1. Scope This standard specifies the determination method of furazolidone residues in aquatic products-high performance liquid chromatography.
This standard applies to the determination of furazolidone residues in the edible parts of aquatic products.
2. Normative cited documents The clauses in the following documents become the clauses of this standard through the quotation of this standard. For dated reference documents, all subsequent amendments (not including errata content) or revisions are not applicable to this standard. However, parties that have reached an agreement based on this standard are encouraged to study whether the latest versions of these documents . For the cited documents without date, the latest version is applicable to this standard.
GB / T 6682-1992 Specification and test method of water for analysis laboratory
3. The furazolidone in the sample is extracted with dichloromethane, the extract is concentrated by evaporation, the anhydrous sodium sulfate is used to remove water, the n-hexane liquid-liquid extraction is used to remove fat and other impurities, centrifugation, and the supernatant is removed and passed through a membrane filter for high efficiency Separated by liquid chromatograph, detected by ultraviolet detector, and quantified by external standard method.
4. Reagents and materials
4.1 Acetonitrile: chromatographically pure.
4.2 Dichloromethane: chromatographically pure.
4.3 n-hexane: analytically pure, saturated with acetonitrile.
4.4 85% phosphoric acid: analytically pure.
4.5 Acetonitrile aqueous solution: acetonitrile + water (volume fraction is 80 + 20).
4.6 Anhydrous sodium sulfate: analytically pure; after burning at 640 ° C for 4.0h, store in a closed container for later use
4.7 Anhydrous sodium sulfate column: 200mm × 24mm (id), containing anhydrous sodium sulfate 50mm ~ 100mm high.
4.8 Furazolidone standard product: purity ≥99.5%.
4.9 Furazolidone standard stock solution: Weigh 10.0mg of furazolidone, dissolve with acetonitrile aqueous solution and dilute to a fixed volume to a 50mL brown volumetric flask, and store it in the refrigerator for one month. 1 mL of this solution is equivalent to 200 μg furazolidone.
4.10 Test water: comply with GB / T 6682 first-class water standard.
5. Instruments and equipment
5.1 High-performance liquid chromatograph with UV detector.
5.2 High-speed organization masher.
5.3 Rotary evaporator.
5.3 Centrifuge.
5.4 Oscillator.
5.5 Evaporation flask with stopper.
5.6 Scale centrifuge tube.
5.7 Membrane pore filter: 0.45μm.
6. Chromatographic conditions
6.1 Chromatographic column: C18 column, 250mm × 4.6mm (id), particle size 5μm.
6.2 Mobile phase: acetonitrile + water + phosphoric acid (volume fraction 40 + 60 + 0.1).
6.3 Flow rate: 0.8mL / min.
6.4 Detection wavelength: 365nm.
6.5 Column temperature: room temperature.
6.6 Injection volume: 20 μL.
7. Sample determination
7.1 Sample preparation Take the edible parts of aquatic products such as fish, shrimp, crab, turtle, etc., cut them into small pieces no larger than 5mm × 5mm × 5mm, mix well, and crush them by high-speed tissue masher.
7.2 Extraction Weigh about 10g of crushed sample (accurate to 0.01g), add 25mL of dichloromethane, stir and disperse, soak for 15min, shake on a shaker for 5min, extract the solution through an anhydrous sodium sulfate column and filter into an evaporation flask. Repeat the extraction with 25mL and 15mL dichloromethane as described above, then rinse the anhydrous sodium sulfate column with 15mL dichloromethane, and blow out the liquid in the column with an ear wash ball. The filtrate is filtered into the same evaporation flask, and the filtrate is rotated On the evaporator, the solvent was evaporated under reduced pressure in a water bath at about 45 ° C, and the evaporation rate was controlled at one drop per second.
7.3 Purify with 1.0mL mobile phase and 1.0mL n-hexane to dissolve the residue and fully wash the evaporation flask, move the liquid into a centrifuge tube, centrifuge at 4000r / min for 5min, remove the upper n-hexane layer with a pointed pipette, and then into the centrifuge tube Add 1.0 mL of n-hexane, mix for 2 min, centrifuge, and remove the upper n-hexane layer with a pointed pipette. The lower supernatant is filtered through a 0.45 μm membrane pore filter and then analyzed by HPLC.
7.4 Determination
7.4.1 Before preparing the standard working fluid of furazolidone, take the standard stock solution of furazolidone and dilute with mobile phase to a standard working fluid with a concentration of 0.01μg / mL ~ 1.00μg / mL.
7.4.2 Liquid chromatographic determination According to the residual amount of furazolidone in the sample solution, select a standard working solution with a similar peak height. The response values ​​of furazolidone in standard working solution and sample solution should be within the linear range of instrument detection. The same volume of standard working solution and sample solution is inserted into the sample for measurement. Under the above chromatographic conditions, the retention time of furazolidone is about 5.8 min.
7.5 The blank test shall be carried out according to steps 7.2 to 7.4 except that no sample is added.
8. Calculation and expression of results According to the peak height of the standard working solution and the sample solution, calculate the residual amount of furazolidone in the sample according to formula (1): (h-h0) × V × 1000
C = Cs × ································ (1)
hs × m
In the formula:
C── The residual amount of furazolidone in the sample, the unit is microgram per kilogram (μg / kg);
h──The peak height of furazolidone in the sample solution;
hs-peak height of furazolidone in standard working solution;
h0-peak height of blank test;
Cs-the concentration of furazolidone in the standard working solution in micrograms per milliliter (μg / mL);
V──The final volume of the sample solution, the unit is milliliter (mL);
m──Weighing the sample, the unit is gram (g).
9. Linear range, detection limit, recovery rate
9.1 Linear range The linear range of the furazolidone standard working solution in this method is 0.01μg / mL ~ 1.00μg / mL.
9.2 Detection limit The detection limit of this method is 1 μg / kg.
9.3 Recovery rate The recovery rate of this method is 72.8% ~ 96.2%.
Wine Display Rack,Wine Rack,Wall Wine Rack,Acrylic Display Rack
Apache Industry Ltd. , http://www.apachedisplay.com