CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63/381,584, filed October 31, 2022, which is incorporated by reference herein in its entirety.

TECHNICAL FIELD

[0002] This application relates to methods for reducing lameness in swine that includes feeding to the swine a feed that includes a nitrate compound.

BACKGROUND

[0003] Feeding calcium nitrate and other ingredients to livestock is one way to mitigate methane production. For example, U.S. Patent No. 8,771,723 titled “Compositions For Reducing Gastro-Intestinal Methanogenesis In Ruminants” issued to Hindrik Bene Perdok et al. discloses one such method. Further, Van Den Bosch et al. (International Patent Application Publication No. WO 2016/090366, published June 9, 2016) describes animal feed compositions containing a nitrate compound and methods for using such a composition in an animal’s gestation phase and/or lactation phase to improve the health of offspring. Ascensao et al. (International Patent Application Publication No. WO 2018/237233) describes methods of improving meat quality (such as meat color and carcass yield (%)) in an animal by feeding an effective amount of nitrate to the animal.

[0004] In humans, dietary nitrate, as presented in beetroot juice, has been shown to reduce blood pressure and the risk of adverse cardiovascular events in healthy individuals after single dose of 500 ml of beetroot juice. It is hypothesized that nitrate might represent a source of vasoprotective nitric oxide (NO) via bioactivation (Webb et al. 2008). A non-enzymatic pathway for the generation of NO has been proposed (nitrate-nitrite-NO pathway) for humans. Dietary inorganic nitrate molecules may be reduced by facultative anaerobic bacteria on the dorsal surface of the tongue to nitrite which can be chemically and enzymatically be further reduced to NO (Lundberg et al. 2009). The endothelial isoform of the NO synthase uses arginine and molecular oxygen as precursors to tonically release NO in the endothelium, which is important for the control of vascular tone, smooth muscle growth, platelet aggregation and inflammation (Umans and Levi, 1995; Bruckdorfer 2005). This induces vasodilation and increased blood flow (Siervo et al. 2011; Keim 1999).

[0005] Pig farm efficiency can be defined in different ways. For reproducers, one indicator is longevity, measured with respect to the culling age for a sow. Longevity is reduced when sows are culled for lameness. Lameness is the second cause for culling in modem sow farms (Supakorn et al., 2022; Phuym et al., 2013) in modern genetic breed. Leg pain is one contributor to lameness, which is a result of heavy weight in swine. Gilts and primiparous sows are highly sensitive to this problem due to the fast growth rate of modern genetics. (Belkova and Rozkot, 2022) The present invention is directed to methods of feeding pigs at key moments in their life cycle a specific dose of a swine feed or feed supplement that includes nitrate compounds with or without polyphenols. As a result, bones development is stimulated, which supports higher sow longevity.

SUMMARY

[0006] In one aspect, the present technology provides a method for method of feeding swine, the method includes: feeding swine a swine feed that includes about 0.01 weight percent (wt%) to about 0.5 wt% per day of a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant that includes polyphenols; wherein swine fed the swine feed exhibit a reduced occurrence of joint lesions compared to a control swine not fed the swine feed. In any aspect, reference to weight percent (wt%) per day is based on total weight of swine fed as part of the swine’s daily feed intake.

[0007] In one aspect, the present technology provides a method for improving bone development in swine, the method comprising: feeding swine a swine feed, wherein the swine feed comprises: a nitrate compound, hydrates thereof, salts thereof, or combinations thereof; and optionally an antioxidant comprising polyphenols; wherein: the swine is fed a daily feed intake comprising an amount of nitrate in a range of about 0.01% by weight to less than about 0.5% by weight based on total weight of the swine feed; the method improves bone mineral deposition in swine fed the swine feed compared to a control swine not fed the swine feed. [0008] In another aspect, the present technology provides the use of a swine feed as described herein in any aspect for increasing bone mineral deposition and/or reducing the occurrence of joint lesions in swine compared to a control swine not fed the swine feed, wherein the swine feed comprises a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant that include polyphenols, and wherein the daily feed intake amount of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof is about 0.01% by weight to less than about 0.5% by weight based on total weight of the swine feed.

DETAILED DESCRIPTION

[0009] Reference will now be made in detail to certain aspects of the disclosed subject matter. While the disclosed subject matter will be described in conjunction with the enumerated claims, it will be understood that the exemplified subject matter is not intended to limit the claims to the disclosed subject matter. One aspect described in conjunction with a particular aspect is not necessarily limited to that aspect and can be practiced with any other aspect(s).

[0010] Throughout this document, values expressed in a range format should be interpreted in a flexible manner to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited. For example, a range of “about 0.1% to about 5%” or “about 0.1% to 5%” should be interpreted to include not just about 0.1% to about 5%, but also the individual values (e.g., 1%, 2%, 3%, and 4%) and the sub-ranges (e.g., 0.1% to 0.5%, 1.1% to 2.2%, 3.3% to 4.4%) within the indicated range. The statement “about X to Y” has the same meaning as “about X to about Y,” unless indicated otherwise. Likewise, the statement “about X, Y, or about Z” has the same meaning as “about X, about Y, or about Z,” unless indicated otherwise.

[0011] As used herein, the singular forms "a," "an," and "the" and similar referents in the context of describing the elements (especially in the context of the following claims) include plural referents unless the context clearly dictates otherwise. For example, reference to "a substituent" encompasses a single substituent as well as two or more substituents, and the like. It is understood that any term in the singular may include its plural counterpart and vice versa, unless otherwise indicated herein or clearly contradicted by context.

[0012] Nothing in the cited references teaches or suggests the claimed process. In addition, it is to be understood that the phraseology or terminology employed herein, and not otherwise defined, is for the purpose of description only and not of limitation. Any use of section headings is intended to aid reading of the document and is not to be interpreted as limiting; information that is relevant to a section heading may occur within or outside of that particular section.

[0013] As used herein, the terms "for example," "for instance," "such as," or "including" are meant to introduce examples that further clarify more general subject matter. Unless otherwise specified, these examples are provided only as an aid for understanding the applications illustrated in the present disclosure, and are not meant to be limiting in any fashion. [0014] In the methods described herein, the acts can be carried out in a specific order as recited herein. Alternatively, in any aspect(s) disclosed herein, specific acts may be carried out in any order without departing from the principles of the disclosure, except when a temporal or operational sequence is explicitly recited. Furthermore, specified acts can be carried out concurrently unless explicit claim language recites that they be carried out separately or the plain meaning of the claims would require it. For example, a claimed act of doing X and a claimed act of doing Y can be conducted simultaneously within a single operation, and the resulting process will fall within the literal scope of the claimed process.

[0015] The term “about” as used herein can allow for a degree of variability in a value or range, for example, plus or minus within 10%, within 5%, or within 1% of a stated value or of a stated limit of a range, and includes the exact stated value or range.

[0016] The term “substantially” as used herein refers to a majority of, or mostly, as in at least about 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.9%, 99.99%, or at least about 99.999% or more, or 100%.

[0017] As used herein, the following terms have the following meanings unless expressly stated to the contrary.

[0018] The term “animals” as used in this disclosure includes monogastric and ruminant animals. As used in this disclosure, the term “monogastric” means any organism having a simple single-chambered stomach. Such monogastric animals include, but are not limited to, porcine, equine, avian animals, seafood (aquaculture) animals.

[0019] Porcine monogastric animals refers to domesticated swine (or pigs) and wild boars. Swine include, but are not limited to, feeder pigs and breeder pigs, including piglets, sows, gilts, barrows, and boars. “Gilts” as used herein refers to female pigs intended for slaughter or breeding purposes that have not yet farrowed a litter. “Barrows” as used herein refers to castrated male pigs intended for slaughter, whereas “boars” as used herein refers to intact, sexually mature, male pigs intended for breeding purposes.

[0020] Swine production can be logically separated into a number of phases. Typically, the stages include, breeding and gestation, farrowing, weaning, growing or growth, and finishing or finisher phases. Generally, the stages described herein may vary across geographies as understood by persons skilled in the art, and the definitions provided herein provide exemplary descriptions, that include but are not limited to, weights, ages, and development stages in swine production.

[0021] Breeding and Gestation. The sow is bred during her estrous period. In the phase between weaning the sow and breeding, a special breeding feed can be fed to the sow. After breeding, the sow “gestates” her litter for 113 to 116 days before the piglets are born or “farrowed.” As used herein, the term “gestation” or gestation phase means the 113 to 116 day period when the sow is pregnant from breeding until farrowing.

[0022] Farrowing. The process of giving birth is called farrowing. Typically, sows are moved to a farrowing room a few days prior to farrowing and farrow about eight to fourteen piglets (as a group called a “litter”). The piglets are born weighing about 1.5 kg at birth. As used herein, the term “farrowing” means birth, and the term “farrowing phase” means the period from birth to weaning. The period of time called the transition period is typically seven days prefarrowing until one to five days post farrowing. The piglets stay with the lactating sow for about 19-35 days after farrowing, during which time the piglets drink milk produced by the lactating sow.

[0023] Weaning. The piglets are weaned from the sow at anywhere from five days to five weeks, with most operations weaning pigs at two to four weeks after farrowing. As used herein, the term “weaning” means the process of separating the piglets from the sow. During a weaning or nursery phase, the piglets remain in the nursery — for example, until about four to eight weeks after farrowing (or in other instances, up to about 18 kg to about 30 kg). “Postweaning” as used herein means the period following the weaning or nursery phase — typically at about six to ten weeks of age where the pig is fed a transitioning diet and before being fed a suitable grower/finishing phase diet based on the pigs age and weight.

[0024] Growing and Finishing. Pigs are normally placed in a grow-fini shing building until they reach market weight, following the weaning (or nursery) phase. “Growing phase” or “growth phase” as used herein refers to the stage where pigs starting from about 35 pounds to 60 pounds (-16-27 kg) are fed to a body weight of about 120 to about 165 pounds (-54-75 kg). “Finishing phase” or “finisher phase” as used herein refers to the stage where pigs are fed from about 120 to 150 pounds (-54-68 kg) are fed to a market body weight — typically about 200 to about 309 pounds (-91-140 kg), and preferably about 243 pounds to about 309 pounds (—110- 140 kg).

[0025] As used herein, the term “nitrate compound” refers to any material including one or more nitrate-containing compounds suitable for use in swine feed.

[0026] The nitrate compounds as described in any aspect herein may be any suitable nitrate-containing compound. As would be understood by a person of ordinary skill in the art, a suitable nitrate-containing compound may be defined as any physiologically acceptable or tolerated nitrate compound. For example, the nitrate compound may be well-soluble in water, z.e., the compound has sufficient solubility for bioavailability after administration to the swine. In some aspects, the nitrate compound may be an ionic nitrate compound, preferably an inorganic nitrate salt. Typically, the ionic nitrate salts may include, but are not limited to, sodium nitrate, potassium nitrate, calcium nitrate, magnesium nitrate, ammonium nitrate, or mixtures thereof, all of which are readily soluble in water at standard temperature (e.g., 0°C (32°F)) and pressure (e.g., 1 bar). Preferably, the ionic nitrate salt may be calcium nitrate. The ionic nitrate salts may include different hydrated forms. For example, the ionic nitrate salts may also include double salts (e.g., calcium nitrate and ammonium nitrate). In any aspect described herein, a mixture of two or more different ionic nitrate salts or salt forms may be used in the method.

[0027] While not intending to be limited by any particular theory, it is believed that nitrate is a source for the biological messenger nitric oxide (NO) according to a non-enzymatic pathway for the generation of NO (nitrate-nitrite-NO pathway). Unlike arginine conversion to nitric oxide, nitrate conversion to nitric oxide via the nitrate-nitrite-NO pathway is not dependent upon oxygen levels. It is believed that the release of NO in the animal is important for the control of vascular tone, smooth muscle growth, platelet aggregation and inflammation. The release of NO in the animal is believed to induce vasodilation and increased blood flow and exchange of oxygen. While vasodilation is one biological pathway affected by nitrate supplementation, vasodilation does not account for all improvements observed following nitrate supplementation and can vary based on a number of factors (e.g., species, age, feed, etc.). For example, nitric oxide has also been shown increase bone formation and bone mineral density (van’t Hof and Ralston, 2001). Accordingly, outcomes can vary across species (or within a species) fed a feed containing nitrates. [0028] In any aspect described herein, the nitrate compound may be provided as calcium nitrate having the formula Ca(NCh)2. Calcium nitrate is also referred to as calcium dinitrate, Kalksalpeter, nitrocalcite, Norwegian saltpeter, and lime nitrate. Calcium nitrate may be produced by treating limestone with nitric acid, according to the reaction: CaCCh + 2HNCh Ca(NCh)2 + CO2 + H2O. Additionally or alternatively, in some aspects, the one or more nitrates may be provided as an inorganic salt of magnesium nitrate hexahydrate having the formula (Mg(NO3)2*6H2O). This product contains 10.8% N from nitrate and 9.5% Mg.

[0029] A variety of related complex inorganic salts of calcium nitrate include calcium ammonium nitrate decahydrate and calcium potassium nitrate decahydrate. Calcium ammonium nitrate is a double salt (calcium nitrate and ammonium nitrate) having the formula 5Ca(NO3)2*NH4NO3*10H2O. For example, the calcium ammonium nitrate may be pentacalcium ammonium nitrate decahydrate commercially available from Bri-Chem Supply Limited with the following specification: Ammonium-N (NH4-N): 1.1%; Nitrate-N (NO3-N): 14.4%; Total N: 15.5%; Calcium (Ca): 18.8%. In another example, the calcium nitrate may be BOLIFOR CNF calcium nitrate feed grade having the formula SCafNChC’NILNCh’ I OFhO commercially available from Yara Phosphates Oy of Helsingborg Sweden. In yet another example the calcium nitrate may have the following specification: Calcium (Ca): 18.9%; Nitrogen (N) 15.5%; pH (10% solution): 6; bulk density (kg/m ³ ): 1050; appearance: prilled; size: <1.0 mm: 2%; 1.0-2.0 mm: 78%; >2 mm: 20%. Exemplary formulations of calcium nitrate lacking ammonia include Ca(NO3)2*4H2O. An exemplary anhydrous air-stable derivative of calcium nitrate may include urea complex Ca(NO3)2*4[OC(NH2)2].

[0030] Antioxidants, when included in swine diets, are known to protect cells from damage caused by free radicals and peroxides. There are different types of antioxidants available for feeding swine, including natural dietary antioxidants. For example, natural antioxidants may include, but are not limited to, vitamins, minerals, carotenoids, and polyphenols.

[0031] In any aspect described herein, the antioxidant preferably includes polyphenols. Suitable polyphenol sources may include, but are not limited to, fruits (such as apples, apricots, currants, blackberries, blueberries, cherries, chokeberries, cranberries, dates, elderberries, gooseberries, grapes, kiwis, lemons, lingonberries, limes, mangoes, marionberries, nectarines, olives, oranges, peaches, pears, plums, pomegranates, quinces, raspberries, rhubarb, strawberries, tomatoes, or mixtures thereof), vegetables (such as artichokes, broccoli, celery, corn, eggplant, fennel, garlic, greens, kohlrabi, leeks, lovage, onions, parsnips, peppers, spinach, red cabbage, rutabagas, scallions, shallots, sweet potatoes, watercress, or mixtures thereof), legumes (such as chick peas, beans, lentils, snap beans, English peas, green peas, fava beans, or mixtures thereof), nuts (such as almonds, cashews, hazelnuts, pecans, peanuts, pistachios, walnuts, or mixtures thereof), seeds (such as fruit seeds, flax seeds, pumpkin seeds, sunflower seeds, or mixtures thereof), teas (such as green tea, black tea, oolong tea, Earl Gray tea, Ceylon tea, Darjeeling tea, or mixtures thereof), herbs and spices (such as basil, chives, capers, cinnamon, curry, dill weed, horseradish, oregano, parsley, rosemary, sage, tarragon, thyme, or mixtures thereof), tree barks (such as arjuna bark, aspen bark, birch bark, conifer bark, eucalyptus bark, maple bark, or mixtures thereof), or mixtures thereof. Preferably, the antioxidant may include polyphenols from polyphenol sources including onion extract, grape seed extract, rosemary extract, or mixtures thereof.

[0032] A polyphenol source suitable for use in any aspect of the present technology may include commercially available sources. For example, a polyphenol source is commercially available under the trade name PROVIOX 50, available from Cargill, Incorporated, Wayzata, MN, USA. The formulation of PROVIOX 50 includes a blend of polyphenol grape seed and skin extracts, onion extracts, and rosemary extracts. PROVIOX 50 is a standardized product for its total polyphenol content. PROVIOX 50 contains sources of polyphenols with antioxidant properties. In any aspect, the antioxidant that includes polyphenols is PROVIOX 50.

[0033] In any aspect described herein, the term “swine feed” refers to a feed ration produced for consumption by swine. The term “compound feed” as used in any aspect described herein refers to a swine feed blended to include two or more ingredients which assist in meeting certain daily nutritional requirements of swine, preferably swine in one or more of a postweaning, growth, or finishing phase. The swine feed may be a complete swine feed according to any aspect described herein. The term “complete feed” as used in any aspect described herein refers to a swine feed having a nutritionally balanced blend of ingredients designed as the sole ration to provide all the daily nutritional requirements of a swine to maintain life and promote production without any additional substances being consumed except for water. The swine feed may also be a concentrate swine feed according to any aspect described herein. The term “concentrate feed” or “concentrate swine feed” as used herein in any aspect described herein refers to a swine feed that typically includes a protein source blended with supplements or additives or vitamins, trace minerals, other micro ingredients, macro minerals, etc. to provide a part of the ration for the swine. The concentrate feed may be fed along with other ingredients. The swine feed may include a premix according to any aspect described herein. The term “premix” as used in any aspect described herein refers to a blend of primarily vitamins and/or minerals along with appropriate carriers. The swine feed may also include a base mix according to any aspect described herein. The term “base mix” as used in any aspect described herein refers to a blend containing vitamins, trace minerals and/or other micro ingredients plus macro minerals such as calcium, phosphorus, sodium, magnesium, and potassium, vitamins, or combinations thereof. The swine feed may be a feed “supplement.” As used herein in any aspect, the terms “feed supplement” or “swine feed supplement” refers to a concentrated additive premix that includes the active ingredients, which premix or supplement may be added to a swine’s feed or ration to form a supplemented feed in accordance with the present technology. As used herein in any aspect, the term “additive” or “feed additive” refers to an ingredient such as a protein source, salt, mineral, additive, or buffer that is added to a swine feed. For example, an additive may include, but is not limited to, calcium, zinc, manganese, copper, iodine, cobalt, selenium, other trace ingredients, or mixtures thereof.

[0034] Evidence from humans suggest that consumption of nitrate may reduce resting metabolic rate, which suggests there is a potential for more efficient energy utilization (e.g., growth, exercise). (Larsen et al., Am. J. Clin. Nutr., 2014, 99:843-50). However, a meta-analysis found that dietary nitrate supplementation did not significantly decrease resting metabolic rate. (Pawlak-Chaouch et al., Nitric Oxide, 2016, 53:65-67). While there is not clarity in differences in outcome of the two studies, it is believed that the amount of nitrate supplementation or nitrate content of individuals daily diet may have contributed to the differences observed. Based upon a review by Ghasemi (EXCLI Journal, 2022, 21 :470-486), up to 10% of nitric oxide produced in human bodies come from an exogenous source (e.g., diet, environment).

[0035] While the above studies provide limited, yet inconclusive, insight on the potential effects of nitrate consumption in humans, much is unknown with respect to the effect of exogenous nitrate consumption on the growth of pigs, including appropriate intake levels of exogenous nitrate. Although pigs are omnivores like humans, most domestic pigs (z.e., pigs in swine production) are fed grain and oilseed diets devoid of leafy-green vegetables that may serve as nitrate sources, which suggests that overall nitrate intake is lower than historic values in both feral pigs and humans. Moreover, much is unknown with respect to the effect of exogenous nitrate on the bone development in swine, such as swine in a post-weaning, growth, and/or finishing phase.

[0036] The present inventors discovered inclusion of nitrate in the daily feed intake of swine significantly improved bone development (i.e., bone mineralization measured as metacarpals bone ash content(published method according to Adeola et al., 2015)) of the swine compared to swine fed a conventional feed that does not include exogenous nitrate. In addition, the inventors surprisingly discovered the occurrence of joint lesions is significantly reduced in swine fed a daily amount of nitrate compared to swine fed a conventional feed. As used herein, “joint lesions” refers to swelling and puffiness around the joints of the pigs. While not all pigs that have joint lesions will have lameness, the likeness of lameness in pigs increases with increasing and persistent occurrences of joint lesions. Described herein are methods for improving bone development and reducing lameness in swine.

[0037] In one aspect, the present technology provides a method for method of feeding swine, the method includes: feeding swine a swine feed that includes about 0.01 weight percent (wt%) to about 0.5 wt% per day of a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant that includes polyphenols; wherein swine fed the swine feed exhibit a reduced occurrence of joint lesions compared to a control swine not fed the swine feed. In any aspect, reference to weight percent (wt%) per day is based on total weight of swine fed as part of the swine’s daily feed intake.

[0038] Generally, swine may be in one or more of a post- weaning, growth, or finishing phase encompasses gilts, barrows, boars, or combinations thereof. During swine production, gilts may be housed separately from barrows and/or boars during post-weaning, growth, and/or finishing phases. In any aspect, the swine may include gilts, barrows, boars, or combinations thereof. In any aspect, the swine may be gilts. In any aspect, the swine may be barrows. In any aspect, the swine may be boars. In any aspect, the swine may be a reproducing swine. For example, the reproducing swine may be a sow (such as nullparous sow, primiparous sow, or multiparous sow).

[0039] In any aspect, the swine may be fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof during a growth phase, finishing phase, or combination thereof. In any aspect, the swine may be fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof during a growth phase. In any aspect, the swine may be fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof during a finishing phase. [0040] In any aspect, the swine may be fed a daily swine feed intake that includes an amount of nitrate in a range of about 0.01% by weight to about 0.50% by weight based on total weight of a swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be in a range of about 0.02% by weight to about 0.35% by weight based on total weight of the swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be about 0.03% by weight to about 0.25% by weight based on total weight of the swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be about 0.05% by weight to about 0.2% by weight based on total weight of the swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be about 0.07% by weight to about 0.15% by weight based on total weight of the swine feed. For example, suitable amounts of nitrate compound present in the swine’s daily feed intake may be about 0.01wt%, about 0.015 wt%, about 0.02 wt%, about 0.025 wt%, about 0.03 wt%, about 0.035 wt%, about 0.04 wt%, about 0.045 wt%, about 0.05 wt%, about 0.055 wt%, about 0.06 wt%, about 0.065 wt%, about 0.07 wt%, about 0.075 wt%, about 0.08 wt%, about 0.085 wt%, about 0.09 wt%, about 0.1 wt%, about 0.15 wt%, about 0.2 wt%, about 0.25 wt%, about 0.3 wt%, about 0.35 wt%, about 0.4 wt%, about 0.45 wt%, about 0.5 wt%, or any range including and/or in between any two of the preceding values.

[0041] In any aspect, the nitrate compound may include an ionic nitrate salt as described herein. For example, the nitrate compound may include an ionic nitrate salt selected from sodium nitrate, potassium nitrate, calcium nitrate, magnesium nitrate ammonium nitrate, or mixtures thereof. In any aspect, the nitrate compound may be calcium nitrate, complex inorganic salts of calcium nitrate, hydrates thereof (e.g., calcium nitrate dihydrate, calcium nitrate tetrahydrate, calcium nitrate decahydrate), or mixtures thereof. Preferably, the nitrate compound may be calcium ammonium nitrates, or hydrates thereof (e.g. ₄ 5Ca(NO3)2*NH4NO3*10H2O).

[0042] In any aspect, the nitrate compound may also be provided by a variety of plant ingredients according to alternative aspects. For example, such plant ingredients may include, but are not limited to, leafy greens such as spinach, arugula, beetroot, or mixtures thereof. Preferably, the plant ingredient is beetroot. Beetroot has an inorganic nitrate content typically ranging from 110 to 3670 mg nitrate/kg.

[0043] In any aspect, the method reduces the occurrence of joint lesions in swine fed the swine feed by up to about 65% compared to a control swine not fed the swine feed. In any aspect, the method reduces the occurrence of joint lesions in swine fed the swine feed by at least about 1% to about 65% compared to a control swine not fed the swine feed. In any aspect, the method reduces the occurrence of joint lesions in swine fed the swine feed by about 5% to about 50% compared to a control swine not fed the swine feed. In any aspect, the method reduces the occurrence of joint lesions in swine fed the swine feed by about 40% to about 60% compared to a control swine not fed the swine feed. For example, the method may reduce the occurrence of joint lesions in swine fed the swine feed by at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40% and up to about 65%, 64%, 63%, 62%, 61%, 60%, 59%, 58%, 57%, 56%, 55%, 54%, 53%, 52%, 51%, 50%, 49%, 48%, 47%, 46%, or up to about 45%, or any range including and/or in between any two of the preceding values.

[0044] In any aspect, the method may reduce the occurrence of lameness in swine. [0045] The method may further include feeding the swine antioxidants that include polyphenols (e.g., quercetin). In any aspect, the swine feed may further include antioxidants that include the polyphenols. Polyphenols are known to those of skill in the art for their free radical scavenging function. Without being bound by any particular theory, it is believed that dietary polyphenols at gastric pH may enhance the formation of nitric oxide. (Rocha et al., Food Funct., 2014, 5: 1646-1652). For example, it has been reported that dietary polyphenols activate the endothelial nitric oxide synthase pathway, which may contribute to the formation of nitric oxide. (Vilahur et al., Rev. Esp. Cardiol., 2015, 68[3]:216-225).

[0046] Suitable sources of polyphenols, as described herein, may include polyphenols extracted from fruits, vegetables, legumes, nuts, seeds, tea, herbs, spices, tree barks, or mixtures thereof. In any aspect, the polyphenols may be from polyphenol sources including onion extract, grape seed extract, grape skin extract, rosemary extract, or mixtures thereof. For example, the polyphenols may be PROVIOX 50 (Cargill, Incorporated).

[0047] The polyphenols as described herein may be fed to the swine in an amount of about 1 ppm to about 500 ppm per day. Suitable amounts of the polyphenols may include about 1 ppm to about 500 ppm, about 5 ppm to about 500 ppm, about 10 ppm to about 500 ppm, about 50 ppm to about 300 ppm, about 75 ppm to about 250 ppm, about 90 ppm to about 150 ppm, or any range including and/or in between any two of the preceding values.

[0048] In any aspect, the antioxidants may further include natural antioxidants, other than polyphenols. For example, the antioxidants may include vitamins, minerals, carotenoids, or mixtures thereof.

[0049] Vitamins may include, but are not limited to, vitamin A (including vitamin A sources such as vitamin A supplement, vitamin A oil, etc.) vitamin Bl, vitamin B2, vitamin B3, vitamin B4, vitamin B5, vitamin B6, vitamin B7, vitamin B8, vitamin B9, vitamin B 12, vitamin C, Vitamin D, (including vitamin D sources such as vitamin D supplement, etc.), vitamin E (including vitamin E sources such as Vitamin E supplement), vitamin K, and other vitamin product ingredients (including riboflavin, vitamin D3 supplement, niacin, betaine, choline chloride, tocopherol, inositol, etc.). In any aspect, the vitamins may preferably be vitamins E, vitamins C, or combinations thereof.

[0050] Minerals may include, for example and without limitation, calcium, chlorine (as chloride ions), magnesium, phosphorus, potassium, sodium, sulfur, cobalt, copper, chromium, iodine, manganese, molybdenum, nickel, selenium vanadium, zinc, or mixtures thereof. In any aspect, the minerals may preferably be selenium, zinc, manganese, copper, or mixtures thereof. [0051] In another aspect, the present technology provides a method for improving bone development in swine, the method comprising: feeding swine a swine feed, wherein the swine feed comprises: a nitrate compound, hydrates thereof, salts thereof, or combinations thereof; and optionally an antioxidant comprising polyphenols; wherein: the swine is fed a daily feed intake comprising an amount of nitrate in a range of about 0.01% by weight to less than about 0.5% by weight based on total weight of the swine feed; the method improves bone mineral deposition in swine fed the swine feed compared to a control swine not fed the swine feed.

[0052] In any aspect, the swine may be fed a daily swine feed intake that includes an amount of nitrate in a range of about 0.01% by weight to about 0.50% by weight based on total weight of a swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be in a range of about 0.02% by weight to about 0.35% by weight based on total weight of the swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be about 0.03% by weight to about 0.25% by weight based on total weight of the swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be about 0.05% by weight to about 0.2% by weight based on total weight of the swine feed. In any aspect, the daily feed intake amount of nitrate fed to the swine may be about 0.07% by weight to about 0.15% by weight based on total weight of the swine feed. For example, suitable amounts of nitrate compound present in the swine’s daily feed intake may be about 0.01wt%, about 0.015 wt%, about 0.02 wt%, about 0.025 wt%, about 0.03 wt%, about 0.035 wt%, about 0.04 wt%, about 0.045 wt%, about 0.05 wt%, about 0.055 wt%, about 0.06 wt%, about 0.065 wt%, about 0.07 wt%, about 0.075 wt%, about 0.08 wt%, about 0.085 wt%, about 0.09 wt%, about 0.1 wt%, about 0.15 wt%, about 0.2 wt%, about 0.25 wt%, about 0.3 wt%, about 0.35 wt%, about 0.4 wt%, about 0.45 wt%, about 0.5 wt%, or any range including and/or in between any two of the preceding values.

[0053] In any aspect, the nitrate compound may include an ionic nitrate salt as described herein. For example, the nitrate compound may include an ionic nitrate salt selected from sodium nitrate, potassium nitrate, calcium nitrate, magnesium nitrate ammonium nitrate, or mixtures thereof. In any aspect, the nitrate compound may be calcium nitrate, complex inorganic salts of calcium nitrate, hydrates thereof (e.g., calcium nitrate dihydrate, calcium nitrate tetrahydrate, calcium nitrate decahydrate), or mixtures thereof. Preferably, the nitrate compound may be calcium ammonium nitrates, or hydrates thereof (e.g. ₄ 5Ca(NO3)2*NH4NO3*10H2O). [0054] In any aspect, the nitrate compound may also be provided by a variety of plant ingredients according to alternative aspects. For example, such plant ingredients may include, but are not limited to, leafy greens such as spinach, arugula, beetroot, or mixtures thereof. Preferably, the plant ingredient is beetroot. Beetroot has an inorganic nitrate content typically ranging from 110 to 3670 mg nitrate/kg.

[0055] The method of the present technology may increase bone mineral deposition in swine fed the swine feed, as described herein in any aspect, compared to a control swine not fed the swine feed. In any aspect, bone mineral deposition may be measured as metacarpals bone ash content. For example, bone mineral deposition may be measured as metacarpals bone ash content after slaughter.

[0056] In any aspect, the method increases bone mineral deposition in swine fed the swine feed by at least about 0.5% compared to a control swine not fed the swine feed. In any aspect, the method may increase bone mineral deposition in swine fed the swine feed by at least about 0.5% to about 10% compared to a control swine not fed the swine feed. In any aspect, the method may increase bone mineral deposition in swine fed the swine feed by about 1% to about 8% compared to a control swine not fed the swine feed. In any aspect, the method may increase bone mineral deposition in swine fed the swine feed by about 4% to about 7% compared to a control swine not fed the swine feed. For example, the method may increase bone mineral deposition in swine fed the swine feed by at least about 0.05%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or up to about 10%, or any range including and/or in between any two of the preceding values.

[0057] In any aspect, the method may further reduce the occurrence of or prevent lameness in swine fed the swine feed compared to a control swine not fed the swine feed. [0058] In any aspect, the swine feed may be a premix, a feed additive, a feed supplement, a compound feed, or a complete feed as described herein. In any aspect, the swine feed may be a composition that includes an amount of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof fed to the swine before, during, or after providing the swine a feed that does not include nitrate. For example, the swine feed comprising the nitrate compound, hydrates thereof, salts thereof, or combinations thereof may be provided in a supplement separately from the feed or is provided in a supplement that is mixed with the feed. As would be understood by a person skilled in the art, the amount of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof would typically be different from the final swine feed, z.e., the supplement may include a higher concentration of the nitrate compound compared to the final swine feed itself such that the daily feed intake of the swine fed the final swine feed is from about 0.01% by weight to about 0.5% by weight based on total weight of the swine feed.

[0059] The method may further include feeding the swine antioxidants that include polyphenols (e.g., quercetin). Polyphenols are known to those of skill in the art for their free radical scavenging function. Suitable sources of polyphenols, as described herein, may include polyphenols extracted from fruits, vegetables, legumes, nuts, seeds, tea, herbs, spices, tree barks, or mixtures thereof. In any aspect, the polyphenols may be from polyphenol sources including onion extract, grape seed extract, grape skin extract, rosemary extract, or mixtures thereof. For example, the polyphenols may be PROVIOX 50 (Cargill, Incorporated).

[0060] The polyphenols as described herein may be fed to the swine as part of the swine feed in an amount of about 1 ppm to about 500 ppm per day. Suitable amounts of the polyphenols may include about 1 ppm to about 500 ppm, about 5 ppm to about 500 ppm, about 10 ppm to about 500 ppm, about 50 ppm to about 300 ppm, about 75 ppm to about 250 ppm, about 90 ppm to about 150 ppm, or any range including and/or in between any two of the preceding values.

[0061] In any aspect, the antioxidants may further include natural antioxidants, other than polyphenols. For example, the antioxidants may include vitamins, minerals, carotenoids, or mixtures thereof.

[0062] Vitamins may include, but are not limited to, vitamin A (including vitamin A sources such as vitamin A supplement, vitamin A oil, etc.) vitamin Bl, vitamin B2, vitamin B3, vitamin B4, vitamin B5, vitamin B6, vitamin B7, vitamin B8, vitamin B9, vitamin B 12, vitamin C, Vitamin D, (including vitamin D sources such as vitamin D supplement, etc.), vitamin E (including vitamin E sources such as Vitamin E supplement), vitamin K, and other vitamin product ingredients (including riboflavin, vitamin D3 supplement, niacin, betaine, choline chloride, tocopherol, inositol, etc.). In any aspect, the vitamins may preferably be vitamins E, vitamins C, or combinations thereof.

[0063] Minerals may include, for example and without limitation, calcium, chlorine (as chloride ions), magnesium, phosphorus, potassium, sodium, sulfur, cobalt, copper, chromium, iodine, manganese, molybdenum, nickel, selenium vanadium, zinc, or mixtures thereof. In any aspect, the minerals may preferably be selenium, zinc, manganese, copper, or mixtures thereof. [0064] In any aspect, the swine may include gilts, barrows, boars, or combinations thereof. In any aspect, the swine may be gilts. In any aspect, the swine may be barrows. In any aspect, the swine may be boars. In any aspect, the swine may be a reproducing swine. For example, the reproducing swine may be a sow (such as nullparous sow, primiparous sow, or multiparous sow).

[0065] In any aspect, the swine may be fed the swine feed during a growth phase, finishing phase, or combination thereof. In any aspect, the swine may be fed the feed during a growth phase. In any aspect, the swine may be fed the feed during a finishing phase.

[0066] In another aspect, the present technology provides the use of a swine feed as described herein in any aspect for increasing bone mineral deposition and/or reducing the occurrence of joint lesions in swine compared to a control swine not fed the swine feed, wherein the swine feed comprises a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant that include polyphenols, and wherein the daily feed intake amount of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof is about 0.01% by weight to less than about 0.5% by weight based on total weight of the swine feed.

[0067] The present invention, thus generally described, will be understood more readily by reference to the following examples, which are provided by way of illustration and are not intended to be limiting of the present invention.

[0068] EXEMPLARY ASPECTS

[0069] The following exemplary aspects are provided, the numbering of which is not to be construed as designating levels of importance:

[0070] Para. A: A method of feeding swine, the method comprising: feeding swine a swine feed comprising about 0.01 weight percent (wt%) to about 0.5 wt% per day of a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant comprising polyphenols; wherein swine fed the swine feed exhibit a reduced occurrence of joint lesions compared to a control swine not fed the swine feed.

[0071] Para. B: The method of Para. A, wherein the swine is in one or more of a postweaning phase, growth phase, or finishing phase.

[0072] Para. C: The method of Para. A or B, wherein the swine comprises gilts, barrows, boars, or combinations thereof.

[0073] Para. D: The method of Para. A or B, wherein the swine comprises gilts.

[0074] Para. E: The method of Para. A or B, wherein the swine comprises barrows or boars.

[0075] Para. F: The method of any one of Paras. A-E, wherein the swine is in a growth phase or a finishing phase.

[0076] Para. G: The method of Para. A or B, wherein the swine comprises reproducing swine.

[0077] Para. H: The method of Para. G, wherein the reproducing swine is a sow.

[0078] Para. I: The method of any one of Paras. A-H, wherein the daily feed diet comprises about 0.05 wt% to about 0.2 wt% per day of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0079] Para. J: The method of any one of Paras. A-I, comprising feeding the swine about 0.07 wt% to about 0.15 wt% per day of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0080] Para. K: The method of any one of Paras. A- J, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises ionic nitrate salts.

[0081] Para. L: The method of any one of Paras. A-K, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises sodium nitrate, potassium nitrate, calcium nitrate, magnesium nitrate, ammonium nitrate, or mixtures thereof. [0082] Para. M: The method of any one of Paras. A-L, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises calcium nitrate, complex inorganic salts of calcium nitrate, or mixtures thereof.

[0083] Para. N: The method of any one of Paras. A-M, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises calcium ammonium nitrate, pentacalcium ammonium nitrate decahydrate, or combinations thereof.

[0084] Para. O: The method of any one of Paras. A-N, wherein the method reduces joint lesions in swine by up to about 65% compared to a control swine not fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0085] Para. P: The method of any one of Paras. A-O, wherein the method reduces joint lesions in swine by about 1% to up to about 30% compared to a control swine not fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0086] Para. Q: The method of any one of Paras. A-O, wherein the method reduces joint lesions in swine by about 5% to about 25% compared to a control swine not fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0087] Para. R: The method of any one of Paras. A-O, wherein the method reduces joint lesions in swine by about 15% to about 25% compared to a control swine not fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0088] Para. S: The method of any one of Paras. A-R further comprising feeding the swine about 1 ppm to about 500 ppm per day of an antioxidant comprising polyphenols based on total weight of the swine feed.

[0089] Para. T: The method of Para. S, wherein the polyphenols comprise polyphenols sources selected from the group consisting of onion extract, grape seed extract, grape skin extract, rosemary extract, and mixtures thereof.

[0090] Para. U: A method for improving bone development in swine, the method comprising: feeding swine a swine feed, wherein the swine feed comprises: a nitrate compound, hydrates thereof, salts thereof, or combinations thereof; and optionally an antioxidant comprising polyphenols as part of the feed; wherein: the swine is fed a daily feed intake comprising an amount of nitrate is in a range of about 0.01% by weight to less than about 0.50% by weight based on total weight of the swine feed; the method increases bone mineral deposition in swine fed the swine feed compared to a control swine not fed the swine feed.

[0091] Para. V: The method of Para. U, wherein the daily feed intake amount of nitrate is in a range of about 0.05% by weight to about 0.25% by weight of based on total weight of a swine feed.

[0092] Para. W: The method of Para. U or V, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises ionic nitrate salts.

[0093] Para. X: The method of any one of Paras. U-W, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises sodium nitrate, potassium nitrate, calcium nitrate, magnesium nitrate, ammonium nitrate, or mixtures thereof.

[0094] Para. Y: The method of any one of Paras. U-X, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises calcium nitrate, complex inorganic salts of calcium nitrate, or mixtures thereof.

[0095] Para. Z: The method of any one of Paras. U-Y, wherein the nitrate compound, hydrates thereof, salts thereof, or combinations thereof comprises calcium ammonium nitrate, pentacalcium ammonium nitrate decahydrate, or combinations thereof.

[0096] Para. AA: The method of any one of Paras. U-Z, wherein the method increases bone mineralization in the swine by at least about 0.5% compared to a control swine not fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0097] Para. AB: The method of any one of Paras. U-AA, wherein the method improves the feed to gain ratio of the swine by about 0.5% to about 10% compared to a control swine not fed the nitrate compound, hydrates thereof, salts thereof, or combinations thereof.

[0098] Para. AC: The method of any one of Paras. U-AB, wherein bone mineralization in swine is measured as total amount of metacarpals bone ash content present after slaughter. [0099] Para. AD: The method of any one of Paras. U-AC further comprising feeding the swine about 1 ppm to about 500 ppm per day of an antioxidant comprising polyphenols based on total weigh of the swine feed.

[0100] Para. AE: The method of Para. AD, wherein the polyphenols comprise polyphenols sources selected from the group consisting of onion extract, grape seed extract, grape skin extract, rosemary extract, and mixtures thereof.

[0101] Para. AF: The method of any one of Paras. U-AE, wherein the swine is in a growth phase or a finishing phase.

[0102] Para. AG: The method of any one of Paras. U-AF, wherein the swine comprises gilts, barrows, boars, or combinations thereof.

[0103] Para. AH: The method of any one of Paras. U-AF, wherein the swine feed is a premix, a feed additive, a feed supplement, a compound feed, or a complete feed.

[0104] Para. Al: The method of any one of Paras. A-T or U-AH, wherein the method reduces the occurrence of or prevents lameness in swine fed the swine feed compared to a control swine not fed the swine feed.

[0105] Para. AJ: Use of a swine feed for increasing bone mineral deposition in swine fed the swine feed compared to a control swine not fed the swine feed, wherein the swine feed comprises a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant comprising polyphenols, and wherein the daily feed intake amount of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof is about 0.01% by weight to less than about 0.5% by weight based on total weight of the swine feed.

[0106] Para. AK: Use of a swine feed for reducing joint lesions in swine fed the swine feed compared to a control swine not fed the swine feed, wherein the swine feed comprises a nitrate compound, hydrates thereof, salts thereof, or combinations thereof, and optionally an antioxidant comprising polyphenols, and wherein the daily feed intake amount of the nitrate compound, hydrates thereof, salts thereof, or combinations thereof is about 0.01% by weight to less than about 0.5% by weight based on total weight of the swine feed. EXAMPLES

Example 1: Calcium Nitrate Safety Study Trial.

[0107] Study Design. Forty-eight pigs (119.1 ± 5.3 kg) were stratified by weight and sex and randomly allotted to one of four treatments (n=12) in a randomized complete block design.

[0108] Materials and Methods. Diets were formulated to meet or exceed nutrient requirements determined by the Cargill Nutrition System (Table 1).

Table 1. Experimental diets and expected nutrient analysis.

Provided per kilogram of diet: vitamin A, 5,000 IU; vitamin D3, 800 IU; vitamin E, 50 IU; vitamin B12, 17.5 pg; menadione, 1.75 mg; folic acid, 0.55 mg; niacin, 25 mg, pantothenic acid, 30 mg; pyroxidine, 1 mg; riboflavin, 4 mg; thiamine, 0.75 mg; copper, 12.6 mg; iron, 77 mg; iodine, 0.49 mg; manganese, 35 mg; zinc, 77 mg; selenium 0.21 mg; phytase, 391 FTU.

² CP, crude protein; NE, Net Energy; SID, Standardized Ileal Digestible; Ca, calcium; P, phosphorus; Dig, digestible

[0109] Experimental diets were made by inclusion of calcium nitrate (CAS # 10124-37- 5; ADOB, Ponzan, PL) at the expense of com and provided as pellets (4 mm). Calcium nitrate was included at 0%, 0.08%, 0.16% and 0.32% of the diet which is equivalent to 0 wt%, 0.05 wt%, 0.10 wt% and 0.20 wt% (0 ppm, 500 ppm, 1000 ppm, and 2000 ppm) supplemental dietary nitrate inclusion, respectively. Experimental diets were fed ad libitum for 28 days. Bodyweights were measured at dO and d28.

[0110] At the end of the 28 day feeding experiment, animals were fasted 14-16 hours overnight. On the morning of day 29, animals were euthanized by penetrating captive bolt and exsanguination. The front left hoofs above the metacarpals were removed, frozen on dry ice, and stored at -20°C.

[0111] Bone ash was measured at the University of Wisconsin (Madison, WI) according to the method of Adeola et al. (2015). Briefly, the hooves were thawed, and the 3rd metacarpal was dissected out and cleaned. Bones were dried, defatted with petroleum ether, and dried again. Thereafter, bones were ashed in a muffle furnace. Ash weight was recorded as percent of dry fat free bone.

[0112] Statistical Methods. All data were analyzed in R (version 4.1.1). Data was analyzed as a general linear model with treatment as the fixed effect and weight block and sex as random effects. Linear and Quadratic contrasts were constructed to determine the effect of nitrate dose on variables of interest. Statistical relevance was defined when the contrasts -value was less than 0.05.

[0113] Results and Discussion. The results of this study indicated that inclusion of dietary nitrate at 0.1 wt% and 0.2 wt% of the daily feed intake increased the bone ash of pigs consuming nitrate for 28 days (Table 2).

Table 2. Effect of Dietary Nitrate on Fat-Free Bone Ash.

Example 2: Evaluation of Dietary Nitrate Effect on Bone Mineralization and Joint Lesion Occurrence.

[0114] Study Design'.

456 gilts and boars starting at 87 days of age, 40 kg live weight , ending at 187 days of age with a weight around -132 kg. 2 phase feeding program : 40-100 kg with 16% crude protein, 1.02% total lysine, and 1983 kcal/kg net energy; 100 kg -Slaughter with 14.5% crude protein, 0.90% total lysine, and 1945 kcal/kg (gilts) or 1856 kcal/kg (boars) net energy.

12 pigs per pen, separated genders.

10 pens of boars and 9 pens of gilts per treatment.

2 groups : control and treatment, same wheat and barley based basal diet.

Treatment group fed with a premixture containing polyphenols (PROVIOX) and the equivalent of 0.19% calcium nitrate (-0.12 wt% nitrate) in complete feed.

Monitoring: metacarpals bone ash content at -130 kg, joint lesions indicating joints inflammation at -40, -100, and -130 kg.

[0115] Bone Ash Results. Following a similar procedure for metacarpals bone ash content analysis described in Example 1, samples were collected from pigs in both treatment groups after slaughter and evaluated for metacarpals bone ash content. Pigs in the nitrate (0.12 wt%) treatment group showed a 6.31% increase in metacarpals bone ash content compared to pigs fed the control diet (49.67 vs. 46.72%, respectively, P=0.043).

[0116] Joint Lesion Results. Reductions in joint inflammation symptoms were measured as probability of observing joint lesions (observed joint lesions/total pigs) total occurrence of joint lesions at 40, 100, and 130 kg BW, gilts and boars. Inclusion of 0.12 wt% nitrate reduced the probability of observing joint lesions for the entirety of the trial, including a 24% reduction at 100 kg-

[0117] Discussion. The data suggests that pigs fed a combination of calcium nitrate and polyphenols exhibit a reduced risk of developing lameness, and, in particular, the pigs fed a swine feed in keeping with the present invention exhibited a reduction of joint lesions (less inflammation) and increased bone mineral deposition.

[0118] The Example 2 demonstrates nitrate increases ash content of bones to a larger extent than Example 1 when fed to pigs at 0.12% dietary nitrate for 100 days (-40 - 132 kg). Pigs were fed during 28 days in the trial described in Example 1. The difference between Example 1 and Example 2 is believed to be due to a difference in the length of feeding or feeding age, and/or a synergistic effect of the dietary polyphenols contained in PROVIOX that were added to the nitrate source in Example 2. Prophetic Example: Evaluation of Dietary Nitrate Effect on Bone Mineral Deposition and Nitrate Toxicity.

[0119] The proposed trial design will include six treatments, plus a negative control with 12 replicates per treatment and 13 pigs per replicate — a total of 936 pigs will receive a swine feed that includes calcium nitrate. The study will evaluate three levels of calcium nitrate supplementation, beginning at either weaning or the end of the nursery phase of production. Pigs will be allotted to treatment based on weight and sex at weaning. Supplemental dietary nitrate levels being considered are 400 ppm, 600 ppm, and 1200 ppm. Animals will be harvested at typical commercial weights. Diets will be formulated to meet or exceed nutrient requirements determined by the Cargill Nutrition System as appropriate for the growth phase. Experimental diets will be made by inclusion of Calcium Nitrate (CAS # 10124-37-5; ADOB, Ponzan, PL) at the expense of corn and provided as meal.

[0120] Samples of bones (front left hoofs above the metacarpals) will be collected at harvest for bone ash analysis as described in Example 1.

[0121] References.

1. Adeola O. et al.. A cooperative study on the standardized total -tract digestible phosphorus requirement of twenty-kilogram pigs. J Anim Sci. 2015. (12):5743-53.

[0122] The present technology is also not to be limited in terms of the particular aspects described herein, which are intended as single illustrations. Many modifications and variations of this present technology can be made without departing from its spirit and scope, as will be apparent to those skilled in the art from the foregoing descriptions. Such modifications and variations are intended to fall within the scope of the appended claims. It is to be understood that this present technology is not limited to particular methods, reagents, compounds, or compositions, which can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only, and is not intended to be limiting. Thus, it is intended that the specification be considered as exemplary only with the breadth, scope and spirit of the present technology indicated only by the appended claims, definitions therein and any equivalents thereof. [0123] The aspects, illustratively described herein may suitably be practiced in the absence of any element or elements, limitation or limitations, not specifically disclosed herein. Thus, for example, the terms “comprising,” “including,” “containing,” etc. shall be read expansively and without limitations. Additionally, the terms and expressions employed herein have been used as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the claimed technology. Additionally, the phrase “consisting essentially of’ will be understood to include those elements specifically recited and those additional elements that do not materially affect the basic and novel characteristics of the claimed technology. The phrase “consisting of’ excludes any element not specified.