防风药材化学成分及其与根际土壤中无机元素含量的相关性研究
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摘要
本文通过对我国9个省份14个产地的防风药材中主要有效成分及8种无机元素的含量,防风根际土壤的基本养分及8种无机元素的含量进行测定,并对防风总色原酮含量测定方法进行筛选,试图阐明土壤对药材中各类成分的影响,以及建立一种简便、快捷的测定防风总色原酮含量的方法。主要研究结果如下：
1.防风挥发油的含量随产地及采集时间的不同差异显著,栽培防风中挥发油含量变幅为0.7～4.5mg/g,野生防风为0.8～5.3mg/g,均以1-甲酰基-4-(1-丁烯-3基)-苯、没药烯为主要成分,但野生防风挥发油中所含成分的种类要多于栽培防风,陇西水防风的挥发油含量和成分均与正品防风相似。药材中多糖的含量随采集时间变化显著,总体上8月份多糖含量较高,且正品防风多糖含量要高于习用防风。防风色原酮及香豆素总含量均以左家最高,分别为2.5%和0.2%；安国防风总色原酮含量最低,为0.2%。
2.防风中8种无机元素的平均含量由高到低依次为：Ca (36243.1mg/kg)> Mg (18095.2mg/kg)> Na (16847.5mg/kg)> K (9461.7mg/kg)>>Fe (184.3mg/kg)> Zn (97.2mg/kg)> Mn (24.8mg/kg)> Cu (15.7mg/kg)。8种元素的最高含量依次出现在通辽(Ca)、白城(Mg)、大庆(Na和K)、侯马(Fe和Mn)、四平(Zn)、(Cu)防风中,最低含量依次出现在大兴安岭(Ca)、鞍山(Mg)、四平(Na)、通辽(Cu、Mn、Zn、Fe和K)防风中。而不同地区防风中无机元素含量间没有较好的规律性。
3.通过对不同地区2个时间采集的防风根际土壤样品理化性质进行分析,结果表明,各地区土壤基本养分差异较大,其中；土壤含水量变幅为1.5～7.9%；pH由酸性到碱性,变化较大,变幅为5.3-8.4；碱解氮的含量由49.0mg/kg到488.8mg/kg;有效磷含量的变幅为5.0～384.8mg/kg;速效钾含量的变幅为47.9～575.9mg/kg;有机质含量变幅为9.7～319.6g/kg;全氮含量变幅为1.3～9.1g/kg。总体上,栾川和大兴安岭2个地区的土壤养分极其丰富,除大连地区碱解氮含量偏低外,其他地区均不存在营养缺乏的情况。
4.不同地区土壤样品中8种元素的平均含量间差异较大,按平均含量由高到低的顺序为：Na (14923.9mg/kg)> Fe (10024.5mg/kg)> Ca (9047.0mg/kg)> K (4357.7mg/kg)> Mg (1854.3mg/kg)> Mn (1386.1mg/kg)> Zn (119.7mg/kg)> Cu (16.2mg/kg)。其中侯马土壤中含Zn、Fe、Mg、K和Na最高,但含Mn最低；鞍山土壤含Cu最高,含Mg最低；白城土壤含Ca最高,K、Na和Zn最低；通辽土壤中Mn含量最高,Cu最低：大庆土壤中含Fe最低,Ca的最低含量则出现在大兴安岭地区。
5.防风各成分含量的相关分析表明,挥发油和多糖的含量总体上不受其他有机成分的影响；色原酮(除升麻素外)及其总含量和香豆素及其总含量相互之间均存在着不同程度的正相关,彼此之间具有良好的协同作用,总色原酮的含量亦与锌含量呈正相关。无机元素中锰与铜和铁之间呈显著的正相关；而铜与镁,钙与钾之间呈显著的负相关。其他无机元素相互之间亦有不同程度的相互抑制或协同作用,但均受时间的影响较大。
6.防风有效成分与土壤间的相关性分析表明,防风挥发油含量及多糖含量随采集时间的不同受土壤的影响无规律性。色原酮及香豆素类化合物与土壤的理化性质无显著相关,但受土壤中无机元素含量的影响较大：总色原酮与土壤中钾和镁含量之间,欧前胡素含量与土壤中锰含量之间呈显著的正相关；总香豆素含量与土壤中铜含量之间,亥茅酚苷含量与土壤钾、钠含量间呈显著的负相关。野生防风中的铜、钠、镁含量分别随土壤中的铜、钠、镁含量的增加而显著增加,但锰却随土壤中锰含量的增加而显著降低,防风中钙与土壤锰含量之间呈显著的正相关。
首次对防风生长的土壤环境进行了研究,初步明确了影响防风有效成分含量的主要土壤因子是有效磷和有机质的含量、碱解氮和有机质的含量、Fe和K的含量。
7.本文首次建立了苯酚-硫酸-紫外分光光度法测定防风总色原酮的含量。通过实验确定最佳测定条件：5%苯酚加入量为0.8mL,浓硫酸为3.5mL,迅速振荡均匀后置40℃水浴上加热40min,冷却15min后于324nm处测定吸光度值。以5-0-甲基维斯阿米醇苷为标准对照品,其回归方程为Y=7.2305X+0.1889(r=0.9992),在11.5～115μg范围内线性关系良好。经过方法学考察,其精密度(RSD为1.1%)、稳定性(RSD为1.0%)、重现性(RSD为1.1%)、加样回收率(RSD为1.0%)均符合样品含量测定的方法学要求。该方法具有准确、简便、经济、适用的特点,可用于防风总色原酮含量的测定。
研究结果为防风的栽培及质量调控措施的建立提供了理论依据。
In this article, contents of main active components and8inorganic elements in the root of Saposhnikovia divaricata (S. divaricata), a raw material for medicine from14areas in9provinces of China, were measured, and those of basic nutrients and8inorganic elements in the rhizosphere soil of S. divaricata were also determined. Moreover, methods choice of the determination of total chromones in S. divaricata was studied. The purpose of our study was to clarify the influence of the rhizosphere soil on all kinds of ingredients in the herbs, and establish a simple and quick method of determination of the total chromnones content in S. divaricata.
The main findings by us are summarized as follows:
1. The content of volatile oil in S. divaricata varied with the place of origin and the collecting time significantly. The amplitude of variation in the volatile oil in the cultivated S. divaricata ranged from0.7to4.5mg/g, while that of the wild S. divaricata from0.8to5.4mg/g; the main components of all volatile oils were1-formyl-4-(1-buten-3-yl)-benzene and bisabolene, but the varieties of components of volatile oil from the wild S. divaricata were more than those from the cultivated S. divaricata. The content and the variedties of components of volatile soil form Carum carvi L were similar to S. divaricata. The polysaccharide contents in the herbs varied significantly with the change of the collecting time. The results showed that in general, higher polysaccharide content was found on August and the polysaccharide content in the sertificated S. divaricata was higher than that in the traditional S. divaricata. The contents of total chromones and total coumarins in S. divaricata were highest in Zuojia area, which were2.5%and0.2%, respectively; the content of total chromones in S. divaricata from Anguo area was lowest, which was0.2%.
2. The descending order of the average contents of8inorganic elements in S. divaricata was Ca (36243.1mg/kg)> Mg (18095.2mg/kg)> Na (16847.5mg/kg)> K (9461.7mg/kg)>> Fe (184.3mg/kg)> Zn (97.2mg/kg)> Mn (24.8mg/kg)> Cu (15.7mg/kg). The highest content of eight elements was found in S. divaricata in turn from Tongliao (Ca), Baicheng (Mg), Daqing (Na and K), Houma (Fe and Mn), Siping (Zn) and Anshan (Cu), while the lowest content of them was found in S. divaricata in turn from Daxinganling (Ca), Anshan (Mg), Siping (Na) and Tongliao (Cu, Mn, Zn, Fe, and K). However, the differences in contents of inorganic elements among the different regions were great and no good regularity was shown.
3. The physical and chemical properties of rhizosphere soil samples collected from different areas at two different times were analyzed. The results showed that there were significant differences in the basic nutrients of soils from the different regions, including:the amplitude of variation of water content ranged from1.5to7.9%; that of pH from acidic to alkaline ranged from5.3to8.4, which changed more significantly; that in the content of hydrolyzable nitrogen was from49.0mg/kg to488.8mg/kg; that in the content of available phosphorus ranged from5.0to384.8mg/kg; that of available potassium ranged from47.9-575.9mg/kg; organic matter content ranged from9.7～319.6g/kg; the total nitrogen content ranged from1.3to9.1g/kg. As a whole, the nutrients in soils from two areas, Luanchuan and Daxinganling, were extremely rich, and those of soils from the other areas were not deficient except for the lower content of hydrolyzable nitrogen in the soil from Dalian area.
4. There were significant differences in the average content of eight elements in the soil samples from different regions. Based on the average content, the descending order was:Na (14923.9mg/kg)> Fe (10024.5mg/kg)> Ca (9047.0mg/kg)> K (4357.7mg/kg)> Mg (1854.3mg/kg)> Mn (1386.1mg/kg)> Zn (119.7mg/kg)> Cu (16.2mg/kg), in which average contents of Zn, Fe, Mg, K and Na in the soil from Houma area were the highest, but the average content of Mn was the lowest; the average content of Cu in the soil from Anshan was the highest, but that of Mg was the lowest; the average content of Ca in the soil from Baicheng was the highest, but average contents of K, Na, and Zn were the lowest; the average content of Mn in the soil from Tongliao was the highest, but that of Cu was the lowest; the average content of Fe in the soil from Daqing was the lowest; the average content of Ca in the soil from Daxinganling was the lowest.
5. The correlation analysis on contents of components in S. divaricata showed that the contents of volatile oil and polysaccharides were generally not influenced by the other organic ingredients in it; there was a positive correlation in various degrees between chromones (except cimicifugin) and its total content, and coumarins and its total content, they had a good synergy with each other, and the total chromones content was positively correlated with the Zn content. Mn, which is an inorganic element, was significant positive correlated with Cu and Fe; but the correlations between Cu and Mn contents, and Ca and K contents were significantly negative. There were also mutual inhibitory or synergistic effects between the other inorganic elements, but they were affected by the collecting time.
6. The analysis on the correlation between the active ingredients in S. divaricata and the soils showed that there were no regularities in the volatile oil and polysaccharide contents affected by the soil at the different collecting times. Chromone and coumarin compounds had nothing to do with the physicochemical properties of the soil, but were affected greatly by the inorganic element content in the soil:significantly negaitive correlations between the total chromone in S.divaricata and the K, Na contents in the soil, between the total coumarin content in S. divaricata and the Cu content in the soil, and between the sec-O-glucosylhamaudol content in S. divaricata and the K or Na content in the soil could be seen; it was also found that there was significantly positive correlation between the imperitorin content in S. divaricata and the Mn content in the soil. The Cu, Na and Mg contents in wild S. divaricata were positively affected by the increase of themselves in the soil, while Mn content was negatively affected by the Mn content in the soil. Ca content in plant would increase with the increase of Mn content in the soil.
The soil environment of S. divaricata was investigated and the nutritional elements of soil were determined in this paper. Factor analysis results showed that concentrations of available phosphorus and organic matter, hydrolizable nitrogen and organic matter, Fe and K in rhizosphere soil were the main factors according to the quality of S. divaricata.
7. It should be the first time that a phenol-sulfuric acid-UV spectrophotometric method was established to determine the total chromones content in S. divaricata. The optimal conditions were established by experiment as followed:the added amount of5%phenol was0.8mL and that of concentrated sulfuric acid was3.5mL, it was rapidly oscillated and then heated in a water bath at40℃for40min, and finally was used for the determination of absorbance value at324nm after it was cooled for15min.5-O-methylvisammioside was used as the standard reference substance, the regression equation was Y=7.2305X+0.1889(r=0.9992), and there was a good linear relationship in the range of11.5～115μg. The methodological study indicated that the precision (RSD1.1%), stability testing (RSD1.0%), reproducibility (RSD1.1%), and average recovery (RSD1.0%) were in line with the requirement for the determination of sample content proposed by the methodology. It could be believed that the method should be accurate, simple, cheap, and applicable for the determination of the total content of chromones.
The results of this study provide theoretical foundation for cultivation and foundation of measures for regulating and controlling quality of S. divariata
1.防风挥发油的含量随产地及采集时间的不同差异显著,栽培防风中挥发油含量变幅为0.7～4.5mg/g,野生防风为0.8～5.3mg/g,均以1-甲酰基-4-(1-丁烯-3基)-苯、没药烯为主要成分,但野生防风挥发油中所含成分的种类要多于栽培防风,陇西水防风的挥发油含量和成分均与正品防风相似。药材中多糖的含量随采集时间变化显著,总体上8月份多糖含量较高,且正品防风多糖含量要高于习用防风。防风色原酮及香豆素总含量均以左家最高,分别为2.5%和0.2%；安国防风总色原酮含量最低,为0.2%。
2.防风中8种无机元素的平均含量由高到低依次为：Ca (36243.1mg/kg)> Mg (18095.2mg/kg)> Na (16847.5mg/kg)> K (9461.7mg/kg)>>Fe (184.3mg/kg)> Zn (97.2mg/kg)> Mn (24.8mg/kg)> Cu (15.7mg/kg)。8种元素的最高含量依次出现在通辽(Ca)、白城(Mg)、大庆(Na和K)、侯马(Fe和Mn)、四平(Zn)、(Cu)防风中,最低含量依次出现在大兴安岭(Ca)、鞍山(Mg)、四平(Na)、通辽(Cu、Mn、Zn、Fe和K)防风中。而不同地区防风中无机元素含量间没有较好的规律性。
3.通过对不同地区2个时间采集的防风根际土壤样品理化性质进行分析,结果表明,各地区土壤基本养分差异较大,其中；土壤含水量变幅为1.5～7.9%；pH由酸性到碱性,变化较大,变幅为5.3-8.4；碱解氮的含量由49.0mg/kg到488.8mg/kg;有效磷含量的变幅为5.0～384.8mg/kg;速效钾含量的变幅为47.9～575.9mg/kg;有机质含量变幅为9.7～319.6g/kg;全氮含量变幅为1.3～9.1g/kg。总体上,栾川和大兴安岭2个地区的土壤养分极其丰富,除大连地区碱解氮含量偏低外,其他地区均不存在营养缺乏的情况。
4.不同地区土壤样品中8种元素的平均含量间差异较大,按平均含量由高到低的顺序为：Na (14923.9mg/kg)> Fe (10024.5mg/kg)> Ca (9047.0mg/kg)> K (4357.7mg/kg)> Mg (1854.3mg/kg)> Mn (1386.1mg/kg)> Zn (119.7mg/kg)> Cu (16.2mg/kg)。其中侯马土壤中含Zn、Fe、Mg、K和Na最高,但含Mn最低；鞍山土壤含Cu最高,含Mg最低；白城土壤含Ca最高,K、Na和Zn最低；通辽土壤中Mn含量最高,Cu最低：大庆土壤中含Fe最低,Ca的最低含量则出现在大兴安岭地区。
5.防风各成分含量的相关分析表明,挥发油和多糖的含量总体上不受其他有机成分的影响；色原酮(除升麻素外)及其总含量和香豆素及其总含量相互之间均存在着不同程度的正相关,彼此之间具有良好的协同作用,总色原酮的含量亦与锌含量呈正相关。无机元素中锰与铜和铁之间呈显著的正相关；而铜与镁,钙与钾之间呈显著的负相关。其他无机元素相互之间亦有不同程度的相互抑制或协同作用,但均受时间的影响较大。
6.防风有效成分与土壤间的相关性分析表明,防风挥发油含量及多糖含量随采集时间的不同受土壤的影响无规律性。色原酮及香豆素类化合物与土壤的理化性质无显著相关,但受土壤中无机元素含量的影响较大：总色原酮与土壤中钾和镁含量之间,欧前胡素含量与土壤中锰含量之间呈显著的正相关；总香豆素含量与土壤中铜含量之间,亥茅酚苷含量与土壤钾、钠含量间呈显著的负相关。野生防风中的铜、钠、镁含量分别随土壤中的铜、钠、镁含量的增加而显著增加,但锰却随土壤中锰含量的增加而显著降低,防风中钙与土壤锰含量之间呈显著的正相关。
首次对防风生长的土壤环境进行了研究,初步明确了影响防风有效成分含量的主要土壤因子是有效磷和有机质的含量、碱解氮和有机质的含量、Fe和K的含量。
7.本文首次建立了苯酚-硫酸-紫外分光光度法测定防风总色原酮的含量。通过实验确定最佳测定条件：5%苯酚加入量为0.8mL,浓硫酸为3.5mL,迅速振荡均匀后置40℃水浴上加热40min,冷却15min后于324nm处测定吸光度值。以5-0-甲基维斯阿米醇苷为标准对照品,其回归方程为Y=7.2305X+0.1889(r=0.9992),在11.5～115μg范围内线性关系良好。经过方法学考察,其精密度(RSD为1.1%)、稳定性(RSD为1.0%)、重现性(RSD为1.1%)、加样回收率(RSD为1.0%)均符合样品含量测定的方法学要求。该方法具有准确、简便、经济、适用的特点,可用于防风总色原酮含量的测定。
研究结果为防风的栽培及质量调控措施的建立提供了理论依据。
In this article, contents of main active components and8inorganic elements in the root of Saposhnikovia divaricata (S. divaricata), a raw material for medicine from14areas in9provinces of China, were measured, and those of basic nutrients and8inorganic elements in the rhizosphere soil of S. divaricata were also determined. Moreover, methods choice of the determination of total chromones in S. divaricata was studied. The purpose of our study was to clarify the influence of the rhizosphere soil on all kinds of ingredients in the herbs, and establish a simple and quick method of determination of the total chromnones content in S. divaricata.
The main findings by us are summarized as follows:
1. The content of volatile oil in S. divaricata varied with the place of origin and the collecting time significantly. The amplitude of variation in the volatile oil in the cultivated S. divaricata ranged from0.7to4.5mg/g, while that of the wild S. divaricata from0.8to5.4mg/g; the main components of all volatile oils were1-formyl-4-(1-buten-3-yl)-benzene and bisabolene, but the varieties of components of volatile oil from the wild S. divaricata were more than those from the cultivated S. divaricata. The content and the variedties of components of volatile soil form Carum carvi L were similar to S. divaricata. The polysaccharide contents in the herbs varied significantly with the change of the collecting time. The results showed that in general, higher polysaccharide content was found on August and the polysaccharide content in the sertificated S. divaricata was higher than that in the traditional S. divaricata. The contents of total chromones and total coumarins in S. divaricata were highest in Zuojia area, which were2.5%and0.2%, respectively; the content of total chromones in S. divaricata from Anguo area was lowest, which was0.2%.
2. The descending order of the average contents of8inorganic elements in S. divaricata was Ca (36243.1mg/kg)> Mg (18095.2mg/kg)> Na (16847.5mg/kg)> K (9461.7mg/kg)>> Fe (184.3mg/kg)> Zn (97.2mg/kg)> Mn (24.8mg/kg)> Cu (15.7mg/kg). The highest content of eight elements was found in S. divaricata in turn from Tongliao (Ca), Baicheng (Mg), Daqing (Na and K), Houma (Fe and Mn), Siping (Zn) and Anshan (Cu), while the lowest content of them was found in S. divaricata in turn from Daxinganling (Ca), Anshan (Mg), Siping (Na) and Tongliao (Cu, Mn, Zn, Fe, and K). However, the differences in contents of inorganic elements among the different regions were great and no good regularity was shown.
3. The physical and chemical properties of rhizosphere soil samples collected from different areas at two different times were analyzed. The results showed that there were significant differences in the basic nutrients of soils from the different regions, including:the amplitude of variation of water content ranged from1.5to7.9%; that of pH from acidic to alkaline ranged from5.3to8.4, which changed more significantly; that in the content of hydrolyzable nitrogen was from49.0mg/kg to488.8mg/kg; that in the content of available phosphorus ranged from5.0to384.8mg/kg; that of available potassium ranged from47.9-575.9mg/kg; organic matter content ranged from9.7～319.6g/kg; the total nitrogen content ranged from1.3to9.1g/kg. As a whole, the nutrients in soils from two areas, Luanchuan and Daxinganling, were extremely rich, and those of soils from the other areas were not deficient except for the lower content of hydrolyzable nitrogen in the soil from Dalian area.
4. There were significant differences in the average content of eight elements in the soil samples from different regions. Based on the average content, the descending order was:Na (14923.9mg/kg)> Fe (10024.5mg/kg)> Ca (9047.0mg/kg)> K (4357.7mg/kg)> Mg (1854.3mg/kg)> Mn (1386.1mg/kg)> Zn (119.7mg/kg)> Cu (16.2mg/kg), in which average contents of Zn, Fe, Mg, K and Na in the soil from Houma area were the highest, but the average content of Mn was the lowest; the average content of Cu in the soil from Anshan was the highest, but that of Mg was the lowest; the average content of Ca in the soil from Baicheng was the highest, but average contents of K, Na, and Zn were the lowest; the average content of Mn in the soil from Tongliao was the highest, but that of Cu was the lowest; the average content of Fe in the soil from Daqing was the lowest; the average content of Ca in the soil from Daxinganling was the lowest.
5. The correlation analysis on contents of components in S. divaricata showed that the contents of volatile oil and polysaccharides were generally not influenced by the other organic ingredients in it; there was a positive correlation in various degrees between chromones (except cimicifugin) and its total content, and coumarins and its total content, they had a good synergy with each other, and the total chromones content was positively correlated with the Zn content. Mn, which is an inorganic element, was significant positive correlated with Cu and Fe; but the correlations between Cu and Mn contents, and Ca and K contents were significantly negative. There were also mutual inhibitory or synergistic effects between the other inorganic elements, but they were affected by the collecting time.
6. The analysis on the correlation between the active ingredients in S. divaricata and the soils showed that there were no regularities in the volatile oil and polysaccharide contents affected by the soil at the different collecting times. Chromone and coumarin compounds had nothing to do with the physicochemical properties of the soil, but were affected greatly by the inorganic element content in the soil:significantly negaitive correlations between the total chromone in S.divaricata and the K, Na contents in the soil, between the total coumarin content in S. divaricata and the Cu content in the soil, and between the sec-O-glucosylhamaudol content in S. divaricata and the K or Na content in the soil could be seen; it was also found that there was significantly positive correlation between the imperitorin content in S. divaricata and the Mn content in the soil. The Cu, Na and Mg contents in wild S. divaricata were positively affected by the increase of themselves in the soil, while Mn content was negatively affected by the Mn content in the soil. Ca content in plant would increase with the increase of Mn content in the soil.
The soil environment of S. divaricata was investigated and the nutritional elements of soil were determined in this paper. Factor analysis results showed that concentrations of available phosphorus and organic matter, hydrolizable nitrogen and organic matter, Fe and K in rhizosphere soil were the main factors according to the quality of S. divaricata.
7. It should be the first time that a phenol-sulfuric acid-UV spectrophotometric method was established to determine the total chromones content in S. divaricata. The optimal conditions were established by experiment as followed:the added amount of5%phenol was0.8mL and that of concentrated sulfuric acid was3.5mL, it was rapidly oscillated and then heated in a water bath at40℃for40min, and finally was used for the determination of absorbance value at324nm after it was cooled for15min.5-O-methylvisammioside was used as the standard reference substance, the regression equation was Y=7.2305X+0.1889(r=0.9992), and there was a good linear relationship in the range of11.5～115μg. The methodological study indicated that the precision (RSD1.1%), stability testing (RSD1.0%), reproducibility (RSD1.1%), and average recovery (RSD1.0%) were in line with the requirement for the determination of sample content proposed by the methodology. It could be believed that the method should be accurate, simple, cheap, and applicable for the determination of the total content of chromones.
The results of this study provide theoretical foundation for cultivation and foundation of measures for regulating and controlling quality of S. divariata
引文
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