Chondroprotective dietary supplement compositions and methods of use thereof

Abstract

A chondroprotective dietary supplement composition and method are disclosed that can increase calcium absorption and retention in mammalian skeletal structure. The chondroprotective dietary supplement composition contains collagen. In addition to improving calcium intake, the chondroprotective dietary supplement composition of the present disclosure can also improve various other bone health markers. The composition can be used in the form of a supplement to prevent bone disease and / or treat bone disease.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to the benefit of U.S. Provisional Patent Application No. 63 / 257,170, filed October 19, 2021, which is expressly incorporated by reference herein in its entirety. [Background technology]

[0002] The mammalian skeleton provides support, allows walking, sitting, and bending, and also protects the brain and various organs housed within the body. Poor bone health can affect all different types of mammals, including dogs, livestock, and humans. However, poor bone health can be difficult to detect in that individuals usually do not notice the problem until stress is placed on the bone, which causes a fracture or break. Especially as mammals age, bones can wear down faster than the body can repair them, which can lead to a fragile bone structure that can occur without any noticeable side effects until the condition causes an injury.

[0003] For example, humans can lose up to 2% of bone density each year after reaching age 50. Loss of bone density can lead to osteoporosis. Osteoporosis is a disabling disease characterized by reduced bone mass and microarchitectural deterioration of the skeletal structure, leading to reduced bone strength and predisposing mammals, such as humans, to an increased risk of fragility fractures. Osteoporosis affects over 70 million people in the United States, Europe, and Japan alone, with millions of patients suffering bone fractures each year. Osteoporosis can affect over 25% of all postmenopausal women, increasing as the population ages. For example, over 30% of all women over the age of 50 will have an osteoporotic fracture. Older men are similarly affected by osteoporosis, which can lead to multiple hip fractures.

[0004] Osteoporosis can be caused by a variety of different factors, including metabolic changes in the body combined with a diet low in bone-healthy nutrients. Osteoporosis can also occur as a result of medication or as a side effect of contracting a disease. Indeed, the long-term effects of the COVID-19 pandemic on bone health are still unknown.

[0005] Throughout life, bone is continually remodeled by resorption of old bone (catabolic process) performed by osteoclasts and deposition of new bone (anabolic process) performed by osteoblasts. Bone remodeling is not a random process but takes place in local bone multicellular units (BMUs), remodeling units that contain osteoblasts, osteoclasts, and their precursors, where resorption and formation are coupled. Bone resorption is likely the initial event that occurs in response to local mechanical stress signals. The loss of bone density seen in osteoporosis is due to an imbalance between resorption and formation, where the rate of resorption exceeds the rate of formation. Osteoporosis represents a continuum in which multiple pathogenic mechanisms converge to cause loss of bone mass and microstructural deterioration of skeletal structure. Osteoporosis is likely caused by complex interactions between local and systemic regulators of bone cell function. Heterogeneity in osteoporosis may be due to differences in the production of systemic and local regulators, as well as alterations in receptors, signaling mechanisms, nuclear transcription factors, and enzymes that produce or inactivate the local regulators.

[0006] Bone strength reflects the integration of two major features: bone density and bone quality. Bone density is expressed as grams of mineral per area or volume and is determined in any given individual by the peak bone mass reached and the amount of bone loss thereafter. Bone quality refers to structure, turnover, accumulation of damage (i.e., microfractures) and mineralization. Fractures occur frequently when trauma is applied to osteoporotic bones with low bone density. Osteoporosis is therefore a significant risk factor for fractures.

[0007] As noted above, osteoporosis can be difficult to diagnose. Typically, osteoporosis is detected by bone mineral density testing. However, most affected patients do not realize that they have low bone density or are at risk until they suffer a fracture.

[0008] In view of the above, there remains a need in the art for methods and supplements to prevent the onset of bone diseases such as osteoporosis. In another aspect, there is also a need for methods and supplements that can improve bone and joint health, including tendon and ligament health associated with the mammalian skeletal system. Summary of the Invention

[0009] In general, the present disclosure relates to a chondroprotective dietary supplement composition that can increase the absorption of at least one bone health additive into the skeletal system of a mammal, including associated tendons and ligaments. The mammal may be, but is not limited to, a human, a dog, a cat, a cow, a horse, etc. The present disclosure is also directed to a method for increasing the absorption of at least nutrients, such as bone health additives, into the bones, tendons, and ligaments of a mammal by administering a therapeutically effective amount and / or a nutritionally effective amount of the chondroprotective dietary supplement composition to the mammal. The bone health additive may already be present in the mammal's diet and / or may be added in conjunction with the chondroprotective dietary supplement composition. According to the present disclosure, the chondroprotective dietary supplement composition contains at least one collagen source.

[0010] For example, in one embodiment, the present disclosure is directed to a method for increasing calcium absorption in a mammal. The mammal can be fed a diet that includes calcium. The method includes administering to the mammal a therapeutically effective amount of a chondroprotective nutraceutical composition sufficient to increase calcium absorption in the mammal. The chondroprotective nutraceutical composition can include type II collagen.

[0011] The chondroprotective nutraceutical composition can be administered to a mammal to increase or improve various bone health parameters. For example, the chondroprotective nutraceutical composition can be administered to a mammal in an amount sufficient to increase the percent absorption and / or percent retention of calcium in the mammal's diet. In addition to calcium, the chondroprotective nutraceutical composition can also increase the intake of various nutrients, including vitamin D, vitamin K2, magnesium, herbs, and the like. The chondroprotective nutraceutical composition can also be administered to a mammal in an amount sufficient to increase bone weight, such as dry femur weight. The chondroprotective nutraceutical composition can also be administered to a mammal in an amount sufficient to increase various bone health markers, such as an increase in osteocalcin. In another embodiment, the chondroprotective nutraceutical composition can be administered to a mammal in an amount sufficient to increase bone morphogenetic protein 2.

[0012] The composition can contain any suitable collagen source, including type 1, type II, or type III collagen. In one embodiment, the collagen can include undenatured type II collagen. The collagen can be obtained from chicken cartilage, such as chicken sternal cartilage.

[0013] The chondroprotective dietary supplement composition can be administered to a mammal at least once a week, for example at least every three days. In one embodiment, the chondroprotective dietary supplement composition can be administered to a mammal daily. The chondroprotective dietary supplement composition can contain collagen, such as type II collagen, at a dosage level of about 10 mg to about 3,000 mg of collagen. Any suitable delivery form can be used, including capsules, tablets, suspensions, and the like.

[0014] In one embodiment, the method of the present disclosure further comprises the step of administering to the mammal a calcium supplement, which may, for example, be contained within the chondroprotective nutraceutical composition or may be administered separately to the mammal.

[0015] In one embodiment, the method is administered to a mammal having a calcium deficiency in its diet.

[0016] The present disclosure is also directed to a chondroprotective supplement for improving bone health in a mammal, including improving associated cartilage, tendons and ligaments. The chondroprotective supplement comprises a bone health additive in combination with a bone health supplement. The bone health supplement comprises type II collagen, such as undenatured type II collagen. The bone health supplement is present in the supplement in a therapeutically effective amount sufficient to increase absorption of the bone health additive into the skeletal structure or bone of the mammal.

[0017] In one embodiment, the bone health additive comprises calcium. The calcium may be present in the chondroprotective supplement in an amount of, for example, about 250 mg to about 2,500 mg. The calcium may be in any suitable form, such as calcium carbonate, calcium citrate, calcium gluconate, calcium lactate, calcium hydroxyapatite, or mixtures thereof.

[0018] Collagen may be present in the chondroprotective supplement in an amount greater than about 20 mg, such as greater than about 30 mg, such as greater than about 40 mg, such as greater than about 80 mg, such as greater than about 100 mg, and in an amount less than about 1000 mg, such as less than about 350 mg.

[0019] Other features and aspects of the disclosure are discussed in more detail below.

[0020] A full and enabling disclosure of the present invention, including the best mode thereof, directed to one of ordinary skill in the art, is set forth more particularly in the remainder of the specification, which makes reference to the accompanying figures. [Brief description of the drawings]

[0021] [Figure 1] 1 is a graph illustrating some of the results obtained in the examples described below. [Diagram 2] 1 is a graph illustrating some of the results obtained in the examples described below. [Diagram 3] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 4] 1 is a graph illustrating some of the results obtained in the examples described below. [Diagram 5] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 6] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 7] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 8] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 9] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 10] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 11] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 12] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 13] 1 is a graph illustrating some of the results obtained in the examples described below. [Figure 14] 1 is a graph illustrating some of the results obtained in the examples described below.

[0022] Repeat use of reference characters in the present specification and drawings is intended to represent same or analogous features or elements of the invention.

[0023] definition As used herein, the terms "about," "approximately," or "generally," when used to modify a value, indicate that the value can be increased or decreased by 10% in one embodiment, such as 8% in one embodiment, such as 5%, such as 4%, such as 3%, such as 2%, such as 1% and still remain within the range of the disclosed embodiments.

[0024] As used herein, the terms "therapeutically effective amount" and / or "nutraceutical effective amount" shall mean that dosage or amount of a composition that provides a particular pharmacological or nutritional response when the composition is administered or delivered to a mammal in need of such treatment. It is emphasized that an "effective amount" administered to a particular subject in a particular case is not necessarily effective in treating a disease or otherwise improving health as described herein, even if such a dosage is considered to be an "effective amount" by those skilled in the art. Certain subjects may in fact be "refractory" to an "effective amount". For example, refractory subjects may have low bioavailability or genetic variation in certain receptors, metabolic pathways, or response capabilities such that clinical efficacy is not obtained. It is further understood that in certain cases, the composition or supplement may be measured as an oral dosage or in terms of component levels that can be measured in the blood. In other embodiments, the dosage may be measured in the amount applied to the skin when the composition is included with a topical formulation.

[0025] The term "supplement" refers to a product that is in addition to the mammal's normal diet, but may be combined with the mammal's normal food or beverage composition. The supplement may be in any form, including but not limited to solid, liquid, gel, capsule, tablet, or powder. The supplement may also be administered simultaneously with or as a component of a food composition, which may include food, beverage, pet food, snack, or treat. In one embodiment, the beverage may be an activity drink.

[0026] The term "nutraceutical" refers to any compound added to a nutritional source (e.g., a food, beverage, or dietary supplement) that provides a health or medical benefit in addition to its basic nutritional value.

[0027] As used herein, the term "delivering" or "administering" refers to any route for providing a composition, product, or nutritional functional food to a subject, as accepted as standard by the medical community. For example, the present disclosure contemplates routes of delivery or administration including oral ingestion and any other suitable delivery route, including transdermal, intravenous, intraperitoneal, intramuscular, topical, and subcutaneous.

[0028] As used herein, the term "mammal" includes any mammal that can benefit from improved joint health, resilience, and recovery, and may include, but is not limited to, canine, equine, feline, bovine, human, or porcine mammals.

[0029] As used herein, "healthy" refers to the absence of disease or injury.

[0030] Unless otherwise indicated, as used herein, "collagen" refers to all forms of collagen, with or without denaturation, with or without salts or stabilizers, as well as fibrillar and nonfibrillar collagens, including but not limited to fibril-associated collagens with interrupted triple helices (FACIT, IX, XII, XIV, XIX, XXI), including short chain collagens (commonly types VII and X), basement membrane (type IV), Multiplexin (multiple triple helical domains with interruptions) (types XV, XVIII), and other types of collagens (types VI, VII). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0031] Those skilled in the art should understand that this discussion is merely a description of exemplary embodiments and is not intended to limit the broader aspects of the disclosure.

[0032] In general, the present disclosure relates to a chondroprotective dietary supplement composition for improving bone health, including improving the health of cartilage, tendons and ligaments associated with the skeletal system. The present disclosure is also directed to a method of administering the chondroprotective dietary supplement composition to a mammal. In one aspect, the chondroprotective dietary supplement composition can be orally administered to a mammal in the form of a supplement. The chondroprotective dietary supplement composition is, in one embodiment, administered to a mammal in an amount sufficient to increase the absorption of one or more bone health additives into the skeletal system of the mammal. When administered to a mammal in a therapeutically effective amount, for example, the mammal may experience weight gain, increased retention of one or more bone health additives, such as calcium, increased dry bone weight, such as dry femur weight, and the like. The chondroprotective dietary supplement composition of the present disclosure can also increase bone mineral density, increase bone mineral content, and / or increase the amount of calcium in bone.

[0033] The chondroprotective nutraceutical composition of the present disclosure can be administered to healthy mammals and mammals that may have underlying damage or disease of the skeletal system. In addition, the chondroprotective nutraceutical composition is particularly well suited for administration to healthy young adults (e.g., mammals or humans) in the growth stage of bone development. Overall, the chondroprotective nutraceutical composition can enhance the absorption and retention of one or more bone health additives. The composition can also enhance the activity of bioactive additives or bone health additives. The chondroprotective nutraceutical composition, alone or in combination with any mineral or vitamin, can enhance all of these outcomes in healthy and diseased mammalian populations, including mammals in the growth stage of bone development. Conditions that can be treated or prevented according to the present disclosure include co-morbidities such as osteoporosis, any other bone disease, joint disease, inflammatory disease states in bone and joints, diabetes, cardiometabolic syndrome, and / or any other chronic disease.

[0034] The chondroprotective dietary supplement composition of the present disclosure, when administered to a mammal in a therapeutically effective amount, can also positively affect a variety of different bone markers.For example, the mammal can experience an increase in osteocalcin and a decrease in parathyroid hormone.The mammal can also experience an increase in osteoprotegerin, an increase in RANK, which promotes bone formation, and a decrease in RANK-L, which regulates bone resorption.The mammal can also experience a decrease in SOX-9, which is expressed in chondrocytes and is for chondrogenic differentiation.The mammal can also experience an increase in IGF-1, which correlates with bone mineral density, and an increase in bone morphogenetic protein 2, which plays a role in osteoblast differentiation.

[0035] According to the present disclosure, the chondroprotective dietary supplement composition comprises collagen, either alone or in combination with other bone health additives.Although collagen has been found to improve joint health at various concentrations, it was completely unexpected that collagen can have a dramatic and synergistic effect on improving bone health when combined with one or more bone health supplements.One or more bone health additives can be present in the composition, can be present in the mammal's diet, or can be present in both.

[0036] The chondroprotective nutraceutical composition of the present disclosure can be used to prevent or treat a variety of different bone diseases or conditions.For example, the chondroprotective nutraceutical composition of the present disclosure is particularly well suited for administration to mammals that are predisposed to or at risk of developing osteoporosis, osteopenia, bone fractures (including humans such as working animals, horses, athletes, etc.).The composition is also well suited for administration to healthy mammals or humans that are in the growth phase of bone development.The chondroprotective nutraceutical composition of the present disclosure is also well suited for treating mammals that have any of the above bone diseases.For example, the composition can be used to slow or stop the progression of osteoporosis or osteopenia, or to substantially slow or prevent their onset.

[0037] The chondroprotective dietary supplement composition of the present disclosure generally contains a collagen source, which contains collagen alone or in combination with one or more bone health additives. In one embodiment, for example, the chondroprotective dietary supplement composition of the present disclosure does not contain any other bone health additives. Instead, the collagen contained in the composition is synergistically combined with a bone health additive contained in the mammal's diet, such as calcium, to increase the absorption of the bone health additive into the mammal's skeletal system. In this way, the chondroprotective dietary supplement composition of the present disclosure can enhance the activity of various additives already contained in the mammal's diet. For example, the mammal may have a diet that is deficient in one or more bone health additives, such as calcium. The chondroprotective dietary supplement composition of the present disclosure can optimize and maximize the absorption of the bone health additive into the mammal, so that additional amounts of additives are not required in the mammal's diet.

[0038] Alternatively, the cartilage-protecting dietary supplement composition of the present disclosure may comprise collagen in combination with one or more bone health additives.As mentioned above, one bone health additive that may be incorporated into the composition may be an additive that contains calcium.The calcium additive may be any suitable calcium salt, such as, for example, calcium carbonate, calcium citrate, calcium gluconate, calcium lactate, calcium hydroxyapatite, or mixtures thereof.

[0039] Other bone health additives that may be present in the composition include vitamins, such as vitamin D3 and / or vitamin K2, and one or more minerals, either alone or in combination with one or more vitamins. In one embodiment, the bone health composition may be a plant extract including cissus quadrangularis.

[0040] In other embodiments, collagen can be combined with bone resorption inhibitors, anabolic agents, nonsteroidal anti-inflammatory drugs (NSAIDs), any drug for treating osteoarthritis, or combinations thereof. Such drugs include bisphosphonates, denosumab, estrogen, calcitonin, romosozumab, teriparatide, or combinations thereof. Bisphosphonates that may be present include alendronate, ibandronate, risedronate, zoledronic acid, or combinations thereof.

[0041] The chondroprotective dietary supplement compositions of the present disclosure may also contain various other minerals and nutrients, such as a source of magnesium, a source of manganese, and the like.

[0042] When the chondroprotective dietary supplement composition is administered to a mammal in a therapeutically effective amount, it can dramatically and unexpectedly improve the bone health of the mammal by synergistically combining with the bone health additives present in the mammal.Particularly advantageously, it can increase the bone mineral density of the mammal.For example, after a certain period of time, such as 12 weeks, the bone mineral density of the mammal can be at least about 4%, such as at least 8%, such as at least 12%, such as at least 18%, such as at least 22%, such as at least 26%, such as at least 31%, such as at least 35%, and generally less than about 100%, such as less than about 80% (mg / cm 2) may increase. Similarly, the bone mineral content of a mammal may increase by at least about 8%, such as at least about 15%, such as at least about 20%, such as at least about 25%, such as at least about 30%, such as at least about 35%, such as at least about 40%, such as at least about 45%, and generally less than about 110%. Osteocalcin may increase by more than about 10%, such as more than about 15%, such as more than about 25%, such as more than about 35%, such as more than about 45%, such as more than about 55%, such as more than about 65%, such as more than about 75%, such as more than about 85%, and generally less than about 200% over a similar period, such as 12 weeks. On the other hand, parathyroid hormone (PTH) may decrease during administration of the chondroprotective dietary supplement composition. For example, PTH may decrease by more than about 5%, such as more than about 15%, such as more than about 25%, such as more than about 35%, such as more than about 45%. The above percentages can be compared to the mammal before undergoing treatment, or can be compared to the same mammal or a group of the same mammals that have not been administered the chondroprotective nutritional supplement composition.

[0043] It has been found that mammals administered the chondroprotective dietary supplement composition of the present disclosure exhibit a significant increase in calcium absorption and calcium retention when a collagen source is administered to the mammal in combination with calcium, regardless of whether the calcium is part of the mammal's regular diet, is included in the composition administered to the mammal, or both. The amount of absorption of minerals such as calcium is calculated according to the following formula: Absorption (mg / day or μg / day) = intake - fecal excretion Absorption rate (%) = amount absorbed / amount ingested x 100 Mineral retention is calculated as follows: Retention amount (mg / day or μg / day) = absorption amount - urinary excretion amount Retention rate (%) = retention amount / intake amount x 100

[0044] When the chondroprotective nutritional supplement composition is administered to a mammal in a therapeutically effective amount in combination with calcium, calcium absorption and calcium retention can be increased by more than about 15%, such as more than about 20%, for example more than about 25%, such as more than about 30%, for example more than about 35%, such as more than about 40%, for example more than about 45%, such as more than about 50%, for example more than about 55%, such as more than about 60%, for example more than about 65%, for example more than about 70%, relative to the same mammal prior to treatment, or relative to a mammal or group of mammals not administered the chondroprotective nutritional supplement composition.

[0045] In addition, a mammal administered the chondroprotective nutraceutical composition of the present disclosure may show a dramatic increase in femoral dry weight compared to the same mammal or group of mammals not administered the chondroprotective nutraceutical composition.For example, after 12 weeks of administration of the chondroprotective nutraceutical composition in a therapeutically effective amount, femoral dry weight may increase by more than about 10%, for example, more than about 20%, for example, more than about 30%, for example, more than about 40%, for example, more than about 50%, and generally less than about 80%, for example, less than about 70%.

[0046] A variety of different bone markers and / or gene expression markers can also be affected by the chondroprotective nutraceutical composition according to the present disclosure.For example, administering a therapeutically effective amount of a chondroprotective nutraceutical composition to a mammal (relative to the same mammal before treatment, or to a mammal or group of mammals that have not been administered the chondroprotective nutraceutical composition) can increase RANK, which promotes bone formation, by more than about 50%, for example more than about 80%, for example more than about 100%, for example more than about 120%, for example more than about 150%, and generally less than about 250%.RANK-L, a marker of bone resorption, can decrease by at least about 10%, for example at least about 20%, for example at least about 30%, for example at least about 40%, for example at least about 50%. Osteoprotegerin, which protects skeleton from excessive bone resorption, can be increased by at least about 25%, for example, at least about 50%, for example, at least about 75%, for example, at least about 100%, for example, at least about 125%, and generally less than about 300%. SOX-9 is a bone marker associated with sex-determining region Y. It has been found that the chondroprotective dietary supplement composition of the present disclosure can significantly reduce SOX-9 levels in mammals, for example, by more than about 15%, for example, more than about 20%, for example, more than about 25%, for example, more than about 30%, for example, more than about 40%, for example, more than about 50%, and even for example, more than about 55%.

[0047] The chondroprotective nutraceutical composition of the present disclosure can also dramatically affect insulin-like growth factor 1 (IGF-1) and / or bone morphogenetic protein 2 (BMP2) in a mammal. For example, administration of a therapeutically effective amount of the chondroprotective nutraceutical composition to a mammal over a period of time, such as 12 weeks, can increase IGF-1 by more than 100%, such as more than about 150%, such as more than about 200%, such as more than about 250%, and typically less than about 500%, compared to the same mammal not administered the chondroprotective nutraceutical composition.

[0048] Administration of a therapeutically effective amount of the chondroprotective nutritional supplement composition to a mammal over a period of time, such as 12 weeks, can increase BMP2 by at least about 30% or more, such as at least about 50% or more, such as at least about 75% or more, such as at least about 100% or more, such as at least about 125% or more, such as at least about 150% or more, such as at least about 175% or more, and generally less than about 300%, compared to the same mammal not administered the chondroprotective nutritional supplement composition.

[0049] The chondroprotective nutraceutical composition may include any one or more collagens, and / or in one aspect, may include one or more of type I collagen, type II collagen, type III collagen, type IV collagen, or collagen peptides, or a mixture thereof. In one aspect, the chondroprotective nutraceutical composition contains type II collagen alone or in combination with one or more of type I collagen, type III collagen, type IV collagen, or collagen peptides. In one aspect, the chondroprotective nutraceutical composition may include a mixture of type II collagen (sometimes referred to as native type II collagen) and undenatured type II collagen. Additionally or alternatively, the chondroprotective nutraceutical composition may include a mixture of native type II collagen and undenatured type II collagen, in addition to additional collagens such as type I, type III, type IV, or collagen peptides. Furthermore, in one aspect, the chondroprotective nutraceutical composition includes whole collagen protein, biologically active peptide fragments of collagen, or a combination thereof.

[0050] In one aspect, one or more types of collagen in the chondroprotective nutraceutical composition may be heat sterilized, such as by autoclaving, and / or may include salts such as alkalizing or acids, and / or organic or inorganic salts. Thus, in one aspect, a portion of collagen may be at least partially hydrolyzed. In one aspect, collagen is hydrolyzed by any process or compound, including acid-base agents, enzymes, heat or other extreme temperatures, chemicals, UV, salts, or combinations thereof. In one aspect, hydrolyzed collagen is non-denatured collagen that has been at least partially hydrolyzed by any method, and in one aspect, may include non-denatured collagen with any portion that has been denatured. However, as discussed, in one aspect, at least a portion of the chondroprotective nutraceutical composition is non-denatured.

[0051] Type II collagen for use in the present disclosure can be obtained from any suitable source. For example, collagen can be derived from various mammalian sources, avian sources, or can be derived from various fish species or combinations thereof. For example, collagen can be obtained from salmon, shark, poultry, pig, eggshell, turkey cartilage, bovine cartilage, and the like. In one embodiment, for example, type II collagen can be obtained as disclosed in U.S. Patent No. 7,083,820 to Schilling, which is incorporated by reference. For example, undenatured type II collagen is commercially available from InterHealth Nutraceuticals under the UC-II® brand. The UC-II® brand is a natural ingredient that contains glycosylated undenatured type II collagen. The chondroprotective nutraceutical composition can also include hydrolyzed collagen. The chondroprotective nutraceutical composition can also include pure protein or active peptide fragments. In one embodiment, the chondroprotective nutraceutical composition can be free of any bone or bone material. In other embodiments, the chondroprotective nutraceutical composition may be free of any transforming growth factors (TGFs), bone morphogenetic proteins (BMPs), or both. In yet another embodiment, the chondroprotective nutraceutical composition comprises type II collagen and no type I collagen.

[0052] In preparing animal tissue for oral administration, in one embodiment, the collagen type II-containing tissue can first be cut free of surrounding tissue and diced or otherwise ground into particles. The particulate or ground cartilage can be sterilized by means that do not affect or denature the structure of the bulk of the collagen type II in the tissue, such as cryotreatment, and can be formed into doses containing therapeutically effective levels of undenatured collagen type II. Being a natural product, some variation from sample to sample is expected. These variations can be minimized by blending after grinding. Blending can be aided by analytical techniques that allow for the determination of the amount of undenatured collagen type II and other components.

[0053] By forming particles and sterilizing type II collagen as discussed above, undenatured type II collagen can be resistant to gastric acid and digestive enzymes in the stomach.Because of this sterilization process, undenatured type II collagen also retains its three-dimensional shape and preserves the bioactive epitope region.Without wishing to be bound by theory, it is believed that the epitope region contains the ability to induce oral immune tolerance.In particular, the epitope region allows undenatured collagen to bind to Peyer's patches, which have the ability to induce oral immune tolerance process.

[0054] The chondroprotective dietary supplement composition of the present disclosure can be orally administered to mammals in the form of individual dosage containers.Each dose can contain a therapeutically effective amount of collagen based on the type of mammal to be treated, the condition of the mammal, and various other factors.In one aspect, each dose of the chondroprotective dietary supplement composition is at least about 10 mg, such as at least about 20 mg, such as at least about 40 mg, such as at least about 60 mg, such as at least about 80 mg, such as at least about 100 mg, such as at least about 150 mg, such as at least about 200 mg, such as at least about 250 mg, such as at least about 300 mg, such as at least about 350 mg, such as at least about 400 mg, such as at least about 500 mg, such as at least about 600 mg, such as at least about 70 mg. Collagen may be contained in an amount of about 0 mg, such as at least about 800 mg, such as at least about 900 mg, such as at least about 1,000 mg, such as at least about 1,100 mg, such as at least about 1,200 mg, such as at least about 1,300 mg, such as at least about 1,400 mg, such as at least about 1,500 mg, such as at least about 1,600 mg, such as at least about 1,700 mg, such as at least about 1,800 mg, such as at least about 1,900 mg, such as at least about 2,000 mg. Generally, each dose of the chondroprotective dietary supplement composition may contain collagen in an amount of less than about 5,000 mg, such as less than about 3,000 mg, such as less than about 2,000 mg, such as less than about 1,500 mg. It should further be appreciated that in one aspect, the chondroprotective dietary supplement composition may be a type II chondroprotective dietary supplement composition, in which substantially all of the collagen in the chondroprotective dietary supplement composition is type II collagen.

[0055] In one embodiment, the amount of type II chondroprotective dietary supplement composition present in the supplement is based on the type of mammal and / or the body weight of healthy mammal.For example, collagen can be present in each dose in an amount of more than about 0.2mg / kg body weight, such as more than about 0.5mg / kg body weight, such as more than about 1mg / kg body weight, such as more than about 3mg / kg body weight, such as more than about 5mg / kg body weight, and generally less than about 10mg / kg body weight, such as less than about 3mg / kg body weight.

[0056] In one aspect, the undenatured type II collagen can form all or substantially all of the total type II collagen in the chondroprotective nutraceutical composition and thus can be present in the supplement in the amounts discussed above. However, in one aspect, the undenatured type II collagen can comprise from about 0.5% to about 95% of the total type II collagen and / or chondroprotective nutraceutical composition, for example, from about 1% to about 75% of the total type II collagen or the total chondroprotective nutraceutical composition, for example, from about 1.5% to about 50%, for example, from about 2% to about 40%, from about 2.5% to about 15%, or any range or value therebetween.

[0057] In one embodiment, the chondroprotective nutraceutical composition may further comprise a preservative salt, such as potassium chloride. Thus, in one embodiment, the total amount of the chondroprotective nutraceutical composition discussed above may comprise type II collagen and / or non-denatured type II collagen, alone or in combination with additional collagen, a preservative salt, or a combination thereof. For example, in an embodiment, the total type II collagen, including native and non-denatured type II collagen, may comprise about 1% to about 99% of the chondroprotective nutraceutical composition, such as about 2.5% to about 90%, such as about 5% to about 80%, such as about 7.5% to about 70%, such as about 10% to about 60%, such as about 15% to about 50%, such as about 20% to about 35%, or any range or value therebetween.

[0058] In one aspect, when the type II collagen comprises undenatured type II collagen, the undenatured type II collagen may have a high oxygen radical absorbance capacity (ORAC) as measured according to ORAC 6.0. In particular, the ORAC test measures antioxidant scavenging activity against oxygen radicals known to be involved in the pathogenesis of aging and common diseases, and is composed of six types of ORAC assays that evaluate the antioxidant capacity of materials against primary reactive oxygen species, peroxyl radicals, hydroxyl radicals, superoxide anions, and peroxynitrite. In particular, the ORAC assay involves introducing a reactive oxygen species (ROS) introducer into the assay system, which causes the release of specific ROS that degrade the probe and change its emission wavelength or intensity. Thus, when the assay being tested includes an antioxidant, the antioxidant absorbs the ROS and protects the probe from degradation. The degree of probe preservation indicates the antioxidant capacity of the material, and the results are expressed as μmol Trolox equivalents (TE) / g of the test material.

[0059] For example, the ORAC assay for peroxyl radicals measures the antioxidant capacity of a sample to protect a fluorescent protein (fluorescein) from damage by peroxyl radicals generated from 2,2'azobis(2amidinopropane) dihydrochloride (AAPH). The ORAC assay for hydroxyl radicals measures the antioxidant capacity of a sample to protect a fluorescent protein (fluorescein) from damage by hydroxyl radicals generated from the reaction of cobalt with hydrogen peroxide. The ORAC assay for peroxin nitrite measures the antioxidant capacity of a sample to protect dihydrorhodamine-123 from damage by peroxin nitrite radicals generated from 3-morpholinosyndnonimine hydrochloride. The ORAC assay for superoxide measures the antioxidant capacity of a sample to protect hydroethidine from damage by superoxide generated from xanthine oxidase. The ORAC assay for singlet oxygen measures the antioxidant capacity of a sample to protect hydroethidine from damage by singlet oxygen generated from the reaction of lithium molybdate with hydrogen peroxide. Finally, the ORAC assay for hypochlorite measures the antioxidant capacity of a sample to protect the fluorescent protein fluorescein from damage by hypochlorous acid radicals generated from sodium hypochlorite.

[0060] Thus, in one aspect, a chondroprotective nutraceutical composition having undenatured type II collagen according to the present disclosure may have a total ORAC of about 200 μmol TE / g or more, such as about 250 μmol TE / g or more, such as about 300 μmol TE / g or more, such as about 350 μmol TE / g or more, such as about 400 μmol TE / g or more, such as about 450 μmol TE / g or more, such as about 500 μmol TE / g or more, such as about 550 μmol TE / g or more, such as about 600 μmol TE / g or more, such as about 700 μmol TE / g or more, such as about 750 μmol TE / g or more, such as about 800 μmol TE / g or more, such as about 825 μmol TE / g or more, up to about 1000 μmol TE / g, or any range or value therebetween.

[0061] Further, in one aspect, a chondroprotective dietary supplement composition having undenatured type II collagen according to the present disclosure may have an ORAC against peroxyl radicals of about 1 μmol TE / g or more, such as about 2.5 μmol TE / g or more, such as about 5 μmol TE / g or more, such as about 7.5 μmol TE / g or more, such as about 10 μmol TE / g or more, such as up to about 10.5 μmol TE / g or more, up to about 50 μmol TE / g, or any range or value therebetween.

[0062] Similarly, in one aspect, a chondroprotective dietary supplement composition having undenatured type II collagen according to the present disclosure may have an ORAC against hydroxyl radicals of about 10 μmol TE / g or more, such as about 15 μmol TE / g or more, such as about 20 μmol TE / g or more, such as about 25 μmol TE / g or more, such as about 27.5 μmol TE / g or more, such as about 30 μmol TE / g or more, up to about 40 μmol TE / g, or any range or value therebetween.

[0063] Additionally or alternatively, in one aspect, a chondroprotective nutritional supplement composition having undenatured type II collagen according to the present disclosure may have an ORAC to peroxynitrite of about 0.5 μmol TE / g or more, such as about 1 μmol TE / g or more, such as about 1.5 μmol TE / g or more, such as about 2 μmol TE / g or more, such as about 2.25 μmol TE / g or more, up to about 5 μmol TE / g, or any range or value therebetween.

[0064] In one aspect, a chondroprotective dietary supplement composition having undenatured type II collagen according to the present disclosure may have an ORAC to singlet oxygen of about 500 μmol TE / g or more, such as about 550 μmol TE / g or more, such as about 600 μmol TE / g or more, such as about 650 μmol TE / g or more, such as about 700 μmol TE / g or more, such as about 725 μmol TE / g or more, up to about 1000 μmol TE / g, or any range or value therebetween.

[0065] Further, in one aspect, a chondroprotective dietary supplement composition having undenatured type II collagen according to the present disclosure may have an ORAC to hypochlorite of about 25 μmol TE / g or more, such as about 30 μmol TE / g or more, such as about 35 μmol TE / g or more, such as about 40 μmol TE / g or more, such as about 45 μmol TE / g or more, such as up to about 50 μmol TE / g or more, up to about 75 μmol TE / g, or any range or value therebetween.

[0066] When the type II collagen includes undenatured type II collagen, the undenatured type II collagen has a molecular weight of about 10,000 daltons or more, for example, about 15,000 daltons or more, for example, about 20,000 daltons or more, for example, about 25,000 daltons or more, for example, about 30,000 daltons or more, for example, about 35,000 daltons or more, for example, about 40,000 daltons or more, for example, about 45,000 daltons or more, for example, about 50,000 daltons or more, for example, about 55,000 daltons or more. 0 daltons or more, such as about 60,000 daltons or more, such as about 65,000 daltons or more, such as about 70,000 daltons or more, such as about 75,000 daltons or more, such as about 80,000 daltons or more, such as about 85,000 daltons or more, such as about 90,000 daltons or more, such as about 95,000 daltons or more, such as about 100,000 or more, up to about 350,000 daltons or less, or any range or value therebetween.

[0067] Various aspects and advantages have been discussed, in one aspect, the chondroprotective dietary supplement composition is incorporated into a suitable delivery form before being incorporated into a dosage form, as will be discussed below.In one aspect, the composition of the present disclosure can be included as an oil-in-water emulsion as a delivery form.In particular, in one aspect, such a configuration can allow one or more oil-soluble active ingredients and / or one or more water-soluble active ingredients to be contained in the same delivery form.Alternatively, only oil-soluble components can be used (e.g., type II collagen), and the composition can be incorporated into aqueous applications using emulsion.However, it should be understood that in one aspect, water-soluble type II collagen can be used in supplements.

[0068] Nevertheless, the oil-in-water emulsion may also contain at least one functional gum, such as gum arabic. Generally, gum arabic is a complex mixture of glycoproteins and polysaccharides containing arabinose and galactose. Gum arabic is generally soluble in water and edible. In some embodiments, gum arabic may comprise 100% processed gum arabic, such as Ticamulsion® A-2010 gum arabic powder. In certain embodiments, gum arabic may be a mixture or blend of gum arabic and processed gum arabic. For example, in certain embodiments, gum arabic may comprise Ticamulsion® 3020.

[0069] In certain aspects, the oil-in-water emulsion contains about 10% to about 30% gum arabic by weight. In some embodiments, the oil-in-water emulsion contains about 15% to about 25% gum arabic by weight. In some embodiments, the oil-in-water emulsion contains less than about 20% gum arabic by weight, such as less than 15%, such as less than 10%, such as less than 5%.

[0070] The oil-in-water emulsion may also contain water. In certain embodiments, the oil-in-water emulsion contains deionized water. In certain embodiments, the oil-in-water emulsion may contain any water suitable for human consumption and for incorporation into dietary supplements designed for human consumption.

[0071] The amount of water incorporated into the oil-in-water emulsion may vary depending on the desired hygroscopic and water-soluble components incorporated into the oil-in-water emulsion. In certain aspects, the oil-in-water emulsion may contain about 5% to 35% water by weight. In some embodiments, the oil-in-water emulsion may contain about 10% to about 30% water by weight. In some embodiments, the oil-in-water emulsion may contain about 15% to about 20% water by weight. In some embodiments, the oil-in-water emulsion may contain less than about 20% water by weight, such as less than about 15% water by weight, such as less than about 10% water by weight.

[0072] In some embodiments, the oil-in-water emulsion may contain one or more stabilizers or suspension promoters.For example, in certain embodiments, the oil-in-water emulsion may contain one or more gums, such as gellan gum or xantham gum.When included, the gellan gum or xantham gum may be present in an amount of less than about 3.5% by weight of the oil-in-water emulsion, such as less than about 2.5% by weight, such as less than about 1.5% by weight, such as less than about 1.0% by weight, such as less than about 1.0% by weight.

[0073] In other embodiments, the oil-in-water emulsion may contain one or more stabilizers, such as silica. If included, the silica may be present in an amount less than about 2% by weight, such as less than about 1.5% by weight, such as less than about 1% by weight, such as less than about 0.5% by weight.

[0074] Furthermore, in one embodiment, the oil-in-water emulsion can also contain one or more fat-soluble ingredients or nutrients.In certain embodiments, one or more fat-soluble ingredients or nutrients can be incorporated into the oil phase of the oil-in-water emulsion.Suitable fat-soluble ingredients include, but are not limited to, retinol, vitamin E provided by mixed tocopherols, beta-carotene, ubiquinone, lecithin, sunflower lecithin, vitamin D, cannabinoids, hemp extract, vitamin K, phosphatidylcholine, and combinations thereof.

[0075] In certain embodiments, at least one or more fat-soluble components may be incorporated into the oil-in-water emulsion in an amount of about 0% to about 50% by weight. For example, in some embodiments, the oil-in-water emulsion contains less than about 50% by weight of one or more fat-soluble components, such as less than about 40% by weight, such as less than about 30% by weight, such as less than about 20% by weight, such as less than about 10% by weight, such as less than about 5% by weight.

[0076] Additionally, in one embodiment, the oil-in-water emulsion may contain one or more additional antioxidants in one or more of the water-soluble phase or the oil / fat-soluble phase.

[0077] In some aspects, the oil-in-water emulsions disclosed herein may be used in any suitable dosage form, such as tablets, gummy chewables, edible films, lozenges, liquid suspensions, syrups, lipid micelles, spray-dried dispersions, nanoparticles, and the like, which may also be incorporated into additional supplements.

[0078] Alternatively, the oil-in-water emulsions may be included in nutritional products such as supplements, foods, or beverages. For example, in certain aspects, the oil-in-water emulsions may be incorporated into liquid nutritional products such as dietary supplements or infant formula for consumption by a mammal. Additionally, the oil-in-water emulsions provided herein may be added to any liquid nutritional product designed to provide nutritional support to a mammal.

[0079] The nutritional product may comprise any suitable composition for consumption by a mammal. Such compositions include complete foods or beverages intended to supply the necessary dietary requirements for a mammal or food supplements such as sweets and snacks. Food compositions may include pellets, beverages, bars, canned prepared foods, milkshake drinks, juices, dairy products, or any other functional food composition. Food compositions may also include supplements in any form such as pills, soft gels, gummy figurines, wafers, powders, etc.

[0080] In one aspect, the supplement according to the present disclosure may be administered to a mammal, including by oral, enteral or inhalation administration of whole collagen protein or biologically active peptide fragments of collagen.For example, in one aspect, whole collagen protein or biologically active peptide fragments of collagen are believed to increase the trans-L-hydroxyproline content based on the total weight of amino acids contained in collagen type II, allowing efficient production of trans-L-hydroxyproline and enhancing pharmaceutical efficacy.

[0081] The supplement composition of the present disclosure may further comprise one or more excipients as further additives in the composition.Exemplary but non-limiting excipients and / or additives include anti-adherent agents such as magnesium stearate; binders such as sugars, sugar alcohols, gelatin, and synthetic polymers; coating agents such as cellulose ether hydroxypropylmethylcellulose (HPMC), shellac, corn protein zein, gelatin, fatty acids, fats, oils, and / or waxes; colorants such as titanium dioxide and azo dyes; disintegrants such as modified starch sodium starch glycolate and cross-linked polymers, including polyvinylpyrrolidone and sodium carboxymethylcellulose; fillers such as maltodextrin; mint, licorice, and the like. , flavorings such as anise, vanilla, and fruit flavors including peach, banana, grape, strawberry, blueberry, raspberry, and mixed berry; glidants such as fumed silica, talc, and magnesium carbonate; lubricants such as talc, silica, and fats including vegetable stearin, magnesium stearate, and stearic acid; preservatives such as antioxidants, vitamins, retinyl palmitate, selenium, the amino acids cysteine ​​and methionine, citric acid, sodium citrate, and parabens; adsorbents; sweeteners such as sucrose and sucralose; and vehicles such as petrolatum and mineral oil.

[0082] In one aspect, the supplement composition of the present disclosure may be combined with various additives and components that can improve one or more properties of the composition. For example, in one embodiment, the additive composition may be combined with a stabilizer package that can help stabilize at least one property of the composition. In a particular embodiment, for example, the stabilizer package may be added to the composition in an amount sufficient to reduce the hydrophobic properties of the composition and / or prevent the composition from absorbing moisture. The stabilizer package may also be combined with the composition to improve the handling properties of the composition. For example, the stabilizer package may enable the composition to have better flow properties, especially when granular.

[0083] In one aspect, the supplement composition may be combined with a polymer binder in conjunction with a stabilizer package.In addition, a coating material may be applied to the composition after the composition is combined with the polymer binder and stabilizer package.The coating material may, for example, contain at least one fat.According to the present disclosure, the above components may be added to any suitable pharmaceutical composition in addition to the composition of the present disclosure.For example, the above components may be added to any pharmaceutical composition that contains carnitine or amino acid.

[0084] The polymer binder and stabilizer package may be combined with the supplement composition in a manner that uniformly incorporates the stabilizer package into the product. In one embodiment, for example, the composition of the present disclosure is first combined with the polymer binder, such as by a spray drying process, and then combined with the stabilizer package. The polymer binder may include any suitable pharma- ceutically acceptable polymer, such as a film-forming polymer and / or a polysaccharide. Specific examples of polymer binders that may be used according to the present disclosure include starch, maltodextrin, gum arabic, arabinogalactan, gelatin, and mixtures thereof. In one embodiment, the polymer binder is added to the pharmaceutical composition in an amount of at least about 5% by weight, such as at least about 8% by weight, such as at least about 10% by weight, such as at least about 15% by weight. One or more polymer binders are present in the composition in an amount of less than about 50% by weight, such as less than about 45% by weight, such as less than about 40% by weight, such as less than about 35% by weight, such as less than about 30% by weight.

[0085] In one embodiment, the polymeric binder may include a starch, such as a modified starch. The starch may be derived, for example, from corn or waxy corn. In one embodiment, the starch may include HI-CAP 100 starch sold by National Starch and Chemical Company.

[0086] In an alternative embodiment, the polymer binder may comprise arabinogalactan.Arabinogalactan is a soluble polysaccharide that can function as a polymer binder and also provide other benefits.For example, arabinogalactan can enhance adaptive immune response in some situations.Arabinogalactan is described, for example, in U.S. Patent No. 8,784,844, which is incorporated herein by reference.

[0087] In one embodiment, arabinogalactan may be used as the polymer binder. Larch arabinogalactan is a highly branched polysaccharide composed of galactose and arabinose units in an approximately 6:1 ratio. Larch arabinogalactan is extracted from a large tree. The polysaccharide has a galactan backbone with galactose and arabinose side chains. Arabinogalactan is commercially available from Lonza Consumer Health Inc. with offices in Morristown, NJ USA.

[0088] Once the polymer binder is combined with the composition, such as by a spray drying process, the resulting mixture can then be combined with a stabilizer package. In one embodiment, the stabilizer package includes oxide particles combined with a salt of a carboxylic acid. In one particular embodiment, the stabilizer package may include a dry product, such as a powder or granular product, combined with the composition and the polymer binder. It has been found that the combination of oxide particles and salt of a carboxylic acid provides numerous advantages and benefits when combined with the composition. For example, it has been found that the stabilizer package stabilizes the composition and reduces the hydrophobicity of the composition. This composition also produces a product that is easy to handle, granular, and flowable.

[0089] The oxide particles that can be added to the supplement composition can include silica.For example, the oxide particles can include precipitated silica particles.The silica particles can have a particle size (d50, laser diffraction after ISO test 13320) of less than about 55 microns, for example, less than about 40 microns, for example, less than about 30 microns, for example, less than about 25 microns, for example, less than about 20 microns, for example, less than about 15 microns, for example, less than about 12 microns, for example, less than about 10 microns, for example, less than about 8 microns, for example, less than about 6 microns, for example, less than about 4 microns, for example, less than about 2 microns, for example, less than about 1 micron.The particle size is typically more than about 0.5 microns, for example, more than about 1 micron. The particles may have a specific surface area (ISO test 9277) of more than about 120 m2 / g, for example more than about 130 m2 / g, for example more than about 150 m2 / g, for example more than about 170 m2 / g, for example more than about 200 m2 / g, for example more than about 220 m2 / g. The specific surface area is generally less than about 500 m2 / g. Oxide particles such as silica particles can be present in the pharmaceutical composition in an amount of more than about 0.01% by weight, for example more than about 0.05% by weight, for example more than about 0.1% by weight. Oxide particles are generally present in an amount of less than 5% by weight, for example less than about 2% by weight, for example less than about 1.5% by weight, for example less than 0.5% by weight.

[0090] In addition to the oxide particles, the stabilizer package may also include a salt of a carboxylic acid. The salt of a carboxylic acid may include a salt of a fatty acid. The fatty acid may have, for example, a carbon chain length of about 6 carbon atoms to about 40 carbon atoms, for example, about 12 carbon atoms to about 28 carbon atoms. In one embodiment, the salt of a carboxylic acid may include a stearate. Stearates that may be used include calcium stearate, sodium stearate, magnesium stearate, mixtures thereof, and the like. In one embodiment, the salt of a carboxylic acid may include both hydrophilic and hydrophobic groups. The salt of a carboxylic acid may be present in the composition in an amount greater than about 0.5% by weight, for example, greater than about 1% by weight, for example, greater than about 1.5% by weight. The salt of a carboxylic acid is generally present in an amount less than about 5% by weight, for example, less than about 4% by weight, for example, less than about 3% by weight.

[0091] In addition to the polymer binder and stabilizer package, the composition may include various other components and ingredients. In one embodiment, for example, the composition may include a citric acid ester, such as a citric acid ester of a monoglyceride and / or diglyceride of a fatty acid. The composition may also include a lecithin, such as a lecithin obtained from rapeseed, sunflower, etc. The above components may be present in the composition in relatively small amounts, such as less than about 2% by weight, such as less than about 1.5% by weight, such as less than about 1% by weight. The above components are generally present in an amount greater than about 0.05% by weight, such as greater than about 0.1% by weight.

[0092] As mentioned above, the chondroprotective dietary supplement composition or supplement of the present disclosure can contain collagen in combination with one or more bone health additives.In one embodiment, for example, the composition or supplement contains a calcium source.The calcium source can include, for example, calcium carbonate, calcium citrate, calcium gluconate, calcium lactate, calcium hydroxyapatite, or mixtures thereof. Calcium may be present in each dose in an amount greater than about 100 mg, such as greater than about 200 mg, for example greater than about 250 mg, such as greater than about 300 mg, for example greater than about 350 mg, such as greater than about 400 mg, for example greater than about 500 mg, such as greater than about 600 mg, for example greater than about 700 mg, such as greater than about 800 mg, for example greater than about 900 mg, such as greater than about 1,000 mg, for example greater than about 1,200 mg, such as greater than about 1,400 mg, for example greater than about 1,600 mg, such as greater than about 1,800 mg, for example greater than about 2,000 mg, such as greater than about 2,500 mg, and typically less than about 5,000 mg, such as less than about 3,000 mg, for example less than about 2,500 mg.

[0093] Other bone health additives that may be included in the composition include vitamin K2, vitamin D3, bone antiresorptive agents, anabolic agents, or mixtures thereof. Each bone health additive may be present in the composition in a weight ratio relative to collagen of about 5:1 to about 1:1,000. For example, the bone health additive may be present in a weight ratio relative to collagen of about 1:1 to about 1:500, such as about 1:10 to about 1:300, such as about 1:100 to about 1:200.

[0094] In addition to calcium, the composition may contain one or more other minerals, such as potassium, manganese, magnesium, zinc, or mixtures thereof. The minerals may be included in the supplement in an amount of about 1 mg / g to about 50 mg / g, e.g., about 2.5 mg / g to about 45 mg / g, e.g., about 5 mg / g to about 40 mg / g, or any range or value therebetween. The above ranges may be for any one mineral or the total amount of one mineral. In one embodiment, the supplement contains potassium in an amount of about 9.5 mg / g to about 12 mg / g, e.g., about 9.75 mg / g to about 11.5 mg / g, e.g., about 10 mg / g to about 11 mg / g, or any range or value therebetween. Similarly, in one embodiment, the supplement contains magnesium in an amount of about 1 mg / g to about 10 mg / g, e.g., about 2.5 mg / g to about 7.5 mg / g, e.g., about 4 mg / g to about 6 mg / g, or any range or value therebetween. Further, in one embodiment, the supplement contains calcium in an amount of about 1 mg / g to about 50 mg / g, e.g., about 2.5 mg / g to about 47.5 mg / g, e.g., about 5 mg / g to about 45 mg / g, e.g., about 10 mg / g to about 40 mg / g, e.g., about 20 mg / g to about 37.5 mg / g, e.g., about 30 mg / g to about 35 mg / g, or any range or value therebetween.

[0095] The supplements can be administered with or in combination with a variety of other health additives. For example, a chondroprotective nutraceutical composition can contain or be administered with an antioxidant, an anti-inflammatory agent, or a combination thereof.

[0096] The supplement may be suitable for administration to any mammal. For example, the mammal may be a human, a dog, a cat, a cow, a horse, etc. The composition may be provided to a mammal of any age from birth to adulthood in the mammal. In various embodiments, the mammal may be a human, a dog, a cat, a horse, a pig, a sheep, or a cow. EXAMPLES

[0097] The present disclosure may be better understood in accordance with the following examples, which are intended to be non-limiting and exemplary in nature.

[0098] Example 1 Materials and Methods Ten weaned rats of the same sex, approximately 4 weeks old and weighing approximately 60-80 grams, were used in each group. The animals were housed at temperature (22 ± 2°C), humidity (55 ± 5%), and a 12 / 12-h light / dark cycle. After a one-week adaptation period, the weaned rats at 4 weeks of age were fasted for 12 hours, weighed, and randomly grouped according to weight and housed in separate cages. Deionized water was provided to avoid calcium intake from drinking water.

[0099] The following feed formulations were used during the study: [Table 1]

[0100] Experimental design To evaluate the efficacy of native form of type II collagen (UCII) on Ca-resorbing bone tissue, 4-week-old male Wistar rats were fed a normal diet for a 1-week adaptation period prior to the study.

[0101] Rats were randomly assigned to six dietary groups as follows: 1) Negative control (0.150% Ca) 2) Negative control (0.150% Ca) + 0.66 mg / kg BW UCII (40 mg HED based on 60 kg BW) 3) Negative control (0.150% Ca) + 1.32 mg / kg BW UCII (80 mg HED based on 60 kg BW) 4) Negative control (0.150% Ca) + 3.3 mg / kg BW UCII (200 mg HED based on 60 kg BW) 5) Negative control (0.150% Ca) + 6.6 mg / kg BW UCII (400 mg HED based on 60 kg BW) 6) Negative control (0.150% Ca) + 19.8 mg / kg BW UCII (1200 mg HED based on 60 kg BW)

[0102] The test substance was given by oral gavage or, if gavage was unsuccessful, the test substance was mixed into the diet. The body weight of each rat was determined weekly. After 12 weeks, blood, liver, kidney and bone (right femur weight was measured and at the same time the left femur was excised for measurement of BMD) samples were taken.

[0103] Metabolic experiments In the last week of the study, a calcium metabolism experiment was performed for 3 days: food intake was recorded for 3 days, and feces were collected for 72 hours to determine calcium content in the diet and feces.

[0104] Determination of calcium in blood, feed and feces: Determination by atomic absorption spectrophotometry. Feed samples were mixed homogeneously and passed through a 20-mesh sieve, and rat fecal samples were dried in an oven at 60°C, cooled in a desiccator, and ground for mineral analysis. Approximately 0.3 g of ground samples were mineralized in HNO3 (5 ml) using a closed microwave digestion system (Berghof, Eningen, Germany). Feed and fecal samples were then diluted with deionized water to determine the Ca content in feces and diet. Lanthanum chloride (Merck, Darmstadt, Germany) was added as an interference suppressor for Ca g analysis. Ca levels in all samples were measured by atomic absorption spectroscopy (AAS, Perkin-Elmer, Analyst 800, Norwalk, CT, USA) by flame nebulization in acetylene-air at a wavelength of 422.7 nm with Zeeman background correction. Evaluations were performed in triplicate. Serum Ca levels were also measured by atomic absorption spectroscopy.

[0105] calculation Calcium intake (mg / day) = calcium content in feed (mg / g) × feed consumption (g / day)

[0106] Fecal calcium (mg / d) = fecal calcium content (mg / g) × fecal excretion (g / d)

[0107] Apparent calcium absorption rate (%) = (calcium intake - calcium in feces) / calcium intake x 100%

[0108] Femur weight measurement The animals were sacrificed after 3 months of feeding. The right femurs were removed and baked in an oven at 105°C to a constant weight, and the oven-dry weight was measured.

[0109] Determination of femoral bone mineral density Bone mineral density was measured at the midpoint and distal end of the femur by a dual energy X-ray bone densitometer. Marks were drawn at the distal end of the femur and at the midpoint of the femur to determine the measurement points. The entire length of the femur was measured and a straight line was drawn from the midpoint along the cross-sectional direction. The measurement point of the distal end of the femur was determined at the lower end of the articular groove of the distal femur and a straight line was drawn through this point parallel to the mark at the midpoint of the femur. This is the measurement point of the distal end of the femur.

[0110] The bone to be measured was moved so that the marker line of the measurement point coincided with the vertical projection of the movement path of the probe on the measurement table. The measurement was started and repeated twice for each point. If the two results were not parallel (error > 10%) the measurement was repeated. The two results were averaged to determine BMC (bone mineral content), BW (bone width) and consequently BMD (bone mineral density).

[0111] Determination of bone calcium content and calcium in feed ingredients Bone calcium content and dietary calcium content were measured by atomic absorption spectroscopy (AAS). Approximately 0.3 g of ground samples were mineralized in HNO3 (5 ml) using a closed microwave digestion system (Berghof, Eningen, Germany). Samples were then diluted with deionized water to determine tissue and dietary Ca content. Lanthanum chloride (Merck, Darmstadt, Germany) was added as an interference suppressor for Ca analysis. Ca levels in all samples were measured by atomic absorption spectroscopy (AAS, Perkin-Elmer, Analyst 800, Norwalk, CT, USA) by flame nebulization in acetylene-air at a wavelength of 422.7 nm with Zeeman background correction. Evaluations were performed in triplicate.

[0112] Osteocalcin levels in serum were detected with an ELISA (Elx-800, Bio-Tek Instruments Inc, Vermont, USA) device using a rat-specific commercial kit according to the manufacturer's instructions.

[0113] OPG, RANK, RANKL, SOX-9, BMP2, and IGF-1 were measured by WB.

[0114] The results of the body weight measurements are given below and shown in FIG. [Table 2]

[0115] Calcium absorption and retention results are as follows: [Table 3]

[0116] After 12 weeks, the effects of different doses of non-denatured collagen type II on femur dry weight, bone mineral density, bone mineral content, bone calcium, serum calcium, osteocalcin, and parathyroid hormone levels in the rats under study are presented below. Figures 2-8 also illustrate the results. [Table 4]

[0117] The effects of different doses of undenatured collagen II (UCII) on the levels of receptor activator of nuclear factor-κB (RANK, Panel A), receptor activator of nuclear factor-κB ligand (RANKL, Panel B), osteoprotegerin (OPG, Panel C), sex determining region Y box 9 (SOX-9, Panel D), and the RANKL / OPG ratio (Panel E) in bones of rats fed a low Ca (0.15%) diet for 12 weeks are illustrated in Figures 9-12.

[0118] The effect of different doses of undenatured collagen II (UCII) on insulin-like growth factor 1 (IGF-1, panel A) and bone morphogenetic protein 2 (BMP2, panel B) protein levels in bones of rats fed a low Ca (0.15%) diet for 12 weeks is illustrated in Figures 13 and 14.

[0119] These and other modifications and variations to the present invention may be practiced by those of ordinary skill in the art without departing from the spirit and scope of the present invention, which is particularly set forth in the appended claims. In addition, it should be understood that aspects of the various embodiments may be interchanged in whole or in part. Moreover, those skilled in the art will appreciate that the foregoing description is by way of example only, and is not intended to limit the invention as further set forth in such appended claims.

Claims

1. 1. A chondroprotective nutritional supplement composition for use in increasing calcium absorption in a mammal, wherein the mammal is consuming a diet containing calcium, and wherein the chondroprotective nutritional supplement composition comprises collagen.

2. 10. The chondroprotective nutritional supplement composition of claim 1, wherein the mammal has a calcium deficiency.

3. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the mammal is a healthy young adult who is healthy, suffers from a joint injury or skeletal disease, or is in the growth stage of bone development.

4. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the mammal is a postmenopausal female.

5. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for treating bone diseases, joint diseases or co-morbidities including diabetes, cardiometabolic syndrome, inflammatory and immune disease states or any chronic disease state related thereto.

6. 10. The chondroprotective nutritional supplement composition of claim 1, wherein said chondroprotective nutritional supplement composition meets a nutritional need or requirement for therapeutic or pharmaceutical purposes in said mammal.

7. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to a mammal in an amount sufficient to increase bone mineral density in the mammal, for example, by more than about 4%, compared to the mammal before administration of the chondroprotective nutritional supplement composition.

8. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to a mammal in an amount sufficient to increase the percent absorption of calcium in the mammal, for example by more than about 1%, for example by more than about 5%, for example by more than about 10%, for example by more than about 15%, compared to the mammal before administration of the chondroprotective nutritional supplement composition.

9. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to a mammal in an amount sufficient to increase the percent calcium retention in the mammal, for example by more than about 15%, compared to the mammal prior to administration of the chondroprotective nutritional supplement composition.

10. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to a mammal in an amount sufficient to increase femur dry weight in the mammal.

11. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to a mammal in an amount sufficient to increase osteocalcin in the mammal, for example by more than about 10%, compared to the mammal prior to administration of the chondroprotective nutritional supplement composition.

12. 3. The chondroprotective nutritional supplement composition of claim 1, wherein the chondroprotective nutritional supplement composition is for administration to a mammal in an amount sufficient to increase bone morphogenetic protein 2 in the mammal, for example by more than about 30%, compared to the mammal prior to administration of the chondroprotective nutritional supplement composition.

13. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the collagen comprises undenatured type II collagen.

14. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the collagen is obtained from chicken, fish, bovine or porcine cartilage.

15. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the mammal is a human.

16. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to the mammal at least once a week, such as at least every three days, such as at least daily.

17. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to the mammal based on the amount of calcium in the mammal's blood.

18. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the collagen comprises type I collagen, type II collagen, type III collagen, type IV collagen, or type V collagen.

19. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition is for administration to the mammal at collagen dose levels of at least about 10 mg collagen per dose, such as at least about 20 mg collagen per dose, for example at least about 40 mg collagen per dose, such as at least about 60 mg collagen per dose, for example at least about 80 mg collagen per dose, and less than about 3,000 mg collagen per dose, such as less than about 1,000 mg collagen per dose, for example less than about 500 mg collagen per dose, for example less than about 350 mg collagen per dose.

20. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein said chondroprotective nutritional supplement composition is for administration to said mammal in combination with a calcium supplement.

21. 21. The chondroprotective nutritional supplement composition of claim 20, wherein said calcium supplement is contained within said chondroprotective nutritional supplement composition.

22. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the chondroprotective nutritional supplement composition comprises an antioxidant, an anti-inflammatory agent, or a mixture thereof.

23. 3. The chondroprotective nutritional supplement composition of claim 1 or 2, wherein the diet of the mammal is calcium deficient.

24. 1. A chondroprotective supplement for use in improving bone health in a mammal, comprising: Bone health additives and a bone health supplement for said bone health additive, said bone health supplement comprising collagen, said collagen being present in said chondroprotective supplement in a therapeutically effective amount sufficient to increase absorption of said bone health additive into bone of a mammal.

25. 25. The chondroprotective supplement of claim 24, wherein said bone health additive comprises calcium, and wherein calcium is present in said supplement in an amount of from about 250 mg to about 2,500 mg.

26. 26. The chondroprotective supplement of claim 25, wherein the bone health additive comprises calcium carbonate, calcium citrate, calcium gluconate, calcium lactate, or mixtures thereof.

27. 27. The chondroprotective supplement according to any one of claims 24 to 26, wherein the collagen is present in the supplement in an amount of more than about 10 mg, such as more than about 40 mg, for example more than about 60, such as more than about 80 mg, for example more than about 100 mg, such as more than about 120 mg, for example more than about 200 mg, and less than about 500 mg.

28. 27. The chondroprotective supplement of any one of claims 24 to 26, wherein the bone health supplement is present in the supplement in an amount sufficient to increase osteocalcin in the mammal and / or bone morphogenetic protein 2 in the mammal.

29. 27. The chondroprotective supplement according to any one of claims 24 to 26, wherein the supplement is in the form of a tablet, capsule, suspension, or emulsion.

30. 27. The chondroprotective supplement of any one of claims 24 to 26, wherein the bone health supplement is present in the supplement in an amount sufficient to increase the percent absorption or percent retention of the bone health additive in the mammal.

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