CN1895305A - 猫人参挥发油在制药中的应用 - Google Patents
猫人参挥发油在制药中的应用 Download PDFInfo
- Publication number
- CN1895305A CN1895305A CN 200610052004 CN200610052004A CN1895305A CN 1895305 A CN1895305 A CN 1895305A CN 200610052004 CN200610052004 CN 200610052004 CN 200610052004 A CN200610052004 A CN 200610052004A CN 1895305 A CN1895305 A CN 1895305A
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- CN
- China
- Prior art keywords
- volatile oil
- radix actinidiae
- actinidiae valvatae
- valvatae
- oil
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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Abstract
本发明涉及猫人参挥发油在制药中的用途。以猫人参新鲜叶为原料,采用通常的提取方法提取挥发油。挥发油至少包含:芳樟醇、1,2-二甲基-2,3-二氢吲哚、亚麻酸甲酯、叶绿醇、亚麻酸、2-己烯醛、α-松油醇、香叶醇和棕榈酸等。药理研究结果表明,猫人参挥发油具有良好的抗菌抑菌作用,尤其是对真菌的抑制;且毒副作用小。因此其可用于制成各种抗菌药剂,如注射剂、胶囊剂、滴丸剂、片剂、颗粒剂、散剂、口服液或其它剂型。
Description
技术领域
本发明涉及猫人参挥发油的用途,尤其是在制药中的用途。
背景技术
随着人类生活水平的提高,抗菌素、化学药物的大量应用,导致某些菌类产生抗药性,这对人类生产生活造成了严重威胁。世界各国医学专家们不断致力于寻找毒副作用小、菌类不易产生抗药性的防治药剂。我国中草药资源丰富,高效低毒,成本低廉。广大的中药工作者,对中草药防菌抑菌的活性作了大量有益的尝试,积累了不少经验。
猫人参为猕猴桃科猕猴桃属植物大籽猕猴桃Actinidia macrospermaC.F.Liang或对萼猕猴桃Actinidia.valvata Dunn。前者在中药中俗称“红货”,后者俗称“白货”。主要分布于我国华东及华南地区,集中在广东、湖南、湖北、江西、浙江、江苏、安徽等省。根据中药大词典记载:该药味苦、性寒、归肺、胃经;根茎入药;具有清热解毒、祛风除湿、消肿去疖的功效;临床用于治疗皮肤疮疡脓肿、麻风、骨髓炎、风湿痹痛、白带异常、肝硬化黄疸腹水等疾病,民间还用于肺癌和消化道肿瘤的治疗。猫人参在抑制肿瘤生长,改善生命征象方面有独特的效果,经过反复实践,逐渐形成了以猫人参为主的各种治疗肿瘤的验方,引起中医药界的关注。由于猫人参在抗肿瘤提高免疫力方面的良好疗效,以至于其它药理作用被大大忽视了,药典及各种本草中均无有关于猫人参挥发油药用的记载。到目前为止,还未见关于该中药挥发油化学成分、药理作用及其应用的文献报道。对猫人参的应用,也仅仅限于将其整体加工后,作为复方药的原料之一与其他的原料共同制成中成药,尚未发现以猫人参为原料特别是作为单一原料进行加工并/或将加工出的提取物直接作为药物使用的制造方法及其制成品。
引证文献:
①.裘宝林等。浙江植物志(第四卷)。浙江:浙江科学技术出版社,1993:177。
②.江苏新医学院。中药大词典(下)。上海:上海科学技术出版社,1977:2205。
③.姚淦,王铁僧。华东地区猕猴桃属药用植物的整理。中药材,1989,12(2):15。
④.浙江省卫生厅。浙江省中药炮制规范。杭州:浙江科学技术出版社,1994:89。
⑤.金瑞芝,王国强。猫人参治疗臌胀例释。浙江中医杂志,1998,33(1):35。
⑥。倪勤武,诸葛陇。猫人参及其混淆品的鉴别研究。浙江中医学院学报,1999,23(5):60。
⑦.周兴兆.唐福安论肺癌证治。浙江中医学院学报,2000,24(2):45。
⑧.来平凡,章红燕。浙江地区习用中药猫人参研究进展。浙江中医学院学报,2002,26(1):77~78。
发明内容
本发明的目的在于提供猫人参的活性成分挥发油的新用途,即在制药中的用途。
本发明特点是,猫人参挥发油在制备抗菌抑菌药物中的应用。
本发明具体涉及猫人参活性成分挥发油在制备抗菌抑菌单方或复方药物中的新应用。
本发明以猫人参挥发油为原料,可将其作为活性成分与制药学上可接受的载体或药用辅料开发成相宜的抗菌抑菌药物制剂,如注射剂、胶囊剂(软胶囊、硬胶囊)、滴丸剂、片剂、颗粒剂、散剂、口服液等。
为了更好地理解本发明的实质,下面用猫人参挥发油的体外抑菌活性研究和急性毒性实验的结果来说明其在制备抗菌抑菌药物中的用途。
一、体外抑菌实验
1.实验材料与方法
1.1菌株 实验菌株包括两种革兰氏阳性菌:金色葡萄球菌(Staphylococcus aureus ATCC 25923)和枯草杆菌(Bacillus subtilis ATCC26633);两种革兰氏阴性菌:大肠杆菌(Escherichia coli ATCC 25922)和绿脓杆菌(Pseudomonas aeruginosa ATCC 27853);三种真菌:酵母菌(Candida albicans ATCC 10231),烟曲霉(Aspergillus fumigatus ATCC 9142)和犬小孢子菌(Microsporum canis ATCC 36299)。各菌种的临床病症表现见表1。
表1各菌种的临床病症表现
菌种 | 临床表现 | |
革兰氏阳性菌 | 金色葡萄球菌 | 食物中毒、皮肤局部或化脓性感染、败血病、肺炎、脑膜炎,等 |
枯草杆菌 | 食物中毒,等 | |
革兰氏阴性菌 | 大肠杆菌 | 痢疾、肠炎,等 |
绿脓杆菌 | 伤口感染、败血病、尿道感染,等 | |
真菌 | 酵母菌 | 肺炎,阴道、耳、骨等感染,鹅口疮,等 |
烟曲霉 | 过敏性哮喘、膀胱纤维症的并发感染、肺结核,等 | |
犬小孢子菌 | 动物的毛发皮癣,等 |
1.2实验方法 采用纸片琼脂扩散法对体外抑菌活性进行初测。采用微量稀释法测定最小抑制浓度(minimum inhibitory concentration,MIC)和最小杀菌浓度(minimum bactericidal or fungicidal concentration,MBC/MFC)。
2.实验结果
结果见表2、表3。
猫人参挥发油对革兰氏阳性菌有一定抑制作用,对革兰氏阴性菌有较强抑制作用(除绿脓杆菌这一普遍抗药菌株外),最小抑制浓度MIC为0.78~25.50μl/ml,最小杀菌浓度1.56~50.0μl/ml;在真菌的测试中,猫人参挥发油表现出更强的抑制活性,最小抑制浓度MIC为0.78~1.56μl/ml,最小杀菌浓度0.78~3.12μl/ml。
表2猫人参挥发油的体外抑菌活性初测(纸片法)a,b
菌种 | 挥发油抑菌圈直径(mm) | 抗生素抑菌圈直径(mm) | |||
10μl/disc | 20μl/disc | 克霉唑Clotrimazole(10μg/ml) | 氯霉素Chloramphenicol(30μg/ml) | ||
革兰氏阳性菌 | 金色葡萄球菌 | 7 | 12 | nd | 19 |
枯草杆菌 | 9 | 13 | nd | 21 | |
革兰氏阴性菌 | 大肠杆菌 | 18 | 21 | nd | 24 |
绿脓杆菌 | - | - | nd | 10 | |
真菌 | 酵母菌 | 19 | 25 | 12 | nd |
烟曲霉 | 16 | 27 | 9 | nd | |
犬小孢子菌 | 15 | 19 | 6 | nd |
a抑菌圈直径7~13mm表示微弱活性;抑菌圈直径≥14mm表示较强活性
b-表示没有活性;nd表示没有检测。
表3猫人参挥发油的最小抑制浓度(MIC)和最小杀菌浓度(MBC/MFC)
(微量稀释法)
菌种 | 挥发油 | 抗生素 | |||
MIC(μl/ml) | MBC/MFC(μl/ml) | MIC(μg/ml) | MBC/MFC(μg/ml) | ||
左氧氟沙星Levofloxacin | |||||
革兰氏阳性菌 | 金色葡萄球菌 | 3.12 | 3.12 | 0.12 | 4.00 |
枯草杆菌 | 3.12 | 6.25 | 0.50 | 2.00 | |
革兰氏阴性菌 | 大肠杆菌 | 0.78 | 1.56 | 0.02 | 0.62 |
绿脓杆菌 | 25.50 | 50.00 | 0.05 | 8.00 | |
两性霉素Amphotericin | |||||
真菌 | 酵母菌 | 0.78 | 0.78 | 0.25 | 2.00 |
烟曲霉 | 0.78 | 1.56 | 0.38 | 2.00 | |
犬小孢子菌 | 1.56 | 3.12 | 0.04 | 5.00 |
根据对猫人参挥发油化学成分的分析和相关药理文献的报道,我们发现:猫人参挥发油中的芳樟醇具有明显抗菌活性,并有镇痛、解痉、抗白血病的作用;2-己烯醛能作为伤口消毒剂,具抗菌抑菌活性;另外,叶绿醇、α-松油醇、香叶醇和棕榈酸等化合物也具有一定的抑菌活性。
二、急性毒性实验
1.实验材料与方法
1.1动物昆明种小白鼠18~22g,雌雄各半,检疫后备用。
1.2实验方法 小鼠按体重均衡分为5个剂量组,每组20只。实验前禁食不禁水12小时,根据预实验结果,各组剂量分别为2.00ml/kg、2.56ml/kg、3.20ml/kg、4.00ml/kg、5.00ml/kg,将猫人参挥发油用色拉油稀释给小鼠灌胃,给药后连续观察4h、24h、48h、72h各观察一次。72h记录小白鼠毒性反应及死亡数,用改良寇氏法计算小白鼠的半致死剂量LD50及95%的置信区间。
2.实验结果
结果如表4。经测定,LD50值为3.27ml/kg,95%可信限为3.04~3.54ml/kg(P>0.05)。根据测定,1ml挥发油重为1.17259g,因此LD50值为3834mg/kg,95%可信限为3565~4150mg/kg;按急性毒性分级标准判定,该样品属实际无毒类。
表4猫人参挥发油对小鼠急性毒性试验死亡率的影响
给药剂量(ml/kg) | 小鼠总数 | 72h死亡数 | 死亡率(%) |
5.00 | 20 | 20 | 100 |
4.00 | 20 | 15 | 75 |
3.20 | 20 | 10 | 50 |
2.56 | 20 | 3 | 15 |
2.00 | 20 | 0 | 0 |
综上所述,猫人参挥发油在体外抑菌尤其是抑制真菌方面有良好疗效,且毒副作用小,有着良好的开发应用前景。
本发明的猫人参挥发油所用的原料为猫人参新鲜叶子,而非根茎,这将一定程度上缓解猫人参生药材供应的不足的现象,提高猫人参的有效利用率。本发明所述的方法工艺简单,符合中国药典的要求,无须特殊设备。经过药理研究,发现其具有明显的抑菌活性,尤其是对真菌的抑制作用(经实验证实,猫人参挥发油细菌最小抑制浓度MIC为0.78~25.50μl/ml,真菌最小抑制浓度MIC为0.78~1.56μl/ml)。且猫人参挥发油具有毒副作用小的优点,有着良好的开发应用前景,可以与其他中草药成分配伍,或单独直接作为药物,进一步开发成有效的抑菌新药,使得猫人参的药用成分得以充分利用,从而填补了我国中草药利用的一项空白,具有实用价值。
本发明中的猫人参挥发油采用通常的水蒸气蒸馏法、溶剂提取法或超临界萃取法等适合于工业化生产的方法进行制备。得到的猫人参挥发油呈淡黄色,主要含有芳樟醇(Linalool)、1,2-二甲基-2,3-二氢吲哚(1,2-dimethyl,Lindoline)、亚麻酸甲酯(Linolenic acid,methylester)、叶绿醇(Phytol)、亚麻酸(α-Linolenic acid)、2-己烯醛(2-Hexenal)、α-松油醇(α-Terpineol)、香叶醇((E)-3,7-dimethyl--2,6-Octadien-1-ol)和棕榈酸(n-Hexadecanoic acid)等化合物。含油量为0.10~0.25%,即每100g生药含挥发油0.1~0.25g。
上述的水蒸气蒸馏法通过油水分离器分离挥发油,其步骤为:按重量份计算,称取猫人参叶0.1~20份,加水0.5~200份,水蒸气蒸馏3~5小时,温度控制在80~105℃(蒸馏前期温度可设置得高些,后期只要保持小沸即可),进行油水分离;用油水分离器分离得含水猫人参挥发油;乙醚萃取后,冷冻干燥(-20℃下冷冻干燥24小时以上)或无水硫酸钠干燥(24小时以上)、过滤,去除水相,浓缩得到猫人参挥发油。
上述的溶剂提取法,其步骤为:取猫人参叶0.1~20份,加入有机溶剂正己烷或环己烷0.6~120份重量份,进行回流提取,温度控制在60~70℃,回流2~3小时,待回流液温度降至常温时,放出回流液,回收有机溶剂,即可得到挥发油粗品;加入无水乙醇0.01~0.05重量份,与挥发油粗品充分混合,放置-20℃冰柜中放置24小时以上,将浑浊液过滤除蜡;继续放置冰柜中重复除蜡过程,直到液体澄清;将含有挥发油的乙醇液浓缩,即得猫人参挥发油。
上述的超临界萃取法,其步骤为:取猫人参叶加入萃取釜,萃取釜压力10~50Mpa,温度20~70℃,进行超临界CO2萃取,萃取时间为0.5~15小时,从萃取釜出来的CO2进入分离系统,一级分离温度30~50℃,压力10~15Mpa;二级分离温度20~40℃,压力4~7Mpa,得到猫人参挥发油,CO2流速为15~25升/h,分离出来的CO2再经过滤和制冷系统加压后循环利用。
具体实施方式
下述实例用以进一步说明本发明,但并不由此限制本发明范围。
实施例1:
取猫人参鲜叶100g,加10倍量水,水蒸气蒸馏5小时;用油水分离器分离含水挥发油;乙醚少量多次萃取后,用无水硫酸钠干燥过夜,过滤浓缩得到猫人参挥发油。挥发油呈浅黄色透明液样,有特殊清香味,得油率为0.10%,即100g猫人参鲜含挥发油0.10g。
挥发油加少量无水乙醇溶解后进行GC-MS分析,结果如表5。色谱条件:石英毛细管柱HP-5MS(30m×0.25mm×0.25μm),载气为氦气,柱流量1ml/min。汽化室温度:260℃,升温程序为从50℃开始以10℃/min升温至260℃;质谱条件:EI电离源,电离能量70eV,离子源温度230℃,扫描范围:30~500amu,进样量1.0μL,分流比10∶1。
表5猫人参挥发油成分分析结果(水蒸汽蒸馏法)
化合物名称 | 相对含量(%) | 化合物名称 | 相对含量(%) |
2-Hexenal 2-己烯醛 | 0.75 | 含N杂环化合物 | 14.38 |
β-Cyclocitralβ-环柠檬醛 | 0.34 | 6,10,14-trimethyl,2-Pentadecanone6,10,14-三甲基-2-十五酮 | 0.16 |
Nonanal壬醛 | 0.43 | Hexadecanoic acid,methylester棕榈酸甲酯 | 0.52 |
β-Linalool β-芳樟醇 | 48.14 | Hexadecanoic acid棕榈酸 | 1.32 |
Decanal癸醛 | 0.12 | Linoleic acid,methylester亚油酸甲酯 | 0.67 |
α-Terpinenol α-萜品醇 | 0.96 | Linolenic acid,methylester亚麻酸甲酯 | 6.57 |
3,7-dimethyl-2-Octen-1-ol香茅醇 | 0.73 | Phytol植醇 | 5.29 |
3,7-dimethyl-2,6-Octadien-1-ol香叶醇 | 1.59 | Linolenic acid,ethylester亚麻酸乙酯 | 0.29 |
1,2-dimethyl,Lindoline1,2-二甲基-2,3-二氢吲哚 | 7.94 | Eicosane正二十烷 | 0.34 |
4-ethenyl-2-methoxy,Phenol4-乙烯基-2-甲氧基苯酚 | 0.61 | α-Linolenic acidα-亚麻酸 | 0.48 |
β-Lonene 4-(2,6,6-三甲基-1-环己烯)-3-丁烯基-2-酮 | 0.16 | Heneicosane正二十一烷 | 0.82 |
Dihydronepetalactone二氢假荆芥内酯 | 0.79 | Docosane正二十二烷 | 0.59 |
Dihydroactinidiolide二氢猕猴桃内酯 | 0.76 | Tetracosanoid acid,methylester二十四酸甲酯 | 0.08 |
实施例2:
取猫人参新鲜叶100g,加入正己烷600ml,回流2小时;含油正己烷减压浓缩,加少量无水乙醇10ml热溶,放置-20℃冰柜中放置24小时低温除蜡,过滤;除蜡步骤重复3次,直到油液澄清;过滤残留物用无水乙醇重新微热溶,低温放置过滤,最后合并滤液;浓缩含油乙醇液,过滤浓缩得到猫人参挥发油。挥发油呈浅黄色透明液样,有特殊清香味,得油率为0.20%,即100g猫人参鲜叶含挥发油0.20g。
挥发油加少量无水乙醇溶解后进行GC-MS分析,结果如表6。色谱条件:石英毛细管柱HP-5MS(30m×0.25mm×0.25μm),载气为氦气,柱流量1mL/min。汽化室温度:260℃,升温程序为从50℃开始以10℃/min升温至260℃;质谱条件:EI电离源,电离能量70eV,离子源温度230℃,扫描范围:30~500amu,进样量1.0μL,分流比10∶1。
表6猫人参挥发油成分分析结果(溶剂提取法)
化合物名称 | 相对含量(%) | 化合物名称 | 相对含量(%) |
2-Hexenal2-己烯醛 | 0.01 | Dodeca-1,6-dien-12-ol,6,10-dimethyl,6,10-二甲基-1,6-十二二烯-12-醇(异) | 1.51 |
3-Hexen-1-ol 3-己烯醇 | 0.26 | Tetradecane正十四烷 | 0.16 |
2-Hexen-1-ol 2-己烯醇 | 0.06 | Dihydronepetalactone二氢假荆芥内酯 | 1.97 |
1-Hexene 1-己烯 | 0.02 | Iridomyrmecin阿根廷蚁素 | 0.40 |
3-Hexen-1-ol,acetate乙酸3-己烯酯 | 0.03 | Dihydroactinidiolide二氢猕猴桃内酯 | 0.18 |
3-methyl,3-Heptanol3-甲基-3-庚醇 | 0.05 | Hexadecane正十五烷 | 0.08 |
Benzyl Alcohol苯甲醇 | 0.31 | 3,7,11,15-Tetramethyl-2-Hexadecen-1-ol3,7,11,15-四甲基-2-十六烯醇 | 0.24 |
β-Linalool β-芳樟醇 | 0.93 | 2-Pentadecanone,6,10,14-trimethyl6,10,14-三甲基-2-十五酮 | 0.04 |
3-Decyn-2-ol 2-羟基-3-癸炔 | 0.02 | n-Hexadecanoic acid棕榈酸 | 3.46 |
Phenylethyl Alcohol苯乙醇 | 0.04 | Linolenic acid,methylester亚麻酸甲酯 | 0.31 |
Dodecane正十二烷 | 0.24 | Phytol植醇 | 1.64 |
cis-α-Terpineol α-松油醇 | 0.18 | α-Linolenic acid α-亚麻酸 | 60.97 |
(R)-,3,7-dimethyl-,6-Octen-1-ol3,7-二甲基-6-辛烯醇(香茅醇) | 0.13 | Spinacen角鲨烯 | 6.04 |
2,6-Octadien-1-ol,3,7-dimethyl-,3,7-二甲基-2,6-辛二烯醇(香叶醇) | 0.52 | Vitamin E维生素E | 2.05 |
1,2-dimethyl,Lindoline1,2-二甲基-2,3-二氢吲哚 | 0.23 | τ-Sitosterolτ-谷甾醇 | 1.40 |
Dodeca-1,6-dien-12-ol,6,10-dimethyl,6,10-二甲基-1,6-十二二烯-12-醇 | 6.30 | α-Tocopherol维生素E | 1.78 |
3-Hexadecyne 3-十六炔 | 1.41 | β-Amyrin β-香树脂醇 | 0.84 |
实例3:
取猫人参叶加350g投入萃取釜,萃取釜压力20Mpa,温度30℃;一级分离温度30℃,压力10Mpa;二级分离温度20℃,压力4Mpa。超临界流体静态浸泡原料1小时,设备稳定后进行动态萃取,由接收器收集产品。最后得猫人参挥发油0.87g,挥发油呈浅黄色透明液样,有特殊清香味,得油率为0.25%,即100g猫人参鲜叶含挥发油0.25g。
挥发油加少量无水乙醇溶解后进行GC-MS分析,结果类似于实例1、2,发现至少含有芳樟醇、1,2-二甲基-2,3-二氢吲哚、亚麻酸甲酯、叶绿醇、亚麻酸、2-己烯醛、α-松油醇、香叶醇和棕榈酸等成分。
本发明涉及的部分参考文献:
(1)NCCLS-National Committee for Clinical LaboratoryStandards,2001.Performancestandards for antimicrobial susceptibility testing:eleventh informational supplement.Document M 100-S 11.National Committee for Clinical Laboratory Standard,Wayne,PA,USA.
(2)Peana,A.T.,D’Aquila,P.S.,Chessa,M.L.,Moretti,M.D.,Serra,G.,Pippia,P.,2003.(-)-Linalool produces antinociception in two experimental models of pain.EuropeanJournal of Pharmacology 460,37-41.
(3)Chiang,L.C.,Chiang,W.,Chang,M.Y.,Ng,L.T.,Lin,C.C.,2003.Antileukemicactivity of selected natural products in Taiwan.American Journal of Chinese Medicine31.37-46.
(4)Chiang,LC.,Ng,L.T.,Chiang,W.,Chang,M.Y.,Lin,C.C.,2003.Immunomodulatory activities of lavonoids,monoterpenoids,triterpenoids,iridoidglycosides and phenolic compounds of Plantago species.Planta Medica 69,600-604.
(5)Lis-Balchin,M.,Hart,S.,1999.Studies on the mode of action of the essential oil oflavender(Lavandula angustifolia P.Miller).Phytotherapy Research 13,54-542.
(6)Schutz,S.,Weissbecker,B.,Klein,A.,Hummel,H.E.,1997.Host plant selection ofthe Colorado potato beetle as influenced by damage induced volatiles of the potato plant.Natur-wissenschaften 84,212-217.
(7)Findlay,J.A.,Patil,A.D.,1984.Antibacterial constituents of the diatom Naviculadelognei.J Nat Prod 47(5),815-818.
(8)Christina,K.,Martin,K.,Alison,R.,2005.Chemical composition and antibacterialactivity of the essential oil and the Gum of Pistacialentiscus Var.chia.J.Agric.FoodChem.53(20),7681-7685.
(9)Terry,L.A.,Joyce,D.C.,Khambay,B.P.S.,2003.Antifungal compounds in Geraldtonwaxflower tissues.Australasian Plant Pathology 32(3),41-420.
(10)Yff,B.T.,Lindsey,K.L.,Taylor,M.B.,Erasmus,D.G.,Jager,A.K,2002.Thepharmacological screening of Pentanisia prunelloides and the isolation of the antibacterialcompound palmitic acid.J Ethnopharmacol 79(1),101-107.
Claims (1)
1、猫人参挥发油在制备抗菌抑菌药物中的应用。
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CN110946848A (zh) * | 2018-09-27 | 2020-04-03 | 东北师范大学 | 一种具有广谱杀菌作用的复方杀菌剂 |
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CN102984937A (zh) * | 2010-05-23 | 2013-03-20 | 高砂香料工业株式会社 | 抗菌组合物 |
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