CN112316122A - 一种治疗多囊卵巢综合征的组合物及其制备方法与应用 - Google Patents
一种治疗多囊卵巢综合征的组合物及其制备方法与应用 Download PDFInfo
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Abstract
本发明提供了一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物20~40份、鹰嘴豆粉10~20份、Mn‑SOD冻干粉10~30份、富硒茶油微囊粉10~20份、复合益生菌微囊粉1~10份、菊粉10~20份。本发明的治疗多囊卵巢综合征的组合物可以改善胰岛素活性、调节激素水平、降低氧化应激、抑制炎症、增加短链脂肪酸含量和调节肠道菌群,协同达到改善多囊卵巢综合征的效果,并且安全无副作用。
Description
技术领域
本发明涉及一种治疗多囊卵巢综合征的组合物及其制备方法与应用,属于功能食品领域。
背景技术
多囊卵巢综合征(Polycystic ovary syndrome,PCOS)是女性常见的与生殖和代谢紊乱有关的内分泌疾病,能够引起包括内分泌、糖脂代谢、生殖、心理健康等多方面的女性健康问题。PCOS的临床表现复杂多样,异质性极大,主要的临床特征为月经紊乱(主要为月经稀发和不规则子宫出血)、高雄激素血症及卵巢多囊样改变,常常伴有肥胖和胰岛素抵抗,是育龄期女性月经紊乱最常见的原因和导致无排卵性不孕的主要原因;在育龄期,还可能与流产、妊娠糖尿病和先兆子痫等妊娠并发症的患病率增加有关。依据不同的诊断标准,PCOS的全球发病率约为6-20%。PCOS除了引起育龄期妇女生育力降低、月经不调、肥胖、多毛等症状,还具有显着的代谢影响,如果PCOS未得到及时纠正,还会增加患2型糖尿病、心血管疾病、代谢综合征、子宫内膜癌等疾病的风险。
PCOS临床表现多样化,病因复杂化,大量研究表明高雄激素血症、胰岛素抵抗和血脂异常与PCOS的发生发展密切相关。越来越多的研究表明肠道菌群在PCOS的发病机制中起着重要的作用。在来曲唑诱导的PCOS小鼠模型中发现PCOS肠道菌群的厚壁菌门和拟杆菌门发生了显著改变。在另一项用来曲唑诱导的PCO大鼠模型与对照组相比,肠道菌群发生了变化,其乳酸杆菌、梭状芽孢杆菌减少,而普氏菌增多,用健康大鼠的乳酸杆菌和粪菌移植对PCOS大鼠进行干预,发现通过粪菌移植和乳酸杆菌移植的方法有益于PCOS大鼠的治疗,并且研究表明PCOS患者出现雄激素过多、胰岛素抵抗与肠道菌群密切相关。
目前临床上所采用的药物治疗主要是根据PCOS患者的诊断标准,针对患者内分泌异常使用抗雄激素药物来治疗育龄PCOS患者,长期服用后会对身体产生副作用,所以开发天然的、安全无副作用的治疗多囊卵巢综合征的产品显得尤为重要。
发明内容
针对现有技术的不足,尤其是目前治疗多囊卵巢综合征的产品副作用明显的缺陷,本发明提供了一种治疗多囊卵巢综合征的组合物及其制备方法与应用。本发明的治疗多囊卵巢综合征的组合物可以改善胰岛素活性、调节激素水平、降低氧化应激、抑制炎症、增加短链脂肪酸含量和调节肠道菌群,协同达到改善多囊卵巢综合征的效果,并且安全无副作用。
本发明的技术方案如下:
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物20~40份、鹰嘴豆粉10~20份、Mn-SOD冻干粉10~30份、富硒茶油微囊粉10~20份、复合益生菌微囊粉1~10份、菊粉10~20份。
根据本发明优选的,所述治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物25~35份、鹰嘴豆粉12~18份、Mn-SOD冻干粉15~25份、富硒茶油微囊粉12~17份、复合益生菌微囊粉2~8份、菊粉13~18份。
根据本发明优选的,所述治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、复合益生菌微囊粉5份、菊粉15份。
根据本发明优选的,所述角豆提取物中D-手性肌醇的质量含量≥95%。
根据本发明优选的,所述鹰嘴豆粉是按照如下方法制备的:将鹰嘴豆粉碎至60~80目,加入去离子水,得到含水量为5~10%的混合物,之后将混合物进行膨化,将膨化后的产物置于60~70℃烘箱中烘干20~60min,使其水分含量为2~6%,经二次粉碎和微波灭菌后得到鹰嘴豆粉;
进一步优选的,所述二次粉碎的粒径为80~100目;所述微波灭菌的条件为:温度60~70℃,时间2~9min。
根据本发明,所述的膨化采用双螺杆挤压机进行膨化,双螺杆挤压机分三个膨化区:一区、二区、三区,膨化时一区关闭,二区温度为110~120℃,三区温度为140~150℃;双螺杆挤压机的螺杆转速45~50rpm,喂料转速8-13rpm,旋切转速16~20rpm。
根据本发明优选的,所述的富硒茶油微囊粉是按照如下方法制备的:
a.将壁材加入水中,搅拌均匀,得到质量浓度为22.38%的壁材溶液;所述壁材为大豆分离蛋白与麦芽糊精的组合,其中,大豆分离蛋白与麦芽糊精的质量比为0.8~1.2:1;
b.将乳化剂加入水中,搅拌均匀,得到质量浓度为5.71%的乳化剂溶液;所述乳化剂为蔗糖脂肪酸酯与硬脂酸甘油的组合,其中,蔗糖脂肪酸酯与硬脂酸甘油的质量比为2~6:1;
c.将乳化剂溶液加入壁材溶液中,混合均匀后,在加入芯材富硒茶油的同时搅拌进行乳化,所述芯材和壁材的质量比为0.4~1:1,所述乳化剂的添加量为芯材质量的0.5~7%;之后在60~80℃下乳化15~25min,将所得乳化液在35~45MPa下均质3~5次;最后将均质后的乳化液通过喷雾干燥,制得富硒茶油微囊粉固体粉末;喷雾干燥条件为:进风温度为170~200℃、出风温度为60~80℃,风速为4.5~5.5m3/min。
根据本发明优选的,所述的复合益生菌微囊粉中的复合益生菌为乳杆菌、双歧杆菌、嗜粘蛋白-阿克曼氏菌(Akk菌)的组合,其中,乳杆菌、双歧杆菌、嗜粘蛋白-阿克曼氏菌的质量比为3:1:1;所述的乳杆菌、双歧杆菌为普通市售产品;所述的嗜粘蛋白-阿克曼氏菌(Akk菌)参照中国专利文献CN110551658A一种嗜黏蛋白-阿克曼氏菌的低氧培养方法得到;
进一步优选的,所述的乳杆菌为质量比1:1:1的嗜酸乳杆菌、罗伊氏乳杆菌和发酵乳杆菌的组合;所述的双歧杆菌为两歧双歧杆菌或动物双歧杆菌;
更优选的,所述的复合益生菌为质量比为1:1:1:1:1嗜酸乳杆菌、罗伊氏乳杆菌、发酵乳杆菌、两歧双歧杆菌、嗜粘蛋白-阿克曼氏菌(Akk菌)的组合。
根据本发明优选的,所述的复合益生菌微囊粉是按照如下方法制备的:
(ⅰ)大豆蛋白与卡拉胶美拉德反应产物的制备:
按照大豆蛋白和I-卡拉胶质量比为0.5~6:3的比例称取大豆蛋白和I-卡拉胶,溶于pH值为8.0的去离子水中,配成I-卡拉胶浓度为0.3~0.7%(w/v)的溶液,于4℃下搅拌8~24h至混合均匀;将所得大豆蛋白和I-卡拉胶的混合液进行喷雾干燥,喷雾干燥条件:入口温度110~130℃、出口温度70~90℃、泵速2~6rpm;将上述喷雾干燥所得的粉末于相对湿度65%、温度55℃下进行干法反应4~24h,得到大豆蛋白与卡拉胶美拉德反应产物;
(ⅱ)混合菌液的制备:
a.将冷冻保藏的乳杆菌、双歧杆菌,分别在MRS平板上画线培养,经过48h厌氧培养后,挑单菌落于20mL MRS液体培养基中,并在冲氮后37℃厌氧培养24h,然后以体积比为1.0%的接种量转接到MRS液体培养基中,厌氧培养12h后,将处于对数稳定期的菌液,通过冷冻高速离心机在4℃、5000r/min下离心15min,去掉上清液,并用无菌生理盐水洗涤两次,重新悬浮于生理盐水中,菌体浓度调整至4x109CFU/mL,分别得到乳杆菌菌液、双歧杆菌菌液;
b.将处于对数稳定期的嗜粘蛋白-阿克曼氏菌(Akkermansia muciniphila)菌液,通过冷冻高速离心机在4℃、5000r/min下离心15min,去掉上清液,并用无菌生理盐水洗涤两次,重新悬浮于生理盐水中,菌体浓度调整至4x109CFU/mL,得到嗜粘蛋白-阿克曼氏菌菌液;
将制备得到的乳杆菌菌液、双歧杆菌菌液、嗜粘蛋白-阿克曼氏菌菌液混合均匀,得到混合菌液;
(ⅲ)微胶囊的制备:
以步骤(ⅰ)制备的大豆蛋白和I-卡拉胶美拉德反应产物为微胶囊化壁材,将其加入去离子水中,充分搅拌混合均匀,得到浓度为0.3~0.7%(w/v)的壁材溶液;按照混合菌液与壁材溶液体积比1~2:7的比例,向壁材溶液中添加混合菌液,搅拌均匀后,进行冷冻干燥8~24h后,立即收集制备得到的复合益生菌微囊粉末于无菌密封玻璃瓶中,并储存于4℃保藏。
根据本发明,上述治疗多囊卵巢综合征的组合物的制备方法,包括步骤如下:
(1)将角豆提取物、鹰嘴豆粉、富硒茶油微囊粉、菊粉分别过80目筛,混合均匀,得混合物,向混合物中加入水,所述水的加入量为混合物质量的3~15%,制得软材;然后过20目筛进行制粒,制粒后于50~60℃烘干2~3h,再经整粒,筛粉后得到初级混合物;
(2)向步骤(1)得到的初级混合物中加入Mn-SOD冻干粉、复合益生菌微囊粉,混合20~25min,得到治疗多囊卵巢综合征的组合物。
根据本发明,上述治疗多囊卵巢综合征的组合物在制备具有改善多囊卵巢综合征的食品、药品或保健品中的应用。
本发明的技术特点在于:
1、本发明采用的角豆提取物(D-手性肌醇)是一种来源于角豆的天然降糖成分,取自地中海沿岸的豆类植物角豆豆荚,D-手性肌醇可以有效改善多囊卵巢综合征(PCOS)的胰岛素活性、调节内分泌及改善荷尔蒙均衡,使患者排卵正常,激素水平回归,解决多囊卵巢综合征问题。
2、本发明采用的鹰嘴豆营养成分丰富,膳食纤维是鹰嘴豆中精华部分,鹰嘴豆中总膳食纤维含量(DFC)为18-22g/100g,其膳食纤维含量是谷类和油籽豆类的两倍以上,而膳食纤维对衰老大鼠卵巢激素分泌失衡状态有明显的调节作用,且修复受损卵巢组织延缓卵巢衰老,减少对卵巢功能的影响;鹰嘴豆中的膳食纤维,能促进人体中的肠道微生物生成短链脂肪酸,乙酸能够激活小鼠的副交感神经,促进胰岛素、ghrelin的分泌,改善胰岛素抵抗和高雄激素血症;同时鹰嘴豆中富含植物蛋白,会降低排卵性不孕的风险。
3、本发明采用的Mn-SOD冻干粉是一种稳定性锰型超氧化物歧化酶(Mn-SOD),在高温、强酸和高胃蛋白酶含量条件下仍然具有良好的稳定性,可以经口进入机体内发挥作用,使用起来更方便,更容易准确定量。Mn-SOD可以通过降低肠道内细菌产生的超氧阴离子,降低氧化应激反应,纠正紊乱的肠道菌群从而改善糖代谢异常及胰岛素抵抗,改善小鼠卵巢多囊样改变,减少睾酮合成,降低血清睾酮水平,阻断多囊卵巢综合征的发生。
4、本发明采用的富硒茶油微囊粉中含有许多抗氧化物质,如角鲨烯、多酚类物质等,能提高总超氧化物歧化酶、谷胱甘肽过氧化物酶活力,提高其总抗氧化能力,减少丙二醛的产生,通过降低氧化应激反应,调整内分泌与激素水平,改善卵泡发育、卵母细胞成熟及排出,进而改善多囊卵巢综合征的发生;同时茶油中的亚麻酸能减轻PCOS大鼠的卵巢损伤,改善高雄激素血症、胰岛素抵抗、血脂异常和氧化应激水平;硒可减轻由CCl4诱发的肝损伤过程,提谷胱甘肽过氧化物酶活性,增强组织清除自由基的能力,通过降低氧化应激反应与茶油中的抗氧化物质协同改善多囊卵巢综合征。
5、本发明采用的复合益生菌微囊粉由嗜酸乳杆菌、罗伊氏乳杆菌、发酵乳杆菌、双歧杆菌、AKK菌复配而成,复合益生菌微囊粉能调节肠道菌群,促进有益菌生长,抑制有害菌繁殖,并可显著降低总睾酮、高敏C反应蛋白(hs-CRP)、丙二醛(MDA)水平和多毛症发生率,协同阻断多囊卵巢综合征的发生。
6、菊粉作为一种益生元,它是由β-(2-1)连接的果糖残基构成的植物储存多糖之一,是肠内双歧杆菌的活化增殖因子,可减少和抑制肠内腐败物质的产生,抑制有害细菌的生长,调节肠道内平衡,菊粉可以通过调节肠道菌群来改善多种代谢性疾病,同时菊粉可以通过抑制炎症和调节肠道菌群来改善多囊卵巢综合征。
本发明能够带来如下有益效果:
1、本发明提供的治疗多囊卵巢综合征的组合物,角豆提取物通过改善胰岛素活性、调节内分泌、调整激素水平达到改善多囊卵巢综合征的功效;鹰嘴豆调节卵巢激素分泌、修复受损卵巢组织延缓卵巢衰老,进而减少对卵巢功能的影响,同时鹰嘴豆中丰富的膳食纤维,增加肠道中短链链脂肪酸含量,改善胰岛素抵抗和高雄激素血症,达到改善多囊卵巢综合征的功效;Mn-SOD冻干粉通过降低氧化应激反应改善糖代谢异常及胰岛素抵抗,改善卵巢多囊样改变,减少睾酮合成,降低血清睾酮水平,阻断多囊卵巢综合征的发生;富硒茶油微囊粉通过降低氧化应激反应,调整内分泌与激素水平,改善卵泡发育、卵母细胞成熟及排出,进而改善多囊卵巢综合征的发生;复合益生菌微囊粉调节肠道菌群,并可显著降低总睾酮、高敏C反应蛋白(hs-CRP)、丙二醛(MDA)水平和多毛症发生率,进而阻断多囊卵巢综合征的发生;菊粉通过抑制炎症和调节肠道菌群来改善多囊卵巢综合征。
2、本发明提供的治疗多囊卵巢综合征的组合物,角豆提取物和鹰嘴豆协同调节激素水平、并通过改善胰岛素活性及修复受损卵巢组织,达到改善多囊卵巢综合征的功效;Mn-SOD冻干粉和富硒茶油微囊粉协同降低氧化应激反应改善糖代谢异常及胰岛素抵抗,改善卵巢多囊样改变和卵泡发育、卵母细胞成熟及排出,进而改善多囊卵巢综合征的发生;复合益生菌微囊粉与菊粉协同调节肠道菌群,增加有益菌相对丰度,抑制有害菌繁殖,实现治疗多囊卵巢综合征的功效;Mn-SOD冻干粉与复合益生菌微囊粉协同减少睾酮合成,降低血清睾酮水平,阻断多囊卵巢综合征的发生。
3、本发明的鹰嘴豆粉采用混合膨化处理,膨化处理可以有效去除了鹰嘴豆中的毒素和难消化物质,很好的保留了鹰嘴豆中的膳食纤维含量,提高可溶性膳食纤维含量,同时显著提高了鹰嘴豆的消化率及吸收利用率。
4、富硒茶油经过微囊包埋后,角鲨烯含量上升,抗氧化活性增大,降低氧化应激效果显著,进而改善糖代谢异常及胰岛素抵抗,显著改善多囊卵巢综合征。
5、以美拉德反应产物为壁材,制备的复合益生菌微囊粉可以耐受胃酸、胆盐等极端环境的影响,通过胃肠屏障后到达人体肠道仍能保持较高的生物活性,提高了生物利用率。
6、本发明提供的治疗多囊卵巢综合征的组合物综合了以上6种成分的功效,合理配伍,安全有效,各种成分通过不同途径进行协同作用,改善胰岛素抵抗、改善小鼠卵巢多囊样改变、减少睾酮合成、降低血清睾酮水平、调节激素水平、降低氧化应激、抑制炎症、增加短链脂肪酸含量和调节肠道菌群,改善多囊卵巢综合征效果更显著。
附图说明
图1为实验例3对目标菌群相对丰度的分析图。
具体实施方式
下面结合具体实施例对本发明进行详细说明,以下实施例仅为方便本领域技术人员理解本发明技术方案,实现或使用本发明所做的说明,并不以此限制本发明的保护范围。
角豆提取物,广东诚华健康科技有限公司有售;鹰嘴豆,罗庄区亿鑫粮行有售;Mn-SOD冻干粉,杭州药明康德公司有售;富硒茶油,江西御润坊富硒山茶油有限公司有售;乳杆菌和双歧杆菌,润盈生物工程(上海)有限公司有售;菊粉,丰宁平安高科实业有限公司有售。
实施例1
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、复合益生菌微囊粉5份、菊粉15份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉是按照如下方法制备的:将鹰嘴豆粉碎至80目,加入去离子水,得到含水量为8%的混合物,接着采用双螺杆挤压机对上述混合物进行膨化,将膨化后的产物置于65℃烘箱中烘干25min,使其水分含量为3%,经二次粉碎至80目,在60℃下微波灭菌6min后得到;
所述膨化的条件为:膨化时一区关闭,二区温度为110℃,三区温度为140℃;双螺杆挤压机的螺杆转速50rpm,喂料转速13rpm,旋切转速20rpm。
所述的富硒茶油微囊粉是按照如下方法制备的:
a.将壁材加入水中,搅拌均匀,得到质量浓度为22.38%的壁材溶液,所述壁材为大豆分离蛋白与麦芽糊精的组合,大豆分离蛋白与麦芽糊精的质量比为1:1;
b.将乳化剂加入水中,搅拌均匀,得到质量浓度为5.71%的乳化剂溶液,所述乳化剂为蔗糖脂肪酸酯与硬脂酸甘油的组合,其中,蔗糖脂肪酸酯与硬脂酸甘油的质量比为4.5:1;
c.将乳化剂溶液加入壁材溶液中,混合均匀后,在加入芯材富硒茶油的同时搅拌进行乳化,所述芯材和壁材的质量比为0.7:1,乳化剂添加量为芯材质量的5%,之后在70℃下乳化20min,采用高压均质设备将上述乳化液在38MPa下均质5次;最后将均质后的乳化液通过喷雾干燥,制得富硒茶油微胶囊固体粉末。喷雾干燥条件为:进风温度为182℃、出风温度为65℃,风速为5m3/min。
所述的复合益生菌微囊粉中的复合益生菌为质量比为1:1:1:1:1的嗜酸乳杆菌、罗伊氏乳杆菌、发酵乳杆菌、两歧双歧杆菌、嗜粘蛋白-阿克曼氏菌(Akk菌)的组合。
所述的复合益生菌微囊粉是按照如下方法制备的:
(ⅰ)大豆蛋白与卡拉胶美拉德反应产物的制备:
按照大豆蛋白和I-卡拉胶质量比为1.2:3的比例称取大豆蛋白和I-卡拉胶,溶于pH值为8.0的去离子水中,配成I-卡拉胶浓度为0.6%(w/v)的溶液,于4℃下搅拌12h至混合均匀。将所得大豆蛋白和I-卡拉胶的混合液进行喷雾干燥,喷雾干燥条件:入口温度120℃、出口温度80℃、泵速4rpm。将上述喷雾干燥的所得粉末于相对湿度65%、温度55℃下进行干法反应8h,得到大豆蛋白与卡拉胶美拉德反应产物;所述pH值为8.0的去离子水是用氢氧化钠调节去离子水的pH至8.0得到的。
(2)混合菌液的制备:
a.将冷冻保藏的嗜酸乳杆菌、罗伊氏乳杆菌、发酵乳杆菌、两歧双歧杆菌,分别通过螺旋平板接种仪在MRS平板上画线培养,经过48h厌氧培养后,挑单菌落于20mL MRS液体培养基中,并在冲氮后37℃厌氧培养24h,然后以体积比1.0%的接种量转接到MRS液体培养基中,厌氧培养12h后,将处于对数稳定期的菌液,通过冷冻高速离心机在4℃、5000r/min下离心15min,去掉上清液,并用无菌生理盐水洗涤两次,重新悬浮于生理盐水中,菌体浓度调整至4x109CFU/mL,分别得到嗜酸乳杆菌菌液、罗伊氏乳杆菌菌液、发酵乳杆菌菌液、两歧双歧杆菌菌液;
b.嗜粘蛋白-阿克曼氏菌(Akkermansia muciniphila)参照中国专利文献CN110551658A一种嗜黏蛋白-阿克曼氏菌的低氧培养方法得到,将处于对数稳定期的菌液,通过冷冻高速离心机在4℃、5000r/min下离心15min,去掉上清液,并用无菌生理盐水洗涤两次,重新悬浮于生理盐水中,菌体浓度调整至4x109CFU/mL,得到嗜粘蛋白-阿克曼氏菌菌液;
将制备得到的嗜酸乳杆菌菌液、罗伊氏乳杆菌菌液、发酵乳杆菌菌液、两歧双歧杆菌菌液、嗜粘蛋白-阿克曼氏菌菌液混合均匀,得到混合菌液;混合菌液中嗜酸乳杆菌、罗伊氏乳杆菌、发酵乳杆菌、两歧双歧杆菌、嗜粘蛋白-阿克曼氏菌质量比为1:1:1:1:1。
(ⅲ)微胶囊的制备:
以步骤(ⅰ)制备的大豆蛋白和I-卡拉胶美拉德反应产物为微胶囊化壁材,将其加入去离子水中,充分搅拌混合均匀,得到浓度为0.6%(w/v)的壁材溶液;按照混合菌液与壁材溶液体积比2:7的比例,向壁材溶液中添加混合菌液,搅拌均匀后,进行冷冻干燥20h后立即收集制备得到的复合益生菌微囊粉末于无菌密封玻璃瓶中,并储存于4℃保藏;
上述治疗多囊卵巢综合征的组合物的制备方法,包括步骤如下:
(1)将角豆提取物、鹰嘴豆粉、富硒茶油微囊粉、菊粉各原料过80目筛,按比例混合均匀,得混合物,向混合物中加入混合物质量8%的水,制得软材;然后过20目筛进行制粒,制粒后于60℃烘干2h,再经整粒,筛粉后得到初级混合物;
(2)按配比向步骤(1)得到的初级混合物中加入Mn-SOD冻干粉、复合益生菌微囊粉,混合25min,得到治疗多囊卵巢综合征的组合物。
实施例2
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物40份、鹰嘴豆粉10份、Mn-SOD冻干粉30份、富硒茶油微囊粉10份、复合益生菌微囊粉10份、菊粉10份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
上述治疗多囊卵巢综合征的组合物的制备方法如实施例1所述。
实施例3
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物20份、鹰嘴豆粉20份、Mn-SOD冻干粉10份、富硒茶油微囊粉20份、复合益生菌微囊粉1份、菊粉20份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
上述治疗多囊卵巢综合征的组合物的制备方法如实施例1所述。
对比例1
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物20份、鹰嘴豆粉30份、Mn-SOD冻干粉40份、富硒茶油微囊粉30份、复合益生菌微囊粉15份、菊粉30份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
上述治疗多囊卵巢综合征的组合物的制备方法如实施例1所述。
对比例2
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物40份、鹰嘴豆粉5份、Mn-SOD冻干粉5份、富硒茶油微囊粉5份、复合益生菌微囊粉0.1份、菊粉5份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
上述治疗多囊卵巢综合征的组合物的制备方法如实施例1所述。
对比例3
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、复合益生菌微囊粉5份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
本对比例的治疗多囊卵巢综合征的组合物中不含有菊粉,其制备方法如实施例1所述。
对比例4
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、Mn-SOD冻干粉20份、复合益生菌微囊粉5份、菊粉15份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、复合益生菌微囊粉的制备过程同实施例1。
本对比例的治疗多囊卵巢综合征的组合物中不含有富硒茶油微囊粉,其制备方法如实施例1所述。
对比例5
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、复合益生菌微囊粉5份、菊粉15份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
本对比例的治疗多囊卵巢综合征的组合物中不含有鹰嘴豆粉,其制备方法如实施例1所述。
对比例6
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、富硒茶油微囊粉20份、复合益生菌微囊粉5份、菊粉15份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
本对比例的治疗多囊卵巢综合征的组合物中不含有Mn-SOD冻干粉,其制备方法如实施例1所述。
对比例7
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、菊粉15份。
所述角豆提取物中D-手性肌醇的含量≥95%。
所述鹰嘴豆粉、富硒茶油微囊粉的制备过程同实施例1。
本对比例的治疗多囊卵巢综合征的组合物中不含有复合益生菌微囊粉,其制备方法如实施例1所述。
对比例8
一种治疗多囊卵巢综合征的组合物,包括如下重量份的原料:鹰嘴豆粉15份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、复合益生菌微囊粉5份、菊粉15份。
所述鹰嘴豆粉、富硒茶油微囊粉、复合益生菌微囊粉的制备过程同实施例1。
本对比例的治疗多囊卵巢综合征的组合物中不含有角豆提取物,其制备方法如实施例1所述。
实验例1
本发明组合物改善多囊卵巢综合征的功效采用Poretsky等胰岛素(INS)联合HCG造模法进行评价。
试验对象:大鼠试验中选用SPF级雌性SD大鼠,体重为160~180g。
试验方法:
所有试验用大鼠(80只),按体重随机分成空白对照组(10只)和试验组(70只),置于SPF级的动物饲养室中喂养,给予充足的食物和浓度为5%的葡萄糖饮水,每天给予14h的周期性光照。实验的第1~12天给予试验组大鼠逐渐增加剂量的中效胰岛素诺和灵,由开始的0.5IU/d逐渐增加至6.0IU/d,增加梯度为0.5IU/d,第13~24天给予固定剂量6.0IU/d,并加用HCG(人体绒毛膜促性腺激素)6.0IU/d,其中HCG分2次注射,每次注射剂量为3.0IU,对照组以注射相同剂量的生理盐水为空白对照。造模后,连续阴道上皮细胞涂片两个性周期(每个周期5天),阴道角化细胞持续出现,提示造模成功。造模成功后将试验组动物平均分成7组,分别为模型对照组、实验组1~5和阳性对照组,实验组1~3组给予实施例1~3的组合物的配比,实验组4~5组分别给予对比例1~2的组合物的配比,人体推荐量为每天3.33g/60kg.bw(仅主药量),按中剂量组555mg/kg.bw(相当于人体推荐摄入量的10倍),以纯化水为溶剂配至所需浓度,每只大鼠按0.2mL/10g.bw灌胃,空白对照组和模型对照组用纯化水灌胃,阳性对照组:己烯雌酚0.5mg/kg.bw灌胃,持续15d。末次给药后,各组大鼠禁食12h,不禁水,采用酶联免疫法测定大鼠体内相关激素水平。
使用SPSS统计软件进行数据处理,t检验进行各组间显著性差异分析,P<0.05为差异显著,P<0.01为差异极显著。所有检测指标的数据均以均数±标准差表示,测试结果见表1。
表1:实验组1~5、模型对照组、空白对照组及阳性对照组对试验大鼠激素水平的影响
注:*表示与模型对照组比较,*P<0.05,**P<0.01;
孕酮-P、睾酮-T、性激素结合球蛋白-SHBG、促黄体生成素-LH、促卵泡生成激素-FSH。
由表1可知,模型对照组同空白对照组相比,大鼠血清P水平、FSH水平下降,T水平、LH水平上升,差异显著(P<0.05),表明大鼠建模成功。实验组1和阳性对照组与模型对照组相比,大鼠血清P水平、T水平、SHBG水平、LH水平、FSH水平及LH/FSH比值,差异极显著(P<0.01),实验组2~3中的大鼠血清P水平、T水平、SHBG水平、LH水平、FSH水平及LH/FSH比值与模型对照组相比,差异显著(P<0.05);实验组4~5中各指标值(P、T、SHBG、LH、FSH、LH/FSH)对于模型对照组P>0.05,表明无显著差异性。
以上实验结果表明,本发明实施例1~3的组合物相较于对比例1~2的组合物,在调整各成分用量后,对大鼠改善多囊卵巢综合征的作用明显增强。
实验例2
根据实验例1的实验结果,实验样品选取实施例1中的组合物,人体推荐量为每天3.33g/60kg.bw(仅主药量),实验设低、中、高三个剂量组,分别为277.5mg/kg.bw、555mg/kg.bw、1665mg/kg.bw,低、中、高三个组的剂量分别相当于人体推荐摄入量的5倍、10倍、30倍,依然以大鼠实验进行评价,将对比例3~8的组合物按高剂量组灌入。实验方法与实验例1中相同,试验组大鼠按随机平分12组,分别为模型对照组、空白对照组、阳性对照组和实验组1~9,实验组1~3分别以实施例1低中高三个剂量,实验组4~9以对比例3~8中组合物的配比,以纯化水为溶剂配至所需浓度,每只大鼠按0.2mL/10g.bw灌胃,空白对照组和模型对照组纯化水灌胃,阳性对照组:己烯雌酚0.5mg/kg.bw灌胃。大鼠体内相关激素水平的测定和数据处理同实验例1,各组实验指标如表2所示:
表2:实验组1~9、模型对照组、空白对照组及阳性对照组对试验大鼠激素水平的影响
注:*表示与模型对照组比较,*P<0.05,**P<0.01;
孕酮-P、睾酮-T、性激素结合球蛋白-SHBG、促黄体生成素-LH、促卵泡生成激素-FSH。
由表2可知,模型对照组同空白对照组相比,大鼠血清P水平、FSH水平下降,T水平、LH水平上升,差异显著(P<0.05),表明大鼠建模成功。实验组1,除大鼠血清P与模型对照组无差异性,其他指标(T、SHBG、LH、FSH、LH/FSH)与模型对照组相比,差异显著(P<0.05);实验组2~3和阳性对照组与模型对照组相比,大鼠血清P水平、T水平、SHBG水平、LH水平、FSH水平及LH/FSH比值,差异极显著(P<0.01);实验组4~9中各指标值(P、T、SHBG、LH、FSH、LH/FSH)对于模型对照组P>0.05,表明无显著差异性。
以上数据表明,本发明提供的治疗多囊卵巢综合征的组合物角豆提取物、鹰嘴豆粉、Mn-SOD冻干粉、富硒茶油微囊粉、复合益生菌微囊粉及菊粉产生协同,可以改善胰岛素抵抗、改善小鼠卵巢多囊样改变、减少睾酮合成、降低血清睾酮水平、增加短链脂肪酸含量和调节肠道菌群,阻断多囊卵巢综合征的发生。
实验例3对肠道菌群的调节作用
大鼠粪便的收集
在实验例1中大鼠饲养周期末,将各笼大鼠的垫料去除后,分别收集各笼大鼠排出的新鲜粪便中段置于取样盒中,并立即置于干冰上,随即储存在-80℃超低温冰箱中待DNA提取及肠道菌群检测。
Illumina Miseq PE250测序
将保存于-80℃的粪便样本根据文献Molecular microbial diversity of ananaerobic digestor as determined by small-subunit rDNA sequence analysis的描述,提取DNA。所有DNA样本在Illumina Miseq平台使用Miseq试剂盒V3(600循环数)进行测序,并对测序数据进行生物信息学分析,结果如图1所示。
肠道菌群主要有9个门的菌群,其中98%的细菌可归于以下4个,拟杆菌门、厚壁菌门、变形菌门、放线菌门。通过各实验组及对照组肠道菌群的不同菌门的相对丰度分析,很明显看出,厚壁菌门和拟杆菌门在所有样品中都占据了绝对优势,约为肠道菌群的90%左右。相对于空白对照组,模型对照组F/B比值下降,与空白对照组具有显著差异性。实验组1~3中F/B比值与模型对照组比较发生明显升高,具有显著差异性(P<0.05),实验组4~5中F/B比值与模型对照无显著差异性,表明实验组1~3组合物配比可以对大鼠肠道菌群结构发挥调节作用,进而改善多囊卵巢综合征的发生。
以上所述仅为本申请的实施例而已,并不用于限制本申请。对于本领域技术人员来说,本申请可以有各种更改和变化。凡在本申请的精神和原理之内所作的任何修改、等同替换、改进等,均应包含在本申请的权利要求范围之内。
Claims (10)
1.一种治疗多囊卵巢综合征的组合物,其特征在于,所述的组合物包括如下重量份的原料:角豆提取物20~40份、鹰嘴豆粉10~20份、Mn-SOD冻干粉10~30份、富硒茶油微囊粉10~20份、复合益生菌微囊粉1~10份、菊粉10~20份。
2.根据权利要求1所述的治疗多囊卵巢综合征的组合物,其特征在于,所述治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物25~35份、鹰嘴豆粉12~18份、Mn-SOD冻干粉15~25份、富硒茶油微囊粉12~17份、复合益生菌微囊粉2~8份、菊粉13~18份;优选的,所述治疗多囊卵巢综合征的组合物,包括如下重量份的原料:角豆提取物30份、鹰嘴豆粉15份、Mn-SOD冻干粉20份、富硒茶油微囊粉15份、复合益生菌微囊粉5份、菊粉15份。
3.根据权利要求1所述的治疗多囊卵巢综合征的组合物,其特征在于,所述角豆提取物中D-手性肌醇的质量含量≥95%。
4.根据权利要求1所述的治疗多囊卵巢综合征的组合物,其特征在于,所述鹰嘴豆粉是按照如下方法制备的:将鹰嘴豆粉碎至60~80目,加入去离子水,得到含水量为5~10%的混合物,之后将混合物进行膨化,将膨化后的产物置于60~70℃烘箱中烘干20~60min,使其水分含量为2~6%,经二次粉碎和微波灭菌后得到鹰嘴豆粉;
所述二次粉碎的粒径为80~100目;所述微波灭菌的条件为:温度60~70℃,时间2~9min。
5.根据权利要求1所述的治疗多囊卵巢综合征的组合物,其特征在于,所述的富硒茶油微囊粉是按照如下方法制备的:
a.将壁材加入水中,搅拌均匀,得到质量浓度为22.38%的壁材溶液;所述壁材为大豆分离蛋白与麦芽糊精的组合,其中,大豆分离蛋白与麦芽糊精的质量比为0.8~1.2:1;
b.将乳化剂加入水中,搅拌均匀,得到质量浓度为5.71%的乳化剂溶液;所述乳化剂为蔗糖脂肪酸酯与硬脂酸甘油的组合,其中,蔗糖脂肪酸酯与硬脂酸甘油的质量比为2~6:1;
c.将乳化剂溶液加入壁材溶液中,混合均匀后,在加入芯材富硒茶油的同时搅拌进行乳化,所述芯材和壁材的质量比为0.4~1:1,所述乳化剂的添加量为芯材质量的0.5~7%;之后在60~80℃下乳化15~25min,将所得乳化液在35~45MPa下均质3~5次;最后将均质后的乳化液通过喷雾干燥,制得富硒茶油微囊粉固体粉末;喷雾干燥条件为:进风温度为170~200℃、出风温度为60~80℃,风速为4.5~5.5m3/min。
6.根据权利要求1所述的治疗多囊卵巢综合征的组合物,其特征在于,所述的复合益生菌微囊粉中的复合益生菌为乳杆菌、双歧杆菌、嗜粘蛋白-阿克曼氏菌(Akkermansiamuciniphila,Akk菌)的组合,其中,乳杆菌、双歧杆菌、嗜粘蛋白-阿克曼氏菌的质量比为3:1:1。
7.根据权利要求6所述的治疗多囊卵巢综合征的组合物,其特征在于,所述的乳杆菌为为质量比1:1:1的嗜酸乳杆菌、罗伊氏乳杆菌和发酵乳杆菌的组合;所述的双歧杆菌为两歧双歧杆菌或动物双歧杆菌;优选的,所述的复合益生菌为质量比为1:1:1:1:1嗜酸乳杆菌、罗伊氏乳杆菌、发酵乳杆菌、两歧双歧杆菌、嗜粘蛋白-阿克曼氏菌(Akk菌)的组合。
8.根据权利要求1所述的治疗多囊卵巢综合征的组合物,其特征在于,所述的复合益生菌微囊粉是按照如下方法制备的:
(ⅰ)大豆蛋白与卡拉胶美拉德反应产物的制备:
按照大豆蛋白和I-卡拉胶质量比为0.5~6:3的比例称取大豆蛋白和I-卡拉胶,溶于pH值为8.0的去离子水中,配成I-卡拉胶浓度为0.3~0.7%(w/v)的溶液,于4℃下搅拌8~24h至混合均匀;将所得大豆蛋白和I-卡拉胶的混合液进行喷雾干燥,喷雾干燥条件:入口温度110~130℃、出口温度70~90℃、泵速2~6rpm;将上述喷雾干燥所得的粉末于相对湿度65%、温度55℃下进行干法反应4~24h,得到大豆蛋白与卡拉胶美拉德反应产物;
(ⅱ)混合菌液的制备:
a.将冷冻保藏的乳杆菌、双歧杆菌,分别在MRS平板上画线培养,经过48h厌氧培养后,挑单菌落于20mL MRS液体培养基中,并在冲氮后37℃厌氧培养24h,然后以体积比为1.0%的接种量转接到MRS液体培养基中,厌氧培养12h后,将处于对数稳定期的菌液,通过冷冻高速离心机在4℃、5000r/min下离心15min,去掉上清液,并用无菌生理盐水洗涤两次,重新悬浮于生理盐水中,菌体浓度调整至4x109CFU/mL,分别得到乳杆菌菌液、双歧杆菌菌液;
b.将处于对数稳定期的嗜粘蛋白-阿克曼氏菌(Akkermansia muciniphila)菌液,通过冷冻高速离心机在4℃、5000r/min下离心15min,去掉上清液,并用无菌生理盐水洗涤两次,重新悬浮于生理盐水中,菌体浓度调整至4x109CFU/mL,得到嗜粘蛋白-阿克曼氏菌菌液;
将制备得到的乳杆菌菌液、双歧杆菌菌液、嗜粘蛋白-阿克曼氏菌菌液混合均匀,得到混合菌液;
(ⅲ)微胶囊的制备:
以步骤(ⅰ)制备的大豆蛋白和I-卡拉胶美拉德反应产物为微胶囊化壁材,将其加入去离子水中,充分搅拌混合均匀,得到浓度为0.3~0.7%(w/v)的壁材溶液;按照混合菌液与壁材溶液体积比1~2:7的比例,向壁材溶液中添加混合菌液,搅拌均匀后,进行冷冻干燥8~24h后,立即收集制备得到的复合益生菌微囊粉末于无菌密封玻璃瓶中,并储存于4℃保藏。
9.权利要求1-8任一项所述的治疗多囊卵巢综合征的组合物的制备方法,包括步骤如下:
(1)将角豆提取物、鹰嘴豆粉、富硒茶油微囊粉、菊粉分别过80目筛,混合均匀,得混合物,向混合物中加入水,所述水的加入量为混合物质量的3~15%,制得软材;然后过20目筛进行制粒,制粒后于50~60℃烘干2~3h,再经整粒,筛粉后得到初级混合物;
(2)向步骤(1)得到的初级混合物中加入Mn-SOD冻干粉、复合益生菌微囊粉,混合20~25min,得到治疗多囊卵巢综合征的组合物。
10.权利要求1-8任一项所述的治疗多囊卵巢综合征的组合物在制备具有改善多囊卵巢综合征的食品、药品或保健品中的应用。
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