CN105758959A - Method for detecting salinomycin and monensin in animal product - Google Patents

Abstract

The invention discloses a method for detecting salinomycin and monensin in an animal product. Salinomycin and monensin in the animal product are synchronously detected once by a high performance liquid chromatograph-tandem triple quadrupole mass spectrometer, and the method comprises the detection steps of extraction, purification and detection. Compared with a traditional or national standard method with liquid chromatograph as a detection instrument, the method adopting the liquid chromatograph-tandem triple quadrupole mass spectrometer has the advantages that the detection limit is lower; the detection accuracy is higher; and the related detection limit reaches 2 microgram/kg of salinomycin and 10 microgram/kg of monensin. Meanwhile, salinomycin and monensin are merged for detection, so that pretreatment and instrument analysis are performed simultaneously, and the efficiency is higher.

Description

A kind of detect the method for Salinomycin and monensin in livestock products

Technical field

The invention belongs to detection of veterinary drugs in food technical field, be specifically related to one and utilize the high performance liquid chromatography triple level Four bar GC-MS of series connection, the method for Salinomycin and monensin in detection livestock products.

Background technology

Along with living standards of the people improve constantly, the demand of domestic animal product is increased by people day by day, residue of veterinary drug is as the key factor affecting animal product edible safety, having become a social hotspots topic of people's common concern, wild animal resources not only needs fine micromanipulation means but also need sensitive trace detection technology.

Salinomycin and monensin are all polyether antibiotics, have antibacterial and anticoccidial effect, also can promote animal growth simultaneously and improve food conversion ratio.Although it is safe that the recommended dose of Salinomycin and monensin uses, but in actual production, poisoning that is excessive due to using dosage and that cause happens occasionally.

At present, in animal tissue, the detection of Salinomycin and monensin mostly is LC-MS detection method, but it is rare with the report of monensin to detect Salinomycin for the high performance liquid chromatography triple level Four bar mass spectrums of series connection, therefore, set up a kind of method of Salinomycin and monensin in disposable synchronous detecting domestic animal product, it is achieved detection limit is lower, detection accuracy is higher, detection efficiency is higher very necessary.

Summary of the invention

It is an object of the invention to overcome the deficiency of existing detection technique, it is provided that the method for Salinomycin and monensin in the domestic animal product that a kind of detection limit is lower, detection accuracy is higher.

A kind of detect the method for Salinomycin and monensin in livestock products, it is characterised in that utilize Salinomycin and monensin in the high performance liquid chromatography series connection disposable synchronous detecting domestic animal product of triple level Four bar GC-MS, comprise the steps:

(1) extract: weigh 2.00-2.50g sample, be placed in centrifuge tube, add the mixing of 10-12mL acetonitrile, add 3.00-3.20g anhydrous sodium sulfate, concussion mixing, supersound extraction 10-12min；

Centrifuging and taking supernatant is in 50mL centrifuge tube, and residue adds 5-6mL acetonitrile to be repeated to extract, and merges twice supernatant, is settled to 20mL, adds 5mL acetonitrile and 5mL normal hexane, and concussion is centrifugal, to be clean.

(2) purify: taking off layer (acetonitrile layer) 5mL in round-bottomed flask, rotary evaporation is to 1mL, and nitrogen dries up；

Common meat matrix: add 1mL acetonitrile ultrasonic dissolution in round-bottomed flask；

Internal organs class substrate: add 3mL methanol+water (1:1) ultrasonic dissolution in round-bottomed flask, cross WatersHLB post (watersoasisHLB6cc/200mg is successively with the activation of 5mL methanol 5mL water), use 5mL water wash, 5mL methanol-eluted fractions is in 100mL round-bottomed flask, 40 DEG C of evaporated under reduced pressure, add 1mL acetonitrile ultrasonic dissolution.

(3) detection: adopting High Performance Liquid Chromatography/Mass Spectrometry that sample is detected, testing conditions is as follows:

Mass spectrometry parameters: ion source ESI source；Atomization gas flow 3L/min；Thermal current 10L/min；Interface temperature 300 DEG C；Desolventizing pipe temperature 250 DEG C；Heating block temperature 400 DEG C；Dry gas stream amount 10L/min.

Liquid chromatograph parameter: mobile phase forms: A:0.1% formic acid B: acetonitrile；Flow velocity: 0.35Ml/min；A-10%B-90% constant current is analyzed；Sample size: 2uL；Chromatographic column model: ODS-III1.6 μm.

The effect of the present invention:

The liquid chromatograph used compared with tradition and national standard method is detecting instrument, this forwarding method uses liquid chromatogram-triple tandem quadrupole GC-MS, and detection limit is lower, and detection accuracy is higher, relevant detection limit reaches Salinomycin 2 μ g/kg, monensin 10 μ g/kg.For common meat matrix, it is possible to the samples such as extraction concentrated solution assistant director higher level analyzes simply, liver use HLB decontaminating column pre-treatment, relatively easy.Meanwhile, Salinomycin is merged with monensin detection, reach pre-treatment simultaneously and carry out Instrumental Analysis, effect in hgher efficiency.

Detailed description of the invention

Below in conjunction with specific embodiment, technical scheme is elaborated; but do not limit protection scope of the present invention; any change that the present invention is done by those possessing an ordinary skill in the pertinent arts; without departing from the essence of the present invention, all equivalence is dropped in claims of the present invention limited range.

Embodiment 1

A kind of detect the method for Salinomycin and monensin in beef, including step once:

Extract: weigh beef 2.00g to be detected, add the mixing of 10ml acetonitrile, add 3.00g anhydrous sodium sulfate, concussion mixing.Supersound extraction 10min, centrifuging and taking supernatant is in 50ml centrifuge tube；Residue adds 5ml acetonitrile to be repeated to extract, and merges twice supernatant, is settled to 20mL, adds 5mL acetonitrile and 5mL normal hexane, and concussion is centrifugal, to be clean.

Purify:

Taking off layer (acetonitrile layer) 5mL in round-bottomed flask, rotary evaporation is to 1mL, and nitrogen dries up.

Detection:

Adopting High Performance Liquid Chromatography/Mass Spectrometry that sample is detected, testing conditions is as follows:

Mass spectrometry parameters: ion source ESI source；Atomization gas flow 3L/min；Thermal current 10L/min；Interface temperature 300 DEG C；Desolventizing pipe temperature 250 DEG C；Heating block temperature 400 DEG C；Dry gas stream amount 10L/min.

Liquid chromatograph parameter: mobile phase forms: A:0.1% formic acid B: acetonitrile；Flow velocity: 0.35Ml/min；A-10%B-90% constant current is analyzed；Sample size: 2uL；Chromatographic column model: ODS-III1.6 μm.

Instrument linear analysis:

The mixed mark (retarder thinner: acetonitrile) of preparation Salinomycin, monensin, five concentration point of 0.1ug/kg, 1ug/kg, 5ug/kg, 10ug/kg, 20ug/kg, Instrumental Analysis linear analysis:

Salinomycin, monensin standard curve R2 are all higher than 0.99, and standard substance goes out peak Salinomycin 1 μ g/kg and goes out peak response value well, and to-noise ratio S/N is far longer than 10, and monensin 5 μ g/kg response value is good, and to-noise ratio S/N is far longer than 10;Each concentration point precision is good；Instrumental Analysis standard substance are linear, stability meets requirement of experiment.

Really spend analysis:

Blank beef being carried out pre-treatment, adopts blank sample extract and solvent dilution unified standard product standard substance, under same concentration, the percentage ratio of both peak area ratios is as really spending evaluating.

Experimental result is as follows:

More than in experiment, beef substrate is to Salinomycin without obvious matrix effect, the matrix effect that monensin generation is certain, and matrix effect is about 70%, and within tolerance interval, routine analysis accurate quantitative analysis can do spiked correction.

Selectivity is analyzed:

Choosing beef sample to carry out blank, add recovery experiment, every level adds recovery sample size N=6, shown in table specific as follows:

By experimental data it can be seen that Salinomycin three level adds the repeatability reclaiming the response rate and precision is all good, meet requirement of experiment；The repeatability that monensin three level, six parallel interpolations are reclaimed is good but the response rate is on the low side about 60%, the matrix effect of beef substrate existence about 70%, and after spiked correction, the response rate is normal

。

Lower limit of quantitation:

Comprehensive standard material goes out peak Salinomycin 1 μ g/kg and goes out peak response value well, and to-noise ratio S/N is far longer than 10, and monensin 5 μ g/kg response value is good, and to-noise ratio S/N is far longer than 10;Empty adds recovery selectivity and separating degree is good；Repeatability is reclaimed in the interpolation of varying level and precision is good；By Salinomycin, monensin sample in detection limit be set to following:

Compound title	Lower limit of quantitation (μ g/kg)
		Salinomycin	2
Monensin	10

Claims (1)

1. one kind is detected the method for Salinomycin and monensin in livestock products, it is characterised in that utilizes Salinomycin and monensin in the high performance liquid chromatography series connection disposable synchronous detecting domestic animal product of triple level Four bar GC-MS, comprises the steps:

(1) extract: weigh 2.00-2.50g sample, be placed in centrifuge tube, add the mixing of 10-12mL acetonitrile, add 3.00-3.20g anhydrous sodium sulfate, concussion mixing, supersound extraction 10-12min；Centrifuging and taking supernatant is in 50mL centrifuge tube, and residue adds 5-6mL acetonitrile to be repeated to extract, and merges twice supernatant, is settled to 20mL, adds 5mL acetonitrile and 5mL normal hexane, and concussion is centrifugal, to be clean；

(2) purify: taking off layer (acetonitrile layer) 5mL in round-bottomed flask, rotary evaporation is to 1mL, and nitrogen dries up；

Common meat matrix: add 1mL acetonitrile ultrasonic dissolution in round-bottomed flask；

Internal organs class substrate: add 3mL methanol+water (1:1) ultrasonic dissolution in round-bottomed flask, cross WatersHLB post (watersoasisHLB6cc/200mg is successively with the activation of 5mL methanol 5mL water), use 5mL water wash, 5mL methanol-eluted fractions is in 100mL round-bottomed flask, 40 DEG C of evaporated under reduced pressure, add 1mL acetonitrile ultrasonic dissolution；

(3) detection: adopting High Performance Liquid Chromatography/Mass Spectrometry that sample is detected, testing conditions is as follows:

Mass spectrometry parameters: ion source ESI source；Atomization gas flow 3L/min；Thermal current 10L/min；Interface temperature 300 DEG C；Desolventizing pipe temperature 250 DEG C；Heating block temperature 400 DEG C；Dry gas stream amount 10L/min；

Liquid chromatograph parameter: mobile phase forms: A:0.1% formic acid B: acetonitrile；Flow velocity: 0.35Ml/min；A-10%B-90% constant current is analyzed；Sample size: 2uL；Chromatographic column model: ODS-III1.6 μm.