CN107281463A - A kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99% - Google Patents
A kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99% Download PDFInfo
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Abstract
The invention discloses a kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99%, comprise the following steps：Prepare free radical scavenger, ensure radicals scavenging agent concentration, screening free radical scavenger solvent and intake free radical scavenger solvent；The present invention is played a role necessary three conditions by meeting free radical scavenger, i.e., and first, free radical scavenger will have certain concentration；Second, because free radical activity is extremely strong, once producing will work with neighbouring life macromolecule at once, so free radical scavenger must be near free radical, and it can at a terrific speed try to be the first and combined with free radical；3rd, in most cases, new free radical can be become after scavenger and radical reaction, the toxicity that the toxicity of this new free radical should be less than original free radical just has defense reaction；And then 99% free radical generating source can be removed, so as to reach defence disease, the purpose of anti-aging.
Description
Technical field
The present invention relates to radicals scavenging technical field, specially it is a kind of can remove 99% free radical generating source it is comprehensive
Mould assembly radicals scavenging method.
Background technology
Free radical is the material with elevated chemical activity, is the intermediate product of a variety of biochemical reactions in vital movement, people
Vital movement be unable to do without free radical, but interior free yl is excessive or removes slow, and free radical can then be attacked and damage macromolecular,
Damage is caused to cell membrane, nucleic acid and body protein etc., is the basic reason for causing body aging, is also to induce malignant tumour
When the important cause of many diseases.The oxidant that radical reduction is non-free radical can be referred to as free radical scavenger.Freely
Base scavenger (FRS) refers to the material with delay, suppression and blocking ROS (active oxygen)/OFR (oxygen radical) oxidative damage
General name, is the body protective agent that can be combined and be allowed to remove with OFR.Therefore, in body normal processes and protection cell
With tissue from playing an important roll in oxidative damage；Free radical scavenger keeps strong to the normal activities of maintenance body
Health plays an important role.But.With advancing age, the ability that free radical is produced in body is gradually reduced, so as to weaken
To the defence capability of free radical damage, body tissue organ is easily damaged, accelerate the aging of body, trigger a series of
Disease.
So, a kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99% how is designed, is turned into
The problem of we will currently solve.
The content of the invention
The present invention provides a kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99%, by full
Sufficient free radical scavenger plays a role necessary three conditions, i.e., and first, free radical scavenger will have certain concentration；Second,
Because free radical activity is extremely strong, once producing will work with neighbouring life macromolecule at once, so radicals scavenging
Agent must be near free radical, and can at a terrific speed try to be the first and combined with free radical；3rd, in most cases, clearly
Except that can become new free radical after agent and radical reaction, the toxicity of this new free radical should be less than the toxicity of original free radical
Just there is defense reaction；And then 99% free radical generating source can be removed, and disease, the purpose of anti-aging are defendd so as to reach, can
The problem of effectively to solve in above-mentioned background technology.
To achieve the above object, the present invention provides following technical scheme：It is a kind of to remove 99% free radical generating source
Comprehensive radicals scavenging method, comprises the following steps：
1）Prepare free radical scavenger：The fresh ginkgo nuts of 1-2kg are taken to divest pulp outer layer duricrust, by except the ginkgo nut after shell and going
Ionized water by volume 1：1 mixed after be put into refiner and crush homogenate, measure 200-250mL it is broken after ginkgo nut it is even
Slurry is fitted into 1L beakers, then adds the phosphate buffer that 2-3 times of pH of homogenate volume is 7.6 into beaker, is stirred with glass bar
Uniformly, mixed liquor is obtained；Supernatant is prepared again：Mixed liquor after above-mentioned stir is moved into Soxhlet extraction device and extracts 3-5h,
Measure 300-400mL extracts and 100-150mL concentration 0.5mol/L ammonium sulfates are placed on magnetic stirrer and stir equal
Move into ice-water bath, staticly settled overnight at 4-6 DEG C after even, then centrifuged with horizontal centrifuge with 5000-7000r/min rotating speeds
Isolated supernatant；Finally prepare ginkgo nut extract：Supernatant obtained above is fitted into bag filter, flowed with 1-2L
Absolute ethyl alcohol as dialyzate, circulation dialysis 12-15h is carried out to supernatant, the absolute ethyl alcohol after circulation dialysis is collected and is placed on
In rotary evaporator, the 1/10 of original volume is reduced in 60-70 DEG C of backspin inspissation, obtains concentrating ginkgo nut extract；Weigh 400-
It is 1 that 500g gen-seng haulms, which are put into after airslide disintegrating mill is crushed by solid-to-liquid ratio,:3 mix cauline leaf powder and n-butanol, are fallen after mixing
Enter in ultrasonic abstraction instrument, ultrasonic extraction 1-2h, is separated by filtration and obtains extract under 100-200W power, by gained extract mistake
Large pore resin absorption column, then polymeric adsorbent is washed with the absolute ethyl alcohol of mass concentration 75% of 5-8 times of macroporous absorbent resin volume
Take off and collect eluent；The above-mentioned eluent being collected into is put into distilling apparatus, ethanol is distilled to recover at 60-70 DEG C, obtains
To panaxoside, by 100-200mL panaxosides and 200-300mL vitamin Es, vitamin C, beta carotene, trace element
Selenium, the mixed liquor of glutathione are mixed, and obtain mixing free radical scavenger, are placed in brown bottle and are preserved.
2）Ensure radicals scavenging agent concentration：By step 1）In obtained free radical scavenger according to circumstances configure certain dense
The free radical scavenger solvent of degree；
3）Screen free radical scavenger solvent：To step 2）In obtained free radical scavenger solvent screened, most of
In the case of, can become new free radical after scavenger and radical reaction, the toxicity of this new free radical should be less than it is original from
Just there is defense reaction by the toxicity of base, the small free radical scavenger solvent of new free radical toxicity is produced so needing to screen；
4）Take in free radical scavenger solvent：, at once will be with neighbouring life once producing because free radical activity is extremely strong
Macromolecular works, so free radical scavenger must be taken near free radical, speed that thus can and can be to be exceedingly fast
Degree is tried to be the first to be combined with free radical, and then can remove 99% free radical.
According to above-mentioned technical proposal, the extracting method of the glutathione is boiling water bath extraction method：By yeast cells with going
Ionized water mixing is made into the yeast bacteria suspension of various concentrations (/V) loaded in test tube, and then test tube is positioned in boiling water bath and heated
For a period of time, be put in rapidly in ice bath cooling can or microwave―assisted extraction：Yeast cells is mixed with deionized water and is made into
The yeast bacteria suspension of various concentrations (/V) is contained in beaker, and then beaker is positioned in micro-wave oven, is 700W in power output
Under conditions of carry out microwave radiation exaraction, take out afterwards it is rapid be put in ice bath it is cold go Ij can or ultrasonic fragmentation：By ferment
Mother cell is mixed with deionized water to be made into the yeast bacteria suspensions of various concentrations (/V) and is contained in beaker, is then positioned over beaker super
In sonicated sound proof box, adjustment pallet height makes 1/2 liquid height below probe immersed in liquid level, and beaker is placed in into ice bath groove
In, it is 800W in power output, interval time takes out for progress under conditions of 2s/lS and caught.
According to above-mentioned technical proposal, the step 2）In free radical scavenger solvent in include Momordia grosvenori aglycone.
According to above-mentioned technical proposal, the preparation method of the Momordia grosvenori aglycone is：First dissolve the Fructus Monordicae extract of Momordica grosvenori
Solubilizer dissolves, and the concentration for making the Momordia grosvenori aglycone in solution is 0.025-0.300g/mL；Hydrolyze again：By in above-mentioned resulting solution
Beta-glucosidase enzyme hydrolysis is added, and adjusts enzymolysis pH value and temperature that the pH value and temperature of solution make it with beta-glucosidase
It is adapted；It is finally separating, purifies, that is, obtains Momordia grosvenori aglycone.
According to above-mentioned technical proposal, the step 2）Temperature during operation is room temperature.
According to above-mentioned technical proposal, the step 3）Middle screening free radical scavenger solvent is exactly to screen radicals scavenging
Agent, by test sample solution point sample on lamellae, volatilizing solvent or/and use exhibition specially after free radical scavenger is determined
Solvent is volatilized after opening agent expansion；Lamellae after processing is impregnated in the substrate solution formed by Fe2+- developers-complexing agent
In；Take out lamellae and wipe its excess surface drop away, then reacted 5-20 minutes under 30-45 °C；Described developer is certainly
The reagent that it fades can be made by base；Reacted lamellae is impregnated in H2O2 reaction solutions；Take out lamellae and wipe its table away
Face residual droplets, then react 20-40 minutes under 40-55 °C；The colour developing situation of lamellae is observed under visible light：Occur
It is the spot as free radical scavenger of the developer intrinsic colour under white background.
According to above-mentioned technical proposal, the lamellae selects silica gel plate, polyamide board or cellulose plate.
Compared with prior art, beneficial effects of the present invention：The present invention must by meeting that free radical scavenger plays a role
Three conditions of palpus, i.e., first, free radical scavenger will have certain concentration；Second, because free radical activity is extremely strong, once
Producing will work with neighbouring life macromolecule at once, so free radical scavenger must be near free radical, and energy
Try to be the first and combined with free radical at a terrific speed；3rd, in most cases, scavenger is with that can become new after radical reaction
Free radical, the toxicity that the toxicity of this new free radical should be less than original free radical just has defense reaction；And then can remove
99% free radical generating source, so as to reach defence disease, the purpose of anti-aging.
Brief description of the drawings
Accompanying drawing is used for providing a further understanding of the present invention, and constitutes a part for specification, the reality with the present invention
Applying example is used to explain the present invention together, is not construed as limiting the invention.
In the accompanying drawings：
Fig. 1 is the preparation flow figure of the present invention.
Embodiment
The preferred embodiments of the present invention are illustrated below in conjunction with accompanying drawing, it will be appreciated that preferred reality described herein
Apply example to be merely to illustrate and explain the present invention, be not intended to limit the present invention.
Embodiment：As shown in figure 1, the present invention provides a kind of comprehensive freedom for the free radical generating source that can remove 99%
Base sweep-out method, comprises the following steps：
1）Prepare free radical scavenger：The fresh ginkgo nuts of 1-2kg are taken to divest pulp outer layer duricrust, by except the ginkgo nut after shell and going
Ionized water by volume 1：1 mixed after be put into refiner and crush homogenate, measure 200-250mL it is broken after ginkgo nut it is even
Slurry is fitted into 1L beakers, then adds the phosphate buffer that 2-3 times of pH of homogenate volume is 7.6 into beaker, is stirred with glass bar
Uniformly, mixed liquor is obtained；Supernatant is prepared again：Mixed liquor after above-mentioned stir is moved into Soxhlet extraction device and extracts 3-5h,
Measure 300-400mL extracts and 100-150mL concentration 0.5mol/L ammonium sulfates are placed on magnetic stirrer and stir equal
Move into ice-water bath, staticly settled overnight at 4-6 DEG C after even, then centrifuged with horizontal centrifuge with 5000-7000r/min rotating speeds
Isolated supernatant；Finally prepare ginkgo nut extract：Supernatant obtained above is fitted into bag filter, flowed with 1-2L
Absolute ethyl alcohol as dialyzate, circulation dialysis 12-15h is carried out to supernatant, the absolute ethyl alcohol after circulation dialysis is collected and is placed on
In rotary evaporator, the 1/10 of original volume is reduced in 60-70 DEG C of backspin inspissation, obtains concentrating ginkgo nut extract；Weigh 400-
It is 1 that 500g gen-seng haulms, which are put into after airslide disintegrating mill is crushed by solid-to-liquid ratio,:3 mix cauline leaf powder and n-butanol, are fallen after mixing
Enter in ultrasonic abstraction instrument, ultrasonic extraction 1-2h, is separated by filtration and obtains extract under 100-200W power, by gained extract mistake
Large pore resin absorption column, then polymeric adsorbent is washed with the absolute ethyl alcohol of mass concentration 75% of 5-8 times of macroporous absorbent resin volume
Take off and collect eluent；The above-mentioned eluent being collected into is put into distilling apparatus, ethanol is distilled to recover at 60-70 DEG C, obtains
To panaxoside, by 100-200mL panaxosides and 200-300mL vitamin Es, vitamin C, beta carotene, trace element
Selenium, the mixed liquor of glutathione are mixed, and obtain mixing free radical scavenger, are placed in brown bottle and are preserved.
2）Ensure radicals scavenging agent concentration：By step 1）In obtained free radical scavenger according to circumstances configure certain dense
The free radical scavenger solvent of degree；
3）Screen free radical scavenger solvent：To step 2）In obtained free radical scavenger solvent screened, most of
In the case of, can become new free radical after scavenger and radical reaction, the toxicity of this new free radical should be less than it is original from
Just there is defense reaction by the toxicity of base, the small free radical scavenger solvent of new free radical toxicity is produced so needing to screen；
4）Take in free radical scavenger solvent：, at once will be with neighbouring life once producing because free radical activity is extremely strong
Macromolecular works, so free radical scavenger must be taken near free radical, speed that thus can and can be to be exceedingly fast
Degree is tried to be the first to be combined with free radical, and then can remove 99% free radical.
According to above-mentioned technical proposal, the extracting method of the glutathione is boiling water bath extraction method：By yeast cells with going
Ionized water mixing is made into the yeast bacteria suspension of various concentrations (/V) loaded in test tube, and then test tube is positioned in boiling water bath and heated
For a period of time, be put in rapidly in ice bath cooling can or microwave―assisted extraction：Yeast cells is mixed with deionized water and is made into
The yeast bacteria suspension of various concentrations (/V) is contained in beaker, and then beaker is positioned in micro-wave oven, is 700W in power output
Under conditions of carry out microwave radiation exaraction, take out afterwards it is rapid be put in ice bath it is cold go Ij can or ultrasonic fragmentation：By ferment
Mother cell is mixed with deionized water to be made into the yeast bacteria suspensions of various concentrations (/V) and is contained in beaker, is then positioned over beaker super
In sonicated sound proof box, adjustment pallet height makes 1/2 liquid height below probe immersed in liquid level, and beaker is placed in into ice bath groove
In, it is 800W in power output, interval time takes out for progress under conditions of 2s/lS and caught.
According to above-mentioned technical proposal, the step 2）In free radical scavenger solvent in include Momordia grosvenori aglycone.
According to above-mentioned technical proposal, the preparation method of the Momordia grosvenori aglycone is：First dissolve the Fructus Monordicae extract of Momordica grosvenori
Solubilizer dissolves, and the concentration for making the Momordia grosvenori aglycone in solution is 0.025-0.300g/mL；Hydrolyze again：By in above-mentioned resulting solution
Beta-glucosidase enzyme hydrolysis is added, and adjusts enzymolysis pH value and temperature that the pH value and temperature of solution make it with beta-glucosidase
It is adapted；It is finally separating, purifies, that is, obtains Momordia grosvenori aglycone.
According to above-mentioned technical proposal, the step 2）Temperature during operation is room temperature.
According to above-mentioned technical proposal, the step 3）Middle screening free radical scavenger solvent is exactly to screen radicals scavenging
Agent, by test sample solution point sample on lamellae, volatilizing solvent or/and use exhibition specially after free radical scavenger is determined
Solvent is volatilized after opening agent expansion；Lamellae after processing is impregnated in the substrate solution formed by Fe2+- developers-complexing agent
In；Take out lamellae and wipe its excess surface drop away, then reacted 5-20 minutes under 30-45 °C；Described developer is certainly
The reagent that it fades can be made by base；Reacted lamellae is impregnated in H2O2 reaction solutions；Take out lamellae and wipe its table away
Face residual droplets, then react 20-40 minutes under 40-55 °C；The colour developing situation of lamellae is observed under visible light：Occur
It is the spot as free radical scavenger of the developer intrinsic colour under white background.
According to above-mentioned technical proposal, the lamellae selects silica gel plate, polyamide board or cellulose.
Based on above-mentioned, it is an advantage of the current invention that played a role necessary three conditions by meeting free radical scavenger,
I.e. first, free radical scavenger will have certain concentration；Second, at once will once producing because free radical activity is extremely strong
Worked with neighbouring life macromolecule, so free radical scavenger must be near free radical, and can be at a terrific speed
Try to be the first and combined with free radical；3rd, in most cases, new free radical can be become after scavenger and radical reaction, this
The toxicity that the toxicity of individual new free radical should be less than original free radical just has defense reaction；And then 99% free radical can be removed
Generating source, so as to reach defence disease, the purpose of anti-aging.
Finally it should be noted that：The preferred embodiment of the present invention is the foregoing is only, is not intended to limit the invention, to the greatest extent
The present invention is described in detail with reference to the foregoing embodiments for pipe, for those skilled in the art, and it still can be with
Technical scheme described in foregoing embodiments is modified, or equivalent substitution is carried out to which part technical characteristic.It is all
Within the spirit and principles in the present invention, any modification, equivalent substitution and improvements made etc., should be included in the guarantor of the present invention
Within the scope of shield.
Claims (5)
1. a kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99%, it is characterised in that：Including as follows
Step：
1）Prepare free radical scavenger：The fresh ginkgo nuts of 1-2kg are taken to divest pulp outer layer duricrust, by except the ginkgo nut after shell and going
Ionized water by volume 1：1 mixed after be put into refiner and crush homogenate, measure 200-250mL it is broken after ginkgo nut it is even
Slurry is fitted into 1L beakers, then adds the phosphate buffer that 2-3 times of pH of homogenate volume is 7.6 into beaker, is stirred with glass bar
Uniformly, mixed liquor is obtained；Supernatant is prepared again：Mixed liquor after above-mentioned stir is moved into Soxhlet extraction device and extracts 3-5h,
Measure 300-400mL extracts and 100-150mL concentration 0.5mol/L ammonium sulfates are placed on magnetic stirrer and stir equal
Move into ice-water bath, staticly settled overnight at 4-6 DEG C after even, then centrifuged with horizontal centrifuge with 5000-7000r/min rotating speeds
Isolated supernatant；Finally prepare ginkgo nut extract：Supernatant obtained above is fitted into bag filter, flowed with 1-2L
Absolute ethyl alcohol as dialyzate, circulation dialysis 12-15h is carried out to supernatant, the absolute ethyl alcohol after circulation dialysis is collected and is placed on
In rotary evaporator, the 1/10 of original volume is reduced in 60-70 DEG C of backspin inspissation, obtains concentrating ginkgo nut extract；Weigh 400-
It is 1 that 500g gen-seng haulms, which are put into after airslide disintegrating mill is crushed by solid-to-liquid ratio,:3 mix cauline leaf powder and n-butanol, are fallen after mixing
Enter in ultrasonic abstraction instrument, ultrasonic extraction 1-2h, is separated by filtration and obtains extract under 100-200W power, by gained extract mistake
Large pore resin absorption column, then polymeric adsorbent is washed with the absolute ethyl alcohol of mass concentration 75% of 5-8 times of macroporous absorbent resin volume
Take off and collect eluent；The above-mentioned eluent being collected into is put into distilling apparatus, ethanol is distilled to recover at 60-70 DEG C, obtains
To panaxoside, by 100-200mL panaxosides and 200-300mL vitamin Es, vitamin C, beta carotene, trace element
Selenium, the mixed liquor of glutathione are mixed, and obtain mixing free radical scavenger, are placed in brown bottle and are preserved.
2.2）Ensure radicals scavenging agent concentration：By step 1）In obtained free radical scavenger according to circumstances configure finite concentration
Free radical scavenger solvent；
3）Screen free radical scavenger solvent：To step 2）In obtained free radical scavenger solvent screened, most of
In the case of, can become new free radical after scavenger and radical reaction, the toxicity of this new free radical should be less than it is original from
Just there is defense reaction by the toxicity of base, the small free radical scavenger solvent of new free radical toxicity is produced so needing to screen；
4）Take in free radical scavenger solvent：, at once will be with neighbouring life once producing because free radical activity is extremely strong
Macromolecular works, so free radical scavenger must be taken near free radical, speed that thus can and can be to be exceedingly fast
Degree is tried to be the first to be combined with free radical, and then can remove 99% free radical.
3. a kind of comprehensive radicals scavenging method of free radical generating source that can remove 99% according to claim 1,
It is characterized in that：The extracting method of the glutathione is boiling water bath extraction method：Yeast cells is mixed with deionized water and is made into
The yeast bacteria suspension of various concentrations (/V) is loaded in test tube, and then test tube is positioned in boiling water bath and heats a period of time, rapidly
Be put in ice bath cooling can or microwave―assisted extraction：Yeast cells is mixed with deionized water and is made into various concentrations (/V)
Yeast bacteria suspension is contained in beaker, and then beaker is positioned in micro-wave oven, is carried out under conditions of power output is 700W micro-
Ripple assisted extraction, take out afterwards it is rapid be put in ice bath it is cold go Ij can or ultrasonic fragmentation：By yeast cells and deionization
The yeast bacteria suspension that water mixing is made into various concentrations (/V) is contained in beaker, and beaker then is positioned over into ultrasonication sound proof box
In, adjustment pallet height makes 1/2 liquid height below probe immersed in liquid level, and beaker is placed in ice bath groove, in power output
For 800W, interval time takes out for progress under conditions of 2s/lS and caught.
4. a kind of comprehensive radicals scavenging method of free radical generating source that can remove 99% according to claim 1,
It is characterized in that：The step 2）In free radical scavenger solvent in include Momordia grosvenori aglycone.
5. a kind of comprehensive radicals scavenging method of free radical generating source that can remove 99% according to claim 3,
It is characterized in that：The preparation method of the Momordia grosvenori aglycone is：First the Fructus Monordicae extract solubilizer dissolving of dissolving Momordica grosvenori, makes molten
The concentration of Momordia grosvenori aglycone in liquid is 0.025-0.300g/mL；Hydrolyze again：Beta-glucosidase will be added in above-mentioned resulting solution
Hydrolysis, and adjusting the pH value and temperature of solution is adapted the enzymolysis pH value and temperature of itself and beta-glucosidase；Be finally separating,
Purifying, that is, obtain Momordia grosvenori aglycone.
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| CN201710728673.2A CN107281463A (en) | 2017-08-23 | 2017-08-23 | A kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99% |
| KR1020180097936A KR20190022369A (en) | 2017-08-23 | 2018-08-22 | A comprehensive method of producing a free-radicar that can eliminate 99% of free-radical sources |
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| CN201710728673.2A CN107281463A (en) | 2017-08-23 | 2017-08-23 | A kind of comprehensive radicals scavenging method for the free radical generating source that can remove 99% |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102477455A (en) * | 2010-11-26 | 2012-05-30 | 成都普瑞法科技开发有限公司 | Method for preparing mogroside IV |
| CN105748350A (en) * | 2016-03-04 | 2016-07-13 | 仇颖超 | Method for preparing coated type free-radical scavenger |
| CN106324175A (en) * | 2015-06-24 | 2017-01-11 | 上海中医药大学 | Method for screening hydroxyl free radical scavengers |
-
2017
- 2017-08-23 CN CN201710728673.2A patent/CN107281463A/en active Pending
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2018
- 2018-08-22 KR KR1020180097936A patent/KR20190022369A/en not_active Application Discontinuation
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102477455A (en) * | 2010-11-26 | 2012-05-30 | 成都普瑞法科技开发有限公司 | Method for preparing mogroside IV |
| CN106324175A (en) * | 2015-06-24 | 2017-01-11 | 上海中医药大学 | Method for screening hydroxyl free radical scavengers |
| CN105748350A (en) * | 2016-03-04 | 2016-07-13 | 仇颖超 | Method for preparing coated type free-radical scavenger |
Non-Patent Citations (3)
| Title |
|---|
| 万艳娟等: "功能性甜味剂罗汉果甜苷的生理功能及食品应用研究进展", 《食品与发酵科技》 * |
| 董永胜: "《谷胱甘肽生产技术》", 31 August 2015 * |
| 黄鹏: "自由基和抗氧化营养素", 《中国社区医师》 * |
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