CN111050775A - Method for improving bioavailability of glucosamine - Google Patents
Method for improving bioavailability of glucosamine Download PDFInfo
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- CN111050775A CN111050775A CN201880056066.9A CN201880056066A CN111050775A CN 111050775 A CN111050775 A CN 111050775A CN 201880056066 A CN201880056066 A CN 201880056066A CN 111050775 A CN111050775 A CN 111050775A
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- glucosamine
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Abstract
The invention aims to improve the bioavailability of glucosamine. The present invention relates to a method for improving the bioavailability of glucosamine, which is characterized by comprising orally ingesting glucosamine within 6 hours after fasting for 3 hours or more and after the time when the mRNA expression level of a Period gene reaches the maximum value in 1 day.
Description
Technical Field
The present invention relates to methods for increasing the bioavailability of glucosamine.
Background
Glucosamine (2-amino-2-deoxy-D-glucose) is an amino sugar in which the hydroxyl group at the 2-position of glucose is substituted with an amino group. Glucosamine has been used in health foods and the like to improve the symptoms of degenerative arthritis (OA), but absolute bioavailability (oral AUC/intravenous AUC) has been reported to be low for oral administration (non-patent document 1).
Documents of the prior art
Non-patent document
Non-patent document 1: Aghazadeh-Habashi A et al, J Pharm Pharmaceut Sci,2002,5(2), p.181-184.
Disclosure of Invention
The invention aims to improve the bioavailability of glucosamine.
The present inventors have conducted intensive studies to solve the above problems and found conditions for oral ingestion that can improve the bioavailability of glucosamine. Specifically, oral ingestion of glucosamine in consideration of the circadian rhythm of specific gene expression after fasting for a predetermined period of time can improve the absorption rate of glucosamine, thereby improving the bioavailability thereof, and the present invention has been completed.
That is, the present invention includes the following methods for improving the bioavailability of glucosamine.
[1] A method for improving the bioavailability of glucosamine, comprising orally administering glucosamine within 6 hours after fasting for 3 hours or more and after the time at which the mRNA expression level of a Period gene reaches the maximum in 1 day.
[2] The method according to the above [1], wherein the improvement in the bioavailability of glucosamine is based on an improvement in the absorption rate of glucosamine.
[3] The method according to the above [1] or [2], wherein the fasting time is 3 to 24 hours.
[4] The method according to any one of the above [1] to [3], wherein the subject is orally administered glucosamine from the time of getting up to the 1 st meal.
[5] The method according to any one of the above [1] to [4], wherein the oral ingestion of the glucosamine is followed by a fasting period of 15 minutes or more.
[6] A method for improving the bioavailability of glucosamine, comprising orally ingesting glucosamine in a subject between 2 hours and 8 hours after fasting for 3 hours or more and after a time when the mRNA expression level of a glucose transporter gene in the small intestine reaches the maximum level among 1 day.
[7] The method according to [6], wherein the improvement in the bioavailability of glucosamine is based on an improvement in the absorption rate of glucosamine.
[8] The method according to the above [6] or [7], wherein the glucose transporter is at least 1 selected from the group consisting of GLUT2 as glucose transporter 2, GLUT5 as glucose transporter 5, and SGLT1 as sodium-glucose cotransporter 1.
[9] The method according to any one of the above [6] to [8], wherein the fasting time is 3 to 24 hours.
[10] The method according to any one of the above [6] to [9], wherein the subject is orally ingested with glucosamine from the time of getting up to the 1 st meal.
[11] The method according to any one of the above [6] to [10], wherein the oral ingestion of the glucosamine is followed by a fasting period of 15 minutes or more.
[12] A method for indicating the time of ingestion of glucosamine for improving bioavailability, which is characterized by using the mRNA expression level of a Period gene as an indicator.
[13] A method for indicating a time of ingestion of glucosamine for improving bioavailability, characterized by using an mRNA expression level of a glucose transporter gene in the small intestine as an index.
[14] A preventive or ameliorating agent for arthralgia which comprises glucosamine and which is for oral ingestion by a subject in the case of (i) or (ii) below,
(i) after fasting for more than 3 hours, the time when the mRNA expression quantity of the Period gene reaches the maximum in 1 day is less than 6 hours later;
(ii) after fasting for more than 3 hours, and between 2 hours and 8 hours after the time when the mRNA expression level of the glucose transporter in the small intestine reaches the maximum of 1 day.
[15] The prophylactic or ameliorating agent for arthralgia according to [14], characterized in that the agent is indicated by the gist of "ingestion from the time of getting up to the 1 st meal".
According to the present invention, the bioavailability of glucosamine can be improved.
Drawings
FIG. 1 is a graph showing the change over time in the plasma glucosamine (GlcN) concentration in rats after a single administration of glucosamine hydrochloride (GlcN-HCl).
FIG. 2 is a graph showing the change over time of GlcN concentration in plasma of rats after a single administration of GlcN-HCl.
FIG. 3 is a graph showing the change over time of GlcN concentration in plasma of rats after a single administration of GlcN-HCl.
Fig. 4 is a graph showing the change over time of the GlcN concentration in the plasma of the free-feeding group and the fasting group at the time of administration 3 hours before administration ((a) is a group of 4:00 drugs, (b) is a group of 10:00 drugs, (c) is a group of 16:00 drugs, and (d) is a group of 22:00 drugs).
FIG. 5 is a graph showing the 24-hour rhythm of the expression levels of the mRNA of GLUT2 gene, GLUT5 gene, and SGLT1 gene in rat duodenal tissue ((a): GLUT2, (b): SGLT1, and (c): GLUT 5).
Detailed Description
The method for improving the bioavailability of glucosamine according to the first aspect of the present invention comprises orally ingesting glucosamine within 6 hours after fasting for 3 hours or more and after the time when the mRNA expression level of a Period gene (hereinafter also referred to as "Period gene") reaches the maximum level in 1 day. Hereinafter, the method for improving the bioavailability of glucosamine will also be referred to as the first method of the present invention. In this specification, the time at which the expression level of mRNA of Per gene reaches the maximum level in 1 day is also referred to as the peak expression time of Per gene. The expression level of the mRNA of the Per gene is within 6 hours after the time when the expression level of the mRNA of the Per gene reaches the maximum level in 1 day, and means from the time of the peak expression level of the Per gene to 6 hours after the peak expression level of the Per gene.
The method for improving the bioavailability of glucosamine according to the second aspect of the present invention comprises orally ingesting glucosamine in a range from 2 hours to 8 hours after fasting for 3 hours or more and after the time when the mRNA expression level of the glucose transporter gene in the small intestine reaches the maximum level in 1 day. Hereinafter, the method for improving the bioavailability of glucosamine will also be referred to as the second method of the present invention. In the present specification, the time at which the expression level of mRNA of the glucose transporter gene in the small intestine reaches the maximum level in 1 day is also referred to as the peak expression time of the glucose transporter gene.
In the present specification, the term "method" refers to the first and second methods of the present invention.
In the method of the present invention, the subject can take glucosamine orally in the above-mentioned specific time period after fasting for a predetermined time period, whereby the absorption rate of glucosamine can be improved and the bioavailability thereof can be improved. The improvement of bioavailability is helpful for enhancing the effect of glucosamine such as improvement of arthralgia. Further, it can contribute to the effect of the glucosamine function or the like with a smaller intake amount. In the present invention, it is preferable that the subject orally ingests glucosamine 1 time at the above-mentioned time.
In the method of the present invention, the fasting time is preferably 4 hours or more, more preferably 5 hours or more, further preferably 6 hours or more, and particularly preferably 7 hours or more. If the subject orally takes glucosamine in the specific time period after fasting for the above time period, the absorption rate of glucosamine is further improved, and the bioavailability thereof is further improved. During the fasting period in the present invention, water can be freely taken. Sleep is also included in the fasting time. The upper limit of the fasting time is appropriately set according to the subject, and is preferably 24 hours or less, more preferably 18 hours or less, further preferably 15 hours or less, and particularly preferably 12 hours or less. In one embodiment, the fasting time is preferably 3 to 24 hours, more preferably 3 to 18 hours, further preferably 3 to 15 hours, further preferably 4 to 12 hours, further preferably 5 to 12 hours, particularly preferably 6 to 12 hours, and most preferably 7 to 12 hours.
In the method of the present invention, in view of further improving the bioavailability of glucosamine, fasting is preferably performed for 15 minutes or more, and more preferably for 30 minutes or more after oral ingestion of glucosamine. In one embodiment, the upper limit of the fasting time after oral ingestion of glucosamine is preferably 4 hours or less.
In the present specification, the bioavailability of glucosamine (BA) is the bioavailability of orally ingested glucosamine, and can be evaluated by considering AUC (area under time curve of drug concentration in blood) at the time of intravenous administration as the bioavailability (absolute bioavailability) of the amount of uptake.
As shown in the test examples described later, a rhythm (circadian rhythm) showing a 24-hour cycle is present in the mRNA expression of the glucose transporter gene in the small intestine. And, by orally ingesting glucosamine to rats after fasting for 3 hours or more and between 2 hours and 8 hours after the peak time of expression of the glucose transporter gene, the absorption rate of glucosamine is improved, and the bioavailability thereof is improved.
In addition, it has been reported that in most organisms, circadian rhythms of various physiological phenomena such as metabolism are regulated by a group of genes called clock genes. The circadian rhythm of mRNA expression of the glucose transporter gene is also correlated with the circadian rhythm of mRNA expression of the clock gene.
In mammals, for Period genes and the like, their functions as clock genes are known. For example, in rodent mice, when raised in the environment of the light/dark phase with a 12-hour cycle, the peak time of expression of Per gene is usually around the start time of the dark phase belonging to the active phase (Yang X et al, Cell,2006,126(4), 801-10.). As shown in test examples described later, when rats were raised in the environment of the bright phase/dark phase with a 12-hour cycle, the peak expression time of the glucose transporter gene was about 3 hours before the start time of the dark phase, which is an active phase. Thus, the peak expression time of the glucose transporter gene is considered to be about 2 to 3 hours before the peak expression time of the Per gene. When considered as the Per gene, the time interval between 2 hours and 8 hours after the peak expression time of the glucose transporter gene is considered to correspond to a time zone within about 6 hours after the peak expression time of the Per gene. Thus, the bioavailability of glucosamine can be improved by orally taking glucosamine within 6 hours after the peak time of expression of Per gene after fasting for 3 hours or more.
Although the details of the reason why the bioavailability of glucosamine in a subject is improved by the method of the present invention are not known, it is considered that the possibility that the activity of the glucose transporter in the small intestine (for example, duodenum) is in a higher (preferably, maximum) time zone of 1 day may be considered between 2 hours and 8 hours after the peak time of expression of the glucose transporter gene in the small intestine. Glucosamine has been reported to be a substrate for glucose transporters such as glucose transporter 2(GLUT2) (Uidry et al, FEBS Lett.,2002,524, p.199-203). For example, in rodents such as rats and mice, and humans, fasting for 3 hours or more is considered to result in the substantial absence, or a very small amount, of monosaccharides such as glucose, which are substrates for glucose transporters, in the duodenum of the upper small intestine. It is presumed that, in the method of the present invention, in a time zone in which the activity of the glucose transporter in the small intestine is high, monosaccharides such as glucose in the small intestine (e.g., duodenum) are reduced, glucosamine can be increased, and the absorption rate of glucosamine can be improved.
In the first method of the present invention, the subject may orally ingest glucosamine at any time point within a time band of 6 hours after the peak time of expression of Per gene. From the viewpoint of further improving the bioavailability of glucosamine, the time period during which a subject orally ingests glucosamine is preferably within 5 hours, more preferably within 4 hours, and still more preferably within 3.5 hours after the expression peak time of the Per gene. In one embodiment, the time zone for oral ingestion of glucosamine is preferably 0.5 to 5 hours after the peak expression time of Per gene (0.5 to 5 hours after the peak expression time), more preferably 0.5 to 4 hours after the peak expression time, still more preferably 1 to 3.5 hours after the peak expression time, and particularly preferably 1.5 to 3 hours after the peak expression time.
Examples of Per genes include Period1(Per1), Period2(Per2) and Period3(Per3) in the Period gene family. The Per gene may be any of these.
The peak time of expression of Per gene can be confirmed, for example, by analyzing the change in the mRNA expression level of Per gene in a subject. The mRNA expression level of Per gene can be analyzed by any method known to those skilled in the art. For example, Northern blot analysis using any of the Per gene sequences as a probe, Real-time RT-PCR analysis, and expression level analysis using a DNA microarray can be suitably used. As a sample for gene expression analysis, any substance may be used as long as it contains cells, and for example, hair containing hair follicle cells, biopsy such as skin, blood, saliva, and the like may be used. Samples such as hair are easy to collect, and have the advantage of low sharing of the subject. The first method of the present invention may further comprise analyzing the change in the mRNA expression level of the Per gene using a sample obtained from the subject to determine the expression peak time of the Per gene.
For example, in humans, the peak time of expression of Per gene is usually around the time of getting up, more specifically, about 1 to 2 hours before the time of getting up (Akashi M et al, "Noninvasive method for assisting the human circa circular clock using hair follicles.", Proc Natl Acad Sci USA.2010Aug 31; 107(35): 15643-8.). More specifically, the peak time of Per gene expression in humans is usually about 4 to 10 am, more specifically about 5 to 9 am (Akashi M et al). For example, it is preferable to take glucosamine orally within 6 hours after the time. In one embodiment, the human is preferably 5 am to 2 pm, more preferably 6 am to 12 am, as a time zone within 6 hours after the peak expression time of the Per gene.
In the second method of the present invention, the subject may orally ingest glucosamine at any time point in a time band between 2 hours and 8 hours after the peak expression time of the glucose transporter gene (after 2 to 8 hours after the peak expression time). From the viewpoint of further improving the bioavailability of glucosamine, the time period for oral ingestion of glucosamine by a subject is preferably between 3 hours and 7 hours after the peak time of expression of the glucose transporter gene, more preferably between 3.5 hours and 7 hours after the peak time, still more preferably between 3.5 hours and 6.5 hours after the peak time, and particularly preferably between 4 hours and 6.5 hours after the peak time. In the second method of the invention, the small intestine is preferably the duodenum.
Examples of the glucose transporter include glucose transporter 2(GLUT2), glucose transporter 5(GLUT5), and sodium-glucose cotransporter 1(SGLT 1). The glucose transporter gene of the present invention may be any of these genes. In one form, GLUT2 is preferred. In the present invention, glucosamine is preferably orally taken between 2 hours and 8 hours after the expression peak time of preferably 1 or more genes among the GLUT2 gene, GLUT5 gene, and SGLT1 gene, more preferably the expression peak time of GLUT2 gene, GLUT5 gene, and SGLT1 gene. In one embodiment of the present invention, glucosamine is preferably orally ingested between 2 hours and 8 hours after the peak time of expression of GLUT2 gene.
The expression peak time of the glucose transporter gene can be confirmed, for example, by a change in the mRNA expression level of the glucose transporter gene in the small intestine to be analyzed. The mRNA expression level of the glucose transporter gene can be analyzed by any method known to those skilled in the art. For example, Northern blot analysis using any of the gene sequences as a probe, Real-time RT-PCR analysis, expression analysis using a DNA microarray, and the like can be suitably used. The sample to be used for gene expression analysis may be any sample containing small intestinal tissue (preferably, duodenal tissue). The second method of the present invention may further comprise analyzing a change in the expression level of the glucose transporter gene in the small intestine using a sample taken from the subject to determine the expression peak time of the glucose transporter gene.
As shown in test examples described later, when rats were raised in the environment of the light phase/dark phase of a 12-hour cycle, the peak time of the expression of the glucose transporter gene was around 3 hours before the start time of the dark phase. In the normal life of human, the daytime and nighttime of sleep and food intake are generally opposite to those of rats, and it is presumed that the peak expression time of glucose transporter genes (e.g., GLUT2 gene, GLUT5 gene, SGLT1 gene, etc.) in human is generally about 1 to 7 am, more specifically about 3 to 6 am, and even more specifically about 4 to 5 am. For example, glucosamine is preferably orally ingested between 2 hours and 8 hours thereafter. In one embodiment, the human preferably orally ingests glucosamine between 5 am and 2 pm, and more preferably between 6 am and 12 am. In general, the time zone is between 2 hours and 8 hours after the peak time of expression of the glucose transporter gene, and is one example of a preferred embodiment of the present invention.
In the method of the present invention, glucosamine is orally ingested in the above-mentioned specific time zone after fasting for 3 hours or more. As an example of a preferable mode of the time zone for oral ingestion of glucosamine, there is a mode from the time of getting up to the 1 st meal in 1 day of life of animals such as humans. Orally ingesting glucosamine from a subject after fasting for 3 hours or more and from the time of getting up to the 1 st meal is an example of a preferred embodiment of the present invention.
As described above, the peak time of expression of Per gene in humans is usually around the time of getting up. In one embodiment of the first method of the present invention, for example, in the case of human, glucosamine can be orally taken within 3.5 hours, more preferably within 3 hours after the time of getting up as a time band within 6 hours after the peak expression time of Per gene. In this case, the time period for oral ingestion of glucosamine is, for example, preferably 0.5 to 3.5 hours after the time of getting up, and more preferably 1 to 3 hours after the time of getting up. In the case of humans, it is preferable that glucosamine be taken in the time period from the time of getting up to the 1 st meal (e.g., breakfast) (e.g., within 3.5 hours after getting up). The above time zone is an example of a preferable embodiment of the time between 2 hours and 8 hours after the peak time of expression of the glucose transporter gene in the second method of the present invention.
In addition, in rodents such as mice and rats, the time zone within 6 hours after the peak time of expression of the Per gene is, for example, preferably between 0.5 and 6 hours after the start of the dark phase, more preferably between 0.5 and 5 hours after the start of the dark phase, still more preferably between 0.5 and 4 hours after the start of the dark phase, still more preferably between 1 and 3.5 hours after the start of the dark phase, and particularly preferably between 1.5 and 3 hours after the start of the dark phase. The above time zone is also an example of a preferable embodiment of the time between 2 hours and 8 hours after the peak time of expression of the glucose transporter gene in the second method of the present invention.
By taking glucosamine over the above time zone, the absorption rate of glucosamine is improved, and the bioavailability thereof is improved.
The first method and the second method of the present invention are further described below.
The glucosamine in the present invention is glucosamine, a derivative thereof, or a salt thereof, and 1 kind thereof may be used, or 2 or more kinds thereof may be used in combination. Examples of the glucosamine derivative include N-acetylglucosamine and N-methyl-L-glucosamine. The salt of glucosamine or a derivative thereof includes hydrochloride, sulfate, lactate, and the like, and the hydrochloride is preferred. Among these, 1 or 2 or more of glucosamine, a salt of glucosamine, and N-acetylglucosamine can be suitably used as the glucosamine. The glucosamine is more preferably glucosamine, glucosamine hydrochloride, glucosamine sulfate, and acetylglucosamine, and still more preferably glucosamine and glucosamine hydrochloride.
The glucosamine usable in the present invention is not particularly limited with respect to its origin, production method, and the like. For example, glucosamine can be obtained by hydrolyzing chitin contained in crustaceans such as crabs, shrimps, and krill, or cartilage of cuttlefish, etc. with an acid or an enzyme, and separating and purifying the chitin. Commercially available glucosamine can also be used.
The amount of glucosamine to be orally taken (intake amount) is, for example, 50 to 5000mg, more preferably 500 to 3000mg per day per individual in the case of adult human. The amount of intake per kg of body weight per day may be, for example, 10 to 100mg/kg, and more preferably 10 to 60 mg/kg. In the method of the present invention, oral ingestion of the above amount of glucosamine is preferably accomplished 1 time. Further, in one embodiment of the present invention, oral ingestion of glucosamine is preferably 1 time a day.
In the method of the present invention, glucosamine can be directly orally ingested. Further, other components may be formulated into glucosamine and orally taken as a composition as desired. In one embodiment, the composition containing glucosamine is preferably ingested orally. The glucosamine-containing composition is preferably a composition containing glucosamine as an active ingredient. The glucosamine-containing composition is preferably in a form to be orally ingested, such as a food or drink (food or drink composition), a medicine for oral administration (oral medicine composition), or an animal feed, and more preferably a food or drink or an animal feed. The food or drink is not particularly limited, and examples thereof include general food or drink, health food, functional marker food, specific health food, patient food, and food additives.
The method of the present invention includes allowing the subject to ingest a food or drink containing glucosamine indicated by the expression "ingest from the time of getting up to the 1 st meal" in the time zone. In the present invention, the indication of the substance "intake from the time of getting up to the 1 st meal" includes, as an example, indications such as "intake at the same time of waking up", "intake first in the morning", "intake at the same time of waking up", "intake first in waking up", "intake at the start of 1 day", "intake before breakfast", "intake with energy supply first as breakfast", "intake at the time of getting up", "intake before 1 day of activity", and "intake before 1 day of meal". Examples of labeling methods include labeling of packages, containers, instructions, advertisements, and the like for foods and beverages containing glucosamine.
The composition containing glucosamine may contain any additives and any components as desired within a range not impairing the effects of the present invention. As these additives and components, substances generally usable in oral compositions such as foods, drinks, oral medicines, quasi drugs for oral administration, and feeds can be used. For example, a sweetener, an acidulant, etc. may be contained. In addition, excipients, binders, emulsifiers, tonicity agents (isotonicity agents), buffers, solubilizers, preservatives, stabilizers, antioxidants, colorants, coagulants, coating agents, flavors, and the like, which are formulated in the formulation, may be contained. The composition may further contain active ingredients other than glucosamine, such as chondroitin, enzyme-treated rutin, collagen, hyaluronic acid, vitamins, etc. These optional components can be used in 1 kind, or can be used in combination of 2 or more. In one embodiment, the glucosamine-containing composition preferably does not contain a substrate for a glucose transporter (preferably GLUT2), such as a monosaccharide such as glucose, or even contains a small amount of glucosamine.
The form of the glucosamine-containing composition is not particularly limited. For example, in the case of food and drink, liquid food such as drink, semisolid food, solid food, and the like may be used. In addition, it can also be made into oral solid preparation such as tablet, capsule, powder, chewable tablet, etc.; various dosage forms such as oral liquid, syrup, etc. The oral pharmaceutical product and the quasi-oral pharmaceutical product may be those suitable for oral administration.
The method for producing a glucosamine-containing composition such as foods, drinks, medicines, quasi drugs, and feeds is not particularly limited, and the composition can be produced by a general method using glucosamine.
The blending ratio of glucosamine in the glucosamine-containing composition is not particularly limited and can be appropriately set. For example, the total amount of glucosamine in the composition is preferably 0.1 to 95% by weight, more preferably 10 to 80% by weight.
In the method of the present invention, the subject to which glucosamine is orally ingested is preferably a mammal (human or non-human mammal), more preferably a human. Examples of the non-human mammal include cows, horses, goats, dogs, cats, rabbits, mice, rats, guinea pigs, and monkeys. As the object in the method of the present invention, an object that desires or requires an improvement in the bioavailability of glucosamine is preferable. In addition, glucosamine has an effect of improving the symptoms of degenerative arthritis and the like, and thus can be used for prevention or improvement of joint pain, for example. In the present invention, a subject who feels uncomfortable in the knee joint or a subject who desires or needs prevention or improvement of degenerative arthritis is preferable. Prevention includes preventing, delaying and reducing the incidence of disease. The improvement includes alleviation of symptoms, inhibition of symptom development, and cure of symptoms. The improvement of the bioavailability of glucosamine can contribute to, for example, the prevention or improvement of knee joint pain and the like, and the prevention or improvement of degenerative arthritis.
The methods of the invention may be therapeutic or non-therapeutic. By "non-therapeutic" is meant a concept that does not encompass medical activities, i.e., surgery, therapy, or diagnosis.
As described above, the mRNA expression level of the Per gene in the subject or the mRNA expression level of the glucose transporter gene in the small intestine can be used as an index for obtaining the time taken for glucosamine to be taken (the time taken for glucosamine to be taken) for improving the bioavailability. It is known that a time period for increasing the bioavailability of glucosamine is useful for subjects desiring or requiring more sufficient effect based on glucosamine, such as prevention or improvement of arthralgia.
The present invention also includes a method for indicating the time taken to take glucosamine for improving bioavailability, using the mRNA expression level of Per gene as an indicator. In this method, the expression level of mRNA of Per gene is used as an index, and the time period of 6 hours after the time when the expression level reaches the maximum of 1 day is indicated as the time period for improving the bioavailability of glucosamine. In one embodiment, the method preferably presents the time taken for the glucosamine to be taken up for improving bioavailability after fasting for 3 hours or more and within 6 hours after the time taken for the expression level to reach the maximum value in 1 day, using the mRNA expression level of the Per gene and the fasting time as indices. A preferable mode of the time zone within 6 hours after the peak time of expression of Per gene is as described above. The analysis of the mRNA expression level of Per gene in the subject can be carried out by the above-mentioned method. The method of the present invention may further comprise determining the expression peak time of Per gene by analyzing the change in the expression level of Per gene using a sample taken from the subject.
The present invention also includes a method for indicating the time taken to take glucosamine for improving bioavailability, using the amount of mRNA expression of the glucose transporter gene in the small intestine as an indicator. In this method, the expression level of mRNA of the glucose transporter gene in the small intestine is used as an index, and the time between 2 hours and 8 hours after the time when the expression level reaches the maximum of 1 day is indicated as the time for improving the uptake of glucosamine. In one embodiment, the method preferably suggests, as the time for increasing the glucosamine bioavailability uptake, an amount of mRNA expressed from the glucose transporter gene and a fasting time as indices, between 2 hours and 8 hours after fasting for 3 hours or more and after the time at which the expression amount reaches the maximum of 1 day. Preferred modes of the time zone between 2 hours and 8 hours after the peak time of expression of the glucose transporter gene are as described above. Analysis of the expression level of the glucose transporter gene in the small intestine of the subject and the like can be carried out by the above-described method. The method of the present invention may further comprise analyzing a change in the expression level of the glucose transporter gene in the small intestine using a sample obtained from the subject to determine the expression peak time of the glucose transporter gene.
The present invention also encompasses the following preventive or ameliorating agents for arthralgia which contain glucosamine.
A preventive or ameliorating agent for arthralgia which comprises glucosamine and which is for oral ingestion by a subject in the case of (i) or (ii) below,
(i) after fasting for more than 3 hours, the time when the mRNA expression quantity of the Period gene reaches the maximum in 1 day is less than 6 hours later;
(ii) after fasting for more than 3 hours, and between 2 hours and 8 hours after the time when the mRNA expression level of the glucose transporter in the small intestine reaches the maximum of 1 day.
The preventive or ameliorating agent for joint pain according to the present invention contains glucosamine as an active ingredient.
By orally ingesting glucosamine in the time zone of (i) or (ii), the bioavailability of glucosamine is improved, and the effect of preventing or ameliorating joint pain due to glucosamine is enhanced. Preferred modes of taking the glucosamine are as described above. The preventive or ameliorating agent for arthralgia of the present invention is preferably orally ingested 1 time a day.
The preventive or ameliorating agent for arthralgia according to the present invention may be a composition containing any of the above additives and any of the above components. The composition containing glucosamine and preferred embodiments thereof are the same as those described above. The amount of glucosamine to be taken is also as described above.
The preventive or ameliorating agent for arthralgia of the present invention can be formulated into the above-mentioned oral pharmaceuticals, oral quasi drugs, foods, drinks, feeds, and the like. The preventive or ameliorating agent for arthralgia of the present invention may be one in which 1 or 2 or more of the uses, types of active ingredients, methods of use, and the like are indicated in packages, containers, instructions, additional documents, advertisements, and the like. In one embodiment, the prophylactic or ameliorating agent for arthralgia according to the present invention may be indicated by the above-mentioned "ingestion from the time of getting up to the 1 st meal".
Examples
The present invention will be described in further detail below with reference to test examples, but the scope of the present invention is not limited thereto. The time is expressed in 24 hours (0: 00-24: 00). In addition, a series of animal experiments are performed in accordance with the animal care management law and other relevant laws and under the examination of animal experiment committees in companies and with the plan of long-term approval.
< Experimental animals >
7-week-old Lewis male rats were purchased from Japan SLC corporation. Rats were housed in groups of 2-3 rats in plastic cages, and were kept for 1 week or more indoors under conditions of free drinking water, 12 hours light-dark cycle (bright period: time 7: 00-19: 00), and constant temperature (24. + -. 1 ℃ C.) for experiments.
< glucosamine >
D (+) -glucosamine hydrochloride (hereinafter, GlcN-HCl) (manufactured by Protein Chemical Co., Ltd.) was used by dissolving it in an optionally used Otsuka distilled water (Otsuka pharmaceutical factory). Rats were dosed with GlcN-HCl aqueous solution at 2 mL/kg.
In the following experiments, the glucosamine (GlcN) concentration in plasma was measured according to the following method.
< determination of GlcN concentration in plasma >
Blood was collected from the tail vein. The collected blood was immediately subjected to EDTA treatment, and then centrifuged at 3,000 Xg at 15 ℃ for 10 minutes to obtain plasma. Plasma was stored frozen at-80 ℃ until assayed.
For the measurement of GlcN concentration in plasma, an internal standard method is used. Using 13C-glucosamine as an internal standard, 5. mu.L of the internal standard solution and 200. mu.L of acetonitrile were added to 40. mu.L of plasma and stirred, and after leaving on ice for 10 minutes, centrifugation was carried out (4 ℃,12,000 rpm, 10 min). The GlcN concentration was measured for 200. mu.L of the obtained supernatant using LC-MS/MS under the analysis conditions described later. The GlcN concentration in plasma before administration of GlcN-HCl was set to the GlcN concentration in plasma at the time of 0 hour of administration.
LC-MS/MS analysis conditions for GlcN concentration in plasma
< HPLC Condition >
A chromatographic column: asahipak NH 2P-502D 5.0 μm, 2.0X 150mm (Shodex) (Showa Denko K.K.)
Protection of the column: asahipak NH2P-50G 2A 5.0 μm, 2.0X 30mm (Shodex) (Showa Denko K.K.)
Mobile phase: 10mM aqueous acetic acid (adjusted to pH7.5 with ammonia): acetonitrile 20: 80(v/v)
Flow rate: 0.4mL/min
Injection amount: 10.00 mu L
Column box: 28 +/-5 DEG C
A sample cooler: 10 +/-5 DEG C
< MS/MS Condition >
An MS detector: quatro micro LC/MS/MS (Waters Corp.)
Detecting ions: 180.1 → 162.1
Ion spray voltage: 3,500V
Temperature: 350 deg.C
< statistical analysis >
One-way ANOVA was used for comparisons between groups, and Scheffe's test was used for comparisons between specific 2 groups. Student's t-test was used in the independent 2-group comparisons. The Cosiner method was used in the periodic assessment of the presence or absence of 24 hours. The significance level was set at 5%.
< test example 1 >
(influence of fasting feeding environment on the in vivo kinetics of GlcN)
The effect of ingestion on the GlcN concentration in plasma was examined. At time 10: 00A single oral administration of GlcN-HCl 500mg/kg was given to rats fed either ad libitum or under a fasting condition (time 22: 00-10: 00) 12 hours prior to administration. The free feeding group (n ═ 3) was kept in a free feeding environment from the start of the test to the end of the test. The fasting group (n ═ 3) 12 hours before administration was fed under fasting conditions from 12 hours before GlcN-HCl administration to the end of the experiment. Both groups were raised in an environment of free drinking water at any time.
Blood was collected from the tail vein of rats before and after administration of GlcN-HCl for 0.083, 0.25, 0.5, 1, 2, 3, and 4 hours, and the GlcN concentration in plasma was measured by the above method. The results are shown in FIG. 1.
Fig. 1 is a graph showing the change over time of the GlcN concentration in the plasma of rats after a single administration of GlcN-HCl, the values shown in fig. 1 are the mean ± s.d. (n-3), in fig. 1, ○ is the free-feeding group, ● is the fasting group 12 hours before administration, and the horizontal axis is the elapsed time after the administration of GlcN-HCl.
No significant difference was observed between the dual drug groups for GlcN concentrations in plasma at each assay time point.
AUC was calculated from the GlcN concentration in plasma at each measurement time point0-4hr. AUC in each group0-4hrThe mean value of (A) was 12.2 (. mu.g/mL. hr) in the free-feeding group, 14.1 (. mu.g/mL. hr) in the fasted group 12 hours before administration, and AUC between the two groups0-4hrNo significant difference was observed. From these results, fasting administration of GlcN-HCl failed to increase GlcN uptake.
< test example 2 >
(influence of the timing of administration on the GlcN concentration in plasma)
At time 4: 00. 10: 00. 16:00 or 22: 00A single oral administration of GlcN-HCl 500mg/kg (n-6 for each group) was given to rats in a free feeding environment. Rats were fasted following GlcN-HCl administration. Further, the breeding was performed in a free-drinking environment from the start of the test to the end of the test.
Blood was collected from the tail vein of rats before and after administration of GlcN-HCl for 0.083, 0.25, 0.5, 1, 2, 3, and 4 hours, and the GlcN concentration in plasma was measured by the above method. The results are shown in FIG. 2.
Fig. 2 is a graph showing the change over time of the GlcN concentration in the plasma of rats after a single administration of GlcN-HCl, the values shown in fig. 2 are the average ± s.d. (n is 6), in fig. 2, ▲ is the group of 4:00, ○ is the group of 10:00, △ is the group of 16:00, ● is the group of 22:00, and the horizontal axis of fig. 2 is the elapsed time after the administration of GlcN-HCl.
No significant difference was observed in plasma GlcN concentration at each assay time point between all drug groups.
AUC was calculated from the GlcN concentration in plasma at each measurement time0-4hr. AUC of each group0-4hrIs in the range of 4: the composition of 00 medication is 19.5 (mu g/mL & hr), and the dosage is 10: the composition of 00 drugs is 15.5 (mu g/mL. hr), and the ratio is 16: the composition of 00 drugs is 16.8 (mu g/mL. hr), and the ratio is 22: the drug group of 00 was 16.4(μ g/mL. hr), AUC between all groups0-4hrNo significant difference was observed. From these results, it was not possible to improve the GlcN absorption rate depending on the administration timing in the free ingestion environment.
< test example 3>
(diurnal variation of spontaneous locomotion and feeding action in rats)
The feeding action and activity rhythm of the rat were measured by the following methods.
The spontaneous exercise amount and food intake amount of rats were measured every 15 minutes for rats who had free access to drinking water and had a 12-hour light-dark cycle (light period: time 7: 00-19: 00). For the measurement, rats were individually housed in plastic measurement cages using cFDM-700AS (a device for measuring amount of bait with a bait restriction function for rats, MELQUEST).
19: activity began at around 00 f, and active activity was intermittently observed during the dark period. The average of the activity amounts in the dark phase and the light phase on each measurement day was 8095.3(counts/12hr), the light phase was 2090.1(counts/12hr), and the average of the food intake was 12.8(g/12hr) in the dark phase and 5.4(g/12hr) in the light phase. The ratio of the amount of the total amount of the activity in the dark phase to the amount of the activity in the dark phase was about 80%, and the ratio of the amount of the total amount of the food intake in the dark phase to the amount of the food intake in the dark phase was about 70%, and it was confirmed that spontaneous exercise and feeding action were actively performed during the dark phase.
The average values of spontaneous exercise amount and food intake amount per3 hours on each measurement day are shown in table 1. As a result, the peak values of spontaneous exercise amount and food intake amount were 19: 00 to 22: time zone 00. Further, with 19: 00 to 22: the diurnal variation of the spontaneous exercise amount and the food intake amount in the time zone of 00as the peak was continuously confirmed in each measurement day during the measurement period.
[ Table 1]
< test example 4 >
(influence of the difference in the administration timing of the drug to the GlcN concentration in plasma of rats fed in a fasting environment 3 hours before administration)
Rats fed in a fasted environment 3 hours before self-administration were used as subjects at time 4: 00. 10: 00. 16:00 or 22: 00A single oral administration of GlcN-HCl 500 mg/kg. Rats were fed in a fasted environment 3 hours before administration of GlcN-HCl until the end of the experiment, but were allowed free water uptake.
Blood was collected from the tail vein of rats before and after administration of GlcN-HCl for 0.083, 0.25, 0.5, 1, 2, 3, and 4 hours, and the GlcN concentration in plasma was measured by the above method. The results are shown in FIG. 3.
FIG. 3 is a graph showing the change over time of GlcN concentration in plasma of rats after a single administration of GlcN-HCl. The values shown in fig. 3 are the average ± s.d. (n ═ 6) (. P <0.05 vs.4: 00 drugs used;. P <0.05,. P <0.01 vs.10:00 drugs used;
：P＜0.05，
p <0.01 vs.16:00 (Scheffe's test)), in FIG. 3, ▲ is 4:00, ○ is 10:00, △ is 16:00, ● is 22:00, and the horizontal axis in FIG. 3 is the elapsed time after GlcN-HCl administration.
Regarding the GlcN concentration in plasma after 0.25 hours of administration of GlcN-HCl, 22: group 00 and 16: the group 00 showed significantly higher values (P <0.05) compared to the group. Further, regarding the GlcN concentration in the plasma after 0.5 hour and 1 hour of the administration of GlcN-HCl, 22: the 00 group showed significantly higher values (P <0.05 and P <0.01, respectively) compared to the other groups. Further, regarding the GlcN concentration in plasma after 2 hours of administration of GlcN-HCl, 4:00 medicine group and 22: group 00 and 16: the 00 dose group showed significantly higher values (P <0.05, respectively).
AUC was calculated from the GlcN concentration in plasma at each measurement time0-4hr. AUC of each group0-4hrIs in the range of 4: the composition of 00 medication is 19.5 (mu g/mL & hr), and the dosage is 10: the composition of 00 drugs is 16.1 (mu g/mL. hr), and the ratio of 16: the composition of 00 medication is 12.5 (mu g/mL. hr), and the ratio is 22: the drug combination for 00 is 27.8 (mu g/mL. multidot.hr). 22: AUC of 00 drug group0-4hrRelative to the weight ratio of 10:00 drug group and 16: AUC of 00 drug group0-4hr1.73 times and 2.22 times respectively (vs.10:00 medication group: P)<0.05, vs.16:00 dosing group: p<0.01)。
(influence of differences in the conditions of food intake at the time of administration on the GlcN concentration in plasma)
Using the data shown in fig. 2 and 3, the change over time of GlcN concentration in plasma in the free-feeding group and the fasting group 3 hours before administration at each administration time is shown in fig. 4((a) is a group of drugs of 4:00, (b) is a group of drugs of 10:00, (c) is a group of drugs of 16:00, (d) is a group of drugs of 22: 00). the values shown in fig. 4(a) to (d) are average ± s.d. (n is 6) (.: P <0.05 vs. each of the feeding groups (Student's t-test)). in fig. 4(a) to (d), □ is the free-feeding group, and ■ is the fasting group 3 hours before administration.
22:00 medicine (fig. 4(d))The GlcN concentration in plasma of the fasted group at 3 hours before the administration of the drug showed significantly higher values (P, respectively) at all the measured time points from 0.083 hour to 1 hour after the administration, as compared with the free-feeding group<0.05), AUC of fasting group 3 hours before administration0-4hr1.70 times (P) compared to the free-feeding group<0.05)。
In the following step 4:00 (fig. 4(a)) and 10:00 (FIG. 4(b)), the GlcN concentration and AUC in plasma at each measurement time point0-4hrNo significant difference was observed between the dual drug groups. At 16: in 00 (FIG. 4(c)), the GlcN concentration in plasma of the fasting group at 3 hours before administration showed significantly lower values (P) at 0.5 hour and 1 hour after administration, respectively, than that of the free-feeding group<0.05), AUC of fasting group 3 hours before administration0-4hrCompared with the free feeding group, the ratio is 0.74 times (P)<0.05)。
As such, only at 22: GlcN concentration in plasma was increased in the fasting group 3 hours before administration of 00 drugs. From the above, it was found that the blood concentration of GlcN significantly increases when glucosamine is taken without ingestion for several hours at a specific administration time.
< test example 5 >
(evaluation of circadian rhythm of mRNA expression of Transportin in Small intestine tissue)
Changes in the mRNA expression levels of glucose transporters (GLUT2, GLUT5, and SGLT1) in the small intestine of rats were examined.
(Collection of Small intestinal tissue)
Rats fed in an environment fasted 3 hours before self-organizing collection (free water intake) were treated with the following formula at time 4: 00. 10: 00. 16: 00. 22:00 under general anesthesia PBS (-) was injected from the lower aorta and duodenum was taken after exsanguination to death. Tissue samples were cryopreserved at-80 ℃.
(Total RNA extraction)
To the collected tissue, 500. mu.L of RNAioso (TAKARA Bio Inc.) was added, and homogenization was performed under ice-cooling. To the homogenate was added 100. mu.L of chloroform, the mixture was shaken and allowed to stand at room temperature for 5 minutes, and then centrifuged at 12,000 Xg at 4 ℃ for 15 minutes. The supernatant was collected, 300. mu.L of isopropyl alcohol was added thereto, and the mixture was mixed with inversion and then allowed to stand at room temperature for 10 minutes. After centrifugation at 12,000 Xg at 4 ℃ for 10 minutes, the supernatant was removed. After the residue was centrifuged at 7,500 Xg and 4 ℃ for 5 minutes with 75% ethanol added thereto, the supernatant was removed and air-dried, and the residue was dissolved in 22. mu.L of TE buffer. After the RNA concentration was measured using a minispectrophotometer (LMS-Nanodrop 1000, Thermo Fisher Scientific Co., Ltd.), TE buffer was added to adjust the concentration to 500 ng/. mu.L, and the Total RNA sample was obtained.
(genomic DNA (gDNA) removal and cDNA Synthesis)
The following experiments were performed according to the guidelines of the PrimeScript RT reagent Kit (TAKARA Bio Inc.). Total RNA sample 1. mu.L, 5 XgDNA Eraser Buffer 1. mu.L, gDNA Eraser 0.5. mu.L, RNase freewater 2.5. mu.L were mixed. The gDNAse treatment was performed using PCR Thermal Cycler (Applied Biosystems) at 42 ℃ for 2 minutes. Then, 2.5. mu.L of 5 XPrimeScript Buffer 2(for Real Time), 0.5. mu.L of RT Primer Mix 0.5. mu. L, PrimeScript (registered trademark), 0.5. mu.L of RT Enzyme Mix I, and 2.5. mu.L of RNase free water were mixed and reacted at 37 ℃ for 15 minutes, 85 ℃ for 5 seconds using a PCR Thermal Cycler to synthesize cDNA.
(Real-time RT-PCR method)
The expression level of each mRNA was measured by Real-time RT-PCR. For Real-time PCR reaction, KODSYBR qPCR Mix (TOYOBO CO., LTD), GAPDH, GLUT2, GLUT5, and SGLT1 primers were used, and the sequences shown below were used.
Rat GAPDH(GenBank accession number:NM_017008.4)
Forward: 5'-AAAGCTGTGGCGTGATGG-3' (SEQ ID NO: 1)
Reverse: 5'-TTCAGCTCTGGGATGACCTT-3' (SEQ ID NO: 2)
Rat GLUT2(GenBank accession number:NM_012879.2)
Forward: 5'-TGTGGGCTAATTTCAGGACTGG-3' (SEQ ID NO: 3)
Reverse: 5'-AAGAGCCAGTTGGTGAAGAGTG-3' (SEQ ID NO: 4)
Rat GLUT5(GenBank accession number:NM_031741.1)
Forward: 5'-ATCTTCTCCTCATCGGCTTCTC-3' (SEQ ID NO: 5)
Reverse: 5'-CAATGACACAGACGATGCTGAC-3' (SEQ ID NO: 6)
Rat SGLT1(GenBank accession number:NM_013033.2)
Forward: 5'-TGGAGTCTACGTAACAGCACAG-3' (SEQ ID NO: 7)
Reverse: 5'-GTCATATGCCTTCCTGAAGCAC-3' (SEQ ID NO: 8)
mu.L of cDNA sample, 0.2. mu.L of forward primer (final concentration 0.2. mu.M), 0.2. mu.L of reverse primer (final concentration 0.2. mu.M), 0.4. mu.L of ROX reference Dye, 10. mu.L of KOD SYBR qPCR Mix, and 7.2. mu.L of RNase freewater were mixed. Initial denaturation was carried out at 98 ℃ for 2 minutes using StepOneNus (applied biosystems), and then PCR was carried out for 40 cycles at 98 ℃ for 10 seconds for dissociation, 60 ℃ for 10 seconds for annealing, 68 ℃ for extension, and 30 seconds for extension. Further, the melting temperature was measured after the PCR reaction.
The mRNA expression level of GAPDH gene was measured as an internal standard, and the relative mRNA expression level of each gene relative to GAPDH gene was calculated.
FIG. 5 is a graph showing the 24-hour rhythm of the expression levels of the mRNA of the GLUT2 gene, the GLUT5 gene, and the SGLT1 gene in rat duodenal tissue ((a): GLUT2, (b): SGLT1, and (c): GLUT 5). The values shown in fig. 5(a) to (c) are the average ± s.d. (n is 4). The results shown in FIGS. 5(a) to (c) are relative mRNA amounts to the mRNA amount of the GAPDH gene.
Circadian rhythms were observed in the mRNA expression levels of GLUT2, GLUT5, and SGLT1 (GLUT 2: F from ANOVA 10.149, P <0.01, P from Cosinor < 0.01; GLUT 5: F from ANOVA 23.143, P <0.01, P from Cosinor < 0.01; SGLT 1: F from ANOVA 33.928, P <0.01, P from Cosinor < 0.01).
In the mRNA expression levels of GLUT2 and GLUT5, it was observed that at time 16:00 shows the highest value, in 4:00 showed the lowest significant circadian rhythm (P <0.01, respectively). In addition, in the mRNA expression level of SGLT1, the expression level was observed to be 16:00 shows the highest value, at 10:00 showed a significant circadian rhythm of lowest value (P < 0.01).
< test example 6 >
value for bioavailability of glucosamine
Rats fed in a fasting environment 3 hours before free feeding or self-administration were treated at time 22: GlcN-HCl (50mg/kg) was administered intravenously at the 00 tail. Blood was collected from the tail vein at each time from the administration of GlcN-HCl to 0.083, 0.25, 0.5, 1, 2, 3, and 4 hours, and the GlcN concentration in plasma was measured by the above method.
AUC was calculated from GlcN concentration in plasma at the time of intravenous injection after 3-hour fasting with drug0-4hr. AUC of intravenous drug (GlcN-HCl 50mg drug)0-4hrThe average value ± SD of (intravenous AUC) was 29.2 ± 1.6(μ g/mL · hr) (n ═ 3).
From the results of test example 4, the rats fed in an environment fasted for 3 hours before free feeding or self-administration were treated at 22: AUC when 00 GlcN-HCl (500mg/kg) was orally administered0-4hr(AUC for oral administration), AUC for oral drug group of free ingestion group0-4hrThe mean value ± SD of (g) ± 3.6(μ g/mL · hr), and the fasting group 3 hours before administration of the drug was 27.8 ± 8.0(μ g/mL · hr) (both n ═ 6).
From the oral AUC and the intravenous AUC of the fasted group, 22 was calculated according to the following formula: bioavailability (BA) of GlcN in the free-feeding group at 00 hours of administration and the fasting group 3 hours before administration.
Bioavailability (%) 100 x AUC orally administered/AUC intravenously administered in fasted group
In the free feeding group, the bioavailability of GlcN was 6%. The bioavailability of the fasted group 3 hours before administration was 10% which was 1.70 times that of the free-fed group.
From the above test examples, it was clarified that the bioavailability of glucosamine can be improved by orally taking glucosamine after a predetermined time from the time point when the expression level of mRNA of glucose transporter in the small intestine shows a high value among 1 day in the fasting state after a predetermined time.
In the above experiment, the day in the light phase belongs to the sleeping time and the night in the dark phase belongs to the feeding time for rats. In the above experiment, the mRNA expression of the glucose transporter gene in the small intestine showed the highest value at 16 points (16: 00) before the start of the dark phase. In the case of humans, the day and night of sleep time and food intake time are generally the fasting time from the time of dinner to the time of getting up, in contrast to rats. Therefore, it is considered that when the results of the rats are applied to humans, in the normal life of humans, the absorption rate of glucosamine can be improved by orally taking glucosamine from the time of getting up to breakfast.
Sequence listing
<110> Sandeli's stock control corporation
<120> method for improving bioavailability of glucosamine
<130>SP2018-0032
<150>JP 2017-165804
<151>2017-08-30
<160>8
<170>PatentIn version 3.5
<210>1
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aaagctgtgg cgtgatgg 18
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tgtgggctaa tttcaggact gg 22
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Claims (15)
1. A method for improving the bioavailability of glucosamine, comprising orally administering glucosamine within 6 hours after fasting for 3 hours or more and after the time at which the mRNA expression level of a Period gene reaches the maximum in 1 day.
2. The method of claim 1, wherein the increase in the bioavailability of glucosamine is based on an increase in the rate of absorption of glucosamine.
3. The method of claim 1 or 2, wherein the fasting time is 3 to 24 hours.
4. The method according to any one of claims 1 to 3, wherein the subject is allowed to take glucosamine orally from the time of getting up to the 1 st meal.
5. The method according to any one of claims 1 to 4, wherein the oral ingestion of the glucosamine is followed by a fasting period of more than 15 minutes.
6. A method for improving the bioavailability of glucosamine, comprising orally ingesting glucosamine in a subject between 2 hours and 8 hours after fasting for 3 hours or more and after a time when the mRNA expression level of a glucose transporter gene in the small intestine reaches the maximum level among 1 day.
7. The method of claim 6, wherein the increase in the bioavailability of glucosamine is based on an increase in the rate of absorption of glucosamine.
8. The method according to claim 6 or 7, wherein the glucose transporter is 1 or more selected from the group consisting of glucose transporter 2(GLUT2), glucose transporter 5(GLUT5), and sodium-glucose cotransporter 1(SGLT 1).
9. The method of any one of claims 6 to 8, wherein the fasting time is 3 to 24 hours.
10. The method of any one of claims 6 to 9, wherein the subject is allowed to take glucosamine orally from the time of getting up to the 1 st meal.
11. The method according to any one of claims 6 to 10, wherein the oral ingestion of the glucosamine is followed by a fast of more than 15 minutes.
12. A method for indicating the time of ingestion of glucosamine for improving bioavailability, which is characterized by using the mRNA expression level of a Period gene as an indicator.
13. A method for indicating a time of ingestion of glucosamine for improving bioavailability, characterized by using an mRNA expression level of a glucose transporter gene in the small intestine as an index.
14. A preventive or ameliorating agent for arthralgia which comprises glucosamine and which is for oral ingestion by a subject in the case of (i) or (ii) below,
(i) after fasting for more than 3 hours, the time when the mRNA expression quantity of the Period gene reaches the maximum in 1 day is less than 6 hours later;
(ii) after fasting for more than 3 hours, and between 2 hours and 8 hours after the time when the mRNA expression level of the glucose transporter in the small intestine reaches the maximum of 1 day.
15. The agent for preventing or ameliorating arthralgia according to claim 14, wherein the indication of "ingestion from the time of getting up to the 1 st meal" is provided.
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| PCT/JP2018/030422 WO2019044524A1 (en) | 2017-08-30 | 2018-08-16 | Method for improving bioavailability of glucosamines |
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Non-Patent Citations (4)
| Title |
|---|
| BETH A BIGGEE ET AL.: "Changes in serum levels of glucosamine and sulphate after", 《ANN RHEUM DIS.》 * |
| MARC ULDRY ET AL.，: "GLUT2 is a high affinity glucosamine transporter", 《FEBS LETT》 * |
| S. PERSIANI ET AL.: "Glucosamine oral bioavailability and plasma pharmacokinetics after increasing doses of crystalline glucosamine sulfate in man", 《OSTEOARTHRITIS AND CARTILAGE》 * |
| YUBING ZHU ET AL.，: "Bioequivalence of Two Formulations of Glucosamine Sulfate SOO-mg Capsules in Healthy Male Chinese Volunteers: An Open-Label, Randomized-Sequence, Single-Dose, Fasting, Two-Way Crossover Study", 《CLINICAL THERAPEUTICS》 * |
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