TW201306760A - Use of nutritional compositions including lactoferrin in supporting resistance to diseases and conditions - Google Patents

Abstract

The present disclosure relates to the use of nutritional compositions including lactoferrin from a non-human source in supporting resistance to diseases or conditions. The method includes administering to a human a nutritional composition comprising a fat or lipid source, a protein source and lactoferrin produced by a non-human source.

Description

Use of a nutritional composition comprising lactoferrin to support resistance to disease and condition

The present disclosure generally relates to the field of nutritional compositions, such as infant formulas, human milk fortified supplements, children's dietary supplements, and the like, which have lactoferrin, particularly lactoferrin produced by non-human sources. More specifically, the present disclosure relates to a method of supporting a resistance to diseases and conditions caused by bacterial and viral pathogens by administering a nutritional composition comprising lactoferrin.

There are currently a variety of dietary compositions for people, especially young people, to provide supplemental or major nutrition at certain stages of life. In general, commercially available dietary compositions for infants seek to mimic the composition of human milk as much as possible and the degree of related functionality. By combining some of the physiologically active proteins and blended fat components, the dietary compositions are formulated so that they mimic human milk for use as a complete or partial replacement. Other ingredients commonly utilized in infant dietary compositions may include, for example, carbohydrate sources of lactose and vitamins, minerals, and elements present in human milk for absorption by infants.

Lactoferrin is one of the major proteins in human milk and is considered to be a glycoprotein having an average molecular weight of about 80 kilodaltons. It is an iron-binding protein that has the ability to bind two iron molecules in a reversible manner and promotes iron uptake in the human small intestine. Functionally, lactoferrin regulates iron uptake and thereby binds iron-based free radicals and supplies iron for immune response.

In a commercially viable dietary composition, lactoferrin is added Often limited, due to expected loss of activity during processing. For example, the temperature and pH requirements in processing infant formulas and other such as human milk fortified supplements and various children's products typically reduce the specific function of lactoferrin, resulting in the absence of lactoferrin in the final formulation. Furthermore, lactoferrin is often only considered for its iron binding; therefore, when such properties are considered to be reduced by processing conditions, lactoferrin may generally be excluded from the preparation.

Again, as is known in the art, human milk has a relatively low iron content (about 0.3 milligrams of iron per liter of human milk). Although low in content, human infants have a high absorption rate and absorb about half of the iron in human milk. However, for example, when a human infant is given a prior art formulation with a high amount of iron fortification of about 10 mg to about 12 mg per liter, the infant absorbs less than about 5% of the total iron. With the high iron content in this prior art formulation, although lactoferrin is a known iron transport protein, substantially all of the lactoferrin iron binding sites are expected to be occupied.

Other difficulties with this prior art formulation include the inability to provide bacteriostatic action. This is due in part to the use of lactoferrin having a blocked or damaged binding site, and the bacteriostatic effect is at least partially related to the degree of iron binding of the lactoferrin present in the formulation.

Therefore, it is advantageous to provide a nutritional composition containing lactoferrin, particularly lactoferrin from a non-human source, such as an infant formula, a human milk nutritional supplement, a dietary supplement for children, and the like. Given the prevalence of viral respiratory infections in humans, especially infants and children, it would be particularly beneficial to provide a nutritional composition that supports resistance to viral respiratory infections. Preferably, even After processing at elevated temperatures and low pH, the lactoferrin included in the composition is capable of supporting resistance to diseases and conditions caused by bacterial and viral pathogens.

Summary of invention

Briefly, in one embodiment, the present disclosure is directed to a method for the body to support resistance to a disease or condition, such as a viral respiratory infection. In another embodiment, the present disclosure relates to a method for human body support against a disease or condition caused by at least one pathogen selected from the group consisting of enterotoxic Escherichia coli (ETEC), intestinal pathogenicity Escherichia coli (EPEC), Haemophilus influenza , Shiga toxin-producing Escherichia coli (STEC), intestinal agglutinating Escherichia coli (EAEC), Salmonella typhimurium serotype ( Salmonella ser. Typhimurium) , Shigella flexneri , rotavirus, norovirus, respiratory syncytial virus (RSV), adenovirus, and combinations thereof. The method comprises administering to a human nutritional composition comprising (1) a source of fat or lipid, (2) a source of protein, and (3) lactoferrin produced by a non-human source. In certain embodiments, the disease or condition is a viral lower respiratory tract infection.

In one embodiment, the nutritional composition comprises: a. a fat or lipid source of up to about 7 g/100 kcal, more preferably a fat or lipid source of from about 3 g/100 kcal to about 7 g/100 kcal; b. A protein source of about 5 g/100 kcal, preferably about 1 a source of protein from g/100 kcal to about 5 g/100 kcal; and c. lactoferrin of at least about 10 mg/100 kCal, more preferably from about 70 mg to about 220 mg/100 kCal of lactoferrin, and most Preferably it is from about 90 mg to about 190 mg/100 kCal of lactoferrin. Optionally, in certain embodiments, the nutritional composition may further comprise from about 0.1 g/100 kcal to about 1 g/100 kcal of a probiotic composition, for example comprising polydextrose and/or galactooligosaccharide The beneficial composition of the sugar. More preferably, the nutritional composition comprises from about 0.3 g/100 kcal to about 0.7 g/100 kcal of a probiotic composition comprising a combination of polydextrose and galactooligosaccharide. In certain embodiments, the disclosure relates to a method for human support for resistance to a disease or condition caused by at least one pathogen selected from the group consisting of: ETEC, EPEC, Shiga toxin-producing Escherichia coli, EAEC , S. typhimurium serum S. serovar, dysentery bacilli or a combination thereof.

Preferably, the lactoferrin is a non-human lactoferrin and/or human lactoferrin produced by a genetically modified organism. In a particularly preferred embodiment, the lactoferrin used is such that an effective amount of the lactoferrin-containing nutritional composition can be administered to an individual to support resistance to a disease or condition caused by a viral or bacterial pathogen. Even during processing, the nutritional composition has been exposed to typical pH and temperature fluctuations similar to those of Pasteur's sterilization processing conditions.

Detailed description of the invention

In one embodiment, the present disclosure provides a disease or condition by bacterial or viral pathogens by administering a human nutritional composition for human support (eg, A method of resistance, such as a viral respiratory infection, comprising a fat or lipid source, a source of protein, and lactoferrin from a non-human source.

As used herein, "lactoferrin produced by a non-human source" and "lactoferrin derived from a non-human source" each mean lactoferrin produced by or taken from human milk other than human milk. For example, lactoferrin for use in the present disclosure includes human lactoferrin and non-human lactoferrin produced by genetically modified organisms. The term "organism" as used herein refers to any persistent living system, such as an animal, plant, fungus or microorganism. As used herein, the term "non-human lactoferrin" refers to lactoferrin having an amino acid sequence that differs from the human lactoferrin amino acid sequence.

Lactoferrin is a single-chain polypeptide of about 80 kD, containing 1-4 glycans depending on the species. The lactoferrin 3-D structures of different species are very similar, but not identical. Each lactoferrin contains two isoform leaf-like structures, called N-leaf and C-leaf, which refer to the N-terminus and C-terminus, respectively. Each leaf-like structure is further composed of two secondary leaves or domains which form a fissure in which the iron ion (Fe ³⁺ ) system and the (di)carbonate anion cooperate in close cooperation. These domains are referred to as N1, N2, C1, and C2, respectively. The N-terminus of lactoferrin has a strong cationic peptide region that is responsible for many important binding properties. Lactoferrin has a very high isoelectric point (~pI 9) and its cationic nature plays a major role in its ability to defend against bacterial, viral and fungal pathogens. Several groups of cationic amino acid residues in the N-terminal region of lactoferrin mediate the biological activity of lactoferrin against various microorganisms. For example, the N-terminal residues 1-47 of human lactoferrin (bovine lactoferrin is 1-48) are critical for the iron-independent biological activity of lactoferrin. In human lactoferrin, residues 2 to 5 (RRRR) (ie Arg-Arg-Arg-Arg; SEQ ID NO: 1) and 28 to 31 (RKVR) (ie Arg-Lys-Val-Arg; SEQ ID NO: 2) is rich in arginine cation domain at the N-terminus, which is particularly important for the antimicrobial activity of lactoferrin. A similar region in the N-terminus was found in bovine lactoferrin (residues 17 to 42; FKCRRWQWRMKKLGAPSITCVRRAFA; ie Phe-Lys-Cys-Arg-Arg-Trp-Gln-Trp-Arg-Met-Lys-Lys-Leu-Gly- Ala-Pro-Ser-Ile-Thr-Cys-Val-Arg-Arg-Ala-Phe-Ala; SEQ ID NO: 3).

As described in "The Perspectives on Interactions Between Lactoferrin and Bacteria " (this article appears in BIOCHEMISTRY AND CELL BIOLOGY, pp 275-281 (2006)), lactoferrin from different host species usually has a relatively high isoelectric point. There is a positively charged amino acid in the end region of the inner leaf, but the amino acid sequence is different. Lactoferrin suitable for use in the present disclosure includes the HLf (349-364) fragment having the amino acid sequence AVGEQELRKCNQWSGL (ie, Ala-Val-Gly-Glu-Gln-Glu-Leu-Arg-Lys-Cys-Asn-Gln -Trp-Ser-Gly-Leu; SEQ ID NO: 4) at least 48% homology. The lactoferrin has an amino acid sequence AVGEQELRKCNQWSGL in the HLf (349-364) fragment (ie, Ala-Val-Gly-Glu-Gln-Glu-Leu-Arg-Lys-Cys-Asn-Gln-Trp-Ser-Gly -Leu, SEQ ID NO: 4) at least 65% homology, and in an embodiment, at least 75% homology. For example, non-human lactoferrin acceptable for use in the present disclosure includes, but is not limited to, bovine lactoferrin, porcine lactoferrin, equine lactoferrin, buffalo lactoferrin, goat lactoferrin, murine lactoferrin, and Camel lactoferrin.

Lactoferrin for use in the present disclosure can be produced, for example, from non-human animal milk or genetically modified organisms. Okonogi et al. disclose a method for making high purity bovine lactoferrin, for example, in U.S. Patent No. 4,791,193, the disclosure of which is incorporated herein by reference. Typically, the method, such as the exposer, includes three steps. First, the raw milk raw material is contacted with a weakly acidic cation exchanger to adsorb lactoferrin, followed by a second step in which rinsing is performed to remove unadsorbed material. The adsorption step is then carried out, at which time the lactoferrin is removed to produce purified bovine lactoferrin. Other methods may include the steps as described in U.S. Patent Nos. 7,368,141, 5,849,885, 5,919,913, and 5,861,491, the entireties of each of

The benefits of lactoferrin, as used in certain embodiments of the present disclosure, support the use of certain bacterial and viral pathogens (including ETEC, EPEC, Haemophilus influenzae, STEC, EAEC, typhoid fever) The ability of S. serovar serotypes, dysentery bacilli, rotavirus, norovirus, RSV, adenovirus, and combinations thereof to be resistant to disease or condition.

In one embodiment, the amount of lactoferrin present in the nutritional composition is at least about 10 mg/100 kCal, particularly when it is desired that the nutritional composition is intended for use in a child. In certain embodiments, the upper limit of lactoferrin is about 240 mg/100 kCal. In another embodiment, wherein the nutritional composition is an infant formula, the amount of lactoferrin present in the nutritional composition is about 70 mg to about 220 mg/100 kCal; in yet another embodiment, the lactoferrin is present in an amount from about 90 mg to about 190 mg/100 kCal. The nutritional composition for the infant can include a lactoferrin amount of from about 0.5 mg to about 1.5 mg per liter of the formulation. In the nutritional composition of the human milk, the lactoferrin content may range from about 0.6 mg to about 1.3 mg per liter of the formulation.

As described, in one embodiment, the nutritional composition supports resistance to viral respiratory infections. People, especially babies and children, are vulnerable to viral respiratory infections. Toxic lower respiratory tract infections, especially those caused by respiratory fusion viruses, are particularly troublesome. In fact, respiratory fusion viruses are the cause of hospitalization for major infants and children and cause, for example, bronchiolitis, pneumonia and long-term complications such as asthma. Thus, in a preferred embodiment, the present disclosure is directed to a method of supporting resistance to viral lower respiratory tract infections. In yet another preferred embodiment, the present disclosure is directed to a method of supporting resistance to a viral infection of the respiratory tract. For example, the nutritional compositions of the present disclosure can be used to support immune development to more effectively prevent or reduce the severity of such infections against viral respiratory infections.

In certain embodiments, the nutritional composition is administered prophylactically to a person not suffering from a disease or condition, such as a viral respiratory infection and/or by at least one pathogen selected from the group consisting of Diseases or conditions: ETEC, EPEC, Haemophilus influenzae, STEC, EAEC, Salmonella typhimurium serotype, Paragonimus, rotavirus, norovirus, RSV, adenovirus and their combination. In other embodiments, the person to whom the nutritional composition is administered has a viral respiratory sensation Dyeing, or the nutritional composition, when administered, has a disease or condition caused by at least one pathogen.

Preferably, the person to whom the nutritional composition is administered is an infant or a child. As used herein, the term "infant" is generally defined as a person born to 12 months of age. "Children" is defined as a person who is over 12 months to about 12 years old.

Preferably, the lactoferrin in the nutritional composition has the same or similar activity against the disease or condition as the free form of lactoferrin. In other words, it is preferred that the activity of the lactoferrin is not significantly reduced in the nutritional composition. It is also preferred that the lactoferrin included in the composition is capable of supporting resistance to diseases and conditions caused by bacterial and viral pathogens even after processing under high temperature and low pH conditions. In one embodiment of the present disclosure, the lactoferrin used is non-human lactoferrin.

Surprisingly, for example, even exposure to low pH (ie, below about 7, and even as low as about 4.6 or lower) and/or elevated temperatures (ie, in excess of about 65 ° C, and up to about 120 ° C) Bovine lactoferrin maintains relevant activity for conditions expected to disrupt or severely limit human lactoferrin stability or activity. These low pH and/or high temperature conditions, such as Pasteurization, are contemplated during certain processing modes for the nutritional composition types described herein. However, the amino acid composition of bovine lactoferrin has about 70% sequence homology with human lactoferrin and is stable and still active under conditions in which human or recombinant human lactoferrin becomes unstable or inactive. At the same time, bovine lactoferrin has bactericidal activity against undesired bacterial pathogens found in the human gut.

In some embodiments, the nutritional composition can be an infant formula. The term "infant formula" refers to a composition that is a human milk substitute, in liquid or powder form, that meets the nutritional needs of the infant. In the United States, infant formula levels are specified in Sections 100, 106, and 107 of Title 21 of the US Code of Federal Regulations. These regulations define large amounts of nutrients, vitamins, minerals and other ingredients to mimic the nutritional and other properties of human milk. In another embodiment, the nutritional product may be a human milk fortified nutritional product, meaning that it is a composition added to human milk to enhance the nutritional value of human milk. As a human milk fortified nutritional product, the disclosed composition may be in powder form or in liquid form. In yet another embodiment, the nutritional product of the present disclosure can be a nutritional composition for a child.

The nutritional composition of the present disclosure provides minimal, partial or complete nutritional support. The nutritional composition can be a nutritional supplement or a meal replacement. In some embodiments, the nutritional composition can be administered in conjunction with food or another nutritional composition. In this embodiment, the nutritional composition can be intermixed with the food or other nutritional composition prior to ingestion by the individual, or can be administered to the individual before or after the individual ingests the food or nutritional composition. The nutritional composition can be administered to premature babies who receive infant formula, breast milk, human milk fortified nutrition, or a combination thereof. For the purposes of this disclosure, "premature babies" are babies born after less than about 37 weeks of gestation, and "full-term babies" means babies born at least about 37 weeks after gestation.

The nutritional composition can be (but not necessarily) nutritionally intact. Those skilled in the art will appreciate that "nutrition integrity" will vary depending on a number of factors including, but not limited to, the age, clinical condition, and dietary intake of the individual to which the term applies. In general, "nutritional integrity" means the nutritional composition of the present disclosure. Provides all the right amount of carbohydrates, lipids, essential fatty acids, proteins, essential amino acids, conditionally essential amino acids, vitamins, minerals and energy required for normal growth. If referring to nutrients, the term "essential" means that any body cannot synthesize nutrients that are sufficient for normal growth and to maintain health and therefore must be supplied by the diet. The term "conditional necessity", if referring to a nutrient, means that the nutrient must be supplied by the meal under conditions in which the body is unable to obtain an appropriate amount of precursor compound for endogenous synthesis.

For premature babies, the "nutritious" composition is defined as providing the right amount of carbohydrates, lipids, essential fatty acids, proteins, essential amino acids, conditionally essential amino acids, etc. required for all growth of the premature baby. Vitamins, minerals and energy. For term infants, the "nutritional" composition is defined as the amount of carbohydrate, lipid, essential fatty acid, protein, essential amino acid, and essential amino groups required to provide all the growth of the full-term infant. Acid, vitamins, minerals and energy. For children, the “nutritional” composition is defined as providing the right amount of carbohydrates, lipids, essential fatty acids, proteins, essential amino acids, conditionally essential amino acids, vitamins, Minerals and energy.

The nutritional composition can be provided in any form known in the art, including powders, gels, suspensions, pastes, solids, liquids, liquid concentrates or ready-to-finish products. In a preferred embodiment, the nutritional composition is an infant formula that is particularly suitable for use as an infant formula that is the sole source of nutrition for the infant.

In a preferred embodiment, the nutritional product disclosed herein can be administered enterally. As used herein, "intestinal" means through the gastrointestinal tract or digestive tract or In the gastrointestinal or digestive tract, and "intestinal administration" includes oral feeding, intragastric feeding, transpyloric administration or any other introduction into the digestive tract.

Suitable fat or lipid sources for practicing the present invention can be any of those known or used in the art, including but not limited to animal sources such as milk fat, butter, butter fat, Egg yolk lipids; sources of aquatic products, such as: fish oil, aquatic oils, single-cell oils; vegetables and vegetable oils, such as: corn oil, rapeseed oil, sunflower oil, soybean oil, palm oil, palm oil, high oleic acid Sunflower oil, evening primrose oil, canola oil, olive oil, linseed oil, cottonseed oil, high oleic safflower oil, palm stearin, palm kernel oil, wheat germ oil; medium chain triglyceride and Emulsifiers and esters of fatty acids; and any combination thereof.

In certain embodiments, the source of protein included in the nutritional composition comprises cow's milk protein. Milk protein sources useful in the practice of the present disclosure include, but are not limited to, milk protein powder, milk protein concentrate, milk protein isolate, skim milk solids, skim milk, skimmed milk powder, whey protein, whey protein isolate, whey protein Concentrate, sweet whey, acid whey, casein, acid casein, casein salt (eg, casein sodium, casein sodium calcium, casein calcium) and any combination thereof.

In one embodiment, the protein is provided as an intact protein. In other embodiments, the protein is provided as both intact protein and partially hydrolyzed protein, with a degree of hydrolysis ranging from about 4% to 10%. In yet another embodiment, the protein source may be supplemented with a branic acid peptide.

In a particular embodiment of the present disclosure, the protein source comprises whey and casein, and the ratio of whey to casein is similar to the ratio of whey to casein in human milk. For example, in certain embodiments, the weight ratio of whey to casein is about 20% whey: 80% casein to about 80% whey: 20% casein.

In certain embodiments of the present disclosure, the nutritional composition may contain one or more probiotics. The term "probiotic" means a low pathogenic or non-pathogenic microorganism that has a positive effect on host health. Any probiotic known in the art is acceptable in the present disclosure as long as the desired result is achieved. In a specific embodiment, the probiotic may be selected from the group consisting of Lactobacillus , Lactobacillus rhamnosus GG, Bifidobacterium , Bifidobacterium longum. ), Bifidobacterium brevis and Bifidobacterium animalis subsp . lactis BB-12.

If included in the composition, the probiotic amount can be from about 10 ⁴ to about 10 ¹⁰ colony forming units (cfu) per kg body weight per day. In another embodiment, the probiotic amount can be from about 10 ⁶ to about 10 ⁹ cfu per kg body weight per day. In yet another aspect of the embodiment, the amount of the probiotic per kg body weight per day may be at least about 10 ⁶ cfu. Again, the disclosed compositions can also include probiotic-conditioned media components.

In one embodiment, one or more probiotics are viable bacteria. In another embodiment, one or more probiotic strains are non-viable. As used herein, the term "live bacteria" refers to a living microorganism. The term "non-living bacteria" or "non-living" "Probiotics" means non-living probiotic microorganisms, their cellular composition and their metabolites. Such non-living probiotics may be heat-killed or otherwise inactivated, but remain capable of beneficially affecting the health of the host. In the present disclosure, useful probiotics can be developed naturally, synthetically, or through genetically manipulated organisms, whether such new sources are known to date or later.

In one embodiment of the present disclosure, the nutritional composition can include a probiotic composition comprising one or more beneficial bacteria. As used herein, the term "probiotics" means to positively affect a non-digestible food component of a host by selectively stimulating one or a few bacterial growth and/or activities in the colon that improve the health of the host. "Probiotics composition" is a composition containing one or more beneficial bacteria. Such probiotics can be developed naturally, synthetically or through genetically manipulated organisms and/or plants, whether such new sources are known to date or later. In certain embodiments, the disclosure of the composition of the present disclosure includes the teachings of U.S. Patent No. 7,572,474, the disclosure of which is incorporated herein by reference.

The probiotics used in the present disclosure can be developed naturally, synthetically or through genetically manipulated organisms and/or plants, whether such new sources are known today or later. Probiotics useful in the present disclosure may include oligosaccharides, polysaccharides, and other probiotics containing fructose, xylose, soy, galactose, glucose, and mannose. More specifically, the probiotics useful in the present disclosure may include lactulose, lactosucrose, raffinose, glucose oligosaccharides, inulin, polydextrose, polydextrose powder, galactose Oligosaccharide, fructose oligosaccharide, isomaltose oligosaccharide, soybean oligosaccharide, lactulose oligosaccharide, xylose oligosaccharide, chito-oligosaccharide, mannose Sugar, arabinose oligosaccharide, sialic oligosaccharide, trehalose oligosaccharide, and gentio-oligosaccharide. Preferably, the nutritional composition comprises polydextrose (PDX) and/or galactooligosaccharide (GOS). Optionally, in addition to polydextrose and/or galactooligosaccharide, the nutritional composition comprises one or more additional probiotics.

If included in the nutritional composition, the total amount of beneficial bacteria present in the nutritional composition may range from about 0.1 g/100 kcal to about 1 g/100 kcal. More preferably, the total amount of beneficial bacteria present in the nutritional composition may range from about 0.3 g/100 kcal to about 0.7 g/100 kcal. At least 20% of the probiotics should contain galactose oligosaccharides and/or polydextrose.

In one embodiment, if polydextrose is used in the probiotic composition, the amount of polydextrose in the nutritional composition can range from about 0.1 g/100 kcal to about 1 g/100 kcal. In the range. In another embodiment, the amount of polydextrose in the nutritional composition can range from about 0.2 g/100 kcal to about 0.6 g/100 kcal.

In one embodiment, if galactose oligosaccharide is used in the probiotic composition, the amount of galactooligosaccharide in the nutritional composition may range from about 0.1 g/100 kcal to about 1 g/100 kcal. In the range. In another embodiment, the amount of galactooligosaccharide in the nutritional composition is in the range of from about 0.2 g/100 kcal to about 0.5 g/100 kcal. In certain embodiments, the ratio of polydextrose to galactooligosaccharide in the probiotic composition is between about 9:1 and about 1:9.

In some embodiments, the nutritional compositions of the present disclosure may additionally comprise a source of long chain polyunsaturated fatty acids (LCPUFA). Preferably, the LCPUFA The source contains docosahexaenoic acid (DHA). Other suitable LCPUFAs include, but are not limited to, alpha-linolenic acid, gamma-linolenic acid, linoleic acid, linoleic acid, eicosapentaenoic acid (EPA), and arachidonic acid (ARA).

In one embodiment, the nutritional composition is supplemented with both DHA and ARA. In this embodiment, the ARA:DHA weight ratio can range from about 1:3 to about 9:1. In one embodiment of the present disclosure, the ARA:DHA weight ratio is from about 1:2 to about 4:1.

The amount of long chain polyunsaturated fatty acids in the nutritional composition may range from about 5 mg/100 kcal to about 100 mg/100 kcal, more preferably from about 10 mg/100 kcal to about 50 mg/100 kcal.

The nutritional composition can be supplemented with DHA and ARA containing grease using standard techniques known in the art. For example, DHA and ARA can be added to the composition by substituting an equivalent amount of a fat (eg, high oleic sunflower oil) normally present in the composition. As another example, a grease containing DHA and ARA can be added to the composition by substituting an equivalent amount of the remaining bulk fat blend typically present in the DHA and ARA compositions.

If used, the DHA and ARA sources can be of any known origin in the art, such as aqua oils, fish oils, single cell fats, egg yolk lipids, and brain lipids. In some embodiments, the DHA and ARA are derived from single cell Martek oil, DHASCO®, and ARASCO®, respectively, or variations thereof. The DHA and ARA may be in a natural form, but the remainder of the LCPUFA source will not cause any substantial deleterious effects on the infant. Alternatively, the DHA and ARA can be used in a refined form.

In one embodiment of the present disclosure, the DHA and ARA sources are the single cell greases taught in U.S. Patent Nos. 5,374,567, 5, 550, 156, and 5, 397, 591, the entireties of each of which are incorporated herein by reference. However, the disclosure is not limited to such fats and oils.

In certain embodiments, the nutritional composition comprises from about 0.5 mg/100 kcal to about 5 mg/100 kcal of iron, including iron in combination with lactoferrin.

Instance

The following examples are provided to illustrate some embodiments of the nutritional compositions of the present disclosure, but should not be construed as limiting any of them. Other embodiments within the scope of the patent application scope herein will be apparent to those skilled in the art in view of the specification or practice of the invention. The description and the examples are intended to be illustrative only, and the scope and spirit of the disclosure are defined by the appended claims.

Example 1

This example illustrates the effect of lactoferrin and infant formula on the growth of diarrhea E. coli strains in vitro.

Obtained fifteen clinical diarrhea from various DEC groups (ETEC, Enteropathogenic Escherichia coli (EPEC), Shiga toxin-producing Escherichia coli (STEC), and Enteroagglutinin Escherichia coli (EAEC) E. coli (DEC) strain. As mentioned previously, the bacteria Strains were isolated from children with diarrhea in the cohort study and identified by an instant polymerase chain reaction (PCR) against the diarrhea E. coli population. In addition, fifteen species of clinical isolates of Salmonella typhimurium serotypes and fifteen species of dysentery bacilli were obtained.

All clinical strains and standard laboratory control strains were grown on McConekey vegetable medium for 24 hours at 37 °C. For survival assays, the strains were grown in lysogeny broth (LB) medium at 37 ° C for 18 hours. Next, the strains were washed twice with PBS and centrifuged at 4500 x g for 5 minutes. Use only bacteria in the mid-log phase.

0, 0.6, 1, 2, 4, 6, 8, and 10 mg/ml stock solutions of lactoferrin and infant formulas containing 2.0 g/100 kcal of milk protein and 1.8 mg/100 kcal of iron 0, 0.6, 1 The 2, 4, 6, 8 and 10 mg/ml stock solutions were prepared in distilled water.

A clinical strain culture inoculum containing about 2 x 10 ⁸ log phase cells was inoculated into a 96-well dish containing 1% bacterial peptone (bactopeptone). Each well plate was also injected into the infant formula or lactoferrin stock solution via microdilution to test each strain against 0, 0.6, 1, 2, 4, 6, 8 and 10 mg/ml lactoferrin or infant formula. . The wells were incubated at 37 ° C and the growth kinetics were monitored every 30 minutes by serially culturing the inoculum. Growth is then monitored by a spectrophotometer and/or ELISA reader. After a total of 18-20 hours of culture, the lowest concentration of the infant formula or lactoferrin that caused complete bacterial inhibition against each strain was recorded as MIC (minimum inhibitory concentration).

Co-assay is also tested to test the resistance of both lactoferrin and infant formula The activity of each strain, as well as the effect of the lactoferrin/infant formula combination on bacterial growth, was measured as described above. For these synergistic assays, lactoferrin and infant formula concentrations were determined for each reagent based on MIC studies and growth kinetics results.

Example 2

This example illustrates the effect of lactoferrin and infant formula on adhesion of diarrhea E. coli strains to human intestinal epithelial cells in vitro.

Infected with enterotoxic Escherichia coli, enteropathogenic Escherichia coli, Shiga toxin-producing Escherichia coli, intestinal agglutinating Escherichia coli, Salmonella typhimurium serotype or Paratuberculosis isolate described in Example 1 Subconfluent) Hep2 cell layer (approximately 5 x 10 ⁴ cells/well in a 24-well plate). The infant formula with or without 10 mg/ml lactoferrin was then added to achieve a bacteria to infant formula concentration of 100:1. Next, the infected Hep2 cell layer was cultured for 4 hours at 37 ° C under 5% CO ₂ . Thereafter, the Hep2 cells were vigorously washed to remove unadhered bacteria. The cells were fixed with 70% methanol, stained with 10% Giemsa solution and microscopically examined under a microscope. In addition, the fluorescent actin staining assay (FAS assay) was performed as previously reported for enteropathogenic Escherichia coli, Shiga toxin-producing Escherichia coli, Paratuberculosis, and Salmonella typhimurium serotypes.

Example 3

This example illustrates the use of lactoferrin and infant formula in vivo to support The effect of the disease caused by the bacterial pathogen or the resistance of the disease.

Healthy female Balb/c strain mice aged 6 to 8 weeks and weighing between 20 and 24 g were obtained and divided into experimental group and control group. The test group was then fed 200 μl of an infant formula containing 75 or 165 mg/ml lactoferrin prior to infection, and the control group was fed a 200 μl infant formula prior to infection. The amount of infant formula administered was adjusted so that the amount received by the mice was equivalent to 600 mg/kg and 1333 mg/kg of lactoferrin per day. Thereafter, 300 μL of 10 ⁸ colony forming units (cfu) of Salmonella typhimurium serovar serotype, 200 μL of 10 ⁸ cfu of Citrobacter rodentium (EPEC rat model) or 200 μL of 10 ⁸ The mice were infected with Cfu's dysentery bacilli. For infection and pretreatment inoculation, a gavage needle is used. After infection, the mice received either an infant formula containing 75 or 165 mg/ml lactoferrin or a separate infant formula for 7 days each. In addition, the amount of infant formula administered was adjusted so that the amount received by the mice was equivalent to 600 mg/kg and 1333 mg/kg of lactoferrin per day. Seven days after infection, all mice were monitored daily for death, body weight, and clinical signs (piloerection, back arched posture, and reduced movement). The incidence of clinical signs was determined by comparing the behavior of individual infected mice for 15 minutes. On the 10th day after infection, the mice were sacrificed and subjected to cardiac puncture for blood culture. Organs (colon, liver, and spleen) were removed for histopathological analysis. Studying the degree of inflammation and necrosis in the organ with the pathologist, the pathologist assigns blindness to the group to avoid bias.

Example 4

This example illustrates the effect of lactoferrin and infant formulas in vivo to support resistance to conditions or diseases caused by viral pathogens.

Example 3 was carried out as described above, but the mice were infected with rotavirus, norovirus, astrovirus, adenovirus and calicivirus instead of bacterial strains. The monitoring and research of mice is as described above.

Example 5

This example illustrates an embodiment of a nutritional product in accordance with the present disclosure.

Example 6

This example illustrates another embodiment of a nutritional product in accordance with the present disclosure.

Example 7

This example illustrates an embodiment of an ingredient that can be used to prepare a nutritional product in accordance with the present disclosure.

Example 8

This example illustrates another embodiment of an ingredient that can be used to prepare a nutritional product in accordance with the present disclosure.

Preferably, the nutritional composition is administered to a human and supports resistance to a human disease or condition caused by a bacterial or viral pathogen, including a viral respiratory infection.

All references cited in this manual, including but not limited to all papers, publications, patents, patent applications, publications, texts, reports, manuscripts, pamphlets, books, online articles, journal articles, periodic publications, etc. All of them are incorporated herein by reference. The discussion of the references in this article is intended only to summarize the claims of its authors and not to act as Recognize that any reference material constitutes prior art. Applicants reserve the right to challenge the accuracy and pertinence of the cited references.

Although the preferred embodiments of the present disclosure have been described using specific terms, devices, and methods, this description is for illustrative purposes only. The text used is explanatory text and not restrictive text. It is to be understood that changes and modifications may be made by those skilled in the art without departing from the spirit and scope of the disclosure. In addition, it should be understood that various embodiments may be substituted in whole or in part. For example, while methods for making commercially available sterile liquid nutritional supplements made according to such methods have been illustrated, other uses are also contemplated. Therefore, the spirit and scope of the appended claims should not be limited to the preferred aspects contained in the description.

<110> Mead Johnson Nutrition Company

<120> Use of a nutritional composition comprising lactoferrin to support resistance to diseases and conditions

<140>

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<150> US 12/980,825

<151> 2010-12-29

<150> US 12/980,833

<151> 2010-12-29

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Claims (15)

A use of a nutritional composition for the preparation of a medicament for the body to support resistance to a disease or condition, the nutritional composition comprising: a) a source of fat or lipid; b) a source of protein; and c) manufactured by a non-human source Lactoferrin, wherein the lactoferrin has at least 48% homology to the amino acid sequence AVGEQELRKCNQWSGL of the HLf (349-364) fragment. The use of the first aspect of the patent application, wherein the disease or condition is caused by at least one pathogen selected from the group consisting of enterotoxic Escherichia coli, enteropathogenic Escherichia coli, Haemophilus influenzae ( Haemophilus influenza) ), Shiga toxin-producing Escherichia coli, intestinal agglutinating Escherichia coli, Salmonella typhimurium serotype ( Salmonella ser. Typhimurium), Shigella flexneri , Rotavirus, Norovirus, Respiratory fusion virus , adenovirus and their combination. The use of the first aspect of the patent application, wherein the disease or condition is a viral respiratory infection. For example, the application of the scope of claim 1 wherein the person is an infant or a child. The use of claim 1, wherein the fat or lipid source is present in an amount from about 3 g/100 kcal to about 7 g/100 kcal, and the protein source is from about 1 g/100 kcal to about 5 The amount of g/100 kcal exists. The use of claim 1 wherein the lactoferrin is present in an amount of at least about 10 mg/100 kcal. The use of claim 6 wherein the lactoferrin is present in an amount from about 70 mg/100 kcal to about 220 mg/100 kcal. The use of the first aspect of the invention, wherein the lactoferrin is selected from the group consisting of non-human lactoferrin, human lactoferrin produced by genetically modified organisms, or a combination thereof. The use of the first aspect of the patent application, wherein the lactoferrin is stable and still active under conditions in which human lactoferrin becomes unstable or inactive. The use of the ninth aspect of the patent application, wherein the nutritional composition has been treated under pasteurization conditions. The use of claim 1, wherein the nutritional composition further comprises a prebiotic composition comprising a compound selected from the group consisting of galactose oligosaccharides, polydextrose, or a combination thereof. The use of claim 1, wherein the nutritional composition comprises from about 0.5 mg/100 kcal to about 5 mg/100 kcal of iron comprising iron bound to lactoferrin. The use of claim 1 wherein the lactoferrin has at least 65% homology to the amino acid sequence AVGEQELRKCNQWSGL of the HLf (349-364) fragment. The use of claim 1, wherein the person has the disease or condition when the nutritional composition is administered. Such as the use of the first item of the patent scope, wherein the nutritional composition The system is administered prophylactically, and when the nutritional composition is administered, the person does not have the disease or condition.