WO2006016595A1 - Lactoferrin complex and method of producing the same - Google Patents

Abstract

It is intended to provide a lactoferrin complex which has an excellent storage stability in the presence of water owing to improvements in the stability of lactoferrin in the presence of water and gastric juice, its solubility in the intestinal tract and its thermal stability during a heating treatment in an aqueous system, in which lactoferrin can get to the intestinal tract without being digested by pepsin in the gastric juice and is efficiently absorbed in vivo to thereby fully exert its physiological activity, and which can be advantageously employed in a heating step such as thermal pasteurization in the production of foods, drugs, feeds and so on; a material composition containing the same; and a method of producing the same. This lactoferrin complex is a complex of lactoferrin with a high-molecular weight electrolyte.

Description

 Specification

 Ratatofu phosphorus complex and method for producing the same

 Technical field

 The present invention relates to a complex of ratatopherin and a polyelectrolyte, a method for producing the same, and a lactoferrin material composition which can be used for medicine, food, feed, etc. containing this complex, and further, the material Relating to medicine, food, feed etc. More specifically, a ratatopheline complex capable of efficiently exerting a physiological activity, a ratatopheline material composition, having enhanced stability in the presence of water and in gastric juice, entericity, and stability to heat treatment in an aqueous system. The present invention relates to a method for producing them, and medicines, foods and feeds using this composition. Background art

 Ratatophelin is a glycoprotein with a molecular weight of about 80,000 which is mainly present in the milk of mammals and is also found in neutrophils, tears, saliva, nasal discharge, bile, semen and the like. Ratatoferrin belongs to the transferrin family because it binds iron. Physiological activities of ratatofurin include antibacterial activity, iron absorption control activity, cell proliferation activation activity, hematopoietic activity, anti-inflammatory activity, antioxidative activity, phagocytosis activity, antiviral activity, bifidobacteria growth promotion activity, antifungal activity. It is known to have effects on cancer, cancer metastasis, and translocation. Furthermore, it has recently been reported that ratatophelin has lipid metabolism improving and analgesic effects. Thus, ratatophelin is a multifunctional physiologically active protein that exhibits various functions, and is expected to be used for applications such as medicines and foods for the recovery or improvement of health, and it contains ratatopheline. Food is already commercially available.

 [0003] Until now, ratatofu phosphorus used in food and the like is a powder which is mainly separated from milk or dairy products and is spray-dried or freeze-dried. In dry powder form, ratatopherin is stable at room temperature for about 1 to 2 years.

However, ratatophelin is unstable when dissolved in water. When ratatoferin is used in medicine, food or feed, etc. in the presence of water, modification due to its three-dimensional structure being broken during storage or modification due to aggregation with a coexistent substance occurs, resulting in ratatopherin It loses its physiological activity. Therefore, ratatohuelin has water Can not be stored for a long time in the existing state.

 Therefore, when ratatopheline is used for medicine, food, feed, pet food, etc., ratatopheline powder is used as it is as a powder in order to enable long-term storage of several months or more. It was limited to any one of whether it was used as a tablet formulated in a dry powder state with the addition of excipients and the like.

[0006] In addition, when ratatofurin is used in solution, for food, pharmaceuticals, feed, etc., sterilization by heat treatment is essential. Ratatopherin is unstable to heat and is denatured by heating at 60 ° C or higher. And precipitate. Therefore, when heat-sterilizing ratatoferin aqueous solution at a temperature of 60 ° C. or higher, a method has been developed to improve the thermal stability of ratatoferrin by adjusting the ionic strength of the aqueous solution according to the heating temperature (Patent Document 1) JP-A-2-10862). According to this method, for example, in the case of heat sterilization at 80 ° C., it is necessary to desalt and adjust ion strength to be 10 ⁻³ or less (NaCl conversion concentration: about 0.006%) . However, in general, this method is not practical because a certain level of salt concentration is indispensable for food, medicine, feed and the like.

 Further, a method of improving the thermal stability of the ratatopheline-containing solution by the heat treatment by performing the heat treatment at a temperature of 60 ° C. or higher after adjusting the ratatopheline-containing solution to pH 10.5 to 6.5. Have been developed (Patent Document 2: Patent No. 2688098, Non-Patent Document 1). However, in general, solutions for food, medicine, feed and the like are around neutral pH around pH 7, and this method can not improve the thermal stability of the ratatopheline-containing solution at near neutral pH.

 [0008] Therefore, there has been a demand for a practical technique for improving the thermal stability of ratatophelin which can be applied to food, medicine, feed and the like.

Furthermore, when ratatophelin is orally taken, it is hydrolyzed by the acid protease pepsin, which is present in gastric juice, to be decomposed into a peptide, so that ratatophelin molecules can almost reach the intestinal tract as the ratatopheline molecule. I can not However, it is known that ratatophelin receptor is present in the small intestine mucosa in the digestive tract, and recently, ratatophelin is taken up by intestinal epithelial cells and is involved in gene expression in the nucleus, and inflammatory site is It has been reported that it controls the development. Therefore, the physiological activity of ratatophelin In order to exert it, it is necessary to allow ratatopheline to reach the intestinal tract without undergoing hydrolysis by pepsin in gastric juice.

 From the above point of view, enteric ratatopherin tablets obtained by coating enteric films on uncoated tablets pressed with tablet in dry powder state without using water by adding an excipient etc. to health support It is sold as a food.

 However, in the presence of water, storage stability is good even when used for applications such as pharmaceuticals, food or feed, and in gastric juice, it reaches the intestinal tract without being degraded by pepsin, and ratatopheline is reached. Ratatofu-phosphorin raw material yarn, which is stable to heat treatment near neutral pH while being able to sufficiently exert its physiological activity, and pharmaceuticals, food and feed using it. He was an unknown force.

 [0012] Polyelectrolytes have been used as thickeners, dispersion stabilizers, adhesives, protective colloids, and the like in various technical fields including food, pharmaceuticals, and formulated feeds. For some high molecular weight electrolytes, it is known that gelled ones adsorb ratatopherin, and this property has been utilized for the purification of extranuclear phosphorus.

 [0013] For example, a method for purifying a protein such as ratatophein is reported, in which proteins present in milk such as ratatophelin and lactoperoxidase are adsorbed to a gelled acidic polysaccharide (alginate, carrageenin) and then eluted. (Patent Document 3: Japanese Patent Application Laid-Open No. 61-246198).

 Also, a method of preparing a gel composition of a protein with excellent thermal stability that can be used as a food and drink material by heating a protein such as casein and a gelling agent containing ratatoferin as an active ingredient (patented) Reference 4: JP-A-8-98655), a method for producing a folate-lactophrins complex useful as a food or drink or a pharmaceutical material by mixing folate or the like with ratatophelin (Patent Document 5: JP-A-2001-106) 238640), ratatofurin by adding a polyvalent inorganic acid (phosphoric acid or sulfuric acid) or an organic acid (taenic acid, maleic acid, fumaric acid, tartaric acid, succinic acid, ethylenediamine tetraacetic acid or chondroitin sulfate) to a solution of ratatofurin. A method for stabilizing (Patent Document 6: WO99Z06065) has been reported.

[0015] However, the techniques described in these documents use only gelled acidic polysaccharides as an adsorption carrier for purifying ratatofurin, and use only calcium phosphate, ratatofelin and others. It is used only as a gelling agent for proteins or for improving the photostability, acid resistance and solubility of folic acid, or a polyvalent organic acid is added for stabilization of aqueous ratatopheline solution. In any of these documents, the complex of these polyelectrolytes and ratatofurin is inactivated in the physiological activity of ratatofurin, changes in heat stability upon heat treatment, and resistance to gastric juice and intestine. The statement about solubility is unknown at all.

 In addition, it is known that a polyion complex is formed by mixing polymer electrolyte solutions having charges of opposite signs to each other (Non-patent Document 4) Force The heat stability, pepsin resistance, and the like of these polyion complexes. It is unknown if it is enteric.

Patent Document 1: Japanese Patent Application Laid-Open No. 2-108629

 Patent Document 2: Patent No. 2688098

 Patent Document 3: JP-A-61-246198

 Patent Document 4: Japanese Patent Application Laid-Open No. 08-098655

 Patent Document 5: Japanese Patent Application Publication No. 2001-238640

 Patent Document 6: WO 99 Z 606 6065

 Patent Document 7: Japanese Patent Application Laid-Open No. 2002-161050

 Patent Document 8: Japanese Patent Application Laid-Open No. 2002-322088

 Patent Document 9: Japanese Patent Application Laid-Open No. 2002-326950

 Patent Document 10: Japanese Patent Application Laid-Open No. 2003-089629

 Patent Document 11: Japanese Patent Application Laid-Open No. 2003-137808

 Patent Document 12: Japanese Patent Application Laid-Open No. 2003-137809

 Non Patent Literatures: Abe, H "baito, H" iya awa, H "Tamura, Y .: Journal of dairy science (1991) 74 (1), 65-71

 Non-patent literature 2: New food material effective utilization technology series No. 6 “Ratatofu ヱ リ ン”, Inc. (Inc.) Confectionery General Technology Center published, March 2000

 Non-Patent Document 3: Biochemical Dictionary, Tokyo Chemical Co., Ltd., April 1984

 Non-patent document 4: Polymer assembly, published by Academic Press Center, January 1983

Disclosure of the invention [0018] The present invention provides improved stability in the presence of water and in gastric juice, entericity, and thermal stability of ratatofu phosphorus to heat treatment in an aqueous system, resulting in preservation in the presence of water. The stability is excellent, and ratatoferin reaches the intestinal tract and is eluted or released without being (or reduced in degradation) degraded by pepsin in gastric juice, so it is efficiently absorbed by the body and sufficiently bioactive. A ratatopheline material composition having a feature that it can be used to perform a process involving heating such as heat sterilization in food, medicine, feed and the like, and a method for producing the same In addition, it is an object of the present invention to provide foods, medicines and feeds containing the material composition. In particular, it is an object of the present invention to provide a ratatophelin raw material composition which can be stably stored for a long time in a form other than a dry powder state.

 The present inventors have intensively studied to solve the above problems, and as a result, ratatopherin is mixed with a polyelectrolyte in an aqueous solution to form a water-insoluble complex with the polyelectrolyte. As a result, they have found that the stability of ratatofurin in the presence of water and in gastric fluid, the entericity, and the heat treatment in an aqueous system are significantly increased, and the present invention has been completed.

 That is, the present invention

 (1) A complex of ratatopheline and a polyelectrolyte;

 (2) The complex according to the above (1), wherein the polyelectrolyte is a polymeric acid which dissociates in an aqueous solution to form polyanion;

 (3) The complex according to (2), wherein the polyelectrolyte further comprises a polymer salt group that dissociates in an aqueous solution to form a polycation.

 (4) The complex according to any one of the above (1) to (3), which is capable of releasing lactoferrin in bile.

(5) The polyelectrolytes are casein, gelatin, acidic egg white proteins, acidic whey proteins, poly L-glutamic acid, carboxymethylcellulose, carrageenan, alginic acid, fucoidan, galactan sulfate, xanthan gum, pectin, gum arabic Lyacrylic acid, polymethacrylic acid, dielan gum, natto bacteria gum, soybean water-soluble polysaccharide, agar, starch, furcellane, tracanth gum, heparin, hyaluronic acid, chondroitin, Heparan sulfate, dermatan sulfate, keratan sulfate, chondroitin sulfate, carboxymethyl starch, carboxymethyl amylose, buttermilk powder, acid soluble soy proteins, soy milk, chitosan, glycol chitosan, polylysine, polyglutamine, polyarginine, polyethylene The complex according to any one of the above (1) to (4), which is one or more selected from the group consisting of immin, polyvinylamine, and salts thereof;

 (6) Any one of the above (1) to (5), which is in the form of a colloidal precipitate, a sol, a gel, a powder, a form encapsulated in a capsule, a tablet, a granule, a pill or an emulsified suspension. A compound of the description;

 (7) A powder obtained by dispersing a lipid coating-coated powder containing outer ferritin and a powder coating containing a lipid coating-coated polymer electrolyte in an aqueous solution, the above-mentioned (1) to (5) 5) The complex according to any one of 1);

 (8) An aqueous solution of a polymer base is dispersed in an aqueous solution by dispersing a powder containing a lipid coating coated exoferrin and a powder containing a lipid coating coated polymeric acid in an aqueous solution. The compound according to any one of the above (3) to (5), which is formed by adding

 (9) A ratatophile material composition comprising the complex according to any one of the items (1) to (8);

 (10) The rattoferrin raw material composition according to the above (9), which is in the form of powder or granules containing a basic substance and a binder.

 (11) The basic substance is also selected from soy milk, chitosan, polylysine, polyarginine, polyarginine, polydaltamin, sodium hydrogencarbonate, arginine, glutamine, histidine, lysine, dulcosamine, sodium carbonate, or a group of one or two. (10) The ratatofelin material composition as described in (10) above.

 (12) The above binder is one or more selected from the group consisting of zein, shellac, water-soluble soybean polysaccharide, carboxymethyl cellulose, alginic acid, guar gum and carboxymethyl cellulose. (10) or the composition of the ratatofelin raw material as described in (11).

(13) A physiological activity of ratatophelin as a raw material composition according to any one of the above (9) to (12). A food containing an amount effective to exert the property;

 (14) A pharmaceutical composition comprising the material composition according to any one of the above (9) to (12) in an amount effective to exert the physiological activity of ratatopherin;

 (15) A feed comprising the material composition according to any one of the above (9) to (12) in an amount effective to exert the physiological activity of ratatopherin;

 (16) A method for producing a complex of ratatopherin and a polymer electrolyte, which is obtained by mixing an aqueous solution containing ratatopherin and an aqueous solution containing a polymer electrolyte at normal temperature or under heating. A method characterized by recovering the body;

 (17) A method for producing a complex of ratatofuline and a polymer electrolyte, which comprises mixing an aqueous solution containing ratatofurin with an aqueous solution containing a polymer electrolyte which is a polymeric acid at room temperature or under heating, Then, an aqueous solution containing a polymer electrolyte which is a polymer base is added to form a complex of ratatopherin, a polymer acid and a polymer base, and the obtained complex is recovered. Method;

 (18) A method for producing a complex of ratatopherin and a polymer electrolyte, which comprises preparing a powder containing ratatopherin and a polymer electrolyte, dispersing the powder in an aqueous solution, and heating at normal temperature or under heating. A method comprising mixing to form a complex, and recovering the obtained complex;

 (19) A method for producing a complex of ratatopherin and a polymer electrolyte, which comprises preparing a powder containing ratatopherin and a polymer electrolyte which is a polymer acid, and dispersing the powder in an aqueous solution to obtain a mixture at room temperature. Alternatively, they are mixed under heating, and then an aqueous solution or powder containing a polymer electrolyte which is a polymer base is added, and mixed under normal temperature or heating to form a complex of ratatophenin, a polymer acid and a polymer base. A method characterized in that it forms and recovers the complex obtained;

 (20) The method according to (18) or (19), wherein the powder containing ratatopherin and the polymer electrolyte is prepared by mixing at normal temperature or under heating;

 (21) The method according to the above (18) or (19), wherein both the outer ferritin and the polyelectrolyte are previously coated with a lipid film;

(22) A powder coating containing a lipid coating and a powder coating containing LaFerrin and a lipid coating Raw material for preparing a complex of ratatofluorine and a polyelectrolyte, which is contained in a container separately or in a mixed state with a powder containing a coated polyelectrolyte.

 I will provide a.

 [0021] The ratatopherin complex and the raw material composition of the present invention are water insoluble in a normal state, and have good storage stability even in the presence of water. In addition, ratatophein contained in the complex and the material composition of the present invention does not elute in gastric juice and does not undergo degradation by pepsin or its degradation is reduced. In the digestive tract containing bile acids such as lipase and deoxycholic acid, that is, in the intestine containing bile etc., the complex can be easily eluted or released, so it is efficiently absorbed by the body. It is possible to exert physiological activity.

 Bile is classified into hepatobile and gallbladder bile, which generally have the following composition. The term "bile" in the context of the present invention also includes both of these and experimentally used model bile (3 wt.% Bile (Wako Pure Chemical Industries or equivalent bile powder), 50 mM sodium bicarbonate).

 [Table 1] General composition of bile

さらに、本発明の複合体及び素材組成物は、酸性 pH付近 (pH3. 5)〜中性 pH付 近での加熱処理に対して安定であるので、食品、医薬、飼料などの製造においてカロ 熱殺菌処理などの加熱を伴う工程を行う上で好都合に利用することができ、製造上 の制限が緩和される結果、多種多様な製品に応用することができる。また、本発明の 複合体及び素材組成物は、シンプルな工程で簡便に製造することができる点におい て、製造上も有利である。

Furthermore, the composite and material composition of the present invention is stable to heat treatment at around the acidic pH (pH 3.5) to near neutral pH, and therefore, it can be used in the production of food, medicine, feed and the like. It can be advantageously used in carrying out processes involving heating, such as heat sterilization, and as a result of the restriction on production being relaxed, it can be applied to a wide variety of products. In addition, the composite and the material composition of the present invention are advantageous in terms of production because they can be easily produced in a simple process.

 Therefore, the food, medicament and feed containing the ratatopheline complex or ratatophelin raw material composition of the present invention are not limited to those in a dry state, and even in the form in which water is present, the physiological activity of ratatofurin is obtained. It can be maintained for a long period of time, and even if it is heat-treated, it can sufficiently retain the physiological activity of ratatopheline, so the storage stability and usefulness of food, medicine and feed itself are improved. Significantly improved.

 Brief description of the drawings

FIG. 1 is a view showing the pH dependence of pepsin degradation of ratatophelin.

 BEST MODE FOR CARRYING OUT THE INVENTION

[0027] In the present invention, "latatopherin" also includes the natural ratatopherin molecule itself, gene recombinant ratatofurin, and functional equivalents of ratatofurin such as active fragments of ratatofurin. The presence or absence of iron ion or its content, regardless of the species from which it originates. Further, in the present invention, “the ratatophelin raw material composition” is a composition containing ratatopheline, which can be used as a raw material of other products such as food, medicine, feed and the like, and the physiological activity or characteristics of ratatopherin. Means a material composition to be used for at least one purpose of exerting it in the product in which it is used.

In the present invention, “polyelectrolyte” refers to a substance that dissociates in an aqueous solution to form polyions (macroions) and counterions (microions), and is a polymeric acid, a polymeric base or ampholytic polymer. Molecular Electrolyte! /, Misalignment May be Synthetic or Natural! /, Misalignment! / ,. The polymer electrolyte used in the composite, material composition and the like of the present invention includes, among arbitrary polymer electrolytes, physiologically acceptable polymer electrolytes or salts thereof, especially for food and medical use etc. Depending on the application, one or two or more may be selected as appropriate.

In general, polyelectrolytes used as food additives are preferred in terms of availability, price and Z or safety. Specifically, for example, casein, gelatin, Acidic egg white proteins (ovalbumin, conalbumin, ovomucoid, ovoglobulin

(Eg, ovomucin), acid whey proteins (eg, 8-lactoglobulin, α-lactoalbumin), poly-L-glutamic acid, carboxymethylcellulose, carrageenan, arginate, fucoidan, galactan sulfate, xanthan gum, pectin, Gum arabic, polyatarlic acid, polymethacrylic acid, dielan gum, natto bacteria gum, soybean water soluble polysaccharide, agar, starch, fermenter, tracanthum gum, carboxymethyl starch, carboxymethyl starch, carboxymethylcellulose, buttermilk powder, soya bean protein (Especially acid soluble soy protein), chitosan, glycol chitosan, polylysine, and salts thereof (physiologically acceptable salts such as sodium salt, potassium salt, hydrochloride); and acids used in medical applications Mucopolysaccharides, for example Palin, hyaluronic acid, chondroitin, heparan sulfate, dermatan sulfate, keratan sulfate, chondroitin sulfate, polyglutamine, polyarginine, polyethyrimin, polybilamin, and their salts (sodium salt, potassium salt, hydrochloride, etc. 1) or 2 or more types can be selected and used from the group which consists of salts which can be used.

 In general, as the polymer electrolyte used in the present invention, when there is one type of polymer electrolyte, a polymer acid which dissociates in an aqueous solution to form polyanion (for example, an acidic protein, Acidic polypeptides and acidic polysaccharides) are preferred. Among these polyelectrolytes, the use of a strongly acidic polyelectrolyte is particularly resistant to acid dissociation and pepsin degradation as compared with a complex with a weakly acidic electrolyte, and the complex is can get. On the other hand, weakly acidic polyelectrolytes are particularly preferred. This is because the complex with the weakly acidic electrolyte has better elution or release of ratatofulin in complex power in bile as compared with the complex with strong acid electrolyte. When there are two or more types of polyelectrolytes, a polymeric acid that dissociates in an aqueous solution to form polyanion and a polymeric base that dissociates in an aqueous solution to form a polycation (for example, basic protein, base It is preferred to include both a sex polypeptide and a basic polysaccharide). In the present invention, a complex of ratatopheline and a polymeric acid may be abbreviated as a binary complex in particular. In addition, a complex of ratatopheline, a polymeric acid and a polymeric base may be abbreviated as a ternary complex.

[0031] A complex of ratatofulin and a polyelectrolyte (herein, simply referred to as The term “complex” or “complex” is sometimes referred to as polyelectrolyte salt of ratatopheline. This is formed, for example, by mixing an aqueous solution containing ratatopherin with an aqueous solution containing polyelectrolyte at normal temperature or under heating. The binary composite is formed by mixing an aqueous solution containing ratatopheline with an aqueous solution containing a polymeric acid at normal temperature or under heating. In the ternary complex, an aqueous solution containing ratatoferin and an aqueous solution containing a polymeric acid are mixed under normal temperature or heating, and then an aqueous solution containing a polymeric base is added to the mixture. It is formed by mixing.

 An "aqueous solution" in the context of the present invention also includes water itself, examples being water as well as saline, physiologically acceptable buffers etc. Also, "solution" does not require that the solute be completely dissolved, and does not need to be liquid in a strict sense. Thus, in the context of the present invention, a "solution" may contain dispersed or suspended substances or it may be in the form of a yogurt or sherbet etc.

 Here, the temperature for heating is generally about 40 ° C. to 120 ° C., preferably 50 ° C. to 90 ° C., and most preferably in the range of 50 ° C. to 65 ° C. is there. Below 40 ° C., certain polyelectrolytes such as agar tend to solidify. On the other hand, when the temperature exceeds 65 ° C., the physiological activity of ratatoferin is easily inactivated for a long period of time when the temperature exceeds 100 ° C., the physiological activity is almost lost. The specific temperature can be appropriately selected according to the selected polymer electrolyte.

 [0033] The time for carrying out the mixing is not particularly limited as long as the outer shell phosphorus and the polymer electrolyte are sufficiently mixed to form a complex. Temperature conditions to be adopted, size of reactor, shape of mixing means It can be selected appropriately according to the situation.

 The state of the complex thus formed varies depending on the type, concentration, mixing ratio, pH and the like of ratatopherin and polyelectrolyte, and is in the form of a colloidal precipitate, sol or gel. Generally, the binary complex is in the form of a hydrophilic hydrogel, and the ternary complex is in the form of a dewatering precipitate.

The two-component complex is formed by binding to the basic group of ratatopheline molecule by a high molecular weight acid bond, and the acidic group of ratatopheline becomes free, so the complex as a whole is considered to be a negative charge. There is. Therefore, the binary complex hydrogen bonds water molecules It is thought that it takes in to form a hydrogel and does not precipitate at high concentrations. On the other hand, in the ternary complex, a polymer base is formed by ionic bonding to a free acidic group of the binary complex, charge is lost and water molecules are released. It is considered to be in the form of a well-dehydrated precipitate, so that the precipitate fraction and the supernatant fraction will be separated even by natural sedimentation. These complexes are in the form of a powder, for example, through a drying process such as lyophilization or spray drying. The ternary complex is excellent in separation property and drying efficiency, and is advantageous in production.

 Recovery of the complex will be described later.

 Alternatively, the complex of the present invention is prepared as a powder containing ratatopherin and a polyelectrolyte, and the powder is dispersed in an aqueous solution to form a complex, and the obtained complex is recovered. It can also be manufactured by collecting. For example, in the case of a two-component complex, a powder containing ratatofuran and a polymeric acid can be prepared, and this powder can be dispersed in an aqueous solution to form a complex. In the case of a ternary complex, a powder containing extraferrin and a polymeric acid is prepared, this powder is dispersed in an aqueous solution, mixed at normal temperature or under heating, and then containing a polymeric base. The aqueous solution or powder can be added and mixed at normal temperature or under heating to form a complex.

 Specifically, a powder containing ratatopheline and a polyelectrolyte may be prepared, for example, by mixing a powder containing lactoferrin and a powder containing a polyelectrolyte at normal temperature or under heating. Can. In this case, it is preferable that the powder containing ratatophenin and the powder containing the polymer electrolyte are each coated with a lipid film in advance. The coating with a lipid film may be separately performed before mixing the ratatopherin-containing powder and the polymer electrolyte-containing powder, or may be performed after mixing both powders.

That is, preferably, ratatopherin in a powder containing ratatopherin is coated with a film (sometimes referred to as “lipid film”) consisting of a film material containing a lipid as a main component, and a polymer The polyelectrolyte in the powder containing the electrolyte is also coated with a lipid coating. As the lipid, any of fats and oils, waxes and complex lipids may be acceptable as long as they are acceptable for food and medicine use. For example, naturally occurring animal or vegetable fats or oils or Examples thereof include hydrogenated oils (such as hydrogenated beef tallow oil, hydrogenated fish oil, hydrogenated rapeseed oil, hydrogenated soybean oil, hydrogenated palm oil, olive cured oil, and hydrogenated peanut oil). Preferably, a lipid having a melting point in the range of 30 to 90 ° C., more preferably a lipid having a melting point of 40 to 80 ° C. is used. In addition, as the lipid of ratatopherin powder and the lipid of polyelectrolyte powder, preferably, lipids having similar melting points, more preferably, lipids of the same composition are used.

 Therefore, according to this method, the composite of the present invention is a powder containing ratatofurin and a polymer electrolyte, or a mixture of a powder containing ratatopherin and a powder containing a polymer electrolyte (preferably a lipid film). The powder is stored in the form of a mixture of powder containing extra-ferrin particles coated with a powder and powder containing a polyelectrolyte coated with a lipid film, and the powder is dispersed in an aqueous solution at the time of use. The complexes of the present invention can be prepared at the time of use and used. In this way, the step of separately preparing the ratatoferrin aqueous solution and the polymer electrolyte aqueous solution can be omitted, and the formation of dams when Z or the powder is dispersed in the aqueous solution can be prevented, and the operability can be improved. It is advantageous.

 [0039] According to this method, the ratatofelin-containing powder and the polymer electrolyte-containing powder are separately stored in a container, mixed when necessary, and combined with the aqueous solution. The present invention can be provided as a raw material product for preparing a complex of ratatopherin according to the present invention and a polyelectrolyte. Alternatively, the powder containing ratatopherin and the powder containing the polymer electrolyte may be provided as a raw material product which is contained in a container in an already mixed state and, if necessary, combined with an aqueous solution, . With respect to these raw materials, in particular, powders containing ratatoferin coated with a lipid coating and powders containing a polyelectrolyte coated with a lipid coating are preferred.

[0040] Recovery of the formed complex can be performed by conventional means.

In the preparation of the ratatopheline complex of the present invention, for example, ratatopheline: polymer electrolyte = 4: 1 to 1: 4 (weight ratio) can be used. In the production of the two-component complex, for example, ratatopheline: polymer acid = 5: 1 to L: 5 (weight ratio) can be used. In addition, in the production of the ternary complex, the binary complex: polymer base = 20: 1 to 3 Lactofuran, a polymeric acid, and a polymeric base that can be about 1 (weight ratio) can be used.

 [0042] The complex formed by the method as described above, after being recovered from the reaction vessel, is in the form as it is, ie, in the form of a colloidal precipitate, sol or gel, particularly in the form of hydrogel or precipitate. Although it can be used, the complexes formed may be further processed to other forms such as powders, emulsified suspensions and the like. For example, after forming a complex as described above, the complex formed in the form of a colloidal precipitate is separated by centrifugation or the like, and this complex is further lyophilized, a sol or The formed complex in the form of a gel can be lyophilized as it is. Also, for example, after forming a ternary complex, the precipitate collected by centrifugation etc. is freeze-dried and pulverized to form powder, or the binary complex in the form of hydrogel is freeze-dried as it is, It can be milled into powder form.

 Furthermore, the ratatoferrin complex of the present invention can be used in the form of a powder and further in the form of being enclosed in a capsule, specifically in the form of being enclosed in a microcapsule, a soft capsule or a hard capsule. The capsule of the material of the present invention can be carried out by known microcapsule technology, soft capsule technology, hard capsule technology and the like.

 [0043] The material composition of the present invention may be a complex of various forms as described above, or may be a tablet or a granule with or without the additional component described later in the complex. The latatopherin material composition of the present invention can be made into any form such as pill and the like. Methods of processing to such forms are known in the art of formulation technology.

 [0044] The material composition of the present invention may contain, in addition to the ratatofelin-polyelectrolyte complex, other additional components generally used in the art, Those skilled in the art may appropriately select the type of the components and the amount to be added according to the form of the specific material composition to be manufactured, the application, etc. and the properties of the product to which the material composition is to be applied. it can.

 [0045] For example, plasticizers, excipients, emulsifiers, stabilizers; vitamin A, vitamin B, vitamin B, vitamin

12 Vitamins such as Tamine B, Vitamin B, Vitamin C, and Vitamin E; Cozym Q, α-li Lipid-soluble or water-soluble general nutritional ingredients such as norenic acid, EPA, DHA; medicinal ingredients with analgesic activity such as morphine, codeine, dihydrocodin, cetylmorphine, other medicinal ingredients; and Z or lactic acid bacteria, La Examples include components for improving intestinal flora such as oligosaccharides.

 [0046] Examples of the raw material composition include a raw material composition in the form of powder or granules containing the binary composite (or ternary composite) of the present invention, a basic substance, and a binder. The basic substance referred to here may be a low molecular weight polymer or a high molecular weight substance as long as it has an effect of neutralizing gastric acid and raising the pH to 3 or more when the complex is brought into contact with gastric juice. Specific examples of the basic substance suitable for use in the present invention include soymilk, chitosan, polylysine, polyarginine, polyglutamine, sodium hydrogencarbonate, arginine, glutamine, histidine, lysine, darcosamine, carbonate Sodium etc. are mentioned. Also, the binder may be one generally used, and examples thereof include Tween, shellac, water-soluble soybean polysaccharide, carboxymethylcellulose, alginic acid, guar gum, carboxymethyl cellulose and the like. . The basic substance and the binder can be used by appropriately selecting one or two or more kinds.

 The ratatofu-phosphorus raw material composition of the present invention can be contained in food, medicine, feed and the like. The appropriate content of lactoflavin when it is contained in medicine, food, feed and the like is known.

 Hereinafter, the present invention will be described in detail by way of examples, but the present invention is not limited to the examples.

 Example

 Example 1: Production of a lactoferrin-pectin complex

 A solution of 5 g of Pectin (Hakobo Shokai) in 1 L of pure water is added to a solution of 10 g of ratatofelin (Tatza 'milk' BIO mouth Zix's) in 1 L of pure water, and mixed for 30 minutes. A complex was formed. The obtained gel-like solution was dried as it was with a lyophilizer (coalescence vacuum technology) to obtain 14 g of a pyrophylline complex.

[0050] Example 2: Notice of Lactopheline-CMC combination

12 g of ratatofelin (Tatza Milk Bio-Zix) was dissolved in 500 mL of pure water A solution of 6 g of CMC sodium (Wako Pure Chemical Industries, Ltd.) dissolved in 500 mL of pure water was added to the solution, and mixed for 30 minutes to form a complex. The obtained gel-like solution was dried as it was with a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 14 g of the exo-ferrin complex of the present invention.

 [0051] Example 3: Production of a lactoferrin-one strength raginan complex

 A solution of 6 g of carrageenan (Ina Foods) in 500 mL of pure water is added to a solution of 10 g of ratatofelin (Tatza 'milk' BIO mouth Zix's) in 1 L of pure water, and mixed for 30 minutes. Formed. The obtained gel-like solution was dried as it was with a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 14 g of the extraferrin complex of the present invention.

Example 4: Fat covered lactoferrin powder and fat covered height> Early lightning-free powder Raw material Review of lactoferrin-carrageenan complex

 2g of mixed ratatoferrin and carrageenan powder coated with rapeseed oil (60% by weight ratatoferrin, 30% carrageenan and 10% by weight rape oil) in a mixer containing 100 mL of RO water Thus, ratatopheline-carrageenan complex was formed.

 At this stage (neutral pH), the complex formed a water-insoluble hydrophilic gel, so 1 mL of 1 N hydrochloric acid was added to adjust to an acidic pH (about pH 2). The complex immediately turned almost water insoluble and formed a precipitate.

 The precipitate was dried as it was with a lyophilizer (Kyowa Vacuum Technology Co., Ltd.) to obtain about 1.7 g of the lactoferrin complex of the present invention.

Example 5: Production of a capsule containing a lactoferrin material composition

 Ratatohurin complex in a dry powder form produced in the same manner as in Examples 1 to 4 is used as a ratatopheline raw material composition as it is, 1 part by weight of ratatopheline raw material composition, 10 parts by weight of microcrystalline cellulose, and rapeseed oil 0.5 parts by weight was mixed, and 150 mg of this powder mixture was filled in a node capsule. The capsule can be used as a pharmaceutical composition or a health supplement.

Example 6: Production of an ice cream containing a lactoferrin material composition

Each of ratatofutorin complex in the form of a dry powder prepared in the same manner as in Examples 1 to 4 was used. As it is, it was used as ratatohuelin raw material composition. A small amount of butter, emulsion stabilizer and flavor was added to 1 L of milk, 300 g of skimmed milk powder, 300 g of sugar and 1 L of water, and after mixing and dissolving, it was homogenized with a homogenizer. An ice cream was prepared by adding 2 g of any ratatopheline material composition and subjecting it to an ice cream freezer.

 Example 7 Production of Yogurt Containing Lactoferrin Material Composition

 1 L of milk, 100 g of skimmed milk powder, 10 g of sugar, ratatopherin complex in dry powder form prepared as in Example 3 (used as it is as a ratatopherin raw material composition) 2 g of butter, stabilizer and flavor added to a small amount of water Then, the mixture was dissolved and homogenized with a homogenizer. It was cooled after heat sterilization at 78 ° C. for 3 minutes and then at 98 ° C. for 2 seconds. After holding at 39 ° C., a lactic acid bacteria starter was added and fermented to prepare yogurt.

 [0056] Example 8: Lactoferrin material for containing pet ^! Notice of fees

 Ratatofurin complex (in the form of ratatofurin ingredient composition) 3g, corn flour 100g, fish meal 30g, yeast extract 10g, beef tallow 2g, soy flour 25g, Fratatooligosaccharide 20g, dextrin 10g, as dried powder form prepared in Example 3. A small amount of sodium chloride was mixed, sufficiently mixed while adding water little by little using a kneader, and granulated with a twin-screw extrusion granulator. Furthermore, the obtained granules were dried at 40 ° C. overnight to produce a pet food.

 [0057] Test, Test Example ί: Heat resistance test, test

10 mg of ratatophein complex 10 mg prepared in Example 1 was dispersed in 10 mM Tris-HCl buffer (pH 7.0) 1. Om, and heated at 80 ^° C. for 30 minutes. After heat treatment with heat, centrifugation at 10,000 rpm for 10 minutes was carried out, the precipitate was dissolved in 1 M NaCl, and ratatopherin in the supernatant liquid was analyzed by high performance liquid chromatography to find that it corresponds to about 6 mg ZmL of ratatopherin. Was detected.

On the other hand, naked ratatopherin as a control (lactopherin used as a raw material in Example 1 (Tatza 'milk' Bio-Physics Co., Ltd.)) 10 mg of 10 mM Tris-HCl buffer (pH 7.0) 1. Dissolve in O mL, 80 ° Heated at C for 30 minutes. After the formed precipitate was removed by centrifugation, the supernatant liquid was measured for ratatopherin by high performance liquid chromatography, and the peak for ratatopherin was hardly detected. In addition, the precipitate was extracted with 1 M NaCl, and the rat's extract of ratatopheline was measured by high performance liquid chromatography. Foremost was an undetectable force.

Thus, a complex with Ratatofuerin the polymer electrolyte of the present invention, to be stable became apparent to heat treatment at around neutral _P H.

Test Example 2: Stabilization with gastric juice PH

 The ratatofurin complex 1 g prepared in Example 1 is dispersed in 100 mL of a model gastric fluid (0.2% sodium chloride, 70 mM HCl, pH 1.2), and left at 37 ° C. for 2 hours, and then the supernatant liquid is recovered at a high speed. It was measured by liquid chromatography. As a result, almost no peak generated by elution of ratatophein was observed, and it was confirmed that the ratatophelin complex of the present invention was stable in gastric juice pH.

[0059] Test, Test Example 3: Stabilizing needle needle for pepsin treatment

 After 0.15 g of the ratatopheline complex prepared in Example 3 is added to 100 g of commercially available plain yoghurt and thoroughly stirred and mixed in a mixer, 500 mg of the mixture is placed in a 1.5 mL Ezbend Ruf tube to obtain 0.4 mg ZmL pepsin solution (Η 2 ) Carried out at 500 ° C for 30 minutes at 37 ° C. Thereafter, centrifugation (10, OOO rpm, 10 minutes) was performed to recover the supernatant and the precipitate, respectively. The precipitate was dispersed in 500 L of 40 mM deoxycholic acid. SDS-PAGE was performed using this precipitate and the supernatant as samples, and the presence or absence of degradation of ratatopheline was examined.

 As a result, it was found that the ratatopheline complex of the present invention is stable to pepsin. As a control, 0.1 mg of naked ratatopherin was subjected to the same treatment as above, and the degradation of ratatoferrin was observed in SDS-PAGE.

Test Example 4: Efficacy and safety of lactoferrin

 Sixteen 5-week-old ICR male mice were randomly divided into two groups of eight each, and a control group was fed with a standard diet for rat 'mouse (CE, 2 from CLEA Japan), and the experimental group was A feed containing 0.2% of ratatopheline complex prepared in the same manner as in Example 1 or 2 was given to CE-2 and the animals were reared for 4 weeks. Body weight was measured every three days during this breeding period.

As for body weight, there was no significant difference between the two groups. In addition, the weights of liver, spleen, spleen, small intestine, cecum, visceral fat, and epididymal adipose tissue weighed at the time of necropsy after 4 weeks were also significantly different. In addition, the length per unit weight and the length of the intestine are both groups There was no significant difference between

 The blood fat content was 25% (P 0 0. 05) and the blood free fatty acid value was 30% (P 0 0. 05), respectively, as determined by oral administration of ratatopheline complex, as measured for blood components. It has dropped significantly. Next, when the blood cholesterol level was measured, the experimental group to which the ratatofu phosphate material composition was administered showed a tendency to increase the total blood cholesterol level compared to the control group, but the increase was due to the HDL cholesterol strength 0% It became clear that it was because it rose (P <0. 01).

 From the above results, it has been found that the ratatophelin raw material composition of the present invention is safe for the living body, and at least exerts the physiological action of Lafrin, which improves lipid metabolism, sufficiently in a small amount.

 [0061] Example 9: Production of extra-ferlin-xanthan gum complex

 A solution of 5 g of xanthan gum (Taiyo Engineering Co., Ltd.) in 500 mL of pure water is added to a solution of 10 g of ratatofelin (Tatza 'milk', BIOguchi Zix Co., Ltd.) dissolved in 1 L of pure water, Mix for minutes to allow complex formation. The obtained gel was dried as it was with a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 14 g of the La complex of the present invention.

 [0062] mio fat covered with lactoferrin powder and fat covered with powdered xanthan gum

Mixed powder (Ratatofuerin 60 weight _^0/0, xanthan gum 30 weight _^0/0 and 10 wt% rapeseed hardened oil) of Ratatofuerin coated with rapeseed hardened oil (Kawaken Fine Chemicals Co., Ltd.) and hexa Ntangamu a 100g, 0. 001N Kuen The mixture was charged into a mixer containing 1 L of acid solution and mixed by stirring to form a complex. The obtained gel was dried as it was with a lyophilizer (Koiwa Sky Technology Co., Ltd.) to obtain about 86 g of the La complex of the present invention.

Example 11 Production of Lactoferrin-Alginate Complex Using Fat-Coated Lactopherin Powder and Fat-Coated Sodium Alginate Powder as Raw Materials

Mixed powder (Ratatofuerin 60 weight _^0/0, sodium alginate 30 wt% and rapeseed hard oils 10 weight _^0/0) of Ratatofuerin and sodium alginate coated with rapeseed hardened oil (Co. Kimi force) to 100g, 0. 001N The mixture was charged into a mixer containing 1 L of a citric acid solution and mixed to form a complex by stirring. The obtained gel is directly freeze-dried. The product was dried by Technology Co., Ltd. to obtain about 87 g of the extraneous phosphorus complex of the present invention.

Example 12: Fat-coated lactoferrin powder and fat-coated CMC sodium powder Preparation of lactoferrin-CMC complex

Mixed powder (Ratatofuwerin 60 wt%, 10 wt CMC Sodium 30 wt% and rapeseed hardened oils _^0/0) of Ratatofuerin and sodium CMC coated with rapeseed hardened oil (Dai-ichi Kogyo Seiyaku Co.) to 100g, 0. 001N Kuen The mixture was charged into a mixer containing 1 L of acid solution and mixed by stirring to form a complex. The obtained gel was dried as it was with a lyophilizer (Kyowa Vacuum Technology Co., Ltd.) to obtain about 87 g of the La complex of the present invention.

Example 13: Lipid-coated lactoferrin powder and lipid-coated vectin powder as raw materials

 100 g of mixed powder of ratatoferin and vectin (3 crystals of rapeseed oil) coated with rapeseed oil (60% by weight ratatopheline, 30% by weight pectin and 10% by weight rapeseed oil), 1 L of 0.10 N citric acid solution The mixture was charged into a mixer and mixed by stirring to form a complex. The obtained gel was dried as it was with a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 87 g of the ratatophelin complex of the present invention.

 [0066] mi 4 fat-coated lactoferrin powder, fat-coated xanthan gum powder, and highly soluble large-protein raw material: lactoferrin-xanthan gum as acid material-capable of acidity ^ protein complex: 3⁄43⁄4 告

Mixed powder (Ratatofuerin 60 weight _^0/0, xanthan gum 30 weight _^0/0 and 10 wt% rapeseed hardened oil) of Ratatofuerin coated with rapeseed hardened oil (Kawaken Fine Chemicals Co., Ltd.) and hexa Ntangamu a 10 g, 0 of LOOmL. The mixture was charged into a mixer containing a 001N lactic acid solution, and stirred and mixed to form a binary complex. Next, add 100 mL of 0. 001N citric acid solution containing 5% by weight of acidic soluble soy protein (Fuji Oil Co., Ltd.), and stir and mix to form a ternary complex (latatopheline-xanthan gum-acidic soluble) Soy protein complex was formed. The resulting precipitate was collected by centrifugation and dried with a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 1 l g of ratatopherin complex of the present invention.

Example 15 Production of Lactoferrin Xanthan Gum Chitosan Complex Starting from Fat Coated Lactoferrin Powder, Fat Coated Xanthan Gum Powder and Chitosan Mixed powder (Ratatofuerin 60 weight _^0/0, xanthan gum 30 weight _^0/0 and 10 wt% rapeseed hardened oil) of Ratatofuerin coated with rapeseed hardened oil (Kawaken Fine Chemicals Co., Ltd.) and hexa Ntangamu a 10 g, 0 of LOOmL. The mixture was charged into a mixer containing a 001N lactic acid solution, and stirred and mixed to form a binary complex. Next, 100 mL of a 0. 001 N citric acid solution containing 1 wt% chitosan (Co. Co. Technics Co., Ltd.) was added and mixed with stirring to form a ternary complex (latatophenin-xanthan gum-chitosan complex). The resulting precipitate was collected by centrifugation and dried with a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 9 g of the ratatofuran complex of the present invention.

 Example 16: Report of Lactoferrin-Pectin-Chitosan Complex Starting from Fat-Coated Lactoferrin Powder, Fat-Coated Vectin Powder, and Chitosan

 100 g of mixed powder of ratatopherin and vectin (3 crystal, Inc.) coated with rapeseed oil (60% by weight ratatopherin, 30% by weight pectin and 10% by weight rapeseed oil), 1 L of 0.10 N aqueous citric acid solution The mixture was charged into a mixer and stirred and mixed to form a binary complex (latatopherin-pectin complex). Next, add 1 L of 0. 001N citric acid solution containing 3 wt% chitosan (Coco Technos Co., Ltd.) and stir and mix to form a ternary complex (La extraferrin-pectin-chitosan complex). The The resulting precipitate was collected by centrifugation and dried by a lyophilizer (Kowa Vacuum Technology Co., Ltd.) to obtain about 90 g of the ratatophrin complex of the present invention.

 [0069] m: Production of a lactoferrin- 1-Pectin-acid-soluble soybean protein complex, which is based on a lipid-coated lactoferrin powder, a lipid-coated 蹬 pectin powder, and a highly acceptable protein.

 Mixed powder of ratatofelin and bectin (Crystal Co., Ltd.) coated with rapeseed oil

100 g of ratatopheline 60% by weight, pectin 30% by weight and rapeseed oil 10% by weight are introduced into a mixer containing 1 L of 0.10 N aqueous solution of 001 N citric acid, and the mixture is stirred to mix to form a binary composite (latatopheline- (Pectin complex) was formed. Next, add 1 L of 0. 001N citric acid solution containing 5% by weight of acidic soluble soybean protein (Fuji Oil Co., Ltd.), and stir and mix to make a ternary complex (latatophein-pectin-acidic soluble soybean Protein). The resulting precipitate is collected by centrifugation and dried with a lyophilizer (Kyowa Vacuum Technology Co., Ltd.), About 120 g of ratatophelin complex of the present invention was obtained.

Example 18 Production of a Lactoferrin-Containing Lactoferrin Powder, a Lipid-Coated Vectin Powder and Soymilk The manufacture of a lactoferrin-containing granule

 Mixed powder of ratatoferin and bectin (Tricrystal Co., Ltd.) coated with rapeseed hydrogenated oil (67% by weight of ratatopheline, 17% by weight of pectin, and 16% by weight of rapeseed hydrogenated oil) 450 g, soy milk powder prepared by Fuji Oil ( The mixture was mixed with 1,560 g of dextrin, 940 g of dextrin, and 50 g of an excipient and granulated with a fluid bed granulator (Freund Sangyo Co., Ltd.) to obtain about 2,700 g of extra-ferlin-containing granules.

 [0071] Test, Test Example 5: Enteric dissolution test, Test

 10 mg of ratatoferin complex (ratatoferin content: 60%) prepared in Example 10 was dispersed in 10 ml of model bile (3 wt% bile (Wako Pure Chemical Industries, Ltd., 50 mM aqueous sodium hydrogen carbonate solution (pH 8)) at 37 ° C. I processed it for 1 hour. After centrifugation, ratatopherin in the supernatant fluid was measured by high performance liquid chromatography, and a peak corresponding to about 5.9 mg ZmL of ratatopherin was detected. As a control, 100 mg of ratatoferrin complex was dispersed in 10 mL of pH 6. 8 (phosphate buffer), treated in the same manner, and ratatopholine in the supernatant fluid was measured by high performance liquid chromatography. It was a force.

 Therefore, the complex of ratatopherin of the present invention and polyelectrolyte liberates ratatopheline in bile near neutral pH and exhibits enteric solubility, but does not contain bile! /, Ratatopherin near neutral pH liberates It became clear that it was water insoluble.

Test Example 6: Heat resistance test (2)

 1. Take lmg of ratatopheline complex prepared in Example 10 in an Eppendorf tube, 1 OmM Tris-HCl buffer (pH 7.0) 1. Add OmL, and block heater (IWAKI; THERM 0 A to UMI BATH) It was heated at 75 ° C. for 30 minutes. After heat treatment with heat, centrifugation at 10,000 rpm for 10 minutes was carried out to dissolve the precipitate in model bile, and ratatopherin of the supernatant fluid was measured by high performance liquid chromatography to find that ratatopherin was about 4.8 mg ZmL (recovery: about 80% The peak corresponding to) was detected.

On the other hand, 10 mg of naked ratatopherin (as a raw material used in Example 9 as a raw material in Tatza 'milk' BIO-ZUKICS Co., Ltd.) as a control was treated with 1 mg of 10 mM Tris-HCl buffer solution (pH 7.0). Dissolve in OmL and heat at 75 ° C for 30 minutes. After the formed precipitate was removed by centrifugation, the supernatant liquid was subjected to ratatopherin measurement by high performance liquid chromatography, and the lactoferrin peak was almost undetectable. Moreover, when the precipitate was extracted with model bile and ratatopheline of the extract was measured by high performance liquid chromatography, the peak of ratatopheline was hardly detected.

Thus, a complex with Ratatofuerin and polymer electrolyte of the present invention are stable to heat treatment at around neutral _P H, it revealed that enteric also maintained.

Test Example 7: A sputum-dependent test for pepsin degradation of lactoferrin

 Prepare a solution of pH 2.0, pH 3.0, pH 3.5, pH 4.0, pH 4.5, pH 5.0, containing ratatoferin (Tatza 'milk' BIO Mouth Dix) lOmg ZmL or pepsin 0.4 mg / mL. did.

 For each pH, 500 μL of ratatopherin solution was added to 500 μL of each pepsin solution of Η and treated at 37 ° C. for 30 minutes. As a control, in place of pepsin solution, each pH solution not containing pepsin was added and treated in the same manner. After the reaction, centrifugation was performed, and ratatopherin in the supernatant was measured by HPLC.

[0074] Similarly, ratatophein-l-pectin-chitosan complex lOOmg prepared in the same manner as in Example 16 had a pH of 2.0, a pH of 3.0, a pH of 3.5, a pH of 4.0, a pH of 4.5, and a pH of 5.0. The solution was dispersed in 5 mL, the pH was measured, and adjusted to each pH. These solutions were covered with 5 mg of 0.4 mg Z mL of pepsin solution of each pH and treated at 37 ° C. for 30 minutes. As a control, instead of pepsin solution, each pH solution not containing pepsin was coated and treated in the same manner. After the reaction, centrifugation was performed to recover the supernatant and the precipitate. The precipitate was dissolved in 10 mL of model bile and centrifuged to recover the supernatant. The ratatopherin concentration in the above sample was measured by HPLC.

 The residual rate of ratatophein in pepsin treatment was determined from the ratio of the concentration of residual ratatofurin after pepsin treatment to the concentration of ratatophein in the control at each pH.

The results are shown in Table 2 and Figure 1. As shown in Table 2 and FIG. 1, naked ratatopheline is almost completely degraded by pepsin below pH 4 but is reduced below pH 4 and hardly affected by pepsin above pH 5. In contrast, ratatohuelin of the present invention The complex was degraded by pepsin at pH 3 or less, but hardly affected by pepsin at pH 3 or more, and it was revealed that pepsin was hardly degraded at pH 3.5 or more. Thus, it is clear that the ratatophelin complex of the present invention is pepsin resistant.

 [Table 2] of lactoferrin and lactoferrin complex

 P H dependence test in pepsin degradation

Test Example 8: Stability against pepsin treatment (2)

 Fifty mg of the ratatofulin complex prepared in Example 10 was dispersed in 5 mL of a 0.2 mg / mL pepsin solution (pH 3, hydrochloric acid) and treated at 37 ° C. for 30 minutes. After centrifugation, the precipitate was extracted with model bile, and the ratatopherin of the extract was measured by high performance liquid chromatography to obtain a peak corresponding to about 3.6 mg ZmL of ratatopherin (about 60% recovery). On the other hand, 50 mg of naked ratatopherin powder is dissolved in 5 ml of 0.2 mg / mL pepsin solution (pH 3, hydrochloric acid), treated at 37 ° C. for 30 minutes, and centrifuged for the supernatant liquid. The peak corresponding to lactoferrin was almost completely degraded.

 Therefore, it was revealed that, for the ratatofurin-polyelectrolyte complex of the present invention, the degradation of ratatofurin by pepsin is reduced and that ratatofurin in the complex is liberated in bile. .

Test, Test Example 9: Heat resistance test, Test (3) 100 mg of the ratatopheline complex prepared in Example 16 was placed in an Eppendorf tube, 1 mL of a 0. 001 N aqueous solution of citric acid was added, and heat treatment was performed at 75 ° C. for 30 minutes using a block heater. After cooling, centrifugation was performed, and 10 mL of model bile was added to the obtained precipitate, and elution was performed at 37 ° C. for 1 hour. Thereafter, centrifugation was carried out, and the lactoferrin in the supernatant liquid was measured by high performance liquid chromatography to obtain a peak corresponding to about 3.3 mg ZmL (recovery rate: 72%) of ratatopherin.

 Therefore, it was revealed that the complex of ratatopherin of the present invention and a polyelectrolyte is stable to heat treatment in the acidic region, and the enteric property is also maintained.

Test Example 10: Stability against pepsin treatment (3)

 After 4 mL of pure water was suspended in 1 g of the ratatophein-containing granule prepared in Example 18 and suspended, it was allowed to stand for 30 minutes, and 4 mL of 0.4 mg ZmL pepsin solution (pH 2, hydrochloric acid) was suspended at 37 ° C. The treatment was carried out for 30 minutes. After centrifugation, 10 mL of model bile was added to the obtained precipitate, and elution was performed at 37 ° C. for 1 hour. Thereafter, centrifugation was performed, and the lactoferrin of the supernatant fluid was measured by high performance liquid chromatography. As a result, a peak corresponding to 1.8 mg ZmL of ratatopheline (recovery rate: 72%) was obtained.

 Therefore, it was revealed that, for the granules containing the ratatopheline complex of the present invention, the degradation of ratatophein by pepsin is reduced and that ratatopheline in the granules is released in the bile.

Test, Test: In Rats: Intestinal Absorption Kinetics Test in Rats

 Ten 8-week-old male Wistar rats were divided into two groups of five each, and naive lactoferrin was dissolved in physiological saline for intragastric administration (300 mg Z kg) in the control group. The extracellular ferritin-containing granules produced in the above were suspended in physiological saline and administered intragastrically (30 mg Z kg of ratatofelin). One hour before administration and also four hours after administration, thoracic duct lymph fluid and venous blood were collected every hour, and ratatoferin was quantified by ELISA method.

Comparing the total amount of ratatofu-phosphorin transferred into the thoracic duct lymphatic fluid, the control group peaked at 1 hour after administration and then decreased gradually, while the experimental group maintained high level until 4 hours. As a result, the total amount in the experimental group from immediately after administration until 4 hours after was about twice the value of the control group. 1 dose of ratatofuelin containing granules of naked ratatophelin It was found that ratatoferin-containing granules were absorbed from the intestinal tract about 20 times as much as naked ratatopheline because it was 1/0.

 When ratatopheline in plasma was measured, ratatopheline was detected in the plasma in only 1 case out of 5 in the control group and in 3 cases out of 5 in the experimental group (concentration of about 50 to 200 ng ZmL).

 From the above results, it was found that the complex of the present invention is resistant to digestion in the stomach and has an effect of improving the absorption efficiency of ratatoferrin from the small intestine.

Example 19: Raw material of lactoferrin-Pectin complex soymilk powder

 Ratatopherin-Pectin complex lOOg prepared in the same manner as in Example 13 and prepared soybean milk powder (Fuji Protein Engineering Co., Ltd.) lOOg is placed in a No-speed mixer (Fukae Powder Tech Co., Ltd.), 25 wt% tween (84% by weight ethanol solution containing Showa Sangyo Co., Ltd. (Kouka Chemical Co., Ltd.) (75 g of 84% by weight ethanol in which 25 g of tween is dissolved) 10 g of 10 g was sprayed and granulated. It was dried at 40 ° C. for 8 hours to obtain about 210 g of a raw material composition in the form of granules.

 3⁄4 row 20: lactoferrin-xanthan gum ^! Coalesced ^ 3⁄4, powder 萌 形 形 ⁄ ⁄

 Ratatopherin- xanthan gum complex lOOg prepared in the same manner as in Example 9 and conditioned soybean milk powder (Fuji Protein Engineering Co., Ltd.) lOOg was placed in a No-Speed Mixer (Fukae Bowtech Co., Ltd.), and 25 wt. An 84% by weight ethanol solution (Showa Sangyo Co., Ltd.) was mixed and granulated by spraying 100 g of ethanol solution. It was dried at 40 ° C. for 8 hours to obtain about 215 g of a raw material composition in the form of granules.

 Example 21: A material in the form of granules obtained from a lactoferrin-one pectin complex chitosan

Ratatoferin-Pectin complex lOOg prepared in the same manner as in Example 13 and chitosan (Kyowa Technos Co., Ltd.) lOOg are placed in an i-speed mixer (Fukae Powder Technology Co., Ltd.), 25 wt% tween (Showa Sangyo Co., Ltd.) Granulation was carried out by spraying 100 g of an 84% by weight ethanol solution (Sanka Kagaku Co., Ltd.) containing K.K. Dry at 40 ° C for 8 hours to obtain about 21 lg of material composition in granular form The

 Example 22: Production of a complex of a chew-shaped bear made from La ferella ferin-Xanthan gum complex and chitosan

 Ratatopheline-Xanthan gum complex lOOg prepared in the same manner as in Example 9 and Kitosan (Kohan Technos Co., Ltd.) lOOg are placed in a speed mixer (Fukae Powder Tech Co., Ltd.), 25 wt% tween (Showa Sangyo The solution was granulated by spraying 1 OOg of an 84 wt% ethanol solution (Sanka Chemical Co., Ltd.) containing Drying at 40 ° C. for 8 hours gave about 210 g of a complex in the form of granules

Example 23: Preparation of chewable particulate form using a complex of exo-ferin, 1-pectin-chitosan and chitosan

 Ratatopherin- 1-Pectin-chitosan complex 100g produced in the same manner as in Example 16 and chitosan (Kohan Technos Co., Ltd.) 1 OOg is placed in an I-speed mixer (Fukae Powder Tech. Co., Ltd.) An 84% by weight ethanol solution containing (Showa Sangyo Co., Ltd.) (Glass Tank Science Co., Ltd.) was granulated by spraying lOOg. The mixture was dried at 40 ° C. for 8 hours to obtain about 21 Og of a material composition in the form of granules.

 Example 24: Lactoferrin-Pectin-Chitosan Complex ^ 3⁄4, Powder Preparation of Granule Particles 3⁄4

 Ratatoferin- 1-Pectin-chitosan complex lOOg prepared in the same manner as in Example 16 and prepared soymilk powder (Fuji Protein Engineering Co., Ltd.) lOOg was placed in a No-Speed Mixer (Fukae Bauttech Co., Ltd.), An 84% by weight ethanol solution containing Twain (Showa Sangyo Co., Ltd.) was granulated by spraying 100 g of a 100 g ethanol solution. It was dried at 40 ° C. for 8 hours to obtain about 210 g of a material composition in the form of granules.

 [0087] Example 25: Powdered powder made from a complex of exo-ferin, pectin-chitosan and chitosan as raw materials

Ratatopherin-one-Pectin-chitosan complex 100g produced in the same manner as in Example 16 and chitosan (Kohan Technos Co., Ltd.) 1 OOg is placed in an I-speed mixer (Fukae Powder Co., Ltd.), 30% by weight shellac 50 g of an anhydrous ethanol solution containing (Gifu Shellac Co., Ltd.) (30 g of shellac dissolved in 70 g of anhydrous ethanol) was sprayed and granulated. 40 ° C, 8 hours The resultant was dried to obtain about 210 g of a raw material composition in powder form.

 Example 26: 13⁄4% of the grain-shaped particles made from the exo-pherin-pectin-chitosan complex and chitosan

 Ratatopherin- 1-Pectin-chitosan complex lOOg and chitosan (Kohan Technos Co., Ltd.) 100g prepared in the same manner as in Example 16 were placed in an I-speed mixer (Fukae Powder Technology Co., Ltd.), and 30% by weight shellac ( It was granulated by spraying 100 g of an anhydrous ethanol solution containing Gifu Shellac Co., Ltd.). It was dried at 40 ° C. for 8 hours to obtain about 210 g of a raw material composition in the form of granules.

 Test Example 12: Stability against pepsin treatment (4)

 EXAMPLE 1 100 mg of ratatopherin-xanthan gum complex prepared in the same manner as in Example 9 was dispersed in 5 mL of a pH 3.5 solution, and the pH was measured and adjusted to pH 3.5. This solution was covered with 5 mL of 0.4 mg / mL pepsin solution (pH 3.5) and treated at 37 ° C. for 30 minutes. As a control, instead of pepsin solution, pH 3.5 solution was added and treated similarly. After the reaction, centrifugation was performed to recover the supernatant and the precipitate. The precipitate was dissolved in 10 mL of model bile and centrifuged to collect supernatant. The concentration of ratatopherin in the above sample was measured by HPLC. As a result, the residual rate of ratatopheline in the ratatopheline complex at pepsin treatment at pH 3.5 was about 80%. Therefore, it has been revealed that the ratatophelin complex of the present invention is pepsin-quenching.

 This application is based on Japanese Patent Application No. 2004-233347 filed on Aug. 10, 2004, and is described in the specification and claims of Japanese Patent Application No. 2004-233347. Are all included in this application.

Claims

The scope of the claims  [1] A complex of ratatopheline and a polyelectrolyte.  [2] The complex according to claim 1, wherein the polyelectrolyte is a polymeric acid which dissociates in an aqueous solution to form polyanion. [3] The composite according to [2], wherein the polyelectrolyte further comprises a polymer base which dissociates in an aqueous solution to form a polycation. [4] The complex according to any one of claims 1 to 3, which can release lactofrin in bile. [5] The polyelectrolytes are casein, gelatin, acidic egg white proteins, acid whey proteins, poly-L-glutamic acid, carboxymethylcellulose, carrageenan, alginate, fucoidan, galactan sulfate, xanthan gum, pectin , Gum arabic, polyacrylic acid, polymethacrylic acid, dielan gum, natto bacteria gum, soybean water-soluble polysaccharide, agar, starch, fercellan, tracanthum gum, heparin, hyaluronic acid, chondroitin, heparan sulfate, dermatan sulfate , Keratan sulfate, chondroitin sulfate, carboxymethyl starch, carboxymethyl amylose, buttermilk powder, acid soluble soy proteins, soy milk, chitosan, glycol chitosan, polylysine, polyglutamine, polyarginine, polyethyleneimine The complex according to any one of claims 1 to 4, which is one or two or more selected from a group consisting of polyvinylamine and their salt power.  [6] The method according to any one of claims 1 to 5, which is in the form of a colloidal precipitate, a sol, a gel, a powder, a form encapsulated in a capsule, a tablet, a granule, a pill or an emulsified suspension. Complex of  [7] Any one of claims 1 to 5, which is formed by dispersing, in an aqueous solution, a powder containing ratatofelin coated with a lipid film and a powder containing a polyelectrolyte coated with a lipid film. Or a complex according to clause 1; [8] A powder containing ratatofelin coated with a lipid film and a powder containing a polymer acid coated with a lipid film are dispersed in an aqueous solution and mixed, and then an aqueous solution of polymer base is prepared. The compound according to any one of claims 3 to 5, which is additionally formed. [9] A ratatofelin material composition comprising the composite according to any one of claims 1 to 8.  [10] The rattoferrin raw material composition according to claim 9, which is in the form of a powder or granules containing a basic substance and a binder. [11] The group force in which the basic substance is also soymilk, chitosan, polylysine, polyarginine, polyglutamine, sodium hydrogen carbonate, arginine, glutamine, histidine, lysine, dulcosamine, sodium carbonate is also selected 1 or 2 The lactoferrin raw material composition according to claim 10, which is a species or more.  [12] The binding agent is one or more selected from the group consisting of zeain, syrup, water-soluble soybean polysaccharide, carboxymethyl cellulose, alginic acid, guar gum, and carboxymethyl cellulose. The ratatophelin raw material composition according to claim 10 or 11.  [13] A food comprising the material composition according to any one of claims 9 to 12 in an amount effective to exert the physiological activity of ratatophelin. [14] A pharmaceutical composition comprising the material composition according to any one of claims 9 to 12 in an amount effective to exert the physiological activity of ratatophelin. [15] A feed comprising the material composition according to any one of claims 9 to 12 in an amount effective to exert the physiological activity of ratatophelin. [16] A method for producing a complex of ratatophenrin and a polymer electrolyte, which is obtained by mixing an aqueous solution containing ratatopheline and an aqueous solution containing a polymer electrolyte under normal temperature or heating, Recovery method. [17] A method for producing a complex of ratatofulin and a polymer electrolyte, which comprises mixing an aqueous solution containing ratatopherin with an aqueous solution containing a polymer electrolyte which is a polymeric acid at normal temperature or under heating. Then, an aqueous solution containing a polymer electrolyte which is a polymer base is added to form a complex of ratatopheline, a polymer acid and a polymer base, and the obtained complex is recovered. Method. [18] A method for producing a complex of ratatopherin and a polymer electrolyte, which comprises preparing a powder containing ratatopherin and a polymer electrolyte, dispersing the powder in an aqueous solution, A method comprising mixing under heating to form a complex, and recovering the resulting complex.  [19] A method for producing a complex of ratatopherin and a polymer electrolyte, which comprises preparing a powder containing ratatopherin and a polymer electrolyte which is a polymer acid, and dispersing the powder in an aqueous solution, Mix at room temperature or under heating, then add an aqueous solution or powder containing a polymer electrolyte which is a polymer base, and mix at room temperature or under heating to form a complex of ratatofelin, a polymer acid and a polymer base. And collecting the resulting complex.  [20] The method according to claim 18 or 19, wherein the powder containing ratatopherin and a polyelectrolyte is prepared by mixing at normal temperature or under heating. [21] The method according to claim 18 or 19, wherein each of the extraferlin and the polyelectrolyte is previously coated with a lipid film. [22] A powder containing Latatopherin coated with a lipid film and a powder containing a polyelectrolyte coated with a lipid film separately or in a mixed state in a container. Raw material product for preparing a complex of water and polymer electrolyte.