肪醇聚氧乙烯醚磷酸单酯乙醇胺盐、烷基酚聚氧乙烯醚磷酸单酯及其盐、壬基酚醚磷酸单酯乙醇胺盐、烷基磷酸酯盐、邻苯二甲酸单月桂醇酯盐、十一碳烯酸锌、阴离子型氨基酸表面活性剂、N-C12-18酰基谷氨酸钠、N-月桂酰L-丙氨酸钠、N-硬脂酰基谷氨酸单钠盐、N-椰子酰基谷氨酸单钠盐、N-混合脂肪酰基谷氨酸单钠盐、长直链烷基芳醚磺酸钠TH;2)非离子表面活性剂,如:油醇聚氧乙烯醚(如乳化剂VO系列)、烷基酚聚氧乙烯醚(如乳化剂OPE系列)、硬脂酸聚氧乙烯酯、乙二醇单硬脂酸酯、二硬脂酸乙二醇酯、聚乙二醇单硬脂酸酯、聚乙二醇双硬脂酸酯、聚乙二醇双月桂酸酯、双硬脂酸甘油酯、三硬脂酸甘油酯、蔗糖硬脂酸酯、聚氧乙烯甘油醚单硬脂酸酯、N-月桂酰基谷氨酸双烷醇酯、苯甲酸脂肪醇酯、山梨醇单脂肪酸酯(司盘,Span,如司盘-20、司盘60、司盘65、司盘-80、司盘-83、司盘-85)、聚氧乙烯醚山梨醇单脂肪酸酯(吐温,Tween,如吐温-20、吐温-40、吐温-60、吐温-61、吐温-80)、二乙醇胺、三乙醇胺、脂肪酸单乙醇酰胺(如椰子油酸单乙醇酰胺、月桂酸单乙醇酰胺、棕榈酸单乙醇酰胺)、椰子油酸二乙醇酰胺、月桂酸二乙醇酰胺、甲基葡萄糖苷硬脂酸酯、聚氧乙烯甲基葡萄糖苷硬脂酸酯、乙二醇葡萄糖苷硬脂酸酯、丙二醇基葡萄糖苷硬脂酸酯、C8-16烷基葡萄糖苷、甘油葡萄糖苷硬脂酸酯、琥珀酸月桂醇醚单酯、月桂醇甘油酯聚氧乙烯醚、硬脂酸甘露醇酐酯、聚氧乙烯貂油、貂油酸甲酯、貂油酸异丙酯;3)阳离子表面活性剂,如氯化C12-18烷基三烷基铵(如氯化十六烷基二甲基烷基铵、氯化十八烷基三甲基铵、氯化十六烷基三甲基铵)、氯化油酰胺丙基二羟丙基二甲基铵、氯化二羟丙基二甲基十二烷基铵、氯化二甲基二烯丙基铵/丙烯酰胺共聚物、氯化油酰胺丙基-2,3-二羟丙基二甲基铵、乙基硫酸油酰胺丙基二甲基乙基铵、氯化双C12-18基二甲基铵、聚季铵-11调理剂、M-505聚季铵-7调理剂、貂油酰胺丙基胺-壳聚糖、貂油酰胺丙基胺-水解蛋白、阳离子蛋白肽、瓜尔胶-羟丙基三甲基氯化铵盐、聚纤维素醚季铵盐、单油酸三乙醇胺酯、DNP系列;4)两性离子表面活性剂,如C12-18烷基二羟乙基甜菜碱、N-C12-18烷基-N-(2-羟乙基)-N-(2-甲酰胺基乙基)铵乙酸盐、C12-18烷氧基羟丙基甜菜碱、氧化椰油酰胺基丙基胺、椰油羟乙基磺酸钠、氧化十八烷基二甲基胺、椰油酰二乙醇胺氧化胺、N-烷基-β-氨基丙酰二乙醇胺、氧化月桂酰基丙胺、N-C12-18酰基谷氨酸、羟乙基癸酸咪唑啉甜菜碱、羟乙基肉豆蔻酸咪唑啉甜菜碱、羟乙基棕榈酸咪唑啉甜菜碱。
其中,所述香精与香料可以是选自:1)天然香料,如苦艾油、桉叶油、八角茴香油、白柠檬油、白丁香浸膏、丁香油、白兰花油、白兰花浸膏、白兰叶油、柏木油、薄荷油、苍术硬脂、草木犀浸膏、橙叶油、丁香罗勒油、大花茉莉浸膏、冬青油、当归油、当归浸膏、地衣酸、地檀香油、玳玳花油、玳玳叶油、风信子浸膏、枫香浸膏、芳樟油、黄樟油、甘松油、桂花浸膏、红橘油、胡萝卜籽油、海狸香、黄葵油、黄兰油、藿香油、黄蒿油、九里香浸膏、金合欢浸膏、姜油、橘子油、椒样薄荷油、菊花浸膏、枯茗油、龙涎香、留兰香油、灵猫香、腊梅油、铃兰浸膏、木香根油、米兰花油、玫瑰香叶油、玫瑰油、茉莉浸膏、牡蛎油、墨红浸膏、墨红油、柠檬桉叶油、柠檬油、七里香浸膏、楠叶油、芹菜籽油、肉豆蔻油、肉桂油、山苍子油、水貂香膏酊、水仙浸膏、松针油、树苔浸膏、素馨浸膏、麝香、麝鼠香膏、甜橙油、檀香油、晚香玉油、晚香玉浸膏、万寿菊精油、小茴香油、血桧木油、香茅油、香叶油、康乃馨油、紫苏油、香紫苏油、香附油、香柠檬薄荷油、岩兰草油、香薷油、香薷草浸膏、悬钩子蔷薇花油、悬钩子蔷薇花浸膏、薰衣草油、月桂叶油、依兰依兰油、芫荽籽油、鱼香草油、岩蔷薇浸膏、岩桂油、柚皮油、鸢尾油、烟草花油、香橼叶油、栀子花浸膏、紫穗槐油、紫罗兰叶浸膏、愈创木酚、4-甲基愈创木酚、4-乙基愈创木酚、天然香兰素、樟脑;2)合成香精,如柠檬烯、长叶烯、乙位石竹烯、异长叶烯、异长叶烯酮、异长叶烷酮、溴代苏合香烯、二苯甲烷、二苯醚、间甲基二苯醚、丁香酚、异丁香酚、乙位萘甲醚、乙位萘***、对甲基茴香醚、异丁香酚苄醚、玫瑰醚、檀香醚、水仙醚(或茉莉醚)、降龙涎香醚、甲基柏木醚、环氧柏木烷、埃林塔尔、埃林塔尔N、3-己烯醇、癸醇、月桂醇、苯甲醇、苯乙醇、对甲氧基苄醇、肉桂醇、二甲基苄基甲醇、香叶醇、橙花醇、芳樟醇、玫瑰醇、松油醇、香茅醇、消旋薄荷脑、薄荷脑、冰片、柏木醇、柏木烷酮、甲基柏木酮、3,4-二氧亚甲基苯甲醇、α-甲基-3,4-二氢亚甲基苯丙醛、2-叔丁基-4-甲基环己醇、香紫苏醇、檀香醇、3-甲基-5-(2’,2’,3’-三甲基环戊烯-1’-基)戊-2-醇、苯甲醛、苯乙酮、苯乙酸、对甲基苯乙酮二苯甲酮、月桂醛、肉桂醛、肉桂酸、α-戊基-β-苯基丙烯醛、α-苯基丙醛、铃兰醛、新铃兰醛、兔耳草醛、3,4-二氧亚甲基苯甲醛、3,7-二甲基-6-辛烯醛、3,7-二甲基-7-羟基辛醛、柠檬醛、乙位环高柠檬醛、艾薇醛、柑青醛、4-羟基-3-甲氧基苯甲醛、α-二氢突厥酮、β-突厥烯酮、紫罗兰酮、6-甲基紫罗兰酮、橙花酮、香芹酮、胡椒基丙酮、覆盆子酮、药草酮、顺式茉莉酮、庚醛乙二醇缩醛、苯乙醛二甲缩醛、辛醛二甲缩醛、α-戊基桂醛二甲缩醛、柠檬醛二甲缩醛、柠檬醛二乙缩醛、茴香醛二甲缩醛、蘑菇醛、叶青素、风信子素、苹果酯、吉普缩酮、甲酸香叶酯、甲酸香茅酯、阿弗曼酯、乙酸叶醇酯、乙酸苄酯、乙酸苯乙酯、乙酸肉桂酯、乙酸松油酯、乙酸龙脑酯、乙酸芳樟酯、乙酸香叶酯、乙酸对叔丁基环己酯、乙酸邻叔丁基环己酯、乙酸香根酯、乙酸琥珀酯、乙酸阿弗曼酯、乙酸十氢萘酯、乙酸柏木酯、丙酸香叶酯、乙酸三氯甲基苯甲酯、丁酸苄酯、丁酸叶醇酯、丁酸香叶酯、2-甲基戊酸苯乙酯、异丁酸对甲酚酯、2-壬烯酸甲酯、苯甲酸乙酯、苯甲酸甲酯、苯甲酸叶醇酯、苯甲酸苄酯、苯乙酸乙酯、苯乙酸对甲酚酯、肉桂酸乙酯、水杨酸丁酯、水杨酸异丁酯、水杨酸异戊酯、水杨酸叶醇酯、水杨酸己酯、水杨酸苄酯、水杨酸苯乙酯、水杨酸环己酯、茉莉素、茉莉酯、2-戊基环戊酮醋酸甲酯、橙花素、γ-壬内酯、γ-十一内酯、香豆素、吲哚、3-甲基吲哚、3,7-二甲基-2,6-辛二烯腈、对甲氧基苯甲腈、二甲苯麝香、酮麝香、2,6-二硝基-3-甲氧基-4-叔丁基甲苯、1,1-二甲基-4-乙酰基-6-叔丁基茚满、六甲基三环异色满麝香、1,1,2,3,3,6-六甲基-5-乙酰基茚满、麝香-T、麝香-M、麝香-L、麝香-F、11-氧杂环十六内酯、吐纳麝香;3)香精,玫瑰香精、茉莉香精、檀香香精、白兰香精。
其中,所述色素可以是选自如下组分中的一种或几种:二氧化锆、醋酸铅、硝酸银、叶绿素、叶绿素铜、亚铁氰化铁、鸟嘌呤、坚牢红、食品红I、还原红I、胭脂红、食用杨梅红、食用樱桃红、食品红17、苋菜红、坚牢绿、食品黄3、食用柠檬黄、食用苏丹黄、食用靛蓝、食品蓝2、荧光桃红、β-胡萝卜素、指甲花、栀子红色素、栀子绿色素、栀子蓝色素、栀子黄色素、钛云母珠光颜料、氧化铁红、氧化铁黄、氧化铁黑、氧化铬绿、氧氯化铋、碱性桃红、银朱R、喹啉黄、紫草宁、紫胶红色素、群青、酸性红87、酸性绿25、酸性荧光黄、酸性橙7、金光红53、立索尔宝红BK、立索尔宝红2R、永固橙。
其中,所述防腐剂可以是选自如下组分中的一种或几种组合:山梨酸及其盐、丙酸盐、对羟基苯甲酸酯、苯甲醇、苯甲酸及其盐、5-氯-2-甲基-4-异噻唑啉-3-酮、-2-甲基-4-异噻唑啉-3-酮、卡松、咪唑烷基脲、脱氢醋酸及其盐、亚硫酸钠、亚硫酸氢钠、焦亚硫酸钠、2,2-溴-2-硝基-1,3-丙二醇(布罗波尔)、异抗坏血酸及其盐、曲酸及其酯、壬二酸、松萝酸、间苯二酚。
其中,所述抗氧剂可以是选自如下组分中的一种或几种组合:SOD、丁基羟基茴香醚、没食子酸酯、抗坏血酸及其盐、异抗坏血酸及其盐、谷甾醇、芦丁、生育酚。
其中,所述保湿剂可以是选自如下组分中的一种或几种组合:1,2-丁二醇、山梨醇、木糖醇、甘油、乳酸盐、聚乙二醇、DL-吡咯烷酮羧酸盐、D-葡萄糖、曲酸及其酯、尿酸、乳清酸、果胶酸、昆布氨酸、胶原蛋白、海藻糖、12、羟基硬脂酸胆甾醇酯。
其中,所述紫外线吸收剂可以是选自如下组分中的一种或几种组合:吡哆醇盐酸盐、曲酸及其酯、芦丁、芦荟苷、咖啡氨酸、橙皮素、2-乙基己基水杨酸酯、2-乙基己基对二甲基氨基苯酸酯、2-乙基己基对甲氧基肉桂酸酯、水杨酸苯酯、基邻氨基苯甲酸酯、2-羟基-4-甲氧基二苯甲酮、2-羟基-4-甲氧基二苯甲酮-5-磺酸、2-羟基-4-辛氧基二苯甲酮。
其中,所述螯合剂可以是选自如下组分中的一种或几种组合:EDTA二钠盐、双吡啶硫酮锌、硫代硫酸钠。
其中,所述收敛剂可以是选自如下组分中的一种或几种组合:苯酚对磺酸锌、羟基氯化铝、硫酸铝、硫酸铝钾、硫酸锌、氯化铝、氯化锌、卡拉明。
其中,所述助渗剂可以是选自如下组分中的一种或几种组合:二甲基亚砜、1-正十二烷基氮杂环庚-2-酮、2-氨基乙醇。
其中,所述pH值调节剂可以是选自如下组分中的一种或几种组合:酒石酸、乳酸、硼酸、琥珀酸。
其中,所述营养性添加剂可以是选自如下组分中的一种或几种组合:甲壳素、壳聚糖、丝肽、丝素、卵磷脂、海藻糖、乳糖、珍珠粉、胶原蛋白、胶原水解物、透明质酸及其盐、蛋膜素、植物甾醇、氯化溶菌酶、硫酸软骨素钠、蜂乳、蜂胶、蜂蜜、甘草酸、甘草次酸、γ-亚麻酸、曲酸及其酯、赤霉酸、甘氨酸、L-天冬氨酸及其盐、L-胱氨酸及其盐、DL-丙氨酸、L-丝氨酸、DL-丝氨酸、L-蛋氨酸、DL-蛋氨酸、L赖氨酸盐酸盐、L苏氨酸、肌醇、麦芽醇、泛酸盐、D-泛醇、视黄醇醋酸酯、尿囊素、吡哆醇盐酸盐、可的松、烟酰胺、维生素、***、苏木素、尿酸、乳清酸、芦荟苷、壬二酸、松香酸、松萝酸、昆布氨酸、胸腺素、茜草素、柠檬烯、黄芩黄素、脱氧胆酸、愈疮木萸、橙皮素、鞣酸、二硫化硒、茶皂苷、柠檬酸三乙酯、丁基羟基甲苯、植酸、聚氯化二甲基二烯丙基铵。
实验例
1 材料与方法
1.1 试剂与仪器
实验设备:
分析天平(TB-2002,北京赛多利斯仪器***有限公司);恒温水浴锅(上海一恒科技有限公司HWS24型);超净工作台(北京世安科林净化技术有限公司TP-3102);摇床(上海福玛实验设备有限公司KYC-100B);立式压力蒸汽灭菌锅(合肥华泰医疗设备有限公司XY-280D);电热恒温鼓风干燥箱(上海森信实验仪器有限公司DGG-9070B);酶标仪(TECANInfinite M200PRO)实验室仪器:移液枪、量筒、玻璃棒、滤板、抽滤瓶、药匙、烧杯、试管、移液枪。
试剂:
葡萄糖(BR北京奥博星生物技术公司),胰蛋白胨(BR北京奥博星生物技术公司)、氯化钠(AR西陇化工股份有限公司)、福林酚(1mol/mL源叶生物)、酵母浸粉(BR北京奥博星生物技术公司)。
1.2制备龙胆水提液及龙胆水提液提取物
制备龙胆水提液:粉碎机将龙胆粉碎,过100目筛,备用。按料液比1:40(g/ml)的比例,称取原料龙胆粉40g加入1600mL超纯水,浸泡2h后在功率560W、温度80℃下进行超声辅助提取50min,过滤,取滤液,得到龙胆水提液。
制备龙胆水提液提取物:将水提液在-80℃冷冻12h后,放入真空冷冻干燥机中24h,制得龙胆水提液提取物。
1.3菌种扩培
将酿酒酵母接种到YPD培养基(葡萄糖20g/L;胰蛋白胨20g/L;酵母浸粉10g/L),类干酪乳酸菌接种到LB培养基(酵母浸粉10g/L;胰蛋白胨10g/L;氯化钠5g/L)在适宜温度的摇床中培养,至其OD值=0.5~1.0时,菌种处在对数期,即为合适的接种浓度。
1.4龙胆发酵
将龙胆水提液分成100mL/份,接入菌种发酵。考虑各因素之间的交互影响,选择发酵温度、菌株结合、发酵时间作为考察因素,选取3个水平,用L9(33)正交表设计试验,以龙胆苦苷作为结果指标,选择B发酵时间(24小时、36小时、48小时)、A菌株种类(酿酒酵母,类干酪乳杆菌,酿酒酵母+类干酪乳杆菌)、C发酵温度(27℃、32℃、37℃)作为考察因素,见表1、2。
表1正交因素表
表2正交试验分组
1.5制备龙胆发酵液提取物
对正交试验中各试验组制备得到的龙胆发酵液进行如下步骤的提取:
将龙胆发酵液在-80℃冷冻12h后,放入真空冷冻干燥机中24h,制得龙胆发酵液提取物。
1.6龙胆发酵液提取物中龙胆苦苷含量测定
通过高效液相色谱分别对各试验组制备得到的龙胆发酵液提取物中的龙胆苦苷含量进行检测,色谱柱:Agilent C18色谱柱(4.6×250mm,5μm),流动相为水:乙腈(75:25);流速1mL/min;检测波长:270nm;柱温35℃;检测器:二极管阵列检测器。
各正交试验组的龙胆发酵液提取物中龙胆苦苷含量检测结果如表3所示。
表3龙胆发酵液提取物中龙胆苦苷含量检测结果
根据表3数据可知,采用类干酪乳杆菌与酿酒酵母结合发酵,发酵时间为24h,温度在32h,在此条件下发酵,所获得的发酵液中龙胆苦干含量达到最高。1.7龙胆水提液提取物、龙胆发酵液提取物中的总多酚含量的测定
对试验组6制备得到的龙胆发酵液提取物中的总多酚含量进行测定:配置26.7%碳酸钠溶液:称取26.7g碳酸钠固体粉末,加入蒸馏水至100g,搅拌溶解。取1mL龙胆发酵液提取物加到10mL比色管中,加入1mL蒸馏水,0.5mL2倍稀释的福林酚试液,1.5mL26.7%碳酸铵溶液,用水定容至10mL,室温下反应2h,于760nm下测定吸光值。
采用上述相同的方法,对1.2中所制备的龙胆水提液提取物中的多酚含量进行测定,以示对比。
对比情况如下:龙胆发酵液提取物和龙胆水提液提取物中的总多酚含量如表4所示。
表4总多酚含量对比表
组别 | 龙胆发酵液提取物组 | 龙胆水提液提取物组 |
总多酚含量 | 81.22μg/mg | 61.45μg/mg |
由上表可知:龙胆发酵液提取物中的总多酚含量显著高于未发酵前的龙胆水提液提取物中的总多酚含量。
1.8龙胆水提液提取物、龙胆发酵液提取物中的蛋白质含量的测定
对试验组6制备得到的龙胆发酵液提取物中的蛋白质含量进行测定:根据BCA蛋白定量(碧云天;货号:P0012)试剂盒试验方法进行。配置工作液:50体积BCA试剂A加1体积BCA试剂B(50:1),充分混匀,每孔20μL龙胆发酵液提取物,200μL BCA工作液,37℃下静置30min,在562nm波长下测定吸光度。
采用上述相同的方法对1.2中所制备的龙胆水提液提取物中的蛋白质含量进行测定,以示对比。
对比情况如下:龙胆发酵液提取物和龙胆水提液提取物中的蛋白质含量如表5所示。
表5蛋白质含量对比表
组别 | 龙胆发酵液提取物组 | 龙胆水提液提取物组 |
蛋白质含量 | 815.1667μg/mg | 693.6687μg/mg |
由上表可知:龙胆发酵液提取物中的蛋白质含量显著高于龙胆水提液提取物中的蛋白质含量。
1.9龙胆发酵液的抗糖化功效检测
配置PBS:配置pH为7.3-7.4的PBS,并在其中加入0.02%(质量百分含量)叠氮化钠。
BSA(牛血清白蛋白)-果糖反应液:20mL PBS作为溶剂的BSA溶液(20mg/mL)与20mL果糖溶液(0.5mol/L)混匀。
配置待测样品:先用PBS对龙胆发酵液、水提液分别做稀释,稀释至龙胆发酵液(或龙胆水提液)在待测样品中的质量浓度为10%、1%,分别得到具有浓度梯度的龙胆发酵液待测样品和龙胆水提液待测样品。
抑制组:将各龙胆发酵液待测样品、龙胆水提液待测样品分别与BSA-果糖反应液混合,作为龙胆发酵液抑制组、龙胆水提液抑制组。
阴性对照组:选用PBS代替待测样品,与BSA-果糖反应液混合,作为阴性对照组。阴性对照组反映了体系中对非酶糖基化反应无抑制作用的情况。
阳性对照组:选用氨基胍作为阳性对照物,与BSA-果糖反应液混合,作为阳性对照组。
空白组:将各龙胆发酵液待测样品、龙胆水提液待测样品分别与PBS混合,不添加反应液,作为空白组。
以下表6所示不同试验组作为孵育体系,于37℃恒温培养箱内避光孵育,选择第5天检测各试验组的荧光强度,激发波长370nm,发射波长为440nm。
表6反应体系建立
数据处理:
每个样品设定3个平行,检测结果为3次测定的算数平均值,计算RSD(相对标准偏差)。3个平行测定的荧光值RSD应≤3%,有时其中一个数值偏差大,导致RSD>3%,应适当选取另两个平行测定的算数平均值作为检测结果。而后分别计算得到龙胆水提液和龙胆发酵液的抑制率。
抑制率(IR)的计算公式:
IR(%)=[1-(RFU抑制组-RFU空白组):RFU阴性对照组)]×100%。
表7龙胆提取液体外非酶糖基化抑制结果(抑制率单位:%)
如上表所示,龙胆提取液在所测浓度范围内,其抑制率都成正相关性。随着提取液浓度越高,其体外非酶糖基化抑制率越高。在100%浓度下抑制率相差不大,接近100%,在较低浓度10%浓度下,发酵后抑制率为73.03%,水提液即发酵前为58.75%,发酵后的抑制率浓度提升为14.28%。在1%浓度下,发酵后的抑制率浓度提升约18%。在较低浓度下,发酵后的龙胆非酶糖基化抑制率远远高于发酵前。
本领域的普通技术人员可以理解,上述各实施方式是实现本发明的具体实施例,而在实际应用中,可以在形式上和细节上对其作各种改变,而不偏离本发明的精神和范围。
Claims (8)
1.龙胆发酵液在制备抗糖化化妆品中的应用,其特征在于,所述抗糖化为抑制非酶糖基化反应,所述龙胆发酵液的制备工艺包括如下步骤:
(1)制备龙胆水提液:将龙胆粉碎过筛,加水浸泡后,进行超声辅助提取,过滤,取滤液,得到龙胆水提液;
(2)菌种扩培:将酿酒酵母接种到YPD培养基,将类干酪乳酸菌接种到LB培养基,在摇床中培养;
(3)龙胆发酵:向所述龙胆水提液中接入培养后的酿酒酵母和类干酪乳杆菌进行发酵,发酵温度为32℃,发酵时间为24小时,得到龙胆发酵液。
2.根据权利要求1所述的应用,其特征在于,所述龙胆粉碎过筛后,按照料液比1:40g/ml的比例加水浸泡。
3.根据权利要求1所述的应用,其特征在于,所述超声辅助提取的功率为550~570W,提取温度为70~80℃,提取45~60分钟。
4.根据权利要求1所述的应用,其特征在于,所述扩培步骤中,培养至培养基的OD值为0.5~1.0。
5.根据权利要求1所述的应用,其特征在于,所述化妆品施用于皮肤外表面。
6.根据权利要求5所述的应用,其特征在于,所述化妆品选自眼霜、面霜、精华、乳液、面膜中的一种或几种。
7.根据权利要求5所述的应用,其特征在于,所述龙胆发酵液作为添加剂分散于基质原料中,所述基质原料选自油质原料、粉质原料、胶质原料中的一种或几种。
8.根据权利要求7所述的应用,其特征在于,所述化妆品还包括:表面活性剂、香精、色素、保湿剂、防腐剂、抗氧剂、紫外线吸收剂、螯合剂、收敛剂、助渗剂中的一种或几种。
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