Method for using prescribed quantitative value of collagen contained in teeth of mammal

Abstract

[Problem] To provide a method for using a prescribed quantitative value and / or weight ratio value of collagen contained in a tooth of a mammal as an index for evaluating a potential risk of developing a disease in the mammal and / or whether the mammal is potentially healthy. [Solution] At least one of PYD, PEN, T-GLYC, PpP, and PpG is compared with a reference value thereof with respect to each of: collagen contained in a tooth of a mammal to be evaluated for a potential risk of developing a disease and / or whether the mammal is potentially healthy; and collagen contained in a tooth of one or more mammals selected from a mammal that has not developed a disease, a mammal that has developed a disease, a healthy mammal, and a non-healthy mammal.

Description

Method using a predetermined quantitative value of collagen contained in mammalian teeth

[0001] The present invention provides a method of using a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in mammalian teeth, a method of using a quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in mammalian teeth, a method of using a quantitative value of the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in mammalian teeth, and a method of using a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in mammalian teeth as an index for assessing the potential risk of disease in mammals and / or whether the mammal is potentially healthy. and a method using a weight ratio (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)) calculated from the quantitative value of the amount of pyridinoline contained in collagen in mammalian teeth and the quantitative value of the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained).

[0002] Various collagens (actually, "collagen-rich organic compositions" would be more accurate, but they are often simply referred to as "collagen") have been developed. Most of these are atelocollagens produced by extraction from soft tissues in living organisms using enzymatic treatment, in which the telopeptides present at the N-terminus and C-terminus of the collagen amino acid sequence are cleaved, and poorly soluble collagen fibers are removed during a purification process. Collagens produced by extraction from hard tissues such as bones and teeth, such as demineralized freeze-dried bone allograft (DFDBA) and demineralized dentin matrix (DDM), have also been developed. These collagens are produced by demineralization and are primarily used as biomaterials.

[0003] The quality of atelocollagen is evaluated based on the fact that telopeptides, which are said to have antigenicity, are cleaved by enzymatic treatment, while the quality of undenatured collagen derived from soft tissues (collagen with a triple helix structure due to the retention of telopeptides) is evaluated based on how highly pure collagen is produced by setting purification conditions such as pH. However, when producing collagen from hard tissues, decalcification, i.e., acid treatment, is required to remove minerals and extract the collagen, which limits the collagen yield and requires a lot of effort. Therefore, almost all collagen products currently being developed are atelocollagen derived from soft tissues, undenatured collagen, or products utilizing them.

[0004] Most collagen contained in biological tissues is type I collagen. Type I collagen has both physiological and non-physiological crosslinks, with pyridinoline crosslinks known as physiological crosslinking molecules and crosslinks derived from advanced glycation end products (AGEs), such as pentosidine, known as non-physiological crosslinking molecules (Non-Patent Document 1). Pyridinoline crosslinks are physiological, enzymatic crosslinks formed by the action of lysyl oxidase, and are regular crosslinking molecules at specific sites in the amino acid sequence of collagen, contributing to improved elasticity and strength of collagen fibers. On the other hand, pentosidine crosslinks are non-physiological, non-enzymatic crosslinks (aging crosslinks (AGEs crosslinks)) involving blood glucose. When blood glucose is heated by body temperature, they are crosslinking molecules formed between arginines and lysines scattered throughout the amino acid sequence of collagen. Therefore, they are crosslinking molecules formed randomly between adjacent arginines and lysines in the amino acid sequence of collagen, which disrupts the physiological three-dimensional structure of collagen and contributes to a decrease in the elasticity and strength of collagen fibers. As mentioned above, atelocollagen contains almost no pyridinoline crosslinking molecules because the telopeptides are cleaved by enzymatic treatment. Furthermore, since the insoluble collagen fibers resulting from pentosidine crosslinking are eliminated during the purification process, atelocollagen contains almost no pentosidine crosslinking molecules.

[0005] It has been known that an increase in pyridinoline crosslink molecules in collagen increases the strength and elasticity of the collagen (Non-Patent Document 2), and that pentosidine crosslinks are pathological crosslinks (aging crosslinks (AGEs crosslinks)) (Non-Patent Document 3). It has also been reported that the amount of pentosidine per unit collagen of human articular cartilage increases linearly with age, and that the ratio of pentosidine to pyridinoline per unit collagen (pentosidine content / pyridinoline content) increases accelerated with age (Non-Patent Documents 4 and 5). It has also been reported that the ratio of pentosidine to pyridinoline per unit collagen (pentosidine content / pyridinoline content) significantly increases in non-calcified lesions of dystrophic aortic calcification (Non-Patent Document 6).

[0006] M. Saito, CLINICIAN Vol. 554, 1141-1146, 2006M. Saito, THE BONE Vol. 21, No. 1, 53-58, 2007Sell, D. R. et al. ,V. M. , J. Biol. Chem. Vol. 284, 21597-21602, 1989A. Uchiyama et al. , J. Biochem. Vol. 110, 714-718, 1991M. TAKAHASHI et al. , Arthritis Rheum. Vol. 37, No. 5, 724-728, May 1994. H. HOSHINO et al. , Atherosclerosis Vol. 112, 39-46, 1995

[0007] The quality of the above-mentioned collagen (collagen-rich organic composition) is generally evaluated based on the purity and characteristics of the collagen in the collagen-rich organic composition, and is not evaluated based on any molecular-level indicator of collagen, i.e., any indicator of the amino acid sequence level of collagen. Therefore, it is difficult to say that the quality of the collagen itself has been properly evaluated. Furthermore, the quality of collagen has not been evaluated based on the amount of pyridinoline contained, which contributes to improving collagen quality, such as improving the elasticity and strength of collagen fibers, the amount of pentosidine contained and the amount of total glycation products contained, the weight ratio of the amount of pyridinoline contained and the amount of pentosidine contained, and the weight ratio of the amount of pyridinoline contained and the amount of total glycation products contained, which are factors that disrupt the physiological three-dimensional structure of collagen, causing a decrease in the elasticity and strength of collagen fibers and ultimately contributing to a decrease in collagen quality.

[0008] Furthermore, the above-mentioned Non-Patent Documents 1 to 6 certainly make various references to pyridinoline crosslink molecules and pentosidine crosslink molecules in collagen, and also discuss the ratio of the amount of pentosidine and the amount of pyridinoline contained per unit collagen in diseases (amount of pentosidine contained / amount of pyridinoline contained), but none of these documents are particularly concerned with collagen derived from teeth, from which collagen is not easy to extract. Furthermore, in tissues that require physiological strength, such as hard tissues such as bones and cartilage, and soft tissues such as tendons and ligaments, the amount of pyridinoline crosslink molecules contained in the collagen tends to be large, and therefore the amount of pentosidine crosslink molecules or crosslink molecules of total glycation products tends to be large (M. TAKAHASHI et al., Anal Biochem Vol. 232, 158-62, 1995). However, because these tissues are metabolically active, the pyridinoline crosslink molecules, pentosidine crosslink molecules, and crosslink molecules of total glycation products contained therein are also metabolized, and therefore do not reflect the total amount of pyridinoline crosslink molecules, pentosidine crosslink molecules, and crosslink molecules of total glycation products that have accumulated in the mammalian host. In contrast, teeth, which are hard tissues, are the only hard tissues that are not incorporated into the metabolic cycle, and therefore the pyridinoline crosslink molecules, pentosidine crosslink molecules, and crosslink molecules of total glycation products contained in the collagen contained therein are not metabolized, so the total accumulated amount of pyridinoline crosslink molecules, pentosidine crosslink molecules, and crosslink molecules of total glycation products that have accumulated in the mammalian host to date is reflected. Therefore, in order to accurately evaluate the potential disease risk in mammals and whether the mammal is potentially healthy or not, it is clear that tooth-derived collagen (collagen-rich organic composition) that reflects the total accumulated amount of pyridinoline crosslink molecules, pentosidine crosslink molecules, and crosslink molecules of total glycation products that have accumulated in the mammalian host to date should be used as an indicator. However, the above-mentioned Non-Patent Documents 1 to 6 do not state, or even suggest, such a statement.

[0009] The present invention relates to a method for using a quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value PYD; quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of one or more mammals selected from non-disease mammals, diseased mammals, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value R-PYD; and comparing the PYD with the R-PYD. The present invention also relates to a method for using a quantitative value of the amount of pentosidine contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy; a step of quantifying the amount of pentosidine (ng / mg) contained in collagen in the teeth of a mammal that is at risk of a disease to obtain a quantitative value of PEN; a step of quantifying the amount of pentosidine (ng / mg) contained in collagen in the teeth of one or more mammals selected from mammals that are not at risk of a disease, mammals that are at risk of a disease, healthy mammals, and unhealthy mammals to obtain a reference quantitative value of R-PEN; and a step of comparing the PEN and the R-PEN; The method uses a quantitative value of the total amount of glycation end products contained in collagen contained in the teeth of a mammal as an index for assessing whether the mammal is potentially healthy or not, the method comprising the steps of quantifying the amount of total glycation end products contained in collagen (ng / mg: converted into the amount of pentosidine contained) contained in the teeth of a mammal to be evaluated for potential disease risk and / or whether the mammal is potentially healthy or not, to obtain a quantitative value of T-GLYC;a method comprising a step of quantifying the amount of total glycation products contained in collagen contained in teeth (ng / mg: converted into the amount of pentosidine contained) to obtain a reference quantitative value of R-T-GLYC, and a step of comparing the T-GLYC with the R-T-GLYC; a method using a weight ratio calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in teeth of a mammal and the quantitative value of the amount of pentosidine contained therein as an index for assessing the potential disease risk in the mammal and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of total glycation products contained in collagen contained in teeth (ng / mg: converted into the amount of pentosidine contained therein) to obtain a reference quantitative value of R-T-GLYC; a step of quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal to be evaluated as to whether the mammal is potentially healthy or not, to obtain a quantitative value of PYD; a step of quantifying the amount of pentosidine (ng / mg) contained in collagen contained in the teeth of the mammal to be evaluated, to obtain a quantitative value of PEN; a step of calculating a weight ratio of PpP (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) from the obtained PYD and the obtained PEN; The present invention relates to a method for determining the amount of pyridinoline (ng / mg) contained in collagen in the teeth of one or more mammals selected from the group consisting of mammals suffering from diseases, healthy mammals, and unhealthy mammals, thereby obtaining a reference quantitative value of R-PYD; a method for determining the amount of pentosidine (ng / mg) contained in collagen in the teeth of the one or more mammals, thereby obtaining a reference quantitative value of R-PEN; and a method for determining the amount of pentosidine (ng / mg) contained in collagen in the teeth of the one or more mammals, thereby obtaining a reference quantitative value of R-PEN from the obtained R-PYD and R-PEN. and a method for evaluating the potential risk of a disease in a mammal and / or the potential health of a mammal, the method comprising the steps of: calculating a weight ratio of the amount of pyridinoline contained in collagen to the amount of total glycation products contained in collagen contained in the teeth of a mammal; and comparing the amount of pentosidine contained in the teeth of a mammal with the amount of pyridinoline contained in collagen and the amount of total glycation products contained in collagen contained in the teeth of a mammal, the method comprising the steps of: calculating a weight ratio of the amount of pyridinoline contained in collagen to the amount of total glycation products contained in collagen contained in the teeth of a mammal; and comparing the amount of pentosidine contained in the teeth of a mammal with the amount of pyridinoline contained in collagen contained in the teeth of a mammal with the amount of total glycation products contained in collagen contained in the teeth of a mammal.The method comprises a step of quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in teeth to obtain a quantitative value of PYD; a step of quantifying the amount of total glycation end products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in teeth of the mammal to be evaluated to obtain a quantitative value of T-GLYC; a step of calculating a weight ratio value of PpG (amount of pyridinoline contained (ng / mg) / total amount of glycation end products contained (ng / mg: converted into the amount of pentosidine contained)) from the obtained PYD and T-GLYC; and a step of measuring the amount of collagen contained in teeth of one or more mammals selected from mammals not affected by a disease, mammals affected by a disease, healthy mammals, and unhealthy mammals. The object of the present invention is to provide at least one of the following methods, which include a step of quantifying the amount of pyridinoline (ng / mg) contained in collagen to obtain R-PYD, which is a reference quantitative value; a step of quantifying the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in the teeth of the one or more mammals to obtain R-T-GLYC, which is a reference quantitative value; a step of calculating R-PpG, which is a reference value for weight ratio (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained)), from the obtained R-PYD and R-T-GLYC; and a step of comparing the PpG and the R-PpG.

[0010] As a result of extensive research into solving the above-mentioned problems, the present inventors have found that it is possible to evaluate the potential disease risk in a mammal and / or whether a mammal is potentially healthy by comparing the values ​​of at least any of the above-mentioned PYD, PEN, T-GLYC, Ppp and PpG for collagen contained in the teeth of a mammal to be evaluated for its potential disease risk and / or whether the mammal is potentially healthy with the collagen contained in the teeth of one or more mammals selected from mammals not affected by a disease, mammals affected by a disease, healthy mammals and unhealthy mammals, with their respective reference values, and have completed the following inventions.

[0011] (1) A method for using a quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a mammal contracting a disease and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal that is to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value, PYD; quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of one or more mammals selected from mammals that are not affected by a disease, mammals that are affected by a disease, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value, R-PYD; and comparing the PYD with the R-PYD.

[0012] (2) A method for using a quantitative value of the amount of pentosidine contained in collagen contained in the teeth of a mammal as an indicator for assessing the potential risk of a mammal contracting a disease and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of pentosidine contained in collagen (ng / mg) contained in the teeth of a mammal that is the subject of an assessment of the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value, PEN; quantifying the amount of pentosidine contained in collagen (ng / mg) contained in the teeth of one or more mammals selected from mammals that are not affected by a disease, mammals that are affected by a disease, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value, R-PEN; and comparing the PEN with the R-PEN.

[0013] (3) A method for using a quantitative value of the total amount of glycation products contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a disease in the mammal and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of total glycation products contained in collagen (ng / mg: converted into the amount of pentosidine contained) in the teeth of a mammal that is the subject of an assessment of the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value of T-GLYC; quantifying the amount of total glycation products contained in collagen (ng / mg: converted into the amount of pentosidine contained) in the teeth of one or more mammals selected from mammals that are not affected by a disease, mammals that are affected by a disease, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value of R-T-GLYC; and comparing the T-GLYC with the R-T-GLYC.

[0014] (4) A method for using a weight ratio calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, the method comprising the steps of quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy to obtain a quantitative value of PYD; quantifying the amount of pentosidine (ng / mg) contained in collagen contained in the teeth of the mammal to be assessed to obtain a quantitative value of PEN; and calculating a weight ratio of PpP (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) from the obtained PYD and the obtained PEN. a step of quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of one or more mammals selected from mammals not afflicted with a disease, mammals afflicted with a disease, healthy mammals, and unhealthy mammals to obtain a reference quantitative value R-PYD; a step of quantifying the amount of pentosidine (ng / mg) contained in collagen contained in the teeth of the one or more mammals to obtain a reference quantitative value R-PEN; a step of calculating the weight ratio reference value R-PpP (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) from the obtained R-PYD and R-PEN; and a step of comparing the PpP and the R-PpP.

[0015] (5) A method for using a weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for evaluating the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal to be evaluated for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value of PYD; quantifying the amount of total glycation end products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in the teeth of the mammal to be evaluated, to obtain a quantitative value of T-GLYC; and calculating a weight ratio value of PpG (amount of pyridinoline contained (ng / mg) / total amount of glycation end products (ng / mg)) from the obtained PYD and the obtained T-GLYC. quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of one or more mammals selected from non-diseased mammals, diseased mammals, healthy mammals, and unhealthy mammals to obtain a reference quantitative value of R-PYD; quantifying the amount of total glycation end products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in the teeth of the one or more mammals to obtain a reference quantitative value of R-T-GLYC; calculating a weight ratio reference value of R-PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation end products contained (ng / mg: converted into the amount of pentosidine contained)) from the obtained R-PYD and R-T-GLYC; and comparing the PpG and the R-PpG.

[0016] (6) The method according to (1), wherein the quantitative determination step is a step of determining the amount of the fluorometrically active substance by using a high performance liquid chromatograph equipped with a fluorescence detector (HPLC-Flu) with heptafluorobutyric acid and formic acid as ion-pairing reagents.

[0017] (7) The method according to any one of (1) to (6), wherein the mammal is one or more mammals selected from the group consisting of humans, cows, horses, pigs, wild boars, sheep, deer, dogs, and cats.

[0018] According to the present invention, by comparing the values ​​of at least any of the above-mentioned PYD, PEN, T-GLYC, Ppp and PpG for collagen contained in the teeth of a mammal that is the subject of an assessment of the potential risk of disease and / or whether the mammal is potentially healthy with the collagen contained in the teeth of one or more mammals selected from mammals that are not afflicted with a disease, mammals that are afflicted with a disease, healthy mammals and unhealthy mammals, with the respective reference values, it is possible to assess the potential risk of disease in a mammal and / or whether the mammal is potentially healthy.

[0019] This figure shows the distribution of PYD, the quantitative value of pyridinoline content (ng / mg) in collagen hydrolysates contained in teeth (37 teeth total) from 14 non-diabetic human patients (●), 4 diabetic human patients (▲), and 19 bovine (specified risk materials (SRM)) (◆). The PYD was determined by high-performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents. The dashed line in the figure indicates an approximation curve (linear approximation) based on the PYD of 14 non-diabetic human patients, which corresponds to R-PYD. This figure shows the distribution of pentosidine (PEN), a quantitative value of the amount of pentosidine (ng / mg) contained in the collagen hydrolysate contained in the teeth of 14 non-diabetic human patients (●), 4 diabetic human patients (▲), and 19 bovine (specified risk materials (SRM)) teeth (◆) (37 teeth in total), as determined by high-performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents. In the figure, the dashed line indicates an approximation curve (linear approximation) based on the PEN of 14 non-diabetic human patients, which corresponds to R-PEN. This figure shows the distribution of T-GLYC, the quantitative value of total glycation end products (ng / mg: converted to pentosidine content), contained in collagen hydrolysates contained in teeth (37 teeth in total) from 14 non-diabetic human patients (●), 4 diabetic human patients (▲), and 19 bovine (specified risk materials (SRM)) (◆). The total amount of T-GLYC was quantified by high-performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents. The dashed line in the figure indicates an approximation curve (linear approximation) based on the T-GLYC of 14 non-diabetic human patients, which corresponds to R-T-GLYC.This figure shows the distribution of Ppp (pyridinoline content (ng / mg) / pentosidine content (ng / mg)) by weight, calculated from PYD, which is the quantitative value of pyridinoline content (ng / mg), and PEN, which is the quantitative value of pentosidine content (ng / mg), of collagen hydrolysates contained in teeth (37 teeth in total) from 14 non-diabetic human patients (●), 4 diabetic human patients (▲), and 19 bovine (specified risk materials (SRM)) teeth (◆). The dashed line in the figure indicates an approximation curve (linear approximation) based on the Ppp values ​​of the 14 non-diabetic human patients, which corresponds to R-Ppp. Figure 1 shows the distribution of PpG (amount of pyridinoline contained (ng / mg) / total amount of glycation end products contained (ng / mg: equivalent to amount of pentosidine contained)) by weight, calculated from PYD, which is the quantitative value of the amount of pyridinoline contained (ng / mg), and T-GLYC, which is the quantitative value of the amount of total glycation end products contained (ng / mg: equivalent to amount of pentosidine contained), of collagen hydrolysates contained in teeth (37 samples in total) from 14 non-diabetic human patients (●), 4 diabetic human patients (▲), and 19 bovine (specified risk materials (SRM)) (◆). The PpG weight ratio was calculated from PYD, which is the quantitative value of the amount of pyridinoline contained (ng / mg), and T-GLYC, which is the quantitative value of the amount of total glycation end products contained (ng / mg: equivalent to amount of pentosidine contained), as quantified by high-performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents. In the figure, the dashed line indicates an approximate curve (linear approximation) based on the PpG of 14 human non-diabetic patients, which corresponds to R-PpG.

[0020] The following describes a method of using a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy according to the present invention; a method of using a quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy according to the present invention; a method of using a quantitative value of the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy according to the present invention; Alternatively, a method using a weight ratio (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) calculated from the quantitative value of the amount of pyridinoline (ng / mg) and the quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in a mammal's tooth as an index for evaluating whether a mammal is potentially healthy, and a method according to the present invention using a weight ratio (amount of pyridinoline (ng / mg) / amount of total glycation products (ng / mg: converted into the amount of pentosidine)) calculated from the quantitative value of the amount of pyridinoline (ng / mg) and the amount of total glycation products (ng / mg: converted into the amount of pentosidine) contained in collagen contained in a mammal's tooth as an index for evaluating a mammal's potential disease risk and / or whether a mammal is potentially healthy will be described in detail. In this specification, a numerical range expressed using "to" or "from" means a range that includes the numerical values ​​before and after "to" or "from" as the lower and upper limits.

[0021] In the present invention, the term "mammal" refers to an animal that basically reproduces sexually, with many extant species being viviparous and raising its young by milk. The term is not particularly limited as long as it is such an animal. Examples of such animals include, but are not limited to, humans; apes such as orangutans, gorillas, chimpanzees, bonobos, and monkeys belonging to the Gibbon family; monkeys other than those belonging to the Gibbon family; large mammals such as cows, horses, pigs, wild boars, sheep, goats, deer, serows, bears, sea lions, seals, walruses, sea lions, fur seals, and whales; and small mammals such as dogs, cats, rabbits, mice, squirrels, weasels, raccoons, and mongooses. Preferred mammals include humans, cows, horses, pigs, wild boars, sheep, deer, dogs, and cats. More preferred mammals include humans, cows, horses, pigs, dogs, and cats. The term "mammal" is also applicable to animals that are not particularly limited.

[0022] In the present invention, the term "potential risk of contracting a disease" does not refer to a state in which the patient is infected with a disease, but rather refers to a state in which the host's defense response against the disease is at its maximum and the patient is barely able to avoid contracting the disease, and refers to a state in which the presence of the disease cannot be confirmed by subjective symptoms, objective findings, or conventional general test findings.

[0023] In addition, in the present invention, "potentially healthy" refers to a state that does not lead to an unhealthy state, and means a state in which it is hardly possible to confirm that one is unhealthy based on subjective, objective, or conventional general test findings. In the present invention, "potentially unhealthy" refers to a state that does not lead to a healthy state, and means a state in which it is hardly possible to confirm that one is healthy based on subjective, objective, or conventional general test findings.

[0024] Collagen is one of the proteins that form the framework of various tissues in living organisms, mainly constituting various organs, blood vessels, nerves, skin, ligaments, tendons, bones, cartilage, and dental dentin of vertebrates, and is the main component of the extracellular matrix of multicellular animals. In the present invention, "collagen" mainly refers to type I collagen. Furthermore, the "collagen" of the present invention refers to dental collagen, and teeth are tissues that are subjected to a strong load, namely, occlusal force. Therefore, unlike soft tissues, dental collagen is required to have extremely high strength to withstand occlusal force.

[0025] Hard tissue collagen can be determined to be derived from hard tissue by detecting, for example, BMP-2, BMP-4, BMP-7, and osteocalcin, and can be determined to be derived from teeth by detecting dentin sialoprotein (DSP), dentin glycoprotein (DGP), and dentin phosphoprotein (DPP).

[0026] In the present invention, the pyridinoline, pentosidine, and total glycation products contained in collagen are, respectively, pyridinoline constituting pyridinoline cross-linking molecules, which are physiological cross-linking molecules in collagen, pentosidine constituting pentosidine cross-linking molecules, which are non-physiological cross-linking molecules in collagen, and total glycation products constituting cross-linking molecules of total glycation products, which are non-physiological cross-linking molecules in collagen. In the present invention, the amount of pyridinoline (ng / mg) contained in collagen, the amount of pentosidine (ng / mg) contained in collagen, and the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen are each quantified to obtain quantitative values ​​of PYD, PEN, and T-GLYC. Furthermore, by dividing the PYD by the PEN, PpP (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)), which is the weight ratio of the PYD to the PEN, is calculated, and by dividing the PYD by the T-GLYC, PpG (amount of pyridinoline (ng / mg) / total glycation products (ng / mg: converted into the amount of pentosidine contained)), which is the weight ratio of the PYD to the T-GLYC, is calculated.

[0027] It is no exaggeration to say that pyridinoline crosslinks, which are physiological enzymatic crosslinking molecules, were primarily designed to improve the fundamental strength of hard tissues. As mentioned above, they are said to be crosslinking molecules formed by the action of the enzyme lysyl oxidase between two lysine residues in the telopeptides located at the N- and C-termini of the collagen amino acid sequence and one lysine residue in the collagen amino acid sequence excluding both termini. The pyridinoline crosslinking molecules formed between the two lysine residues in the telopeptides located at the N- and C-termini of the collagen amino acid sequence are crosslinking molecules between collagen fibers, and therefore contribute greatly to maintaining the three-dimensional structure of tropocollagen (the entire collagen sequence), which has a triple helix structure.

[0028] Therefore, when the pyridinoline cross-linking molecules increase in collagen contained in biological tissues, the quality of the collagen contained in the biological tissues becomes flexible and tenacious, the strength and calcification of the collagen increase, and it can also be a factor that improves the activity of the cells that make up the biological tissues, i.e., it can be a factor that makes it easier to maintain individual homeostasis by increasing the activity of tissue stem cells involved in tissue repair. Therefore, the amount of pyridinoline contained is higher in collagen derived from teeth and bones than in collagen from soft tissues such as skin, and collagen derived from teeth and bones is flexible, strong, and highly elastic.

[0029] On the other hand, crosslinks by advanced glycation end products (AGEs), represented by non-physiological crosslinking molecules such as pentosidine crosslinking molecules and crosslinking molecules of total glycation end products containing pentosidine crosslinking molecules, are pathological crosslinks (aging crosslinks (AGE crosslinks)) caused by blood sugar, and as described above, are crosslinking molecules formed by various combinations of arginine and lysine scattered throughout the amino acid sequence of collagen when blood sugar is heated by body temperature. When arginine and arginine, lysine and lysine, and arginine and lysine are close to each other in the amino acid sequence of collagen, pentosidine crosslinking molecules and crosslinking molecules of total glycation end products containing pentosidine crosslinking molecules are formed nonspecifically (randomly), which can cause collagen fibers to become entangled, causing the collagen fibers to lose their regularity in arrangement and disrupting the three-dimensional structure of tropocollagen. Therefore, an increase in pentosidine crosslinking molecules and crosslinking molecules of total glycation end products containing pentosidine crosslinking molecules in the amino acid sequence of collagen leads to a decrease in the flexibility, strength, and elasticity of collagen fibers. Because the formation of these pathological cross-linking molecules is caused by blood sugar, it is said that their accumulation increases depending on the host's dietary habits (carbohydrate intake habits). Furthermore, because carbohydrates, which are the raw materials for pentosidine cross-linking molecules and cross-linking molecules of total glycation end products containing pentosidine cross-linking molecules, are supplied via the blood vessels, collagen, which serves as the supportive matrix for tissues throughout the body, is cross-linked according to the amount of circulating blood flow, regardless of the location of the tissue. In other words, excessive sugar intake can be said to uniformly reduce the quality of collagen, which serves as the supportive matrix for the body, regardless of whether it is hard or soft tissue.

[0030] Furthermore, when the number of pentosidine crosslink molecules and crosslink molecules of total glycation products containing pentosidine crosslink molecules in collagen increases, the collagen contained in the biological tissue becomes brittle and chalk-like, and the strength and degree of calcification of the collagen contained in the biological tissue decrease, because the pentosidine crosslink molecules and crosslink molecules of total glycation products containing pentosidine crosslink molecules are glycation crosslink molecules formed by the Maillard reaction mediated by sugars. The mature products of the sugar-mediated Maillard reaction are known as advanced glycation end products (AGEs), and the crosslinking molecules of pentosidine crosslinking molecules and the total glycation end products including pentosidine crosslinking molecules are AGEs. Collagen from mammals whose blood sugar levels are constantly maintained at a high level due to a dietary habit with frequent carbohydrate intake contains more pentosidine crosslinking molecules and the total glycation end products including pentosidine crosslinking molecules than collagen from mammals of the same age (months) with a well-balanced diet. Therefore, it can be said that pathological crosslinking (aging crosslinking (AGE crosslinking)) is progressing in the collagen of mammals whose blood sugar levels are constantly high. The generated pentosidine crosslinking molecules and the crosslinking molecules of the total glycation end products including pentosidine crosslinking molecules disrupt the regular arrangement of collagen fibers, impairing the flexibility of collagen, making it hard and brittle, and inhibiting collagen metabolism, resulting in a deterioration in the quality of collagen that constitutes the entire body and causing a decrease in the flexibility of tissues. Collagen is the main component of the supporting tissues that support not only all hard tissues such as bones and teeth, but also all soft tissues, including nerves and blood vessels. Therefore, a decline in collagen quality can lead to a deterioration in the quality of hard and soft tissues throughout the body. For example, when bone quality declines due to bone glycation, bones become brittle and osteoporosis occurs. In addition, when blood vessel quality declines due to glycation of blood vessel walls, arteries harden, leading to arteriosclerosis, which causes tissue damage such as elevated blood pressure.The same is true for the three major complications of diabetes: (1) diabetic retinopathy (which can lead to blindness if it worsens), (2) diabetic nephropathy (which can lead to dialysis if it worsens), and (3) diabetic neuropathy (which can lead to, for example, necrosis of the fingers and toes due to impaired circulation if it worsens).The causes of these complications are said to be (1) impaired circulation in the retina, (2) capillary damage in the glomeruli of the kidneys, which are the blood filtering system, and (3) impaired blood flow in the extremities due to capillary neuropathy and arteriosclerosis.It can be said that a major factor in all of these complications is the deterioration of blood vessel quality due to glycation of the collagen that makes up the blood vessels.

[0031] In addition, when AGEs, such as pentosidine crosslink molecules and crosslink molecules of total glycation end products containing pentosidine crosslink molecules, increase, they bind to RAGE (receptor of age: AGE receptor), which is expressed in a wide range of tissues, including vascular cells such as vascular endothelial cells and immune cells such as macrophages, creating an in vivo environment that is conducive to signal activation. This promotes inflammation-related NF-κB signaling, and AGEs, such as pentosidine crosslink molecules and crosslink molecules of total glycation end products containing pentosidine crosslink molecules, function as chronic inflammation trigger factors in excessively glycated tissues, thereby inducing chronic inflammation in a wide range of tissues. This mechanism is thought to be a factor in exacerbating chronic inflammatory diseases, such as inflammatory bowel disease (IBD) and collagen diseases. As described above, collagen glycation can gradually erode the body in various ways, unwittingly reducing quality of life (QOL) and becoming a major factor in preventing healthy longevity. It has also been reported that the pyridinoline and pentosidine crosslinks in collagen contained in human biological tissues both increase with age (Shimizu, "Aging-Related Nonenzymatic Glycation of Dentin Collagen Protein," Osaka University Knowledge Archive, 2015; Walters C. et al., Calcif. Tissue. Int. Vol. 35, 401-405, 1983).

[0032] The method of the present invention, which uses the quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk of the mammal and / or whether the mammal is potentially healthy, includes the following steps: (i) quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal that is to be evaluated for the potential disease risk and / or whether the mammal is potentially healthy, to obtain a quantitative value of PYD (evaluation target pyridinoline amount quantification step); (ii) quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of one or more mammals selected from disease-free mammals, diseased mammals, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value of R-PYD (reference pyridinoline amount quantification step); (iii) a step of comparing the PYD obtained in the step (i) of quantifying the amount of pyridinoline contained in the evaluation target with the R-PYD obtained in the step (ii) of quantifying the amount of pyridinoline contained in the reference (PYD / R-PYD comparison step).

[0033] Furthermore, the method of the present invention, which uses the quantitative value of the amount of pentosidine (ng / mg) contained in collagen in the teeth of a mammal as an index for assessing the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, includes the following steps: (iv) quantifying the amount of pentosidine (ng / mg) contained in collagen in the teeth of a mammal that is to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value PEN (quantification step of pentosidine contained in the target mammal); (v) quantifying the amount of pentosidine (ng / mg) contained in collagen in the teeth of one or more mammals selected from non-diseased mammals, diseased mammals, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value R-PEN (quantification step of pentosidine contained in the reference mammal); (vi) a step of comparing the PEN obtained in the step (iv) of quantifying the amount of pentosidine contained in the evaluation target with the R-PEN obtained in the step (v) of quantifying the amount of pentosidine contained in the reference (PEN / R-PEN comparison step).

[0034] Furthermore, the method of the present invention, which uses the quantitative value of the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, includes the following steps: (vii) quantifying the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen in the teeth of a mammal that is a target for assessment of the potential disease risk and / or whether the mammal is potentially healthy, to obtain a quantitative value of T-GLYC (a step of quantifying the amount of total glycation products contained in the target); (viii) quantifying the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen in the teeth of one or more mammals selected from disease-free mammals, diseased mammals, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value of R-T-GLYC (a step of quantifying the amount of total glycation products contained in the reference); (ix) a step of comparing the T-GLYC obtained in the step (vii) of quantifying the amount of total glycation products contained in the evaluation subject with the RT-GLYC obtained in the step (viii) of quantifying the amount of total glycation products contained in the reference (a T-GLYC / RT-GLYC comparison step). The method comprises the steps (vii) to (ix).

[0035] Furthermore, the method of the present invention, which uses a weight ratio calculated from the quantitative value of the pyridinoline content of collagen contained in the teeth of a mammal and the quantitative value of the pentosidine content as an index for assessing the potential disease risk of a mammal and / or whether the mammal is potentially healthy, includes: (x) a step of quantifying the pyridinoline content (ng / mg) of collagen contained in the teeth of a mammal that is to be evaluated for the potential disease risk and / or whether the mammal is potentially healthy, to obtain a quantitative value PYD (a step of quantifying the amount of pyridinoline contained in the evaluation target); (xi) a step of quantifying the pentosidine content (ng / mg) of collagen contained in the teeth of the mammal that is to be evaluated, to obtain a quantitative value PEN (a step of quantifying the amount of pentosidine contained in the evaluation target); (xii) a step of calculating the weight ratio PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)) from the PYD obtained in the step (x) of quantifying the amount of pyridinoline contained in the evaluation target and the PEN obtained in the step (xi) of quantifying the amount of pentosidine contained in the evaluation target (evaluation target weight ratio PpP calculation step); (xiii) a step of quantifying the amount of pyridinoline contained (ng / mg) in collagen contained in the teeth of one or more mammals selected from non-diseased mammals, diseased mammals, healthy mammals, and unhealthy mammals, to obtain R-PYD, which is a reference quantitative value (reference pyridinoline contained amount quantification step); (xiv) a step of quantifying the amount of pentosidine contained (ng / mg) in collagen contained in the teeth of the one or more mammals, to obtain R-PEN, which is a reference quantitative value (reference pentosidine contained amount quantification step); (xv) a step of calculating a reference value of weight ratio R-PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)) from the R-PYD obtained in the reference pyridinoline-containing amount determination step (xiii) and the R-PEN obtained in the reference pentosidine-containing amount determination step (xiv) (reference weight ratio R-PpP calculation step);(xvi) A step of comparing the Ppp calculated in the step (xii) of calculating the evaluation target weight ratio PppP with the R-Ppp calculated in the step (xv) of calculating the reference weight ratio R-PppP (a step of comparing weight ratios PppP and R-PppP).

[0036] Furthermore, the method according to the present invention, which uses a weight ratio calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, includes the following steps: (xvii) quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal that is to be evaluated for the potential disease risk and / or whether the mammal is potentially healthy, to obtain the quantitative value PYD (quantification step of pyridinoline contained in the evaluation target); (xviii) quantifying the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in the teeth of the mammal that is to be evaluated, to obtain the quantitative value T-GLYC (quantification step of total glycation products contained in the evaluation target); (xix) a step of calculating a weight ratio value of PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted to amount of pentosidine contained)) from PYD obtained in the step (xvii) of quantifying the amount of pyridinoline contained in the evaluation target and T-GLYC obtained in the step (xviii) of quantifying the amount of total glycation products contained in the evaluation target (evaluation target weight ratio PpG calculation step); (xx) a step of quantifying the amount of pyridinoline contained (ng / mg) in collagen contained in the teeth of one or more mammals selected from mammals not afflicted with a disease, mammals afflicted with a disease, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value of R-PYD (reference step of quantifying amount of pyridinoline contained); (xxi) quantifying the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in the one or more mammalian teeth to obtain a reference quantitative value of R-T-GLYC (reference total glycation product amount quantifying step); (xxii) calculating a reference weight ratio value of R-PpG (amount of pyridinoline contained (ng / mg) / total glycation product contained (ng / mg: converted into the amount of pentosidine contained)) from R-PYD obtained in the reference pyridinoline amount quantifying step (xx) and R-T-GLYC obtained in the reference total glycation product amount quantifying step (xxi) (reference weight ratio R-PpG calculation step);(xxiii) a step of comparing the PpG calculated in the step (xix) of calculating the weight ratio PpG to be evaluated with the R-PpG calculated in the step (xxii) of calculating the reference weight ratio R-PpG (a step of comparing the weight ratio PpG and R-PpG).

[0037] The "step (i) of quantifying the amount of pyridinoline contained in an evaluation target" in the method according to the present invention, which uses a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen in the teeth of a mammal as an index for assessing the potential disease risk of a mammal and / or whether the mammal is potentially healthy; the "step (x) of quantifying the amount of pyridinoline contained in an evaluation target" in the method according to the present invention, which uses a weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen in the teeth of a mammal and the quantitative value of the amount of pentosidine contained, as an index for assessing the potential disease risk of a mammal and / or whether the mammal is potentially healthy; and the "step (xvii) of quantifying the amount of pyridinoline contained in an evaluation target" in the method according to the present invention, which uses a weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen in the teeth of a mammal and the quantitative value of the amount of total glycation products contained, as an index for assessing the potential disease risk of a mammal and / or whether the mammal is potentially healthy, are the same step.

[0038] Furthermore, the "reference-containing pyridinoline amount quantifying step (ii)" in the method according to the present invention, which uses a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in a mammal's teeth as an index for assessing a mammal's potential disease risk and / or whether the mammal is potentially healthy; the "reference-containing pyridinoline amount quantifying step (xiii)" in the method according to the present invention, which uses a weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in a mammal's teeth and the quantitative value of the amount of pentosidine contained as an index for assessing a mammal's potential disease risk and / or whether the mammal is potentially healthy; and the "reference-containing pyridinoline amount quantifying step (xx)" in the method according to the present invention, which uses a weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in a mammal's teeth and the quantitative value of the amount of total glycation products contained as an index for assessing a mammal's potential disease risk and / or whether the mammal is potentially healthy, are the same step.

[0039] Furthermore, the "step (iv) of quantifying the amount of pentosidine contained in an evaluation target" in the method of the present invention, which uses the quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, and the "step (xi) of quantifying the amount of pentosidine contained in an evaluation target" in the method of the present invention, which uses the weight ratio calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal and the quantitative value of the amount of pentosidine contained, as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, are the same step.

[0040] Furthermore, the "reference-containing pentosidine amount quantification step (v)" in the method according to the present invention, which uses the quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in mammalian teeth as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, and the "reference-containing pentosidine amount quantification step (xiv)" in the method according to the present invention, which uses the weight ratio calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth and the quantitative value of the amount of pentosidine contained, as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, are the same step.

[0041] Furthermore, the "step (vii) of quantifying the amount of total glycation products contained in an evaluation target" in the method of the present invention, which uses the quantitative value of the total amount of glycation products contained in collagen in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, and the "step (xviii) of quantifying the amount of total glycation products contained in an evaluation target" in the method of the present invention, which uses the weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen in the teeth of a mammal and the quantitative value of the total amount of glycation products contained, as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, are the same step.

[0042] Furthermore, the "reference-containing total glycation product amount quantification step (viii)" in the method according to the present invention, which uses the quantitative value of the total glycation product amount (ng / mg: converted into the pentosidine amount) contained in collagen in a mammal's tooth as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, and the "reference-containing total glycation product amount quantification step (xxi)" in the method according to the present invention, which uses the weight ratio value calculated from the quantitative value of the pyridinoline amount in collagen in a mammal's tooth and the quantitative value of the total glycation product amount as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy, are the same step.

[0043] Preparation of samples of pyridinoline, pentosidine and total glycation products to be subjected to the quantification of the amount of pyridinoline (ng / mg) contained in the "step of quantifying the amount of pyridinoline contained in the evaluation subject" and the "step of quantifying the amount of pyridinoline contained in the reference", the quantification of the amount of pentosidine contained (ng / mg) in the "step of quantifying the amount of pentosidine contained in the evaluation subject" and the "step of quantifying the amount of pentosidine contained in the reference", and the quantification of the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained) in the "step of quantifying the amount of total glycation products contained in the evaluation subject" and the "step of quantifying the amount of total glycation products contained in the reference" can be performed using conventional preparation means known in the art, and can be performed, for example, generally as follows.

[0044] (1) Dried collagen powder produced by the method for producing a collagen-rich organic composition according to the present invention is weighed into a test tube and swelled with a small amount of 6N hydrochloric acid under degassing. (2) 6N hydrochloric acid is added to the dried collagen, and the tube is then degassed and sealed. (3) Hydrolysis is carried out by heating at 110°C overnight. (4) The sample is cooled to room temperature. (5) Excess hydrochloric acid is removed by drying under reduced pressure while heating to obtain a collagen hydrolysate. (6) The obtained collagen hydrolysate is solubilized in a 10% aqueous methanol solution. (7) The sample is centrifuged at 12,000 rpm for 5 minutes at room temperature, and the supernatant is used as the sample. (8) A known amount of a standard substance is added to the sample to determine the extraction rate and calibration curve, allowing the collagen content to be appropriately corrected.

[0045] Furthermore, the method for quantifying the amount of pyridinoline (ng / mg) contained in the "step of quantifying the amount of pyridinoline contained in the evaluation subject" and the "step of quantifying the amount of pyridinoline contained in the reference", the method for quantifying the amount of pentosidine contained (ng / mg) in the "step of quantifying the amount of pentosidine contained in the evaluation subject" and the "step of quantifying the amount of pentosidine contained in the reference", and the method for quantifying the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained) in the "step of quantifying the amount of total glycation products contained in the evaluation subject" and the "step of quantifying the amount of total glycation products contained in the reference" are not particularly limited as long as they can be quantified, and as such a quantification method, for example, a high performance liquid chromatograph with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents can be used.

[0046] The quantitative determination using a high performance liquid chromatograph equipped with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents can be carried out using conventional quantitative determination means known in the art, but can generally be carried out, for example, as follows.

[0047] (1) Detection was performed using a gradient configuration of ultrapure water containing 0.05% heptafluorobutyric acid (mobile phase A) and a 1:1 mixture of methanol and ethanol containing 0.05% formic acid (mobile phase B). (2) Specific pumping conditions included a detection column temperature of 40°C, a flow rate of 0.5 mL / min, and a starting ratio of 90% mobile phase A and 10% mobile phase B for 0.5 minutes. (3) After that, the concentration of mobile phase B was increased to 22% over 1 minute to create a concentration gradient, and the pumping continued for 7.5 minutes. (4) Next, the concentration of mobile phase B was increased to 80% over 1 minute, and the pumping continued for another 1 minute. (5) Finally, the pumping continued at a ratio of 10% mobile phase A and 90% mobile phase B, followed by equilibration for 4 minutes at a ratio of 90% mobile phase A and 10% mobile phase B. (6) Steps (1) to (5) were performed for a total of 15 minutes per sample. The separation column used was a Cadenza CD-C18, 150 mm long and 3 mm in inner diameter, and a fluorescence detector was used for detection. Monitoring was performed for the first 8 minutes at an excitation wavelength of 295 nm and a fluorescence wavelength of 395 nm, and for the last 7 minutes at an excitation wavelength of 325 nm and a fluorescence wavelength of 385 nm. By these monitoring steps, pyridinoline was detected at a retention time of approximately 6.5 minutes, and pentosidine was detected at a retention time of approximately 9.1 minutes.

[0048] The quantification of the amount of pyridinoline (ng / mg) contained in the "step of quantifying the amount of pyridinoline contained in the evaluation subject" and the "step of quantifying the amount of pyridinoline contained in the reference", the quantification of the amount of pentosidine contained (ng / mg) in the "step of quantifying the amount of pentosidine contained in the evaluation subject" and the "step of quantifying the amount of pentosidine contained in the reference", and the quantification of the amount of total glycation products contained (ng / mg: converted to the amount of pentosidine contained) in the "step of quantifying the amount of total glycation products contained in the evaluation subject" and the "step of quantifying the amount of total glycation products contained in the reference" can be performed on extracted teeth of mammals that are the subject of evaluation of their potential disease risk and / or whether the mammal is potentially healthy, disease-free mammals, diseased mammals, healthy mammals, and unhealthy mammals, and can also be performed in a manner such as cutting, contact, or non-contact.

[0049] Furthermore, the mammal that is the subject of an assessment of potential risk of a disease and / or whether the mammal is potentially healthy, the mammal that is not affected by the disease, the mammal that is affected by the disease, the healthy mammal, and the unhealthy mammal may all be the same individual.

[0050] The method of the present invention, which uses the quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in a mammal's teeth as an index for evaluating the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, may include not only the above-mentioned "step (i) of quantifying the amount of pyridinoline contained in the evaluation target," "step (ii) of quantifying the amount of pyridinoline contained in the reference," and "step (iii) of comparing PYD and R-PYD," but also other steps within the scope that does not impair the features of the present invention.

[0051] Furthermore, the method according to the present invention, which uses the quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in a mammal's tooth as an index for evaluating the potential risk of a disease in the mammal and / or whether the mammal is potentially healthy, may include not only the above-mentioned "step (iv) of quantifying the amount of pentosidine contained in the evaluation target," "step (v) of quantifying the amount of pentosidine contained in the reference," and "step (vi) of comparing PEN and R-PEN," but also other steps within the scope that does not impair the features of the present invention.

[0052] Furthermore, the method according to the present invention, which uses the quantitative value of the amount of total glycation products contained in collagen contained in a mammal's teeth (ng / mg: converted into the amount of pentosidine contained) as an index for evaluating the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, may include not only the above-mentioned "step (vii) of quantifying the amount of total glycation products contained in an evaluation target," "step (viii) of quantifying the amount of total glycation products contained in a reference," and "step (ix) of comparing T-GLYC and R-T-GLYC," but also other steps within the scope that does not impair the features of the present invention.

[0053] Furthermore, the method according to the present invention, which uses the quantitative value of the pyridinoline content of collagen contained in a mammal's tooth and the weight ratio value calculated from the quantitative value of the pentosidine content, as an index for evaluating the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, may include not only the above-mentioned "step (x) of quantifying the amount of pyridinoline contained in an evaluation target," "step (xi) of quantifying the amount of pentosidine contained in an evaluation target," "step (xii) of calculating the weight ratio PpP of an evaluation target," "step (xiii) of quantifying the amount of pyridinoline contained in a reference," "step (xiv) of quantifying the amount of pentosidine contained in a reference," "step (xv) of calculating the reference weight ratio R-PpP," and "step (xvi) of comparing the weight ratios PpP and R-PpP," but may also include other steps as long as the features of the present invention are not impaired.

[0054] Furthermore, the method according to the present invention, which uses a weight ratio value calculated from the quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for evaluating the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, may include not only the above-mentioned "step (xvii) of quantifying the amount of pyridinoline contained in an evaluation target," "step (xviii) of quantifying the amount of total glycation products contained in an evaluation target," "step (xix) of calculating the weight ratio PpG of an evaluation target," "step (xx) of quantifying the amount of pyridinoline contained in a reference," "step (xxi) of quantifying the amount of total glycation products contained in a reference," "step (xxii) of calculating the reference weight ratio R-PpG," and "step (xxiii) of comparing the weight ratio PpG and R-PpG," but also other steps as long as the features of the present invention are not impaired.

[0055] A method according to the present invention using a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in mammalian teeth as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy; a method according to the present invention using a quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in mammalian teeth as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy; a method according to the present invention using a quantitative value of the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in mammalian teeth as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy; a method according to the present invention using a quantitative value of the amount of collagen contained in mammalian teeth as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy. In the method of using a weight ratio value calculated from the quantitative value of the pyridinoline amount and the quantitative value of the pentosidine amount contained, and in the method of the present invention of using a weight ratio value calculated from the quantitative value of the pyridinoline amount and the quantitative value of the total glycation end products amount of collagen contained in a mammal's teeth as an index for evaluating the potential disease risk in a mammal and / or the potential health of the mammal, the reason for limiting to "teeth" is that pyridinoline crosslinks, pentosidine crosslinks, and crosslinks of total glycation end products in type I collagen are all crosslinking reactions accelerated by factors such as internal heat due to body temperature and sugar supplied via the blood vessels, and are not localized crosslinking reactions that are induced according to circulatory blood flow, and among these, collagen contained in teeth (particularly dentin) is thought to most accurately reflect the accumulation of pyridinoline crosslink molecules, pentosidine crosslink molecules, and crosslink molecules of total glycation end products including pentosidine crosslink molecules. Teeth are the only tissues in the body that are not incorporated into the metabolic cycle, and once formed, they are not absorbed or decomposed. Therefore, pyridinoline crosslinks, pentosidine crosslinks, and crosslinks of total glycation products including pentosidine crosslinks accumulate in the collagen contained in teeth (especially dentin).

[0056] Furthermore, collagen in teeth (especially dentin) is more insoluble than that in bone. Pepsin digestion in 0.01 N hydrochloric acid at 4°C for 72 hours solubilizes approximately 35% of the collagen in adult bovine bone, but only 5.6% of the collagen in adult bovine dentin. Furthermore, at pH 2, insoluble collagen from skin and Achilles tendon swells to 4-8 times its volume, while insoluble collagen in adult bovine bone swells to 1.2 times its volume, whereas insoluble collagen in adult bovine dentin does not swell at all (Nagai Yutaka and Fujimoto Daizaburo, eds., Collagen Experimental Methods, Kodansha Scientific, pp. 21-22). Dentin collagen also has the highest content of pyridinoline crosslinks in living organisms. Therefore, PYD (ng / mg) is the quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, PEN (ng / mg) is the quantitative value of the amount of pentosidine contained in collagen contained in mammalian teeth, T-GLYC (ng / mg: converted into the amount of pentosidine contained) is the quantitative value of the amount of total glycation products contained in collagen contained in mammalian teeth, and PpP (pyridinoline content) is the weight ratio calculated from PYD (ng / mg) is the quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth and PEN (ng / mg) is the quantitative value of the amount of pentosidine contained. It is reasonable to use as indicators the weight ratio PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained)), which is calculated from the weight ratio PYD (ng / mg), which is a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, and T-GLYC (ng / mg: converted into the amount of pentosidine contained), which is a quantitative value of the amount of total glycation products contained, to evaluate the potential risk of disease in mammals and / or whether the mammal is potentially healthy.

[0057] On the other hand, the rationale for excluding bone is explained below. The values ​​of type I collagen C-terminal telopeptide (ICTP) in blood (serum) and NTx (type I collagen cross-linked N-telopeptide) in urine are widely used as indicators of osteoporosis. However, these are substances released into the blood and urine as collagen degradation products due to the activity of matrix metalloproteinases (MMPs), and contain pyridinoline cross-link molecules. This indicates that the pyridinoline cross-link molecules, pentosidine cross-link molecules, and cross-link molecules of total glycation end products, including pentosidine cross-link molecules, contained in bone-containing collagen are partially cleared by metabolism. In addition, when using extracted bone, bone is an organ that contains a large amount of lipids derived from yellow bone marrow, making defatting difficult and purifying highly pure collagen extremely difficult.

[0058] The present invention also relates to a method of using PYD, which is a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, as an index for assessing the potential risk of disease in mammals; a method of using PEN, which is a quantitative value of the amount of pentosidine contained in collagen contained in mammalian teeth, as an index for assessing the potential risk of disease in mammals; a method of using T-GLYC, which is a quantitative value of the total amount of glycation end products contained in collagen contained in mammalian teeth, as an index for assessing the potential risk of disease in mammals; a method of using PpP (amount of pyridinoline contained / amount of pentosidine contained), which is a weight ratio calculated from PYD, which is a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, and PEN, which is a quantitative value of the amount of pentosidine contained, as an index for assessing the potential risk of disease in mammals; and a method of using PYD, which is a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, and a quantitative value of the total amount of glycation end products contained, as an index for assessing the potential risk of disease in mammals. a method of using PYD, which is a quantitative value for the amount of pyridinoline contained in collagen contained in mammalian teeth, as an index for assessing whether or not a mammal of the present invention is potentially healthy; a method of using PEN, which is a quantitative value for the amount of pentosidine contained in collagen contained in mammalian teeth, as an index for assessing whether or not a mammal of the present invention is potentially healthy; a method of using T-GLYC, which is a quantitative value for the amount of total glycation products contained in collagen contained in mammalian teeth, as an index for assessing whether or not a mammal of the present invention is potentially healthy; a method of using PpP, which is a weight ratio value calculated from PYD, which is a quantitative value for the amount of pyridinoline contained in collagen contained in mammalian teeth, and PEN, which is a quantitative value for the amount of pentosidine contained in collagen contained in mammalian teeth, as an index for assessing whether or not a mammal of the present invention is potentially healthy;The methods using the value of PpG (amount of pyridinoline contained / total amount of glycation products contained), which is a weight ratio calculated from PYD, which is a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, and T-GLYC, which is a quantitative value of the amount of total glycation products contained, may be independent of each other or may be related to each other.

[0059] The following describes a method of using a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy according to the present invention; a method of using a quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy according to the present invention; a method of using a quantitative value of the amount of total glycation products contained in collagen contained in the teeth of a mammal as an index for assessing the potential disease risk in a mammal and / or whether the mammal is potentially healthy according to the present invention; A method of using the weight ratio (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) calculated from the quantitative value of the amount of pyridinoline (ng / mg) and the quantitative value of the amount of pentosidine (ng / mg) contained in collagen contained in a mammal's tooth as an index for evaluating whether a mammal is potentially healthy, and a method of using the weight ratio (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg: converted into the amount of pentosidine)) calculated from the quantitative value of the amount of pyridinoline and the amount of total glycation products contained in collagen contained in a mammal's tooth as an index for evaluating a mammal's potential disease risk and / or whether a mammal is potentially healthy according to the present invention are described based on examples. Note that the technical scope of the present invention is not limited to the embodiments shown in these examples.

[0060] Example 1 Measurement of pyridinoline, pentosidine, and total glycation product amounts in collagen derived from teeth (mainly dentin) of human non-diabetic patients, human diabetic patients, and bovine patients Using the procedure below, the pyridinoline amounts (ng / mg), pentosidine amounts (ng / mg), and total glycation product amounts (ng / mg: converted into the amount of pentosidine contained) in collagen derived from teeth (mainly dentin) of 14 human non-diabetic patients, 4 human diabetic patients, and 19 bovine specified risk materials (SRM)) were measured.

[0061] 1. Preparation of Collagen Powder [1-1] Hard tissue was pulverized to particle diameters of approximately 1 mm using a grinder. [1-2] The tissue was stirred in a 1% aqueous sodium carbonate solution at 70°C for several hours to remove any attached soft tissue. [1-3] Decalcification was performed. Specifically, the tissue was decalcified for several hours under reduced pressure using a decalcification solution containing 1M phosphoric acid and ethanol, and then replaced with a solution diluted several times and decalcified for several hours under reduced pressure. Subsequently, the tissue was completely decalcified by shaking for approximately one day at room temperature using the decalcification solution diluted 5 times. The tissue was then washed several times with 5% ethanol at room temperature. [1-4] The decalcified hard tissue was transferred to a mortar and mortar and ground in 70% ethanol to produce a fine powder. [1-5] The tissue was stirred in 100% ethanol, washed, degreased, and dehydrated. [1-6] The remaining ethanol in the collagen was completely removed by drying, yielding a dry collagen powder.

[0062] 2. Preparation of Analytical Samples [2-1] The dried collagen powder obtained in [1-6] was hydrolyzed. Specifically, the obtained dried collagen powder was weighed into a test tube and thoroughly swollen with a small amount of 6N hydrochloric acid under degassing. An appropriate amount of 6N hydrochloric acid was then added to the dried collagen, and the test tube was sealed under degassing. The sealed test tube was then heated overnight at 110°C to perform hydrolysis, after which it was cooled to room temperature, cut, and dried under reduced pressure while heating to remove excess hydrochloric acid, yielding a collagen hydrolysate. [2-2] The obtained collagen hydrolysate was solubilized in 10% aqueous methanol and centrifuged at 12,000 rpm for 5 minutes at room temperature. The supernatant was used as a sample. [2-3] A known amount of a standard substance was added to the sample to determine the extraction rate and calibration curve, and the collagen content was appropriately corrected.

[0063] 3. Quantification of the amount of pyridinoline, pentosidine, and total glycation products contained by a high-performance liquid chromatograph (HPLC-Flu) with a fluorescence detector using heptafluorobutyric acid and formic acid as ion-pair reagents, and calculation of their weight ratios. Using ultrapure water containing 0.05% heptafluorobutyric acid (mobile phase A) and a 1:1 mixture of methanol and ethanol containing 0.05% formic acid (mobile phase B), the mobile phase was pumped and detected using a gradient setting. Specifically, the detection column temperature was 40 ° C., and the flow rate was 0.5 mL / min, starting with a ratio of 90% mobile phase A and 10% mobile phase B, and pumped for 0.5 minutes. Thereafter, mobile phase B was increased to 22% over 1 minute to apply a concentration gradient, and then pumped as is for 7.5 minutes. Subsequently, mobile phase B was increased to 80% over 1 minute, and then pumped for another 1 minute. Finally, the mobile phase A was pumped at a ratio of 10% mobile phase B to 90% mobile phase B, followed by reducing the mobile phase B to 10% and equilibrating for 4 minutes at a ratio of 90% mobile phase A to 10% mobile phase B. Subsequently, detection was performed for a total of 15 minutes per sample. A Cadenza CD-C18 separation column, 150 mm in length and 3 mm in inner diameter, was used, and a fluorescence detector was used for detection. Monitoring was performed for the first 8 minutes at an excitation wavelength of 295 nm and a fluorescence wavelength of 395 nm, and for the last 7 minutes at an excitation wavelength of 325 nm and a fluorescence wavelength of 385 nm. By this monitoring, the amount of pyridinoline contained was detected at a retention time of approximately 6.5 minutes, and the amounts of pentosidine and total glycation products contained were detected at a retention time of approximately 9.1 minutes. From the obtained amounts of pyridinoline contained and total glycation products contained, the weight ratio thereof (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into amount of pentosidine contained)) was calculated.

[0064] FIG. 1 shows a distribution diagram of PYD, which is the quantitative value of the amount of pyridinoline (ng / mg) contained in the collagen contained in the teeth of the 14 non-diabetic human patients, 4 diabetic human patients, and 19 bovine animals, with respect to the age of the teeth. FIG. 2 shows a distribution diagram of PEN, which is the quantitative value of the amount of pentosidine (ng / mg) contained in the collagen contained in the teeth of the 14 non-diabetic human patients, 4 diabetic human patients, and 19 bovine animals, with respect to the age of the teeth. FIG. 3 shows a distribution diagram of T-GLYC, which is the quantitative value of the amount of total glycation end products (ng / mg: converted into the amount of pentosidine contained) contained in the collagen contained in the teeth of the 14 non-diabetic human patients, 4 diabetic human patients, and 19 bovine animals, with respect to the age of the teeth. Figure 4 shows a distribution diagram of the weight ratio PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)), calculated from PYD, which is a quantitative value of the amount of pyridinoline (ng / mg), and PEN, which is a quantitative value of the amount of pentosidine contained (ng / mg), over time for tooth age. Figure 5 shows a distribution diagram of the weight ratio PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained)), calculated from PYD, which is a quantitative value of the amount of pyridinoline contained (ng / mg) in collagen contained in the teeth of 14 non-diabetic human patients, 4 diabetic human patients, and 19 bovine animals, and calculated from T-GLYC, which is a quantitative value of the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained). In FIG. 1 , the dashed line indicates an approximation curve (linear approximation) based on PYD from 14 human non-diabetic patients, which corresponds to R-PYD; in FIG. 2 , the dashed line indicates an approximation curve (linear approximation) based on PEN from 14 human non-diabetic patients, which corresponds to R-PEN; in FIG. 3 , the dashed line indicates an approximation curve (linear approximation) based on T-GLYC from 14 human non-diabetic patients, which corresponds to R-T-GLYC; in FIG. 4 , the dashed line indicates an approximation curve (linear approximation) based on PpP from 14 human non-diabetic patients, which corresponds to R-PpP; and in FIG. 5 , the dashed line indicates an approximation curve (linear approximation) based on PpG from 14 human non-diabetic patients, which corresponds to R-PpG.

[0065] Although the measurement targets human permanent teeth, taking into account the period of dentin formation in permanent teeth, it can be assumed that dentin formation begins in tooth germs within the jawbone on average around age 10, and that glycation cross-link formation also begins accordingly (Study on the eruption period of deciduous and permanent teeth in Japanese children II, Journal of the Japanese Society of Pediatric Dentistry, 57(3), 363-373, 2019, 363). On the other hand, the same can be said for the mandibular teeth of bovine SRM, starting on average 18 months after birth. Therefore, for certain teeth in human and bovine SRM, Table 1 below was used to convert ages to months.

[0066] [Table 1]

[0067] Furthermore, the collagen contained in the teeth of 14 non-diabetic human patients was quantified by high performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents. The following data were obtained: PYD, which is the quantitative value of the amount of pyridinoline (ng / mg), PEN, which is the quantitative value of the amount of pentosidine (ng / mg), T-GLYC, which is the quantitative value of the amount of total glycation end products (ng / mg: converted into the amount of pentosidine), PYD, which is the quantitative value of the amount of pyridinoline (ng / mg), and PEN, which is the quantitative value of the amount of pentosidine. Table 2 below shows PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)), which is a weight ratio value calculated from PEN, which is a quantitative value of the amount of pyridinoline contained (ng / mg), and PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained)), which is a weight ratio value calculated from PYD, which is a quantitative value of the amount of pyridinoline contained (ng / mg), and T-GLYC, which is a quantitative value of the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained).

[0068] [Table 2]

[0069] Next, the collagen contained in the teeth of four human diabetic patients was quantified by high performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents. The following were determined: PYD, which is the quantitative value of the amount of pyridinoline (ng / mg), PEN, which is the quantitative value of the amount of pentosidine (ng / mg), T-GLYC, which is the quantitative value of the amount of total glycation end products (ng / mg: converted into the amount of pentosidine contained), PYD, which is the quantitative value of the amount of pyridinoline (ng / mg), and PEN, which is the quantitative value of the amount of pentosidine contained. Table 3 below shows PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)), which is a weight ratio value calculated from PEN, which is a quantitative value of the amount of pyridinoline contained (ng / mg), and PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained)), which is a weight ratio value calculated from PYD, which is a quantitative value of the amount of pyridinoline contained (ng / mg), and T-GLYC, which is a quantitative value of the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained).

[0070] [Table 3]

[0071] Next, the following data were obtained by high performance liquid chromatography with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pair reagents: PYD, which is the quantitative value of the pyridinoline content (ng / mg) of collagen contained in the teeth of 19 bovine animals; PEN, which is the quantitative value of the pentosidine content (ng / mg); T-GLYC, which is the quantitative value of the total glycation end products (ng / mg: converted into the pentosidine content); PYD, which is the quantitative value of the pyridinoline content (ng / mg) and the pentosidine content (ng / mg). PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)), which is a weight ratio value calculated from PEN, which is a quantitative value of the amount of pyridinoline contained (ng / mg), and PpG (amount of pyridinoline contained (ng / mg) / amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained)), which is a weight ratio value calculated from PYD, which is a quantitative value of the amount of pyridinoline contained (ng / mg), and T-GLYC, which is a quantitative value of the amount of total glycation products contained (ng / mg: converted into the amount of pentosidine contained), are shown in Table 4 below.

[0072] [Table 4]

[0073] As shown in Figure 1, the amount of pyridinoline (ng / mg) in collagen contained in the teeth of diabetic human patients increased slightly with age compared to the amount of pyridinoline (ng / mg) in collagen contained in the teeth of non-diabetic human patients and bovine teeth.

[0074] Furthermore, as shown in Figure 2, it was revealed that the amount of pentosidine (ng / mg) contained in collagen in the teeth of human diabetic patients increased with age compared to the amount of pentosidine (ng / mg) contained in collagen in the teeth of non-diabetic human patients and bovine teeth.

[0075] Furthermore, as shown in Figure 3, it was revealed that the total amount of glycation products contained in collagen (ng / mg: converted into the amount of pentosidine contained) in the teeth of human diabetic patients increased with age compared to the total amount of glycation products contained in collagen (ng / mg: converted into the amount of pentosidine contained) in the teeth of human non-diabetic patients and bovine teeth.

[0076] Furthermore, as shown in Figure 4, it was revealed that PpP (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) calculated from PYD, which is a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen, and PEN, which is a quantitative value of the amount of pentosidine (ng / mg) contained in collagen, decreased with age compared to PpP (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) calculated from PYD, which is a quantitative value of the amount of pyridinoline (ng / mg) contained in collagen, and PEN, which is a quantitative value of the amount of pentosidine (ng / mg) contained in collagen, in the teeth of human diabetic patients.

[0077] Furthermore, as shown in Figure 5, compared with PpG (amount of pyridinoline contained (ng / mg) / total amount of glycation products contained (ng / mg: converted to the amount of pentosidine contained)), which is a weight ratio calculated from PYD, which is a quantitative value of the amount of pyridinoline contained (ng / mg) in collagen contained in the teeth of human non-diabetic patients, and T-GLYC, which is a quantitative value of the amount of total glycation products contained (ng / mg: converted to the amount of pentosidine contained), it was revealed that PpG (amount of pyridinoline contained (ng / mg) / total amount of glycation products contained (ng / mg: converted to the amount of pentosidine contained)), which is a weight ratio calculated from PYD, which is a quantitative value of the amount of pyridinoline contained (ng / mg) in collagen contained in the teeth of human diabetic patients, and T-GLYC, which is a quantitative value of the amount of total glycation products contained (ng / mg: converted to the amount of pentosidine contained), decreased with age.

[0078] From the above, it is clear that these can be used as an index for assessing the potential risk of disease in mammals and / or whether a mammal is potentially healthy. Therefore, it is clear that a method using a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth, such as the method of the present invention, is effective as an index for assessing the potential risk of disease in mammals and / or whether a mammal is potentially healthy. It is clear that a method using a quantitative value of the amount of pentosidine contained in collagen contained in mammalian teeth, such as the method of the present invention, is effective as an index for assessing the potential risk of disease in mammals and / or whether a mammal is potentially healthy. It was shown that a method using a quantitative value of the total amount of glycation end products contained in collagen contained in mammalian teeth, such as the method of the present invention, is effective as an index for evaluating the potential risk of disease in a mammal and / or whether a mammal is potentially healthy; a method using a weight ratio value calculated from a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth and a quantitative value of the amount of pentosidine contained in collagen, such as the method of the present invention, is effective as an index for evaluating the potential risk of disease in a mammal and / or whether a mammal is potentially healthy; and a method using a weight ratio value calculated from a quantitative value of the amount of pyridinoline contained in collagen contained in mammalian teeth and a quantitative value of the total amount of glycation end products contained in collagen, such as the method of the present invention, is effective as an index for evaluating the potential risk of disease in a mammal and / or whether a mammal is potentially healthy.

Claims

1. A method of using a quantitative value of the amount of pyridinoline contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, comprising the steps of: quantifying the amount of pyridinoline contained in collagen contained in the teeth of a mammal that is to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value PYD; quantifying the amount of pyridinoline contained in collagen contained in the teeth of one or more mammals selected from non-disease mammals, diseased mammals, healthy mammals and unhealthy mammals, to obtain a reference quantitative value R-PYD; and comparing the PYD with the R-PYD.

2. A method of using a quantitative value of the amount of pentosidine contained in collagen contained in the teeth of a mammal as an index for assessing the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, comprising the steps of: quantifying the amount of pentosidine contained in collagen contained in the teeth of a mammal that is to be assessed for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value, PEN; quantifying the amount of pentosidine contained in collagen contained in the teeth of one or more mammals selected from non-disease mammals, diseased mammals, healthy mammals and unhealthy mammals, to obtain a reference quantitative value, R-PEN; and comparing the PEN with the R-PEN.

3. A method of using a quantitative value of the total amount of glycation products contained in collagen contained in the teeth of a mammal as an index for evaluating the potential risk of a disease in a mammal and / or whether the mammal is potentially healthy, comprising the steps of: quantifying the total amount of glycation products contained in collagen contained in the teeth of a mammal that is to be evaluated for the potential risk of a disease and / or whether the mammal is potentially healthy, to obtain a quantitative value of T-GLYC; quantifying the total amount of glycation products contained in collagen contained in the teeth of one or more mammals selected from non-disease mammals, diseased mammals, healthy mammals and unhealthy mammals, to obtain a reference quantitative value of R-T-GLYC; and comparing the T-GLYC with the R-T-GLYC.

4. A method of using a weight ratio calculated from a quantitative value of the amount of pyridinoline contained in collagen contained in a tooth of a mammal as an index for evaluating the potential risk of a disease in the mammal and / or whether the mammal is potentially healthy, comprising the steps of: quantifying the amount of pyridinoline contained in collagen contained in the tooth of a mammal that is to be evaluated for the potential risk of a disease and / or whether the mammal is potentially healthy to obtain a quantitative value PYD; quantifying the amount of pentosidine contained in collagen contained in the tooth of the mammal that is to be evaluated to obtain a quantitative value PEN; and calculating a weight ratio value PpP (amount of pyridinoline contained (ng / mg) / amount of pentosidine contained (ng / mg)) from the obtained PYD and PEN; The method includes the steps of: quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in teeth of one or more mammals selected from mammals not afflicted with a disease, mammals afflicted with a disease, healthy mammals, and unhealthy mammals, to obtain a reference quantitative value R-PYD; quantifying the amount of pentosidine (ng / mg) contained in collagen contained in teeth of the one or more mammals, to obtain a reference quantitative value R-PEN; calculating a weight ratio reference value R-Ppp (amount of pyridinoline (ng / mg) / amount of pentosidine (ng / mg)) from the obtained R-PYD and R-PEN; and comparing the Ppp and the R-Ppp.

5. A method for using a weight ratio calculated from a quantitative value of the amount of pyridinoline contained in collagen contained in a tooth of a mammal as an index for evaluating a potential disease risk in a mammal and / or whether the mammal is potentially healthy, the method comprising the steps of: quantifying the amount of pyridinoline contained in collagen contained in the tooth of a mammal that is to be evaluated for a potential disease risk and / or whether the mammal is potentially healthy to obtain a quantitative value of PYD; quantifying the amount of total glycation products contained in collagen contained in the tooth of the mammal that is to be evaluated (ng / mg: converted into the amount of pentosidine contained) to obtain a quantitative value of T-GLYC; and calculating a weight ratio value of PpG (amount of pyridinoline contained (ng / mg) / total amount of glycation products contained (ng / mg: converted into the amount of pentosidine contained)) from the obtained PYD and the obtained T-GLYC; The method includes the steps of: quantifying the amount of pyridinoline (ng / mg) contained in collagen contained in teeth of one or more mammals selected from non-diseased mammals, diseased mammals, healthy mammals, and unhealthy mammals to obtain a reference quantitative value R-PYD; quantifying the amount of total glycation products (ng / mg: converted into the amount of pentosidine contained) contained in collagen contained in teeth of the one or more mammals to obtain a reference quantitative value R-T-GLYC; calculating a weight ratio reference value R-PpG (amount of pyridinoline (ng / mg) / total amount of glycation products (ng / mg: converted into the amount of pentosidine contained)) from the obtained R-PYD and R-T-GLYC; and comparing the PpG and the R-PpG.

6. The method according to claim 1, wherein the step of quantifying is a step of quantifying by utilizing high performance liquid chromatography equipped with a fluorescence detector (HPLC-Flu) using heptafluorobutyric acid and formic acid as ion-pairing reagents.

7. The method according to any one of claims 1 to 6, wherein the mammal is one or more mammals selected from the group consisting of humans, cows, horses, pigs, wild boars, sheep, deer, dogs and cats.

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