超临界CO_2萃取—超滤浓缩技术提取分离鹿茸中有效成分的研究
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摘要
鹿茸是传统名贵中药,具有温肾壮阳、益精补血、强健筋骨等功效。鹿茸中含有多种重要的活性成分,其中次黄嘌呤等生物碱基、甾体化合物、磷脂、胆固醇等具有抑制单胺氧化酶(MAO)的抗衰老作用;胰岛素样生长因子-1(IGF-1)可促进生长发育,尤其对糖尿病及其并发症有特殊的疗效。因此采用先进技术提取分离鹿茸中的有效部位对科学利用中国珍贵中药鹿茸资源及促进中药现代化具有重要意义。
本文在课题组前期研究的基础上,基于鹿茸综合利用的角度,首次以鹿茸冻干粉为原料,考察了超临界CO_2萃取技术提取鹿茸中MAO抑制剂的工艺条件,优化结果表明,采用萃取压力35MPa和萃取温度35℃的工艺条件,MAO抑制剂提取率可以达到3.92%。
萃取物对MAO-B活性具有明显的抑制作用,随萃取物浓度增大,抑制作用增强,呈明显的量效关系。在优化条件下得到的萃取物,当其磷酸盐溶液浓度为238.15 mg/L时,对MAO-B的抑制作用达到94.87%;而对MAO-A的抑制作用只有18.92%;表现出萃取物对MAO抑制具有一定的选择性。
利用GC-MS法对萃取物中的化学组成进行了分析,结果表明,萃取物中含有29种具有重要生物活性的饱和脂肪酸、不饱和脂肪酸、甾体化合物和胆固醇等物质。
本文还以超临界萃余物为原料,运用碱液超声法提取其中的IGF-1和蛋白质,结果表明,采用萃取压力35MPa、萃取温度不超过40℃的工艺条件,萃余物中IGF-1和蛋白质提取率较高。在上述优化条件下得到的萃余物,IGF-1提取率可以达到20.86μg/g,蛋白质提取率达到130.73mg/g。与鹿茸冻干粉直接进行碱液超声提取相比,用超临界萃余物作原料对IGF-1和蛋白质的提取效果更好。
采用超滤技术对IGF-1的超滤浓缩工艺进行了考察,以PES(聚醚砜)膜和改性PES膜进行对比实验,通过考察截留分子量、压力、pH、浓缩倍数等因素对鹿茸冻干粉超声提取液超滤浓缩过程的影响,得出适宜的工艺条件,在此条件下,PES膜IGF-1回收率达到68.32%,蛋白质回收率达到69.20%;改性PES膜IGF-1回收率达到57.17%,蛋白质回收率达到87.43%。
采用四种不同的方法对超滤膜的清洗方法进行了考察,结果表明,超滤膜在4‰NaOH-1‰NaClO溶液中浸泡24h,PES膜水通量恢复率达到98.65%,改性PES膜水通量恢复率达到93.75%,膜通量接近完全恢复。
Antler velvet, as a kind of traditional Chinese medicine, has long been recognized as one of the most effective and powerful invigorants for strengthening the kidney, sexual-reinforcing and pro-longing life. Many substances from antler velvet were identified and claimed to contain active components. Among these components, some biological bases(e.g. hypoxanthine), steroidal compounds, phospholipid and cholesterol have inhibitory effect on monoamine oxidase(MAO); insulin-like growth factor-1(IGF-1), which can promote the body's growth and development, has special effect on diabetes mellitus and its complications. Therefore, it is important to extract and separate active components from antler velvet by advanced separation technology for promoting the modernization of Chinese traditional medicine production.
Based on the previous research results in our laboratory and the idea of comprehensive utilization, supercritical CO_2 extraction of MAO inhibitor was firstly investigated with antler velvet freeze-dried powder as raw material. The optimum results show that the extraction yield can reach 3.92% under pressure of 35MPa and temperature of 35℃.
The extracts of antler velvet obtaind by supercritical fluid (EAS) inhibit MAO-B activity in an apparently dose-dependent manner. The activity of MAO-B and MAO-A were inhibited up to 94.87% and 18.92% respectively at 238.15mg/L EAS phosphate solution, which shows that EAS has certain inhibition selectivity on MAO.
The chemical composition of EAS was analyzed by GC-MS. The analytical results show that there are 29 kinds of important active components in EAS, including saturated fatty acids, unsaturated fatty acids, steroids and cholesterol.
Ultrasonic extraction was further used to extract IGF-1 and protein from the residues of SFE. The experimental results show that the extraction yields from residues can reach a higher degree under pressure of 35MPa and temperature not exceeding 40℃. The extraction yields of IGF-1 and protein from residue can reach 20.86μg/g and 130.73mg/g respectively at the optimal condition. The extraction yields of IGF-1 and protein with ultrasonic extraction from SFE residue are higher than those from antler velvet freeze-dried powder directly.
Polyethersulfone (PES) membrane and modified PES membrane were used for further concentration. The effect of MWCO, pressure, pH and concentration rate on IGF-1 ultrafiltration process was investigated, and the optimal conditions were obtained. The recovery rate of IGF-1 and protein can reach 68.32% and 69.20% respectively by PES membrane, while the recovery rate of IGF-1 and protein can reach 57.17% and 87.43% respectively by modified PES membrane.
Different methods were employed to clean membrane. The results show that membrane immersed in 4‰NaOH-1‰NaClO solution for 24h is the appropriate cleaning method, in which PES membrane water flux recovery rate can reach 98.65% and modified PES membrane water flux recovery rate can reach 93.75%.
本文在课题组前期研究的基础上,基于鹿茸综合利用的角度,首次以鹿茸冻干粉为原料,考察了超临界CO_2萃取技术提取鹿茸中MAO抑制剂的工艺条件,优化结果表明,采用萃取压力35MPa和萃取温度35℃的工艺条件,MAO抑制剂提取率可以达到3.92%。
萃取物对MAO-B活性具有明显的抑制作用,随萃取物浓度增大,抑制作用增强,呈明显的量效关系。在优化条件下得到的萃取物,当其磷酸盐溶液浓度为238.15 mg/L时,对MAO-B的抑制作用达到94.87%;而对MAO-A的抑制作用只有18.92%;表现出萃取物对MAO抑制具有一定的选择性。
利用GC-MS法对萃取物中的化学组成进行了分析,结果表明,萃取物中含有29种具有重要生物活性的饱和脂肪酸、不饱和脂肪酸、甾体化合物和胆固醇等物质。
本文还以超临界萃余物为原料,运用碱液超声法提取其中的IGF-1和蛋白质,结果表明,采用萃取压力35MPa、萃取温度不超过40℃的工艺条件,萃余物中IGF-1和蛋白质提取率较高。在上述优化条件下得到的萃余物,IGF-1提取率可以达到20.86μg/g,蛋白质提取率达到130.73mg/g。与鹿茸冻干粉直接进行碱液超声提取相比,用超临界萃余物作原料对IGF-1和蛋白质的提取效果更好。
采用超滤技术对IGF-1的超滤浓缩工艺进行了考察,以PES(聚醚砜)膜和改性PES膜进行对比实验,通过考察截留分子量、压力、pH、浓缩倍数等因素对鹿茸冻干粉超声提取液超滤浓缩过程的影响,得出适宜的工艺条件,在此条件下,PES膜IGF-1回收率达到68.32%,蛋白质回收率达到69.20%;改性PES膜IGF-1回收率达到57.17%,蛋白质回收率达到87.43%。
采用四种不同的方法对超滤膜的清洗方法进行了考察,结果表明,超滤膜在4‰NaOH-1‰NaClO溶液中浸泡24h,PES膜水通量恢复率达到98.65%,改性PES膜水通量恢复率达到93.75%,膜通量接近完全恢复。
Antler velvet, as a kind of traditional Chinese medicine, has long been recognized as one of the most effective and powerful invigorants for strengthening the kidney, sexual-reinforcing and pro-longing life. Many substances from antler velvet were identified and claimed to contain active components. Among these components, some biological bases(e.g. hypoxanthine), steroidal compounds, phospholipid and cholesterol have inhibitory effect on monoamine oxidase(MAO); insulin-like growth factor-1(IGF-1), which can promote the body's growth and development, has special effect on diabetes mellitus and its complications. Therefore, it is important to extract and separate active components from antler velvet by advanced separation technology for promoting the modernization of Chinese traditional medicine production.
Based on the previous research results in our laboratory and the idea of comprehensive utilization, supercritical CO_2 extraction of MAO inhibitor was firstly investigated with antler velvet freeze-dried powder as raw material. The optimum results show that the extraction yield can reach 3.92% under pressure of 35MPa and temperature of 35℃.
The extracts of antler velvet obtaind by supercritical fluid (EAS) inhibit MAO-B activity in an apparently dose-dependent manner. The activity of MAO-B and MAO-A were inhibited up to 94.87% and 18.92% respectively at 238.15mg/L EAS phosphate solution, which shows that EAS has certain inhibition selectivity on MAO.
The chemical composition of EAS was analyzed by GC-MS. The analytical results show that there are 29 kinds of important active components in EAS, including saturated fatty acids, unsaturated fatty acids, steroids and cholesterol.
Ultrasonic extraction was further used to extract IGF-1 and protein from the residues of SFE. The experimental results show that the extraction yields from residues can reach a higher degree under pressure of 35MPa and temperature not exceeding 40℃. The extraction yields of IGF-1 and protein from residue can reach 20.86μg/g and 130.73mg/g respectively at the optimal condition. The extraction yields of IGF-1 and protein with ultrasonic extraction from SFE residue are higher than those from antler velvet freeze-dried powder directly.
Polyethersulfone (PES) membrane and modified PES membrane were used for further concentration. The effect of MWCO, pressure, pH and concentration rate on IGF-1 ultrafiltration process was investigated, and the optimal conditions were obtained. The recovery rate of IGF-1 and protein can reach 68.32% and 69.20% respectively by PES membrane, while the recovery rate of IGF-1 and protein can reach 57.17% and 87.43% respectively by modified PES membrane.
Different methods were employed to clean membrane. The results show that membrane immersed in 4‰NaOH-1‰NaClO solution for 24h is the appropriate cleaning method, in which PES membrane water flux recovery rate can reach 98.65% and modified PES membrane water flux recovery rate can reach 93.75%.
引文
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