CN111239313A - Rapid contour analysis method of human milk oligosaccharide - Google Patents
Rapid contour analysis method of human milk oligosaccharide Download PDFInfo
- Publication number
- CN111239313A CN111239313A CN201811441066.9A CN201811441066A CN111239313A CN 111239313 A CN111239313 A CN 111239313A CN 201811441066 A CN201811441066 A CN 201811441066A CN 111239313 A CN111239313 A CN 111239313A
- Authority
- CN
- China
- Prior art keywords
- oligosaccharide
- human milk
- concentration
- sample
- centrifuging
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
- G01N2030/8809—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample
- G01N2030/8813—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample biological materials
- G01N2030/8836—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample biological materials involving saccharides
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
Abstract
The invention relates to a rapid contour analysis method of human milk oligosaccharide. The method is characterized in that: the sample is subjected to low-temperature centrifugal degreasing, organic solvent is added for precipitation and protein removal, and then the human milk oligosaccharide composition, structure and content information is obtained by separating through a hydrophilic interaction chromatographic column and taking a mass spectrum as a detector. The method has the characteristics of high selectivity, good stability, simplicity and controllability in operation, high flux and the like, and is suitable for rapid and high-flux analysis of human milk samples.
Description
Technical Field
The invention relates to a rapid contour analysis method for human milk oligosaccharide. The method is characterized in that: and (3) carrying out low-temperature centrifugal degreasing on the sample, adding an organic solvent for precipitation and protein removal, and carrying out separation analysis by using a hydrophilic chromatographic column and mass spectrometry to obtain structural composition analysis. The method has the characteristics of good selectivity, good stability, simple and controllable operation, high flux and the like, and is suitable for rapid analysis of a large number of human milk samples.
Technical Field
Human milk is the sole food source for infants at birth, on the one hand it contains almost all the nutrients necessary for infants, such as proteins, fats, carbohydrates and trace amounts of mineral elements and vitamins etc., on the other hand it has an important role in the improvement of the immune system, growth and development etc. of infants, and the achievement of these functions is closely related to the oligosaccharide components in human milk the oligosaccharide structure of human milk consists of 5 basic saccharide units, i.e. glucose (Glc), galactose (Gal), N-acetylglucosamine (GlcNAc), fucose (Fuc) and sialic acid (N-acetylneuraminic acid-Neu 5 Ac. these oligosaccharides generally have a lactose at the reducing end, which is extended by different numbers of Gal β 1-3GlcNAc (type i) or Gal β 1-4 ac (type ii) units to linear or branched oligosaccharides at the non-reducing end of lactose, with the fucose and sialic acid being extended as modified saccharides in the backbone, respectively β -3 or β -6.
It has been found that the differences in the type and amount of oligosaccharides in different populations are related to the Lewis blood type and secretion type, which depends mainly on the type and concentration variations of fucosylated oligosaccharides in human milk and the in vivo gene-regulated glycosyltransferases.Fuc is present in the body of the secretory population and transfers Fuc α 1-2 to the terminal Gal.thus, in the breast milk of the secretory population, there are complex oligosaccharides containing Fuc α 1-2Gal β 1-3GlcNAc residues, while in the non-secretory population, there are no oligosaccharides of this type.for the Lewis expression population, there are FucT 3 genes in the body encoding α 1-4Fuc onto the GlcNAc residues of the type I chain, and for the non-Lewis expression population, there are no oligosaccharides of this type.
The precondition for deep understanding of the biological functions of human milk oligosaccharides is that the composition, structure and content of the oligosaccharides are analyzed, and particularly, the differences in the types and content of the oligosaccharides can be rapidly analyzed for different human population samples. Therefore, a simple, rapid, high-throughput rapid profiling method for human milk oligosaccharides was developed. However, since there are very many kinds of human milk oligosaccharides and the content of different oligosaccharides varies greatly, how to rapidly characterize the differences of oligosaccharides in different breast milks is a very challenging task.
At present, the method for analyzing the human milk oligosaccharide mainly comprises (1) a high-efficiency anion exchange chromatography method, which mainly depends on an oligosaccharide standard substance, and a sample with the standard substance can be qualitatively and quantitatively analyzed, but the oligosaccharide standard substance is very difficult to obtain, and the structure of the oligosaccharide can not be known aiming at unknown oligosaccharide; (2) the molecular weight of the oligosaccharide of the sample is characterized by matrix-assisted laser ionization mass spectrometry (MALDI-MS), but the method can only obtain the molecular weight information, cannot obtain the specific oligosaccharide structure information, and is difficult to express the sample difference; (3) a liquid chromatography-mass spectrometry (LC-MS) combined technology is adopted, particularly a method based on a graphitized carbon chromatographic column, but the method needs to perform derivatization on a sample, peaks of different oligosaccharides on the graphitized carbon column are overlapped seriously, and the method is difficult to be used as an intuitive sample characterization method to compare differences of human samples.
The invention designs a novel contour analysis method of human milk oligosaccharides, which adopts a hydrophilic chromatographic method to separate oligosaccharides, wherein the oligosaccharides sequentially flow out on a chromatographic column according to the sequence of acid sugar, lactose, neutral sugar and polymerization degree from small to large, the classification of the oligosaccharides is realized according to the retention time and molecular weight of the oligosaccharides, meanwhile, the structural characteristics of the oligosaccharides are described according to characteristic fragment ions given by a secondary mass spectrum, the types of the oligosaccharides are assigned according to the characteristics, and the secretion state of a mother and the Lewis blood type can be distinguished according to the characteristics.
Disclosure of Invention
The invention relates to a human milk oligosaccharide contour analysis method. The invention is characterized in that: the sample is subjected to centrifugal degreasing and precipitation deproteinization, and the sample is directly subjected to hydrophilic chromatographic column analysis and mass spectrometric detection to obtain structure, composition and content analysis.
In order to achieve the purpose, the invention adopts the technical scheme that:
and (3) centrifugally degreasing a sample, adding an organic solvent for precipitation and protein removal, separating by using a hydrophilic chromatographic column, and performing mass spectrum detection. The mobile phase is organic solvent and water or organic solvent and additive water solution, and isocratic or gradient elution conditions are used by optimizing chromatographic parameters.
1. The centrifugal degreasing step comprises the following steps: taking a breast milk sample, centrifuging at 500-50000g for 5-500 minutes at the temperature of 0-10 ℃, and removing upper-layer lipid. Centrifuging for 1-5 times, and collecting the lower layer to obtain defatted breast milk sample.
2. The precipitation protein-removing step comprises the following steps: taking a defatted breast milk sample, adding 1-5 times of organic solvent by volume, mixing uniformly, standing for 2-48 hours at 0-10 ℃, centrifuging for 5-500 minutes at 500-50000g at 0-10 ℃, collecting supernatant, centrifuging for 1-5 times, and evaporating to dryness to obtain a defatted deproteinized sample. The organic solvent is one or more of methanol, acetonitrile, ethanol and acetone.
3. The stationary phase of the HILIC column is polar chromatographic packing and comprises silica gel or polar bonded silica gel packing, and the bonded group comprises one or more of amino acid, amide, amino, carboxyl, glycosyl, zwitter-ion and the like. The organic solvent in the eluent is one or more of methanol, acetonitrile, ethanol and acetone.
a) The concentration and pH of the additive in the mobile phase were as follows:
b) ammonium formate buffer salt, concentration 0-200mM, pH 2.0-7.0;
c) ammonium acetate buffer salt, concentration 0-200mM, pH 2.0-7.0;
d) ammonium bicarbonate buffer salt with concentration of 0-200mM and pH of 6.0-9.0;
e) formic acid additive, concentration 0-10%; the pH value is 1-6;
f) acetic acid additive, concentration 0-10%; the pH value is 1-6;
g) trifluoroacetic acid additive, concentration 0-10%; the pH value is 1-6.
4. The mobile phase gradient was optimized as follows: and (3) using mobile phase water or an aqueous solution added with an additive and an organic solvent as eluent, wherein the mixing ratio is 5/95-95/5.
5. The chromatographic operating parameters were optimized as follows: the inner diameter of the chromatographic column is 2.1-10 mm; the flow rate is 0.1-2 BV/min; the column temperature is 15-60 ℃.
6. The mass spectrum detector comprises one or more of a mass spectrum with electrospray as an ionization source, a quadrupole mass analyzer, a time-of-flight mass analyzer, an ion mobility spectrometry, a quadrupole ion trap mass analyzer and an ion cyclotron resonance mass analyzer.
7. The primary and secondary mass spectrum information including molecular weight and ion intensity is collected by mass spectrum, monosaccharide composition information of the oligosaccharide can be obtained through the molecular weight, oligosaccharide structure information can be obtained through the secondary spectrogram, and content information can be obtained through peak intensity.
8. In the identification of different oligosaccharides, a four-digit code is used to indicate that the first digit represents the number of six-carbon sugars contained in the oligosaccharide (which is the sum of the numbers of glucose and galactose), the second digit represents the number of N-acetylglucosamine, the third digit represents the number of fucose, and the fourth digit represents the number of sialic acid. For example, as shown in FIG. 1, 4221 represents a nonasaccharide consisting of four six carbon sugars, two N-acetylglucosamines, two fucosidases and one sialic acid.
9. And (3) extracting secondary fragments by mass spectrometry, judging the secretory type state by extracting m/z 325, and judging the Lewis blood type state by extracting m/z 348.
The method has the characteristics of high selectivity, good stability, simplicity and controllability in operation, high flux and the like, and is suitable for rapid and high-flux analysis of human milk samples.
The invention has the following advantages:
1. the flux is high. The purification and analysis process is simple and rapid.
2. The repeatability is good. The chromatographic column stationary phase used in the invention has good stability and is easy to realize automation.
3. The method has wide application range and can be used for analyzing different human milk.
4. According to the content of different types of neutral oligosaccharides in the breast milk, the secretion type and Lewis blood type phenotype of the mother are deduced through mass spectrum and characteristic fragment ions.
Drawings
Figure 1 is a graph of oligosaccharide EICs in human milk samples of example 1.
FIG. 2 shows the judgment of blood types of different mothers by secondary mass spectrometry fragments in example 1.
Detailed Description
Example 1:
a certain amount of human milk is taken, centrifuged at 4000g at 4 ℃ for 60min, and upper lipid is removed. And (3) taking the lower water layer, adding 2 times of volume of absolute ethyl alcohol, uniformly mixing, standing at 4 ℃ for 12 hours, centrifuging at 4 ℃ for 60min at 4000g, centrifuging for 1 time, and collecting the upper liquid to obtain the defatted protein-removed oligosaccharide sample.
And directly taking a liquid sample of the human milk oligosaccharide subjected to degreasing and protein removal for analysis. The sample injection amount is 20 mu L, the HILIC analysis column adopts a zwitterionic bonded chromatographic column, the inner diameter of the chromatographic column is 2.1mm, and the flow rate is 2.0 mL/min; the column temperature was 30 ℃, the mobile phase a was acetonitrile, the mobile phase B was water, and the gradient condition was 80% a to 50% a over 40 min. The mass spectrum is electrospray-quadrupole time-of-flight mass spectrum (ESI-Q-Tof), and the mass spectrum conditions are as follows: negative ion mode, capillary voltage 3500V, collision energy 35, and simultaneous acquisition of primary and secondary mass spectra.
Example 2:
centrifuging a certain amount of human milk at 10 deg.C 10000g for 30min, and removing upper layer lipid. And (3) taking the lower water layer, adding acetonitrile with the volume of 3 times, uniformly mixing, standing at 10 ℃ for 2 hours, centrifuging at 10 ℃ for 30min at 10000g, centrifuging for 3 times, and collecting the upper liquid to obtain the degreased protein-removed oligosaccharide sample.
And directly taking a liquid sample of the human milk oligosaccharide subjected to degreasing and protein removal for analysis. The sample injection amount is 20 mu L, the HILIC analysis column adopts a zwitterionic bonded chromatographic column, the inner diameter of the chromatographic column is 2.1mm, and the flow rate is 2.0 mL/min; the column temperature was 30 ℃, the mobile phase a was acetonitrile, the mobile phase B was water, and the gradient condition was 80% a to 50% a over 40 min. The mass spectrum is electrospray-quadrupole time-of-flight mass spectrum (ESI-Q-Tof), and the mass spectrum conditions are as follows: negative ion mode, capillary voltage 3500V, collision energy 35, and simultaneous acquisition of primary and secondary mass spectra.
Example 3:
a certain amount of human milk is taken, centrifuged at 4000g at 4 ℃ for 60min, and upper lipid is removed. And (3) taking the lower water layer, adding 2 times of volume of absolute ethyl alcohol, uniformly mixing, standing at 4 ℃ for 12 hours, centrifuging at 4 ℃ for 4000g for 60min, centrifuging for 1 time, and collecting the upper liquid to obtain the defatted protein-removed oligosaccharide sample.
And directly taking a liquid sample of the human milk oligosaccharide subjected to degreasing and protein removal for analysis. The sample injection amount is 20 mu L, an amide bonding chromatographic column is used as an HILIC analytical column, the inner diameter of the chromatographic column is 2.1mm, and the flow rate is 2.0 mL/min; the column temperature was 30 ℃, the mobile phase a was acetonitrile, the mobile phase B was water, and the gradient condition was 80% a to 50% a over 40 min. The mass spectrum is electrospray-quadrupole time-of-flight mass spectrum (ESI-Q-Tof), and the mass spectrum conditions are as follows: negative ion mode, capillary voltage 3500V, collision energy 35, and simultaneous acquisition of primary and secondary mass spectra.
Example 4:
a certain amount of human milk is taken, centrifuged at 4000g at 4 ℃ for 60min, and upper lipid is removed. And (3) taking the lower water layer, adding 2 times of volume of absolute ethyl alcohol, uniformly mixing, standing at 4 ℃ for 12 hours, centrifuging at 4 ℃ for 4000g for 60min, centrifuging for 1 time, and collecting the upper liquid to obtain the defatted protein-removed oligosaccharide sample.
And directly taking a liquid sample of the human milk oligosaccharide subjected to degreasing and protein removal for analysis. The sample injection amount is 20 mu L, an amide bonding chromatographic column is used as an HILIC analytical column, the inner diameter of the chromatographic column is 2.1mm, and the flow rate is 2.0 mL/min; the column temperature was 30 ℃, mobile phase a was acetonitrile, B was 0.1% formic acid water, and the gradient condition was 80% a to 50% a over 40 min. The mass spectrum is electrospray-quadrupole time-of-flight mass spectrum (ESI-Q-Tof), and the mass spectrum conditions are as follows: negative ion mode, capillary voltage 3500V, collision energy 35, and simultaneous acquisition of primary and secondary mass spectra.
Example 5:
a certain amount of human milk is taken, centrifuged at 4000g at 4 ℃ for 60min, and upper lipid is removed. And (3) taking the lower water layer, adding 2 times of volume of absolute ethyl alcohol, uniformly mixing, standing at 4 ℃ for 12 hours, centrifuging at 4 ℃ for 4000g for 60min, centrifuging for 1 time, and collecting the upper liquid to obtain the defatted protein-removed oligosaccharide sample.
And directly taking a liquid sample of the human milk oligosaccharide subjected to degreasing and protein removal for analysis. The sample injection amount is 20 mu L, an amide bonding chromatographic column is used as an HILIC analytical column, the inner diameter of the chromatographic column is 2.1mm, and the flow rate is 2.0 mL/min; column temperature 30 ℃, mobile phase a acetonitrile, B water, C100 mmol ammonium formate (pH 3.2) gradient conditions a: b: c is 80%: 10%: 10% changed to A over 40 min: b: c is 50%: 40%: 10 percent. The mass spectrum is electrospray-quadrupole time-of-flight mass spectrum (ESI-Q-Tof), and the mass spectrum conditions are as follows: negative ion mode, capillary voltage 3500V, collision energy 35, and simultaneous acquisition of primary and secondary mass spectra.
Example 6:
a certain amount of human milk is taken, centrifuged at 4000g at 4 ℃ for 60min, and upper lipid is removed. And (3) taking the lower water layer, adding 2 times of volume of absolute ethyl alcohol, uniformly mixing, standing at 4 ℃ for 12 hours, centrifuging at 4 ℃ for 4000g for 60min, centrifuging for 1 time, and collecting the upper liquid to obtain the defatted protein-removed oligosaccharide sample.
And directly taking a liquid sample of the human milk oligosaccharide subjected to degreasing and protein removal for analysis. The sample injection amount is 20 mu L, the HILIC analysis column adopts a zwitterionic bonded chromatographic column, the inner diameter of the chromatographic column is 2.1mm, and the flow rate is 2.0 mL/min; the column temperature was 30 ℃, the mobile phase a was acetonitrile, the mobile phase B was water, and the gradient condition was 80% a to 50% a over 40 min. The mass spectrum is electrospray quadrupole ion Trap mass spectrum (ESI-Q-Trap), and the mass spectrum conditions are as follows: the negative ion mode, the declustering voltage of 100V, the collision energy of 35, and the primary and secondary mass spectrograms are collected simultaneously.
Claims (10)
1. A rapid contour analysis method of human milk oligosaccharide is characterized in that: directly feeding the whey liquid obtained after centrifugal degreasing and protein removal by precipitation of a breast milk sample into a high performance liquid chromatography-mass spectrometer for analysis to obtain the composition, structure and relative content information of the human milk oligosaccharide;
high performance liquid chromatography uses hydrophilic chromatography columns for human milk oligosaccharide separation.
2. The analytical method as defined in claim 1, wherein the centrifugal degreasing step comprises: taking a breast milk sample, centrifuging for 5-500 minutes at 500-50000g under the condition of 0-10 ℃, and removing upper-layer lipid; centrifuging to remove upper lipid layer for 1-5 times, and collecting the lower layer to obtain defatted breast milk sample.
3. The assay method of claim 1, wherein the step of removing proteins by precipitation comprises: taking a defatted breast milk sample, adding 1-10 times of organic solvent by volume, mixing uniformly, standing for 1-48 hours at 0-10 ℃, centrifuging for 5-500 minutes at 500-50000g at 0-10 ℃, collecting supernatant, centrifuging, collecting supernatant, evaporating to dryness after 1-5 times, and obtaining a sample of defatted and deproteinized milk supernatant;
the organic solvent is one or more of ethanol, acetone, methanol and acetonitrile.
4. The method according to claim 1, wherein the hydrophilic chromatographic column packing is silica gel or polar group-bonded phase silica gel;
the polar group in the polar group bonded phase silica gel is one or more of amino acid, amide, amino, carboxyl, glycosyl, zwitter ion and the like.
5. The analytical method as defined in claim 1, wherein the high performance liquid chromatography mobile phase is water and an organic solvent, or a buffered saline solution and an organic solvent, and the elution is carried out by an isocratic or gradient method;
the organic solvents in the eluent are: one or more of methanol, acetonitrile, ethanol, isopropanol and acetone.
6. An assay method according to claim 5, wherein the buffer salt type is selected and its concentration and pH in the mobile phase is one of:
a) ammonium formate buffer salt, concentration 0-200mM, pH 6-10;
b) ammonium acetate buffer salt, concentration 0-200mM, pH 6-10;
c) ammonium bicarbonate buffer salt with concentration of 0-200mM and pH of 6-10;
d) formic acid additive, concentration 0-10%; the pH value is 1-6;
e) acetic acid additive, concentration 0-10%; the pH value is 1-6;
f) trifluoroacetic acid additive, concentration 0-10%; the pH value is 1-6.
7. The analytical method according to claim 5 or 6, wherein the volume ratio of the mobile phase water or the buffered saline solution to the organic solvent as the eluent is 5/95 to 95/5.
8. The analytical method according to claim 7, wherein the inner diameter of the column is 2.1 to 4.6mm and the flow rate of the mobile phase is 0.1 to 2 BV/min.
9. The method of claim 1, wherein the mass spectrometry is electrospray ionization mass spectrometry comprising one or more of quadrupole mass analyser, ion mobility spectrometry, time-of-flight mass analyser, quadrupole ion trap mass analyser and ion cyclotron resonance mass analyser;
acquiring primary and secondary mass spectrum information through mass spectrum, wherein the primary and secondary mass spectrum information comprises molecular weight and ion intensity, monosaccharide composition information of the oligosaccharide can be obtained through the molecular weight, oligosaccharide structure information can be obtained through a secondary spectrogram, and content information can be obtained through peak intensity; meanwhile, according to the characteristic fragments, the secretory type state of the mother can be obtained by extracting m/z 325, and the Lewis blood type information of the mother can be obtained by extracting m/z 348.
10. The method of claim 9, wherein the monosaccharide composition of the oligosaccharide is represented by four digits, the first digit representing the number of six carbons (which is the sum of the numbers of glucose and galactose) contained in the oligosaccharide, the second digit representing the number of N-acetylglucosamine, the third digit representing the number of fucose, and the fourth digit representing the number of sialic acid.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811441066.9A CN111239313A (en) | 2018-11-29 | 2018-11-29 | Rapid contour analysis method of human milk oligosaccharide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811441066.9A CN111239313A (en) | 2018-11-29 | 2018-11-29 | Rapid contour analysis method of human milk oligosaccharide |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111239313A true CN111239313A (en) | 2020-06-05 |
Family
ID=70877496
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201811441066.9A Pending CN111239313A (en) | 2018-11-29 | 2018-11-29 | Rapid contour analysis method of human milk oligosaccharide |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111239313A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114594169A (en) * | 2020-12-03 | 2022-06-07 | 中国科学院大连化学物理研究所 | Quantitative detection method for oligosaccharide content in breast milk |
| CN114965750A (en) * | 2021-07-19 | 2022-08-30 | 山东恒鲁生物科技有限公司 | Qualitative and quantitative detection method for human lactooligosaccharide in dairy product |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20040096933A1 (en) * | 2002-11-15 | 2004-05-20 | Yunping Huang | Method of preparation of oligosaccharides |
| CN102101875A (en) * | 2009-12-16 | 2011-06-22 | 中国科学院大连化学物理研究所 | Method for purifying and separating oligosaccharides |
| CN106866749A (en) * | 2015-12-13 | 2017-06-20 | 中国科学院大连化学物理研究所 | A kind of preparation method of breast milk oligosaccharides |
| CN107621399A (en) * | 2016-07-14 | 2018-01-23 | 北京三元食品股份有限公司 | A kind of method of oligosaccharide in detection breast milk |
-
2018
- 2018-11-29 CN CN201811441066.9A patent/CN111239313A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20040096933A1 (en) * | 2002-11-15 | 2004-05-20 | Yunping Huang | Method of preparation of oligosaccharides |
| CN102101875A (en) * | 2009-12-16 | 2011-06-22 | 中国科学院大连化学物理研究所 | Method for purifying and separating oligosaccharides |
| CN106866749A (en) * | 2015-12-13 | 2017-06-20 | 中国科学院大连化学物理研究所 | A kind of preparation method of breast milk oligosaccharides |
| CN107621399A (en) * | 2016-07-14 | 2018-01-23 | 北京三元食品股份有限公司 | A kind of method of oligosaccharide in detection breast milk |
Non-Patent Citations (4)
| Title |
|---|
| CONNIE A. REMOROZA ET AL: "Creating a Mass Spectral Reference Library for Oligosaccharides in Human Milk", 《ANAL. CHEM. 》 * |
| MARKO MANK ET AL: "Label-free targeted LC-ESI-MS2 analysis of human milk oligosaccharides (HMOS) and related human milk groups with enhanced structural selectivity", 《ANALYTICAL AND BIOANALYTICAL CHEMISTRY》 * |
| SARAH M. TOTTEN ET AL: "Comprehensive Profiles of Human Milk Oligosaccharides Yield Highly Sensitive and Specific Markers for Determining Secretor Status in Lactating Mothers", 《J. PROTEOME RES.》 * |
| WENGANG CHAI ET AL: "Negative-Ion Electrospray Mass Spectrometry of Neutral Underivatized Oligosaccharides", 《ANAL. CHEM.》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114594169A (en) * | 2020-12-03 | 2022-06-07 | 中国科学院大连化学物理研究所 | Quantitative detection method for oligosaccharide content in breast milk |
| CN114965750A (en) * | 2021-07-19 | 2022-08-30 | 山东恒鲁生物科技有限公司 | Qualitative and quantitative detection method for human lactooligosaccharide in dairy product |
| WO2023000769A1 (en) * | 2021-07-19 | 2023-01-26 | 山东恒鲁生物科技有限公司 | Qualitative and quantitative detection method for human milk oligosaccharides in dairy product |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Xu et al. | Revisiting monosaccharide analysis–quantitation of a comprehensive set of monosaccharides using dynamic multiple reaction monitoring | |
| Martín-Ortiz et al. | Characterization of goat colostrum oligosaccharides by nano-liquid chromatography on chip quadrupole time-of-flight mass spectrometry and hydrophilic interaction liquid chromatography-quadrupole mass spectrometry | |
| JP5116837B2 (en) | MS method for evaluating glycans | |
| Kottler et al. | Development of a high‐throughput glycoanalysis method for the characterization of oligosaccharides in human milk utilizing multiplexed capillary gel electrophoresis with laser‐induced fluorescence detection | |
| Karch et al. | Identification and quantification of histone PTMs using high-resolution mass spectrometry | |
| WO2024001249A1 (en) | Method for qualitatively detecting neutral oligosaccharides in breast milk | |
| WO2024001248A1 (en) | Method for qualitatively testing acidic oligosaccharides in breast milk | |
| Gautam et al. | Glucose unit index (GUI) of permethylated glycans for effective identification of glycans and glycan isomers | |
| WO2011038874A1 (en) | Method for releasing and labelling 0-glycans | |
| NL2029775B1 (en) | Method for detecting multiple advanced glycation end products in dairy products | |
| CN111239313A (en) | Rapid contour analysis method of human milk oligosaccharide | |
| Rehulka et al. | Microgradient separation technique for purification and fractionation of permethylated N‐glycans before mass spectrometric analyses | |
| CN114509509A (en) | Method for detecting full-spectrum vitamins in serum | |
| CN109613144B (en) | Detection method of catecholamine hormone | |
| CN109937363A (en) | The mass spectrometry method and kit of microorganism for identification | |
| KR101672717B1 (en) | Verifying method for human saliva using human saliva-specific glycan | |
| Wang et al. | A new method for purifying N-Glycans released from milk glycoprotein | |
| Strum et al. | Coupling flash liquid chromatography with mass spectrometry for enrichment and isolation of milk oligosaccharides for functional studies | |
| CN110879256B (en) | Mass spectrum method for high-throughput detection of primary metabolites in corn pollen | |
| CN112924563B (en) | Derivatization-based breast milk oligosaccharide characterization and quantitative analysis method | |
| Raymond et al. | Derivatization of carbohydrates for analysis by liquid chromatography and capillary electrophoresis | |
| Albaladejo et al. | Rapid-Throughput Analysis of Human Milk Oligosaccharides from Filipino Breastmilk | |
| CN107807185B (en) | Method for analyzing endotoxin lipid A by liquid chromatography-mass spectrometry | |
| CN112461959A (en) | Method for identifying selfheal honey and application thereof | |
| CN113237982A (en) | Derivatization method of hydroxyl metabolites and efficient non-targeted metabonomics analysis method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20200605 |