CN105651915A

CN105651915A - Method for detecting ochratoxin A in wine

Info

Publication number: CN105651915A
Application number: CN201510989885.7A
Authority: CN
Inventors: 张鑫; 雷纪锋; 吴祖芳; 翁佩芳
Original assignee: Ningbo University
Current assignee: Ningbo University
Priority date: 2015-12-24
Filing date: 2015-12-24
Publication date: 2016-06-08

Abstract

The invention discloses a method for detecting ochratoxin A in wine. The main steps include: taking a wine sample, adding a methanol solution, extracting by a homogenizer at high speed stirring, centrifuging to obtain a supernatant, treating by immunoaffinity column combined with glass microfiber filter paper, and detecting by a liquid chromatography-quadrupole static electric field orbitrap mass spectrometry method. The immunoaffinity column combined with glass microfiber filter paper is selected for pretreatment; liquid chromatography-quadrupole static electric field orbitrap mass spectrometry method is used to detect content of ochratoxin A in wine; and the method has the advantages of high recovery rate, low detection limit and low cost for pretreatment.

Description

The detection method of ochratoxin A in a kind of wine

Technical field

The invention belongs to food inspection technique, relate to the detection method of ochratoxin A in a kind of wine.

Background technology

Wine is internationally recognized health drink, is increase one of very fast, most active industry in the national economic development. In recent years, domestic Wine Market consumption annual rate of growth continued to keep higher level, and growth space is big. Compared to import wine, there is the defects such as product homogeneity, security-hidden trouble be big in domestic wine, has badly influenced the sound development of domestic Grape Wine Industry. Therefore, improving Production of Wine level, eliminating security-hidden trouble is the main policies improving China's wine international competitiveness, the key of Ye Shi China Grape Wine Industry sustainable and stable development.

Ochratoxin is mainly by the mycetogenetic class secondary metabolite of aspergillus (Aspergillussp.) and Penicillium (Penicilliumsp.), and ochratoxin A is that most important of which is a kind of. Research finds, ochratoxin A has the features such as nephrotoxicity, liver toxicity, genotoxicity, teratogenecity, immunotoxicity and fetal toxicity, and toxicity is only second to aflatoxin. Ochratoxin A was divided into IIB class carcinogen by international cancer research institution in 1993, big quantity research confirms that ochratoxin A is widely present in corn, fruit, coffee and wine, and the pollution of ochratoxin A producing strains becomes the important factor threatening the edible safeties such as Fructus Vitis viniferae, dried Fructus Vitis viniferae, Sucus Vitis viniferae and wine. As a kind of health drink, wine is subject to consumers in general more and more to be favored, and in wine, consumer safety is constituted significant threat by the existence of ochratoxin A. For promoting quality safety vinous on root, need badly and set up the detection method that in a kind of wine, ochratoxin A is accurate, efficient and economic,

Summary of the invention

The technical problem to be solved is to provide in a kind of wine the detection method of ochratoxin A, it is intended to lays the foundation for researchs such as the formation source of ochratoxin A in food and transportion and transformation, provides reference for coherent detection simultaneously.

For solving the problems referred to above, the invention provides the detection method of ochratoxin A in a kind of wine, comprise the steps:

1) wine samples is added in methanol solution;

2) utilizing homogenizer, high-speed stirred extracts the ochratoxin A in wine samples;

3), after stirring is extracted, it is centrifuged and obtains supernatant;

4) supernatant is taken after glass microfibre filter paper filters, loading immune affinity column, use methanol-eluted fractions immune affinity column after collecting eluent, twice eluent merges rotary evaporated to dryness, collects by filtration filtrate after redissolving with water.

5) liquid chromatograph-level Four bar electrostatic field Orbitrap mass method is adopted to detect for the ochratoxin A content in filtrate.

Compared with prior art, it is an advantage of the current invention that the detection method of ochratoxin A in a kind of wine, by wine samples is added methanol solution, homogenizer high-speed stirred is extracted, supernatant is obtained after centrifugal, after processing in conjunction with glass microfibre filter paper with immune affinity column, adopt the ochratoxin A content in liquid chromatograph-level Four bar electrostatic field Orbitrap mass method detection filtrate. This detection method response rate is high, and detection limit is low and pre-treatment cost is low, can promote as the detection method of ochratoxin A in a kind of relatively accurate, efficient and economic wine.

Detailed description of the invention

1) adding in methanol solution by wine samples, it is 1% that wine samples accounts for the ratio of methanol solution, and in methanol solution, methanol is 80: 20 with the volume fraction of water;

2) utilizing homogenizer, high-speed stirred extracts the ochratoxin A in wine samples, and homogenizer running speed is 8000r/min, and stirring extraction time is 5min;

3) stirring is centrifuged after extracting, and the centrifugal 5min of rotating speed 3000r/min obtains supernatant;

4) 1mL centrifuged supernatant is taken, with the speed loading immune affinity column of 2mL/min after glass microfibre filter paper filters, after collecting eluent, take the 5mL methanol speed eluting immune affinity column with 2mL/min, twice eluent merges rotary evaporated to dryness, after adding the redissolution of 1mL water, with 0.45 ��m of filtering with microporous membrane, collect filtrate;

5) adopting liquid chromatograph-level Four bar electrostatic field Orbitrap mass method to detect for the ochratoxin A content in filtrate, ion source is Negative electrospray ionization pattern, and capillary temperature is 320 DEG C, atomization temperature is 300 DEG C, sheath gas is 40Arb, and assisted gas is 10Arb, and voltage is 3800V.

Claims

1. a detection method for ochratoxin A in wine, comprises the steps:

1) wine samples is added in methanol solution;

3), after stirring is extracted, centrifuge is utilized to obtain supernatant;

2. the detection method of ochratoxin A in a kind of wine according to claim 1, it is characterised in that step 1) in wine samples to account for the ratio of methanol solution be 1-2%, in methanol solution, methanol is 80: 20 with the volume fraction of water.

3. the detection method of ochratoxin A in a kind of wine according to claim 1, it is characterised in that step 2) in homogenizer running speed be 8000-10000r/min, stirring extraction time is 3-5min.

4. the detection method of ochratoxin A in a kind of wine according to claim 1, it is characterised in that step 3) in the centrifuge time be 5-8min, rotating speed 3000-4000r/min.

5. the detection method of ochratoxin A in a kind of wine according to claim 1, it is characterized in that step 4) in take 1mL centrifuged supernatant, with the speed loading immune affinity column of 2-3mL/min after glass microfibre filter paper filters, the 5mL methanol speed eluting immune affinity column with 2-3mL/min is taken after collecting eluent, twice eluent merges rotary evaporated to dryness, after adding the redissolution of 1mL water, with 0.45 ��m of filtering with microporous membrane, collect filtrate.

6. the detection method of ochratoxin A in a kind of wine according to claim 1, it is characterised in that step 5) in, ion source is Negative electrospray ionization pattern, capillary temperature is 320 DEG C, and atomization temperature is 300 DEG C, and sheath gas is 40Arb, assisted gas is 10Arb, and voltage is 3800V.