CN110464717B

CN110464717B - Application of 2, 4-dihydroxybenzoic acid in preparation of blood sugar reducing medicine

Info

Publication number: CN110464717B
Application number: CN201910899911.5A
Authority: CN
Inventors: 张建国; 任武贤; 韩应兵; 堐榜琴
Original assignee: Ya Bangqin
Current assignee: Ya Bangqin
Priority date: 2019-09-23
Filing date: 2019-09-23
Publication date: 2022-02-18
Anticipated expiration: 2039-09-23
Also published as: CN110464717A; WO2021057680A1

Abstract

The invention discloses application of 2, 4-dihydroxy benzoic acid in food and medicine for treating related diseases caused by iron overload. The 2, 4-dihydroxybenzoic acid is replaced by an isomer of 2, 4-dihydroxybenzoic acid. The isomers of 2, 4-dihydroxybenzoic acid include 2, 3-dihydroxybenzoic acid, 2, 5-dihydroxybenzoic acid, 3, 4-dihydroxybenzoic acid, and 3, 5-dihydroxybenzoic acid. The invention has reasonable design, and directly takes the 2, 4-dihydroxy benzoic acid and the isomer to achieve the aim of directly reducing the 'iron ions' in the non-protein complex iron in the cells, thereby quickly removing the redundant iron in the body and achieving the aim of treating various related diseases (particularly diabetes) caused by iron overload.

Description

Application of 2, 4-dihydroxybenzoic acid in preparation of blood sugar reducing medicine

Technical Field

The invention relates to the field of iron-reducing medicines, in particular to application of 2, 4-dihydroxybenzoic acid in food and medicines for treating related diseases caused by iron overload.

Background

More and more research results show that some major diseases are associated with intracellular iron overload, such as: diabetes, hypertension, hyperlipemia, liver cirrhosis, hyperuricemia, gout disease, pulmonary fibrosis, rheumatoid arthritis, heart disease, angiosclerosis, cerebral infarction, senile dementia, tumor, hepatitis B virus, muscular atrophy, aging, depression, blood disease, etc. The experts point out that: in humans 60% of the common diseases are associated with iron overload, and more researchers predict: if the problem of iron overload in cells can be solved, the life of human can be prolonged by 10-15 years.

Iron overload is a common phenomenon, which is determined by the metabolic pattern of iron. Iron has no obvious excretion mechanism in human body and can be recycled, the total amount of iron is controlled mainly by controlling absorption by intestinal mucosa epithelial cells, and normally, the iron in cells is increased with the age. Clinically, absolute iron deficiency is very rare and can only occur under the conditions of gastrointestinal tract resection, chronic hemorrhage, hemolysis, pregnancy and the like, and common anemia patients have iron overload and mainly have problems in the conversion mechanism of stored iron. The iron in hemoglobin and some functional enzymes is divalent, while the iron stored and transported is all trivalent, and thus iron overload is primarily for trivalent iron. On one hand, ferric iron overload can directly react with macromolecular compounds to form insoluble particles, so that iron deposition is caused, and severe consequences caused by ferric iron deposition are caused in liver cirrhosis, pulmonary fibrosis, heart disease and the like; on the other hand, the ferric iron overload can directly induce the generation of free radicals, thereby triggering lipid peroxidation and seriously damaging the cell function. At present, few medicines clinically applied to treatment of iron overload are available, and only three medicines, namely deferoxamine, deferiprone and snout deferasirox, can not enter cells, and mainly complex and replace ferric iron in serum ferritin, so that the three medicines have the characteristics of large dosage, long time and large side effect, can not be used as general medicines for treating iron overload in cells, and are only clinically used for treating iron overload caused by excessive blood transfusion of thalassemia patients at present. Insulin resistance in diabetic patients is clearly known to be associated with iron overload, but no iron removal drugs are clinically available. Therefore, the iron reduction treatment of diabetes and other related diseases still belongs to a blank field clinically at present.

The reason why the iron-reducing medicine cannot be used for treating related diseases clinically until now, and the actual situation is that no medicine is available, so the iron-reducing medicine has not been developed for a long time because people have misdistinguishing about iron overload: since the discovery of ferritin, it was considered more as the most important form of iron storage in the body, equating "iron overload" to "iron storage overage". The intracellular ferritin concentration cannot be measured at present, but ferritin in blood is released by each tissue cell and is the aggregate of ferritin released by each tissue cell, so that the concentration of ferritin in serum can basically reflect the storage condition of iron in vivo under normal conditions. Based on this recognition, serum ferritin is the main indicator for iron overload and is the only target for reducing iron overload. This presents two problems: firstly, the iron removal place is limited to be in blood, the concentration of ferritin in the blood is reduced, and cells can naturally release again, so that the aim of reducing the concentration of ferritin in the cells is fulfilled, the dosage is very large, the iron-infusion complexing agent is more than 6g per day, the required time is very long, and generally more than one year is needed; secondly, the capacity of complexing iron of the medicament is limited to be stronger than that of ferritin, and the iron in the ferritin can be removed only by the capacity. In this way, some of the main enzymes have their iron complexes removed, which can cause significant side effects. Under the frame of inertial thinking, the research and development of iron removal medicines have no great breakthrough.

Iron overload induces related diseases, mainly free iron ions play a dominant role. The ferric iron in ferritin, however, is inactive and does not induce free radical reactions. Under normal conditions, the average loading capacity of the ferritin in human plasma only reaches 20% -30% of the capacity of the ferritin human plasma, so that the free iron salt in the plasma is practically zero. Ferritin is an antioxidant, and its presence inhibits the presence of free ferric ions, so that high serum ferritin concentrations represent only iron overload, do not damage cells, and only free ferric ions can damage cells and cause disease. It appears that reducing the concentration of free ferric ions by reducing the serum ferritin concentration is a round trip, and even a very wasteful practice.

How the free iron ions in the cells are present, first the transferrin in the blood is analyzed for how to deliver iron to the ferritin in the cells: the iron in transferrin is also inactive, by first binding to cell membrane receptors, forming a pit, and then lowering the local pH to 6, at which point ferric ions are released and return to the blood itself. The released ferric ions are not directly combined with ferritin, but are firstly combined with citric acid, ATP, ADP, CTP, phospholipid and the like in cytoplasm, and the low molecular weight 'non-protein complex iron' has the property of free iron and can catalyze Haber Weiss reaction and is called as flowing biological free iron. The iron plays an important role in the growth, development and metabolism of cells, and is most closely related to diseases or is a main cause of cell function reduction, and the iron is removed to treat the diseases mainly by removing the iron. The presence of this complex iron has been identified in synovial fluid of rheumatoid arthritis, which may be the major cause of the difficulty in curing the disease. The non-protein complex iron also has a very important function, and can be complexed with the iron regulatory protein to ensure that the activation and the conformation of the iron regulatory protein are changed, finally the ferritin and the transferrin gene are induced to express, and in turn, the ferritin can further compete for the ferric iron in the non-protein complex iron to ensure that the concentration of the ferric iron is further reduced. Thus, there is a balance between ferritin and the non-protein complex iron, and it is this balance that maintains the normal function of the cell. The aim of reducing non-protein complexed iron can be finally achieved by reducing the concentration of serum ferritin, but the iron is faster and easier to complex because the iron is far from entering cells directly. Aspirin can specifically induce the expression of ferritin gene, has time and dose effect, so aspirin has the function of removing non-protein complex iron, and can be used for treating cardiovascular and cerebrovascular diseases, etc., but its iron-removing method is indirect, and can promote the combination of free iron and regulatory protein, so that its action is very limited, and it is only applicable to old people with high free iron. With the aging, insoluble macromolecular compounds in cytoplasm are more and more, the fluidity of non-protein complex iron is poorer and more, and in order to ensure the continuous generation of antioxidant ferritin, a higher concentration of the non-protein complex iron is needed for stimulation, so that the more aged cells are, the higher the concentration of the non-protein complex iron is, which is not transferred by people's will, and is the essence of aging.

It has been proposed that deferoxamine is insufficient to treat some serious diseases due to iron overload and the compound sought must satisfy two conditions: firstly, the iron complex can enter cells, and secondly, the iron complex capacity is weaker. The weak ability of complexing iron is a general concept, mainly considering the problem of side effects, but the degree of the complexing ability is the key to the success of reducing iron. Through intensive research, the applicant refines the two conditions into three conditions, namely that the iron complex can enter cells, the iron complexing capacity of the iron complex is smaller than that of ferritin, and the iron complexing capacity of the iron complex is larger than that of non-protein iron complex in the cells. Only if the three conditions are met, the aims of rapid and accurate iron removal and small side effect can be achieved.

Disclosure of Invention

Based on the above, the treatment of iron overload is essentially to reduce the trivalent iron in the non-protein complex iron entering the cell, thereby achieving the purpose of reducing the free iron in the cell.

In this regard, the applicant has found a compound which fully satisfies the above "three conditions": 2, 4-dihydroxybenzoic acid and isomers.

1. Gross CE experiments confirmed that the compound was able to enter the cell.

2. 2, 4-dihydroxy benzoic acid is added into the solution of citric acid complex iron, so that the citric acid can be dissociated, and the fact that the capacity of the 2, 4-dihydroxy benzoic acid complex iron is stronger than that of intracellular non-protein complex iron is shown.

3. The 2, 4-dihydroxy benzoic acid complexed with iron in the dilute alkali solution has no ferric hydroxide separated out, which indicates that the complexed iron is basically inactivated. It has far less iron complexing ability than ferritin.

4. Mice taking 2, 4-dihydroxybenzoic acid detected iron-complexed 2, 4-dihydroxybenzoic acid in the urine, indicating that iron complex could be excreted in the urine. The compound is an iron reagent, has no toxicity or side effect, and is listed as a food additive or a spice in the United states and Europe.

Therefore, the 2, 4-dihydroxy benzoic acid and the isomer can be applied to food and medicines for treating related diseases caused by iron overload.

Isomers of 2, 4-dihydroxybenzoic acid include 2, 3-dihydroxybenzoic acid, 2, 5-dihydroxybenzoic acid, 3, 4-dihydroxybenzoic acid, 3, 5-dihydroxybenzoic acid.

The compound has the main function of reducing intracellular non-protein complex iron, cannot be directly measured at present, but has a linear relation with serum ferritin, and can be indirectly represented by the level of the serum ferritin, so that the change of the intracellular non-protein complex iron can be accurately judged by measuring the serum ferritin value under the conditions of no infection, no inflammation and no alcoholism. Thus, by reducing intracellular non-protein complexed iron, and thus serum ferritin concentrations, a variety of diseases resulting from intracellular non-protein complexed iron overload can be treated, including: diabetes, hypertension, hyperlipemia, liver cirrhosis, pulmonary fibrosis, hyperuricemia, gout, rheumatoid arthritis, heart disease, angiosclerosis, cerebral infarction, senile dementia, tumor, hepatitis B virus, depression, aging, blood disease, etc.

The 2, 4-dihydroxy benzoic acid and isomer raw materials are derived from food additives, spices and medical raw materials, and are safe and free of toxic and side effects.

The specific dosage is as follows: 2, 4-dihydroxybenzoic acid and isomers, adult dose: the medicine is taken by 0.5-3 g every day in three times. The preparation formulation is as follows: various dosage forms such as oral preparation and injection.

The invention has reasonable design, and directly takes the 2, 4-dihydroxy benzoic acid and the isomer to achieve the aim of directly reducing free iron in non-protein complex iron in cells, thereby quickly removing redundant iron in the body and achieving the aim of treating various related diseases (particularly diabetes) caused by iron overload.

Drawings

FIG. 1 shows a control diagram of fasting blood glucose in a mouse experiment.

FIG. 2 shows a control diagram of serum ferritin values in a mouse experiment.

FIG. 3 is a graph showing a control of HBA1C (glycated hemoglobin value) in a mouse experiment.

Detailed Description

Specific examples of the present invention (clinical application in the treatment of diabetes) are described in detail below.

The relationship between iron overload and diabetes is clear in the current research, and the main conclusions are two points: 1. iron overload can lead to insulin resistance; 2. iron overload can reduce islet beta cell function and decrease insulin secretion.

The nature of insulin resistance has not yet been elucidated, and therefore, elimination of insulin resistance is only a concept clinically. The mechanism of the blood sugar reducing effect of the insulin is as follows: it binds first to cell membrane receptors and then transports glucose from the extracellular compartment to the intracellular compartment under the influence of glucose tolerance factors, which is the core function of insulin. In this process, glucose tolerance factor plays a crucial role, and the active center of the substance is trivalent chromium ion, so that the trivalent chromium ion in blood of diabetics is always reduced, and people adopt various methods to supplement chromium, but the clinical effect is very little. The original trivalent chromium ion and the trivalent iron ion are used in blood as the same transport protein, namely transferrin, under normal conditions, the saturation of transferrin does not exceed 35%, when iron is overloaded, the saturation of transferrin is increased, even exceeds 50%, so that the transport of the trivalent chromium ion is difficult, the activity of glucose tolerance is influenced, and even if the structure of insulin is normal, the generated amount is large, the function cannot be normally performed. If the problem of iron overload is not solved, the additional supplement of trivalent chromium is not good for, because the absorption of chromium ions can be influenced by the difficulty in running, which is the essential reason for insulin resistance.

The oxygen stress response caused by iron overload can severely impair islet function and reduce the amount of insulin produced. If it is prolonged, type II diabetes spontaneously transforms into type I diabetes. Therefore, iron removal is the fundamental method for solving the problem of diabetes, and besides, the iron removal can only be controlled and delayed, and is accompanied with lifetime medication, and has large side effects.

The clinical test is as follows:

the applicant selects a plurality of diabetes patients who are injected with insulin but have high blood sugar values, then detects the serum ferritin values, and selects 10 patients with excessive serum ferritin from the serum ferritin values. 2, 4-dihydroxy benzoic acid capsules (food grade) were taken 3 times a day, 2 capsules (0.5 g/capsule) each time. Three months later, serum ferritin, glycated hemoglobin, and blood glucose levels were examined as controls (see Table 1).

The normal values of serum ferritin men are: 15-130 ng/ml; the female normal values were: 15-80 ng/ml.

TABLE 1 comparison of serum ferritin, glycated hemoglobin and blood glucose levels before and after administration

As can be seen from Table 1, for 5 men, four of them recovered to normal values and the other decreased by 45.3% after re-administration of serum ferritin concentration; the blood sugar value is also obviously reduced after the two indexes of fasting and after meal are taken; glycated hemoglobin also decreases significantly to normal levels after administration. For 5 women, after the serum ferritin concentration is taken again, three women recovered to normal values, and one of the other two women also reduced 56.9% to almost normal level, and the other reduced 48.5%; the blood sugar value is also obviously reduced after the two indexes of fasting and after meal are taken; glycated hemoglobin also decreases significantly to normal levels after administration.

The applicant selects 10 diabetic patients with less than two years of medical history, no complication and mild disease, and takes 2, 4-dihydroxy benzoic acid capsules (food grade) 3 times a day with 2 capsules (0.5 g/capsule) each time. After taking for 3 months, other hypoglycemic drugs are completely stopped to take, and blood sugar values are detected and compared. The results are shown in Table 2 (only 7 patients were on demand for 3 months).

TABLE 2 comparison of blood sugar level before and after administration (unit: nmol/l)

As can be seen from Table 2, the blood glucose values of at least 5 of 7 patients after taking the medicine for 3 months are obviously reduced in the two indexes of fasting and postprandial; the other two patients also had a decline, but the effect was not very significant.

In this regard, clinical trials conclude that:

(1) the compound can quickly reduce serum ferritin and solve the problem of iron overload;

(2) the compound has the functions of reducing blood sugar and stabilizing blood sugar;

(3) the compound does not have any adverse reaction in the experimental dosage.

Secondly, animal experiments are as follows:

the applicant entrusts Shanxi university of medicine to perform mouse (diabetes model mouse) blood sugar reduction experiment on the compound:

1. grouping experiments:

1.1, WT control group: 8C 57 mice;

1.2, diabetes model mice: db/db mice 10;

1.3, low dose group: db/db mice, 62.5 mg/kg/time;

1.4, high dose group: db/db mice 10, 125 mg/kg/time.

2. Mode of administration

Gavage was performed 2 times per day at 9 am and 4 pm each day.

The actual dose was administered once per week based on the actual body weight of each group of mice in the week.

8C 57 mice on 29 days of 5 months, and 30 db/db diabetic mice arrive at the laboratory, all the vital signs of the animals are normal through preliminary detection, and the experimental requirements are met. Animals were housed in the SPF-grade laboratory animal center, university of Shanxi medical sciences. One week after acclimatization, the experiment was started. Basal indicators were measured for each group of animals and dosing intervention was initiated on day 16/6.

3. Results of the experiment

And counting by taking the initial value of each measurement index before administration as a reference base number.

As shown in fig. 1, in fasting blood glucose concentration, the high dose group was almost equal to the control group, even lower than the control group, and the low dose group was between the model group and the control group, and it can be seen that the better the blood glucose lowering effect, the closer to the normal value, the higher the blood glucose lowering effect.

As shown in fig. 2, the serum ferritin concentration was increased and then decreased in the high dose group, and finally was significantly lower than that in the control group, and the low dose group was substantially equal to that in the control group.

As shown in FIG. 3, the glycated hemoglobin concentration was found to be stable in the high dose group, which was significantly lower than that in the low dose group, and almost equal to that in the control group.

4. Conclusion

(1) The compound has the function of reducing serum ferritin, and has dosage effect.

(2) The compound has the effects of reducing fasting blood sugar and postprandial blood sugar, and stabilizing glycosylated hemoglobin.

(3) In the experimental process, the compound has no abnormal reaction in mice and no toxic or side effect in dosage.

Finally, it should be noted that the above embodiments are only for illustrating the technical solutions of the present invention and not for limiting, and although the detailed description is made with reference to the embodiments of the present invention, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, which shall all fall within the protection scope of the claims of the present invention.

Claims

1. Application of 2, 4-dihydroxybenzoic acid in preparing medicine for reducing blood sugar is provided.