CN101446574A - LC-MS quantitative detection method for melamine and tricyanic acid in human urine - Google Patents
LC-MS quantitative detection method for melamine and tricyanic acid in human urine Download PDFInfo
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Abstract
The invention discloses an LC-MS quantitative detection method for melamine and tricyanic acid in human urine. The method is characterized in that the isotope dilution technology is adopted, and in pre-processing of an urine sample, 13C3, 15N3-melamine or deuterated melamine, 13C3 and 15N3-tricyanic acid are respectively added in interior label solution; melamine and tricyanic acid in human urine are respectively extracted by a liquid-liquid extraction method; an LC-MS/MS method is established to respectively and quantitatively detect the concentration of melamine and tricyanic acid in human urine; a liquid chromatogram takes filled columns such as phenyl, fluorophenyl, silica gel and the like as immobile phase; methanol, acetonitrile, water, formic acid solution or mixed liquid as mobile phase; in chromatographic resolution, isocratic elution or gradient elution is carried out; and in mass spectrometric detection, an ESI Source is used for detecting the content of melamine and tricyanic acid in human urine. The minimum limit of quantification of melamine and tricyanic acid can reach 10ng/ml. The processing of a sample by the method in the invention is economical, simple and convenient, in addition, the detection has strong specificity and high sensitivity, and various methodology indicators can reach related technological requirements of the Chinese Pharmacopoeia.
Description
Technical field
The present invention relates to medical detection technique field, be specifically related to LC-MS (the liquid chromatography tandem mass spectrum technology) quantitative detecting method of melamine and cyanuric acid in the people urine.
Background technology
Melamine (Melamine, 2,4,6-triamido-1,3,5-triazines) is a kind of triazines nitrogen heterocyclic ring organic compound, molecular formula C
3N
3(NH
2)
3, molecular weight 126.12.Structural formula is:
Melamine is met strong acid or strong alkali aqueous solution hydrolysis, and amido is progressively replaced by hydroxyl, generates ammeline earlier, and further hydrolysis generates ammelide, generates cyanuric acid at last:
Because protein mainly is made up of amino acid, its nitrogen content generally is no more than 30%, and the molecular formula (C of melamine
3N
6H
6) nitrogen content is 66.7%.The protein method of testing " Kjeldahl " that present stage is general is to estimate protein content by measuring nitrogen content, and therefore, the lawless person adds melamine in food makes the protein test content of food rise.Newton in 1978 and Utley find that melamine has this effect, contain a large amount of melamines in the Sanlu baby milk powder of exposure in the recent period and also are based on same cause.
Toxicity study studies show that melamine toxicity in vivo is very faint, the LD that rat is oral
50(median lethal dose) is 3161mg/kg." international chemicals security manual " the 3rd volume and international chemicals safety card that IPCS in 1994 and European Commission EC compile in collaboration with are pointed out: taking in melamine for a long time or repeatedly in a large number may exert an influence to kidney and bladder, causes producing calculus.Zoopery shows that melamine forms calcium oxalate crystal in urine, with calculi in urinary system tangible relevance is arranged, and can cause renal failure, causes animal dead; In China the ID incident taking place also supports this viewpoint in the recent period.
It is reported that melamine metabolism in different plant species there are differences.Its its half life period in the pig body is approximately 4.04 ± 0.37h, is 1.5 times of mouse body interior (2.71h), and it is former because kidney clearance rate of mouse is 5 times of pig.Melamine purge mechanism in vivo and indeterminate, but according to it as the micromolecular physicochemical property of polarity, and clearance rate (27ml/min) and glomerular filtration rate(GFR (31-42ml/min) in the pig body can be thought and mainly remove through kidney.The poison of male Fischer rat surpassed 99% in 28 hours for studies show that the oral dose of 24h 90% is removed in urine, and the amount of melamine is higher than its hetero-organization in kidney and the bladder.Removing speed is so fast in the body, illustrates that melamine may assemble seldom in vivo.Animal experiment study shows that the apparent branch volume of melamine is 0.61 (± 0.04) L/kg in the pig body, near the total amount of body fluid, so infer that melamine mainly is distributed in extracellular fluid in the pig body, does not exist in the most organs tissue.The existing literature data shows that the metabolic characteristics of melamine in different mammals has bigger difference; Relevant melamine does not appear in the newspapers as yet in the metabolism situation of human body.
Common melamine detection method has high performance liquid chromatography (HPLC method), rp-hplc method, liquid chromatography tandem mass spectrometry, combined gas chromatography mass spectrometry, sublimed method, ion-exchange chromatography-uv detection method at present.Sample source is mainly animal foodstuff, feed, dairy products.Sample treatment needs steps such as Solid-Phase Extraction, filtration, derivatization treatment more, there is no target use in the isotope in the assay method.
Detect melamine in the media report Sanlu baby milk powder in September, 2008, and cause the infant of edible this milk powder that the great food security incident of kidney stone takes place, and caused the concern of the whole society.Liquid chromatography-tandem mass spectrometry (LC-MS/MS) quantitative detecting method of melamine and cyanuric acid during the reliable sensitive people of foundation urinates, so that can in time understand residual condition, the clinical trail observation of melamine in the human body and take effective diagnosis and treatment measure etc., all have very great clinical meaning.
Detection method in people's urine is not appeared in the newspapers as yet about melamine and cyanuric acid.
Summary of the invention
Technical matters to be solved by this invention, be to utilize liquid chromatography-tandem mass spectrometry and isotopic dilution mass spectrum principle, for melamine and cyanuric acid quantitative detecting method in clinical clinical people's urine that a kind of easy and simple to handle, single-minded sensitivity is provided, to satisfy clinical demand.
Quantitative detection method disclosed by the invention comprises: adopt the isotopic dilution technology, add respectively when the pre-service of urine sample and contain 13C3; 15N3-melamine or deuterium substituted melamine and 13C3; The inner mark solution of 15N3-cyanuric acid; Liquid-liquid extraction method extracts melamine and cyanuric acid in the urine respectively; Set up melamine and cyanuric acid concentration in the LC-MS/MS method difference detection by quantitative people urine, liquid chromatography is a stationary phase with filled columns such as phenyl, fluorophenyl, silica gel, methyl alcohol, acetonitrile, water, formic acid solution or mixed liquor are moving phase, chromatographic resolution is carried out isocratic elution or gradient elution, and Mass Spectrometer Method adopts electron spray ionisation source (ESI) to detect melamine and cyanuric acid content in people's urine.The minimum quantitative limit of melamine and cyanuric acid can reach 10ng/ml.
The inventive method may further comprise the steps:
1, sample pretreatment
According to the soda acid character of melamine and cyanuric acid, adopt the sample pretreating method of liquid-liquid extraction to handle melamine and cyanuric acid sample respectively.Melamine alkalizes with NaOH or ammoniacal liquor, and the used extraction solvent of liquid-liquid extraction is an ethyl acetate.Cyanuric acid formic acid acidifying, the used extraction solvent of liquid-liquid extraction is isopropyl alcohol, ethyl acetate or the mixed solvent of the two.
2, the liquid phase separation melamine carries out liquid chromatography respectively with cyanuric acid and separates:
(1) moving phase: be made up of A and B, flow velocity is 0.2～0.5ml, and wherein A is for containing 0.0001～0.1% aqueous formic acid, and B contains the methyl alcohol of 0.0001～0.1% formic acid or the mixed liquor of acetonitrile or methyl alcohol and acetonitrile.
(2) chromatographic column: adopt filled columns such as phenyl, fluorophenyl, cyano group, silica gel, the chromatographic column internal diameter is 1～5mm, and length is 50～250mm.
(3) elution process: adopt degree such as grade or gradient elution mode.When adopting isocratic elution, mobile phase A is 0.02% formic acid water, and Mobile phase B is that the volume ratio that contains 0.02% formic acid is 1: 1 methyl alcohol and an acetonitrile solution, and A:B is volume ratio (20～50): (80～50).When adopting gradient elution, mobile phase A: methyl alcohol (containing 0.001% formic acid), Mobile phase B: water, gradient ratio A is 10～100%, B is 0～90%.Use chromatographic column as XbridgeTMPhenyl (150 * 2.1mm, 5 μ m, Waters), but gradient program reference table 1; Flow velocity 0.35ml/min.
Table 1 eluent gradient program list
| Time (min) | Mobile phase A (%) | Mobile phase B (%) |
| 0 | 96 | 4 |
| 0.1 | 40 | 60 |
| 1 | 40 | 60 |
| 1.1 | 20 | 80 |
| 3 | 20 | 80 |
| 3.1 | 96 | 4 |
| 7 | 96 | 4 |
3, Mass Spectrometer Method
Melamine: adopt electron spray ionisation source (ESI), positive ion multiple-reaction monitoring scanning (MRM), ion selector channel (Q1/Q3): melamine m/z 127/68 (quantitatively) and 127/85 (qualitative), interior mark passage is 13C3; 15N3-melamine m/z 133/72.The ion channel scope can be ± 1.0amu.
Cyanuric acid: adopt electron spray ionisation source (ESI), negative ion multiple-reaction monitoring scanning (MRM), ion selector channel (Q1/Q3): m/z 128/42 (quantitatively) and 128/85 (qualitative), interior mark 13C3; 15N3-cyanuric acid m/z 134/44.The ion channel scope can be ± 1.0amu.
4, density calculating method and detectability
With peak area or peak height method, calculate melamine and cyanuric acid content in the urine by internal standard method respectively.
The detection of melamine of the present invention and cyanuric acid is limited to 1ng/ml, and quantitatively scope is 10～5000ng/ml, and linearly dependent coefficient is more than 0.9990, in batch, betweenrun precision is less than 10%, between the accuracy 85～115%.Sample stability is good.
Useful result of the present invention is:
1. the physicochemical property different with cyanuric acid according to melamine, the present invention carries out sample preparation, chromatographic resolution and Mass Spectrometer Method to it respectively.Set up the preprocess method of melamine and cyanuric acid in the easy people's urine of relatively economical.The sample pretreating method of liquid-liquid extraction has been adopted in this research, under the prerequisite that satisfies the every index of methodology, compares with existing Solid-Phase Extraction method, and is more quick, easy and economical.
2. adopt the isotopic dilution standard measure, when the pre-service of urine sample, just add in melamine and the cyanuric acid stable isotope and mark, realized the tracking correction of the overall process of test is guaranteed that measurement result accurately and reliably.
3. the inventive method is strong, highly sensitive, easy and simple to handle to the selectivity of sample detection, and every methodology index all can reach the correlation technique requirement of Chinese Pharmacopoeia.
Embodiment
The invention will be further elaborated below in conjunction with specific embodiment, but do not limit the present invention.
Non-special declaration is concentration expressed in percentage by volume among the embodiment.
Embodiment 1
Testing conditions and method example
1. reagent preparation:
(1) typical curve: it is an amount of to draw melamine and cyanuric acid standard operation liquid, adds dual distilled water, makes respectively to contain that melamine and cyanuric acid concentration are 10,50,100,500,1000,2500, the standard water solution of 5000ng/ml.Contain 7 concentration point, each concentration preparation 1ml.
(2) quality-control product: it is an amount of to draw melamine and cyanuric acid standard operation liquid, adds dual distilled water, makes respectively to contain melamine and cyanuric acid concentration is 10,20,2000, the melamine of 4000ng/ml and cyanuric acid are as quality-control product, contain 4 concentration point, each concentration preparation 2ml.
(3) mark in: draw and mark (100 μ g/ml) solution in the 250 μ l melamines to the 10ml volumetric flask, with 50% methanol aqueous solution constant volume, be mixed with 2.5 μ g/ml working solutions, 4 ℃ of refrigerations are standby, get 2.5 μ g/ml melamine inner mark solutions.Simultaneously, draw in addition and mark (100 μ g/ml) solution in the 250 μ l cyanuric acids to the 10ml volumetric flask, with 50% methanol aqueous solution constant volume, be mixed with 2.5 μ g/ml working solutions, 4 ℃ of refrigerations are standby, get 2.5 μ g/ml cyanuric acid inner mark solutions.
(4) moving phase (methyl alcohol: acetonitrile is 1:1, contains 0.02% formic acid solution): 500ml methyl alcohol is mixed with the 500ml acetonitrile, get 200 μ l formic acid solutions in mixed liquor, mixing, promptly.
(5) moving phase (containing 0.02% aqueous formic acid): get 200 μ l formic acid solutions in 1000ml water, mixing, promptly.
(6) moving phase (containing 0.001% formic acid methanol solution): get 10 μ l formic acid solutions in 1000ml methyl alcohol, mixing, promptly.
(7) 1% aqueous formic acids: get the 1ml formic acid solution in 100ml water, mixing, promptly.
(8) 0.1% aqueous formic acids: get 100 μ l formic acid solutions in 100ml water, mixing, promptly.
(9) 1% sodium hydroxide solutions: take by weighing 1g NaOH, be dissolved in the 100ml water, mixing, promptly.
(10) extract solvent: get isopropyl alcohol and ethyl acetate, the ratio of 5:95 is mixed by volume.Preparation cumulative volume 1000ml.
2. urine sample gathering:
Gather the about 2ml of clinical urine sample, then-80 ℃ of preservations.During sample determination, get under the frozen sample room temperature and thaw, standby.
3. sample pretreatment:
3.1 measure melamine treatment step in people's urine sample
Get 100 μ l urine samples, add mark working fluid (5 μ g/ml) and 20 μ l 1N NaOH solution in the 10 μ l melamine isotopes successively, vortex 10s adds 1ml ethyl acetate, vortex vibration 10min, the centrifugal 3min of 18000rpm; Draw upper organic phase, add the aqueous solution that 100 μ l contain 0.1% formic acid, vortex vibration 10min, the centrifugal 3min of 18000rpm; Drawing lower layer of water phase transfer to 300 μ l plastic tube sample introduction analyzes.
3.2 measure cyanuric acid treatment step in people's urine sample
Get 100 μ l urine samples, add the aqueous solution that 10 μ l cyanuric acid isotopes interior mark working fluid (5 μ g/ml) and 20 μ l contain 1% formic acid successively, vortex 10s adds the ethyl acetate solution that 1ml contains 5% isopropyl alcohol, vortex vibration 10min, the centrifugal 3min of 18000rpm; Draw upper organic phase, pure air dries up for 37 ℃, and residue adds 100 μ L and contains 70% methanol in water, vortex vibration 3min, and the centrifugal 3min of 18000rpm is transferred to 300 μ l plastic tube sample introduction analyses.
4. liquid chromatography tandem mass spectrum (LC-MS/MS) condition determination
4.1 chromatographic condition
4.1.1 measure the melamine chromatographic condition
Chromatographic column: Allure PFP Propyl (100 * 2.1mm, 5 μ m, RESTEK)
Guard column: C18 (4.0 * 3.0mm, 5 μ m, Phenomenex)
Moving phase: mobile phase A: distilled water (containing 0.02% formic acid)
Mobile phase B: methyl alcohol/acetonitrile (1:1 contains 0.02% formic acid)
Moving phase ratio: A:B=70:30
Flow velocity: 0.4ml/min
Column temperature: room temperature
Sample size: 5 μ l
Typical case's retention time: melamine: 1.9min; Interior mark: 1.9min
4.1.2 measure the cyanuric acid chromatographic condition
Chromatographic column: XbridgeTM Phenyl (150 * 2.1mm, 5 μ m, Waters)
Guard column: C18 (4.0 * 3.0mm, 5 μ m, Phenomenex)
Moving phase: methyl alcohol (containing 0.001% formic acid)
Flow velocity: 0.35ml/min
Column temperature: room temperature
Sample size: 5 μ l
Typical case's retention time: cyanuric acid: 1.3min; Interior mark: 1.3min
4.2 mass spectrum condition
4.2.1 measure melamine mass spectrum condition
Instrument API3000
Ion gun: electric spray ion source
Scan mode: positive ion MRM scanning
Source parameters: atomization gas: 60psi (10*); Gas curtain gas: 40psi (10*); Collision gas: 42psi
(7*); Auxiliary gas: 7L/min; Voltage: 5500V; Temperature: 450 ℃
(annotate: * is the instrument setting value)
4.2.2 measure cyanuric acid mass spectrum condition
Instrument 3200 Q Trap
Ion gun: electric spray ion source
Scan mode: negative ion MRM scanning
Source parameters: atomization gas: 50*; Gas curtain gas: 20*; Collision gas: High*; Auxiliary gas: 50*;
Press :-4500V; Temperature: 500 ℃
(annotate: * is the instrument setting value)
5. density calculating method
Press internal standard method with peak area or peak height method, the concentration of melamine and cyanuric acid in the calculating urine.
6. result
Melamine and cyanuric acid method for measuring comprise that linear relationship, accuracy, precision, stability etc. all meet the regulation of Chinese Pharmacopoeia.
Embodiment 2
Chromatographic condition
The method of reagent preparation, sample collecting and sample preparation, testing conditions are with embodiment 1.In the chromatographic condition melamine use instead Thermo Hypersil Silica (150mm * 2.1mm i.d., 5 μ m, Thermo) analytical column, isocratic elution, moving phase (A) 0.0001% formic acid water, (B) 0.0001% formic acid acetonitrile, A:B is volume ratio 15:85.With this understanding, melamine and interior target retention time are 2.6min.Cyanuric acid employing RESTEK Allure PFP Propyl (100mm * 2.1mm i.d., 5 μ m, RESTEK) analytical column, isocratic elution, moving phase (A) 0.1% formic acid water, (B) 0.1% formic acid, 50% acetonitrile, 50% methyl alcohol, A:B is volume ratio 30:70.With this understanding, cyanuric acid and interior target retention time are 1.3min.The chromatographic peak peak shape of melamine and cyanuric acid is good, does not see to disturb at corresponding chromatographic retention to exist.
Embodiment 3
The methodology checking
Adopt embodiment 1 listed testing conditions and method, assay method has been carried out the methodology checking.In the concentration range of 10-5000ng/ml melamine and cyanuric acid, linear relationship good (r 〉=0.9990), lowest detectable limit can reach 1ng/ml.In batch, betweenrun precision RSD respectively≤3.26% and≤5.43%, accuracy is 93.7～102.8%.Extraction recovery is 78.0～83.3%, and RSD≤4.6%, medium effect are 96.4～97.6%.The presentation of results LC-MS/MS of methodology checking can be sensitive, accurately and rapidly clinical sample in enormous quantities is detected.
1. typical curve and quantitatively scope
Press embodiment 1 method preparation typical curve, handle in three batches and sample introduction is measured, altogether 3 overlap typical curves.With concentration is the x axle, and the ratio of mark peak area is the y axle in melamine and cyanuric acid and the isotope separately, carries out linear regression, sets up regression equation.The result shows: the typical curve equation is Y=0.0304X+0.04036 (melamine), Y=0.00406X+0.01367 (cyanuric acid).The r value of 3 batches of bioassay standard curves is respectively 0.9994～0.9998.See table 2 for details.The minimum 10ng/ml that quantitatively is limited to of this law melamine and cyanuric acid.
Table 2 standard curve determination result (3 batches, every crowd of n=2)
2. accuracy and precision
Press embodiment 1 method preparation quality-control product.Every kind of concentration is prepared 6 parts, handles and analyzes by above-mentioned sample treatment and liquid chromatography tandem mass spectrum condition on the same day, calculates withinrun precision and accuracy.As stated above, divide 3 days preparation 3 batch samples to handle and analyze, calculate betweenrun precision and accuracy.The results are shown in Table 3, batch interior, betweenrun precision CV%≤10% are between the accuracy 88.91～111.6%.
Table 3 accuracy and precision result (3 batches, every crowd of n=6)
Annotate: MLM: melamine, CA: cyanuric acid
3. extraction recovery
Melamine, cyanuric acid and interior separately target extraction recovery see Table 4, and the result shows that the average extraction recovery of the melamine of basic, normal, high concentration quality-control sample is respectively 17.69%, 20.23% and 20.88%; The target extraction recovery is 19.68% in the melamine; The average extraction recovery of cyanuric acid is respectively 70.88%, 71.51% and 64.32%; The target extraction recovery is 64.59% in the cyanuric acid.
Table 4 melamine, cyanuric acid and interior target extraction recovery
4. stable
4.1 sample stability of (room temperature) in automatic sampler
Three concentration quality-control samples after the operation of sample preparation step, are put into automatic sampler (room temperature), measure at different time, and it the results are shown in Table 5, show that the sample after the extraction is stable in the 6h in automatic sampler.
Table 5 automatic sampler stability result
4.2 the stability that the sample room temperature is placed
The concentration determination value that the quality-control sample room temperature is placed in the 24h the results are shown in Table 6.The result shows that sample keeps stable in room temperature is placed 24h.
The stability result that table 6 sample room temperature is placed
4.3 many freezing-thawing tests
The results are shown in Table 7 with concentration determination value after the freeze thawing without the quality-control sample of freeze thawing.The result shows that the still maintenance after three concentration quality-control samples are through 3 freeze thawing is stable.
Table 7 freezing-thawing test stability result
Embodiment 4
Measurement result in the clinical urine sample
Press condition and the method for embodiment 1, during September 19 to 1 day October in 2008 in 2008, collect " milk powder outpatient service " infant's urine sample 86 examples, age 0-8 year.Collect adult's health examination urine sample 110 example age 25-57 year.Melamine and cyanuric acid measurement result see Table 8 in the urine sample.
In table 8 " milk powder outpatient service " infant and the health examination urine sample
The CONCENTRATION DISTRIBUTION scope and the ratio of melamine and cyanuric acid
As seen from Table 8,96 examples (87.2%) detect melamine in the 110 routine health examination urine samples, and 109 examples (99.1%) detect cyanuric acid.Illustrate adopt isotopic dilution liquid chromatography tandem mass spectrometry can be sensitive, accurately melamine in the urine sample and cyanuric acid are carried out detection by quantitative.
Claims (5)
1. the LC-MS detection by quantitative method of melamine and cyanuric acid in people's urine is characterized in that, with the isotopic dilution standard measure, adds the inner mark solution that contains melamine and cyanuric acid stable isotope when the pre-service of urine sample; Liquid-liquid extraction method extracts melamine and cyanuric acid in the urine respectively; Melamine and cyanuric acid content in the liquid chromatography tandem mass spectrum technology for detection people urine; Press internal standard method with peak area or peak height method, the concentration of melamine and cyanuric acid in the calculating urine.
2. according to the described quantitative detecting method of claim 1, it is characterized in that mark is respectively 13C3 in the stable isotope that adds when the urine sample pre-service; 15N3-melamine or deuterium substituted melamine, 13C3; The 15N3-cyanuric acid.
3. according to the described quantitative detecting method of claim 1, it is characterized in that described liquid-liquid extraction is that sample is with NaOH or ammoniacal liquor alkalization, then with extracting solvent ethyl acetate extraction melamine; Sample formic acid acidifying, again with extraction solvent isopropyl alcohol or ethyl acetate, or the mixed solvent of isopropyl alcohol and ethyl acetate extracts cyanuric acid.
4. according to the described quantitative detecting method of claim 1, it is characterized in that, be stationary phase with filled columns such as phenyl, fluorophenyl, silica gel during chromatographic resolution, and methyl alcohol, acetonitrile, water, formic acid solution or mixed liquor are moving phase, chromatographic resolution, isocratic elution or gradient elution.
5. according to the described quantitative detecting method of claim 1, it is characterized in that Mass Spectrometer Method electron spray ionization source; (1) melamine scans (MRM) pattern with the positive ion multiple-reaction monitoring, ion selector channel (Q1/Q3): quantitative passage m/z 127/68 and qualitative passage m/z 127/85, and interior mark passage is 13C3,15N3-melamine m/z 133/72;
(2) cyanuric acid scans (MRM) pattern with the negative ion multiple-reaction monitoring, ion selector channel (Q1/Q3): quantitative passage m/z 128/42 and qualitative passage m/z 128/85, interior mark passage 13C3,15N3-cyanuric acid m/z 134/44.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102133488A (en) * | 2010-01-26 | 2011-07-27 | 上海安谱科学仪器有限公司 | Solid-phase extraction column for synchronously extracting melamine and cyanuric acid as well as preparation method and application thereof |
| CN104965035A (en) * | 2015-04-27 | 2015-10-07 | 公安部物证鉴定中心 | Method for screening toxic substances in sample by using solid phase support liquid-liquid extraction-GC MS |
| CN105277634A (en) * | 2015-09-30 | 2016-01-27 | 中国兽医药品监察所 | Method for simultaneously and quantitatively detecting contents of melamine and cyanuric acid and application thereof |
| CN105866281A (en) * | 2016-04-25 | 2016-08-17 | 广西壮族自治区梧州食品药品检验所 | Method for detecting melamine in milk powder through gas chromatographic method |
| CN108982375A (en) * | 2012-03-02 | 2018-12-11 | 博格有限责任公司 | For detecting the method and kit of Co-Q10 |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101122589B (en) * | 2007-09-05 | 2011-01-12 | 刘忠发 | High performance liquid chromatography analysis method of urea and its impurity carbamylurea, methylene biuret |
-
2008
- 2008-10-10 CN CN2008102009880A patent/CN101446574B/en not_active Expired - Fee Related
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102133488A (en) * | 2010-01-26 | 2011-07-27 | 上海安谱科学仪器有限公司 | Solid-phase extraction column for synchronously extracting melamine and cyanuric acid as well as preparation method and application thereof |
| CN102133488B (en) * | 2010-01-26 | 2013-09-04 | 上海安谱科学仪器有限公司 | Solid-phase extraction column for synchronously extracting melamine and cyanuric acid as well as preparation method and application thereof |
| CN108982375A (en) * | 2012-03-02 | 2018-12-11 | 博格有限责任公司 | For detecting the method and kit of Co-Q10 |
| CN104965035A (en) * | 2015-04-27 | 2015-10-07 | 公安部物证鉴定中心 | Method for screening toxic substances in sample by using solid phase support liquid-liquid extraction-GC MS |
| CN105277634A (en) * | 2015-09-30 | 2016-01-27 | 中国兽医药品监察所 | Method for simultaneously and quantitatively detecting contents of melamine and cyanuric acid and application thereof |
| CN105866281A (en) * | 2016-04-25 | 2016-08-17 | 广西壮族自治区梧州食品药品检验所 | Method for detecting melamine in milk powder through gas chromatographic method |
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