Method for detecting sphingolipid in zebra fish brain tissue based on UHPLC-QTrap MS

[0040] In some embodiments, the preparation method of the zebrafish brain tissue protein comprises:

[0041] 1) lyse brain tissue and precipitate protein by protein precipitant;

[0042] 2) adding chloroform to vibrate and stratify, and take the chloroform layer;

[0043] Optional, repeat step 2);

[0044] 3) Blow dry the chloroform layer with nitrogen.

[0045] In some embodiments, the protein precipitation agent is selected from methanol and / or acetonitrile.

[0046] In some embodiments, the preparation method of the zebrafish brain tissue protein also includes:

[0047] Reconstitute with methanol, and pass through a 0.20-0.24 μm microporous membrane, or 0.22 μm.

[0048] Since there are many impurities in animal tissues and the physical and chemical properties of organic solvents are quite different, sample pretreatment is a very critical part. The workload and error sources of pre-processing accounted for more than 60% of the entire detection process. The focus is on t

[0058] Detection of sphingolipids in zebrafish brain tissue

[0059] 1. Preparation of physical samples: Take 5 fish brains (about 20 mg) and perform a blank subtraction and addition recovery test. At the same time, set a blank control and add samples, and add concentrations of 5 μg / kg, 10 μg / kg and 50 μg / kg. Evenly, fully stand. Add 180 μL of methanol to the 2 mL centrifuge tube containing the zebrafish brain tissue sample, and then grind it thoroughly on ice. Transfer the uniformly ground brain tissue to a 5mL centrifuge tube, then add 180 μL of methanol to wash the tube wall of the 2mL centrifuge tube, and transfer it to a 5mL centrifuge tube as well. Add 1.2 mL of chloroform, vortex for 1 min, and ultrasonically extract for 3 min. After the sonication, 240 μL of water was added and shaken to make it appear stratified. The chloroform layer was taken and extracted after repeated addition of chloroform. Then the chloroform layers obtained twice were combined, and blown