赤芍与白芍质量差异及其形成机制研究
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摘要
赤芍Radix Paeonia Rubra和白芍Radix Paeonia Alba同来源于芍药Paeonia lactifloraPall.的干燥根,但功效各异,其中赤芍具有清热凉血、散瘀止痛的功效；而白芍具有养血调经,敛阴止汗,柔肝止痛,平抑肝阳的功效。简而言之,赤芍是清热药,而白芍是补益药。然而,赤芍和白芍在功效上是否存在差异?其差异形成的化学物质基础为何?产生这种差异的主导因素是什么?这一直是中药学领域研究的经典问题。近年来的研究表明,赤芍与白芍在药理作用上确实存在一定的差异,这初步给出了赤芍和白芍功效差异的药理学证据；在化学成分差异物质基础上,也有报道成分含量和组成比例上存在一定差异。而对于药材质量的差异程度的数量化分析,以及导致这种差异形成相关的遗传、生态环境、加工方法三大要素的贡献度,还有待于设计更为系统的试验加以揭示。从查阅文献和课题组到药材市场和药材主产区调查的情况看,有关赤芍和白芍药材分化形成的本草学证据也有待于考证。基于上述目的,本文主要以采集于代表性产区的野生芍药(赤芍主要来源)和栽培芍药(白芍主要来源)为材料,通过遗传学的多地点栽培对比试验、分子群体遗传学研究以及交替加工处理分析,从性状指标及分子遗传本质两个方面研究二者是否存在差异,并分析人为加工因素和客观环境因素对其差异的影响规律,以探讨形成二者质量差异的机制。主要结论如下：
1.本草考证结果表明：我国自唐宋开始所用的白芍,其来源及道地产区就与现代相同,即为芍药P. lactiflora,道地产区位于浙江杭州。而赤芍的基源植物可能为多种植物,最有可能的是芍药P. lactiflora、草芍药P. obovata、川赤芍P.anomala subsp. veitch、美丽芍药P.mairei四种。对赤芍道地产区为内蒙多伦的认识,形成于现代。自古以来,赤芍与白芍的划分依据共有根色、花色、产地、加工与否、栽培或野生五种。
2.野生和栽培芍药群体内的形态指标变异丰富,采用无性繁殖栽培一年后,二者之间部分形态指标存在显著差异,野生芍药表现为为株幅大、叶片多而大、发根数多的特征,而栽培芍药表现为地茎粗、叶片厚而墨绿的特征。
3.野生和栽培芍药群体间光合生理指标存在显著差异,表现为栽培芍药较野生芍药的叶绿素a、叶绿素b、总叶绿素a+b和类胡萝卜素叶含量均高,但类胡萝卜素／叶绿素低于野生芍药。栽培芍药对弱光的利用效率高于野生芍药,但在高光强条件下,野生芍药的光合性能更强；野生芍药在低浓度CO2即可启动光合反应,但是其在弱光区的光合羧化酶活性却不如栽培芍药高。
4.野生和栽培芍药群体内化学成分指标含量的变异丰富,但栽培芍药中芍药内酯苷含量显著高于野生芍药,野生芍药中儿茶素、芍药苷的含量显著高于栽培芍药。二者主要含有的芍药苷和芍药内酯苷两种成分的含量总和没有显著差异,但这两种成分的比例存在显著差异。
5.赤芍和白芍不同居群材料的ITS序列对比结果表明,野生芍药群体内遗传多样性较栽培芍药丰富；野生芍药和栽培芍药群体间存在显著的遗传分化。
6.加工方法对赤芍与白芍中化学成分的影响研究结果表明：采用赤芍与白芍的加工方法同时处理赤芍和白芍基源植物后,不同加工方法会改变二者中化学成分的含量,但不论加工前后,二者主之间的芍药苷和芍药内酯苷两种化学成分的含量均存在显著差异。说明不能仅以加工方法作为划分赤芍和白芍的依据。
7.环境对赤芍和白芍性状指标的影响结果表明,与环境因子具有显著相关性的性状指标主要是形态指标,总体呈现出随着纬度和日照时数的增高,植株趋于矮小、茎枝和根部分枝密集；光合色素指标中类胡萝卜素含量,以及化学成分中没食子酸、五没食子酰葡萄糖、苯甲酸和丹皮酚含量与环境因子具有不同程度的相关性；其余指标和环境因子之间没有显著相关性。
8.基因型、环境及互作效应分析结果表明,赤芍与白芍之间多数形态指标的差异受到环境因素的影响较大,但叶部形态指标的差异受基因的影响大于环境；二者光合生理指标的差异主要来自于遗传；二者之间显著差异性化学成分中,儿茶素、芍药内酯苷和芍药苷三种成分均受遗传的影响占主导。
综上所述,赤芍和白芍之间形态、光合生理及化学成分三方面性状均存在部分指标的显著性差异,在分子遗传本质上也存在显著的遗传分化。二者之间部分指标受到人为加工和客观环境的影响,但二者主要含有的化学成分芍药苷和芍药内酯苷存在显著差异,且受到加工和环境的影响较小,其差异主要来自于野生和栽培芍药群体间遗传本质上的差异。
According to Chinese Pharmacopoeia(2010ed), the Radix Paeonia Rubra and Radix aeonia Alba are both from the root of Paeonia lactiflora Pall. The Radix Paeonia Rubra has the unction of clearing heat to cool blood and eliminating stasis to stop pain, and the Radix Paeonia lba has the function of enriching blood to regulate menstruation, gathering Yin to stop sweat, ourishing the liver to relieve pain and balancing the liver Yang. In recent years, studies have hown that the Radix Paeonia Rubra and the Radix Paeonia Alba must have the differences in ie pharmacological effects, and the chemical composition and proportion are also different, this onfirm that their function is different, though they derive from the same original plants. Correlational studies indicated that the difference has related to the ecological environment, rocessing methods and genetic factor. The analysis of the relationship in the Radix Paeonia Rubra and the Radix Paeonia Alba on characteristic index and the genetical background and the nalysis of the effects of the process methods and environmental factors on the differences etween them have been made in this thesis based on the cultivation test in different ecological nvironments with the different sources of the materials and the analysis gene research on the Radix Paeonia Rubra and the Radix Paeonia Alba that from different sources.
Herbal textual research indicated that the study on the functions of the Radix Paeonia Rubra and the Radix Paeonia Alba have been done from the ancient time. The original herb and ie authentic region of Radix Paeonia Alba used in the Tang and Song Dynasties was the same s that today, the original herb is P. lactiflora, the authentic region is Hangzhou, Zhejiang 'rovince. And the original herbs of Radix Paeonia Rubra are many, the conventional herbs are P. actiflora,P. obovata,P.anomala subsp. veitch and P.mairei, and the authentic region of Duolun, nner Mongolia was just known in modern time. The color of root and flower, the producing rea, processing or not and cultivation or not have been the basis to distinguish them, and has een done from ancient time, but the final conclusion is not be made.
2. The variation of morphological characteristics in the wild and cultivated P. lactiflora groups are both great. And some morphological characteristics is significant different between he wild and the cultivated, the leaf and the plant width of the wild is bigger, the stem is lenderer, and the root is more than that in the cultivated.
3. The difference of the photosynthetic physiological indexes in the in the wild and ultivated P. lactiflora groups is significant:the chlorophyll a, the chlorophyll b, the total hlorophyll and the carotenoid in the cultivated are higher than that the wild, but the rate of arotenoid to total chlorophyll is lower in the cultivated. The utilization efficiency of weak light in the cultivated is higher than that in the wild, but under the strong light the photosynthesis of the wild is stronger than that of the cultivated. The wild can start the photo-reaction in the lower concentration of the CO2, but in the disphotic zone the photosynthetic carboxylase of the cultivated is stronger than that of the wild.
4. The variation of the chemical composition content in the wild and cultivated P. lactiflora groups are both great. And the albiflorin content in cultivated is obviously higher than that in the wild, but the contents of catechin, and the paeoniflorin in the wild are significantly higher than that in the cultivated, respectively.
5. The comparison results of the ITS in the different population of the Radix Paeonia Rubra and Radix Paeonia Alb indicated that the genetic diversity in the population of the wild is rich, and there are certain genetic distance between the wild and the cultivated.
6. The effect of different process methods on the chemical composition content in the Radix Paeonia Rubra and Radix Paeonia Alb have been studied and the result showed that the chemical composition contents have some difference after the treatment on the original plants with the methods of the Radix Paeonia Rubra and Radix Paeonia Alb,respectively, no matter processing or not, there are some the chemical composition content difference between that. And it indicated that the distinguishing of the Radix Paeonia Rubra and Radix Paeonia Alb is not only basic on the process mrthods.
7. The effects of the environment on the characteristic index indicated that significant correlation mainly exist in the environmental factors and morphological index, it generally shows that as the increasing of the Latitude and sunshine duration, the stem become shorter, and the branching of the stem and root become more. There are different relativity between environmental factors and the content of the carotenoid in photosynthetic pigment, gallic acid, PGG, benzoic acid and paeonol in chemical composition content,respectively. And other characteristic indexes have no relativity relationship with the environmental factors.
8. The analysis results of the gene, environment and its interaction indicated that the difference in the most morphological index of the Radix Paeonia Rubra and Radix Paeonia Alb is mainly affected by the environmental factors, but the difference of leaf morphological index is mainly affected by the gene. The Photosynthetic Physiological Indexes in both Radix Paeonia Rubra and Radix Paeonia Alb is mainly affected by the gene. There are 4 kinds chemical composition content are significantly different between them, the catechin, albiflorin and paeoniflorin are mainly affected by the gene.
In conclusion, Some indexes of the morphology, photosynthetic physiology and chemical constituents are significantly different between the Radix Paeonia Rubra and Radix Paeonia Alb, and there are some genetic differentiation in the gene. Some indexes are affected significantly by the environment and the processing methods. But some are not significantly affected by the environment and the process methods, those differences are mainly from gene. The results of this thesis can provide some reference basic for the rational clinic medicine of the Radix Paeonia Rubra and Radix Paeonia Alb, and provide theoretical support for the sustainable utilization of the wild Radix Paeonia Rubra.
1.本草考证结果表明：我国自唐宋开始所用的白芍,其来源及道地产区就与现代相同,即为芍药P. lactiflora,道地产区位于浙江杭州。而赤芍的基源植物可能为多种植物,最有可能的是芍药P. lactiflora、草芍药P. obovata、川赤芍P.anomala subsp. veitch、美丽芍药P.mairei四种。对赤芍道地产区为内蒙多伦的认识,形成于现代。自古以来,赤芍与白芍的划分依据共有根色、花色、产地、加工与否、栽培或野生五种。
2.野生和栽培芍药群体内的形态指标变异丰富,采用无性繁殖栽培一年后,二者之间部分形态指标存在显著差异,野生芍药表现为为株幅大、叶片多而大、发根数多的特征,而栽培芍药表现为地茎粗、叶片厚而墨绿的特征。
3.野生和栽培芍药群体间光合生理指标存在显著差异,表现为栽培芍药较野生芍药的叶绿素a、叶绿素b、总叶绿素a+b和类胡萝卜素叶含量均高,但类胡萝卜素／叶绿素低于野生芍药。栽培芍药对弱光的利用效率高于野生芍药,但在高光强条件下,野生芍药的光合性能更强；野生芍药在低浓度CO2即可启动光合反应,但是其在弱光区的光合羧化酶活性却不如栽培芍药高。
4.野生和栽培芍药群体内化学成分指标含量的变异丰富,但栽培芍药中芍药内酯苷含量显著高于野生芍药,野生芍药中儿茶素、芍药苷的含量显著高于栽培芍药。二者主要含有的芍药苷和芍药内酯苷两种成分的含量总和没有显著差异,但这两种成分的比例存在显著差异。
5.赤芍和白芍不同居群材料的ITS序列对比结果表明,野生芍药群体内遗传多样性较栽培芍药丰富；野生芍药和栽培芍药群体间存在显著的遗传分化。
6.加工方法对赤芍与白芍中化学成分的影响研究结果表明：采用赤芍与白芍的加工方法同时处理赤芍和白芍基源植物后,不同加工方法会改变二者中化学成分的含量,但不论加工前后,二者主之间的芍药苷和芍药内酯苷两种化学成分的含量均存在显著差异。说明不能仅以加工方法作为划分赤芍和白芍的依据。
7.环境对赤芍和白芍性状指标的影响结果表明,与环境因子具有显著相关性的性状指标主要是形态指标,总体呈现出随着纬度和日照时数的增高,植株趋于矮小、茎枝和根部分枝密集；光合色素指标中类胡萝卜素含量,以及化学成分中没食子酸、五没食子酰葡萄糖、苯甲酸和丹皮酚含量与环境因子具有不同程度的相关性；其余指标和环境因子之间没有显著相关性。
8.基因型、环境及互作效应分析结果表明,赤芍与白芍之间多数形态指标的差异受到环境因素的影响较大,但叶部形态指标的差异受基因的影响大于环境；二者光合生理指标的差异主要来自于遗传；二者之间显著差异性化学成分中,儿茶素、芍药内酯苷和芍药苷三种成分均受遗传的影响占主导。
综上所述,赤芍和白芍之间形态、光合生理及化学成分三方面性状均存在部分指标的显著性差异,在分子遗传本质上也存在显著的遗传分化。二者之间部分指标受到人为加工和客观环境的影响,但二者主要含有的化学成分芍药苷和芍药内酯苷存在显著差异,且受到加工和环境的影响较小,其差异主要来自于野生和栽培芍药群体间遗传本质上的差异。
According to Chinese Pharmacopoeia(2010ed), the Radix Paeonia Rubra and Radix aeonia Alba are both from the root of Paeonia lactiflora Pall. The Radix Paeonia Rubra has the unction of clearing heat to cool blood and eliminating stasis to stop pain, and the Radix Paeonia lba has the function of enriching blood to regulate menstruation, gathering Yin to stop sweat, ourishing the liver to relieve pain and balancing the liver Yang. In recent years, studies have hown that the Radix Paeonia Rubra and the Radix Paeonia Alba must have the differences in ie pharmacological effects, and the chemical composition and proportion are also different, this onfirm that their function is different, though they derive from the same original plants. Correlational studies indicated that the difference has related to the ecological environment, rocessing methods and genetic factor. The analysis of the relationship in the Radix Paeonia Rubra and the Radix Paeonia Alba on characteristic index and the genetical background and the nalysis of the effects of the process methods and environmental factors on the differences etween them have been made in this thesis based on the cultivation test in different ecological nvironments with the different sources of the materials and the analysis gene research on the Radix Paeonia Rubra and the Radix Paeonia Alba that from different sources.
Herbal textual research indicated that the study on the functions of the Radix Paeonia Rubra and the Radix Paeonia Alba have been done from the ancient time. The original herb and ie authentic region of Radix Paeonia Alba used in the Tang and Song Dynasties was the same s that today, the original herb is P. lactiflora, the authentic region is Hangzhou, Zhejiang 'rovince. And the original herbs of Radix Paeonia Rubra are many, the conventional herbs are P. actiflora,P. obovata,P.anomala subsp. veitch and P.mairei, and the authentic region of Duolun, nner Mongolia was just known in modern time. The color of root and flower, the producing rea, processing or not and cultivation or not have been the basis to distinguish them, and has een done from ancient time, but the final conclusion is not be made.
2. The variation of morphological characteristics in the wild and cultivated P. lactiflora groups are both great. And some morphological characteristics is significant different between he wild and the cultivated, the leaf and the plant width of the wild is bigger, the stem is lenderer, and the root is more than that in the cultivated.
3. The difference of the photosynthetic physiological indexes in the in the wild and ultivated P. lactiflora groups is significant:the chlorophyll a, the chlorophyll b, the total hlorophyll and the carotenoid in the cultivated are higher than that the wild, but the rate of arotenoid to total chlorophyll is lower in the cultivated. The utilization efficiency of weak light in the cultivated is higher than that in the wild, but under the strong light the photosynthesis of the wild is stronger than that of the cultivated. The wild can start the photo-reaction in the lower concentration of the CO2, but in the disphotic zone the photosynthetic carboxylase of the cultivated is stronger than that of the wild.
4. The variation of the chemical composition content in the wild and cultivated P. lactiflora groups are both great. And the albiflorin content in cultivated is obviously higher than that in the wild, but the contents of catechin, and the paeoniflorin in the wild are significantly higher than that in the cultivated, respectively.
5. The comparison results of the ITS in the different population of the Radix Paeonia Rubra and Radix Paeonia Alb indicated that the genetic diversity in the population of the wild is rich, and there are certain genetic distance between the wild and the cultivated.
6. The effect of different process methods on the chemical composition content in the Radix Paeonia Rubra and Radix Paeonia Alb have been studied and the result showed that the chemical composition contents have some difference after the treatment on the original plants with the methods of the Radix Paeonia Rubra and Radix Paeonia Alb,respectively, no matter processing or not, there are some the chemical composition content difference between that. And it indicated that the distinguishing of the Radix Paeonia Rubra and Radix Paeonia Alb is not only basic on the process mrthods.
7. The effects of the environment on the characteristic index indicated that significant correlation mainly exist in the environmental factors and morphological index, it generally shows that as the increasing of the Latitude and sunshine duration, the stem become shorter, and the branching of the stem and root become more. There are different relativity between environmental factors and the content of the carotenoid in photosynthetic pigment, gallic acid, PGG, benzoic acid and paeonol in chemical composition content,respectively. And other characteristic indexes have no relativity relationship with the environmental factors.
8. The analysis results of the gene, environment and its interaction indicated that the difference in the most morphological index of the Radix Paeonia Rubra and Radix Paeonia Alb is mainly affected by the environmental factors, but the difference of leaf morphological index is mainly affected by the gene. The Photosynthetic Physiological Indexes in both Radix Paeonia Rubra and Radix Paeonia Alb is mainly affected by the gene. There are 4 kinds chemical composition content are significantly different between them, the catechin, albiflorin and paeoniflorin are mainly affected by the gene.
In conclusion, Some indexes of the morphology, photosynthetic physiology and chemical constituents are significantly different between the Radix Paeonia Rubra and Radix Paeonia Alb, and there are some genetic differentiation in the gene. Some indexes are affected significantly by the environment and the processing methods. But some are not significantly affected by the environment and the process methods, those differences are mainly from gene. The results of this thesis can provide some reference basic for the rational clinic medicine of the Radix Paeonia Rubra and Radix Paeonia Alb, and provide theoretical support for the sustainable utilization of the wild Radix Paeonia Rubra.
引文
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