CN107014969A - Detect the agent prescription of urea and ammonia content - Google Patents
Detect the agent prescription of urea and ammonia content Download PDFInfo
- Publication number
- CN107014969A CN107014969A CN201610054761.4A CN201610054761A CN107014969A CN 107014969 A CN107014969 A CN 107014969A CN 201610054761 A CN201610054761 A CN 201610054761A CN 107014969 A CN107014969 A CN 107014969A
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- urea
- content
- agent prescription
- ammonia
- detection
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/02—Food
- G01N33/04—Dairy products
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
Abstract
The present invention develops a kind of agent prescription and detection method, is characterized in, can be used together with commercialization blood glucose meter, the content of quick and easy detection urea (or urea nitrogen).Use the agent prescription of the present invention, can be by the interaction between specific coenzyme, zymolyte and enzyme, by urea content in sample it is quantitative be converted into the signal output that commercialization blood glucose meter can be detected, the large-scale instrument testing result of testing result and inspection center is basically identical.
Description
Technical field
The invention belongs to technical field of analytical chemistry, it is related to the quick detection of urea and nitrogen content.
Background technology
Urea nitrogen content in milk（To express urea content from the content of the nitrogen of urea）Detection be evaluate milk
Ox dietary protein plastidome and the important indicator of cows improvement, help to adjust full price, the milk cow of lactating cow diet nutrient
The reasonability of high efficiency and the feed cost input of output.Milk urea nitrogen index, not only can correctly evaluate lactating cow day
The reasonable disposition of degradable albumen and rumen bypass protein in grain albumen system, while one can be entered by milk urea nitrogen index
The resonable degree that lactation cows reproductive efficiency, health status and lactating cow dietary protein are utilized is evaluated and determined to step.
Milk urea nitrogen content is too low to show that degradable protein content is too low in daily ration, and dietary protein digests in cud to be obstructed, and can lead
Cause the decline of dry matter intake and the decline of the output of milk；Milk urea nitrogen content is too high, shows that dietary protein part is unrestrained
Take, cost input is unreasonable.
At present determine milk urea nitrogen main method have diacetyl-oxime method, urase-Podbielniak colorimetric method, enzyme-linked performance rate method,
Infra-red sepectrometry.First three detection method complex operation, process are cumbersome, result accuracy is easily disturbed by extraneous factor；Infrared light
Spectrometry is simple and quick, result is more accurate but higher to equipment requirement.The present invention uses a kind of agent prescription, with commodity
Change blood glucose meter to be used together, measure milk urea nitrogen content that can be quick and easy, testing cost is low, it is easy to operate it is quick, by
The result that external interference is small, result and milk Spot detection method are determined is basically identical.
Detected in addition, the detection method of the present invention also can apply to urea content in the biological samples such as blood, urine.Blood urine
It is to evaluate the whether normal Testing index of renal function that element and urine urea, which are determined, plasma wrea, the rapid and handy for urinating urea content
Detection contributes to relevant disease to investigate and physiology monitor.Meanwhile, detection method of the invention can be used for these biological specimens
The detection of middle ammonia content.
The content of the invention
The invention provides a kind of agent prescription and application method for detecting urea and ammonia content.Pass through reagent in formula
The quantitative relationship that enzyme reaction is set up between urea and ammonia content and commercialization blood glucose meter output result, is successfully realized quick, simple
Urea and ammonia content in milk are measured, measurement result and current common testing methods measurement result are basically identical.This method also may be used
The detection of urea content suitable for the samples such as urine, blood.
1. the agent prescription of the present invention contains following material：
1) coenzyme（Including but not limited to：Reduced nicotinamide adenine dinucleotide（NADH）, reduced form nicotinamide adenine
Dinucleotides phosphoric acid（NADPH）And the salt form such as NADH and NADPH lithium salts, sodium salt, sylvite）
2) α -one class material（Including but not limited to：The salt form such as α-ketoglutaric acid and its lithium salts, sodium salt, sylvite）
3) urease
4) amino acid dehydrogenase（Including but not limited to：Glutamte dehydrogenase, glycine dehydrogenase etc.）Or amino acid transaminase
（Including but not limited to：Alanine aminotransferase, glutamic-pyruvic transaminase etc.）
2. the agent prescription of the present invention generally is prepared into freeze-dried powder or solution, it is easy to use.For example be when being prepared into solution, can be with
Use following several materials and concentration range：
1)0.5~500 mM NADH（Or the salt form such as its lithium salts, sodium salt, sylvite）
2) 1 ~ 1000 mM α-ketoglutaric acids（Or the salt form such as its lithium salts, sodium salt, sylvite）
3) 0.001 ~ 1000 U/mL ureases
4) 0.001 ~ 1000 U/mL glutamte dehydrogenases
5) pH 5 ~ 9 phosphate buffer, phosphorus content is 1 ~ 2000 mM
3. using the method for urea and ammonia content in above-mentioned 1 or 2 reagent detection milk, blood, urine, comprise the following steps：
1) reagent and the sample copy containing urea or ammonia are pressed 1:9~9:After the mixing of 1 volume ratio, after 4 ~ 90 DEG C are reacted 1 ~ 60 minute
Stand to room temperature
2) commodity in use blood glucose meter is measured to reacted sample solution, is obtained signal and is calculated urea or ammonia
Content
Brief description of the drawings
Coenzyme in the reagent that Fig. 1 present invention is provided（By taking NADH as an example）With the linear relationship chart of detector electric signal reading
Fig. 2 present invention provides reagent stability with time chart
Reagent and detecting instrument schematic diagram that Fig. 3 present invention is provided
Fig. 4 application method schematic diagrames of the present invention（By taking milk urea nitrogen as an example）
Embodiment
1. embodiment one：
Urea nitrogen content in ternary plain chocolate is determined with the formula and method of the present invention：1）By external standard method, into milk sample
Be separately added into 0mM, 0.5mM, 1mM, 1.5mM urea, determine milk urea content, and with Beijing Milk Cow Center's test result ratio
Compared with.2）Different urea nitrogen content milk are directly tested, and are compared with 5 kinds of milk samples that Beijing inspection center demarcates.Test
Reagent is matched：0.5 ~ 500 mM NADH sodium salts, 1 ~ 1000 mM α-ketoglutaric acid disodiums salt hydrate, 0.001 ~ 1000 U/
The phosphate buffer of ml ureases, 0.001 ~ 1000 U/ml glutamte dehydrogenases and pH 5 ~ 9（The mM of phosphorus content 1 ~ 2000）.
Method of testing：Buffer solution and sample milk are pressed 1:9~9:1 add freeze-dried reagent in, mixing after 4 ~ 90 DEG C of heating responses 1 ~
60 minutes, after room temperature to be restored, a drop is taken to carry out urea nitrogen content measure with commercialization blood glucose meter.Measure blood glucose meter reading process
It is as follows that standard curve calculating obtains urea nitrogen content result（Each three measurement average values of sample）：
1）External standard method tests milk sample
2）Directly test different urea nitrogen content milk samples
| Sequence number | Certain inspection center tests urea nitrogen content（mg/dL） | Present invention test urea nitrogen content（mg/dL) | Deviation |
| 1 | 9.1 | 9.8 | 7.69% |
| 2 | 12.1 | 12.2 | 1.01% |
| 3 | 15.6 | 15.5 | -0.06% |
| 4 | 17.7 | 17.1 | -3.39% |
| 5 | 21.3 | 21.4 | 0.05% |
2. embodiment two：
Survey with urea nitrogen content in the former breast of the different milk cows of 21 kinds of formula and method measure of the present invention and with Beijing inspection center
Determine result to be compared.Test agent is matched：0.5 ~ 500 mM NADH sodium salts, 1 ~ 1000 mM α-ketoglutaric acid disodium salts
Hydrate, 0.001 ~ 1000 U/ml ureases, 0.001 ~ 1000 U/ml glutamte dehydrogenases and pH 5 ~ 9 phosphate-buffered
Liquid（The mM of phosphorus content 1 ~ 2000）.Method of testing：Buffer solution and sample milk are pressed 1:9~9:1 adds in freeze-dried reagent, mixing
After after 4 ~ 90 DEG C of heating responses 1 ~ 60 minute, room temperature to be restored, a drop is taken to carry out urea nitrogen content survey with commercialization blood glucose meter
It is fixed.Measure blood glucose meter reading and calculate that to obtain urea nitrogen content result as follows by standard curve（Each three measurements of sample are average
Value）：
| Sequence number | Certain inspection center tests urea nitrogen content（mg/dL） | Present invention test urea nitrogen content（mg/dL) | Deviation |
| 1 | 13.23 | 13.2 | 0.23% |
| 2 | 13.10 | 13.4 | -2.24% |
| 3 | 13.37 | 13.6 | -1.69% |
| 4 | 13.25 | 13.4 | -1.12% |
| 5 | 13.42 | 13.7 | -2.04% |
| 6 | 13.52 | 13.3 | 1.65% |
| 7 | 13.41 | 13.6 | -1.40% |
| 8 | 13.66 | 14.3 | -4.48% |
| 9 | 13.71 | 14.0 | -2.07% |
| 10 | 13.68 | 13.2 | 3.64% |
| 11 | 13.72 | 14.4 | -4.72% |
| 12 | 13.66 | 13.7 | -0.29% |
| 13 | 13.46 | 13.2 | 1.97% |
| 14 | 13.62 | 13.3 | 2.41% |
| 15 | 13.69 | 13.4 | 2.16% |
| 16 | 13.69 | 13.6 | 0.66% |
| 17 | 13.67 | 13.2 | 3.56% |
| 18 | 13.85 | 13.6 | 1.84% |
| 19 | 13.60 | 13.2 | 3.03% |
| 20 | 12.66 | 12.8 | -1.09% |
| 21 | 10.60 | 10.9 | -2.75% |
Embodiment
1. NADH concentration and the linear relationship of commercialization blood glucose meter reading are as shown in Figure 1 in the formula of the present invention, it was demonstrated that can
There is quantitative relationship with coenzyme concentration such as blood glucose meter reading and NADH, and after enzyme reaction occurs, NADH consumption and sample
The concentration of middle urea or ammonia is related, therefore can be by blood glucose meter reading come the concentration of quantitative urea or ammonia.
2. the reagent stability of freeze-dried powder changes over time curve as shown in Fig. 2 its method of testing in the formula of the present invention
It is as follows：Reagent is lyophilized to be placed in 4 DEG C of preservations of refrigerator, from freeze-drying time, takes a freeze-dried reagent to add buffering at regular intervals
Liquid, is tested with electrical signal detector and records result.
3. test agent provided by the present invention and detecting instrument schematic diagram are as shown in Figure 3.
4. to detect in milk exemplified by urea content, formula of the invention includes following compound method and application method：Such as
Shown in Fig. 4, reagent each component is included：0.5 ~ 500 mM NADH sodium salts, the hydration of 1 ~ 1000 mM α-ketoglutaric acids disodium salt
Thing, 0.001 ~ 1000 U/ml ureases, 0.001 ~ 1000 U/ml glutamte dehydrogenases and pH 5 ~ 9 phosphate buffer
（The mM of phosphorus content 1 ~ 2000）.Buffer solution, which is kept separately after other components are prepared with water, to be freezed as preservation reagent.The reagent can
More than half a year never degenerated in being preserved at 4 DEG C.A freeze-dried reagent is taken, 1 is added:9~9:The buffer solution and milk of 1 volume ratio, mixing are equal
4 ~ 90 DEG C of heating responses are placed in after even 1 ~ 60 minute, it is to be restored to room temperature, a drop mixed liquor is taken, is directly entered with commercialization blood glucose meter
Row analysis is determined.
Advantages and positive effects of the present invention：
1. reagent prepared by the formula of the present invention, can be used in conjunction with commercialization blood glucose meter simple and easy to get, simple and rapid to determine
The content of urea and ammonia in milk, blood, urine, measurement result and conventional test methodologies result are basically identical
2. the present invention detection method it is with low cost, it is simple to operate, available for milk cow or patient's field portable detection urea and
Ammonia content, operating personnel need not possess professional skill.
Claims (6)
1. a kind of agent prescription, the content for detecting urea and ammonia, it is characterised in that include following material：
1) coenzyme, can be one or more of mixing in following material：Reduced nicotinamide adenine dinucleotide
(NADH), NADPH (NADPH) and NADH and NADPH lithium salts, sodium salt, sylvite
Deng salt form.
2) α -one class material, can be one or more of mixing in following material：α-ketoglutaric acid and its lithium salts, sodium salt,
The salt forms such as sylvite.
3) urease, can be the mixture for one or more of enzymes that urea can be converted into ammonia.
4) amino acid dehydrogenase or amino acid transaminase, can be one or more of mixing in following material：Glutamic acid takes off
Hydrogen enzyme, glycine dehydrogenase, alanine aminotransferase, glutamic-pyruvic transaminase and other amino acid dehydrogenases or transaminase.
2. the solution that according to claim 1 prepared by agent prescription, the combination of several compositions comprising following concentration range：
0.5~500mM coenzyme.
1~1000mM α -one class materials.
0.001~1000U/mL ureases.
0.001~1000U/mL amino acid dehydrogenases.
PH 5~9 phosphate buffer, phosphorus content is 1~2000mM.
3. solution removes the dry powder obtained after a certain amount of moisture or concentrated solution by drying or concentrating according to claim 2.
4. agent prescription according to claims 1 to 3 is used for the method for detecting urea and ammonia content, comprise the following steps：
1) material in agent prescription described in claims 1 to 3 is mixed with testing sample.
2) the blood glucose meter detection said mixture of commodity in use, obtains testing result.
5. formula according to claims 1 to 4 and detection method, the detection for urea content in milk.
6. formula according to claims 1 to 4 and detection method, the detection for urea and ammonia content in blood, urine.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610054761.4A CN107014969A (en) | 2016-01-27 | 2016-01-27 | Detect the agent prescription of urea and ammonia content |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610054761.4A CN107014969A (en) | 2016-01-27 | 2016-01-27 | Detect the agent prescription of urea and ammonia content |
Publications (1)
| Publication Number | Publication Date |
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| CN107014969A true CN107014969A (en) | 2017-08-04 |
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ID=59438846
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| Application Number | Title | Priority Date | Filing Date |
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| CN201610054761.4A Pending CN107014969A (en) | 2016-01-27 | 2016-01-27 | Detect the agent prescription of urea and ammonia content |
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Cited By (3)
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| CN111707631A (en) * | 2020-06-16 | 2020-09-25 | 深圳市锦瑞生物科技有限公司 | Preparation method of reagent ball for urea determination, reagent ball and detection chip |
| CN112126674A (en) * | 2020-08-18 | 2020-12-25 | 兰州百源基因技术有限公司 | Urea detection kit, preparation method and use method thereof |
| CN113075139A (en) * | 2021-03-29 | 2021-07-06 | 迪瑞医疗科技股份有限公司 | Stable double-reagent blood ammonia determination kit |
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| CN111707631A (en) * | 2020-06-16 | 2020-09-25 | 深圳市锦瑞生物科技有限公司 | Preparation method of reagent ball for urea determination, reagent ball and detection chip |
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| CN113075139A (en) * | 2021-03-29 | 2021-07-06 | 迪瑞医疗科技股份有限公司 | Stable double-reagent blood ammonia determination kit |
| CN113075139B (en) * | 2021-03-29 | 2022-10-11 | 迪瑞医疗科技股份有限公司 | Stable double-reagent blood ammonia determination kit |
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Application publication date: 20170804 |