WO1999051251A1 - Use of extracts of bearberry leaves (arctostaphylos uva ursi), birch leaves (betula), horsetail (equisetum) and stinging nettle (urtica) - Google Patents

Abstract

The invention relates to the use of extracts of bearberry leaves (Arctostaphylos uva ursi), birch leaves (Betula), horsetail (Equisetum) and stinging nettle (Urtica), especially dry extracts from the fresh or dried plants, to produce medicaments for treating or preventing (protecting against) damage caused to organs and tissue by radiation (radioprevention, radioprotection), infection (infection prophylaxis) and chemical action (chemoprevention, chemoprotection), especially damage to the O-MALT system of the small bowel (inflammatory diseases, enteritis regionalis Crohn), the lungs (inflammation, disease), the bronchial tubes and the rhinopharynx (rhino-pharyngo-bronchitis, rhinitis, pharyngitis, sinusitis), the bone marrow (aplasia of the bone marrow), the thymus (dysfunction, aplasia or hypoplasia resulting from damage caused by medicaments or radiation), the liver (atrophy, necrosis), for treating/preventing hepatitis A, B and C, for damage to the pancreas, insufficiency of the exocrine, secretory functions of proteases, esterases, carbohydrases and nucleases and impairment of the carbohydrate metabolism (Langerhans' islands), damage to the kidneys (insufficiency), and generally immunosuppressed conditions, for cellular immunostimulation, for treating leucocytopenia, granulocytopenia, lymphoctopenia, thrombocytopenia, erythrocytopenia (infect anaemia, tumour anaemia i.a.), immunoglobulin deficiency conditions, and for conditions resulting from AIDS, tumours and diseases with a suppressed immune system.

Description

USE OF BEAR GRAPE LEAVES (ARCTOSTAPHYLOS UVA

URSI) -, BIRCH LEAVES (BETULA) -, BROWNWALK HERB (EQUISE-

TUM) AND NETTLE (URTICA) EXTRACTS

The invention relates to the use of bearberry leaves (Arcostaphylos uva ursi, AU), birch leaves (Betula, BP), horsetail herb (Equisetum, EM) and nettle (Urtica, UD) extracts, in particular dry extracts, if appropriate in combination with other phytopharmaceuticals and / or chemically defined pharmaceuticals for the production of pharmaceuticals and orally administrable pharmaceuticals and for use in the context of high-dose phytopharmaceutical therapy.

AU, BP, EM and UD are treated together due to their comparable potential for diuretic (aquaretic) analgesic, antirheumatic, antibacterial, antiviral, antitumoral, hypoglycemic, hypolipidemic, nephrokurative, -protective as well as hepatocurative and protective effects. AU, BP, EM and UD also show clearly pronounced immunostimulating effects (paramunization) in the context of high-dose therapy.

From Römpp Chemielexikon, 9th edition, p. 334 (1989) it can be seen that under the term "Bärentrau" an evergreen heather plant (Arctostaphylos uva-ursi, Ericaceae) with white-red flower clusters and red berries (sand, flour or cranberry ) is understood that grows especially in dry pine forests and mountain heaths and whose leaves contain the hydroquinone glycosides, arbutin and methylarbutin as well as tannins and flavones. The hydroquinone split off from the glycosides in the organism is excreted as a conjugate (with glucuronic or sulfuric acid) and released again in alkaline urine (which can be achieved by taking sodium hydrogen carbonate if necessary), which is the basis for the antibacterial effect of the bearberry. Bearberry leaves (as tea) are diuretic and diuretic, but not as real diuretics.

Arbutin, better known as arbutoside, is a mono-ß-D-glycoside of hydroquinone and occurs in quantities of 5 to 12% during the harvest. The clearly astringent, tart taste of the leaves is due to the high content of tannins.

A combination preparation consisting of a bearberry extract and other plant extracts has been proposed in the prior art. This preparation was used to treat patients with sulfonamide-resistant urinary tract infections. Shortly after taking the preparation, the therapy led to an increase in the number of leukocytes in the blood and to an increased phagocytosis effect of the leukocytes in the urine sediment. However, since a combination preparation consisting of several drug extracts was assumed, it is not possible to assign the leukocyte-stimulating and anti-inflammatory effect to a specific plant.

After overdosing several grape leaf preparations, there is inflammatory irritation of the bladder and urethral mucosa, accompanied by urinary urgency and blood urine, the latter possibly up to the haemorrhagic-purulent stage. The ancient peoples used birch leaf (Betula) species as a remedy.

From Rompp Chemielexikon, 9th edition, p. 434 (1989) it can be seen that birch oils are oil-like distillates obtained from different parts of the birch and are usually differentiated into birch bud oil, birch bark oil, birch tar and birch tar oil.

The use of birch trees in medicine is based on a centuries-old tradition. Since the Middle Ages, the use of birch sap, which is obtained by drilling young birch trunks, has been used for stone disease and jaundice, as well as for hair loss. In traditional medicine, the bark is used as a bath additive for skin diseases. Nowadays, however, the foliar drug is used far more frequently in folk medicine as a diuretic and in gout and rheumatism, which is why most pharmacopoeias for birch leaves have included a monograph. The leaf drug contains a large variety of flavon and flavone compounds as well as a number of other relatively unknown natural substances. The specialties betula aar® and aar®diu coated tablets, which contain native extracts of birch leaves, are commercially available.

Birch leaves consist of whole or cut, dried leaves of various types. The drug contains at least 1.5% flavonoids, calculated as hyperoside, based on the dried drug.

The Betula dry extract is made from shredded birch leaves and water according to a process described for dry extracts in the monograph "Extracts".

From the point of view of a diuretic and anti-inflammatory effect of horsetail herb (Equisetum), the following, in part centuries-old, experience-therapeutic applications are of scientific interest:

diuretic effect

Inflammation of the urinary organs such as nephritis and cystopyelitis, renal colic, kidney stones internal and external wounds, ulcers febrile processes

Bleeding

Field horsetail is used to increase the flow of urine in catarrhs in the kidney and bladder, as a hemostatic agent in the case of excessive menstrual bleeding in the woman, in nasal, pulmonary and gastric bleeding, as an adjuvant in the case of tuberculous diseases, in the case of cracked fingernails and hair loss, in the form of Baths used for female abdominal disorders, rheumatic diseases, gout, poorly healing wounds and ulcers, swelling and broken bones. The large nettle (Urtica dioica L.) has already been found in Neolithic pile dwellings in Switzerland (around 3000 BC), in ships from Kralsund (Norway) dating from the early Iron Age and in the mud of the Roman fort Zugmantel in the Taunus. It is only in later times that information about their use as medicinal, vegetable and fodder plants is found.

The nettle has been attributed to diuretic, menstrual, hemostatic, digestive, cough-relieving, sore and "boil-healing", blood-cleaning and strengthening effects.

Folk medicine used U. dioica and U. urens for dropsy, cough, rash, for hemostasis in the case of lung diseases (tuberculosis), for sleeplessness, as envelopes for swelling, the fresh juice of pressed leaves as a means of stopping pulmonary bleeding as well as hemorrhoidal and excessive Menstrual bleeding, for kidney and liver problems, against kidney stones and with weakened blood formation.

Liquid extracts from nettles were found to be cardiotonic. Favorable diuretic effects with considerable excretion of chlorides and urea were also found. Satisfactory test results were obtained with nettle preparations for arthritis, gout diathesis, joint (muscle) rheumatism and with reduced lactation.

Some authors have achieved stimulation of the pancreas by injecting nettle juice. Favorable hypoglycemic effects in nettle preparations were also found. Nettles have also been used in bronchial asthma.

From Römpp Chemie Lexikon, 10th edition, keyword "nettle" it is known to use nettle extracts against rheumatic complaints or urination disorders. It is also known that nettle extracts on the skin and

The scalp has a hyperemic effect, which is related to the content of nettle poisons Histamine, serotonin and acetylcholine are attributed.

There are only a few phytochemical studies against the diverse therapeutic indications of nettle, which examine the spectrum of ingredients of the Urtica species, in particular Urtica dioica L. and Urtica urens L., the parent plants of the drugs Urticae herba, Uricae folia and Urticae radix. There are currently no indications of which substances have a specific effect.

Urtica dioica L. is rich in minerals (up to 20%), based on the dry matter (DAB 6, Erg.B.). The mineral content is directly related to the nature of the soil and the season. The organic ingredients are characterized by essentially ubiquitous natural substances that are widespread in green plants. Saponins could not be detected, at least when examining Chilean urtica species. Nicotine was only found in Urtica dioica L., but not in Urtica urens L.

In addition to the pentacyclic triterpene derivative oleanolic acid, the substances identified from the root include the well-known sterols 3b-sitosterol, sitosterol-3-O-ß-D-glucoside, (6'-O-palmitoyl) -sitosterol-3-ObD-glucoside , 7a-hydroxysitosterol and 24R-ethyl-5a-cholestan-3b, 6a-diol, 7b-hydroxysitosterol-3-ObD-glucoside and 7a-hydroxysitosterol-3-ObD-glucoside.

Furthermore, two phenylpropanes were identified, the homovanillyl alcohol, which is relatively rare in free form, and the homovanillyl alcohol-4'-O-b-D-glucoside.

It was also possible to isolate a number of lignans, dimeric phenylpropanes, which are derived from three basic types: - Seciosolariciresinol-9-ObD-glucoside, of the 1,4-butanediol type; 8-O-4'-aryl ether-type substances are derived, according to Gottlieb D7 ' ^(E) -7-O-bD-glucopyranosyl-4,4', 7,9,9'-pentahydroxy-3,3 'dimethoxy-8-O-4'-lignans, 6

presumably erythro / threo isomers, and their aglycones and derivatives of the so-called monoepoxylignans, which have a 2,3,4,5-tetrasubstituted tetrahydrofuran ring. These include the well-known Neo-Olivil as well as compounds 9-Acetyl-Neo-olivil, 9,9'-Disacetyl-Neo-Olivil, Neo-Olivil-4-ObD-glucoside, 9-Acetyl-Neo-Olivil-4-ObD -glucoside, 9'-acetyl-

Neo-Olivil-4-O-b-D-glucoside and the predominant lignan 9,9'-bisacetyl-Neo-Olivil-4-O-b-D-glucoside.

In addition to the sitosterol found in the roots, its glucoside and adenosine, 7 flavonol glycosides could be isolated from the flowers of the large nettle.

These are the quercetin derivatives, isoquercitrin and rutin, the kaempferol dehvate, astragalin and kaempferol rutinoside as well as isorhamnetin neohesperidoside.

The occurrence of flavonoids in Urtica dioica L. is thus secured.

The DC comparison of the methanol extracts of male and female flowers gave a nearly identical picture. The isolated substances occur in both, but there is a quantitative difference with regard to the quercetin and isorhamnetin derivatives. The former appear to be present in higher concentrations in female flowers, whereas the latter predominate in quantity in male flowers.

The different parts of Urtica dioica L. and Urtica urens L. plants have almost identical ingredient spectra in the above-ground organs. They only seem to differ in the occurrence of the neo-olivial derivatives in the underground organs.

From a chemotaxonomical point of view, the examined species Urtica dioica L. and Urtica urens L. can be classified qualitatively as the same in terms of their management of non-polar sterols. The extracts of Urtica urens L. indicate a possible closer relationship in terms of the lignan spectrum to Urtica dioica L. From the aqueous root extract of Urtica dioica 5 polysaccharides became pure Shown form. A quantitative determination of polysaccharides in the commercial preparation Bazoton ^® showed a neutral sugar content of 0.96%, which consisted mainly of galactose, glucose, arabinose, rhamnose and mannose. A determination of uronic acid showed a share of uronic acids of 0.74%. This results in a total content of approx. 1.7% polysaccharides in the Bazoton ^® .

With the ingredients from Urtica, there is a sufficiently large number of compounds for seduction that are characteristic of the drug and that could therefore be used sensibly as key substances in terms of standardization.

The commercially available specialties urtica aar ^® (150 mg) and aar ^® mic (300 mg) coated tablets contain the standardized dry extracts from nettle herb (DAC). This extract is produced from the comminuted drug using a process described for dry extracts in the monograph "Extracts".

Medication with immunoregulatory substances are substances that nonspecifically intervene to promote or inhibit immune processes (Mayer, A. et al. 1979).

The so-called immunostimulants seem to have in common that they are surface-active and activate the lymphocytes and macrophages responsible for the formation of immunity, e.g. gram-positive bacteria or their components. Listeria monocytogenes preferentially stimulates the B lymphocytes, Comeybacterium parvum the macrophages and BCG the T lymphocytes (Hahn, 1978).

Even with numerous vaccinations there are parasitic inhibitory effects

Non-pathogen and non-antigen-related infections have become known, e.g. Smallpox vaccine for the therapy of herpes simplex or BCG vaccination as

Additional therapy for tumors. Paraspecific effects are induced by: Increase in phagocytosis a. Activation of the lymphopoietic cell system b. Formation of interferon c. Stimulation of humoral factors.

The local administration of the vaccine, especially orally and nasally, is apparently better for parasites than parenteral vaccination.

Increasing the defense against infections, stimulating the mechanisms responsible for immunity, inducing interferon and the parasite-specific effects of certain vaccinations have one thing in common: They create a rapidly developing pathogen and antigen-unspecific, more or less resilient protection against one Majority of infections (= Paramunity, Mayer, A. et al., 1979).

The formation of a paramunity leads to an increase in phagocytosis activity, to a better recognition, uptake and mediation of antigens - especially in T-lymphocytes - and thus to a faster development of the killer cells, effector cells derived from the antigen-specified T cells. Memory cells, helper cells and repressor cells, which together with the B lymphocytes regulate the immune mechanism centrally.

In connection with these induction, stimulation of the lymphopoietic cell system - especially that of the T lymphocytes - not only leads to a faster and stronger reaction with specific antigens, but also to an increase in the function of already antigen-specified T and B lymphocytes, whereby the stimulation of the T cell-dependent repressor cells has a favorable regulatory effect (Mayer, A. et al., 1979).

An infiltration of macrophages in the spleen, liver and lymph nodes and an increase in monocytes in the circulation run parallel to these processes. The increased responsiveness of the phagocytic and lymphopoietic cell system seems not to be based on a lymphopoietic multiplication of the phagocytes or lymphocytes, but rather to be the result of an increase in the metabolism (Mayer, A., et al., 1979).

A paramunity can be acquired naturally and artificially, both systemically and locally via the mucous membranes of the respiratory, digestion and urogenital tracts. The locally acquired paramunity is of particular importance (Mayer, A., et al, 1979).

In contrast, the object of the present invention is to provide new forms of use for the aforementioned extracts.

The above-mentioned object is achieved according to the invention by the use of bearberry leaves (Arcostaphylos uva ursi, AU) -, birch leaves (Betula, BP) -, horsetail herb (Equisetum, EM) - and nettle (Urtica, UD) extracts, in particular dry extracts from the fresh or dried plant for the manufacture of pharmaceuticals for the treatment or prevention (protection) of radiation (radio prevention, protection), infection (infection prophylaxis) and chemically related organ and tissue damage (chemoprevention, protection), in particular of the O-MALT system of the small intestine (inflammatory diseases, enteritis regionalis Crohn), the lungs, (inflammation, disease), the bronchi and the nasopharynx (rhinopharyngeal bronchitis, thinitis, pharyngitis, sinusitis), the bone marrow (bone marrow aplasia), the thymus udysfunction , Aplasia or hypoplasia as a result of damage caused by medication or radiation), the liver (athropia, necrosis), hepatitis A , B and C, the pancreas, (insufficiency of the exocrine, secretory function of proteases, esterases, carbohydrases and nucleases as well as the endocrine dysfunction of carbohydrate metabolism (Langernhans Islands) and the kidneys (insufficiency) as well as of generally immunosuppressed conditions, for cellular immunotherapy of leukocytopenia, granulocytopenia, lymphocytopenia, thrombocytopenia, erythrocytopenia (infection, tumor anemia etc.) and in immunoglobulin deficiency states, also as a result of AIDS, tumors 10

and diseases with suppressed immune systems.

According to the invention, a dry extract of bearberry leaves was examined. The defined function of the circulatory and immune systems as well as of the liver, pancreas and kidneys could be influenced with a dry extract in the galenical preparation standardized as a drageekern granulate.

According to the invention, with a birch leaf dry extract - parameters defined in the galenic preparation as drageekern granules of the circulatory and immune systems as well as kidney and liver function in the

Animal testing can be influenced.

The dry birch leaf extracts used (100 mg / kg body weight) induced:

- a slightly negative effect of the average body weight development that differs from the control group, which was attributed to an aquaretic effect

an increase in thymus weight as an expression of cellular immune stimulation

no decrease in erythrocyte count, rather a slight one

Increase and thus no confirmation of a possible haemolytic effect of the total extract tested, although such an effect has been described in the literature for dammanan esters.

an increase in the cell count of segmented granulocytes, leukocytes and lymphocytes, T, B, helper, suppressor and NK cells as an expression of cellular immune stimulation 11

an increase in platelet counts, hemoglobin levels and hematocrit levels

a decrease in the levels of bilirubin, urea, GOT, GPT and AP, as well as cholesterol, triglycerides and protein

no morphofunctional substrate, which should be interpreted as indicating a mutagenic potential of the test substance.

The following are considered to be therapeutically desirable:

the lowering of cholesterol and triglyceride values and thus a positive influence on the risk factor of fat metabolism disorder

the increase in the number of erythrocytes, the hemoglobin content and the hematocrit value as an expression of a qualitative improvement in the physiological blood potential and thus also the oxygen supply to the organism

the absolute increase in defined immunocompetent cells as an adjuvant for an anti-inflammatory effect: lymphocytes, T-lymphocytes, B * lymphocytes, suppressor cells and NK cells with permanent relative cell count ratios, which can generally be interpreted as an expression of a physiological non-specific immune stimulation

the decrease in bilirubin and the liver-specific activity values of GOT, GPT and AP as an expression of a hepatocurative effect

the decrease in urea as a sign of an improvement in 12

renal function and, in addition, the detoxification of the ammonia generated in the protein metabolism and in connection with this reaction cycle an increased mitochondrial performance of the liver cells

lack of signs of adverse effects.

From these experimental results, it was possible to conclude that the registered changes in defined haematological and clinical-chemical analysis values should be viewed in two ways: on the one hand, the results of the parameters used speak for the aquaretic, antipyretic, antirheumatic, antityphoral and bakeriostatic effects of the birch leaf dry extracts, on the other hand for new biological effects, which can be used under therapeutic aspects in disorders of the fat metabolism, liver and kidney function as well as deficits of the cellular immune status.

The following effects of the drageekern granulate with standardized dry extract from horsetail herb DAB 10 (EK) are considered therapeutically desirable:

the general increase in defined lymphatic cells in the terminal current pathway and in the primary and secondary lymphatic organs with a view to improving the non-specific immune defense in the treatment of inflammatory disorders of various pathogenesis, but also as scientific evidence of a "hyperleukocytosis" described in the 1930s "as a result of oral administration of resorbable silica, the decrease in the serum values of bilirubin and urea and the slight morphofunctional expansion of draining urinary tubules in the medullary zone as a sign of a specific or a general improvement in renal function; ie promoting the detoxification of the ammonia generated in the protein metabolism and the diuretic activity 13

the lowering of the cholesterol and triglyceride values in the sense of a positive influence on derailments of the fat metabolism and consequently reduction of atherogenic risk factors, but also as a recent proof for the hypolipidemic effects listed in the literature the increase of the erythrocyte number and the hemoglobin content in the peripheral blood and the cell production of the peripheral blood red bone marrow in the spongiosa of the stem as an expression of an improved oxygen supply, but also as current evidence of one of the components of hemostyptic activity described in the literature; These obviously differ from the mode of action of local and parenteral hemostatic agents, as there are no signs of micromorphological structural disorders or pathologically altered functions of the organs examined in the circulatory system (aorta, heart) of the digestive tract (stomach, duodenum, hay, liver and pancreas) and the urogenital system (kidneys).

From these experimental results, the conclusion can be drawn that the registered changes in defined haematological and clinico-chemical analysis values as well as morphofunctional status images of primary and secondary lymphatic organs are to be considered under two aspects: on the one hand, the present "results speak for the diuretic, hemostypical, erythropoietic described in the literature and anti-inflammatory effects of the test substance EK, on the other hand for new - as yet unpublished - biological effects, which can be used under therapeutic aspects for deficits in the cellular immune status as well as for disorders of the fat metabolism and kidney function.

No signs of micromorphological structural disorders or pathologically altered functions of the organs examined were found.

The object of the present invention is also to

Dry extracts for the production of orally administrable drugs for the treatment of radiation, infection, tumor and chemically related organ and 14

Odor damage, leukocyto, granulocyto, lymphocyto, thrombocytopenia, erythrocytopenia also as a result of AIDS, tumors and diseases with suppressed immune system, also for the treatment of hyperlipoprotein (hypercholesterolemia, hypertriglyceridaemia), diabetes, kidney and liver diseases to provide.

The known anti-inflammatory efficacy in the rat paw edema test could also be demonstrated after p.o. application. One of the polysaccharides was found to be highly anti-complementary in both the classic and alternative ways of complement testing, which in turn means that the inflammatory process is strongly influenced. Other immunological parameters were influenced by other polysaccharides: one of the polysaccharides was able to increase lymphocyte proliferation considerably, while another was able to activate effector cells for cytotoxicity against tumor cells as well as to increase the TNF release of macrophages.

The lectin mixture Urtica dioica agglutinin (UDA) isolated from Urticae radix by a Belgian working group (Peumans et al., 1984) has an antiviral effect by induction of human gamma-interferon. Extensive studies of UDA on mouse lymphocytes from thymus and spleen have been carried out in the prior art. It was shown that UDA caused a selective proliferation of T-lymphocytes, which, however, was strictly dependent on the presence of adherent and accessory cells. Interleukin-1 and interleukin-4 stimulation by UDA approximately matched that of Con A. Interleukin-1 is the first interleukin to play a role in T cell activation.

Rheumatic diseases are characterized, among other things, by inflammatory reactions close to the joints, which in the long term lead to wear of cartilage and bone. A causal treatment does not yet exist. The aim of symptomatic, drug therapy with non-steroidal anti-inflammatory drugs is to suppress the painful and movement-restricting inflammatory reactions by inhibiting the cascade of arachidonic acid. 15

To date, at best, incomplete data are available on possibly therapeutic secondary ingredients and the clinical-therapeutic effects of nettle, which are essentially limited to the diuretic effect of the drug. Recent studies for the first time quantified the potential secondary ingredients of the "phenol carboxylic acid type" responsible for pharmacological effects. According to the invention, the anti-inflammatory effects of the nettle leaf extract (1 capsule contains 335 mg nettle leaf dry extract 6, 4-8: 1) (Extractum Urticae dioicae foliorum) or its dominant secondary ingredient, the caffeine in vitro. The anti-inflammatory effects of the nettle leaf extract and the dominant secondary ingredient, coffeefluoric acid, were investigated in vitro with regard to their inhibitory potential on the biosynthesis of arachidonic acid metabolites.

In the in vitro test system chosen here, the nettle leaf extract inhibited the 5-lipoxygenase-dependent leukotriene synthesis by 20.8%, the course of the inhibition not showing a strict concentration dependence. In contrast, coffeoylic acid inhibited the reaction chain in a strictly concentration-dependent manner. The half-maximum inhibitory concentration was 83 mg / ml in the batch.

The content of caffeine in Urticae herba varied between 0.03 and 1.6% of the dry weight, depending on the quality of the drug. The inhibition of the 5-lipoxygenase-dependent reactions detected for the extract was comparable to that after administration of the same concentrations of pure coffeefluoric acid. Nettle leaf extract caused a significant, concentration-dependent inhibition of cyclooxygenase-mediated prostaglandin synthesis.

The half-maximum inhibitory concentration for nettle leaf extracts was 92 mg / ml. The caffeine acid also showed linearity, the IC ₅₀ was 38 mg / ml. Nettle leaf extract contained approximately 10 mg / caffeoyl malic acid / mg

Extract, that is, the 50% inhibition of the cyclooxygenase-dependent prostaglandin synthesis occurred at a computed concentration of coffee 16

of approximately 1 mg / ml batch of the selected assay. Nettle leaf extract therefore showed a 40% greater inhibition compared to pure coffeefluoric acid, which could not be explained by this substance itself. At the same time, in the thin-layer chromatographic separation of the cyclooxygenase-dependent reaction products, coffeoylefluoric acid showed a different inhibitory behavior than nettle leaf extract (PGD ₂ plus PGF _2a vs. PGD ₂ ).

In order to explain the proven superiority of nettle leaf extracts, other previously unidentified active substances must be available. Clinically, the inhibition of leukotriene synthesis by nettle leaf extracts became more important because it inhibited the immigration of immunocompetent cells in the inflamed tissue, as well as their activity, ie the expression of cytokines etc. A positive effect on the progression of rheumatic diseases is thus conceivable, which is still lacking in the case of nonsteriodal anti-inflammatory drugs. The inhibition of the arachidonic acid cascade can be regarded as part of the overall effect of nettle leaf extracts in the sense of antiphlogesis based on the available findings.

In a further investigation, it was examined whether nettle leaf extracts ex-vivo influence the secretion of pro-inflammatory cytokines after lipopolysaccharide (LPS) stimulation in human whole blood from healthy volunteers. LPS-stimulated whole human blood showed an increase in tumor necrosis factor a - / (TNF-a) and interleukin-1 b- (IL-1 b) concentrations that peaked within 24 h followed by a stable plateau, or a slight drop after 65 h. The concentrations of TNF-a and IL-1 b correlated with the number of monocytes / macrophages in the whole blood of the test subjects. The LPS-stimulated TNF-a and IL-1 b concentrations were reduced in a strict dose-effect relationship by the simultaneous addition of nettle leaf extract. After 24 hours of incubation, the inhibitions at the highest IDS 23 concentration (5 mg / ml batch) were 50.8% for TNF-a and 99.7 for IL-1 b (p <0.001). After 65 h, the corresponding inhibition of these cytokines was 38.9% or 99.9% (p <0.001). In contrast 17

The IL-6 secretion was not inhibited by nettle leaf extract, but stimulated to an extent corresponding to the LPS. The synchronous administration of both substances caused no additional increase in IL-6 concentrations.

The effectiveness of the nettle leaf dry extract in patients with inflammatory and degenerative rheumatic diseases was tested in a multicentre three-week application observation by general practitioners (general practitioners and orthopedic surgeons). Inclusion criteria: an analgesic effect not yet sufficiently achieved by NSAIDs with a current pain intensity> 5, which was queried on the visual analog scale. At the time of the initial examination and after a therapy period of 7 and 21 days, the patients documented the current pain intensity using the scale.

A total of 219 patients took part, 142 women (65%) with an average age of 61 years and 77 men (35%), on average 53 years old. 60% of the patients suffered from degenerative diseases of the joints. Approximately 16% of patients with inflammatory joint diseases were represented. Taking multiple answers into account, approx. 22% showed soft tissue rheumatism or rheumatic diseases not specified in more detail. 153 patients already took NSAIDs before the start of use observation, mostly long-term (pretreatment). In principle, the therapy should be tailored to the individual needs of the individual patient. 217 patients could be considered for the evaluation. As it turned out, 72 patients were treated exclusively with nettle leaf extracts over the entire observation period, the others received the nettle leaf extract in addition to the NSAIDs prescribed for them. The pain intensity of different pain qualities was evaluated by means of a total score, which recorded the values for rest, movement and pressure pain given by the patients. In addition, the patients assessed the degree of restricted movement.

121 patients previously treated with NSAIDs received an adjuvant pre-post comparison 18th

Pain intensity decreased by 49% and movement restricted by 48%. The combination therapy NSAID plus nettle leaf extract reduced the pain score by 60% and the movement restriction by 56% in patients not previously treated (n = 5). Patients without previous treatment responded to monotherapy with nettle leaf extracts (n = 41) with a decrease in

Pain intensity from 51%. The movement restriction was reduced by 52%. If the patients who received nettle leaf extracts regardless of any NSAID pretreatment were summarized (n = 71), they all showed an average decrease in pain intensity and restricted movement of 47% at the end of the observation period. In comparison with previously treated patients who were treated with NSAID plus nettle leaf extract, an essentially identical therapy result was obtained (decrease by 49%). Untreated patients who only received nettle leaf extracts tended to react less well than those treated with NSAID plus nettle leaf extract. The effectiveness of the three-week therapy was assessed by approximately 80%, its tolerance by more than 95% of the patients as "very good" or "good".

The subgroup analysis of the 106 patients who had received pretreatment before the start of the observation and continued this during the observation period showed a decrease in the pain intensity comparable to the total group. In the patient group without pretreatment in the last three weeks before the start of the observation, but with a combination therapy from the nettle leaf extract and standard therapy (n = 19), there was a 10% higher, i.e. an average 61% decrease the pain parameter observed. The pain was less relieved for patients who had not been pretreated and who received only the capsules containing nettle leaf dry extracts within the three-week observation period (n = 12) (decrease: 43%).

The positive influence on the clinical symptoms was also reflected in the final overall assessments of the effectiveness of the doctor and patient 19

again. Doctors and patients alike assessed the effectiveness of the supportive therapy with nettle leaf extract as good or very good in approximately 80% of cases. Tolerance was rated good or very good by over 95% of doctors and patients. One patient stopped treatment with nettle leaf extracts before the three-week observation period due to an allergic reaction. While the effectiveness of the previous monotherapy with NSAIDs was rated as good or very good by the doctors in only 17% of the cases, the doctor's judgments after three weeks of adjuvant therapy with nettle leaf extract were significantly better with 78% good or very good response (n = 106) .

Urtica extracts from herbs, leaves and roots are rich in minerals and trace elements, predominantly contain carboxylic acids, phenol carboxylic acid derivatives, amino acids, phenols, lignans, phytosterols, glucosides, polysaccharides, sugars and lectins.

Of the substances known to date, the lectins (agglutinating, cytotoxic), the polysaccharides (anti-inflammatory, immunomodulating) and caffeine (anti-inflammatory, inhibition of cyclooxygenase-dependent prostaglandin synthesis) were recognized as biologically active in specific tests. However, the Uritca extract showed a greater inhibition in the in vitro test than with pure coffee malic acid (equivalent dose). In order to explain this proven superiority of the Urtica extract, it is assumed that other previously unidentified active substances synergistically increase the inhibitory effect. This connection showed that the totality of certain extractive substances was obviously the dominant carrier of defined therapeutic effects.

In vivo studies with nettle leaf dry extract:

The tasks of the experimental in vivo study were:

Testing the effectiveness of the coated tablet granules with standardized drying 20th

ken extract from stinging nettle herb and from stinging nettle leaves as well as film-coated tablet granules with standardized dry extract from stinging nettle root on the circulatory and immune system and on defined functions of the liver and kidneys after oral administration;

Studies of bioequivalence and

Evidence of possible signs of undesirable effects from these test substances.

The rat species was very suitable as a test animal for carrying out in vivo haematological examinations of the peripheral blood and bone marrow. In contrast to the mouse, the minimal blood withdrawals required for modern microanalysis remain without major changes in haematological parameters. In the morphological comparison of the hematopoietic and circulating blood cells, human and murine blood differ only very slightly. In addition, the rat model is biologically suitable for carrying out investigations in the field of human as well as murine cytokines, since these are equally haematologically immune-active. Results of immunological investigations with defined active substances in the rat correlate very well with the corresponding studies in humans due to the very related morphology and kinetics of circulating leukocytes.

The test substance used

had no influence on the percentage increase in the average body weight as well as the organ weights of the spleen and thymus of the test animals

- induced an increase in the number of leukocytes, segmented granulocytes and lymphocytes, T, B, helper, suppressor and NK cells in peripheral blood 21

Expression of cellular immune stimulation; It should be particularly emphasized that there are no changes in the physiological relationships of these tested immunocompetent cell types

led to an increased tendency in the bone marrow to form cell-rich nests of granulo-, erythro-, lympho- and thrombopoiesis, whereby it was conspicuous that the number of mature myelocytes with ring or ring nuclei was significantly reduced

caused an increase in lymphoblastic cell elements in the cortex and medulla in the thymus

led to widening of the marginal zones in the spleen, particularly in the area of the periarterial vagina (PALS), which corresponds to an increase in lymphocytic cell elements

induced a significant increase in lymphocytic cell elements in the B- and T- in the mesenteric lymph nodes

Lymphocyte areas as well as in the market strands

caused a decrease in the levels of creatinine, bilirubin, urea, GLDH, GOT, GPT, AP, cholesterol, triglycerides, glucose, protein, calcium and in the peripheral blood

Potassium.

The following are considered to be therapeutically desirable:

- The general increase in defined immunocompetent cells as a possibility of non-specific treatment of inflammatory processes of different causal pathogenesis 22

the decrease in bilirubin and the liver-specific activity values of GLDH, GOT, GPT and AP under the aspect of a hepatocurative effect

the decrease in the serum values of creatinine and urea as a sign of an improvement in renal function and thus the detoxification of the ammonia produced in the protein metabolism and in connection with this reaction cycle an increased mitochondrial performance of the hepatocytes

the lowering of cholesterol and triglyceride values in the sense of a positive influence on pathological fat metabolism disorders and thus a reduction in atherogenic risk factors

the decrease in serum glucose as a functional equivalent for a hypoglycemic effect

missing signs of micromorphological structural disorders or pathologically altered organ functions in the sense of undesirable, in particular carcinogenic or mutagenic, effects due to the results of the hematological, clinical-chemical and histological examinations required for evaluation.

From these experimental results, it was possible to conclude that the registered changes in defined hematological and clinico-chemical analysis values as well as morphofunctional status images of primary and secondary lymphatic organs can be viewed under two aspects: on the one hand, the present results speak for the antibacterial and leukocyte-stimulating effects described in the literature the test substance, on the other hand for new - as yet unpublished - biological effects, the therapeutic aspects of deficits in the cellular immune status as well as disorders of carbohydrate metabolism, liver and kidney function

Can find use. 23

The test substance administered orally showed an increased release of segmented granulocytes, monocytes, T and B lymphocytes, helper cells, suppressor cells, NK cells and thrombocytes into the peripheral blood with regard to cell growth, the physiological relation of these cell qualities in comparison mi those of the control group remained almost unchanged.

This increased release was accompanied by an increased cell regeneration rate of granulo-, erythro-, lympho- and thrombopoiesis in the bone marrow, whereby the number of mature myelocytes with nuclear or toroidal nuclei was significantly reduced (morphological substrate of a youthful bone marrow). In contrast, an exogenous supply of adrenaline, for example, led to an increase in the number of neutrophils in peripheral blood, but to stimulate the formation of neutrophils.

The circulating lymphocytes mainly come from the thymus and peripheral lymphoid organs (spleen, lymphocytes, etc.). The lymphocyte stem cells are located in the bone marrow. Some of these relatively undifferentiated lymphocytes migrate to the thymus, where they become T lymphocytes.

Under the influence of bearberry leaf extracts, the cell count of the lymphocytes in the bark zone of the thymus was significantly increased, which on the other hand led to a relative reduction in the marrow zone. In the thymus marrow there are usually predominantly mature T lymphocytes, which are about to leave the thymus via the bloodstream. The increased secretion of T-lymphocytes as well as the other lymphocyte subsets could be measured in the in-vivo study after specific monoclonal incubation by means of fluorescence-activated cell sorting in the blood.

The circulating lymphocyte count is also increased under the influence of the substance acetylsalicylic acid. In contrast to bearberry extracts, the chemically defined substance clearly led to the thymus marrow being stored 24

delayed resettlement tendency.

The T-lymphocytes then colonized specific regions in the peripheral lymphatic organs, for example the paracortical zone of the lymph nodes and periarteriolar accompanying sheaths in the spleen. The situation was different for B lymphocytes. These remained in the bone marrow, where they differentiated. From here they migrated to the peripheral lymph organs, where they settled in regions intended for this cell quality (for example in the lymph follicle).

Micromorphologically, the bearberry extract induced a widening of the marginal zone in the spleen, particularly in the area of the periarteriolar lymphatic sheaths (PALS), without inducing an increased proliferation of immunoblasts. PALS is the T cell region of the spleen. Lymphocytes with T cell properties predominated, of which 70 - 90% were T helper lymphocytes and 10 to 30% were suppressor cells. After antigen stimulation, there was a vigorous proliferation of immunoblasts in the area of PALS, characterized by the increased occurrence of mitotic core division figures of the lymphocytes. The marginal zone, which contained around 50-70% T-lymphocytes and around 40% B-lymphocytes, was the border section between white and red pulp. This zone plays an important role in the immunological activity of the spleen.

Under the influence of bearberry extracts, the lymphocytic cell elements in the B and T lymphocyte areas and in the medullary canals were stimulated in the mesenteric lymph nodes. The paracortical zone mainly had T lymphocytes and was therefore called the thymus-dependent zone. The intensive cell accumulation, in particular of medium, small, and naked nucleus lymphocytes (without macrophage and germ center activation) in the corresponding areas and market strands had to be an expression of an antigen-independent, side effect-free immune stimulation in the mesenteral lymph nodes by the

Bearberry extracts can be interpreted. In contrast, antigens (microorganisms, toxins), for example, caused immune reactions in the B lymphocytes 25th

out. The processes began in the peripheral zone (corona) of the lymphatic follicles and continued in the germinal centers with the involvement of the dentritic and T helper cells. There the cells multiplied, and immunoblasts and immunocytes and finally plasma cells emerged, which migrated into the market strands. There other T, helper and suppressor cells had the opportunity to regulate the immune response. The synthesized specific antigens were then released into the lymph of the marrow sinus. After antigenic stimulation, a large number of lymphocytes left the lymph node; among these were many newly formed sensitized T lymphocytes.

The results obtained according to the invention allow the conclusion that, as a result of the induction of non-specific granulo- and lymphopoiesis - an increase in the cell numbers of phagocytosis-capable and immunocompetent cells in primary (bone marrow) and secondary lymphatic organs (thymus, spleen, lymph nodes) and in peripheral blood - in a pathogen - and antigen-unspecific state of an increased immune defense developed in a healthy organism. In the event of the action of pathogenic microorganisms or antigens, for example in the case of an acute inflammatory disease of the urinary tract - known indication of bearberry extracts - the stimulated effect of the physiological activity of the lymphatic cell system (parameterization) could explain the curative effect of these bearberry leaves.

The test substances UK, ÜB and UW influenced the noradrenaline and adrenaline content in comparison with the control group. Compared to the control group, noradrenaline levels were reduced by 64.4% in treatment group II (UK), by 49.4% in group III (ÜB) and by 62.0% in group IV (UW). The adrenaline values were decreased in Group II (UK) by 11% in the group III (ÜB) to 44.4% (only 1 sample recyclable) and in the group IV of 2.8% ^'.

The light microscopic micromorphological analysis of bone marrow, thymus,

Spleens, lymph nodes and Peyer's plaques of the animals from treatment groups II (UK), III (ÜB) and IV (UW) showed in comparison with the corresponding organs 26

the animals from group I (control) indicate substance-related stimulation of immunologically relevant cell compartments. The micromorphological evidence of an increased immunological activity correlated with the increase in the tested immunocompetent cell types in the peripheral blood. The micro-morphological substrate of the immune system on the one hand and the quantitative behavior of immune-competent cells in peripheral blood on the other hand showed no usable differences in a direct comparison of the three urtica treatment groups, which could be interpreted in terms of a bioequivalent active potential of the urtica extracts from herbs, leaves and roots.

The histological examination of the aorta and heart showed no usable micromorphological changes when the organs from the treatment groups were compared with those from the control group.

Stomach, duodenum, lieum, life and pancreas of the treatment groups impressed by an intact microarchitecture, both in the treatment groups and in the control group.

In the kidneys of the animals from the three groups of active substances, signs of a slight morphofunctional expansion of draining urinary tubules in the marrow zone were occasionally observed, which was not found in the kidneys of the control animals.

In the present in vivo study in rats - treatment duration 7 days - with the help of coated tablet granules with standardized dry extract from stinging nettle herb (UK) and from stinging nettle leaves (ÜB) as well as film-coated tablet granules with standardized dry extract from stinging nettle root (UW), defined functions of the immune and Circulatory system, fat metabolism and the liver and kidneys are affected after oral administration.

Because when evaluating the group data of the tested parameters - with the exception of the quantitative behavior of the erythrocytes - no usable differences 27

In a direct comparison between groups II (UK), III (ÜB) and IV (UW), the urtica groups were subsequently regarded as equivalent units and compared as a whole with the control group.

The test substances used UK, ÜB and UW - related to the application of the same native extract equivalents - induced changes in the following physiological parameters:

Increase in the number of cells of leukocytes, segmented granulocytes as well as T, B, helper, suppressor and NK cells as an expression of a cellular immune stimulation; It is particularly noteworthy that there were no changes in the physiological relationships of these tested immunocompetent cell types

- Increase in MCH, glucose and chloride

Decrease in platelets and levels of creatinine, urea, GOT, cholesterol, triglycerides, noradrenaline, calcium and potassium

- increase in the number of erythrocytes, the hemoglobin content and the hematocrit; bioequivalent only in the treatment groups UK and ÜB, but not in the treatment group UW

Stimulation of immunologically relevant cell compartments of the primary and secondary lymphatic organs

The following effects of UK, ÜB and UW were considered therapeutically desirable:

- The general increase in defined immune competent cells in the peripheral

Blood and in the primary as well as secondary lymphatic organs with the aspect of an improvement of the non-specific immune defense in the 28

Therapy of inflammatory affections of different pathogenesis

the decrease in the serum values of creatinine and urea and the slight morphofunctional expansion of draining urinary tubules in the medullary zone as a sign of a specific or a general improvement of the renal functions, that is to say, favoring the poisoning of the ammonia and the diuretic activity arising in the protein metabolism

the lowering of cholesterol and triglyceride values in the sense of a positive influence on pathological fat metabolism disorders and thus a reduction in atherogenic risk factors

the increase in the number of erythrocytes, the hemoglobin content and the hematocrit value (for UK and ÜB) as an expression of an improved oxygen supply

Reduction of the circulating neurotransmitter norepinephrine as a possible component of the clinically proven analgesic effects of urtica aar ^® .

missing signs of micromorphological structural disorders or pathologically altered functions of the examined organs of the circulatory system (aorta, heart), the digestive tract (stomach, duodenum, hay, liver and pancreas) and the urogenital system (kidneys).

From these experimental results, the conclusion can be drawn that the registered changes in defined haematological and clinico-chemical analysis values as well as morphofunctional conditioners of primary and secondary lymphatic organs can be viewed under two aspects: on the one hand, the available results speak for the diuretic (aquaretic) described in the literature, immunological, antiviral, antitumor, antiphlogistic, analgesic and antirheumatic effects of 29

Test substances UK, ÜB and UW, on the other hand for new, biological effects, which are used from a therapeutic point of view in deficits of the cellular immune status as well as in disorders of the fat metabolism and kidney function.

The fresh plant, in particular pressed juice, in dried form, in particular powder or infusion, as a tincture, in particular drops, granules, in particular capsule, and as a tablet, in particular dragee or lozenge, are suitable as application forms. For the purposes of the present invention, methanol is particularly preferably used as an extracting agent for the plant.

Example 1 :

Animal and animal husbandry

Male Wistar rats with an average body weight of 322.5 g were kept under conventional conditions at room temperature (21 ± 1 ° C) in a relative humidity of about 60% and a 12 hour day / night rhythm. Before the start of the experiment, they were subject to an acclimatization period of 12 days.

The diet was based on a standard pelleted Altromin C1000 Misch diet. No. 100 (Altrogge company, location).

The animals received tap water ad libidum as drinking water.

Test substances

Drageekerngranulat with dry extracts from bearberry leaves standardized to hydroquinone derivatives (3-4: 1) extracting agent: purified water

Parent drug: Bearberry leaves (Uva ursi folium) DAB10

1 coated tablet with 420 mg contained bearberry leaf extract 3-4: 1) 228 - 315 mg 30th

Native extract corresponding to 63 mg hydroquinone derivatives calculated as anhydrous

Arbutin;

Application form: Bearberry leaf granules 1, 547 mg (≥ 0.232 mg arbutin, BC) were in 0.02 ml distilled water. suspended (BCS).

The investigations were carried out with the standardized extracts in granular form containing calcium carbonate and saccharide. (Excipients for the granulate form: calcium carbonate, iron oxide, magnesium stearate, cellulose acetate phthalate, sodium carboxymethyl cellulose, talc, triacetin, arabic gum, carnauba wax, sucrose, lactose and silicon dioxide, potato starch)

Application:

BC was administered intragastral to the non-sedated animals once a day for 7 days using a rigid button probe. The suspension was freshly prepared immediately before application and applied homogeneously. The control animals received physiological NaCl solution in an equivalent volume.

Dose and treatment groups:

10 male Wistar rats were divided into the following treatment groups for examination:

Group: Dose ^" : number of animals: (mg / kg body weight)

I control group (comparison) - 5

II BC (invention) 23.2 5

Bearberry leaf preparations based on arbutin in mg (see above)

Blood sampling: 31

On the 7th day of treatment, 2 hours after the application, the animals were taken 2 × 500 μl of blood from the tetroorbital venous plexus, filled into sodium EDTA-coated vessels and mixed well.

Analytical method:

The aim of the investigations according to the invention was to use a defined in vivo model to simultaneously capture specific markers which indicate the therapeutic effect as a synergistic effect of differently influenced physiological control loops at an early stage.

The following were recorded in peripheral blood:

combined: erythrocytes (RBC), leukocytes (WBC), platelets (PLT), hemoglobin

(HGB), hematocrit (HCT), erythrocyte indices: mean cell volume (MCV), mean corpuscular hemoglobin (MCH), mean hemoglobin concentration of erythrocytes (MCHC) and erythrocyte distribution width (RDW)

- flow cytometric:

Leukocyte differentiation: granulocytes, monocytes, lymphocytes, B, T cells, helper and suppressor cells

combined: GPT, GOT,

Glucose,

Cholesterol, triglycerides Na, K, Cl, Ca, creatinine, urea, total protein

With the help of the fully automatic hematology analyzer Sysmex K-1000 it was possible to determine: WBC, RBC, PLT, HGB, HCT, MCV, MCH, MCHC. The 32

The main unit of this device essentially consists of a hydraulic (HS) and an electronic system (ES). The HS is used to aspirate, pipette, dilute, mix and lyse. The ES analyzed and converted the signals from the HS and transmitted the results to the printer. With the help of microprocessors, the ES also monitored the test procedures, the test station and carried out the quality control.

The hematocrit value indicated the percentage volume of the erythrocytes in the blood. Hemoglobin, the chromoprotein contained in the erythrocytes, was an important criterion for the diagnosis of anemia in addition to the number of erythrocytes and the hematocrit value. The classification was based on the erythrocyte indices. Erythrocyte size and hemoglobin content are characterized by the erythrocyte volume (MCV = mean corpuscular volume), the hemoglobin content of the erythrocytes (MCH = mean corpuscular hemoglobin) and the mean corpuscular hemoglobin concentration (MCHC = mean corpuscular heamoglobin concentration. The red cell distribution width (RDW) was a measure of anisocytosis.

The parameters such as enzymes, glucose, lipids, electrolytes, creatinine, urea and protein were recorded selectively, method-oriented, photometrically or ion-selectively using the Cobas Mira analyzer. The additional report provided analysis-specific data from quality control and statistics.

The leucocyte differentiation was carried out using the flow cytometer FACScan ^® after appropriate lysing of the whole blood sample (scattered light measurement).

Lymphocyte differentiation was carried out after specific monoclonal incubation using fluorescence-activated cell sorting (fluorescence measurement).

Quantitatively, m 7 treatment days were analyzed: leukocytes (total), lymphocytes (total), 33

T lymphocytes (CD2 + / CD45 RA-), B lymphocytes (CD2- / CD45 RA +), helper lymphocytes (CD4- / CD8b +).

To phenotype the lymphocytes, they were incubated with the following antibodies from Pharmingen, San Diego USA, to which a fluorochrome is coupled:

Fluorescein Isothiocyanate (FITC) conjugated Mouse Anti-Rat DC 2 Monoclonal Antibody, Phycoerythrin (R-PE) conj. Mouse Anti-Rat CD 4 Monoclonal Antibody,

Fluorescein Isothiocyanate (FITC) Mouse Ant-Rat Monoclonal Antibody, R-Phycoerythrin (R-PE conj.Mouse Anti-Rat CD 8 (ß-Chaim) Monoclonal Antibody, Fluorescein Isothiocyanate (FITC) conj.Mouse Anti-Rat CD8a Monoclonal Antibody.

Approach: a) CD2 / CD45RA = T and B cells b) CD4 / CD8b = T ₄ and T ₈ cells c) CD8a / CD8b = NK cells

5 μl antibody each were incubated with 50 μl Na EDTA blood at room temperature in the dark for 20 min. The suspension was shaken with 2 ml of Lyses reagent from Becton-Dickinson and incubated for 10 minutes as described. The mixture was then centrifuged at 400 G for 6 min and the supernatant was poured off. The sediment was washed with 3 ml cell-wash and centrifuged at 400 G for 6 min. The sediment was taken up in 100 ul cell-wash. The suspension was analyzed using the flow cytometer.

Clinical observations

The general well-being of the animals was checked during acclimatization before the start of the experiment and throughout the entire duration of the experiment. In addition 34

body weight was determined daily.

All animals survived the intragstral application without impairments. The animals that were treated with the test substances showed no adverse behavior.

The animals of all groups were painlessly killed and dissected at the end of the experiment. The abdominal, pelvic and thoracic organs were examined macroscopically and the spleen and thymus were weighed.

The histological examinations were carried out after the organ specimens had been fixed to the paraffin sections (Medim-Plast ^® ) with hematoxylin-eosin staining (HE).

The following selected organs are available for light microscopic examination: bone marrow (sternal), thymus, spleen, lymph nodes (mesenteric catchment area) from the area of the immune system and the large parenchyma liver and kidneys.

During the treatment period, the average body weight increased only slightly in the groups.

The immune potential of the test substance in group II was indirectly determined by its stimulating effect on the number of immune-competent cells in the peripheral blood.

In treatment group II according to the invention, the mean cell count values increased by 27.1% in the leukocytes and by 21.6% in the thrombocytes in comparison with the control group.

The test substance BC showed no physiologically significant influence on HCT, MCV and MCHC. 35

The following clinical-chemical metabolic parameters were combined in a method-specific analysis:

Bilirubin, creatinine, urea, glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), gamma glutamyl transferase (GGT), glucose, cholesterol, triglycerides, calcium, sodium, potassium, chloride and total protein.

The following was measured for the test substance: a clear decrease of:

Bilirubin by 10.8%

Creatinine by 37.5%

Urea by 35.3%

GLDH by 21.6%

GOT by 29.8%

GPT by 25.0%

Cholesterol by 27.8%

Triglycerides by 34.0%

16.4% glucose

Caicium by 22.9%

Protein by 20.9%

slight decrease of:

Potassium by 1.9%

Leu cyzytend ifferencing

In treatment group II, the cell numbers of the granular granulocytes, monocytes, T and B lymphocytes, helper, suppressor and NK cells were reproduced in comparison with the control group.

The changes were measured for the test substances BC: 36

Leukocyte increase of 27.1% ¹⁾

28.2% increase in lymphocytes

22.6% increase in T lymphocytes

28.1% increase in helper cells - 25.8% increase in suppressor cells

9.3% increase in NK cells

¹⁾ corresponding values of the control group = 100%

Like all parts of the hematopoietic system, the matrix of the red bone marrow consists of reticular cells and reticular fibers, which together form a loose network. Erythropoiesis and leukopoiesis (granulopoiesis, monocytopoiesis, thrombopoiesis and, to a lesser extent, lymphopoiesis) contain cells in the mesh. There are also mature blood cells, macrophages and different amounts of fat cells. The bone marrow is traversed by numerous endothelial-lined sinusoidal capillaries, including the supply and discharge vessels.

The reticular cells are highly branched. Their processes are interconnected and their surface nestles against reticular fibers. Reticulum cells form an adventitia around the sinusoidal capillaries. Recticular cells, together with macrophages, have the ability to phagocitate and degrade foreign and endogenous substances that have entered the bone marrow.

The free cells in the mesh of the bone marrow matrix are predominantly blood formation cells. They tend to form nests, with the formation of a type of blood cell predominating in each network. Due to their cell and nucleus size - despite their small number - and the many normoblasts with a dense cell nucleus, the megakaryocytes are conspicuously exposed between the blood formation cells by light microscopy.

When the blood cells are mature, they enter the sinusoids of the bone marrow 37

and from here - usually in batches - into the blood. The wall of these vessel sections and the capillaries consists of a delicate, fenestrated endothelium, which is covered by a loose mesh fiber network. A basement membrane is largely missing. Adventitia cells (reticulum cells) capable of phagocytosis are attached. The mature blood cells enter the bloodstream through the endothelial cells, the granulocytes are an active process, the details of the discharge have not yet been clarified in the case of the erythrocytes.

The terminal current pathway in the bone marrow is connected on the one hand to supplying arterioles through which blood reaches the bone marrow and on the other hand to venules that drain the blood.

The most important tasks of the red bone marrow are therefore the formation of blood cells including immunologically incompetent progenitor cells of B and T lymphocytes and their release into the terminal current pathway

Participation in erythrocyte degradation and storage of the iron released as ferritin, (released from the storage reticulum cells to neighboring erythroblasts, taken up by them by phagocytosis and used to build up hemoglobin).

KriαcώsnmarJk lrjjpp ^

The light microscopic examination of the sternal bone marrow in the H.-E.-stained paraffin section impressed the colorfulness of the marrow image with the juxtaposition of the easily identifiable cell elements of a functional bone marrow. All cell qualities from the development series of normal hematopoiesis could be differentiated. When you look at the magnifying glass, the most conspicuous bone marrow cells due to their size - namely the megakaryocytes - file. Fat cells were present. The sinusoidal capillaries and the supply and discharge vessels were normal. 38

Bone marrow group II (according to the invention)

In 4 out of 5 animals in this group, the bone marrow completely filled the respective marrow space. In comparison with the corresponding control preparations, a higher cell density was noticeable, with the blood formation activity taking place in the individual regions without any discernible differences. Fat cells were no longer found. When the magnification was increased, it became apparent that the number of mature myelopoese cells with holes or rings in the nucleus was reduced in these 4 animals.

The thymus is divided into lobes, which are formed as side branches of a central market strand and are held together. The microscopic sectional view of a thymus lobe reveals a central medulla and the surrounding, much more cell-rich cortex. The medulla and cortex have a cellular network (reticulum as the basis. In the mesh of this epithelogenic reticulum lie thymocytes (lymphoid cells), which are morphologically indistinguishable from small lymphocytes. The thymocytes are in the cortex, which is to be regarded as a germ layer extremely tightly stored.

The lymphocytes of the cortex are of different sizes. Large lymphoblasts are preferably located in the outer zone of the cortex. These cells are derived from immigrated lymphoid stem cells. Proliferation gives rise to small, immature T-lymphocytes, the majority of which transfer to the inner thymus cortex and remain sedentary for long periods. However, most of them die and are not used to colonize the secondary lymphoid organs. There are only a few, also small, but mostly mature T lymphocytes in the thymus marrow. These cells come from the inner thymus cortex and are about to leave the thymus via the bloodstream.

The reticulum cells of the thymus are branched, cytoplasmic cells connected by desmosomes with a large nucleus, the chromatin 39

is dispersed.

Small membrane-bound granules are observed in the cytoplasm of some reticular cells. The so-called thymus hormones (thymus factors) are stacked in them, which are of great importance for the differentiation of T-lymphocytes.

In addition, interdigitating cells (from the bone marrow) are found in the epithelial network, especially in the medullary zone. These cells, which contain many class II antigens of the main histocompatibility complex, are said to play an important role in the learning process of recognizing self-antigens in the thymus: Main histocompatibility complex - (MHC) antigens of class II are for the interaction of the cells of the immune system is important, as is the recognition of an antigen by helper T cells.

The parenchyma of the cortex and medulla of the thymus is traversed by blood vessels. Lymphatic vessels are restricted to the connective tissue components. Blood capillaries and venules have an unfenestrated endothelium. A coat of reticular cells clings to its basal lamina. The endothelium of this vascular segment is impermeable to antigens in the thymus cortex, similar to the blood-brain barrier (blood-thymus barrier). However, thymocytes are able to overcome this barrier. In the thymus medulla, however, antigens can pass through the endothelial bandage and come into contact with the thymus cells.

Histological assessment of the thymus requires special attention to the structural elements of the cortex and medulla, since active substances that have an effect on the immune functions can change the morphology of these zones, both in terms of regression and stimulation.

Thymus Gr. I (comparison) 40

The light microscopic examination of the thymus generally showed a basophilic basic tone in the H.E.-stained section. The cortex, which appeared densely packed with large lymphoblasts, immature, proliferating cells and small lymphocytes, stood out clearly from the medulla, which was relatively sparsely populated by comparison with small, mature T-lymphocytes.

Thymus, _Gruppe_lL (according to the invention)

In the histological examination, 4 out of 5 animals in this group noticed that in numerous thymus lobules the marrow zone no longer differs so clearly from the

Bark zone took off, as was the case with the control animals. With stronger

A micro-morphological explanation was found for the enlargement. Starting from the inner cortex zone, bright lymphoblast conglomerates pulled into the medulla zone, which on the one hand no longer made the transition between the cortex and medulla zone sharp, on the other hand there was a relative reduction in the medulla zone or an irregular one

Enlargement of the bark area. The absolute average thymus weight of group II treated according to the invention was not increased in comparison with group I.

The light microscopic examination of this organ showed the typical micromorphology of the rat spleen: the parenchyma is composed of white and red pulp, which are distinctly stained in terms of color, the basophilic basic tone predominating in the former and eosinophilic in the latter. Thorough studies of the spleens of these animal species have recently shown that predominantly the T lymphocytes of the thymus are located in the periarterial sheaths made of lymphoreticular tissue (PALS), whereas the B lymphocytes are located in the splenic follicles (Malpighian bodies). The T lymphocytes are primarily small and medium-sized cell forms. Light centers (germ or reaction centers) in the splenic nodules (secondary follicles) are occasionally found. 41

The marginal or mantle zone ("nodule margin zone") around the splenic follicles and PALS is free of sinus and blood vessels. It contains large mononuclear cells with a round nucleus and little chromatin, most of which are addressed as juvenile lymphocytes. In the rat, the marginal zone is separated from the actual follicles by a narrow border of collagenous connective tissue and also sets itself apart relatively sharply from the red pulp. It is of special significance for rats. It is a barrier that separates the white pulp from the liquid and corpuscular components that come from the blood, since the spleen is involved in the bloodstream and not in the lymphatic system. Cooperative interactions between B and T lymphocytes begin in this marginal zone, which therefore fulfills an important function for the localization of antigens. In the histological assessment of the spleen, it is therefore necessary to pay special attention to the components of the white pulp, since active substances that have an effect on the immune function can change the morphology of these zones. The red pulp from the sinus and the intersinuous reticular meshwork shows plenty of stored cells, especially erythrocytes. In addition, minor signs of extramedullary hematopoiesis are found in the red pulp in the control animals.

Spleen, jJ q2p_eJ (V jgJeictL)

The light microscopic examination of the spleen of all control animals showed the characteristic micromorphology of this lymphatic organ of the rat. In terms of staining, the follicles and the periarterial lymphoreticular sheaths - white pulp - clearly stood out from the surrounding red pulp with an eosinophilic basic tone due to a basophilic basic tone. The red pulp showed signs of minimal focal extramedullary hematopoiesis in all animals.

Spleen, group IL (according to the invention)

In the micro-morphological examination of the spleen in this group was 42

A widening of the marginal zones of the periarterial vagina (PALS) was observed in 3 animals. An unchanged microarchitecture of this organ was found in one animal. Another animal in this group showed atrophy of the white pulp. There was no evidence of any significant extramedullary hematopoiesis in any animal.

The lymph node is enclosed in a thin connective tissue capsule. Trabeculae (bars) move from the inner surface of the capsule to the hilus. The reticular connective tissue stiffened by lattice fibers spreads out in the space enclosed by the capsule, the meshes of which are largely filled by lymphocytes and plasma cells. The periphery of the lymphatic tissue condenses to the bark layer of the lymph node, in which the primary and secondary follicles lie.

Between the bark and the hilus is the medulla of the lymph node, into which lymphatic tissue continues in the form of the medullary strands. The market strands and the entire subcapsular bark area with primary and secondary follicles contain B lymphocytes and plasma cells that have developed here.

T-lymphocytes, on the other hand, preferentially colonize the paracortical zone and are only occasionally found in the germinal centers of the secondary follicles.

From the marginal sinus, the lymph is led through the bark via an intermediate sinus and then introduced into a wide and richly branched marrow sinus. All sinuses of the lymph node are lined with endothelium and interspersed with reticulum cells with lattice fibers and macrophages so that a trap-like system is created.

Through the wall of the venules, lymphocytes and monocytes or markophages pass from the bloodstream into the lymph of the lymph node parenchyma. Non-body components (bacteria, viruses, inanimate foreign substances) can be retained and macrophages can be phygocyted. 43

Even tumor cells cannot easily overcome these filters. Thymopoetin, which regulates T-lymphocyte maturation in the lymph nodes, has been detected in some reticulum cells and macrophages. When histologically assessing the mesenteric lymph nodes, it is therefore necessary to pay special attention to the structural elements of the cortical B-lymphocyte area, the paracortical T-lymphocyte area and the market strands, since active substances that have an effect on the immune function considerably influence the morphology of these zones can change, both in terms of regression and stimulation.

Mesenteric lymph nodes (MLK) group I (comparison)

The light microscopic examination of the mesenteric lymph node (MLK) of the rat showed in the H.E. - colored cut generally a basophilic basic tone. This resulted on the one hand from the staining cells in the B lymphocyte area (bark), on the other hand from the cells in the T lymphocyte area (paracortex) and from the cells in the medullary cord. The subcapsular B lymphocyte area was rich in B cells. The same applies to the primary follicles above. Secondary follicles with peripheral B-lymphocyte walls and central accumulation of lymphoblasts were found less frequently. The paracortical area was characterized by the accumulation of T cells, the market stems were characterized by the presence of plasma cells and B lymphocytes.

Mes „eDteriaJej_Lymp kno_tejι_ (MLK) Group II (according to the invention)

During treatment with the test substance BC over a period of 7 days, a much stronger basophilic staining of the lymphatic tissue was observed in the light microscopic examination when viewed with a magnifying glass. In comparison with the mesenteric lymph nodes of the control animals, an intensive cell accumulation in the B and T lymphocyte area as well as in the medullary strands impressed, so that the boundaries between these areas were lost. This finding was as 44

Expression of a significantly increased number of cells of the lymphatic group. It was also remarkable that there was no activation of secondary follicles.

The structure of the liver results from the close relationship between the intrahepatic vascular system and liver cells. The 1 to at most 2 cells wide, anastomosing, perforated liver cell plates are closely related to thin-walled sinusoids. This gives metabolites in the blood almost unlimited access to the liver cells. Liver cell plates with their accompanying sinusoids come together to form microscopic liver lobes.

Histological liver sections are characterized by periportal fields (canales centrales) with a triad of at least one interlobular vein, one interlobular artery and one bile duct (interlobular duct)

Vv. Centrales

Rows of liver cells, which are oriented radially towards the central vein, anastomise and lead between them blood vessels with sinusoidal capillaries - sublobular veins, which run individually.

A micro-anatomical structure of the liver can be done according to different criteria. The central vein, the periportal field or the terminal portal veinoles and hepatic arterioles can be regarded as centers of morphological structural units. This results in the division into classic lobules, periportal lobules,

Liver Azini.

Classic lobules: These are descriptive units in which the central vein

Forms the center. This is followed by radially placed liver cell plates and sinusoids. The peripheral corners of the classic lobules form periportal fields that 45

spatially represent a multi-surface, prismatic structure.

Periportal lobule: With this approach, the periportal field is the focus, with the corners being formed by the central veins. As a concept, this approach is based on the fact that the bile formed by the liver cells flows in bile capillaries to the bile ducts running in the periportal fields. The periportal lobule focuses on the glandular character of the liver.

Leberazinus (according to Rappaport): The terminal portal veinoles and

Liver arterioles an axis that is surrounded by liver cells. In the corners of the Leberazini are the neighboring periportal fields and Vv. Centrales. For the construction of the liver from Azini, a functional view is in the foreground. The liver cells form 3 zones around the terminal liver vessels.

-Zone I: Immediately surrounds the terminal liver vessels, the liver cells located here are the first to come into contact with the blood supplied to the liver. -Zone II: The liver cells located in this zone receive blood that had previously had contact with the cells of Zone I.

-Zone III: The blood has passed the two pre-zones and has been changed there.

The zoning plays an important role in numerous metabolic processes in the liver. In the periportal (peripheral) zone are the cells that are best equipped for the breakdown of amino acids and fatty acids to acetyl-CoA and for the endoxidative metabolism and in which the endergonic process of gluconeogenesis and urea formation from amino acids can run optimally. On the other hand, the enzymes of glucose metabolism, lipogenesis and urea formation from ammonia and biotransformation (detoxification processes) occur mainly perivenous (central). The area between 46

these two zones are considered to be a not clearly defined transition zone.

According to the concept of metabolic zoning, this structure must not be understood statically, but must be understood dynamically. Thanks to their enzyme equipment, every liver cell is in principle able to perform any metabolic function. However, the heterogeneous distribution of the enzyme activities indicates that certain metabolic steps preferentially take place in one zone due to the existing functional structure. The transitions of the enzyme activities between and within the zones take place gradually.

The sinusoid capillaries (liver sinusoids) are of particular importance for liver function. They slow blood flow and improve the exchange of substances with the environment. The walls of the liver sinusoids are formed by endothelial cells and Kupffer cells.

Furthermore, the liver sinusoids are characteristic of the wall

- openings and ports,

- reticular fibers on the outer surface,

- the lack of a basement membrane.

The endothelial cells are flat and form most of the wall of the liver sinusoid. They are poor in organelles. Intercellular openings occur between endothelial cells and the endothelial cells also have pores. However, blood components cannot leave the current pathway through openings and pores and enter the perisinusoidal space. The endothelial cells of the liver sinuoid lack a basement membrane.

The Kupffer cells (after v. Kupffer, 1892-1902, Anatom in Dorpat, Kiel, Königsberg, Munich) are cells that are capable of phagocytosis. For example, they take fragments of cells (including erythrocytes or

Erythrocyte parts), bacteria and foreign bodies. Kupffer cells are partly part of the wall of the liver sinusoids, partly they are located on the endothelial cells 47

on. They always have long processes (hence Kupffer star cells) that connect them to the endothelial cells on the same side as on the opposite side of the capillary wall. As a result, extensions of the Kupffer cells cross the current path. Kupffer cells can detach from the sinus wall and reach the lungs with the blood. The Kupffer cells differ cytologically from the endothelial cells by their relative abundance of organelles (granular endoplasmic reticulum, Golgi apparatus, lysosomes, bright vacuoles), but above all by peroxisomes and high peroxidase activity. The Kupffer cells belong to the reticuloendothelial or reticulohistiocytic or mononuclear phagocytotic system. They are modified monocytes. They originate from the bone marrow, which also supplies replacement cells, although Kupffer cells can also mitotically multiply on site.

The Disse space (perisinuoid space) is a wide gap between endothelial cells and surrounding liver cells (J. Diss, 1852-1912, anatomist in Göttigen, Halle, Marburg). Non-cellular parts of the butt get into this disse space. The perisinusoidal space is important for the metabolism between liver cells and blood.

eJιej ^ τup J V-eτgieJcJι)

The liver of the control group was characterized by the microarchitecture typical for this organ: indistinct lobule drawing, network connection of the liver cell plates and a delicate interstitium.

The hepatocytes were characterized by changing functional states. The deposition of clumpy glycogen in the H.E.-stained paraffin section is characterized by more or less large, optically empty cavities perinuclear (cytoplasmic rarefaction). In the vicinity of the central veins, this took

Glycogen and the density of the cytoplasm too. 48

Leber_Gruppeil_ (according to the invention)

The characteristic microarchitecture of the liver remained unchanged during treatment with BC over a period of 7 days. In comparison with the liver preparations of the control animals, there were no signs of functional alterations. The level of glycogen deposits was appropriate. In particular, no micromorphological evidence of degenerative cell or core changes in the hepatocytes could be detected.

The renal parenchyma is divided into medula renalis, (renal medulla), cortex renalis, (renal cortex). The rat's unilobular renal medulla consists only of a conical pyramid with associated bark (pyrames renales), the tip of which (papilla renalis) protrudes into the cali renalis and the ready base of which faces the renal cortex. At the top of the renal papillae there are openings (foramina papillaria) of collecting ducts (ductus papillares) that release urine into the kidney pelvis. The openings form an area cribrosa at each papilla tip. Marrow rays (pars radiata) penetrate from the base of the pyramid into the renal cortex. The medullary rays consist of bundles of collecting tubes and elongated tubule sections. The labyrinth of the renal cortex fills the space between the medulla rays. It essentially consists of niral bodies and tortuous canal sections. The bark tissue between neighboring pyramids forms the Colunae renales (Bertini columns).

The bark can be divided into kidney lobes (Lobuli corticales). Each lobule has a medulla ray and surrounding bark maze.

Niece, _GruppeJL (comparison)

The light microscopic examination of the H.E.-stained kidney preparations in this group showed clear morphological structures of the bark and marrow zone including the papilla and the renal pelvis. They showed in the bark zone

Glomerula a physiological fullness of blood. Epithelium and lumen of the draining tubules 49

were real.

Kidney, _Grupp, eJ (inventive device)

The light-microscopic examination of the H.E.-stained kidney preparations of this treated group showed no evidence of usable differences in morphofunctional condition pictures in comparison with the kidneys of the control animals. In particular, no signs of a disturbed function or changed structure were found in the sensitive cell elements of the capillary loops. The same was true for the entirety of the epithelium of the draining tubules and collecting channels. The uepathel of the renal pelvis was normal, and no pathological content was found in the lumen.

Example ^;

Animals and animal husbandry

Male Wistar rats with an average body weight of 333 g were kept under conventional conditions at room temperature (21 "1 ° C) in a relative humidity of about 60% and a 12 hour day / night rhythm. They underwent acclimatization before the start of the experiment from 12 days on these keeping conditions.

Under the in vivo test conditions according to the invention, under the influence of the orally administered complex Betula extractive substances, the measured values of defined parameters did not allow any interpretation in terms of undesirable effects. Neither hemolytic effects, as observed with certain isolated betulin ingredients, nor signs of weak mutagenicity could be observed.

The birch leaf dry extract was applied intragastral once a day for 7 days. Subsequently, retroorbital blood was taken from the animals and 50

subsequently analyzed in vitro hematologically: leukocytes, erythrocytes, hemoglobin, hematocrit, mean cell volume (MCV), mean corpuscular hemoglobin (MCH), mean hemoglobin concentration of the erythrocytes (MCHC).

In addition, leucocyte differentiation by means of scattered light measurement, lymphocyte differentiation after specific monoclonal incubation by means of fluorescence-activated cell sorting and in vitro were carried out by flow cytometry.

The clinical-chemical parameters bilirubin, creatinine, urea, GOT, GPT, AP, glucose, cholesterol, triglycerides, calcium, sodium, potassium, chloride and total protein were recorded selectively, photometrically or with the help of ion-selective electrodes.

The test substance used (100 mg / kg body weight) induced a slightly reduced body weight gain, an increase in the thymus weight, an increase in the cell numbers of erythrocytes, thrombocytes, segmented granulocytes, leukocytes and the lymphocytes, T, B, helper, suppressor and NK cells, an increase in hemoglobin content and hematocrit value and a decrease in bilirubin, urea, GOT, GPT, AP, cholesterol, triglycerides and protein.

As an application form, the fresh plant, in particular pressed juice, is available in dried form, in particular powder or infusion, as a tincture, in particular drops, granules, in particular capsule, and as a tablet, in particular dragee or lozenge. For the purposes of the present invention, methanol is particularly preferably used as an extracting agent for the plant.

Macromorphological assessment of the spleen and thymus

The mean organ weights of the spleen (+7.65%) and thymus (+22.50%) of the 51

Groups treated according to the invention differed from the corresponding values in the control group.

The immune potentials of the birch leaf dry extracts in the group treated according to the invention were determined indirectly by their induction effect on the increase in the number of immunocompetent cells in the peripheral blood. The samples were analyzed in vitro by flow cytometry, the measurement of the cell size and the granularity based on the registration of corresponding scattering intensity allowing the discrimination of lymphocytes, monocytes and granulocytes.

In the treatment group according to the invention, in comparison with the control group, the mean cell count values increased by 22.2% in the leukocytes, by 12.2% in the thrombocytes and by 1.2% in the erythrocytes.

Hemoglobin, hematoma, _ erythrocyte indices

The percentage increase in the hemoglobin content as well as the hematocrit value in the treatment group (BT) correlates with the corresponding increase in the erythrocytes.

Under the influence of the test substance BT, the index value MCV (quotient of hematocrit and erythrocyte number) was slightly lower with regard to the percentage changes, and slightly higher for MCH and MCHC.

Bilirubin, creatinine tlarastoff, GJukose Jpide, electrolytes and protein

The following clinical-chemical metabolic parameters were combined in a method-specific analysis:

Bilirubin, creatinine, urea, glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), alkaline phosphatase (AP), glucose, 52

Cholesterol, triglycerides, calcium, sodium, potassium, chloride and total protein.

The following was measured for the birch leaf dry extracts:

a clear decrease of:

Bilirubin by 71.0% o

Urea by 16.2%

GOT by 28.2%

GPT by 19.0%

Cholesterol by 26.5%

Triglycerides by 18.5%

a slight decrease in: AP by 4.3% o glucose by 1.8%

a significant increase of: GLDH by 31.1%.

LeuJφ ^ yiejicüflejr ^ nzieTurjg

In treatment group II, the cell numbers of the leukocytes, lymphocytes, T and B lymphocytes, helper, suppressor, NK cells and the segmented granulocytes were reproduced in comparison with the control group.

The following was measured for the birch leaf dry extracts:

Leukocyte increase of 27.5% ¹⁾

25.0% increase in lymphocytes - 21.7% increase in T-lymphocytes

B Lymphocyte Increase 39.2%

Helper cells increase by 8.7% 53

21.5% increase in suppressor cells

Increase in NK cells by 22.7% increase in segmental nuclei

Granulocytes by 37.0%

Although there was a significant increase in defined immune-competent cells under the influence of birch leaf dry extracts, the relative cell number ratios remained almost at the same level for the following cell qualities.

- lymphocytes (I, 77.6%; II, 75.4%)

T lymphocytes (I, 50.6%; II, 50.6%) B lymphocytes (I, 36.8%; II, 40.6%) suppressor cells (I, 16.0%; II, 15 , 4%) NK cells (I, 7.0%; II, 7.0%).

The diet was based on a standard pelleted Altromin C 1000 Misch diet. No. 100 (Altrogge company, location).

The animals received tap water ad libidum as drinking water.

Test substances

Drageekerngranulat with dry extract from birch leaves (4-8: 1) extract: purified water starting drug: birch leaves (Betulae folium) DAB 10

1 coated tablet with 240 mg contained dry birch leaf extract (4-8: 1)

150mg

Application form: Drageekerngranulat 1, 6 mg (≥ 1 mg native dry extract were suspended in 0.02 ml distilled water (BT)).

The above-mentioned investigations were carried out with the corresponding standardized plant extracts in granulate containing calcium carbonate and saccharide. 54

form carried out. (Excipients for the granulate form: calcium carbonate, iron oxide, magnesium stearate, cellulose acetate phthalate, sodium carboxymethyl cellulose, talc, triacetin, arabic gum, carnauba wax, sucrose, lactose and silicon dioxide, potato starch.)

Application:

Birch leaf dry extract was the non-sedated once a day for 7 days

Intragastric animals administered with a rigid button probe. The suspension was freshly prepared immediately before application and applied homogeneously.

Dose and treatment groups:

10 male Wistar rats were divided into the following treatment groups for examination:

Group: Dose: Number of animals:

(mg / kg body weight) I control group (comparison) - 5 II BT (invention) 100 5

The aim of the investigations was to use a defined in vivo model to simultaneously acquire specific markers that indicate the therapeutic effect as a synergistic effect of differently influenced physiological control loops at an early stage.

Analytical methods: see example 1

Example 3:

Male Wistar rats with an average body weight of 329 g were tested under conventional conditions at a room temperature of 21 +/- 55

1 ° C, a relative humidity of around 60% and a 12 hour day / night rhythm. Before the start of the experiment, they were subject to an acclimatization period of 12 days.

The diet was carried out using the pelleted standard diet Altromin C1000, Mix No. 100 (Altrogge, Lage). The animals received tap water ad libidum as drinking water. Drageekerngranulat with dry extract from field horsetail herb (4-7: 1) extracting agent: purified water DAB 10, Ch.-Nr. 049439 extract drug: horsetail herb (Equiseti herba) DAB 10.2. Natr. 1993; 1 Drageekem aar ^® ren contains 300 mg native horsetail herb dry extract (4-7: 1); 1 drageekern equisetum aar ^® contains 150 mg dry horsetail herb extract (4-7: 1).

Application form: equisetum aar ^® granules 1, 86 mg f≥1mg native horsetail herb extract, EK) are in 0.02 ml aqua. least suspended (EKS).

D_osis_uαd_BeharιdJüiιgsgtuppejD

10 male Wistar rats were divided into the following treatment groups for examination:

Group dose number of animals (mg / kg body weight)

I control group - 5

II EK 100 5

The test substance EK is administered intragastral to the non-sedated animals once a day for 7 days using a rigid button probe. The suspensions are freshly prepared immediately before application and applied homogeneously. The control animals receive physiological NaCI solutions in an equivalent volume, based on the respective volume of the test substance / KGW. 56

Analytical methods: see example 1

The potential of the test substance EK was indirectly determined by its stimulating effect on the number of leukocytes (WVC), erythrocytes (RBC) and thrombocytes (PLT).

The test substance EK induced an increase:

in leukocytes by 10.9% in erythrocytes by 1.6% in thrombocytes by 0.6% in hemoglobin by 2.7% by MCH by 0.7% by MCHC by 4.7%

The test substance EK induced an increase

of leukocytes by 11.5% of lymphocytes by 14.0% of T-lymphocytes by 25.8% of B-lymphocytes by 23.1% of helper cells by 14.2%.

The test substance EK induced a decrease:

of NK cells by 221,% of segmented granulocytes by 5.7%

(corresponding values of the control group = 100%) 57

The test substance used Horsetail extract induced changes in the following physiological parameters:

an effect of the average body weight development that deviates slightly from the control group, which is attributed to an aquaretic effect, a slight increase in the average thymus weight and a histologically recognizable broadening of the cortex and medulla of this organ as an expression of a cellular immune stimulation increase in the cell numbers of leukocytes and in T-, B, helper and suppressor cells in the terminal current pathway as an expression of lymphocytic immune stimulation Increase in MCH, MCHC, creatinine, glucose, potassium and chloride Decrease in the levels of bilirubin, urea, GOT, GPT, AP, GGT cholesterol, triglycerides, Caicium, and sodium increase in the number of erythrocytes and hemoglobin in peripheral blood and a histologically detectable cell production of the red bone marrow stimulation of immunologically relevant cell compartments of the primary and secondary lymphatic organs with formation of ovn secondary follicles and signs of increased cell production in the paracortical zone of the lymph nodes.

Examples:

Male Wistar rats with an average body weight of 292.5 g were kept under conventional conditions at a room temperature of 21 +/- 1 ° C, a relative humidity of about 60% and a 12 hour day / night rhythm. Before the start of the experiment, they were subject to an acclimatization period of 12 days.

The diet was carried out using the pelleted standard diet Altromin C1000 (Altrogge, Lage). The animals received tap water ad libidum as drinking water. 58

Test substances and dosage:

Drageekem granules with dry extract from nettle herb (5-10: 1); Extracting agent: purified water DAB 10, Ch.-Nr. 1283936, parent drug: nettle herb (Urticae herba) DAC 1989.3.- Erg. 1991;

1 coated tablet with 600 mg contained 300 mg native nettle dry extract (5-10: 1);

1 300 mg coated tablet contained 150 mg of native nettle dry extract (5-10: 1);

Application form: Granules 2 mg (≥ 1 mg native nettle herb extract) were in 0.02 ml distilled water. suspended, 0.5 ml = 5.8 mg UK / 250 g KGW.

Drageekem granules with dry extract from nettle leaves (5-10.); Extracting agent: purified water DAB 10, Ch.-No .: 049409; Starting drug: Nettle leaf granules 2 mg (= 1 mg native nettle leaf extract, ÜB) were in 0.02 ml distilled water. suspended, 0.5 ml _s 25 mg UB / 250 g KGW.

14 male Wistar rats were divided into the following treatment groups for examination:

Group dose number of animals (mg / kg body weight)

I control group ~ 5

II UK 100 5

III ÜB 100 4

The test substances UK, ÜB and UW were not once a day for 7 days 59

sedated animals intragastral with the help of a rigid button probe. The suspensions were freshly prepared immediately before application and applied homogeneously. The control animals received physiological NaCl solution in an equivalent volume.

The aim of the present own investigations was to use a defined in vivo model to simultaneously acquire specific markers that indicated the therapeutic effect as a synergistic effect of differently influenced physiological control loops at an early stage.

Analytical methods: see example 1

The general well-being of the animals was checked during acclimatization before the start of the experiment and throughout the entire duration of the experiment. In addition, the body weight was determined daily.

The animals of all groups were killed and dissected painlessly at the end of the experiment. The organs of the abdominal, pelvic and thoracic cavities were examined macroscopically and the spleen and thymus were weighed.

The histological examinations were carried out after formalin fixation of the organ samples on paraffin sections (Medim-Plast ^® ) with hematoxylin-eosin staining (HE).

The following selected organs were examined by light microscopy: - Defense system: bone marrow (sternal), thymus, spleen, lymph nodes

(Mesenteric catchment area), Peyersche Plaques circulatory system: aorta, heart

Digestive system: stomach, duodenum, ileum, liver, pancreas urinary organs: kidneys

All animals survived the intragastric application without impairments. The animals treated with the test substance showed no clinical disadvantage 60

Behavior until the end of the experiment. During the treatment period, the average body weight increased by approximately 10%.

The deviation of the mean organ weights of the spleen and thymus in group II UK was -13.8% and 0%,

Group III ÜB - 2.7% or -15% and in the

Group IV UW + 1, 4% and +13.5%.

The potentials of the test substances of UK, ÜB and UW were indirectly determined by their stimulating effect on the number of leukocytes (WBC), erythrocytes (RBC) and platelets (PLT).

Comparable bioequivalent changes in type and intensity were measured for the test substances UK, ÜB and UW:

Leukocyte increase in group II by 32.0% ¹⁾ in group III by 25.9% in group IV by 38.3%

MCH increased by 1.4% in Group II by 1.2% in Group III by 2.4% in Group IV

Platelet decrease in group II by 2.1% in group III by 3.9% in group IV by 10.8% 61

In addition, further bioequivalent changes were registered for the test substances UK and ÜB:

Red blood cells increased by 1.2% in group II by 0.4% in group III

Hemoglobin increased by 2.6% in group II by 1.7% in group III

¹ 'corresponding values of the control group = 100%

Development of the blood count:

Average cell count values of the erythrocytes in 10 ² / l, the leukocytes and the platelets ind 10 ⁹ / l, concentration of the amount of hemoglobin in mol / l, hematocrit in l / l, MCV in fl, MCH in amol, MCHC in mmol / i. Percentage changes of the listed values in relation to the corresponding values of the control group

Group mg / kg body weight / day

I control group

II UK 100

III ÜB 100

IV UW 100

The test substances UK, ÜB and UW showed no physiologically significant influences on the erythrocyte indices MCV, MCH and MCHC. While the influence of UK and ÜB on the hematocrit was insignificant, it fell by 50% after treatment with UW. 62

The following clinical-chemical metabolic parameters were combined in a method-specific analysis:

Bilirubin, creatinine, urea, glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), gamma glutamyl transferase (GGT), glucose, cholesterol, triglycerides, calcium, sodium, potassium, chloride and total protein.

For the test substance UK, ÜB and UW, comparable bioequivalent changes were measured in type and intensity:

- a decrease in creatinine in group II by 21.6% in group III by 22.4% in group IV by 25.9%

- a decrease in urea in group II by 3.4% in group III by 1.4% in group IV by 0.5%

- a decrease in GOT in group II by 11.3% in group III by 6.1% in group IV by 13.9%

- a decrease in cholesterol in group II by 18.3% in group III by 17.2% in group IV by 8.9%

- a decrease in triglycerides in group II by 15.1% in group III by 50.5% 63

in group IV by 43.2%

a decrease in calcium in group II by 19.2% in group III by 32.3% in group IV by 34.8%

a decrease in potassium in group II by 12.1% in group III by 9.6% in group IV by 3.5%

an increase in glucose in group II by 1.5% in group III by 3.0% in group IV by 9.5%

an increase in chloride in group II by 2.8% in group III by 1.9% in group IV by 2.6%

The individual measurements of the number of leukocytes, monocytes, granulocytes and lymphocytes per ml of blood with the corresponding percentages, based on the total number of leukocytes (100%), and of the lymphocyte subpopulation, based on the total number of lymphocytes (100%), are shown below. specified.

In treatment groups II, III and IV, the cell numbers of the segmented granulocytes, monocytes, T and B lymphocytes,

Helper, suppressor and NK cells reproduced. 64

Comparable bioequivalent changes in type and intensity were measured for the test substances UK, ÜB and UW:

Leukocyte increase in group II by 32.0% ¹⁾ in group III by 25.9% in group IV by 38.3%

Lymphocyte increase in group II by 26.5% in group III by 27.0% in group IV by 18.2%

T-lymphocyte increase in group II by 33.8% in group III by 32.0% in group IV by 11.0%

B lymphocyte increase in group II by 66.7% in group III by 47.0% in group IV by 43.0%

Helper cells increased by 15.9% in group II by 36.9% in group III by 15.8% in group IV

corresponding values of the control group = 100%

Claims

65 PATENT REQUIREMENTS

1. Use of bearberry leaves (Arcostaphylos uva ursi, AU) -, birch leaves (Betula, BP) -, horsetail (Equisetum, EM) - and nettle (Urtica, UD) extracts, especially dry extracts from the fresh or dried plant for the manufacture of medicinal products for the treatment or prevention (protection) of radiation (radio prevention, protection), infection (infenion prophylaxis) and chemically-related organ and tissue damage (chemoprevention, protection), in particular of the O-MALT system of the small intestine (inflammatory diseases, regional enteritis) Crohn), the lungs (inflammation, disease), the bronchi and the nasopharynx (rhinopharyngeal bronchitis, thinitis, pharyngitis, sinusitis), the bone marrow (bone marrow aplasia), the thymus udysfunction, aplasia or hypoplasia due to medication or radiation-related damage), the liver (atrophy, necrosis), hepatitis A, B and C, the pancreas, (insufficiency of the exocrine, secretory Chen function of proteases, esterases, carbohydrases and nucleases as well as the endocrine dysfunction of the carbohydrate metabolism (Langemhans Islands) and the kidneys (insufficiency) as well as of generally immunosuppressed conditions, for cellular immune stimulation, for the therapy of leukocytopenia, granulocytopenia, thrombocytopenia, never lymphocytopenia, lymphocytopenia, never lymphocytopenia, lymphocytopenia, never (Infectious, tumor anemia, etc.) and in immunoglobulin deficiency states, also as a result of AIDS, tumors and diseases with suppressed immune systems.

2. Use according to claim 1, characterized in that the cellular immune stimulation (paramunization), in particular after an immunosuppression by radiation or chemotherapy treatments, ie the increase in the number of cells of leukocytes, segmented granulocytes and lymphocytes, T-, B-, helpers -, suppressor and NK cells, in particular of B lymphocytes in the terminal current pathway, cause an increase in vital lymphocytes in the bark zone and the follicles in the Peyer plaques (small intestine) to be recognized, which is associated with a secretory induction of sIGA in the small intestine is connected to the fact that an increase in all blood formation elements in the bone marrow is detectable 66

cellular stimulation of immunocompetent cells in the thymus, spleen and mesenteric lymph nodes is detectable and / or that the metabolic functions of the liver, pancreas and kidneys (lowering of the serum values of creatinine, uric acid, glucose, bilirubin, cholesterol and trigiycerides as well as GOT, GPT and GGT) are clearly evident to be activated.

3. Use according to claim 1 and 2, characterized in that the cellular immune stimulation (paramunization) an increase in the cell count of leukocytes, segmented granulocytes, lymphocytes, T, B, helper, suppressor and NK cells in the bone marrow increased Tendency to form cell-rich nests of granulo-, erythro-, lympho- and thrombopoiesis, however, a decrease in the number of mature pithy and / or toroidal myelocytes, in the thymus an increase in lymphatic cell elements in the cortex and medulla, in the spleen to widening of the marginal zones, particularly in the area of the periarterial vagina and in the mesenteric lymph nodes, includes an increase in lymphocytic cell elements in the B and T lymphocyte areas and the medullary strands.

4. Use of AU, BP, EA and UD extracts according to claim 1, as high-dose phytopharmaceuticals, in particular in one

Dose for AU-Extrak 600-5400 mg / day «2-3 x 1 to 6 drg. Ä 300 mg native extract dose for BP extract 600-5400 mg / day ^ 2-3 x 1 to 6 drg. Λ 300 mg native extract dose for EA ax extract 600-5400 mg / day * 2-3 x 1 to 6 Drg, ä 300 mg native extract extract for US extract 600-5400 mg / day * 2-3 x 1 to 6 Drg, a 300 mg native extract, also in combination with

Artichoke (Cynara) extract (CY) 600-5400 mg native extract / day Mistletoe (Viscum) extract (VI) 300-3600 mg native extract / day, St. John's wort (Hypericum) extract (HY) 300-3600 mg native extract / Day Putamen ovi (eggshell components of the pallisade and mammill zone 600-5400 mg / day; the extracts AU, BP, EA, US, CY, VI and HY have multiantigenic effects after oral administration and consequently lead to paramunization of the organism (non-specific stimulation) find them as paramunity inducers with antiviral, antibacterial 67

and anti-tumor effects use.

5. Use according to claim 1 or 2 for the treatment of disorders of carbohydrate metabolism and liver and kidney function, zuer treatment

5 of prediabetic, especially senile forms for the treatment of infections in combination with chemically defined anti-diabetic agents.

6. Use according to claim 1 or 2 as oral adjuvants of malignant tumor treatment, in particular the treatment of breast, portio, o colon or prostate cancer, also in combination with cystostatics, in particular cyclophosphamide or radiation therapy, and the treatment of AIDS and diseases with suppressed immune system.

7. Use according to claim 1 to increase the tolerance 5 of chemo-, cytostatic, radiological therapy, in particular in combination with chemically defined substances, selected from cytostatic agents, such as cisplatin, cyclophosphamide, methotrexate, fluorouracil, bleomycin and / or clofibrate, ACE inhibitors , α-2 receptor antagonists, Ca antagonists and / or diuretics, as an adjuvant for analgesia, in particular as a pharmacological component of the positive influence on the interactions between the endocrine, nervous and immune systems.

8. Use according to claim 1 or 2 as an adjuvant in the treatment of bacterially and virally induced clinical pictures, in particular inflammatory diseases of the small intestine (Enteritis regionalis Crohn), the lungs, the pancreas and the kidneys, liver atrophy, hepatitis A, B and C, Skin lesions (leg ulcers), as well as herpes simplex I and II and herpes zoster, AIDS.

9. Use according to one or more of claims 1 to 7 as a fresh or dried plant, in particular pressed juice or in extracted form 0 with the aid of aqueous, organic and / or supercritical solvents, in particular carbon dioxide; in dried form, in particular powder, dry extracts and the like of the fresh plant or drug (dried plant, granules, in particular 68

as a capsule and as a tablet and as a dragee or lozenge.

10. Use according to one or more of claims 1 to 8, characterized in that AU, BP, EA and UD extracts are preferably used with a drug / extract ratio (DEV native) of 4-9: 1.

11. Use according to one of the claims 1 to 10, characterized in that the AU, BP, EA and UD extracts are combined individually or with one another or in each case in combination with Cynara and / or mistletoe (Viscum) extracts and / or with Echinacea extracts, furthermore with lion number (Taraxacum) extracts as well as with Putamen ovi preparations based on eggshell components, in particular the central pallisade and mammalian zone, optionally in combination with chemically defined chemotherapeutic agents, antibiotics, diuretics, cytostatics to inhibit bone demineralization and / or depression for the treatment of malignant tumors, in particular the treatment of primary and secondary bone tumors, breast, protio, colon or prostate cancers, as well as rumora anemia and infectious anemia.

12. Orally administrable medicament containing AU, BP, EA and UD dry extracts as defined in one or more of claims 1 to 11 in

Calcium carbonate and saccharide-containing granules.