CN105486647A - 侧柏叶中乌发活性成分的提取和性能测定 - Google Patents

侧柏叶中乌发活性成分的提取和性能测定 Download PDF

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CN105486647A
CN105486647A CN201510630744.6A CN201510630744A CN105486647A CN 105486647 A CN105486647 A CN 105486647A CN 201510630744 A CN201510630744 A CN 201510630744A CN 105486647 A CN105486647 A CN 105486647A
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extraction
solution
cacumen biotae
resin
blackening
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CN105486647B (zh
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包海鹰
高旦
刘叶
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Jilin Agricultural University
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Abstract

<b>本发明公开了侧柏叶中乌发活性成分树脂的提取和对酪氨酸激活率测试：称取新鲜的侧柏叶</b><b>2.5kg</b><b>，用石油醚（</b><b>30</b><b>℃</b><b>-60</b><b>℃）进行回流提取，提取时间</b><b>3</b><b>小时，料液比</b><b>1:12.5</b><b>，合并提取液浓缩，干燥。取干燥后的适量提取物，放入挥发油提取器中，用水蒸气蒸馏法进行提取，得黄色透明液体</b><b>1.2g</b><b>，剩余的胶状液体干燥后即为树脂，采用蘑菇酪氨酸酶多巴速率氧化法，对树脂进行酪氨酸酶激活率测试，药物浓度为</b><b>30mg/ml</b><b>时，激活率达</b><b>102.13%</b><b>。</b>

Description

侧柏叶中乌发活性成分的提取和性能测定
技术领域
本发明公开了侧柏叶的乌发活性成分树脂的提取分离，以及对酪氨酸酶活性测试。
背景技术
侧柏叶(CacluneuPlatycladi)又名丛柏叶，是侧柏的干燥嫩枝梢及叶。中医认为侧柏叶气清香，味苦涩，性微寒.无毒入心`,肝、大肠经。具有凉血、止血、除内湿、散肿毒之功。可治吐血、鼻出血、尿血、血痢、崩漏、风湿痛咳嗽、烫伤等症。明代李时珍《本草纲目》中记载侧柏叶“乃多寿之木，所以可以入服食。道家以之点汤常饮。”侧柏叶作为传统的中药，其乌发功能一直被人忽略。而在诸多古代文献中记载其主要功能为乌发和生发，在《本草纲目》中谓其：浸油，生发，烧汁，黑发。和猪脂，沐发长黑。然而侧柏叶乌发的机理尚不明确。
发明内容
本发明公开了侧柏叶的乌发活性成分树脂的提取和分离，以及对酪氨酸酶活性的测试。1材料与方法
1.1材料
侧柏叶购自山东省济宁市侧柏种植基地，经吉林农业大学中药材学院中药学专业包海鹰教授鉴定。蘑菇酪氨酸酶(tyrosinase)，L-多巴（L-Dopa），均购自美国Sigma公司：磷酸酶氢二钠，磷酸二氢钠，乙醇，石油醚，乙酸乙酯均为分析纯。
1.2仪器
721型分光光度计为上海第三仪器分析厂产品,SY21-K型恒温水浴锅，RE-52AA旋转蒸发仪上海亚荣生化仪器厂。
实验内容与方法
1.3.1磷酸缓冲溶液（PBS）的配制：称取0.2mol/ml磷酸二氢钠溶液250ml,加0.2mol/ml的氢氧化钠溶液118ml,用水稀释至1000ml，定容，即得。
1.3.2酪氨酸酶溶液的配制：称取13.28g(25KU)蘑菇酪氨酸酶（1881U/mg）,用PBS定容至25ml,即1KU/ml，分装-20℃保存，临用时室温平衡，即得，稀释至实验所需浓度143U/ml。
1.3.3L-多巴溶液的配制：精密称取197.19mg，用PBS溶液定容至100ml,须现用现配，浓度为197.19g/mol。
3.4称取新鲜的侧柏叶2.5kg，用石油醚（30℃-60℃）进行回流提取，提取时间3小时，料液比1:12.5，合并提取液浓缩，干燥。取干燥后的适量提取物，放入挥发油提取器中，用水蒸气蒸馏法进行提取，提取挥发油后剩余的提取液，干燥浓缩，即为侧柏叶中的树脂成分。
1.3.5将上述提取所得到的树脂，用适量二甲基亚砜溶解，然后用磷酸缓冲溶液定容，制成浓度分别为10mg/ml、20mg/ml、30mg/ml的三个浓度梯度。以L-多巴溶液为底物，在3ml的磷酸缓冲液的测活体系中，先加入0.1ml不同浓度的提取物于比色杯中，再加入2.8ml预先在37℃恒温水浴保温的底物溶液，然后加入0.1ml蘑菇酪氨酸酶溶液，立刻混匀，在37℃恒温条件下测定波长为475的吸光度值。
酪氨酸酶活性测试结果
3结论
通过对侧柏叶中的树脂类成分进行酪氨酸酶活性测试，结果树脂对酪氨酸酶的激活率高达102.13%，可能是侧柏叶的乌发活性成分，为进一步侧柏叶乌发产品的开发提供了很好的理论依据，很好的诠释了在《本草纲目》中谓其：“浸油，生发，烧汁，黑发。和猪脂，沐发长黑”的记载。

Claims (2)

1.侧柏叶（ CacumenPlatycladi ）具有乌发生发的功能，并且价格低廉适合作为化妆品、保健品的开发。
2.根据权利要求1所述的应用，其中侧柏叶中乌发活性成分的提取和性能测定如下步骤：
一称量侧柏叶2.5㎏，按1:12.5的量加石油醚（60℃-90℃）回流提取3个小时，提取两次，合并提取液浓缩、干燥，得到34.6g混合固体；
二称取10g混合固体，用水蒸气蒸馏法除去混合物中的挥发油，得到混合物固体9.2g，即为树脂；
三称取13.28g(25KU)蘑菇酪氨酸酶（1881U/mg）,用PBS定容至25ml,即1KU/ml，分装-20℃保存，临用时室温平衡，即得，稀释至实验所需浓度143U/ml；
四称取13.28g(25KU)蘑菇酪氨酸酶（1881U/mg）,用PBS定容至25ml,即1KU/ml，分装-20℃保存，临用时室温平衡，即得，稀释至实验所需浓度143U/ml；
五将上述提取所得到的树脂，用适量二甲基亚砜溶解，然后用磷酸缓冲溶液定容，制成浓度分别为10mg/ml、20mg/ml、30mg/ml的三个浓度梯度；
六以L-多巴溶液为底物，在3ml的磷酸缓冲液的测活体系中，先加入0.1ml不同浓度的提取物于比色杯中，再加入2.8ml预先在37℃恒温水浴保温的底物溶液，然后加入0.1ml蘑菇酪氨酸酶溶液，立刻混匀，在37℃恒温条件下测定波长为475的吸光度值。
CN201510630744.6A 2015-09-29 2015-09-29 侧柏叶中乌发活性成分的提取和性能测定 Expired - Fee Related CN105486647B (zh)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112057371A (zh) * 2020-09-27 2020-12-11 新疆大德恒生物股份有限公司 一种杏仁育发乳、其组合物及其制备方法
CN112842972A (zh) * 2021-04-02 2021-05-28 广州莎莎化妆品制造有限公司 一种乌发液及其制备方法

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CN102028759A (zh) * 2011-01-14 2011-04-27 霸王（广州）有限公司 中药乌发经验方及其制备方法
CN103450136A (zh) * 2013-08-19 2013-12-18 南京标科生物科技有限公司 一种高从侧柏叶中提取扁柏双黄酮的方法

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CN102028759A (zh) * 2011-01-14 2011-04-27 霸王（广州）有限公司 中药乌发经验方及其制备方法
CN103450136A (zh) * 2013-08-19 2013-12-18 南京标科生物科技有限公司 一种高从侧柏叶中提取扁柏双黄酮的方法

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112057371A (zh) * 2020-09-27 2020-12-11 新疆大德恒生物股份有限公司 一种杏仁育发乳、其组合物及其制备方法
CN112842972A (zh) * 2021-04-02 2021-05-28 广州莎莎化妆品制造有限公司 一种乌发液及其制备方法

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