CN116650546A - 一种苣荬菜水提物在防治溃疡性结肠炎中的应用 - Google Patents
一种苣荬菜水提物在防治溃疡性结肠炎中的应用 Download PDFInfo
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- CN116650546A CN116650546A CN202310015917.8A CN202310015917A CN116650546A CN 116650546 A CN116650546 A CN 116650546A CN 202310015917 A CN202310015917 A CN 202310015917A CN 116650546 A CN116650546 A CN 116650546A
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- 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
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- 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
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- Medicines Containing Plant Substances (AREA)
Abstract
本申请公开了一种苣荬菜水提物在防治溃疡性结肠炎中的应用。本申请的苣荬菜水提物在防治溃疡性结肠炎中能够有效治疗和缓解溃疡性结肠炎的症状：包括体重变化、结肠缩短、DAI评分、病理损伤等情况；16SrRNA检测显示，苣荬菜水提物处理后的肠道微生物群在门、纲、目、科、属水平明显升高，各组间肠道菌群组成存在显著差异。因此，苣荬菜水提物能够作为治疗或者预防溃疡性结肠炎的有效药物，其对小鼠结肠炎的保护作用可能与肠道微生物群多样性有关。
Description
技术领域
本申请涉及医疗医药技术领域，尤其涉及一种苣荬菜水提物在防治溃疡性结肠炎中的应用。
背景技术
苣荬菜为菊科(Compositae)苦苣菜属(Sonchus)植物苣荬菜(Sonchus arvensisL.)的干燥全草，又名苦菜、曲麻菜等。苣荬菜为多年生草本，主要在中国西北、华北、东北等地的荒山坡地、海滩、路旁野生。苣荬菜营养成分丰富，含有丰富的钙、镁、锰、钛、硒和锌等多种无机元素。全草均可入药。苣荬菜性味苦寒，具有清热解毒、活血化瘀、利湿排脓、凉血止血、祛瘀消肿等功效，常用于急性咽炎、急性细菌性痢疾、尿血、痔疮肿痛等症，民间也作野菜食用。苣荬菜中含有的化学成分有黄酮、生物碱、多糖类以及丰富的萜类成分。药理研究表明其具有降血压和降胆固醇作用、抗菌作用、抗炎活性、抗心率失常作用、抗肿瘤作用、保肝作用、免疫调节活性、抗疟活性、肾脏保护作用、解毒作用，具有较强的清除自由基和抗氧化的能力。苣荬菜在药用和保健方面均有使用价值。因此，苣荬菜的进一步研究开发具有重要意义，为之后的天然资源开发利用提供理论参考。
溃疡性结肠炎(UC)是一种胃肠道的慢性炎症性疾病，病变主要局限于大肠黏膜及黏膜下层，呈连续性弥漫性分布。病变大多自直肠开始，呈逆行性发展，累及直肠和乙状结肠多见。主要表现为腹泻、黏液脓血便、腹痛、里急后重等，可伴随发热、营养不良等全身表现。病程迁延不愈，轻重不等。目前，病因尚不清楚。近年来，UC的发病率呈明显升高趋势，与直结肠癌的发生密切相关，几率大大增加。相关数据显示，UC患者患直肠癌的风险要高出常人2-5倍，已被世界卫生组织(WHO)列为现代难治疾病之一。但关于其病因及发病机制尚未完全阐明，多认为与免疫异常、遗传、感染、环境、精神等因素有关。对于UC的治疗，西医主要运用水杨酸制剂、微生态制剂、糖皮质激素、抗生素及生物制剂等。西医UC的治疗虽然具有较强的针对性，但疗程长，成本高，副作用大，容易复发且有部分患者对药物不敏感。而手术治疗过程存在一定的死亡风险，术后并发症也很难避免，尤其是对于顽固性UC，更是治标不治本。因此，从天然资源中开发有效的药物对减少副作用和改善治疗效果尤为重要。而中药具有疗效确切，不良反应少，能有效减少病情复发等优势。所以，中药治疗UC具有很好的应用前景。
目前，国内外均未见苣荬菜水提取物在治疗溃疡性结肠炎中的应用的报道。
发明内容
鉴于现有技术中的上述缺陷或不足，期望提供一种苣荬菜水提物在防治溃疡性结肠炎中的应用。
本申请提供的一种苣荬菜水提物在防治溃疡性结肠炎中的应用，包括苣荬菜水提物在制备防治溃疡性结肠炎的药物中的应用。
优选的，苣荬菜水提物的提取制备方法包括以下步骤:
步骤一：称取苣荬菜干药材并剪碎、清洗，加入蒸馏水浸泡2小时；
步骤二：将浸泡后的混合液煎煮第一次，过滤并倾倒出水煎液，药渣加入蒸馏水煎煮第二次，过滤并倾倒出水煎液，合并两份水煎液；
步骤三：将合并后的水煎液离心，用旋转蒸发仪浓缩，真空冷冻干燥48小时得到苣荬菜水提物。
优选的，步骤一中苣荬菜干药材与蒸馏水的质量比为1:15～30。
优选的，苣荬菜水提物在防治溃疡性结肠炎中的应用为苣荬菜水提物能够减少溃疡性结肠炎症状中的一种以上，即缓解或治疗体重降低、腹泻、便血等公认的溃疡性结肠炎症状中的至少一种。
优选的，苣荬菜水提物能够改善结肠长度，改善或降低DAI评分。
优选的，苣荬菜水提物能够保护结肠粘膜屏障及修复DSS诱导的结肠损伤。
优选的，苣荬菜水提物能够调节肠道菌群组成及促进肠道菌群恢复。
优选的，药物是以苣荬菜提取物为活性成分，单独或与其他药物组方，加入药学上可接受的辅料、辅助性成分或载体制备成药学上可接受的剂型。
优选的，药物的剂型为散剂、胶囊剂、片剂、丸剂、颗粒剂、口服液体制剂、注射剂或靶向制剂。
优选的，药学上可接受的载体是指药学领域常规的药物载体，例如：稀释剂、赋形剂如水等，填充剂如淀粉、蔗糖等；黏合剂如纤维素衍生物、藻酸盐、明胶及聚乙烯吡咯烷酮；湿润剂如甘油；崩解剂如琼脂、碳酸钙及碳酸氢钠；吸收促进剂如季铵化合物；表面活性剂如十六烷醇；吸附载体如高岭土及皂黏土；润滑剂如滑石粉、硬脂酸钙、硬脂酸镁及聚乙二醇等。另外还可以加入其它药学领域常规的辅剂如甜味剂、香味剂等。
相对于现有技术而言，本申请的有益效果是：
本发明中的苣荬菜水提物，能够有效治疗和缓解溃疡性结肠炎的症状：包括体重降低、腹泻和便血等情况；能够有效缓解结肠缩短现象，降低DAI评分；能够调节肠道菌群组成、促进菌群恢复，改善肠道健康；以及能够进一步保护粘膜屏障和修复结肠损伤。因此，苣荬菜水提物可以作为溃疡性结肠炎的有效治疗药物或有效治疗成分。
应当理解，本发明中的药学上可接受的辅料、辅助性成分或载体是指与活性成分一同给药的稀释剂、辅剂、赋形剂或媒介物，并且其在合理的医学判断的范围内适于接触人类和/或其它动物的组织而没有过度的毒性、刺激、过敏反应、并发症或其它问题，并且具有合理的益处/风险比。
应当理解，本发明中，所用术语“治疗”，包括缓和、抑制或改善疾病的症状或状况；抑制并发症的产生：改善或预防潜在代谢综合征；抑制疾病或症状的产生，如控制疾病或情况的发展；减轻疾病或症状；使疾病或症状减退；减轻由疾病或症状引起的并发症，或预防或治疗由疾病或症状引起的征兆。如本文所用，给药后，可以使某一疾病、症状或情况得到改善，尤指其严重度得到改善，延迟发病，减缓病情进展，或减少病情持续时间。
应当理解，发明内容部分中所描述的内容并非旨在限定本申请的实施例的关键或重要特征，亦非用于限制本申请的范围。本申请的其它特征将通过以下的描述变得容易理解。
附图说明
通过阅读参照以下附图所作的对非限制性实施例所作的详细描述，本申请的其它特征、目的和优点将会变得更明显：
图1为本发明实施例中苣荬菜水提物降低DSS诱导的C57BL/6小鼠结肠炎的效果示意图，其中，A图：各组小鼠的体重变化；B图：疾病活动性指数(DAI)；C图和D图：各组小鼠的结肠的长度；
图2为本发明实施例中苣荬菜水提物对DSS诱导的小鼠结肠HE染色图；
图3为本发明实施例中苣荬菜水提物对DSS诱导的小鼠肠道菌群结构的影响示意图，其中，A图：稀疏曲线图；B图：维恩图；C图：等级聚类曲线图；D图：物种相对丰度柱形图；
图4为本发明实施例中苣荬菜水提物对DSS诱导的小鼠肠道菌群物种差异性的分析示意图，其中，包括：PCoA分析图及Beta多样性的箱形图。
具体实施方式
下面结合附图和实施例对本申请作进一步的详细说明。可以理解的是，此处所描述的具体实施例仅仅用于解释相关申请，而非对该申请的限定。另外还需要说明的是，为了便于描述，附图中仅示出了与申请相关的部分。
需要说明的是，在不冲突的情况下，本申请中的实施例及实施例中的特征可以相互组合。实施例中所使用的试验方法如无特殊说明，均为常规方法；所使用的材料、试剂等，如无特殊说明，均可从商业途径得到的试剂和材料。下面将参考附图并结合实施例来详细说明本申请。
实施例一：
苣荬菜水提物的制备：
步骤一：称取苣荬菜干药材100g并剪碎，用蒸馏水清洗，加入2500ml蒸馏水浸泡2小时；
步骤二：将浸泡后的混合液煎煮第一次40分钟，过滤并倾倒出水煎液，药渣加入蒸馏水煎煮第二次40分钟，过滤并倾倒出水煎液，合并两份水煎液；
步骤三：将合并后的水煎液离心，用旋转蒸发仪浓缩，真空冷冻干燥48小时得到深褐色苣荬菜水提物。
实施例二：
苣荬菜水提物对大鼠结肠组织的作用研究：
1.动物分组及给药方案
C57BL/6小鼠，雄性，8周龄，体质量20±2g，SPF级，购自北京维通利华实验动物技术有限公司。小鼠适应性饲养7天后，将小鼠随机分为4组，每组5只，分别设置为对照组(CL)、模型组(ML)、药物治疗组(MS)、药物组(SA)。
自实验第一天起，药物治疗组和药物组每天灌胃苣荬菜水提物，对照组和模型组每天给予等体积的生理盐水，连续21天，期间各组小鼠自由饮用灭菌水，第22天开始采用DSS诱导法建立溃疡性结肠炎模型：模型组和药物治疗组小鼠从第22天自由饮用2％DSS溶液，连续七天。
实施例三：
2.实验方法及数据处理
2.1疾病活动指数(Disease activity index，DAI)评分
每天观察并记录小鼠体重、饮食饮水状况、粪便黏稠情况和便血情况。按照DAI评分标准进行评分，按动物体质量下降百分率(体质量未下降为0；下降≤5％，且＞0为1分；下降≤10％，且＞5％为2分；下降≤15％，且＞10％为3分；下降＞15％为4分)、大便黏稠度(正常为0，松散的大便为2分，腹泻为4分)和大便出血(正常为0分，隐血阳性为2分，显性出血为4分)情况进行综合评分，将3项结果的总分除以3即得到DAI。
DAI＝(体质量指数+大便黏稠度指数+出血情况指数)/3。
2.2结肠长度
在实验第29天，采用颈椎脱臼法处死各组小鼠，分离整段结肠组织，用标尺测量各组小鼠结肠长度，进行统计学分析(**p<0.01)。
2.3结肠病理组织学观察
取各组小鼠病变结肠组织常规石蜡包埋，切片，HE染色。然后光学显微镜观察。
2.4结果
在实验过程中，每天监测小鼠的体重减轻、腹泻和血便。与对照组CL组相比，ML组和MS组小鼠的体重明显下降，尤其是ML组(图1A)。而MS组小鼠的体重减轻情况显著优于ML组(图1A)。
ML组和MS组小鼠的DAI评分较对照组CL组有显著升高。而MS组小鼠的DAI评分明显低于ML组(图1B)。如图1C和图1D所示，ML组小鼠的结肠变化呈现明显的缩短，而MS组小鼠的结肠变化较ML组呈现明显的改善。
如图2所示，对DSS诱导的UC小鼠给予苣荬菜水提物后，病理变化有明显改善。
实施例四：
3.肠道菌群分析：
3.1粪便样品采集及处理
在实验第29天处死小鼠，剖开腹腔，取出肛门至盲肠的完整结肠组织并置于冰盘上，取出盲肠内粪便收集于冻存管中，立即放于液氮内保存，待提取粪便中细菌的DNA。
3.2粪便中细菌DNA的提取及测序
收集盲肠内容物，采用CTAB法提取所有样本的总基因组DNA。扩增不同区域的16SrRNA/18SrRNA/ITS基因。然后对产物进行纯化和定量。序列分析采用Uparse软件进行。相似性≥97％的序列被分配到相同的OTUs上。筛选每个OTU的代表性序列以进行进一步注释。OTUs丰度信息使用序列最少的样本对应的序列号标准进行归一化。随后的alpha多样性和beta多样性分析都基于归一化数据进行分析。
3.3结果
如图3A所示，稀疏曲线结果显示测序数据量合理。如图3B维恩图显示，所有组共享670OTUs，CL组和ML组之间共享774OTUs，CL组和MS组之间共享904OTUs，CL组和SA组之间共享837OTUs，ML组和MS组之间共享760OTUs，ML组和SA组之间共享794OTUs，SA组和MS组之间共享785OTUs。如图3C等级聚类曲线显示，细菌的相对丰度彼此非常接近。科水平上最大丰度排名前10的物种如图3D。
如图4所示，CL组肠道菌群组成与SA组相似，而MS组与SA组肠道菌群组成不同。与其他3个组相比，ML组的微生物群组成特别集中。如图4BBeta多样性分析结果显示，各组间肠道菌群的组成存在显著差异。
如表1所示，ANOSIM和MRPP进一步证实CL组、ML组、MS组和SA组之间的差异。为了寻找不同的物种，在分类水平(门、纲、目、科、属和种)下进行了t检验。如表2所示，各组间属水平存在差异。
本发明通过建立DSS诱导的急性小鼠结肠炎模型来评估本发明的效果。该动物模型常用于研究溃疡性结肠炎的病理机制和药物疗效评价。通过DSS诱导的结肠炎小鼠模型，我们对小鼠一般体质量特征、DAI评分、结肠长度、结肠组织病理学评分进行检测。以上实验结果表明：苣荬菜水提物，能够有效治疗和缓解溃疡性结肠炎的症状：包括体重降低、腹泻和便血等情况；能够调节肠道菌群组成、促进菌群恢复，调节肠道健康；以及能够进一步保护粘膜屏障和修复结肠损伤。因此，苣荬菜水提物可以作为溃疡性结肠炎的有效治疗药物，制备成各种剂型药物或药物组合物用于溃疡性结肠炎的治疗。
表1CL组、ML组、MS组、SA组肠道微生物菌群ANOSIM和MRPP分析
表2属水平组间物种差异
在本说明书的描述中，术语“一个实施例”、“一些实施例”等的描述意指结合该实施例或示例描述的具体特征、结构、材料或特点包含于本申请的至少一个实施例或示例中。在本说明书中，对上述术语的示意性表述不一定指的是相同的实施例或实例。而且，描述的具体特征、结构、材料或特点可以在任何的一个或多个实施例或示例中以合适的方式结合。
以上仅为本申请的优选实施例而已，并不用于限制本申请，对于本领域的技术人员来说，本申请可以有各种更改和变化。凡在本申请的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本申请的保护范围之内。
Claims (8)
1.一种苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，包括苣荬菜水提物在制备防治溃疡性结肠炎的药物中的应用。
2.根据权利要求1所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述苣荬菜水提物的提取制备方法包括以下步骤:
步骤一：称取苣荬菜干药材并剪碎、清洗，加入蒸馏水浸泡2小时；
步骤二：将浸泡后的混合液煎煮第一次，过滤并倾倒出水煎液，药渣加入蒸馏水煎煮第二次，过滤并倾倒出水煎液，合并两份水煎液；
步骤三：将合并后的水煎液离心，用旋转蒸发仪浓缩，真空冷冻干燥48小时得到所述苣荬菜水提物。
3.根据权利要求2所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述苣荬菜水提物能够减少溃疡性结肠炎症状中的一种以上。
4.根据权利要求3所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述苣荬菜水提物能够改善结肠长度，改善或降低DAI评分。
5.根据权利要求4所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述苣荬菜水提物能够保护结肠粘膜屏障及修复DSS诱导的结肠损伤。
6.根据权利要求5所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述苣荬菜水提物能够调节肠道菌群组成及促进肠道菌群恢复。
7.根据权利要求6所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述药物是以所述苣荬菜提取物为活性成分，单独或与其他药物组方，加入药学上可接受的辅料、辅助性成分或载体制备成药学上可接受的剂型。
8.根据权利要求7所述的苣荬菜水提物在防治溃疡性结肠炎中的应用，其特征在于，所述药物的剂型为散剂、胶囊剂、片剂、丸剂、颗粒剂、口服液体制剂、注射剂或靶向制剂。
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