CN1822827A

CN1822827A - Use of alpha-ketoglutaric acid for the treatment of malnutrition or high plasma glucose condition

Info

Publication number: CN1822827A
Application number: CNA2004800203227A
Authority: CN
Inventors: S·G·皮尔茨诺夫斯基; D·布林
Original assignee: ESSENTYS AB
Current assignee: ESSENTYS AB
Priority date: 2003-07-01
Filing date: 2004-07-01
Publication date: 2006-08-23
Anticipated expiration: 2024-07-01
Also published as: JP2007529408A; JP4847323B2; CN1822827B; SE0301947D0; CN101301285A

Abstract

A method for improving adsorption of amino acids in a vertebrate, including mammal and bird, is included. The method comprises administering to a vertebrate, including mammal and bird, in a sufficient amount and/or at a sufficient rate to enable desire effect. AKG, AKG derivatives or metabolites, AKG analogues or mixture thereof. Also conteplated is a method for decreasing adsorption of glucose in a vertebrate, including mammal and bird, in the need thereof, AKG, AKG derivates or metabolites, AKG analogues or mixture thereof, for decreasing glucose adsorption as well as compositions for use in treatment.

Description

α-Tong Wuersuan is used for the treatment of the purposes of malnutrition or high plasma glucose symptom

Technical field

The present invention relates to be used for improving the method for amino acid whose absorption the vertebrates that comprises mammal and birds, and the method that is used to reduce glucose absorption.The invention still further relates to the production that is used for improving the compositions that described vertebrates aminoacid absorbs.

Background of invention

Diabetes are serious metabolic diseases, and it is to determine by the appearance of the plasma glucose content of chronic rising.The classical symptom of adult's diabetes is polyurias, excessive thirst, beta-oxybutyria, the plasma glucose of weight loss and higher level fast.

Normal fasting plasma glucose concentration is for being lower than every liter of 115mg/dl.In diabetics, find that fasting concentration is above 140 milligrams every liter.Generally, diabetes are that β cells injury owing to pancreas occurs.This damage may be owing to primary diabetes mellitus occurs, wherein, the β cell is by the autoimmune system destruction or as the secondary diabetes reaction at other primary diseases, as pancreatic diseases, not owing to lack the hormone abnormality of insulin action, medicine or chemical induction, Insulin receptor INSR is unusual, hereditary symptom or other.

Primary diabetes mellitus can be classified as type i diabetes (be known as not only insulin-dependent diabetes or IDDM) and type ii diabetes (but also being known as noninsulindependent diabetes or NIDDM).

The I type, teenager outbreak or insulin-dependent diabetes are well-known hormone defect states.Wherein, as if pancreatic beta cell is destroyed by the immune defence mechanism of health self.It is rare or do not have an endogenous insulin secretion ability to suffer from the patient of type i diabetes.Extreme hyperglycemia can appear in these patients.Type i diabetes is fatal, introduced insulin substitution and treat-use the insulin from animal origin first before about 70 years, recently, uses the insulin human by the recombinant DNA technology preparation.Clear now, the destruction of β cell has caused two kinds of hormones, the defective of the combination of insulin and dextrin in the type i diabetes.When pancreatic cell is destroyed, the Disability of excreting insulin and dextrin.

The character of pancreatic beta cell damage it be unclear that in type ii diabetes.Different with type i diabetes patient's pancreatic beta cell, type ii diabetes patient's β cell has kept the ability of synthetic and excreting insulin and dextrin.Type ii diabetes is a feature with insulin and resistance, that is, normal metabolic response can not be made to the effect of insulin by peripheral tissues.In other words, insulin resistance is a kind of like this symptom, and wherein, the circulation insulin has produced subnormal biologically.Clinically, but show at insulin and show when continuing normal or higher plasma glucose content and have insulin resistance in the face of normal or higher level.The hyperglycemia relevant with type ii diabetes be sometimes can by diet recover or alleviate or lose weight and be enough to recover the sensitivity of peripheral tissues insulin.In fact, type ii diabetes is usually being feature in the hyperglycemia that occurs being higher than under the plasma insulin condition of normal level.The development of type ii diabetes is relevant with the high concentration plasma glucose, and with the relative reduction of the insulin secretion speed of glucose induction.Therefore, for example,, may there be insulin deficit in the late period of type ii diabetes.

Known treating diabetes and prevention

The main purpose of treatment all types diabetes is identical, being about to plasma glucose concentration is reduced to as much as possible near normal level, thereby reduce the short-term and the long-term complications (Tchobroutsky, Diabetologia 15:143-152 (1978)) of this disease.

Relation between the order of severity of the hyperglycemia of diabetics and the caused long-term complications is verified further by the DCCT (DCCT) that National Institutes of Health (The Diabetes Control andComplications Trial Research Group, N.Eng.J.Med.329:977 (1993)) finishes recently.DCCT carried out in the U.S. and Canadian 29 clinical center with 10 years, had confirmed to reduce type i diabetes patient's average blood plasma concentration of glucose, had reduced final organ complication.Retinopathy reduced by 76%, the development of retinopathy has reduced by 54%, and the index of nephropathy (albuminuria, albuminuria) is alleviated to some extent.Significantly the generation of neuropathy reduces equally.

The treatment of type i diabetes must relate to uses the insulin that recovers dosage, uses by parenteral route.Combine with correct diet and the monitoring of self-plasma glucose, most of type i diabetes patient can obtain the control to a certain degree to plasma glucose.

Opposite with type i diabetes, the treatment of type ii diabetes does not need to use insulin usually.The therapeutic scheme of type ii diabetes is usually directed to Diet Therapy test and life style adjustment, at first carries out the 6-12 time-of-week usually.

It is suitable that the feature of diabetics diet comprises, but be not excessive total calorie picked-up, clocklike has meal, and limits the content of saturated fat, increases the content of polyunsaturated fatty acid simultaneously, and the picked-up that increases dietary fiber.

Life style changes the motion that comprises that maintenance is frequent, as the supplementary means of start-up control and minimizing insulin resistance degree.

If after suitable diet test and life style change, still there is the fasting hyperglycemia, just can make the diagnosis of " constitutional dietary deficiency ", and the Steering Body that needs oral blood sugar lowering therapeutic test or directly insulinize need produce plasma glucose control, and therefore reduces the complication of this disease.The type ii diabetes patient that can not react to diet and losing weight may be to using oral hypoglycemic, reacts as the treatment that sulfonylureas or biguanide are carried out.But, insulinize is used to treat other patients that suffer from type ii diabetes, constitutional dietary deficiency and NO patient have particularly occurred or the constitutional dietary deficiency occurred and the patient of Secondary cases blood sugar lowering failure.

At U.S. Patent number 5,124, disclosed in 314 and 5,175,145 the dextrin stimulant has been used for the treatment of diabetes.The new Therapeutic Method that is used as of excessive dextrin that Simulation with I type i diabetes and dextrin are suppressed has been proposed.

For example, known treatment has the diabetes pill, and for example, based on the pill of sulfonylureas, it helps pancreas to produce more insulin, and helps health to use insulin better.Possible side effect: hypoglycemia, stomach-ache, erythra or pruritus and weight increase.

Can limit the glucose generation of liver based on other pills of biguanide, and can reduce the amount of insulin in the health equally, improve blood fat and cholesterol state.Possible side effect has the vomiting with the ethanol combination, has increasing the weight of of kidney and disease now, weakness, and dizziness, dyspnea is felt sick, and diarrhoea.

The enzyme that can suppress digestible starch based on other pills of alpha-glucosidase inhibitor.Possible side effect has gastrointestinal problems.

Other pills based on thiazolidinedione help cell more responsive for insulin.Possible side effect is, they can not be used in combination (making regular check on) with hepatic disease, and hypoglycemia and can only making up with other treatment can not pass through pill birth control, weight increase, anemia danger, swelling (edema) effectively.

Help pancreas with producing more insulin after the meal based on other pills of meglitinide.Possible side effect is a hypoglycemia, and weight increase.

In addition, exist the mouth of combination to use medicine, for example, based on glyburide (sulfonylureas) and metformin (biguanide), for example, " Glucovance ".Possible side effect is a hypoglycemia, can not use simultaneously with nephropathy, and can not be used in combination with ethanol.

US 5,234, and 906 have disclosed the compositions that comprises glucagon and dextrin stimulant, and they are used to control or treat the purposes of hyperglycemia symptom.

WO 93/10146 has disclosed the dextrin stimulant, and uses it for treatment or prevention hypoglycemia symptom, comprises the symptom that needs insulin, as the purposes of diabetes.

Renal failure and malfunction

Renal failure or kidney malfunction are the states that kidney can not be removed intravital refuse.Renal failure has caused the accumulation of deleterious refuse in blood.Kidney has extra removing ability usually, and before symptom occurring, the ability of kidney have 50% normal.Symptom comprises pruritus, and fatigue is felt sick, vomiting, and inappetence causes malnutrition.Renal failure is relevant with diabetes and hypertension usually.In suffering from the object of renal failure, above-mentioned symptom appears, that is and, vomit and cause underfed inappetence.

Dialysis process can alleviate from the pressure of refuse to kidney.In addition, it is the method for spended time.The patient may need to carry out weekly the several treatment.Carry out the needs of patients medical attention of dialysis treatment, and this method is cost and time loss type.

Glucose oxidation

From the original position rat studies that Windmueller and Spaeth (1) carry out, known glutamic acid and glutamine are the important metabolism fuel of small intestinal.Windmueller and Spaeth have reported during absorbing first by the metabolism of intestinal to the vast scale of glutamic acid (95%) and glutamine (70%).Their result has obtained checking in vivo on piglets (2) and people (3).

During glucose oxidation, first step is the transamination of being undertaken by any amount of enzyme, and by the deaminizating that glutamte dehydrogenase (GDH) carries out, a lot of enzymes are wherein expressed (4,5) in gastrointestinal tract.The deaminizating that is undertaken by GDH has produced AKG and free ammonia.During the transamination of being undertaken by branched-amino transferring enzyme (BCAT), glutamic acid partly offers branched-chain alpha-ketoacid with amino, has formed AKG and corresponding branched-chain amino acid.

α-Tong Wuersuan

Glutamine and derivant thereof, for example, α-Tong Wuersuan (AKG) is the molecule that has pivotal role in systematicness of being undertaken by tricarboxylic acid cycle and intestinal metabolism.But, its mechanism of action is still not exclusively understood (Pierzynowski, S.G., and Sjodin, A. (1998) J.Anim.a.Feed Sci.7:79-91; With Pierzynowski S.G., wait Eds:KBK Knutsen and J-E Lindberg., Uppsala 19-21 June, 2001).

Theoretically, AKG (2-oxygen-1,3-propanedicarboxylic acid, 2-oxoglutaric acid, α-Tong Wuersuan, α-oxygen-1,3-propanedicarboxylic acid, 2-oxoglutaric acid, 2-oxygen-1,5-1,3-propanedicarboxylic acid, 2-oxygen-1,3-propanedicarboxylic acid, 2-oxoglutaric acid) may be glutamine, glutamate, Glu, the product of glutamic acid internal metabolism degraded.In addition, it can also not only play a part glutamine and arginic precursor, but also can be used as some other amino acid whose precursor, therefore, is considered to the metabolic protective agent of albuminolysis.Olin etc., 1992 when having confirmed in fish meal to add AKG, the carbamide of discharge has reduced.Similarly; in human body; in total parenteral absorption (TPN) solution, add AKG and amino-acid mixed fashionable with other; after operation, observed good protective action (Pierzynowski to nitrogen loss; S.G.; and Sjdin, A. (1998) J.Anim.a.Feed Sci.7:79-91).For the mankind, AKG may degrade with muscle protein and combine, to satisfy the demand of intestinal during coercing after the so-called operation, for example, catabolism, hunger etc.

Function of intestinal canal has obtained the confirmation (1996 of Reeds etc. recently to the demand of the relevant metabolite of glutamine family, Am.J.of Physiol.-Endocrinology and Metabolism270:413-418), this author has reported in operation 100% glutamic acid/glutamine utilization rate almost during by newborn piglet small intestinal.

AKG may be important energy donor, by a few path for transformation, for example forms GABA or succinate by ornithine and putrescine.Theoretically, AKG can also play a part the ammonium ion scavenger, may be by transforming to glutamic acid/glutamine.

Have realized that-but still unexposed mistake-enterocyte depends on their growths from ammonium.

Therefore, consider the problems referred to above, the needs exploitation is used for the treatment of and prevents mammal especially, for example, and cat, Canis familiaris L. or human hypoglycemia symptom, as diabetes, and the malnutrition relevant with diabetes, for example apparatus and method of renal failure, wherein, can avoid with the problem or the side effect of existing apparatus and method.In addition, except the nutritional status of nephrotic and diabetes, also need to improve health status.At this on the one hand, the present invention has satisfied above-mentioned needs and advantage.

Summary of the invention

Consider above-mentioned in prevention, known problem and be used for the treatment of and be used to proofread and correct the malnutrition relevant with renal failure in treatment and/or diabetes-alleviating and other hypoglycemia relevant disease fields with for example diabetes, the invention provides and be used for prevention, treatment and/or diabetes-alleviating and underfed new and method and composition improvement.

An object of the present invention is to provide the method that is used to improve the vertebrate aminoacid absorption that comprises mammal and birds.This method comprises to the vertebrates that comprises mammal and birds, uses AKG with enough quantity and/or enough speed, AKG derivant or metabolite, and AKG analog or its mixture are so that can absorb the effect of wishing that produces to aminoacid.

A kind of embodiment of described method is such, wherein, described AKG, AKG derivant or metabolite, AKG analog or its mixture are selected from following one group: one of the salt of other of α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, chitosan-AKG, AKG and aminoacid and amino acid derivativges, AKG-and two-slaine, as CaAKG, Ca (AKG) 2 and NaAKG.

Another embodiment is such, and wherein, described vertebrates is a rodent, as mice, and rat, Cavia porcellus or rabbit; Birds, as turkey, hen, chicken or other chickling; Farm animal, as cow, horse, pig, the farm animal that piglets or other are freely walked about; Or house pet, as Canis familiaris L. or cat.

Another embodiment is such, and wherein, described vertebrates is human.

Another embodiment is such, and wherein, described aminoacid is any essential amino acids.

A kind of embodiment in sea is such, and wherein, described essential amino acids is an isoleucine, leucine, lysine, and proline.

The present invention also comprises the method that is used to reduce the vertebrate glucose absorption that comprises mammal and birds.This method comprises to the vertebrates that comprises mammal and birds uses AKG with enough quantity and/or enough speed, AKG derivant or metabolite, and AKG analog or its mixture are so that can produce the influence that needs to glucose absorption.

The present invention also comprises and is used for preventing, suppresses or alleviate the method for the vertebrate high glucose symptom that comprises mammal and birds.This method comprises to the vertebrates that comprises mammal and birds, uses AKG with enough quantity and/or enough speed, AKG derivant or metabolite, and AKG analog or its mixture are so that can produce the influence that needs to described symptom.

A kind of embodiment is such, and wherein, described high glucose symptom is I type or type ii diabetes.

The present invention also comprises AKG, AKG derivant or metabolite, and AKG analog or its mixture production are used to prevent, alleviate or treat the purposes of the compositions of high glucose symptom.

The invention still further relates to AKG, AKG derivant or metabolite, AKG analog or its mixture production are used to prevent, alleviate or treat the purposes of underfed compositions.

A kind of embodiment of described purposes is such, and wherein, described compositions is a Pharmaceutical composition, and optionally comprise can be medicinal carrier and/or additive.

The another embodiment of described purposes is such, and wherein, described compositions is food or feed additive.

Another embodiment is such, and wherein, described food or feed additive are the compositions of food additive and/or solid food and/or drink form.

Another embodiment is such, and wherein, at AKG described in the compositions of producing, AKG derivant or metabolite, AKG analog or its mixture are to treat effective dose.

Another embodiment is such, and wherein, described treatment effective dose is 0.01-0.2g/kg body weight dosage every day.

Brief description of drawings

Fig. 1 is illustrated in contrast and the intravital whole leucine kinetics of AKG infusion pig.Value is mean value SEM; N=9, each pig is accepted contrast and AKG.When carrying out error analysis (ANOVA), value and the contrast of AKG do not have difference.AKG represents α-Tong Wuersuan; On behalf of non-oxidizable leucine, NOLD handle; On behalf of leucine, Ra speed occurs; Balance, promptly Ra deducts from NOLD, expression albuminous body leucine balance.

The detailed explanation of invention

Definition

In the context of the application and invention, used to give a definition:

Herein, term " medicinal composition " expression the present invention's the effective composition for the treatment of.

Herein, term " treatment is effectively measured " or " effectively amount " or " treatment effectively ", expression can be to the consumption of specific symptoms and application program generation result for the treatment of. It is the scheduled volume that calculates the active material that can produce the desirable treatment effect relevant with diluent with the additive of needs; That is, carrier or application media. In addition, it also can be used for expression is enough to alleviate, and most preferably prevents host's activity, the consumption of the remarkable defective on the clinical medicine of function and reaction. In addition, treatment effectively amount be the improvement that is enough to cause host's clinical remarkable symptom. As is understood by persons skilled in the art, the consumption of compound can change according to its ratio is active. Suitable dosage can comprise the active compound of scheduled volume, and it is calculated and can produces the desirable result for the treatment of relevant with the diluent of needs; That is, carrier or additive. In method and purposes for the production of the present invention's composition, provide the effectively active component of amount for the treatment of. Treating effective amount can be determined according to patient's feature by common doctor or animal doctor worker, such as the age, and body weight, sex, symptom, complication, other diseases etc. are as known in the field.

Herein, term " derviate " or " derivative " expression is directly or by modifying or part replaces chemical substance from parent.

Herein, similar each other on term " similar body " or " similar thing " the expression structure, but the compound of isomers not necessarily. Similar thing has similar function, but different on structure or the origin of evolving.

Herein, the processing of " treatment " expression take healing as purpose, it may be symptom fully/comprehensively or part cure.

Herein, the intensity of mitigation symptoms is not only in term " alleviation " expression, but also has postponed the outbreak of symptom.

Herein, term " prevention " expression guarantees that some event can not occur, and for example, the symptom relevant with immature GIT can not occur. By preventing some symptom, can postpone the generation of described symptom.

Herein, term " increase amino acid absorb " expression and the treatment of not accepting the present invention or the vertebrate of using are compared, change has occured in the amino acid whose clean absorption of vertebrate, if compare with the vertebrate of the identical type of not accepting described treatment, larger in the clean amount that absorbs described in the described vertebrate, just think that described change is to strengthen.

Herein, the amino acid of vertebrate and the reading of glucose absorption are monitored or are measured in term " dynamics " expression continuously or often, in order to determine its absorption speed.

Herein, term " sodium-AKG " can with term " AKG-Na ", " Na-AKG ", " sodium salt of AKG ", " AKG (sodium salt) " exchange is used.

Herein, term " chitosan-AKG " can with term " AKG-chitosan ", " AKG (chitosan salts) " exchange is used.

The diagnosis of I type and type ii diabetes

The diagnosis of suffering from the patient of I type and type ii diabetes belongs to those skilled in the art's ability and the scope of knowledge. For example, the age surpasses 35 years old, has polydipsia, polyuria, and voracious disease (being with or without body weight alleviates) and relevant with higher plasma glucose concentration, and do not have the individuality of the medical history of ketone acid usually to be diagnosed as type ii diabetes. Fat appearance, the Positive family history of type ii diabetes, and normal or higher fasting plasma insulin and c-peptide concentration are other features of suffering from the Most patients of type ii diabetes. The consumption of the one or more dosage of " treatment is amount effectively " expression, it preferably can reduce the plasma glucose concentration of the object of suffering from type ii diabetes.

The inventor finds startledly that absorption has impact on AKG in infusion site. After duodenum is inculcated AKG, observed in surprise the glucose absorption that the amino acid that strengthened absorbs and weakened.

Therefore, the present invention can be used for reducing the blood plasma glucose of non--insulin-absorption type ii diabetes object.

Malnutritive diagnosis

Suffer from underfed patient, promptly can not suitably take in nutrition or underfed diagnosis, belong to those skilled in the art's the ability and the scope of knowledge.Usually, the general health state of individuality is assessed, so that determine malnutrition.

The diagnosis of renal failure

The diagnosis of suffering from the patient of renal failure belongs to those skilled in the art's the ability and the scope of knowledge.

The renal failure that two kinds of forms are arranged, acute and chronic renal failure (ACF and CRF).Acute renal failure normally can be recovered, and chronic renal failure normally develops.CRF can be distinguished into pre-dialysis and uremic active treatment, for example, uses dialysis or transplanting.Do not have the definite restriction of the starting point of pre-dialysis, but, pre-usually dialysis is confirmed as diagnosing out the time between renal failure and the beginning active treatment.

Dialysis and transplanting are considered to active treatment.

Improve the method that aminoacid absorbs

According to the present invention, disclosed the method that is used to improve the vertebrate aminoacid absorption that comprises mammal and birds.This method comprises to the vertebrates that comprises mammal and birds, uses AKG with enough quantity and/or enough speed, AKG derivant or metabolite, and AKG analog or its mixture produce the influence that needs so that can absorb aminoacid.

Described aminoacid is absorbed in and does not accept described AKG, AKG derivant or metabolite, and the vertebrate aminoacid that comprises mammal and birds of AKG analog or its mixture absorbs and is considered to improve when comparing.

The another embodiment of described method is such, wherein, described AKG, AKG derivant or metabolite, AKG analog or its mixture are selected from following one group: α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, chitosan-AKG, the salt of other of AKG and aminoacid and amino acid derivativges; One of AKG-and two-slaine, as CaAKG, CaAKG ₂, and NaAKG.

Other embodiments are such, and wherein, described vertebrates is a rodent, as mice, rat, Cavia porcellus or rabbit; Birds is as turkey, hen, chicken or other chickling; Farm animal is as cow, horse, pig, piglets or the farm animal of freely walking about; Or house pet, as Canis familiaris L. or cat.

Another embodiment is such, and wherein, described vertebrates is human.The described mankind need the underfed patient of treatment, described malnutrition be because, for example, renal failure, diabetes, the athlete, age (child and old people), pregnancy, anorexia nervosa, bulimia, binge eating, the eating disorder that mandatory binge eating or other are not pointed out (EDNOS).

In another embodiment, described vertebrates as the people, can be any vertebrates that needs to increase amino acid whose utilizability and availability, for example, and essential amino acids or condition aminoacid, particularly isoleucine, leucine, lysine and proline.

The example of essential amino acids is an a-amino acid, as being isoleucine (Ileu) in human body, leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), threonine (Thr), tryptophan (Try) and valine (Val).Essential amino acids between the species is different.Rat needs two kinds of other aminoacid, i.e. arginine (Arg) and histidine (His).

Other embodiments are such, and wherein, described aminoacid is any aminoacid, as alanine, valine, leucine, isoleucine, proline, phenylalanine, tryptophan, methionine, threonine, cystine, tyrosine, glutamine, histidine, lysine, arginine, aspartic acid, agedoite, glutamic acid, glutamine, glycine and serine.

Other embodiments are such, and wherein, described aminoacid is any essential or condition essential amino acids.Example essential or the condition essential amino acids is as shown in table 2.

Another embodiment is such, wherein, described must or the condition essential amino acids be selected from following one group: isoleucine, leucine, lysine and proline.

Be used to reduce the method for glucose absorption and be used for prevention, suppress or alleviate the method for plasma glucose increase

Plasma glucose content is the content of glucose in the blood (sugar).It is also called serum glucose level.The content of glucose is expressed as every liter of mM (mmol/l) or mg/dL in the blood.

Usually, in human body, plasma glucose content remained in one day in the narrow bound scope, approximately 4-8mmol/l.Glucose content after having meal is higher, and minimum in the morning.Be typically about 70-110mg/dL (3.9-6.1mmol/L) during fasting, and after having meal 2 hours, be typically about 80-140mg/dL (4.4-7.8mmol/L).(＞10.0mmol/l) plasma glucose content is considered to the high plasma glucose value to 2 hours＞180mg/dL after having meal usually.When fasting, when plasma glucose value＞140mg/dL, also be this situation.

If a people for example suffers from, diabetes, their plasma glucose content sometimes may exceed the above-mentioned limit.It is lower that all diabetes patients' basic defect is that insulin-induced soma is removed the ability of glucose (sugar) molecule from blood.No matter this lower insulin active be because the minimizing (for example, type i diabetes) of the amount of the insulin that is produced or since cell to the insulin insensitivity of normal contents, consequently identical, that is, the plasma glucose too high levels.It is known as " hyperglycemia " or " hyperglycemia " or " hyperglycemia ", and it represents " the high glucose in the blood ".Usually, when plasma glucose＞240mg/dL (＞just be considered to hyperglycemia 13.4mmol/L) time.

According to the present invention, disclosed and be used to weaken the method that the vertebrates that comprises mammal and birds absorbs at interior plasma glucose.This method comprises to the vertebrates that comprises mammal and birds, uses AKG with enough quantity and/or enough speed, AKG derivant or metabolite, and AKG analog or its mixture are so that can produce the influence that needs to the absorption of glucose.

Using AKG, AKG derivant or metabolite, after AKG analog or its mixture, the minimizing of glucose absorption can be for the 5-50% of initial plasma glucose value, as 5,10,15,20,25,30,35,40,45 or 50%.

In another embodiment, the 20-40% that is reduced to initial plasma glucose value of described absorption.

In another embodiment, described 30% of the initial plasma glucose value that is reduced to.

In addition, disclosed and be used for prevention, suppressed or alleviated the method for the vertebrate high plasma glucose symptom that comprises mammal and birds.Described method comprises to the vertebrates that comprises mammal and birds, use AKG with enough quantity and/or enough speed, AKG derivant or metabolite, AKG analog or its mixture are so that can produce ideal effect to described high glucose symptom.

In another embodiment, described high glucose symptom is the hyperglycemia symptom.

The described method relevant with high glucose or hypoglycemia comprises other embodiments, wherein, described AKG, AKG derivant or metabolite, AKG analog or its mixture, be selected from following one group: α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, the salt of other of chitosan-AKG, AKG and aminoacid and amino acid derivativges; One of AKG-and two-slaine, as CaAKG, CaAKG ₂And NaAKG.

Other embodiments are such, and wherein, described vertebrates is human.

Other embodiments are such, wherein, described high glucose symptom be because, for example, acromegaly, the glad syndrome of Ke Xing Shi, hyperthyroidism, cancer of pancreas, pancreatitis, pheochromocytoma, insulin content is not enough or too much food intake is too much.

Other embodiments are such, and wherein, described high glucose level causes owing to I type or type ii diabetes.

AKG is used for the treatment of diabetes and is used for the treatment of malnutrition

According to the present invention, disclosed AKG, AKG derivant or metabolite, AKG analog or its mixture production are used to prevent, alleviate or treat the purposes of the compositions of high glucose symptom.

High glucose symptom and hyperglycemia symptom are described in front the paragraph.

Other embodiments comprise that wherein, described hyperglycemia symptom is I type or II diabetes.

According to the present invention, disclose AKG, AKG derivant or metabolite, AKG analog or its mixture production are used to prevent, alleviate or treat the purposes of underfed compositions.

In other embodiments of described purposes, described compositions is a Pharmaceutical composition, it optionally comprise can be medicinal carrier and/or additive.

In other embodiments, described compositions is food or feed additive.

In other embodiments, described food or feed additive are the compositions of food additive and/or solid food and/or drink form.

In other embodiments, the described AKG in the compositions of producing, AKG derivant or metabolite, AKG analog or its mixture are to treat effective dose.

In other embodiments, described treatment effective dose is 0.01-0.2g/kg body weight dosage every day.

Use AKG, AKG derivant or metabolite, AKG analog or its mixture

According to above disclosed method, give the vertebrates that comprises mammal and birds; Rodent is as mice, rat, Cavia porcellus or rabbit; Birds is as turkey, hen, chicken or other chickling; Farm animal is as cow, horse, pig, piglets or the farm animal of freely walking about; Or house pet, use AKG as Canis familiaris L. or cat, AKG derivant or metabolite, AKG analog or its mixture.

Use and can be undertaken, depend on the vertebrate species that will treat, need the vertebrate symptom of described method by diverse ways, and the specific targets that will treat.

In one embodiment, described using with food or feed additive form carried out, as the composition of food additive and/or solid food and/or drink form.Other embodiments can be suspension or solution form, as the beverage of following further disclosure.

In addition, dosage form can comprise capsule or tablet, as masticable or solubility, and for example effervescent tablet, and powder and other dried forms known in those skilled in the art, as bead, as microsphere, granule and powder.

As above disclosed, described using can be parenteral, rectum or oral cavity food or feed additive form.The parenteral medium comprises sodium chloride solution, Ringer ' s glucose, glucose and sodium chloride, lactic acid Ringer ' s or fixed oil.

Described food and feed additive can also be emulsive.Then can be with active treatment composition and mixed with excipients, described excipient can be medicinal, and compatible with described active component.Suitable excipient has, for example, and water, saline, glucose, glycerol or ethanol etc., and their combination.In addition, if necessary, described compositions can contain the auxiliary substance of trace, as wetting agent or emulsifying agent, and the pH buffer agent, they can strengthen the effect of described active component.

Can replenish the different dosage form of parenteral food or feed additive, as solid food, liquid or cryodesiccated or other exsiccant preparations.It can comprise diluent (for example, Tris-HCl, the acetic acid with various cushions, phosphoric acid), pH and ionic strength, additive, as albumin or gel, so that prevent absorption to the surface, detergent (for example, Tween 20, and Tween 80, Pluronic F68, cholate), solubilizing agent, (for example, glycerol, Polyethylene Glycol), antioxidant is (for example, ascorbic acid, sodium pyrosulfite), antiseptic is (for example, thimerosal, benzylalcohol, parabens), filler or osmotic pressure regulator are (for example, lactose, mannitol), such as the polymer of Polyethylene Glycol and the covalent bond of described compositions.Arrive as polylactic acid with the complexation or the described material sorting of metal ion, polyglycolic acid in the granular formulations of the polymer of hydrogel etc. or on it, or is incorporated into liposome, micro emulsion, and micelle, the single or multiple lift vesicle is on erythrocyte umbra or the spheroplast.

Beverage

In one embodiment, described food or feed additive are with beverage or its dry compositions form, use by any method of the present invention.

Described beverage comprises the AKG of effective dose, AKG derivant or metabolite, and AKG analog or its mixture, and acceptable water-solubility carrier in the nutrition, as mineral, vitamin, carbohydrate, fat and albumen.AKG, AKG derivant or metabolite, the example of AKG analog or its mixture have α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, the salt of other of chitosan-AKG, AKG and aminoacid and amino acid derivativges; One of AKG-and two-slaine, as CaAKG, CaAKG ₂And NaAKG.

If described beverage is to provide with dried forms, above-mentioned all the components all provides with dried forms.The beverage that provides with the form that can directly consume also comprises water.Final drink soln can also have in check osmotic pressure and acidity, as the buffer solution of conduct according to the explanation of the generality in above paragraph preparation.

Preferably in the scope of about 2-5, particularly about 2-4 is so that suppress antibacterial and conk for pH.Can also use sterilized beverage, its pH is about 6-8.

Described beverage can use separately or use with one or more treatment effective composition.

AKG, AKG derivant or metabolite, the purposes of AKG analog or its mixture

According to the present invention, disclosed and used AKG, AKG derivant or metabolite, AKG analog or its mixture production are used for prevention, alleviate or treatment hyperglycemia symptom, as I type and type ii diabetes, and the purposes that is used for the treatment of underfed compositions.

Another embodiment of the present invention comprises purposes, and wherein, described compositions is a Pharmaceutical composition.This Pharmaceutical composition can with carrier and/or additive that can be medicinal, as diluent, antiseptic, solubilizing agent, emulsifying agent, adjuvant and/or carrier one are used from the present invention disclosed method and purposes.

In addition, in this article, " carrier that can be medicinal " is well known to those skilled in the art, and includes, but are not limited to 0.01-0.05M phosphate buffer or 0.8% saline.In addition, described carrier that can be medicinal can be water or non-aqueous solution, suspension and emulsion.The example of nonaqueous solvent has propylene glycol, Polyethylene Glycol, and vegetable oil is as olive oil and the organic ester that can inject, as ethyl oleate.Aqueous carrier comprises water, alcohols/aqueous solution, and emulsion or suspension comprise saline and buffering medium.The parenteral medium comprises sodium chloride solution, Ringer ' s glucose, glucose and sodium chloride, newborn acidifying Ringer ' s or fixed oil.Can also use antiseptic and other additives, for example, anti-microbial agents, antioxidant, chelating agen and noble gas etc.

Another embodiment of the present invention comprises purposes, and wherein, described compositions is the composition of food additive and/or solid food and/or drink form,

The compositions of this production, as Pharmaceutical composition or food or feed additive, comprise compositions of the present invention, and comprise that optionally carrier and/or a certain amount of second kind maybe can influence any hyperglycemia symptom, as I type and type ii diabetes, and underfed other active component.

The dosage of the Pharmaceutical composition of using

According to the present invention, AKG, AKG derivant or metabolite, AKG analog or its mixture are used to produce the purposes of compositions of the present invention, comprise to needs it such as birds or mammiferous vertebrates administering therapeutic effective dose.Described treatment effective dose is about 0.01-0.2g/kg body weight dosage every day.

AKG, AKG derivant or metabolite, AKG analog or its mixture

According to the present invention, comprise AKG, AKG derivant or metabolite, AKG analog or its mixture.AKG, AKG derivant or metabolite, the example of AKG analog or its mixture have α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, chitosan-AKG, the salt of other of AKG and aminoacid and amino acid derivativges; One of AKG-and two-slaine, as CaAKG, CaAKG ₂, NaAKG.

Application target

As the fine understanding of those of ordinary skills, method of the present invention and Pharmaceutical composition are particularly suitable for using to its any vertebrates of needs, as birds, include, but are not limited to turkey, hen or chicken and other chickling and the animal or the mammal of freely walking about include, but are not limited to, domestic animal is as cat or dog object, farm animal, for example, but be not limited to cattle, horse, he-goat, sheep and pig object, wild animal, no matter be to live in open-air or live in the zoo, zoologize, as mice, rat, rabbit, goat, sheep, pig, Canis familiaris L., cats etc. promptly are used for veterinary's goals of medicine.

In addition, in treatment any high glucose level or hyperglycemia symptom, as I type and type ii diabetes, and for example, when renal failure, any symptom relevant after I type and the type ii diabetes, comprise the mankind as subject with malnutrition.

In addition, described subject can also be any vertebrates, and as one of above-mentioned animal, it need improve amino acid whose effectiveness and utilizability, described aminoacid for example is essential amino acids or condition aminoacid, particularly isoleucine, leucine, lysine and proline.The described mankind can also be the patients who needs the treatment malnutrition or increase amino acid whose effectiveness and utilizability, for example owing to renal failure, and operative treatment, for example, pancreatectomy or transplanting, old symptom, diabetes, athlete, age (child and old people), pregnancy, anorexia nervosa, bulimia, binge eating, mandatory binge eating, nutrition disease, the feed disorder (EDNOS) that metabolism disorder or other are not pointed out, decubital ulcer, the vertebrates of inappetence or because wasting disease.

List of references

(all documents that this paper quoted are all received with their full text form and are done this paper reference).

1.Windmueller，H.G.，&Spaeth，A.E.(1975)Intestinalmetabolism of glutamine and glutamate from the lumen as comparedto glutamine from blood.Arch.Biochem.Biophys.171：662-672.

2.Stoll，B.，Burrin，D.G.，Henry，J.，Hung，Y.，Jahoor，F.，&Reeds，P.J.(1999)Substrate oxidation by the portaldrained viscera of fed piglets.Am.J.Physiol.277：E168-E175.

3.Matthews，D.E.，Marano，M.A.，&Campbell，R.G.(1993)Splanchnic bed utilization of glutamine and glutamic acid inhumans.Am.J.Physiol.264：E848-E854.

4.Madej，M.，Lundh，T.，&Lindberg，J.E.(1999)Activitiesof enzymes involved in glutamine metabolism in connection withenergy production in the gastrointestinal tract epithelium ofnewborn，suckling and weaned piglets.Biol.Neonate 75：250-258.

5.Suryawan，A.，Hawes，J.W.，Harris，R.A.，Shimomura，Y.，Jenkins，A.E.，&Hutsun，S.M.(1998)Amolecular modelof human branched-chain amino acid metabolism.Am.J.Clin.Nutr.68：72-81.

6.Lambert，B.D.，Stoll，B.，Niinikoski，H.，Pierzynowski，S.，&Burrin，D.G.(2002)Net portal absorption of enterallyfedalpha-ketoglutarate is limited in young pigs.J.Nutr.132：3383-3386.

7.Kristens en，N.B.，Jungvid，H.，Fernandez，J.A.，&Pierzynowski，S.G.(2002)Absorption and metabolism ofa-ketoglutarate in growing pigs.J.Anim.Physiol.Anim.Nutr.86：239-245.

8.Bergmeyer，H.U.，&Bemt，E.(1974)2-oxoglutarate.UVspectrophotometric determination.In：Methods of enzymaticanalysis，2nd Ed.(Bergmeyer，H.U.，ed.).Academic Press，NewYork，NY

9.Pajor，A.M.(1999)Sodium-coupled transporters forkrebs cycle intermediates.Annu.Rev.Physiol.61：663-682.

10.Murphy，J.M.，Murch，J.M.，and Ball，R.O.(1996)Proline is synthesized from glutamate duringintragastricinfusion but not during intravenous infusion in neonatalpiglets.J.Nutr.126：878-886.

Embodiment

Below will the present invention will be described by a plurality of indefiniteness embodiment.Although the embodiment in conjunction with some disclosure describes the present invention, the technical staff can predict change that still belongs to the appended claims scope or the combination of specifically not mentioning.

Material and the method part of example 1-2

Animal is kept

Stable breeding of animal and nursing meet the regulation of U.S.Department of Agricultureguidelines.

Research design

Female piglets (n=9) is from Texas Department of Criminal Justice, Huntsville, and TX buys.

These pigs (14 days big) arrive Children ' s Nutrition Research Center, and carry out 7 days the phase of adjustment, (Middleton WI) raises for LitterLife, Merrick with liquid formula with the speed of 50g/ (kg.d).

The prescription of described milk replacer (every kilogram of dry): 500g lactose, 100g fat and 250g albumen.

After 7 days, cancel the food of described piglets, spend the night, and allow them perform the preparation (2) that undergos surgery as stated above.

Say that simply under isoflurane anesthesia and aseptic condition, (external diameter is 1.27mm with the polyethylene intubate, Becton Dickinson, Sparks, MD) the common door vein of insertion piglets, and with silicone rubber intubate (external diameter 1.78mm) insertion external jugular vein and carotid artery.

(Ithaca NY) is placed on around the portal vein for internal diameter 8-10mm, Transonic with the ultrasonic flow probe.

With the silicon intubate (external diameter is 2.17mm, Baxter Healthcare, McGaw Park IL) inserts duodenal inner chamber, described intubate is filled and is contained heparin (2.5 * 10 ⁴U/L) Sterile Saline, and from abdominal part left side (portal vein and duodenal intubation, flow probe guiding) or take out by abdomen from (jugular vein and carotid artery intubate) between the scapula.

Before being about to carry out surgical operation, animal is carried out intramuscular injection antibiotic (20mg/kg anthracene Flucloxacillin, Bayer, Shawnee Mission, KS), and intramuscular injection analgesics (0.1mg/kg butorphenol tartrate, Fort Dodge Labs, Fort Dodge, IA).

After operation, recover before the intestinal feed, these pigs were kept 24 hours by comprehensive parenteral absorption, and feed speed is 5mL kg ^-1h ^-1Allow these pigs from operation, recover with 7 day time.Food intake and weight increase speed return to preoperative level in all piglets bodies.

Sample preparation

Blood sample is placed on ice at once and centrifugal.

Collect blood plasma, at once in liquid nitrogen, and under-80 ℃, preserve up to analysis.

Amino acid analysis

In order to carry out the blood plasma amino acid analysis, the equal portions plasma sample of 0.2mL is mixed with the aqueous solution (4mmol/L) of isopyknic methionine sulfone, and blocked filter centrifugal 120 minutes by 10-kDa with the speed of 10,000 * g.

The described filtrate sample of equal portions of dry 50 μ L, and by reversed-phase HPLC to amino acid whose phenylisothiocyanate, PITC salt derivative analyze (Pico Tag, Waters, Woburn, MA).

Method by improved slightly Brgmeyer and Bernt (8) is measured plasma A KG.

Say that simply mensuration is to carry out in the working solution of 0.5mL, this solution comprises 100mmol/L phosphate buffer (pH 7.6), 4mmol/L ammonium chloride and 50 μ mol/L NADH.

In described working solution, add an amount of blood plasma of the AKG that contains 1-10nmol.

Under the 340nm wavelength, obtain initial absorption reading.

After described initial absorption reading, in each test tube, add the cattle GDH (G2501 of about 6 units (volume is 10 μ L); Sigma-Aldrich, St.Louis, MO).

After cultivating 10 minutes, under the 340nm wavelength, obtain second and absorb reading.

In the sample content of AKG directly with first and second readings between absorb weaken and be directly proportional.

By using standard curve to calculate AKG concentration.

Plasma ammonia is measured

Plasma ammonia is to use the spectrophotometry test kit, and (St.Louis MO) measures for 171C, Sigma-Aldrich.

Plasma glucose is measured

Plasma glucose is to use spectrophotometry test kit (315-100; Sigma-Aldrich, St.Louis MO) measures.

The blood bicarbonate is measured

In order to estimate the concentration of blood bicarbonate, with whole blood etc. duplicate samples (1.0mL) be placed on 10-mL Vacutainer (Becton Dickinson, Franklin Lakes, NJ) in, and add the perchloric acid (10%wt/wt) of 0.5mL.

Will be by soda lime (Sodasorb; Grace Container Products, Lexington, MA) filtering room air (10mL) injects Vacutainer, is drawn in the air-tightness syringe, and puts into second Vacutainer.

The isotopes concentration of carbon dioxide is than mass spectrograph (ANCA at the continuous-flow gas isotope in the gaseous sample; Europa Instruments, Crewe U.K.) upward measures.

The mensuration of blood plasma ketoisocaproate

Blood plasma ketoisocaproate (KIC) be by cation-exchange chromatography (AG-SOV resin, Bio-Rad) isolating.

With sodium hydroxide (100 μ L; 10N) with azanol HCl (200 μ L; 0.36M) handle eluent, and heat (60 ℃; 30 minutes).After cooling, the pH of sample is adjusted to＜2.

With the ethyl acetate extraction keto acid of 5mL, and dry at ambient temperature in nitrogen.

The derivatization of KIC is to finish by the N-methyl-N-t-butyl-dimetylsilyl-trifluoroacetamide that adds 50 μ L+1%t-butyl-dimethyl chlorosilane.

By EI GC-MS (Hewlett Packard 5970GC-mass spectrograph is equipped with HewlettPackard 5890 Series II GC), under the condition of 316m/z and 317m/z, pass through the isotopes concentration of monitoring ion measurement KIC.

The mensuration of carbamide isotopes concentration

By EI GC-MS assay determination blood plasma carbamide isotopes concentration.

With the iced acetone protein precipitation from the blood plasma of 50 μ L of 200 μ L.

After vortex stirs, by centrifugalize albumen, and take out supernatant, dry under nitrogen.

In described exsiccant supernatant, add 1: 20 diluent and the concentrated HCl (30wt%) of the propionic aldehyde bid (dimethyl-acetal) of 250 μ L.At room temperature cultivated described sample 2 hours, be evaporated to then drying (Speedvac, Savant Insturments, FormaScientific, Marietta, OH).

N-methyl-N-t-butyl-dimetylsilyl-trifluoroacetamide+1%t-butyl-dimethyl chlorosilane with 50 μ L carries out derivatization to described carbamide, and uses EI GC-MS to analyze by monitoring ion measurement blood plasma isotopes concentration under the condition of 153-155m/z.

Calculate

The clean portal vein balance of metabolite [μ mol/ (kg h)] is calculated by the following method:

(Conc. _PORT-Conc. _ART)×PBF (1)

When Conc. was haemoconcentration (μ mol/L), PORT and ART represented portal vein and arterial blood, and PBF is portal vein flow [L/ (kg h)].

Calculate the whole leucine flow of whole blood [Q by the following method; μ mol/ (kg h)]:

Q=R*[(IE _{Inculcate notes}/ IE _{Blood plasma})-1] (2)

R is tracer infusion rate [μ mol/ (kg h)], and IE _{Inculcate annotate and}IE _{Blood plasma}Be respectively the tracer inculcated and the isotopes concentration (being expressed as mol%) of Plasma K IC.

Health CO ₂Output is calculated as follows:

Wherein, IE inculcates that to annotate be to inculcate H in the notes ¹³CO ₃ ^-The concentration of (molar percentage that exceeds), IE tremulous pulse bicarbonate is the concentration (molar percentage that exceeds) in the arterial blood, and each the processing stage tracer infusion rate e[μ mol/ (kg h) during the intravenous perfusion bicarbonate that carries out].Whole formula is divided by 0.82, so that proofread and correct the recovery of the labelling carbon in the bicarbonate of inculcating.

Whole leucine oxidation [μ mol/ (kg h)] is calculated by the following method:

[IE _CO2/ IE _KIC] * formula 3 (4)

Wherein, IE _CO2Be at 1- ¹³The isotopes concentration of bicarbonate during the C-leucine is inculcated, and IE _LEUBe at 1- ¹³1-during the C-leucine is inculcated ¹³The isotopes concentration of C-KIC.

It is to being incorporated into the leucic estimation in the muscle that whole non-oxide leucine is handled (NOLD).NOLD[μ mol/ (kg h)] calculate by following formula:

NOLD=formula 2-formula 4 (5)

Whole leucine occurrence rate (Ra) [μ mol/ (kg h)] is to the metabolic estimation of albuminolysis, and calculates as follows:

Ra=formula 2-leucine picked-up (6)

Whole carbamide flow calculates as follows:

The carbamide flow=[([ ¹⁵N ₂] carbamide IE/[ ¹⁵N ₂] carbamide PE)-1] * [ ¹⁵N ₂] urine

Plain IR (7)

Wherein, IE inculcates notes concentration, and PE is the plasma concentration under the steady statue during the infusion carbamide, and IR is an infusion velocity.

Statistical analysis

Concerning all statistical test, 0.05 p value is used to indicate significance,statistical.

In example 1, AKG is to tremulous pulse, pylic effect, and single amino acids, AKG, glucose, analyzing appears in the dynamic (dynamical) clean portal vein of ammonia and leucine, use the general linear model method (Minitab.Inc., State College, PA).Described model comprises the effect and the pig of adding AKG.Pig is included as stochastic variable.Use LSMEANS option computing meansigma methods.Utilize the clean portal vein balance of unidirectional Student ' s T inspection AKG whether obviously greater than the null value during control treatment.

Example 1-plasma A KG, glucose, ammonia, the measurement of blood flow and whole carbamide flow.

Purpose

The purpose of present embodiment is to assess the AKG infusion to plasma A KG, glucose, ammonia, the influence of blood flow and whole carbamide flow.

Zoopery

Before this experiment of beginning, deprive the food 15 hours of piglets.

Testing the same day,, begin continuous duodenum and pour into preprepared milk replacer [Litter Life, Merrick, Middleton, WI at-1 hour; 7.75mL/ (kgh)] (7.75mL/kg; The 25%wt/wt aqueous solution; Oral), it is as the preparation of 25% (wt/wt) aqueous solution, and it can provide about 920kJ and 12.5g albumen/(kg d).

With saline (contrast; 930mmol/L NaCl) or sodium-AKG (Na-AKG), 930mmol/L, available from Sigma-Aldrich, St.Louis MO) is dissolved in the described milk replacer.

According to former data (6), select AKG content by the laboratory of outside, wherein,, need picked-up to surpass 2.5% diet dry in order to observe detectable AKG portal vein balance.

Pig is also accepted ¹⁵N ₂Successive 6 hours infusions (98% of intravenous (200 μ mol/kg) of-carbamide [20 μ mol/ (kg h)]; Cambridge Isotope Laboratories).

In the 0h time, begin 2 hours successive NaH ¹³CO ₂Preprepared (the 15 μ mol/ (kg h) of infusion (15 μ mol/kg); 99%; Cambridge IsotopeLaboratories, Andover, MA).

At beginning NaH ¹³CO ₂Obtain the tremulous pulse sample in 0,90,105 and 120 minutes after the infusion, be used to measure whole CO ₂Output.

At 2 hours, stop NaH ¹³CO ₂Infusion, and beginning 1- ¹³C-leucine (40 μ mol/ (kg h); 99%; Cambridge Isotope Laboratories) continuous 4 hours infusions preprepared (40 μ mol/kg h).

Obtain tremulous pulse and portal vein sample at 4,5 and 65 hours, be used to measure leucine and carbamide kinetics, and ammonia, AKG, glucose and amino acid whose mass balance.

All pigs all accept contrast in the mode of completely random design and AKG handles, and the interval is at least 24 hours between handling.

The result

Plasma A KG has been shown, glucose, ammonia, blood flow and whole carbamide flow in table 1.

Show the 1.AKG infusion to metabolite concentration, clean portal vein balance and whole 1- ¹³The C-leucine and ¹⁵N ₂-carbamide effect of kinetics.

	AKG ¹(dry %)
	AKG ¹(dry %)			0	3.75	P
	The AKG rate of flooding, and μ mol/ (kg, h)	0	930	0	3.75	P	-
Portal vein flow L/ (kg h)	The AKG rate of flooding, and μ mol/ (kg, h)	0	930	3.21±0.28 ²	3.36±0.27	0.34	-
Portal vein flow L/ (kg h)	Artery A KG, μ mol/L	13.8±1.7	27.4±3.6	3.21±0.28 ²	3.36±0.27	0.34	＜0.01
Portal vein AKG, μ mol/L	Artery A KG, μ mol/L	13.8±1.7	27.4±3.6	22.0±1.4	64.6±5.9	＜0.001	＜0.01
Portal vein AKG, μ mol/L	The clean portal vein balance of AKG, and μ mol/ (kg, h)	19.7±2.8	95.2±12	22.0±1.4	64.6±5.9	＜0.001	＜0.001
The clean portal vein balance of AKG, the % of infusion	The clean portal vein balance of AKG, and μ mol/ (kg, h)	19.7±2.8	95.2±12	-	10.23±0.57	-	＜0.001
The clean portal vein balance of AKG, the % of infusion	The clean portal vein balance of glucose mg/ (kg h)	303.1±61	203.9±69	-	10.23±0.57	-	＜0.05
The clean portal vein balance of ammonia μ mol/ (kg h)	The clean portal vein balance of glucose mg/ (kg h)	303.1±61	203.9±69	520.1±66	561.1±53	0.91	＜0.05
The clean portal vein balance of ammonia μ mol/ (kg h)	Whole carbamide flow μ mol/ (kg h)	398.3±35	377.8±39	520.1±66	561.1±53	0.91	0.56

¹AKG, alpha Ketoglutarate; ²SEM

[(930 μ mol/ (kg.h)) improved (P＜0.01) tremulous pulse and portal vein AKG concentration to the AKG infusion, and the clean portal vein balance of AKG.Even not having AKG to beat under the duodenal situation, the clean portal vein of AKG absorbs [19.7 ± 2.8 μ mol/ (kg.h)] also significantly greater than 0.But, compared with the control, handle the clean portal vein that has significantly improved AKG with AKG and absorb (P＜0.001).The clean portal vein balance of AKG is 95 μ mol/ (kg.h), and it only is equivalent to 10.23% of infusion amount.

10.23% clean portal vein balance has been over-evaluated the AKG that inculcates in fact slightly and has been absorbed, because when only inculcating saline, exists the statistics of AKG significantly to absorb.If according to control diet gauged words are carried out in the absorption of AKG, the ratio of the AKG that inculcates that occurs in portal vein is drained is reduced to 8.12%.

What is interesting is, handle, reduced the clean portal vein balance (P＜0.05) of glucose with AKG.The influence that portal vein flow, clean portal vein balance of ammonia and whole carbamide flow are not handled by AKG.

Handle by AKG, tremulous pulse and pylic concentration of proline have improved (P＜0.05), and the portal vein leucine tends to (P＜0.01) increase (data are not delivered).Amino acid whose portal vein mass balance has been shown in table 2.AKG handles and has improved leucine, the portal vein mass balance of lysine and proline (P＜0.05), and tend to the portal vein mass balance that (P＜0.10) improves isoleucine.

The amino acid whose clean portal vein balance of the pig of the duodenum infusion of table 2. acceptance 0 or 930 μ mol/ (kgh) AKG (n=5).

Aminoacid	Contrast		AKG
Aminoacid	Contrast		AKG			The portal vein balance		The portal vein balance
	μmol/(kg h)	Take in %	μmol/(kg h)	Take in %		The portal vein balance		The portal vein balance
	μmol/(kg h)	Take in %	μmol/(kg h)	Take in %	Essential amino acids
Isoleucine	164.5±26	100.1	230.2 ^b±28	140.0	Essential amino acids
Isoleucine	164.5±26	100.1	230.2 ^b±28	140.0	Leucine	294.9±44	76.3	438.6 ^a±50	113.4
Phenylalanine	80.4±11	83.3	95.2±11	98.7	Leucine	294.9±44	76.3	438.6 ^a±50	113.4
Phenylalanine	80.4±11	83.3	95.2±11	98.7	Valine	218.5±33	85.2	279.3±32	108.9
Histidine	27.7±11	43.1	45.9±3.8	71.4	Valine	218.5±33	85.2	279.3±32	108.9
Histidine	27.7±11	43.1	45.9±3.8	71.4	Threonine	185.0±40	66.4	210.9±18	75.7
Lysine	237.7±35	72.3	324.5 ^a±37	98.8	Threonine	185.0±40	66.4	210.9±18	75.7
Lysine	237.7±35	72.3	324.5 ^a±37	98.8	Tryptophan	38.6±6.4	-	47.2±4.3	-
The condition essential amino acids					Tryptophan	38.6±6.4	-	47.2±4.3	-
The condition essential amino acids					Arginine	95.2±24	85.8	109.0±19	98.3
Proline	216.4±25	69.9	354.5 ^a±32	114.5	Arginine	95.2±24	85.8	109.0±19	98.3
Proline	216.4±25	69.9	354.5 ^a±32	114.5	Tyrosine	85.7±12	100.6	115.8±17	135.9
Non essential amino acid					Tyrosine	85.7±12	100.6	115.8±17	135.9
Non essential amino acid					Alanine	539.6±61	182.9	557.8±48	189.0

Aspartic acid	28.2±4.6	9.2	29.7±6.0	9.6
Aspartic acid	28.2±4.6	9.2	29.7±6.0	9.6	Agedoite	169.9±23	-	185.6±18	-
Glutamic acid	64.2±23	14.9	80.1±17	18.6	Agedoite	169.9±23	-	185.6±18	-
Glutamic acid	64.2±23	14.9	80.1±17	18.6	Glutamine	17.2±12	-	25.5±45	116.0
Glycine	167.0±27	109.4	177.2±20	-	Glutamine	17.2±12	-	25.5±45	116.0
Glycine	167.0±27	109.4	177.2±20	-	Serine	213.3±89	94.9	244.7±64	108.3

^aDifference (P≤0.05) with contrast; ^bDifference (P＜0.10) with contrast

¹AKG, α-Tong Wuersuan; ²LS meansigma methods ± SEM

Whole leucine kinetics as shown in Figure 1.Whole flow NOLD, the influence that Ra and oxidation are not handled by AKG.

The average enteric cavity that example 2-measures AKG disappears

Purpose

The purpose of present embodiment is that the average enteric cavity of assessing the AKG medicine group that inculcates disappears.

Zoopery

(LitterLife Merrick) carries out duodenum medicine group to pig (n=7) and inculcates (7.75mL/kg with the liquid formula that contains 25mg/mL AKG sodium (1040 μ mol/kg BW); 25% (wt/wt) aqueous solution).

After 1 hour, slaughter pig.

Carefully small intestinal is clamped at duodenum near-end and caecum far-end, taken out, and with 2 * 50mL normal saline washing, so that clean small intestinal.

Collect cleaning mixture, merge, and with 15mL etc. duplicate samples quick freezing in liquid nitrogen, and preserve down at-80 ℃, the AKG that is used for subsequently analyzes.

The result

Inculcate the AKG medicine group of 1040 μ mol/kg.Average enteric cavity disappearance is 663 ± 38 μ mol/kg in 1 hour.63.8 of the 1040 μ mol/kg AKG that this is equivalent to inculcate.

The discussion and the common conclusions of experiment 1 and 2

In example 1, the AKG continuous irrigation is passed in the duodenum, and has only 10% the AKG that inculcates to appear in the portal vein discharge liquor.

Have only 10% the AKG that inculcates to appear at the some kinds of probabilities that this discovery in the portal vein blood plasma has produced the destiny of enteric cavity AKG.A kind of possible explanation that low AKG portal vein is occurred is that enteric cavity AKG transhipment is limited.Sodium/dicarboxylic acids can be transported the cotransporter of AKG, and it is present in (9) on the pig brush border membrane, as if therefore, AKG can be absorbed by enterocyte.In order to verify this probability, we have inculcated the single medicine of the duodenum group of 1040 μ mol/kg, and find to surpass 660 μ mol/kg disappeared from the small intestinal of piglets in 1 hour (example 2).Therefore, about 64% AKG medicine group is only just disappearing in duodenal enteric cavity within an hour.

The influence (6) of AKG infusion appears not being subjected to as the clean portal vein of former viewed glutamic acid and glutamine.If the AKG that absorbs changes into glutamic acid, it can be discharged in the portal vein blood or change into other aminoacid.

But, can estimate that the release of glutamic acid and glutamine can not increase because of AKG, even occurred to these amino acid whose remarkable conversions, because under normal raising condition, seldom there are food glutamic acid or glutamine to discharge (list of references 1,2) by PDV.Verified (10), proline can be synthetic by enteral glutamic acid by intestinal tissue.Suppose in the pig that AKG handles, the equilibrated 138.1 μ mol/ (kg h) that increase to of the clean portal vein of proline, and the AKG that surpasses 800 μ mol/ (kg h) does not participate in the portal vein balance, and the clean portal vein balance of proline is because the conversion of AKG causes probably fully.But, AKG should cause the reduction of portal vein nitrogen balance to this extensive conversion of proline in intestinal cell, and still, the portal vein nitrogen balance remains unchanged.Shortage to the influence of portal vein nitrogen balance is embodied on the similar speed of whole urea synthesis in two groups equally.

Reaction between branched-chain amino acid (BCAA) transaminase energy catalysis AKG and the branched-chain amino acid (leucine, isoleucine and valine).BCAA is changeed ammonia, forms glutamic acid by AKG, and forms corresponding keto acid by BCAA.The AKG that replenishes may form glutamic acid and cause the minimizing to the clean release of BCAA by PDV by the transamination that promotes BCAA.But, AKG can increase leucic portal vein and discharge, but it can not influence whole leucine kinetics.The clean portal vein balance of lysine also can improve with AKG.Because handle concerning a lot of aminoacid with AKG, a lot of amino acid whose clean portal vein balances are near 100%, and unclear is because AKG has saved described aminoacid or owing to the Proteolytic enzyme in the internal organs of portal vein-drain has increased amino acid whose release.

The destiny of the another possibility of AKG in enterocyte is by the TCA cyclic oxidation.If all carbon of inculcating with the AKG form is oxidized to CO ₂, can estimate to increase CO by PDV ₂Output, yet CO on the whole ₂Output do not increase because AKG inculcates.What is interesting is, handle that by AKG the clean portal vein balance of glucose has been reduced.

Because a large amount of AKG has disappeared from the enteric cavity of small intestinal, and do not cause the portal vein drainage of AKG or the clean balance of the metabolic amino acid product of AKG, the destiny of the AKG of intestinal picked-up it be unclear that.But, when AKG is beaten into duodenum, have only 10% enteric cavity quantity delivered to appear in the portal vein discharge liquor, yet the AKG of this quantity is enough to improve the circulation composition of portal vein balance and described chemical compound.Therefore, although the definite metabolism destiny of AKG in enteric cavity is still uncertain, described result shows that the intestinal utilizability of diet AKG is limited.

The increase of the circulation A KG that is caused is to glutamic acid, glutamine, and ammonia, not influence appears in the clean portal vein of BCAA.

In addition, increase the AKG of system to PDV or whole leucine kinetics or the not influence of carbamide flow.The above results and former data are coincide, and wherein, AKG provides by the gastrointestinal tract approach.

Na-AKG and chitosan-AKG that example 3-intestinal is used absorb in enterocyte and the blood plasma aminoacid and keto acid, and the comparison of their metabolic effect

Purpose

The purpose of present embodiment is that the Na-AKG (or Na salt of AKG) that uses of comparison intestinal and chitosan-AKG are to absorbing aminoacid and keto acid and their metabolic effect in enterocyte and the blood plasma.In addition, by monitoring of blood blood plasma amino acid content mensuration Na-AKG and chitosan-AKG keto acid is changed into amino acid whose influence.This research has been checked AKG to influence keto acid and has been transformed and improve the synthetic hypothesis of albumen to amino acid whose intestinal.

Zoopery

To have three pigs altogether and be used for this experiment.The body weight of these pigs is approximately 20kg.These pigs separately are housed in the box, and feed standard food 4-5 days, so that adapt to new facility.Insert intubate and intestinal sleeve pipe by operation to pig then, and recovered with 3-7 days time.The operation method that is adopted is the method that is usually used in this area, and is conventionally known to one of skill in the art.

After operation, under this occasion, allow 3 days convalescent period, and use standard feed (3% body weight) feed once (10.00) every day.After convalescent period, using Na-AKG ((ii)) referring to experiment, use chitosan-AKG (experiment is (iii)) and do not use AKG (experiment (i)); Control experiment) measures the amino acid content in the blood plasma under the condition, further details is provided below.

The AKG application conditions:

Experiment (i): inculcate (i.d.) by duodenum, use keto acid or aminoacid (amine) (cumulative volume is 50ml), * " feed equivalent in early morning " 1 hour.

Provided 10 parts, (50ml dosage+50ml saline) with 1 hour.

It represents control experiment.

(described animal obtains and the aminoacid that is present in the same amount in the food that is equivalent to the feedstuff in early morning usually in * " feed equivalent in early morning " expression).Blood sample (being background * * level in the time of 0 hour) is 0,1,2, and 4 disappearances are gathered.

(* * background sample is defined in aminoacid/keto acid and inculcates the sample of 0 time before)

Blood sample (the 5ml whole blood is used for amino acid analysis, from tremulous pulse, and portal vein, hepatic vein is captured on the ethylenediaminetetraacetic acid (EDTA) that contains aprotinin, so that stop blood coagulation and proteinase activity.

(processing method relates to 5 EDTA+ aprotiniies of use, and is centrifugal, and-20 ℃ of following frozen plasma).

Experiment (ii)

Inculcate (i.d.) by duodenum and use and blended keto acid of Na-AKG or aminoacid (amine) (cumulative volume is 50ml), dosage is * " feed equivalent in early morning ", with 1 hour time (provided 10 parts with 1 hour time, 50ml dosage optionally uses saline).

0,1,2,4 hours collection blood samples (in the time of 0 hour, being background level).

Blood sample (the 5ml whole blood is used for amino acid analysis, from tremulous pulse, portal vein, hepatic vein) is captured on the ethylenediaminetetraacetic acid (EDTA) that contains aprotinin, so that stop blood coagulation and proteinase activity.

Experiment (iii)

Inculcating (i.d.) by duodenum uses and blended keto acid of chitosan-AKG or aminoacid (amine) (cumulative volume is 50ml), dosage is * " feed equivalent in early morning ", with 1 hour time (provided 10 parts with 1 hour time, 50ml dosage optionally uses saline).

The result

The result of following this research of table 3 expression:

Table 3. is the increment of free amino acid increase in the blood after using aminoacid:

Time (hour)	ΔI(mmol/L)	ΔII(mmol/L)	ΔI(mmol/L)	ΔII(mmol/L)
Time (hour)	ΔI(mmol/L)	ΔII(mmol/L)	ΔI(mmol/L)	ΔII(mmol/L)		Tremulous pulse	Tremulous pulse	Hepatic vein	Hepatic vein
1	-0.5 ^a	0.65 ^c	0.43 ^A	0.23 ^A		Tremulous pulse	Tremulous pulse	Hepatic vein	Hepatic vein
1	-0.5 ^a	0.65 ^c	0.43 ^A	0.23 ^A	1.5	0.04 ^b	1.11 ^d	1.61 ^B	1.85B ^C
2.5	-0.48 ^a	1.69 ^d	1.59 ^B	1.94 ^C	1.5	0.04 ^b	1.11 ^d	1.61 ^B	1.85B ^C

I represents Na-AKG salt

II represents chitosan-AKG salt

Delta time increment=(at the Δ aminoacid of 0 time-) at 1,1.5 and 2.5 hour amino acid content

Provide result's difference to represent significant difference when the p＜0.05 with the lower case or upper case letter.

The discussion of example 3 and general conclusion

Present embodiment has confirmed that chitosan-AKG salt can improve the absorption of essential amino acids, and this improvement is better than the result who obtains with Na-AKG.

This discovery is important, and with the gastrointestinal tissue that diet aminoacid is used to improve better impaired, for example, the amino acid whose absorption that is common in diabetes or old people patient's the gastrointestinal tissue is relevant.

Claims

1. a vertebrates that is used to improve comprise mammal and birds is to the method for amino acid whose absorption, this method comprises to the vertebrates that comprises mammal and birds, use AKG with enough quantity and/or enough speed, AKG derivant or metabolite, AKG analog or its mixture produce the influence that needs so that can absorb aminoacid.

2. method as claimed in claim 1, wherein, described AKG, AKG derivant or metabolite, AKG analog or its mixture are selected from following one group: α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, chitosan-AKG, the salt of other of AKG and aminoacid and amino acid derivativges; One of AKG-and two-slaine, as CaAKG, Ca (AKG) ₂And NaAKG.

3. as method any among the claim 1-2, wherein, described vertebrates is a rodent, as mice, rat, Cavia porcellus or rabbit; Birds is as turkey, hen, chicken or other chickling; Farm animal is as cow, horse, pig, piglets or the farm animal of freely walking about; Or house pet, as Canis familiaris L. or cat.

4. as method any among the claim 1-2, wherein, described vertebrates is human.

5. as method any among the claim 1-4, wherein, described aminoacid is any essential amino acids.

6. method as claimed in claim 5, wherein, described essential amino acids is isoleucine, leucine, lysine and proline.

7. a vertebrates that is used to reduce comprise mammal and birds is to the method for the absorption of plasma glucose, this method comprises to the vertebrates that comprises mammal and birds uses AKG with enough quantity and/or enough speed, AKG derivant or metabolite, AKG analog or its mixture are so that can produce the influence that needs to glucose absorption.

8. one kind is used for prevention, suppress or alleviate the method for the vertebrate high plasma glucose symptom that comprises mammal and birds, this method comprises to the vertebrates that comprises mammal and birds, use AKG with enough quantity and/or enough speed, AKG derivant or metabolite, AKG analog or its mixture are so that can produce the influence that needs to described symptom.

9. as method any among the claim 7-8, wherein, described AKG, AKG derivant or metabolite, AKG analog or its mixture are selected from following one group: α-Tong Wuersuan (AKG), ornithine-AKG, arginine-AKG, glutamine-AKG, glutamic acid-AKG, leucine-AKG, chitosan-AKG, the salt of other of AKG and aminoacid and amino acid derivativges; One of AKG-and two-slaine, as CaAKG and NaAKG.

10. as method any among the claim 7-9, wherein, described vertebrates is a rodent, as mice, rat, Cavia porcellus or rabbit; Birds is as turkey, hen, chicken or other chickling; Farm animal is as cow, horse, pig, piglets or the farm animal of freely walking about; Or house pet, as Canis familiaris L. or cat.

11. as method any among the claim 7-10, wherein, described vertebrates is human.

12. as method any among the claim 8-11, wherein, described high plasma glucose symptom is I type or type ii diabetes.

13.AKG, AKG derivant or metabolite, AKG analog or its mixture production are used to prevent, alleviate or treat the purposes of the compositions of high plasma glucose symptom.

14. as the purposes of claim 13, wherein, described high plasma glucose symptom is I type or type ii diabetes.

15.AKG, AKG derivant or metabolite, AKG analog or its mixture production are used to prevent, alleviate or treat the purposes of underfed compositions.

16. as purposes any among the claim 13-15, wherein, described compositions is a Pharmaceutical composition, optionally comprise can be medicinal carrier and/or additive.

17. as purposes any among the claim 13-15, wherein, described compositions is food or feed additive.

18. as the purposes of claim 17, wherein, described food or feed additive are the compositions of food additive and/or solid food and/or drink form.

19. as purposes any among the claim 13-18, wherein, in the compositions of described production, described AKG, AKG derivant or metabolite, AKG analog or its mixture treat effective dose.

20. as the purposes of claim 19, wherein, described treatment effective dose is 0.01-0.2g/kg body weight dosage every day.