Title of Invention

INFANT FORMULAS CONTAINING DOCOSAHEXAENOIC ACID AND LUTEIN

Abstract Disclosed are infant formulas and corresponding methods of using them to promote retinal health and vision development in infants. The formulas, which are free of egg phospholipids and comprise fat, protein, carbohydrate, vitamins, and minerals, including docosahexaenoic acid and, on a ready-to-feed basis, at least about 50 mcg/liter of lutein, wherein the weight ratio of lutein (mcg) to docosahexaenoic acid (mg) is from about 1:2 to about 10:1. The formulas are also believed to be especially useful in reducing the risk of retinopathy of prematurity in preterm infants.
Full Text FORM 2
The Patents Act, 1970
(39 of 1970)
&
The Patent Rules, 2003
COMPLETE SPECIFICATION
(See section 10 and rule 13)
"INFANT FORMULAS CONTAINING DOCOSAHEXAENOIC ACID AND LUTEIN"
ABBOTT LABORATORIES, a corporation organized and existing under the laws of State of Illinois, United States of America, having its Registered Office at Dept.377/AP6A-1, 100 Abbott Park Road, Abbott Park, Illinois 60064 (US)
The following specification particularly describes the invention and the manner in which it is to be performed

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INFANT FORMULAS CONTAINING DOCOSAHEXAENOIC ACTD AND LUTEIN
TECHNICAL FIELD [0001] The present invention relates to infant formulas containing select combinations of docosahexaenoic acid arid lutein for promoting retinal health and vision development in infants.
BACKGROUND OF THE INVENTION
[0002] Infant formulas are commonly used today to provide supplemental or sole source nutrition early in life. These formulas contain protein, carbohydrate, fat, vitamins, minerals, and other nutrients. They are commercially available as powders, ready-to-feed liquids, and liquid concentrates.
[0003] Although many infant formulas provide a quality alternative to human milk, they still do not provide the same high level of nutrition as found in human milk. As such, much of the research effort into infant formulas over the past several years has been directed to better understanding the natural constituents of human milk, and then modifying infant formulas accordingly, or at least to the extent possible with currently available technology.
[0004] Arachidonic acid and docosahexaenoic acid, for example, have been identified in human milk and subsequently added to synthetic infant formulas. These fatty acids support brain and vision development in infants, and are now commonly found in commercially available formulas such as Similac® Advance® Infant Formula, Isomil® Advance® Infant formula, and Similac® Special Care ® Advance® infant formula, all of which are available from Ross Products Division, Abbott Laboratories, Columbus, Ohio, USA.
[0005] Lutein has also been identified in human milk. Although it is not currently added to infant formulas as an isolated ingredient, lutein can be found at low concentrations in infant formulas as an inherent ingredient in some of the natural oils commonly used make such formulas. Lutein is an antioxidant that also happens to concentrate within the retina of the eye. It is generally known that dietary lutein may provide individuals with eye health benefits, and it is speculated that such benefits may be extended to infants receiving lutein from either human milk or supplemented infant formula.
[0006] ft is now believed that a combination of lutein and docosahexaenoic acid may be particularly important in promoting retinal hearth and vision development in infants. Both materials are present in
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human milk and both are known to concentrate in the retina in otherwise healthy subjects. Docosahexaenoic acid (DHA), as a polyunsaturated fatty acid, is highly susceptible to damage by oxidation and degradation within the eye, while lutein is a known antioxidant It is believed that by adding lutein to infant formulas, not only will it concentrate within me retina, it may also reduce oxidative degradation of the retinal DHA and thus further promote retinal health and vision development in the infant
[0007] ft has now been found, however, that lutein concentrations in infant fonnula must be much higher than me lutein concentrations found in human milk in order to achieve the same plasma lutein concentrations found in breast fed infants due to a lower relative bioavailability of lutein from infant formula. Although infant formulas today typically contain less than about 20 mcg/hter of lutein, most of which comes inherently from added fats and oils, it has now been found mat such lutein concentrations must exceed about 50 meg/liter, preferably from about 100 meg/ liter to about 200 meg/liter, in order to duplicate plasma lutein concentrations found in exclusively in breast fed infants.
[0008] Consequently, it has also been found that infant formulas containing combinations of lutein and DHA, as described above, should now be formulated with higher ratios (lutein to DHA) than pre commonly found in human milk. These weight ratios of lutein (meg) to DHA (mg) should now range from about 1:2 to about 10:1.
SUMMARY OF THE INVENTION [0009] The present invention is directed to infant formulas comprising fat, protein, carbohydrate, vitamins, and minerals, including docosahexaenoic acid and at feast about 50 meg/liter of lutein, wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 1:2 to about 10:1. The present invention is also directed to methods of using the fonnulasto promote retinal health and vision development in infants, including reducing the risk of retinopathy of prematurity in infants, and protecting against the damaging effects of excessive natural or artificial light on the infants' eyes.
[0010] It has been found that infant formulas should be prepared with lutein concentrations of at least 50 meg/liter if they are to produce the same plasma lutein concentrations found in breast fed infants, even though human milk itself typically contains no more than about 30 meg/liter of lutein. It has also been found, consequently, that the weight ratio of lutein (meg) to DHA (mg) in the infant fonnula should range from about 1:2 to about 10:1. It is believed that the combination of lutein and docosahexaenoic acid are particularly useful in promoting retinal health and vision development in infants, provided that sufficient quantities of each are designed into the infant formula as described herein.
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BRIEF DESCRIPTION OF THE DRAWINGS
[0011] Fig. 1 is a graph of lutein intake (meg/day) and corresponding plasma lutein concentrations (mcg/dl) in infant groups fed human milk (HM) or infant formulas containing varied concentrations of lutein [CTRL with 14.6 meg Iutein/liter (no added lutein, all lutein inherent in ingredients); LI with 32.6 meg ratem/Iiter (approximately 18 meg/liter added lutein, remainder inherent), L2 with 52.6 meg lutein/liter (approximately 38 mcg/Iiter added lutein, remainder inherent).
[0012] Fig. 2 is a graph showing visual acuity as measured by sweep visual evoked potential (logMAR) in monkeys at 4, 8, and 12 weeks of age. The monkeys are fed infant formula with either DHA and added lutein (n=8) or DHA without added lutein (n=8) during a 12 week feeding period.
DETAILED DESCRIPTION OF THE INVENTION [0013] The infant formulas of the present invention comprise fat, protein, carbohydrate, minerals, and vitamins, and include a novel combination of lutein and docosahexaenoic acid. These and other essential elements or limitations of the infant formulas and corresponding methods of the present invention are described in detail hereinafter.
[0014] The term "infant" as used herein refers to individuals not more than about one year of age, and includes infents from 0 to about 4 months of age, infants from about 4 to about 8 months of age, infants from about 8 to about 12 months of age, low birth weight infants at less than 2,500 grams at birth, and preterm infants born at less man about 37 weeks gestational age, typically from about 26 weeks to about 34 weeks gestational age.
[0015J The term "infant formula" as used herein refers to a nutritional composition, free of egg phospholipids, which is designed for infants to contain sufficient protein, carbohydrate, fat, vitamins, and minerals to potentially serve as the sole source of nutrition when provided in sufficient quantity.
[0016] The term "ready-to-feed" as used herein, unless otherwise specified, refers to infant formulas in liquid form suitable for administration to an infant, including reconstituted powders, diluted concentrates, and manufactured liquids.
[0017] As used herein, all concentrations expressed as either "mcg/Iiter" or "mg/Iiter" refer to ingredient concentrations within the infant formulas of the present invention as calculated on a ready-to-feed or as fed basis, unless otherwise specified.
[0018] Allpercentages, 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
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and, therefore, do not include solvents or by-products that may be included in commercially available materials, unless otherwise specified.
[0019] All references to singular characteristics or limitations of the present invention 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.
[0020] 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.
[0021] The infant formulas of the present invention may also be substantially free of any optional or selected essential ingredient or feature described herein, provided that the remaining formula still contains all of the required ingredients or features as described herein. In this context, the term "substantially free" means mat the selected composition contains less than a functional amount of the optional ingredient, typically less than 0.1% fay weight, and also mcluding zero percent by weight of such optional or selected essential ingredient
[0022] The infant formulas and corresponding methods of the present invention can comprise, consist of or consist essentially of the essential elements and limitations of the invention described herein, as well as any additional or optional ingredients, components, or limitations described herein or otherwise useful in nutritional formula applications.
Lutein
[0023] The infant formulas of the present invention comprise lutein, concentrations of which must be at least about 50 mcg/titer of lutein. Any source of lutein is suitable for use herein provided that such a source is also known for or otherwise suitable for use in infant formulas and is compatible with the other selected ingredients in the formula, wherein the weight ratio of lutein (meg/liter) to docosahexaenoic acid (mg/liter) in the formulas ranges from about 1:2 to about 10:1.
[0024] Lutein concentrations in the infant formulas of the present invention range from about 50 to about 1150 meg /liter, including from about 75 to about 230 meg/liter, and also including from about 100 to about 200 meg/liter, as calculated on a ready-to-feed basis. All lutein concentrations and ratios referenced herein are calculated on a free lutein basis, unless o&erwise specified.
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[0U25J I"he amount of lutein in trie lnlant formulas must also be selected so that the weight ratio of lutein (meg) to docosahexaenoic acid (mg) ranges from about 1:2 to about 10:1, including from about 1.5:1 to about 9:1, also including from about 1.7:1 to about 5:1.
[0026] The term "lutein" as used herein, unless otherwise specified, refers to one or more of free lutein, lutein esters, lutein salts, or other lutein derivatives or related structures as described or otherwise suggested herein. Lutein or lutein sources suitable for use in the infant formulas of the present invention include free lutein as well as esters, salts or other derivatives or related structures thereof, including those that conform to the formula:

The above formula includes the general structure of lutein and related derivatives or structures. Free lutein, for example, corresponds to the formula wherein Rj and R2 are both hydrogen, and includes cis and trans isomers thereof as well as salts thereof, e.g^ sodium, potassium.
[0027J Lutein esters suitable for use in the infant formulas of the present invention include any lutein ester of the above formula wherein Rt and R2 are the same or different, and are nutritionally acceptable monovalent salts, hydrogen or an acyl residue of a carboxylic acid, provided that at least one of Rj or Rj is an acyl residue of a carboxylic acid. Suitable lutein esters include, as well, both cis and trans isomers. The Ri and R2 moieties are residues of a saturated or unsaturated Ci to C22 fatty carboxylic acids, non-limiting examples of which include formic, acetic, propionic, butyric, valeric, caproic, caprylic, capric, lauric, myristic, palmitic, stearic, and oleic acids.
[0028] Lutein for use herein includes any natural or synthetic source that is known for or is otherwise an acceptable source for use in oral nutritionals, including infant formulas. Lutein sources can be provided as individual ingredients or in any combination with other materials or sources, including sources such as multivitamin premixes, mixed carotenoid premises, pure lutein sources, and inherent lutein from other fat or oil components in the infant formula. The lutein concentrations and ratios as described herein are calculated based upon added and inherent lutein sources. The infant formulas of the present invention preferably comprise at least about 25%, more preferably from about 50% to about 95%, by weight of total lutein as added lutein, the remainder being inherent lutein that accompanies added fats and oils.
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(0029] Non-limiting examples of some suitable lutein sources for use herein include FIoraGLO® Crystalline Lutein, available from Kemin Foods, Des Moines, Iowa, USA; and Xangold ® Lutein Esters provided by Cognis, Cincinnati, Ohio, USA.
[0030] The infant formulas of the present invention include those preferred embodiments comprising a single source combination of free lutein and zeaxanthin, in a purified crystalline extract from the marigold flower (Tagetes erecta), wherein the free lutein represents from 85% to 95% by weight of tie combination and the zeaxanthin represents from about 5% to about 15% by weight of the combination. The preferred lutem-zeaxanfhin combination is available from Kemin Foods, Des Moines, Iowa, USA, under the FIoraGLO® brand.
Docosahexaenoic Acid (DHA)
[0031] The infant formulas of the present invention comprise docosabexaenoic acid, an organic carboxylic acid having a chain length of 22 carbons with 6 double bonds beginning with the third carbon from the methyl end (22:6 n-3). Any source of docosahexaenoic acid is suitable for use herein provided that such a source is also known for or otherwise suitable for use in infant formulas and is compatible with the other selected ingredients in the formula.
[0032] Docosahexaenoic acid concentrations in the infant formulas of the present invention must be selected so that the resulting weight ratio of lutein to docosahexaenoic acid falls within the range as defined herein. Such concentrations most typically range from about 36 to 360 mg/liter, including from about 50 to about 144 mg/liter, and also including from about 72 to about 130 mg/liter, as calculated on a ready-to-feed basis.
[0033] The docosahexaenoic acid may be added to the infant formula as free fatty acids or as compounds or materials that can otherwise provide a source of such free fatty acids upon or following administration to the infant, including non-egg phospholipids and glyceride esters (mono-, di-, tri-) of docosahexaenoic acids. Polyunsaturated fatty acids and sources thereof are described in U.S. Patent 6,080,787 (Carlson, et al.) and U.S. Patent 6,495,599 (Auestad, et al.), which descriptions are incorporated by reference herein. Some non-limiting examples of suitable docosahexaenoic acid sources include fish oils, algal oils, other single cell oils, and combinations thereof
[0034] The infant formulas of the present invention may further comprise, in addition to the docosahexaenoic acid as described herein, other long chain polyunsaturated fatty acids such as arachidonic acid (20:4 n-6), eicosapentaenoic acid or EPA (20:5 n-3), linoleic acid (18:2 n-6), y-
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linolenic acid or GLA (18:3 n-6), a-linolenic acid (18:3 n-3), dftomo-v-linolenic or DHGLA (20:3 n-6), a-linolenic (18:3 n-3), stearidonic acid (18:4 n-3), and combinations thereof Snch optional long chain polyunsaturated fatty acids may likewise be formulated into the infent formula as free fatty acids or as compounds or materials that can otherwise provide a source of such free fatty acids upon or following administration to the infant, including non-egg phospholipids and glyceride esters (mono-, di-, tri-) of docosahexaenoic acids.
Other Nutrients
[0035] The infant formulas of the present invention comprise fat, protein, carbohydrate, minerals, and
vitamins, all of which are selected in kind and amount to meet the dietary needs of the intended infant
population.
[0G36J Many different sources and types of carbohydrates, fats, proteins, minerals and vitamins are known and can be used in the infant formulas of the present invention, provided mat such nutrients are compatible with the added ingredients in the selected formulation and are otherwise suitable for use in an infant formula.
[0037] Carbohydrates suitable for use in the infant formulas of the present invention may be simple or complex, lactose-containing or lactose-free, or combinations thereof, non-limiting examples of which include hydrolyzed, intact, naturally and/or chemically modified cornstarch, maltodextrin, glucose polymers, sucrose, corn syrup, corn syrup solids, rice or potato derived carbohydrate, glucose, fructose, lactose, high fructose corn syrup and indigestible oligosaccharides such as fractooligosaccharides (FOS), galactooligosaccharides (GOS), and combinations thereof.
[0038] Proteins suitable for use in the infant formulas of the present invention include hydrolyzed, partially hydrolyzed, and non-hydrofyzed or intact proteins or protein sources, and can 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 tiiereof.
[0039] Proteins for use herein can also include, or be entirely or partially replaced by, free amino acids known for or otherwise suitable for use in infant formulas, non-limiting examples of which include alanine, arginine, asparagine, carnitine, aspartic acid, cystine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, taurine, threonine, tryptophan, taurine, tyrosine, valine, and combinations thereof. These amino acids are most typically used in their L-forms, although the corresponding D-isomers may also be used when nutritionally equivalent Racemic or isomeric mixtures may also be used.
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[0040] Fats suitable for use in theinfantformulas of the present invention include coconut oil, soy oil, corn oil, olive oil, safflower oil high oleic safSower oil, algal oil, MCT oil (medium chain triglycerides), sunflower oil, high oleic sunflower oil, palm and palm kernel oils, palm olein, canola oil, marine oils, cottonseed oils, and combinations thereof
10041] Vitamins and similar other ingredients suitable for use in the infant formulas of the present invention include vitamin A, vitamins D, vitamin E, vitamin K, thiamine, riboflavin, pyridoxine,
derivatives thereof, and combinations thereof.
10042] Minerals suitable for use in the infant formulas of the present invention include calcium, phosphorus, magnesium, iron, zinC manganese, copper, chromium, iodine, sodium, potassium, chloride, and combinations thereof.
[0043] The infant formulas preferably comprise nutrients in accordance with the relevant infant formula guidelines for the targeted consumer or user population, an example of which would be the Infant Formula Act, 21 U.S.C. Section 350(a).
[0044] The infant formulas of the present invention also include those enibodiments containing the carbohydrate, fat, and protein concentrations described in the following table.

Nutrient
Carbohydrate
Fat
Protein
Table 1: Infant Formula Nntriens1,.
Range gm/lOOkcal gm/titer2
1st embodiment 8-16 54-108
2nd embodiment 9-13 61-88
la embodiment 3-8 20-54
2ntl embodiment 4-6.6 27-45
1st embodiment 1-3.5 7-24
2nd embodiment' 1.5-3.4 10-23
1. All numerical values may be modified by the term "about"
2. From ready-to-feed liquid, reconstituted powder, or dilated concentrate
[0045] The infant formulas of the present invention include those embodiments mat comprise per 100 Kcal of formula one or more of the following: vitamin A (from about 250 to about 750IU), vitamin D (from about 40 to about 100 IU), vitamin K(greater than about 4 meg), vitamin E (at least about 0.3 IU), vitamin C (at least about 8 mg), thiamine (at least about 8 g), vitamin B12 (at least about 0.15 g), niacin (at least about 250 g), folic acid (at least about 4 g), pantothenicacid (at least about 300 9), biotin(atleast about 1.5g)choline(atleast about 7mg) and inositol(atleast about 4mg),
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[0046] The infant formulas of the present invention also include those embodiments that comprise per 100 kcal of formula one or more of the following: calcium (at least about 50 mg), phosphorus (at least about 25 mg), magnesium (at least about 6 mg), iron (at least about 0.15 mg), iodine (at least about 5 g), zinc (at least about 0.5 mg), copper (at least about 60 g), manganese (at least about 5 g), sodium (from about 20 to about 60 mg), potassium (from about 80 to about 200 mg), and chloride (from about 55 to about 150 mg).
Optional Ingredients
[0047] The infant formulas of the present invention may further comprise other optional ingredients that may modify the physical, chemical, aesthetic or processing characteristics of the compositions or serve as pharmaceutical or additional nutritional components when used in the targeted infant population. Many such optional ingredients are known or are otherwise suitable for use in nutritional products and may also be used in hie infant formulas of the present invention, provided that such optional materials are compatible with the essential materials described herein and are otherwise
suitable for use in an infant formula.
[0048] Non-limiting examples of such optional ingredients include preservatives, additional anti¬oxidants, emulsifying agents, buffers, colorants, flavors, nucleotides and nucleosides, probiotics, prebiotics, lactoferrin and related derivatives, thickening agents and stabilizers, and so forth.
Prodnct Form
[0049] The infant formulas of the present invention may be prepared as any prodnct form suitable for use in infants, including reconstitutable powders, ready-4o-feed liquids, and dilutable liquid concentrates, which product forms are all well known in the nutrition and infant formula arts.
[0050] The infant formulas of the present invention may have any caloric density suitable for the intended infant population, or provide such a density upon reconstitntion of a powder embodiment or upon dilution of a liquid concentrate embodiment Most common caloric densities for the infant formulas of the present invention are generally at least about 18 kcal/fl oz (609 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 22-30 kcal/fl oz, most typically from about 22-24 kcal/fl oz, formulas are more commonly used in pre-term of 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. Higher caloric feedings may be used with pre¬term infants of low birth weight; such feedings are typically from about 27 kcal/fl oz (90-95 kcal/liter) to about 30 kcal/fl oz (1000-1015 kcal/liter).
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[0051] For powder embodiments of the present invention, such powders are typically in the form of flowable or substantially flowable particulate compositions, or at least particulate compositions mat can be easily scooped and measured with a spoon or similar other device, wherein the compositions can easily be reconstituted by the intended user with a suitable aqueous fluid, typically water5 to form a liquid nutritional formula 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. These powder embodiments include spray dried, agglomerated, dry mixed or other known or otherwise effective particulate form. The quantity of a nutritional powder required to produce a volume suitable for one serving can vary.
[0052] The infant formulas of the present invention may be packaged and sealed in single or multi-use containers, and then stored under ambient conditions for up to about 36 months or longer, more typically from about 12 to about 24 months. For multi-use containers, these packages can be opened and then covered for repeated use by the ultimate user, provided that the covered package is men stored under ambient conditions (e.g., avoid extreme temperatures) and the contents used within about one month or so.
Retinal Health and Vision Development [0053] The present invention is also directed to methods of administering the formulas to promote retinal health and vision development in infants. In this particular method, the infant formulas are administered to term or preterm infants as a sole source, primary source, or supplemental source of nutrition, wherein the formulas comprise fat, protein, carbohydrate, vitamins, and minerals, including docosahexaenoic acid and at least about 50 meg/liter of lutein, wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 112 to about 10:1. Such a method may be applied to any formula embodiments described or otherwise suggested herein.
[0054] This particular method should therefore provide the infant with an effective amount of lutein to provide the stated benefits, including from about 7 to about 300 meg/kg/day, including from about 14 to about 220 meg/kg/day, and also including from about 22 to about 150 meg/kg/day (of lutein per kg of body weight of the infant), wherein the weight ratio of lutein to docosahexaenoic acid is maintained within the ratios described herein.
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[0055] Eye and vision development occurs at a rapid rate during the first year of life. At birth, infants can only see high-contrast objects at perhaps 25-30 cm away. During the next 6 months, the infants retina develops enough to see and discern small details. And as an infant* s vision develops, most of which will occur during the first year, the infant becomes better able to learn through visual stimulation now made possible with a newly developed sight For infants, this visual learning then plays a key role in brain and cognitive development, especially during the first 2-3 years of life.
(0056) By promoting retinal health and vision development in infants, the infant formulas of the present invention may also help children develop their ability to visually learn as soon as possible, and to potentially accelerate brain and cognitive development associated with early visual stimulation through the developing retina of the eye. The infant formulas of the present invention are therefore useful in promoting vision development in infants, and consequently are useful in promoting secondary benefits such as associated cognitive and brain development through early visual stimulation.
[0057] This particular method of the present invention may be particularly useful in preterm infants to help accelerate the development of normal vision, to thus reduce the time needed to cateh-up with development milestones set by their term infant counterparts.
Retinopathy of Prematurity [0058] The infantformulas of the present invention are especially useful when administered to preterm infants to reduce the risk of retinopathy of prematurity. In accordance with such a method, the formulas are administered as a sole source, primary source, or supplemental source of nutrition, wherein the formulas comprise fat, protein, carbohydrate, vftarnins, and minerals, mcluding docosahexaenoic acid and at least about 50 meg/liter of lutein, wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 1:2 to about 10:1. Such a method may be applied to any formula embodiments described or otherwise suggested herein.
[0059] Retinopathy of prematurity is a condition that often affects preterm infants and is most commonly characterized by abnormal development of retinal vessels in the eye possibly as a result of oxidative stress secondary to high oxygen tension. This affliction can occur to varying degrees, from slight vessel involvement with minimal or no impact on vision, to partial or complete retinal detachment leading to blindness. Historically, therapy for appropriate cases included laser treatment as well as cryotherapy.
[0060] This particular method should therefore provide the infant with an effective amount of lutein to provide the stated benefits, mcluding from about 7 to about 300 meg/kg/day, including
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from about 14 to about 220 mcg/kg/day, and also including from about 22 to about 150 mcg/kg/day (of lutein per kg of body weight of the infant), wherein the weight ratio of lutein to docosahexaenoic acid is maintained within the ratios described herein.
Method of Manufacture

[0061]
The infant formulas of the present invention may be prepared by any known or otherwise effective technique suitable for making and formulating an infant formula or similar other formula, variatioas of which may depend upon variables such as the selected product form, ingredient combuntion, packaging and container selection, and so forth, for the desired infant formula. Such techniques and variations for any given formula are easily determined and applied by one of ordinary skill in the infant nutrition formulation or manufacturing arts.
[0062] The infant formulas of the present invention, includmg the exemplified formulas described hereinafter, can therefore be prepared by any of a variety of known or otherwise effective formulation or manufacturing methods. These methods most typically involve the initial formation of an aqueous slurry containing carbohydrates, proteins, lipids, stabilizers or other formulation aids, vitamins, minerals, or combinations thereof. The slurry is emulsified, pasteurized, homogenized, and cooled. Various other solutions, mixtures, or other materials may be added to the resulting emulsion before, during, or after further processing. 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 reconsiitatable powder, e.g., spray dried, dry mixed, agglomerated.
[0063] Other suitable methods for making nutritional formulas are described, for example, in U.S. Patent 6,365,218 (BorscheL et al.), U.S Patent 6,589,576 (BorscheL et aL), U.S. Patent 6,306,908 (Carlson, et al.), U.S. Patent Application 20030118703 Al (Nguyen, et aL), which descriptions are incorporated herein by reference.

EXPERIMENT
10064]
The purpose of this experiment is to evaluate changes in visual acuity in animals fed infant formulas comprising either DHA or DHA with added lutein. Sixteen monkeys are fed one of two defined infant formulas during their first 12 weeks of life. One is a control formula - Similac® Advance ® Infant formula, available from Abbott Laboratories, Columbus Ohio, and the other is an experimental formula including Similac® Advance® Infant Formula as a base, but with added carotenoids comprising lutein. The formulas include the following:
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Nutrient Control formula Experimental formula
DHA 50mg/L 60mg/L
Lutein 0 mcg/L added (18 mcg/L inherent) 117 mcg/L
Zeaxan hin 0 meg/ L added (4 mcg/L inherent) 36 mcg/L
Zeaxan hin/lutein 22% 31%
[0065] The base formula (Similac® Advance® Infant formula) contains water, nonfat milk, lactose, high oleic safflower oiL soy oil, coconut oil, whey protein concentrate; C. cohnii oil, M. alpina oil, potassium citrate, calcium carbonate, ascorbic acid, mono- and digh/cerides, soy lecithin, carrageenan, potassium chloride, magnesium chloride, sodium chloride, ferrous sulfate, choline chloride, choline bhartrate, taurine, m-inositoL d-alpha-tocopheryl acetate, LKarnitme, zinc sulfate, niacinamide, calcium pantothenate, riboflavin, vitamin A pahnitate, cupric sulfate, thiamine chloride hydroclloride, pyridoxine hydrochloride, beta-carotene, folic acid, manganese sulfate, phylloquinone, biotin, sodium selenate, vitamin D3, cyanocobalamin and nucleotides (adenosine ^-monophosphate,

cytidine

y-monophosphate, disodium guanosine 5'-nionophosphate, disodium uridine 5'-

monophosphate).
[0066J The monkeys are randomized to receive either the experimental (n=8) or control (n=8) formulas from birth to 12 weeks of life. The pntmak do not receive any milk from their mothers. Infants aid mothers are separated at birth. Ehiring the study, the monkeys are exposed to light having the intensity and spectral characteristics of sunlight for 12 hours per day to simulate the light-induced oxidative stress potentially experienced by infants. During the study, the monkeys are evaluated for several parameters, including plasma lutein concentrations and sweep visual evoked potential (YEP).
Plasma' Lutein
[0067] Plasma concentrations of lutein, lycopene, and beta-carotene are not significantly different between the monkeys fed the control and experimental formulas at birth (0 weeks of age). Plasma lutein concentrations are significantly higher in monkeys fed the experimental formula man monkeys fed the control formula at 4 (p
"Visual

Acuity

[0068] The monkeys are assessed for changes in visual acuitv at 4,8, and 12 weeks of life. Visual acuityis measured by sweep visual evoked potential (VEP), a method well known in the art for measuring visual evoked potential in infants. Visual acuity is measured by determining the smallest
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spatial frequency high contrast grating that evokes a measurable response from the visual cortex. The YEP from the primary cortical visual area is recorded using small silver disk EEG electrodes placed on the scalp with water-soluble electrode paste. The infant is held in an experimenter's lap while h gazes at a video monitor displaying phase-reversing black and white gratings. When necessary, the infant's attention will be drawn to the center of the screen with small dangling toys. During each "sweep', the spatial frequency of the grating will be decreased stepwise from above to below the subject's acuity threshold during a recording period of several seconds. The amplitude of the second harmonic of the VEP response, which reflects the response linked to the stimulus reversal rate, will be plotted as a function of spatial frequency to define the subject's acuity threshold (Neuringer M, Jeffrey BG: Visual development neural basis and new assessment methods. JPediatr 2003;143:S87-S95).
[0069J VEP scores from the study are summarized in the Fig. I graph. Lower VEP (logMAR) scores are indicative of better visual acuity. Although VEP scores decreased (Le., visual acuity improved) for all monkeys during the 12 week testing period, as expected, the VEP scores at 8 weeks were surprisingly lower in the experimental group (added lutein+DHA formula) than in the control group (DHA without added lutein formula) (4 weeks, p=0.412, etc.)
[0070] The data suggest accelerated development in infant monkeys fed the experimental formula at 8 weeks of life — specifically in visual acuity as measured by VEP values. To extrapolate the data to human infants, the eye development in monkeys at ages of 4,8, and 12 weeks corresponds to the eye develop potent in human infants at 4, 8, and 12 months, respectively. The data therefore suggests that evenin a human infant, the experimental formula would improve visual acuity at between about 4 and about 12 months of life.

EXAMPLES
[0071]
The following examples represent specific embodiments within the scope of the present invention, each of which is given solely for the purpose of illustration and is not to be construed as limitations of the present invention, as many variations thereof are possible without departing from the spirit and scope of the invention. All exemplified amounts are weight percentages based upon the total weight of the composition, unless otherwise specified.
Examples 1.1-13
[0072] The following are examples of milk-based, ready-to-feed, infant formulas of the present invention, including a method of using and making the formulas. The formula ingredients for each batch are listed in the table below.
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Ingredient Example 1.1 Example \2 Example 1.3

Amount per 454 kg Quantity per 454 kg Quantity per 454 kg
Water QS OS OS
Lactose 27 kg 27 kg 27 kg
ARA-containing oil (40% ARA) 0.167 kg 0.167 kg 0.167 kg
BHA-containing oil (40% DHA) 0.063 kg 0.095 kg 0.145 kg
Non-fit dry milk 1133 kg 1133 kg 1133 kg
High oleic saffiower oil 6.5 kg 6.5 kg 6 J kg
Mono- and di-giycerides 0.162 kg 0.162 kg 0.162 kg
Soybean oil 5kg 5kg 5kg
Whey protein 2.8 kg 2.8 kg 2-8 kg
Calchim carbonate 0211kg 0211kg 0.211kg
Coconut oil 4.6 kg 4.6 kg 4.6 kg
Citric acid 0.014 kg 0.014 kg 0.014 kg
Potassium citrate 0245 kg 0.245 kg 0.245 kg
Ascorbic acid 178 g Il78g 178 g
Lecithin 162 g | 162g 162 g
Magnesium chloride 25 g 25g 25g
Potassium chloride 88 g 88g 88g
jFenons sulfate 26g 26g 26g
iCanageenan 136 g 136 g 136 g
Choline chloride 25g t25g 25g
Nucleotide and choline premix 3 133 g 133 g 133 g
[Riboflavin lg lg lg
L-Camitine 1-5 g 1-5 g 1-5 g
Potassium hydroxide 998 g 998 g 998 g
Lutein solution (5% active) 4 0.882 g 1323 g 1.764 g
Water soluble vitamin premix' 65- 65 g 65 g
Vitamin ADEK premix 2 21 s 21 g 21 g
VitaminA 0.4 g 0.4 g 0.4 g
[Beta-carotene soln (30% active) 0.0485 g 0.0485 g 0.0485 g

[Total Lutein (meg/liter) 100 150 200
Total DHA (mg/Iiter) 50 75 115
(Ratio - Lutein (meg) : DHA (mg) 2 2 1.74
taurine, 14.4 g inositol, 6.7 g zinc sulfite, 42 g niacinamide, 2.6 g calcium pantothenate, 23 g ferrous sulfite, 0.8 g enpric sulfite, 0.6 g thiamine, 03 g riboflavin, 0.26 g pyridoxins, 0.1 g folic acid, 0.07 g manganese sulfate, 0.03 g biotin, 0.025 g sodium selenate, 0.002 g cyanocobalamin
premix contains (per 21 g) 4.0 g alpha-tocopheiol acetate, 0& g vitamin A palmitale, 0.05 g pnylloquinone, 0.006 g vitamin D3
premix contains (per 133 g): 23 g choline bitartrate, 15 g 5'-CMP, 11 g 5'-C»ff, 10 g 5*-UMP,6g5'-AMP FJoraGLO® Crystalline Lutein, Kemin Foods, Des Moines, Icrwa, USA
3.
4.
1 premix contains (per 65 g) 19.8 g
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[0073] The exemplified formulas may be prepared by making at least three separate slurries that are later blended together, heat treated, standardized, packaged and sterilized. Initially, a carbohydrate-mineral slurry is prepared by dissolving lactose in water at 65-71 °C, followed by the addition of magnesium chloride, potassium citrate, potassium chloride, choline chloride, and citric acid. The resulting slurry is held with agitation at 55-65°C for not longer than eight hours until it is later blended with the other prepared slurries.
[0074j| A protein-fat slurry is prepared by combining high oleic safflower oil, soybean oil, and coconut oil at 55-60°C, followed by fee addition of vitamin ADEK premix, mono- and diglycerides, lecithin, carrageenan, vitamin A, ARA oil, and DHA ofl. Whey protein and calcium carbonate are then added. The resulting protein-oil slurry is held under moderate agitation at 40-43°C for no longer than two hours until it is later blended with the other formed slurries.
[0075] The carbohydrate-mineral slurry is then combined with water and non-fat dry milk and agitated for 10 minutes. The protein-oil slurry is then added and the resulting mixture agitated for at least 10 minutes. Lutein and beta-carotene are then added to the blend and agitated for at least 15 minutes. The pH of the resulting blend is adjusted to 6.6&-6.75 with IN potassium hydroxide.
[0076] After waiting for a period of not less than one minute nor greater than two hours, the resulting blend is heated to 71-82°C and dearated under vacuum, emulsified through a single stage homogenizer at 900-1100 psig, and then heated to 99-110°C, and then heated again to 146°C for about 5 seconds. The heated blend is passed through a flash cooler to reduce the temperature to 99-110°C and then through a plate cooler to further reduce the temperature to 71-76°C. The cooled blend is then homogenized at 3900-4100/ 400-600 psig, and men held at 74-80°C for 16 seconds, and then cooled to 1-7°C. Samples are taken for microbiological and analytical testing. The mixture is held under agitation.
[0077] A water-soluble vitamin (WSV) solution and an ascorbic acid solution are prepared separately and added to the processed blended slurry. The vitamin solution is prepared by adding the following ingredients to 9.4 kg of water with agitation: potassium citrate, ferrous sulfate, WSV premix, L-canirrine, riboflavin, and the nucleotide-choline premix. The ascorbic acid solution is prepared by adding potassium citrate and ascorbic acid to a sufficient amount of water to dissolve the ingredients. The vitamin and ascorbic acid solutions are then added to the blend, and the pH of the blend adjusted to 7-10 with 45% potassium hydroxide solution.
[0078] Based on the analytical results of the quality control tests, an appropriate amount of water is added to the batch with agitation to achieve the desired total solids. The product pH may be adjusted
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to achieve optimal product stability. The completed product is then placed in suitable containers and subjected to terminal sterilization.
[0079] The resulting formulas are fed to infants as a sole source of nutrition during the first 6 to 12 months of life to provide each infant with 7-300 ug/kg/day of lutein. The formulas provide improved retinal health and vision development as described herein.
Examples 2.1-23
[0080] The following are examples of soy-based, powder, infant formulas of the present invention, including a method of using and making the formulas. The formula ingredients for each batch are listed in the table below.

INGREDIENT Example 2.1 Example 22 Example 23

Amount per 454kg Amount per 454 kg Amount per
High oleic safHower oil 52.1kg 52.1kg 52.1kg
Coconut oil 35.2 kg 352 kg 352 kg
Soy oil __, 38.1 kg 38.1kg 38.1kg
ARA-conteining oil (40% ARA) 13kg 13kg 13kg
DBA-containing oil (40% DHA) 0381kg 0.762 kg 0.876 kg
Ofl soluble vitamin premix 0.173kg 0.173kg 0.173kg
P-carotene solution (30% active) 0.0004 kg 0.0004 kg 0.0004 kg
Ascorbyl palmitate 0.162 kg 0.162 kg 0.162 kg
Soy protein isolate 66.1kg 66.1kg 66.1kg
Cornsyrop 236.0kg |236.0kg 236.0kg
Calcium phosphate (di and tribasic) 8.0 kg 8.0 kg 8.0 kg
Ferrous sulfate 0.138kg 0.138kg 0.138kg
Lutein solution (5.0% active) ' 7-06 g 10.5903 g 14200 g
Water soluble vitamin premix trace minerals/taurine 0.65 kg 0.65 kg 0.651s
Choline chloride 0.23 kg 023 kg 023 kg
Potassium iodide 0.0005 kg 0.0005 kg 0.0005 kg
Methionine 0.722jg 0.722 kg 0.722 kg
Ascorbic acid 0.72 kg 0.72 kg 0.72 kg
Potassium hydroxide (45% solution) 12 kg 1.2 kg 12 kg
Potassium chloride 0.87 kg 0.87 kg 0.87 kg
Magnesium Chloride 0.4 kg 0.4 kg 0.4 kg
Carnitine 0.05 kg 0.05 kg 0.05 kg

Total Lutein (mcg/Iiter as fed) 100 150 J200
Total DHA (mg/Iiter as fed) 50 100 115
Ratio - Lutein (meg): DHA (mg) as fed 2 2 I 1.74
1. FloraGLO® Crystalline Lutein, Kemin Foods, Des Moines, Iowa, USA
[0081] The first step in the preparation of the exemplified powder is the preparation of the oil blend. Soy oil, coconut oil and high oleic safHower oil are combined in a suitable container or tank at 60-65°C with agitation. Ascorbyl palmitate and mixed tocopherols are added to the tank, followed by the oil soluble vitamin premix, all with agitation. Beta-carotene (BASF, Mount Olive, New Jersey), and lutein (Kemin, Des Moines, Iowa) are added to the oil blend and agitated until well dispersed. Soy
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protein isolate and methionine are then added to the oil blend, and the resulting mixture agitated and held at 54.0-60°C until used later during the manufacturing process.
10082] The carbohydrate-mineral slurry is then prepared. Potassium chloride, sodium chloride, magnesium chloride, and potassium iodide are added to water 60-65cC, followed by di- and tri-caicium phosphates, all with agitation. Corn syrup is then added with agitation, and the slurry held at 54-6Q°C until used later during the manufacturing process.
[0083] The carbohydrate-mineral slurry is added to the oil blend. Additional water is added as necessary. The ARA and DHA oils are added to the blend. The pH of the resulting mixture is adjusted to 6.75-6.85 using KOH solution. The adjusted mixture is then held at 54-60C under agitation for at least 15 minutes.
{0084] The resulting mixture is then heated to 74-79C and dearated under vacuum, emulsified through a single stage homogenizer at 0-2.76 Mpa, passed through a two-stage homogenizer at 62.-7.6 MPa and 2.1-3.4 MPa. The homogenized mixture is held at 73-79C for 16 seconds and then cooled to 1-7C. Samples are taken for microbiological and analytical testing. The mixture is held under agitation.
[0085] A calcium carbonate solution may be prepared for use in adjusting the calcium level of the mixture if outside of specification.
[0086] A vitamin stock solution containing a water soluble vitamin premix with trace minerals and taurine is prepared. Potassium citrate and ferrous sulfite are added to water at 37-66°C. The vitamin premix s then added and the mixture agitated. The choline chloride and carnitine are added and then the required amount of this vitamin mixture is added to the batch.
[0087] An ascorbic acid solution is prepared and added slowly to the batch with agitation for at least 10 minutes. The batch is then preheated to 74-79°C. The batch is then held for 5 seconds at 107-111°C using direct steam injection. The batch is then cooled to 71-82°C before being pumped to a spray dryer and dried to a flowable powder. The batch is thai packaged in suitable containers and sealed under a headspace of less than 2.0% oxygen.
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10088] The exemplified powders are reconstituted with water to a caloric density of 676 kcal/lher. The resulting liquid formulas are fed to infants as a sole source of nutrition during the first 6 to 12 months of life to provide 7-300 jig/kg/day of lutein. The formula provides improved retinal health and vision development as described herein.
Examples 3.1-33
[0089] The following are examples of milk-based, powder, infant formulas of the present invention, including a method of using and making the fonnula. The formula ingredients for each batch are listed in the table below.
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Ingredient Name Example 3.1 Example 3.2 Example 33

Amount per 454kg Amount per 454kg Amount per 454kg
Soy oil 35.8 kg 35.8 kg 35.8 kg
Coconut oil 23.8 kg 23.8 kg 23.8 kg
MCT oil (medium chain triglyceride) 32.1kg 32.1kg . 32.1 kg
High Oleic Safflower oil 34.6 kg 34.6 kg 34.6 kg
Ascorbyl pahnitate 0.157 kg 0.157 kg 0.157 kg
Vitamin A pahnitate 0.002 kg 0.002 kg 0.002 kg
1 Vitamin ADEK premix ' 0.192 kg 0.192 kg 0.192 kg
Mixed tocopherols 0.075 kg 0.075 kg 0.075 kg
Lutein solution (20 5% active) 4 7.060 g 10.590 g 13.714.200g


Whey protein concentrate 32.7 kg 32.7 kg 32.7 kg
Calcium carbonate l-2kg L2kg 1.2 kg
Lactose 54.5 kg 54.5 kg 54 3 kg
Corn syrup solids 117.1kg j 117.1kg 117.1 kg
Magnesium chloride 0.724 kg 0.724 kg 0.724 kg
Potassium citrate 2.8 kg 2.8 kg 2.8 kg
Sodium chloride 039 kg 039 kg 039 kg
Sodium citrate 0.001 kg 0.001kg 0.001kg
Non-fat dried milk 116.9 kg 116.9 kg 116.9 kg
Calcium phosphate tribasic 1.8 kg 1.8 kg 18 kg
ARA-contaMng oil (40% ARA) 13 kg 13 kg 13 kg
DHA^XHitaining oil (40% DHA) 0.43 kg 0.65 kg 1.00 kg
Ascorbic acid 129 kg |L29 kg 1.29 kg
Potassium hydroxide IN solution 9.8 kg 9.8 kg 9.8 kg
Ferrous sulfate 0.168 kg [0.168 kg 0.168 kg
Carnitine 0.136 kg 0.136 kg 0.136 kg
Choline chloride 0.182 kg 0.182 kg 0.182 kg
Vh. and trace mineral premix 2 0.825 kg 0.825 kg 0.825 kg
Inositol 0.734 kg 0.734 kg 0.734 kg
Nucleotide, choline hitartrate premix 3 1-1 kR 1-1 kg 1-lkg

Total Lutein (meg/liter as fed) 100 J150 200
Total DHA (mg/liter as fed) 50 75 115
Ratio - Lutein (meg) : DHA (mg) as fed 2 P 1.74
1. premix provides 71 gm d-alpha-tocopheryl acetate, 7.29 gm Vitamin A pahnitate, 0.422 gm
phyOoquinone, and 0.051 gm Vitamin D3 to the product
2. premix provides 252 gm taurine, 183 gm inositol, 843 gm zinc sulfate, 53.8 gm niacinamide, 32.6 gm calcium pantothenate, 29 gm ferrous sulfate, 10.1 gm cupric sulfate, 8.4 gm thiamine, 3.7 gm riboflavin, 3.4 gm pyridoxine (HO), 1.1 gm folic acid, 1.0 gm manganese sulfate, 03 gm biotin, 0.2 gm sodium selenate, and 0.03 gm cyanocobalamin to the product.
3. premix provides 188 gm choline bftartrate, 118 gm cytidine 5'-monophosphate, 92 gm disodium guanosine 5'-monophosphate, 80 gm disodium uridine 5'-monophospfeate, and 51 gm adenosine 5'-monophosphate to the product
4. FloraGLO® Crystalline Lutein, Kemin Foods, Des Moines, Iowa, USA
[0090] This powder formula is manuractured by preparing at least two slurries that are later blended together, heat treated, standardized, spray dried and packaged. Initially, a carbohydrate-mineral slurry is prepared (45-50% solids) by dissolving lactose in water at 66-76°C. Com syrup solids are then added and allowed to dissolve, followed by the addition of magnesium chloride, potassium citrate,
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sodium chloride, choline chloride, and sodium citrate, all with agitation. The resulting carbohydrate-minral slurry is held at 54-60°C under agitation until used later during the manufacturing process.
[0091] A protein-fat slurry is prepared by combining high oleic saffiower oil, coconut oil, soy oiL and MCT oil at 40.5-49°C, followed by ascorbyl palmitate, mixed tocopherols, vitamin A palmitate, and the vitamin ADEK premix, all with agitation. Lutein (Kemin, Des Moines, Iowa), is then added with agitation. Whey protein concentrate is then added to the slurry, followed by calcium carbonate and calcium phosphate tribasic, all with agitation. The completed protein-fat slurry is held under moderate agitation at 54-60°C for no longer than twelve hours until it is blended with the other prepared slurries.
[0092] The carbohydrate-mineral slurry is transferred to a tank in which a sufficient amount of water is added to create a final blend slurry of approximately 50% solids. Non-fat dry milk is then added to the blend and allowed to solubilize. The protein-fat slurry is then added and the entire blend slurry is allowed to agjtate for at least 15 minutes. The resulting blend is maintained at 60-65°C. The blend pH is adjusted to 6.7 - 6.9 with IN KOH.
[0093] After waiting for a period of not less than one minute nor greater than two hours, the resulting blend is heated to 71-79°C, emulsified at 2.75-4.1 Mpa, and then heated to 115-127°C for about 5 seconds using direct steam injection. The heated emulsion is then flash cooled to 87-99°C, and homogenized at 9.7-11.0 / 2.75-4.1 MPa. The homogenized slurry is then cooled to 1.6-7.2°C. Samples are taken for microbiological and analytical testing. The rnixmre is held under agitation.
[0094] A vitamin-trace mineral solution is prepared by adding the following ingredients to the required amount of water, under agitation: potassium citrate, ferrous sulfate, carnitine, vitamin and the trace rnineral premix, inositol, and nucleotide and choline bitartrare premix. The vitamin-trace mineral solution is then added to the homogenized slurry under agitation.
[0095] An ascorbic acid solution is prepared by adding potassium citrate and ascorbic acid to water with agitation, and then adding the aqueous mixture to the homogenized slurry under agitation.
[0096] The product is preheated to 65.5-77°C. The product is then held at 82-90.5°C for 5 seconds before being flash cooled to 71 — 82°C and pumped to the spray dryer. The product is spray dried to produce a desired free-flowing powder. The resulting powder is packaged under nitrogen to maximize product stability and flavor.
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[0097] The exemplified powders are reconstituted with water to a caloric density of 676 kcal/liter. The resulting liquid formulas are fed to infants as a sole source of nutrition during the first 6 to 12 months of life to provide 7-300 µg/kg/day of lutein. The formula provides improved retinal health and vision development as described herein.
Examples 4.1-43
10098} The following are examples of concentrated human milk fortifier liquids of the present
invention, including a method of using and making the formula. The formula ingredients for each
batch are listed in the following table.
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Ingredient Name Example 4.1 Example 42. Example 43

Gm per kg Cm per kg Gm per kg
Sucrose 125.5 125.5 125.5
Milk protein isolate 64.6 64.6 64.6
ICocomrtoil 30.2 302 302
Whey protein concentrate 24.4 24.4 24.4
MCTOii 21.9 21.9 21.9
Soy Oil 21.9 21.9 21.9
Tricalcium phosphate 14.4 14.4 14.4
Potassium chloride 5.18 5.18 5.18
Calcium carbonate 3.44 3.44 3.44
Magnesium phosphate 3.05 3.05 3.05
Potassium citrate 132 132 132
DBA (docosahexaenoic acid) oil 02 02 02
Soy lecithin 0.756 i 0.756 0.756
ARA (arachidonic acid) oil 0.729 0.729 0.729
Dipotasshnn phosphate 0.596 0.596 0.596
Monopotasshnn phosphate 0.466 0.466 0.466
VhaminE 0357 0357 0357
Sodium chloride _j 0.170 0.170 0.170
KOH 5% solution Q.S. Q.S. Q.S.
Lutein (from 20% solution) ' 0.00018 0.00064 0.00091
m-Inositol 0.0698 0.0698 0.0698
Ascorbic acid 0.913 0.913 0.913
Taurine 0.0663 0.0663 0.0663
Niacinamide 0.0582 0.0582 0.0582
VhaminA 0.0494 0.0494 0.0494
Zinc sulfate 0.0461 0.0461 0.0461
Calcium pantothenate 0.0286 0.0286 0.0286
Ferrous sulfate 0.0136 0.0136 0.0136
Cupric sulfate 0.00836 0.00836 0.00836
Riboflavin 0.00763 0.00763 0.00763
Thiamine chloride HCL 0.00507 0.00507 0.00507
Pyridoxine HCL 0.00459 0.00459 0.00459
Folic add 0.000778 0.000778 0.000778
Manganese Sulfate 0.000573 0.000573 0.000573
Biotin 0.000507 0.000507 0.000507
Vitamin K 0.000835 0.000835 0.000835
Vitamin D3 0.000235 0.000235 0.000235
Sodium selenate 0.0000491 0.0000491 0.0000491
Potassium iodide 0.0000105 0.0000105 | 0.0000105
Cyanocobalamin 0.0000103 0.0000103 1 0.0000103


Total Lutein (meg/liter as fed) 200 ] 700 1000
Total DHA (mg/liter as fed) 200 200 200
Ratio - Lutein (meg) : DHA (nig) as fed 1-5 1 32 4.5
1. FIoraGLO® Crystalline Lutein, Kemin Foods, Des Moines, Iowa, USA
The ingredients listed in the preceding table are combined and processed to form a concentrated human milk fortifier embodiment of the present invention. One method of preparing such an embodiment is described below.
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(0099] An initial intermediate blend is prepared by heating to 32-37°C the specified amounts of coconut oil, MCT oil, soy oil, DHA oil and AA oil, all with agitation. A soy lecithin emulsifier is added with agitation to the heated blend and allowed to dissolve. Vitamins A, D, and K, Natural Vitamin E, and lutein are then added with agitation to the developing blend. Milk protein isolate (25.8 kg) and the specified amounts of ultra roicronized tricalcium phosphate and calcium carbonate are added to the blend. The resulting intermediate blend is maintained at 26-48°C under moderate agitation for a period of time not to exceed six hours before being added to the aqueous protein blend described below.
[0100] An aqueous protein blend is then prepared by hearing 573 kg of ingredient water at 48-60°C, and then adding to it with agitation milk protein isolate (38.8 kg) and the specified amount of whey protein concentrate. Thereafter, and with agitation, the entire intermediate blend described above is added to the aqueous protein blend. The following ingredients are then added to the resulting blend in the following order: potassium citrate, dipotassium phosphate, monopotassnnn phosphate, magnesium phosphate, sodium chloride, potassium chloride, potassium iodide and sucrose. After no less than five minutes, the blend pH is adjusted to 6.60-6.80 using a IN KOH solution, and thereafter maintained at 51-60°C, for a period of time not to exceed two hours before further processing.
[0101] The pH adjusted blend is then homogenized using one or more in-line homogenizers at pressures from 1000-4000 psig with or without a second stage homogenizafion from 100-500 psig followed by heat treatment using a HTST (high temperatnre short time, 74°C for 16 seconds) or UHTST (ultra-high temperature short time, 132-154°C for 5-15 seconds) process. The choice of UHTST or HTST is normally made based upon a review of the bioborden of each of the ingredients in the formulation. After the appropriate heat treatment, the batch is cooled in a plate cooler to 1.0-5.0°C and then transferred to a refrigerated holding tank, where it is subjected to analytical testing and then standardized to finished product specifications, which includes the addition of an ascorbic acid solution and a water-soluble vitamin and trace mineral solution, all of which is prepared separately before adding to the previously described refrigerated batch.
[0102] The ascorbic acid solution is prepared by adding the specified amount of ascorbic acid to 11.1 kg of IN KOH solution with agitation. The water-soluble vitamin and trace mineral solution is prepared by heating 25.2 kg of ingredient water to 37°C to 48°C. The water soluble vitamins and trace minerals are added to the water as a premix which contains m-inositoL taurine, niacinamide, zinc sulfate, calcium pantothenate, ferrous sulfate, cupric sulfate, riboflavin, thiamine hydrochloride, pyridoxine hydrochloride, folic acid, manganese sulfate, biotin, sodium selenate, and cyanocobalarnin. As noted above, both solutions are then added to the refrigerated batch, all with agitation. As part of batch standardization, the appropriate amount of ingredient dilution water is then
24

WO 2007/050521 PCT/US2006/041303
added to the batch for a target total solids level of 31%, and the pH adjusted to 7.1 with a IN KOH solution. The batch is filled into suitably sized containers containing 5 ml of product
[0103] The exemplified human milk fortifier concentrates are combined with human milk (5 ml concentrate with 20-25 ml human milk). The fortified human milk is then fed to pre-term infants to provide 7-300 ug/kg/day of lutein. The formula provides improved retinal health and vision development as described herein, including reduced risk of retinopathy of prematurity.
Examples 5.1-5.3
(0104) This example illustrates a ready-to-feed, preterm infant formula embodiment of the present invention. This formula is similar to Similac® Special Care® Advance ® with Iron Premature Infant formula, a preterm infant formula available from Abbott Laboratories, Columbus, Ohio, except for the increased lutein concentrations and subsequent lutein to docosahexaenoic acid ratios.
[01€5J The preterm infant formula includes nonfat milk, com syrup solids, lactose, medium chain triglycerides, whey protein concentrate, soy oil, coconut oiL C. cohnii oil (source of docosahexaenoic acid), M. afpina oil (source of arachidonic acid), calcium phosphate, calcium carbonate, potassium citrate, ascorbic acid, magnesium chloride, soy lecrfliin, mono- and digfycerides, m-inositol, sodium citrate, carrageenan, ferrous sulfate, choline bitartrate, taurine, choline chloride, niacinamide, d-alpha-tocopheryJ acetate, L-camrrine, zinc sulfate, potassium chloride, potassium phosphate dibasic, calcium pantothenate, cupric sulfete, vitamin A rmlrnitate, riboflavin, thiamine chloride hydrochloride, pyridoxine hydrochloride, folic acid, beta-carotene, manganese sulfate, biotin, phylloquinone, sodium selenate, vitamin D3, cyano«>ba3amin and nucleotides (cytidine ^-monophosphate, disodhrm goanosioe 5,-monophosphate, disodium uridine 5-raonophosphale, adenosine ^-monophosphate).
[0106] The above-referenced ingredients are formulated together by conventional methods to provide the following nutrition profile:
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I Nntrients Example 5.1 Example 5.2 Example 5.3
Amount per 100 kcal (or 123 ml) Amount per 100 kcal (or 123 ml) Amount per 100 kcal (or 123 ml)
Protein {from nonfat milk, whey protein concentrate) 3-00 g 3.00 g 3.00 g
Fat (form 5030:18.3 mix of MCT oil, soy oil, and coconut oils;
mgDHA, mgARA;700mg
linoleic acid) 5.43 g 5.43 g 5.43 g
Carbohydrate (source - 50:50 mix of com syrup solids, Lactose) 103 g 103 g 103 g
Lutein 18.4 meg 37 meg 9141 meg
VitaininA 1250IU 1250IU 1250 IU
"Vitamin D 150IU 150 IU 150 IU
VitaminE 4.0 IU 4.0 IU 4.0 IU
Vitamin K 12 meg 12 meg 12 meg
Thiamine 250 meg 250 meg 250 meg
Riboflavin 620 meg 620 meg 620 meg
Vitamin B6 250 meg 250 meg 250 meg
Vitamin B12 0.55 meg 0.55 meg 0.55 meg
Niacin 5000 meg 5000 meg 5000 meg
Folic acid 37 meg 37 meg 37 meg
Pantothenic acid 1900 meg 1900 meg 1900 meg
Biotin 37.0 meg 37.0 meg 37.0 meg
Ascorbic acid 37 mg I 37 mg 37 mg
Choline 10 mg 10 mg 10 mg
Inositol 40.0 mg 40.0 mg 40.0 mg
Calcium 180 mg ( 9.0 mEq) 180 mg( 9.0 mEq) 180 mg( 9.0 mEq)
Phosphorus 100 mg 100 mg 100 mg
Magnesium 12.0 mg 12.0 mg 12.0 mg
Iron 1.8 mg 1.8 mg 1.8 mg
Zinc 1.50 meg 1.50 meg 1.50 meg
Manganese 12 meg 12 meg 12 meg
Copper 250 meg 250 meg 250 meg
Iodine 6 meg 6mcg 6 meg
Selenium 1.8 meg 1.8 meg 1.8 meg
Sodium 43 mg (1.9 mEq) 43 mg (1.9 mEq) 43 mg (1.9 mEq)
Potassium 129 mg (33 mEq) 129 mg (33 mEq) 129 mg (33 mEq)
Chloride 81 mg (23 mEq) 81 mg (23 mEq) 81 mg (23 mEq)

Total Lutein (meg/liter as fed) 150 300 1150
Total DHA (mg/liter as fed) 112 112 115
Ratio - Lutein (meg) : DHA (mg) as fed 13:1 3:1 10:1
10107] The exemplified ready-to-feed formulas (caloric density of 812 fccaMiter) are administered to preterm infants to provide from 7-300 mcg/kg of lutein per day. The administered formula improves eye health as described herein, and are especially usefol as applied to preterm mfents to reduce the risk of retinopathy of prematurity and helps protect the eyes from natural or artificial light, especially biliary lights.
EXPERIMENT
[0108] A study is conducted to compare plasma lutein concentrations in breastfed infants with plasma lutein concentrations in formula fed infants. The latter received one of three formulas defined by
26

WO 2007/050521 PCT/US2006/041303
lutein concentrations of 32.6 meg/liter (LI), 52.6 mcg/Titer (L2), or 14.6 meg/liter (CTRL). The study groups and resulting plasma lutein concentrations are summarized in the following table.

Feeding Group Plasma Lutein (mcg/dL)*

Study Day 1 Study Day 56
Control (CTRL) - no added lutein Total inherent lutein 14.6 mcg/lher lutein) 137 ±029"
(0.40-5.10)
20 2.17 ±0.12*
(1.16-325)
18
Formula LI
Added lutein approximately 18 meg/liter
Inherent lutein approximately 14.6 mcg/lher
Total lutein 32.6 mcg/L 0.78 ±0.09"
(027-2.09)
24 221 ± 0.16 a
(020-3.61)
22
Formula 12
Added lutein approximately 3Smcg/riter
Inherent lutein approximately 14.6 mcg/lher
Total ratek 52.6 mcg/L 0.97 ±0.13*
(020-231)
21 325 ±026*
(0.6O4.70)
19
Human Milk (HM) 6.5-107.8 meg/liter lutein 6.53 ±0.54* (1.69-14.12) 24 5.88 ± 0.77 e (0.49-20.09) 26
*V~aioes are presented as mean ± SEM, (range) n Values in a column with superscripts -without a common letter
difier(p [O109J Plasma lutein concentrations (at day 56 of the study) are used as the primary variable in the sfndy. The primary comparison is the difference in plasma lutein concentrations between the L2 and CTRL formula groups. Secondary comparisons included differences in plasma lutein concentrations among the formula groups (CTRL, LI, L2) and differences between the formula groups and the human milk group. Plasma lutein concentrations on day 56 fiom a total of 85 infants (CTRL, n=l 8; LI, n=22; L2, n=19; HM, n=26) is used in these analyses and results reported as mean ± SEM in the following table.
[0110] Infants in the L2 formula group have significantly higher (p [0111] The data from the study show that lutein is surprisingly less bioavailable from infant formula than from human milk (see Figure 1). As such, in order for an infant formula to produce plasma
27

WO 2007/050521 PCT/US2006/041303
lutein concentrations in infants similar to that produced by feeding human milk, an infant formula must be formulated to contain at least about 50 meg/liter of lutein, preferably from 100 meg/liter to about 200 mcg/Iiter.
Dated this 5th day of May 2008 To


The controller of Patents


The Patents office at mumbai
28

WO 2007/050521




WE CLAIM:-

PCT/US2006/041303


1- An infant fonnula comprising fat, protein, carbohydrate, vitamins, and minerals, including docosahexaenoic acid and, on a ready-to-feed basis, at least about 50 mcg/lher of lutein, wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 1:2 to about 10:1 and the formula is free of egg phospholipids.
2. The infant formula of Claim 1 wherein the composition comprises from about 50 to about 1150 meg/liter of added lutein.
3. The infant fonnula of Claim 1 wherein the formula comprises from about 100 to about 200 meg/liter of lutein.
4. The infant formula of Claim 1 wherein the formula emprises from about 100 to about 200 meg/liter of lutein.
5. The infant formula of claim 1 wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 1.5:1 to about 10:1.
6. The infant formula of claim 1 wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 2:1 to about 5:1.
7. The infant formula of claim 1 wherein the fonnula has a caloric density of from 20 to 30 kcal/fiuid ounce.
8. The infant formula of claim 1 wherein the fonnula comprises a combination of free lutein and
zeaxanthm from a single source, that source being a crystalline extract of Tagetes erecta in which the
free lutein represents from 85% to 95% by weight of the combination and the free zeaxanthin
represents from about 5% to about 15% by weight of the combination.
9. The infant formula of claim 1 wherein the formula is a powder.
10. The infant formula of claim 1 wherein the formula is a liquid.
11. The infant fonnula of claim 1 wherein the lutein comprises at least 25% by weight of added
lutein.
12. An infant formula comprising, on a ready-to-feed basis,
29

WO 2007/050521

PO7US2006/041303

(a) from about 54 to about 108 gm/liter of carbohydrate,
(b) from about 20 to about 54 gm/liter of fat,
(c) from about 7 to about 24 gm/liter of protein,
(d) from about 100 to about 200 meg/liter of lutein, and
(e) docosahexaenoic acid,
wherein the weight ratio of lutein (meg) to docosahexaenoic acid (mg) is from about 2:1 to about 5:1 and the formula is free of egg phospholipids.
13. The infant formula of claim 12 wherein the formula comprises a combination of free lutein and
zeaxanthin from a single source, that source being a crystalline extract of Tagetes erecta in which the
free lutein represents from 85% to 95% by weight of the combination and the free zeaxanthin
represents from about 5% to about 15% by weight of the combination.
14. A method to reduce the risk of retinopathy of prematurity in preterm infants, said method comprising administering to a preterm infant in need thereof a formula according to claim 1 to provide the infant from about 7 to about 300 meg/kg/day of lutein.
15. A method to reduce the risk of retinopathy of prematurity in preterm infants, said method comprising administering to a preterm infant in need thereof a formula according to claim 1 to provide the infant from about 14 to about 220 meg/kg/day of lutein.
16. A method to promote retinal health and vision development in infants, said method comprising administering to an infant in need thereof a formula according to claim 1 to provide the infant from about 7 to about 300 meg/kg/day of lutein.
17. A method to promote retinal health and vision development in infants, said method comprising administering to an infant in need thereof a formula according to claim 1 to provide the infant from about 7 to about 300 meg/kg/day of lutein.
30

Documents:

993-MUMNP-2008-ASSIGNMENT (10-10-2011).pdf

993-MUMNP-2008-ASSIGNMENT(10-10-2011).pdf

993-MUMNP-2008-CANCELLED PAGES(10-10-2011).pdf

993-MUMNP-2008-CLAIMS(15-5-2008).pdf

993-MUMNP-2008-CLAIMS(AMENDED)-(10-10-2011).pdf

993-MUMNP-2008-CLAIMS(AMENDED)-(16-6-2011).pdf

993-MUMNP-2008-CLAIMS(GRANTED)-(1-11-2011).pdf

993-MUMNP-2008-CLAIMS(MARKED COPY)-(10-10-2011).pdf

993-MUMNP-2008-CLAIMS(MARKED COPY)-(16-6-2011).pdf

993-mumnp-2008-claims.doc

993-mumnp-2008-claims.pdf

993-MUMNP-2008-CORRESPONDENCE (12-10-2012).pdf

993-MUMNP-2008-CORRESPONDENCE(10-10-2011).pdf

993-MUMNP-2008-CORRESPONDENCE(12-10-2012).pdf

993-MUMNP-2008-CORRESPONDENCE(18-5-2011).pdf

993-MUMNP-2008-CORRESPONDENCE(20-6-2011).pdf

993-MUMNP-2008-CORRESPONDENCE(21-3-2011).pdf

993-MUMNP-2008-CORRESPONDENCE(8-11-2011).pdf

993-MUMNP-2008-CORRESPONDENCE(IPO)-(1-11-2011).pdf

993-mumnp-2008-correspondence(ipo)-(10-8-2009).pdf

993-mumnp-2008-correspondence.pdf

993-mumnp-2008-descprition(complete).doc

993-MUMNP-2008-DESCRIPTION(COMPLETE)-(15-5-2008).pdf

993-mumnp-2008-description(complete).pdf

993-MUMNP-2008-DESCRIPTION(GRANTED)-(1-11-2011).pdf

993-MUMNP-2008-DRAWING(GRANTED)-(1-11-2011).pdf

993-mumnp-2008-drawing.pdf

993-MUMNP-2008-FORM 1(15-5-2008).pdf

993-MUMNP-2008-FORM 1(21-3-2011).pdf

993-mumnp-2008-form 1.pdf

993-mumnp-2008-form 13(21-3-2011).pdf

993-mumnp-2008-form 18(15-5-2008).pdf

993-MUMNP-2008-FORM 2(COMPLETE)-(15-5-2008).pdf

993-MUMNP-2008-FORM 2(GRANTED)-(1-11-2011).pdf

993-MUMNP-2008-FORM 2(TITLE PAGE)-(15-5-2008).pdf

993-MUMNP-2008-FORM 2(TITLE PAGE)-(GRANTED)-(1-11-2011).pdf

993-mumnp-2008-form 2(title page).pdf

993-mumnp-2008-form 2.doc

993-mumnp-2008-form 2.pdf

993-MUMNP-2008-FORM 3(15-5-2008).pdf

993-MUMNP-2008-FORM 3(18-5-2011).pdf

993-mumnp-2008-form 3.pdf

993-mumnp-2008-form 5.pdf

993-MUMNP-2008-GENERAL POWER OF ATTORNEY(21-3-2011).pdf

993-MUMNP-2008-OFFICE ACTIONS OF EP & US PATENT(18-5-2011).pdf

993-mumnp-2008-pct request.pdf

993-mumnp-2008-pct-ib-304.pdf

993-mumnp-2008-pct-isa-237.pdf

993-MUMNP-2008-PETITION UNDER RULE 137(10-10-2011).pdf

993-MUMNP-2008-PETITION UNDER RULE 137(18-5-2011).pdf

993-mumnp-2008-power of attorney.pdf

993-MUMNP-2008-REPLY TO EXAMINATION REPORT(16-6-2011).pdf

993-MUMNP-2008-REPLY TO EXAMINATION REPORT(18-5-2011).pdf

993-MUMNP-2008-REPLY TO HEARING(10-10-2011).pdf

993-MUMNP-2008-REPLY TO HEARING(5-10-2011).pdf

993-MUMNP-2008-US DOCUMENT(16-6-2011).pdf

993-mumnp-2008-wo international publication report a2.pdf

993-mumnp-2008-wo international publication report a3.pdf

993-mumnp-2008-wo international publication report(15-5-2008).pdf

abstract1.jpg


Patent Number 249657
Indian Patent Application Number 993/MUMNP/2008
PG Journal Number 44/2011
Publication Date 04-Nov-2011
Grant Date 01-Nov-2011
Date of Filing 15-May-2008
Name of Patentee ABBOTT LABORATORIES
Applicant Address DEPT. 377/AP6A-1, 100 ABBOTT PARK ROAD, ABBOTT PARK, ILLINOIS 60064,
Inventors:
# Inventor's Name Inventor's Address
1 BARRETT-REIS, BRIDGET 9401 TRACEYTON DRIVE, DUBLIN, OHIO 43017,
2 PRICE, PAMELA, T. 630 W.BROADWAY,GRANVILLE,OHIO 43023,
3 MACKEY, AMY 409 E COMO AVENUE, COLUMBUS,OHIO 43202,
PCT International Classification Number A23L1/29
PCT International Application Number PCT/US2006/041303
PCT International Filing date 2006-10-23
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 60/730283 2005-10-26 U.S.A.