CN110470662A - A method of measurement drug and copper ion complexing power - Google Patents
A method of measurement drug and copper ion complexing power Download PDFInfo
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- CN110470662A CN110470662A CN201910752379.4A CN201910752379A CN110470662A CN 110470662 A CN110470662 A CN 110470662A CN 201910752379 A CN201910752379 A CN 201910752379A CN 110470662 A CN110470662 A CN 110470662A
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- copper ion
- rhodamine
- hydrazides
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- 229910001431 copper ion Inorganic materials 0.000 title claims abstract description 46
- JPVYNHNXODAKFH-UHFFFAOYSA-N Cu2+ Chemical compound [Cu+2] JPVYNHNXODAKFH-UHFFFAOYSA-N 0.000 title claims abstract description 45
- 238000000034 method Methods 0.000 title claims abstract description 34
- 229940079593 drug Drugs 0.000 title claims abstract description 26
- 239000003814 drug Substances 0.000 title claims abstract description 26
- 230000000536 complexating effect Effects 0.000 title claims abstract description 19
- 238000005259 measurement Methods 0.000 title description 5
- 229940042795 hydrazides for tuberculosis treatment Drugs 0.000 claims abstract description 51
- PYWVYCXTNDRMGF-UHFFFAOYSA-N rhodamine B Chemical compound [Cl-].C=12C=CC(=[N+](CC)CC)C=C2OC2=CC(N(CC)CC)=CC=C2C=1C1=CC=CC=C1C(O)=O PYWVYCXTNDRMGF-UHFFFAOYSA-N 0.000 claims abstract description 50
- 229940043267 rhodamine b Drugs 0.000 claims abstract description 50
- 239000007788 liquid Substances 0.000 claims abstract description 41
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 42
- 239000000243 solution Substances 0.000 claims description 34
- 239000003960 organic solvent Substances 0.000 claims description 16
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 14
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 claims description 12
- 239000011259 mixed solution Substances 0.000 claims description 12
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 claims description 9
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 9
- 238000003756 stirring Methods 0.000 claims description 9
- 238000001514 detection method Methods 0.000 claims description 7
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 6
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 6
- 239000013641 positive control Substances 0.000 claims description 6
- 239000013642 negative control Substances 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- 229960004063 propylene glycol Drugs 0.000 claims description 5
- 238000002835 absorbance Methods 0.000 claims description 4
- ORTQZVOHEJQUHG-UHFFFAOYSA-L copper(II) chloride Chemical compound Cl[Cu]Cl ORTQZVOHEJQUHG-UHFFFAOYSA-L 0.000 claims description 4
- 102100021587 Embryonic testis differentiation protein homolog A Human genes 0.000 claims description 3
- 101000898120 Homo sapiens Embryonic testis differentiation protein homolog A Proteins 0.000 claims description 3
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 claims description 3
- OPQARKPSCNTWTJ-UHFFFAOYSA-L copper(ii) acetate Chemical compound [Cu+2].CC([O-])=O.CC([O-])=O OPQARKPSCNTWTJ-UHFFFAOYSA-L 0.000 claims description 2
- UGWKCNDTYUOTQZ-UHFFFAOYSA-N copper;sulfuric acid Chemical compound [Cu].OS(O)(=O)=O UGWKCNDTYUOTQZ-UHFFFAOYSA-N 0.000 claims description 2
- 238000002845 discoloration Methods 0.000 claims description 2
- 238000000825 ultraviolet detection Methods 0.000 claims description 2
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 238000010521 absorption reaction Methods 0.000 abstract description 5
- BEJNERDRQOWKJM-UHFFFAOYSA-N kojic acid Chemical compound OCC1=CC(=O)C(O)=CO1 BEJNERDRQOWKJM-UHFFFAOYSA-N 0.000 description 36
- 229960004705 kojic acid Drugs 0.000 description 36
- WZNJWVWKTVETCG-UHFFFAOYSA-N kojic acid Natural products OC(=O)C(N)CN1C=CC(=O)C(O)=C1 WZNJWVWKTVETCG-UHFFFAOYSA-N 0.000 description 36
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 18
- 150000001875 compounds Chemical class 0.000 description 11
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 10
- 239000010949 copper Substances 0.000 description 10
- 229910052802 copper Inorganic materials 0.000 description 10
- 229910021645 metal ion Inorganic materials 0.000 description 7
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 5
- 239000002689 soil Substances 0.000 description 5
- 238000001917 fluorescence detection Methods 0.000 description 4
- 239000002904 solvent Substances 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 230000003287 optical effect Effects 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 229910001385 heavy metal Inorganic materials 0.000 description 2
- 230000031700 light absorption Effects 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- PUKLDDOGISCFCP-JSQCKWNTSA-N 21-Deoxycortisone Chemical compound C1CC2=CC(=O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@@](C(=O)C)(O)[C@@]1(C)CC2=O PUKLDDOGISCFCP-JSQCKWNTSA-N 0.000 description 1
- QJZYHAIUNVAGQP-UHFFFAOYSA-N 3-nitrobicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid Chemical compound C1C2C=CC1C(C(=O)O)C2(C(O)=O)[N+]([O-])=O QJZYHAIUNVAGQP-UHFFFAOYSA-N 0.000 description 1
- 208000024827 Alzheimer disease Diseases 0.000 description 1
- 102000012437 Copper-Transporting ATPases Human genes 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- FCYKAQOGGFGCMD-UHFFFAOYSA-N Fulvic acid Natural products O1C2=CC(O)=C(O)C(C(O)=O)=C2C(=O)C2=C1CC(C)(O)OC2 FCYKAQOGGFGCMD-UHFFFAOYSA-N 0.000 description 1
- 208000002972 Hepatolenticular Degeneration Diseases 0.000 description 1
- 208000018737 Parkinson disease Diseases 0.000 description 1
- 208000024777 Prion disease Diseases 0.000 description 1
- 206010039966 Senile dementia Diseases 0.000 description 1
- 208000018839 Wilson disease Diseases 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 150000001408 amides Chemical class 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000010668 complexation reaction Methods 0.000 description 1
- 210000002808 connective tissue Anatomy 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- -1 dimethyl methyl Chemical group 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 239000005446 dissolved organic matter Substances 0.000 description 1
- 238000003934 electrogravimetry Methods 0.000 description 1
- 230000002124 endocrine Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000003344 environmental pollutant Substances 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 239000002509 fulvic acid Substances 0.000 description 1
- 229940095100 fulvic acid Drugs 0.000 description 1
- 230000011132 hemopoiesis Effects 0.000 description 1
- 239000004021 humic acid Substances 0.000 description 1
- GPRLSGONYQIRFK-UHFFFAOYSA-N hydron Chemical compound [H+] GPRLSGONYQIRFK-UHFFFAOYSA-N 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-M hydroxide Chemical compound [OH-] XLYOFNOQVPJJNP-UHFFFAOYSA-M 0.000 description 1
- 208000033065 inborn errors of immunity Diseases 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 231100000719 pollutant Toxicity 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 230000001376 precipitating effect Effects 0.000 description 1
- 208000028529 primary immunodeficiency disease Diseases 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000003900 soil pollution Methods 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 230000016776 visual perception Effects 0.000 description 1
Classifications
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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
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
- G01N21/33—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using ultraviolet light
-
- 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
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N21/643—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" non-biological material
-
- 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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- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Pathology (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Engineering & Computer Science (AREA)
- Plasma & Fusion (AREA)
- Molecular Biology (AREA)
- Optics & Photonics (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
- Investigating Or Analyzing Non-Biological Materials By The Use Of Chemical Means (AREA)
Abstract
Inventor provide a kind of method for measuring drug and copper ion complexing power, the following steps are included: 100 μ l drug to be measured and 100 μ l 0.1mmol/L-10mmol/L copper ion solutions are mixed, stirred evenly, stand 1-5min, it is in 0.1mmol/L-5mmol/L rhodamine B hydrazides solution that 2ml concentration, which is added, 3-10min is reacted, prepare liquid is obtained.Above-mentioned technical proposal is then added in the solution of rhodamine B hydrazides after being pre-mixed copper ion and drug to be measured.If drug to be measured can be complexed with copper ion, copper ion competition is formed with rhodamine B hydrazides, so that the rhodamine B hydrazides quantity of open loop reduces, the pink for finally resulting in generation shoals, the intensity of UV absorption and fluorescent emission all declines, and the variation of these signals just embodies the complexing power of drug and copper ion.
Description
Technical field
The present invention relates to a kind of detection method, in particular to a kind of method for measuring drug and copper ion complexing power.
Background technique
Although soil has certain self-purification capacity to pollutant, will cause when harmful substance is more than soil environment capacity
Soil and its relevant ecological system are affected and destroy, at present both at home and abroad to heavy metal in soil pollution and its influence factor
It conducts extensive research, copper can generate toxicity to plant as a kind of necessary microelement of plant, high concentration, toxicity
Depend mainly on the size of the activity of free state copper ion rather than the total amount of copper due to metal ion itself have a fixed number
The positive charge of amount, easily in conjunction with the dissolved organic matter of negatively charged group, the cohesive process of the two will change metal ion and soil
Adsorption equilibrium between earth influences the form of metal ion in the soil and the mobility in pedosphere or even bioavailable
Property.Substance of different nature (drug) is studied to exist to the complexing of copper ion to the heavy metal ion such as copper ion are further clarified
Form, behavior in environment have certain reference role.
Copper is the active constituent of many enzymes of body, is the medium of organism metabolism；Copper can influence the metabolism of iron, join indirectly
With hematopoiesis；Copper also plays key player in the formation of bone and connective tissue and the health of blood vessel.In body, copper
Content must be in a kind of balance state, it is excessive or very few, can all cause a series of lesions of body.When body is lacked in copper
Under weary situation, anoxic, endocrine imbalance, slow and hypoimmunity of development of tissue etc. will cause.Conversely, working as body
Copper overload, can equally induce a variety of diseases, such as: senile dementia, Wilson disease, Parkinson's disease, prion disease and door
Gram this disease etc..Food or drug can form complex compound with copper ion, so that the content of the intracorporal free copper ion of people is interfered, from
And play prevent and treat disease purpose, therefore develop it is a kind of screening drug and copper ion complexing power method have very
Important meaning.
The method of measurement drug and copper ion complexing power has following several at present, but has certain defect.For example, pH drop
Fixed test, for containing the ligand of a large amount of acidic-groups, such as humic acid and fulvic acid have in complex compound forming process it is several
A hydrogen ion is replaced out, usually along with the reduction of solution ph, and usually determines in this, as one kind that complex compound is formed
Property instruction, but this method is not intuitive enough, and only drug contain it is just usable when a large amount of acidic-groups.For another example, AOAC
Method, principle are the multiple pH alkalinity for adjusting solution, precipitate free metal ion shape hydroxide, by precipitating centrifugation point
It is complexing metal amount from, the metal ion content in clear liquid, this method both free metal ion to be separated measures again
Complexation of metal ions content generally requires and uses Atomic absorption etc. costly, operates more complex instrument.For another example, atom is inhaled
Transmit/receive that penetrate instrument analytical methods, the equipment used such as spectroscopic methodology, inductively coupled method, electrochemical process (electrogravimetry) more multiple
It is miscellaneous, and free state and complexed copper ion cannot be distinguished when detecting copper in it.
Summary of the invention
For this reason, it may be necessary to provide a kind of simple and easy, direct visual perception to drug and copper ion complexing power can be used, simultaneously
It again can be in the method for quantitative detection.To achieve the above object, a kind of measurement drug and copper ion complexing power are inventor provided
Method, comprising the following steps:
100 μ l drug to be measured and 100 μ l 0.1mmol/L-10mmol/L copper ion solutions are mixed, stirred evenly, is stood
1-5min, it is to react 3-10min in 0.1mmol/L-5mmol/L rhodamine B hydrazides solution, obtain to be measured that 2ml concentration, which is added,
Liquid；
The discoloration of naked-eye observation prepare liquid；
And/or the ultraviolet absorptivity of detection prepare liquid, when the OD value of prepare liquid is higher than 0.2, by surveyed absorbance and sun
Property control and/or negative control compare, obtain the complexing power of drug and copper ion to be measured；
When the OD value of prepare liquid is less than or equal to 0.2, then detect the fluorescence intensity of prepare liquid, and with positive control and/
Or negative control compares, and obtains the complexing power of drug and copper ion to be measured.
Further, the preparation steps of the rhodamine B hydrazides solution are as follows: weigh rhodamine B hydrazides and mixed solution is added
In, stirring and dissolving；The mixed solution is that water is mixed with organic solvent, and the organic solvent is selected from acetonitrile, methanol, dimethyl methyl
One of amide, isopropanol, ethylene glycol, 1,2- propylene glycol, normal propyl alcohol, glycerol are a variety of.
Further, the mass ratio of the organic solvent and water is 1:1.
Further, the organic solvent is acetonitrile.
Further, the copper ion solution is copper chloride, copper acetate or copper-bath.
Further, the method is negative right with water or the organic solvent for preparing prepare liquid using ETDA as positive control
According to.
Further, the concentration of the copper ion solution is 2.5mmol/L, and the concentration of the rhodamine B hydrazides solution is
2mmol/L。
It is different from the prior art, above-mentioned technical proposal, after copper ion and drug to be measured are pre-mixed, is then added to Luo Dan
In the solution of bright B hydrazides.The solution of rhodamine B hydrazides is that colourless and unstressed configuration then makes rhodamine B after copper ion is added
Hydrazides open loop, color becomes pink and has UV absorption, and can emit fluorescence after by ultraviolet excitation.If drug to be measured
It can be complexed with copper ion, copper ion competition be formed with rhodamine B hydrazides, so that the rhodamine B hydrazides quantity of open loop reduces, most
The pink generated is caused to shoal afterwards, the intensity of UV absorption and fluorescent emission all declines, and the variation of these signals just embodies
The complexing power of drug and copper ion.
Detailed description of the invention
Fig. 1 is the prepare liquid ultraviolet absorptivity figure that different organic solvents are prepared；
Fig. 2 is the prepare liquid fluorescence intensity figure that different organic solvents are prepared；
Fig. 3 is the prepare liquid OD value figure of the copper ion of the hydrazides of rhodamine B containing various concentration and various concentration；Wherein A is
1mmol/L rhodamine B hydrazides is reacted with 2.5mmol/L copper sulphate；B is 1mmol/L rhodamine B hydrazides and 1.25mmol/L sulfuric acid
Copper reaction；C is that 2mmol/L rhodamine B hydrazides is reacted with 2.5mmol/L copper sulphate；D be 2mmol/L rhodamine B hydrazides with
1.25mmol/L copper sulphate reacts.
Fig. 4 is the OD value figure that various concentration EDTA mixes prepare liquid with copper ion；Wherein the concentration of EDTA is in A
The concentration of EDTA is the concentration that the concentration of EDTA in 1.25mmol/L, C is EDTA in 2.5mmol/L, D in 0.625mmol/L, B
The concentration that concentration for EDTA in 5mmol/L, E is EDTA in 10mmol/L, F is 20mmol/L.
Fig. 5 is the OD value figure that various concentration kojic acid mixes prepare liquid with copper ion；Wherein the concentration of kojic acid is in A
The concentration of kojic acid is that the concentration that the concentration of kojic acid in 20mmol/L, C is kojic acid in 10mmol/L, D is in 40mmol/L, B
It is 0.625mmol/L that the concentration of kojic acid, which is the concentration of kojic acid in 1.25mmol/L, F, in 2.5mmol/L, E, the concentration of kojic acid in G
For 0mmol/L.
Fig. 6 is the fluorescence detection figure that various concentration kojic acid mixes prepare liquid with copper ion；Wherein the concentration of kojic acid is in A
The concentration of kojic acid is that the concentration that the concentration of kojic acid in 0.625mmol/L, C is kojic acid in 1.25mmol/L, D is in 0mmol/L, B
The concentration of kojic acid is that the concentration that the concentration of kojic acid in 10mmol/L, F is kojic acid in 20mmol/L, G is in 2.5mmol/L, E
40mmol/L。
Specific embodiment
Technology contents, construction feature, the objects and the effects for detailed description technical solution, below in conjunction with specific reality
It applies example and attached drawing is cooperated to be explained in detail.
OD is the abbreviation of optical density (optical density), indicates the optical density that detected material sponges.
1, influence of the different solvents to the ultraviolet absorptivity and fluorescence signal intensity of rhodamine B hydrazides:
Water is mixed with organic solvent 1:1, prepares mixed solution；The organic solvent is respectively acetonitrile, methanol, dimethyl
Formamide, isopropanol, ethylene glycol, 1,2- propylene glycol, normal propyl alcohol, glycerol.It weighs rhodamine B hydrazides to be added in mixed solution, stir
Dissolution is mixed, preparation concentration is 1mmol/L rhodamine B hydrazides solution；It is with 2ml concentration by 100 μ l 2mmol/L copper-baths
The mixing of 1mmol/L rhodamine B hydrazides solution, reacts 5min, obtains prepare liquid.
Prepare liquid naked-eye observation is pink liquid；Detect the absorbance and fluorescence intensity of prepare liquid, different organic solvents
The prepare liquid absorbance and fluorescence signal intensity of preparation are as depicted in figs. 1 and 2.In remaining condition it can be seen from Fig. 1 and Fig. 2
In identical situation, organic solvent is different, and ultraviolet absorptivity and fluorescence signal intensity are also different, illustrate organic solvent influence copper from
The combination and open loop of son and rhodamine B hydrazides.
And in the reaction system that acetonitrile is solvent, the rhodamine B hydrazides quantity of open loop is most, also shows it in Fig. 1
At 555nm, ultraviolet absorptivity highest reacts susceptibility highest.
In Fig. 2, the fluorescence intensity of acetonitrile system is medium, is lower than propylene glycol and methanol system.But inventor sends out after study
It is existing: this is because rhodamine B hydrazides open loop quantity is excessive, and fluorescence intensity is too high in the reaction system that acetonitrile is solvent, so that
Fluorescence self-extinguishment phenomenon occurs in test.
And but in actual use, only in the case where OD is less than 0.2, i.e. the rhodamine B hydrazides quantity of open loop is very little,
So that just will use fluorescence detection method, therefore also not in the case that ultraviolet light absorption photometer not can be carried out accurate measurement
It can there is a situation where excessively lead to fluorescence self-extinguishment due to rhodamine B hydrazides open loop quantity.
Meanwhile by as it can be seen that propylene glycol and methanol are also preferably solvent selection, being appropriate for ultraviolet light absorption in Fig. 1 and Fig. 2
The detection of degree and fluorescence intensity.
Select ultraviolet detection wavelength of the wavelength of the absorption peak of 555nm in Fig. 1 as subsequent detection.
2, the ultraviolet absorptivity of the rhodamine B hydrazides of the copper sulphate of various concentration and various concentration:
It weighs rhodamine B hydrazides to be added in mixed solution (water is mixed with acetonitrile 1:1), stirring and dissolving, compound concentration is
1mmol/L rhodamine B hydrazides solution is mixed with 100 μ l 2.5mmol/L copper-baths, is reacted 5min, is obtained prepare liquid A.
It weighs rhodamine B hydrazides to be added in mixed solution (water is mixed with acetonitrile 1:1), stirring and dissolving, compound concentration is
1mmol/L rhodamine B hydrazides solution is mixed with 100 μ l 1.25mmol/L copper-baths, is reacted 5min, is obtained prepare liquid B.
It weighs rhodamine B hydrazides to be added in mixed solution (water is mixed with acetonitrile 1:1), stirring and dissolving, compound concentration is
2mmol/L rhodamine B hydrazides solution is mixed with 100 μ l 2.5mmol/L copper-baths, is reacted 5min, is obtained prepare liquid C.
It weighs rhodamine B hydrazides to be added in mixed solution (water is mixed with acetonitrile 1:1), stirring and dissolving, compound concentration is
2mmol/L rhodamine B hydrazides solution is mixed with 100 μ l 1.25mmol/L copper-baths, is reacted 5min, is obtained prepare liquid D.
The OD value that various concentration rhodamine B hydrazides is reacted with the copper ion of various concentration is as shown in Figure 3.By in Fig. 3, see with
The ultraviolet absorptivity and concentration for finding out solution are positively correlated.Wherein, when rhodamine B hydrazides concentration is 2mmol/L, copper sulphate is dense
When degree is 2.5mmol/L, the rhodamine B hydrazides open loop concentration of generation is higher, and OD value significantly improves.
3, difference ETDA concentration influences the ultraviolet absorptivity of rhodamine B hydrazides:
It weighs rhodamine B hydrazides to be added in mixed solution (water is mixed with acetonitrile 1:1), stirring and dissolving, compound concentration is
2mmol/L rhodamine B hydrazides solution；
The EDTA solution of 100 μ l various concentrations is mixed with 100 μ l 2.5mmol/L copper chloride solutions, it is dense to add 2ml
Degree is 2mmol/L rhodamine B hydrazides solution, reacts 5min, obtains prepare liquid.
The concentration of EDTA be respectively 0.625mmol/L, 1.25mmol/L, 2.5mmol/L, 5mmol/L, 10mmol/L,
When 20mmol/L, prepare liquid A, B, C, D, E, F.
The OD value that various concentration EDTA mixes prepare liquid with copper chloride is as shown in Figure 4.By seeing in Fig. 4 with find out with
The OD value of the increase of EDTA concentration, prepare liquid gradually declines, and illustrates the complexing power of EDTA and copper ion with the raising of concentration
And increase；EDTA and copper ion are 1:1 chelating.And when EDTA concentration is 2.5mmol/L and 5mmol/L, the OD value of EDTA connects
Closely, it may be possible to which, since this stage EDTA- copper ion complex compound largely generates, the colour developing of the complex compound causes OD value to rise.
Therefore, as positive control, the optional 0.1-2.5mmol/L of EDTA concentration avoids the area of 2.5mmol/L-5mmol
Between.Meanwhile when the concentration of EDTA is 0.625mmol/L, 1.25mmol/L as seen from Figure 4, OD value is 0.2 or more.
Therefore, the positive control as the detection of ultraviolet OD value, the concentration of EDTA is preferably 0.1-1.25mmol/L.
4, influence of the various concentration drug to be measured (kojic acid) to rhodamine B hydrazides:
It weighs rhodamine B hydrazides to be added in mixed solution (water is mixed with acetonitrile 1:1), stirring and dissolving, compound concentration is
2mmol/L rhodamine B hydrazides solution；
The kojic acid solution of 100 μ l various concentrations is mixed with 100 μ l 2.5mmol/L copper ion solutions, it is dense to add 2ml
Degree is 2mmol/L rhodamine B hydrazides solution, reacts 5min, obtains prepare liquid.Negative controls are prepared with water.
Prepare liquid, it is kojic acid in 20mmol/L, C that wherein the concentration of kojic acid, which is the concentration of kojic acid in 40mmol/L, B, in A
Concentration is that the concentration that the concentration of kojic acid in 10mmol/L, D is kojic acid in 2.5mmol/L, E is the dense of kojic acid in 1.25mmol/L, F
Degree is 0.625mmol/L, and the concentration of kojic acid is 0mmol/L in G.
Naked-eye observation prepare liquid, the color of prepare liquid is from A to G, from light to dark.What various concentration kojic acid and copper ion were complexed
OD value is as shown in Figure 5.By in Fig. 5, it can be seen that with the increase of kojic acid concentration, the OD value of prepare liquid gradually declines, and illustrates song
The complexing power of acid and copper ion increases with the raising of concentration.By Fig. 5 it can also be seen that kojic acid concentration be 0,
When 0.625mmol/L, 1.25mmol/L, 2.5mmol/L, 10mmol/L, OD value is 0.2 hereinafter, needing with carrying out fluorescence detection.
Prepare liquid also carries out fluorescence detection with Fluorescence Spectrometer, and wherein the concentration of kojic acid is kojic acid in 0mmol/L, B in A
Concentration is that the concentration that the concentration of kojic acid in 0.625mmol/L, C is kojic acid in 1.25mmol/L, D is kojic acid in 2.5mmol/L, E
Concentration be the concentration of kojic acid in 10mmol/L, F be the concentration of kojic acid in 20mmol/L, G be 40mmol/L.Testing result is as schemed
Shown in 6, with the rising of kojic acid concentration, fluorescence intensity is gradually reduced.
It should be noted that, in this document, relational terms such as first and second and the like are used merely to a reality
Body or operation are distinguished with another entity or operation, are deposited without necessarily requiring or implying between these entities or operation
In any actual relationship or order or sequence.Moreover, the terms "include", "comprise" or its any other variant are intended to
Non-exclusive inclusion, so that the process, method, article or the terminal device that include a series of elements not only include those
Element, but also including other elements that are not explicitly listed, or further include for this process, method, article or end
The intrinsic element of end equipment.In the absence of more restrictions, being limited by sentence " including ... " or " including ... "
Element, it is not excluded that there is also other elements in process, method, article or the terminal device for including the element.This
Outside, herein, " being greater than ", " being less than ", " being more than " etc. are interpreted as not including this number；" more than ", " following ", " within " etc. understand
Being includes this number.
It should be noted that being not intended to limit although the various embodiments described above have been described herein
Scope of patent protection of the invention.Therefore, it based on innovative idea of the invention, change that embodiment described herein is carried out and is repaired
Change, or using equivalent structure or equivalent flow shift made by description of the invention and accompanying drawing content, it directly or indirectly will be with
Upper technical solution is used in other related technical areas, is included within scope of patent protection of the invention.
Claims (8)
1. a kind of method for measuring drug and copper ion complexing power, which comprises the following steps:
100 μ l drug to be measured and 100 μ l 0.1mmol/L-10mmol/L copper ion solutions are mixed, stirred evenly, 1- is stood
5min, it is to react 3-10min in 0.1mmol/L-5mmol/L rhodamine B hydrazides solution, obtain prepare liquid that 2ml concentration, which is added,；
The discoloration of naked-eye observation prepare liquid；
And/or the ultraviolet absorptivity of detection prepare liquid, it is when the OD value of prepare liquid is higher than 0.2, surveyed absorbance and the positive is right
According to and/or negative control compare, obtain the complexing power of drug and copper ion to be measured；
When the OD value of prepare liquid is less than or equal to 0.2, then detect the fluorescence intensity of prepare liquid, and with positive control and/or yin
Property control compare, obtain the complexing power of drug and copper ion to be measured.
2. the method according to claim 1, wherein the preparation steps of the rhodamine B hydrazides solution are as follows: weigh
Rhodamine B hydrazides is added in mixed solution, stirring and dissolving；The mixed solution is that water is mixed with organic solvent, described organic molten
Agent is selected from one of acetonitrile, methanol, dimethylformamide, isopropanol, ethylene glycol, 1,2- propylene glycol, normal propyl alcohol, glycerol or more
Kind.
3. according to the method described in claim 2, it is characterized in that, the organic solvent is acetonitrile.
4. according to the method described in claim 2, it is characterized in that, the mass ratio of the organic solvent and water is 1:1.
5. the method according to claim 1, wherein the copper ion solution is copper chloride, copper acetate or sulfuric acid
Copper solution.
6. the method according to claim 1, wherein the method is using ETDA as positive control, with water or preparation
The organic solvent of prepare liquid is negative control.
7. the method according to claim 1, wherein the concentration of the copper ion solution be 2.5mmol/L, it is described
The concentration of rhodamine B hydrazides solution is 2mmol/L.
8. the method according to claim 1, wherein ultraviolet detection wavelength is 555nm.
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