A chromatographic detection method for simultaneous analysis of ammonium ions, amino acids and biogenic amines 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 poor stability of derivative products, slow derivative reaction, high price, etc., to improve sample detection throughput and data accuracy, less by-products, cheap effect
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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), and then add 1.8 Microwave (800W) assisted extraction of mL 0.1% formic acid solution for 40s, centrifuged at 3000rpm for 5min, and the supernatant was passed through a 0.45μm aqueous filter membrane;
[0030] (2) Take 1mL of the upper layer filtrate in a centrifuge tube, add 8.5μL of 3M sodium hydroxide solution, vortex, add 1mL of 0.8M boric acid buffer solution (pH=9.0-9.2), then add 0.3mL of methanol or acetonitrile and 30uL of acetonitrile Diethyl oxymethylene malonate, vortex for 1 min, and then sonicate for 35 min at room temperature to react. After the reaction is completed, add 60 μL of 3M sodium hydroxide solution, vortex, add to a water bath at 80°C...
Embodiment 2
[0043] 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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