Method for measuring contents of tyrosine, tryptophan and 5-hydroxytryptamine in blood serum by high-efficiency liquid phase chromatography-fluorescence method

A technology of high-performance liquid chromatography and fluorescence method, which is applied in the direction of measuring devices, fluorescence/phosphorescence, instruments, etc., can solve problems such as unseen and achieve high sensitivity

Inactive Publication Date: 2011-10-12
CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there are many methods for the determination of Tyr, Trp and 5-HT reported at home and abroad. Among them, the most reported method is to use HPLC to measure one or two of them, but there is no simultaneous determination of Tyr in serum by HPLC-FLD. , Trp and 5-HT content reports, the present invention utilizes Tyr, Trp and 5-HT to have the characteristic of natural fluorescence, discloses a kind of HPLC-FLD that simultaneously measures serum Tyr, Trp and 5-HT content at home and abroad for the first time, serum After deproteinizing the sample, take the supernatant and directly inject it into the sample. Tyr, Trp and 5-HT are separated by the chromatographic column, and then detected by wavelength conversion at a specific time period with their respective optimal fluorescence excitation and emission wavelengths.

Method used

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  • Method for measuring contents of tyrosine, tryptophan and 5-hydroxytryptamine in blood serum by high-efficiency liquid phase chromatography-fluorescence method
  • Method for measuring contents of tyrosine, tryptophan and 5-hydroxytryptamine in blood serum by high-efficiency liquid phase chromatography-fluorescence method
  • Method for measuring contents of tyrosine, tryptophan and 5-hydroxytryptamine in blood serum by high-efficiency liquid phase chromatography-fluorescence method

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Experimental program
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Effect test

Embodiment 1

[0051] Accurately weigh Tyr10.0mg, Trp10.0mg, 5-HT10.0mg, dissolve and dilute to 10ml with 2.5% (v / v) perchloric acid solution, and prepare 5500μmol / L, 4900μmol / L and 117.6μmol / L respectively L of standard stock solution, aliquoted and stored in a -20°C refrigerator for later use. Dilute it with 2.5% perchloric acid to form a mixed standard working solution containing Tyr (55 μmol / L), Trp (24.5 μmol / L) and 5-HT (1.176 μmol / L) before use.

Embodiment 2

[0053] Take 2ml of fasting venous blood (8-9 hours in the morning) from the research subjects in a coagulation-promoting tube, centrifuge at normal temperature and speed to separate the serum, take an appropriate amount of serum in an eppendorf tube, add an equal amount of 5% perchloric acid solution, mix well, and place it at room temperature for 10 minutes. Centrifuge at 10000r / min for 10min, and take 20μL of the supernatant for analysis.

[0054] Working principle and working process

[0055] Use a manual injector to draw 20 μL of the substance to be tested into the mobile phase, and the substance to be tested is mixed with the mobile phase, 0.1mol / LKH 2 PO 4 It flows into the chromatographic column together with methanol, and the Atlantis C18 chromatographic column separates Tyr, Trp, and 5-HT, and they arrive at the fluorescence detector in turn for identification and detection. The sample processing of this method is simple, and the detection can be completed within 10...

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Abstract

The invention provides a method for measuring the contents of tyrosine, tryptophan and 5-hydroxytryptamine in blood serum by adopting a high-efficiency liquid phase chromatography-fluorescence method. The method is accurate and rapid, and comprises the following steps of: removing proteins from blood serum with perchloric acid with the concentration of 5 percent and detecting by adopting a high-efficiency liquid phase chromatography technology, wherein a chromatography condition is that: a chromatography column is an Atlantis C18 column of the specification of 4.6 millimeters*150 millimeters ID; mixing 0.1 mol/L of KH2PO4 with methanol in the volume ratio 85:15 at the flow rate 1.0 ml/min to obtain a mixed solution serving as a moving phase, wherein the excitation wavelength and the emitting wavelength of a fluorescence detector are 228 nanometers and 306 nanometers respectively within 0-4minutes, 278 nanometers and 338 nanometers within 4-7.5 minutes and 285 nanometers and 353 nanometers after 7.5 minutes; and adjusting the wavelengths within a specific period of time to perform separate detection on the tyrosine, the 5-hydroxytryptamine and the tryptophan.

Description

Technical field: [0001] The invention belongs to the field of analytical chemistry and relates to a research method for quantitatively measuring the contents of tyrosine, tryptophan and serotonin in serum, in particular to a method for simultaneously measuring tyrosine, tryptophan and serotonin in serum by using high performance liquid chromatography-fluorescence method. Related technology of tryptophan and serotonin content. Background technique: [0002] Tyrosine (Tyr) is a non-essential amino acid for the human body. It is converted from phenylalanine (Phe) by dehydroxylation of phenylalanine hydroxylase (PAH). It is a synthetic catecholamine Neurotransmitters—precursors of dopamine, epinephrine, and norepinephrine—are important in neuronal transmission. L-tryptophan (L-tryptophan, L-Trp) is one of the essential amino acids in the human body, participates in human protein synthesis and metabolic regulation, and is the precursor of 5-hydroxytryptamine (5-hydroxytryptamine...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/88G01N21/64
Inventor 唐爱国李影穆萨
Owner CENT SOUTH UNIV
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