CN116942750B - 一种治疗阴道炎的药物 - Google Patents
一种治疗阴道炎的药物 Download PDFInfo
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- CN116942750B CN116942750B CN202310962238.1A CN202310962238A CN116942750B CN 116942750 B CN116942750 B CN 116942750B CN 202310962238 A CN202310962238 A CN 202310962238A CN 116942750 B CN116942750 B CN 116942750B
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- repair liquid
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Abstract
本发明公开了一种用于生殖系统的细胞修复液及其制备方法,该细胞修复液的主要成分包括益母草、藏红花、鱼腥草、山药、水杨酸、烟酰胺、壬二酸、罗红霉素、月桂氮酮和透明质酸钠。该修复液能显著改善生殖道的微环境,抑制致病微生物和改善炎症,修复受损的阴道上皮细胞和促进细胞再生,同时还能显著抑制衣原体和支原体感染,使生殖道恢复到正常pH范围;同时,该细胞修复液能显著增加阴道润滑度和湿润度,改善外阴色泽,改善暗沉和色素沉积,本发明能应用于制备女性下生殖系统修复剂、抗妇科炎症药物、以及预防性传播疾病的药物,具有显著的应用效果。
Description
技术领域
本发明涉及医药技术领域,具体涉及一种用于生殖系统的细胞修复液及其制备方法。
背景技术
女性的生殖系统器官包括腺体、卵巢、子宫、宫颈、阴道和外阴,而阴道是体内重要的微生态区,阴道微生态对于女性生殖健康和生理健康具有重要的意义。整个生殖系统生理功能在三十岁左右达到高峰,随着年龄和激素水平的变化,生殖系统功能也开始呈现衰退趋势,
随着生活方式的改变和年龄的增长,大多数女性生殖系统都会发生衰老或者疾病的影响,女性生殖系统衰老不仅体现在生殖器官上,例如阴道萎缩、阴道上皮粘膜变薄抵抗力下降、容易感染细菌或真菌引发炎症、阴道干涩松弛等,还会导致外阴色素沉着,色泽度降低等等,但由于对这种退行性病变不够重视,目前关于各种不适症状还没有针对性的治疗技术和产品。目前最常用的治疗手段是采用雌激素补充治疗,而炎症和病菌入侵又会导致阴道局部免疫、微环境发生变化,而抗菌冲洗液、医用阴道凝胶功能敷料在阴道炎治疗中较为常用,但临床关于上述两者联用或多种症状同时治疗的研究报道较少,且现有临床中常用的甲硝唑凝胶在备孕或妊娠阶段是不适用的,因此,迫切需要开发出临床上关于生殖系统的安全、综合的修复剂或用药。
近年来,中西药结合在多种疾病的治疗中占比越来越重,也得到了显著的应用效果,进一步说明中国传统的中草药的优势,因此,在中西药结合在药物的开发中被应用的愈发广泛。
发明内容
本发明的目的是提供一种用于生殖系统的细胞修复液及其制备方法,该修复液能显著改善生殖道的微环境,抑制致病微生物和改善炎症,修复受损的阴道上皮细胞和促进细胞再生,同时还能显著抑制衣原体和支原体感染,使生殖道恢复到正常pH范围;同时,该细胞修复液能显著增加阴道润滑度和湿润度,改善外阴色泽,改善暗沉和色素沉积,本发明能应用于制备女性下生殖系统修复剂、抗妇科炎症药物、以及预防性传播疾病的药物,具有显著的应用效果。
为了达到上述目的,本发明提供了一种用于生殖系统的细胞修复液,该细胞修复液的主要成分包括益母草、藏红花、鱼腥草、山药、水杨酸、烟酰胺、壬二酸、罗红霉素、月桂氮酮、透明质酸钠和pH调节剂。该细胞修复液的pH为3.8-4.5。
本发明还提供了一种上述细胞修复液的制备方法,包含如下步骤:
1)分别称取重量比为25-35%的益母草、30-40%的藏红花、25-35%的鱼腥草和10-15%的山药共4种中药材,加入所有中药材10倍重量的水,沸水煮30min,过滤后再加入10倍重量的水,沸水煮30min,过滤,合并两次滤液并烘干至稠膏;
2)取上述稠膏质量分数为2%的水杨酸、3.5%的烟酰胺、2%的壬二酸、1%的月桂氮酮、1%的透明质酸钠和1.5%的罗红霉素,加入稠膏中搅拌均匀,并再次调节含水量与步骤1)保持相同;
3)通过pH调节剂调节稠膏的pH至3.8-4.5,即得细胞修复液。
进一步地,上述步骤1)中稠膏的含水量为40%。
进一步地,上述pH调节剂为枸橼酸、苹果酸、酒石酸、乳酸、富马酸中的一种或多种。
本发明还提供了一种由上述细胞修复液制备的生殖系统用药。
本发明提供的细胞修复液可应用于女性生殖系统中。
进一步地,上述的应用包括阴道炎的治疗、外阴色泽的改善、阴道的干涩和润滑度的改善等。
本发明的细胞修复液,解决了阴道微生物感染,抑制了阴道炎症和促进细胞再生等,具有以下优点:
本发明提供的细胞修复液多采用天然药草,副作用小,可同时适用于备孕或妊娠期的女性,适用性更广,具有显著的经济价值。
本发明提供的细胞修复液能抑制多种病原微生物,且对衣原体和支原体感染也具有显著的抑制效果,液相比现有的女性生殖系统用药具有更广、更优的治疗效果,应用价值显著。
本发明提供的细胞修复液能显著增加阴道润滑度和湿润度,改善外阴色泽,改善暗沉和色素沉积,在女性的特殊护理领域具有显著的应用效果,为女性的保养和功能性保健具有一定的参考意义。
附图说明
图1为本发明中的细胞修复液对阴道上皮细胞的处理中其炎症和凋亡靶标分子的变化结果。
图2为本发明中的细胞修复液对阴道上皮细胞的促生长增殖测定结果。
具体实施方式
下面将对本发明实施例中的技术方案进行清楚、完整地描述,显然,所描述的实施例仅仅是本发明一部分实施例,而不是全部的实施例。基于本发明中的实施例,本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例,都属于本发明保护的范围。
部分涉及的培养基:
真菌平板培养基:麦芽糖40g、蛋白胨10g、琼脂20g、蒸馏水1L;
细菌平板培养基:蛋白胨10g、酵母粉5g、NaCl10g、琼脂粉20g、蒸馏水1L。
市售的DMEM培养基、胎牛血清FBS、青链霉素、0.25%的胰酶。
说明:本发明涉及的微生物均来自中国微生物菌种保藏管理中心,本发明未具体说明的方法均为本领域的常规方法,未具体说明的试剂均为本领域常规试剂。
一种细胞修复液的制备:
制备对比例1
1)制备稠膏:分别称取重量比为30%的益母草、30%的藏红花、30%的鱼腥草和10%的山药中药材,加入中药材的10倍重量的水,沸水煮30min,过滤后再加入10倍重量的水,沸水煮30min,过滤,合并两次滤液并烘干至稠膏含水量约40%;
2)取稠膏质量分数为2%的水杨酸、3.5%的烟酰胺、2%的壬二酸、1%的月桂氮酮、1%的透明质酸钠和1.5%的罗红霉素,加入稠膏中搅拌均匀,并再次调节含水量约40%;
3)通过pH调节剂-枸橼酸调节稠膏的pH至3.8-4.5,即得半流体的细胞修复液1。
制备对比例2
1)制备稠膏:分别称取重量比为25%的益母草、35%的藏红花、30%的鱼腥草和10%的山药中药材,加入中药材的10倍重量的水,沸水煮30min,过滤后再加入10倍重量的水,沸水煮30min,过滤,合并两次滤液并烘干至稠膏含水量约40%;
2)取稠膏质量分数为2%的水杨酸、3.5%的烟酰胺、2%的壬二酸、1%的月桂氮酮、1%的透明质酸钠和1.5%的罗红霉素,加入稠膏中搅拌均匀,并再次调节含水量约40%;
3)通过pH调节剂-枸橼酸调节稠膏的pH至3.8-4.5,即得半流体的细胞修复液2。
制备对比例3
1)制备稠膏:分别称取重量比为30%的益母草、25%的藏红花、35%的鱼腥草和10%的山药中药材,加入中药材的10倍重量的水,沸水煮30min,过滤后再加入10倍重量的水,沸水煮30min,过滤,合并两次滤液并烘干至稠膏含水量约40%;
2)取稠膏质量分数为2%的水杨酸、3.5%的烟酰胺、2%的壬二酸、1%的月桂氮酮、1%的透明质酸钠和1.5%的罗红霉素,加入稠膏中搅拌均匀,并再次调节含水量约40%;
3)通过pH调节剂-苹果酸调节稠膏的pH至3.8-4.5,即得半流体的细胞修复液3。
实验例 细胞修复液的效果测定
在制备细胞修复液的同时,还制备了不含上述稠膏的细胞修复液4,具体为取质量分数为2%的水杨酸、3.5%的烟酰胺、2%的壬二酸、1%的月桂氮酮、1%的透明质酸钠和1.5%的罗红霉素配置成水溶液,经枸橼酸调节水溶液的pH至3.8-4.5,即为细胞修复液4。
一、抑菌效果测定
抑菌圈测定法:取制备的真菌平板和细菌平板,其中,在真菌平板上接种假丝酵母菌,在细菌平板上接种加德纳菌,每种平板各5个重复。待平板长满后,每隔平板上均匀打5个孔,5个孔分别接种甲硝唑凝胶(阳性对照)、细胞修复液1、细胞修复液2、细胞修复液3、细胞修复液4。进一步培养并观察平板情况,在5-8日进行抑菌圈直径测定,统计结果如下表1所示:
表1 抑菌效果测定统计
由表1可知,上述方法制备的细胞修复液,其对真菌和细菌的抑制效果显著,且优于甲硝唑凝胶的抑制效果。
对上述制备的细胞修复液进一步测定其抑菌效果,得知,该细胞修复液不仅可以抑制假丝酵母菌(CCTCC AB 204047)、霉菌(CPCC 200181)、念珠菌(ATCC 10231)等真菌,还可以抑制加德纳菌(ATCC14018)、沙门氏菌(CMCC 50094)、葡萄球菌(ATCC 6538)等细菌,具有广谱的抑菌效果,但该细胞修复液对乳杆菌抑制不明显,能保护生殖道有益菌的同时,显著抑制致病微生物,抑菌效果显著。
二、对生殖道上皮细胞的效果测定
本实验选择人阴道上皮细胞细胞系VK2/E6E7进行验证,培养VK2/E6E7细胞至传代状态良好,分别接种于培养皿中进行培养,待加药进行效果测定,根据实验例1的‘一’得到的结果选择细胞修复液3进一步验证其他效果。
1、抗氧化测定
取对数生长的VK2/E6E7细胞,消化后接种于6孔板中,共接种3个重复的6孔板(测定三次取3次结果的平均数记为测定结果),在细胞生长至约70%时随机选择5个孔分别接种DMEM-FBS、叔丁基过氧化氢(tBHP)、tBHP+细胞修复液3、tBHP+细胞修复液4、tBHP+甲硝唑凝胶,并进行标记。
采用MDA检测试剂盒(碧云天,S0131M),参考使用说明制作标准曲线后,再进行各孔细胞的氧化应激水平检测,检测结果如下表2所示:
表2 抗氧化效果测定
由表2可知,在加入了tBHP后,细胞的氧化应激水平显著升高,而不加中草药提取稠膏的细胞修复液4可以降低氧化应激水平,但其效果逊色于常规用药的甲硝唑凝胶,而在添加有中草药提取稠膏的细胞修复液3的作用下,由tBHP造成高氧化应激水平显著降低,且显著优于常规用药的甲硝唑凝胶,可知,通过本发明制备的上述细胞修复液具有显著的抗氧化效果。
2、抗炎、抗凋亡测定
取对数生长的VK2/E6E7细胞,消化后接种于6孔板中,共接种3个重复的6孔板,在细胞生长至约70%时随机选择5个孔分别接种DMEM-FBS、叔丁基过氧化氢(tBHP)、tBHP+细胞修复液4、tBHP+细胞修复液3、tBHP+甲硝唑凝胶,并进行标记。在加入药物的第48h提取各孔细胞的全蛋白,定量后进行常规的WB检测,检测指标为IL-1β、和Caspase3,以GAPDH为内参进行WB检测,对目标分子进行ECL发光成像得到WB结果如图1所示,其中,泳道M为DNAmarker,泳道1-5分别为细胞中接种了DMEM-FBS、叔丁基过氧化氢(tBHP)、tBHP+细胞修复液4、tBHP+细胞修复液3、tBHP+甲硝唑凝胶,可知,该细胞修复液能显著降低炎症因子的产生,同时能降低凋亡蛋白的表达,表明,该细胞修复液具有显著的抗炎和抗凋亡效果。
3、促生长效果测定
取对数生长的VK2/E6E7细胞,消化后接种于96孔板中,每孔2*103 cell/100 μL,共设置3个重复孔,在细胞贴壁且生长至约70%时分别加入5μL的细胞修复液、5μL的甲硝唑凝胶,再次置于培养箱进行培养,分别在0、3、6、12、24、48h加入CCK8,孵育后进行A450nm处的吸光度,测定结果见图2所示,可知该细胞修复液相比甲硝唑凝胶对VK2/E6E7细胞具有更优的促细胞生长效果。
综上可知,本发明提供的制备方制备的细胞修复液具有显著的抑菌、抗氧化、抗炎、抗凋亡和促细胞生长的效果,对生殖系统的药物开发和临床治疗具有指导意义和潜在的应用价值。
三、细胞修复剂在临床上的应用验证
收集患有女性下生殖系统相关疾病的患者9例,具体年龄和症状如下:
年龄:25-35岁的3例、35-45岁的5例、45-55岁的1例。
经多项检查,具体包含:妇科常规查体、微生物培养检查、支原体与衣原体的检测、白带检查、药物敏感试验、胺实验、阴道镜检查等。
综合症状如下:9例患者均患有不同程度的阴道炎症状,其中有6例患有细菌和霉菌感染,2例滴虫感染且1例伴有支原体感染,5例患有不同程度的宫颈糜烂;同时,有8例患者出现不同程度的外阴颜色暗沉,6例出现阴道干涩和松弛等。
用药治疗:先用温水清洗外阴,将上述制备的半流体状的细胞修复液3摇匀,带上无菌的一次性PE手套,将其均匀涂抹于阴道内壁和外阴,涂膜厚度为3mm左右,每天睡前用药一次,连续用药一个月,并每7天去医院复查。
治疗结果:在治疗过程中均未出现皮肤刺激、过敏等不良反应,经过4周治疗后,临床症状几乎得到有效缓解,阴道炎症状消失,阴道pH全部恢复至3.8-4.4之间。其中,在用药7天后,经检查微生物感染得到显著改善,致病菌(包括细菌和霉菌)数量均得到显著的控制,滴虫感染也得以恢复正常,并有效清除了支原体;在用药14天后检查,其中有7例阴道炎症状几乎消失,有2例宫颈糜烂患者的症状显著改善;在用药21天后的,所有患者的阴道炎症状全部消失,2例宫颈糜烂患者的症状也恢复正常,且有2例患者的外阴颜色得到改善,4例患者的阴道干涩症状得到改善;在用药4周后,所有患者的阴道炎症状全部消失,阴道的pH也恢复至正常生理范围,有3例患者停用药物,并持续观察连续2个月以上未出现症状反复,表明该细胞修复液作为一种妇科用药在阴道炎症状的治疗上具有显著的疗效。其余6例患者在用药4周后,持续用药2个月,在持续涂抹一个月后,有3例患者的外阴颜色出现明显好转,由开始的色泽暗沉、皮肤干皱转为皮肤平滑、细胞状态好转,并出现组织弹性,阴道干涩显著改善,阴道内壁按压恢复部分弹性;在持续用药2个月后,有5例患者外阴颜色出现明显好转,外阴色泽由暗沉变淡并微泛红,且阴道干涩显著改善,阴道润滑度显著增加,阴道内壁恢复弹性,表明该细胞修复液能修复阴道内壁受损的细胞,促进细胞再生,使阴道润滑度明显改善并恢复阴道组织的弹性;同时还能改善外阴的色素沉积,显著改善外阴皮肤干皱等,对女性的私处保养和改善效果显著。
综上可知,该修复液能显著改善生殖道的微环境,抑制致病微生物和改善炎症,修复受损的阴道上皮细胞和促进细胞再生,同时还能显著抑制衣原体和支原体感染,使生殖道恢复到正常pH范围;同时,该细胞修复液能显著增加阴道润滑度和湿润度,改善外阴色泽,改善暗沉和色素沉积,本发明能应用于制备女性下生殖系统修复剂、抗妇科炎症药物、以及预防性传播疾病的药物,具有显著的应用效果和经济价值。
尽管本发明的内容已经通过上述优选实施例作了详细介绍,但应当认识到上述的描述不应被认为是对本发明的限制。在本领域技术人员阅读了上述内容后,对于本发明的多种修改和替代都将是显而易见的。因此,本发明的保护范围应由所附的权利要求来限定。
Claims (3)
1.一种治疗阴道炎的药物,其特征在于,该药物包括益母草、藏红花、鱼腥草、山药、水杨酸、烟酰胺、壬二酸、罗红霉素、月桂氮酮、透明质酸钠和pH调节剂,
其中,所述药物的pH为3.8-4.5,
所述药物的制备方法步骤如下:
1)分别称取重量比为25-35%的益母草、30-40%的藏红花、25-35%的鱼腥草和10-15%的山药共4种中药材,加入所有中药材10倍重量的水,沸水煮30min,过滤后再加入10倍重量的水,沸水煮30min,过滤,合并两次滤液并烘干至稠膏;
2)取所述稠膏质量分数为2%的水杨酸、3.5%的烟酰胺、2%的壬二酸、1%的月桂氮酮、1%的透明质酸钠和1.5%的罗红霉素,加入稠膏中搅拌均匀,并再次调节含水量与步骤1)保持相同;
3)通过pH调节剂调节稠膏的pH至3.8-4.5,即得。
2.根据权利要求1所述的治疗阴道炎的药物,其特征在于,所述步骤1)中稠膏的含水量为40%。
3.根据权利要求1所述的治疗阴道炎的药物,其特征在于,所述pH调节剂为枸橼酸、苹果酸、酒石酸、乳酸、富马酸中的一种或多种。
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