CN105699462B - A kind of method for quantitatively detecting L-cysteine - Google Patents
A kind of method for quantitatively detecting L-cysteine Download PDFInfo
- Publication number
- CN105699462B CN105699462B CN201410714046.XA CN201410714046A CN105699462B CN 105699462 B CN105699462 B CN 105699462B CN 201410714046 A CN201410714046 A CN 201410714046A CN 105699462 B CN105699462 B CN 105699462B
- Authority
- CN
- China
- Prior art keywords
- cysteine
- open circuit
- glucose
- biological
- circuit potential
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Landscapes
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
Detection method, in particular to a kind of method using glucose/oxygen biological fuel cell quantitative determination L cysteines the present invention relates to a kind of physiologically substance.Confession using glucose/oxygen biological fuel cell is electrical, while in Cu2+In the presence of inhibit the open circuit potential of battery, and in L cysteines and Cu2+Cu S keys are formed, according to open circuit potential with quantitative detection of the variation realization of L semicystinol concentrations to L cysteines in sample to be tested.The present invention establishes the variable quantity of open circuit potential and the linear relationship of L semicystinol concentrations using the principle, realizes its highly sensitive detection to L cysteines.The method of the present invention is easy to operate, high sensitivity, specificity it is good, be applicable in the detection of L cysteines in food service industry and physiology course and clinical medicine domain.
Description
Technical field
Detection method, in particular to a kind of utilization glucose/oxygen bio-fuel the present invention relates to a kind of physiologically substance
The method that battery quantitative determines L-cysteine.
Background technology
L-cysteine is a kind of important amino acid, is present in many protein, glutathione.It is in many biologies
There is very important function, and its contents level exception and hepar damnification, skin lesion, senile dementia and the heart in the process
Many important diseases such as vascular diseases are related.In addition L-cysteine is widely used in food processing industry, main to use
In baked goods, the required ingredient as flour-dough improver.Though therefore the quantitative detection of L-cysteine to physiology course and
Clinical diagnosis still to food processing industry, is respectively provided with highly important meaning.
At present, quantitatively the common method of detection L-cysteine has spectrophotometry, high performance liquid chromatography, gas phase color
Spectrum-mass spectrography and electrochemical method etc..But these methods have its shortcoming, for example, spectrophotometry, high-efficient liquid phase color
Spectrometry and gas chromatography-mass spectrography are although the precision of detection is higher, and expensive equipment, sample treatment is cumbersome, technology requirement
It is high.Electrochemical sensor developed in recent years, such as the half Guang ammonia of electrode detection L- of graphene oxide/gold nano particle modification
The range of linearity and detection limit of acid are respectively 0.5-20 μM and 0.02 μM, half Guangs of electrode detection L- that Zinc oxide nanoparticle is modified
The range of linearity and detection limit of propylhomoserin are respectively 0.2-20 μM and 0.05 μM.Although this electrochemical sensor employs relatively honest and clean
The instrument of valency, but the defects of its technology is more demanding, is inevitable.And electrochemical workstation is difficult to realize be miniaturized.Cause
This, it is imperative to establish a kind of simple, sensitive, special L-cysteine detection method.
Invention content
It is a kind of based on glucose/oxygen biological fuel cell high sensitivity quantitation detection L- half present invention aims at providing
The method of cystine.
To achieve the above object, the technical solution adopted by the present invention is：
A kind of method for quantitatively detecting L-cysteine, the confession using glucose/oxygen biological fuel cell is electrical, together
When in Cu2+In the presence of inhibit the open circuit potential of battery, and in L-cysteine and Cu2+Formed Cu-S keys, according to open circuit potential with
The quantitative detection to L-cysteine in sample to be tested is realized in the variation of L-cysteine concentration.The glucose/oxygen biology
The glucose dehydrogenase (FAD-GDH) that fuel cell is relied on flavin adenine dinucleotide (FAD) acts on biological anode, and laccase is made
For biological-cathode.
Technical solution of the present invention is further preferred that, in Cu2+In the presence of, utilize Cu2+Inhibit biological anode FAD-GDH
Activity, reduce glucose/oxygen bio-fuel open circuit potential, then recycle sample to be tested in L-cysteine and Cu2+
Cu-S keys are formed, and reduce its inhibiting effect, the variation of front and rear open circuit potential is added in by L-cysteine, it is quantitative so as to reach
Detect the purpose of L-cysteine.
The biological anode of the glucose/oxygen biological fuel cell uses vitamin K3For electron mediator, biological-cathode
2,2 '-connection nitrogen-two (3- ethyls-benzothiazole -6- sulfonic acid) di-ammonium salts (ABTS) are then used as electron mediator.
The present invention effect be：
1. the present invention utilization biological anode based on FAD-GDH and the biological-cathode based on laccase, and biological anode and life
Vitamin K is respectively adopted in object cathode3It is electron mediator with ABTS, so as to construct high-performance glucose/oxygen bio-fuel electricity
Pond.The quantitative detection to L-cysteine of the battery is utilized simultaneously；Wherein, battery maximum power density is 98 μ W cm-2, open
Road current potential 0.78V.The method high sensitivity of L-cysteine is measured, there is wider detection range (20nM-3 μM) and relatively low
Detection limit (10nM).And the amino acid of non-sulfydryl is to detecting almost without interference.
2. the present invention utilizes L-cysteine and Cu2+Between formed Cu-S keys effect, reduce Cu2+To battery open circuit electricity
The inhibiting effect of position, while realize that the indirect quantification of L-cysteine detects using the self-powered of biological fuel cell.It can be bright
The aobvious sensitivity for improving detection.
3. highly sensitive detection L-cysteine is established by glucose/oxygen biological fuel cell that the present invention constructs
Method.The method high sensitivity, specificity are good, easy to operate, at low cost, available for food service industry, physiology course and clinic
The monitoring of L-cysteine in the fields such as medical diagnosis.
Description of the drawings
Fig. 1 is the detection mechanism of glucose provided in an embodiment of the present invention/oxygen biological fuel cell and L-cysteine
Figure.It is L-cysteine and Cu2+After effect, battery open circuit current potential can be effectively replied, realizes the indirect quantification of L-cysteine
Detection.
Fig. 2 is the power density curve graph of glucose provided in an embodiment of the present invention/oxygen biological fuel cell.
Fig. 3 is the canonical plotting of L-cysteine provided in an embodiment of the present invention detection.
Specific embodiment
The embodiments will be further described with reference to the accompanying drawings for the object of the invention, features and benefits.
Embodiment 1
Glucose/oxygen biological fuel cell is constructed
1. the preparation of biological anode
By 4 μ L multi-walled carbon nanotubes (2mg/mL), 5 μ L vitamin Ks3It is dripped successively with 10 μ LFAD-GDH solution (5mg/mL)
It is coated on glass-carbon electrode, 5 μ LNafion solution (0.05wt%) is added dropwise after dry.The life based on FAD-GDH is made in drying at room temperature
Object anode.
2. the preparation of biological-cathode
Drop coating on glass-carbon electrode and is done successively for 4 μ L multi-walled carbon nanotubes (2mg/mL) and 10 μ L laccase solutions (5mg/mL)
It is dry.Then 2 μ L glutaraldehyde solutions (2wt%) are added dropwise, and the biological-cathode based on laccase is made with dry at 4 DEG C.
Biological anode and biological-cathode are inserted into assembling bio-fuel electricity in 5mL Acetic acid-sodium acetates buffer solution (pH 5.5)
Pond, 30mM glucose make fuel, and add in 2mM ABTS as electron mediator, using CHI660E type electrochemical workstations, use
Linear sweep voltammetry (LSV) sweeps speed with 1mV/s and obtains i-E curves to 0V from open circuit potential scanning, battery is obtained after conversion
Power density curve (referring to Fig. 2), i.e. battery maximum power density are 98 μ W cm-2, open circuit potential 0.78V.
Embodiment 2
Using the glucose constructed/oxygen biological fuel cell, in Cu2+In the presence of, realize the Gao Ling of L-cysteine
Sensitivity detects：
4 μM of Cu is added in the battery of above-described embodiment2+, then with the open circuit potential of multimeter detection battery.Then
It adds in the L-cysteine of various concentration and records corresponding open circuit potential value.Finally with a concentration of abscissa of L-cysteine, open
The variable quantity of road current potential is ordinate, draw standard curve, can obtain straight line, be obtained its slope, as L-cysteine
Working curve slope, the variable quantity of open circuit potential and the linear relationship of L-cysteine concentration (referring to Fig. 3).
The working curve of L-cysteine is as shown in Fig. 2, as seen from the figure, the R of the working curve2Value reaches 0.997, linearly
Degree of fitting is fine.
I.e. detection range is 20nM-3 μM, and relatively low detection is limited to 10nM.
Embodiment 3
The measure of L-cysteine content in certain local brand L-cysteine composite sheet：
1. sample treatment：The L-cysteine composite sheet of certain appropriate local brand is taken, grinds and is dissolved in ultra-pure water, use
0.22 μm of disposable filter membrane is filtered processing, and constant volume is diluted with 100mM Acetic acid-sodium acetates buffer solution (pH 5.5).
The detection process of 2.L- cysteines：It is buffered with the 5mL Acetic acid-sodium acetates containing 30mM glucose, 2mM ABTS
Liquid (pH 5.5) is bottom liquid assembling fuel cell, and 4mM Cu are added in the liquid of bottom2+Measure the open circuit potential OCP of system1, then
The L-cysteine sample of different volumes is sequentially added in the liquid of bottom, and surveys its open circuit potential OCP respectively2。
The calculating of 3.L- semicystinol concentrations：Take the variable quantity (OCP of open circuit potential2–OCP1) in standard curve range
Dilution ratio is calculated, and is substituted into the calculation formula that standard curve obtains, is finally multiplied by corresponding extension rate, so as to obtain
The content of L-cysteine is 43.1 ± 2.8 (w/w, %) in the L-cysteine composite sheet of certain brand.L-cysteine content
Calculation formula is：
The preferred embodiments of the disclosure and embodiment are described in detail above in conjunction with attached drawing, but this hair
It is bright to be not limited to the above-described embodiment and examples, made under other any Spirit Essences and principle without departing from the present invention
Change, modification, substitute, combination, simplify, should be equivalent substitute mode, be included within protection scope of the present invention.
Claims (4)
- A kind of 1. method for quantitatively detecting L-cysteine, it is characterised in that：Using glucose/oxygen biological fuel cell from It is electron-donating, while in Cu2+In the presence of inhibit the open circuit potential of battery, and in L-cysteine and Cu2+Cu-S keys are formed, according to Open circuit potential realizes the quantitative detection to L-cysteine in sample to be tested with the variation of L-cysteine concentration.
- 2. the method as described in claim 1 for quantitatively detecting L-cysteine, it is characterised in that：The glucose/oxygen life The glucose dehydrogenase that object fuel cell is relied on flavin adenine dinucleotide (FAD)（FAD-GDH）Act on biological anode, laccase Act on biological-cathode.
- 3. the method as described in claim 1 or 2 for quantitatively detecting L-cysteine, it is characterised in that：In Cu2+In the presence of, Utilize Cu2+Inhibit the activity of biological anode FAD-GDH, inhibit the open circuit potential of battery, then recycle L- half in sample to be tested Cystine and Cu2+Cu-S keys are formed, reduce Cu2+To the inhibiting effect of battery open circuit current potential, before and after being added in by L-cysteine The variation of open circuit potential, so as to achieve the purpose that quantitative detection L-cysteine.
- 4. the method as described in claim 3 for quantitatively detecting L-cysteine, it is characterised in that：The glucose/oxygen life The biological anode of object fuel cell uses vitamin K3For electron mediator, biological-cathode then using 2,2 '-connection nitrogen-two (3- ethyls- Benzothiazole -6- sulfonic acid) di-ammonium salts（ABTS）For electron mediator.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410714046.XA CN105699462B (en) | 2014-11-28 | 2014-11-28 | A kind of method for quantitatively detecting L-cysteine |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410714046.XA CN105699462B (en) | 2014-11-28 | 2014-11-28 | A kind of method for quantitatively detecting L-cysteine |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN105699462A CN105699462A (en) | 2016-06-22 |
| CN105699462B true CN105699462B (en) | 2018-06-08 |
Family
ID=56230457
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410714046.XA Expired - Fee Related CN105699462B (en) | 2014-11-28 | 2014-11-28 | A kind of method for quantitatively detecting L-cysteine |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105699462B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108645805B (en) * | 2018-05-02 | 2020-10-27 | 天津科技大学 | Novel method for label-free rapid detection of cysteine |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101329279A (en) * | 2008-07-18 | 2008-12-24 | 山西大学 | Method for rapidly testing cysteine in water solution |
| CN101509892A (en) * | 2009-03-20 | 2009-08-19 | 山东大学 | Method for rapidly identifying cysteine and cystine by utilizing electrochemical technique |
| CN201852813U (en) * | 2010-10-22 | 2011-06-01 | 东南大学 | Blue-green algae concentration sensor |
| CN103149257A (en) * | 2013-03-05 | 2013-06-12 | 中国科学院大学 | Nano-gold/graphene nano compound-based method for rapidly determining cysteine based on |
| CN103207230A (en) * | 2013-04-02 | 2013-07-17 | 北京林业大学 | Method for constructing dual-chamber microbial fuel cell-type BOD (biochemical oxygen demand) sensor by using potassium permanganate as cathode electron acceptor |
| CN104064791A (en) * | 2014-02-26 | 2014-09-24 | 武汉科技大学 | Biological inverted-conversion reactor of microbial fuel cell, purification method of CO2 in gas and preparation method of CO2 biological synthetic fuel |
-
2014
- 2014-11-28 CN CN201410714046.XA patent/CN105699462B/en not_active Expired - Fee Related
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101329279A (en) * | 2008-07-18 | 2008-12-24 | 山西大学 | Method for rapidly testing cysteine in water solution |
| CN101509892A (en) * | 2009-03-20 | 2009-08-19 | 山东大学 | Method for rapidly identifying cysteine and cystine by utilizing electrochemical technique |
| CN201852813U (en) * | 2010-10-22 | 2011-06-01 | 东南大学 | Blue-green algae concentration sensor |
| CN103149257A (en) * | 2013-03-05 | 2013-06-12 | 中国科学院大学 | Nano-gold/graphene nano compound-based method for rapidly determining cysteine based on |
| CN103207230A (en) * | 2013-04-02 | 2013-07-17 | 北京林业大学 | Method for constructing dual-chamber microbial fuel cell-type BOD (biochemical oxygen demand) sensor by using potassium permanganate as cathode electron acceptor |
| CN104064791A (en) * | 2014-02-26 | 2014-09-24 | 武汉科技大学 | Biological inverted-conversion reactor of microbial fuel cell, purification method of CO2 in gas and preparation method of CO2 biological synthetic fuel |
Non-Patent Citations (2)
| Title |
|---|
| Direct Current Generation from NADH and L-Cysteine Using Carbon Fiber: Possible Uses in Biofuel Cells;Hidekatsu Maeda等;《Bull. Chem. Soc. Jpn》;20110507;第84卷;第544-551页 * |
| Enhanced Performance of a Glucose/O2 Biofuel Cell Assembled with Laccase-Covalently Immobilized Three-Dimensional Macroporous Gold Film-Based Biocathode and Bacterial Surface Displayed Glucose Dehydrogenase-Based Bioanode;Chuantao Hou等;《Analytical Chemistry》;20140512;第86卷;第6057-6063页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN105699462A (en) | 2016-06-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Sanati et al. | Electrochemical determination of methyldopa by graphene quantum dot/1-butyl-3-methylimidazolium hexafluoro phosphate nanocomposite electrode | |
| Lin et al. | Electrochemical non-enzymatic glucose sensors based on nano-composite of Co3O4 and multiwalled carbon nanotube | |
| Amiri et al. | Electrocatalytic determination of sumatriptan on the surface of carbon-paste electrode modified with a composite of cobalt/Schiff-base complex and carbon nanotube | |
| Li et al. | Electrochemical dual signal sensing platform for the simultaneous determination of dopamine, uric acid and glucose based on copper and cerium bimetallic carbon nanocomposites | |
| Wei et al. | Fabrication of CeO2 nanoparticles modified glassy carbon electrode and its application for electrochemical determination of UA and AA simultaneously | |
| Wen et al. | Simultaneous analysis of uric acid, xanthine and hypoxanthine using voltammetric sensor based on nanocomposite of palygorskite and nitrogen doped graphene | |
| Ensafi et al. | Determination of isoproterenol and uric acid by voltammetric method using carbon nanotubes paste electrode and p-chloranil | |
| Chawla et al. | An amperometric hemoglobin A1c biosensor based on immobilization of fructosyl amino acid oxidase onto zinc oxide nanoparticles–polypyrrole film | |
| Chekin et al. | Preparation and characterization of Ni (II)/polyacrylonitrile and carbon nanotube composite modified electrode and application for carbohydrates electrocatalytic oxidation | |
| KR101721572B1 (en) | Biosensors for Glycosylated Hemoglobin using electrochemically active nano particles | |
| Sun et al. | Simultaneous electrochemical determination of guanosine and adenosine with graphene–ZrO2 nanocomposite modified carbon ionic liquid electrode | |
| Fan et al. | Smartphone-based electrochemical system with multi-walled carbon nanotubes/thionine/gold nanoparticles modified screen-printed immunosensor for cancer antigen 125 detection | |
| CN205067415U (en) | Uric acid test test paper | |
| Liu et al. | A facile electrochemical uricase biosensor designed from gold/amino acid nanocomposites | |
| CN103175884A (en) | High-sensitivity glucose biosensor and preparation method thereof | |
| Zhao et al. | An ultrasensitive photoelectrochemical sensor with layer-by-layer assembly of chiral multifarene [3, 2, 1] and g-C3N4 quantum dots for enantiorecognition towards thyroxine | |
| Mohammadi et al. | Simultaneous determination of droxidopa and carbidopa by carbon paste electrode functionalized with NiFe2O4 nanoparticle and 2-(4-ferrocenyl-[1, 2, 3] triazol-1-yl)-1-(naphthalen-2-yl) ethanone | |
| Dong et al. | A disposable printed amperometric biosensor for clinical evaluation of creatinine in renal function detection | |
| Qiu et al. | A low-cost wireless intelligent portable sensor based on disposable laser-induced porous graphene flexible electrode decorated by gold nanoshells for rapid detection of sulfonamides in aquatic products | |
| Murthy et al. | Application of derivative voltammetry in the quantitative determination of alloxan at single-walled carbon nanotubes modified electrode | |
| Beitollahi et al. | Electrochemical sensor for selective determination of N-acetylcysteine in the presence of folic acid using a modified carbon nanotube paste electrode | |
| CN105699462B (en) | A kind of method for quantitatively detecting L-cysteine | |
| Dehdashtian et al. | Fabrication of a novel electrochemical sensor based on an electrosynthesized indolyldihydroxyquinone as a bio-based modifier for sensitive and selective direct electrochemical determination of tryptophan | |
| Chen et al. | Direct electrochemical behavior of cytochrome c on sodium dodecyl sulfate modified electrode and its application to nitric oxide biosensor | |
| CN109187678A (en) | Utilize the nitrite detection device of nanogold graphene modified electrochemical method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20180608 Termination date: 20181128 |