CN110658292B - Method for detecting geranyl ester pyrophosphate content based on prenylation reaction - Google Patents
Method for detecting geranyl ester pyrophosphate content based on prenylation reaction Download PDFInfo
- Publication number
- CN110658292B CN110658292B CN201810702345.XA CN201810702345A CN110658292B CN 110658292 B CN110658292 B CN 110658292B CN 201810702345 A CN201810702345 A CN 201810702345A CN 110658292 B CN110658292 B CN 110658292B
- Authority
- CN
- China
- Prior art keywords
- sample
- dansyl
- geranyl pyrophosphate
- detected
- recovery
- 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.)
- Active
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
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
- G01N30/08—Preparation using an enricher
-
- 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
- G01N30/62—Detectors specially adapted therefor
- G01N30/74—Optical detectors
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)
- Spectroscopy & Molecular Physics (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a detection method of geranyl pyrophosphate content, which comprises the steps of purifying and enriching a sample to be detected, designing and synthesizing two fluorescent polypeptides which are subjected to prenylation reaction with geranyl pyrophosphate according to the principle of prenylation reaction, adding the two polypeptides into a sample solution to be detected and a solution containing a quantitative geranyl pyrophosphate pure product respectively to perform prenylation reaction, obtaining a target compound and an internal standard substance for indirectly detecting geranyl pyrophosphate respectively, detecting the target compound and the internal standard substance by using a high performance liquid chromatograph-fluorescence detector, obtaining an initial detection value of geranyl pyrophosphate, designing a recovery rate experiment, calculating the mass of geranyl pyrophosphate lost during the purification of the sample to be detected, establishing a recovery rate correction curve to correct the initial measurement value, and finally obtaining the geranyl pyrophosphate content. The method provided by the invention is stable and reliable, has high reproducibility, and the application range covers the detection of geranyl pyrophosphate in different organisms.
Description
Technical Field
The invention belongs to the technical field of pathophysiology and metabolite detection, relates to a method for detecting the content of metabolites in organisms, and particularly relates to a method for detecting the content of geranyl pyrophosphate as a metabolite in organisms.
Background
Geranyl pyrophosphate (GGPP) is a 20-carbon isoprenoid derivative that is synthesized in vivo via the mevalonate pathway from the sole source in the organism. Geranyl pyrophosphate participates in post-translational modification of guanosine triphosphate superfamily protein (Rho protein) via prenylation reaction in vivo. These guanosine triphosphate superfamily proteins play important roles in physiological processes such as cell proliferation, cell division, differentiation, etc., such as maintaining the integrity of cell membranes (cholesterol), serving as substrates for protein prenylation reactions (FPP and GGPP), and providing energy sources (coenzyme Q). See Hooff, G.P., et al, A rapid and reactive assay for determining human branched peptides of farnesyl- (FPP) and geranylgeranyl phosphate (GGPP) and transferase activating using UHPLC-MS/MS, analytical and analytical Chemistry,2010.398(4): p.1801-1808.
Various studies show that the content and the effect of geranyl pyrophosphate in a human body have certain correlation with treatment of pulmonary hypertension. In order to detect the content of geranyl ester pyrophosphate, an accurate and reliable detection method is needed, but the detection methods reported at present have technical defects in different aspects, such as poor reproducibility and low accuracy.
Disclosure of Invention
The invention aims to provide a method for detecting the content of geranyl ester pyrophosphate based on prenylation reaction, so as to overcome the defects of poor accuracy and low reproducibility in the detection of geranyl ester pyrophosphate in the prior art.
The technical scheme for realizing the invention is as follows:
the invention provides a method for detecting geranyl ester pyrophosphate content based on prenylation reaction, which comprises the following steps:
(1) firstly, purifying and enriching a sample to be detected by using a liquid-liquid extraction method to obtain a sample solution to be detected containing trace geranyl pyrophosphate;
(2) then, by utilizing the principle of prenylation reaction, two polypeptides connected with dansyl and capable of carrying out prenylation reaction with geranyl pyrophosphate are designed and synthesized: Dansyl-GCVLL and Dansyl-CVIF;
(3) adding polypeptide Dansyl-GCVLL into a sample solution to be detected to perform prenylation reaction with geranyl pyrophosphate to obtain a mixed solution containing a reaction product, wherein the reaction product is GG-Dansyl-GCVLL, GG is a part of geranyl pyrophosphate connected with the polypeptide, the rest part of the geranyl pyrophosphate is a phosphate group, and removing the phosphate group in the reaction, and the reaction product GG-Dansyl-GCVLL in the mixed solution is used as a target compound for indirectly detecting the geranyl pyrophosphate;
(4) adding Dansyl-CVIF into a solution containing a geranyl pyrophosphate pure product with a known concentration to perform a prenylation reaction to obtain a mixed solution containing a reaction product GG-Dansyl-CVIF, wherein the reaction product GG-Dansyl-CVIF in the mixed solution is used as an internal standard substance;
the two polypeptides are:
Dansyl-GCVLL: dansyl (Dansyl) -glycine (Gly/G) -cysteine (Cys/C) -valine (Val/V) -leucine (Leu/L);
Dansyl-CVIF: dansyl (Dansyl) -cysteine (Cys/C) -valine (Val/V) -isoleucine (IIE/I) -phenylalanine (Phe/F);
(5) and (3) sucking the mixed solution containing GG-Dansyl-CVIF obtained in the step (4), adding the mixed solution containing GG-Dansyl-GCVLL obtained in the step (3), wherein the volume ratio of the mixed solution containing GG-Dansyl-CVIF to the mixed solution containing GG-Dansyl-GCVLL is 1:5, mixing, detecting by using a high performance liquid chromatograph and a fluorescence detector to obtain a chromatographic peak diagram of the target compound GG-Dansyl-GCVLL in the step (3) and the internal standard substance GG-Dansyl-CVIF in the step (4), reading peak areas of the two peak diagrams from a machine, and dividing the peak area of the target compound by the peak area of the internal standard substance to obtain a working value.
(6) Detecting by using the solution containing a pure geranyl ester pyrophosphate product with a known concentration by using the method in the steps (3), (4) and (5), establishing a concentration working curve of the sample solution to be detected, and substituting the working value obtained in the step (5) into the concentration working curve to obtain an initial detection value of the sample solution to be detected;
(7) designing a recovery rate experiment to calculate the mass of geranyl pyrophosphate lost in the purification of a sample to be detected, establishing a recovery rate correction curve, correcting the initial detection value in the step (6) by using the curve, and calculating the corrected value through mathematical transformation, namely calculating according to the sampling volume or mass and the sample injection volume to finally obtain the content of geranyl pyrophosphate in the sample to be detected;
the sample to be tested in the step (1) is a sample of plasma or tissue derived from a human being, or a sample of plasma or tissue derived from an animal, or a sample of food, wherein the animal can be a mouse, a rat or a beagle dog; the food may be beef, tomato, rice or soybean.
The specific method for purifying and enriching the sample to be detected by using the liquid-liquid extraction method in the steps comprises the following steps: dissolving a sample to be detected in 1ml of water to be fully dissolved, if the sample is plasma, directly sucking 1ml of plasma without adding water, then adding 7ml of organic solvent, shaking for 5 minutes by using a shaking table, centrifuging for 10 minutes at a rotating speed of 2500 rpm by using a centrifuge, dissolving most of geranyl pyrophosphate in the organic solvent by utilizing the difference of the solubility of the geranyl pyrophosphate in an aqueous solution and the solubility of the geranyl pyrophosphate in the organic solvent, taking an upper layer of organic solution, and carrying out rotary evaporation at 15-20 ℃ until the organic solvent is volatilized, wherein geranyl pyrophosphate remains in a tube wall in the form of crystals, and finally adding water again to dissolve the crystals to achieve the purposes of purification and enrichment, wherein the organic solvent is a mixture of analytically pure hexane and absolute ethanol, and the volume ratio of the analytically pure hexane to the absolute ethanol is 98.5: 1.5.
The recovery rate experiment in the method can adopt a sample standard addition recovery rate experiment, and the specific method can be as follows: preparing a batch of samples with the same source as the sample to be detected, namely samples with the same source but not to be detected, namely a recovery experiment sample group to be detected, dividing the recovery experiment sample group to be detected into two groups, wherein one group is added with a geranyl pyrophosphate pure product with a known concentration to be used as a recovery experiment group, and the other group is added with water with the same volume to be used as a control group. And then carrying out the same purification and enrichment processes on the two groups of samples to be detected in the recovery experiment, and carrying out high performance liquid chromatography detection to obtain a working value. And then subtracting the working value of the control group from the working value of the recovery experimental group to obtain a recovery working value. And then substituting the recovery working value into a concentration working curve to obtain the recovery concentration. The recovery concentration is the concentration of the geranyl ester pyrophosphate pure product added in the recovery experiment group sample, and regression analysis is carried out on the recovery concentration and the actual addition concentration to obtain a recovery rate correction curve. This curve can be corrected for all initial detection values of the present invention.
The invention discloses a detection method of geranyl pyrophosphate content, geranyl pyrophosphate is an isoprenoid derivative synthesized by mevalonic acid in an organism, and the method comprises the following steps: (1) purifying and enriching a plasma or tissue sample to obtain a sample solution containing geranyl pyrophosphate; (2) by utilizing the principle of prenylation reaction, two fluorescent polypeptides Dansyl-GCVLL and Dansyl-CVIF which can carry out prenylation reaction with geranyl pyrophosphate are designed and synthesized; (3) adding Dansyl-GCVLL into a sample solution to be detected to perform prenylation reaction with geranyl pyrophosphate, and taking a reaction product (Dansyl-GG-GCVLL) as a target compound for indirectly detecting geranyl pyrophosphate; adding Dansyl-CVIF into a solution containing a quantitative geranyl pyrophosphate pure product to perform a prenylation reaction, and taking a reaction product (Dansyl-GG-CVIF) as an internal standard substance; (4) detecting a target compound and an internal standard substance by using a high performance liquid chromatograph-fluorescence detector to obtain an initial detection value of geranyl pyrophosphate; (5) and designing a recovery rate experiment to calculate the mass of geranyl pyrophosphate lost during the purification of the plasma or tissue sample, establishing a recovery rate correction curve, and correcting the initial measurement value according to the curve to finally obtain the content of geranyl pyrophosphate. The method provided by the invention has the advantages of stability, reliability, high reproducibility, wide application range, simplicity and convenience in operation and the like, the application range covers the detection of geranyl pyrophosphate in different organisms, and the accuracy and the reproducibility are obviously improved compared with the prior art.
Drawings
FIG. 1 is a high performance liquid chromatogram of a target compound (Dansyl-GG-GCVLL) obtained by binding geranyl pyrophosphate to a polypeptide according to example 2 of the present invention.
FIG. 2 is a high performance liquid chromatogram of an internal standard (Dansyl-GG-CVIF) after geranyl pyrophosphate is bound to a polypeptide, provided in example 2 of the present invention.
FIG. 3 is a high performance liquid chromatogram obtained after mixing a target compound (Dansyl-GG-GCVLL) and an internal standard substance (Dansyl-GG-CVIF) after geranyl pyrophosphate is bound to a polypeptide, provided in example 2 of the present invention.
FIG. 4 is a graph and formula of recovery correction provided in example 3 of the present invention: the curve equation is that y is 1.3058x +4.1179
The Y axis is: real concentration, X-axis: measurement concentration.
FIG. 5 is a target compound as an indirect detection of geranyl pyrophosphate: chemical structure schematic diagram of GG-Dansyl-GCVLL.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention will be described in further detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
Example 1: liquid-liquid extraction method for purifying blood plasma or tissue samples
(1) Taking 1mL of plasma or tissue grinding fluid, and adding the plasma or tissue grinding fluid into a 10mL Ep tube;
(2) to the above solution was added 350. mu.L of Tris-HCl solution, followed by 13.5. mu.L of alkaline phosphatase inhibitor (100X);
(3) placing the mixture in a constant temperature water bath box, and incubating for 40min at 37 ℃;
(4) taking out a sample from the constant-temperature water bath box, adding 7mL of liquid-liquid extracting agent, uniformly mixing, wherein the liquid-liquid extracting agent is a mixture of normal hexane and absolute ethyl alcohol, and uniformly mixing;
(5) placing the Ep tube on a shaking bed for 5min, then placing the Ep tube in a desk type low-speed centrifuge, and centrifuging the Ep tube for 5min at 2000 rpm;
(6) demixing the solution after centrifugation, sucking an organic layer, namely an upper layer solution, into a new 4mL Ep tube, dividing the tube into two tubes if the tube is not filled, and storing the tube at 4 ℃ for subsequent drying;
(7) turning on a rotary evaporator, and setting the temperature of a water bath kettle below the rotary evaporator to be 37 ℃;
(8) connecting a 4mL Ep tube to a small adapter made of glass, connecting the adapter with a rotary evaporator, and adjusting the height to immerse the bottom of the Ep tube in water;
(9) and (3) taking down the Ep tube after the organic solvent in the tube is completely volatilized, wherein geranyl pyrophosphate remains at the bottom of the tube wall and cannot be seen by naked eyes, sucking 48.5 mu L of Tris-HCl to the bottom of the tube, continuously blowing and flushing the bottom of the tube wall, transferring the tube to a 1.5mL Ep tube, and storing at 4 ℃.
Example 2: obtaining target compound and internal standard substance to be detected through prenylation reaction
(1) Putting 48.5 μ L of sample on ice, adding 1 μ L of polypeptide solution (Dansyl-GCVLL), adding 0.5 μ L of GGTase (prenylation reaction specific enzyme), and mixing to obtain reaction solution of target compound;
(2) putting 48.5 mu L of 2000ng/mL pure geranyl ester pyrophosphate on ice, adding 1 mu L of polypeptide solution (Dansyl-CVIF), then adding 0.5 mu L of GGTase (prenylation reaction specific enzyme), and uniformly mixing, wherein the reaction solution is reaction solution of an internal standard substance;
(3) putting the two groups of reaction liquid into a constant-temperature water bath box, reacting for 90min at 37 ℃, taking out, and then placing on ice to terminate the reaction;
(4) and (3) sucking 10 mu L of internal standard substance reaction liquid, adding the internal standard substance reaction liquid into the reaction liquid of the target compound, mixing, and keeping on-machine detection.
Example 3: measurement of initial detection value of geranyl pyrophosphate
High performance liquid chromatography detection
(1) Opening the workstation, the fluorescence detector, the pump, the column incubator, the sample injection system and the sample injector until the fluorescence detector is adjusted;
(2) connecting the C18 chromatographic column to the inside of the sample injection system, paying attention to the outflow sequence of the chromatographic column, and closing the column temperature box;
(3) respectively pouring the pre-filtered aqueous phase mobile phase (ammonium acetate solution, attention is paid to the existing filtration) and the organic phase mobile phase (chromatographic grade acetonitrile) into a large-caliber solution bottle, and winding a sealing film at the bottle mouth to prevent the mobile phase from volatilizing;
(4) detection conditions are as follows:
(5) after the detection conditions are set on the workstation, washing the chromatographic column for 30-60 min by using a mobile phase to balance the chromatographic column and stabilize the state of the fluorescence detector;
(6) the workstation can record the change of fluorescence intensity all the time and draw a curve, after the operation is finished, the workstation can automatically integrate the peak area, and record the retention time and the peak area to finish the whole detection process.
Establishment of (II) content working curve
(1) Taking out a GGPP pure product from an ultra-clean workbench, carefully unscrewing a cover, diluting the pure product with 1mg/mL of concentration by using prepared Tris-HCl to obtain 1ng/mL of mother solution;
(2) sucking the mother liquor, diluting the mother liquor into 10ng/mL, 30ng/mL, 50ng/mL, 100ng/mL, 300ng/mL and 500ng/mL respectively by using Tris-HCl, subpackaging the gradient standard substance in a volume of 48.5 mu L, and storing the gradient standard substance at the temperature of minus 20 ℃ for a subsequent experiment;
(3) a reaction solution was obtained in the same manner as in example 2. And (5) detecting by using a computer, recording numerical values and drawing a curve.
(III) detection of sample content
(1) Reacting the sample obtained by purification and enrichment in the example 1 according to the method in the example 2 to obtain a reaction solution;
(2) and (5) performing detection on the machine, recording the numerical value and bringing the numerical value into a content working curve to obtain an initial detection value.
Example 4 establishment of recovery Curve
(1) Enough plasma was prepared and mixed together to maintain homogeneity, and 42mL was aspirated and divided into 3 groups, numbered a, B, C. Each group was 14mL, 3 groups of plasma were purified and tested separately on different 3 days as independent replicates;
(2) dividing 14mL of group A into 14 parts of 1mL, and respectively transferring the parts into ep tubes with numbers of A1-A14;
(3)14 portions of plasma were added with different amounts of pure GGPP (when the amount is 0, Tris-HCl solution is added instead), and the amount of GGPP added was as follows:
(4) after the addition is finished, 14 parts of blood plasma are purified and enriched together;
(5) carrying out prenylation reaction on the samples, and adding 10 mu L of internal standard substance reaction solution into each sample after the reaction;
(6) opening a high performance liquid chromatography instrument, and performing on-machine detection;
(7) subtracting the detection result of A1-A12 from the control group (average value of A13 and A14) to obtain the measurement value of the addition amount of GGPP, and performing regression analysis on the measurement value and the actual value;
(8) b, C the experimental procedure described above was repeated.
It will be understood by those skilled in the art that the foregoing is only a preferred embodiment of the present invention, and is not intended to limit the invention, and that any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the scope of the present invention.
Claims (6)
1. A method for detecting geranyl ester pyrophosphate content based on prenylation reaction comprises the following steps:
(1) purifying and enriching a sample to be detected by using a liquid-liquid extraction method to obtain a sample solution to be detected, which contains geranyl pyrophosphate;
(2) two polypeptides which are connected with dansyl and can carry out the prenylation reaction with geranyl pyrophosphate are designed and synthesized by utilizing the principle of the prenylation reaction: Dansyl-GCVLL and Dansyl-CVIF;
(3) adding polypeptide Dansyl-GCVLL into a sample solution to be detected to perform prenylation reaction with geranyl pyrophosphate to obtain a mixed solution containing a reaction product, wherein the reaction product is GG-Dansyl-GCVLL, GG is a part of geranyl pyrophosphate connected with the polypeptide, the rest part of the geranyl pyrophosphate is a phosphate group, and removing the phosphate group in the reaction, and the reaction product GG-Dansyl-GCVLL in the mixed solution is used as a target compound for indirectly detecting the geranyl pyrophosphate;
(4) adding Dansyl-CVIF into a solution containing a geranyl pyrophosphate pure product with a known concentration to perform a prenylation reaction to obtain a mixed solution containing a reaction product GG-Dansyl-CVIF, wherein the reaction product GG-Dansyl-CVIF in the mixed solution is used as an internal standard substance;
(5) sucking the mixed solution containing GG-Dansyl-CVIF obtained in the step (4), adding the mixed solution containing GG-Dansyl-CVIF and the mixed solution containing GG-Dansyl-GCVLL obtained in the step (3) in a volume ratio of 1:5, mixing, and detecting by using a high performance liquid chromatograph and a fluorescence detector, wherein the high performance liquid chromatograph adopts a C18 chromatographic column, the phase A of the high performance liquid chromatograph is a water phase, the phase B of the high performance liquid chromatograph is an organic phase, the flow rate is 1mL/min, the sample injection volume is 10 mu L, the operation time of each needle is 20min, the column temperature is 25 ℃, the column pressure is 840psi, the gain of the fluorescence detector is 10, the excitation wavelength is 340nm, and the emission wavelength is 525 nm; obtaining chromatographic peak images of the target compound GG-Dansyl-GCVLL in the step (3) and the internal standard substance GG-Dansyl-CVIF in the step (4), reading peak areas of the two peak images from a machine, and dividing the peak area of the target compound by the peak area of the internal standard substance to obtain a working value;
(6) detecting by using the solution containing a pure geranyl ester pyrophosphate product with a known concentration by using the method in the steps (3), (4) and (5), establishing a concentration working curve of the sample solution to be detected, and substituting the working value obtained in the step (5) into the concentration working curve to obtain an initial detection value of the sample solution to be detected;
(7) designing a recovery rate experiment, calculating the mass of geranyl pyrophosphate lost during the purification of a sample to be detected, establishing a recovery rate correction curve, correcting the initial detection value in the step (6) by using the curve, and calculating the corrected value through mathematical transformation, namely calculating according to the sampling volume or mass and the sample injection volume to finally obtain the content of geranyl pyrophosphate in the sample to be detected;
the specific method of the recovery rate experiment is as follows: preparing a batch of samples with the same source as a sample to be detected, dividing the sample group to be detected of a recovery experiment into two groups, adding a geranyl pyrophosphate pure product with a known concentration into one group of samples to be detected as a recovery experiment group, adding water with the same volume into the other group of samples to be detected as a control group, carrying out the same purification and enrichment processes on the samples to be detected of the two groups of recovery experiments, carrying out high performance liquid chromatography detection to obtain a working value, subtracting the working value of the control group from the working value of the recovery experiment group to obtain a recovery working value, substituting the recovery working value into a concentration working curve to obtain a recovery concentration, wherein the recovery concentration is the concentration of the geranyl pyrophosphate pure product added into the samples of the recovery experiment group, carrying out regression analysis on the recovery concentration and the actual added concentration to obtain a recovery correction curve, and correcting the initial detection value by using the curve.
2. The method according to claim 1, wherein the sample to be tested in step (1) is a sample of human-derived plasma or tissue, or a sample of animal-derived plasma or tissue, or a sample of food-derived sample.
3. The method of claim 2 wherein the animal is a mouse, rat, or beagle dog.
4. The method of claim 2 wherein the food is beef, tomato, rice or soy.
5. The method according to claim 1, wherein the specific method for purifying and enriching the sample to be tested by using the liquid-liquid extraction method comprises: dissolving a sample to be detected in 1ml of water to fully dissolve the sample, if the sample is plasma, directly sucking 1ml of plasma without adding water, then adding 7ml of organic solvent, shaking for 5 minutes by using a shaking table, then centrifuging for 10 minutes at a rotating speed of 2500 rpm by using a centrifuge, taking an upper layer of organic solution, carrying out rotary evaporation at 15-20 ℃ until the organic solvent is volatilized, at the moment, geranyl pyrophosphate remains in a tube wall in a crystal form, and finally adding water again to dissolve the crystal to finish purification and enrichment.
6. The method of claim 5, wherein the organic solvent is a mixture of analytically pure hexane and absolute ethanol at a volume ratio of 98.5: 1.5.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810702345.XA CN110658292B (en) | 2018-06-29 | 2018-06-29 | Method for detecting geranyl ester pyrophosphate content based on prenylation reaction |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810702345.XA CN110658292B (en) | 2018-06-29 | 2018-06-29 | Method for detecting geranyl ester pyrophosphate content based on prenylation reaction |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN110658292A CN110658292A (en) | 2020-01-07 |
| CN110658292B true CN110658292B (en) | 2021-03-26 |
Family
ID=69027672
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810702345.XA Active CN110658292B (en) | 2018-06-29 | 2018-06-29 | Method for detecting geranyl ester pyrophosphate content based on prenylation reaction |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN110658292B (en) |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1327686A4 (en) * | 2000-10-06 | 2005-01-19 | Sapporo Breweries | Farnesyl pyrophosphate synthase protein, nucleic acid and promoter region thereof |
| CN103898037B (en) * | 2014-03-11 | 2017-11-17 | 中国科学院青岛生物能源与过程研究所 | A kind of coproduction geraniol and the genetic engineering bacterium and its construction method of nerol and application |
-
2018
- 2018-06-29 CN CN201810702345.XA patent/CN110658292B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN110658292A (en) | 2020-01-07 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Moore et al. | [117] Chromatographic determination of amino acids by the use of automatic recording equipment | |
| CN108663462B (en) | Method for measuring vitamin A, D and E in milk powder | |
| CN101273267A (en) | Method for analysis of albumin in sample solution | |
| CN112526051B (en) | Fmoc-lysine high performance liquid chromatography determination method | |
| CN111175405A (en) | Method for simultaneously detecting multiple fat-soluble vitamins in blood sample and application thereof | |
| CN111208220A (en) | Preparation and application of quality control substance of catecholamine substances and metabolites of plasma type freeze-dried powder | |
| CN107973787A (en) | A kind of coumarin derivative DMAC and its preparation method and application | |
| CN113533565A (en) | Method for detecting concentrations of 8 flavonoid compounds in human urine by UPLC-MS/MS method | |
| CN110658292B (en) | Method for detecting geranyl ester pyrophosphate content based on prenylation reaction | |
| CN111505184A (en) | Method for determining components of freeze-dried powder injection containing multiple vitamins | |
| CN102081078A (en) | Method for measuring residual quantities of four fluoroquinolone medicaments in animal food | |
| CN107976499A (en) | A kind of method of alpha-aminobutyric acid content in efficiently binary liquid chromatography detection livestock and poultry organization | |
| CN108872454A (en) | A kind of method of VFA in rapid and accurate determination anaerobic digestion solution | |
| CN108709942B (en) | Method for determining vitamin A and vitamin E in milk powder | |
| CN113759039A (en) | Synchronous detection method and device for beta-carotene and vitamin A | |
| CN110045053A (en) | A kind of QuEChERS pre-treating method of the analysis of the amphetamines suitable for blood | |
| CN114324647B (en) | Simultaneous determination of vitamin K in milk powder 1 And K 2 Methods and uses of (2) | |
| Van den Berg et al. | A rapid and sensitive method for the determination of short chain fatty acids in serum | |
| CN108535400B (en) | Thin-layer chromatography analysis and detection method for impurity 5-methoxytryptamine in melatonin bulk drug | |
| CN103926368A (en) | Method for extracting biotin from corn steep liquor and thin layer chromatography (TLC) scanning detection method of biotin | |
| CN106443028B (en) | Method that is a kind of while measuring shellfish GSH-PX activity and free amino acid | |
| JPH01229972A (en) | Determining method of low carboxylic acid in body fluid material | |
| CN114563504B (en) | Method and kit for determining content of free aldosterone in blood plasma | |
| CN110361525B (en) | Method for measuring content of phosphatidylserine in blood | |
| CN111323497B (en) | Optical purity analysis method of pasireotide starting material |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |