Leaflet

Kits and are test systems for enzyme immunoassay. They are mass-produced in accordance with ISO 9000, complete with the necessary reagents, and are intended for the quantitative determination of ochratoxin in cereals, feed, grain products, beer and blood serum. Methodical instructions on the use of test systems RIDASCREEN ® FAST Ochratoxin A and RIDASCREEN ® Ochratoxin A approved by the Veterinary Directorate of the Federal Agency for agriculture Ministry of Agriculture of Russia under the number MUK 5-1-14 / 1001. The systems are included in the "List of regulatory documents permitted for use in state veterinary laboratories in the diagnosis of diseases of animals, fish, bees, as well as control over the safety of raw materials of animal and vegetable origin." Test systems RIDASCREEN ® FAST Ochratoxin A correspond GOST 34108-2017"Feed, compound feed, compound feed raw materials. Determination of the content of mycotoxins by direct solid-phase competitive enzyme immunoassay".

Determination of ochratoxin A in grain, feed, grain products, beer and blood serum

Ochratoxin is a poisonous substance formed as a result of the vital activity of molds of the genus Aspergillus and Penicillium... Along with pronounced nephrotoxicity, ochratoxin has hepatotoxicity, teratogenic, carcinogenic and immunosuppressive properties. With products of plant and animal origin, ochratoxin can enter the human body. It is found not only in cereals (13% of positive samples) and animal feed, but also in pig blood (60% of positive samples) and kidneys (21% of positive samples).

Technical Regulations of the Customs Union TR CU 021/2011 "On food safety" regulate the following limiting level of ochratoxin A content: in food grains, cereals, flour - 0.005 mg / kg (5 μg / kg); in baby food, food for preschoolers and schoolchildren, food for pregnant and lactating women - not allowed (<0,0005 мг/кг).

Draft federal law № 349084-5 "Technical Regulations for Wine Products" establishes requirements for the content of ochratoxin A in wine not more than 0.002 mg / l.

The draft Federal Law "On Requirements for the Safety of Food Products and the Processes of Their Production, Storage, Transportation, Sale and Utilization" also includes the requirement for mandatory control of food grain for ochratoxin A, the content of which should not exceed 0.005 mg / kg. The current legal regulations can be found on the website compact24.com.

Until recently, chromatographic methods (high performance liquid chromatography, thin layer chromatography) were mainly used to control ochratoxin. The method of enzyme-linked immunosorbent assay (ELISA, or ELISA), which has a very high sensitivity, is much more convenient.

Specification:	RIDASCREEN ® FAST Ochratoxin A	RIDASCREEN ® Ochratoxin A 30/15
Format:	Striped plate, 48 holes (6 strips of 8 holes)	Stripped plate, 96 wells (12 strips of 8 wells)
Standards:	0/5/10/20/40 μg / l	0/50/100/300/900/1800 ng / l
Sample preparation:	Sample grinding, extraction, filtration	extraction, centrifugation / filtration (grain, feed); extraction, centrifugation, filtration, shaking, re-extraction, centrifugation, evaporation (beer / blood serum)
Time spent:
Detection limit:	0.005 mg / kg	0.001250 mg / kg (grain, feed) 0.000050 mg / kg (beer, blood serum)

Related Products:

pour one patch at a time, and in the middle part - two pads with a minimum pitch between them, which indicate the boundaries of the oncoming and off-going tape branches. The step between the overlays on the oncoming branch of the tape is determined using the dependence of the arithmetic progression, and on the running down - according to the dependence of the geometric progression. In this case, the first member of the arithmetic progression must be specified and take into account that the last gap between the linings of the oncoming belt branch is the first member of the geometric progression, and the difference and the denominator of the progressions are determined from the total gap between the linings of each belt branch.

In a drum-shoe brake, the leveling of specific loads is achieved by placing in a multi-section brake shoe on its incoming and outgoing parts of the radially movable linings connected by a balancer, i.e., the principle of conventional weights is used. This technical solution is protected by the inventor's certificate.

Stabilization of surface temperatures in friction pairs of the aforementioned braking devices is achieved due to the thermoelectric effect using thermopiles operating in thermoelectric cooler and thermoelectric generator modes in the linings of the running and running branches of the tape, as well as the running and running sections of the friction linings of the block in the main and additional servo load depending on the servo load. their friction units. At the same time, it is provided that

redistribution of thermal energy between the surfaces of the frictional units of the brakes, which leads to its quasi-stabilization. The operation of thermopiles in the above modes is theoretically justified.

Rational control of the band-shoe brake operation modes is considered provided that the level of heat load of the surface layers of the friction linings does not exceed the permissible temperature for their materials. Combined cooling (thermoelectric with a heat pipe) of brake friction pairs can be used to implement control of braking modes.

As a result of the application of this technical solution, an increase in the efficiency of the band-shoe brake of the U2-5-5 winch was achieved.

Thus, the ways of controlling the dynamic and thermal loading of the friction units of the braking devices are indicated.

LITERATURE

1. Declaration Pat. 63418A (Ukraine). A method of controlling specific loads on the incoming and outgoing branches of the brake band of the band-shoe brake / A.I. Volchenko, V.V. Dyachuk, N.A. Volchenko and others - B.I. - 2004. - No. 1. - In Ukrainian. lang.

2.A.S. 1682675 A1 USSR. Drum-shoe brake / A.I. Volchenko, V.V. Moskalev, P.A. Skorokhod and others - B.I. - 1991. -

3. Pat. 2221944 C1 Russia. Cooling systems of the brake mechanism with servo action and a method for its implementation / A.I. Volchenko, A.A. Petrik, N.A. Volchenko and others - B.I. - 2004. - No. 2.

Department of Technical Mechanics

Received November 22, 2004

DETERMINATION OF OCHRATOXIN A IN GRAPE WINE

E.N. RIKUNOVA, T.I. GUGUCHKIN

North Caucasian Zonal Research Institute of Horticulture and Viticulture

Among mycotoxins, ocher toxins occupy a special place. They are produced by some types of microscopic molds of the genera Penicillium, Aspergillus, in particular A. ochraceus, P. viridicatum. These molds are ubiquitous, mostly in warm and humid conditions, causing grapes to rot during lingering rains. Ochratoxins have a general toxic effect, affect the kidneys, liver, reduce productivity, have embryotoxic, mutagenic and carcinogenic effects.

The International Agency for Research on Cancer has classified ochratoxin as a potential carcinogen and classified it as a hazard class 2B. When foodstuffs are contaminated with ochratoxins, a person becomes ill with Balkan endemic nephropathy.

Patu-lin was previously found in wine products, and more recently, information has appeared on the content of ochratoxin A. It contaminates grain, vegetables, fruits and their products, feed, malt, beer, juices and wine.

We have developed a method for the determination of ochratoxin in grape wine by thin layer chromatography (TLC).

Thin-layer chromatography is a type of liquid chromatography in a flat sorbent layer on one side applied to a flat solid substrate. The main features of TLC are due to the movement of the eluent (solvent) along the sorbent layer due to capillary forces, which simplifies and facilitates the chromatographic process. The use of a universal sorbent - silica gel and an open layer provide ease of sample application, the possibility of simultaneous analysis of several samples, ease of monitoring the elution process

Thin layer chromatography provides for the purification and concentration of mycotoxins. To do this, use a two-dimensional or step chromatography

fiyu. The first stage of elution is purification, the separation of substances interfering with the determination, the second stage is the separation of mycotoxins.

TLC analysis of a wine sample includes the stages of sample preparation, a plate, a chromatographic chamber and eluents, as well as a Diapak S16MT concentrating cartridge; then the actual chromatography, evaporation of the eluent from the plate, identification, quantification and documentation.

The advantage of the method lies not only in its simplicity, accessibility, the possibility of using specific developing agents confirming that the substance belongs to the desired one, lower requirements for the purification of extracts, but also in the possibility of determining small amounts of ochratoxin - the detection limit is 0.1 μg / cm3.

To determine mycotoxin ochratoxin A in wine and wine materials, 10 cm3 of the sample is passed through a diapak C16MT concentrating cartridge, concentrating the sample 10 times, and finally 1 cm3 of acetonitrile is purified. The resulting extract in an amount of 5 μl and the standard are applied to TLC plates and chromatographic separation (elution) is carried out in a prepared chromatographic chamber with appropriate eluents. The most optimal solvent system for the separation of mycotoxin was in the form of isopropanol and ammonia. It is quite volatile and has a low retention coefficient.

Rf on the sorbent. Mycotoxin spots were developed by irradiation with long wavelength (365 nm) ultraviolet light. Under the influence of UV rays, the mycotoxin spots glow with a green-blue light.

The identification and quantitative determination of ochratoxin was carried out by scanning densometry on a Sorbfil densitometer with a specialized program for processing the analysis results and calculating the chromatogram parameters.

The use of a densitometer makes the TLC method quantitative, comparable in resolution to HPLC, while retaining all the advantages of TLC.

The proposed technique was tested on wine samples with preliminary addition of certain amounts of ochratoxin. The method allows you to quickly and accurately control the content of ochratoxin in wine products.

LITERATURE

1. Kretova L. G, Lunev L. I. Mycotoxins. Product contamination and analytical control. - M .: Agrprogress, 2000.

2. Materials of the OIV assembly. - Paris, 2000 .-- S. 57-59.

3. Guide to modern thin-layer chromatography / Ed. O.G. Larionova // Based on the materials of the school-seminar on thin layer chromatography. - M., 1994.

Winemaking technology laboratory

Received 09/08/04

N.T. SIYUKHOVA

Maikop State Technological University

At present, serious attention is paid to the issues of contamination of agricultural crops with toxic substances of various natures, including pesticides. Among the crops most treated with chemical means of protection against pests and diseases, the vine stands out. Due to multiple protective treatments in each growing season, vineyards have long been considered to be a kind of accumulators of environmentally hazardous chemicals.

These include organophosphate compounds characterized by an increased risk of accumulation in the treated areas and leading in terms of practical application. These drugs accumulate in the cells of the plant. Berries are most dangerous and intensively contaminated with them, which ultimately affects the quality and environmental safety of the products produced from grapes. Taking into account the high toxicity and stability of organophosphorus compounds and their metabolites, the determination of their contamination of grape products is of great scientific and practical importance.

On the production sites of the specialized farm "Fanagoria" (Temryuk district), toxicological control of red grapes was carried out (1999-2002). Samples were taken during harvesting, and the analysis of products for the content of residual amounts of organochlorine and organophosphorus insecticides in it was carried out in an accredited toxicological testing laboratory of the NKZNIISiV. The principle of choosing grape sites for sampling was based on the fact that the grape harvest, harvested from them, was used for factory processing and preparation of dry red wines in the microvincy shop of the grape processing laboratory of the NKZNIISiV.

When planning experiments to study the preservation of toxic substances in grapes, the possible influence of two factors was taken into account, which summarize the manifestation of the potential danger of the ingress of insecticides into cultivated grapes: the intake of toxic residues from the soil of plantations and from the plant itself as a result of current seasonal treatments

Ochratoxins are produced by some types of fungi Aspergillus and Penicillium... The main producers are A.ochraceus and P.viridicatum... These mushrooms are ubiquitous. Aspergillus produces ochratoxins at elevated temperature and humidity, and Penicillium already at 5 ° C. Ochratoxins are highly toxic compounds with a pronounced teratogenic effect.

Ochratoxins A, B, and C are a group of structurally similar compounds that are isocoumarins associated with L-phenylalanine peptide bond. Depending on the nature of the radicals, various types of ochratoxins are formed (Table 2.3.).

Ochratoxin A is a colorless crystalline substance, slightly soluble in water, moderately soluble in polar organic solvents (methanol, chloroform), as well as in an aqueous solution of sodium carbonate. In chemically pure form, it is unstable and very sensitive to the effects of light and air, but in ethanol solution it can remain unchanged for a long time. In UV light, it has green fluorescence.

Ochratoxin B is a crystalline substance, analogous to ochratoxin A, which does not contain a chlorine atom. It is about 50 times less toxic than ochratoxin A. It exhibits blue fluorescence under UV light.

Ochratoxin C is an amorphous substance, ethyl ester of ochratoxin A, which is close to it in toxicity, but has not been found as a natural contaminant in food and feed. In U-light, it has a pale green fluorescence.

Ochratoxins belong to toxic mycotoxins, have high toxicity for the liver, kidneys, teratogenic and immunosuppressive properties, pronounced hemolytic effect. Of the ochratoxins, the most toxic is ochratoxin A (LD 50 = 3.4 mg / kg, (day-old chickens, orally)). It is more toxic than aflatoxins. Other mycotoxins in this group are an order of magnitude less toxic.

Biochemical, molecular, cellular mechanisms of action of ochratoxins have not been studied enough. It is known that ochratoxin A inhibits protein synthesis and carbohydrate metabolism, in particular glyconogenosis, by inhibiting the activity of phenylalanine - t-RNA - a specific enzyme that plays a key role in the initial stage of protein synthesis.

Ochratoxin A is found in corn, barley, wheat, oats, barley. An important and dangerous fact is that with high contamination of feed grains and compound feeds, ochratoxin A is found in livestock products (ham, bacon, sausages). Ochratoxin B is rare. Ochratoxins also affect all fruits of horticultural crops. Apples are especially affected: up to 50% of the crop can be contaminated with mycotoxins.

It should be noted that ochratoxins are stable compounds. For example, during prolonged heating of wheat contaminated with ochratoxin A, its content decreased by only 32% (at a temperature of 250–300 ° C). Thus, the prevalence of ochratoxins in food, the toxicity and persistence of ochratoxins pose a real threat to human health.

Analysis methods

Ochratoxin A is found in oxidized foods. It dissolves easily in many organic solvents used for extraction. The most commonly used extraction is chloroform and aqueous phosphoric acid, followed by column purification and quantification using TLC.

An HPLC method has also been developed. Before HPLC analysis, a sample is prepared as follows. The crushed sample is treated with a mixture of 2 M hydrochloric acid and 0.4 M magnesium chloride solution. After homogenization, the mixture is extracted with toluene for 60 minutes. The mixture is centrifuged. The centrifugate is passed through a silica gel column and washed with a mixture of toluene and acetone (mobile phase). Ochratoxin A is eluted with a mixture of toluene and acetic acid (9: 1) and dried at 40 ° C. The residue is dissolved and filtered. The analysis is carried out using HPLC.

In addition, a number of biological assays on shrimp and bacteria have been developed, but the results obtained did not allow the use of these methods for the determination of ochratoxins.



Fungi - producers of ochratoxins - are fungi of the genera Aspergillus and Penicillium. The first reports on the toxicity of the waste products of the A. ochraceus fungus to ducks were made by Scott in 1965. In subsequent years, a large number of studies were carried out to isolate the waste products of this fungus in a pure form, decipher the chemical structure of the isolated mycotoxins, study their biological activity, conditions of toxin formation, determination methods in various biological substrates. Ochratoxins were named after the type of fungus - the first producer of this mycotoxin.

Four ochratoxins, A, B, C, and D, were isolated from the culture of the fungus A. ochraceus. Ochratoxin A has the greatest sanitary and toxicological significance. It dissolves well in acetone, benzene, acetonitrile, chloroform, and alcohols. When interacting with iron chloride, it forms a red-colored complex, strong complexes with alkalis.

The main ochratoxin-producing fungi are A. ochraceus and P. veridicatum. In countries with a warm climate, feed is most often contaminated with ochratoxin from the fungus A. ochraceus, in countries with a temperate climate - by the fungus P. veridicatum. The optimum substrate temperature, at which the greatest toxin formation occurs during cultivation, is 28 ° C for A. ochraceus fungi and 20 ° C for P. veridicatum fungi.

According to N.V. Volkov (1980), out of 316 fungal isolates isolated in five pig breeding complexes in Ukraine, 106 strains (33.5%) were attributed to A. ochraceus fungi. Of this amount, four isolates formed ochratoxin A.

Mushrooms - producers of ochratoxin A - are quite often found in feed in Russia, however, very few cases of animal disease have been registered. This is due to the lack of a sensitive and specific method for the determination of ochratoxin A in feed. Recently, ochratoxicosis A was established in a number of pig farms in the Kursk and Belgorod regions (G.P. Kononenko et al., 1999).

Ochratoxins, like other mycotoxins, are destroyed relatively quickly in the body of animals. However, there are reports that mycotoxin, depending on the dose, can be retained in the muscle tissue and in the muscles of pigs for up to 2 weeks, in the liver up to 3 weeks, and in the kidneys up to 4 weeks. Therefore, it is necessary to establish the deadline for slaughtering animals after the last case of mycotoxin intake into the body at 4 weeks. It is also possible that mycotoxin will be excreted in milk if it enters the body with feed in relatively large quantities.

Ochratoxin A belongs to highly toxic compounds - LD5o for laboratory animals with a single oral administration of 20-28 mg / kg of animal weight, for chickens 7 days old 11-15 mt / kg. Mycotoxin has a pronounced cumulation. Pigs are most sensitive to it, especially young ones, then birds.

With the content of mycotoxin in feed 0.2-0.4 mg / kg of feed in pigs, even with prolonged feeding, there were no clinical signs of intoxication, but a decrease in the weight gain of animals and polyuria was observed. For chickens, the subtoxic dose is 0.6-0.8 mg / kg of feed, the toxic dose is 1.5-2.0 mg / kg. With an increase in the content of ochratoxin A in feed up to 5 mg / kg, signs of poisoning and death of individual animals were expressed in pigs and chickens.

Toxicodynamics. Not clarified enough. Ochratoxin A predominantly acts on the kidneys, so in Denmark, where this mycotoxicosis in pigs was first reported, it was called “porcine mycotoxic nephropathy”. It was found that ochratoxins entering the blood relatively quickly bind to its proteins. Perhaps, getting into the acidic environment of the kidneys, the mycotoxin is released and manifests its nephropathic effect.

Clinic. Chronic ochratoxicosis, which often occurs in practical conditions, is very weak. In animals, thirst increases, polyuria is expressed, and a decrease in weight gain. In the blood, in some cases, leukocytosis, an increase in the number of lymphocytes, and a decrease in basophils are noted. In chickens, general depression, ruffled feathers, and a decrease in productivity are observed. According to some authors, yellow spots appear on the eggshell.

Treatment. There are no specific treatment methods. First of all, feeds containing ochratoxin A or those affected by mold should be excluded from the diet. Effectively introducing various adsorbents into feed, such as zeolites, glauconites, etc.

Pathological changes. Kidney damage is most typical in ochratoxicosis. As a rule, they are enlarged, the capsule is connected in places with the cortical layer. On the cut, the cortical layer is pale; under the capsule there may be cysts 1 - 2 mm in size. Histological examination reveals necrosis of cells of the proximal tubules, proliferation of connective tissue in the cortex.

An increased fluid content is sometimes found in the abdominal cavity. In some cases, the liver is enlarged and its cells are necrotic.

Vetsanexpertiza. In the case of forced slaughter of animals in the case of ochratoxicosis, organs and tissues, and especially the kidneys, must be examined for the presence of mycotoxin. The MRL for ochratoxin in meat and offal has not been established. If residues of mycotoxin are found, the carcass and internal organs are disposed of.

Concentration of ochratoxin A in the sample, mg / kg

Limits of relative error (accuracy index) (± d),%, R = 0,95

Repeatability standard deviation (s r), %

Repeatability limit ( r), %

Completeness of extraction of substances,%

4.2... Auxiliary equipment

Apparatus for shaking samples, type AVU-6S or similar
Rotary evaporator IR-1M with trap or similar
Laboratory drying electrical cabinet with an error of maintaining the temperature of ± 2.5 in the range from 50 to 350 ° С
Household refrigerator
Laboratory electric mill EM-3A or similar pH meter	TU 46-22-236-79
Magnetic stirrer type MM 5 with stirring bar	TU 25-11.834-80
Flat-bottomed conical flasks, 250 cm3 with NSh 29, type KnKSh 250-29 / 32	GOST 10394-74
Screwed glass vials made of dark glass (vile), volume 7 cm3
Volumetric flasks, with a capacity of 100, 500, 1000 cm3 type 2-100-2,2-500-2
Funnels laboratory
Pear-shaped flasks, 10 cm3 with NSh 14.5, type GrKSh-10-14 / 23	GOST 10394-72

4.3... Reagents and materials

... Preparing to take measurements

5.1... Preparation of standard solutions of ochratoxin A

To prepare a standard storage solution (concentration of ochratoxin A - 10 ng / μl), a sample of crystalline ochratoxin A weighing 5 mg is placed in a volumetric flask with a volume of 500 cm3, 50 cm3 of a mixture of toluene-acetic acid (98: 2% vol.) Is added, thoroughly mixed until complete dissolution of the substance and bring the same mixture of solvents to the mark. To establish the exact concentration of the storage solution, measure its optical density at a wavelength of 333 nm (D333). The concentration of the solution is calculated by the formula:

For the preparation of working solutions of ochratoxin A with a concentration of 0.005; 0.05 and 0.1 ng / μl, respectively, 50, 500 and 1000 μl of a solution with a concentration of 0.5 ng / μl are taken, evaporated to dryness and dissolved in 5 cm3 of the mobile phase.

The storage solution of ochratoxin A is kept in a glass container with a ground stopper in a cool dark place (at a temperature of about 0 ° C) for up to one year and is used to prepare working standard solutions. The working standard solutions are stored in dark glass vials in a cool dark place (at a temperature of about 0 ° C) for 1 month.

Before using the working standard solutions, bring them to room temperature and only then open the caps.

5.2... Preparation of phosphate buffer solution, pH = 7.4

A weighed portion of sodium phosphate disubstituted 12-water weighing 1.15 g, a weighted portion of sodium monosubstituted 2-water weighing 0.124 g and a weighed portion of sodium chloride weighing 1.74 g are transferred into a volumetric flask with a capacity of 100 cm3, 10-20 cm3 of distilled water is added. Stir and bring the volume of the solution in the flask to the mark. Shelf life is 1 month in the refrigerator.

5.3... Preparation of solvent mixtures

Toluene-acetic acid (98:2 % about.).

In a volumetric flask of 1000 cm3, add 20 cm3 of acetic acid and, stirring, bring toluene to the mark. Shelf life - 1 month in a cool, dark place.

Acetonitrile-water (60:40 % about.).

In a volumetric flask of 1000 cm3, add 600 cm3 of acetonitrile and, stirring, add water to the mark. Shelf life - 1 month in a cool, dark place.

Acetonitrile-water (60:40 % about.; NS = 3 ,0 ).

In a volumetric flask of 1000 cm3, add 600 cm3 of acetonitrile and, stirring, bring up to the mark with bidistilled water. By introducing phosphoric acid, the pH of the mixture is adjusted to a value of 3.0. Shelf life - 1 month in a cool, dark place.

Methanol-acetic acid (98:2 % about.).

In a volumetric flask of 1000 cm3, add 20 cm3 of acetic acid and, stirring, bring up to the mark with methanol. Shelf life - 1 month in a cool dark place.

... Sampling and preparation of samples for analysis

6.1... Sample selection

To take into account the specifics of sampling of certain types of products, one should be guided by the current regulatory and technical documentation:

"Corn. Acceptance rules and sampling methods "GOST 13586.3-83;

“Groats. Acceptance rules and sampling methods "GOST 26312.1-84;

“Flour and bran. Acceptance and sampling methods "GOST 27668-88;

“Canned food products. Sampling and preparing them for testing "GOST 8756.0-70.

Samples for analysis, representative of the concentration of mycotoxins for the entire batch, should be taken from a preliminary homogenized average (initial) sample weighing 2 kg.

6.2... Sample preparation for analysis

The selected samples are ground for 1 - 2 minutes in a laboratory mill. In this case, two parallel samples are used.

6.2.1... Extraction

A weighed portion of 25 g of the crushed sample is placed in a flat-bottomed conical flask of 250 cm, 100 cm3 of an acetonitrile-water mixture (60:40% vol.) Are added. Extract on a shaker for 30 min. The resulting mixture is filtered through a blue ribbon pleated paper filter. Take 10 cm3 of the filtrate and add 90 cm of phosphate buffer solution, pH = 7.4.

6.2.2... Extract purification

On the immunoaffinity column, 100 ml of the resulting mixture is applied at a rate of 1 - 2 drops per second, washed with 20 cm3 of phosphate buffer solution, pH = 7.4. Ochratoxin A was eluted with 3 cm3 of a methanol-acetic acid mixture (98: 2% by volume).

... Taking measurements

7.1... Test sample preparation

The eluate is evaporated to dryness. The dry residue is dissolved in 400 μl of the mobile phase (solution A).

7.2... Chromatography conditions

HPLC conditions: mobile phase - acetonitrile-water (60:40 vol.%; PH = 3.0); the speed of the mobile phase is 1.5 cm3 / min.

The fluorometric detector is set to the excitation wavelength of 333 nm, and an emission filter with a passband of 466 nm is installed on the emission line.

To analyze samples, 50 μl of a test sample (solution A) is injected into the chromatograph injector using a microsyringe. If there is a peak coinciding in retention time with ochratoxin A, calculate the mass of ochratoxin A in the injection using the calibration graph.

... Processing of measurement results

8.1... Building a graded dependency

To build a calibration graph, chromatographic analysis of a series of working standard solutions is carried out. Using a microsyringe, 50 μl of the working standard solution with a concentration of 0.005 ng / μl, which corresponds to 0.25 ng of ochratoxin A. corresponds to 2.5 and 5.0 ng of ochratoxin A in the injection. Under these conditions, the retention times for ochratoxin A are in the range of 4 to 5 minutes. Based on the data obtained, a calibration graph is plotted (the dependence of the area of ​​the chromatographic peak on the mass of ochratoxin A in the injection).

The analysis result is presented in the form (with the probability R = 0,95):

D is the limit of the absolute error:

d is the limit of the relative error of the method (accuracy indicator),% (Table 1).

* 0.0001 mg / kg is the detection limit.

... Requirements for the qualification of the performer

To perform the analysis of ochratoxin A in grain and grain products, persons with a special higher education or secondary specialized education, possessing the technique of HPLC analysis, who have undergone appropriate training and have experience in a chemical laboratory, are allowed.

... Measurement conditions

Ambient temperature from 15 to 25 ° С.

Relative air humidity not more than 80% at 25 ° С.

Atmospheric pressure 730 - 760 mm Hg

Power supply voltage: 210 - 220 V. AC frequency: 45 - 50 Hz.