GOST 26669-85

INTERSTATE STANDARD

FOOD AND TASTING PRODUCTS

SAMPLE PREPARATION FOR MICROBIOLOGICAL
ANALYZES

Introduction date 01.07.86

This International Standard applies to food and flavor products and specifies the preparation of samples for microbiological analysis.

The terms used in the standard and their explanations are given in the appendix.

1. EQUIPMENT, REAGENTS AND MATERIALS

membrane filtration device; gas or alcohol burner according to GOST 25336;

metal funnels; punch;

bottle opener; can opener;

scissors, scalpel, tweezers according to GOST 21241, spatula, spoon;

stencils (template); test tubes according to GOST 25336;

* On the territory of the Russian Federation, GOST R 51652-2000 is valid.

70%; plastic bags; detergent;

peptone for bacteriological purposes according to GOST 13805.

Tools and surfaces of devices in direct contact with the product must be sterilized by one of the methods specified in GOST 26668.

1.2. Preparation of peptone-salt solution

Peptone-salt solution is prepared as follows: 8.5 g of sodium chloride and 1.0 g of peptone are dissolved in 1 dm 3 of distilled water with slow heating. The resulting solution, if necessary, is filtered through a paper filter, adjusted to pH 7.0 ± 0.1, poured into flasks, test tubes or other vessels, sealed and sterilized at a temperature of (121 ± 1) °C for 30 minutes.

The solution is stored in a dark place at a temperature of (4 ± 2) °C for no more than 30 days under conditions that exclude moisture evaporation.

The temperature of the peptone-salt solution should correspond to the temperature of the analyzed product.

1.3. Preparation of peptone water

Peptone water is prepared similarly to a peptone-brine solution without the addition of sodium chloride.

2. PREPARATION OF SAMPLES FOR ANALYSIS

2.1. The sample packaging is inspected and matched with the inscription on the lithographic print or on the label indicated in the accompanying document.

2.3. Microbiological analysis of normal in appearance samples of the product is carried out in a box under aseptic conditions. The packaging of a sample of a suspicious-looking or spoiled product is opened in a separate room.

Boxing preparation is set out in the appendix.

2.2, 2.3. (Changed edition, Rev. No. 1).

2.4. Samples with a frozen product are thawed at a temperature of (4 ± 2) °C before weighing. The sample is taken immediately after thawing, but no later than 18 hours after the start of thawing.

It is allowed to defrost a sample of the product at a temperature of 18 - 20 ° C for 1 hour.

Samples of the product of homogeneous consistency may be thawed in a thermostat at 35 °C, provided that complete defrosting is achieved in no more than 15 minutes.

2.5. Opening the packaging with a product sample

2.5.1. Immediately before opening the package with a sample of the product in consumer containers, free-flowing or having a liquid phase, mix by 10-fold inversion of the container from the bottom to the lid or in a circular motion.

2.6.1. From each sample of the product, depending on the indicators to be determined, one or several portions are taken for the preparation of dilutions and/or inoculation into nutrient media.

2.6.2. The mass (volume) of the sample intended for sowing into nutrient media and/or for preparing its dilutions must be established in the regulatory and technical documentation for a specific type of product or analysis methods.

2.6.3. A sample for sowing is taken by weight or volumetric method immediately after opening the product sample. The opening is carried out under conditions that exclude contamination of the product by microorganisms, in the immediate vicinity of the burner flame with sterile instruments.

2.6.4. A sample of the product is selected so that it contains all its components and in the same ratio as in the analyzed sample.

2.6.5. To prepare dilutions of a sample of the product, a peptone-salt solution is used.

It is allowed to prepare the initial dilutions of products with a mass fraction of NaCl more than 5% using peptone water, the initial dilutions of meat, fish and dairy products - using saline.

The mass (volume) of the sample of the product intended for the preparation of the initial dilution or homogenate must be at least (10 ± 0.1) g/cm 3 .

The ratio between the mass (volume) of the sample of the product and the volume of the peptone-salt solution for the initial and subsequent dilutions is:

1:9 - for 10-fold dilution (for products containing a large amount of fat without surfactants 1:10);

1:5 - for 6-fold dilution;

1:3 - for 4-fold dilution;

1:1 - for 2-fold dilution.

If it is necessary to dilute a sample of products containing a large amount of fat, it is allowed to use surfactants (sodium bicarbonate, etc.) that do not have antimicrobial activity.

To prepare a dilution of a sample of products with high osmotic pressure, it is allowed to use peptone or distilled water.

(Revised edition, Rev. No. 1).

2.6.6. The initial dilution of a sample of the product is prepared under aseptic conditions in one of the following ways:

dissolution of products;

dilution of products having a liquid phase;

suspension of powders, pasty products and the surface of microbially contaminated product pieces;

homogenization of solid products.

The portion of the product remaining on the surface of the pipette is allowed to drain to the tip of the pipette. The resulting drop is removed by touching the inner wall of the dish or consumer container above the surface of the product.

Viscous products are removed from the surface of the pipette with a sterile swab.

A portion of the product is transferred to a container with peptone-brine solution to prepare the initial dilution so that the pipette does not touch the surface of the peptone-brine solution. With another sterile pipette, thoroughly mix the product with the peptone-salt solution by tenfold filling and ejection of the mixture.

When working with viscous products, it is advisable to place several glass balls in a container for faster mixing with a peptone-salt solution.

2.6.8. The liquid product saturated with carbon dioxide (CO 2 ) is transferred to a sterile, cotton-stopped conical flask or other container and heated with frequent stirring in a circular motion in a water bath at a temperature of 30 to 37 ° C until no more gas bubbles will come out.

A sample of the product is taken and processed according to p.

A sample from samples of water-insoluble solid products is homogenized in the cases specified in the regulatory and technical documentation. When homogenizing the product, the total number of revolutions of the homogenizer should be 15 - 20 thousand. The number of revolutions of the homogenizer should not be less than 8000 and more than 45000 revolutions per minute.

If a heterogeneous mass is obtained during homogenization of the product, then it is settled for 15 minutes and the supernatant liquid is used for inoculation and (or) preparation of dilutions.

It is allowed to homogenize an unsterilized product by grinding it until a homogeneous consistency is achieved in a sterile mortar under aseptic conditions.

(Revised edition, Rev. No. 1).

2.6.12. A sample of pasty products is taken after thorough mixing with a spoon or glass rod and then processed according to p.

2.6.13. A sample of liquid fat samples is taken with a warm pipette heated by flambing. After filling the pipette with the product, the remains of it are removed from the surface of the pipette with a sterile swab.

The product from the pipette is introduced into a glass-stoppered dish and diluted with the required amount of peptone-salt solution heated to 40-45 °C; when psychrophilic microorganisms are detected, the temperature should not exceed 37 °C. Fat residues adhering to the pipette are rinsed with a peptone-salt solution, which is sucked up several times and released from the pipette.

2.6.14. A sample of solid fats is taken after cutting the product with a knife or wire into several parts. If necessary, remove the top layer.

A sample of the product is taken from the surface of the cuts from different places with a scalpel and transferred to a weighed dish with a lid.

A certain mass of sample is transferred to a wide-mouthed dish with ground glass stopper. The remains of fat adhering to the walls of the dish are rinsed in the same dish with a certain amount of peptone-salt solution heated to 40 - 45 ° C, which is added to the dish in the amount necessary to obtain the initial dilution.

From solid fats, a sample can be selected by volume. Fats are melted in a wide-necked dish in a water bath at a temperature not exceeding 45 ° C; when psychrophilic microorganisms are detected, the temperature should not exceed 37 °C.

After mixing the melted fat, it is transferred with a warm pipette to a wide-necked dish with a ground glass lid containing the required amount of peptone-salt solution to prepare the initial dilution. Peptone-salt solution is preheated to 40 - 45 °C; when detecting psychrophilic microorganisms up to 37 °C.

2.6.15 Samples from samples of whipped products or a mushy consistency containing a large amount of fat, after mixing with a glass rod, are taken with a spoon into a weighed dish and a peptone-salt solution, heated to 40 - 45 ° C, is added in the amount necessary to prepare the initial dilution.

2.6.16 Determination of microbial contamination of the surface of product samples is carried out by flushing with cotton swabs.

A sterile cotton swab is moistened with a peptone-salt solution and wiped with it in different places on the surface of various pieces of the analyzed product with a total area of ​​100 cm 2 .

The surface area to be analyzed is measured using sterile templates with holes of the appropriate size.

The swab is placed in a test tube containing 10 cm 3 peptone-salt solution. The contents of the tube are thoroughly mixed with a pipette. The resulting suspension is considered the original dilution.

(Revised edition, Rev. No. 1).

2.7. Preparation of tenfold dilutions

2.7.1. The first tenfold dilution of the sample is the initial one, the initial dilution is prepared in accordance with p. Subsequent dilutions are obtained from it.

2.7.2. The subsequent second dilution is prepared from one part of the original dilution and nine parts of peptone-saline solution by mixing in a test tube.

If a pipette was used to mix the initial dilution, then with the same pipette add 1 cm 3 of the initial dilution to 9 cm 3 of the peptone-salt solution, without touching the surface of the solution with the pipette. The dilution is mixed with another pipette by sucking and blowing out the contents of the tube ten times.

2.7.3. The third and subsequent dilutions are prepared in a similar way.

2.7.4. The interval between the preparation of weighed portions of the product, their dilutions and sowing in nutrient media should not exceed 30 minutes.

ATTACHMENT 1
Reference

TERM USED IN THE STANDARD AND EXPLANATIONS TO IT

(Revised edition, Rev. No. 1).

APPENDIX 2
Reference

CANNED DEFECTS

Defects in a canned product are:

visible to the naked eye signs of the development of microorganisms: fermentation, mold, mucus, etc .;

sediment at the bottom of the can or at the border of the surface of the product with the container (“ring”);

turbidity of the liquid phase;

coagulation;

souring;

extraneous, not characteristic of the product, smell and (or) taste;

color change.

Defects in the appearance of containers with products packaged in it are considered:

signs of leaks visible to the naked eye: holes, through cracks, smudges or traces of the product flowing out of the can;

cans with vibrating ends;

incorrectly designed seam of cans (tongues, teeth, undercut, false seam, rolled seam);

rust, after the removal of which shells remain;

deformation of the body, ends or longitudinal seam of cans in the form of sharp edges and "birds";

skewed lids on glass jars, undercutting the corrugations of lids along a rolled field, a protruding rubber ring (“loop”);

cracks or chipped glass at the seam, incomplete fit of the lids relative to the neck of the can;

deformation (indentation) of the lids of glass jars, which caused a violation of the seaming seam;

convex elastic membrane (button) on the cover.

APPENDIX 3
Reference

BOXING PREPARATION

Canned food is opened in a box-room specially adapted for microbiological analysis. In the box, there should be no surfaces inaccessible for wet disinfection and air movement created by drafts should be excluded. Walls, floors and ceilings must be lined with material or painted with disinfectants resistant to wet treatment. For air sterilization, the box is equipped with ultraviolet lamps at the rate of 1.5 - 2.5 W per 1 m 3.

Only the microbiologist conducting the analysis and, if necessary, an assistant should be in the box.

The box must have a table and a stool. There should be no extra items, except for those required for the analysis of canned food.

On the table should be:

spirit stove or gas burner;

a jar with a ground stopper with alcohol;

a jar closed with a lid with pre-prepared dense sterile cotton swabs measuring 3 ´ 3 cm or cotton rings;

jars with a disinfectant solution (layer height 3 cm) for placing pipettes or tubes used after analysis;

a small metal or enameled tray on which the analyzed cans are placed;

sterile pipettes or tubes with which the sample is taken.

Auxiliary equipment should be stored in the drawer of the table: tweezers and a punch. The punch should have the shape of a spear with a cross section in the form of a rhombus with diagonals of 1 ´ 1.5 cm or with a cross section in the form of an isosceles triangle.

When opening a large number of cans, a punch mounted on a tripod is used. In this case, the opening is performed by pressing the punch on the lid of the jar with a lever.

Before opening the jar, the punch is flambéed in the flame of a swab.

The box is washed and disinfected immediately before the analysis (not earlier than 24 hours before the start) and after its completion. Disinfection is carried out by wiping all surfaces with chlorine or other disinfectants according to the instructions corresponding to each preparation. 45 minutes before the start of work in the box, bactericidal lamps are turned on for (30 ± 5) minutes.

At present, laminar flow cabinets (protective cabins of ultrapure air) are used for microbiological analysis. Laminar boxes are produced by the Uzhgorod plant of medical equipment "Laminar", boxes of the BPV 1200 brand are produced in Hungary, boxes of the TVG-S II 1.14.1 brand are produced by Babcock - BSH (Germany).

APPENDICES 2, 3. (Introduced additionally, Amendment No. 1).

INFORMATION DATA

1. DEVELOPED AND INTRODUCED by the State Agro-Industrial Committee of the USSR

2. APPROVED AND INTRODUCED BY Decree of the USSR State Committee for Standards No. 3810 dated 04.12.85

3. The standard fully complies with ST SEV 3014-81

4. International standards are introduced into the standard: ISO 6887-83 (E) and ISO 7218-85

5. REPLACE GOST 10444.0-75

6. REFERENCE REGULATIONS AND TECHNICAL DOCUMENTS

Conducted purposeful sanitary and bacteriological control of food products, washings from environmental objects, water allows you to effectively carry out current sanitary and epidemiological surveillance, gives an objective assessment of compliance with the regime. Helps in finding out the ways of transmission of the infectious beginning. Errors in sampling can lead to incorrect hygienic assessment of the studied samples using the most sensitive and accurate research methods, and as a result, to an inadequate assessment of the object itself.

Therefore, one of the basic principles of microbiological research is the correct sampling, strictly adhering to the rules for sampling and their quantitative ratio.

The main documents for sampling food products for microbiological studies are:

GOST R 54004-2010 Food products. Sampling methods for microbiological tests"
GOST R 53430-2009 “Milk and milk products. Methods of microbiological analysis»
GOST R ISO 707 - 2010 “Milk and dairy products. Sampling Guide"

Features of sampling food products according to GOST R 54004-2010:

1. Before sampling, on the basis of visual inspection, packaging units or products are divided into 3 groups, while sampling is carried out for each group separately:

Normal in appearance (no signs of microbial spoilage)
- suspicious (with signs of deviations that can occur both due to microbial spoilage and due to chemical or biochemical reactions in the product)
- spoiled products during inspection of which obvious defects of the product were found (bombing, molding, sliming, etc.). Products with an expired shelf life are not selected for research.

2. Samples are taken using sterile instruments into sterile dishes, the throat of which is burned in a burner flame (sterile jars or sterile bags, sterile plastic containers).

If routine sampling is carried out and one sample is formed, then sampling for microbiological analysis should precede sampling for organoleptic and physico-chemical examination, observing the rules of asepsis, excluding contamination at the time of sampling.

3. The volume (mass) of the sample is determined in accordance with the NTD for this type of product. The number of packaging units is established by the current standards, OST, TU, etc. for the respective products.

4. If the mass of the sample is equal to the mass of the product in consumer packaging, then use the entire package. If the mass of the sample is greater than one package, then several packages are taken, otherwise (there is no package), the sample is taken by taking incremental samples from different places.

5. If the mass (volume) of the product is not established by the NTD, at least 1 sample is taken from products in consumer packaging and up to 1000 g (cm3) from products in transport containers (lumpy, liquid, pasty, loose and mixed consistency). When sampling from lumpy products weighing more than 1000 g, use one of the following methods:

• cut off or cut out a part of the product with a knife or other tool, while for rectangular products the cut is made perpendicular to the longitudinal axis, and for spherical products it is wedge-shaped;
• the product is cut in several places with a knife, and then the required number of pieces are taken from the cut surface and from the depth with a scalpel, which are transferred with tweezers into a wide-mouthed container;
• cut off the surface layer of the product to a thickness of 0.5 - 1 cm and, using a probe or a special tool, squeeze the product into a wide-mouthed container.

6. Samples of frozen products are placed in isothermal containers or with refrigerants. The temperature of such samples during transportation should not exceed minus 150C. Samples of perishable products are transported at 50C in refrigerated bags for no more than 6 hours. In other cases, the NTD for each type of product is guided.

7. Sampling of milk and dairy products is carried out in accordance with: GOST 26809-86 “Milk and dairy products. Acceptance rules, sampling methods and preparation of samples for analysis. If the products are presented in consumer packaging, 1 unit of consumer packaging is selected. When compiling a combined sample, for example, cottage cheese: 3 point samples are taken from each unit of transport packaging: 1 from the center, 2 others at a distance of 5 cm from the side wall. The selected mass is transferred into a sterile dish, making up a combined sample weighing 500 g. When determining the amount of bifidobacteria in fermented milk products, 3 consumer packaging units are selected by random sampling. Microbiological studies should be started no more than 4 hours after sampling, if the samples were transported at a temperature not higher than 6 0 С, and ice cream samples - not higher than 2 0 С.

8. Sampling of fish products - according to GOST 31339-2006 "Fish, non-fish objects and products from them"

9. Meat products in accordance with GOST R 51447-99 "Meat and meat products"

10. Poultry meat, poultry by-products and semi-finished products according to GOST R 50396.0-92 "Poultry meat, poultry by-products and semi-finished products".

9. When sampling products at catering establishments, one should be guided by MU No. 2657 “On sanitary and bacteriological control at catering establishments and food trade”.

If the sample of the dish is taken in the dispenser, then the entire portion is transferred from the plate to the jar; if a sample is taken in the kitchen from a large mass of the product (from a pan, from a large piece of meat), then a sample weighing about 200 g is taken (liquid dishes - after thorough mixing; dense ones - from different places in the depth of the piece). Mineral, non-alcoholic drinks and beer are selected in the amount of 1 bottle of factory packaging or 200 ml of a drink made at the enterprise.

When sampling a product of complex consistency, it should include all components in the same ratio as in the original product. If necessary, each component is selected separately.

Bulk products before taking samples are thoroughly mixed, or the sample is made up of point samples.

10. All samples are supplied with labels, on which, in addition to the sample number, product name, the date and hour of sampling, as well as the date and hour of production, and the product's shelf life must be indicated. Samples are sealed or sealed.

11. In the process of sampling, a sampling protocol is drawn up and sent for research, indicating the reason for sampling (scheduled, unscheduled, epid. situation research, etc.) and the purpose of the compliance test is indicated:

Uniform sanitary-epidemic and hygienic requirements for goods approved. 05/28/2010 No. 299

TR TS 02\2011 "on food safety"

SanPiN 2.3.2.1078-01 "Hygienic requirements for the safety and nutritional value of food products"

Federal law Technical regulations for milk and dairy products No. 88-FZ dated June 12, 2008

Federal law Technical regulations for oil and fat products

Federal law Technical regulations for juice products from fruits and vegetables No. 178-FZ dated October 27, 2008

Instructions on the procedure for investigating, recording and conducting laboratory tests in institutions of the sanitary and epidemiological service in case of food poisoning No. 1135-73 g

Uniform sanitary and epidemiological and hygienic requirements for goods subject to sanitary and epidemiological supervision (control) have been developed in order to implement the provisions of the Customs Union Agreement on Sanitary Measures dated December 11, 2009.

Washouts.

According to MU No. 2657 of December 31, 1982 "On sanitary and bacteriological control at public catering and food trade enterprises."

In the practice of the current sanitary supervision of the food units of children's, preschool and adolescent institutions, as well as buffets, the flush method is widely used to control the effectiveness of the sanitization of inventory, equipment, utensils, sanitary clothing and staff hands. The flushing method makes it possible to objectively assess the sanitary content of the examined institutions.

Particular attention during flushing is paid to the control of equipment and apparatus that are used during the technological process of preparing products that are not further heat treated (cold shop).

Bacteriological control by the method of washings from the surfaces of inventory, equipment, hands and sanitary clothing of personnel can pursue two goals:

a) to establish the effectiveness of sanitization, for this, washings from inventory, equipment, hands and sanitary clothing of personnel are carried out before starting work, or, if this is not possible, during breaks, after hands and equipment have been sanitized, i.e. swabs are produced from clean objects.

b) determine the role of equipment and personnel hands in bacterial contamination of a product or a finished dish during the production process, paying special attention to the production of products and ready meals that have undergone heat treatment or are eaten without pre-treatment (some vegetables, gastronomic products, salads, vinaigrettes, etc.). To solve the problem, at the same time as taking swabs, repeated samples of food products are taken (washouts are taken from untreated hands and surfaces).

Flushing is carried out from the surface with a moistened sterile cotton swab, mounted on a glass or metal holder, mounted in a cotton-gauze stopper of the test tube. The test tube contains a sterile medium. Immediately before taking a flush, the swab is moistened by lowering the swab into the liquid. When taking swabs, the following recommendations should be considered:

• Of the equipment, you should pay attention to cutting boards, meat grinders, production tables for finished products.
• Washes from hands, sanitary clothes, towels are taken from workers who deal with products that are not further heat treated.
• Washouts from large equipment are taken from a surface of 100 sq.cm. A stencil 25 sq. cm is applied in 4 different places on the surface of the controlled object.

When taking swabs from small objects, the entire surface is washed off. Washouts are taken:

• one tampon with 3 items of the same name (plates, spoons, etc.). Glasses are wiped from the side of the inner surface and the upper outer edge 2 cm down.
• When taking swabs from the hands, wipe the palmar surfaces of both hands with a swab, swiping at least 5 times on each palm and fingers, then wipe the interdigital spaces, nails and subungual spaces.
• When taking swabs from sanitary clothing, wipe 4 areas of 25 sq. cm - the lower part of each sleeve and 2 areas from the upper and middle parts of the front floor. Towels - 4 pads of 25 sq.cm.

When taking swabs, record the sample number in order, the place where the swab was taken. An act of taking swabs is drawn up in 2 copies.

Delivery time - no more than 2 hours. With an increase in time, delivery in thermal containers.

Water sampling for microbiological research

Selection, conservation, storage and transportation of water samples is carried out:

According to GOST R 53415-2009 “Water. Sampling for microbiological analysis”;

According to GOST 31942-2012 “Water. Sampling for microbiological analysis”;

According to GOST R 51592-2000 “Water. General requirements for sampling”, all water is taken, which is delivered to the microbiological laboratory for analysis;

According to GOST R 51593-2000 “Drinking water. Sampling”, applies only to tap water from centralized water supply systems;

In accordance with the requirements of standards and other normative documents for determination methods;

Specific indicators and intended for certain types of waters.

For example, water sampling of centralized drinking water supply systems is carried out according to three regulatory documents:

- GOST R 51593-2000 “Drinking water. Sample selection",
- MUK 4.2.1018-01 "Sanitary and microbiological analysis of drinking water".

The conditions under which water sampling for microbiological studies is carried out should be close to aseptic, i.e. do not forget to burn the tap, drain the water from this tap for 10 minutes and only then draw water into a sterile container. The container is opened immediately before sampling by removing the stopper together with a sterile cap. During sampling, the cork and the edges of the container should not touch anything. Currently, disposable water sampling bags are used with and without sodium thiosulfate tablet. Rinsing dishes is prohibited. The sample is taken directly from the tap without rubber hoses, water distribution grids and other nozzles. If a constant outflow of water occurs through the sampling cock, sampling is carried out without preliminary firing, without changing the water pressure and the existing structure (in the presence of silicone or rubber hoses).

Water samples from sources of centralized and non-centralized water supply are taken in accordance with GOST R 51592-2000 “Water. General requirements for sampling”.

Water from the swimming pool bowl is taken according to the following documents:

GOST R 51592-2000 “Water. General requirements for sampling”,
- SanPiN 2.1.2.1188-03 “Swimming pools. Hygienic requirements for the device, operation and water quality. Quality control".

Water sampling for analysis is carried out at least at 2 points: a surface layer 0.5–1.0 cm thick and at a depth of 25–30 cm from the surface of the water mirror. Control of the quality of water in the bath of the swimming pool according to the main microbiological indicators should be carried out 2 times a month.

Samples should be analyzed in the laboratory as soon as possible after collection. In the absence of cooling, the analysis is performed no later than 2 hours after sampling, and when cooled to 4-10˚ C, the sample storage time increases to 6 hours. Therefore, it is necessary to transport samples to the laboratory in thermal containers (do not allow freezing, since more than 99% of bacteria die when the sample is frozen).

Since the number of microorganisms in the sample can be reduced by half in less than 20 minutes due to the action of residual amounts of disinfectants (chlorine - in a matter of seconds), they are used in a container with sodium thiosulfate (at the rate of 10 mg per 500 ml of water) to neutralize chlorinated and brominated water.

The sample volume is set depending on the number of indicators to be determined and the type of analysis in accordance with the normative documents for the method for determining indicators. For example, when analyzing tap water and water from a well, 350 ml of water is enough for indicator microorganisms, and 1350 ml for indicator microorganisms and pathogenic flora, the volume of swimming pool water samples is 500 ml and 1500 ml, respectively.

SanPiN 2.1.4.1116-02 “Drinking water. Hygienic requirements for the quality of water packaged in containers. Quality control”, MU 2.1.4.1184-03 “Guidelines for the implementation and application of sanitary and epidemiological rules and regulations SanPiN 2.1.4.1116-02 “Drinking water. Hygienic requirements for the quality of water packaged in containers. Quality control"

Drinking water packaged in containers is taken in a volume of 2.5 liters, because only for the determination of Pseudomonas aeruginosa and coliphages, 1.0 l of water is required.

Soil sampling is carried out in accordance with GOST 17.4.3.01-83 "General requirements for soil sampling", GOST 17.4.4.02-84 "Methods of sampling and sample preparation for chemical, bacteriological, helmitological analysis".

A trial plot is a part of the study area characterized by similar conditions (relief, uniformity of soil structure and vegetation cover, and the nature of economic use).

The test site should be located in a typical location for the study area. On an area of ​​100 m2, one test site with a size of 25 m is laid.

Point sample - material taken from one place of a horizon or one layer of a soil profile, typical for a given horizon or layer.

Point samples are taken on a trial plot from one or more layers or horizons using the envelope method. They dig a hole 0.3m x 0.3m and a depth of 0.2m. The surface of one of the walls of the pit is cleaned with a sterile knife. Then a soil sample is cut out of this wall, the size of which is determined by a given sample, so if it is necessary to select 200 g of soil, the sample size is 20cm x 3cm x 3cm, 500g - 20cm x 5cm x 3cm.

Point samples are taken with a knife, spatula or soil drill.

The pooled sample is made up by mixing incremental samples taken from the same sample site.

For bacteriological analysis, 10 combined samples are made from one trial site. Each combined sample is made up of three point samples weighing from 200 to 250 g each, taken in layers from a depth of 0 to 5 cm, from 5 to 20 cm.

Soil samples intended for bacteriological analysis, in order to prevent their secondary contamination, should be taken in compliance with aseptic rules: sterile instruments, mixed on a sterile surface, placed in a sterile container. The time from sampling to the start of their study should not exceed 1 day.

When monitoring the sanitary condition of soils in the territories of children's institutions and playgrounds, sampling is carried out separately from sandboxes and from the general territory from a depth of 0–10 cm.

One pooled sample is taken from each sandbox, made up of 5 incremental samples. If necessary, it is possible to select one combined sample from all sandboxes of each age group, composed of 8-10 point samples.

Soil samples are taken either from the playing territories of each group (one combined of at least five point samples), or one combined sample from a total area of ​​10 point ones, while taking into account the most likely places of soil contamination.

When monitoring soils in the area of ​​point sources of pollution (cesspools, waste bins, etc.), test plots no larger than 5 x 5 m are laid at different distances from the source and in a relatively clean place (control).

When studying soil pollution by transport routes, test sites are laid on roadside strips, taking into account the terrain, vegetation cover, meteorological and hydrological conditions.

Soil samples are taken from narrow strips 200-500 m long at a distance of 0-10, 10-50, 50-100 m from the roadbed. One mixed sample is made up of 20-25 point samples taken from a depth of 0-10 cm.

When assessing the soils of agricultural areas, soil samples are taken 2 times a year (spring, autumn) from a depth of 0-25 cm. For every 0-15 ha, at least 1 site of 100-200 m2 is laid, depending on the terrain and land use conditions.

To prepare an average sample with a volume of 0.5 kg, the soil of all samples of one area is poured onto a sterile, dense sheet of paper, thoroughly mixed with a sterile spatula, stones and other solid objects are discarded. Then the soil is spread on the sheet in an even thin layer in the shape of a square.

The soil is divided diagonally into 4 triangles, the soil from two opposite triangles is discarded, and the rest is mixed again, again distributed in a thin layer and divided diagonally until about 0.5 kg of soil remains.

Then the sample is delivered to the laboratory with the direction and act of sampling.

1. The sample packaging is inspected and matched with the inscription on the lithographic print or on the label indicated in the accompanying document.

2. The package with the sample is cleaned from contamination. If hermetically sealed samples of the product were received for analysis, then the tightness of the container is checked. Hermetically sealed glass, metal or polymer containers with the product are washed with water and detergent, then rinsed with clean water and dried. Leaky packaging with the product is wiped with a swab moistened with ethyl alcohol.

3. Microbiological analysis of samples of the product normal in appearance is carried out in a box under aseptic conditions. The packaging of a sample of a suspicious-looking or spoiled product is opened in a separate room.

4. Samples with a frozen product are thawed at a temperature of (4 ± 2) ° C before preparing a sample. The sample is taken immediately after thawing, but no later than 18 hours later. It is allowed to defrost a sample of the product at a temperature of 18-20°C for 1 hour. Samples of a product of a homogeneous consistency can be thawed in a thermostat at 35°C, provided that complete defrosting is achieved in no more than 15 minutes.

5. Opening the packaging with a product sample

5.1. Immediately before opening the package with a product sample in a consumer container, free-flowing or liquid-phase products are mixed by turning the container 10 times from the bottom to the lid or in a circular motion.

5.2. The package with a sample of the product (except for canned food) is wiped with a swab soaked in 70% ethyl alcohol, the alcohol is then removed by free evaporation. Then the package is opened, the neck of metal or glass jars is fired and the mass (volume) of the product is taken in the amount necessary to prepare one or more samples.

5.3. The package with the sample (bags made of foil, polymeric materials or paper) is opened in a place previously treated with a swab soaked in alcohol. The opening of the package with the sample of the product is carried out in such a way that the possibility of contamination of the product and surrounding objects is excluded.

The surface of canned food that is normal in appearance is treated with ethyl alcohol in one of the following ways:

The surface of the lid is wiped with an alcohol swab, the swab is left on the surface and set on fire before opening the canned food;

Rubber caps and crown caps, Bekelite and plastic closures are treated in the same way, but the tampon is not set on fire;

The metal cover (end), depending on the purpose of the analysis, is opened or pierced with a punch 1-4 times in the immediate vicinity of the burning swab. The size of the hole (diameter or length) should be 1-3 cm.

5.4. Selected weighed portions of the product are immediately sown in nutrient media or transferred to a peptone-salt solution to prepare a dilution.

Before opening bottles or tubes with screw caps, the finished cap or bush is unscrewed. The edges of the bottle or the membrane of the tube are fired in the flame of a burner; the membrane is pierced with a sterile scalpel.

Before opening a bottle sealed with a crown or foil stopper, the shutter is fired in the flame of a burner, the cork is removed with a sterile key, the edges of the bottle are again burned in the flame of the burner.

When opening bottles with a rubber closure, the closure treated with ethyl alcohol is removed without preliminary firing, and the edges of the bottle are burned with a burner flame.

5.5. Canned food that is defective in appearance is placed on a metal tray. Immediately before taking a sample of the product, the surface of the lid (end) is treated in the manner specified in paragraph 5.2, but ethyl alcohol is not ignited. The treated lid (or end) is covered with an inverted sterile metal funnel so that the funnel completely covers the surface. Through the narrow opening of the funnel, carefully pierce the lid (end) with a sterile punch, forming a needle hole.

Instead of a metal funnel, it is allowed to use a plastic bag. After processing the lid (end), canned food is placed in a plastic bag previously wiped with ethyl alcohol so that the bottom of the bag covers the surface to be opened. The bag is tightly tied at the bottom. Carefully, with a light pressure of the punch, a hole is made simultaneously in the lid of the can and in a plastic bag tightly pressed against it.

After the gas and product stop escaping from the can with the product, the funnel and bag are removed, the lid is wiped again with a sterile swab, the hole is expanded with a punch, and a sample of the product is immediately taken from the can for sowing or for preparing its dilutions.

6. Sampling and preparation of initial dilution

6.1. Depending on the indicators to be determined, one or several portions are taken from each sample of the product for the preparation of dilutions and / or inoculation into nutrient media.

6.2. The mass (volume) of the sample intended for sowing into nutrient media and / or for preparing its dilution should be established in the regulatory and technical documentation for a specific type of product or analysis methods, but be at least 10 ± 0.1 g (cm 3).

6.3. A sample for sowing is taken by weight or volumetric method immediately after opening the product sample. The opening is carried out under conditions that exclude contamination of the product by microorganisms, in the immediate vicinity of the burner flame with sterile instruments.

6.4. A sample of the product is selected so that it contains all its components and in the same ratio as in the analyzed sample.

6.5. To prepare dilutions of a sample of the product, a peptone-salt solution is used. The ratio between the mass (volume) of the sample of the product and the volume of the peptone-salt solution for the initial and subsequent dilutions is:

1: 9 - for 10-fold dilution (for products containing a large amount of fat without surfactants 1:10);

1:5 - for 6-fold dilution;

1:3 - for 4-fold dilution;

1:1 - for 2-fold dilution.

If it is necessary to dilute a sample of products containing a large amount of fat, it is allowed to use surfactants (sodium bicarbonate, etc.) that do not have antimicrobial activity.

To prepare a dilution of a sample of products with high osmotic pressure, it is allowed to use peptone or distilled water.

6.6. The initial dilution of a sample of the product is prepared under aseptic conditions in one of the following ways:

dissolution of products;

dilution of products having a liquid phase;

suspension of powders, pasty products and microbially contaminated product pieces; homogenization of solid products.

6.7. Samples of liquid and viscous products are taken with a sterile pipette with a cotton plug by inserting the pipette into the depth of the product.

The portion of the product remaining on the surface of the pipette is allowed to drain to the tip of the pipette. The resulting drop is removed by touching the inner wall of the dish or consumer container above the surface of the product.

Viscous products are removed from the surface of the pipette with a sterile swab.

A portion of the product is transferred to a container with peptone-brine solution to prepare the initial dilution so that the pipette does not touch the surface of the peptone-brine solution. With another sterile pipette, thoroughly mix the product with the peptone-salt solution by tenfold filling and ejection of the mixture.

When working with viscous products, it is advisable to place several glass balls in a container for faster mixing with a peptone-salt solution.

6.8. The liquid product saturated with carbon dioxide (CO 2 ) is transferred to a sterile, cotton-stopped conical flask or other container and heated with frequent stirring in a circular motion in a water bath at a temperature of 30 to 37 ° C until no no more gas bubbles will come out.

A sample of the product is taken and processed according to clause 6.7.

6.9. Samples of powdered or bulk products are taken with a sterile spoon or spatula from different places of the product (if necessary, 2 cm of the top layer of the product is removed with a sterile spoon before sampling), then the sample is transferred to a pre-weighed sterile container with a lid, weighed. A peptone-salt solution is added to the sample in the amount necessary to prepare the initial dilution. The mixture is stirred or shaken 25 times in a circular motion with a radius of 30 cm until a homogeneous product consistency is obtained.

If the powdered product did not dissolve in water, then after mixing it with a peptone-salt solution, the resulting suspension was allowed to settle for 10 minutes and again shaken vigorously for 1 minute.

6.10. A portion of samples of water-swellable products is taken and the initial dilution is prepared in accordance with the requirements of regulatory and technical documentation for a specific type of product.

6.11. A sample of solid water-soluble products is taken with a spatula or spoon after they have been crushed, ground or ground under aseptic conditions and then processed according to clause 6.9.

A sample from samples of water-insoluble solid products is homogenized in the cases specified in the regulatory and technical documentation. When homogenizing the product, the total number of revolutions of the homogenizer should be 15-20 thousand. The number of revolutions of the homogenizer should not be less than 8000 and more than 45000 revolutions per minute.

It is allowed to homogenize an unsterilized product by grinding it until a homogeneous consistency is achieved in a sterile mortar under aseptic conditions.

6.12. A sample of pasty products is taken after thorough mixing with a spoon or glass rod and then processed according to clause 6.9.

6.13. A sample of liquid fat samples is taken with a warm pipette heated by flambing. After filling the pipette with the product, the remains of it are removed from the surface of the pipette with a sterile swab.

The product from the pipette is introduced into a glass-stoppered dish and diluted with the required amount of peptone-salt solution heated to 40-45°C; when psychrophilic microorganisms are detected, the temperature should not exceed 37 ° C. Fat residues adhering to the pipette are rinsed with a peptone-salt solution, which is sucked up several times and released from the pipette.

6.14. A sample of solid fats is taken after cutting the product with a knife or wire into several parts. If necessary, remove the top layer.

A sample of the product is taken from the surface of the cuts from different places with a scalpel and transferred to a weighed dish with a lid.

A certain mass of sample is transferred to a wide-mouthed dish with ground glass stopper. The remains of fat adhering to the walls of the dish are rinsed in the same dish with a certain amount of peptone-salt solution heated to 40-45 ° C, which is added to the dish in the amount necessary to obtain the initial dilution.

From solid fats, a sample can be selected by volume. Fats are melted in a dish with a wide neck in a water bath at a temperature not exceeding 45 ° C; when psychrophilic microorganisms are detected, the temperature should not exceed 37 ° C.

After mixing the melted fat, it is transferred with a warm pipette to a wide-necked dish with a ground glass lid containing the required amount of peptone-salt solution to prepare the initial dilution. Peptone-salt solution is preheated to 40-45°C; when detecting psychrophilic microorganisms up to 37°C.

6.15. Samples from samples of whipped products or a mushy consistency containing a large amount of fat, after mixing with a glass rod, are taken with a spoon into a weighed dish and a peptone-salt solution heated to 40-45 ° C is added in the amount necessary to prepare the initial dilution.

6.16. Determination of microbial contamination of the surface of product samples is carried out by flushing with cotton swabs.

A sterile cotton swab is moistened with a peptone-salt solution and wiped with it in different places on the surface of various pieces of the analyzed product with a total area of ​​100 cm 2 .

The surface area to be analyzed is measured using sterile templates with holes of the appropriate size.

The swab is placed in a container containing at least 100 cm 3 of peptone-salt solution. The dishes are closed with a rubber stopper and shaken until the swabs disintegrate into individual fibers. The resulting suspension is considered the original dilution.

7. Preparation of tenfold dilutions

7.1. The first tenfold dilution of the sample is the initial one, the initial dilution is prepared in accordance with paragraph 6. Subsequent dilutions are obtained from it.

7.2. The subsequent second dilution is prepared from one part of the original dilution and nine parts of peptone-saline solution by mixing in a test tube.

If a pipette was used to mix the initial dilution, then with the same pipette add 1 cm 3 of the initial dilution to 9 cm 3 of the peptone-salt solution, without touching the surface of the solution with the pipette. The dilution is mixed with another pipette by sucking and blowing out the contents of the tube ten times.

7.3. The third and subsequent dilutions are prepared in a similar way.

7.4. The interval between the preparation of weighed portions of the product, their dilutions and sowing in nutrient media should not exceed 30 minutes.

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METHODS OF MICROBIOLOGICAL SOIL CONTROL - METHODOLOGICAL RECOMMENDATIONS (approved by the Chief State Sanitary Doctor of the Russian Federation ... Relevant in 2018

4. Sampling for bacteriological analysis

The control of soil pollution in settlements is carried out taking into account the functional zones of the city. Sampling sites are preliminarily marked on a map showing the structure of the urban landscape. Sampling is carried out in accordance with GOST 17.4.4.01-83 "General requirements for soil sampling"; GOST 17.4.4.02-84 "Methods for sampling and sample preparation for chemical, bacteriological, helminthological analysis". For the territory to be controlled, a description is made indicating the address, sampling point, general relief of the microdistrict, location of sampling sites and pollution sources, vegetation cover, land use, groundwater level, soil type and other data necessary for the correct assessment and interpretation of the analysis results. samples.

Sampling for bacteriological analysis is carried out at least once a year in places where people and animals are likely to be located, in places of pollution with organic waste. When studying the dynamics of soil self-purification, selection is carried out weekly during the first month, and then monthly during the growing season until the end of the active phase of self-purification.

Trial site - a part of the study area, characterized by similar conditions (relief, uniformity of soil structure and vegetation cover, the nature of economic use).

The test site should be located in a typical location for the study area. On an area of ​​100 sq. m, one test site with a size of 25 m is laid. In case of heterogeneity of the relief, the sites are selected according to the elements of the relief.

Point sample - material taken from one place of the horizon or one layer of the soil profile, typical for this horizon or layer.

Point samples are taken on a trial plot from one or more layers or horizons using the envelope method. A pit 0.3 m x 0.3 m and a depth of 0.2 m is excavated. The surface of one of the walls of the pit is cleaned with a sterile knife. Then a soil sample is cut out of this wall, the size of which is determined by a given sample, so if 200 g of soil is to be taken, the sample size is 20 cm x 3 cm x 3 cm, 500 g is 20 cm x 5 cm x 3 cm.

Point samples are taken with a knife, spatula or soil drill.

The pooled sample is made up by mixing incremental samples taken from the same sample site.

For bacteriological analysis, 10 combined samples are made from one trial site. Each combined sample consists of three point samples weighing from 200 to 250 g each, taken in layers from a depth of 0 to 5 cm, from 5 cm to 20 cm.

Soil samples intended for bacteriological analysis, in order to prevent their secondary contamination, should be taken in compliance with aseptic rules: they are taken with sterile instruments, mixed on a sterile surface, and placed in a sterile container. The time from sampling to the start of their study should not exceed 1 day.

When studying the impact of pesticides and other chemicals on microflora and self-purification processes in deeper soil layers, a pit up to 1 m deep is used for soil sampling. Samples are taken from the pit wall with a sterile instrument every 10 cm.

To control the sanitary condition of soils in preschool, school and medical institutions, playgrounds and recreation areas, sampling is carried out at least 2 times a year - in spring and autumn. The size of the trial site should not exceed 5 x 5 m.

When monitoring the sanitary condition of soils in the territories of children's institutions and playgrounds, sampling is carried out separately from sandboxes and from the general territory from a depth of 0 - 10 cm.

One pooled sample is taken from each sandbox, made up of 5 incremental samples. If necessary, it is possible to select one combined sample from all sandboxes of each age group, composed of 8 - 10 point samples.

Soil samples are taken either from the playing territories of each group (one combined of at least five point samples), or one combined sample from a total area of ​​10 point ones, while taking into account the most likely places of soil contamination.

When monitoring soils in the area of ​​point sources of pollution (cesspools, waste bins, etc.), test plots no larger than 5 x 5 m are laid at different distances from the source and in a relatively clean place (control).

When studying soil pollution by transport routes, test sites are laid on roadside strips, taking into account the terrain, vegetation cover, meteorological and hydrological conditions.

Soil samples are taken from narrow strips 200 - 500 m long at a distance of 0 - 10, 10 - 50, 50 - 100 m from the roadway. One mixed sample is made up of 20 - 25 point samples taken from a depth of 0 - 10 cm.

When assessing the soils of agricultural areas, soil samples are taken 2 times a year (spring, autumn) from a depth of 0 - 25 cm. m depending on the terrain and land use conditions.

On the territory of large cities with numerous sources of pollution, geochemical mapping is carried out using the testing network. To identify sources of contamination, a sampling density of 1–5 samples per 1 sq. km is recommended. km with a distance between sampling points of 400 - 1000 m. km with a distance between sampling points of about 200 m. Samples are taken from a depth of 0 - 5 cm.

The samples taken must be numbered and registered in a journal, indicating the following data: serial number and place of sampling, terrain, soil type, purpose of the territory, type of pollution, date of sampling.

Samples must have a label indicating the place and date of sampling, the number of the soil section, soil difference, the horizon and depth of sampling, and the name of the researcher.

7. The limitation of the validity period was removed by the Decree of the State Standard of the USSR of 01.23.91 N 38

8. EDITION (April 2010) with Amendment No. 1 approved in September 1989 (IUS 12-89)


This International Standard applies to food and flavor products and specifies the preparation of samples for microbiological analysis.

The terms used in the standard and their explanations are given in Appendix 1.

1. EQUIPMENT, REAGENTS AND MATERIALS

1.1. When preparing samples for analysis, the following equipment, reagents and materials are used:

water bath;

homogenizer, laboratory mixer or porcelain mortar according to GOST 9147;

membrane filtration device;

gas or alcohol burner according to GOST 25336;

metal funnels;

punch;

bottle opener;

can opener;

scissors, scalpel, tweezers according to GOST 21241, spatula, spoon;

stencils (template);

test tubes according to GOST 25336;

flasks according to GOST 25336;

pipettes according to NTD;

rubber stoppers;

glass balls;

rectified ethyl alcohol according to GOST 5962*; 70%;
________________
* On the territory of the Russian Federation, GOST R 51652-2000 applies.


plastic bags;

detergent;

sodium chloride according to GOST 4233;

peptone for bacteriological purposes according to GOST 13805.

Tools and surfaces of devices in direct contact with the product must be sterilized by one of the methods specified in GOST 26668.

1.2. Preparation of peptone-salt solution

Peptone-salt solution is prepared as follows: 8.5 g of sodium chloride and 1.0 g of peptone are dissolved in 1 dm of distilled water with slow heating. The resulting solution, if necessary, is filtered through a paper filter, adjusted to pH 7.0 ± 0.1, poured into flasks, test tubes or other vessels, sealed and sterilized at a temperature of (121 ± 1) ° C for 30 minutes.

The solution is stored in a dark place at a temperature of (4 ± 2) ° C for no more than 30 days under conditions that exclude moisture evaporation.

The temperature of the peptone-salt solution should correspond to the temperature of the analyzed product.

1.3. Preparation of peptone water

Peptone water is prepared similarly to a peptone-brine solution without the addition of sodium chloride.

2. PREPARATION OF SAMPLES FOR ANALYSIS

2.1. The sample packaging is inspected and matched with the inscription on the lithographic print or on the label indicated in the accompanying document.

2.2. The packaging with the sample is cleaned of contaminants. If hermetically sealed samples of the product were received for analysis, then the tightness of the container is checked. The tightness of canned food is determined according to GOST 8756.18, the tightness of polymer containers with the product, as well as canned food sealed with lids with an elastic membrane (button) - visually. The surface of the elastic membrane should be concave inwards. Hermetically sealed glass, metal or polymer containers with the product are washed with water and detergent, then rinsed with clean water and dried. Leaky packaging with the product is wiped with a swab moistened with ethyl alcohol.

Canned food is thermostated immediately before microbiological analysis.

Canned food is subject to temperature control:

hermetically sealed, defect-free in appearance, designed to determine the industrial sterility of a canned product and the microbiological stability of canned food;

with vibrating ends and crackers in hermetically sealed containers, designed to identify the causes of these defects.

Canned food intended for the detection of botulinum toxins in them, bombed, with signs of microbiological spoilage and leaky thermostats are not subject to thermostating.

For the manifestation of the vital activity of mesophilic aerobic, facultative anaerobic and anaerobic microorganisms, canned food is thermostated at 30-37 ° C in a container with a capacity of up to 1 dm inclusive for at least 5 days, in a container with a capacity of more than 1 dm - for at least 7 days.

For the manifestation of the vital activity of thermophilic aerobic, facultative anaerobic and anaerobic microorganisms, canned food in a container of any capacity is thermostated at 55-62 ° C for at least 3 days. During thermostatting, canned food is inspected daily. Canned food with manifested container defects immediately after their detection is removed from the thermostat and kept for 24 hours at room temperature, after which the state of the container and, if possible, the appearance of the product are noted. Canned food in containers that take on a normal appearance after cooling at room temperature are considered defect-free and continue their thermostating.

After thermostating canned food and cooling for 24 hours to room temperature, note the condition of the container and, if possible, the appearance of the product.

Defects in canned food are listed in Appendix 2.

2.3. Microbiological analysis of normal in appearance samples of the product is carried out in a box under aseptic conditions. The packaging of a sample of a suspicious-looking or spoiled product is opened in a separate room.

The preparation of the box is described in Appendix 3.

2.2, 2.3. (Changed edition, Rev. N 1).

2.4. Samples with a frozen product are thawed at a temperature of (4 ± 2) °C before preparing a sample. The sample is taken immediately after thawing, but no later than 18 hours after the start of thawing.

It is allowed to defrost a sample of the product at a temperature of 18-20 °C for 1 hour.

Samples of the product of homogeneous consistency may be thawed in a thermostat at 35 °C, provided that complete defrosting is achieved in no more than 15 minutes.

2.5. Opening the packaging with a product sample

2.5.1. Immediately before opening the package with a sample of the product in consumer containers, free-flowing or having a liquid phase, mix by 10-fold inversion of the container from the bottom to the lid or in a circular motion.

2.5.2. The package with a sample of the product (except for canned food) is wiped with a swab soaked in 70% ethyl alcohol, the alcohol is burned or removed by free evaporation. Then the package is opened, the neck of metal or glass jars is fired and the mass (volume) of the product is taken in the amount necessary to prepare one or more samples.

2.5.3. The package with the sample (bags made of foil, polymeric materials or paper) is opened in a place previously treated with a swab soaked in alcohol. Opening of the package with a sample of the product is carried out in such a way that the possibility of contamination of the product, surrounding objects and the environment is excluded.

2.5.4. Before opening, the surface of canned food that is normal in appearance is treated with ethyl alcohol in one of the following ways:

for glass jars, the lid is processed, for metal cans - the end opposite to the marked one.

The surface of the lid is wiped with an alcohol swab, the swab is left on the surface and lit before opening the canned food;

rubber caps and crown caps, bekelite and plastic closures are also processed, but the tampon is not lit;

the metal cover (end), depending on the purpose of the analysis, is opened or pierced with a punch 1-4 times in the immediate vicinity of the burning swab. The size of the hole (diameter or length) should be 1-3 cm.

The selected sample of the product is immediately sown in nutrient media or transferred to a peptone-salt solution to prepare a dilution;

before opening bottles or tubes with screw caps, the finished cap or bush is unscrewed. The edges of the bottle or the membrane of the tube are fired in the flame of a burner; the membrane is pierced with a sterile scalpel.

Before opening a bottle sealed with a crown or foil stopper, the shutter is fired in the flame of a burner, the cork is removed with a sterile key, the edges of the bottle are again burned in the flame of the burner.

When opening bottles with a rubber stopper, the stopper treated with ethyl alcohol is removed without preliminary firing and the edges of the bottle are burned with a burner flame.

2.5.5. Canned food that is defective in appearance is placed on a metal tray. Immediately before taking a sample of the product, the surface of the lid (end) is treated in the manner specified in clause 2.5.2, but ethyl alcohol is not ignited. The treated lid (or end) is covered with an inverted sterile metal funnel so that the funnel completely covers the surface. Through the narrow opening of the funnel, carefully pierce the lid (end) with a sterile punch, forming a needle hole.

Instead of a metal funnel, it is allowed to use a plastic bag. After processing the lid (end), canned food is placed in a plastic bag previously wiped with ethyl alcohol so that the bottom of the bag covers the surface to be opened. The bag is tightly tied at the bottom. Carefully, with a light pressure of the punch, a hole is made simultaneously in the lid of the can and in a plastic bag tightly pressed against it.

After the gas and product stop escaping from the can with the product, the funnel and bag are removed, the lid is wiped again with a sterile swab, the hole is expanded with a punch, and a sample of the product is immediately taken from the can for sowing or for preparing its dilutions.

2.6. Sampling and preparation of initial dilution

2.6.1. From each sample of the product, depending on the indicators to be determined, one or several portions are taken for the preparation of dilutions and/or inoculation into nutrient media.

2.6.2. The mass (volume) of the sample intended for sowing into nutrient media and/or for preparing its dilutions must be established in the regulatory and technical documentation for a specific type of product or analysis methods.

2.6.3. A sample for sowing is taken by weight or volumetric method immediately after opening the product sample. The opening is carried out under conditions that exclude contamination of the product by microorganisms, in the immediate vicinity of the burner flame with sterile instruments.

2.6.4. A sample of the product is selected so that it contains all its components and in the same ratio as in the analyzed sample.

2.6.5. To prepare dilutions of a sample of the product, a peptone-salt solution is used.

It is allowed to prepare the initial dilutions of products with a mass fraction of NaCl more than 5% using peptone water, the initial dilutions of meat, fish and dairy products - using saline.

The weight (volume) of the sample of the product intended for the preparation of the initial dilution or homogenate must be at least (10±0.1) g/cm.

The ratio between the mass (volume) of the sample of the product and the volume of the peptone-salt solution for the initial and subsequent dilutions is:

1:9 - for 10-fold dilution (for products containing a large amount of fat without surfactants 1:10);

1:5 - for 6-fold dilution;

1:3 - for 4-fold dilution;

1:1 - for 2-fold dilution.

If it is necessary to dilute a sample of products containing a large amount of fat, it is allowed to use surfactants (sodium bicarbonate, etc.) that do not have antimicrobial activity.

To prepare a dilution of a sample of products with high osmotic pressure, it is allowed to use peptone or distilled water.

(Changed edition, Rev. N 1).

2.6.6. The initial dilution of a sample of the product is prepared under aseptic conditions in one of the following ways:

dissolution of products;

dilution of products having a liquid phase;

suspension of powders, pasty products and the surface of microbially contaminated product pieces;

homogenization of solid products.

2.6.7. Samples of liquid and viscous products are taken with a sterile pipette with a cotton plug by inserting the pipette into the depth of the product.

The portion of the product remaining on the surface of the pipette is allowed to drain to the tip of the pipette. The resulting drop is removed by touching the inner wall of the dish or consumer container above the surface of the product.

Viscous products are removed from the surface of the pipette with a sterile swab.

A portion of the product is transferred to a container with peptone-brine solution to prepare the initial dilution so that the pipette does not touch the surface of the peptone-brine solution. With another sterile pipette, thoroughly mix the product with the peptone-salt solution by tenfold filling and ejection of the mixture.

When working with viscous products, it is advisable to place several glass balls in a container for faster mixing with a peptone-salt solution.

2.6.8. The liquid product saturated with carbon dioxide (CO) is transferred to a sterile, cotton-stopped conical flask or other container and heated with frequent stirring in a circular motion in a water bath at a temperature of 30 to 37 ° C until it stops gas bubbles are released.

A sample of the product is taken and processed according to clause 2.6.7.

2.6.9. Samples of powdered or bulk products are taken with a sterile spoon or spatula from different places of the product (if necessary, 2 cm of the top layer of the product is removed with a sterile spoon before sampling), then the sample is transferred to a pre-weighed sterile container with a lid, weighed. A peptone-salt solution is added to the sample in the amount necessary to prepare the initial dilution. The mixture is stirred or shaken 25 times in a circular motion with a radius of 30 cm until a homogeneous product consistency is obtained.

If the powdered product is insoluble in water, then after mixing it with a peptone-salt solution, the resulting suspension is allowed to stand for 10 minutes and again shaken vigorously for 1 minute.

2.6.10. A portion of samples of water-swellable products is taken and the initial dilution is prepared in accordance with the requirements of regulatory and technical documentation for a specific type of product.

2.6.11. Samples of solid water-soluble products are taken with a spatula or spoon, after they are crushed, milled or rubbing under aseptic conditions, and then processed according to clause 2.6.9.

A sample from samples of water-insoluble solid products is homogenized in the cases specified in the regulatory and technical documentation. When homogenizing the product, the total number of revolutions of the homogenizer should be 15-20 thousand. The number of revolutions of the homogenizer should not be less than 8000 and more than 45000 revolutions per minute.

If a heterogeneous mass is obtained during homogenization of the product, then it is settled for 15 minutes and the supernatant liquid is used for inoculation and (or) preparation of dilutions.

It is allowed to homogenize an unsterilized product by grinding it until a homogeneous consistency is achieved in a sterile mortar under aseptic conditions.

(Changed edition, Rev. N 1).

2.6.12. A sample of pasty products is taken after thorough mixing with a spoon or glass rod and further processed according to clause 2.6.9.

2.6.13. A sample of liquid fat samples is taken with a warm pipette heated by flambing. After filling the pipette with the product, the remains of it are removed from the surface of the pipette with a sterile swab.

The product from the pipette is introduced into a container with a ground glass stopper and diluted with the required amount of peptone-salt solution heated to 40-45 ° C; when psychrophilic microorganisms are detected, the temperature should not exceed 37 °C. Fat residues adhering to the pipette are rinsed with a peptone-salt solution, which is sucked up several times and released from the pipette.

2.6.14. A sample of solid fats is taken after cutting the product with a knife or wire into several parts. If necessary, remove the top layer.

A sample of the product is taken from the surface of the cuts from different places with a scalpel and transferred to a weighed dish with a lid.

A certain mass of sample is transferred to a wide-mouthed dish with ground glass stopper. The remains of fat adhering to the walls of the dish are rinsed in the same dish with a certain amount of peptone-salt solution heated to 40-45 ° C, which is added to the dish in the amount necessary to obtain the initial dilution.

From solid fats, a sample can be selected by volume. Fats are melted in a wide-necked dish in a water bath at a temperature not exceeding 45 ° C; when psychrophilic microorganisms are detected, the temperature should not exceed 37 °C.

After mixing the melted fat, it is transferred with a warm pipette to a wide-necked dish with a ground glass lid containing the required amount of peptone-salt solution to prepare the initial dilution. Peptone-salt solution is preheated to 40-45 °C; when detecting psychrophilic microorganisms up to 37 °C.

2.6.15. Samples from samples of whipped products or a mushy consistency containing a large amount of fat, after mixing with a glass rod, are taken with a spoon into a weighed dish and a peptone-salt solution, heated to 40-45 ° C, is added in the amount necessary to prepare the initial dilution.

2.6.16. Determination of microbial contamination of the surface of product samples is carried out by flushing with cotton swabs.

A sterile cotton swab is moistened with a peptone-salt solution and wiped with it in different places on the surface of various pieces of the analyzed product with a total area of ​​100 cm2.

The surface area to be analyzed is measured using sterile templates with holes of the appropriate size.

The swab is placed in a test tube containing 10 ml of peptone-salt solution. The contents of the tube are thoroughly mixed with a pipette. The resulting suspension is considered the original dilution.

(Changed edition, Rev. N 1).

2.7. Preparation of tenfold dilutions

2.7.1. The first tenfold dilution of the sample is the initial one, the initial dilution is prepared in accordance with clause 2.6. Subsequent dilutions are obtained from it.

2.7.2. The subsequent second dilution is prepared from one part of the original dilution and nine parts of peptone-saline solution by mixing in a test tube.

If a pipette was used to mix the initial dilution, then the same pipette is used to add 1 cm of the initial dilution to 9 cm of peptone-salt solution, without touching the surface of the solution with the pipette. The dilution is mixed with another pipette by sucking and blowing out the contents of the tube ten times.

2.7.3. The third and subsequent dilutions are prepared in a similar way.

2.7.4. The interval between the preparation of weighed portions of the product, their dilutions and sowing in nutrient media should not exceed 30 minutes.

APPENDIX 1 (informative). The term used in the standard and its explanations

ATTACHMENT 1
Reference

(Changed edition, Rev. N 1).

APPENDIX 2 (informative). CANNED DEFECTS

APPENDIX 2
Reference

Defects in a canned product are:

visible to the naked eye signs of the development of microorganisms: fermentation, mold, mucus, etc .;

sediment at the bottom of the can or at the border of the surface of the product with the container ("ring");

turbidity of the liquid phase;

coagulation;

souring;

extraneous, not characteristic of the product, smell and (or) taste;

color change.

Defects in the appearance of containers with products packaged in it are considered:

signs of leaks visible to the naked eye: holes, through cracks, smudges or traces of the product flowing out of the can;

bombing;

crackers;

cans with vibrating ends;

incorrectly designed seam of cans (tongues, teeth, undercut, false seam, rolled seam);

rust, after the removal of which shells remain;

deformation of the body, ends or longitudinal seam of cans in the form of sharp edges and "birds";

skewed lids on glass jars, undercutting the corrugations of lids along a rolled field, a protruding rubber ring ("loop");

cracks or chipped glass at the seam, incomplete fit of the lids relative to the neck of the can;

deformation (indentation) of the lids of glass jars, which caused a violation of the seaming seam;

convex elastic membrane (button) on the cover.

APPENDIX 3 (informative). BOXING PREPARATION

APPENDIX 3
Reference

Canned food is opened in a box-room specially adapted for microbiological analysis. In the box, there should be no surfaces inaccessible for wet disinfection and air movement created by drafts should be excluded. Walls, floors and ceilings must be lined with material or painted with disinfectants resistant to wet treatment. For air sterilization, the box is equipped with ultraviolet lamps at the rate of 1.5-2.5 W per 1 m.

Only the microbiologist conducting the analysis and, if necessary, an assistant should be in the box.

The box must have a table and a stool. There should be no extra items, except for those required for the analysis of canned food.

On the table should be:

spirit stove or gas burner;

a jar with a ground stopper with alcohol;

a jar closed with a lid with pre-prepared dense sterile cotton swabs 3x3 cm in size or cotton rings;

jars with a disinfectant solution (layer height 3 cm) for placing pipettes or tubes used after analysis;

a small metal or enameled tray on which the analyzed cans are placed;

sterile pipettes or tubes with which the sample is taken.

Auxiliary equipment should be stored in the drawer of the table: tweezers and a punch. The punch should have the shape of a spear with a cross section in the form of a rhombus with diagonals of 1x1.5 cm or with a cross section in the form of an isosceles triangle.

When opening a large number of cans, a punch mounted on a tripod is used. In this case, the opening is performed by pressing the punch on the lid of the jar with a lever.

Before opening the jar, the punch is flambéed in the flame of a swab.

The box is washed and disinfected immediately before the analysis (not earlier than 24 hours before the start) and after its completion. Disinfection is carried out by wiping all surfaces with chlorine or other disinfectants according to the instructions corresponding to each preparation. 45 minutes before the start of work in the box for (30 ± 5) minutes, bactericidal lamps are turned on.

At present, laminar flow cabinets (protective cabins of ultrapure air) are used for microbiological analysis. Laminar boxes are produced by the Uzhgorod plant of medical equipment "Laminar", boxes of the BPV 1200 brand are produced in Hungary, boxes of the TVG-S II 1.14.1 brand are produced by Babcock - BSH (Germany).


APPENDICES 2, 3. (Introduced additionally, Rev. N 1).

Electronic text of the document
prepared by Kodeks JSC and verified against:
official publication
Food products, canned food.
Methods of microbiological analysis:

Sat. GOSTs. - M.: Standartinform, 2010