CN105274044A - Inactivated lactic acid bacterium cell and application thereof to food - Google Patents
Inactivated lactic acid bacterium cell and application thereof to food Download PDFInfo
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Abstract
The invention discloses an inactivated lactic acid bacterium and application thereof to food (including drinking water). The inactivated lactic acid bacterium is fermented and cultivated by the aid of culture media, which are made of food-grade raw materials, until later periods of log phases and is thermally inactivated and treated after the number of cells reaches a certain numerical value; the cells are separated from one another to obtain wet cell mud, namely, inactivated lactic acid bacterium cells, or the obtained wet cell mud is further dried to obtain dry powder of the inactivated lactic acid bacterium cells. The inactivated lactic acid bacterium and the application have the advantages that the prepared inactivated lactic acid bacterium cells can be used as food ingredients instead of active lactic acid bacteria, effects identical to effects of active lactic acid bacterium products can be completely realized, and the inactivated lactic acid bacterium has powerful healthcare functions; effects of effectively reducing and removing possible harmful ingredients such as mycotoxins, alga-toxin and heavy metal in food can be realized, and accordingly the safety of the food can be further guaranteed; the inactivated lactic acid bacterium is high in resistance to high-temperature, dry and radiation environments and the like when used as food or a food additive, and has excellent popularization and application prospects, and stable effects can be realized.
Description
Technical field
The invention belongs to technical field of food additives, edible microorganism.More specifically, a kind of deactivation lactic-acid bacteria cells is related to and the application of (comprise tap water, the present patent application book be referred to as " food ") in food.
Background technology
Living lactic acid bacteria goods separately as protective foods, or add in bread and cheese or infant or baby food as a kind of beneficial and are more and more admitted by industry and human consumer.Milk-acid bacteria not only can give the functional of food, as improved digestion level, improving intestinal health etc., can also eliminate the antinutritional factor in food.But in the course of processing of food, or in use procedure, food often can be in high temperature, dry environment, thus cause living lactic acid bacteria dead and lose activity.As added in milk powder or in formula milk, the temperature of people's water used when modified milk powder generally at 60 DEG C or more, even reaches boiling point (100 DEG C).At this temperature, living lactic acid bacteria can be killed rapidly, causes losing efficacy.
For solving this problem, some product is by living lactic acid bacteria cell bag quilt at present, even derma, to reach heat resistanceheat resistant, resist drying, radiation-resistant object.But this kind of mode makes production cost rise, and benefit is low, and popularizing application prospect is poor.
Summary of the invention
The technical problem to be solved in the present invention overcomes the defect of existing living lactic acid bacteria in food applications and technical deficiency, the production technique of a kind of deactivation milk-acid bacteria is provided, and substitute living lactic acid bacteria as food ingredients, reach effect that living lactic acid bacteria goods are same completely, very strong resistance is had to environment such as high temperature, drying and radiation, thus more stable; Its nourishing function of lactic-acid bacteria cells simultaneously after inactivation treatment of the present invention is stronger, and to (comprising tap water) in food, possible oxious component such as mycotoxin, algae toxin and heavy metal etc. have potent subduction effect, thus improve food safety further.
The object of this invention is to provide a kind of production technique of deactivation lactic-acid bacteria cells.
Another object of the present invention is to provide the application of deactivation milk-acid bacteria in food (comprising tap water) that above-mentioned production technique obtains, except effect of general probiotic bacterium, the mycotoxin in food, algae toxin and heavy metal can also be reduced, thus invention also provides a kind of method of mycotoxin, algae toxin and heavy metal in subduction food completely newly, make an addition in food (comprising tap water) by the deactivation milk-acid bacteria prepared by the present invention.
Above-mentioned purpose of the present invention is achieved through the following technical solutions:
A kind of deactivation lactic-acid bacteria cells, obtains by the following method:
S1. by milk-acid bacteria liquid fermentation and culture to the logarithmic phase later stage, cell count reaches 10
8cFU/mL or 10
8during more than CFU/mL, carry out hot inactivation treatment;
S2. cell and separation of fermentative broth obtain wet cell mud, are deactivation lactic-acid bacteria cells;
Or step S2 is that cell and separation of fermentative broth obtain wet cell mud, obtains deactivation lactic-acid bacteria cells dry powder after drying.
Further preferably, add protective material in the wet cell mud that above-mentioned steps S2 obtains, vacuum lyophilization or 70 ~ 75 DEG C of vacuum-dryings (being preferably 75 DEG C of vacuum-dryings), obtain deactivation lactic-acid bacteria cells dry powder.
Preferably, described protective material is trehalose or skimming milk etc.
Preferably, described in step S1, cell count reaches 10
8~ 10
10cFU/mL.
More preferably, described in step S1, cell count reaches 10
9cFU/mL.
In addition, the substratum that above-mentioned steps S1 fermentation culture milk-acid bacteria is used all adopts food grade materials to prepare.
Preferably can embodiment as one, described in step S1, the formula of fermentation culture used medium is: based on MRS substratum, adds vegetables juice and originates as somatomedin, then add calcium carbonate.
Preferably, in often liter of MRS substratum, add vegetables juice 10 ~ 50mL, add calcium carbonate 1 ~ 2g.
Preferably, described vegetables juice is Radix Dauci Sativae juice or tomato juice.
Fermentation described in step S1 is constant temperature or temperature-variable fermentation.Preferably can embodiment as one, described in step S1, the technique of fermentation culture is as follows:
S11. when temperature 38 ~ 40 DEG C, milk-acid bacteria seed is accessed liquid nutrient medium, cultivate 4 hours;
S12. after, broth temperature is reduced to 30 ~ 37 DEG C, maintains 6 hours;
S13. temperature is raised to 40 ~ 45 DEG C again and cultivates more than 2 hours or 2 hours again, make cell count reach 10
9cFU/mL or 10
9more than CFU/mL.
Preferably, milk-acid bacteria seed is accessed liquid nutrient medium when temperature 38 DEG C by step S11.
Preferably, broth temperature is reduced to 35 DEG C by step S12.
Preferably, temperature is raised to 40 DEG C and cultivates by step S13.
More preferably, in above fermentation culture process, fermentor tank passes into nitrogen, and keeps static or stirring at low speed.
Further preferably, described hot inactivation treatment is after fermentation culture terminates and fermented liquid is warming up to 60 ~ 120 DEG C of maintenances 5 ~ 40 minutes, and stirs.
More preferably, hot inactivation treatment is adopted after fermentation culture described in step S1.
More preferably, described hot inactivation treatment fermented liquid is warming up to 60 DEG C to maintain 30 minutes, and stir.
Preferably, being separated described in step S2 is centrifugal 10 ~ 20 minutes of 8000 ~ 12000rpm, collects thalline.More preferably centrifugal 15 minutes of 10000rpm.
In addition, the bacterial strain that the described deactivation lactic-acid bacteria cells of above-mentioned production uses is probiotic bacterium, belong to the edible bacterial strain that national health and Family Planning Committee announce, comprise the lactic bacterium strains that can be used for food, the lactic bacterium strains that can be used for infant or baby food or can be used for protective foods milk-acid bacteria bacterial strain in one or more.
Preferably, as one or more in Bacterium lacticum, galactococcus or bifidus bacillus.
Specifically preferably, as Lactobacterium acidophilum, bifidus bacillus, lactobacillus salivarius, lactobacillus rhamnosus or thermophilus streptococcus etc.
Cell count contained by the deactivation lactic-acid bacteria cells dry powder utilizing aforesaid method of the present invention to prepare is 10,000,000,000/gram ~ 1,000 hundred million/gram.
Above-mentioned deactivation lactic-acid bacteria cells is as the application of food or the application that makes an addition to as food ingredients in food, or the application in processing drinking water, also all should within protection scope of the present invention.
In a kind of subduction food completely newly, the method for mycotoxin, algae toxin and heavy metal is also within protection scope of the present invention, and described method makes an addition in food (comprising tap water) by the deactivation milk-acid bacteria prepared by the present invention.
Wherein, the type of described deactivation milk-acid bacteria for determine to belong to probiotic bacterium, and is ratified to audit by national health and Family Planning Committee and can directly to be eaten or for one or more in the bacterial strain of food fermentation food-processing.Preferably, as one or more in Bacterium lacticum, galactococcus or bifidus bacillus.
Specifically preferably, make an addition to as food ingredients apply in food time, the mass ratio of described deactivation lactic-acid bacteria cells and food is 0.5 ~ 1.0:1000 ~ 2000, wherein, cell count contained by described deactivation milk-acid bacteria is 10,000,000,000/gram ~ 1,000 hundred million/gram, be preferably 10,000,000,000/gram.
In addition, described deactivation lactic-acid bacteria cells, when processing drinking water, is make an addition in tap water by deactivation lactic-acid bacteria cells, the oxious component in planar water, centrifugal or filter removes the cell in drinking water and the oxious component that adsorbs thereof afterwards.
Preferably, described oxious component is mycotoxin, algae toxin and/or heavy metal.Preferably, described heavy metal is plumbous or cadmium, but is not limited to this two heavy metal species.
Further, make an addition to as food ingredients apply in food time, described food refer to can high temperature, radiation and/or drying conditions process food.Described food can be any base edible product, preferably includes milk-product, soya products, rice food, flour product, corn product, fruit preserves, other cereal and vegetable products, candy, beverage or tap water etc.
Further preferably, described milk-product comprise ordinary powdered milk, baby formula milk powder, liquid milk (any packaging), cheese or other food made containing animal milk composition;
Described soya products comprises the food such as soya-bean milk, bean curd, soya-bean milk, fermented bean products (as soy sauce, sauce, fermented soya bean), soybean oil or the eight treasure gruel made primarily of beans;
Other food etc. that described rice food comprises rice, ground rice, rice and flour, rice cake, rice cake or makes primarily of rice;
Described flour product comprises bread, noodles, face cake or is other food that main raw material is made with flour;
Described corn product comprises corn-dodger, maize gruel, Semen Maydis powder, corn crisp or is other food that main raw material is made with corn;
Described fruit preserves comprises fruit juice, preserved fruit or preserved fruit etc., and is not limited to the above food;
Other cereal described and vegetable products comprise the food of other cereal such as peanut butter, peanut oil, seed sesame paste, seed sesame oil and plant processing, but are not limited to above kind.
The present invention, by large quantity research and exploration, provides the production technique of a kind of deactivation milk-acid bacteria, and the deactivation lactic-acid bacteria cells prepared can be used as food ingredients and substitutes living lactic acid bacteria, reaches effect that living lactic acid bacteria goods are same, and ensures safety; The more important thing is, more stable, there is very strong resistance to environment such as high temperature, drying and radiation; We also find simultaneously, its nourishing function of lactic-acid bacteria cells after inactivation treatment of the present invention is stronger, to oxious component possible in food as the potent subduction effect as plumbous cadmium etc. has of mycotoxin, algae toxin and heavy metal, thus effectively improve the security of food.
The present invention has following beneficial effect:
(1) the present invention cultivates in the culture medium prescription that milk-acid bacteria adopts and all adopts food grade materials, the deactivation lactic-acid bacteria cells prepared substitutes living lactic acid bacteria as food ingredients, while reaching the same effect of living lactic acid bacteria goods, can ensure food safety utterly, this is also the primary guarantee of the object of the invention.
(2) the present invention is only using the cell of deactivation milk-acid bacteria as effective ingredient, only utilizes the milk-acid bacteria thalline after deactivation (i.e. the cell of deactivation), does not comprise fermentating metabolism product.We find, deactivation lactic-acid bacteria cells prepared by the present invention just can promote the growth of the intrinsic milk-acid bacteria of people's enteron aisle, and need not its fermentating metabolism product; To the objectionable constituent that may exist in food, as mycotoxin (as zearalenone), algae toxin and heavy metal lead and cadmium etc., also there is potent subduction effect, this subduction effect is the surface adsorption effect relying on as killed cells, thus the present invention also provides a kind of method of mycotoxin, algae toxin and heavy metal in subduction food completely newly simultaneously, make an addition in food (comprising tap water) by the deactivation milk-acid bacteria prepared by the present invention, food safety can be ensured further.
And the present invention only utilizes the deactivation lactic-acid bacteria cells not comprising fermentating metabolism thing, and on the one hand, cell concn is higher, in food, usage quantity is less, does not take food formulations space; On the other hand, the pure as killed cells removed after meta-bolites is almost tasteless, therefore can not change food or drink water original local flavor and nutrition.
(3) the deactivation lactic-acid bacteria cells for preparing of the present invention, the promotion intestinal health that can not only bring into normal play, stablize outside microecological balance, immunoregulation effect, more stress the promoter action to lactobacter growth intrinsic in people's enteron aisle, and to the absorption of the mycotoxin that may exist in food, algae toxin and heavy metal and removal effect, improve the safety of food.
So deactivation lactic-acid bacteria cells of the present invention can be applied to any food, comprise various beverage, be particularly useful for dairy products, bean product, rice made products, flour products and tap water etc. by the food of mycotoxin, algae toxin and heavy metal contamination, therefore to have great importance.
(4) the deactivation lactic-acid bacteria cells prepared of the present invention is as food or when using as foodstuff additive, compared with living lactic acid bacteria, use more simple, its performance is more stable, there is very strong resistance to environment such as high temperature, drying and radiation, there is good popularizing application prospect.
Accompanying drawing explanation
Fig. 1 is that high temperature, uv irradiating and drying treatment are on the impact of milk-acid bacteria as killed cells on Caco-2 cell adhesion.
Embodiment
Further illustrate the present invention below in conjunction with specific embodiment, but embodiment does not limit in any form to the present invention.Unless stated otherwise, the present invention adopts reagent, method and apparatus are the art conventional reagent, method and apparatus.Unless stated otherwise, agents useful for same of the present invention and material are commercial.
embodiment 1 prepares deactivation milk-acid bacteria
1, deactivation lactic-acid bacteria cells preparation method is as follows:
S1. ordinary method prepares milk-acid bacteria seed, when temperature 38 DEG C, milk-acid bacteria is accessed fermention medium, cultivates 4 hours; Temperature is reduced to 35 DEG C, maintains 6 hours; Again temperature is raised to 40 DEG C to cultivate 2 hours; In above culturing process, fermentor tank passes into nitrogen, and keeps static or stirring at low speed;
Treat that lactobacter growth is to the logarithmic phase later stage, cell count reaches 10
8~ 10
10during CFU/mL, carry out hot inactivation treatment, specifically fermented liquid is warming up to 60 DEG C and maintains 30 minutes, and stir, and guarantee that all cells is inactivated;
Centrifugal 15 minutes of S2.10000rpm, collects thalline and obtains wet cell mud, be milk-acid bacteria as killed cells.
Wherein, step S1 milk-acid bacteria used is Lactobacterium acidophilum.
Described in step S1, the formula of fermentation culture used medium is: based on MRS substratum, adds tomato juice and originates as somatomedin, then add calcium carbonate.Wherein, in often liter of MRS substratum, add tomato juice 10mL, add calcium carbonate 2g.
2, cell count contained by the deactivation lactic-acid bacteria cells dry powder utilizing aforesaid method to prepare can reach 10,000,000,000/gram ~ 1,000 hundred million/gram or more.
embodiment 2 prepares deactivation milk-acid bacteria
1, deactivation lactic-acid bacteria cells preparation method is as follows:
S1. ordinary method prepares milk-acid bacteria seed, when temperature 38 DEG C, milk-acid bacteria is accessed fermention medium, cultivates 4 hours; Temperature is reduced to 30 DEG C, maintains 6 hours; Again temperature is raised to 40 DEG C to cultivate 2 hours; In above culturing process, fermentor tank passes into nitrogen, and keeps static or stirring at low speed;
Treat that lactobacter growth is to the logarithmic phase later stage, cell count reaches 10
8~ 10
10during CFU/mL, carry out hot inactivation treatment, specifically fermented liquid is warming up to 80 DEG C and maintains 20 minutes, and stir, and guarantee that all cells is inactivated;
Centrifugal 15 minutes of S2.10000rpm, collects thalline and obtains wet cell mud, be milk-acid bacteria as killed cells.
Wherein, step S1 milk-acid bacteria used is bifidus bacillus.
Described in step S1, the formula of fermentation culture used medium is: based on MRS substratum, adds Radix Dauci Sativae juice and originates as somatomedin, then add calcium carbonate.Wherein, in often liter of MRS substratum, add Radix Dauci Sativae juice 30mL, add calcium carbonate 1.5g.
2, cell count contained by the deactivation lactic-acid bacteria cells dry powder utilizing aforesaid method to prepare can reach 10,000,000,000/gram ~ 1,000 hundred million/gram or more.
embodiment 3 prepares deactivation milk-acid bacteria
1, deactivation milk-acid bacteria preparation method is as follows:
S1. ordinary method prepares milk-acid bacteria seed, when temperature 40 DEG C, thermophilus streptococcus is accessed substratum, cultivates 4 hours; Temperature is reduced to 37 DEG C, maintains 6 hours; Again temperature is raised to 45 DEG C to cultivate 2 hours;
Now, streptococcus thermophilus fermentation is cultured to the logarithmic phase later stage, and cell count reaches 10
8~ 10
10after CFU/mL, 100 DEG C process 10 minutes;
S2. at 10000rpm centrifugal 15 minutes, collect thalline and obtain wet cell mud, be deactivation thermophilus acid mycetocyte.
Described in step S1, the formula of fermentation culture used medium is: based on MRS substratum, adds Radix Dauci Sativae juice and originates as somatomedin, then add calcium carbonate.Wherein, in often liter of MRS substratum, add Radix Dauci Sativae juice 50mL, add calcium carbonate 1g.
2, cell count contained by the thermophilus streptococcus as killed cells dry powder utilizing aforesaid method to prepare is 10,000,000,000/gram ~ 1,000 hundred million/gram.
embodiment 4 prepares deactivation milk-acid bacteria
1, deactivation milk-acid bacteria preparation method is as follows:
S1. ordinary method prepares milk-acid bacteria seed, when temperature 38 DEG C, milk-acid bacteria is accessed fermention medium, cultivates 4 hours; Temperature is reduced to 35 DEG C, maintains 6 hours; Again temperature is raised to 40 DEG C to cultivate 2 hours; In above culturing process, fermentor tank passes into nitrogen, and keeps static or stirring at low speed;
Now, lactobacillus-fermented is cultured to the logarithmic phase later stage, and cell count reaches 10
8~ 10
10cFU/mL, carries out hot inactivation treatment, specifically fermented liquid is warming up to 60 DEG C and maintains 30 minutes, and stir, and guarantee that all cells is inactivated;
S2. at 10000rpm centrifugal 15 minutes, collect thalline and obtain wet cell mud, add protective material in wet cell mud, vacuum lyophilization or 75 DEG C of vacuum-dryings, obtained deactivation lactic-acid bacteria cells dry powder.
Wherein, step S1 milk-acid bacteria used is lactobacillus rhamnosus or lactobacillus salivarius.
The formula of fermentation culture used medium described in step S1 is with embodiment 2.
Protective material described in step S2 is trehalose and skimming milk.
2, cell count contained by the lactobacillus rhamnosus utilizing aforesaid method to prepare or lactobacillus salivarius as killed cells dry powder is 10,000,000,000/gram ~ 1,000 hundred million/gram.
embodiment 5 prepares deactivation milk-acid bacteria
1, deactivation milk-acid bacteria preparation method is as follows:
S1. when temperature 40 DEG C, milk-acid bacteria is accessed substratum, cultivate 4 hours; Temperature is reduced to 37 DEG C, maintains 6 hours; Again temperature is raised to 45 DEG C to cultivate 2 hours;
Now, lactobacillus-fermented is cultured to the logarithmic phase later stage, and cell count reaches 10
9cFU/mL, carries out hot inactivation treatment, specifically fermented liquid is warming up to 60 DEG C and maintains 30 minutes, and stir;
S2. at 10000rpm centrifugal 15 minutes, collect thalline and obtain wet cell mud, add protective material in wet cell mud, vacuum lyophilization or 70 DEG C of vacuum-dryings, obtained deactivation lactic-acid bacteria cells dry powder.
Wherein, step S1 milk-acid bacteria used is Lactobacterium acidophilum.
The formula of fermentation culture used medium described in step S1 is with embodiment 1.
Protective material described in step S2 is skimming milk.
2, cell count contained by the deactivation lactic-acid bacteria cells dry powder utilizing aforesaid method to prepare is 10,000,000,000/gram ~ 1,000 hundred million/gram.
embodiment 6 deactivation lactic-acid bacteria cells is to intestinal health and somatotrophic effect
1, take weanling pig as animal model, the deactivation lactic-acid bacteria cells of research prepared by the present invention adds in feed or food, to animal and human's body intestinal health and somatotrophic effect.Test arranges as follows:
(1) subjects: eutocous 20 sow litters (average 10/nest) are experiment piglet, count 60 sows and produce 600 piglets.According to the childbirth situation of sow, and the size of the pig that farrows and healthy state, select as far as possible consistent piglet 450, be divided into 3 groups at random.
Experiment totally 3 batches, results averaged.
(2) test combinations:
Control group: daily ration based on 311 material of feeding.
Experimental group 1: add the Heat-killed Bifidobacterium On Microflora cell prepared by 0.5kg/t embodiment 2 on basal diet.
Experimental group 2: add the deactivation lactobacillus acidophilus cells prepared by 0.5kg/t embodiment 5 at basal diet.
(3) daily feeding and management is formulated by company of raising pigs " pig farm feeding and management handbook " carries out, free choice feeding, fully drinks water, and conventional epidemic prevention, often sterilizes.Every day reinforced four times of piglet, every day feeds by standard weighing, and the next morning reclaims residue material and weighs, every day observed and recorded piglet diarrhoea situation, piglet searches for food and healthy state.
Experimental data: the record condition of production of sow, piglet Birth weight, 26 day age weaning weight, 35 day age be heavy; By day entry addition material amount and residue doses, record diarrhoea head number and each stage respectively organize weight gain of piglets, analyze Incidence of Diarrhea.
2, the data of test-results statistics are as shown in table 1.
Table 1
From table 1 data, in feed, add Heat-killed Bifidobacterium On Microflora prepared by the present invention and deactivation lactobacillus acidophilus cells can significantly reduce grice diarrhoea rate, test 1,2 group and reduce 88.18% and 87.64% than control group diarrhea rate respectively.
And experimental group piglet healthy state is better, and the colour of skin is ruddy, good bloom, and within 35 days, survival rate is to 98.14% and 96.85%, improves 7.5% and 6.3% respectively than control group.
Average daily gain is analyzed, and from birth to 35 ages in days, control group average daily gain is 186 grams/day/head, and experimental group 1 and 2 is respectively 225 grams/day/and 219 grams/day/head, and experimental group 1 and 2 average daily gain improves 17.3% and 15.1% than control group respectively.
The price of deed is analyzed, and from the 25 to the 35 day, the daily material consumption of control group, experimental group 1 and 2 was respectively 286.44 grams/day/, 227.25 grams/day/, 238.71 grams/day/head, and feedstuff-meat ratio is respectively 1.54,1.01,1.09, and experimental group is significantly better than control group.
3, above result shows, in pig starter feed, add deactivation bifidus of the present invention and deactivation lactobacillus acidophilus cells significantly can improve sucking pig intestinal health situation, suppress pathogenic bacterium in GI breeding, thus reduce diarrhoea, also can improve food conversion ratio, improve weightening finish.
Demonstrated by the animal model of this test, deactivation milk-acid bacteria (bifidus and the Lactobacterium acidophilum) cell utilizing method of the present invention to prepare can substitute living lactic acid bacteria as food ingredients, reach effect that living lactic acid bacteria goods are same, promote humans and animals intestinal health, stablize microecological balance, immunity moderation effect, promotion growth of animal; Meanwhile, food safety can be ensured.
embodiment 7 deactivation milk-acid bacteria is to the subduction effect of objectionable impurities in food
1, the aflatoxin in milk is removed
The deactivation lactobacillus acidophilus cells 0.1g that respectively prepared by Example 1 and 5 joins 10mL and contains 3 μ g/L aflatoxin (AFB
1) milk in, mix, be placed in 30 DEG C, after shaking 1h under the condition of 200r/min, the supercentrifuge of 3000r/min is centrifugal, gets the content that supernatant liquor utilizes high effective liquid chromatography for measuring aflatoxin.
Result shows, and records AFB after process
1concentration is 0.876 μ g/L, and adsorption rate reaches 70.8%.
2, the zearalenone in flour is removed
Deactivation lactobacillus salivarius 0.005g prepared by Example 4 joins 0.5g and contains in the edible flour of 3 μ g/kg zearalenones (ZEA), after above-mentioned flour is mixed with deactivation milk-acid bacteria, add the distilled water of 10mL, as for 30 DEG C, after shaking 1h under the condition of 200r/min, the supercentrifuge of 3000r/min is centrifugal, gets the content that supernatant liquor utilizes high effective liquid chromatography for measuring zearalenone.
Result shows, and recording zearalenone concentration after process is 1.305 μ g/L, and adsorption rate reaches 73.9%.
3, the lead in tap water is removed
It is in the tap water of 10mg/L that deactivation lactobacillus rhamnosus 0.02g prepared by Example 4 joins 10mL lead content, is shaken up by deactivation lactic-acid bacteria cells in water, and the method with reference to GB7475-1987 measures content plumbous in water.
Result shows, and do not detect plumbous existence after process, adsorption rate reaches 100%.
4, the cadmium in tap water is removed
It is in the tap water of 1mg/L that deactivation thermophilus streptococcus 0.03g prepared by Example 3 joins 10mL cadmium content, is shaken up by deactivation lactic-acid bacteria cells in water, and the method with reference to GB7475-1987 measures the content of cadmium in water.Recording cadmium concentration after process is 0.05mg/L, and adsorption rate reaches 95.0%.
5, the algae toxin in tap water is removed
It is in the tap water of 150 μ g/L that each 0.1g of Heat-killed Bifidobacterium On Microflora+embodiment 1 or 5 preparation deactivation Lactobacterium acidophilum prepared by Example 2 joins 10mL Microcystins, mix, as for 30 DEG C, after shaking 4h under the condition of 175r/min, the supercentrifuge of 11000g is centrifugal, get supernatant liquor, utilize the content of high effective liquid chromatography for measuring Microcystin.
Result shows, and recording Microcystins Concentration after process is 92.25 μ g/L, and adsorption rate is 38.5%.
embodiment 8 deactivation lactic-acid bacteria cells is to the tolerance of temperature, radiation and drying
1, treatment process:
S1. respectively by lactobacillus bulgaricus, thermophilus streptococcus, Lactobacterium acidophilum or lactobacillus rhamnosus fermentation culture respectively;
S2. fermented liquid is warmed up to 60 DEG C and maintains 30 minutes, hot deactivation, collected by centrifugation as killed cells, brine cell 2 ~ 3 times, obtains as killed cells bacterium mud; Vacuum-drying obtains milk-acid bacteria as killed cells dry powder, moisture about 10%;
S3. S2 as killed cells dry powder is resuspended in physiological saline, carries out following two kinds of process respectively:
(1) 100 DEG C of water bath processing 10min(pyroprocessing);
(2) 5mL re-suspension liquid bacterium liquid is placed in glass culture dish, and 5min is irradiated at distance 30W ultraviolet lamp 5cm place, carries out ultraviolet process;
S4. respectively by the bacterium liquid after pyroprocessing and ultraviolet process 70 DEG C of vacuum-dryings, final moisture, below 5%, places testing product performance after 30d.
2, the deactivation milk-acid bacteria after above-mentioned high temperature, uv irradiating and drying treatment is to the characterization of adsorption of human colon adenocarcinoma cell line Caco-2 cell strain
(1) cultured Caco-2 cell strain is digested, make 10
5the cell suspension of individual/mL, hatches to unicellular adherent for 37 DEG C in cell culture incubator, adds the deactivation lactic-acid bacteria cells liquid of above-mentioned each process gained, makes bacterium number be about 10
7individual/mL, mixes deactivation lactic-acid bacteria cells liquid with Caco-2 cell culture fluid, 37 DEG C of 5%CO
2hatch 2h in-95% air, the lactic acid bacteria number that microscopic counting Caco-2 cell sticks, what calculate deactivation lactic-acid bacteria cells sticks index (stick index=stick total count * 100% in bacterium number/visual field).
(2) result as shown in Figure 1, milk-acid bacteria as killed cells after 100 DEG C of pyroprocessing, 100 DEG C of pyroprocessing+drying treatment, uv irradiating process, uv irradiating process+drying treatment is sticked index and is respectively 45.7%, 40.1%, 51.5%, 47.4%, variant between each group, but difference is not remarkable; Same, individual group also not obvious with the difference of control group, illustrate that the deactivation milk-acid bacteria utilizing method of the present invention to prepare can have very strong resistance to environment such as high temperature, drying and radiation, effect is more stable, than viable cell more resistant against high temperatures, radiation and drying conditions, performance is more reliable and more stable, and the application in food is more extensive.
3, the deactivation milk-acid bacteria after above-mentioned high temperature, uv irradiating and drying treatment is to AFB
1removal effect
(1) 100 DEG C of high temperature+drying treatment, uv irradiating+drying treatment is got respectively) the deactivation lactic-acid bacteria cells 0.1g of gained joins in the milk of 10mL aflatoxin (AFB1) content 3 μ g/L, mix, as for 30 DEG C, after shaking 1h under the condition of 200r/min, the supercentrifuge of 3000r/min is centrifugal, gets the content that supernatant liquor utilizes high effective liquid chromatography for measuring aflatoxin.
(2) result display, records AFB1 adsorption rate and is respectively 55.3% and 61.5% after process.With deactivation milk-acid bacteria before treatment to compared with AFB1 adsorption rate, have and slightly decline, but still can remain on more than 55%.Show that the impacts of condition on deactivation milk-acid bacteria prepared by the present invention such as high temperature, radiation and drying are little.
The present invention includes but be not limited to test example to the result drawn, the variation of all non-essential within theory of the present invention is all in protection scope of the present invention.
Claims (10)
1. a deactivation lactic-acid bacteria cells, is characterized in that, preparation method is as follows:
S1. by milk-acid bacteria liquid fermentation and culture to the logarithmic phase later stage, cell count reaches 10
8cFU/mL or 10
8during more than CFU/mL, carry out hot inactivation treatment;
S2. cell and separation of fermentative broth obtain wet cell mud, are deactivation lactic-acid bacteria cells;
Or step S2 is that cell and separation of fermentative broth obtain wet cell mud, obtains deactivation lactic-acid bacteria cells dry powder after drying.
2. deactivation lactic-acid bacteria cells according to claim 1, it is characterized in that, add protective material in the wet cell mud obtained, vacuum lyophilization or 70 ~ 75 DEG C of vacuum-dryings, obtain deactivation lactic-acid bacteria cells dry powder.
3. deactivation lactic-acid bacteria cells according to claim 1, it is characterized in that, described in step S1, the formula of fermentation culture used medium is: based on MRS substratum, adds vegetables juice and originates as somatomedin, then add calcium carbonate.
4. deactivation lactic-acid bacteria cells according to claim 1, it is characterized in that, described in step S1, the technique of fermentation culture is as follows:
S11. when temperature 38 ~ 40 DEG C, milk-acid bacteria seed is accessed liquid nutrient medium, cultivate 4 hours;
S12. after, broth temperature is reduced to 30 ~ 37 DEG C, maintains 6 hours;
S13. temperature is raised to 40 ~ 45 DEG C again and cultivates more than 2 hours or 2 hours again, make cell count reach 10
9cFU/mL or 10
9more than CFU/mL.
5. deactivation lactic-acid bacteria cells according to claim 1, is characterized in that, hot inactivation treatment described in step S1 is and after fermentation culture terminates, fermented liquid is warming up to 60 ~ 120 DEG C and maintains 5 ~ 40 minutes, and stirs; Cell described in step S2 and separation of fermentative broth are centrifugal 10 ~ 20 minutes of 8000 ~ 12000rpm, collect thalline.
6. according to the arbitrary described deactivation lactic-acid bacteria cells of Claims 1 to 5, it is characterized in that, producing the bacterial strain that described deactivation lactic-acid bacteria cells uses is probiotic bacterium, belong to the edible bacterial strain that national health and Family Planning Committee announce, comprise the lactic bacterium strains that can be used for food, the lactic bacterium strains that can be used for infant or baby food or can be used for protective foods milk-acid bacteria bacterial strain in one or more.
7. the arbitrary described deactivation lactic-acid bacteria cells of claim 1 ~ 6 is in the application as food or the application that makes an addition to as food ingredients in food, or the application in processing drinking water.
8. apply according to claim 7, it is characterized in that, make an addition to as food ingredients apply in food time, the mass ratio of described deactivation lactic-acid bacteria cells and food is 0.5 ~ 1.0:1000 ~ 2000, cell count contained by described deactivation milk-acid bacteria is 10,000,000,000/gram ~ 100,000,000,000/gram.
9. apply according to claim 7, it is characterized in that, described deactivation lactic-acid bacteria cells, when processing drinking water, is make an addition in tap water by deactivation lactic-acid bacteria cells, oxious component in planar water, centrifugal or filter removes the cell in drinking water and the oxious component that adsorbs thereof afterwards.
10. apply according to claim 7, it is characterized in that, make an addition to as food ingredients apply in food time, described food refer to can high temperature, radiation and/or drying conditions process food.
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| CN108968025A (en) * | 2018-06-13 | 2018-12-11 | 江苏新申奥生物科技有限公司 | Adjust the high concentration lactic acid bacteria freeze drying powder of blood pressure and blood lipoid |
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