CN112807322A - 低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用 - Google Patents
低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用 Download PDFInfo
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Abstract
本发明提供了一种低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用。本发明中低聚甘露糖醛酸盐是以褐藻胶为原料,经化学降解法或物理降解法或两种降解法结合制得不同分子量的低聚甘露糖醛酸;低聚甘露糖醛酸经中和制备得到不同分子量的低聚甘露糖醛酸盐,其分子骨架为D‑甘露糖醛酸(M)通过β‑1,4糖苷键连接形成的线性寡糖化合物。本发明产品来源于海藻多糖,具有资源丰富、易于产业化,安全有效等诸多优点,在用于制备延缓皮肤衰老、调节免疫的药物和功能性食品中方面具有广阔的开发应用前景。
Description
技术领域
本发明属于医药技术领域,具体涉及低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用。
背景技术
患有皮肤光损伤的病人数量急剧增加,皮肤光损伤已经成为一个重要的公共健康问题。皮肤作为一种生理屏障,具有复杂的酶和非酶抗氧化防御网络,可将挥发性活性氧(ROS)/自由基清除为毒性较低的物质,从而维持细胞氧化还原稳态。长波紫外线(UVA)穿透力强于中波紫外线(UVB),可穿透到真皮深处,UVA也是导致皮肤过早衰老的主要原因。皮肤过度暴露于UVA会诱发细胞内ROS的生成,直接或间接损伤细胞蛋白结构、脂质膜和核酸,导致细胞炎症、光老化和皮肤癌等多种皮肤病。近年来,海洋藻类因含有丰富的具有抗氧化活性的多糖而引起广泛关注。
不同来源的海藻酸盐及其衍生物有多种生物和药用作用,如神经保护作用、抗炎作用、免疫刺激作用、抗癌作用和抗氧化作用等。海藻酸盐也因其稳定性、生物降解性和低毒性而广泛应用于食品、制药、化妆品和纺织业。低聚甘露糖醛酸盐是从褐藻中分离出的一种海藻酸盐多糖,目前,国内外尚未见有将其应用于皮肤抗衰老和抵抗炎症过敏方面的的报道。
发明内容
本发明的目的在于提供了低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,本发明通过实验结果证明低聚甘露糖醛酸盐可对UVA引起的HaCaT细胞损伤及HSF细胞炎症有明显的调节作用,可使细胞产生有效的抗紫外损伤效果,从而起到延缓皮肤衰老和调节免疫的作用。本发明的技术方案为临床提供了适合光敏性皮肤病、皮肤炎症病人以及亚健康等人群服用的药品或功能性食品。
为实现上述发明目的,本发明采用以下技术方案予以实现:
本发明提供了低聚甘露糖醛酸盐在制备延缓皮肤衰老、免疫抗炎的药物和功能性食品中的应用。
进一步的:所述低聚甘露糖醛酸盐是以褐藻胶为原料,经酶降解法、化学降解法或物理降解法得不同分子量低聚甘露糖醛酸;低聚甘露糖醛酸经中和制备得到不同分子量的低聚甘露糖醛酸盐,其分子骨架为D-甘露糖醛酸通过β-1,4糖苷键连接形成的线性寡糖化合物,重均分子量为0.3kDa~15kDa;其中,低聚甘露糖醛酸及其盐的分子结构通式如下:
式中,R=H或Li,Na, K,Ca,Mg,n≤60。
进一步的:所述低聚甘露糖醛酸盐和褐藻胶及褐藻胶寡糖制成复配制剂,或将低聚甘露糖醛酸硫酸化或磷酸化、乙酰化制得衍生制剂。
进一步的:所述低聚甘露糖醛酸盐与甘油、丁二醇、香精、丙二醇、乳酸、黄原胶、透明质酸钠、烟酰胺、珍珠粉、维生素C、维生素E、中草药提取物、去离子水制备得到抗衰老护肤品或化妆品。
进一步的:所述低聚甘露糖醛酸盐提高UVA辐射HaCaT细胞存活率,提高UVA辐射HaCaT细胞线粒体膜电位,降低UVA辐射HaCaT内活性氧含量,提高UVA辐射HaCaT细胞内线粒体复合物I和II的活性。
进一步的:所述低聚甘露糖醛酸盐提高HaCaT细胞内SirT1蛋白的表达量,降低UVA辐射后成纤维细胞内NO的含量,降低致炎白酶COX-2酶活力,通过靶向激活线粒体和抑制COX-2从而达到延缓皮肤衰老、免疫抗炎的作用。
进一步的:所述低聚甘露糖醛酸盐采用下述制备方法:将褐藻胶配成5~15wt%水溶液,采用0.5~1.5wt%稀盐酸加热到80~90℃后搅拌降解4~5 小时,冷却后用5~15wt%的碳酸钠水溶液中和,再用2~5wt%稀盐酸调节pH 到3~4,离心收集沉淀,用1.5~2.5mol/LNaOH溶解,加入95wt%乙醇后收集沉淀,经无水乙醇脱水后干燥,得到聚甘露糖醛酸钠盐;将聚甘露糖醛酸用纯水配置成不同浓度的水溶液,采用Fenton法降解,得不同分子量低聚甘露糖醛酸;低聚甘露糖醛酸经氢氧化锂、氢氧化钠、氢氧化钾、氢氧化钙或氢氧化镁中和后,经超滤分级、冷冻干燥,得到不同分子量的低聚甘露糖醛酸的锂盐、钠盐、钾盐、钙盐或镁盐。
进一步的:所述低聚甘露糖醛酸盐的浓度为50μmol/L以上。
进一步的:所述褐藻胶为褐藻酸或褐藻酸钠。
进一步的::所述化学降解法为酸降解法或Fenton降解法,所述物理降解法为微波或超声降解法;所述酶降解法使用褐藻胶酶。
本发明的优点及有益效果是:
1、本发明采用HaCaT细胞UVA损伤模型和HSF细胞UVA炎症模型,证明低聚甘露糖醛酸盐对UVA引起的HaCaT细胞损伤及HSF细胞炎症的保护作用。实验结果证明低聚甘露糖醛酸盐可显著提高UVA辐射后HaCaT细胞存活率,显著提高UVA辐射后HaCaT细胞线粒体膜电位,显著降低UVA辐射后HaCaT细胞内活性氧含量,显著提高UVA辐射后HaCaT细胞内线粒体复合物I和II的活性,并显著提高HaCaT细胞内SirT1蛋白的表达量;可显著降低UVA辐射后HSF细胞内NO的含量,显著降低UVA辐射后HSF细胞内COX-2酶活力。因此,D-聚甘露糖醛酸钠对UVA引起的HaCaT细胞损伤及HSF细胞炎症有明显的调节作用,可细胞产生有效的抗紫外损伤效果,从而起到延缓皮肤衰老和调节免疫的作用。
2、本发明产品来源于海洋天然多糖,具有资源丰富、易于产业化,安全有效等诸多优点,可开发成抗皮肤衰老、调节免疫和抗紫外损伤的药品、护肤品及保健品,因此具有较好的市场应用前途。
具体实施方式
下面结合具体实施例对本发明的技术方案做进一步详细说明。
实施例1:低聚甘露糖醛酸盐的提取制备及分析
本发明中的低聚甘露糖醛酸盐可以采用本领域内常规技术手段制得,本实施例中的具体步骤为:
将褐藻胶(本实施例为褐藻酸钠)配成10wt%水溶液,采用1wt%稀盐酸加热到80~90℃后搅拌降解4~5小时,冷却后用10wt%的碳酸钠水溶液中和,再用5wt%稀盐酸调节pH到3.65左右,离心收集沉淀,用2mol/L NaOH溶解,加入3倍体积95wt%乙醇后收集沉淀,经无水乙醇脱水后干燥,得到聚甘露糖醛酸钠盐。将聚甘露糖醛酸钠盐用纯水配置成不同浓度的水溶液,采用Fenton法降解,所得不同分子量低聚甘露糖醛酸。低聚甘露糖醛酸经氢氧化锂、氢氧化钠、氢氧化钾、氢氧化钙或氢氧化镁中和后,双层滤纸过滤,超滤分级浓缩,冷冻干燥,得到不同分子量的低聚甘露糖醛酸的锂盐、钠盐、钾盐、钙盐或镁盐。
所制得的低聚甘露糖醛酸或其盐的结构式如下:
式中,R=H或Li,Na, K,Ca,Mg,n≤60,其中单糖组成中4位和5位为含有双键或者不含双键。
本实施例中,用Tosoh TSK Gel3000 PWXL柱色谱(内径7.8 mm*柱长300 mm)和右旋糖酐标准品(中国药品生物制品检定所)作为标准,测得低聚甘露糖醛酸盐的重均分子量为0.3kDa~15kDa。
实施例2:低聚甘露糖醛酸盐对UVA诱导HaCaT细胞存活率的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。具体步骤为:
(1)细胞培养:将HaCaT细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔8000个细胞种植于96孔板中,放入恒温细胞培养箱孵育12h后,加入不同浓度的低聚甘露糖醛酸钠放入恒温细胞培养箱继续孵育48h。孵育结束后更换培养基,给予30J/cm2 UVA照射。放入恒温细胞培养箱继续孵育12h后,每孔加入20μL (5mg/ml)MTT溶液,培养箱继续孵育4h。结束后,除去液体,每孔加入150μLDMSO溶液,混匀放入37℃培养箱20孵育20min。最后,酶标仪A540nm处测量吸光值。每次三个平行,实验重复三次。
计算公式:细胞存活率(Cell viability)=实验组吸光值/对照组吸光值。
实验结果如表1所示,模型组和对照组存在显著性差异,UVA损伤模型构建成功。当给药浓度为100μmol/L、200μmol/L、400μmol/L、500μmol/L时,细胞存活率与模型组存在显著性差异,低聚甘露糖醛酸盐可对UVA损伤HaCaT细胞产生较好保护作用。
表1低聚甘露糖醛酸盐对UVA诱导HaCaT细胞存活率影响实验结果
组别(μmol/L) | 细胞存活率 |
对照组 | 1.0±0.003*** |
模型组 | 0.4±0.0002 |
10 | 0.7±0.004* |
100 | 0.8±0.01* |
200 | 0.7±0.09* |
300 | 0.5±0.03 |
400 | 0.8±0.002* |
500 | 0.8±0.004* |
600 | 0.51±0.02 |
注: n=9,x±SD,与模型组相比,*P<0.05, ***P<0.001
实施例3:低聚甘露糖醛酸盐对UVA诱导HaCaT细胞线粒体膜电位的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。具体实验步骤为:
(1)细胞培养:将HaCaT细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔8000个细胞种植于96孔板中,放入恒温细胞培养箱孵育12h后,加入不同浓度的低聚甘露糖醛酸钠放入恒温细胞培养箱继续孵育48h。孵育结束后更换培养基,给予30J/cm2 UVA照射。放入恒温细胞培养箱继续孵育12h后,吸除培养基,用PBS溶液清洗一次,加入150μL染色工作液,充分混匀,细胞培养箱孵育40 min。孵育结束后,吸除液体,用100μL JC-1(1X)染色缓冲液清洗两次,每孔加入150μL培养基,用酶标仪检测。检测波长为(488/585和488/535),结果为红绿色荧光比值。
实验结果由表2所示,模型组与对照组存在显著性差异,当给药浓度为50μmol/L,100μmol/L,400μmol/L,500μmol/L,600μmol/L时,线粒体膜电位与模型组相比显著升高。
表2低聚甘露糖醛酸盐对UVA诱导HaCaT细胞线粒体膜电位影响实验结果
组别(μmol/L) | 线粒体膜电位 |
对照组 | 0.7±0.02*** |
模型组 | 0.4±0.009 |
10 | 0.6±0.004** |
100 | 0.6±0.004* |
200 | 0.5±0.003 |
300 | 0.5±0.00 |
400 | 0.6±0.004* |
500 | 0.6±0.01** |
600 | 0.7±0.007* |
注:n=9,x±SD,与模型组相比,*P<0.05, **P<0.01, ***P<0.001
实施例4:低聚甘露糖醛酸盐对UVA诱导HaCaT细胞内线粒体复合物I活性的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。具体实验步骤为:
(1)细胞培养:将HaCaT细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔50万个细胞种植于6孔板中,放入恒温细胞培养箱孵育12h,然后加入200μL (10μmol/L) 低聚甘露糖醛酸钠,放入恒温细胞培养箱继续孵育48h。孵育结束后更换培养基,给予30 J/cm2 UVA照射。放入恒温细胞培养箱继续孵育12h后收集细胞,加入1ml线粒体复合物I提取液,用冰浴匀浆器匀浆,取匀浆600 g,4 ℃离心5 min。弃沉淀,将上清液移至另一离心管中,11000 rmp,4 ℃离心10 min。弃上清,沉淀中加入500μL提取液,超声波破碎(冰浴,功率20%或200 W,超声3 s,间隔10s,重复30次),用于复合体Ⅰ酶活性测定。测定体系:样本(10μL)+试剂一(154μL)+工作液(20μL)+试剂四(16μL),将上述试剂依次加入96孔板,迅速吹打均匀,酶标仪测定其吸光值。记录第10 s的吸光值为A1,迅速将96孔板连同反应液放入恒温细胞培养箱中准确反应2 min,记录2 min时的吸光度为A2,计算ΔA=A1-A2。
复合物I活力单位的计算:
单位定义:每mg组织蛋白每分钟消耗1 nmol的NADH定义为一个酶活力单位。
复合物I活力(U/mg prot)=1608*ΔA/Cpr (Cpr:样本蛋白浓度,mg/ml)。
实验结果如表3所示,模型组与对照组存在显著性差异,低聚甘露糖醛酸钠(10μmol/L)使UVA损伤HaCaT细胞内线粒体复合物I的活性相较于模型组显著上升,且正常HaCaT细胞内线粒体复合物I的活性相较于模型组有显著上升。
表3低聚甘露糖醛酸盐对UVA诱导HaCaT细胞内线粒体复合物I活性影响实验结果
组别 | 线粒体复合物I活性 |
对照组 | 100±8.0**** |
模型组 | 31.9±7.9 |
模型组加药(10μmol/L) | 105.5±25.3**** |
正常细胞加药(10μmol/L) | 75.0±21.4**** |
注:n=9,x±SD,与模型组相比,***P<0.001,****P<0.0001
实施例5:低聚甘露糖醛酸盐对UVA诱导HaCaT细胞内线粒体复合物II活性的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。
(1)细胞培养:将HaCaT细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔50万个细胞种植于6孔板中,放入恒温细胞培养箱孵育12h,然后加入200μL (200μg/ml) 低聚甘露糖醛酸钠,放入恒温细胞培养箱继续孵育48h。孵育结束后更换培养基,给予30 J/cm2 UVA照射。放入恒温细胞培养箱继续孵育12h后收集细胞,加入1ml线粒体复合物II提取液,用冰浴匀浆器匀浆,取匀浆600 g,4 ℃离心5 min。弃沉淀,将上清液移至另一离心管中,11000 rmp,4 ℃离心10 min。弃上清,沉淀中加入500μL提取液,超声波破碎(冰浴,功率20%或200 W,超声3 s,间隔10s,重复30次),用于复合体II酶活性测定。测定体系:样本(10μL)+试剂一(154μL)+工作液(20μL)+试剂四(16μL),将上述试剂依次加入96孔板,迅速吹打均匀,酶标仪测定其吸光值。记录第10 s的吸光值为A1,迅速将96孔板连同反应液放入恒温细胞培养箱中准确反应2 min,记录2 min时的吸光度为A2,计算ΔA=A1-A2。
复合物II活力单位的计算:
单位定义:每mg组织蛋白每分钟消耗1 nmol的FADH2定义为一个酶活力单位。
复合物II活力(U/mg prot)=476.2*ΔA/Cpr (Cpr:样本蛋白浓度,mg/ml)。
实验结果如表4所示,模型组与对照组存在显著性差异,低聚甘露糖醛酸盐(10μmol/L)使UVA损伤HaCaT细胞内线粒体复合物II的活性相较于模型组显著上升,且正常HaCaT细胞内线粒体复合物II的活性相较于模型组有显著上升。
表4低聚甘露糖醛酸盐对UVA诱导HaCaT细胞内线粒体复合物II活性影响实验结果
组别 | 线粒体复合物II活性 |
对照组 | 100±5.1**** |
模型组 | 37.3±1.17 |
模型组加药(10μmol/L) | 128.6±24.6**** |
正常细胞加药(10μmol/L) | 83.4±41.3*** |
注:n=9,x±SD,与模型组相比,***P<0.001,****P<0.0001
实施例6:低聚甘露糖醛酸盐对UVA诱导HaCaT细胞内SirT1蛋白表达量的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。
(1)细胞培养:将HaCaT细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔50万个细胞种植于6孔板,放入恒温细胞培养箱孵育12h,然后加入200μL (10μmol/L) 低聚甘露糖醛酸钠,放入恒温细胞培养箱继续孵育48h,孵育结束后更换培养基,给予30 J/cm2 UVA照射,放入恒温细胞培养箱继续孵育12h。结束后,吸除培养基,加入预冷的PBS溶液冲洗三次。将6孔板置于冰上,每孔加200μL细胞裂解液,充分裂解10 min后,收集裂解液放入1.5 ml EP管中。12000 rmp,4 ℃离心10 min后,取上清液,用BCA试剂盒定量。在收集的蛋白样品中加入适量浓缩的SDS-PAGE蛋白上样缓冲液,100 ℃或沸水浴加热3-5min,以充分变性蛋白。冷却至室温后,将蛋白样品直接上样至SDS-PAGE胶加样孔内即可。待电泳结束后,转膜,将PVDF膜放入4 ℃孵育一抗。12 h后取出PVDF膜放置于1×TBST溶液中于摇床上洗涤六次,每次10 min。室温孵育1 h,取出PVDF膜放置于1×TBST溶液中于摇床上洗涤六次,每次10 min。将200 μl显色液均匀滴加到PVDF膜上,避光显色5 min,凝胶显色仪显色,获取图像。数据处理:SirT1活性=模型组(实验组)灰度积分值/对照组灰度积分值。
实验结果如表5所示,模型组与对照组相比较,模型组SirT1蛋白表达量明显降低,此现象揭示经过30 J/cm2 UVA照射后,HaCaT细胞内SirT1蛋白表达量发生显著变化,模型组细胞加入低聚甘露糖醛酸盐可以使HaCaT细胞内SirT1蛋白表达量显著上升,相反对照组细胞加入低聚甘露糖醛酸盐后细胞内SirT1蛋白表达量无显著性变化。
表5低聚甘露糖醛酸盐对UVA诱导HaCaT细胞内SirT1蛋白表达量影响实验结果
组别 | SirT1蛋白含量 |
对照组 | 1.0±0.05* |
模型组 | 0.5±0.03 |
模型组加药(10μmol/L) | 1.2±0.1*** |
正常细胞加药(10μmol/L) | 0.6±0.006 |
注:n=9,x±SD,与模型组相比,**P<0.01,***P<0.001,****P<0.0001
综合以上指标,本发明所采用的低聚甘露糖醛酸盐对UVA损伤HaCaT细胞有保护作用,可对细胞产生有效的抗紫外损伤效果,从而起到延缓皮肤衰老的作用。
实施例7: 低聚甘露糖醛酸盐对UVA诱导HSF细胞内NO含量的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。
(1)细胞培养:将HSF细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔8000个细胞种植于96孔板中,放入恒温细胞培养箱孵育24h,然后加入200μL(50μmol/L)低聚甘露糖醛酸钠,放入恒温细胞培养箱继续孵育48h。孵育结束后,更换培养基,给予15 J/cm2能量UVA照射后,放入恒温细胞培养箱孵育12h。结束后,每孔加入200μL PBS溶液,冲洗两次。每孔加入200μL DAF-FM DA溶液,培养箱继续孵育20min后,除去液体,每孔加入无酚红的MEM培养基溶液150μL,冲洗两次。最后,酶标仪检测荧光值,检测波长(495/515)。实验重复三次。
计算公式:NO(Fluorescence per mg protein)=荧光值/细胞存活率。
实验结果如表6和7所示,模型组与对照组存在显著性差异,UVA损伤模型构建成功。低聚甘露糖醛酸盐(50μmol/L)使UVA损伤HSF细胞内NO含量相较于模型组显著降低,且使正常HSF细胞内NO含量相较于模型组有显著降低。
表6不同浓度低聚甘露糖醛酸盐对UVA诱导HSF细胞内NO含量影响实验结果
组别 | NO含量 |
对照组 | 32662.8±253.2**** |
模型组 | 43571.5±676.9 |
2μmol/L | 37190.2±225.6**** |
10μmol/L | 37018.4±525.6**** |
50μmol/L | 28028.8±238.5**** |
注:n=9,x±SD,与模型组相比,****P<0.0001
表7低聚甘露糖醛酸盐对UVA诱导HSF细胞内NO含量影响实验结果
组别 | NO含量 |
对照组 | 47.5±6.2**** |
模型组 | 100±17.5 |
模型组加药(50μmol/L) | 23.7±7.2**** |
正常细胞加药(50μmol/L) | 32.1±6.6**** |
注:n=9,x±SD,与模型组相比,****P<0.0001
实施例8: 低聚甘露糖醛酸盐对UVA诱导HSF细胞内COX-2酶活力的影响
本实施例采用实施例1所制备的低聚甘露糖醛酸钠盐。
(1)细胞培养:将HSF细胞接种于MEM完全培养液中(含有100U/mL青霉素、100U/mL链霉素和10%FBS),置于37℃、5%CO2培养箱中培养。
(2)将每孔8000个细胞种植于96孔板中,放入恒温细胞培养箱孵育24h,然后加入200 μg/ml 低聚甘露糖醛酸钠,放入恒温细胞培养箱继续孵育48h。孵育结束后,更换培养基,给予15 J/cm2能量UVA照射后,放入恒温细胞培养箱孵育12h。孵育结束后,每孔加入200μL PBS溶液,清洗两次。每孔加入350μL细胞裂解液,置于冰上裂解20min后,收集裂解液。13000 rpm,4℃离心5min,收集上清。最后,取20μL提取蛋白,加入底物和辅助因子在37 ℃孵育10min后,用酶标仪检测荧光值,检测波长(565/590)。实验重复三次。
计算公式:COX-2酶活力=实验组荧光值/100%酶活组荧光值。
实验结果如表8所示,对照组和模型组COX-2酶活存在显著性差异,低聚甘露糖醛酸盐(50μmol/L)可使HSF细胞内的COX-2酶活显著降低。正常组细胞给予相同量的低聚甘露糖醛酸盐,COX-2酶活与对照组相比显著降低。
表8低聚甘露糖醛酸盐对UVA诱导HSF细胞内COX-2酶活力影响实验结果
组别 | COX-2酶活 |
对照组 | 0.8±0.002**** |
模型组 | 1.1±0.001 |
模型组加药(50μmol/L) | 0.8±0.001**** |
正常细胞加药(50μmol/L) | 0.9±0.0003*** |
注:n=9,x±SD,与模型组相比,***P<0.001,****P<0.0001
综合以上指标,本发明所采用的低聚甘露糖醛酸盐对UVA引起的HaCaT细胞损伤和HSF细胞炎症有明显的调节作用,具有免疫抗炎的作用。所述的低聚甘露糖醛酸盐能够用于制备延缓皮肤衰老和调节免疫的药物和功能食品;或者所述低聚甘露糖醛酸盐用于饮食补充剂;或与甘油、丁二醇、香精、丙二醇、乳酸、黄原胶、透明质酸钠、烟酰胺、珍珠粉、维生素C、维生素E、中草药提取物、去离子水相关抗衰老护肤品或化妆品;或者所述低聚甘露糖醛酸盐和褐藻胶及褐藻胶寡糖之一的复配制剂或制备以低聚甘露糖醛酸为母核的硫酸化或磷酸化、乙酰化等衍生应用制剂。
以上实施例仅用以说明本发明的技术方案,而非对其进行限制;尽管参照前述实例对本发明进行了详细的说明,对本领域的普通技术人员来说,依然可以对前述实施例所记载的技术方案进行修改,或者对其中部分技术特征进行等同替换;而这些修改或替换,并不使相应技术方案的本质脱离本发明所要求保护的技术方案的精神和范围。
Claims (9)
1.低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用。
3.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,其特征在于:所述低聚甘露糖醛酸盐和褐藻胶及褐藻胶寡糖制成复配制剂。
4.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老、免疫抗炎的药物和功能性食品中的应用,其特征在于:所述低聚甘露糖醛酸盐与甘油、丁二醇、香精、丙二醇、乳酸、黄原胶、透明质酸钠、烟酰胺、珍珠粉、维生素C、维生素E、中草药提取物、去离子水制备得到抗衰老护肤品或化妆品。
5.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,其特征在于:所述低聚甘露糖醛酸盐提高UVA辐射HaCaT细胞存活率,提高UVA辐射HaCaT细胞线粒体膜电位,降低UVA辐射HaCaT内活性氧含量,提高UVA辐射HaCaT细胞内线粒体复合物I和II的活性。
6.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,其特征在于:所述低聚甘露糖醛酸盐提高HaCaT细胞内SirT1蛋白的表达量,降低UVA辐射后成纤维细胞内NO的含量,降低致炎白酶COX-2酶活力,通过靶向激活线粒体和抑制COX-2从而达到延缓皮肤衰老和免疫抗炎的作用。
7.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,其特征在于:所述低聚甘露糖醛酸盐采用下述制备方法:将褐藻胶配成5~15wt%水溶液,采用0.5~1.5wt%稀盐酸加热到80~90℃后搅拌降解4~5 小时,冷却后用5~15wt%的碳酸钠水溶液中和,再用2~5wt%稀盐酸调节pH 到3~4,离心收集沉淀,用1.5~2.5mol/L NaOH溶解,加入95wt%乙醇后收集沉淀,经无水乙醇脱水后干燥,得到聚甘露糖醛酸钠盐;将聚甘露糖醛酸用纯水配置成不同浓度的水溶液,采用Fenton法降解,得不同分子量低聚甘露糖醛酸;低聚甘露糖醛酸经氢氧化锂、氢氧化钠、氢氧化钾、氢氧化钙或氢氧化镁中和后,经超滤分级、冷冻干燥,得到不同分子量的低聚甘露糖醛酸的锂盐、钠盐、钾盐、钙盐或镁盐。
8.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,其特征在于:所述褐藻胶为褐藻酸或褐藻酸钠。
9.根据权利要求1或2所述的低聚甘露糖醛酸盐在制备延缓皮肤衰老和免疫抗炎的药物和功能性食品中的应用,其特征在于:所述化学降解法为酸降解法或Fenton降解法,所述物理降解法为微波或超声降解法;所述酶降解法使用褐藻胶酶。
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CN115708834A (zh) * | 2022-11-25 | 2023-02-24 | 中国科学院海洋研究所 | 一种褐藻胶寡糖衍生物的应用 |
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