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Chromatographic detection method for simultaneously analyzing ammonium ions, amino acids and biogenic amine in tobacco leaves

A chromatographic detection and ammonium ion technology, applied in the field of determination of tobacco chemical components, can solve the problems of slow derivatization reaction, poor stability of derivative products, high price, etc., achieve less by-products, improve sample detection throughput and data accuracy, cheap effect

Active Publication Date: 2016-10-26
GUIZHOU TOBACCO SCI RES INST
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The main liquid-phase derivatization reagents are o-phthalaldehyde, ninhydrin, 9-fluorenylmethyl chloroformate, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate and benzene isothiocyanate. Esters, etc., but these derivatization reagents all have defects such as slow derivatization reaction, poor stability of derivatized products, and relatively expensive prices.
[0007] In summary, in the technical field of determination of tobacco chemical components, there is currently no simple, rapid, high-sensitivity and stable method for the simultaneous detection of ammonium ions, amino acids and polyamines in tobacco leaves.

Method used

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  • Chromatographic detection method for simultaneously analyzing ammonium ions, amino acids and biogenic amine in tobacco leaves
  • Chromatographic detection method for simultaneously analyzing ammonium ions, amino acids and biogenic amine in tobacco leaves
  • Chromatographic detection method for simultaneously analyzing ammonium ions, amino acids and biogenic amine in tobacco leaves

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Embodiment 1

[0028] refer to figure 1 , the detection method of the present invention comprises the following steps:

[0029] (1) Weigh 100 mg of flue-cured tobacco powder or 50 mg of fresh tobacco leaves (freeze-dried sample) into a centrifuge tube, then add 40 μL of internal standard (aminoadipic acid-1 mg / mL; norvaline-1 mg / mL ), then add 1.8 mL of 0.1% formic acid aqueous solution for microwave (800 W) assisted extraction for 40 s, centrifuge at 3000 rpm for 5 min, and pass the supernatant through a 0.45 μm water filter membrane;

[0030] (2) Take 1 mL of the upper layer filtrate in a centrifuge tube, add 8.5 μL of 3 M sodium hydroxide solution, vortex, add 1 mL of 0.8M boric acid buffer (pH=9.0-9.2), and then add 0.3 mL of methanol or hexanonitrile and 30 uL of diethyl ethoxymethylene malonate, vortexed for 1 min, and then sonicated at room temperature for 35 min to react. After the reaction was completed, 60 μL of 3 M sodium hydroxide solution was added, vortexed and placed in a w...

Embodiment 2

[0045] In order to compare the changes of ammonium ions, amino acids and polyamines during the curing process of tobacco leaves, the method as in Example 1 was used to analyze the fresh tobacco leaves before curing and the tobacco leaves after curing respectively. The sample variety is K326. The sample was taken before the 12th leaf position (counting from bottom to top) of the tobacco plant was ripe and roasted. The leaf was separated from the main vein by the half-leaf method, and half of the leaf was immediately put into liquid nitrogen and brought back to the laboratory for vacuum freezing. Dry; the other half is baked normally with the main veins. Take 8 slices for each repetition and mix, 5 repetitions in total. In order to avoid the difference in moisture content of tobacco leaf samples before and after curing, the main veins were removed from the tobacco leaf samples after curing, and they were dried in a constant temperature oven at 40°C until they reached a constant ...

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Abstract

The invention discloses a chromatographic detection method for simultaneously analyzing ammonium ions, amino acids and biogenic amine in tobacco leaves. Thechromatographic detection method comprises the following steps of A, extracting the ammonium ions, amino acids and biogenic amines in the tobacco leaves; B, deriving the ammonium ions, amino acids and biogenic amines in the tobacco leaves; C, performing the chromatographic analysis on the derived products of ammonium ions, amino acids and biogenic amines in the tobacco leaves; D, quantitatively analyzing the ammonium ions, amino acids and biogenic amines in the tobacco leaves. The chromatographic detection method has the advantagesthattheammonium ions, 25 types of amino acids and 8 types of biogenic amines in tobacco leaves can be simultaneously measured by microwave-assisted extraction, ethoxy methylene malonic diethyl esterderiving, ultra-high performance liquid chromatography and ultraviolet detectors; the simplicity and rapidityare realized, the sensitivity is high, thestability is good, and the like; the detection flux and data accuracy of the sample can be improved.

Description

technical field [0001] The invention relates to a simple, stable and accurate chromatographic detection method for simultaneously analyzing ammonium ions, amino acids and biogenic amines in tobacco, and belongs to the technical field of determination methods for chemical components of tobacco. Background technique [0002] Ammonium ion is a nitrogen source that plants can directly use, and is one of the important raw materials for the synthesis of important nitrogen-containing compounds in tobacco - proteins, amino acids, alkaloids, nucleotides, etc. Although nitrate nitrogen can also be directly absorbed by plants, it cannot direct use. After nitrate nitrogen enters the plant, it must be converted into ammonium ions by nitrate reductase and nitrite reductase before it can enter the metabolic synthesis pathway, so that it can be used by plants and truly play the role of nutrients. [0003] Amino acids are a class of amino acids (-NH 2 ) or imino (-NH) organic acids, which ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/027G01N2030/067
Inventor 蔡凯赵会纳向章敏蔡斌雷波潘文杰高维常
Owner GUIZHOU TOBACCO SCI RES INST
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