盐穗木总生物碱的提取及其抑菌活性的研究
摘要
盐穗木(Halostachyscaspica)隶属于藜科(Chenopodiaceae)盐穗木属(Halostachys C.A.Mey),对盐分适应性极强,是盐土荒漠主要植物之一。盐穗木化学成分复杂,目前有关盐穗木抑菌活性成分的研究很少。本论文针对盐穗木中的生物碱成分进行了初步研究,包括生物碱的提取、纯化、结构鉴定和抑菌作用。
本试验首先确定了盐穗木所含主要化学成分种类;选择了四种不同提取方法(酸水提取法、乙醇加热提取法、超声辅助提取法和索氏提取法)提取盐穗木中的总生物碱并对提取效率进行评价;通过紫外波段扫描确定盐穗木总生物碱的特征吸收波长;采用正交试验考察了溶媒量(A)、溶媒浓度(B)和超声波处理时间(C)三个因素对提取效率的影响;研究了盐穗木提取物对微生物生长的抑制作用;利用薄层色谱分析和质谱分析简单鉴定了盐穗木提取物中生物碱的种类。
结果表明:盐穗木中含有酚类、鞣质、生物碱、黄酮、糖、多糖及其甙类、氨基酸及多肽、皂苷、有机酸、甾体和三萜类物质、挥发油等多种化学成分。乙醇加热法效果最差,总碱含量仅为0.404mg/g;索氏提取法提取效率次之(0.477mg/g),酸水提取法提取时间短、效率较高(0.520mg/g);超声波提取法效果最好,总碱含量为0.993mg/g。盐穗木特征吸收波长为280nm。进一步优化方法,得到最佳工艺为A2B2C3,即溶媒量为0.5g/mL、溶媒浓度为85%、超声波处理时间为90min。根据抑菌试验结果可知,盐穗木的抑菌活性物质主要在总生物碱的乙酸乙酯相中。盐穗木提取物对枯草芽孢杆菌、大肠杆菌、金黄色葡萄球菌有较强的抑菌作用。乙酸乙酯相对金黄色葡萄球菌和枯草芽孢杆菌的最小抑菌浓度为7.8mg/mL,对大肠埃希杆菌的最小抑菌浓度为15.6mg/mL,说明乙酸乙酯相对金黄色葡萄球菌的抑菌作用要大于对于大肠埃希杆菌的抑菌作用。总浸膏对大肠埃希杆菌、金黄色葡萄球菌和枯草芽孢杆菌的最小抑菌浓度为31.2mg/mL;对链球菌、普通变形杆菌、面包酵母无抑菌作用。通过对盐穗木总生物碱进行TLC和GC-MS分析,鉴定出9种生物碱类化合物。
Caspica (Halostachyscaspica) is belonging to Chenopodiaceae (Chenopodiaceae) caspica (Halostachys CAMey).It is one of the main plant in the saline desert which is strong adaptability to the salt. Caspica contains complex chemical compositions. Currently,there are little study of antibacterial activity of constituents in the caspica. This thesis aim at caspica alkaloids with a preliminary study, including the alkaloid extraction, purification, structure identification and antimicrobial activities.
Firstly, this study identified the main kind of chemical components in the caspica ; chosen four different extraction methods (acidic water extraction, ethanol heating extraction, ultrasound-assisted extraction and Soxhlet extraction) to extract the total alkaloids and evaluation of extraction efficiency; use ultraviolet scanning to determine the total alkaloids caspica of characteristic absorption wavelength; investigated by orthogonal test solvent volume (A), solvent concentration (B) and ultrasonic treatment time (C) 3 factors on the extraction efficiency; study caspica extract on microbial growth inhibition; use of thin layer chromatography and mass spectrometry simply identified the types of alkaloids in the caspica.
The results showed that: the caspica contains phenols, tannins, alkaloids, flavonoids, sugars, polysaccharides and their glycosides, amino acids and peptides, saponins, organic acids, steroids and triterpenoids, a variety of chemical constituents of volatile oil ,etc. Ethanol heating is the worst, with a total alkali content of only 0.404mg / g; Soxhlet extraction followed by extraction efficiency (0.477mg / g);Acid extraction extraction time, higher efficiency (0.520mg / g); Ultrasonic extraction was the best, with a total alkali content of 0.993mg / g. Caspica characteristic absorption wavelength is 280nm.
Further optimization, the best technology is the A2B2C3, the solvent volume 0.5g/mL, 85% solvent concentration, ultrasonic processing time is 90min.
According to inhibition experiment showed that, caspica antibacterial active substances is mainly in the ethyl acetate phase of total alkaloids. Caspica extract has strong antimicrobial effect to the Bacillus subtilis, Escherichia coli, Staphylococcus aureus. Ethyl acetate relative of Staphylococcus aureus and Bacillus subtilis the minimum inhibitory concentration are 7.8mg/mL, to the Escherichia coli minimum inhibitory concentration is 15.6mg/mL.The result shows that ethyl acetate relative inhibition of Staphylococcus aureus greater than the bacteria of Escherichia coli antibiotic effects. The total extract to the Escherichia coli, Staphylococcus aureus and Bacillus subtilis the minimum inhibitory concentration are 31.2mg/mL; there are no inhibitory effect to the streptococcus, Proteus vulgaris and baker's yeast. Caspica identified nine kinds of alkaloids by TLC of total alkaloids and GC-MS analysis.
本试验首先确定了盐穗木所含主要化学成分种类;选择了四种不同提取方法(酸水提取法、乙醇加热提取法、超声辅助提取法和索氏提取法)提取盐穗木中的总生物碱并对提取效率进行评价;通过紫外波段扫描确定盐穗木总生物碱的特征吸收波长;采用正交试验考察了溶媒量(A)、溶媒浓度(B)和超声波处理时间(C)三个因素对提取效率的影响;研究了盐穗木提取物对微生物生长的抑制作用;利用薄层色谱分析和质谱分析简单鉴定了盐穗木提取物中生物碱的种类。
结果表明:盐穗木中含有酚类、鞣质、生物碱、黄酮、糖、多糖及其甙类、氨基酸及多肽、皂苷、有机酸、甾体和三萜类物质、挥发油等多种化学成分。乙醇加热法效果最差,总碱含量仅为0.404mg/g;索氏提取法提取效率次之(0.477mg/g),酸水提取法提取时间短、效率较高(0.520mg/g);超声波提取法效果最好,总碱含量为0.993mg/g。盐穗木特征吸收波长为280nm。进一步优化方法,得到最佳工艺为A2B2C3,即溶媒量为0.5g/mL、溶媒浓度为85%、超声波处理时间为90min。根据抑菌试验结果可知,盐穗木的抑菌活性物质主要在总生物碱的乙酸乙酯相中。盐穗木提取物对枯草芽孢杆菌、大肠杆菌、金黄色葡萄球菌有较强的抑菌作用。乙酸乙酯相对金黄色葡萄球菌和枯草芽孢杆菌的最小抑菌浓度为7.8mg/mL,对大肠埃希杆菌的最小抑菌浓度为15.6mg/mL,说明乙酸乙酯相对金黄色葡萄球菌的抑菌作用要大于对于大肠埃希杆菌的抑菌作用。总浸膏对大肠埃希杆菌、金黄色葡萄球菌和枯草芽孢杆菌的最小抑菌浓度为31.2mg/mL;对链球菌、普通变形杆菌、面包酵母无抑菌作用。通过对盐穗木总生物碱进行TLC和GC-MS分析,鉴定出9种生物碱类化合物。
Caspica (Halostachyscaspica) is belonging to Chenopodiaceae (Chenopodiaceae) caspica (Halostachys CAMey).It is one of the main plant in the saline desert which is strong adaptability to the salt. Caspica contains complex chemical compositions. Currently,there are little study of antibacterial activity of constituents in the caspica. This thesis aim at caspica alkaloids with a preliminary study, including the alkaloid extraction, purification, structure identification and antimicrobial activities.
Firstly, this study identified the main kind of chemical components in the caspica ; chosen four different extraction methods (acidic water extraction, ethanol heating extraction, ultrasound-assisted extraction and Soxhlet extraction) to extract the total alkaloids and evaluation of extraction efficiency; use ultraviolet scanning to determine the total alkaloids caspica of characteristic absorption wavelength; investigated by orthogonal test solvent volume (A), solvent concentration (B) and ultrasonic treatment time (C) 3 factors on the extraction efficiency; study caspica extract on microbial growth inhibition; use of thin layer chromatography and mass spectrometry simply identified the types of alkaloids in the caspica.
The results showed that: the caspica contains phenols, tannins, alkaloids, flavonoids, sugars, polysaccharides and their glycosides, amino acids and peptides, saponins, organic acids, steroids and triterpenoids, a variety of chemical constituents of volatile oil ,etc. Ethanol heating is the worst, with a total alkali content of only 0.404mg / g; Soxhlet extraction followed by extraction efficiency (0.477mg / g);Acid extraction extraction time, higher efficiency (0.520mg / g); Ultrasonic extraction was the best, with a total alkali content of 0.993mg / g. Caspica characteristic absorption wavelength is 280nm.
Further optimization, the best technology is the A2B2C3, the solvent volume 0.5g/mL, 85% solvent concentration, ultrasonic processing time is 90min.
According to inhibition experiment showed that, caspica antibacterial active substances is mainly in the ethyl acetate phase of total alkaloids. Caspica extract has strong antimicrobial effect to the Bacillus subtilis, Escherichia coli, Staphylococcus aureus. Ethyl acetate relative of Staphylococcus aureus and Bacillus subtilis the minimum inhibitory concentration are 7.8mg/mL, to the Escherichia coli minimum inhibitory concentration is 15.6mg/mL.The result shows that ethyl acetate relative inhibition of Staphylococcus aureus greater than the bacteria of Escherichia coli antibiotic effects. The total extract to the Escherichia coli, Staphylococcus aureus and Bacillus subtilis the minimum inhibitory concentration are 31.2mg/mL; there are no inhibitory effect to the streptococcus, Proteus vulgaris and baker's yeast. Caspica identified nine kinds of alkaloids by TLC of total alkaloids and GC-MS analysis.
引文
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