CN107823239A - 胭脂萝卜提取物的制备方法以及其在制备防治阿尔茨海默症食品、保健品或药物中的应用 - Google Patents
胭脂萝卜提取物的制备方法以及其在制备防治阿尔茨海默症食品、保健品或药物中的应用 Download PDFInfo
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Abstract
本申请涉及阿尔茨海默症治疗技术领域,公开了胭脂萝卜提取物在制备防治阿尔茨海默症药物、保健品或食品中的应用。通过动物体内试验发现,胭脂萝卜提取物能够改善老年痴呆模型SAMP8小鼠和D‑半乳糖模型小鼠的认知和记忆能力,可用于阿尔茨海默症食品、保健品和药物等领域。
Description
技术领域
本申请涉及阿尔茨海默症治疗技术领域,尤其涉及一种胭脂萝卜提取物的制备方法以及其在制备防治阿尔茨海默症药物、保健品或食品中的应用。
背景技术
阿尔茨海默症(Alzheimer's disease,AD)是一种多发生在老年期的进行性认知功能障碍和行为损害为特征的中枢神经系统退行性疾病,是老年期痴呆最常见的类型。其主要临床特征包括进行性记忆减退、认知功能障碍、行为障碍及日常生活能力下降等,主要病理特征为大脑中海马区β淀粉样蛋白(amyoid β peptide,Aβ)在细胞外沉积形成老年斑(senile plaque,sp);脑神经细胞内tau蛋白过度磷酸化形成神经细胞内神经元纤维缠结(neurofibrillary tangles,NFTs);脑皮质神经细胞减少,以及新皮层及脑血管的改变。
国际阿尔茨海默病协会发布的《全球阿尔茨海默病2011年报告》显示:全球现有AD患者3600万人,并且以每20年翻一番的速度增加,到2050年能达到1.15亿。2010年治疗、护理AD等造成的社会成本达到了6040亿美元,超过了2010年全球GDP的1%。目前抗AD药物研究存在的最主要问题有两点:一是可供选择的药物类型过少,迄今只有5个上市药物,分别为乙酰胆碱酯酶抑制剂:他克林(tacrine)、多奈哌齐(donepezil)、利凡斯的明(rivastigmine)、和加兰他敏(galanthamine),以及N-甲基-D-天冬氨酸受体阻滞剂美金刚(memantine);二是药物疗效不佳。已上市药物仅适用于轻、中度AD的对症治疗,只能缓解AD的症状,却不能阻断AD的发病进程。
发明内容
有鉴于此,本发明的目的在于克服现有技术的不足,提供一种胭脂萝卜提取物的制备方法,以及胭脂萝卜提取物在制备治疗阿尔茨海默症食品、保健品或药物的应用。
为了解决上述技术问题,本发明采用如下方案实现:
一种胭脂萝卜提取物的制备方法,将胭脂萝卜先后经过粉碎、浸提、过滤澄清、精制、浓缩以及干燥制得胭脂萝卜提取物。
具体步骤如下:
S1.粉碎:将胭脂萝卜清洗干净后粉碎;这里的粉碎为将胭脂萝卜切成薄片、小块或机器粉碎;
S2.浸提:将步骤S1的所得物使用溶剂进行浸提;所述溶剂为甲醇、乙醇或水,浸提过程也可使用微波或超声波进行辅助;
S3.过滤澄清:将步骤S2的所得物进行过滤,后对滤液进行澄清得胭脂萝卜粗提液;过滤操作可采用离心过滤或压榨过滤等方式;澄清操作可采用絮凝沉降后砂滤或管式离心等方式;
S4.精制:将步骤S3所得的胭脂萝卜粗提液加入至大孔树脂柱中进行静态吸附,然后依次用洗涤剂洗涤以及用洗脱剂洗脱;所述大孔树脂柱为DM130、 AB-8、X-5或NKA-9大孔树脂柱;所述洗涤剂为纯水;所述洗脱剂为乙醇溶液;
S5.浓缩:对步骤S4的所得物进行浓缩得胭脂萝卜浓缩液;浓缩操作可采用常规的减压浓缩;
S6.干燥:将步骤S5得到的胭脂萝卜浓缩液进行干燥,收集干燥粉末,得胭脂萝卜提取物;所述干燥方式为喷雾干燥或真空冷冻干燥。
一种用于防治阿尔茨海默症的药物,包括上述胭脂萝卜提取物以及药学上可接受的载体。所述药物的剂型为片剂、颗粒剂、粉剂、胶囊剂或口服液。
与现有技术相比,本发明具有如下有益效果:本发明经动物体内药效试验发现,胭脂萝卜提取物能显著改善老年痴呆模型SAMP8小鼠和D-半乳糖模型小鼠的认知和记忆能力,可用于阿尔茨海默症药物、保健品和食品等领域。
具体实施方式
为了让本领域的技术人员更好地理解本发明的技术方案,下面对本发明作进一步阐述。
实施例
胭脂萝卜提取物的制备,包括如下步骤:
S1:将胭脂萝卜洗干净,用粉碎机粉碎;
S2:将步骤1所得的胭脂萝卜碎块加入提取罐,提取罐中加入纯水,其中胭脂萝卜碎块质量(kg):纯水体积(L)=1:4;将胭脂萝卜碎块进行搅拌,用稀盐酸将pH调为2~3,维持搅拌30min(更好地对有效成分进行提取);
S3:将提取罐中的胭脂萝卜碎块悬浮液用平板离心机进行过滤,转速为1500r/min,过滤10min,得到第一次过滤液;将滤渣重新倒入提取罐,加入纯水,其中滤渣重量(kg):纯水体积(L)=1:1。将滤渣搅拌均匀后转入平板离心机过滤,转速为1500r/min,过滤10min,得到第二次过滤液。将第一次过滤液和第二次过滤液合并,通入管式离心机,得到澄清的胭脂萝卜粗提液;
S4:将胭脂萝卜粗提液泵入DM130大孔树脂柱,在流速为4BV/h的条件下,直至流出液出现红色为止。用3倍柱体积的纯水对树脂柱进行冲洗,收集洗涤液。再用体积比为60%的乙醇水溶液进行洗脱,直至洗脱液无红色为止,收集洗脱液;
S5:将收集的洗脱液置于真空浓缩机中,在真空度为0.04Mpa、温度为80℃的条件下,浓缩至体积为原洗脱液的1/4,得到胭脂萝卜浓缩液;
S6:将胭脂萝卜浓缩液泵入喷雾干燥机,进风温度为160℃,出风温度为 80℃,转速为10000r/min,收集干燥粉末,即得胭脂萝卜提取物。
对上述胭脂萝卜提取物进行动物体内药效试验:
试验例一:通过Barnes迷宫、避暗实验检测胭脂萝卜提取物改善SAMP8 小鼠学习记忆能力
以SPF级雄性SAMP8小鼠进行试验,随机分为模型组、阳性对照组、胭脂萝卜提取物低剂量组(RE-L)、胭脂萝卜提取物高剂量组(RE-H),同时以SAMR1 小鼠作为对照组。分组后第二天,阳性对照组按400mg/(kg·d)的剂量给予脑复康(脑复康溶于生理盐水制成混悬液)灌胃治疗;RE-L和RE-H分别按 151.7mg/(kg·d)、455.1mg/(kg·d)的剂量给予胭脂萝卜提取物(胭脂萝卜提取物分别溶于生理盐水制成混悬液)灌胃治疗;对照组、模型组分别灌胃等量溶剂(生理盐水),每日上午以0.1ml/10g连续灌胃给药8周。8周后,通过Barnes 迷宫、避暗实验来评估小鼠的空间学习记忆能力。结果见表1~3。
表1胭脂萝卜提取物对SAMP8小鼠Barnes迷宫测试潜伏期的影响
注:与对照组相比**P<0.01,***P<0.001;与模型组相比#P<0.05,###P<0.001
表2胭脂萝卜提取物对SAMP8小鼠Barnes迷宫测试错误次数的影响
注:与对照组相比*P<0.05,***P<0.001;与模型组相比##P<0.01,###P<0.001
由表1、表2可以看出,与对照组小鼠相比,模型组小鼠找到目标箱的潜伏期明显延长、错误次数明显增多,差异具有统计学意义(P<0.05或P<0.01或 P<0.001),表明模型组小鼠具有比较差的学习记忆能力;与模型组小鼠比较,阳性对照组、RE-L和RE-H小鼠找到目标箱的潜伏期明显缩短,错误次数明显减少,差异具有统计学意义(P<0.05或P<0.01或P<0.001)。试验结果表明,胭脂萝卜提取物和脑复康干预后,SAMP8小鼠的学习记忆能力得到改善,且胭脂萝卜提取物与脑复康具有相似的疗效。
表3胭脂萝卜提取物对SAMP8小鼠避暗实验的影响
注:与对照组相比**P<0.01,***P<0.001;与模型组相比#P<0.05,###P<0.001
由表3可以看出,与对照组小鼠相比,模型组小鼠逃避潜伏期明显较少,错误次数明显增加,差异具有统计学意义(P<0.01或P<0.001),说明模型组小鼠的学习记忆能力明显下降;与模型组小鼠相比,RE-H和阳性对照组小鼠逃避潜伏期明显增加,错误次数明显减少,差异具有统计学意义(P<0.05或P <0.001)。试验结果表明,胭脂萝卜提取物和脑复康干预后,SAMP8小鼠的学习记忆能力得到一定改善,且高剂量胭脂萝卜提取物与脑复康具有相似的疗效。
试验例二:通过Barnes迷宫、避暗实验检测胭脂萝卜提取物改善D-半乳糖模型小鼠学习记忆能力
以SPF级雄性昆明小鼠进行试验,随机分为对照组、模型组、阳性对照组、胭脂萝卜提取物低剂量组(RE-L)、胭脂萝卜提取物高剂量组(RE-H)。分组后第二天,阳性对照组按400mg/(kg·d)的剂量给予脑复康(脑复康溶于生理盐水制成混悬液)灌胃治疗;RE-L和RE-H分别按151.7mg/(kg·d)、455.1mg/(kg·d) 的剂量给予胭脂萝卜提取物(胭脂萝卜提取物分别溶于生理盐水制成混悬液) 灌胃治疗;对照组、模型组分别灌胃等量溶剂(生理盐水),每日上午以0.1ml/10g 连续灌胃给药30天。灌胃的同时,除对照组外,其余各组按120mg/(kg·d)的剂量颈部皮下注射D-半乳糖(D-半乳糖溶于生理盐水制成溶液),对照组颈部皮下注射等量生理盐水。给药结束后,通过Barnes迷宫、避暗实验来评估小鼠的空间学习记忆能力,结果见表4~6。
表4胭脂萝卜提取物对D-半乳糖模型小鼠Barnes迷宫测试潜伏期的影响
注:与对照组相比**P<0.01,***P<0.001;与模型组相比#P<0.05,###P<0.001
表5胭脂萝卜提取物对D-半乳糖模型小鼠Barnes迷宫测试错误次数的影响
注:与对照组相比*P<0.05,***P<0.001;与模型组相比##P<0.01,###P<0.001
由表4、表5可以看出,与对照组小鼠相比,模型组小鼠找到目标箱的潜伏期明显延长、错误次数明显增多,差异具有统计学意义(P<0.05或P<0.01或 P<0.001),表明模型组小鼠具有比较差的学习记忆能力,造模成功;与模型组小鼠比较,阳性对照组、RE-L和RE-H小鼠找到目标箱的潜伏期明显缩短,错误次数明显减少,差异具有统计学意义(P<0.05或P<0.01或P<0.001)。试验结果表明,胭脂萝卜提取物和脑复康干预后,D-半乳糖模型小鼠的学习记忆能力得到改善,且胭脂萝卜提取物与脑复康药效相似。
表6胭脂萝卜提取物对D-半乳糖模型小鼠避暗实验的影响
注:与对照组相比**P<0.01,***P<0.001;与模型组相比#P<0.05,###P<0.001
由表6可以看出,与对照组小鼠相比,模型组小鼠逃避潜伏期明显较少,错误次数明显增加,差异具有统计学意义(P<0.01或P<0.001),说明模型组小鼠的学习记忆能力明显下降,造模成功;与模型组相比,RE-H和阳性对照组小鼠逃避潜伏期明显增加,错误次数明显减少,差异具有统计学意义(P<0.05或 P<0.001)。试验结果表明,通过胭脂萝卜提取物和脑复康干预后,D-半乳糖模型小鼠的学习记忆能力得到改善,且高剂量胭脂萝卜提取物与脑复康具有相似的疗效。
由上述可以看出,胭脂萝卜提取物具有类似于脑复康的作用,能够改善SAMP8小鼠、D-半乳糖模型小鼠的认知、记忆能力,具有防治阿尔茨海默症的应用前景。
上述实施例仅为本发明的其中具体实现方式,其描述较为具体和详细,但并不能因此而理解为对本发明专利范围的限制。应当指出的是,对于本领域的普通技术人员来说,在不脱离本发明构思的前提下,还可以做出若干变形和改进,这些显而易见的替换形式均属于本发明的保护范围。
Claims (10)
1.胭脂萝卜提取物在制备防治阿尔茨海默症食品中的应用。
2.胭脂萝卜提取物在制备防治阿尔茨海默症保健品中的应用。
3.胭脂萝卜提取物在制备防治阿尔茨海默症药物中的应用。
4.一种用于防治阿尔茨海默症的药物,其特征在于,包括胭脂萝卜提取物和药学上可接受的载体。
5.根据权利要求4所述的用于防治阿尔茨海默症的药物,其特征在于,所述药物的剂型为片剂、颗粒剂、粉剂、胶囊剂或口服液。
6.一种胭脂萝卜提取物的制备方法,其特征在于,具体包括如下步骤:
S1.粉碎:将胭脂萝卜清洗干净后粉碎;
S2.浸提:将步骤S1的所得物使用溶剂进行浸提;
S3.过滤澄清:将步骤S2的所得物进行过滤,然后对滤液进行澄清得胭脂萝卜粗提液;
S4.精制:将步骤S3所得的胭脂萝卜粗提液加入至大孔树脂柱中进行静态吸附,然后依次用洗涤剂洗涤以及用洗脱剂洗脱;
S5.浓缩:对步骤S4的所得物进行浓缩得胭脂萝卜浓缩液;
S6.干燥:将步骤S5得到的胭脂萝卜浓缩液进行干燥,收集干燥粉末,得胭脂萝卜提取物。
7.根据权利要求6所述的胭脂萝卜提取物的制备方法,其特征在于,步骤S2中,所述溶剂为甲醇、乙醇或水。
8.根据权利要求6所述的胭脂萝卜提取物的制备方法,其特征在于,步骤S3中,过滤方式为离心过滤或压榨过滤;澄清方式为絮凝沉降后砂滤或管式离心。
9.根据权利要求6所述的胭脂萝卜提取物的制备方法,其特征在于,步骤S4中,所述大孔树脂柱为DM130、AB-8、X-5或NKA-9大孔树脂柱。
10.根据权利要求6所述的胭脂萝卜提取物的制备方法,其特征在于,步骤S4中,所述洗涤剂为纯水;所述洗脱剂为乙醇溶液。
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