基于声致化学发光的茶汤抗活性氧能力和药物溶出分析研究
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摘要
近年来,活性氧研究成为现代生命科学的前沿和热点,评价和筛选具有强抗氧化活性的天然产物己成为生物学、医学和食品科学研究的一个重要研究领域。
本论文以声致化学发光方法为基础,利用声能在水溶液中产生活性氧,活性氧氧化鲁米诺产生化学发光;还原性分子或抗氧化剂清除活性氧使鲁米诺发光强度下降,通过测量鲁米诺声致化学发光强度的变化,就可以对还原性分子或抗氧化剂的抗活性氧能力进行评价。该声致化学发光方法具有分析速度快,操作简单、快捷,容易实现自动化等优点.
本论文主要由综述和实验部分组成。第一章综述介绍了活性氧的产生、危害、抗氧化剂的分类及作用及抗氧化能力的评价方法和研究现状。二至五章是实验部分,主要内容如下:
1.将流动技术与声致化学发光方法结合,研究了五种茶叶在不同冲泡水温、冲泡时间及冲泡次数等条件下,茶汤的抗活性氧能力。考察了隔夜茶汤抗活性氧能力的变化。实验结果表明:冲泡茶叶的最佳条件为冲泡水温96℃,冲泡时间为30 min,冲泡次数最多三次。五种茶叶茶汤抗活性氧能力由高到低顺序为紫阳毛尖>乌龙茶>寿眉>祁门红茶>普洱茶。
2.利用流动一声致化学发光方法,以铁观音为研究对象,考察了不同价位的铁观音茶汤抗活性氧能力与价位的相关性。测定了在不同冲泡水温、冲泡时间及冲泡次数等条件下,各茶汤的抗活性氧能力。实验结果表明:在冲泡时间为40 min,冲泡水温为96℃时,首次冲泡获得的茶汤抗活性氧能力最强;在不同冲泡条件下,价位与对应茶汤的抗活性氧能力之间不存在显著相关性。
3.基于流动-声致化学发光方法,以天然产物茶叶,药物维生素C片和异烟肼片为研究对象,进行溶出度研究。实验结果表明:茶叶在10 min内溶出较快,之后溶出平缓,17 min时完全溶出;异烟肼片在10 min内溶出较快,之后溶出平缓,30 min时溶出完全;维生素C片溶出速度较快,在6 min内溶出较快30 min时已完全溶出。流动-声致化学发光方法操作简单快速,分析频率可达356h~(-1)能便捷在线监测抗活性氧物质溶出的全过程。
4.以罗丹明B为母体,合成对金属离子具有选择性络合的荧光探针。试验了罗丹明B内酰肼与钌离子作用产生强的荧光,初步建立了将荧光分析与流动注射技术相结合测定钌离子的分析方法。结果表明,该方法的线性范围为2.0×10~(-6)～1.0×10~(-5)mol/L,检出限为6×10~(-7)mol/L。对3.0×10.6mol/L的钌离子进行11次平行测定,相对标准偏差为2.1%
In the last 20 years, the study on reactive oxygen species has become the hot natural topics ofmodern life science. Appraise and selection of resources with strong antioxidative activities havebecome the important trends of biologymedical science,food science and technology.
Based on the principle of that a strong sonochemiluminescence signal is generated by ROSwhich are produced in the solution of water by ultrasound, but the reactive oxygen species could beeliminated by the antioxidants or reductant,which quench sonochemiluminescence signal. Themethod can be applied to estimate anti-reactive oxygen species (ROS) capacity of antioxidants orreductant. The method can approach high detection frequency, simplicity and automatic operation.
Five chapters are included in this thesis. The first chapter is review, and the rest are researchreports. In the first chapter, a brief review was given on the generation harmness of ROS, sorts ofantioxidant and analytical methods for the determination antioxidation capacity. In the researchreports, main content is as follows:
1. Various factors including extracting temperature, extracting time and times of extraction, thataffect on the anti-reactive oxygen species (ROS) capacity of extracted tea from five kinds tea areinvestigated by the flow-sonochemiluminescence method. Experiment results show that the optimumtea infusing condition are: 96? water, 30 minutes infusing time and three times infusing frequency.Tea can effectively restrain the sonochemiluminescence of luminol induced by reactive oxygenspecies on line. Descending order of extracted tea anti-ROS capcity of studied teas is Zi yang maojian> Oolong tea>Shou mei>Keemun black tea>Pu'er tea.
2. Various factors including extracting temperature, extracting time and times of extraction, thataffect on the anti-reactive oxygen species (ROS) capacity of extracted tea from different price of teasare investigated by the flow-sonochemiluminescence method. Experiment results show: Thestrongest anti-reactive oxygen species ROS of tea can gain at the following conditions: 96°Cinfusing temperature, 40 minutes infusing time and one times infusing frequency; under differentconditions, relationship between the price and the anti-reactive oxygen capacity is no obvious.
3. Based on flow-sonochemiluminescence method,dissolution test of tea, Isoniazd and Vctablets are studied. Experiment results show: The system responds linearly to Isoniazd concentrationin the range of 0.01～1 mg/ml. The system responds logarithm to L-Ascorbic concentration in therange of2.0×10～(-4) mg/ml～1.0×10～(-2) mg/ml. Dissolution testing of substance are described: dissolutionof tea is quick in ten minutes and dissolution is complete at seventeen minutes; dissolution ofisoniazd tablet is relatively quick in ten minutes and dissolution has completed at thirty minutes;isoniazd tablet is relatiVely quick in ten minutes and diSS01utiOll has completed at thirty minutes； dissolution of Vc tablet is very quick in six minutes and dissolution has completed at thirty minutes.Experiment demonstrate that automatic operation and high throughput, in which analysis frequencyis 356 samples h～(1) can be achieved by flow-sonochemiluminescence method. It can determine thedissolution profiles of the tested substances. It can be used as on line dissolution monitoring of thetested substances easily and exhaustivly.
4. This research is based on the triphenylmethane class compound as the parent substance.Fluorescence probe synthesized has the selective to the metallic ion. Rhodamine B hydrazine, acolorless and non-fluorescent substance, gave weak fluorescence. When Ru～(3+) was introduced into asolution of Rhodamine B hydrazine, a strong fluorescence and absorption can be detected. Basedon this, the method of determination of Ru～(3+) by flow-injection analysis was primary established.The system responds linearly to Ru～(3+) concentration in the range of 2.0×～10(-6)～1.O×10～(-5) mol/L withcorrelation coefficient of 0.9983 and a detection of 6×10～(-7) mol/L. The relative standard deviation(RSD) was 2.1% for 3.0×10～(-6) mol/L (n=11).
本论文以声致化学发光方法为基础,利用声能在水溶液中产生活性氧,活性氧氧化鲁米诺产生化学发光;还原性分子或抗氧化剂清除活性氧使鲁米诺发光强度下降,通过测量鲁米诺声致化学发光强度的变化,就可以对还原性分子或抗氧化剂的抗活性氧能力进行评价。该声致化学发光方法具有分析速度快,操作简单、快捷,容易实现自动化等优点.
本论文主要由综述和实验部分组成。第一章综述介绍了活性氧的产生、危害、抗氧化剂的分类及作用及抗氧化能力的评价方法和研究现状。二至五章是实验部分,主要内容如下:
1.将流动技术与声致化学发光方法结合,研究了五种茶叶在不同冲泡水温、冲泡时间及冲泡次数等条件下,茶汤的抗活性氧能力。考察了隔夜茶汤抗活性氧能力的变化。实验结果表明:冲泡茶叶的最佳条件为冲泡水温96℃,冲泡时间为30 min,冲泡次数最多三次。五种茶叶茶汤抗活性氧能力由高到低顺序为紫阳毛尖>乌龙茶>寿眉>祁门红茶>普洱茶。
2.利用流动一声致化学发光方法,以铁观音为研究对象,考察了不同价位的铁观音茶汤抗活性氧能力与价位的相关性。测定了在不同冲泡水温、冲泡时间及冲泡次数等条件下,各茶汤的抗活性氧能力。实验结果表明:在冲泡时间为40 min,冲泡水温为96℃时,首次冲泡获得的茶汤抗活性氧能力最强;在不同冲泡条件下,价位与对应茶汤的抗活性氧能力之间不存在显著相关性。
3.基于流动-声致化学发光方法,以天然产物茶叶,药物维生素C片和异烟肼片为研究对象,进行溶出度研究。实验结果表明:茶叶在10 min内溶出较快,之后溶出平缓,17 min时完全溶出;异烟肼片在10 min内溶出较快,之后溶出平缓,30 min时溶出完全;维生素C片溶出速度较快,在6 min内溶出较快30 min时已完全溶出。流动-声致化学发光方法操作简单快速,分析频率可达356h~(-1)能便捷在线监测抗活性氧物质溶出的全过程。
4.以罗丹明B为母体,合成对金属离子具有选择性络合的荧光探针。试验了罗丹明B内酰肼与钌离子作用产生强的荧光,初步建立了将荧光分析与流动注射技术相结合测定钌离子的分析方法。结果表明,该方法的线性范围为2.0×10~(-6)～1.0×10~(-5)mol/L,检出限为6×10~(-7)mol/L。对3.0×10.6mol/L的钌离子进行11次平行测定,相对标准偏差为2.1%
In the last 20 years, the study on reactive oxygen species has become the hot natural topics ofmodern life science. Appraise and selection of resources with strong antioxidative activities havebecome the important trends of biologymedical science,food science and technology.
Based on the principle of that a strong sonochemiluminescence signal is generated by ROSwhich are produced in the solution of water by ultrasound, but the reactive oxygen species could beeliminated by the antioxidants or reductant,which quench sonochemiluminescence signal. Themethod can be applied to estimate anti-reactive oxygen species (ROS) capacity of antioxidants orreductant. The method can approach high detection frequency, simplicity and automatic operation.
Five chapters are included in this thesis. The first chapter is review, and the rest are researchreports. In the first chapter, a brief review was given on the generation harmness of ROS, sorts ofantioxidant and analytical methods for the determination antioxidation capacity. In the researchreports, main content is as follows:
1. Various factors including extracting temperature, extracting time and times of extraction, thataffect on the anti-reactive oxygen species (ROS) capacity of extracted tea from five kinds tea areinvestigated by the flow-sonochemiluminescence method. Experiment results show that the optimumtea infusing condition are: 96? water, 30 minutes infusing time and three times infusing frequency.Tea can effectively restrain the sonochemiluminescence of luminol induced by reactive oxygenspecies on line. Descending order of extracted tea anti-ROS capcity of studied teas is Zi yang maojian> Oolong tea>Shou mei>Keemun black tea>Pu'er tea.
2. Various factors including extracting temperature, extracting time and times of extraction, thataffect on the anti-reactive oxygen species (ROS) capacity of extracted tea from different price of teasare investigated by the flow-sonochemiluminescence method. Experiment results show: Thestrongest anti-reactive oxygen species ROS of tea can gain at the following conditions: 96°Cinfusing temperature, 40 minutes infusing time and one times infusing frequency; under differentconditions, relationship between the price and the anti-reactive oxygen capacity is no obvious.
3. Based on flow-sonochemiluminescence method,dissolution test of tea, Isoniazd and Vctablets are studied. Experiment results show: The system responds linearly to Isoniazd concentrationin the range of 0.01～1 mg/ml. The system responds logarithm to L-Ascorbic concentration in therange of2.0×10～(-4) mg/ml～1.0×10～(-2) mg/ml. Dissolution testing of substance are described: dissolutionof tea is quick in ten minutes and dissolution is complete at seventeen minutes; dissolution ofisoniazd tablet is relatively quick in ten minutes and dissolution has completed at thirty minutes;isoniazd tablet is relatiVely quick in ten minutes and diSS01utiOll has completed at thirty minutes； dissolution of Vc tablet is very quick in six minutes and dissolution has completed at thirty minutes.Experiment demonstrate that automatic operation and high throughput, in which analysis frequencyis 356 samples h～(1) can be achieved by flow-sonochemiluminescence method. It can determine thedissolution profiles of the tested substances. It can be used as on line dissolution monitoring of thetested substances easily and exhaustivly.
4. This research is based on the triphenylmethane class compound as the parent substance.Fluorescence probe synthesized has the selective to the metallic ion. Rhodamine B hydrazine, acolorless and non-fluorescent substance, gave weak fluorescence. When Ru～(3+) was introduced into asolution of Rhodamine B hydrazine, a strong fluorescence and absorption can be detected. Basedon this, the method of determination of Ru～(3+) by flow-injection analysis was primary established.The system responds linearly to Ru～(3+) concentration in the range of 2.0×～10(-6)～1.O×10～(-5) mol/L withcorrelation coefficient of 0.9983 and a detection of 6×10～(-7) mol/L. The relative standard deviation(RSD) was 2.1% for 3.0×10～(-6) mol/L (n=11).
引文
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