JPH0351686B2 - - Google Patents

Description

[Detailed Description of the Invention] (Industrial Application Field) The present invention is based on the formula () The present invention relates to a pharmaceutical composition for regulating mitochondria and conserving oxygen consumption, containing as an active ingredient a compound having the following. (Prior art) The active ingredient of the present invention corresponding to the above formula () is:
Described in British Patent No. 1360426. The pharmaceutical compositions of the present invention contain as active ingredient a compound of formula () together with an inert solid or liquid pharmaceutical carrier and optionally further additives. The compounds of formula () have mitochondria-modulating and oxygen consumption-sparing properties that are useful in the treatment of humans, eg, diseased individuals. The pharmaceutical activity of compounds of formula () is shown in the Examples. Compounds of formula () have the great advantage of very low toxicity. The favorable effects exhibited on mitochondrial activity allow for dual therapeutic uses. This type of preparation can be used not only to treat hypoxic heart diseases, but also to provide favorable therapeutic effects in the case of bone metabolic disorders. In this connection, reference should be made to some similar literature data:
Today's topic in Bioenergetics, Volume 6, Byrave FL: Mitochondrial calcium
Transport, pp. ^259-318 (1976), p. 300, states: ``Muscle contraction-relaxation [Carafoli E. Recent，Adv.Stud.
Card.Struct.Metab., 5 , 151-163, 1975] and bone formation [Lehninger AL, J.Biochem.,
119, 129-138, 1970]. According to Shapiro et al. [Schapiro JM and Greenspan JS,
Calc. Tiss. Res., 3 , 100, 1969], mitochondria play an important role in the mineralization of living organisms. ”. The compositions of the invention are prepared by methods known in the pharmaceutical industry by mixing the active ingredient with a suitable solid or liquid carrier and formulating the mixture in a form suitable for direct pharmaceutical use. I can do it. The pharmaceutical composition of the present invention can be used in a form suitable for oral administration. The compositions of the invention may contain the usual carriers such as talc, starch, gelatin, water, polyethylene glycol, magnesium stearate, calcium carbonate, and the like. The compositions of the invention may also contain customary additives, such as wetting agents, buffers, etc. Additionally, pharmaceutically active substances may also be included in the composition. Preferred unit dosage forms are tablets and capsules. To manufacture these,
The crushed and sifted ingredients are mixed and the homogeneous mixture is packed into capsules or made into tablets. The daily dosage of 7-isopropoxy-isoflavone can be from about 600 mg to about 1200 mg orally. The content of active ingredient in the compositions may also vary within a wide range. Advantageous dosage units in tablets, capsules may have an active ingredient content of about 100 to about 300 mg. Particularly advantageous are dosage units of about 200 mg. Activities described in the present invention: (a) increase in ATP content, (b) glycerophosphate dehydrogenase activity, and (c) conservation of oxygen consumption are closely related to calcium content in mitochondria and bone formation (e.g., biochemistry,
Vol.12, No.7, 1438-1444, 1973; Biochem.I
(1970) 119, 129-138; Calc.Tiss.Res., 3,
100−102 (1969); Current Topics in
(Bioenergetics, Vol. 6, 303, 1977) The present invention will be explained in more detail with reference to Examples.
However, the scope of the claims shall not be limited by these examples. EXAMPLE 1 Capsules are prepared by methods known in the pharmaceutical industry by mixing the following ingredients in the proportions indicated and filling the well-mixed mixture into capsules. 7-Isopropoxy-isoflavones 200 mg Colloidal silicic acid 20 mg Talc 20 mg Magnesium stearate 40 mg Potato starch 80 mg Lactose 80 mg Total weight 440 mg Example 2 Tablets of the following composition are prepared by methods of the pharmaceutical industry. 7-isopropoxy-isoflavone 200mg Potato starch 36mg Lactose 60mg PVP/Polyvinylpyrrolidone 13mg Esmaspreng (Dinamit Nobel A G
Witten, DBR.) 30mg Magnesium stearate 4mg Talc 7mg Total weight 350mg 200Kg of 7-isopropoxy-isoflavone
Knead with 36Kg potato starch and 60Kg lactose,
Inject 13Kg of polyvinylpyrrolidone. The wet mass is granulated and 30 Kg Esmaspreng, 4 Kg magnesium stearate and 7 Kg talc are added. The coated granules are dried and compressed into tablets in a conventional manner. The characteristics of the tablet are as follows. Diameter 10mm Average weight 350mg±5% Disintegration time within 15 minutes Abrasion strength 3% or less Example 3 Prepare a suppository with the following composition. 7-isopropoxy-isoflavone 0.30g Witepsol W (Dynamit Nobel A G
Witten D B R) 200g Total weight 2.13g The above Witepsol W suppository base is melted and added to a container for forming eyelashes through a cotton filter. The sifted active ingredient is carefully wiped off with a small amount of the above melt cooled to 40-45° C. and added to the homatsu-forming container. Continue stirring until completely cooled. Suppositories start from 34
Mold at a temperature of 35 degrees C. Freeze at -5 to -8 degrees Celsius. This oral composition exhibits excellent stability and can be stored at room temperature for 5 years in suitable packaging, protected from light. Pharmaceutical and Biochemical PropertiesFunctional and Oxygen Consumption Tests [Table] Male rats of the same strain, body weight and age are used for the experiments. When examining O ₂ consumption during rest in male rats, we found that
Ipriflavone 1/3 mg/100 body weight/day orally 4
After a week of treatment, _O2 consumption is significantly reduced compared to their starting values and untreated controls (P<0.001 for the former, P<0.01 for the latter). [Table] The experimental group was administered 3.3 mg/Kg body weight of ipriflavone in the diet for 6 weeks. The load during swimming was 4 g/100 g body weight, the water temperature was 29 degrees C, and the average weight of the animals was 180 g. Rats are allowed to swim in glass baths in water tanks in an oxygen environment. Absorb the carbon dioxide gas produced. P<0.05. During the experiment, it seemed necessary to exclude the possibility of decreased oxygen consumption due to decreased thyroid function.
the State Institute of Public Hygiene
Tests at the Iodine Isotope Laboratory found that ipriflavone had no effect on the thyroid gland. Studies on liver mitochondria The effect of ipriflavone on oxidative processes at the mitochondrial level was investigated at the Institute of Biochemistry, Faculty of Medicine, Semmelweis University, Budapest. Study on Swelling of Hepatic Mitochondria The degree of swelling during the experiment was determined from the decrease in the absorbance value of the mitochondrial suspension at 520 nm using a Specord UV-VIS spectrophotometer. Increased hepatic mitochondrial swelling was observed as a result of ipriflavone treatment (Tables 3 and 4). This increase is particularly pronounced with thyroxine-induced swelling in the presence of 3-hydroxybutyrate. It can therefore be concluded that ipriflavone provides favorable conditions for the substrate to penetrate into the hepatocyte mitochondria. [Table] Control: Normal feed Treatment: Added ipriflavone at a rate of 5 g/q (quarts), 6 weeks, P < 0.01 [Table] Glycerophosphate activity in rat liver mitochondria From Table 5, ipriflavone has an effect on glycerophosphate in liver mitochondria. It is clear that it increases acid activity. The increase in glycerophosphate activity is believed to be due to increased activity of the circuit system as a result of ipriflavone administration, and the reduced NAD hydrogen generated in the cytoplasm can be oxidized to a greater extent in the mitochondria. [Table] Study on oxidation of rabbit liver mitochondria Mitochondria were prepared by the method of Schneider et al. The animals were fed normal feed alone or ipriflavone-added feed, and examined after 6 weeks. Oxygen consumption of isolated liver mitochondria is measured with a Clark type oxygen electrode (Radiometer) in the presence of 10mM 3-oxybutyrate. (Table 6) [Table] PCP is pentachlorophenol. The composition of the incubation medium in a volume of 4 c.c. is: 3 to 6 mg of mitochondrial protein;
0.225M sucrose, 10μM potassium phosphate buffer, PH
7.4, 5 μM MgCl ₂ , 20 μM Tris, PH7.4. 200μMADP for measurement in state 3. Based on oxygen consumption, the value of Qo ₂ (μlO ₂ /mg protein/hour) that characterizes the substrate in question is Chance.
Calculate according to the method. State 3 is the unregulated or activated state with excess ADP, and state 4 is the regulated state without ADP. To measure the maximum oxidation capacity of mitochondria, 3.
Oxygen consumption is also determined in the presence of 10 ^-6 and 5.10 ^-6 mol of pentachlorophenol. The mitochondrial Qo ₂ value in the presence of glutamate + maleate or hydroxyl butyrate is
It decreases in state 4 (adjustment state) and increases in state 3 (activation state). Thus, mitochondrial respiratory control values from ipriflavone-treated animals are higher than those of untreated controls. Tests on mitochondrial enzymes Hungary, KeResteto, the National
Conducted at Therapeutic Institute. Absorption spectrum of cytochrome enzyme used to assay the effect of ipriflavone on oxidation processes in mitochondria
(M/30) in phosphate buffer. Test at concentrations of 1.0 mg/cc, 0.5 mg/cc and 0.25 mg/cc. Measure absorption at wavelengths from 510 to 570 nm at 5 nm intervals in a 1 cm cube using SPectromom361. Prepare a solution of ipriflavone using Triton X100 and phosphate buffer simultaneously. Use Triton 0.05c.c. for enzyme solution 3c.c. Since 5.0 mg and 2.5 mg of substance are used, the ratio of enzyme to test substance is 1:1 with an enzyme concentration of 1 mg/cc.
1.63, and 3 c.c. of enzyme solution contains 3 mg of enzyme protein. At 2.5 mg, the ratio is 1:0.83.
At a concentration of 0.5mg/cc the ratio is 1:3.25 and 1:1
It becomes 1.63. On the other hand, the values for 0.25 mg/cc are 1:6.50 and 1:3.25. Since the reaction tends to appear with some delay, changes in the absorption spectrum were observed over several days. [Table] [Table] The following can be seen from the results in Table 7. a Increased enzyme activity in liver tissue with 5 mg ipriflavone. About each mg of cytochrome C enzyme protein
1.63mg of compound reacted. b Enzyme activity increased by approximately 10% with ipriflavone. c At 2.5 mg of ipriflavone, the activity of cytochrome oxidase remains unchanged when 1 mg of cytochrome C enzyme protein reacts with 0.82 mg of the compound. d The oxidation of reduced cytochrome C is promoted as an effect of the compound (ipriflavone). Summary: Ipriflavone was found to be an electron acceptor in the terminal part of the respiratory system. The decrease in the absorption maximum at 550 nm indicates that the reduced enzyme cytochrome C is reduced by the effect of the compound. Similarly, manganese-citrate-diethylparaphenylenediamine complexes are affected. This is an artificial electron transport system based on cytochromes. The compound increases the color intensity of the composite measured at 550 nm. This effect corresponds to weak oxidation. Ipriflavone increases cytochrome oxidase activity in liver tissue by 10%. This effect changes depending on the amount used. Determination of acid-labile phosphates in rabbit striated muscle. Performed at the Institute of Biochemistry, Faculty of Medicine, Semmelweis University, Budapest. The acid-labile phosphate content of muscles and, potentially, their ATP-producing capacity.
increases as a result of ipriflavone treatment. The ability to generate high-energy phosphates was investigated by measuring acid-labile phosphates. [Table] Effect of ipriflavone on anaerobic glycolysis Conducted at the Institute of Biochemistry, Semmelweis University, Budapest An increase in the glycolytic capacity of muscles with tonic spasms is advantageous in terms of muscle function. Fast for 24 hours and measure the presence of endogenous substrate only. The lactate content of the blood and the production of lactate in the semimembrane muscles (semimembrane, tonic muscle) are measured, respectively. Lactic acid is measured using an enzymatic method. HU Bergmeyer,
According to Methods of Enzyme Analysis. The blood lactate content of rabbits fed ipriflavone-containing diets was significantly higher than that of normal controls. Lactic acid production in the tonic muscles of these animals is also high. [Table] [Table] Muscle and liver glycogen content tested in rabbits and rats [Table] In these experiments, rats were fasted for 12 hours before fructose loading. Process the rabbits after 6 hours. Remarkably, the average muscle glycogen content of ipriflavone-treated animals remains unchanged, and liver glycogen content increases by more than 40% on average. [Table] Expensive.
These results in rats indicate that ipriflavone treatment does not change muscle glycogen content but increases hepatic glycogen storage capacity. The following conclusions can be drawn from the above results. In isolated, intact mitochondria, administration of ipriflavone affects NAD-dependent substrate oxidation. Decreases oxidizing power in the non-activated state and increases oxidation in the activated state. This alone indicates that ipriflavone administration improves oxidation efficiency. Even if the mechanism of action is unavoidably descriptive, it is probably due to the strengthening of the circuit mechanism.
This is concluded from the swelling of the mitochondria, which increases energy expenditure. It can also be seen from an increase in glycerophosphate activity. These two phenomena, increased hydrogen transport in the cytoplasm and increased mitochondrial oxidative capacity, indicate an increased economy of mitochondrial energy conversion. Increased tonic muscle glycolysis, on the other hand, promotes anaerobic energy production and enhances the oxygen-sparing effect. Therapeutic Effect 1 Antianginal Effect Test No. Tested on 10 patients in total. Check the ECG to confirm that an angina attack is occurring. That is, ST suppression and T wave negativity. In four cases,
Coronalography also showed angina. Two weeks of observation confirms the patient's request for Nitromint. In the second week, Nitromint requirements for patients receiving 7-isopropoxy-isoflavone treatment are half of those given placebo.
Here, the isoflavone dose was 600 mg/day. When patients treated with 7-isopropoxy-isoflavone had a reduction in angina attacks, they were given a beta-receptor blocker and Rigedal (isosorbitone dinitrate), respectively, which further reduced the number of attacks. When a new placebo was given in place of 7-isopropoxy-isoflavone, the number of attacks increased. The number of attacks during the second week of treatment was as follows. [Table] Study No. Experimental design: A total of 15 angina patients were investigated. The experiment consisted of a drug-free period, followed by a placebo period, and then a drug-free period. 200 mg of 7-isopropoxy-isoflavone active substance in tablet form was then administered three times daily. The treatment period is 4 weeks. [Table] Number

Claims (1)

[Scope of Claims] 1. A pharmaceutical composition for increasing ATP content, increasing glycerophosphate dehydrogenase, and saving oxygen consumption in mitochondria, which is characterized by containing 7-isopropoxy-isoflavone as an active ingredient. 2. The pharmaceutical composition according to claim 1, which is used for treating heart failure and lung failure. 3. Pharmaceutical composition according to claim 1 or 2, in the form of a tablet or capsule, containing 100 to 300 mg of 7-isopropoxy-isoflavone. 4. The pharmaceutical composition according to any one of claims 1 to 3, wherein the daily dosage is about 600 to 1200 mg.