CN101071099A - Method for detecting fat-soluble azo dyes in food or fodder and its kit - Google Patents
Method for detecting fat-soluble azo dyes in food or fodder and its kit Download PDFInfo
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Abstract
The invention discloses a the fat-soluble azo dye detection method and kit in the food or feed, which includes the following steps: (1) Preparation for the test solution: to be examined samples using organic solvent dilution, dissolved or extraction, as evidence test solution; (2) adding the test solution of excessive oxidant; (3) adding reducing agent to remove excess oxidizer, (4) static hierarchical, according to the color of organic layer judge whether the seized samples containing Sudan, and other fat-soluble azo dyes. The detection method of this invention is low cost, specific, sensitive and simple rapid, and does not require expensive equipment, and it can be used for the scene rapid detection of food or feed whether added illegal Sudan, and other fat-soluble azo dyes. At the same time it provides an apid screening test to see whether the food or feed contain Sudan or fat-soluble azo dyes illegally.
Description
Technical field
The present invention relates to the food hygiene detection field, specifically, relate to fat-soluble azo dye detection method and kit such as tonyred in a kind of food or the feed.
Background technology
At present, chemical synthetic dye is existing several thousand kinds.Azo dyes accounts for 2/3rds of sum.Azo dyes is the maximum class of quantity in the synthetic dyestuffs, kind, is widely used industrial.Azo dyes can be divided into fat-soluble azo dyes (or being called the lipophilicity azo dyes) and soluble azo dyes (or being called the water wettability azo dyes) again.Azo-compound non-natural in food exists.Because the azo dyes great majority have stronger toxicity, can as food additives use for number seldom.In China, have only lemon yellow a few water-soluble dye that waits to be allowed to use, and its consumption is had strict regulation as food additives.Fat-soluble azo dyes is inhibited in the food and adds.
In food, use pigment can change " appearance " of commodity, improve its value.Because natural colouring matter exists and to cost an arm and a leg, colouring power is poor, shortcomings such as fugitive color, and azo dyes has characteristics such as strong coloring force, price are low, bright-colored, so azo dyes often is added in the food by illegal.
Fat-soluble azo dyes is cheap and easy to get, the industrial dyeing that is usually used in products such as floor wax, shoe polish, machine oil.In recent years, the food security incident that is caused about the fat-soluble azo dyes of illegal interpolation in food and feed happens occasionally.These dyestuffs comprise Sudan red 1, Sudan red 1 I, Sudan red III, Sudan red 1 V, sudan orange G, sudan red 7B, tonyred G, para red etc.
In April, 2005, No. 43 bulletin of General Administration of Quality Supervision, Inspection and Quarantine o of the People's Republic of China's issue pointed out: " the international cancer research institution (IARC) according to the World Health Organization (WHO) determines nineteen ninety-five; tonyred belongs to the 3rd class carcinogen; it can make mouse and rabbit cancer stricken, but does not also have positive evidence to show that the people is had carcinogenesis.Be higher than human body possibility intake far away because tonyred is brought out the dosage of animal tumor, therefore, it is less to the possibility that human body works the mischief to take in the food that contains a small amount of tonyred accidentally, if but long-term heavy dose of absorption meeting increases human carcinogen's danger.Thereby China and many countries all forbid using it for food production.”
State General Administration for Quality Supervision organized national quality inspection system that pungent goods, the catsup goods that may contain tonyred have been carried out special examination in 2005.Find that by checking minority Additive Production enterprise pretends to be the edible additive capsicum red pigment to be sold to pungent production of articles enterprise with the tonyred chemical dyestuff, sees that State Administration for Quality Supervision and Inspection and Quarantine announces No. 54 (2005-04-11).
As far back as 2003, for preventing in import pimiento and the product doping carcinogen tonyred No. one, European Commission, determines taking urgent measure from import pimiento of non-member countries and products thereof about " European Union is about checking the 2003/460/EC resolution of a chemical dyestuff of tonyred in import pimiento and products thereof severely " in June 20 issue.
For guaranteeing that China defeated Europe pimiento and products thereof meets European Union's requirement, State Administration for Quality Supervision and Inspection and Quarantine has sent " about forbidding to use the emergency notice of a chemical dyestuff of tonyred in defeated European Union pimiento and products thereof " (No. [2003] 620, state's quality inspection food letter, on August 1st, 2003), require each to strengthen conscientiously failing inspection and quarantine and the supervision and management of European Union pimiento and products thereof directly under inspection and quarantine bureau.With this regulation written notice local government, each relevant enterprise of European Commission, require export enterprise to strengthen the quality automatic control work of links such as purchase, processing as early as possible, guarantee to export pimiento and product and do not have tonyred No. one.
2005, FSA (FSA) issue warning reminded the public to watch out for the pigment of a kind of crying " para red ".This office claims, has found " para red " in the pungent enchilada of their certain plate and the burrito.Subsequently, FSA is found 69 kinds of food that contain " para red ", comprises flavouring such as curry paste that people use always, thick chilli sauce, barbecue, and this thing one goes out, and has caused the concern of Britain society at once." para red " is that a kind of chemical constitution is similar to the industrial dye of " No. 1, tonyred ".
In November, 2006, State Administration for Quality Supervision and Inspection and Quarantine has issued " about reporting the emergency notice of tonyred special examination result in the egg products " in (No. [2006] 146, quality inspection food prison letter).Emergent Circular claims: recently, the medium wide coverage some raisers of some areas, Hebei province use the forage feed egg duck that contains Sudan red 1 V number and output " red heart " duck's egg, indivedual egg products enterprises are as the illegal activities of raw material production and processing Salted duck egg goods, and this accident has at present caused the concern of more than ten provinces and cities.General bureau sends " about carrying out the emergency notice of tonyred special examination in the egg products immediately " (bright: state's quality inspection bright [2006] No. 62) morning November 14, requires quality testing departments at different levels to carry out the special examination of egg products manufacturing and processing enterprise immediately.
On January 17th, 2007, State General Administration for Quality Supervision sends notice of warning: in the product quality special project government supervision selective examination of State General Administration for Quality Supervision to pre-packing chilli products such as chilli powder, thick chilli sauce, oily capsicum, thick broad-bean sauce and chilli powder in bulk, 137 kinds of product mean sample qualification rates of selective examination only are 74.5%, and defective item all concentrates on Sudan red 1～IV number.80 kinds of pre-packing capsicum products in 8 provinces such as Guizhou, Sichuan, Chongqing, Hunan, Hubei, Guangxi, Shandong, Henan (district, city) manufacturing enterprise, market, supermarket have this time been spot-check altogether, qualification rate is 98.75%, have a kind of product defective, nonconformance is Sudan red 1 V.To the 57 kinds of chilli products in bulk (55 kinds of chilli powders) that extract on 23 market of farm produces, ground such as Guizhou, Sichuan, Chongqing, qualified 23 kinds, qualification rate only is 40%, and underproof 34 kinds of products all detect tonyred.State General Administration for Quality Supervision requires chilli products manufacturing and processing enterprise to set up the forced examination system, and tonyred is essential items for inspection, and product export must be up to the standards.Enterprise does not possess detectability voluntarily, must entrust the inspection body with legal qualification to test.
Above-mentioned situation shows that food is subjected to chemical synthetic dye pollution problems such as tonyred that generation is often arranged, and is subjected to paying attention to widely at home and abroad.For banning the poisoned food of mixing chemical synthetic dyes such as tonyred effectively, help manufacturing enterprise to carry out from performing check, it is very necessary to set up simple and easy to do method for quick.
Detect the method for whether having added fat-soluble azo dyes such as tonyred in the food at present and mainly contain high performance liquid chromatography (LC method) and high performance liquid chromatography-mass spectroscopy (LC-MS method).The former is existing national standard method (the detection method high performance liquid chromatography of tonyred dyestuff in the food; State Standard of the People's Republic of China GB/T 19681-2005), the latter is the conclusive evidence method that European Union uses.
The advantage of high performance liquid chromatography is chromatographic resolution efficient height, and is highly sensitive.Its shortcoming is: (1) instrument costs an arm and a leg, and time for sample pretreatment is long, chromatographic column vulnerable to pollution, analysis cost height; (2) when the chromatographic retention of chromatographic retention and the target component of coexistence composition near the time judgement that makes mistake of do easily; (3) instrument requires height to environment for use, needs fixedly put, and the mobile big scene of uncomfortable cooperation is detected; (4) to reviewer's technical requirement height.
Be suitable for complicated component, sample analysis that background interference serious as separation vessel, mass spectrometer as the high performance liquid chromatography liquid-mass spectroscopy of analyzer with high performance liquid chromatograph, this analytical approach can improve the reliability of testing result.High performance liquid chromatography liquid-mass spectroscopy is that a kind of analysis cost is higher than high performance liquid chromatography, operation is than high performance liquid chromatography complex analysis methods more.Instrument requires higher to environment for use, need fixedly to put, and the mobile big scene of uncomfortable cooperation is detected.These checks can only be carried out in a few experiments chamber, can't popularize use.
Government regulator is investigated and prosecuted added this illegal activities of fat-soluble azo dyes such as tonyred in food, will be a long-term task.Its coverage is wide, need carry out a large amount of checks, if detect with efficient liquid-phase chromatography method all, then survey fees are with huge, administrative law enforcement cost height; In vast rural area and small and medium-sized cities underdeveloped, because do not have instrument and relevant detection technician, and can't carry out this detection, thereby make these areas become the no-man's-land of supervision.A lot of manufacturing enterprises are not because have instrument and relevant detection technician yet, and can't raw material and finished product be carried out from performing check, have stayed food security hidden danger.
For ensuring food safety, guarantee people's life health, press for set up a kind of simple, rapidly and efficiently, the detection food with low cost method of fat-soluble azo dyes such as illegal interpolation tonyred whether.
In view of can be a lot of by the kind of the fat-soluble azo dyes of illegal interpolation, only obviously there be deficiency in the instrument analytical method that detects at one or more compounds in Sudan red 1, Sudan red 1 I, Sudan red III, Sudan red 1 V, sudan orange G, sudan red 7B, tonyred G and the para red.Be subjected to effects limit such as chromatographic resolution rate, coexistence chaff interference, standard items, instrument analytical method generally once can only carry out examination to several or a ten or twenty target compound, and the kind of fat-soluble azo dyes that can be used for illegal interpolation is numerous, when the dyestuff that adds not in preset range, omission will appear in instrument analytical method.Therefore need set up a kind of rapid screening detection method that can contain most of fat-soluble azo dyes.
Summary of the invention
The objective of the invention is to overcome the shortcoming that prior art exists, provide a kind of analysis cost low, do not need to use valuable analytical instrument, specificity strong, highly sensitive, be easy to popularize and be applicable to and on-the-spotly detect food or whether feed has added fat-soluble azo dye detection method such as tonyred.
Another object of the present invention provides utilizes above-mentioned detection method to detect the kit of fat-soluble azo dyes such as tonyred.
To achieve these goals, detection method of the present invention comprises the steps:
(1) preparation of test liquid: sample to be checked adopts organic solvent diluting, dissolving or extraction, makes test liquid;
Oily liquids sample (as chilli oil) is made test liquid after with an amount of organic solvent diluting; Solid or semifluid oil product are made test liquid behind organic solvent dissolution; Other sample (as chilli powder, thick chilli sauce, catsup, cold cuts, preserved radish strip, ham sausage, yolk, cake, pungent dried fish, powdery and granulated meal etc.) is then used organic solvent extraction, and extract is made test liquid.
(2) add excessive oxidant in test liquid, oxidation reaction is removed coloured interfering material.
(3) add reductive agent and remove superfluous oxygenant, eliminate of the influence of superfluous oxygenant later observation.
(4) standing demix judges according to the color of organic layer whether sample to be checked contains fat-soluble azo dyes; If it is colourless or almost colourless that organic layer is, then negative, do not contain fat-soluble azo dyes; If organic layer is color is arranged, then positive, contain fat-soluble azo dyes probably.
In above-mentioned detection method, organic solvent is meant the organic solvent with following character: (a) immiscible with water; (b) can not be by potassium permanganate oxidation; (c) tangible hydrolysis reaction can not take place in the presence of sulfuric acid.The potpourri of one or more in homolog, benzene, toluene, substituted benzene, chloroform, methylene chloride or the sherwood oil of the homolog of the preferred cyclohexane of these organic solvents, cyclohexane, normal hexane, normal hexane.
In above-mentioned detection method, the preferred liquor potassic permanganate of described oxygenant, as preferably also adding sulfuric acid solution, the purpose that adds sulfuric acid is to improve the oxidizing potential of potassium permanganate, makes it enough oxidabilities, improves the speed of oxidation reaction.
In above-mentioned detection method, the oxalic acid solution of the oxammonium hydrochloride of preferred 5～12 weight % of described reductive agent, the ascorbic acid solution of 5～12 weight % or 5～12 weight %.
The kit that utilizes above-mentioned detection method to detect comprises organic solvent, Oxidizing and Reducing Agents.
Azo dyes is artificial chemosynthesis material, and the azo dyes azo dyes can be divided into water-soluble dye and fat-soluble azo dyes, and the present invention only relates to fat-soluble azo dyes.Azo-compound all is a colored compound.Understand for convenient, name and the dissolubility of listing part azo dyes commonly used are as follows:
The chemical name of Sudan red 1 (SudanI) is 1-phenylazo-beta naphthal, (1-phenylazo-2-naphthalenol), is raw material with aniline, carries out diazo-reaction with sodium nitrite in hydrochloric acid, generates diazo salt, carries out coupling with beta naphthal again and makes.Dissolubility: water insoluble, be dissolved in grease, cyclohexane, mineral oil, acetone and benzene.
Sudan red 1 I (SudanII) chemical name is 1-[(2, the 4-dimethyl benzene) azo]-beta naphthal, (1-[(2,4-dimethylphenyl) azo]-2-naphthalenol); By 2, after the 4-xylidin diazotising, make with the betanaphthol coupling.Dissolubility: be dissolved in ether, benzene, volatile oil, cyclohexane, fat, the oil water insoluble, alkali, weak acid.
Sudan red III (SudanIII) chemical name is 1-[4-(phenylazo) phenyl] azo-beta naphthal, (1-[4-(phenylazo) phenyl] azo]-2-naphthalenol); Make by aminoazabenzol diazotising and betanaphthol coupling.Dissolubility: be dissolved in ether, acetone, sherwood oil, grease, cyclohexane, chloroform, glacial acetic acid, ethanol, water insoluble, aqueous slkali.
Sudan red 1 V (SudanIV) chemical name is 1-2-methyl-4-[(2-methylbenzene) azo] phenylazo-beta naphthal,
(1-2-methyl-4-[[(2-methylphenyl) azo] phenylazo]-2-naphthalenol), make by o-aminoazotoluene diazotising and betanaphthol coupling.Dissolubility: be dissolved in ether, ethanol, cyclohexane, grease, phenol, water insoluble.
Tonyred G (Sudan red G) chemical name is 1-(2-methoxyphenyl) azo-beta naphthal, 1-(2-Methoxyphenylazo)-2-naphthol.Dissolubility: be dissolved in ether, cyclohexane, ethanol, grease, phenol, water insoluble.
Sudan red 7B (Sudan Red 7B) chemical name is: 2-Naphthalenamine, N-ethyl-1-[[4-(phenylazo) phenyl] azo]-; CAS Number 6368-72-5; Dissolubility: be dissolved in ether, acetone, cyclohexane, grease, water insoluble.
Sudan orange G (Sudan Orange G) chemical name is 2,4-dihydroxy azobenzene; 2,4-Dihydroxyazobenzene.Dissolubility: be dissolved in ether, ethanol, cyclohexane, grease, phenol, water insoluble.
Para red (Para Red) chemical name is 1-p-nitrophenyl azo-beta naphthal, and 1-(4-Nitrophenylazo)-2-naphthol also claims that paranitraniline is red.Dissolubility: be dissolved in ether, acetone, grease, ethanol (solution takes on a red color), water insoluble.
The chemical structure characteristic of azo dyes is one or more azo groups to be arranged in the molecule (N=N-), the one or both ends of azo group link to each other with aromatic ring, and for example: azo group one end of Sudan red 1 links to each other with phenyl ring, and the other end links to each other with naphthalene nucleus; What the azo group two ends of sudan orange G linked to each other all is phenyl ring; Lemon yellow (C
16H
9N
4O
9S
2Na
3) azo group have only an end to link to each other with aromatic ring.Connect different substituting groups at the aromatic ring that links to each other with azo group and then derive different azo-compounds, therefore can derive a variety of azo aromatics.Usually, on aromatic ring, connect one or more sulfonic groups, then the water-soluble enhancing of azo-compound.
Most of fat-soluble azo dyes have at least the two ends of an azo group all to connect aromatic ring in its molecular structure.Form a big conjugated system because aromatic ring links to each other by azo group, pi-electron has bigger delocalization scope, the certain wavelength in the visible light absorbing, so azo-compound all is a colored compound.Making the key that the azo dyes degraded is faded is to destroy its azo group.
Two keys of N=N on the azo group and aromatic ring form the delocalization conjugated system, the result of conjugation has strengthened on azo group nitrogen-atoms and the aromatic ring connection stability between carbon atom, also strengthened the stability of azo group itself, simultaneously also make the lone pair electrons on the nitrogen-atoms be not easy to lose, so azo group itself is not easy oxidized.When the azo group two ends all were connected with aromatic ring, this conjugative effect was more obvious.Be that compound is more stable, more difficult oxidized, color is darker.Because the p electronics on the carbon atom of aromatic ring own has participated in conjugation, has formed rock-steady structure, so aromatic ring also is not easy oxidized dose of destruction.Under there was situation in oxygenant, some substituting group on the aromatic ring may be oxidized, but can not make compound become colorless (this is that big conjugated system also exists because aromatic ring and azo group also exist).Reason to sum up, under certain concentration and acidity, liquor potassic permanganate can not destroy the chromophoric group of azo dyes, and azo dyes is faded.
Natural colouring matters such as beta carotene, capsorubin and other carotenoid extensively exist in plant, are fat-soluble pigments.Compounds such as beta carotene, capsorubin and other carotenoid contain a plurality of conjugated double bonds, and this is that they show coloured reason.These pairs key is easily ruptured by the strong oxidizer oxidation, and oxidized back conjugated system has not existed.For example, contain a plurality of conjugated double bonds in the carrotene molecular structure, have strong reducing property, two keys are easily oxidized and rupture, and are generated as colourless substance.
Water-soluble colored compound in the natural products as flavones saponins pigment, is insoluble to lipophilic solvents such as grease and cyclohexane.By the extraction of lipophilic solvents such as cyclohexane, water-soluble colored compound is separated with fat-soluble azo dyes.
Cardinal principle of the present invention is as follows:
(1) utilizes lipophilicity organic solvent diluting oily liquids food (as chilli oil), make it when jolting, be dispersed into droplet easily, fully contact, the finishing of accelerated reaction with oxygenant.
Utilize lipophilicity organic solvent (as cyclohexane) from sample, to extract fat-soluble azo dyes.
(2) utilize difficult oxidized the fading of azo dyes and the easy oxidized characteristics of fading of natural fat-soluble pigment, under the condition that sulfuric acid exists, adopt excessive strong oxidizer potassium permanganate oxidation natural colouring matter, reach the purpose that the natural colouring matter of eliminating coexistence disturbs.
(3) utilize the superfluous oxygenant of reductive agent (example hydrochloric acid azanol, oxalic acid or ascorbic acid) reduction, eliminate the influence that later observation is judged.
(4) characteristic of utilizing azo dyes self to show color is judged.
Compared with prior art, the present invention has following beneficial effect:
The method of inspection of the present invention has overcome the shortcoming that prior art exists, have that analysis cost is low, specificity is strong, highly sensitive, easy and simple to handle fast, do not need to use advantages such as expensive instrument, whether be applicable in field quick detection food and the feed fat-soluble azo dyes such as illegal interpolation tonyred, satisfy the needs of food and feed supervision and inspection; Simultaneously for check food and feed whether the specialized laboratories of fat-soluble azo dyes such as illegal interpolation tonyred screening test method fast is provided, can reach the purpose that reduces analysis cost, improves checkability.
Reagent for quickly examining box manufactured according to the present invention can satisfy the needs of vast rural area and economically underdeveloped area's conducting food and feed supervision and inspection, also can satisfy manufacturing enterprise and commercial unit carry out self check to relevant food and feed needs.
Embodiment
Embodiment 1
Chilli oil 1 (commercially available bottled, it is orange red that chilli oil is, and known every mL contains the Sudan I 0.3mg)
The first step: draw the about 0.2mL of chilli oil with suction pipe, place tool plug test tube, (purpose that adds cyclohexane is to reduce the viscosity of oil to add cyclohexane 0.8mL, make and when the test tube jolting, split into droplet easily, increase the oxidation reaction contact area, make in the oil coloured interfering material faster oxidized);
Second step: the adding concentration expressed in percentage by weight is 10% sulfuric acid solution 5mL (purpose that adds sulfuric acid is to improve the oxidizing potential of potassium permanganate, makes it enough oxidabilities, improves the speed of oxidation reaction);
The 3rd step: adding volumetric molar concentration is the liquor potassic permanganate 1mL of 0.1mol/L, and firmly jolting is about 5 minutes, leaves standstill 2 minutes, and the water layer in the test tube still is tangible aubergine.
The 4th step: splash into concentration expressed in percentage by weight again and be 10% oxammonium hydrochloride solution,, become colorless to water layer while splash into the jolting test tube.
The 5th step: leave standstill, treat the color of observing organic layer after the layering.
This experimental observation is significantly orange-yellow to the color of organic layer, may contain fat-soluble azo dyes in the judgement sample thus.
Embodiment 2
Chilli powder 1 (commercially available in bulk, take on a red color, detect the known dyestuffs such as the Sudan I, the Sudan II, soudan III, Sudan IV, sudan orange G, sudan red 7B, tonyred G, para red that do not contain with high performance liquid chromatography).
The first step: get about 2 grams of chilli powder, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being orange-yellow) after the layering, places another test tube; All the other operations are according to～the five step of the step of second among the embodiment 1.
This experimental observation is clarification, colourless to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 3
Chilli powder 2 (chilli powder of getting among the embodiment 2 is an amount of, adds the Sudan I standard solution, mixes, gets the positive that every gram sample contains the Sudan I0.3mg thoroughly).
All operation is identical with embodiment 2.
This experimental observation organic layer of obtaining of single stepping to the end is orange-yellow, contains fat-soluble azo dyes thus in the judgement sample probably.
Embodiment 4
Chilli powder 3 (chilli powder of getting among the embodiment 2 is an amount of, adds the Sudan I standard solution, mixes, gets the positive that every gram contains the Sudan I 1mg thoroughly).
All operation is identical with embodiment 2.
This experimental observation organic layer of obtaining of single stepping to the end is orange red, contains fat-soluble azo dyes thus in the judgement sample probably.
Among comparing embodiment 3 and the embodiment 4 shade of observed organic layer as can be known, the addition of the Sudan I is many more in the positive, the color of organic layer is dark more.
Embodiment 5
Thick chilli sauce 1 (commercially available bottled, detect the known dyestuffs such as the Sudan I, the Sudan II, soudan III, Sudan IV, sudan orange G, sudan red 7B, tonyred G, para red that do not contain with high performance liquid chromatography).
The first step: get about 8 grams of thick chilli sauce, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being orange-yellow) after the layering, places another test tube; All the other operations are with reference to～the five step of the step of second among the embodiment 1.
This experimental observation is colourless for clarification to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 6
Thick chilli sauce 2 (thick chilli sauce of getting among the embodiment 5 is an amount of, adds the Sudan IV standard solution, mixes, gets the positive that every gram sample contains Sudan IV 1mg thoroughly).
All operation is identical with embodiment 5.
This experimental observation organic layer of obtaining of single stepping to the end is a salmon pink, contains fat-soluble azo dyes thus in the judgement sample probably.
Embodiment 7
Catsup 1 (commercially available bottled, detect the known dyestuffs such as the Sudan I, the Sudan II, soudan III, Sudan IV, sudan orange G, sudan red 7B, tonyred G, para red that do not contain with high performance liquid chromatography).
The first step: get about 4 grams of catsup, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being orange-yellow) after the layering, places another test tube; All the other operations are according to～the five step of the step of second among the embodiment 1.
This experimental observation is colourless for clarification to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 8
Catsup 2 (catsup of getting among the embodiment 7 is an amount of, adds the Sudan IV standard solution, mixes, gets the positive that every gram sample contains Sudan IV 1mg thoroughly).
All operation is identical with embodiment 7, and extract takes on a red color.
This experimental observation organic layer of obtaining of single stepping to the end is a salmon pink, contains fat-soluble azo dyes thus in the judgement sample probably.
Embodiment 9
Thick broad-bean sauce (commercially available bottled)
The first step: get about 8 grams of thick broad-bean sauce, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being orange-yellow) after the layering, places another test tube; All the other operations are according to～the five step of the step of second among the embodiment 1.
This experimental observation is colourless for clarification to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 10
Thick broad-bean sauce (the bottled sample that uses among the embodiment 9 adds the para red standard solution, mixes, gets the positive that every gram contains Sudan IV 0.3mg thoroughly)
All operation is identical with embodiment 9, and extract is orange red.
This experimental observation organic layer of obtaining of single stepping to the end is a salmon pink, contains fat-soluble azo dyes thus in the judgement sample probably.
Embodiment 11
Cut green pepper (commercially available bottled)
The first step: get and cut about 8 grams of green pepper, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being orange-yellow) after the layering, places another test tube; All the other operations are according to～the five step of the step of second among the embodiment 1.
This experimental observation is colourless for clarification to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 12
Cut green pepper (commercially available bottled sample among the embodiment 11 adds the soudan III standard solution, mixes, gets the positive that every gram contains soudan III 0.3mg thoroughly)
All operation is identical with embodiment 11, and extract is orange red.
This experimental observation organic layer of obtaining of single stepping to the end is a salmon pink, contains fat-soluble azo dyes thus in the judgement sample probably.
Embodiment 13
Feed (brown particle shape)
The first step: get about 5 grams of feed, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being yellow) after the layering, places another test tube; All the other operations are according to～the five step of the step of second among the embodiment 1.
This experimental observation is colourless for clarification to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 14
Feed (get the brown particle shape of embodiment 13, add the Sudan IV standard solution, mix, get the positive that every gram contains Sudan IV 0.5mg thoroughly)
All operation is identical with embodiment 13, and extract takes on a red color.
This experimental observation organic layer of obtaining of single stepping to the end contains fat-soluble azo dyes thus probably for red in the judgement sample.
Embodiment 15
Cold cuts (redness): cut the about 4g in sample top layer with cutter, shred, placing specification is the color comparison tube of 50mL, add entry 10mL, add cyclohexane 8mL again, firmly jolting is 5 minutes, leave standstill, the organic layer milkiness that is white in color is drawn organic layer 3mL and is put another test tube, adds saturated nacl aqueous solution 12mL, jolting, leave standstill, treat layering or draw the 1mL organic layer and make test liquid, all the other are operated with embodiment 1.
This experimental observation organic layer of obtaining of single stepping to the end is colourless, does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 16
Cream cake (redness): sample thief 4g, placing specification is the color comparison tube of 50mL, adds entry 10mL, add cyclohexane 8mL again, firmly jolting is 5 minutes, leaves standstill, and organic layer is the blush milkiness, draw organic layer 3mL and put another test tube, add saturated nacl aqueous solution 12mL, jolting is left standstill, treat layering or draw the 1mL organic layer and make test liquid that all the other are operated with embodiment 1.
This experimental observation organic layer of obtaining of single stepping to the end is colourless, does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 17
Ovum Anas domestica yolk (yellow): take by weighing the sample 4g that stirs evenly, placing specification is the color comparison tube of 50mL, adds entry 10mL, add cyclohexane 8mL again, firmly jolting is 5 minutes, leaves standstill, and organic layer is yellow milkiness, draw organic layer 3mL and put another test tube, add saturated nacl aqueous solution 12mL, jolting is left standstill, treat layering or draw the 1mL organic layer and make test liquid that all the other are operated with embodiment 1.
This experimental observation organic layer of obtaining of single stepping to the end is colourless, does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 18
Ovum Anas domestica yolk (Ovum Anas domestica yolk of getting embodiment 17 is an amount of, adds the Sudan IV standard solution, mixes, gets the positive that every gram contains Sudan IV 0.3mg thoroughly)
Take by weighing the sample 4g that stirs evenly, placing specification is the color comparison tube of 50mL, adds entry 10mL, add cyclohexane 8mL again, firmly jolting is 5 minutes, leaves standstill, and organic layer is orange red milkiness, draw organic layer 3mL and put another test tube, add saturated nacl aqueous solution 12mL, jolting is left standstill, treat layering or draw the 1mL organic layer and make test liquid that all the other are operated with embodiment 1.
This experimental observation organic layer of obtaining of single stepping to the end is a salmon pink, contains fat-soluble azo dyes thus in the judgement sample probably.
Embodiment 19
Ham sausage (rose)
Take by weighing the sample 4g that shreds, placing specification is the color comparison tube of 50mL, adds entry 10mL, add cyclohexane 8mL again, firmly jolting is 5 minutes, leaves standstill, and organic layer is the blush milkiness, draw organic layer 3mL and put another test tube, add saturated nacl aqueous solution 12mL, jolting is left standstill, treat layering or draw the 1mL organic layer and make test liquid that all the other are operated with embodiment 1.
This experimental observation organic layer of obtaining of single stepping to the end is colourless, does not contain fat-soluble azo dyes thus in the judgement sample.
Embodiment 20
Preserved radish strip (having the red chili powder)
The first step: get the sample 8g that shreds, placing specification is the color comparison tube of 50mL, adds entry 10mL, adds cyclohexane 8mL again, and firmly jolting is 5 minutes, treats to make test liquid with suction pipe absorption supernatant liquor 1mL (being orange-yellow) after the layering, places another test tube; All the other operations are according to～the five step of the step of second among the embodiment 1.
This experimental observation is colourless for clarification to the organic layer that the operation of the 5th step obtains, and does not contain fat-soluble azo dyes thus in the judgement sample.
In the aforesaid operations, if there is the vortex oscillation device to replace manual jolting, better effects if; When emulsion appears in organic layer, it can be changed in the centrifuge tube, centrifugal treating can be quickened breakdown of emulsion and layering.
Embodiment 21 oxygenants are to the influence of fat-soluble azo dyes color:
Sudan red 1, Sudan red 1 I, Sudan red III, Sudan red 1 V, sudan orange G, sudan red 7B, tonyred G and para red are respectively got in right amount, with the cyclohexane dissolving, make the standard solution of series concentration, and each standard solution is done following processing:
Draw above-mentioned standard solution 1mL with suction pipe, place test tube, the adding concentration expressed in percentage by weight is 10% sulfuric acid solution 5mL, adding volumetric molar concentration again is the liquor potassic permanganate 1mL of 0.1mol/L, firmly jolting is about 5 minutes, leaves standstill, and the water layer in the test tube still is aubergine; Splash into concentration expressed in percentage by weight and be 10% oxammonium hydrochloride solution,, become colorless to water layer while splash into the jolting test tube.Leave standstill, treat to observe after the layering color of organic layer, the results are shown in Table 1.Test findings shows that liquor potassic permanganate can not make azo dyes fade under this test condition.
Chilli oil (orange red) with the AZO dyestuff replaces the cyclohexane give solvent to make the standard solution of series concentration, and each standard solution is done following processing:
Get standard solution (0.5mL) and place same test tube with cyclohexane (0.5mL), the adding concentration expressed in percentage by weight is 10% sulfuric acid solution 5mL, and adding volumetric molar concentration again is the liquor potassic permanganate 1.5mL of 0.1mol/L, and firmly jolting is about 5 minutes, leave standstill, the water layer in the test tube still is aubergine; Splash into concentration expressed in percentage by weight and be 10% oxammonium hydrochloride solution,, become colorless to water layer while splash into the jolting test tube.Leave standstill, treat to observe after the layering color of organic layer, the results are shown in Table 2.Experimental result shows, can be detected as long as the incorporation of fat-soluble azo dyes is not too low.
Embodiment 22 oxygenants to sample copy at the bottom of the influence of color:
Get the chilli oil (0.2mL of known AZO dyestuff, orange red) place same test tube with cyclohexane (0.8mL), the adding concentration expressed in percentage by weight is 10% sulfuric acid solution 5mL, observe organic layer and be orange red, adding volumetric molar concentration again is the liquor potassic permanganate 1.5mL of 0.1mol/L, firmly jolting is about 5 minutes, leaves standstill, and the water layer in the test tube still is aubergine; Splash into concentration expressed in percentage by weight and be 10% oxammonium hydrochloride solution,, become colorless to water layer while splash into the jolting test tube.Leave standstill, treat that the color of observing organic layer after the layering is: orange red disappearance, almost colourless, easily with the fat-soluble azo dyes differentiation.
The cyclohexane extract of samples such as chilli powder, thick chilli sauce, catsup, preserved radish strip, yolk, pungent dried fish also is orange red, and after handling with reference to said method extraction, oxidation and reduction, organic layer becomes colourless.Experimental result shows that fat-soluble natural pigment can oxidizedly fade.
Embodiment 23 adds the experiment of different cultivars and different amount fat-soluble azo dyes
Samples such as chilli powder, thick chilli sauce, catsup, cold cuts, preserved radish strip, ham sausage, yolk, cake, pungent dried fish, powdery and granulated meal, add Sudan red 1, Sudan red 1 I, Sudan red III, Sudan red 1 V, sudan orange G, sudan red 7B, tonyred G and the para red dyestuff of different amounts respectively, experimentize with reference to said method, the result shows: need only fat-soluble azo dyes incorporation 〉=25mg level in every kilogram of (or every liter) sample, then can be detected, phenomenon is obvious; As long as the fat-soluble azo dyes incorporation also can be detected in the 10mg level in every kilogram of (or every liter) sample, phenomenon is still more obvious.
8 kinds of dyestuffs are with the color before and after the reactant in table 1 cyclohexane
| Dye strength | 0.01mg/mL | 0.03mg/mL | 0.05mg/mL | 0.1mg/mL | 0.3mg/mL | 0.5mg/mL | ||||||
| Kind | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant |
| The Sudan I | Yellow | Yellow | Orange | Orange | Orange | Orange | Orange | Orange | Orange red | Orange red | Orange red | Orange red |
| The Sudan II | Orange | Orange | Orange | Orange | Orange | Orange | Orange red | Orange red | Orange | Orange | Orange | Orange |
| Soudan III | Pale red | Pale red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange | Orange | Orange | Orange |
| Sudan IV | Pale red | Pale red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange | Orange | Orange | Orange |
| Sudan orange G | Yellow | Yellow | Orange | Orange | Orange | Orange | Orange red | Orange red | Orange red | Orange red | Orange red | Orange |
| Sudan red 7B | Yellow | Yellow | Orange | Orange | Orange | Orange | Orange | Orange | Orange red | Orange red | Orange | Orange |
| Tonyred G | Pale yellow | Pale yellow | Orange | Yellow | Orange | Orange | Orange | Orange | Orange red | Orange red | Red | Red |
| Para red | Orange | Orange | Orange | Orange | Orange red | Orange red | Orange red | Orange red | Orange | Orange | Orange | Orange |
8 kinds of dyestuffs are with the color before and after the reactant in table 2 chilli oil
| Dye strength | 0.01mg/mL | 0.025m/mL | 0.05mg/mL | 0.1mg/mL | 0.5mg/mL | 1.0mg/mL | ||||||
| Kind | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant | Before reactant | After reactant |
| The Sudan I | Orange red | Yellowish | Orange red | Orange | Orange red | Orange | Orange red | Orange | Orange red | Orange red | Orange red | Orange red |
| The Sudan II | Orange red | Greenish orange Huang | Orange red | Orange red | Orange red | Red | Orange red | Red | Orange red | Red | Orange red | Red |
| Soudan III | Orange red | Orange red | Orange red | Red | Orange red | Red | Orange red | Red | Orange red | Red | Orange red | Red |
| Sudan IV | Orange red | Orange red | Orange red | Red | Orange red | Red | Orange red | Red | Orange red | Red | Orange red | Red |
| Sudan orange G | Orange red | Greenish orange Huang | Orange red | Orange | Orange red | Orange | Orange red | Orange | Orange red | Orange red | Orange red | Orange red |
| Sudan red 7B | Orange red | Greenish orange Huang | Orange red | Orange | Orange red | Orange | Orange red | Orange | Orange red | Orange | Orange red | Orange red |
| Tonyred G | Orange red | Yellowish | Orange red | Orange | Orange red | Orange | Orange red | Orange | Orange red | Orange red | Orange red | Orange red |
| Para red | Orange red | Orange | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red | Orange red |
Claims (6)
1. fat-soluble azo dye detection method in food or the feed is characterized in that comprising the steps:
(1) preparation of test liquid: sample to be checked adopts organic solvent diluting, dissolving or extraction, makes test liquid;
(2) in test liquid, add excessive oxidant;
(3) add reductive agent and remove superfluous oxygenant;
(4) standing demix judges according to the color of organic layer whether sample to be checked contains fat-soluble azo dyes; If it is colourless or almost colourless that organic layer is, then negative, do not contain fat-soluble azo dyes; If organic layer is color is arranged, then positive, contain fat-soluble azo dyes probably.
2. detection method as claimed in claim 1 is characterized in that described organic solvent is one or more the potpourri in homolog, benzene, toluene, substituted benzene, chloroform, methylene chloride or the sherwood oil of homolog, normal hexane, normal hexane of cyclohexane, cyclohexane.
3. detection method as claimed in claim 1 is characterized in that described oxygenant is a liquor potassic permanganate.
4. detection method as claimed in claim 3 is characterized in that adding sulfuric acid solution when adding liquor potassic permanganate.
5. detection method as claimed in claim 1 is characterized in that described reductive agent is the oxammonium hydrochloride of 5～12 weight %, the ascorbic acid solution of 5～12 weight % or the oxalic acid solution of 5～12 weight %.
6. the kit that utilizes the described detection method of claim 1 to detect is characterized in that comprising organic solvent, Oxidizing and Reducing Agents.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102433072A (en) * | 2011-09-22 | 2012-05-02 | 晨光生物科技集团股份有限公司 | Method for removing rhodamine B in capsicum oleoresin |
| CN103149208A (en) * | 2013-03-09 | 2013-06-12 | 福州大学 | Method for rapidly detecting multiple components of fat-soluble azo dyes |
| CN106324156A (en) * | 2016-11-02 | 2017-01-11 | 百奥森（江苏）食品安全科技有限公司 | Detection method for azo dyestuffs in fodder |
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| CN102023109A (en) * | 2009-09-23 | 2011-04-20 | 帝斯曼知识产权资产管理有限公司 | Method for the extraction and detection of fat-soluble components from biological materials |
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| US20030087324A1 (en) * | 1998-02-26 | 2003-05-08 | Yukio Toyoda | Monoclonal antibody for immunologically analyzing or concentrating endocrine disruptor or its degradation product and utilization of the same |
| CN100370254C (en) * | 2005-09-05 | 2008-02-20 | 河南工业大学 | Thin-layered chromatography detection method for Sudan pigment in red pepper products |
| CN100403030C (en) * | 2006-05-10 | 2008-07-16 | 北京望尔生物技术有限公司 | ELISA kit for detecting Sudan red medicines and detection method thereof |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102433072A (en) * | 2011-09-22 | 2012-05-02 | 晨光生物科技集团股份有限公司 | Method for removing rhodamine B in capsicum oleoresin |
| CN103149208A (en) * | 2013-03-09 | 2013-06-12 | 福州大学 | Method for rapidly detecting multiple components of fat-soluble azo dyes |
| CN103149208B (en) * | 2013-03-09 | 2015-03-04 | 福州大学 | Method for rapidly detecting multiple components of fat-soluble azo dyes |
| CN106324156A (en) * | 2016-11-02 | 2017-01-11 | 百奥森（江苏）食品安全科技有限公司 | Detection method for azo dyestuffs in fodder |
| CN106324156B (en) * | 2016-11-02 | 2018-12-18 | 百奥森（江苏）食品安全科技有限公司 | The detection method of azo dyes in a kind of feed |
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