CN102692409B - Method and kit for detecting concentration of chiral amino acid - Google Patents
Method and kit for detecting concentration of chiral amino acid Download PDFInfo
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- CN102692409B CN102692409B CN201210211371.5A CN201210211371A CN102692409B CN 102692409 B CN102692409 B CN 102692409B CN 201210211371 A CN201210211371 A CN 201210211371A CN 102692409 B CN102692409 B CN 102692409B
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- reaction
- amino acid
- concentration
- luminol
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Abstract
The invention discloses a method and a kit for detecting concentration of chiral amino acid. The method comprises the following steps of: 1) adding D-amino acid oxidase or L-amino acid oxidase into a sample to be tested for carrying out an enzyme catalysis reaction; 2) sequentially adding buffer solution, luminol and horse radish peroxidase into the obtained reaction solution in the step 1) for carrying out a chemiluminiscence reaction; and 3) measuring the luminous intensity of the reaction solution in the step 2). The method can be used for measuring chiral amino acid in an enantiomer. The method is not interfered by another chiral amino acid in the enantiomer or is not interfered by common metal ions. The method is easy to operate and high in sensitivity, and the content of the chiral amino acid in the enantiomer can be detected.
Description
Technical field
The invention belongs to analytical chemistry field, particularly a kind of chemoluminescence method detects method and the kit thereof of chiral amino acid.
Background technology
In biosome, amino acid whose level reflects the many biological phenomenas in biosome, and closely related with numerous disease.Therefore, particularly the amino acid whose detection of D type has great importance in clinical diagnosis and clinical basic.But because this compounds lacks uv absorption, fluorescence activity and electrochemical activity, most analytical approach needs to derive this compounds; What is more important often also needs chiral derivatizing agent or other chiral resolution technology to realize for the analysis of enantiomorph, is separated etc. as adopted chiral chromatographic column.The detection of chemiluminescence (CL) method has highly sensitive, and method is simple, and without any need for features such as additional light sources, thus receives the concern of people in recent years.Although more existing luminescence system is used for amino acid whose detection: (1) peroxoxalate luminescence-producing reaction measures amino acid fluorescent derivative.Though this method can obtain higher sensitivity, but still need to derive, and complicated operation, condition is harsh.(2) luminol-hypobromite luminescence system, in the basic conditions, amino acid can suppress luminol-hypobromite luminescence-producing reaction, easy and simple to handle with this method of the amino acid whose content of the attenuation measurement of luminous signal, but sensitvity constraint.(3) luminol-hydrogen peroxide luminescence system, amino acid can form complex compound with copper ion, and cataluminescence is active.But these analytical approachs are also not suitable for the content analysis of the chiral amino acid in enantiomorph.
Amino acid oxidase is widely used in the fractionation of chiral amino acid because it has stereocpecificity highly.Amino acid oxidase is combined with chemoluminescence method, though have bibliographical information for the analysis of chiral amino acid.But this analytical approach many employings Using Flow Injection Analysis, and luminescence system is metal ion catalysis luminol-hydrogen peroxide luminescence system.This analytical approach needs special instrument and equipment on the one hand, and on the other hand, because the another kind of amino acid itself existed can suppress this luminescence system, the analysis that therefore neither be applicable to chiral amino acid in enantiomorph detects.For this reason, develop a kind of easy and simple to handle, highly sensitive luminescence system, the content that can be used for measuring chiral amino acid in enantiomorph seems very necessary.
Summary of the invention
The technical problem to be solved in the present invention is exactly the content analysis that can not realize chiral amino acid in enantiomorph for existing chemoluminescence method when detecting amino acid, a kind of simple to operate, highly sensitive chemoluminescence method is provided, can be used for detecting the chiral amino acid in enantiomorph.
The technical scheme that the present invention solves the problems of the technologies described above is as follows: a kind of method detecting concentration of chiral amino acid, comprises the following steps:
1) in measuring samples, D-AAO is added or L-amino acid oxidase carries out enzymic catalytic reaction;
2) in the reactant liquor of step 1) gained, add damping fluid successively, luminol, horseradish peroxidase carry out chemiluminescence reaction;
3) determination step 2) luminous intensity of reactant liquor of gained.
Know-why of the present invention is: chiral amino acid to be determined is fully converted into hydrogen peroxide under the effect of corresponding amino acid oxidase, further interpolation horseradish peroxidase enzyme catalytic hydrogen peroxide oxidation luminol is luminous, and luminous signal is linearly relevant within the specific limits to the amino acid concentration of mensuration, therefore may be used for the mensuration of chiral amino acid.The present invention, when measuring D-amino acid, adopts D-AAO; When measuring L-amino acid, then adopt L-amino acid oxidase.
In the present invention, step 1) is: in measuring samples, add D-AAO or L-amino acid oxidase carries out enzymic catalytic reaction.Preferably, in the reaction system of the enzymic catalytic reaction described in step 1), also comprise conventional buffer solution, preferably phosphoric acid damping fluid (PBS).The pH value of reaction system preferably 7.5 ~ 9.5, more preferably pH 8.3.The total concentration of damping fluid preferably 1.0 × 10
-21.0mol/L.The concentration of D-AAO or L-amino acid oxidase is preferably 2.6 ~ 4.4U/ml.The temperature of reaction of enzymic catalytic reaction is 20 ~ 45 ° of C, and the reaction time is greater than 1 minute, preferably 1 ~ 15 minute.
Wherein, add D-AAO, measure D-amino acid, the best is: D-AAO 2.6 ~ 4.4U/ml, and damping fluid is the phosphate buffer of 10mM, and pH is 8.3, and enzyme reaction temperature is 25 ° of C, and the reaction time is 10 minutes.Sensing range is preferred: the concentration of primary sample is 1.0 × 10
-5~ 2.0 × 10
-3mol/L.
Wherein, add L-amino acid oxidase, measure L-amino acid, the best is: L-amino acid oxidase 2.6 ~ 4.4U/ml, and damping fluid is the phosphate buffer of 100mM, and pH is 8.8, and enzyme reaction temperature is 37 ° of C, and the reaction time is 10 minutes.Sensing range is preferred: the concentration 2.0 × 10 of primary sample
-4~ 1.0 × 10
-2mol/L.
In the present invention, step 2) be: in the reactant liquor of step 1) gained, add damping fluid successively, luminol, horseradish peroxidase carry out chemiluminescence reaction.Described damping fluid, for adjusting the pH of system in certain scope, preferably, makes the pH value of reaction system be greater than 9.0.Described damping fluid is conventional, preferably phosphoric acid damping fluid.The total concentration preferably 0.01 ~ 1.0mol/L of the damping fluid added.The final concentration of luminol preferably 1.0 × 10
-5~ 1.0 × 10
-3mol/L; Horseradish peroxidase is 0.1 ~ 5.0U/ml preferably.Preferably, the temperature of reaction of luminescence-producing reaction is 10 ~ 45 ° of C, and the reaction time is 1 ~ 10 minute.The best is: luminol 1.0 × 10
-4mol/L, horseradish peroxidase 0.36 ~ 0.6U/ml; Damping fluid is the phosphate buffer of 0.1mol/L, and pH is 10; The temperature of luminescence-producing reaction is 25 ° of C, and the reaction time is 1 ~ 10 minute.
In the present invention, step 3) is: determination step 2) luminous intensity of the reactant liquor of gained.The method of described mensuration luminous intensity is the conventional method of this area, generally measures on Chemiluminescence Apparatus.
In the present invention, preferably, measuring samples also carries out pre-treatment before carrying out step 1), namely adopt in cation exchange solid phase extraction removing sample as Cu
2+, Co
2+, Fe
3+deng metallic ion.Concrete grammar preferably comprises: the pH of sample is adjusted to 5.0, adopts conventional method to be activated by SCX SPE pillar, by measuring samples by SCX SPE pillar, with 4% ammonia spirit wash-out, collects eluent, namely can be used for follow-up analysis design mothod.
For luminol-hydrogen peroxide luminescence system, common metallic ion is as Cu
2+, Co
2+, Fe
3+all there is catalysis Deng to this system, thus interference is existed to mensuration.For this reason, usually need when working sample to carry out purified treatment to sample.For eliminating the interference of common metal ion, the impurity that cation exchange solid phase extraction separating metal ions etc. impacts enzymatic reaction and luminol luminescence can be adopted.
The present invention also provides second technical scheme: a kind of chemoluminescence method detects the kit of concentration of chiral amino acid, comprises following reagent:
1) D-AAO or L-amino acid oxidase;
2) luminol solution;
3) horseradish peroxidase; With
4) damping fluid.
In the present invention, reagent 2) described in luminol solution preferably concentration 1.0 × 10
-3the luminol aqueous solution of mol/L.In the present invention, reagent 4) described in damping fluid be the buffer solution of this area routine, the phosphate buffer of preferred 10-100mM, pH 8-10.Preferably comprise 10mM, the phosphate buffer of pH 8.3, the phosphate buffer of 100mM, pH 8.0 and the phosphate buffer of 100mM, pH 10.Damping fluid can be used for the solution preparing D-AAO or L-amino acid oxidase and horseradish peroxidase, and as the damping fluid of chemiluminescence reaction system.
In the present invention, described kit preferably also comprises cation exchange solid-phase extraction column, preferred SCX SPE pillar.And can further include the activating reagent, washing reagent, elution reagent etc. of this extraction column, as methyl alcohol, distilled water and phosphate buffer solution (0.01M, pH 5.0), ammonia spirit etc.
The raw material that the present invention is used or reagent except special instruction, all commercially.
Compared to prior art, beneficial effect of the present invention is as follows:
In the present invention, when being fully converted into hydrogen peroxide under the effect of chiral amino acid at amino acid oxidase, luminol can be made under the effect of horseradish peroxidase luminous, and luminous signal is linearly relevant to the amino acid of mensuration.The metallic ion of other routines, as Cu
2+, Co
2+, Fe
3+though the luminescence of catalysis luminol also may be used for the mensuration of the hydrogen peroxide produced, but cannot be used for the content analysis for the chiral amino acid in enantiomorph.This is because such as when the amino-acid oxidase enzymatic conversion D-type amino acid selecting D-type is hydrogen peroxide, when adopting the luminescence of metal ion catalysis luminol to detect, the amino acid of the L-type of existence can suppress the luminescence of metal ion catalysis luminol.Generally speaking that this luminescence system can not measure D-type amino acid when L-type amino acid exists.And there is lower Background luminescence and the catalytic effect of Geng Gao due to horseradish peroxidase, thus detection sensitivity is higher.For this reason, the present invention is luminous with horseradish peroxidase enzyme catalytic luminol-hydrogen peroxide, builds the method for the chiral amino acid detected in enantiomorph.
Horseradish peroxidase-luminol-hydrogen peroxide is used for as luminescence system the chiral amino acid that detects in enantiomorph by the present invention first.The present invention is also the hybrid mode of chemical luminous system, composition and ratio and is applicable to the sample treatment of this detection system.Adopt detection method provided by the present invention, may be used for amino acid whose detection in biological sample.
Accompanying drawing explanation
Below in conjunction with accompanying drawing, characteristic sum beneficial effect of the present invention is described.
Fig. 1 be in embodiment 3 L-amino acid to the chemiluminescence column diagram (horseradish peroxidase-luminol-hydrogen peroxide luminescence system) in the amino acid whose interference experiment of D-.Sample 1 ~ 4 is respectively:
1, PBS(blank); 2,2.0 × 10
-3the D-alanine (D-Ala) of mol/L; 3, D-alanine (D-Ala) and ALANINE (L-Ala) potpourri, concentration is respectively 2.0 × 10
-3mol/L; 4,2.0 × 10
-3the ALANINE (L-Ala) of mol/L.
Fig. 2 is the chemiluminescence column diagram (horseradish peroxidase-luminol-hydrogen peroxide luminescence system) in embodiment 3 after the D-type Freamine Ⅲ SCX pillar process of copper ions.Sample 1 ~ 3 is respectively: 1, and 1.0 × 10
-2copper ion (the Cu of mol/L
2+), carry out the process of SCX pillar; 2,1.0 × 10
-3the D-alanine (D-Ala) of mol/L, does not carry out sample preparation; 3,1.0 × 10
-3the D-alanine (D-Ala) of mol/L, carries out the process of SCX pillar; 4,1.0 × 10
-3the D-alanine of mol/L, wherein containing 1.0 × 10
-2copper ion (the Cu of mol/L
2+), carry out the process of SCX pillar.
Embodiment
Further illustrate the present invention by embodiment below, but the present invention is not limited.The experimental technique of unreceipted actual conditions in the following example, usually conveniently condition, or according to the condition that manufacturer advises." room temperature " described in embodiment refers to the temperature of carrying out the operation room tested, and is generally 25 DEG C.
Embodiment 1
Sample pre-treatments
1, solution preparation:
1. PBS damping fluid: take a certain amount of sodium hydrogen phosphate and sodium dihydrogen phosphate, preparation total concentration is the phosphate buffer of 10mM, and the pH of adjustment solution is 5.0.
2. Freamine Ⅲ: take D-alanine, with 10mM PBS(pH 5.0) dissolve, being mixed with concentration is 1.0 × 10
-3the solution of mol/L.
3. copper ion solution: take a certain amount of copper sulphate, with 10mM PBS(pH 5.0) dissolve, be mixed with 1.0 × 10
-2the copper ion solution of mol/L.
4. the Freamine Ⅲ of copper ions: add copper sulphate solid in 2. prepared amino acid mixed solution, makes its final concentration be 1.0 × 10
-2mol/L.
2, cation exchange solid phase extraction purification sample
(1) SCX SPE pillar (model is CNWBOND SCX, 500mg, 3ml, CNW company) is activated.
The methyl alcohol of 3ml, distilled water and phosphate buffer solution (0.01M, pH 5.0) is used respectively to wash SPE pillar three times respectively.
(2) loading
Get the upper SPE pillar activated of solution (ie in solution 3., 4.) 3ml of the copper ions of above-mentioned preparation, and wash three times with 3ml phosphate buffer solution (0.01M, pH 5.0).Finally use 3ml 4% ammoniacal liquor wash-out, collect and merge eluent, regulating the pH value of this solution.If measure D-amino acid, adjust ph is 8.3, otherwise is 8.8 when measuring L-amino acid.
Embodiment 2
D-type amino-acid oxidase enzymatic conversion D-alanine (D-Ala) is hydrogen peroxide
1. solution preparation:
1. D-AAO solution: take a certain amount of D-AAO, with 10mM PBS(pH 8.3) to dissolve, concentration is 43U/ml.
2. D-alanine solution: take D-alanine, with 10mM PBS(pH 8.3) dissolve, being mixed with concentration is 5.0 × 10
-6, 1.0 × 10
-5, 5.0 × 10
-5, 1.0 × 10
-4, 5.0 × 10
-4, 1.0 × 10
-3, 2.0 × 10
-3a series of solution of mol/L.
3. ALANINE solution: take ALANINE, with 10mM PBS(pH 8.3) dissolve, being mixed with concentration is 2.0 × 10
-3the solution of mol/L.
4. D-alanine and ALANINE mixed solution: take D-alanine and ALANINE, with 10mMPBS(pH 8.3) dissolve, be mixed with final concentration and be respectively 2.0 × 10
-3the mixed solution of mol/L.
2, the amino acid converting enzyme reaction of D-
Get the above-mentioned D-AAO solution of 3 μ l (ie in solution 1.), be formulated as the reaction system of 50 μ l after solution after the above-mentioned Freamine Ⅲ of 5 μ l (ie in solution 2.-4.) or embodiment 1 purify and 42 μ l 10mM PBS fully mix, the pH value of reactant liquor is 8.3.25 ° of C water-baths, after 10 minutes, add luminescence-producing reaction damping fluid at once.
Embodiment 3
Chemiluminescence detection D-alanine
1, solution preparation:
(1) horseradish peroxidase solution: take a certain amount of horseradish peroxidase, with 100mMPBS(pH 8.0) to dissolve, concentration is 60U/ml.
(2) luminescence-producing reaction damping fluid: the phosphate buffer solution of preparation 0.1mol/L, the pH value to 10 of adjustment solution.
(3) luminol solution: take a certain amount of luminol, by water-soluble solution and to be mixed with concentration be 1.0 × 10
-3the luminol solution of mol/L.The concentration of luminol should not be too high, otherwise not soluble, needs to add more alkali, be unfavorable for the control of the pH of reaction system.
2, horseradish peroxidase enzyme catalytic luminol luminescence-producing reaction
Get the reactant liquor 50 μ l of D-type amino-acid oxidase enzymatic conversion D-alanine, add above-mentioned luminescence-producing reaction damping fluid 397 μ l, luminol solution 50 μ l, horseradish peroxidase solution 3 μ l successively.React 3 minutes under room temperature after abundant mixing, get 150 μ l and add 96 hole ELISA Plate and measure its luminous intensity, every class value measures three times.
3, testing conditions:
This experiment adopts the multi-functional microplate reader (Synergy H1) of Bioteck, and select chemiluminescence pattern, detected parameters Gain value is 100, Read Height 1.00mm.
Adopt the D-alanine solution of a series of variable concentrations gradient to detect, find that concentration is 1.0 × 10
-5~ 2.0 × 10
-3mol/L is linear, and working curve is I=2.0 × 10
6c
d-alanine+ 83.62, linearly dependent coefficient R is 0.998.With 3S
b/ S calculates detectability, the detectability difference 3.5 × 10 of D-alanine (D-ALA)
-6mol/L.
In addition, carried out the mensuration of luminous intensity to the ALANINE containing same concentrations in sample D-alanine, as shown in Figure 1, ALANINE can not bring interference to detection to result.Meanwhile, to the metal copper ion added higher than D-amino acid/11 0 times of concentration, after carrying out the process of SCX pillar, to detection not interference (as shown in Figure 2).
Embodiment 4
Detect ALANINE
Carry out sample pre-treatments with embodiment 1, except obtain solution 2. in D-alanine is replaced with except ALANINE.
Carrying out L-amino acid oxidase conversion ALANINE (L-Ala) with embodiment 2 is hydrogen peroxide, and except D-AAO in solution (1) replaces with L-amino acid oxidase, the D-alanine in solution (2) replaces with ALANINE.Reaction conditions difference is: damping fluid is the phosphate buffer of 100mM, and pH is 8.8, and enzyme reaction temperature is 37 ° of C.
Chemiluminescence detection is with embodiment 3.
As a result, adopt the ALANINE solution of a series of variable concentrations gradient to detect, find that concentration is 2.0 × 10
-4~ 1.0 × 10
-2mol/L is linear, and working curve is I=5.27 × 10
4c
aLANINE+ 20.95, linearly dependent coefficient R is 0.999.With 3S
b/ S calculates detectability, and the detection of ALANINE (L-ALA) is limited to 5.7 × 10
-5mol/L.
In addition, carried out the mensuration of luminous intensity to the D-alanine containing same concentrations in sample ALANINE, D-alanine can not bring interference to detection.Meanwhile, to the metal copper ion added higher than L-amino acid/11 0 times of concentration, after carrying out the process of SCX pillar, to detection not interference.
The all documents mentioned in the present invention are quoted as a reference all in this application, are just quoted separately as a reference as each section of document.In addition should be understood that those skilled in the art can make various changes or modifications the present invention after having read foregoing of the present invention, these equivalent form of values fall within the application's appended claims limited range equally.
Claims (6)
1. detect a method for concentration of chiral amino acid, it is characterized in that, comprise the following steps:
1) in measuring samples, D-AAO is added or L-amino acid oxidase carries out enzymic catalytic reaction;
2) in step 1) add damping fluid in the reactant liquor of gained successively, luminol, horseradish peroxidase carry out chemiluminescence reaction;
3) determination step 2) luminous intensity of reactant liquor of gained;
Wherein, step 1) described in enzymic catalytic reaction reaction system in, also comprise conventional buffer solution, the pH value of described reaction system is 7.5 ~ 9.5, the concentration of D-AAO or L-amino acid oxidase is greater than 0.01U/ml, the temperature of reaction of enzymic catalytic reaction is 20 ~ 45 DEG C, 1 ~ 15 minute reaction time; Step 2) described in the pH value of reaction system of chemiluminescence reaction be greater than 9.0, the final concentration of luminol is 1.0 × 10
-5~ 1.0 × 10
-3mol/L, horseradish peroxidase is 0.1 ~ 5.0U/ml, and the temperature of reaction of luminescence-producing reaction is 10 ~ 45 DEG C, and the reaction time is 1 ~ 10 minute.
2. the method for claim 1, is characterized in that, in step 1) described in enzymic catalytic reaction in, described amino acid oxidase is D-AAO, and described damping fluid is the phosphate buffer of 10mM, pH8.3, described temperature of reaction is 25 DEG C, and the reaction time is 10 minutes.
3. the method for claim 1, is characterized in that, in step 1) described in enzymic catalytic reaction in, described amino acid oxidase is L-amino acid oxidase, and described damping fluid is the phosphate buffer of 100mM, pH8.8, described temperature of reaction is 37 DEG C, and the reaction time is 10 minutes.
4. the method for claim 1, is characterized in that, in step 2) described in chemiluminescence reaction in, described damping fluid is the phosphate buffer of 0.1mol/L, pH10, and in described reaction system, luminol concentration is 1.0 × 10
-4mol/L, horseradish peroxidase concentration is 0.22 ~ 0.36U/ml, and the temperature of described luminescence-producing reaction is 25 DEG C, and the reaction time is 1 ~ 10 minute.
5. the method for claim 1, is characterized in that, described measuring samples carry out step 1) the front metallic ion also adopted in cation exchange solid phase extraction removing sample.
6. method as claimed in claim 5, it is characterized in that, described cation exchange solid phase extraction comprises: the pH of measuring samples is adjusted to 5.0, after adopting conventional method to be activated by SCX SPE pillar, by sample by SCX SPE pillar, with 4% ammonia spirit wash-out, collect eluent.
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| CN106018371B (en) * | 2016-07-06 | 2019-02-15 | 天津大学 | Enzyme coupling nucleic acid-argentum nanometer probe |
| CN113310958B (en) * | 2021-05-19 | 2022-12-23 | 华东理工大学 | Preparation method of hierarchical porous metal organic framework chiral sensing probe, probe obtained by preparation method and application of probe |
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Non-Patent Citations (4)
| Title |
|---|
| 《中性体系中鲁米诺电化学发光行为》;屠一锋等;《分析化学研究简报》;20020630;第30卷(第6期);第729-731页 * |
| 《反相流动注射-化学发光法测定氨基酸》;郎惠云等;《分析试验室》;20030930;第22卷(第5期);第44-46页 * |
| 《基于固定化酶的化学发光停流法测定D-氨基酸》;封满良等;《高等学校化学学报》;19961231;第17卷;第1859-1861页 * |
| 《氨基酸化学发光分析的进展》;李正平等;《河北农业大学学报》;19950131;第18卷(第1期);第104-106页 * |
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