RU2585521C1 - Method and process line for production of nisin - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: group of inventions relates to production of nisin. Disclosed is method and process line for microbiological production of nisin. Method involves culturing of nisin producer in fermenter using nisin producer strain Lactococcus (Streptococcus) lactis, followed by extraction of nisin. Components of nutrient medium represented by brine after drying of dairy milk whey, milk whey, rinsings obtained after process equipment washing at dairy factories, as well as megaterin. Culturing is carried out using automatic pH-stating and thermal regulation, nisin extraction is carried out by acidification with subsequent nisin extraction by centrifugation. Process line contains fermenter with spherical bottom and cover. Fermenter is equipped with bubbler, jacket, mixer and automatic pH-stating and temperature controlling system. Inputs of fermenter are aligned with nutrient medium and seed material feed systems, and output by culture fluid is interconnected through extraction, centrifugation, ultrafiltration and nano filtration units with nisin preparation drying unit.

EFFECT: simultaneously with production of nisin inventions provide recycling of previously not used milk production wastes.

17 cl, 1 dwg

Description

The invention relates to the field of biotechnology, and in particular to the production of the target product using microbiological processes, and can be used in the manufacture of a nisin food additive used as a preservative in the manufacture of processed and hard cheeses, tomato products, canned vegetables, soups and mushrooms, as well as milk products.

Known (RU, patent 2374320, publ. 10.07.2009) a method for producing an antibiotic complex LGS, characterized in that the cultivation of a strain of Lactococcus lactis subsp. lactis 284 - Collection of microorganisms of Moscow State University. M.V. Lomonosov producer of the nizin-like target product in a fermentation medium containing a carbon source, potassium phosphate, magnesium sulfate, sodium chloride, isoleucine and yeast autolysate in an amount that ensures biosynthesis, followed by drying of the culture fluid containing the target product, and, if necessary, separating it into LGS- antibiotics B and LGS-H, which includes a nizin-like product.

The disadvantage of this method should be recognized as the impossibility of obtaining pure nisin.

Known (RU, patent 2151796, publ. 27.06.2000) a method for producing nisin. According to a known method, the strain of Lactococcus lactis VKPM-B-7699 was grown on a nutrient medium. Hydrolyzed milk and whey were used as a nutrient medium. The prepared skimmed milk powder was diluted with water to a solids content of 30 g / l, 20 g / l of whey was added, boiled, cooled to 45-50 ° C, adjusted to a pH of 7.2-7.5 units, and megateria dissolved in a small amount of warm distilled water at the rate of 0.5 g / l of medium. The hydrolysis was carried out at a temperature of 50 ° C for 3 h. At the end of the hydrolysis, the pH of the medium was adjusted to 7.0 units and sterilized for 20 min at 0.8 atm. The nutrient medium can be used for rocking flasks and fermenters of various volumes. The pH of the medium in the flasks and fermenters was adjusted by adding 10% sodium hydroxide solution. The activity of the formation of nisin by the strain of Lactococcus lactis VKPM-B-7699 in a periodic mode in flasks was 10350 IU / ml.

The known method is unsuitable for industrial implementation.

There is a known (SU, ed. St. 707320, publ. 30.11.1981) method for the production of lowlands. According to a known method, a nisin producer is grown on the basis of a nutrient medium comprising whey and fodder yeast extract, followed by extraction of nisin from microbial cells, separation of the extract, its concentration, salting out, filtering and drying, and the purified culture liquid is concentrated with sorbital C-20, filtration is carried out through a perlite filter, the resulting nisin-perlite paste is dissolved in hydrochloric acid, followed by repeated filtration, the obtained concentrate atm dried by spraying it on sodium caseinate.

The disadvantage of this method should recognize its complexity, as well as the low yield of the finished product.

A well-known source of information adopted as the closest analogue to the object method. The processing line used in the implementation of this method is not disclosed.

The technical problem solved by the developed technical solution is to provide the possibility of industrial production of nisin using waste from dairy production.

The technical result achieved by the implementation of the developed technical solution consists in developing an industrial method for producing nisin with the simultaneous utilization of previously unused industrial waste.

To achieve the specified technical result, it is proposed to use the developed method for the production of nisin. According to the developed method, the cultivation of the nisin producer in the fermenter is carried out using the producer strain of nisin Lactococcus (Streptococcus) lactis, followed by extraction of nisin, and brine after drying whey, milk whey and rinses obtained after washing technological equipment with milk are used as components of the nutrient medium enterprises, as well as at least partially hydrolyzed waste from the granary and megateria, cultivation is carried out using an automatic about pH-stating and temperature control, extraction of nisin is carried out by acidification, followed by isolation of nisin by centrifugation.

When implementing the method, a strain of Lactococcus lactis subsp is preferably used. Lactis hybrid strain F-116, a collection of Moscow State University. V.M. Lomonosov, although it is possible to use other strains, including other cultures - producers of nisin.

After drying whey, the brine used in the implementation of the developed method usually contains up to 9.0 g / dm ^{3 of} mineral salts. The composition of the specified brine is well known (http://www.molz.ru/produkty-iz-molochnoi-syvorotki?page=11).

The whey used in the implementation of the method preferably contains not more than 1.4% protein. Used rinses obtained after washing technological equipment in dairy enterprises preferably contain not more than 0.2% protein. Their composition is known (http://www.molz.ru/tekhnologicheskie-tsikly-molochnogo-zavoda?page=6).

The total content of milk protein in the nutrient medium is preferably not more than 1.6%.

During the preparatory operations, a nutrient medium can be used with at least partially hydrolyzed granary waste, the content of nutrients in which corresponds to their amount in 0.05 kg of dry yeast extract.

Usually use a nutrient medium containing 0.07-0.15% megaterina.

In a preferred embodiment of the developed method, nisin is extracted by exposing the culture fluid to a solution of phosphoric acid at a pH of 2.4 ÷ 2.6 with stirring and subsequent heating to 100 ° C, and the fermentation is carried out while maintaining a temperature of 28 ± 1 ° C and pH 6 in the fermenter , 8 with stirring at a speed of 180 rpm

Also, the specified technical result is achieved when using the technological line for the production of nisin. The developed production line contains a fermenter with a spherical bottom and a lid equipped with a bubbler, a shirt, a stirrer and an automatic pH-stat and temperature control system, the inputs of which are combined with the feed medium and seed material supply systems, and the output through the culture fluid is communicated through extraction, centrifugation, ultrafiltration and nanofiltration with the drying unit of the drug nisin.

The specified fermenter may further comprise a means of sampling the culture fluid.

In a preferred embodiment, the processing line may further comprise a nutrient preparation section as well as a seed preparation section.

In a preferred embodiment of the developed production line, the extraction unit is made in the form of a thermostatically controlled vessel equipped with a stirrer, the separation unit is made in the form of a bactofuge, and the line may further comprise a lowland cleaning unit, including an ultrafiltration unit and a nanofiltration unit connected in series to the unit output line separation.

Also in a preferred embodiment for the industrial use of the generated production waste, the line may further comprise an organo-mineral additive production unit connected to the sludge outlet of the separation unit, as well as a lactic acid solution production unit connected to the permeate to the nanofiltration unit.

The developed technological line for the production of nisin is shown in the block diagram, with the following notation used: block 1 for preparing solutions of hydrochloric acid and sodium hydroxide, block 2 for hydrolysis of dairy products, block 3 for introducing a culture of microorganisms, block 4 for preparing uterine flasks, block 5 for sowing preparation nutrient medium, inoculant preparation unit 6, seed preparation unit 7, sterile nutrient preparation unit 8, fermenter 9, extraction unit 10, centrifuge 11, ultrafine unit 12 ltratsii, nanofiltration unit 13, drying unit 14 nisin preparation unit 15 evaporating the permeate unit 16 neutralizing and drying the solid phase centrifugation, lactate deposition unit 17, the unit 18 evaporating the mother liquor lactate unit 19 produce lactic acid.

The process of obtaining the drug nisin consists of the following main steps:

- preparation of equipment, obtaining sterile solutions and media;

- growing seed;

- cultivation of the producer of nisin in a fermenter with automatic pH-statirovanie and thermostating;

- extraction of nisin;

- separation of the extract from the producer cells;

- recycling of lowland production waste.

The developed technical solution can be implemented as follows.

When preparing sterile air, the source air is supplied centrally from the factory air manifold, and in the workshop passes through a heat exchanger for heating the air, the oil separator and through the system for removing mechanical particles enters the places of consumption. Filters and the air manifold are pre-sterilized with hot steam according to the instructions.

When preparing an alkali solution, the operation of weighing and dissolving dry sodium hydroxide is carried out with ventilation turned on or in a well-ventilated room, in personal protective equipment. The dissolution of alkali in water is accompanied by the release of a large amount of heat, which leads to an increase in the temperature of the solution; therefore, this operation is carried out with the observance of precautions required when working with concentrated alkalis.

In particular, to prepare a sterile alkali solution, a 10% (w / v) alkali solution of dry sodium hydroxide is prepared in an amount of 12 l in 2 glass bottles with a capacity of 10 l or in one with a capacity of 15 l.

0.6 kg of dry sodium hydroxide is weighed on scales in a glass or enameled container. 4 l of water is poured into an enameled 10 liter container; dry hydroxide is gradually poured with constant stirring. After dissolution, the volume of the solution is adjusted to 6 liters. The alkali solution is poured into the bottle for sterilization. The following 6 L of solution is prepared similarly. Pour 1 liter of alkali solution for use at the stage of preparation of the nutrient medium TP 2.2.

The solution is sterilized at 0.8 ati for 30 minutes. Sterile alkali is used to maintain pH during fermentation in a fermenter.

A solution of phosphoric acid is prepared in an apparatus with a stirrer and cooling with a capacity of 250 l by diluting concentrated 85% phosphoric acid with water in a ratio of 1: 1 by volume: 25 l of water and 25 l of concentrated acid. 25 l of water is poured into the apparatus and with stirring, acid is gradually poured through the upper hatch, cooling water is supplied to the jacket.

During this operation, all safety precautions necessary when working with concentrated acids are observed: ventilation included and personal protective equipment.

A hydrochloric acid solution is prepared in a container resistant to hydrochloric acid, equipped with a pH sensor. This operation is carried out in a well-ventilated area or under traction. When carrying out this operation, all precautions necessary when working with concentrated acids, in particular with hydrochloric acid, are observed. Concentrated 36% hydrochloric acid is gradually added to 180 l of water until a pH of 2.5 is established.

To obtain the inoculum carry out the following operations.

Prepare 5 l of a suspension of 12% (w / v) milk. To do this, weigh 600 g of milk powder and dissolve it in warm water at 40-45 ° C. A suspension of milk is poured into 100 and 600 ml into sterile flasks with a capacity of 250 and 2000 ml, respectively, and sterilized in an autoclave at 0.5 ati (112 ° C) for 30 minutes. After sterilization, the milk is cooled to a temperature of 30 ° C and thermostated at 30 ° C for 22-24 hours to verify sterility. Flasks in which milk is curled are discarded. Milk for propagation and scaling of the sowing culture is prepared in advance for 4-5 days.

A portion of dry yeast extract 20 g is dissolved in 100 ml of warm water, the solution is poured into a flask with a capacity of 250 ml and sterilized in an autoclave at 0.8 ati (117-118 ° C) for 30 minutes.

Powdered milk can be replaced with whey, brine from drying whey, as well as milk strips, while the content of milk protein should correspond to its content in used milk powder.

Similarly, dry yeast extract can be replaced with at least partially hydrolyzed granular waste, the nutrient content of which corresponds to their amount in the dry yeast extract.

Storage of the resulting culture is carried out mainly in the form of a milk clot in 12% skim sterile milk in a domestic refrigerator at a temperature of 4-6 ° C or in dry (lyophilized) form in a protected environment containing, g / l, gelatin - 140, sucrose - 220 , monosodium glutamate - 44, trisubstituted sodium citrate - 500, in glass ampoules sealed under vacuum. Ampoules with culture stored in the refrigerator at minus 10 ° C for 12 months.

Reseeding is carried out after 30 days. Preferably, 1 ml of milk fermented with this culture is added to 10 ml of sterile 12% (w / v) skim milk with the addition of 0.1 ml of yeast extract from a 20% solution. Then thermostat at 30 ° C for 18-20 hours until a clot forms. The grown culture is stored in a refrigerator at a temperature of 5-7 ° C and used to prepare a working crop at the stage of TP 1.2. Thus, the culture can be stored in a larger volume of 100-1000 ml.

To obtain a working seed crop, 450 ml of sterile milk prepared as previously indicated is aseptically added to a pre-sterilized flask with a capacity of 1 L. Next, 5 ml of a sterile 20% solution of yeast extract and 45 ml of a culture grown previously, whose age is not more than 10 days, are added to the flask. They are placed in a thermostat at 30 ° C and incubated for 18-20 hours until a clot forms. At this stage, you can also use brine from drying whey, whey itself, as well as washing equipment of the dairy industry in appropriate quantities, determined from the calculation of the need for nutrients.

To obtain a working sowing culture, in 3 previously sterilized flasks with a capacity of 2 l, aseptically add 1.2 l of sterile milk prepared previously. Then, 12 ml of a sterile 20% solution of yeast extract and 120 ml of freshly grown culture, the age of which is no more than 10 days, are added to the flasks. They are placed in a thermostat at 30 ° C and incubated for 18-20 hours until a clot forms. Ready inoculum in the amount of 4 l is aseptically transferred to a sterile flask with a tap.

The fermentation is carried out in a fermenter with a geometric volume of 70 l, having a mixing device that provides a mixing speed of up to 1000 rpm, a nozzle for supplying a nutrient medium, a nozzle for supplying seed, a sampler and a line for transferring culture fluid from the fermenter. The device has automatic systems for monitoring and maintaining pH and temperature.

Before loading the fermenter, it is checked for leaks in accordance with the instructions; in case of malfunctions, they are eliminated. After that, the apparatus is sterilized at 130 ° C for 1 h according to the instructions for use of the fermenter.

The fermentation medium is prepared by a fermenter using drinking water. The components are calculated per volume of medium 37.3 liters.

The nutrient medium is prepared from brine after drying whey, whey and rinses obtained after washing technological equipment at dairy enterprises, while the content of nutrients must correspond to their content in a mixture containing, g / l:

Skimmed milk powder thirty Dry whey 40

To the resulting composition add the enzyme megateria in an amount of 0.7-1.0 g / l

The dry enzyme is weighed on a laboratory balance in an amount of 26 g. Thoroughly dissolved in warm water at 40-45 ° C in a volume of 500 ml in an enameled container.

The dry components of the mixture are weighed on a balance, dissolved in an enameled container in 7-10 liters of warm - 40-45 ° C water, and mixed thoroughly. Liquid components are measured using measuring equipment and added to the indicated container. The suspension is poured into the apparatus through the filter material (gauze). The pH of the suspension is adjusted to 8.0 by introducing the previously obtained 10% alkali solution. This operation, like all subsequent ones related to adjusting the pH, is performed with constant stirring of 70-100 rpm.

In the fermenter, the pH is adjusted to 8.0 by introducing a 10% NaOH solution, the volume is brought up to 37.3 l with drinking water, heated to 50 ± 1 ° C by supplying steam to the apparatus jacket with constant stirring. After this, the enzyme is introduced, stirred for 20-30 minutes and hydrolysis is carried out for 2.5 hours, maintaining a constant temperature of 50 ± 1 ° C, by supplying steam to the jacket. After hydrolysis is complete, the medium is quickly heated and sterilization begins.

The medium is sterilized by supplying steam to the jacket, maintaining 25 minutes at a temperature of 113-114 ° C. After sterilization is complete, the medium is cooled to 29 ± 1 ° C by supplying water to the jacket. It is necessary to ensure the fastest possible heating before sterilization and cooling of the medium after sterilization. This is a prerequisite for obtaining a high-quality nutrient medium.

In a sterile medium, a pH of 7.0 is established by applying a sterile 10% NaOH solution from a glass bottle with a pump, the pH value is checked with a control pH meter, the temperature is brought to 28 ° C.

The flask with the inoculum is sterilely attached under a torch to the fermenter via a seed nozzle. Next, the inoculum is pressed with a peristaltic pump into the fermenter with constant stirring.

Fermentation is carried out while maintaining a temperature of 28 ± 1 ° C in the fermenter, overpressure, stirring 300 rpm, a constant pH of 6.8. Maintaining a constant pH level is carried out automatically by a pH-statization system, by supplying a sterile 10% NaOH solution from a glass bottle. In the fermentation process, temperature, pH, the amount of titrant consumed per hour (ml / h), the number of revolutions of the stirrer per minute, and the overpressure in the apparatus are noted every hour. Sterility samples are taken — in empty sterile tubes: after sterilization of the medium, after inoculation of the medium, 7 hours after the start of fermentation and after the completion of fermentation - only 4 samples; for activity of 9.5 ml of culture fluid in test tubes with 0.5 ml of hydrochloric acid: every hour, starting 2 hours after the maximum titration and until the end of fermentation, the samples are stored in the refrigerator. Test tubes with acid are pre-prepared: 0.5 ml of hydrochloric acid diluted 1: 1 with water are added to volumetric tubes, closed with rubber stoppers and put a mark at the level of 10 ml.

Fermentation is stopped when the titration rate drops to zero. The duration of the fermentation process is approximately 12-14 hours.

Fermentation is carried out in an apparatus with a geometric volume of 1 m ³ , with a mixing device that provides up to 180 rpm, a shirt with a supply of steam and cold water, a hatch for supplying a nutrient medium, a fitting for supplying seed, a sampler and a line for transferring culture fluid from fermenter. The device is equipped with an automatic pH control and maintenance system.

Before loading the fermenter, it is checked for leaks in accordance with the instructions; in case of malfunctions, they are eliminated. After that, the apparatus is sterilized at 130 ° C for 1 hour.

The fermentation medium is prepared using drinking water. The components are based on a volume of 505 l.

The nutrient medium is prepared from brine after drying whey, whey and rinses obtained after washing technological equipment at dairy enterprises, while the content of nutrients must correspond to their content in a mixture containing, g / l:

Skimmed milk powder thirty Dry whey 40

The dry components of the mixture are weighed on a balance, dissolved in an enameled container of warm - 40-45 ° C water, mixed thoroughly. Liquid components are measured using measuring equipment and added to the indicated container. The suspension is poured into the apparatus through the filter material (gauze). The pH of the suspension is adjusted to 8.0 by introducing the previously obtained 10% alkali solution. This operation, like all subsequent ones related to adjusting the pH, is performed with constant stirring of 70-100 rpm.

The loading line and the fermenter begin to be sterilized 1 hour before the end of hydrolysis of the medium. Sterilization is carried out at 131 ± 1 ° C for 1 h. After sterilization is completed, the steam supply to the loading line and to the fermenter is turned off. The fermenter is immediately cooled by supplying water to the jacket; when the pressure in the apparatus drops to 0.7 ± 0.2 ati, sterile air is supplied to maintain an excess pressure of 0.5 ati.

After hydrolysis is complete, the medium is quickly transferred to a sterile fermenter through filter material (gauze) and sterilization is started immediately by supplying steam to the jacket. Sterilization mode - exposure for 25 minutes at a temperature of 113-114 ° C. It is necessary to ensure the fastest possible heating before sterilization and cooling of the medium after sterilization. This is a prerequisite for obtaining a high-quality nutrient medium.

In a sterile medium, a pH of 7.0 is established by supplying a previously prepared 10% sterile NaOH solution, the pH value is checked with a control pH meter, the temperature is adjusted to 28 ° C, the overpressure is released, stirring is turned off.

Previously obtained seed along the seed line is sterilized under the torch and introduced into the fermenter through the seed nozzle. Next, the nozzle is closed, the stirrer is turned on, an overpressure in the apparatus of 0.5 ati is established by supplying sterile air.

Fermentation is carried out while maintaining a temperature of 28 ± 1 ° C, pH 6.8 in the apparatus by applying a sterile 10% NaOH solution along a sterile line, with stirring 180 rpm. In the process, temperature, pH, the number of clicks on the counter of the pH-stating device, and the pressure in the fermenter are noted every hour. Fermentation is stopped when the titration rate drops to 1/3 of the maximum amount of alkali consumed per hour. Sterility samples are taken — in empty sterile tubes: after sterilization of the medium, after inoculation of the medium, 7 hours after the start of fermentation and after the completion of fermentation - only 4 samples; for activity of 9.5 ml of culture fluid in test tubes with 0.5 ml of hydrochloric acid: every hour, starting 2 hours after the maximum titration and until the end of fermentation, the samples are stored in the refrigerator. Test tubes with acid are pre-prepared: 0.5 ml of hydrochloric acid diluted 1: 1 with water are added to volumetric tubes, closed with rubber stoppers and put a mark at the level of 10 ml.

The concentration of nisin in the culture fluid is 7000-7500 IU / ml. The duration of the fermentation process is 8-10 hours. The culture fluid is transferred to the extraction stage.

After the fermentation process is over, the culture fluid (QL) is acidified to a final pH of 2.5 with a solution of phosphoric acid. Then, the QOL is heated through a jacket (or simultaneously by supplying steam to the apparatus) to 100 ° C and held for 10 minutes. Cool to 30 ° C by applying cold water to the jacket and transfer to the separation (centrifugation) stage. The operation is carried out with stirring 180 rpm

After extraction, a culture liquid of 700 L is separated in a bactofuge. The feed rate of QOL from the extraction stage is determined experimentally by the quality of the centrate, achieving the cleanest fluid from the cells. Quality is evaluated visually. The centrate is collected in an ultrafiltration apparatus. The precipitate is collected in a neutralization and drying unit.

In a preferred embodiment of the developed technical solution, a centrate containing the target product, nisin, is successively passed through an ultrafiltration unit, a nanofiltration unit, and a nisin drying unit, while equipment used in the processing of milk and milk products can be used as an ultrafiltration unit, as a nanofiltration unit, equipment for the dairy industry can also be used, and equipment can be used for drying, before Assigning to obtain milk powder.

To the neutralization and drying unit, in addition to the bactofuge outlet, the sediment can be connected through the calcium lactate precipitation unit and the mother liquor evaporator unit, the permeate evaporator unit connected to the permeate exit of the nanofiltration unit. At the output of the waste neutralization and drying unit, an organo-mineral supplement for livestock and poultry feed is obtained.

The precipitate output of the calcium lactate precipitation unit can be connected to the inlet of the lactic acid solution production unit.

The developed technology and technological line ensure the industrial production of nisin, as well as the processing of waste products obtained in the production of nisin into useful products.

Claims (17)

1. The method of microbiological production of nisin, characterized in that the cultivation of the producer of nisin in the fermenter using a strain of the producer of nisin Lactococcus (Streptococcus) lactis, followed by extraction of nisin, and as a nutrient medium used brine after drying whey, containing up to 9, 0 g / dm ³ of mineral salts, and rinsing whey obtained after cleaning of process equipment in dairy plants and containing not more than 0.2% protein and megaterin cultured of conducted using an automated pH-statirovaniya and temperature control, extraction is carried out by acidification with nisin followed by isolation by centrifugation nisin. 2. The method according to p. 1, characterized in that the strain of Lactococcus lactis subsp. Lactis hybrid strain F-116, a collection of Moscow State University. V.M. Lomonosov. 3. The method according to p. 1, characterized in that the use of whey production with a protein content of not more than 1.4%. 4. The method according to p. 1, characterized in that the total content of milk protein in the nutrient medium is not more than 1.6%. 5. The method according to p. 1, characterized in that previously, in the process of cultivating the strain, a nutrient medium is used containing at least partially hydrolyzed granary waste, the nutrient content of which corresponds to their amount in 0.05 kg of dry yeast extract. 6. The method according to p. 1, characterized in that they use a nutrient medium containing 0.07-0.15% megaterina. 7. The method according to p. 1, characterized in that nisin is extracted by the action on the culture fluid with a solution of phosphoric acid at a pH of 2.4 ÷ 2.6 with stirring and subsequent heating to 100 ° C. 8. The method according to p. 1, characterized in that the fermentation is carried out, maintaining in the fermenter a temperature of 28 ± 1 ° C and a pH of 6.8, with stirring at a speed of 180 rpm 9. The technological line for the microbiological production of nisin according to claim 1, characterized in that it contains a fermenter with a spherical bottom and a lid, equipped with a bubbler, a shirt, a stirrer and an automatic pH-stating and thermostating system, the inputs of which are combined with the feed medium and seed systems material, and the output of the culture fluid is communicated through blocks of extraction, centrifugation, ultrafiltration and nanofiltration with the drying unit of the nisin preparation. 10. The line according to p. 9, characterized in that the fermenter further comprises a means of sampling the culture fluid. 11. The line according to p. 9, characterized in that it further comprises a site for the preparation of a nutrient medium. 12. The line according to claim 9, characterized in that it further comprises a seed preparation section. 13. The line according to p. 9, characterized in that the extraction unit is made in the form of a thermostatically controlled vessel equipped with a stirrer. 14. The line according to p. 9, characterized in that the separation unit is made in the form of bactofuga. 15. The line according to claim 9, characterized in that it further comprises a nisin purification unit, comprising an ultrafiltration unit and a nanofiltration unit connected in series to the outlet through the centrate of the separation unit. 16. The line according to p. 9, characterized in that it further comprises a unit for producing an organo-mineral additive connected to the sludge outlet of the separation unit. 17. The line according to claim 9, characterized in that it further comprises a lactic acid solution production unit connected to the permeate to the nanofiltration unit.

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