Test items

DB13/T 1384.2-2011 Determination of oxytetracycline, tetracycline and chlortetracycline in feed

Sample and mailing requirements

Samples can be picked up by express delivery/self-delivery/door-to-door sampling, and the quantity and specifications depend on the test items.

What is the content of the determination of oxytetracycline, tetracycline, and chlortetracycline in feed in DB13/T1384.2-2011? Baijian.com has a wide range of tests and can select appropriate test standards and projects according to customer needs. There are nearly a thousand cooperative laboratories across the country with complete CMA/CNAS qualifications, and samples can be sent nearby for testing. Today, Baijian.com will give you a brief introduction to the content of the determination standard of oxytetracycline, tetracycline, and chlortetracycline in feed in DB13/T1384.2-2011.

1. Scope

This standard specifies the liquid chromatography-tandem mass spectrometry method for the determination of oxytetracycline, tetracycline, and chlortetracycline in feed.

This standard applies to the determination of oxytetracycline, tetracycline, and chlortetracycline in feed.

The method detection limit of this standard is 50μg/kg.

2. Normative references

The following documents are essential for the application of this document. For dated references, only the version with the date is applicable to this document. For undated references, the new version (including all amendments) is applicable to this document.

GB/T 6682 Specifications and test methods for water for analytical laboratories (GB/T 6682-1992, new ISO 3696:1987)

3. Principle

After centrifugation, the sample extract is purified by Oasis HLB solid phase extraction column and anion exchange column, and then measured by liquid chromatography-tandem mass spectrometry and quantified by external standard method.

4. Reagents and materials

Unless otherwise specified, all reagents used are analytical grade, and water is first-grade water specified in GB/T 6682.

Methanol: chromatographic grade

Acetonitrile: chromatographic grade.

Hydrochloric acid.

0.4molL hydrochloric acid solution: accurately measure 32.8mL hydrochloric acid in a 1000mL volumetric flask, add water to the mark, and shake well.

Oxalic acid.

0.01Lmol/L oxalic acid methanol solution: accurately weighs 1.26g oxalic acid, dissolves it in methanol, and makes up to volume in a 1000mn volumetric flask.

Formic acid

Formic acid solution: accurately measure 4mL formic acid in a 1000mL volumetric flask, add water to the mark, and shake well.

Oxytetracycline, tetracycline, chlortetracycline standard substances: purity ≥95%.

Oxytetracycline, tetracycline, chlortetracycline standard stock solution: 0.1mgmL. Accurately weigh appropriate amounts of oxytetracycline, tetracycline, chlortetracycline, and standard substances, and use methanol to prepare 0.1mgmL standard stock solutions. The stock solutions are stored in a -18℃ freezer.

Oxytetracycline, tetracycline, chlortetracycline matrix mixed standard working solution: Take appropriate amount of oxytetracycline, tetracycline, chlortetracycline standard stock solution as needed, dilute with blank sample extract to matrix mixed standard working solution of appropriate concentration, and store at 4℃ for three days.

Oasis HLB solid phase extraction column or equivalent: 500mg, 6mL. Pre-treat with 5mL methanol and 10mL water before use to keep the column moist.

5. Instruments

Liquid chromatography-tandem quadrupole mass spectrometer, equipped with an electrospray ion source.

Analytical balance: one with a sensitivity of 0.1mg and one with a sensitivity of 1mg

Vortex mixer.

Solid phase extractor.

High-speed centrifuge

Conical flask with stopper: 50mL.

Centrifuge tube: 50mL, 5mL.

Nitrogen blower

Rotary evaporator.

6. Sample preparation

Extraction:

Weigh 3.0g sample accurately (accurate to 0.01g), place in a 50mL stoppered conical flask, add 10mL methanol, vortex mix for 1min, add 30mL hydrochloric acid solution (4.4), vortex mix for 1min, ultrasonicate for 20min, transfer to a 50mL centrifuge tube, centrifuge at 5000r/min for 10min, take the supernatant, and wait for purification.

Purification:

Take 10mL supernatant and dilute and mix with 40mL water, then transfer to the reservoir connected to the Oasis HLB column. The solution passes through the column at a flow rate of less than 1.0mL/min, then rinse with methanol-water solution (4.9) in turn, and drain under reduced pressure for 20min. Elute with 5 mL of oxalic acid methanol solution (4.6), collect the eluate and evaporate to dryness under reduced pressure at 40°C, dissolve the residue with 1.0 mL of mobile phase, centrifuge at 12000 r/min for 10 min, filter through 0.45 μm organic filter membrane, and then test.

7. Determination steps

Chromatographic column: Inertsil C8, 3.5 μm, 150 mm × 2.1 mm (id) or equivalent;

Mobile phase: acetonitrile + methanol + 0.4% formic acid solution (18 + 4 + 78);

Flow rate: 0.2 mL/min;

Column temperature: 25°C;

Injection volume: 20 μL.

Ion source: electrospray ion source;

Scanning mode: positive ion scanning;

Detection mode: multiple reaction monitoring;

Optimize the electrospray voltage, collision voltage, and other conditions to the optimal value;

Qualitative ion pair.