CN104215721A - Application of phthalic acid in amino acid analysis - Google Patents
Application of phthalic acid in amino acid analysis Download PDFInfo
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- CN104215721A CN104215721A CN201410527881.2A CN201410527881A CN104215721A CN 104215721 A CN104215721 A CN 104215721A CN 201410527881 A CN201410527881 A CN 201410527881A CN 104215721 A CN104215721 A CN 104215721A
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Abstract
The invention discloses application of phthalic acid in amino acid analysis. The application comprises adding the phthalic acid in a mobile phase for performing amino acid liquid chromatography-mass spectrum combined analysis, wherein concentration of the added phthalic acid is 0.005-0.1mmol/L. According to the application of phthalic acid in amino acid analysis, detection sensitivity of the amino acid can be significantly improved, and meanwhile chromatography separating effect can be improved; the application is convenient that only adding proper amount of the phthalic acid into the mobile phase is needed, and thereby analysis result of the amino acid can be significantly improved without changing other analysis conditions; in addition, experiments show that a chromatographic column and a mass spectrum ion source are prevented from being seriously damaged by using the mobile phase added with the phthalic acid for a long time, reproducibility is good, and quantitative analysis is easy to implement, and good application and promotion values are provided.
Description
Technical field
The present invention relates to the application of phthalic acid in amino acid analysis, specifically, relate to phthalic acid application in amino acid liquid phase chromatograph-mass spectrometer coupling is analyzed as Mobile Phase Additives, belong to Analytical Technology of Amino Acid field.
Background technology
Amino acid is not only the basic composition unit of biosome internal protein, is also the important as precursors of synthesizing the nitrogen-containing compounds such as neurotransmitter, porphyrin, polyamines in body.Recent research shows, amino acid also plays important regulating and controlling effect in vivo, as the phosphorylation of gene expression process, protein, hormonal synthetic and secretion etc.Amino acid existence form in vivo mainly contains two kinds, and one is to be present in physiological fluid with free state, and among blood plasma, urine etc., another kind is to be present in peptide and protein in conjunction with state.Amino acid, as the important component part of biosome intracellular metabolite circulation, all plays very important effect to its qualitative and quantitative analysis in the research in the fields such as protein chemistry, biological chemistry, clinical medicine.
Current amino acid whose separation, during analytical approach is constantly being expanded and is being improved, these methods comprise liquid chromatography and fluorescence detector coupling technique (LC methods coupled with optical detection), LC-MS-MS (is called for short: LC-MS), capillary electrophoresis-mass spectrometry coupling technique (is called for short: CE-MS), gas chromatography-mass spectrography technology (is called for short: GC-MS), NMR) etc. nuclear-magnetism method (is called for short: (referring to H.Kaspar, K.Dettmer, W.Gronwald, P.J.Oefne, Anal.Bioanal.Chem.2009, 393, 445～452, C.W.Klampfl, W.Buchberger, M.Turner, G.S.Fritz, J.Chromatogr.A.1998,804,349～355.).
Liquid phase chromatogram-mass spectrometry combination method is one of amino acid whose important analysis method.Due to liquid chromatography at heat of dissociation advantage and the sensitivity of mass spectrum superelevation and the powerful identification capacity aspect unstable and higher-boiling compound, make LC-MS method become the effective means of compartment analysis complex biological sample.Use LC-MS methods analyst amino acid can also be further divided into two classes, one class is the Direct Analysis of free amino acid, mainly comprise that ion pair reverse-phase chromatography-mass spectrometry method (Ion-pair reversed-phase LC-MS) is (referring to P.Chaimbault, K.Petritis, C.Elfakir, M.Dreux, J.Chromatogr.A.1999, 855, 191～202.) and aqueous favoring interaction chromatography-mass spectroscopy method (be called for short: HILIC-MS) (referring to S.Guo, J.A.Duan, D.W.Qian, Y.P.Tang, , Y.F.Qian, D.W.Wu, S.L.Su, E.X.Shang, J.Agric.Food Chem.2013, 61, 2709～2719.), another kind of is derivatization indirect analysis method, mainly refer to before post chemical derivatization in conjunction with reverse-phase chromatography-mass spectrometry combination method (referring to K.Guo, C.Ji, L.Li, Anal.Chem.2007,79,8631-8638, W.-C.Yang, H.Mirzaei, X.Liu, F.E.Regnier, Anal.Chem.2006,78,4702～4708.).
The Direct Analysis of free amino acid is simple to operate, and required analysis time is short, and the error component of introducing when pre-treatment is less, has comparatively significantly advantage in the time analyzing a large amount of, complicated biological specimen.
Hydrophilic interaction chromatography (is called for short: be HILIC) one of focus of chromatographic field research in recent years.It is a kind of chromatographic technique that retains poor strong polar material retention behavior in reverse-phase chromatography that is used for improving.It is by adopting strong polarity stationarity, and the mobile phase of combination organic phase at high proportion/low ratio water composition is realized this purpose.And such mobile phase composition is particularly advantageous in the mass spectrographic sensitivity of raising electro-spray ionization.While adopting aqueous favoring interaction chromatograph-mass spectrometer coupling technical point isolated amino acid, without sample is carried out to derivatization, sample is direct injected after treatment, can in 20 minutes, realize the separation of several amino acids in conjunction with Ultra Performance Liquid Chromatography.Because amino acid is amphoteric compound, in order to be conducive to its separation and to improve peak shape, conventionally need in mobile phase, add the formic acid of higher concentration and the buffer solution of ammonium formate or acetic acid and ammonium acetate formation, and in mobile phase, the buffer salt of higher concentration can suppress the response of determinand, has affected detection sensitivity.
Summary of the invention
For the above-mentioned problems in the prior art, the object of this invention is to provide phthalic acid application in amino acid liquid phase chromatograph-mass spectrometer coupling is analyzed as Mobile Phase Additives, to improve amino acid whose detection sensitivity and chromatographic resolution effect.
For achieving the above object, the present invention adopts following technical scheme:
The application of phthalic acid of the present invention in amino acid analysis, refer to and adding phthalic acid for the mobile phase of amino acid liquid phase chromatograph-mass spectrometer coupling analysis, the interpolation concentration of phthalic acid is 0.005～0.1mmol/L, is preferably 0.08mmol/L.
As preferred version, described mobile phase is made up of water and organic phase, and all contains formic acid and ammonium formate in water and organic phase.
As further preferred version, in described water, contain the ammonium formate of 5～10mmol/L and the formic acid of 0.1V%～0.3V%, in described organic phase, contain the ammonium formate of 1～5mmol/L and the formic acid of 0.1V%～0.3V%.
As preferred version further, described water is formed by 0.005～0.1mmol/L phthalic acid, 5～10mmol/L ammonium formate, 0.1V%～0.3V% formic acid and water; Described organic phase is formed by 0.005～0.1mmol/L phthalic acid, 1～5mmol/L ammonium formate, 0.1V%～0.3V% formic acid, 0.1V% methyl alcohol and acetonitrile.
As preferred plan, described water is formed by 0.08mmol/L phthalic acid, 10mmol/L ammonium formate, 0.2V% formic acid and water; Described organic phase is formed by 0.08mmol/L phthalic acid, 2mmol/L ammonium formate, 0.2V% formic acid, 0.1V% methyl alcohol and acetonitrile.
As preferred version, described amino acid liquid phase chromatograph-mass spectrometer coupling is analyzed, and comprises the steps:
A) prepare a series of working solutions that formed by standard solution and inner mark solution;
B) adopt the mobile phase that is added with phthalic acid of the present invention, a series of activities solution that step a) is prepared carries out Liquid Chromatography-Tandem Mass Spectrometry analysis, draws internal standard method typical curve;
C) adopt and b) identical mobile phase of step, free amino acid sample to be measured is carried out to Liquid Chromatography-Tandem Mass Spectrometry analysis;
D) according to the amino acid concentration in the internal standard method typical curve calculation sample of having set up.
As further preferred version, the separate mode of described liquid chromatography is hydrophilic interaction chromatography (HILIC), and described mass spectrographic detection mode is multiple-reaction monitoring (MRM).
As further preferred version, the compound method of described standard solution is as follows: accurately take phenylalanine, leucine, isoleucine, tryptophane, methionine, valine, proline, glutamine, halfcystine, alanine, glycocoll, threonine, serine, arginine, glutamic acid, asparagine, histidine, γ-aminobutyric acid, lysine, asparatate and citrulline, hydroxyproline, 2-amino-butyric acid, at least one standard items in the each 10mg of Beta-alanine and tyrosinase 15 mg, after the mixed solvent ultrasonic dissolution being formed for 1:1 by volume by acetonitrile and water with 100mL, add again mixed solvent to be diluted to scale, be diluted to as required again the standard solution of variable concentrations.
As further preferred version, the compound method of described inner mark solution is as follows: the interior mark storing solution that accurately preparation contains four kinds of isotope amino acid internal standard compounds, wherein: the concentration of d5-phenylalanine and d3-alanine is 10 μ g/mL, the concentration of d3-serine and d5-glutamic acid is 100 μ g/mL, is diluted to the inner mark solution of desired concn when use.
As further preferred version, the method for preparing free amino acid sample from biological organization sample is as follows: take the every 10mg of biological organization sample and add 100 μ L deionized waters, fully homogenate; Get homogenate 20 μ L, add inner mark solution 20 μ L and 160 μ L methyl alcohol; After vortex is abundant, centrifugal; Get supernatant liquor 40 μ L to new sample hose, nitrogen dries up; Use 200 μ L by acetonitrile and water by volume for the mixed solvent that 1:1 forms dissolves again.
Compared with prior art, the present invention has following conspicuousness beneficial effect:
1) mobile phase that is added with phthalic acid provided by the invention, can significantly improve amino acid whose detection sensitivity, improves its chromatographic resolution effect simultaneously, and compared with not adding the mobile phase of phthalic acid, amino acid whose detectability can reduce by 4～50 times;
2) mobile phase that is added with phthalic acid provided by the present invention, simple and easy to get, preparation and easy to use, only need in mobile phase, add the phthalic acid of suitable number, without changing other analysis conditions, such as: sample preparation methods, chromatographic separation condition, the MS detection parameters etc., amino acid whose analytical effect will significantly improve;
3) experiment shows, the long-term mobile phase that is added with phthalic acid that uses, all can not cause serious harm to chromatographic column and mass ion source, and favorable reproducibility, is easy to quantitative test;
4) experiment showed, that application the inventive method detects amino acid whose content in analysis human body human thyroid carcinoma and the other normal structure of cancer and can obtain good effect, illustrate that the present invention has good application and popularization value.
Brief description of the drawings
Fig. 1 is the MRM spectrogram obtaining while using respectively mobile phase of the present invention (being added with phthalic acid) and contrast mobile phase (not adding phthalic acid) to detect amino acid mixed standard solution; A figure wherein uses mobile phase of the present invention, and b figure uses contrast mobile phase;
Fig. 2 is the MRM spectrogram obtaining while using respectively mobile phase of the present invention (being added with phthalic acid) and contrast mobile phase (not adding phthalic acid) to detect phenylalanine; A figure wherein uses mobile phase of the present invention, and b figure uses contrast mobile phase;
Fig. 3 is the MRM spectrogram obtaining while using respectively mobile phase of the present invention (being added with phthalic acid) and contrast mobile phase (not adding phthalic acid) to detect proline; A figure wherein uses mobile phase of the present invention, and b figure uses contrast mobile phase.
Embodiment
Below in conjunction with embodiment, technical solution of the present invention is described in further detail and completely.
In embodiment, mass spectrometer used is the LCMS-8040 mass spectrometer of Shimadzu company.
Embodiment 1
The object of the present embodiment is to investigate phthalic acid to have or not the relation between amino acid signal response in mobile phase, to verify that phthalic acid applies to the feasibility of amino acid analysis as Mobile Phase Additives.
1, accurately take phenylalanine, leucine, isoleucine, tryptophane, methionine, valine, proline, glutamine, halfcystine, alanine, glycocoll, threonine, serine, arginine, glutamic acid, asparagine, histidine, γ-aminobutyric acid, lysine, asparatate and citrulline, hydroxyproline, 2-amino-butyric acid, the each 10mg of Beta-alanine and tyrosinase 15 mg, be placed in 100mL volumetric flask, add acetonitrile/water (v/v, 1:1) after ultrasonic dissolution, add again mixed solvent to be diluted to scale, make standard items mixed liquor to be measured,
2, use mobile phase of the present invention: A phase (being water) is formed by 0.08mmol/L phthalic acid, 10mmol/L ammonium formate, 0.2V% formic acid and water; B phase (being organic phase) is formed by 0.08mmol/L phthalic acid, 2mmol/L ammonium formate, 0.2V% formic acid, 0.1V% methyl alcohol and acetonitrile; And contrast mobile phase: A phase (being water) is formed by 10mmol/L ammonium formate, 0.2V% formic acid and water; B phase (being organic phase) is formed by 2mmol/L ammonium formate, 0.2V% formic acid and acetonitrile; And adopt gradient elution program to be: 0min:B 85%, 6min:B 80%, 10min:B 70%, 12-14min:B 60%, then B comes back to mutually 85% balance 8min and carries out sample introduction next time again; The overall flow rate of mobile phase is 0.4mL/min; Column temperature is 40 DEG C; Sample size is 2 μ L; Above-mentioned standard items mixed liquor is carried out respectively to Liquid Chromatography-Tandem Mass Spectrometry analysis, obtain the MRM spectrogram shown in Fig. 1, a figure wherein uses mobile phase of the present invention, and b figure uses contrast mobile phase; As seen from Figure 1: in mobile phase the existence of phthalic acid very large on the impact of amino acid signal intensity, in the time containing phthalic acid in mobile phase, all amino acid whose mass signal intensity all has obvious lifting, is on average lifted at 10 times of left and right.
Accurately take the each 10mg of phenylalanine and proline, be placed in respectively 100mL volumetric flask, add after acetonitrile/water (v/v, 1:1) ultrasonic dissolution, add again mixed solvent to be diluted to scale, make phenylalanine standard solution to be measured and proline standard solution to be measured; Respectively by above-mentioned mobile phase (being added with phthalic acid) and contrast mobile phase (not adding phthalic acid) and analysis condition carry out Liquid Chromatography-Tandem Mass Spectrometry analysis, obtain the MRM spectrogram shown in the MRM spectrogram shown in Fig. 2 and Fig. 3, a figure is wherein and uses mobile phase of the present invention, b figure to be use contrast mobile phase; From Fig. 2 and Fig. 3: contain phthalic acid in mobile phase time, amino acid whose half-peak breadth reduces, post effect obviously raises, separate be improved significantly, further illustrate employing the technology of the present invention, can significantly improve amino acid whose detection sensitivity, can obviously improve its chromatographic resolution effect simultaneously.
Embodiment 2
The object of the present embodiment is to use the mobile phase that is added with phthalic acid, sets up the amino acid whose typical curve of internal mark method determination, investigates the quantitative feasibility of the method.
A) accurately take phenylalanine, leucine, isoleucine, tryptophane, methionine, valine, proline, glutamine, halfcystine, alanine, glycocoll, threonine, serine, arginine, glutamic acid, asparagine, histidine, γ-aminobutyric acid, lysine, asparatate and citrulline, hydroxyproline, 2-amino-butyric acid, the each 10mg of Beta-alanine and tyrosinase 15 mg, be placed in 100mL volumetric flask, after the mixed solvent ultrasonic dissolution being formed for 1:1 by volume by acetonitrile and water with 100mL, add again mixed solvent to be diluted to scale, be diluted to as required again the standard solution of variable concentrations,
B) the interior mark storing solution that accurately preparation contains four kinds of isotope amino acid internal standard compounds, wherein: the concentration of d5-phenylalanine and d3-alanine is 10 μ g/mL, the concentration of d3-serine and d5-glutamic acid is 100 μ g/mL, is diluted to the inner mark solution of desired concn when use;
C) standard solution of the variable concentrations obtaining is mixed as serial working solution with the inner mark solution equal-volume of certain concentration, in working solution, the concentration of d5-phenylalanine and d3-alanine is 0.1 μ g/mL, and the concentration of d3-serine and d5-glutamic acid is 1 μ g/mL;
D) use mobile phase: A phase (being water) is formed by 0.08mmol/L phthalic acid, 10mmol/L ammonium formate, 0.2V% formic acid and water; B phase (being organic phase) is formed by 0.08mmol/L phthalic acid, 2mmol/L ammonium formate, 0.2V% formic acid, 0.1V% methyl alcohol and acetonitrile; And adopt gradient elution program to be: 0min:B 85%, 6min:B 80%, 10min:B 70%, 12-14min:B 60%, then B comes back to mutually 85% balance 8min and carries out sample introduction next time again; The overall flow rate of mobile phase is 0.4mL/min; Column temperature is 40 DEG C; Sample size is 2 μ L; Above-mentioned serial working solution is carried out to Liquid Chromatography-Tandem Mass Spectrometry analysis (every kind of concentration is distinguished sample introduction three times) by low concentration to high concentration successively sample introduction;
E) draw internal standard method typical curve, obtain regression equation: y=ax+b, wherein: y is multiple-reaction monitoring signal ratio=amino acid peak area/interior mark amino acid peak area, and x is amino acid whose mass volume ratio concentration (μ g/mL); In a kind of, mark amino acid can corresponding multiple amino acid to be analyzed.
Table 1 is the analysis results such as the typical curve that obtains, related coefficient, the range of linearity.
Table 1 typical curve, related coefficient, the range of linearity
From table 1 result: each amino acid all becomes good linear relationship, coefficient R in range of linearity concentration
2>0.995, further illustrates the present invention and goes for each amino acid to carry out quantitative test.
Application examples 1
1, take the every 10mg of human thyroglobulin cancerous tissue sample (8 increments originally) and add 100 μ L deionized waters, fully homogenate; Get homogenate 20 μ L, add inner mark solution 20 μ L and the 160 μ L methyl alcohol prepared by embodiment 2; After vortex is abundant, centrifugal; Get supernatant liquor 40 μ L to new sample hose, nitrogen dries up; Use 200 μ L water/acetonitriles (v/v, 1:1) again to dissolve, obtain sample solution to be tested;
2, after centrifugal the sample solution of gained, get supernatant liquor, carry out Liquid Chromatography-Tandem Mass Spectrometry analysis by mobile phase described in embodiment 2 and analysis condition;
3, the internal standard method typical curve of setting up according to embodiment 2 calculates the amino acid concentration in testing human parathyroid tissue sample, and (μ g) to be finally converted into the amount of amino acid containing in tissue samples (g).
Analysis result is as shown in Table 2 below, and in table, ND represents not detect.
Amino acid content (8 increments originally) in table 2 human thyroglobulin cancerous tissue sample
From table 2 result: all have this 25 seed amino acid in human thyroglobulin cancerous tissue, halfcystine lower than quantitative limit, does not detect in part sample.
Application examples 2
1, take the every 10mg of the other normal structure sample of human thyroglobulin cancer (8 increments originally) and add 100 μ L deionized waters, fully homogenate; Get homogenate 20 μ L, add inner mark solution 20 μ L and the 160 μ L methyl alcohol prepared by embodiment 2; After vortex is abundant, centrifugal; Get supernatant liquor 40 μ L to new sample hose, nitrogen dries up; Use 200 μ L water/acetonitriles (v/v, 1:1) again to dissolve, obtain sample solution to be tested;
2, after centrifugal the sample solution of gained, get supernatant liquor, carry out Liquid Chromatography-Tandem Mass Spectrometry analysis by mobile phase described in embodiment 2 and analysis condition;
3, the internal standard method typical curve of setting up according to embodiment 2 calculates the amino acid concentration in testing human parathyroid tissue sample, and (μ g) to be finally converted into the amount of amino acid containing in tissue samples (g).
Analysis result is as shown in Table 3 below, and in table, ND represents not detect.
Amino acid content (8 increments originally) in the other normal structure sample of table 3 human thyroglobulin cancer
From table 3 result: in the other normal structure of thyroid cancer, halfcystine does not all detect.With respect to human thyroid carcinoma, the amino acid content in the other normal structure of cancer is obviously on the low side.
Can further illustrate the mobile phase that is added with phthalic acid provided by the invention by above-mentioned two application examples results and can be used for the amino acid quantitative test in complex biological sample, there is good application and popularization value.
Finally be necessary described herein: above embodiment is only for being described in more detail technical scheme of the present invention; can not be interpreted as limiting the scope of the invention, some nonessential improvement that those skilled in the art's foregoing according to the present invention is made and adjustment all belong to protection scope of the present invention.
Claims (9)
1. the application of phthalic acid in amino acid analysis, is characterized in that: adding phthalic acid for the mobile phase of amino acid liquid phase chromatograph-mass spectrometer coupling analysis, the interpolation concentration of phthalic acid is 0.005～0.1mmol/L.
2. application as claimed in claim 1, is characterized in that: described mobile phase is made up of water and organic phase, and all contains formic acid and ammonium formate in water and organic phase.
3. application as claimed in claim 2, is characterized in that: in described water, contain the ammonium formate of 5～10mmol/L and the formic acid of 0.1V%～0.3V%, contain the ammonium formate of 1～5mmol/L and the formic acid of 0.1V%～0.3V% in described organic phase.
4. application as claimed in claim 3, is characterized in that: described water is formed by 0.005～0.1mmol/L phthalic acid, 5～10mmol/L ammonium formate, 0.1V%～0.3V% formic acid and water; Described organic phase is formed by 0.005～0.1mmol/L phthalic acid, 1～5mmol/L ammonium formate, 0.1V%～0.3V% formic acid, 0.1V% methyl alcohol and acetonitrile.
5. application as claimed in claim 1, is characterized in that, described amino acid liquid phase chromatograph-mass spectrometer coupling is analyzed, and comprises the steps:
A) prepare a series of working solutions that formed by standard solution and inner mark solution;
B) adopt the mobile phase that is added with phthalic acid of the present invention, a series of activities solution that step a) is prepared carries out Liquid Chromatography-Tandem Mass Spectrometry analysis, draws internal standard method typical curve;
C) adopt and b) identical mobile phase of step, free amino acid sample to be measured is carried out to Liquid Chromatography-Tandem Mass Spectrometry analysis;
D) according to the amino acid concentration in the internal standard method typical curve calculation sample of having set up.
6. application as claimed in claim 5, is characterized in that: the separate mode of described liquid chromatography is hydrophilic interaction chromatography (HILIC), and described mass spectrographic detection mode is multiple-reaction monitoring (MRM).
7. application as claimed in claim 5, it is characterized in that, the compound method of described standard solution is as follows: accurately take phenylalanine, leucine, isoleucine, tryptophane, methionine, valine, proline, glutamine, halfcystine, alanine, glycocoll, threonine, serine, arginine, glutamic acid, asparagine, histidine, γ-aminobutyric acid, lysine, asparatate and citrulline, hydroxyproline, 2-amino-butyric acid, at least one standard items in the each 10mg of Beta-alanine and tyrosinase 15 mg, after the mixed solvent ultrasonic dissolution being formed for 1:1 by volume by acetonitrile and water with 100mL, add again mixed solvent to be diluted to scale, be diluted to as required again the standard solution of variable concentrations.
8. application as claimed in claim 5, it is characterized in that, the compound method of described inner mark solution is as follows: the interior mark storing solution that accurately preparation contains four kinds of isotope amino acid internal standard compounds, wherein: the concentration of d5-phenylalanine and d3-alanine is 10 μ g/mL, the concentration of d3-serine and d5-glutamic acid is 100 μ g/mL, is diluted to the inner mark solution of desired concn when use.
9. application as claimed in claim 5, is characterized in that, the method for preparing free amino acid sample from biological organization sample is as follows: take the every 10mg of biological organization sample and add 100 μ L deionized waters, fully homogenate; Get homogenate 20 μ L, add inner mark solution 20 μ L and 160 μ L methyl alcohol; After vortex is abundant, centrifugal; Get supernatant liquor 40 μ L to new sample hose, nitrogen dries up; Use 200 μ L by acetonitrile and water by volume for the mixed solvent that 1:1 forms dissolves again.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106841488A (en) * | 2017-03-06 | 2017-06-13 | 辽宁润生康泰生物医药科技有限公司 | A kind of Liquid Chromatography-Tandem Mass Spectrometry method of sulfur-containing amino acid in non-derivative method detection blood plasma |
| CN109239252A (en) * | 2018-09-10 | 2019-01-18 | 吉尔生化（上海）有限公司 | A kind of detection method for continuous multiple proline polypeptides |
| CN109633011A (en) * | 2018-12-29 | 2019-04-16 | 中山百灵生物技术有限公司 | A kind of detection method of new Glycine Levels |
| CN114414710A (en) * | 2021-12-22 | 2022-04-29 | 深圳天祥质量技术服务有限公司 | Method for detecting polyester isophthalic acid |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106841488A (en) * | 2017-03-06 | 2017-06-13 | 辽宁润生康泰生物医药科技有限公司 | A kind of Liquid Chromatography-Tandem Mass Spectrometry method of sulfur-containing amino acid in non-derivative method detection blood plasma |
| CN109239252A (en) * | 2018-09-10 | 2019-01-18 | 吉尔生化（上海）有限公司 | A kind of detection method for continuous multiple proline polypeptides |
| CN109239252B (en) * | 2018-09-10 | 2020-07-03 | 吉尔生化（上海）有限公司 | Detection method for continuous multiple proline polypeptides |
| CN109633011A (en) * | 2018-12-29 | 2019-04-16 | 中山百灵生物技术有限公司 | A kind of detection method of new Glycine Levels |
| CN114414710A (en) * | 2021-12-22 | 2022-04-29 | 深圳天祥质量技术服务有限公司 | Method for detecting polyester isophthalic acid |
| CN114414710B (en) * | 2021-12-22 | 2023-09-26 | 深圳天祥质量技术服务有限公司 | Detection method of polyester intermediate phthalic acid |
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