JP2009273450A - Composition comprising dehydrated cell and hydrophobic solvent - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a composition with cells kept dormant or invariant, mitigating pollinosis through highly depositing and adsorbing allergens such as pollen thereonto, to provide an emulsified product thereof, and to provide methods for producing the composition and emulsified product. <P>SOLUTION: The composition is such as to be obtained by melting dehydrated cells into a hydrophobic solvent. The emulsified product thereof is also provided. Besides, the respective methods for producing the composition and emulsified product are provided. They can be used as food, cosmetics or medical supplies. In the above, the dehydrated cells can be kept dormant by making them lyophilized cells. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a composition obtained by dissolving / melting dehydrated cells in a hydrophobic solvent, an emulsified composition thereof, a production method, and application. More specifically, the present invention relates to a composition, an emulsified composition and a production method in which living cells or microorganisms from which moisture has been removed by freeze-drying or the like are dissolved / melted in a hydrophobic solvent, and foods and cosmetics containing them. And medical supplies.

  Since BC, there have been various fermented foods made from microorganisms, and fermented milk made from milk has been around the world. In recent years, lactic acid bacteria and bifidobacteria are biotic bacteria that are considered to be good for human health such as intestinal regulation, immune activity, antimutagenicity, and cholesterol-lowering action, and fermented foods using these bacteria have been developed. Has been. The biotic lactic acid bacteria used in these fermented foods, such as acidophilus bacteria and casei bacteria derived from the human intestinal tract, produce a strong diacetyl odor as a fermentation odor and are not preferred by the Japanese due to an unpleasant odor. was there. Bifidobacteria, in particular, have an effect of regulating the human intestine and have a positive effect on health, but it is anaerobic and is not suitable for growth and storage in an oxygen-containing environment. If anaerobic, it produces acetic acid in addition to lactic acid. However, there is a drawback that it cannot be said that it is a preferable bacteria in terms of flavor.

  In order to solve this problem, it has been studied to put functional lactic acid bacteria in enteric capsules for direct drinking without being used for food fermentation (Patent Document 1). Capsules that can be propagated have also been studied (Patent Document 2). Moreover, the thing regarding the liquid food containing the lactic-acid-bacteria granule coated with vegetable fat is examined (patent document 3). However, these have shapes such as hard and soft capsules, have a sense of incongruity as foods such as normal fermented milk and beverages, and have problems different from the original fermented foods such as yogurt and drink yogurt. Also, by inoculating the anaerobic Bifidobacterium (Bifidobacterium longum) in a reduced skim milk medium with no growth-promoting substance added and culturing it, it is proliferative even under aerobic conditions. Breeding of strains with high culture coagulation properties are being studied (Patent Document 4). However, this method has a problem of time loss, poor probability and generation of acetic acid.

  Therefore, even if it is an excellent functional bacterium but its fermentation odor is invincible in flavor, it can be used in aerobic environments with fermented foods with a delicious flavor and anaerobic bacteria, and does not produce acetic acid. There is a demand for a technique for producing a delicious product that can be used for a food containing bifidobacteria having a good flavor and is beneficial to human health.

  In addition to food, in Japan, many people suffer from hay fever, which is suffering from tears, runny nose, sneezing and stuffy nose in the spring, and is now being described as a national disease. It is known that allergic reactions such as hay fever are caused by allergens, which are proteins eluted by the action of moisture in pollen attached to the mucosal surfaces of the nose and eyes. In order to alleviate this symptom, various drugs have been developed and provided. However, the antihistamine agent has a side effect such as drowsiness and dry mouth, and the steroid agent has a side effect as a steroid. Masks are used to prevent the inhalation of pollen, which is a causative agent for allergies, but they are troublesome such as breathing when breathing, and have the disadvantage of having to be removed when eating.

  In order to solve this problem, a long chain hydrocarbon petrolatum and a liquid paraffin mixture are applied to the nasal cavity wall to adhere pollen (Patent Document 5). However, there is a problem that even if pollen may adhere, it flows out due to its fluidity and releases allergens when it meets body fluids. Therefore, when pollen reaches the hydrophobic solvent, it has an adsorptive power that keeps it staying in the substance, thereby reducing the amount of pollen reaching the mucous membrane at the back of the nose, and forming a film on the pollen to make contact with body fluids. There is a need for a technique that prevents allergens from occurring.

Foods useful for human health, cosmetics such as skin water retention, and medical supplies that reduce hay fever are a wide range of products, but there are compositions, emulsions, and manufacturing methods that can be used as raw materials for these products. It has been demanded.
Japanese Patent Laid-Open No. 8-242663 JP 2001-245660 A International Publication Number WO2005 / 070221 Japanese Patent Publication No. 2000-245440 JP 2005-314111 A

  Therefore, a wide range of uses such as food, cosmetics and medical supplies can be made, dehydrated cells can be dissolved / melted in a hydrophobic solvent to keep them dormant or unchanged, or allergens such as pollen can be added. There is a need for compositions and emulsions that strongly adhere and adsorb and reduce hay fever, and a method for their production. The medical supplies of the present invention include drugs, quasi drugs, miscellaneous goods related to prevention, and the like.

The first invention relates to a composition comprising dehydrated cells dissolved / melted in a hydrophobic solvent.
The second invention relates to a composition comprising a first dehydrated cell that is a dehydrated microorganism dissolved / melted in a hydrophobic solvent.
The third invention relates to a composition comprising a solution obtained by dissolving / melting a first dehydrated cell, a second dehydrated microorganism which is a freeze-dried cell, and a freeze-dried microorganism in a hydrophobic solvent.
The fourth invention relates to a composition in which the first dehydrated cell, the second dehydrated microorganism, the third freeze-dried cell, and the hydrophobic substance that dissolves / melts the freeze-dried microorganism are fats and oils.
The fifth invention relates to a method for producing the first to fourth compositions.
The sixth invention relates to an emulsion production method for the first to fourth compositions.
7th invention relates to fermented milk, a drink, fermented soymilk, a functional food, moisture retention cosmetics, a pollen adsorbent, and a house dust adsorbent in the foodstuff or cosmetics and medical supplies containing the composition of 1-4.

  Conventionally, it has been difficult to dissolve / melt cultured cells in a hydrophobic solvent, and therefore, fermented products containing animal and plant tissue cultures and bacterial cells have been lyophilized products coated with fats and oils. Since it was impossible to melt cells and bacteria cultured in an aqueous system into a hydrophobic solution, no attempt was made. However, the present inventors have succeeded in melting cells in a hydrophobic solvent for the first time, and they continue dormancy without being killed even though the cells are trapped in the hydrophobic solvent. The present invention has been completed by finding that it can be used for many products by emulsifying the product.

  According to the present invention, pure cells and microorganisms have been found to have two contradictory properties that are hydrophilic when wet and hydrophobic when dry, and are also soluble / melted in hydrophobic solvents. Then, it seems that the hydrophobic solvent seems to stop at the cell wall, and it can be utilized to the maximum extent that it retains the property of being regenerated if water is included again. For example, in the past, functional lactic acid bacteria collected from human intestines and those with strong fermentation odors such as diacetyl are disliked by the Japanese, and are activated immediately when they are put in water containing milk or juice. It can improve the growth and abnormal flavor, and when bifidobacteria that have not been bred to grow aerobically are put into milk or juice, it will die due to aerobic conditions and acidity. If the composition is used, anaerobic bacteria can also be used in foods alive. This makes it easy to produce foods that are good for human health, such as yogurt, with bifidobacteria and functional lactic acid bacteria, and have a great improvement effect. People do not eat food if it is delicious, and deliciousness is an important factor along with nutrition.

  In addition, according to the present invention, pollen adheres to the mucous membrane and causes sneezing, nasal congestion, runny nose, itchy eyes, etc., which are symptoms of hay fever, but lactic acid bacteria are dissolved around the pollen by dissolving / melting them in a hydrophobic solvent. Wrapping up, making it possible to take advantage of the unexpected and remarkable ability of the lactic acid bacteria mass to produce a powerful effect of attaching pollen to the grape.

The weight of the dehydrated bacteria contained in the hydrophobic solvent is about 40%, and more than that is pasty, and the content for maintaining a preferable liquid state in product production is 10% or less. If 10% of the composition containing 10% dehydrated bacteria is added as fermented milk, it becomes 10 ¹¹ to 10 ¹⁰ cells / g of bacteria per gram, which is extremely difficult by fermentation technology in which dehydrated bacteria are not added to a hydrophobic solvent. The utility becomes clearer.

  In the present invention, “dehydrated cells” are those obtained by removing media components other than cells, that is, removing cells that do not dissolve in the hydrophobic solution and drying the cells. As a drying method, freeze-drying is preferable for using cells while they are alive, and when the viable cell rate can be lowered, vacuum drying, drum drying, spray drying, or the like can be used.

  Since there are many hydrophilic substances in the culture solution, it is important to remove them. The method of washing the culture solution is usually used in physiological saline or buffer solution, and salt and inorganic salts remain after drying. Therefore, it is important to wash with clean water sterilized or sterilized / sterilized, for example, distilled water, ion exchange water, membrane filtered water, or tap water. The culture solution is centrifuged at 100 × g to 10000 × g for 5 minutes to 60 minutes in a centrifuge, and the supernatant is decanted and the amount of fresh water substantially divided into the precipitate is added and homogenized by stirring. Hydrophilic substances other than bacterial cells are removed by centrifugal washing under the same conditions for 10 times or more. More preferably, after centrifugation at 500 × g to 5000 × g for 10 minutes to 45 minutes, centrifugal washing may be performed 2 to 5 times under the same conditions. The second washing method is membrane washing, but after culturing, the membrane is washed with fresh water using a filtration device, but the diameter of the filtration hole is about 0.01 to 1000 μm and is removed. Hydrophilic substance is removed by adding clean water and homogenizing and filtering and washing under the same conditions 1 to 10 times, but may be performed continuously. More preferable conditions are slightly smaller than the size of the cell, and a membrane having a pore of 0.1 to 100 μm, and may be filtered and washed 2 to 5 times under the same conditions.

  In the present invention, examples of the “dehydrated cells” include dried animal cells, plant cells, algae, fungi, and bacteria, and may be used according to each application. As the drying method, freeze-drying, reduced-pressure drying, drum drying, spray drying, and the like can be used, and the drying method can be selected in consideration of the rate of living and dead bodies depending on the purpose of the cells to be used. In particular, lyophilization is optimal for obtaining a dried body of living cells, and the freezing temperature is preferably -1 ° C to -90 ° C, and the freezing and thawing temperature is preferably -20 ° C to 50 ° C. More preferable conditions are a freezing temperature of −5 ° C. to −50 ° C. and a thawing temperature of −10 ° C. to 50 ° C. The degree of vacuum may be 100 to 0.001 Torr (Torr), preferably 10 to 0.01 Torr.

  Examples of cells used for food include lactic acid bacteria, bifidobacteria, kefiya bacteria, natto bacteria, edible fungi, unicellular algae, multicellular algae, herbal tissues (eg, herbal cultured tissues), and the like. Examples of cosmetics and medical supplies that can be used include lactic acid bacteria, bifidobacteria, medical bacteria, medical fungi, cellulose, aloe soft tissue, Japanese and Chinese medicine tissues, and the like.

  In the present invention, “hydrophobic solvent” refers to a hydrophilic solvent, and “oils and fats” are biological oils and fats derived from animals and plants and mineral fats and oils derived from petroleum. Specifically, rapeseed oil, sesame oil, soybean oil, olive oil, cacao oil, palm oil, palm oil, palm kernel oil, wheat germ oil, avocado oil, castor oil, milk fat, fish oil, pork fat, beef tallow, tonsil oil, horse Examples include oil, petroleum jelly, paraffin, waxy oil, silicone, plastic base and the like. Moreover, fat-soluble vitamins such as tocopherol and carotene are also included.

  In the method of dissolving / melting dehydrated cells in a hydrophobic solvent, while the hydrophobic solvent is taken in a container and stirred with a stirrer, the dehydrated cells sieved to 3.5 to 500 mesh are dropped little by little to dissolve / melt. When the hydrophobic solvent is gelled at the working process temperature, dissolution / melting is started as a liquid by heating or the like. The sieve mesh is determined depending on the size of the cells, but it is preferable to use a 10-200 mesh sieve. The amount of cells that can be lysed in a hydrophobic solvent varies depending on the cell, but evenly dissolves / melts up to about 40% by weight. If the amount of cells is large, the paste is kneaded by adding a hydrophobic solvent and paste Can be made. When alive, the temperature of the solvent for lysing / melting the cells can be 75 ° C. or less and the solvent can be in a liquid range, but more preferably 0 ° C. to 65 ° C.

  In the present invention, cells melted in a hydrophobic oil and fat can be stored alive and exist in a dormant state even when emulsified in an aqueous system. In the emulsification method according to the present invention, the composition can be emulsified with an emulsified substance contained in food, such as phospholipids such as milk protein, soy protein, lecithin, etc., but mono / diglyceride, polyglycerin ester, sorbitan fatty acid used in food Emulsifiers such as esters, propylene glycolic acid esters and sugar esters can be used, and ionic and nonionic surfactants can be used for cosmetics and medical supplies. Lyophilized cells are dissolved in fats and oils at a temperature up to 75 ° C. and emulsified with a protein, phospholipid or emulsifier at a pressure of 1 to 50 MPa (megapascal), more preferably 10 to 30 MPa. The emulsifier may be a known one, but in the case of food, an aseptic homogenizer is preferred. The emulsion of the present invention may have an average particle size of 0.01 to 100 μm, more preferably an average particle size of 0.1 to 30 μm.

  Examples of the food product obtained by emulsifying the composition or the composition of the present invention include fermented milk and fermented beverages in which lactic acid bacteria and bifidobacteria are dissolved / melted in milk fat or vegetable oil and emulsified in skim milk or soy milk. It is a kefir drink or functional food that does not generate carbon dioxide gas even when there is a yogurt or yeast containing bifidobacteria that can be produced under tempering conditions. In addition, foods containing herbs, ginseng tissue and chlorella can be produced without changing the taste of milk or soy milk. When lactic acid bacteria and cellulose are dissolved in mineral fats and / or biological fats and oils, the fungus coated with fats and oils adheres around the pollen, and the bacterial mass and fine cellulose have the property of adsorbing pollen and house dust. It can be applied near the mucous membrane of the eyes to inactivate allergens and reduce the occurrence of hay fever and house dust allergies. Thus, a moisturizing cream having moisturizing properties can be produced. In addition, the foodstuffs, cosmetics, and medical supplies which add the hydrophobic solvent containing a dehydrated cell, or its emulsion are not limited to these.

  EXAMPLES Hereinafter, although an Example demonstrates this invention concretely, this invention is not restrict | limited to these Examples. In addition, the composition in an Example is shown by weight%.

Comparison of solubility of “wet cells”, “dehydrated cell cultures” and “dehydrated cells” The medium for culturing lactic acid bacteria may be any liquid lactic acid bacteria medium, but here it is cultured in MRS broth for lactic acid bacteria (Dico) The solution was prepared, sterilized at 120 ° C. for 15 minutes, and then cooled to 40 ° C., and (a) Lactobacillus bulgaricus JCM1002, (b) Lactobacillus rhamnosus ATCC 7469, (c) Lactococcus lactis ATCC 15346 2% of each bacterial solution preliminarily cultured at 38 ° C. for 18 hours in each medium was added to each medium and cultured at 38 ° C. for 20 hours using a low-speed intermittent stirring tank to obtain each culture. This culture was freeze-dried to produce a “dehydrated lactic acid bacteria culture”. The culture is centrifuged at 5000 × g for 15 minutes in a centrifuge (5420; Kubota Corporation), decanted, and the supernatant removed from the precipitate is added to the precipitate to make it homogeneous and homogenized. And was washed by centrifugation three times. Things other than the cells were removed to produce “wet lactic acid bacteria”. This wet lactic acid bacterium was freeze-dried at a freezing temperature of −30 ° C. using a freeze dryer (RLE2; Nissei Co., Ltd.), with a degree of vacuum of 0.2 Torr and a freeze-thaw temperature of 30 ° C. to produce “dehydrated lactic acid bacteria”.

  A GAM broth medium for Bifidobacterium was prepared, sterilized at 120 ° C. for 15 minutes, and then cooled. (D) Bifidobacterium infantis ATCC15697 was preliminarily cultured at 37 ° C. for 18 hours in the above medium. %, And an unstirred container was used, and simple anaerobic (Aneropack; Mitsubishi Gas Chemical Co., Ltd.) was cultured at 37 ° C. for 20 hours to obtain a culture. This culture was freeze-dried to produce a “dehydrated bifidobacteria culture”. In addition, the culture was concentrated with a membrane (MLE-7101; Kuraray Co., Ltd.) having a hole size of 0.1 μm, and almost equal amount of sterile water was added to the removed solution to homogenize it, and three times under the same concentration conditions. Filter washed. Except for the bacterial cells, "wet bifidobacteria" was produced. The wet bifidobacteria were freeze-dried in the same manner as lactic acid bacteria to produce “dehydrated bifidobacteria”.

  While stirring, 1 part each of “wet cells”, “cultured dehydrated cells” and “dehydrated cells” was dissolved / melted at 30 ° C. in 9 parts each of ion-exchanged water, purified rapeseed oil and liquid paraffin. . The results of dissolution / melting are shown in Table 1. “Wet lactic acid bacteria a, b, c and wet bifidobacteria d” dissolves well in ion exchange water, does not melt at all in refined rapeseed oil and liquid paraffin, and “dehydrated lactic acid bacteria and bifidobacteria culture” dissolves well in on-exchange water. / Melted and dispersed in refined rapeseed oil and liquid paraffin in the form of mamako. “Dehydrated lactic acid bacteria and dehydrated bifidobacteria” became mamoko-like in on-exchange water, and were uniformly dissolved / melted in purified rapeseed oil and liquid paraffin. This makes it clear that the method of easily dissolving / melting in a hydrophobic solvent requires a step of removing the culture medium, washing the cells and drying.

1 part of each lyophilized lactic acid bacterium (b) in a hydrophobic solvent of dehydrated lactic acid bacteria and dehydrated bifidobacteria was dissolved in 99 parts of purified soybean oil and 99 parts of petroleum jelly, and 1 part of lyophilized Bufidobacterium (d) was milk fat (butter Oil) dissolved in 99 parts, kept at 10 ° C., and the results of examining the number of bacteria after 7 days, 14 days and 28 days are shown in Table 2. From this result, it was proved that lactic acid bacteria and bifidobacteria melted in fats and oils could retain the number of bacteria without dying. From this, it was found that it can be used alive in food.

Vegetable oil and fat-containing fermented soymilk 2.94 parts of purified soybean solidified oil are heated and sterilized at 85 ° C, and then cooled to 40 ° C. I let you. In addition, 97 parts of soymilk was sterilized at 85 ° C. for 15 minutes, cooled to 40 ° C., mixed with the purified soybean solidified oil containing the lactic acid bacteria and mixed with a homogenizer (L-01; Sanwa Machinery Co., Ltd.) at 20 MPa. After emulsifying with pressure, the mixture was packed in a container and contained lactic acid bacteria alive, but a soymilk drink containing lactic acid bacteria without change in flavor could be produced. One week later, 1.9 × 10 ⁹ cells / g of lactic acid bacteria were detected.

Method for Producing Fermented Milk Containing Bifidobacteria After dissolving butter at 85 ° C. and removing water by decantation, it was freeze-dried to 2,98 parts of butter oil cooled to 40 ° C. 0.02 part of the bacterium (d) was dissolved / melted in a sterile room. Also, 96 parts skim milk and 1 part skim milk powder were added to prepare a mix, sterilized at 85 ° C. for 10 minutes, cooled to 40 ° C., mixed with the butter oil containing the previous bifidobacteria, and 15 MPa of homogenizer. After emulsification under pressure, add a savory starter of bulgariya and lactis, put it in a container, ferment it at 40 ° C, contain Bifidas alive, and produce a plain yogurt with good flavor It was. One week later, Bufidobacterium was detected at 4.2 × 10 ⁸ cells / g.

Evaluation of adsorbability of cedar pollen As a liquid A, an appropriate amount of cedar pollen was dispersed in 10 parts of liquid paraffin. As liquid B, 0.6 parts of freeze-dried lactic acid bacteria (b) was dissolved / melted in 9.4 parts of long-chain hydrocarbon (7 parts liquid paraffin + 2.4 parts petrolatum) heated to 50 ° C. As liquid C, 0.6 part of absorbent cotton fiber was dissolved / dissolved in 9.4 parts of long-chain hydrocarbon (7 parts liquid paraffin + 2.4 parts petrolatum) heated to 50 ° C. After adding 1 part of liquid A and 1 part of liquid B and stirring slowly, 0.1 ml was taken with a micropipette and spread flatly to an area of 10 mm × 10 mm, and 20 fields of view were examined with a microscope with a magnification of 200. Moreover, 1 part of A liquid and 1 part of C liquid were added, and it microscopically examined similarly. As a result, there were 64 cedar pollen / 20 visual fields that were neatly dispersed only with the liquid A. There were 35/20 fields of A + B liquid that were aggregated and adsorbed in two or more, and it was confirmed that bacterial clumps were also attached to the pollen alone. There were six A + C liquids that aggregated and adsorbed to two or more.

  Also, an appropriate amount of cedar pollen was dispersed in 10 parts of purified soybean oil as D solution. As E solution, 0.6 parts of freeze-dried lactic acid bacteria (b) was dissolved in 9.4 parts of purified soybean oil, and as absorbent F, 0.6 parts of absorbent cotton fiber was dissolved in 9.4 parts of purified soybean oil. Liquid D alone, liquid D 1 part and liquid E 1 part, liquid D 1 part and liquid F 1 part were mixed, treated in the same manner as long-chain hydrocarbons and microscopically examined.

  These results are shown in Table 3. From this, it was found that long-chain hydrocarbons in which lactic acid bacteria were dissolved / melted adsorbed pollen and aggregated into grapes. By applying a composition in which lactic acid bacteria are dissolved / melted in long-chain hydrocarbons such as liquid paraffin and petrolatum to the skin around the nose and eyes, the lactic acid bacteria dissolved / melted in the hydrophobic solvent adhere around the pollen. It is adsorbed on the lactic acid bacteria and agglomerates in the lump of lactic acid bacteria, and the pollen is inactivated to reduce hay fever, and if moisturized dehydrated aloe soft tissue is added, it becomes a moisturizing cosmetic ingredient and emulsified into eye drops and moisturizing sprays. It can be used and can also be used in cosmetics if emulsified in a cosmetic cream.

Claims (7)

A composition comprising dehydrated cells dissolved / melted in a hydrophobic solvent. A composition wherein the dehydrated cells of claim 1 are dehydrated microorganisms. A composition wherein the dehydrated cells of claim 1 are lyophilized cells and the dehydrated microorganisms of claim 2 are lyophilized microorganisms. The composition whose hydrophobic solvent of Claims 1-3 is fats and oils. The manufacturing method of the composition of Claims 1-4. The emulsification method of the composition of Claims 1-4. Foods, cosmetics and medical supplies containing the composition comprising claims 1, 2, 3, 4 are fermented milk, beverages, fermented soymilk, functional foods, moisturizing cosmetics, pollen adsorbents, house dust adsorbents, etc. is there.