U.S. patent application number 15/866322 was filed with the patent office on 2018-05-17 for nutritional formulations including human milk oligosaccharides and antioxidants and uses thereof.
This patent application is currently assigned to ABBOTT LABORATORIES. The applicant listed for this patent is ABBOTT LABORATORIES. Invention is credited to RACHAEL BUCK, GERALYN O. DUSKA-MCEWEN, JOSEPH P. SCHALLER.
Application Number | 20180133235 15/866322 |
Document ID | / |
Family ID | 45496314 |
Filed Date | 2018-05-17 |
United States Patent
Application |
20180133235 |
Kind Code |
A1 |
BUCK; RACHAEL ; et
al. |
May 17, 2018 |
NUTRITIONAL FORMULATIONS INCLUDING HUMAN MILK OLIGOSACCHARIDES AND
ANTIOXIDANTS AND USES THEREOF
Abstract
Disclosed are nutritional compositions including human milk
oligosaccharides in combination with long chain polyunsaturated
fatty acids and/or carotenoids that can be administered to preterm
infants, term infants, toddlers, and children for reducing
inflammation and the incidence of inflammatory diseases.
Inventors: |
BUCK; RACHAEL; (Gahanna,
OH) ; DUSKA-MCEWEN; GERALYN O.; (Columbus, OH)
; SCHALLER; JOSEPH P.; (Reynoldsburg, OH) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
ABBOTT LABORATORIES |
Abbott Park |
IL |
US |
|
|
Assignee: |
ABBOTT LABORATORIES
Abbott Park
IL
|
Family ID: |
45496314 |
Appl. No.: |
15/866322 |
Filed: |
January 9, 2018 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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15804751 |
Nov 6, 2017 |
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15866322 |
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14195938 |
Mar 4, 2014 |
9808474 |
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15804751 |
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13334995 |
Dec 22, 2011 |
8703737 |
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14195938 |
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61428861 |
Dec 31, 2010 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P 31/12 20180101;
A23V 2002/00 20130101; A61K 31/202 20130101; A61K 31/202 20130101;
A23V 2200/3202 20130101; A61K 31/7016 20130101; A61P 31/00
20180101; A23L 33/40 20160801; A61P 37/00 20180101; A61K 31/702
20130101; A23V 2200/324 20130101; A23V 2250/28 20130101; A61P 39/06
20180101; A61K 45/06 20130101; A23L 33/12 20160801; A61P 1/00
20180101; A61K 2300/00 20130101; A61K 2300/00 20130101; A61K
31/7016 20130101; A61P 11/00 20180101; A61K 31/702 20130101; A61K
2300/00 20130101; A23V 2250/1882 20130101; A23V 2200/324 20130101;
A61K 2300/00 20130101; A23V 2250/28 20130101; A23V 2200/3202
20130101; A23V 2002/00 20130101; A61P 29/00 20180101; A23V
2250/1882 20130101 |
International
Class: |
A61K 31/702 20060101
A61K031/702; A23L 33/00 20060101 A23L033/00; A23L 33/12 20060101
A23L033/12; A61K 31/7016 20060101 A61K031/7016; A61K 31/202
20060101 A61K031/202; A61K 45/06 20060101 A61K045/06 |
Claims
1. A nutritional composition comprising a human milk
oligosaccharide selected from the group consisting of
6'-sialyllactose, 3'-sialyllactose, 2'-fucosyllactose,
3-fucosyllactose, lacto-N-tetraose, lacto-N-neotetraose,
lacto-N-fucopentaose, and combinations thereof, and from 40 mg/L to
185 mg/L of total long chain omega-3 fatty acids.
2. The nutritional composition of claim 1, wherein the long chain
omega-3 fatty acid comprises docosahexaenoic acid.
3. The nutritional composition of claim 2 further comprising from
0.001 .mu.g/mL to 5 .mu.g/mL of a carotenoid.
4. The nutritional composition of claim 2, wherein the composition
further includes long chain omega-6 fatty acids and a ratio of long
chain omega-6 fatty acids to long chain omega-3 fatty acids is from
about 2:1 to about 3:1.
5. The nutritional composition of claim 1 further comprising a
nucleotide in an amount of from 10 mg/mL to 200 mg/mL.
6. The nutritional composition of claim 1, wherein the nutritional
composition is a nutritional powder or ready-to-feed nutritional
liquid.
7. The nutritional composition of claim 6, wherein the nutritional
composition is a nutritional powder and the powder is a spray dried
powder.
8. The nutritional composition of claim 6, wherein the nutritional
composition is a ready-to-feed nutritional liquid.
9. The nutritional composition of claim 1, wherein the human milk
oligosaccharide is derived from a fermentation process, an
enzymatic process, a chemical process, or a combination
thereof.
10. A nutritional composition comprising a human milk
oligosaccharide selected from the group consisting of
6'-sialyllactose, 3'-sialyllactose, 2'-fucosyllactose,
3-fucosyllactose, lacto-N-tetraose, lacto-N-neotetraose,
lacto-N-fucopentaose, and combinations thereof, from 40 mg/L to 185
mg/L of total long chain omega-3 fatty acids, and a source of
protein, wherein the source of protein comprises partially
hydrolyzed protein.
11. The nutritional composition of claim 10, wherein the long chain
omega-3 fatty acid comprises docosahexaenoic acid.
12. The nutritional composition of claim 10 further comprising from
0.001 .mu.g/mL to 5 .mu.g/mL of a carotenoid.
13. The nutritional composition of claim 10, wherein the
composition further includes long chain omega-6 fatty acids and a
ratio of long chain omega-6 fatty acids to long chain omega-3 fatty
acids ranges from about 2:1 to about 3:1.
14. The nutritional composition of claim 10 further comprising a
nucleotide in an amount of from 10 mg/mL to 200 mg/mL.
15. The nutritional composition of claim 10, wherein the
nutritional composition is a nutritional powder or ready-to-feed
nutritional liquid.
16. The nutritional composition of claim 15, wherein the
nutritional composition is a nutritional powder and the powder is a
spray dried powder.
17. The nutritional composition of claim 15, wherein the
nutritional composition is a ready-to-feed nutritional liquid.
18. The nutritional composition of claim 10, wherein the human milk
oligosaccharide is derived from a fermentation process, an
enzymatic process, a chemical process, or a combination
thereof.
19. A human milk fortifier comprising a human milk oligosaccharide
selected from the group consisting of 6'-sialyllactose,
3'-sialyllactose, 2'-fucosyllactose, 3-fucosyllactose,
lacto-N-tetraose, lacto-N-neotetraose, lacto-N-fucopentaose, and
combinations thereof, and from 0.001 .mu.g/mL to 5 .mu.g/mL of a
carotenoid.
20. The human milk fortifier of claim 19 further comprising from 40
mg/L to 185 mg/L of total long chain omega-3 fatty acids.
21. The human milk fortifier of claim 20, wherein the long chain
omega-3 fatty acid comprises docosahexaenoic acid.
22. The human milk fortifier of claim 20, wherein the composition
further includes long chain omega-6 fatty acids and a ratio of long
chain omega-6 fatty acids to long chain omega-3 fatty acids ranges
from about 2:1 to about 3:1.
23. The human milk fortifier of claim 19 further comprising a
nucleotide in an amount of from 10 mg/mL to 200 mg/mL.
24. The human milk fortifier of claim 19, wherein the human milk
oligosaccharide is derived from a fermentation process, an
enzymatic process, a chemical process, or a combination
thereof.
25. The human milk fortifier of claim 19, wherein the human milk
oligosaccharide is derived from a source selected from human,
bovine, ovine, porcine, caprine, or combinations thereof.
26. The human milk fortifier of claim 19, wherein the human milk
oligosaccharide is lacto-N-fucopentaose.
27. The human milk fortifier of claim 19, wherein the human milk
oligosaccharide is present in an amount of 0.01 mg/mL to 5
mg/mL.
28. A method of reducing inflammation in an infant, toddler, or
child in need thereof, the method comprising administering to the
infant, toddler, or child a nutritional composition comprising a
human milk oligosaccharide selected from the group consisting of
6'-sialyllactose, 3'-sialyllactose, 2'-fucosyllactose,
3-fucosyllactose, lacto-N-tetraose, lacto-N-neotetraose,
lacto-N-fucopentaose, and combinations thereof, and from 40 mg/L to
185 mg/L of total long chain omega-3 fatty acids.
29. The method of claim 28, wherein the nutritional composition
further comprises from 0.001 .mu.g/mL to 5 .mu.g/mL of a
carotenoid.
30. The method of claim 29, wherein the carotenoid is selected from
the group consisting of lutein, lycopene, beta-carotene, and
combinations thereof.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims the benefit of U.S. Provisional
Application No. 61/428,861 filed on Dec. 31, 2010, which disclosure
is incorporated by reference in its entirety.
FIELD OF THE DISCLOSURE
[0002] The present disclosure relates to human milk
oligosaccharides (HMOs) for modulating inflammation in an infant,
toddler, or child. More particularly, the present disclosure
relates to human milk fortifiers, preterm and term infant formulas
and pediatric formulas comprising HMOs that can reduce inflammation
and thereby prevent and/or treat inflammatory conditions and
diseases.
BACKGROUND OF THE DISCLOSURE
[0003] The inflammatory response is an attempt by the body to
restore and maintain homeostasis after invasion by an infectious
agent, antigen challenge, or physical, chemical or traumatic
damage. While the inflammatory response is generally considered a
healthy response to injury, the immune system can present an
undesirable physiological response if it is not appropriately
regulated. Specifically, unregulated oxidation and associated
inflammation are major causes of tissue damage and clinically
significant disease in preterm and term infants. This is due in
large part to the immaturity in function of the natural immune
system of infants, and especially preterm infants.
[0004] Breastfeeding has been associated with enhanced development
and balanced growth and maturation of the infant's respiratory,
gastrointestinal and immune systems, thereby providing protection
of the infant to infection and inflammatory diseases. Breast milk
appears to contain endogenous antioxidants, such as superoxide
dismutase, glutathione peroxidase and catalase, or other
non-enzymatic antioxidants such as glutathione, lactoferrin and
polyphenols, in addition to exogenous antioxidants, such as
vitamins A, C, E and selenium. Further, breast milk includes HMOs
that not only act as pathogen receptor analogues, but activate
immune factors by infant intestinal epithelial cells and/or
associated immune cell populations. The function of these breast
milk components, functioning as antioxidants and as immune
modulators, includes not only the protection of breast milk lipids
by peroxidation, but may also assist in the regulation of
inflammatory responses to infection or other injury.
[0005] Not all infants receive human breast milk. Further, no
vaccines are currently available for the prevention of inflammatory
diseases. Therefore, development of safe and efficacious
preventative or therapeutic methods would be beneficial, especially
for infants.
[0006] It would therefore be desirable to provide nutritional
compositions, and synthetic infant formulas in particular, that can
produce nutritional benefits including improved immune system
growth and development. It would additionally be beneficial if the
nutritional compositions could modulate inflammation and enhance
immunity against microbial infections, including bacterial and
viral infections, and other inflammatory diseases.
SUMMARY OF THE DISCLOSURE
[0007] The present disclosure is directed to nutritional
compositions, including synthetic infant formulas, synthetic
pediatric formulas, and synthetic child formulas, including at
least one HMO, alone or in combination with one or more of long
chain polyunsaturated fatty acids (LCPUFAs), antioxidants, and/or
nucleotides, for reducing inflammation in an infant, toddler, or
child, as well as methods of using the compositions.
[0008] One embodiment is a synthetic pediatric formula comprising a
human milk oligosaccharide and from about 0.025 mg/mL to about
0.130 mg/mL of a long chain polyunsaturated fatty acid.
[0009] Another embodiment is a synthetic pediatric formula
comprising a human milk oligosaccharide and from about 0.001
.mu.g/mL to about 10 .mu.g/mL of a carotenoid.
[0010] Another embodiment is a method of reducing inflammation in
an infant, toddler, or child in need thereof. The method comprises
administering to the infant, toddler, or child a composition
comprising a human milk oligosaccharide and from about 0.025 mg/mL
to about 0.130 mg/mL of a long chain polyunsaturated fatty
acid.
[0011] Another embodiment is a method of reducing inflammation in
an infant, toddler, or child in need thereof. The method comprises
administering to the infant, toddler, or child a composition
comprising a human milk oligosaccharide and from about 0.001
.mu.g/mL to about 10 .mu.g/mL of a carotenoid.
[0012] It has been discovered that specific HMOs, such as
3'-sialyllactose, 6'-sialyllactose and others as noted herein, as
well as specific combinations of HMOs as described herein, are
highly effective in dampening inflammation generally in infants,
toddlers, and children, and specifically in dampening virus-induced
inflammation, including respiratory syncytial virus, human
parainfluenza, and influenza A, in infants, toddlers, and children
by reducing the production of some key cytokines from human immune
cells without increasing viral load, which may lead to faster
recovery from infections. Surprisingly, it was determined that the
HMOs demonstrate the desirable dampening effects even at very low
concentrations, including concentrations lower than those found in
breast milk. Also, it was unexpectedly found that 6'-sialyllactose
is immunomodulatory even in the absence of a virus, and induces the
production of monocyte-derived cytokines. It has further been
discovered that although biological reactions often occur within a
30 to 60 minute period, and thus a 30 to 60 minute incubation is
generally used for in vitro procedures, a 24 hour pre-treatment of
cells provides a closer reflection of the daily pre-exposure to
HMOs that a breast-fed infant would receive from breast milk.
[0013] Additionally, it has been found that fucosyllated HMOs,
including 3'-fucosyllactose, alone or in combination with sialic
acid, are highly effective in inhibiting respiratory viruses. Even
at low concentrations, the 3'-fucosyllactose and sialic acid are
effective.
[0014] Moreover, it has been discovered that specific HMOs act in a
synergistic manner against respiratory viruses, including RSV, when
combined with a long chain polyunsaturated fatty acid and/or a
carotenoid. These synergistic actions dampen virus-induced
inflammatory cytokines, and specifically interferon-inducible
protein 10 (IP-10). Additional components including antioxidants,
such as vitamin A and vitamin E, or nucleotides, may also be added
to the HMO and long chain polyunsaturated fatty acid and/or
carotenoid combinations.
[0015] It has further been found that a combination of HMOs
including acidic/sialylated (e.g., 6'-sialyllactose) and/or
neutral/fucosylated (e.g., 2'-fucosyllactose) and/or
n-acetylglucosylated (e.g., LNnT) prevents the development of
necrotizing entercolitis. Also, these HMOs have been found to
decrease the oxidative stress in infants.
BRIEF DESCRIPTION OF THE DRAWINGS
[0016] FIG. 1 is a graph depicting H1N1 virus infectivity of MDCK
cells in the presence of various HMOs as tested in Example 37.
[0017] FIG. 2 is a graph depicting blood plasma levels of
glutathione from piglets as measured in Example 38.
[0018] FIG. 3 is a graph depicting IP-10 levels resulting from
administration of 3'SL and 6'SL as measured in Example 39.
[0019] FIG. 4 is a graph depicting IP-10 levels resulting from
administration of 3'SL and 6'SL as measured in Example 39.
[0020] FIG. 5 is a graph depicting IP-10 levels resulting from
administration of LNnT as measured in Example 39.
[0021] FIG. 6 is a graph depicting IP-10 levels resulting from
administration of LNnT as measured in Example 39.
[0022] FIG. 7 is a graph depicting IL-10 levels resulting from
administration of 3'SL and 6'SL as measured in Example 39.
[0023] FIG. 8 is a graph depicting IL-10 levels resulting from
administration of 3'SL and 6'SL as measured in Example 39.
[0024] FIG. 9 is a graph depicting IL-10 levels resulting from
administration of LNnT as measured in Example 39.
[0025] FIG. 10 is a graph depicting IL-10 levels resulting from
administration of LNnT as measured in Example 39.
[0026] FIG. 11 is a graph depicting RSV NS1 copy levels resulting
from administration of 2'FL and/or lycopene as measured in Example
40.
[0027] FIG. 12 is a graph depicting IP-10 levels resulting from
administration of 2'FL and lycopene as measured in Example 41.
[0028] FIG. 13 is a graph depicting the percent reduction in IP-10
resulting from the administration of 2'FL, 3'SL, and lycopene
individually or in combination as measured in Example 42.
DETAILED DESCRIPTION OF THE DISCLOSURE
[0029] The nutritional compositions and methods described herein
utilize HMOs alone or in combination with long chain
polyunsaturated fatty acids, and/or antioxidants, and in particular
carotenoids, and/or nucleotides for controlling and reducing a
number of diseases and conditions related to inflammation. The
nutritional compositions described herein include synthetic infant
formulas that include HMOs or a combination of HMOs. These and
other features of the nutritional compositions and methods, as well
as some of the many optional variations and additions, are
described in detail hereafter.
[0030] The terms "retort packaging" and "retort sterilizing" are
used interchangeably herein, and unless otherwise specified, refer
to the common practice of filling a container, most typically a
metal can or other similar package, with a nutritional liquid and
then subjecting the liquid-filled package to the necessary heat
sterilization step, to form a sterilized, retort packaged,
nutritional liquid product.
[0031] The term "aseptic packaging" as used herein, unless
otherwise specified, refers to the manufacture of a packaged
product without reliance upon the above-described retort packaging
step, wherein the nutritional liquid and package are sterilized
separately prior to filling, and then are combined under sterilized
or aseptic processing conditions to form a sterilized, aseptically
packaged, nutritional liquid product.
[0032] The terms "fat" and "oil" as used herein, unless otherwise
specified, are used interchangeably to refer to lipid materials
derived or processed from plants or animals. These terms also
include synthetic lipid materials so long as such synthetic
materials are suitable for oral administration to humans.
[0033] The term "human milk oligosaccharide" or "HMO", as used
herein, unless otherwise specified, refers generally to a number of
complex carbohydrates found in human breast milk that can be in
acidic or neutral form, and to precursors thereof. Exemplary
non-limiting human milk oligosaccharides include 3'-sialyllactose,
6'-sialyllactose, 3'-fucosyllactose, 2'-fucosyllactose, and
lacto-N-neotetraose. Exemplary human milk oligosaccharide
precursors includes sialic acid and/or fucose.
[0034] The term "shelf stable" as used herein, unless otherwise
specified, refers to a nutritional product that remains
commercially stable after being packaged and then stored at
18-24.degree. C. for at least 3 months, including from about 6
months to about 24 months, and also including from about 12 months
to about 18 months.
[0035] The terms "nutritional formulation" or "nutritional
composition" as used herein, are used interchangeably and, unless
otherwise specified, refer to synthetic formulas including
nutritional liquids, nutritional powders, nutritional solids,
nutritional semi-solids, nutritional semi-liquids, nutritional
supplements, and any other nutritional food product as known in the
art. The nutritional powders may be reconstituted to form a
nutritional liquid, all of which comprise one or more of fat,
protein and carbohydrate and are suitable for oral consumption by a
human. The terms "nutritional formulation" or "nutritional
composition" do not include human breast milk.
[0036] The term "nutritional liquid" as used herein, unless
otherwise specified, refers to nutritional products in
ready-to-drink liquid form, concentrated form, and nutritional
liquids made by reconstituting the nutritional powders described
herein prior to use.
[0037] The term "nutritional powder" as used herein, unless
otherwise specified, refers to nutritional products in flowable or
scoopable form that can be reconstituted with water or another
aqueous liquid prior to consumption and includes both spraydried
and drymixed/dryblended powders.
[0038] The term "nutritional semi-solid," as used herein, unless
otherwise specified, refers to nutritional products that are
intermediate in properties, such as rigidity, between solids and
liquids. Some semi-solids examples include puddings, gelatins, and
doughs.
[0039] The term "nutritional semi-liquid," as used herein, unless
otherwise specified, refers to nutritional products that are
intermediate in properties, such as flow properties, between
liquids and solids. Some semi-liquids examples include thick shakes
and liquid gels.
[0040] The term "infant" as used herein, unless otherwise
specified, refers to a person 12 months or younger. The term
"preterm infant" as used herein, refers to a person born prior to
36 weeks of gestation.
[0041] The term "toddler" as used herein, unless otherwise
specified, refers to a person greater than one year of age up to
three years of age.
[0042] The term "child" as used herein, unless otherwise specified,
refers to a person greater than three years of age up to twelve
years of age.
[0043] The term "newborn" as used herein, unless otherwise
specified, refers to a person from birth up to four weeks of
age.
[0044] The terms "infant formula" or "synthetic infant formula" as
used herein, unless otherwise specified, are used interchangeably
and refer to liquid, semi-liquid, solid and semi-solid human milk
replacements or substitutes that are suitable for consumption by an
infant. The synthetic formulas include components that are of
semi-purified or purified origin. As used herein, unless otherwise
specified, the terms "semi-purified" or "purified" refer to a
material that has been prepared by purification of a natural
material or by synthesis. The terms "infant formula" or "synthetic
infant formula" do not include human breast milk.
[0045] The term "synthetic pediatric formula" as used herein,
unless otherwise specified, refers to liquid, semi-liquid, solid,
and semi-solid human milk replacements or substitutes that are
suitable for consumption by an infant or toddler up to the age of
36 months (3 years). The synthetic formulas include components that
are of semi-purified or purified origin. As used herein, unless
otherwise specified, the terms "semi-purified" or "purified" refer
to a material that has been prepared by purification of a natural
material or by synthesis. The term "synthetic pediatric nutritional
formula" does not include human breast milk.
[0046] The term "synthetic child formula" as used herein, unless
otherwise specified, refers to liquid, semi-liquid, solid, and
semi-solid human milk replacements or substitutes that are suitable
for consumption by a child up to the age of 12 years. The synthetic
formulas include components that are of semi-purified or purified
origin. As used herein, unless otherwise specified, the terms
"semi-purified" or "purified" refer to a material that has been
prepared by purification of a natural material or by synthesis. The
term "synthetic child nutritional formula" does not include human
breast milk.
[0047] The term "preterm infant formula" as used herein, unless
otherwise specified, refers to liquid and solid nutritional
products suitable for consumption by a preterm infant.
[0048] The term "human milk fortifier" as used herein, unless
otherwise specified, refers to liquid and solid nutritional
products suitable for mixing with breast milk or preterm infant
formula or infant formula for consumption by a preterm or term
infant.
[0049] The terms "absence of a virus" or "absent a virus" as used
herein with respect to inducing production of monocyte-derived
cytokines, unless otherwise specified, refer to an individual
(e.g., an infant) without the virus or having the virus in an
amount less than the amount required to illicit an immune response;
that is, an amount that is less than required for the body's
natural immune response to increase the production of cytokines and
other immune factors.
[0050] The terms "inflammatory disease" or "inflammatory condition"
as used herein, unless otherwise specified, refer to any disease,
disorder, or condition characterized by inflammation. The term
"infection-mediated inflammatory disease" as used herein, unless
otherwise specified, refers to an inflammatory disease associated
or induced by microbial infection, including viral and bacterial
infection.
[0051] The terms "susceptible" and "at risk" as used herein, unless
otherwise specified, mean having little resistance to a certain
condition or disease, including being genetically predisposed,
having a family history of, and/or having symptoms of the condition
or disease.
[0052] The terms "modulating" or "modulation" or "modulate" as used
herein, unless otherwise specified, refer to the targeted movement
of a selected characteristic.
[0053] The terms "growth of a virus" or "growth of bacteria" as
used herein, unless otherwise specified, refer to the production,
proliferation, or replication of a virus or bacteria.
[0054] All percentages, parts and ratios as used herein, are by
weight of the total composition, unless otherwise specified. All
such weights, as they pertain to listed ingredients, are based on
the active level and, therefore, do not include solvents or
by-products that may be included in commercially available
materials, unless otherwise specified.
[0055] Numerical ranges as used herein are intended to include
every number and subset of numbers within that range, whether
specifically disclosed or not. Further, these numerical ranges
should be construed as providing support for a claim directed to
any number or subset of numbers in that range. For example, a
disclosure of from 1 to 10 should be construed as supporting a
range of from 2 to 8, from 3 to 7, from 5 to 6, from 1 to 9, from
3.6 to 4.6, from 3.5 to 9.9, and so forth.
[0056] All references to singular characteristics or limitations of
the present disclosure shall include the corresponding plural
characteristic or limitation, and vice versa, unless otherwise
specified or clearly implied to the contrary by the context in
which the reference is made.
[0057] All combinations of method or process steps as used herein
can be performed in any order, unless otherwise specified or
clearly implied to the contrary by the context in which the
referenced combination is made.
[0058] The nutritional compositions and methods may comprise,
consist of, or consist essentially of the essential elements of the
compositions and methods as described herein, as well as any
additional or optional element described herein or otherwise useful
in nutritional product applications.
Product Form
[0059] The nutritional compositions of the present disclosure may
be formulated and administered in any known or otherwise suitable
oral product form. Any solid, liquid, semi-solid, and semi-liquid,
or powder product form, including combinations or variations
thereof, are suitable for use herein, provided that such forms
allow for safe and effective oral delivery to the individual of the
essential ingredients as also defined herein.
[0060] The nutritional compositions of the present disclosure
include one or more HMOs as described herein. The compositions may
include one or more HMOs alone or in combination with other immune
enhancing factors including, but not limited, to long chain
polyunsaturated acids (LCPUFAs), nucleotides, and antioxidants,
such as carotenoids and vitamins, as discussed below.
[0061] The nutritional compositions may be in any product form
comprising the ingredients described herein, and which is safe and
effective for oral administration. The nutritional compositions may
be formulated to include only the ingredients described herein, or
may be modified with optional ingredients to form a number of
different product forms.
[0062] The nutritional compositions of the present disclosure are
desirably formulated as dietary product forms, which are defined
herein as those embodiments comprising the ingredients of the
present disclosure in a product form that then contains at least
one of fat, protein, and carbohydrate, and preferably also contains
vitamins, minerals, or combinations thereof. The nutritional
compositions will comprise at least HMOs, desirably in combination
with at least one of protein, fat, vitamins, and minerals, to
produce a nutritional composition.
[0063] The nutritional compositions may be formulated with
sufficient kinds and amounts of nutrients to provide a sole,
primary, or supplemental source of nutrition, or to provide a
specialized nutritional product for use in individuals afflicted
with specific diseases or conditions or with a targeted nutritional
benefit as described below.
[0064] Specific non-limiting examples of product forms suitable for
use with the HMO-containing compositions as disclosed herein
include, for example, liquid and powdered dietary supplements,
liquid and powdered human milk fortifiers, liquid and powdered
preterm infant formulas, liquid and powdered infant formulas,
liquid and powdered elemental and semi-elemental formulas, liquid
and powdered pediatric formulas, liquid and powdered toddler
formulas, and liquid and powdered follow-on formulas suitable for
use with infants and children.
Nutritional Liquids
[0065] Nutritional liquids include both concentrated and
ready-to-feed nutritional liquids. These nutritional liquids are
most typically formulated as suspensions or emulsions, although
other liquid forms are within the scope of the present
disclosure.
[0066] Nutritional emulsions suitable for use may be aqueous
emulsions comprising proteins, fats, and carbohydrates. These
emulsions are generally flowable or drinkable liquids at from about
1.degree. C. to about 25.degree. C. and are typically in the form
of oil-in-water, water-in-oil, or complex aqueous emulsions,
although such emulsions are most typically in the form of
oil-in-water emulsions having a continuous aqueous phase and a
discontinuous oil phase.
[0067] The nutritional emulsions may be and typically are shelf
stable. The nutritional emulsions typically contain up to about 95%
by weight of water, including from about 50% to about 95%, also
including from about 60% to about 90%, and also including from
about 70% to about 85%, of water by weight of the nutritional
emulsions. The nutritional emulsions may have a variety of product
densities, but most typically have a density greater than about
1.03 g/mL, including greater than about 1.04 g/mL, including
greater than about 1.055 g/mL, including from about 1.06 g/mL to
about 1.12 g/mL, and also including from about 1.085 g/mL to about
1.10 g/mL.
[0068] The nutritional emulsions may have a caloric density
tailored to the nutritional needs of the ultimate user, although in
most instances the emulsions comprise generally at least 19 kcal/fl
oz (660 kcal/liter), more typically from about 20 kcal/fl oz
(675-680 kcal/liter) to about 25 kcal/fl oz (820 kcal/liter), even
more typically from about 20 kcal/fl oz (675-680 kcal/liter) to
about 24 kcal/fl oz (800-810 kcal/liter). Generally, the 22-24
kcal/fl oz formulas are more commonly used in preterm or low birth
weight infants, and the 20-21 kcal/fl oz (675-680 to 700
kcal/liter) formulas are more often used in term infants. In some
embodiments, the emulsion may have a caloric density of from about
50-100 kcal/liter to about 660 kcal/liter, including from about 150
kcal/liter to about 500 kcal/liter. In some specific embodiments,
the emulsion may have a caloric density of 25, or 50, or 75, or 100
kcal/liter.
[0069] The nutritional emulsion may have a pH ranging from about
3.5 to about 8, but are most advantageously in a range of from
about 4.5 to about 7.5, including from about 5.5 to about 7.3,
including from about 6.2 to about 7.2.
[0070] Although the serving size for the nutritional emulsion can
vary depending upon a number of variables, a typical serving size
is generally at least about 1 mL, or even at least about 2 mL, or
even at least about 5 mL, or even at least about 10 mL, or even at
least about 25 mL, including ranges from about 1 mL to about 300
mL, including from about 4 mL to about 250 mL, and including from
about 10 mL to about 240 mL.
Nutritional Solids
[0071] The nutritional solids may be in any solid form but are
typically in the form of flowable or substantially flowable
particulate compositions, or at least particulate compositions.
Particularly suitable nutritional solid product forms include spray
dried, agglomerated and/or dryblended powder compositions. The
compositions can easily be scooped and measured with a spoon or
similar other device, and can easily be reconstituted by the
intended user with a suitable aqueous liquid, typically water, to
form a nutritional composition for immediate oral or enteral use.
In this context, "immediate" use generally means within about 48
hours, most typically within about 24 hours, preferably right after
reconstitution.
[0072] The nutritional powders may be reconstituted with water
prior to use to a caloric density tailored to the nutritional needs
of the ultimate user, although in most instances the powders are
reconstituted with water to form compositions comprising at least
19 kcal/fl oz (660 kcal/liter), more typically from about 20
kcal/fl oz (675-680 kcal/liter) to about 25 kcal/fl oz (820
kcal/liter), even more typically from about 20 kcal/fl oz (675-680
kcal/liter) to about 24 kcal/fl oz (800-810 kcal/liter). Generally,
the 22-24 kcal/fl oz formulas are more commonly used in preterm or
low birth weight infants, and the 20-21 kcal/fl oz (675-680 to 700
kcal/liter) formulas are more often used in term infants. In some
embodiments, the reconstituted powder may have a caloric density of
from about 50-100 kcal/liter to about 660 kcal/liter, including
from about 150 kcal/liter to about 500 kcal/liter. In some specific
embodiments, the emulsion may have a caloric density of 25, or 50,
or 75, or 100 kcal/liter.
Human Milk Oligosaccharides (HMOs)
[0073] The nutritional compositions of the present disclosure
include at least one HMO, and in many embodiments, a combination of
two or more HMOs. Oligosaccharides are one of the main components
of human breast milk, which contains, on average, 10 grams per
liter of neutral oligosaccharides and 1 gram per liter of acidic
oligosaccharides. The composition of human milk oligosaccharides is
very complex and more than 200 different oligosaccharide-like
structures are known.
[0074] The HMOs may be included in the nutritional compositions
alone, or in some embodiments, in combination with other immune
enhancing factors (e.g., LCPUFAs, antioxidants, nucleotides, etc.)
as described herein. The HMO or HMOs may be isolated or enriched
from milk(s) secreted by mammals including, but not limited to:
human, bovine, ovine, porcine, or caprine species. The HMOs may
also be produced via microbial fermentation, enzymatic processes,
chemical synthesis, or combinations thereof.
[0075] Suitable HMOs for use in the nutritional compositions may
include acidic oligosaccharides, neutral oligosaccharides,
n-acetylglucosylated oligosaccharides, and HMO precursors. Specific
non-limiting examples of HMOs that may be included individually or
in combination in the compositions of the present disclosure
include: sialic acid (i.e., free sialic acid, lipid-bound sialic
acid, protein-bound sialic acid); D-glucose (Glc); D-galactose
(Gal); N-acetylglucosamine (GlcNAc); L-fucose (L-Fuc); D-fucose
(D-Fuc); fucosyl oligosaccharides (i.e., Lacto-N-fucopentaose I;
Lacto-N-fucopentaose II; 2'-Fucosyllactose; 3'-Fucosyllactose;
Lacto-N-fucopentaose III; Lacto-N-difucohexaose I; and
Lactodifucotetraose); non-fucosylated, non-sialylated
oligosaccharides (i.e., Lacto-N-tetraose and Lacto-N-neotetraose);
sialyl oligosaccharides (i.e., 3'-Sialyl-3-fucosyllactose;
Disialomonofucosyllacto-N-neohexaose;
Monofucosylmonosialyllacto-N-octaose (sialyl Lea);
Sialyllacto-N-fucohexaose II; Disialyllacto-N-fucopentaose II;
Monofucosyldisialyllacto-N-tetraose); and sialyl fucosyl
oligosaccharides (i.e., 2'-Sialyllactose; 2-Sialyllactosamine;
3'-Sialyllactose; 3'-Sialyllactosamine; 6'-Sialyllactose;
6'-Sialyllactosamine; Sialyllacto-N-neotetraose c;
Monosialyllacto-N-hexaose; Disialyllacto-N-hexaose I;
Monosialyllacto-N-neohexaose I; Monosialyllacto-N-neohexaose II;
Disialyllacto-N-neohexaose; Disialyllacto-N-tetraose;
Disialyllacto-N-hexaose II; Sialyllacto-N-tetraose a;
Disialyllacto-N-hexaose I; and Sialyllacto-N-tetraose b). Also
useful are variants in which the glucose (Glc at the reducing end
is replaced by N-acetylglucosamine (e.g.,
2'-fucosyl-N-acetylglucosamine (2'-FLNac) is such a variant to
2'-fucosyllactose). These HMOs are described more fully in U.S.
Patent Application No. 2009/0098240, which is herein incorporated
by reference in its entirety. Other suitable examples of HMOs that
may be included in the compositions of the present disclosure
include lacto-N-fucopentaose V, lacto-N-hexaose,
para-lacto-N-hexaose, lacto-N-neohexaose, para-lacto-N-neohexaose,
monofucosyllacto-N-hexaose II, isomeric fucosylated lacto-N-hexaose
(1), isomeric fucosylated lacto-N-hexaose (3), isomeric fucosylated
lacto-N-hexaose (2), difucosyl-para-lacto-N-neohexaose,
difucosyl-para-lacto-N-hexaose, difucosyllacto-N-hexaose,
lacto-N-neoocataose, para-lacto-N-octanose, iso-lacto-N-octaose,
lacto-N-octaose, monofucosyllacto-neoocataose,
monofucosyllacto-N-ocataose, difucosyllacto-N-octaose I,
difucosyllacto-N-octaose II, difucosyllacto-N-neoocataose II,
difucosyllacto-N-neoocataose I, lacto-N-decaose,
trifucosyllacto-N-neooctaose, trifucosyllacto-N-octaose,
trifucosyl-iso-lacto-N-octaose, lacto-N-difucohexaose II,
sialyl-lacto-N-tetraose a, sialyl-lacto-N-tetraose b,
sialyl-lacto-N-tetraose c, sialyl-fucosyl-lacto-N-tetraose I,
sialyl-fucosyl-lacto-N-tetraose II, and disialyl-lacto-N-tetraose,
and combinations thereof. Particularly suitable nutritional
compositions include at least one of the following HMOs or HMO
precursors: sialic acid (SA); 3'-Sialyllactose (3'SL);
6'-Sialyllactose (6'SL); 2'-Fucosyllactose (2'FL);
3'-Fucosyllactose (3'FL); Lacto-N-tetraose and Lacto-N-neotetraose
(LNnT), and in particular, combinations of 6'SL and 3'SL;
combinations of 3'FL and SA; combinations of 2'FL and 3'FL;
combinations of 2'FL, 3'SL, and 6'SL; combinations of 3'SL, 3'FL,
and LNnT; and combinations of 6'SL, 2'FL, and LNnT.
[0076] Other exemplary combinations include: SA, 3'SL, 6'SL, 3'FL,
2'FL, and LNnT; 3'SL, 6'SL, 3'FL, 2'FL, and LNnT; SA, 6'SL, 3'FL,
2'FL, and LNnT; SA, 3'SL, 3'FL, 2'FL, and LNnT; SA, 3'SL, 6'SL,
2'FL, and LNnT; SA, 3'SL, 6'SL, 3'FL, and LNnT; SA, 3'SL, 6'SL,
3'FL, and 2'FL; SA and 3'SL; SA and 6'SL; SA and 2'FL; SA and LNnT;
SA, 3'SL, and 6'SL; SA, 3'SL and 3'FL; SA, 3'SL and 2'FL; SA, 3'SL
and LNnT; SA, 6'SL and 3'FL; SA, 6'SL, and 2'FL; SA, 6'SL, and
LNnT; SA, 3'FL, and 2'FL; SA, 3'FL, and LNnT; SA, 2'FL, and LNnT;
SA, 3'SL, 6'SL, and 3'FL; SA, 3'SL, 6'SL and 2'FL; SA, 3'SL, 6'SL,
and LNnT; SA, 3'SL, 3'FL, and 2'FL; SA, 3'SL, 3'FL, and LNnT; SA,
3'SL, 2'FL, and LNnT; SA, 6'SL, 3'FL, and 2'FL; SA, 6'SL, 2'FL, and
LNnT; SA, 6'SL, 3'FL, and LNnT; SA, 3'FL, 2'FL, and LNnT; SA, 6'SL,
2'FL, and LNnT; SA, 3'SL, 3'FL, 2'FL, and LNnT; SA, 6'SL, 3'FL,
2'FL, and LNnT; SA, 3'SL, 6'SL, 3'FL, and LNnT; SA, 3'SL, 3'FL,
2'FL, and LNnT; SA, 3'SL, 6'SL, 2'FL, and LNnT; 3'SL, 6'SL, 3'FL,
and 2'FL; 3'SL, 6'SL, 2'FL, and LNnT; 3'SL, 3'FL, 2'FL, and LNnT;
3'SL, 6'SL, 3'FL, and LNnT; 3'SL, 6'SL, and 3'FL; 3'SL, 3'FL, and
2'FL; 3'SL, 2'FL, and LNnT; 3'SL, 6'SL, and 2'FL; 3'SL, 6'SL, and
LNnT; 3'SL and 3'FL; 3'SL and 2'FL; 3'SL and LNnT; 6'SL and 3'FL;
6'SL and 2'FL; 6'SL and LNnT; 6'SL, 3'FL, and LNnT; 6'SL, 3'FL,
2'FL, and LNnT; 3'FL, 2'FL, and LNnT; 3'FL and LNnT; and 2'FL and
LNnT.
[0077] The HMOs are present in the nutritional compositions in
total amounts of HMO in the composition (mg of HMO per mL of
composition) of at least about 0.001 mg/mL, including at least
about 0.01 mg/mL, including from about 0.001 mg/mL to about 20
mg/mL, including from about 0.01 mg/mL to about 20 mg/mL, including
from about 0.01 mg/mL to about 10 mg/mL, including from about 0.01
mg/mL to about 5 mg/mL, including from about 0.001 mg/mL to about 1
mg/mL, including from about 0.01 mg/mL to about 1 mg/mL, including
from about 0.001 mg/mL to about 0.23 mg/mL, including from about
0.01 mg/mL to about 0.23 mg/mL of total HMO in the nutritional
composition. Typically, the amount of HMO in the nutritional
composition will depend on the specific HMO or HMOs present and the
amounts of other components in the nutritional compositions.
[0078] In one specific embodiment when the nutritional product is a
nutritional powder, the total concentration of HMOs in the
nutritional powder is from about 0.0005% to about 5%, including
from about 0.01% to about 1% (by weight of the nutritional
powder).
[0079] In another specific embodiment, when the nutritional product
is a ready-to-feed nutritional liquid, the total concentration of
HMOs in the ready-to-feed nutritional liquid is from about 0.0001%
to about 0.50%, including from about 0.001% to about 0.15%,
including from about 0.01% to about 0.10%, and further including
from about 0.01% to about 0.03% (by weight of the ready-to-feed
nutritional liquid).
[0080] In another specific embodiment when the nutritional product
is a concentrated nutritional liquid, the total concentration of
HMOs in the concentrated nutritional liquid is from about 0.0002%
to about 0.60%, including from about 0.002% to about 0.30%,
including from about 0.02% to about 0.20%, and further including
from about 0.02% to about 0.06% (by weight of the concentrated
nutritional liquid).
[0081] In one specific embodiment, the nutritional composition
includes a neutral human milk oligosaccharide in an amount of from
about 0.001 mg/mL to about 20 mg/mL, including from 0.01 mg/mL to
about 20 mg/mL, including from about 0.001 mg/mL to less than 2
mg/mL, and including from about 0.01 mg/mL to less than 2
mg/mL.
[0082] In some embodiments, the HMOs are used in combination to
provide the desired immune enhancing effect. For example, in one
embodiment, the nutritional composition includes 6'SL in
combination with 3'SL in a total amount of HMO of from about 0.001
mg/mL to about 20 mg/mL, including from about 0.01 mg/mL to about
20 mg/mL, including from about 0.001 mg/mL to about 0.23 mg/mL,
including from about 0.01 mg/mL to about 0.23 mg/mL, including from
about 0.001 mg/mL to less than 0.15 mg/mL, and including from about
0.01 mg/mL to less than 0.15 mg/mL of the nutritional composition.
In another embodiment, the nutritional composition includes 6'SL in
combination with 3'SL in a total amount of HMO of from about 0.001
mg/mL to about 20 mg/mL, including from about 0.01 mg/mL to about
20 mg/mL and including greater than 0.65 mg/mL to about 20 mg/mL.
In another embodiment, the nutritional composition includes 3'SL
and 6'SL in a weight ratio of from about 1:20 to about 20:1,
including from about 1:10 to about 10:1, and including from about
1:2 to about 2:1.
[0083] In one specific embodiment, the nutritional composition
includes 6'SL, alone or in combination with other HMOs, in an
amount of from about 0.001 mg/mL to about 20 mg/mL, including from
about 0.01 mg/mL to about 20 mg/mL, including from about 0.001
mg/mL to less than 0.25 mg/mL, including from about 0.01 mg/mL to
less than 0.25 mg/mL, including from greater than 0.4 mg/mL to
about 20 mg/mL, and including from about 0.1 mg/mL to about 0.5
mg/mL.
[0084] In one embodiment, when the nutritional composition includes
6'SL, the total amount of HMOs in the nutritional composition
includes at least about 88% (by total weight HMOs) 6'SL, including
from about 88% (by total weight HMOs) to about 96% (by total weight
HMOs), including from about 88% (by total weight HMOs) to about
100% (by total weight HMOs), and including about 100% (by total
weight HMOs) 6'SL.
[0085] In another embodiment, the nutritional composition includes
3'SL, alone or in combination with other HMOs, in an amount of from
about 0.001 mg/mL to about 20 mg/mL, including from about 0.01
mg/mL to about 20 mg/mL, including from about 0.001 mg/mL to less
than 0.15 mg/mL, including from about 0.01 mg/mL to less than 0.15
mg/mL, and including from greater than 0.25 mg/mL to about 20
mg/mL.
[0086] In one embodiment, when the nutritional composition includes
3'SL, the total amount of HMOs in the nutritional composition
includes at least about 85% (by total weight HMOs) 3'SL, including
from about 85% (by total weight HMOs) to about 88% (by total weight
HMOs), including from about 85% (by total weight HMOs) to about
100% (by total weight HMOs), and including about 100% (by total
weight HMOs) 3'SL.
[0087] In one specific embodiment, the nutritional composition
includes LNnT, alone or in combination with other HMOs, in an
amount of from about 0.001 mg/mL to about 20 mg/mL, including from
about 0.01 mg/mL to about 20 mg/mL, including from about 0.001
mg/mL to less than 0.2 mg/mL, including from about 0.01 mg/mL to
less than 0.2 mg/mL, and including from greater than 0.32 mg/mL to
about 20 mg/mL.
[0088] In another specific embodiment, the nutritional composition
includes 3'FL, alone or in combination with other HMOs, in an
amount of from about 0.001 mg/mL to about 20 mg/mL, including from
about 0.01 mg/mL to about 20 mg/mL, including from about 0.001
mg/mL to less than 1 mg/mL, including from about 0.01 mg/mL to less
than 1 mg/mL, and including from greater than 1.7 mg/mL to about 20
mg/mL.
[0089] In one specific embodiment, the nutritional composition
includes 3'FL in combination with SA in a total amount of HMO of
from about 0.001 mg/mL to about 20 mg/mL, including from about 0.01
mg/mL to about 20 mg/mL. In one embodiment, the nutritional
composition includes 3'FL in an amount of from about 0.001 mg/mL to
less than 1 mg/mL, including from 0.01 mg/mL to less than 1 mg/mL
and SA in an amount of about 1 mg/mL.
[0090] In another embodiment, the nutritional composition includes
2'FL, alone or in combination with other HMOs, in an amount of from
about 0.001 mg/mL to about 20 mg/mL, including from about 0.01
mg/mL to about 20 mg/mL, including from about 0.001 mg/mL to less
than 2 mg/mL, including from about 0.01 mg/mL to less than 2 mg/mL,
including from about 0.001 mg/mL to about 1 mg/mL, and including
from about 0.01 mg/mL to about 1 mg/mL. In another embodiment, the
nutritional composition includes 2'FL, alone or in combination with
other HMOs, in an amount of from about 0.001 mg/mL to about 20
mg/mL, including from about 0.01 mg/mL to about 20 mg/mL and
including greater than 2.5 mg/mL to about 20 mg/mL.
[0091] In one specific embodiment, the nutritional composition
includes 2'FL in combination with 3'FL in a total amount of HMO of
from 0.001 mg/mL to about 20 mg/mL, including from about 0.01 mg/mL
to about 20 mg/mL.
[0092] In yet another embodiment, the nutritional composition
includes a combination of 6' SL, 2'FL, and LNnT in a total amount
of HMO of from about 0.001 mg/mL to about 20 mg/mL, including from
about 0.01 mg/mL to about 20 mg/mL.
Long Chain Polyunsaturated Fatty Acids (LCPUFAs)
[0093] In addition to the HMOs described above, the nutritional
products of the present disclosure may include LCPUFAs. LCPUFAs are
included in the nutritional compositions to provide nutritional
support, as well as to reduce oxidative stress and enhance growth
and functional development of the intestinal epithelium and
associated immune cell populations. In some embodiments, the
nutritional composition includes a combination of one or more HMOs
and one or more LCPUFAs such that the composition provides a
synergistic benefit to the end user, such as a synergistic benefit
in modulating anti-viral immune responses and dampening
inflammation. In some embodiments, the HMO or HMOs used in
combination with the LCPUFAs to provide the synergistic effect are
acidic HMOs.
[0094] Exemplary LCPUFAs for use in the nutritional compositions
include, for example, co-3 LCPUFAs and co-6 LCPUFAs. Specific
LCPUFAs include docosahexaenoic acid (DHA), eicosapentaenoic acid
(EPA), docosapentaenoic acid (DPA), arachidonic acid (ARA),
linoleic acid, linolenic acid (alpha linolenic acid) and
gamma-linolenic acid derived from oil sources such as plant oils,
marine plankton, fungal oils, and fish oils. In one particular
embodiment, the LCPUFAs are derived from fish oils such as
menhaden, salmon, anchovy, cod, halibut, tuna, or herring oil.
Particularly preferred LCPUFAs for use in the nutritional
compositions with the HMOs include DHA, ARA, EPA, DPA, and
combinations thereof.
[0095] In order to reduce potential side effects of high dosages of
LCPUFAs in the nutritional compositions, the content of LCPUFAs
preferably does not exceed 3% by weight of the total fat content,
including below 2% by weight of the total fat content, and
including below 1% by weight of the total fat content in the
nutritional composition.
[0096] The LCPUFA may be provided as free fatty acids, in
triglyceride form, in diglyceride form, in monoglyceride form, in
phospholipid form, in esterfied form or as a mixture of one or more
of the above, preferably in triglyceride form.
[0097] The nutritional compositions as described herein will
typically comprise total concentrations of LCPUFA of from about
0.01 mM to about 10 mM and including from about 0.01 mM to about 1
mM. Alternatively, the nutritional compositions comprise total
concentrations of LCPUFA of from about 0.001 g/L to about 1
g/L.
[0098] In one embodiment, the nutritional compositions include
total long chain .omega.-6 fatty acids in a concentration of from
about 100 to about 425 mg/L or from about 12 to about 53 mg per 100
kcals and/or further include total long chain .omega.-3 fatty acids
in a concentration of from about 40 to about 185 mg/L or from about
5 to about 23 mg per 100 kcals. In one specific embodiment, the
ratio of long chain .omega.-6 fatty acids to long chain .omega.-3
fatty acids in the nutritional compositions ranges from about 2:1
to about 3:1, preferably about 2.5:1.
[0099] In one specific embodiment, the nutritional compositions
include DHA in a concentration of from about 0.025 mg/mL to about
0.130 mg/mL or from about 3 to about 16 mg per 100 kcals. In
another embodiment, the nutritional compositions include ARA in a
concentration of from about 0.080 mg/mL to about 0.250 mg/mL or
from about 10 to about 31 mg per 100 kcals. In yet another
embodiment, the nutritional compositions include combinations of
DHA and ARA such that the ratio of DHA to ARA ranges from about 1:4
to about 1:2.
Antioxidants
[0100] Additionally, the nutritional compositions may comprise one
or more antioxidants in combination with the HMOs (and optionally
LCPUFAs and/or nucleotides also) to provide nutritional support, as
well as to reduce oxidative stress. In some embodiments, the
nutritional composition includes a combination of HMOs and
antioxidants such that the composition provides a synergistic
benefit to the end user, such as a synergistic benefit in
modulating anti-viral immune responses and dampening inflammation.
In some embodiments, the HMO or HMOs is used in combination with
carotenoids (and specifically lutein, beta-carotene, zeaxanthin
and/or lycopene) to provide the synergistic effect.
[0101] Any antioxidants suitable for oral administration may be
included for use in the nutritional compositions of the present
disclosure, including, for example, vitamin A, vitamin E, vitamin
C, retinol, tocopherol, and carotenoids, including lutein,
beta-carotene, zeaxanthin, and lycopene, and combinations thereof,
for example.
[0102] As noted, the antioxidants for use in the nutritional
compositions may be used with the HMOs alone or in combination with
HMOs and LCPUFAs and/or nucleotides. In one specific embodiment,
the antioxidants for use in the nutritional compositions include
carotenoids, and particularly, combinations of the carotenoids
lutein, lycopene, zeaxanthin and/or beta-carotene. Nutritional
compositions containing these combinations, as selected and defined
herein, can be used to modulate inflammation and/or levels of
C-reactive protein in preterm and term infants.
[0103] It is generally preferable that the nutritional compositions
comprise at least one of lutein, lycopene, zeaxanthin, and
beta-carotene to provide a total amount of carotenoid of from about
0.001 .mu.g/mL to about 10 .mu.g/mL. More particularly, the
nutritional compositions comprise lutein in an amount of from about
0.001 .mu.g/mL to about 10 .mu.g/mL, including from about 0.001
.mu.g/mL to about 5 .mu.g/mL, including from about 0.001 .mu.g/mL
to about 0.0190 .mu.g/mL, including from about 0.001 .mu.g/mL to
about 0.0140 .mu.g/mL, and also including from about 0.044 .mu.g/mL
to about 5 .mu.g/mL of lutein. It is also generally preferable that
the nutritional compositions comprise from about 0.001 .mu.g/mL to
about 10 .mu.g/mL, including from about 0.001 .mu.g/mL to about 5
.mu.g/mL, from about 0.001 .mu.g/mL to about 0.0130 .mu.g/mL,
including from about 0.001 .mu.g/mL to about 0.0075 .mu.g/mL, and
also including from about 0.0185 .mu.g/mL to about 5 .mu.g/mL of
lycopene. It is also generally preferable that the nutritional
compositions comprise from about 1 .mu.g/mL to about 10 .mu.g/mL,
including from about 1 .mu.g/mL to about 5 .mu.g/mL, including from
about 0.001 .mu.g/mL to about 0.025 .mu.g/mL, including from about
0.001 .mu.g/mL to about 0.011 .mu.g/mL, and also including from
about 0.034 .mu.g/mL to about 5 .mu.g/mL of beta-carotene. It
should be understood that any combination of these amounts of
beta-carotene, lutein, zeaxanthin, and lycopene can be included in
the nutritional compositions of the present disclosure. Other
carotenoids may optionally be included in the nutritional
compositions as described herein. Any one or all of the carotenoids
included in the nutritional compositions described herein may be
from a natural source, or artificially synthesized. In one
particular embodiment, the nutritional composition comprises a
combination of 2'FL and lycopene.
[0104] Each of the carotenoids in the selected combinations can be
obtained from any known or otherwise suitable material source for
use in nutritional compositions, and each can be provided
individually, or all together, or in any combination and from any
number of sources, including sources such as multivitamin premixes
containing other vitamins or minerals in combination with one or
more of the carotenoids as described herein. Non-limiting examples
of some suitable sources of lutein, lycopene, beta-carotene, or
combinations thereof include LycoVit.RTM. lycopene (available from
BASF, Mount Olive, N.J.), Lyc-O-Mato.RTM. tomato extract in oil,
powder, or bead form (available from LycoRed Corp., Orange, N.J.),
beta-carotene, lutein, or lycopene (available from DSM Nutritional
Products, Parsippany, N.J.), FloraGLO.RTM. lutein (available from
Kemin Health, Des Moines, Iowa), Xangold.RTM. Natural Lutein Esters
(available from Cognis, Cincinnati, Ohio), and Lucarotin.RTM.
beta-carotene (available from BASF, Mount Olive, N.J.).
Nucleotides
[0105] In addition to the HMOs, the nutritional compositions of the
present disclosure may additionally comprise nucleotides and/or
nucleotide precursors selected from the group consisting of
nucleosides, purine bases, pyrimidine bases, ribose and
deoxyribose. The nucleotide may be in monophosphate, diphosphate,
or triphosphate form. The nucleotide may be a ribonucleotide or a
deoxyribonucleotide. The nucleotides may be monomeric, dimeric, or
polymeric (including RNA and DNA). The nucleotide may be present in
the nutritional composition as a free acid or in the form of a
salt, preferably a monosodium salt. In some embodiments, the
nutritional composition includes a combination of HMOs and
nucleotides such that the composition provides a synergistic
benefit to the end user, such as a synergistic benefit in
modulating anti-viral immune responses and dampening inflammation
and/or improving intestinal barrier integrity.
[0106] Incorporation of nucleotides in the nutritional compositions
of the present disclosure improves intestinal barrier integrity
and/or maturation, which is beneficial to preterm and term infants
who have less developed intestinal flora and hence a slower
maturing intestinal barrier.
[0107] Suitable nucleotides and/or nucleosides for use in the
nutritional compositions include one or more of cytidine
5'-monophosphate, uridine 5'-monophosphate, adenosine
5'-monophosphate, guanosine 5'-1-monophosphate, and/or inosine
5'-monophosphate, more preferably cytidine 5'-monophosphate,
uridine 5'-monophosphate, adenosine 5'-monophosphate, guanosine
5'-monophosphate, and inosine 5'-monophosphate.
[0108] The nucleotides are present in the nutritional products in
total amounts of nucleotides of at least about 5 mg/L, including at
least about 10 mg/L, including from about 10 mg/L to about 200
mg/L, including from about 42 mg/L to about 102 mg/L, and including
at least about 72 mg/L of the nutritional product.
[0109] In one specific embodiment when the nutritional composition
is a nutritional powder, the nucleotide may be present at a level
of at least about 0.007%, including from about 0.0078% to about
0.1556%, and including about 0.056% (by weight of the nutritional
powder), or at least about 0.007 grams, including from about 0.0078
grams to about 0.1556 grams, and including about 0.056 grams of
nucleotide per 100 grams of nutritional powder.
[0110] In another specific embodiment, when the nutritional
composition is a ready-to-feed nutritional liquid, the nucleotide
is present at a level of at least about 0.001%, including from
about 0.001% to about 0.0197%, and including about 0.0071% (by
weight of the nutritional liquid), or at least about 0.001 grams,
including from about 0.001 grams to about 0.0197 grams, and
including about 0.0071 grams of nucleotide per 100 grams of
ready-to-feed nutritional liquid.
[0111] In another specific embodiment when the nutritional
composition is a concentrated nutritional liquid, the nucleotide is
present at a level of at least about 0.0019%, including from about
0.0019% to about 0.0382%, and including about 0.0138% (by weight of
the nutritional liquid), or at least about 0.0019 grams, including
from about 0.0019 grams to about 0.0382 grams, and including about
0.0138 grams of nucleotide per 100 grams of concentrated
nutritional liquid.
Macronutrients
[0112] The nutritional compositions including the HMO or HMOs may
be formulated to include at least one of protein, fat, and
carbohydrate. In many embodiments, the nutritional compositions
will include the HMO or HMOs with protein, carbohydrate and
fat.
[0113] Although total concentrations or amounts of the fat,
protein, and carbohydrates may vary depending upon the product type
(i.e., human milk fortifier, preterm infant formula, infant
formula, etc.), product form (i.e., nutritional solid, powder,
ready-to-feed liquid, or concentrated liquid) and targeted dietary
needs of the intended user, such concentrations or amounts most
typically fall within one of the following embodied ranges,
inclusive of any other essential fat, protein, and/or carbohydrate
ingredients as described herein.
[0114] For the liquid preterm and term infant formulas,
carbohydrate concentrations most typically range from about 5% to
about 40%, including from about 7% to about 30%, including from
about 10% to about 25%, by weight of the preterm or term infant
formula; fat concentrations most typically range from about 1% to
about 30%, including from about 2% to about 15%, and also including
from about 3% to about 10%, by weight of the preterm or term infant
formula; and protein concentrations most typically range from about
0.5% to about 30%, including from about 1% to about 15%, and also
including from about 2% to about 10%, by weight of the preterm or
term infant formula.
[0115] For the liquid human milk fortifiers, carbohydrate
concentrations most typically range from about 10% to about 75%,
including from about 10% to about 50%, including from about 20% to
about 40%, by weight of the human milk fortifier; fat
concentrations most typically range from about 10% to about 40%,
including from about 15% to about 37%, and also including from
about 18% to about 30%, by weight of the human milk fortifier; and
protein concentrations most typically range from about 5% to about
40%, including from about 10% to about 30%, and also including from
about 15% to about 25%, by weight of the human milk fortifier.
[0116] The amount of carbohydrates, fats, and/or proteins in any of
the liquid nutritional compositions described herein may also be
characterized in addition to, or in the alternative, as a
percentage of total calories in the liquid nutritional composition
as set forth in the following table. These macronutrients for
liquid nutritional compositions of the present disclosure are most
typically formulated within any of the caloric ranges (embodiments
A-F) described in the following table (each numerical value is
preceded by the term "about").
TABLE-US-00001 Nutrient % Total Cal. Embodiment A Embodiment B
Embodiment C Carbohydrate 0-98 2-96 10-75 Protein 0-98 2-96 5-70
Fat 0-98 2-96 20-85 Embodiment D Embodiment E Embodiment F
Carbohydrate 30-50 25-50 25-50 Protein 15-35 10-30 5-30 Fat 35-55
1-20 2-20
[0117] In one specific example, liquid infant formulas (both
ready-to-feed and concentrated liquids) include those embodiments
in which the protein component may comprise from about 7.5% to
about 25% of the caloric content of the formula; the carbohydrate
component may comprise from about 35% to about 50% of the total
caloric content of the infant formula; and the fat component may
comprise from about 30% to about 60% of the total caloric content
of the infant formula. These ranges are provided as examples only,
and are not intended to be limiting. Additional suitable ranges are
noted in the following table (each numerical value is preceded by
the term "about").
TABLE-US-00002 Nutrient % Total Cal. Embodiment G Embodiment H
Embodiment I Carbohydrates: 20-85 30-60 35-55 Fat: 5-70 20-60 25-50
Protein: 2-75 5-50 7-40
[0118] When the nutritional product is a powdered preterm or term
infant formula, the protein component is present in an amount of
from about 5% to about 35%, including from about 8% to about 12%,
and including from about 10% to about 12% by weight of the preterm
or term infant formula; the fat component is present in an amount
of from about 10% to about 35%, including from about 25% to about
30%, and including from about 26% to about 28% by weight of the
preterm or term infant formula; and the carbohydrate component is
present in an amount of from about 30% to about 85%, including from
about 45% to about 60%, including from about 50% to about 55% by
weight of the preterm or term infant formula.
[0119] For powdered human milk fortifiers the protein component is
present in an amount of from about 1% to about 55%, including from
about 10% to about 50%, and including from about 10% to about 30%
by weight of the human milk fortifier; the fat component is present
in an amount of from about 1% to about 30%, including from about 1%
to about 25%, and including from about 1% to about 20% by weight of
the human milk fortifier; and the carbohydrate component is present
in an amount of from about 15% to about 75%, including from about
15% to about 60%, including from about 20% to about 50% by weight
of the human milk fortifier.
[0120] The total amount or concentration of fat, carbohydrate, and
protein, in the powdered nutritional compositions of the present
disclosure can vary considerably depending upon the selected
composition and dietary or medical needs of the intended user.
Additional suitable examples of macronutrient concentrations are
set forth below. In this context, the total amount or concentration
refers to all fat, carbohydrate, and protein sources in the
powdered product. For powdered nutritional compositions, such total
amounts or concentrations are most typically and preferably
formulated within any of the embodied ranges described in the
following table (each numerical value is preceded by the term
"about").
TABLE-US-00003 Nutrient % Total Cal. Embodiment J Embodiment K
Embodiment L Carbohydrate 1-85 30-60 35-55 Fat 5-70 20-60 25-50
Protein 2-75 5-50 7-40
Fat
[0121] The nutritional compositions of the present disclosure may,
in addition to the LCPUFAs described above, comprise an additional
source or sources of fat. Suitable additional sources of fat for
use herein include any fat or fat source that is suitable for use
in an oral nutritional product and is compatible with the essential
elements and features of such products. For example, in one
specific embodiment, the additional fat is derived from short chain
fatty acids.
[0122] Additional non-limiting examples of suitable fats or sources
thereof for use in the nutritional products described herein
include coconut oil, fractionated coconut oil, soybean oil, corn
oil, olive oil, safflower oil, high oleic safflower oil, oleic
acids (EMERSOL 6313 OLEIC ACID, Cognis Oleochemicals, Malaysia),
MCT oil (medium chain triglycerides), sunflower oil, high oleic
sunflower oil, palm and palm kernel oils, palm olein, canola oil,
marine oils, fish oils, fungal oils, algae oils, cottonseed oils,
and combinations thereof.
Protein
[0123] The nutritional compositions of the present disclosure may
optionally further comprise protein. Any protein source that is
suitable for use in oral nutritional compositions and is compatible
with the essential elements and features of such products is
suitable for use in the nutritional compositions.
[0124] Non-limiting examples of suitable proteins or sources
thereof for use in the nutritional products include hydrolyzed,
partially hydrolyzed or non-hydrolyzed proteins or protein sources,
which may be derived from any known or otherwise suitable source
such as milk (e.g., casein, whey), animal (e.g., meat, fish),
cereal (e.g., rice, corn), vegetable (e.g., soy) or combinations
thereof. Non-limiting examples of such proteins include milk
protein isolates, milk protein concentrates as described herein,
casein protein isolates, extensively hydrolyzed casein, whey
protein, sodium or calcium caseinates, whole cow milk, partially or
completely defatted milk, soy protein isolates, soy protein
concentrates, and so forth. In one specific embodiment, the
nutritional compositions include a protein source derived from milk
proteins of human and/or bovine origin.
Carbohydrate
[0125] The nutritional products of the present disclosure may
further optionally comprise any carbohydrates that are suitable for
use in an oral nutritional product and are compatible with the
essential elements and features of such products.
[0126] Non-limiting examples of suitable carbohydrates or sources
thereof for use in the nutritional products described herein may
include maltodextrin, hydrolyzed or modified starch or cornstarch,
glucose polymers, corn syrup, corn syrup solids, rice-derived
carbohydrates, pea-derived carbohydrates, potato-derived
carbohydrates, tapioca, sucrose, glucose, fructose, lactose, high
fructose corn syrup, honey, sugar alcohols (e.g., maltitol,
erythritol, sorbitol), artificial sweeteners (e.g., sucralose,
acesulfame potassium, stevia) and combinations thereof. A
particularly desirable carbohydrate is a low dextrose equivalent
(DE) maltodextrin.
Other Optional Ingredients
[0127] The nutritional compositions of the present disclosure may
further comprise other optional components that may modify the
physical, chemical, aesthetic or processing characteristics of the
products or serve as pharmaceutical or additional nutritional
components when used in the targeted population. Many such optional
ingredients are known or otherwise suitable for use in medical food
or other nutritional products or pharmaceutical dosage forms and
may also be used in the compositions herein, provided that such
optional ingredients are safe for oral administration and are
compatible with the essential and other ingredients in the selected
product form.
[0128] Non-limiting examples of such optional ingredients include
preservatives, emulsifying agents, buffers, fructooligosaccharides,
galactooligosaccharides, polydextrose, and other prebiotics,
probiotics, pharmaceutical actives, anti-inflammatory agents,
additional nutrients as described herein, colorants, flavors,
thickening agents and stabilizers, emulsifying agents, lubricants,
and so forth.
[0129] The nutritional compositions may further comprise a
sweetening agent, preferably including at least one sugar alcohol
such as maltitol, erythritol, sorbitol, xylitol, mannitol,
isolmalt, and lactitol, and also preferably including at least one
artificial or high potency sweetener such as acesulfame K,
aspartame, sucralose, saccharin, stevia, and tagatose. These
sweetening agents, especially as a combination of a sugar alcohol
and an artificial sweetener, are especially useful in formulating
liquid beverage embodiments of the present disclosure having a
desirable favor profile. These sweetener combinations are
especially effective in masking undesirable flavors sometimes
associated with the addition of vegetable proteins to a liquid
beverage. Optional sugar alcohol concentrations in the nutritional
product may range from at least 0.01%, including from about 0.1% to
about 10%, and also including from about 1% to about 6%, by weight
of the nutritional product. Optional artificial sweetener
concentrations may range from about 0.01%, including from about
0.05% to about 5%, also including from about 0.1% to about 1.0%, by
weight of the nutritional product.
[0130] A flowing agent or anti-caking agent may be included in the
nutritional compositions as described herein to retard clumping or
caking of the powder over time and to make a powder embodiment flow
easily from its container. Any known flowing or anti-caking agents
that are known or otherwise suitable for use in a nutritional
powder or product form are suitable for use herein, non-limiting
examples of which include tricalcium phosphate, silicates, and
combinations thereof. The concentration of the flowing agent or
anti-caking agent in the nutritional composition varies depending
upon the product form, the other selected ingredients, the desired
flow properties, and so forth, but most typically range from about
0.1% to about 4%, including from about 0.5% to about 2%, by weight
of the nutritional composition.
[0131] A stabilizer may also be included in the nutritional
compositions. Any stabilizer that is known or otherwise suitable
for use in a nutritional composition is also suitable for use
herein, some non-limiting examples of which include gums such as
xanthan gum. The stabilizer may represent from about 0.1% to about
5.0%, including from about 0.5% to about 3%, including from about
0.7% to about 1.5%, by weight of the nutritional composition.
[0132] The nutritional compositions may further comprise any of a
variety of other vitamins or related nutrients, non-limiting
examples of which include vitamin A, vitamin D, vitamin E, vitamin
K, thiamine, riboflavin, pyridoxine, vitamin B.sub.12, carotenoids
(e.g., beta-carotene, zeaxanthin, lutein, lycopene), niacin, folic
acid, pantothenic acid, biotin, vitamin C, choline, inositol, salts
and derivatives thereof, and combinations thereof.
[0133] The nutritional compositions may further comprise any of a
variety of other additional minerals, non-limiting examples of
which include calcium, phosphorus, magnesium, iron, zinc,
manganese, copper, sodium, potassium, molybdenum, chromium,
chloride, and combinations thereof.
Methods of Manufacture
[0134] The nutritional compositions of the present disclosure may
be prepared by any known or otherwise effective manufacturing
technique for preparing the selected product solid or liquid form.
Many such techniques are known for any given product form such as
nutritional liquids or powders and can easily be applied by one of
ordinary skill in the art to the nutritional compositions described
herein.
[0135] The nutritional compositions of the present disclosure can
therefore be prepared by any of a variety of known or otherwise
effective formulation or manufacturing methods. In one suitable
manufacturing process, for example, at least three separate
slurries are prepared, including a protein-in-fat (PIF) slurry, a
carbohydrate-mineral (CHO-MN) slurry, and a protein-in-water (PIW)
slurry. The PIF slurry is formed by heating and mixing the oil
(e.g., canola oil, corn oil, etc.) and then adding an emulsifier
(e.g., lecithin), fat soluble vitamins, and a portion of the total
protein (e.g., milk protein concentrate, etc.) with continued heat
and agitation. The CHO-MIN slurry is formed by adding with heated
agitation to water: minerals (e.g., potassium citrate, dipotassium
phosphate, sodium citrate, etc.), trace and ultra trace minerals
(TM/UTM premix), thickening or suspending agents (e.g. avicel,
gellan, carrageenan). The resulting CHO-MIN slurry is held for 10
minutes with continued heat and agitation before adding additional
minerals (e.g., potassium chloride, magnesium carbonate, potassium
iodide, etc.), and/or carbohydrates (e.g., HMOs,
fructooligosaccharide, sucrose, corn syrup, etc.). The PIW slurry
is then formed by mixing with heat and agitation the remaining
protein, if any.
[0136] The resulting slurries are then blended together with heated
agitation and the pH adjusted to 6.6-7.0, after which the
composition is subjected to high-temperature short-time (HTST)
processing during which the composition is heat treated, emulsified
and homogenized, and then allowed to cool. Water soluble vitamins
and ascorbic acid are added, the pH is adjusted to the desired
range if necessary, flavors are added, and water is added to
achieve the desired total solid level. The composition is then
aseptically packaged to form an aseptically packaged nutritional
emulsion. This emulsion can then be further diluted, heat-treated,
and packaged to form a ready-to-feed or concentrated liquid, or it
can be heat-treated and subsequently processed and packaged as a
reconstitutable powder, e.g., spray dried, drymixed,
agglomerated.
[0137] The nutritional solid, such as a spray dried nutritional
powder or drymixed nutritional powder, may be prepared by any
collection of known or otherwise effective technique, suitable for
making and formulating a nutritional powder.
[0138] For example, when the nutritional powder is a spray dried
nutritional powder, the spray drying step may likewise include any
spray drying technique that is known for or otherwise suitable for
use in the production of nutritional powders. Many different spray
drying methods and techniques are known for use in the nutrition
field, all of which are suitable for use in the manufacture of the
spray dried nutritional powders herein.
[0139] One method of preparing the spray dried nutritional powder
comprises forming and homogenizing an aqueous slurry or liquid
comprising predigested fat, and optionally protein, carbohydrate,
and other sources of fat, and then spray drying the slurry or
liquid to produce a spray dried nutritional powder. The method may
further comprise the step of spray drying, drymixing, or otherwise
adding additional nutritional ingredients, including any one or
more of the ingredients described herein, to the spray dried
nutritional powder.
[0140] Other suitable methods for making nutritional products are
described, for example, in U.S. Pat. No. 6,365,218 (Borschel, et
al.), U.S. Pat. No. 6,589,576 (Borschel, et al.), U.S. Pat. No.
6,306,908 (Carlson, et al.), U.S. Patent Application 20030118703 A1
(Nguyen, et al.), which descriptions are incorporated herein by
reference to the extent that they are consistent herewith.
Methods of Use
[0141] The nutritional compositions as described herein can be used
to address one or more of the diseases or conditions discussed
herein, or can be used to provide one or more of the benefits
described herein, to preterm infants, infants, toddlers, and
children. The preterm infant, infant, toddler, or child utilizing
the nutritional compositions described herein may actually have or
be afflicted with the disease or condition described, or may be
susceptible to, or at risk of, getting the disease or condition
(that is, may not actually yet have the disease or condition, but
is at elevated risk as compared to the general population for
getting it due to certain conditions, family history, etc.) Whether
the preterm infant, infant, toddler, or child actually has the
disease or condition, or is at risk or susceptible to the disease
or condition, the preterm infant, infant, toddler, or child is
classified herein as "in need of" assistance in dealing with and
combating the disease or condition. For example, the preterm
infant, infant, toddler, or child may actually have respiratory
inflammation or may be at risk of getting respiratory inflammation
(susceptible to getting respiratory inflammation) due to family
history or other medical conditions, for example. Whether the
preterm infant, infant, toddler, or child actually has the disease
or condition, or is only at risk or susceptible to getting the
disease or condition, it is within the scope of the present
disclosure to assist the preterm infant, infant, toddler, or child
with the nutritional compositions described herein.
[0142] Based on the foregoing, because some of the method
embodiments of the present disclosure are directed to specific
subsets or subclasses of identified individuals (that is, the
subset or subclass of individuals "in need" of assistance in
addressing one or more specific diseases or specific conditions
noted herein), not all preterm infants, infants, toddlers, and
children will fall within the subset or subclass of preterm
infants, infants, toddlers, and children as described herein for
certain diseases or conditions.
[0143] The nutritional compositions as described herein comprise
HMOs, alone or in combination with one or more additional
components, to provide a nutritional source for reducing
inflammation, such as respiratory inflammation (e.g., respiratory
syncytial virus-induced inflammation), enteric inflammation, and
nasopharyngeal inflammation. The nutritional compositions of the
present disclosure comprising HMOs may also provide optimal
development and balanced growth and maturation of the infant's
gastrointestinal and immune systems, thereby enhancing the infant's
ability to resist microbial infection and modulate inflammatory
responses to infection (e.g., increased phagocytosis and increased
production of reactive oxidative species).
[0144] The nutritional compositions also provide growth and
maturation of the intestinal epithelial cells in an infant. In one
specific embodiment, the administration of the nutritional
compositions of the present disclosure including HMOs and
nucleotides can further activate immune activity in or by the
intestinal epithelial cells in a newborn.
[0145] Further, the use of HMOs in nutritional compositions can
reduce the growth of respiratory viruses (e.g., RSV, human
parainfluenza virus type 2, and influenza A virus), and thus,
reduce viral-induced upper respiratory infections. As such, by
utilizing HMOs, alone or in combination with other immune enhancing
factors, in a nutritional product, such as an infant formula, it is
now possible to provide infants with an alternative, or supplement,
to breast milk that more closely mimics the benefits thereof.
[0146] Along with improved growth and maturation of the infant's
immune system as described above, the use of the nutritional
compositions of the present disclosure also functions as an immune
modulator, thereby reducing inflammation induced by infection in
infants, toddlers, and children such as respiratory virus-induced
infection, and particularly, RSV-induced inflammation, and other
infection-mediated inflammatory diseases.
[0147] The addition of HMOs can further increase glutathione levels
in the body and blood of an infant, and in specific embodiments, of
a preterm infant.
[0148] When used in combination with LCPUFAs and/or antioxidants,
and particularly, with carotenoids, the HMOs can reduce oxidative
stress, which is a metabolic condition in which there is an
increased production and accumulation of oxidized biomolecules such
as lipid peroxides and their catabolites, protein carbonyls, and
oxidatively damaged DNA. The outcomes of oxidative stress range
from unwanted changes in metabolism to inflammation and cell and
tissue death. Accordingly, by reducing the incidence of unregulated
inflammation and oxidation in the infant, damage to the tissue
lining and cell death is reduced, further reducing the incidence of
inflammatory diseases, such as necrotizing enterocolitis (NEC).
[0149] In addition to the benefits discussed above, it has been
discovered that nutritional products including HMOs can modulate
production of monocyte-derived cytokines in the infant, even in the
absence of a virus. This production results in improved immunity to
further prevent microbial infection and reduce the growth of
viruses. In one specific embodiment, monocyte-derived cytokines
produced by administration of the nutritional compositions of the
present disclosure include, for example, interleukin-10,
interleukin-8, interleukin-1.alpha., interleukin-1.beta.,
interleukin-1ra, and combinations thereof.
[0150] Another benefit of utilizing HMOs in nutritional
compositions is that it has been discovered that HMOs modulate the
production of IP-10, which is a chemokine that plays an important
role in the inflammatory response to viral infection. Specifically,
a positive correlation exists between RSV clinical infection
severity in children and serum IP-10 levels. Accordingly, a
decrease in IP-10 signals a decrease in severity of RSV infection.
In one specific embodiment, IP-10 production is reduced to the
level found in uninfected controls.
[0151] Along with reducing IP-10, HMOs have been found to reduce
platelet-neutrophil complex (PNC) formation, which is present in
human blood and consists of up to 25% of unstimulated neutrophils.
As PNCs are present in aggregates, they have a greater capacity to
initiate inflammatory processes and can increase the production of
reactive oxidative species. Accordingly, a decrease in PNC
formation can lead to reduced oxidative stress and inflammation in
the infant.
EXAMPLES
[0152] The following examples illustrate specific embodiments
and/or features of the nutritional compositions of the present
disclosure. The examples are given solely for the purpose of
illustration and are not to be construed as limitations of the
present disclosure, as many variations thereof are possible without
departing from the spirit and scope of the disclosure. All
exemplified amounts are weight percentages based upon the total
weight of the composition, unless otherwise specified.
[0153] The exemplified compositions are shelf stable nutritional
compositions prepared in accordance with the manufacturing methods
described herein, such that each exemplified composition, unless
otherwise specified, includes an aseptically processed embodiment
and a retort packaged embodiment.
Examples 1-5
[0154] Examples 1-5 illustrate ready-to-feed nutritional emulsions
of the present disclosure, the ingredients of which are listed in
the table below. All ingredient amounts are listed as kilogram per
1000 kilogram batch of product, unless otherwise specified.
TABLE-US-00004 Ingredient Ex. 1 Ex. 2 Ex. 3 Ex. 4 Ex. 5 Water Q.S.
Q.S. Q.S. Q.S. Q.S. Condensed Skim Milk 86.64 86.64 86.64 86.64
86.64 Lactose 54.80 54.80 54.80 54.80 54.80 High oleic safflower
oil 14.10 14.10 14.10 14.10 14.10 Soybean oil 10.6 10.6 10.6 10.6
10.6 Coconut oil 10.1 10.1 10.1 10.1 10.1 3' sialylallactose (3'SL)
0.0948 0.090 0.085 9.479 9.005 Galactooligosaccharides (GOS) 8.63
8.63 8.63 8.63 8.63 Whey protein concentrate 6.40 6.40 6.40 6.40
6.40 Potassium citrate 478.9 g 478.9 g 478.9 g 478.9 g 478.9 g
Calcium carbonate 448.28 g 448.28 g 448.28 g 448.28 g 448.28 g Soy
lecithin 355.74 g 355.74 g 355.74 g 355.74 g 355.74 g Stabilizer
355.74 g 355.74 g 355.74 g 355.74 g 355.74 g ARA oil 368.01 g
368.01 g 368.01 g 368.01 g 368.01 g Nucleotide/chloride premix
293.26 g 293.26 g 293.26 g 293.26 g 293.26 g Potassium chloride
226.45 g 226.45 g 226.45 g 226.45 g 226.45 g Ascorbic acid 445.94 g
445.94 g 445.94 g 445.94 g 445.94 g Vitamin mineral premix 142.88 g
142.88 g 142.88 g 142.88 g 142.88 g DHA oil 137.8 g 137.8 g 137.8 g
137.8 g 137.8 g Carrageenan 180.0 g 180.0 g 180.0 g 180.0 g 180.0 g
Magnesium chloride 55.0 g 55.0 g 55.0 g 55.0 g 55.0 g Ferrous
sulfate 58.0 g 58.0 g 58.0 g 58.0 g 58.0 g Choline chloride 53.9 g
53.9 g 53.9 g 53.9 g 53.9 g Vitamin A, D.sub.3, E, K.sub.1 premix
47.4 g 47.4 g 47.4 g 47.4 g 47.4 g Citric acid 29.77 g 29.77 g
29.77 g 29.77 g 29.77 g Mixed carotenoid premix 26.40 g 26.40 g
26.40 g 26.40 g 26.40 g Sodium chloride AN AN AN AN AN L-carnitine
3.31 g 3.31 g 3.31 g 3.31 g 3.31 g Tricalcium phosphate 15.65 g
15.65 g 15.65 g 15.65 g 15.65 g Potassium phosphate monobasic 13.67
g 13.67 g 13.67 g 13.67 g 13.67 g Riboflavin 2.42 g 2.42 g 2.42 g
2.42 g 2.42 g Potassium hydroxide AN AN AN AN AN AN = as needed
Examples 6-10
[0155] Examples 6-10 illustrate ready-to-feed nutritional emulsions
of the present disclosure, the ingredients of which are listed in
the table below. All ingredient amounts are listed as kilogram per
1000 kilogram batch of product, unless otherwise specified.
TABLE-US-00005 Ingredient Ex. 6 Ex. 7 Ex. 8 Ex. 9 Ex. 10 Water Q.S.
Q.S. Q.S. Q.S. Q.S. Condensed Skim Milk 86.64 86.64 86.64 86.64
86.64 Lactose 54.80 54.80 54.80 54.80 54.80 High oleic safflower
oil 14.10 14.10 14.10 14.10 14.10 Soybean oil 10.6 10.6 10.6 10.6
10.6 Coconut oil 10.1 10.1 10.1 10.1 10.1 6' sialylallactose (6'SL)
0.0948 0.0901 0.0853 9.479 9.0047 Galactooligosaccharides (GOS)
8.63 8.63 8.63 8.63 8.63 Whey protein concentrate 6.40 6.40 6.40
6.40 6.40 Potassium citrate 478.9 g 478.9 g 478.9 g 478.9 g 478.9 g
Calcium carbonate 448.28 g 448.28 g 448.28 g 448.28 g 448.28 g Soy
lecithin 355.74 g 355.74 g 355.74 g 355.74 g 355.74 g Stabilizer
355.74 g 355.74 g 355.74 g 355.74 g 355.74 g ARA oil 368.01 g
368.01 g 368.01 g 368.01 g 368.01 g Nucleotide/chloride premix
293.26 g 293.26 g 293.26 g 293.26 g 293.26 g Potassium chloride
226.45 g 226.45 g 226.45 g 226.45 g 226.45 g Ascorbic acid 445.94 g
445.94 g 445.94 g 445.94 g 445.94 g Vitamin mineral premix 142.88 g
142.88 g 142.88 g 142.88 g 142.88 g DHA oil 137.8 g 137.8 g 137.8 g
137.8 g 137.8 g Carrageenan 180.0 g 180.0 g 180.0 g 180.0 g 180.0 g
Magnesium chloride 55.0 g 55.0 g 55.0 g 55.0 g 55.0 g Ferrous
sulfate 58.0 g 58.0 g 58.0 g 58.0 g 58.0 g Choline chloride 53.9 g
53.9 g 53.9 g 53.9 g 53.9 g Vitamin A, D.sub.3, E, K.sub.1 premix
47.40 g 47.40 g 47.40 g 47.40 g 47.40 g Citric acid 29.77 g 29.77 g
29.77 g 29.77 g 29.77 g Mixed carotenoid premix 26.40 g 26.40 g
26.40 g 26.40 g 26.40 g Sodium chloride AN AN AN AN AN L-carnitine
3.31 g 3.31 g 3.31 g 3.31 g 3.31 g Tricalcium phosphate 15.65 g
15.65 g 15.65 g 15.65 g 15.65 g Potassium phosphate monobasic 13.67
g 13.67 g 13.67 g 13.67 g 13.67 g Riboflavin 2.42 g 2.42 g 2.42 g
2.42 g 2.42 g Potassium hydroxide AN AN AN AN AN AN = as needed
Examples 11-15
[0156] Examples 11-15 illustrate concentrated liquid emulsions of
the present disclosure, the ingredients of which are listed in the
table below. All ingredient amounts are listed as kilogram per 1000
kilogram batch of product, unless otherwise specified.
TABLE-US-00006 Ingredient Ex. 11 Ex. 12 Ex. 13 Ex. 14 Ex. 15 Water
Q.S. Q.S. Q.S. Q.S. Q.S. Condensed Skim Milk 157.67 157.67 157.67
157.67 157.67 Lactose 108.66 108.66 108.66 108.66 108.66 High oleic
safflower oil 26.82 26.82 26.82 26.82 26.82 Soybean oil 20.16 20.16
20.16 20.16 20.16 Coconut oil 19.24 19.24 19.24 19.24 19.24 3'
sialylallactose (3'SL) 0.1896 0.1802 0.1706 18.958 18.009
Galactooligosaccharides (GOS) 17.67 17.67 17.67 17.67 17.67 Whey
protein concentrate 12.20 12.20 12.20 12.20 12.20 Potassium citrate
1.277 1.277 1.277 1.277 1.277 Calcium carbonate 996.1 g 996.1 g
996.1 g 996.1 g 996.1 g Soy lecithin 685.0 g 685.0 g 685.0 g 685.0
g 685.0 g Monoglycerides 685.0 g 685.0 g 685.0 g 685.0 g 685.0 g
ARA oil 684.2 g 684.2 g 684.2 g 684.2 g 684.2 g Nucleotide/chloride
premix 568.9 g 568.9 g 568.9 g 568.9 g 568.9 g Potassium chloride
429.7 g 429.7 g 429.7 g 429.7 g 429.7 g Ascorbic acid 293.8 g 293.8
g 293.8 g 293.8 g 293.8 g Vitamin mineral premix 276.9 g 276.9 g
276.9 g 276.9 g 276.9 g DHA oil 256.1 g 256.1 g 256.1 g 256.1 g
256.1 g Carrageenan 200.0 g 200.0 g 200.0 g 200.0 g 200.0 g
Magnesium chloride 173.3 g 173.3 g 173.3 g 173.3 g 173.3 g Ferrous
sulfate 112.7 g 112.7 g 112.7 g 112.7 g 112.7 g Choline chloride
104.8 g 104.8 g 104.8 g 104.8 g 104.8 g Vitamin A, D.sub.3, E,
K.sub.1 premix 86.90 g 86.90 g 86.90 g 86.90 g 86.90 g Citric acid
57.50 g 57.50 g 57.50 g 57.50 g 57.50 g Mixed carotenoid premix
41.90 g 41.90 g 41.90 g 41.90 g 41.90 g Sodium chloride 23.50 g
23.50 g 23.50 g 23.50 g 23.50 g L-carnitine 6.40 g 6.40 g 6.40 g
6.40 g 6.40 g Tricalcium phosphate AN AN AN AN AN Potassium
phosphate monobasic AN AN AN AN AN Potassium hydroxide AN AN AN AN
AN AN = as needed
Examples 16-20
[0157] Examples 16-20 illustrate ready-to-feed nutritional
emulsions of the present disclosure, the ingredients of which are
listed in the table below. All ingredient amounts are listed as
kilogram per 1000 kilogram batch of product, unless otherwise
specified.
TABLE-US-00007 Ingredient Ex. 16 Ex. 17 Ex. 18 Ex. 19 Ex. 20 Water
Q.S. Q.S. Q.S. Q.S. Q.S. Condensed Skim Milk 86.64 86.64 86.64
86.64 86.64 Lactose 54.80 54.80 54.80 54.80 54.80 High oleic
safflower oil 14.10 14.10 14.10 14.10 14.10 Soybean oil 10.6 10.6
10.6 10.6 10.6 Coconut oil 10.1 10.1 10.1 10.1 10.1 HMO Mixture
0.0948 0.0901 0.0853 9.479 9.0047 6'SL 0.0316 0.0300 0.0284 3.159
3.002 2'FL 0.0316 0.0300 0.0284 3.159 3.002 LNnT 0.0316 0.0300
0.0284 3.159 3.002 Galactooligosaccharides (GOS) 8.63 8.63 8.63
8.63 8.63 Whey protein concentrate 6.40 6.40 6.40 6.40 6.40
Potassium citrate 478.9 g 478.9 g 478.9 g 478.9 g 478.9 g Calcium
carbonate 448.28 g 448.28 g 448.28 g 448.28 g 448.28 g Soy lecithin
355.74 g 355.74 g 355.74 g 355.74 g 355.74 g Stabilizer 355.74 g
355.74 g 355.74 g 355.74 g 355.74 g ARA oil 368.01 g 368.01 g
368.01 g 368.01 g 368.01 g Nucleotide/chloride premix 293.26 g
293.26 g 293.26 g 293.26 g 293.26 g Potassium chloride 226.45 g
226.45 g 226.45 g 226.45 g 226.45 g Ascorbic acid 445.94 g 445.94 g
445.94 g 445.94 g 445.94 g Vitamin mineral premix 142.88 g 142.88 g
142.88 g 142.88 g 142.88 g DHA oil 137.8 g 137.8 g 137.8 g 137.8 g
137.8 g Carrageenan 180.0 g 180.0 g 180.0 g 180.0 g 180.0 g
Magnesium chloride 55.0 g 55.0 g 55.0 g 55.0 g 55.0 g Ferrous
sulfate 58.0 g 58.0 g 58.0 g 58.0 g 58.0 g Choline chloride 53.9 g
53.9 g 53.9 g 53.9 g 53.9 g Vitamin A, D.sub.3, E, K.sub.1 premix
47.40 g 47.40 g 47.40 g 47.40 g 47.40 g Citric acid 29.77 g 29.77 g
29.77 g 29.77 g 29.77 g Mixed carotenoid premix 26.40 g 26.40 g
26.40 g 26.40 g 26.40 g Sodium chloride AN AN AN AN AN L-carnitine
3.31 g 3.31 g 3.31 g 3.31 g 3.31 g Tricalcium phosphate 15.65 g
15.65 g 15.65 g 15.65 g 15.65 g Potassium phosphate monobasic 13.67
g 13.67 g 13.67 g 13.67 g 13.67 g Riboflavin 2.42 g 2.42 g 2.42 g
2.42 g 2.42 g Potassium hydroxide AN AN AN AN AN AN = as needed
Examples 21-25
[0158] Examples 21-25 illustrate concentrated liquid emulsions of
the present disclosure, the ingredients of which are listed in the
table below. All ingredient amounts are listed as kilogram per 1000
kilogram batch of product, unless otherwise specified.
TABLE-US-00008 Ingredient Ex. 21 Ex. 22 Ex. 23 Ex. 24 Ex. 25 Water
Q.S. Q.S. Q.S. Q.S. Q.S. Condensed Skim Milk 157.67 157.67 157.67
157.67 157.67 Lactose 108.66 108.66 108.66 108.66 108.66 High oleic
safflower oil 26.82 26.82 26.82 26.82 26.82 Soybean oil 20.16 20.16
20.16 20.16 20.16 Coconut oil 19.24 19.24 19.24 19.24 19.24 HMO
Mixture 18.957 18.009 17.061 19.905 20.853 6'SL 6.319 6.003 5.687
6.635 6.951 2'FL 6.319 6.003 5.687 6.635 6.951 LNnT 6.319 6.003
5.687 6.635 6.951 Galactooligosaccharides (GOS) 17.67 17.67 17.67
17.67 17.67 Whey protein concentrate 12.20 12.20 12.20 12.20 12.20
Potassium citrate 1.277 1.277 1.277 1.277 1.277 Calcium carbonate
996.1 g 996.1 g 996.1 g 996.1 g 996.1 g Soy lecithin 685.0 g 685.0
g 685.0 g 685.0 g 685.0 g Monoglycerides 685.0 g 685.0 g 685.0 g
685.0 g 685.0 g ARA oil 684.2 g 684.2 g 684.2 g 684.2 g 684.2 g
Nucleotide/chloride premix 568.9 g 568.9 g 568.9 g 568.9 g 568.9 g
Potassium chloride 429.7 g 429.7 g 429.7 g 429.7 g 429.7 g Ascorbic
acid 293.8 g 293.8 g 293.8 g 293.8 g 293.8 g Vitamin mineral premix
276.9 g 276.9 g 276.9 g 276.9 g 276.9 g DHA oil 256.1 g 256.1 g
256.1 g 256.1 g 256.1 g Carrageenan 200.0 g 200.0 g 200.0 g 200.0 g
200.0 g Magnesium chloride 173.3 g 173.3 g 173.3 g 173.3 g 173.3 g
Ferrous sulfate 112.7 g 112.7 g 112.7 g 112.7 g 112.7 g Choline
chloride 104.8 g 104.8 g 104.8 g 104.8 g 104.8 g Vitamin A,
D.sub.3, E, K.sub.1 premix 86.90 g 86.90 g 86.90 g 86.90 g 86.90 g
Citric acid 57.50 g 57.50 g 57.50 g 57.50 g 57.50 g Mixed
carotenoid premix 41.90 g 41.90 g 41.90 g 41.90 g 41.90 g Sodium
chloride 23.50 g 23.50 g 23.50 g 23.50 g 23.50 g L-carnitine 6.40 g
6.40 g 6.40 g 6.40 g 6.40 g Tricalcium phosphate AN AN AN AN AN
Potassium phosphate monobasic AN AN AN AN AN Potassium hydroxide AN
AN AN AN AN AN = as needed
Examples 26-30
[0159] Examples 26-30 illustrate human milk fortifier liquids of
the present disclosure, the ingredients of which are listed in the
table below. All ingredient amounts are listed as kilogram per 1000
kilogram batch of product, unless otherwise specified.
TABLE-US-00009 Ingredient Ex. 26 Ex. 27 Ex. 28 Ex. 29 Ex. 30 Water
Q.S. Q.S. Q.S. Q.S. Q.S. Non-fat milk 353 353 353 353 353 Corn
Syrup Solids 85.3 85.3 85.3 85.3 85.3 Medium Chain Triglycerides
53.2 53.2 53.2 53.2 53.2 Whey Protein Concentrate 47.2 47.2 47.2
47.2 47.2 HMO Mixture 18.957 18.009 17.061 19.905 20.853 6'SL 6.319
6.003 5.687 6.635 6.951 2'FL 6.319 6.003 5.687 6.635 6.951 LNnT
6.319 6.003 5.687 6.635 6.951 Calcium Phosphate 25.5 25.5 25.5 25.5
25.5 Ascorbic Acid 5.6 5.6 5.6 5.6 5.6 Potassium Citrate 3.1 3.1
3.1 3.1 3.1 Magnesium Chloride 2.8 2.8 2.8 2.8 2.8 Sodium Citrate
1.4 1.4 1.4 1.4 1.4 Sodium Chloride 1.4 1.4 1.4 1.4 1.4 Soy
Lecithin 609 g 609 g 609 g 609 g 609 g M-Inositol 500 g 500 g 500 g
500 g 500 g Niacinamide 400 g 400 g 400 g 400 g 400 g ARA Oil 313 g
313 g 313 g 313 g 313 g Tocopherol Acetate 310 g 310 g 310 g 310 g
310 g Zinc Sulfate 300 g 300 g 300 g 300 g 300 g Calcium
Pantothenate 182 g 182 g 182 g 182 g 182 g Ferrous Sulfate 133 g
133 g 133 g 133 g 133 g DHA Oil 116 g 116 g 116 g 116 g 116 g
Vitamin A Palmitate 100 g 100 g 100 g 100 g 100 g Cupric Sulfate
51.0 g 51.0 g 51.0 g 51.0 g 51.0 g Thiamine Hydrochloride 50.0 g
50.0 g 50.0 g 50.0 g 50.0 g Riboflavin 47.0 g 47.0 g 47.0 g 47.0 g
47.0 g Pyridoxine Hydrochloride 27.0 g 27.0 g 27.0 g 27.0 g 27.0 g
Vitamin D.sub.3 20.0 g 20.0 g 20.0 g 20.0 g 20.0 g Folic Acid 3.5 g
3.5 g 3.5 g 3.5 g 3.5 g Biotin 3.4 g 3.4 g 3.4 g 3.4 g 3.4 g
Manganous Sulfate 1.5 g 1.5 g 1.5 g 1.5 g 1.5 g Phylloquinone 1.2 g
1.2 g 1.2 g 1.2 g 1.2 g Cyanocobalamin 100 mg 100 mg 100 mg 100 mg
100 mg Sodium Selenate 43.0 mg 43.0 mg 43.0 mg 43.0 mg 43.0 mg
Examples 31-35
[0160] Examples 31-35 illustrate spray dried nutritional powders of
the present disclosure, the ingredients of which are listed in the
table below. All ingredient amounts are listed as kilogram per 1000
kilogram batch of product, unless otherwise specified.
TABLE-US-00010 Ingredient Ex. 31 Ex. 32 Ex. 33 Ex. 34 Ex. 35
Condensed Skim Milk 698.5 698.5 698.5 698.5 698.5 Lactose 386.0
386.0 386.0 386.0 386.0 High oleic safflower oil 114.4 114.4 114.4
114.4 114.4 Soybean oil 85.51 85.51 85.51 85.51 85.51 Coconut oil
78.76 78.76 78.76 78.76 78.76 3' sialylallactose (3'SL) 0.3792
0.3604 0.3412 37.916 36.0188 Galactooligosaccharides (GOS) 69.50
69.50 69.50 69.50 69.50 Whey protein concentrate 51.08 51.08 51.08
51.08 51.08 Potassium citrate 9.168 9.168 9.168 9.168 9.168 Calcium
carbonate 4.054 4.054 4.054 4.054 4.054 Soy lecithin 1.120 1.120
1.120 1.120 1.120 ARA oil 2.949 2.949 2.949 2.949 2.949
Nucleotide/chloride premix 2.347 2.347 2.347 2.347 2.347 Potassium
chloride 1.295 1.295 1.295 1.295 1.295 Ascorbic acid 1.275 1.275
1.275 1.275 1.275 Vitamin mineral premix 1.116 1.116 1.116 1.116
1.116 DHA oil 1.113 1.113 1.113 1.113 1.113 Magnesium chloride
1.038 1.038 1.038 1.038 1.038 Sodium chloride 579.4 g 579.4 g 579.4
g 579.4 g 579.4 g Ferrous sulfate 453.6 g 453.6 g 453.6 g 453.6 g
453.6 g Choline chloride 432.1 g 432.1 g 432.1 g 432.1 g 432.1 g
Vitamin A, D.sub.3, E, K.sub.1 premix 377.2 g 377.2 g 377.2 g 377.2
g 377.2 g Ascorbyl Palmitate 361.3 g 361.3 g 361.3 g 361.3 g 361.3
g Mixed carotenoid premix 350.1 g 350.1 g 350.1 g 350.1 g 350.1 g
Mixed Tocopherols 159.2 g 159.2 g 159.2 g 159.2 g 159.2 g
L-carnitine 26.30 g 26.30 g 26.30 g 26.30 g 26.30 g Riboflavin
3.181 g 3.181 g 3.181 g 3.181 g 3.181 g Tricalcium phosphate 0-5.23
0-5.23 0-5.23 0-5.23 0-5.23 Potassium phosphate monobasic 0-5.23
0-5.23 0-5.23 0-5.23 0-5.23 Potassium hydroxide AN AN AN AN AN AN =
as needed
Example 36
[0161] In this Example, the effect of purified human milk
oligosaccharides (HMO) on in vitro inhibition of viral infectivity
is analyzed.
[0162] Samples are prepared by co-incubation of a uniform virus
dose of from about 500 units/mL to about 1,000 units/mL of one of
three respiratory viruses: (1) respiratory syncytial virus (RSV);
(2) human parainfluenza virus (HPIV3); or (3) H1N1 influenza virus
with one of the following HMOs: (1) 3'-sialyllactose (3'SL); (2)
6'-sialyllactose (6'SL); (3) 3'-fucosyllactose (3'FL); (4)
2'-fucosyllactose (2'FL); (5) lacto-N-neotetraose (LNnT); or (6)
sialic acid (SA). The HMOs are added at concentrations of either 1
mg/mL or 10 mg/mL. The antiviral activities of the various HMOs on
the respiratory viruses are evaluated, and the results are shown in
the table below:
TABLE-US-00011 IC50 (mg HMO/mL) HMO RSV HPIV3 H1N1 Influenza 3'SL
>10 >10 ~5 6'SL >10 >10 ~10 3'FL ~5 ~2 ~5 2'FL >10
>10 ~10 LNnT >10 NT >10 SA NT ~2 ~5 NT = Not Tested
[0163] The results show that 3'FL, at a concentration of 1 mg/ML
(IC50 .about.2-5 mg/ML), has anti-viral activity for all three
respiratory viruses. This result is unexpected as previous
published reports show only sialylated oligo forms providing
antiviral activity. SA significantly inhibits HPIV3 and H1N1
viruses at a concentration of 1 mg/mL. H1N1 influenza virus is also
inhibited by 3'SL at a concentration of 1 mg/mL.
Example 37
[0164] In this Example, the ability of various HMOs to block H1N1
influenza virus infectivity in vitro is analyzed.
[0165] Virus infectivity is assessed by observing cytopathic effect
(CPE) and quantifying virus focus forming units. To create virus
stocks, H1N1 influenza virus is purchased from ATCC (VR 1469) and
expanded in Madin-Darby Canine Kidney (MDCK) epithelial cells (ATCC
CCL-34). Cell-free supernatants are frozen in aliquots to maintain
stock virus. During initial virus culture and expansion to create
virus stocks, cell CPE is observed.
[0166] To quantify virus infectivity, an immunocytochemical focus
forming unit (FFU) assay is developed using commercially purchased
mouse monoclonal antibodies against the virus nucleoprotein coupled
with a biotinylated anti-mouse IgG secondary antibody. To visualize
virus-infected cell foci, color development is performed using
Strepavidin HRP (ABC from Vector Laboratories, Inc.). Although the
total number of virus foci appear proportional to the infecting
virus concentration, the foci are quite large, disperse, and there
are numerous individually infected cells that do not form foci,
especially at higher virus concentrations. As this makes
quantifying of virus infectivity difficult and time-consuming, the
FFU assay is further refined by varying virus concentration and by
applying an overlay medium of Tragacanth gum to help reduce
Brownian movement spread of the virus throughout the cell
layer.
[0167] The use of Tragacanth gum improves the assay by reducing the
number of individually infected cells while still allowing for the
formation of readily observable foci. While the foci vary in size,
with some being quite large, they are still easily quantified and
directly proportional to virus concentration or titer by using a
grid technique during the enumeration.
[0168] Once verified, the assay is used with various HMOs for the
ability to block H1N1 virus infectivity. Specifically, the HMOs are
added, at concentrations of 0.01 mg/mL, 0.1 mg/mL, 1.0 mg/mL, and
10 mg/mL, to the inoculating virus suspension, incubated at
37.degree. C. for one hour, and then added to MDCK monolayer cells.
This mixture is allowed to bind to the cell layer for thirty
minutes at 37.degree. C. The cell layer is then washed, and the
cells are further incubated for approximately 18-24 hours before
fixing and processing for immunocytochemical staining. The results
are shown in FIG. 1.
[0169] As shown in FIG. 1, 3'FL, 3'SL, and SA each inhibit virus
infectivity by greater than 90% when used at a concentration of 10
mg/mL. 2'FL and 6'SL inhibit infectivity by approximately 60% at 10
mg/mL.
Example 38
[0170] In this Example, nutritional compositions including various
HMOs are evaluated for their effects on reducing oxidative stress
in preterm piglets.
[0171] Preterm piglets are harvested by caesarian section (CS) at
92% of gestation. Piglets receive total parenteral nutrition (TPN)
for 48 hours. After 48 hours, TPN is ceased and the piglets are
randomized into three groups: a formula group (n=7) that is fed
Enfamil.RTM. Lacto-Free, commercially available from Mead Johnson,
Evansville, Ind.; a treatment group (n=9) that is fed Enfamil.RTM.
Lacto-Free with the addition of a combination of 400 mg/L 6'SL,
1500 mg/L 2'FL, and 200 mg/L LNnT; and a colostrum group (n=5) that
is fed bovine colostrum. Piglets are fed their respective feeding
enterally at a rate of 120 mL formula per kg body weight for the
next 48 hours. Piglets are then euthanized after 48 hours of
enteral nutrition (EN), or earlier if a piglet develops signs of
necrotizing enterocolitis. Blood is collected via an umbilical
artery catheter, and plasma is separated from the blood and stored
at -70.degree. C. until analyzed.
[0172] Glutathione (GSH) concentrations are measured in plasma
taken from the piglets just prior to feeding time (time 0), and at
6 hours, 12 hours, 24 hours, 36 hours, and 48 hours after feeding
using a commercially available assay (NWLSS Glutathione Assay
#NWK-GSH01, Northwest Life Science Specialties, Vancouver, Wash.).
The results are shown in FIG. 2.
[0173] As shown in FIG. 2, the concentration of GSH in blood plasma
from the control group declines from time 0 to 6 hours after
feeding. GSH remains lower in the control group 24 hours after EN.
In contrast, piglets fed the composition with a combination of HMOs
have a pattern of blood plasma GSH levels that are comparable to
the colostrum piglets.
Example 39
[0174] In this Example, the abilities of 3'SL, 6'SL, and LNnT to
reduce virus-induced inflammation in vitro are demonstrated.
[0175] Specifically, either 3'SL or 6'SL is added, at
concentrations of 0.1 mg/mL, 0.2 mg/mL, or 0.5 mg/mL to fresh
peripheral blood mononuclear cells and incubated at 37.degree. C.
in 5% CO.sub.2 to pretreat the cells for approximately 24 hours.
LNnT is added, at concentrations of 0.1 mg/mL, 0.2 mg/mL, or 1
mg/mL to fresh peripheral blood mononuclear cells and incubated at
37.degree. C. in 5% CO.sub.2 to pretreat the cells for
approximately 24 hours. Lactose is included as a carbohydrate
control. Matched endotoxin unit concentration controls are included
to allow differentiation of ingredient effects from inherent low
levels of endotoxin. Some variables are then incubated with RSV at
a multiplicity of infection (MOI) of 0.1 for approximately 1 hour
at 37.degree. C. in 5% in CO.sub.2. Uninfected control variables
are incubated with medium for approximately 1 hour at 37.degree. C.
in 5% CO.sub.2. After approximately one hour, fresh medium alone,
or fresh medium containing the appropriate concentration of 3'SL,
6'SL, LNnT, lactose, or endotoxin is added to the appropriate tubes
and the cells are incubated for 48 hours at 37.degree. C. in 5%
CO.sub.2 Supernatants are collected at 24 and 48 hours
post-infection.
[0176] Cytokines are measured in supernatants for each variable at
24 and 48 hours to assess the effects of HMOs on the early immune
response to RSV. Cytokines are measured using custom Bio-Plex Human
cytokine kits from Bio-Rad. Results for interferon-inducible
protein 10 (IP-10, also known as CXCL 10) are shown in FIGS. 3 and
4 for 3'SL and 6'SL, and in FIGS. 5 and 6 for LNnT. IP-10 is a CXC
chemokine that attracts, binds to and activates the CXCR3 receptor
on natural killer cells and memory T cells. IP-10 is expressed by
monocytes and a number of other cells, and is induced by
interferon. A positive correlation exists between RSV clinical
disease severity in children (as measured by: length of hospital
stay, fever, and number of days supplemental O.sub.2 is required)
and serum IP-10. Therefore, a decrease in IP-10 signals a decrease
in severity of RSV disease experienced.
[0177] IP-10 results for 3'SL and 6'SL are detailed in FIGS. 3 and
4 and show some variability in donor response, but surprisingly,
6'SL clearly downregulates IP-10 in virus-infected variables in
both donors. Note that 6'SL is able to reduce IP-10 to levels found
in uninfected controls. 3'SL is not effective in Donor B, but
downregulates RSV-induced IP-10 in Donor E. These data show both
3'SL and 6'SL dampen RSV-induced IP-10, but that 6'SL is more
effective at downregulation of IP-10. Results also suggest that
levels below 0.1 mg/mL of 6'SL as well as levels greater than 0.5
mg/mL may be effective at reducing IP-10 in some individuals.
[0178] IP-10 results for LNnT are detailed in FIGS. 5 and 6 and
show some variability in donor response, but surprisingly, LNnT
clearly downregulates IP-10 in virus-infected variables in both
donors. Note that LNnT is able to reduce IP-10 to levels found in
uninfected controls. Results also suggest that levels between 0.2
and 1 mg LNnT/mL as well as greater than 1 mg/mL may be effective
at reducing IP-10 in some individuals. Inclusion of matched
endotoxin unit concentration controls clearly indicates that the
decrease in IP-10 is not due to the presence of very low levels of
endotoxin in the LNnT.
[0179] In FIGS. 7 and 8, cytokine results also surprisingly show
6'SL increases interleukin 10 (IL-10) concentration in a
dose-dependent manner in the presence or absence of RSV. IL-10
results for LNnT are shown in FIGS. 9 and 10. Surprisingly, LNnT
increases IL-10 concentration in a dose-dependent manner in the
presence or absence of RSV. IL-10 is produced by activated CD8+
T-cells, by CD4+ T-cells after both antigen-specific and polyclonal
activation, and by monocytes following cell activation by bacterial
lipopolysaccharides. Inclusion of matched endotoxin unit
concentration controls clearly differentiates that the increase in
IL-10 is not due to the presence of very low levels of endotoxin in
the 6'SL or the LNnT.
[0180] Surprisingly, it is found that pretreatment for 24 hours by
6'SL, 3'SL, or LNnT is effective in reducing inflammation caused by
RSV. Moreover, 6'SL and LNnT are shown to be more effective than
3'SL at dampening virus-induced inflammation as measured by a
decrease in IP-10. Further, it is shown that 6'SL is
immunomodulatory in the absence of the virus, as the inclusion of
6'SL induces and/or modifies the production of monocyte-derived
cytokines such as IL-10, MIP-1.beta., Interferon-.gamma., IL-8,
IL-1.alpha., IL-1.beta., and IL-1ra. Surprisingly, 3'SL is also
immunomodulatory in the presence or absence of the virus, as the
inclusion of 3'SL induces and/or modifies the production of
monocyte-derived cytokines such as MIP-1.beta., Interferon-.gamma.,
IL-8, and IL-1ra. Surprisingly, LNnT is also immunomodulatory in
the presence or absence of the virus, as the inclusion of LNnT
induces and/or modifies the production of monocyte-derived
cytokines such as IL-10, MIP-1.beta., Interferon-.gamma., IL-8,
IL-1.alpha., IL-1.beta., and IL-1ra.
Example 40
[0181] In this example, the ability of the combination of 2'FL and
lycopene to reduce viral replication in vitro is demonstrated.
[0182] Specifically, on Day -1, Calu3 monolayers are seeded in
sufficient numbers to reach 95-100% confluence in 24 well plates by
Day 0. On day 0, 2'FL at a concentration of 0.5 .mu.g/mL, 1
.mu.g/mL, or 5 .mu.g/mL alone or in combination with lycopene at a
concentration of 0.5 .mu.g/mL, 1 .mu.g/mL, or 5 .mu.g/mL or
tetrahydrofuran (THF) at a concentration of 0.5 .mu.g/mL, 1
.mu.g/mL, or 5 .mu.g/mL are added and incubated for approximately
24 hours at 37.degree. C. in 5% CO.sub.2. THF is a solvent used to
solubilize the lycopene, and as such, is a control included to
differentiate solvent effects. On day 1, the cell supernatants are
removed and the monolayers are incubated with medium alone or
medium plus Respiratory Syncytial Virus (RSV) for approximately 1
hour at 37.degree. C. in 5% CO.sub.2 at a multiplicity of infection
(MOI) of 1. After approximately 1 hour, fresh medium alone or
containing the appropriate concentrations of 2'FL and lycopene or
2'FL and THF is added to the appropriate wells, and the cells are
incubated for 48 hours at 37.degree. C. in 5% CO.sub.2. On day 3,
supernatants and cell lysates are collected separately, aliquotted
and stored frozen at -70.degree. C. for later analysis. Cell
lysates are analyzed by TaqMan qRTPCR to assess viral replication
through measurement of RSV NS1 copy numbers.
[0183] As shown in FIG. 11, the combination of 2'FL and lycopene at
certain combinations (1 .mu.g and 10 .mu.g of 2'FL+0.5 .mu.g
lycopene; and 0.1 .mu.g and 1 .mu.g of 2'FL+1 .mu.g lycopene) shows
a synergistic effect that results in a dramatic inhibition of viral
load as measured by copies of RSV NS1. Further, as can be seen in
FIG. 11, 2'FL alone showed a modest concentration dependent
decrease in RSV NS1. Surprisingly, the combination of 2'FL and
lycopene at select concentrations can substantially inhibit RSV
replication in vitro.
Example 41
[0184] In this example, the ability of the combination of 2'FL and
lycopene to reduce IP-10, a marker of viral inflammation in vitro
is demonstrated.
[0185] Specifically, 2'FL at a concentration of 0.1 mg/mL, 0.2
mg/mL, or 1 mg/mL in combination with lycopene at a concentration
of 0.5 .mu.g/mL, 1.0 .mu.g/mL, or 5.0 .mu.g/mL or THF at a
concentration of 0.5 .mu.g/mL, 1.0 .mu.g/mL, or 5.0 .mu.g/mL is
added to fresh human peripheral blood mononuclear cells (PBMCs) and
incubated at 37.degree. C. in 5% CO.sub.2 to pretreat the cells for
approximately 24 hours. THF is a solvent used to solubilize the
lycopene, and as such, is a control included to differentiate
solvent effects. After approximately 24 hours, some variables are
then incubated with RSV at a multiplicity of infection (MOI) of 1
for approximately 1 hour at 37.degree. C. in 5% in CO.sub.2. The
uninfected control variable is incubated with medium for
approximately 1 hour at 37.degree. C. in 5% CO.sub.2. After
approximately 1 hour, fresh medium alone or containing the
appropriate concentrations of 2'FL and lycopene or 2'FL and THF is
added to the appropriate variables, and the PBMCs are incubated for
48 hours at 37.degree. C. in 5% CO.sub.2. Supernatants are
collected at 48 hours post-infection. Cytokines are measured in
supernatants for each variable at 48 hours using Luminex human
cytokine kits to assess the effects of HMOs on the early immune
response to RSV.
[0186] Interferon-inducible Protein 10 (IP-10, also known as
CXCL10) is a CXC chemokine that attracts, binds to and activates
the CXCR3 receptor on Natural Killer Cells and Memory T cells.
IP-10 is expressed by monocytes and a number of other cells, and is
induced by interferon. A positive correlation exists between RSV
clinical disease severity in children (as measured by: length of
hospital stay, fever, and number of days supplemental O.sub.2 was
required) and serum IP-10. Therefore, a decrease in IP-10 may
signal a decrease in severity of RSV disease experienced.
[0187] Surprisingly, as shown in FIG. 12, the combination of 2'FL
and lycopene resulted in a stepwise concentration dependent
downregulation of IP-10. Although effects were evident with 2'FL at
the lower lycopene concentrations, the strongest decrease in IP-10
was seen for 2'FL at concentrations of 0.1 mg/mL, 0.2 mg/mL, or 1
mg/mL in combination with the highest lycopene concentration tested
of 5.0 .mu.g/mL. As such, it can be concluded that the combination
of 2'FL and lycopene can decrease the severity of RSV disease
experienced, especially at a lycopene concentration of 5.0
.mu.g/mL.
Example 42
[0188] In this example, the ability of the combination of 2'FL,
3'SL, and lycopene to reduce IP-10, a marker of viral inflammation,
in vitro is demonstrated.
[0189] Specifically, 2'FL (at a concentration of 0.1 mg/mL), 3'SL
(at a concentration of 0.1 mg/mL), and lycopene (at concentrations
of 0.5 .mu.g/mL, 1.0 .mu.g/mL, or 5 .mu.g/mL) or tetrahydrofuran
(THF) (at concentrations of 0.5 .mu.g/mL, 1.0 .mu.g/mL, or 5
.mu.g/mL) alone or in combinations (as shown in FIG. 13) are added
to fresh human peripheral blood mononuclear cells (PBMCs) and
incubated at 37.degree. C. in 5% CO.sub.2 to pretreat the cells for
approximately 24 hours. THF is a solvent used to solubilize the
lycopene, and as such, a THF concentration control is included to
differentiate solvent effects. After approximately 24 hours, some
variables are then incubated with RSV at a multiplicity of
infection (MOI) of 1 for approximately 1 hour at 37.degree. C. in
5% in CO.sub.2. The uninfected control variable is incubated with
medium for approximately 1 hour at 37.degree. C. in 5% CO.sub.2.
After approximately 1 hour, fresh medium alone or containing the
appropriate concentrations of 2'FL, 3'SL, and Lycopene; 2'FL; 3'SL;
2'FL and 3'SL; lycopene; or THF is added to the appropriate
variables, and the PBMCs are incubated for 48 hours at 37.degree.
C. in 5% CO.sub.2. Supernatants are collected at 48 hours
post-infection. Cytokines are measured in supernatants for each
variable at 48 hours using Luminex human cytokine kits to assess
the effects of HMOs on the early immune response to RSV.
[0190] Interferon-inducible Protein 10 (IP-10, also known as
CXCL10) is a CXC chemokine that attracts, binds to and activates
the CXCR3 receptor on Natural Killer Cells and Memory T cells.
IP-10 is expressed by monocytes and a number of other cells, and is
induced by interferon. A positive correlation exists between RSV
clinical disease severity in children (as measured by: length of
hospital stay, fever, and number of days supplemental O.sub.2 was
required) and serum IP-10. Therefore, a decrease in IP-10 may
signal a decrease in severity of RSV disease experienced.
[0191] Surprisingly, the combination of 2'FL, 3'SL, and lycopene
results in a downregulation of IP-10 that increases with increasing
dose of lycopene (See FIG. 13). The synergistic decrease (86%
decrease) in IP-10 for 2'FL, 3'SL, and lycopene is seen with the
highest lycopene concentration (5.0 .mu.g/mL) tested. As such, it
can be concluded that the combination of 2'FL, 3'SL, and lycopene
may have a synergistic effect in decreasing the severity of RSV
disease experienced.
* * * * *