CN107831161A - Small molecule containing acetate is used as the new application of catalyst as class peroxidase - Google Patents
Small molecule containing acetate is used as the new application of catalyst as class peroxidase Download PDFInfo
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- CN107831161A CN107831161A CN201711276406.2A CN201711276406A CN107831161A CN 107831161 A CN107831161 A CN 107831161A CN 201711276406 A CN201711276406 A CN 201711276406A CN 107831161 A CN107831161 A CN 107831161A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N2021/7756—Sensor type
- G01N2021/7759—Dipstick; Test strip
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Abstract
It is used as the new application of catalyst as class peroxidase the invention discloses the small molecule containing acetate, making public for the first time the small molecule containing acetate has class peroxidase activity.The molecular structure of the small molecule containing acetate of the present invention is clear and definite, is readily available, insensitive to heat, and stability is fine, and cheap, is easily commercially exploited.Based on the new application, the present invention also successfully constructs the new method of detection hydrogen peroxide, and is successfully applied in the test paper of detection hydrogen peroxide, glucose, uric acid or cholesterol etc..The class peroxidase activity based on small molecule of the present invention can also be applied to the every field such as food, environment and health, there is extensive and good application prospect.
Description
Technical field
The invention belongs to analogue enztme field, the more particularly to small molecule containing acetate is used as catalysis as class peroxidase
The new application of agent.
Background technology
Horseradish peroxidase, it is by long-term evolution and a kind of caused biocatalyst in nature.It can be
Efficient under conditions of gentle, exclusively catalyzing hydrogen peroxide chemical reaction.Biology biography of the design based on horseradish peroxidase
Sensor is the strategy of current developers' generally use, i.e., can by the specific binding output of antibody and antigen under the auxiliary of enzyme
The electrochemistry and optical signalling of reading, this method are easy to miniaturization and naked eyes readable signal, can meet that people are daily to it
The monitoring of health.As a kind of efficient catalyst, horseradish peroxidase is widely applied to food hygiene, environment
The fields such as protection, legal medical expert.
But horseradish peroxidase is poor to the sensitiveness and stability of heat, and its source is also limited, and these are lacked
Fall into it in terms of biology sensor extensive exploitation and being very restricted property of use.Therefore, new simulation native enzyme is urged
Agent is gradually paid close attention to by people, develops and developed.
The content of the invention
The shortcomings that primary and foremost purpose of the present invention is to overcome prior art and deficiency, there is provided the small molecule conduct containing acetate
Class peroxidase is used as the new application of catalyst.
A further object of the present invention is to provide a kind of small molecule measure content of hydrogen peroxide based on containing acetate or life
The method of thing molecule content.
Another object of the present invention is to provide the new opplication of the described small molecule containing acetate.
The purpose of the present invention is achieved through the following technical solutions：
Small molecule containing acetate is used as the purposes of catalyst as class peroxidase.
The described small molecule containing acetate is acetic acid and/or soluble acetate.
Described soluble acetate is preferably in sodium acetate, potassium acetate, magnesium acetate, calcium acetate, ammonium acetate and barium acetate
One kind or at least two.
Described class peroxidase, refer to the material of catalytic activity for showing peroxidase.Specifically, it is of the invention
Class Catalyzed Synthesis By Peroxidase redox reaction, and using peroxide as electron acceptor, so as to oxidation substrates.
Described substrate is preferably peroxidase substrate, and under the conditions of existing for peroxidase substrate, described contains
The small molecule of acetate can be used as catalyst, the reaction of catalyzing hydrogen peroxide and peroxidase substrate, cause peroxide
Zymolyte produces coloured or luminescent products, content of hydrogen peroxide can be surveyed by determining the coloured or luminescent products of production
It is fixed.
Described hydrogen peroxide can be produced by enzymatic reaction.
When hydrogen peroxide is produced by enzymatic reaction, refer to by biomolecule under specific protein enzyme effect, certain
Under the conditions of caused hydrogen peroxide.Therefore, under the conditions of existing for peroxidase substrate, the small molecule containing acetate
Catalyst can be used as, the reaction of catalyzing hydrogen peroxide and peroxidase substrate, causes peroxidase substrate production coloured
Or luminescent products, described biomolecule content is measured by determining the coloured or luminescent products of production.
Described biomolecule is preferably one kind or at least two in glucose, uric acid and cholesterol.
Described specific proteases are preferably oxidizing ferment；More preferably glucose oxidase, uricase, cholesterol oxygen
Change one kind or at least two in enzyme.
Described peroxidase substrate is preferably 3,3,5,5- tetramethyl benzidines (TMB), 2,2- connection (the 3- second of nitrogen-two
Base-benzothiazole -6- sulfonic acid) di-ammonium salts (ABTS), one kind in o-phenylenediamine or at least two.
Described certain condition can be 5~8,25~40 DEG C of pH.
The condition of the reaction of described hydrogen peroxide and peroxidase substrate is preferably 1~6,10~60 DEG C of pH.
A kind of method based on the small molecule measure content of hydrogen peroxide containing acetate, described method are：With
The described small molecule containing acetate is catalyst, makes the reaction of hydrogen peroxide and peroxidase substrate, produces coloring matter
Or luminescent substance, determine the coloring matter or luminescent substance of generation, so that it is determined that or hydrogen oxide content.
A kind of method based on the small molecule measure biomolecule content containing acetate, described method are：
Make biomolecule oxidation generation hydrogen peroxide under specific protein enzymatic, according to described based on small point containing acetate
The method measure content of hydrogen peroxide of son measure content of hydrogen peroxide, so that it is determined that described biomolecule content.
Application of the described small molecule containing acetate in biological detection or field of biosensors.
The described application in field of biological detection, can be the application in test paper, reagent, kit is prepared；Institute
The test paper stated, reagent, the detection object of kit are preferably one kind or more in hydrogen peroxide, glucose, uric acid, cholesterol
Kind.
Described biology sensor is preferably hydrogen peroxide, glucose, uric acid or cholesterol sensor.
The present invention is had the following advantages relative to prior art and effect：
1. the present invention is based on inventors be surprised to learn that the small molecule containing acetate has a class peroxidase activity, there is provided
Small molecule containing acetate is used as the new application of catalyst as class peroxidase.The molecule knot of small molecule containing acetate
Structure is clear and definite, is readily available, insensitive to heat, and stability is fine, and cheap, is easily commercially exploited.
2. the present invention utilizes the class peroxidase activity of the described small molecule containing acetate, successfully construct and detected
The new method of hydrogen oxide, and it is prepared for detecting the test paper of hydrogen peroxide, glucose, uric acid or cholesterol etc..
It is each that 3. the class peroxidase activity based on small molecule of the present invention can also be applied to food, environment and health etc.
Individual field, is with a wide range of applications.
Brief description of the drawings
Fig. 1 is that the micromolecule catalyst of embodiment 1 is catalyzed peroxide under the conditions of pH 1.0,2.0,3.0,4.0,5.0,6.0
Change hydrogen and the colour developing result photo figure of ABTS reactions.
Fig. 2 is that hydrogen peroxide Test paper made from embodiment 2 develops the color result photo under the hydrogen peroxide of various concentrations
Figure.
Embodiment
With reference to embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention are unlimited
In this.
The measure of the peroxidase activity of embodiment 1
0.82g sodium acetates are dissolved in 10mL water, take 0.1mL above-mentioned solution to be added to 0.9mL differences pH buffering
System (including 0.05mol/L Tris-HCl, 1mmol/L H2O2, 1mmol/L 2,2- connection the (3- ethyl-benzothiazoles-of nitrogen-two
6- sulfonic acid) di-ammonium salts (ABTS)) in.
As a result as shown in figure 1, under the conditions of pH 1.0,2.0,3.0,4.0,5.0,6.0, the small molecule has urges well
Change effect.Color is deeper, represents that catalytic effect is better.
The preparation and application of the hydrogen peroxide Test paper of embodiment 2
(1) maceration extract is prepared：One, 1000mL beakers are taken, 40mL sodium alginate solns (concentration 50g/L) is added, adds
10mL 1% (v/v) Tween 80 solution, add 40mL gelatin solutions (concentration 80g/L), add 5.0 acetic acid of 100mL pH-second
Sour sodium buffer solution (100mM sodium acetates), 10mL KIs (50g/L), 10g polyvinylpyrrolidones, 0.5g light blues, it is fully mixed
It is even, obtain maceration extract；
(2) impregnated paper：The maceration extract that step (1) prepares is poured into glass dish, soaks filter paper, will be unnecessary with glass bar
Solution scrapes off, and is put into 40 DEG C of convection ovens and dries 30 minutes, is pale blue test paper after doing, and produces described hydrogen peroxide detection
Test paper.
As shown in Fig. 2 by various concentrations hydrogen peroxide to be measured (0 μM, 10 μM, 20 μM, 50 μM, 100 μM, 1000 μM,
2000 μM) directly drop on obtained hydrogen peroxide Test paper, different color changes can be shown, with the liter of concentration
There is pale blue in height, color, and green, yellow to brown changes.
The preparation and application of the hydrogen peroxide Test paper of embodiment 3
(1) maceration extract is prepared：One, 1000mL beakers are taken, 40mL sodium alginate solns (concentration 50g/L) is added, adds
10mL 1% (v/v) Tween 80 solution, add 40mL gelatin solutions (concentration 80g/L), add 5.0 acetic acid of 100mL pH-second
Sour potassium buffer solution (100mM potassium acetates), 10mL KIs (50g/L), 10g polyvinylpyrrolidones, 0.5g light blues, it is fully mixed
It is even, obtain maceration extract；
(2) impregnated paper：The maceration extract that step (1) prepares is poured into glass dish, soaks filter paper, will be unnecessary with glass bar
Solution scrapes off, and is put into 40 DEG C of convection ovens and dries 30 minutes, is pale blue test paper after doing, and produces described hydrogen peroxide detection
Test paper.
Various concentrations hydrogen peroxide (0 μM, 10 μM, 20 μM, 50 μM, 100 μM, 1000 μM, 2000 μM) to be measured is dropped in
As a result similar with Fig. 2 on obtained hydrogen peroxide Test paper, concentration difference can show different color changes, with
There is pale blue in the rise of concentration, color, and green, yellow to brown changes.
The preparation and application of the glucose detection test paper of embodiment 4
(1) maceration extract is prepared：One, 1000mL beakers are taken, 40mL sodium alginate solns (concentration 50g/L) is added, adds
10mL 1% (v/v) Tween 80 solution, add 40mL gelatin solutions (concentration 80g/L), add 5.0 acetic acid of 100mL pH-second
Sour potassium buffer solution (100mM potassium acetates), 10mL KIs (concentration 50g/L), 10g polyvinylpyrrolidones, 0.5g light blues,
0.001g glucose oxidases (purchased from Shanghai life work), fully mix, obtain maceration extract；
(2) impregnated paper：The maceration extract that step (1) prepares is poured into glass dish, soaks filter paper, will be unnecessary with glass bar
Solution scrapes off, and is put into 40 DEG C of convection ovens and dries 30 minutes, is pale blue test paper after doing, and produces described glucose detection examination
Paper.
The result of gained is also similar to that embodiment 2, by the glucose to be measured of various concentrations (0 μM, 10 μM, 20 μM, 50 μM,
100 μM, 1000 μM, 2000 μM) directly drop in described glucose detection test paper, different color changes can be shown, with
The rise of concentration, pale blue occurs in color, and green, yellow to brown changes.
The preparation and application of the uric acid Test paper of embodiment 5
(1) maceration extract is prepared：One, 1000mL beakers are taken, 40mL sodium alginate solns (concentration 50g/L) is added, adds
10mL 1% (v/v) Tween 80 solution, add 40mL gelatin solutions (concentration 80g/L), add 5.0 acetic acid of 100mL pH-second
Sour potassium buffer solution (100mM potassium acetates), 10mL KIs (concentration 50g/L), 10g polyvinylpyrrolidones, 0.5g light blues,
0.001g urate oxidases (purchased from Shanghai life work), fully mix, obtain maceration extract；
(2) impregnated paper：The maceration extract that step (1) prepares is poured into glass dish, soaks filter paper, will be unnecessary with glass bar
Solution scrapes off, and is put into 40 DEG C of convection ovens and dries 30 minutes, is pale blue test paper after doing, and produces described uric acid detection examination
Paper.
Uric acid to be determined is dropped on described uric acid Test paper, the result of gained is also similar to that embodiment 2, passes through
The uric acid (0 μM, 10 μM, 20 μM, 50 μM, 100 μM, 1000 μM, 2000 μM) of measure various concentrations can show different colors
Change, with the rise of concentration, there is pale blue in color, and green, yellow to brown changes.
The preparation and application of the cholesterol detection test paper of embodiment 6
(1) maceration extract is prepared：One, 1000mL beakers are taken, 40mL sodium alginate solns (concentration 50g/L) is added, adds
10mL 1% (v/v) Tween 80 solution, add 40mL gelatin solutions (concentration 80g/L), add 5.0 acetic acid of 100mL pH-second
Sour potassium buffer solution (100mM potassium acetates), 10mL KIs (50g/L), 10g polyvinylpyrrolidones, 0.5g light blues, 0.001g courages
Sterol oxidizing ferment (purchased from Shanghai life work), fully mixes, obtains maceration extract；
(2) impregnated paper：The maceration extract that step (1) prepares is poured into glass dish, soaks filter paper, will be unnecessary with glass bar
Solution scrapes off, and is put into 40 DEG C of convection ovens and dries 30 minutes, is pale blue test paper after doing, and produces described cholesterol detection examination
Paper, the result of gained are also similar to that embodiment 2, by the cholesterol of various concentrations (0 μM, 10 μM, 20 μM, 50 μM, 100 μM, 1000
μM, 2000 μM) directly drop on test paper, different color changes can be shown, with the rise of concentration, there is pale blue in color,
Green, yellow to brown change.
Above-described embodiment is the preferable embodiment of the present invention, but embodiments of the present invention are not by above-described embodiment
Limitation, other any Spirit Essences without departing from the present invention with made under principle change, modification, replacement, combine, simplification,
Equivalent substitute mode is should be, is included within protection scope of the present invention.
Claims (10)
1. the small molecule containing acetate is used as the purposes of catalyst as class peroxidase.
2. the small molecule according to claim 1 containing acetate is used as the purposes of catalyst as class peroxidase, its
It is characterised by：
The described small molecule containing acetate is acetic acid and/or soluble acetate.
3. the small molecule according to claim 2 containing acetate is used as the purposes of catalyst as class peroxidase, its
It is characterised by：
Described soluble acetate be sodium acetate, potassium acetate, magnesium acetate, calcium acetate, ammonium acetate and barium acetate in one kind or
At least two.
A kind of 4. method based on the small molecule measure content of hydrogen peroxide containing acetate, it is characterised in that described
Method is：
Using the small molecule containing acetate as catalyst, make the reaction of hydrogen peroxide and peroxidase substrate, have
Color substance or luminescent substance, determine the coloring matter or luminescent substance of generation, so that it is determined that or hydrogen oxide content.
5. the method according to claim 4 based on the small molecule measure content of hydrogen peroxide containing acetate, its
It is characterised by：
Described hydrogen peroxide can be by caused by enzymatic reaction.
6. the method according to claim 4 based on the small molecule measure content of hydrogen peroxide containing acetate, its
It is characterised by：
The condition of the reaction of described hydrogen peroxide and peroxidase substrate is preferably pH 1~6.
A kind of 7. method based on the small molecule measure biomolecule content containing acetate, it is characterised in that described
Method is：
Make biomolecule oxidation generation hydrogen peroxide under albumen enzymatic, it is according to claim 4 to be contained based on described
The method measure content of hydrogen peroxide of the small molecule measure content of hydrogen peroxide of acetate, so that it is determined that described biomolecule contains
Amount.
8. the method according to claim 7 based on the small molecule measure biomolecule content containing acetate, its
It is characterised by：
Described biomolecule is one kind or at least two in glucose, uric acid and cholesterol；
Described protease is oxidizing ferment.
The application of the small molecule containing acetate described in 9., it is characterised in that：
Application in biological detection or field of biosensors.
10. according to the application of the small molecule containing acetate described in claim 9, it is characterised in that：
The described application in field of biological detection, can be the application in test paper, reagent, kit is prepared；
Described biology sensor is hydrogen peroxide, glucose, uric acid or cholesterol sensor.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108760727A (en) * | 2018-04-12 | 2018-11-06 | 广东轻工职业技术学院 | It is used as the purposes of catalyst as class peroxidase containing hydrionic small molecule |
| CN111537463A (en) * | 2020-04-15 | 2020-08-14 | 广东省第二人民医院（广东省卫生应急医院） | Method for quantitatively detecting uric acid in serum |
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|---|---|---|---|---|
| CN108760727A (en) * | 2018-04-12 | 2018-11-06 | 广东轻工职业技术学院 | It is used as the purposes of catalyst as class peroxidase containing hydrionic small molecule |
| CN111537463A (en) * | 2020-04-15 | 2020-08-14 | 广东省第二人民医院（广东省卫生应急医院） | Method for quantitatively detecting uric acid in serum |
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Application publication date: 20180323 |