WO2008037418A1 - Green tea medicinal extract, especially for use in the prophylaxis and treatment of diseases of the central nervous system or metabolic diseases, its use for producing a drug and method for producing the same - Google Patents

Abstract

A green tea extract, especially for use in the prophylaxis and treatment of neurological and psychiatric diseases of the central nervous system, especially of mental concentration disorders, pains, depression and dementia, or for use in the prophylaxis and treatment of metabolic diseases such as pancreatic diabetes, by increasing the glucose absorption in the case of hypo-insulinism or insulin resistance, has an increased theanine and theogallin content and is decaffeinated.

Description

 GREEN TEA MEDICINAL EXTRACT, USE THEREOF

PROPHYLAXIS AND TREATMENT OF DISEASES OF THE CENTRAL NERVOUS SYSTEM OR DIABETES AND METHODS FOR

THEIR PRODUCTION

The invention relates to a green tea drug extract for the prophylaxis and treatment of neurological and psychiatric disorders of the central nervous system, in particular mental concentration performance disorders, pain, depression and dementia, its use for the manufacture of a medicament and a method for producing such a green tea - Medicinal extract. In addition, use of this green tea drug extract has also been recognized for the prophylaxis and treatment of metabolic diseases such as diabetes mellitus as a field of application.

It can be stated in the prior art that theanine as the pure substance is described on the one hand as soothing (see JP 2005 289 948 A1, JP 2005 232 045 A1 or JP 901 2454 A1) and on the other hand as stimulating properties (see JP 9100230 A1). This is especially true for theanine mixed with caffeine, arginine, rice bran extract (see JP 2002 3220 53 Al, KR 10 2005 121 535 Al or US 2005 0020 627 Al).

DE 10 106 216 A1 discloses a green tea extract which is decaffeinated, has low tanning properties but is enriched in terms of L-theanine. It is made by first decaffeinating green tea leaves, green tea blanc or green tea powder and then extracting it in water. The extract is then contacted with polystyrene and then concentrated. The extract is used as an ingredient in beverages or beverage concentrates. EP 10 57 483 A1 discloses a pharmaceutically active composition for the treatment of various symptoms, such as obesity, premenstrual symptoms or hypersensitivity, which contains theanine.

From EP 10 74 252 B1 a composition containing theanine for the suppression of behavioral problems in domestic animals is known.

The currently used drugs for the treatment of diseases of the central nervous system, in particular dementia, depression and concentration performance disorders have a broad spectrum of side effects. There is therefore a very great need for a prophylaxis or an improved drug with a good therapeutic efficacy at the lowest possible rate of side effects. In particular, this also applies and especially for prophylaxis.

Another area relevant here is in the field of metabolic diseases. By way of background, it should be noted that false, e.g. Unilateral diet and lack of exercise in the general view are seen as a major cause of obesity, diabetes and other metabolic diseases. This is reflected in an increase in the prevalence of diabetes II disease in the population.

Decreased insulin sensitivity of the cells or, under certain circumstances, reduced insulin production ultimately lead to higher sugar levels being measured in the blood of the persons affected. There is therefore a need for medicaments which, in the sense of a prophylaxis or therapy, can help to keep the blood sugar level of endangered persons in a physiological range.

The patent literature already describes various plants and plant extracts in this context, e.g. in the Korean patent application KR 1020030056987A a composition with green tea catechin as an active ingredient for the prophylaxis and therapy of cardiovascular diseases associated with diabetes.

On the one hand, the invention is based on the object of specifying a medicament for the treatment of diseases of the central nervous system and a process for its production which, based on green tea as natural starting material, has a good therapeutic activity at a low side-effect rate and is easy to prepare and produce Drug is configurable. On the other hand, the object is to provide a medicament for the prophylaxis and treatment of metabolic diseases, such as diabetes mellitus.

This object is achieved by a green tea drug extract (hereinafter referred to briefly as SGTE = "special green tea extract") of the type specified in claim 1 in terms of product technology. Thus, in addition to an enriched portion of theanine, the SGTE also contains an enriched portion of theogallin and is decaffeinated.

At least six percent and at least three percent have proved to be the preferred degrees of enrichment for theanine and theogallin, with the caffeine content being at most 0.03 percent. A biological or pharmacological effect of said SGTE has not yet been described. On the basis of the previously known literature, an effect of the SGTE on pathological disorders of the central nervous system, in particular of neurological and psychiatric diseases as well as concentration performance disorders, had not been expected. Only the application of the completely "bias" -free examination in the rat telestereo-EEG yielded in the context of experiments with other purposes the indication of the existence of such a central nervous effect. The effect was further substantiated by in vitro experiments on the hippocampal slice in direct interaction between the drug and its target organ, the brain.

The SGTE according to the invention can be used for the preparation of a corresponding medicament for the prophylaxis and treatment of neurological and psychiatric disorders of the central nervous system, in particular of mental concentration performance disorders, pain, depression and dementia. In this context, also disorders with such a drug are treatable, as they are observed in children in the context of attention deficit hyperactivity syndrome.

Surprisingly, studies such as a screening of various plant extracts on the subject of "blood sugar regulation, antidiabetic properties of plant extracts" have shown that the inventive green tea extract also has anti-diabetic properties Theogallin- enriched green tea extract, decaffeinated, is able to significantly increase the glucose uptake by differentiated adipocytes. Further information on the above indications can be found in the description of the embodiment.

In terms of process technology, the invention provides a process for producing a SGTE, which comprises the following process steps: a) aqueous extraction of green tea leaves, b) liquid-liquid extraction of the aqueous green tea extract obtained in step a), c) absorptive enrichment d) elution of the absorber, e) concentration of the eluate to a decaffeinated, enriched with theanine and theogallin green tea extract having a dry matter content of preferably at least 50%.

On the basis of these basic process steps specified in the further dependent claims, the abovementioned proportions of theanine, theogallin and caffeine can be achieved.

Further features, details and advantages of the invention will become apparent from the following description in which the nature of the SGTE, its mechanisms of action and the attempts made therewith are explained in more detail with reference to the attached diagrams. Show it:

Fig. 1 is a comparative diagram showing the effect of SGTE on EEG frequencies during the first hour after drug administration (mean of n = 4 animals); 2 is a graph showing the concentration-dependent increase in the amplitude of the population spike (activity of the pyramidal cells) in the presence of SGTE in the hippocampal slice preparation in vitro (average values of 4 sections with mean error of the mean value);

Fig. 3 is a graph showing the behavior of amplitude behavior of the population spike in the hippocampal slice after theta-burst stimulation in the presence of SGTE, and

Fig. 4 is a diagram showing the glucose uptake of test cells - human, differentiated adipocytes - depending on the concentration of added green tea extract.

Green tea is used as raw material for the production of the SGTE according to the invention. This is subjected to an aqueous extraction in a first step - step a) of the production process.

The aqueous green tea extract thus obtained is subjected to a liquid-liquid extraction step b) in a second step by means of ethyl acetate. Here, caffeine and a part of the polyphenols are removed, which leads to an accumulation of the desired substances theanine and Theogal- Hn.

A further, adsorptive enrichment takes place in a third process stage - step c) - via an adsorber filled with functionalized divinylbenzene. The resulting aqueous run should be discarded. The elution - step d) - is carried out with demineralized water to give two separate fractions. In particular, the second part of tion is rich in theanine, theogallin and green tea amino acids.

In a further step - step e) - the relevant fraction is concentrated to a dry matter content of about 50% and can already be used as such. The concentrated theanine and theogallin-enriched green tea extract thus contained can be spray-dried in a further process step.

The SGTE produced as described above has essentially the following composition based on dry substance:

Water content 3.2%

Ash content 24.1%

Total Amino Acids 3.52%

Caffeine <0.01%

L-Theanine 9.71%

Theogallin 4.65%

The rest is made up of a variety of fabrics. Analyzes have revealed residual proportions of tannins of 2.5% and carbohydrates, acids and acid derivatives of 52.3%.

Surprisingly, it was surprisingly found that the preparation had an effect in the brain during treatment of alterations of rat EEG frequencies after administration of SGTE and direct interaction of SGTE and brain matter in the rat hippocampal slice preparation of neurological and psychiatric Diseases of the brain, in particular dementia, depression, pain and concentration disorders is useful.

SGTE surprisingly shows in the tele-stereo-EEG model in rats changes in EEG frequencies as measured after administration of classical drugs for the treatment of dementias (example galanthamine), depression (example paroxetine) as well as mental dysfunctions (eg amphetamine). The changes, as occurred after administration of classical drugs and after administration of a dose of SGTE (75 mg / kg) and measured according to test method 1, are shown in FIG.

Based on the finding that SGTE causes the same characteristic changes in EEG frequencies as commonly used in the treatment of dementia, depression and impaired concentration, it can be concluded that SGTE is effective in treating the same indications. Dimpfel has shown that more than 40 reference substances can be used to treat similar EEG changes in more than 40 reference substances (Dimpfel W .: "Preclinical data base of pharmaco-specific rat EEG fingerprints (Tele-Stereo-EEG) "in Eur J Med Res (2003) 8: 199-207).

In addition, studies on the hippocampal cut preparation model were performed in vitro. In these studies it was surprisingly found that SGTE causes both an increase in pyramidal cell activity after single irritation and an increase in long-term potentiation (see FIGS. 2 and 3). This phenomenon has been implicated in other stimulant and antidemental drugs such as memantine in the literature (Dimpfel W .: "Effects of Memantine on Sym- netic Transmission in the Hippocampus in Vitro" in Arzneim.-Forsch / Drug Res (1995) 45: 1-5).

The SGTE according to the invention as active ingredient-containing medicaments are preferably administered orally.

For administration, the pharmaceutical composition of the invention containing SGTE as an active ingredient, e.g. in the form of powders, tablets, capsules, pills, dragees, granules, pellets, solutions, syrups, dispersions, the active ingredient optionally being able to be combined with pharmaceutically acceptable excipients and carriers.

The pharmaceutical compositions of the invention containing SGTE as an active ingredient will normally be prepared according to conventional methods and administered in a pharmaceutically acceptable form.

For example, the solid oral forms together with the active ingredient may include diluents, eg lactose, dextrose, sucrose, cellulose, maize starch or potato starch; Lubricants, for example silicate, talc, stearic acid, magnesium or calcium stearate and / or polyethylene glycols; Binders, for example starches, gum arabic, gelatin, methylcellulose, carboxymethylcellulose or polyvinylpyrrolidone: anesthetic, for example starch, alginic acid, alginates or sodium starch glycolates, intumescent mixtures; dyes; sweeteners; Wetting agents such as lecithin, polysorbates, lauryl sulfates and generally non-toxic and pharmacologically inactive substances used in pharmaceutical formulations. The pharmaceutical preparations can be prepared in a known manner, for example by means of mixing, granulation, tableting, sugar coating or coating coating methods.

The liquid dispersions for oral administration may be e.g. Syrups, emulsions and suspensions.

Syrups may be used as carriers e.g. Sucrose or sucrose with glycerol and / or mannitol and / or sorbitol.

The suspensions and the emulsions may be used as carriers e.g. a natural resin, agar, sodium alginate, pectin, methyl cellulose, carboxymethyl cellulose or polyvinyl alcohol.

The dosage unit of the drug may contain, for example, from 1720 to 4300 mg, preferably from 1935 to 2580 mg of SGTE per daily dose in the case of oral dosage forms. The daily dose can be administered, for example, in 1 to 3 single doses, preferably in two single doses, daily.

Test Method 1 (Tele-Stereo EEG)

Changes in EEG frequencies were determined after administration of saline (control) or orally SGTE 75 mg / kg.

The investigations were carried out analogously to the method described by W. Dimpfel (Dimpfel W .: "Preclinical data base of pharmaco-specifc rat EEG fingerprints (tele-stereo-EEG)" in Eur J Med Res (2003) 8: 199-207) as follows carried out: Four male adult Fischer-344 rats (day-night converted) were implanted with 4 bipolar concentric electrodes at the age of 6 months along with a microplug on a common baseplate. The plug was used to record a 4-channel transmitter for the telemetric transmission of the field potentials derived from the frontal cortex, hippocampus, stria- tum and formatio reticularis. The signals were subjected to Fast Fourier Transformation in real time on a computer system ("EEG Analysis" software, OS Science operating system, "LabTeam" lab computer from MediSyst, Linden, DE), and the power density spectra were each averaged over 60 minutes. The subdivision of the spectra into 6 different frequency ranges allowed the detection of pharmaco-specific changes with respect to the pre-values measured before each application within these frequency bands.

For the application protocol of the substance, it should be noted that the substance was administered orally 45 minutes after the start of the measurements (pre-value). Five minutes later, measurements were restarted, continuously analyzed for at least the next 5 hours, and pooled into 60-minute periods. The test substance was applied at a dose of 75 mg / kg (SGTE).

The statistical comparison of the experiments to the results, which was measured after administration of saline solution, was carried out with the aid of a multivariant analysis according to Ahrens and Läuter (see H. Ahrens, J. Läuter: "Multidimensional Analysis of Variance" (1974), Akademie Verlag, Berlin) on the basis of the changes within the individual frequency bands in all brain regions as variables. The administration of saline solution hardly led to any changes in the electrical activity (μV / Ω) compared to the pre-phase values.

Administration of SGTE resulted in marked changes in power density in the frontal cortex (see Fig. 1). Note the similarity to drugs used to treat depression, dementia, and concentration disorders.

Test Method 2 (Long-term potentiation in hippocampal slice preparation in vitro)

Effects of SGTE on the excitability of the pyramidal cells were determined in the rat hippocampal preparation. The analysis of the effects took place on two levels. First of all, the normal excitability was examined by the application of individual stimuli and then the effects on long-term potentiation were measured after a so-called "theta-burst stimulation".

Nine adult male CD rats were used to perform these studies. Isolation of the hippocampus was performed on phosphate-buffered saline (NaCl: 124 mM, KCl: 5 mM, CaCl 2 mM, MgSO 4: 2 mM, NaH 2 PO 4: 1, 25 mM, NaHCO 3: 26 mM: glucose: 10) on ether-anaesthetized and subsequently exsanguinated animals mM; Control Solution: Artificial Cerebral Spinal Fluid (ACSF) The SGTE was supplied by Plantextrakt, Vestenbergsgreuth. The middle part of the hippocampus was then blocked with the aid of a rapid adhesive and cut with a vibratome into 400 μm thick slices. These sections were stored for at least one hour before the start of the experiment (see SJ Schiff, GG Somjen: "The effects of temperature on synaptic transmission in hippocampal tissue slices" in Brain Research (1985), 345: 279-284) in one with carbogen bubbled through incubation chamber.

The experiment itself was performed in a so-called "base unit" with "Haas Top" (Medical Systems Corp., USA) at 35 degrees C. The hippocampus cut superfused with the aid of peristaltic pumps (Infusomat B. Braun Melsungen AG) was on a piece of gauze Introduced Carbogen maintained the required oxygen supply to the solution, and the flow rate was 200 ml / h.

Stimulation of the CA2 region (Schaffer collaterals) was carried out using a stimulus generator (Laboratory Lab Lab team from MediSyst GmbH, Linden, DE) via an isolation unit using a bipolar concentric steel electrode (Rhodes Medical Systems, USA). The pulse width was 200 μs, the current constant 200 μA. The stimulus generator triggered four individual irritations at intervals of 20 seconds each, which on average evoked 4 population spikes. The response was derived in the CA3 region. The system averaged the stimulus response of the 4 spike amplitudes.

The effect of SGTE on the evoked potential was investigated after single irritation as well as after induction of long-term potentiation by a short duration (1 s) tetanic stimulus (90 Hz) (see KG Reymann, HK Matthies, U. Frey, VS Voborbyev, H. Matthies: "calcium-induced long-term potentiation in the hippocampal slice. In Brain Bulletin (1986), 17: 291-296), the cut was first superfused with control solution, and after finding a suitable signal and registering this output signal for several time points, the solution was replaced with SGTE Each cut was only used for one experiment, values are given for n = 4 cuts.

The evoked response was derived 10 minutes extracellularly with a pulled glass capillary (Elektrodenpuller, Rhema Labortechnik, DE). The response signal was amplified (Amplifier "LMI", List Electronics, Darmstadt, Germany) with a laboratory computer system "Labteam" (software NeuroTool, MediSyst GmbH, Linden DE) analog-digitally converted (resolution 12 bits) and evaluated. The amplitude value of the population spike (SAP = sum action potential, popspike) was determined as the starting point, which was the average of the last three amplitude levels measured during superfusion with the ACSF solution (initial value) Superfusion with test substance with the same procedure In subsequent experiments a short-term tetanic stimulation (Theta-burst stimulus ^ TBS) was used to induce long-term potentiation resulting in amplitude changes in% of this initial value (n = 4) Values in the presence of the test substance are in % Reduction of this reference value.

The results of these studies are shown in FIGS. 2 and 3. The first experimental set-up shows a concentration-dependent increase in the amplitude of the population spike in the presence of 0 to 13 mg / 1 SGTE in single challenge (see Fig. 2). The triggering of the long-term Addition by TBS was also concentration-dependent at the same concentration (see Fig. 3), as known from experiments with anti-dementia drugs. From this result it can be concluded that, as can be seen from the EEG examinations, SGTE has a positive effect on depression and dementia.

The SGTE prepared as described above is also suitable for pharmacological use for the prophylaxis or therapy of diabetes diseases, such as diabetes mellitus, as the following studies show.

For a glucose uptake test, 3T3-L1 cells (murine

Fibroblasts) were differentiated into adipocytes in three differentiation steps over 8-9 days with insulin (lμg / ml), dexamethasone (40μM) and IBMX (500μM). In the second differentiation step, the cells (60,000 cells / well) were seeded in cell culture plates. For the assay, the cells were incubated in serum-free medium for 3 hrs. After preincubation of the cells with a 0.16 μM insulin solution and the SGTE test extracts for 25 min at 37 ° C., the assay was started with 2-deoxy-D- [1- ³ H] -glucose (0.1 lμCi / ml) and incubated for 25 min incubated at room temperature. The amount of the captured by the cells 2-deoxy-D- [l- ³ H] glucose was determined after various washing steps and lysis of the cells using a scintillation counter. The measurements were carried out in a Tri-Carb 1900 TR Liquid Scintillation Counter (Packard, USA). Zero controls (t (0)) were performed without addition of insulin. The negative control used was cytochalasin B (200 μM), an inhibitor of the insulin-stimulated glucose transporter system.

The measured values for the samples were determined from three experiments and are given as mean values with the respective standard deviation. The result of the investigations is shown graphically in FIG. It can be clearly seen that the tested green tea extract more than doubled the glucose uptake in the described incubation time compared to the control. This applies to all three test concentrations used of 3 μg / ml, 100 ng / ml, and 300 pg / ml.

Claims

claims 1. green tea drug extract for the prophylaxis and treatment of neurological and psychiatric disorders of the central nervous system, in particular mental concentration performance disorders, Pain, depression and dementia, or for the prophylaxis and treatment of metabolic diseases, such as diabetes mellitus, by increasing the glucose uptake capacity in insulin deficiency situations or in insulin resistance, with an enriched portion of theanine, characterized in that the extract further enriched Contains portion of theogallin and is decaffeinated. 2. green tea-drug extract according to claim I ₅ characterized by the following proportions based on the dry matter: - theanine at least 6.0% - Theogallin at least 3.0% - Caffeine maximum 0.03%. 3. green tea-drug extract according to claim 1 or 2, characterized by the following proportions based on the dry matter: - water content 2.0% to 5.0% Ash 20.0% to 30.0% - amino acids 2.0% to 10.0% Caffeine less than 0.03%, theanine 6.0% to 40.0% and Theogallin 3.0% to 20.0%. 4. green tea drug extract according to claim 1 or 2, characterized in that the extract additionally pharmaceutically acceptable Contains carrier, auxiliary and / or diluent. 5. green tea drug extract according to any one of the preceding claims, characterized in that it is prepared for oral administration. 6. green tea-drug extract according to one of the preceding claims, characterized by the following composition based on the dry matter: - water content 2.5% to 4.0%, in particular 3.2% Ash from 23,0% to 25,0%, in particular 24,1% Amino acids 3.0% to 4.0%, in particular 3.52% - Caffeine below 0.01 Theanine 9.0% to 10.5%, in particular 9.71%, and Theogallin 4.0% to 5.5%, in particular 4.65%. 7. Use of the green tea drug extract according to any one of the preceding claims for the manufacture of a medicament for the prophylaxis and treatment of neurological and psychiatric disorders of the central nervous system, in particular mental concentration performance disorders, pain, depression and dementia. 8. Use of the green tea drug extract according to one of the preceding claims 1 to 6 for the manufacture of a medicament for prophylaxis and treatment of metabolic diseases, such as diabetes mellitus, by increasing glucose uptake in insulin deficiency situations or insulin resistance. 9. A process for producing a green tea drug extract according to any one of claims 1 to 6, characterized by the following process steps: a) aqueous extraction of green tea leaves, b) liquid-liquid extraction of the obtained in step a) aqueous Green tea-drug extract, c) absorptive enrichment of the extract obtained in step b) in an absorber, d) elution of the absorber, e) concentration of the eluate to a decaffeinated, theanin and theogallin-enriched green tea extract with a dry matter extract. Content of preferably at least 50%. 10. The method according to claim 9, characterized in that the liquid-liquid extraction according to step b) is carried out by means of ethyl acetate. 11. The method according to claim 9 or 10, characterized in that the absorptive enrichment in step c) via an filled with functionalized divinylbenzene absorber. 12. The method according to any one of claims 9 to 1 1, characterized in that the elution of the absorber according to step d) is carried out with demineralized water. 13. The method according to any one of claims 9 to 12, characterized in that in the elution of the absorber according to step d), two separate partial fractions are obtained, of which the second, rich in theanine, theogallin and green tea-specific amino acids fraction fraction the far More process steps is subjected. 14. The method according to any one of claims 9 to 13, characterized in that the green tea-drug extract obtained in step e) is spray-dried.