CN107121427B - Reagent for detecting tyrosine phenol metabolites in human urine and preparation method thereof - Google Patents
Reagent for detecting tyrosine phenol metabolites in human urine and preparation method thereof Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 34
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- 238000001514 detection method Methods 0.000 claims abstract description 76
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- ZELCNSAUMHNSSU-UHFFFAOYSA-N 3,5-diamino-2-[(4-sulfamoylphenyl)diazenyl]benzoic acid Chemical compound OC(=O)C1=CC(N)=CC(N)=C1N=NC1=CC=C(S(N)(=O)=O)C=C1 ZELCNSAUMHNSSU-UHFFFAOYSA-N 0.000 claims abstract description 22
- DRXYRSRECMWYAV-UHFFFAOYSA-N mercury(I) nitrate Inorganic materials [Hg+].[O-][N+]([O-])=O DRXYRSRECMWYAV-UHFFFAOYSA-N 0.000 claims abstract description 22
- KBJMLQFLOWQJNF-UHFFFAOYSA-N nickel(ii) nitrate Chemical compound [Ni+2].[O-][N+]([O-])=O.[O-][N+]([O-])=O KBJMLQFLOWQJNF-UHFFFAOYSA-N 0.000 claims abstract description 22
- 238000003756 stirring Methods 0.000 claims description 66
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- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 52
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- 238000006243 chemical reaction Methods 0.000 claims description 22
- 238000000034 method Methods 0.000 claims description 21
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- 229910000372 mercury(II) sulfate Inorganic materials 0.000 claims description 19
- 238000001291 vacuum drying Methods 0.000 claims description 14
- 239000006087 Silane Coupling Agent Substances 0.000 claims description 13
- 239000008367 deionised water Substances 0.000 claims description 13
- 229910021641 deionized water Inorganic materials 0.000 claims description 13
- 238000010992 reflux Methods 0.000 claims description 13
- 238000001132 ultrasonic dispersion Methods 0.000 claims description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 13
- 239000001488 sodium phosphate Substances 0.000 claims description 2
- 229910000162 sodium phosphate Inorganic materials 0.000 claims description 2
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 claims description 2
- 238000003828 vacuum filtration Methods 0.000 claims 1
- 229910000370 mercury sulfate Inorganic materials 0.000 abstract description 3
- 238000000338 in vitro Methods 0.000 abstract description 2
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- CMPGARWFYBADJI-UHFFFAOYSA-L tungstic acid Chemical compound O[W](O)(=O)=O CMPGARWFYBADJI-UHFFFAOYSA-L 0.000 description 12
- IGMNYECMUMZDDF-UHFFFAOYSA-N homogentisic acid Chemical compound OC(=O)CC1=CC(O)=CC=C1O IGMNYECMUMZDDF-UHFFFAOYSA-N 0.000 description 6
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 5
- 238000012360 testing method Methods 0.000 description 5
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- 206010008342 Cervix carcinoma Diseases 0.000 description 2
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- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 2
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- MHUWZNTUIIFHAS-CLFAGFIQSA-N dioleoyl phosphatidic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OCC(COP(O)(O)=O)OC(=O)CCCCCCC\C=C/CCCCCCCC MHUWZNTUIIFHAS-CLFAGFIQSA-N 0.000 description 2
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- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 1
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- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 description 1
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- 230000001225 therapeutic effect Effects 0.000 description 1
- 201000011296 tyrosinemia Diseases 0.000 description 1
- 150000003668 tyrosines Chemical class 0.000 description 1
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- 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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- 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
- 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
- G01N2021/775—Indicator and selective membrane
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Abstract
The invention relates to the field of in-vitro diagnostic reagents, and further particularly relates to a detection reagent for tyrosine phenol metabolites in human urine and a preparation method thereof. The detection reagent for the tyrosine phenol metabolites in the human urine at least comprises: 1-20 mol/L of mercurous nitrate; 1-5 mol/L of mercury sulfate; 0.1-2 mol/L of nickel nitrate; 3-20 mol/L of sulfuric acid; 0.01-1 g/L of phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound.
Description
Technical Field
The invention relates to the field of in-vitro diagnostic reagents, and further particularly relates to a detection reagent for tyrosine phenol metabolites in human urine and a preparation method thereof.
Background
In recent years, the incidence of tumor diseases worldwide is on an increasing trend, and according to the statistical data of the american cancer Society (american cancer Society) on worldwide cancers, the number of newly added cancers worldwide increases from 1270 to 2140 million and the number of cancer deaths increases from 760 to 1320 million from 2008. Meanwhile, recent statistics in our country show that 349 ten thousand new cases of cancer in our country are estimated in 2020, 263 ten thousand death due to malignant tumor are estimated, and 60% [ "8] is increased compared with 2002. It is clear that malignant tumors (cancers) have replaced cardiovascular diseases and become the most deaths in both developed and developing countries. Because cancer is hidden, progresses rapidly and is high in malignancy degree, early detection, early diagnosis and early treatment are still the most important measures for reducing cancer mortality. Because the tumor marker has important guiding significance for the diagnosis, classification, prognosis judgment and treatment of tumors, research on the tumor marker is widely concerned by scholars at home and abroad.
Tyrosine (Tyrosine) belongs to aromatic amino acid, can be lightened from phenylalanine (Phe), can also be directly taken in food, and is an important semi-essential amino acid for human bodies. P-hydroxyphenyllactic acid (PHPLA), DOPA (DOPA) (3, 4-dihydroxylanine) and homogentisic acid (HGA) are important metabolites of tyrosine. Tyrosine metabolic disorder can cause various amino acid metabolic diseases, such as tyrosinemia, homogentisate urinate, and the like, and has certain relevance with liver and kidney diseases, nervous system degenerative diseases, malignant tumor of qi, and the like. Therefore, the research on tyrosine and its metabolites is not only very important in protein chemistry and evaluation of nutritional status of patients, but also important in early diagnosis, therapeutic effect detection, etiology research and the like of various diseases.
The presence of proteins in biological samples can interfere with the determination of tyrosine and its metabolites, which in turn affects the accuracy of the assay.
Disclosure of Invention
In view of the above technical problems, a first aspect of the present invention provides a reagent for detecting tyrosol metabolites in human urine, comprising at least:
in a preferred embodiment, the reagent for detecting a tyrosol metabolite in human urine at least comprises:
in a preferred embodiment, the reagent for detecting a tyrosol metabolite in human urine at least comprises:
in a preferred embodiment, in the phosphomolybdotungstic acid-chitosan-nano silica composite, the weight ratio of the phosphomolybdotungstic acid to the chitosan to the nano silica is (1-10): 1: (5-30).
In a preferred embodiment, in the phosphomolybdotungstic acid-chitosan-nano silica composite, the weight ratio of the phosphomolybdotungstic acid to the chitosan to the nano silica is (3-7): 1: (10-25).
In a preferred embodiment, in the phosphomolybdotungstic acid-chitosan-nano silica composite, the weight ratio of the phosphomolybdotungstic acid to the chitosan to the nano silica is 5: 1: 15.
in a preferred embodiment, the reagent for detecting the tyrosol metabolite in the human urine further comprises sodium phosphate.
The second aspect of the invention provides a preparation method of a reagent for detecting tyrosine phenol metabolites in human urine, which at least comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The third aspect of the invention provides a detection kit for the tyrosine phenol metabolites in the human urine, which comprises the detection reagent for the tyrosine phenol metabolites in the human urine.
The invention provides a method for detecting tyrosine phenol metabolites in human urine, which at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not.
Detailed Description
The disclosure may be understood more readily by reference to the following detailed description of preferred embodiments of the invention and the examples included therein. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In case of conflict, the present specification, including definitions, will control.
When an amount, concentration, or other value or parameter is expressed as a range, preferred range, or as a range of upper preferable values and lower preferable values, this is to be understood as specifically disclosing all ranges formed from any pair of any upper range limit or preferred value and any lower range limit or preferred value, regardless of whether ranges are separately disclosed. For example, when a range of "1 to 5" is disclosed, the described range should be interpreted to include the ranges "1 to 4", "1 to 3", "1 to 2 and 4 to 5", "1 to 3 and 5", and the like. When a range of values is described herein, unless otherwise stated, the range is intended to include the endpoints thereof and all integers and fractions within the range.
The singular forms "a", "an" and "the" include plural referents unless the context clearly dictates otherwise. "optional" or "any" means that the subsequently described event or events may or may not occur, and that the description includes instances where the event occurs and instances where it does not.
Approximating language, as used herein throughout the specification and claims, is intended to modify a quantity, such that the invention is not limited to the specific quantity, but includes portions that are literally received for modification without substantial change in the basic function to which the invention is related. Accordingly, the use of "about" to modify a numerical value means that the invention is not limited to the precise value. In some instances, the approximating language may correspond to the precision of an instrument for measuring the value. In the present description and claims, range limitations may be combined and/or interchanged, including all sub-ranges contained therein if not otherwise stated.
In addition, the indefinite articles "a" and "an" preceding an element or component of the invention are not intended to limit the number requirement (i.e., the number of occurrences) of the element or component. Thus, "a" or "an" should be read to include one or at least one, and the singular form of an element or component also includes the plural unless the stated number clearly indicates that the singular form is intended.
As used herein, each of the following terms has the meaning associated with it in this section. The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. For example, "an element" means one element or more than one element.
As used herein, the term "sensitivity" is defined as a statistical measure of the performance of an assay (e.g., method, test) calculated by dividing the number of true positives by the sum of true positives and false negatives.
As used herein, the term "specificity" is defined as a statistical measure of the performance of an assay (e.g., method, test) calculated by dividing the number of true negatives by the sum of true negatives and false positives.
In view of the above technical problems, a first aspect of the present invention provides a reagent for detecting tyrosol metabolites in human urine, comprising at least:
in a preferred embodiment, the reagent for detecting a tyrosol metabolite in human urine at least comprises:
in a preferred embodiment, the reagent for detecting a tyrosol metabolite in human urine at least comprises:
in the application, mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid are used as main components for developing color of the detection reagent, when the detection reagent is contacted with the tyrosine phenol metabolite, a color reaction occurs, and the positive color is red; weak positive is pink; negatives appeared white or light yellow.
In the application, the content of the mercurous nitrate is 1-20 mol/L, which means that the content of the mercurous nitrate in the whole detection reagent is 1-20 mol/L, and the volume calculation reference is based on the volume of the detection reagent. The content of the mercury sulfate is 1-5 mol/L; the content of mercury sulfate in the whole detection reagent is 1-5 mol/L, and the volume calculation reference is based on the volume of the detection reagent; the content of the nickel nitrate is 0.1-2 mol/L; the content of nickel nitrate in the whole detection reagent is 0.1-2 mol/L, and the volume calculation reference is based on the volume of the detection reagent; the content of the sulfuric acid is 3-20 mol/L; the content of sulfuric acid in the whole detection reagent is 3-20 mol/L, and the volume calculation reference is based on the volume of the detection reagent; the content of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound is 0.01-1 g/L, and the volume calculation reference is based on the volume of the detection reagent.
In the application, the term "phosphomolybdotungstic acid-chitosan-nano silica composite" refers to that chitosan and nano silica are grafted to obtain a chitosan-nano silica composite, and then the chitosan-nano silica composite is loaded with phosphomolybdotungstic acid to obtain a phosphomolybdotungstic acid-chitosan-nano silica composite.
In the application, the phosphomolybdotungstic acid, the chitosan and the nano silicon dioxide are all obtained from markets.
The phosphomolybdic tungstic acid is purchased from Xiamen sea Mark science and technology limited;
the chitosan is purchased from the bioscience and technology company Limited of Xianruin;
the nano silicon dioxide is purchased from Yingchuangdegussa;
in a preferred embodiment, the preparation method of the phosphomolybdotungstic acid-chitosan-nano silica composite at least comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
The inventor unexpectedly discovers that by adding a very small amount of phosphomolybdic tungstic acid-chitosan-nano silicon dioxide compound, on one hand, the alkaloid in urine, such as urea, can be better adsorbed, and the excessive amount of the alkaloid in the urine can be combined with mercurous ions in a system to influence the color reaction and further influence the detection precision; the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and the alkaloid generate physical adsorption and chemical adsorption, and finally precipitate is formed, so that the interference of the alkaloid is accurately eliminated. On the other hand, the addition of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound enables the color of the color reaction to be more obvious, and can accurately judge the positive, weak positive and negative, and the positive is red; weak positive is pink; the negative color is white or light yellow, so that the technical problem that people misjudge the color difference is solved.
The second aspect of the invention provides a preparation method of a reagent for detecting tyrosine phenol metabolites in human urine, which at least comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The third aspect of the invention provides a detection kit for the tyrosine phenol metabolites in the human urine, which comprises the detection reagent for the tyrosine phenol metabolites in the human urine.
The invention provides a method for detecting tyrosine phenol metabolites in human urine, which at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not.
The present invention will be specifically described below by way of examples. It should be noted that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention, and that the insubstantial modifications and adaptations of the present invention by those skilled in the art based on the above disclosure are still within the scope of the present invention.
In addition, the starting materials used are commercially available, unless otherwise specified, and the parts used for the following materials are parts by weight.
Example 1：
Embodiment 1 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 1: 1: 5.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 1.
example 2：
Embodiment 2 provides a reagent for detecting tyrosol metabolites in human urine, which comprises:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 10: 1: 30.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 2.
example 3:
embodiment 3 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 3: 1: 25.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 2.
example 4:
embodiment 4 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 3: 1: 25.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 5.
example 5:
example 5 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 10: 1: 30.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 5.
example 6:
embodiment 6 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 10: 1: 5.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 5.
example 7:
example 7 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 7: 1: 25.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
example 8:
embodiment 8 provides a reagent for detecting tyrosol metabolites in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 3: 1: 10.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 8.
example 9:
example 9 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 5: 1: 15.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 8.
example 10:
example 10 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 5: 1: 15.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
example 11:
example 11 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum drying at 60 ℃ to obtain the chitosan-nano-silica composite.
(2) Dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide composite obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, vacuum filtering, and drying to obtain the phosphomolybdotungstic acid-chitosan-nano silicon dioxide composite.
Wherein the weight ratio of the phosphomolybdic tungstic acid to the chitosan to the nano silicon dioxide is 5: 1: 15.
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
The method for detecting the tyrosine phenol metabolites in the human urine at least comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
comparative example 1：
Comparative example 1 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
and uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid to obtain the reagent for detecting the tyrosine phenol metabolites in the human urine.
The method for detecting the tyrosine phenol metabolites in the human urine comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
comparative example 2：
Comparative example 2 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
and uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate, sulfuric acid and phosphomolybdotungstic acid to obtain the reagent for detecting the tyrosine phenol metabolites in the human urine.
The method for detecting the tyrosine phenol metabolites in the human urine comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
comparative example 3：
Comparative example 3 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
and uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate, sulfuric acid and chitosan to obtain the reagent for detecting the tyrosine phenol metabolites in the human urine.
The method for detecting the tyrosine phenol metabolites in the human urine comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
comparative example 4：
Comparative example 4 provides a reagent for detecting a tyrosol metabolite in human urine, comprising:
the preparation method of the detection reagent for the tyrosine phenol metabolites in the human urine comprises the following steps:
and uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate, sulfuric acid and nano silicon dioxide to obtain the reagent for detecting the tyrosine phenol metabolites in the human urine.
The method for detecting the tyrosine phenol metabolites in the human urine comprises the following steps:
acquiring human urine to be detected;
and (3) contacting and mixing the detection reagent for the tyrosine phenol metabolites in the human urine with the human urine to be detected, and judging whether a color reaction occurs or not. Wherein, the volume ratio of the urine to the detection reagent is as follows: 1: 10.
characterization test
1000 cancer patients with an empty stomach in the morning are selected, and 10mL of urine is respectively taken as human urine to be detected.
The detection method comprises the following steps:
1. the human urine to be tested is respectively contacted and mixed with the human urine in the examples 1-11 and the comparative examples 1-4, and whether the color reaction occurs or not is judged.
Wherein a positive is presented in red; weak positive is pink; negatives appeared white or light yellow.
Table 1 characterization test
The test detection rates of 100 cervical cancer patients, 100 gastric cancer patients, 100 breast cancer patients, 100 liver cancer patients and 100 lung cancer patients were respectively selected from example 5, example 8, example 9, example 10, example 11, comparative example 1, comparative example 2, comparative example 3 and comparative example 4, and the results are shown in table 2.
TABLE 2 cancer detection Rate
As can be seen from tables 1 and 2, the detection reagent for the tyrosine phenol metabolites in the human urine has a very good detection rate, and meanwhile, the applicant unexpectedly finds that the product has a very good detection rate for the cervical cancer.
The foregoing examples are illustrative only, and serve to explain some of the features of the present disclosure. The appended claims are intended to claim as broad a scope as is contemplated, and the examples presented herein are merely illustrative of selected implementations in accordance with all possible combinations of examples. Accordingly, it is applicants' intention that the appended claims are not to be limited by the choice of examples illustrating features of the invention. And that advances in science and technology will result in possible equivalents or sub-substitutes not currently contemplated for reasons of inaccuracy in language representation, and such changes should also be construed where possible to be covered by the appended claims.
Claims (5)
1. A reagent for detecting tyrosine phenol metabolites in human urine, which is characterized by at least comprising:
0.1-0.8 g/L of phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound;
in the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound, the weight ratio of the phosphomolybdotungstic acid to the chitosan to the nano silicon dioxide is 5: 1: 15;
the preparation method at least comprises the following steps:
(1) dissolving nano silicon dioxide in an ethanol solution, performing ultrasonic dispersion for 60min at room temperature, adding a silane coupling agent K560, and stirring for 3h at 45 ℃ to obtain a nano silicon dioxide solution; dissolving chitosan in acetic acid solution, and stirring at room temperature for 3h to obtain chitosan solution; adding the chitosan solution into the nano-silica solution, stirring and reacting for 8h, centrifuging and washing for 3 times, washing for 2 times with ethanol, and vacuum-drying at 60 ℃ to obtain a chitosan-nano-silica compound;
(2) dissolving phosphomolybdotungstic acid in deionized water to prepare a phosphomolybdotungstic acid solution with the mass fraction of 10%, adding the chitosan-nano silicon dioxide compound obtained in the step (1), refluxing and stirring at constant temperature for 20 hours, performing vacuum filtration, and drying to obtain a phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound;
the reagent for detecting the tyrosine phenol metabolites in the human urine also comprises sodium phosphate.
2. The reagent for detecting tyrosine phenol metabolites in human urine as claimed in claim 1, which comprises at least:
0.5g/L of phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound;
in the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound, the weight ratio of the phosphomolybdotungstic acid to the chitosan to the nano silicon dioxide is 5: 1: 15.
3. the method for preparing the reagent for detecting the tyrosol metabolite in the human urine according to any one of claims 1 to 2, characterized by at least comprising the following steps:
the reagent for detecting the tyrosine phenol metabolites in the human urine is obtained by uniformly mixing mercurous nitrate, mercuric sulfate, nickel nitrate and sulfuric acid, then adding the phosphomolybdotungstic acid-chitosan-nano silicon dioxide compound and uniformly stirring.
4. A detection kit for tyrosine phenol metabolites in human urine, which is characterized by comprising the detection reagent for tyrosine phenol metabolites in human urine according to claims 1-3.
5. A method for detecting tyrosine phenol metabolites in human urine is characterized by at least comprising the following steps:
acquiring human urine to be detected;
the reagent for detecting the tyrosol metabolite in the human urine according to any one of claims 1 to 2 is contacted and mixed with the human urine to be detected, and whether a color reaction occurs is judged.
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