DE102015012685A1 - Method for supporting intestinal health - Google Patents

Abstract

Calcium pyruvate (CaPyr) and derivatives or formulations thereof can be used to aid intestinal health and reduce inflammation, such as in chronic inflammatory diseases of the digestive tract and irritable bowel syndrome. Therefore, the invention relates to the use of pyruvate preparations for the treatment of inflammatory diseases, in particular the gastrointestinal tract of humans and mammals, domestic animals and livestock. The digestive tract includes the oral cavity, pharynx, esophagus, and gastrointestinal tract. The inflammatory diseases include irritable bowel syndrome (IBS) and / or inflammatory bowel disease (IBD), but also inflammatory diseases of the group of gingivitis, periodontitis, pharyngitis, esophagitis, gastritis, lymphatic colitis, Ulcerative colitis, diverticulitis, duodenitis and Crohn's disease were understood.

Description

The invention relates to a method for oral ingestion or supplementation of a food, a dietary supplement or a pharmaceutical composition comprising an active ingredient, preferably calcium pyruvates and especially calcium pyruvate monohydrate (CPM) or bioactive functional derivatives and combinations or formulations thereof, and especially of the same Soothing and health-promoting effect for the digestive tract, especially the intestine.

Optimal health of the digestive system is important for vitality and well-being throughout life, and has implications for the health of the whole body, while gastrointestinal disorders severely affect quality of life.

Symptoms of gastrointestinal distress include flatulence, changes in bowel mobility, stool transfer time, and digestive tract pain that can affect well-being. Gastrointestinal complaints may be caused by stress, food composition, microbial distribution or intestinal disorders that cause serious gastrointestinal disorders, including dyspepsia or irritable bowel syndrome (IBS). or chronic inflammatory bowel disease (IBD).

About 20 percent of people, especially in the Western world, are affected by prolonged gastrointestinal problems, such as irritable bowel syndrome, at least once in their lives.

Many gastrointestinal diseases are caused by inflammatory processes, so that the promotion of a healthy digestive process, in particular by anti-inflammatory nutrition or therapeutic approaches that include a reduction in gene expression of inflammatory mediators, can be achieved.

Standard therapeutic procedures include drugs such as corticosteroids (cortisone, prednisone) and aminosalicylic acid derivatives such as sulfasalazine (sulphasalazine) as well as immunomodulators, such as: Azathioprine, methotrexate or natalizumab. These drugs have anti-inflammatory effects, but some have many serious side effects, limiting their use in acute exacerbations. New medicines for reducing inflammation are u. a. of an antioxidant character, such as ethylpyruvate, a synthetic alkyl ester of pyruvic acid.

Therefore, it is important to provide functional food ingredients and nutritional supplements with similar modes of action but less harmful side effects besides medications.

Alternative medical and nutritional concepts include nutrition with probiotics (Escherichia coli Nissle 1917, Saccharomyces boulardii, bifidobacteria and lactic acid bacteria), prebiotics (barley, opuntia fibers, psyllium, oligofructose, inulin, modified starches, botanical or microbiotic derived gums), fish and krill oils (Omega-3 fatty acids) and herbal preparations (frankincense (Boswellia spp.), Turmeric (Curcuma longa), thyme spp., Chamomile, cannabis), essential and aromatic oils such as. Peppermint, chamomile, sage, fennel, cumin, thyme and lavender, and a range of nutritional supplements.

The focus and thus the object of the present invention is to provide new active substances for the prevention and support of functional gastrointestinal complaints and diseases, such as inflammatory bowel disease and RDS, which are used as an ingredient u. a. in functional foods, nutritional supplements and medicines.

Surprisingly, it has been found that a composition containing as active ingredient salts of pyruvic acid (pyruvates) and in particular calcium pruvate (CaPyr), but especially calcium pyruvate monohydrate (CPM), or functionally active derivatives, anti-inflammatory and RDS-alleviating activity by inhibiting the inflammation at the molecular and gene expression level.

Alpha-ketocarboxylic acids, including pyruvic acid (2-oxo-propanoic acid, pyruvic acid) and their derivatives, such as esters and salts, also called pyruvates, are known for their anti-inflammatory activity (US Pat. Fink MP (2008). Ethyl pyruvate. Current opinion in anesthesiology 21 (2): 160-167 ).

Pyruvic acid and pyruvates are reactive chemical compounds which tend to dimerize or undergo polymerization by aldol condensation. This results in the general understanding that the salt form with free pyruvate anions is far less stable than the ester form. Pyruvate salts are said to decompose much faster than pyruvate esters, which has led to the recent development of pyruvic acid esters, such as pyruvate esters. For example, ethyl pyruvate has been used much more frequently in clinical research, and the salts have been more and more neglected ( Fink MP (2008). Ethyl pyruvate. Current opinion in anesthesiology 21 (2): 160-167 ).

On the other hand, even simple pyruvic acid esters, such as ethylpyruvate, are absent in nature, or only in trace amounts, as aroma components and are thus of synthetic and non-physiological character, while salt-form pyruvates and pyruvate anions, as sodium and calcium pyruvate, are prevalent in living organisms because pyruvates (in biochemistry synonymous with pyruvic acid) are intermediates of sugar metabolism and glycolysis.

Surprisingly, it has been found in the context of this invention that the pyruvate salts of the alkali and alkaline earth metal groups (Ia and IIa) and in particular the calcium pyruvate monohydrate, in contradiction to the abovementioned and prevailing scientific opinion, are capable of an even better anti-inflammatory action exercise as pyruvate ester.

This greater anti-inflammatory activity of the pyruvate salts over the pyruvate esters became apparent on a chemically induced animal colitis model. The experimental approach was to determine the effect of calcium pyruvate compared to ethyl pyruvate at doses of 20, 40 and 100 mg / kg body weight after a 7-day oral treatment on the trinitrobenzenesulfonic acid (TNBS) colitis model in rats, particularly for gut histology and existing inflammatory mediators. As a key finding, both forms of pyruvate have been shown to have intestinal anti-inflammatory activity in TNBS-induced colitis. Both pyruvate forms histologically improved the inflamed tissue by restoring mucosal cell architecture as well as reducing neutrophil infiltration and inflammatory mediators (IL-1, IL-6, IL-17, IL-23, iNOS). However, calcium pyruvate, and especially the monohydrate (CPM), appears to be more effective than ethyl pyruvate and even better than the therapeutic drug sulfasalazine, as demonstrated by Examples 1b-1g.

Noteworthy in this context is the positive effect of calcium pyruvate on the mediators IL-17 and IL-23, which play a central role in the development of inflammatory bowel processes (Example 1e).

Basically, the animal experiments were carried out analogously, as described in detail in the publication "Intestinal anti-inflammatory activity of the Serpylli herba extract in experimental models of rodent colitis." Galvez J et al .; J Crohn's Colitis. 2014: 775-88 , is described.

In addition, a relationship between inflammatory bowel disease (IBD) and irritable bowel syndrome (RDS) has recently been seen. Since most people with RDS appear to be physically healthy, a psychosomatic cause has been considered in the past, but recently, "mini-inflammation" has been detected in the gut of people with RDS, and neuronal communication disorders in the intestinal neoplasm are now suspected and added as a cause discussed. These sites of inflammation bring RDS into closer association with CED, which could thus facilitate an anti-inflammatory treatment of the irritable bowel similar to the CED (1. Bercik P, Verdu EF, Collins SM. Is irritable bowel syndrome a low-grade inflammatory bowel disease? Gastroenterol Clin North Am. (2005) Vol. 34, Issue: 2: 235-245 // 2. Mearin F, Perello A, Balboa A; Irritable bowel syndrome and inflammatory bowel disease: Is there a connection? Gastroenterology and Hepatology (2009) Volume: 32, Issue: 5, Pages: 364-372 // 3. Quigley EMM, Shanahan F; Irritable Bowel Syndrome and Inflammatory Bowel Disease: Is There an Overlap? Practical Gastroenterology, November 2010, 31-37 ).

Thus, the effects of various doses of calcium pyruvate (CaPyr) and gabapentin, a drug for RDS and as a control, are positively evaluated in an experimental irritable bowel rat model initiated by intracolonic deoxycholic acid (DCA) administration. Induced abdominal reflexes, the so-called "colorectal distension (CRD)", radiative abdominal pain by Von Frey filaments and gene expression of various markers in the colon tissue by qPCR of IL-1β and COX-2, which proved remarkably effective calcium pyruvate (Example 2 ).

Although the experimental findings of this invention show an obvious positive effect of calcium pyruvate monohydrate on inflammatory gastrointestinal processes, derivatives of calcium pyruvate, e.g. B. other hydrates (n = 0-5 × H ₂ O) and useful combinations and formulations, with known alternative medicines and nutritional approaches, such as diets, probiotics (Escherichia coli Nissle 1917, Saccharomyces boulardii, bifidobacteria and lactic acid bacteria), prebiotics (plant fibers, Psyllium, oligofructose, polydextrose, fructooligo saccharides (FOS), xylooligosaccharides (XOS), galactooligosaccharides (GOS) and mannooligosaccharides (MOS) as well as lactulose and tagatose, inulin, modified starches, botanical or microbiotic derived gums), fish and krill oils (Omega 3-fatty acids) and herbal preparations (frankincense (Boswellia serrata), turmeric (Curcuma longa), thyme spp., Chamomile, cannabis), essential and aromatic oils such as those of peppermint, chamomile, sage, fennel, caraway, thyme and lavender -Oil. Combinations with certain dietary supplements can contribute to the effect or even have a synergistic effect.

In summary, it should be noted that the solution to the stated object of the present invention is therefore to provide a novel functional and sufficiently stable active substance, which can be applied for use in inflammatory diseases throughout the digestive tract in mammals, preferably in humans. Anti-inflammatory effects along the entire digestive tract are just as helpful as the treatment or prevention of inflammatory diseases of individual intestinal sections. Gingivitis, periodontitis, pharyngitis, esophagitis and gastritis describe the inflammatory diseases of the upper part of the digestive tract. Lymphatic colitis, ulcerative colitis, diverticulitis, duodenitis and Crohn's disease are, in addition to CED and RDS, the most prominent representatives of lower intestinal inflammatory diseases.

It should be emphasized that this invention is not limited only to human use, but applies to the group of mammals, especially domestic animals, livestock and livestock.

Surprisingly, it has been found that calcium pyruvate (CaPyr), in particular its monohydrate (CPM), has sufficient stability, and act as active ingredients with antioxidant, antiinflammatory properties. These are based on molecular mechanisms in the inflammatory area, including downregulation of gene expression, which also applies to the mediators IL-17 and IL-23.

The invention is further illustrated by the following non-limiting examples.

embodiments

Example 1a:

TNBS colic model in rats and effects of various doses of calcium pyruvate (CaPyr), ethylpyruvate (EtPyr) and sulfasalzine (SAZ) on macroscopic damage score on rat intestine, weight / length ratio, myeloperoxidase (MPO) Activity and glutathione (GSH) concentration in the experimental 2,4,6-trinitrobenzenesulfonic acid (TNBS) -induced colitis rat model.

All of these studies were conducted in accordance with guidelines for the care and use of laboratory animals, according to the National Institute of Health.

Female Wistar rats (180-200 g) were randomized and assigned to nine groups (n = 10). Three groups received treatment with CaPyr (20, 40 and 100 mg / kg); the other three received treatment with ethylpruvate (20, 40 and 100 mg / kg); and the remaining groups were control groups: non-colitis (natively, completely untreated), treated only with TNBS (control) or with sulfasalazine (SAZ) as standard (30 mg / kg). All compounds were dissolved in 1 ml of water with carboxymethylcellulose (0.2%) and administered daily by oral gavage. An untreated TNBS control group and a non-colitis group were included as reference groups which received only the carrier solution but not the test compounds. The method for initiating the column ignition and further details were in the above publication " Galvez et al. 2014 "Described in detail. Brief Description: Overnight fasting rats were anesthetized with halothane and 0.25 ml of an ethanolic TNBS solution was administered via a teflon cannula inserted 8 cm through the anus. During and after TNBS administration, the rats were kept in a head position until recovering from anesthesia, then returned to their cages. Rats from the non-colitis control group were given 0.25 ml of phosphate buffered saline instead of TNBS into the intestinal lumen. The treatments were performed from day of colitis induction to seven days later, then the rats were sacrificed with halothane overdose. In all attempts Daily was the award of the weight of the animals, the occurrence of diarrhea and the water and food intake. The colon was removed aseptically from the dead animals and placed on an ice-cold plate, then opened longitudinally and freed from the contents with cold saline. Thereafter, the colon was weighed and its length measured under a constant load of 2 grams. Each colon was macroscopically examined by two panelists for visible tissue damage using a scale of 0-10 to specified criteria (analog Galvez et al. 2014 ). Colon samples (0.5 cm ² ) containing all tissue layers were taken from the area of the inflamed colon and adjacent segments and fixed in 4% buffered formaldehyde for histological evaluation of macroscopic damage. Corresponding colon segments were also obtained from the non-colitis group for comparison. The colon preparations were then finely minced, divided and stored frozen at -80 ° C until the measurement of biochemical parameters and RNA extraction was performed.

For histological studies, suitable cross sections were selected and embedded in paraffin. Sections of 5 μm thickness in different planes were obtained and stained with hematoxylin and eosin. The histological damage was evaluated by a pathologist who was blinded to the experimental groups according to the criteria described by Galvez.

For the biochemical parameters in the colon tissue, the myeloperoxidase (MPO) activity was determined according to a described method ( Krawisz et al., 1984 ) measured; the results were reported as MPO units per gram of fresh tissue; One unit of MPO activity is defined as the decomposition of 1 μmol of hydrogen peroxide / min at 25 ° C. The total glutathione (GSH) content was compared to that of Anderson (1985) quantified and the results were expressed as nmol / g wet tissue. To assess tissue IL-1β levels, the colon samples were assayed in phosphate buffered saline containing 0.1% sodium dodecyl sulfate (SDS), 01% sodium deoxycholate, 1% Triton X-100, protease and phosphatase inhibitors (aprotinin, leupeptin, and Phenylmethyl-sulfonyl fluoride) homogenized. The cytokine was quantitated by a corresponding ELISA test (R & D Systems Inc., Minneapolis, MN, USA) and the results expressed as pg / g fresh tissue.

The analysis of gene expression in the colon samples was carried out by RT-qPCR, the total RNA from the colon samples with Trizol ^® (Thermo Fisher Scientific Inc., Waltham, MA USA) was isolated according to the manufacturer's protocol. All RNA samples were quantitated with the Thermo Scientific NanoDrop ^™ 2000 Spectrophotometer (Thermo Fisher Scientific Inc., Waltham, MA USA) and 2 μg RNA was reverse transcribed using oligo (dT) primers (Promega, Southampton, UK).

The real time quantitative PCR (RT qPCR) amplification and detection was performed on 48-well plates in a EcoTM Real-Time PCR System (Illumina, CA, USA) with 20 ng cDNA and the KAPA SYBR ^® FAST qPCR Master Mix (Illumina, CA, USA) and specific primers at their annealing temperatures (Ta). To normalize mRNA expression, the expression of housekeeping genes and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were measured. The relative mRNA quantification was calculated by the ΔΔCt method.

In terms of statistics, all results are expressed as mean ± standard deviation (mean ± SEM). Differences between the mean values were checked for statistical significance using one-way variance analysis (ANOVA), post-hoc significance tests, Chi-square test, and Tukey's post-hoc test, respectively. Nonparametric data (macroscopic and microscopic scores) were analyzed according to the Kruskal-Wallis test. All statistical analyzes were performed with GraphPad Prism Version 5.0 (GraphPad Software Inc., La Jolla, Calif., USA) and statistical significance fixed at p <0.05.

Example 1b:

Effects of various doses of calcium pyruvate (CaPyr), ethylpyruvate (EtPyr), and sulfasalzine (SAZ) on macroscopic colonic damage score scale, weight-to-length ratio, myeloperoxidase (MPO) activity, and glutathione (GSH) concentration in experimentally-induced TNBS colitis in rats (Example 1a).

Table 1b Group with N = 10 Damage Points (0-10) Weight / length (mg / cm) MPO (mU / g tissue) GSH (nmol / g tissue) Non-colitis 0 80.3 ± 4.0 2.0 ± 0.3 1488 ± 141 TNBS control 7.9 ± 0.2 201.8 ± 15.2 83.8 ± 7.7 403 ± 58 CaPyr (20 mg / kg) 7.1 ± 0.4 186.0 ± 22.8 48.2 ± 7.2 * 389 ± 55 CaPyr (40 mg / kg) 5.7 ± 0.4 * 184.6 ± 25.1 48.8 ± 8.2 * 675 ± 67 * CaPyr (100 mg / kg) ^{6 , 6 ± 0 , 6 *} 198.9 ± 15.0 57.7 ± 11.5 * 768 ± 100 * EtPyr (20 mg / kg) 6.6 ± 0.6 * 185.4 ± 31.1 58.1 ± 11.9 560 ± 74 EtPyr (40 mg / kg) 6.1 ± 0.4 * 172.0 ± 15.8 61.5 ± 9.8 715 ± 88 * EtPyr (100 mg / kg) 6.0 ± 0.5 * 186.4 ± 13.2 54.7 ± 10.8 * 870 ± 124 * SAZ (30 mg / kg) 6.7 ± 0.4 * 180.4 ± 16.9 82.8 ± 8.2 900 ± 118 * Data means mean ± standard deviation (mean ± SEM). * P <0.05 vs. TNBS control group. All colitis groups differ statistically from the non-colitis group (P <0.05).

Example 1c:

Effects of different doses of calcium pyruvate (CaPyr), ethylpyruvate (EtPyr) and sulfasalzine (SAZ) on the microscopic damage scale according to Arribas et al. on the experimental TNBS rat model of colitis (Example 1a).

Table 1c Group with N = 10 Microscopic damage (0-59) Non-colitis 0 TNBS control 35 ± 3 CaPyr (20 mg / kg) 27 ± 2 CaPyr (40 mg / kg) 18 ± 5 * CaPyr (100 mg / kg) 13 ± 2 * EtPyr (20 mg / kg) 25 ± 3 EtPyr (40 mg / kg) 17 ± 3 * EtPyr (100 mg / kg) 23 ± 3 * SAZ (30 mg / kg) 30 ± 5 Data means mean ± standard deviation (mean ± SEM). * P <0.05 vs. TNBS control group.

Example 1d:

Effects of different doses of calcium pyruvate (CaPyr), ethylpyruvate (EtPyr), and sulfasalzine (SAZ) on IL-1β concentration in ng / g fresh tissue as measured by ELISA assay and on mRNA expression of IL-1β, IL-6 and IL-12, quantified by real-time PCR and plotted as a fold increase on the experimental TNBS rat colitis model (Example 1a).

Table 1d Group (n = 10) IL-1β (ng / g tissue) IL-1β (fold increase) IL-6 (fold increase) IL-12 (fold increase) Non-colitis 5.4 ± 0.6 1.3 ± 0.03 1.01 ± 0.1 1.02 ± 0.06 TNBS control 1140.0 ± 90.1 9.00 ± 1.20 5.81 ± 0.25 2.34 ± 0.42 CaPyr (20 mg / kg) 1095.0 ± 165.0 3.90 ± 0.61 * 5.41 ± 1.01 1.09 ± 0.13 CaPyr (40 mg / kg) 765.0 ± 60.0 * 2.40 ± 0.61 * 3.20 ± 1.20 * 0.66 ± 0.18 * CaPyr (100 mg / kg) 810.1 ± 120.0 * 4.80 ± 0.90 * 4.61 ± 1.01 1.62 ± 0.30 EtPyr (20 mg / kg) 975.0 ± 90.1 6,90 ± 1,50 5.60 ± 1.41 1.09 ± 0.13 EtPyr (40 mg / kg) 1140.0 ± 150.1 7.20 ± 1.20 5.60 ± 1.20 1.81 ± 0.48 EtPyr (100 mg / kg) 975.0 ± 211.7 10.21 ± 1.50 6.00 ± 1.01 1.62 ± 0.37 SAZ (30 mg / kg) 840.0 ± 120.0 * 9.70 ± 0.90 4.21 ± 0.80 * 1.59 ± 0.24 Data mean mean ± standard deviation (mean ± SEM). * P <0.05 vs. TNBS control group.

Example 1e:

Effects of different doses of calcium pyruvate (CaPyr), ethyl pyruvate (EtPyr) and sulfasalzine (SAZ) on mRNA expression of IL-17, IL-23 and iNOS, quantified by real-time PCR and plotted as relative increase (fold increase) on the experimental TNBS rat model of colitis (Example 1a).

Table 1e Group (n = 10) IL-17 (fold increase) IL-23 (fold increase) iNOS (fold increase) Non-colitis 1.02 ± 0.03 0.99 ± 0.06 1.22 ± 0.06 TNBS control 2.16 ± 0.30 2.37 ± 0.27 19.82 ± 2.14 CaPyr (20 mg / kg) 0.84 ± 0.09 * 1.47 ± 0.24 * 10.37 ± 1.83 * CaPyr (40 mg / kg) 0.63 ± 0.27 * 1.02 ± 0.21 * 8.85 ± 3.36 * CaPyr (100 mg / kg) 1.83 ± 0.48 1.98 ± 0.30 9.15 ± 1.53 * EtPyr (20 mg / kg) 1.17 ± 0.27 * 1.68 ± 0.39 16.17 ± 2.75 EtPyr (40 mg / kg) 1.74 ± 0.33 1.56 ± 0.27 * 12.20 ± 1.53 * EtPyr (100 mg / kg) 2.34 ± 0.30 1.80 ± 0.21 22.88 ± 2.14 SAZ (30 mg / kg) 2.22 ± 0.54 1.26 ± 0.18 * 16.47 ± 2.75 Data means mean ± standard deviation (mean ± SEM). * P <0.05 vs. TNBS control group.

Example 1f:

Effects of different doses of calcium pyruvate (CaPyr), ethyl pyruvate (EtPyr) and sulfasalzine (SAZ) on mRNA expression of CINC-1, MCP-1 and ICAM-1, quantified by real-time PCR and plotted as relative change (fold increase) on the experimental TNBS rat model for colitis (Example 1a).

Table 1f Group (n = 10) CINC-1 (fold increase) MOP-1 (fold increase) ICAM-1 (fold increase) Non-colitis 1.06 ± 0.04 0.98 ± 0.04 1.07 ± 0.01 TNBS control 5.46 ± 0.49 3.20 ± 0.21 4.79 ± 0.44 CaPyr (20 mg / kg) 3.59 ± 0.49 * 3.03 ± 0.49 1.64 ± 0.25 * CaPyr (40 mg / kg) 2.93 ± 0.57 * 1.80 ± 0.37 * 0.95 ± 0.19 * CaPyr (100 mg / kg) 3.50 ± 0.90 * 2.13 ± 0.41 * 2.77 ± 0.44 * EtPyr (20 mg / kg) 4.40 ± 0.65 1.23 ± 0.37 * 1.39 ± 0.19 * EtPyr (40 mg / kg) 4.40 ± 0.57 1.85 ± 0.29 * 2.27 ± 0.38 * EtPyr (100 mg / kg) 4,40 ± 0,90 2.21 ± 0.29 * 2.39 ± 0.50 * SAZ (30 mg / kg) 4.89 ± 0.82 2.62 ± 0.41 * 2.90 ± 0.88 * Data means mean ± standard deviation (mean ± SEM). * P <0.05 vs. TNBS control group.

Example 1g:

Effects of different doses of calcium pyruvate (CaPyr), ethylpyruvate (EtPyr), and sulfasalzine (SAZ) on mRNA expression of MUC-2 and TFF-3, quantified by real-time PCR and plotted as relative increase (fold increase) on experimental TNBS rat model for colitis (Example 1a).

Table 1g Group (n = 10) MUC-2 (fold increase) TFF-3 (fold increase) Non-colitis 1.02 ± 0.08 0.99 ± 0.05 TNBS control 0.26 ± 0.02 0.36 ± 0.05 CaPyr (20 mg / kg) 0.50 ± 0.14 * 0.84 ± 0.23 * CaPyr (40 mg / kg) 0.53 ± 0.16 * 0.67 ± 0.17 * CaPyr (100 mg / kg) 0.56 ± 0.11 * 0.99 ± 0.19 * EtPyr (20 mg / kg) 0.39 ± 0.09 0.59 ± 0.16 EtPyr (40 mg / kg) 0.70 ± 0.08 * 1.01 ± 0.05 * EtPyr (100 mg / kg) 0.50 ± 0.11 * 0.96 ± 0.11 * SAZ (30 mg / kg) 0.53 ± 0.14 * 0.91 ± 0.12 * Data means mean ± standard deviation (mean ± SEM). * P <0.05 vs. TNBS control group.

Example 2a:

The effects of calcium pyruvate monohydrate (CPM) in an experimentally induced irritable bowel model in rats by intracolonic administration of deoxycholic acid (DCA) were determined as follows. All of these studies were conducted in accordance with guidelines for the care and use of laboratory animals, according to the National Institute of Health.

Male Sprague-Dawley rats (240-320 g) were administered once daily DCA on three consecutive days and then distributed to the various experimental groups (n = 8): three groups received daily oral calcium pyruvate monohydrate (40 and 100 mg / kg) or gabapentin (70 mg / kg); As a control, a non-RDS group and an untreated RDS group were classified. One and two weeks afterwards the abdominal distension was corrected by colorectal distension ( CRD; La et al., World J Gastroenterol, 2008; 14: 731-6 ) determined semi-quantitatively. In addition, the radiating pain was scored with "von Frey filaments". After these two weeks of treatment, all rats were sacrificed, and the expression of various markers (IL-1β and COX-2) in the colon tissues assessed by qPCR. Table 2 Group (n = 8) CRD (units) AUC (units) qPCR IL-1β (fold increase) qPCR COX-2 (fold increase) Non-RDS 0.14 ± 0.0 ^a 54.55 ± 19.93 ^a 1.15 ± 0.23 ^a 1.02 ± 0.15 ^a RDS Kontrollel 2.21 ± 2.5 ^b 18883 ± 2730 ^b 1978 ± 345b 5.00 ± 1.16 ^b CPM (40 mg / kg) 1.30 ± 0.14 ^c 104.65 ± 20.48 ^a 7.82 ± 2.07 ^c 2.47 ± 0.58 ^c CPM (100 mg / kg) 0.70 ± 0.14 63.70 ± 22.75 ^a 5.52 ± 1.15 ^c 1.60 ± 0.36 ^c Gabapentin (30 mg / kg) 0.25 ± 0.1 ^a 27.30 ± 6.83 ^a 8.28 ± 1.38 ^c 1.74 ± 0.36 ^c Data means mean ± standard deviation (mean ± SEM). * P <0.05 vs. Control or further group (with different letters).

As results, the treated groups showed more reduced CRD scores after one or two weeks than the RDS control. Also, the radiating pain in these groups with CPM and gabapentin treatment was reduced. Induced irritable bowel syndrome was associated with altered expression of the various markers studied and the treatments resulted in partial normalization of IL-1β and COX-2 expression by CPM.

In conclusion, calcium pyruvate monohydrate shows very beneficial effects on the experimental RDS model, which has similar efficacy to standard Gabapentin therapy. This effect was also clearly associated with an improvement in the altered immune response in RDS.

Example 3: Formulations

Example 3a)

Preparation of a water-soluble preparation for use in drinks

Liquid formulations prepared from a powder mixture serve to provide and orally administer from 20 mg to 5,000 mg per serving of pyruvic acid salts, especially calcium pyruvate. The powder preparations are completely water-soluble and can be added to drink concentrates, sodas, dietetic foods and dairy products in concentrations ranging from 1 to 10% and drunk. The pyruvate salts are mixed with prebiotic or non-digestible carbohydrates and fillers such as maltodextrin, fructose or others, ideally sugar free, fully water soluble, with low glycemic indexes and hydrolyzed proteins (eg gelitasol etc.). Thus, the above-mentioned substances are used as adjuvants, and these preparations can also be used in food products, dietary products, dietetic foods for special medical purposes and drinks for inflammatory bowel disease or irritable bowel. One to five, especially two to three servings per day (20 mg to 25 g / day) are recommended depending on the mammal species, the type of disease and its severity.

Example 3b)

Pektinpastillen

40 grams of commercial citrus pectin and 2 grams of trisodium citrate are mixed with 100 grams of isomalt sugar substitute. This mixture is stirred into 200 ml of water and heated with constant stirring until all the pectin has dissolved. Another 475 g of isomalt and 260 g of fructose syrup are added and the mixture boiled to about 80% dry matter content. Now, 5-50 g of pyruvate of an alkali metal (eg lithium, sodium, potassium) and / or an alkaline earth metal (eg magnesium, calcium) and / or an organic base (eg creatine, carnitine) as well as Added color and flavorings. To adjust the pH to 3.4-3.5, add about 17 ml of a 50% citric acid solution. This mixture is poured at a temperature of about 95 ° C in such molds that 2 g pastilles are obtained. You can make about 500 lozenges of two grams in this way. A pectin paste contains about 10-100 mg of the corresponding pyruvate salts.

Example 3c)

Pastilles based on gum arabic

In a stirring apparatus 15 kg of solution with 33% gum arabic, 5% sorbitol, 20% water and 42% maltitol solution at 65 ° C are prepared. To this solution are added 50-450 grams of pyruvate of an alkali metal (e.g., lithium, sodium, potassium) and / or an alkaline earth metal (e.g., magnesium, calcium) and / or an organic base (e.g., creatine, carnitine ) and 5-50 grams of ethyl pyruvate and flavored flavors and (if necessary) sweetener. After complete homogenization, the mixture is poured into starch molds. After drying at 50 ° C, the final weight of the lozenges is adjusted to 2.0 g containing about 10-100 mg of pyruvate. The pastilles are separated from the starch, treated with a release agent and bottled. There will be about 5000 lozenges.

Example 3d)

chewing gum

100 g of chicle are powdered, mixed with 300 g of sugar substitute isomalt and heated in an evaporating dish until the mass softens. 5 g of a mixture of calcium pyruvate monohydrate and a pyruvate of an alkali metal (eg lithium, sodium, potassium) and / or an alkaline earth metal (eg magnesium, calcium) and / or an organic base (e.g. Creatine, carnitine), the mixture is worked through thoroughly, whereby aromas could be added, then brought to a starch-strewn surface and kneaded to homogeneity. The mixture is then rolled out into thin sheets and cut into flat bars while still warm. Using some starch powder prevents the mixture from sticking to the surface. The chewing gum sticks should weigh 2 grams and contain about 25 mg of pyruvate.

Example 3e)

Preparation of a pharmaceutical tablet composition

The formulations of the invention may also be provided as a tablet. For this purpose, a mixture of a defined amount of calcium pyruvate monohydrate and suitable tabletting excipients is prepared, as shown for example with the following recipe and subjected to direct tabletting
300 mg calcium pyruvate monohydrate
160 mg of microcrystalline cellulose
25 mg sodium carboxymethylcellulose
10 mg fumed silica
5 mg magnesium stearate

Example 3f)

Preparation of a pharmaceutical hard gelatin capsule composition

The formulations according to the invention can also be provided in capsule form, for which purpose a defined amount of a mixture of calcium pyruvate monohydrate and the pyruvate of an alkali metal (eg lithium, sodium, potassium) and / or an alkaline earth metal (eg magnesium, calcium) and / or an organic base (eg, creatine, carnitine) and suitable excipients, as shown for example in the recipe below, and filled into a hard gelatin rinse. The quantities refer to a capsule dosage.
100 mg calcium pyruvate monohydrate
100 mg sodium pyruvate
50 mg of microcrystalline cellulose
2 mg of fumed silica

QUOTES INCLUDE IN THE DESCRIPTION

This list of the documents listed by the applicant has been generated automatically and is included solely for the better information of the reader. The list is not part of the German patent or utility model application. The DPMA assumes no liability for any errors or omissions.

Cited non-patent literature

• Fink MP (2008). Ethyl pyruvate. Current opinion in anesthesiology 21 (2): 160-167 [0011]
• Fink MP (2008). Ethyl pyruvate. Current opinion in anesthesiology 21 (2): 160-167 [0012]
• "Intestinal anti-inflammatory activity of the Serpylli herba extract in experimental models of rodent colitis." Galvez J et al .; J Crohn's Colitis. 2014: 775-88 [0017]
• Bercik P, Verdu EF, Collins SM. Is irritable bowel syndrome a low-grade inflammatory bowel disease? Gastroenterol Clin North Am. (2005) Vol. 34, Issue: 2: 235-245 [0018]
• Mearin F, Perello A, Balboa A; Irritable bowel syndrome and inflammatory bowel disease: Is there a connection? Gastroenterology and Hepatology (2009) Volume: 32, Issue: 5, Pages: 364-372 [0018]
• Quigley EMM, Shanahan F; Irritable Bowel Syndrome and Inflammatory Bowel Disease: Is There an Overlap? Practical Gastroenterology, November 2010, 31-37 [0018]
• Galvez et al. 2014 [0026]
• Galvez et al. 2014 [0026]
• Krawisz et al., 1984 [0028]
• Anderson (1985) [0028]
• CRD; La et al., World J Gastroenterol, 2008; 14: 731-6 [0039]

Claims (10)

A method of treating inflammatory diseases of the mammalian digestive tract by using at least one pyruvic acid pyruvate. The method of claim 1, wherein said at least one pyruvic acid salt is selected from the group of pyruvates of alkali and / or alkaline earth metals, and / or physiologically acceptable and organic bases. The method of claims 1 and 2, wherein the inflammatory diseases of the digestive tract is selected from irritable bowel syndrome (IBS), chronic inflammatory bowel disease (IBD), lymphocytic colitis, ulcerative colitis, diverticulitis, duodenitis, and Crohn's disease. Method according to one of the preceding claims, wherein the at least one pyruvic acid salt of the organic and physiologically acceptable bases is selected from the group of creatine, carnitine, basic amino acids, choline, di / trimethylamines u. a. Process according to claims 1 and 2, wherein calcium is the cation of the at least one pyruvic acid salt. A method according to claim 5, wherein the water content of the at least one pyruvic acid salt is 0 to 5 molecules, preferably 0 to 2.5 molecules, particularly preferably 0.5 to 1.5 molecules. A composition for use in the method of any one of the preceding claims, which is selected from at least one pyruvic acid salt and additionally a prebiotic selected from the group of water soluble carbohydrates, especially selected from the group of prebiotics: cereal fibers and dietary fibers, psyllium, oligofructoses, inulin, modified starches, gums of vegetable or microbial origin. A composition for use in the method of any preceding claim, wherein useful combinations and formulations, known from alternative medical, nutritional and nutritional approaches and diets, include the following: probiotics (Escherichia coli Nissle 1917, Saccharomyces boulardii, bifido and lactic acid bacteria and others), prebiotics (cereal fibers and fibers, psyllium, oligofructoses, inulin, modified starches, gums of vegetable or microbial origin), fish and krill oils (omega-3 fatty acids), herbal preparations and medicinal herbs and plants and extracts thereof ( for example, frankincense (Boswellia serrata), turmeric (Curcuma longa), thyme species, chamomile, cannabis), essential oils and aromatic oils of peppermint, chamomile, sage, fennel, caraway, thyme or lavender, and dietary supplements. Pharmaceutical formulations, medical devices, functional foods, nutritional supplements or foods containing calcium pyruvates, preferably calcium pyruvate monohydrate, or formulations thereof, for use in humans or mammals, domestic animals and livestock according to the preceding claims. Pharmaceutical formulations, medical devices, functional foods, nutritional supplements or foods containing calcium pyruvates, preferably calcium pyruvate monohydrate, in dosages of 10 mg to 25 grams per day for mammals, including humans.