KR20050107043A - A pharamcetutical composition for treating endodoxemia comprising bovine colostrum - Google Patents

Abstract

The present invention relates to a method for treating endotoxin-related diseases using cow colostrum, and provides a pharmaceutical composition comprising cow colostrum or cow colostrum-derived product alone or in combination with an immunotrophic substance as an active ingredient. The pharmaceutical compositions of the present invention are suitable for preventing or treating intestinal bacterial potential, endotoxins or diseases associated with endotoxins due to intestinal wall injury.

Description

A pharamcetutical composition for treating endodoxemia comprising bovine colostrum}

The present invention relates to a method for effectively preventing and treating enterobacterial bacterium potential and endotoxins caused by intestinal injury by using colostrum of cows that are mostly discarded in the dairy field. More specifically, by using colostrum of cow's milk to maintain intestinal wall function to prevent intestinal damage caused by various factors such as drugs and to effectively prevent or treat enterobacterial bacterium and endotoxemia and various related diseases. It is about a method.

Intestinal bacterial potential refers to a phenomenon in which enteric bacteria and various bacterial products spread beyond the intestinal wall to areas other than the intestine. The spread of endotoxin, which is part of the intestinal bacteria, is called endotoxemia. Endotoxins cause systemic inflammatory response syndrome (SIRS) and sepsis. When enterobacterial potential or enterointestinal bleeding occurs, enterobacteriaceae and various products first spread to mesenteric lymph nodes, and also through the lymphatic system through the chest tube to distant organs such as liver, spleen and lungs, and through the liver and spleen through the portal vein. It can also spread to systemic blood flow.

Enterobacterial bacterium potential and endotoxins have been demonstrated in various animal tests since Berg and Garilington first mentioned in 1979 and are now being identified in human diseases. Diseases associated with enterobacterial bacterium potential and endotoxinemia include trauma, aortic aneurysm and intestinal transplantation, obstructive jaundice, acute pancreatitis, intestinal obstruction, cirrhosis, heart failure, neoplastic disease, inflammatory bowel disease, extrapulmonary acute respiratory failure (extrapulomonary ARDS) And systemic inflammatory response syndrome and pneumonia, and endotoxemia has been shown to be correlated with infectious complications of these diseases ( Adv . Exp . Med . Biol 1999; 473: 11-30 ).

Intestinal bacterial potential caused by products such as intestinal gram-negative bacteria and endotoxins and intestinal endotoxins have been increasingly clinically important.

In the United States, for example, more than 10% of hospitalized patients are treated with pathogenic infections each year, costing more than $ 4.5 billion annually, and a significant number of patients are suspected of having an enterobacterial infection. In addition, 70,000 to 330,000 patients are treated with bacteremia caused by Gram-negative bacteria every year, and have a high mortality rate of 20-40% despite intensive care. It cannot be concluded that both of these Gram-negative bacteria and sepsis were caused by intestinal bacteria in the intestinal tract, but survived 30% of those who had bacteremia and died of sepsis and multi-organ failure (MOF). Investigations and autopsies indicate that the cause of infection cannot be found, suggesting that enterobacterial bacterium and intestinal endotoxemia are major or exacerbating factors in patients with a significant number of infectious complications.

Since such endotoxins are derived from the cell membranes of Gram-negative bacteria, endotoxins are known to be derived from the growth of Gram-negative bacteria. In other words, when gram-negative bacteria propagate due to wound infection or inflammatory reactions in the body, endotoxins are induced through the blood flow in the area, which is considered to be overcome to some extent by the administration of antibiotics. Is not uncommon, and the area or amount of endotoxin, if any, does not matter much. Another important cause is endotoxins from Gram-negative bacteria present in the intestine.

Normally, humans have about 1 trillion bacteria per gram of stool, and each person's disease varies from one environment to another, but there is a significant proportion of Gram-negative bacteria. May cause In this case, if the intestinal wall is maintained normally, it will not be a problem, but since most of the critical patients have problems with the function of the intestinal wall due to various causes, intestinal endotoxemia is usually associated with various infectious complications and endotoxins in severely compromised immune function. May cause complications related to

Thus, control of the intestinal permeability of enterobacteriaceae would be quite effective in treating endotoxins and related diseases.

As a method for maintaining the intestinal wall, there are two methods of orally administering a growth factor and a method of concurrently administering a growth factor and a mucosal cell nutrient such as glutamine.

The method of administering nutrients such as glutamine, ω-3 fatty acids, arginine, and nucleic acids to suppress membrane damage is called immunonutrition ( JAMA 2001; 286: 944-953 ). Mainly. To date, new substances such as glycine, antioxidants, inducible carbon monoxide synthase inhibitors, beneficial active microorganisms (probiotics) and trefoil factor have been proposed to suppress intestinal mucosal damage. Results were often unclear or difficult to apply to clinical practice due to problems of mass production ( J Gastroenterol Hepatol 2003; 18: 479-97 ). Recently, the use of epidermal growth factor (EGF), transforming growth factor-beta (TGF-β), and various growth factors extracted from dairy products has been attempted to improve intestinal membrane damage ( J Gastroenterol 1995; 30: 169-176, Cancer Res 1994; 54: 1135-38, J Nutr 1996; 126: 2519-30 ). Bovine colostrums are a rich source of growth factors in dairy products.

Colostrums are milk secreted for the first time after childbirth and contain large amounts of growth factors, immunoglobulins and lactoferrins or lactoperoxidases, antimicrobial peptides and biologically active substances ( Am J Clin Nutr 2000; 72: 5). -14 ). Cow colostrum, in particular, is classified as a functional food due to its intermediate nature between the therapeutic drug and the food that is being sold over the counter without a doctor's prescription. Recently, there have been attempts to treat various digestive and infectious diseases. It is a substance becoming.

Cow colostrum contains a variety of growth factors, including IGF-I (Insulin like growth factor-I), IGF-II, EGF (Epidermal growth factor), TGF-β (Transforming growth factor-alpha), and TGF-β. (Transforming growth factor-beta), Platelet-derived growth factor (PDGF), Vascular endothelial growth factor (VEGF), and particularly, IGF contained in cow colostrum is 25 times higher than that contained in human colostrum, and It contains 50 times higher content than milk, and TGF-β contains about 20 times higher than milk. In addition, it is rich in immunoglobulins such as IgG, IgA, and IgM, which contains three times as much as human colostrum and about 30-100 times as much milk. About 90% is about 80% to 90% IgG, unlike colostrum of human IgA. It is. Clinically, cow's colostrum uses many properties of various immunoglobulins and antimicrobial peptides, such as Shigella flexneri , enterotoxigenic Escherichia coli , Rotavirus (rotavirus), jakeunwa used for infection prevention and treatment of Cryptosporidium (Cryposporidium parvum) or its usefulness is present and is a material that is expected to inhibit the overgrowth of intestinal bacteria in the different diseases by using such a characteristic ( Br J Nutr 2000; 84 (S1): S135-46, Am J Trop Med Hyg 1992; 47: 276-83, NEJM 1988; 12: 1240-43, J Med Virol 1992; 38: 117-23, Clin Infect Dis 1998; 26: 1324-29 ).

The use of such colostrum growth factors and immune factors to prevent or prevent intestinal damage and overgrowth of enterobacteriaceae may be useful for improving intestinal mucosal damage and suppressing enterobacterial potential due to various diseases such as infectious complications.

In addition, there are many clinical reports that glutamine is administered as a method of inhibiting the intestinal mucosa in various diseases, and there are clinical reports that this method is effective ( Cancer Treat Rev 2003; 29: 501-13; Nutrition 2003; 19: 805-). 11 ) Multiple doses have not been reported. Therefore, if colostrum is used in combination therapy with these immunotrophic therapies will be able to maximize the effect.

It is an object of the present invention to effectively prevent or treat endotoxins and related diseases by improving colostral mucosal damage caused by enterobacteria, inhibiting enterobacterial potential and maintaining the intestinal mucosa by using colostrum containing a large amount of growth factors and immune factors. It is to.

It is also an object of the present invention to more effectively prevent or treat endotoxins and related diseases by using colostrum or colostrum derived-products in combination with immunotrophic therapy.

In addition, the objective test is to measure the degree of intestinal damage in various diseases by using a functional test called intestinal permeability test, and to establish an association between changes in intestinal permeability and pathophysiology and improvement and inhibition of intestinal permeability enhancement.

In order to achieve this object, the present invention provides a pharmaceutical composition comprising cow colostrum or cow colostrum-derived product as an active ingredient. The composition is useful for preventing or treating enterobacterial bacterium, endotoxemia and related diseases caused by intestinal damage.

The present invention also provides a pharmaceutical composition for preventing or treating enterobacterial potential and endotoxins and related diseases due to intestinal damage, comprising an immunotrophic substance together with cow colostrum or cow colostrum-derived products as an active ingredient. do.

Colostrum or colostrum-derived product as an active ingredient according to the present invention includes insulin-like growth factor (IGF), epidermal growth factor (EGF), lactoferrin, immunoglobulin, and the like. It may be prepared according to the method known in the art. More specifically, colostrum obtained from healthy non-immunized grazing cows may be prepared by removing the cream, pasteurizing and spray drying to form protein powders, or optionally using colostrum obtained from highly immunized cows. The colostrum powder may optionally be coated with a beta-lipid coating comprising milk phospholipids, in particular phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine and sphingomyelin.

In addition to the cow colostrum itself, colostrum fractions, in particular colostrum fractions prepared to grow factor-riched, are commercially available (Functional Nutraceuticals Ltd., New Zealand; Proper Nutrition, Inc. USA; Immuno). -Dynamics, Inc, USA; Tricom, Korea).

Colostrum according to the present invention, as described below, suppresses intestinal damage (increased intestinal permeability) in experiments using an intestinal damage model using a nonsteroidal anti-inflammatory agent dichlorophenac and an intestinal damage model using an anticancer agent 5-fluorouracil. It has been shown to inhibit intestinal bacterial potential and endotoxinemia.

Therefore, the pharmaceutical composition containing cow's colostrum of the present invention can be effectively used for the prevention and treatment of endotoxins, and furthermore, to improve all diseases related to endotoxins, in particular, to treat alcoholic liver disease, and to progress cirrhosis. Suppression, prevention and recurrence of coronary artery disease, amelioration and treatment of acute pancreatitis, treatment of acute sepsis and Systemic Inflammatory Response Syndrome (SIRS), Multiple Organ Dysfunction Syndrome (MODS) It can be used for therapeutic purposes throughout all fields of medicine, such as improvement and treatment of prognosis, prevention and reduction of postoperative complications, reduction of complications after organ transplantation, prevention of complications such as mucositis after administration of anticancer drugs, treatment of various respiratory diseases, and treatment of immune diseases.

More specifically, the pharmaceutical composition according to the present invention can be used for digestive system diseases such as alcoholic liver disease, viral and nonviral cirrhosis exacerbation, liver cancer, pancreatitis and complications thereof, which are known to correlate with intestinal damage or endotoxemia. Can be used for palliative and therapeutic purposes and can be used for adjuvant mucosal diseases such as Inflammatory Bowel Disease (IBD), Irritable Bowel Syndrome (IBS), and Intestinal Tuberculosis; Can be used for the prevention of recurrent systemic diseases such as coronary artery disease, atherosclerosis, cerebral infarction and for preventing recurrences; Can be used for the prevention and treatment of infectious diseases such as sepsis, systemic inflammatory response syndrome (SIRS), multi-organ dysfunction (MODS); Can be used for the purpose of preventing and treating respiratory diseases such as asthma, Chronic Obstructive Pulmonary Disease (COPD), colds and flu; It can be used before and after surgery to prevent complications after hemodialysis or peritoneal dialysis, postoperative complications, especially complications after intestinal surgery, and to prevent complications after organ transplantation, to prevent complications after chemotherapy, and to burn patients. Can be used for improved survival and treatment; Can be used for the treatment of acquired immunodeficiency syndrome (AIDS), various autoimmune diseases, in particular, atopic dermatitis, allergic rhinitis, allergic keratitis, rheumatoid arthritis; Can also be used for obstetrics and gynecological diseases such as infertility caused by endotoxin and abortions caused by endotoxin; Others can be used for anti-aging, for the treatment of chronic fatigue syndrome, for hangovers and for child development.

The pharmaceutical composition of the present invention may further comprise an immunotrophic substance together with cow colostrum or cow colostrum-derived product as an active ingredient. Immunonutrients have been used to inhibit the intestinal mucosa, and when combined with colostrum containing various bioactive growth factors, it is possible to maximize the effect of inhibiting intestinal damage and endotoxemia.

Examples of such immunotrophs include glutamine, ω-3 fatty acids, arginine, nucleic acids, probiotics or prebiotics.

The effective dosage of colostrum according to the present invention is about 1 g to 50 g per day, and those skilled in the art can appropriately add or subtract effective dosages within this range depending on the prophylactic or therapeutic purpose or depending on the severity of symptoms and the condition of the patient. In addition, even when used in combination with an immunotrophic substance, the effective daily dose is about 1 g to 50 g, but the ratio may be appropriately adjusted according to the degree of symptoms and the condition of the patient.

The pharmaceutical composition of the present invention is preferably administered orally. Preparations for oral administration include, but are not limited to, tablets, capsules, powders, pouches, liquids, and the like. For such formulations, the pharmaceutical compositions of the present invention may include pharmaceutically acceptable adjuvants such as excipients known in the art.

While the foregoing has been described with respect to some aspects of the invention, their equivalents are also within the scope of the invention.

Hereinafter, the effects of preventing or treating endotoxin and related diseases caused by intestinal damage of the pharmaceutical composition of the present invention will be described in more detail. Such examples are to be understood as illustrative of the invention and not as limiting the scope of the invention. Accordingly, modifications and improvements thereof according to the art are included within the scope of the present invention as defined in the claims appended to the present application.

EXAMPLE

Example 1 Effect of Dairy Cow Colostrum in the Long-Injured Model of White Paper by Diclofenac

Intestinal permeability predicted by the small intestine damage mechanism of nonsteroidal anti-inflammatory drugs in non-administered and non-administered cow colostrum using nonsteroidal anti-inflammatory drugs (NSAID Diclofenac (Diclofenac)), which are mainly used as injectables in Korea, to induce small bowel disease in rats. And changes in intestinal bacteria and protein loss and histological changes in the intestine.

Test subject and material

1) Experimental Animals:

Sixty-eight male Sprague-Downeys, weighing 150-170 g, were fed from Orient (Orient, Seoul, Korea) for seven days before the experiment. During breeding of white paper, day and night were distinguished at 12 hour intervals, and breeding temperature was maintained at 22 ± 1 ° C. Solid feed (Basal diet 5755, PMI Nutrition International, Inc., Richmond, USA) and water were freely consumed. The white papers did not show any infection or other abnormalities before the start of the experiment and weighed 180-220 g. The white papers were divided into 4 groups as follows, and 16 animals were assigned to each group.

Experimental group A B C D Control NSAID NSAID + 5% Cow Colostrum NSAID + 10% Cow Colostrum

2) NSAIDs:

Dichlorophenac sodium (Ildong Pharm., Seoul, Korea), which is widely used as injection in Korea, was used.

3) Dairy Cow Colostrum:

A fat-free dairy colostrum, Percoba ™ Liquid (Immuno Dynamics, Ltd., Perry, USA) was used.

Composition of Percoba ™ Liquid Formula (calories = 64 cal / dL) ingredient Content (g / dL) carbohydrate 10 Fat <0.1 Fiber <0.1 Total protein 6.0 albumin 2.0 IgG 1.02

4) Enteric permeability measuring material:

⁵¹ Cr-EDTA (ethylenediaminetetraacetic acid) (Amersham Health, Inc., Buckinghamshir, UK) was used and dispensed in an amount the day before dosing.

Experiment method

1) Dose of cow colostrum and diclofenac sodium:

As shown in FIG. 1, group A and group B were allowed to ingest solid feed and water freely during the experiment, and group C and D were fed 5% and 10% cow colostrum instead of water. In addition, by installing a net to prevent the intake of non-food, such as feces or rugs. On the 5th day of the experiment, groups B, C, and D were injected subcutaneously with 50 mg / kg of diclofenac sodium, and group A, the control group, was injected with the same amount of saline.

2) Enteric permeability measurement:

Diclofenac administered 24 to ⁵¹ Cr-EDTA the 25 micro Ci of time after the white paper by using a metal oral administration tube was mitmang after oral administration is installed and metabolic cages that can be collected by separating the feces (metabolic cage) ⁵¹ Cr-EDTA in Urine was collected for 24 hours from 2 hours after dosing. ^The water and feed were not fed to the white paper for 12 hours prior to ⁵¹ Cr-EDTA administration. To calculate the recovery rate of ⁵¹ Cr-EDTA in 24-hour urine, measure the 24-hour urine volume of white paper, take 4 ml of this solution, and 4 ml of solution containing ⁵¹ Cr-EDTA 2.5 micro Ci within 6 hours after the completion of urine collection. Together, radioactivity (cpm) was measured with a gamma counter (Cobra Auto-gamma counter, Packerd Instrument Comp., Meriden, USA). The measurements were calculated as follows and the amount recovered in the ⁵¹ Cr-EDTA taken was expressed as a percentage ( ⁵¹ Cr-EDTA [%] = [urine cpm x urine volume x 2.5] / standard cpm x 25).

3) Surgery:

After 24 hours of urine collection for intestinal permeability test, the white paper was weighed, anesthetized with ether, and aseptically dissected the abdomen at the center. Immediately after incision, an 18 gauge syringe was inserted in the 20 cm ileum from the cecum and the intestinal contents were collected about 0.5 cc, and removed 3 cm at the same site for histological examination and fixed in 10% formalin. The diaphragm was then dissected and at least 5 cc of blood drawn from the heart and the white paper died afterwards.

4) Intestinal aerobic bacteria count:

100 mg of the intestinal contents obtained from the ileum were continuously diluted with a solution of phosphate buffered saline (PBS) at pH 7.4, followed by blood sheep agar plates and enterogram Gram-negative bacteria to measure the intestinal aerobic bacteria. In order to measure the number was inoculated in MacConkey agar plate (MacConkey agar plate) and incubated for 24 hours at 37 ℃ in an incubator. After incubation, the number of colonies grown in each medium was observed and expressed as CFU (colony-forming unit) / g.

5) Histological examination:

The formalin-fixed ileum was longitudinally excised at the site opposite to the mesentery attachment, dehydrated and penetrated into paraffin to make a slide, and cut to 5 microns thick. Staining was observed by light microscopy by H & E (Hematoxilin & Eosin) staining. Histologic findings were as follows.

Scorecard for NSAID-induced Intestinal Injury 1 point Increased cell infiltration and / or chorionic degeneration and / or epithelial reduction 2 points Ulcers and / or necrosis and / or lymphoid vesicles 3 points Cell wall permeation and / or peritonitis x 2  More serious than each lesion

6) Hematological tests:

To measure protein loss in the intestinal tract due to intestinal damage of nonsteroidal anti-inflammatory drugs, total protein and albumin levels were measured by blood from white paper, and total protein and albumin levels were not due to hepatotoxicity of nonsteroidal anti-inflammatory drugs. In order to compare the ALT (alanine transaminase) ALP (alkaline phosphatase), total bilirubin, albumin and total protein amounts were measured at the same time.

7) Determination of Serum Endotoxin:

In order to break the binding between protein components and endotoxins in serum, the serum obtained from the white paper was diluted 1:10 with BD100 buffer, and heated at 75 ° C. for 15 minutes to remove substances causing interference in serum. The heated serum was diluted 1: 2 with BD100 buffer again after cooling and dispensed 4 times with 100 μl of each sample in 96 microtiter well plates. Two of these wells, as a positive control for judging the presence of interference, was added a predetermined concentration of endotoxin and the endotoxin level was recognized when the amount of recovered endotoxin was in the range of 50-200%. Standard straight lines were made using standardized control standard endotoxin (CSE) of 5, 0.5, 0.05 and 0.005 EU / mL. Limulus Amebocyte Lysate (LAL) reagent was used as Kinetic turbidimetric analysis (KTA) -2 (Charles River Endosafe, Charleston, South Carolina, USA). The LAL reagent was dissolved in ES (endotoxin-specific) buffer to remove the effect of β-D-glucan, and then 100 μl was added to each well, and maintained at 37 ° C. Endoscan-V (Charles River Endosafe, Charleston, M. Carolina, USA) program using the ELX808IU microplate reader (Bio-Tek Instruments, Inc., Winooski, Vermont, USA). In addition, all reagents, vitreous and plastic products used in the test were free of endotoxin.

8) Statistical Processing:

All figures are shown as mean ± standard deviation difference between each group was SPSS 11.0 program to carry out (SPSS Inc, Chicago, IL, USA) Mann-Whitney test, ANOVA test using which the significance of not more than the p value 0.05 It was determined that.

Experiment result

The average daily intake of colostrum in the white paper was 5% cow colostrum 33.6 ± 3.3 ml / day (1.68 ± 0.16 g / day) in group C, and 10% cow colostrum 34.9 ± 2.0 ml / day in group D (3.49 ± 0.10 g / day) There was no difference in the intake of colostrum between the two groups ( p > 0.05), and the colostrum was administered twice as much as the C group. The change of intestinal permeability using ⁵¹ Cr-EDTA increased more than 7 times in group B with diclofenac alone compared with group A, but group C with 10% cow colostrum and 10% cow colostrum together with 5% cow colostrum. In one group D, 4.5 times and 3 times increased, respectively, the increase in enteric permeability by diclofenac was suppressed as the intake of cow colostrum increased ( p <0.01) (Fig. 2).

The results of culture in blood agar medium for measuring intestinal aerobic bacteria were observed to be overproliferated by 400 times more in diclofenac administration group than the control group, and the propagation of the strain was 30 times higher than that in the control group. Hyperproliferation was reduced by 1/10 ( p <0.001), but there was no significant difference between the 5% cow colostrum and 10% cow colostrum groups (FIG. 3A).

In addition, in the culture results in MacConkey agar medium for measuring the intestinal gram negative bacteria number, it was observed to overproliferate on average 1000 times more than the control group ( p <0.001). It was reduced to 1/20 or less of the diclofenac alone administration group by 40 times, but there was no significant difference between the 5% cow colostrum administration group and the 10% cow colostrum administration group as in the blood agar medium (FIG. 3B).

There were no differences in the ALT and total bilirubin levels between the groups, but the total protein and albumin levels in the diclofenac-treated group were significantly lower than those in the control group, and the total protein and albumin levels were higher in the control group. It was found that the recovery was close to the numerical value ( p <0.001) (Table 3).

Biochemicals found in serum 2 days after diclofenac Experimental group Total protein (g / dL) Albumin (g / dL) ALT (IU / L) ALP (IU / L) Total bilirubin (mg / dL) A 5.38 ± 0.19 3.61 ± 0.14 28.3 ± 5.9 202.7 ± 31.9 0.17 ± 0.12 B 4.46 ± 0.38 a 2.88 ± 0.21 a 26.4 ± 5.1 117.8 ± 26.8 0.19 ± 0.09 C 5.04 ± 0.32 ab 3.18 ± 0.19 ab 27.1 ± 3.5 112.8 ± 20.8 0.24 ± 0.19 D 5.38 ± 0.42 abc 5.38 ± 0.42 abc 29.4 ± 7.0 143.9 ± 36.0 0.34 ± 0.31

. All tested on the Mann-Whitney exam.

. The above values are mean ± standard deviation.

. a: compared to group A p <0.001 / b: compared to group B p <0.001 / c: compared to group C p <0.05 /

In addition, histological examination showed increased intestinal mucosal damage in the diclofenac-treated group compared with the control group, and improved mucosal damage in the colostrum-treated group. Mucosal damage was improved ( p <0.05) (FIG. 4).

In addition, the serum endotoxin by LAL test was measured at 0.1 EU / mL or less in all 16 times in the control group, but in the diclofenac-administered group, it was measured at 0.1 EU / mL or more in 12 of 16 cases and 5 times in the diclofenac and colostrum group. Or in two cases, endotoxinemia caused by diclofenac administration was measured to be greater than 0.1 EU / mL ( p <0.05) (Table 4).

Serum endotoxin concentration 2 days after diclofenac administration A group B group C group D group One <0.1 0.1211 <0.1 <0.1 2 <0.1 0.5244 <0.1 <0.1 3 <0.1 0.2988 0.1813 <0.1 4 <0.1 2.4562 <0.1 0.1314 5 <0.1 <0.1 1.3124 <0.1 6 <0.1 2.2433 <0.1 0.1877 7 <0.1 3.6326 <0.1 <0.1 8 <0.1 0.2756 0.1211 <0.1 9 <0.1 <0.1 <0.1 <0.1 10 <0.1 5.3428 <0.1 <0.1 11 <0.1 <0.1 0.2422 <0.1 12 <0.1 0.7632 <0.1 <0.1 13 <0.1 0.8943 <0.1 <0.1 14 <0.1 1.5313 0.2422 <0.1 15 <0.1 5.3427 <0.1 <0.1 16 <0.1 <0.1 <0.1 <0.1

In conclusion, it was confirmed that cow colostrum was effective in inhibiting the increase of enteropermeability, overgrowth of enterobacteria and protein loss and mucosal damage in the intestine in acute intestinal injury model induced by nonsteroidal anti-inflammatory drugs induced in rats. Thus, cow colostrum has not only been applied to the prevention and treatment of chronic and acute intestinal damage by nonsteroidal anti-inflammatory drugs, but also to enterobacterial potentials such as burns, hemorrhagic shock, severe pancreatitis, intestinal ischemia and obstruction, and spontaneous bacterial peritonitis in patients with cirrhosis. And for the treatment of diseases showing an increase in enteric permeability and intestinal bacteria, such as intestinal endotoxin.

Example 2: Effect of Dairy Cow Colostrum on Intestinal Injury and Intestinal Bacterial Potential and Intestinal Endotoxinemia Induced by 5-Fluorouracil (5-FU) in White Paper

Anti-cancer agent 5-fluorouracil (5-FU), which is widely used for the treatment of gastrointestinal tumors such as gastric cancer and colorectal cancer, was used to induce intestinal wall damage and enterobacterial bacterial potential in rats.

Test subject and material

Except for using 30 male rats weighing 250-300 g of 8-week-old as an experimental animal, and using 5-FU (an anti-cancer drug, Seoul, Korea) as an intestinal damage-inducing drug, As described.

Experiment method

1) Cow colostrum and 5-FU administration:

As the experimental group, group A was divided into control group, group B to 5-FU administration group, group C to 5-FU and cow colostrum administration group, and group C forced oral cow colostrum at the dose of 4 g per day 5 days before 5-FU administration. The administration was continued until the end of the experiment, group 1 and group C was administered intraperitoneally with 5-FU at 300 mg / kg dose, and the control group A was administered intraperitoneally with saline as a control drug, in Example 1 According to the experimental method described.

2) Intestinal Permeability Measurement:

Intestinal permeability was measured as described in Example 1 except that oral administration of 10 μCi of 51Cr-EDTA was performed 48 hours after 5-FU administration, and the urine volume of the white paper was measured and 4 ml of this was taken. Shown in percentage. (Cr-EDTA [%] = [urine cpm x urine volume x 1.0] / standard cpm x 10).

3) Surgery:

After 24 hours of urine collection for intestinal permeability, the white paper was weighed, anesthetized with ether, and aseptically dissected the abdomen from the center. Immediately after the incision, 500 mg of the intestinal contents were taken from the cecum at 40 cm above the caecum, and 3 cm were excised from the same site for histological examination and fixed in 10% formalin. The diaphragm was then excised and blood was drawn from the heart to kill the white paper. To determine the intestinal bacterial potential, 100 mg of mesenteric lymph nodes, liver and spleen were collected by aseptic procedure, and the caecum was incised about 1 cm to measure the number of enterobacteriaceae. In order to observe histological changes in the large intestine, 5 cm to 3 cm in the lower abdomen was excised and fixed in 10% formalin.

4) Intestinal aerobic bacteria count and intestinal bacterial potential:

100 mg of intestinal contents obtained from ileum and caecum were serially diluted, and then inoculated in blood agar medium and intestinal gram-negative bacteria in order to measure intestinal aerobic bacteria and inoculated in MacConkey agar medium for 24 hours at 37 ° C. Incubated for After incubation, the number of colonies grown in each medium was observed and expressed as CFU (colony-forming unit) / g. To determine the intestinal bacterial potential, homogenizer (homogenizer, Heidolph-Elektro GmbH & Co., Schwabach, Germany) was used for homogenization (homogenizer, Heidolph-Elektro GmbH & Co., Schwabach, Germany). MacConkey agar medium was inoculated and incubated for 24 hours at 37 ℃ in the incubator. After incubation, the number of colonies grown in each medium was observed and expressed as CFU (colony-forming unit) / g. In addition, 1 mL of blood collected from the heart was incubated with BactecPed-F (Becton Dickinson and Company, Sparks, Maryland, USA).

5) Histological, hematological and serum endotoxin measurements

According to the method described in Example 1, mucosal damage such as atrophy and edema and inflammation of the villi, and detachment of the mucosal cells were observed, and the amounts of ALT, AST (Aspartate aminotransferase) and total bilirubin, ALP, albumin and total protein were simultaneously measured. Serum endotoxin was measured.

Experiment result

Intestinal permeability using ⁵¹ Cr-EDTA to measure intestinal wall damage was more than doubled in group B treated with 5-FU alone ( p <0.001). In administered group C, the increase in enteropermeability was reduced ( p <0.01) (FIG. 6).

Gram-negative bacterial counts in the mesenteric lymph nodes, liver and spleen to monitor the intestinal bacterial potential were observed in the 5-FU administration group, and the number of gram-negative bacteria was increased in the mesenteric lymph nodes, liver and spleen than in the control group ( p < 0.01), significantly less intestinal bacterial potential in mesenteric lymph nodes (A) and spleen (C) than in the 5-FU alone group ( p <0.05) (Fig. 7).

In the hematological test, the total protein and albumin levels of blood were significantly decreased in the 5-FU group without the increase of ALT, AST, ALP and total bilirubin levels ( p <0.01). And albumin levels recovered ( p <0.05) (Table 5).

Biochemicals found in serum 2 days after 5-FU administration Experimental group Total protein (g / dL) Albumin (g / dL) ALT (IU / L) ALP (IU / L) AST (IU / L) Total bilirubin (mg / dL) A 6.16 ± 0.53 3.84 ± 0.21 32.4 ± 6.8 144.0 ± 52.6 184.4 ± 44.6 0.17 ± 0.12 B 5.09 ± 0.72 a 3.30 ± 0.39 b 26.0 ± 4.6 130.3 ± 58.9 108.2 ± 34.3a 0.19 ± 0.09 C 5.67 ± 0.41 c 3.66 ± 0.19 c 27.6 ± 4.4 139.7 ± 44.8 160.2 ± 29.2d 0.24 ± 0.19

. All tested on the Mann-Whitney exam.

. The above values are mean ± standard deviation.

. a: compared with group A p = 0.001 / b: compared with group A p <0.01 / c: compared with group B p <0.05 / d: compared with group B p <0.01

Serum endotoxin by LAL test was measured at 0.1 EU / mL in the control group, but it was measured at 0.1 EU / mL in five cases, which was half in the 5-FU group, and 0.1 in one case in the simultaneous group of 5-FU and colostrum. Endotoxinemia caused by 5-FU administration, measured above EU / mL, was reduced by administration of cow colostrum ( p <0.05) (Table 6)

Serum endotoxin concentration 3 days after 5-FU administration (unit: EU (Endotoxin Unit / mL) A group B group C group One <0.1 <0.1 <0.1 2 <0.1 <0.1 <0.1 3 <0.1 0.1211 <0.1 4 <0.1 3.6326 <0.1 5 <0.1 <0.1 <0.1 6 <0.1 <0.1 <0.1 7 <0.1 0.2989 <0.1 8 <0.1 <0.1 <0.1 9 <0.1 0.2228 <0.1 10 <0.1 5.3427 <0.1

Based on these results, it was confirmed that intestinal bacterial potential and intestinal endotoxemia caused by 5-FU-induced intestinal barrier damage in rats were suppressed by cow colostrum administration. Is useful for ameliorating intestinal wall injury by chemotherapy, such as 5-FU, and suppressing intestinal bacterial potential and intestinal endotoxemia. .

According to the present invention, a pharmaceutical composition comprising cow colostrum or cow colostrum-derived product alone or in combination with an immunotrophic substance is an intestinal bacterium potential caused by drugs or irritant stress, and enterobacterial bacterium potential, It is useful for preventing or treating diseases associated with toxinemia or endotoxinemia.

Figure 1 shows the test plan for determining the effect of cow colostrum in the intestinal injury model of white paper by nonsteroidal anti-inflammatory drugs (NSAID).

FIG. 2 shows the change in intestinal permeability measured by urine excretion of ⁵¹ Cr-EDTA for 24 hours in the experiment of FIG. 1.

Figure 3 shows the total bacterial count (3A) and Gram negative aerobic bacterial count (3B) in the ileum two days after administration of the nonsteroidal anti-inflammatory agent.

Figure 4 shows the biopsy for small intestine damage 2 days after the administration of nonsteroidal anti-inflammatory drugs.

Figure 5 shows the change in intestinal permeability measured by urine excretion of ⁵¹ Cr-EDTA for 24 hours for the effect of colostrum on the intestinal damage of rats by anticancer agents.

Figure 6 shows the effect of cow colostrum on bacterial potential to mesenteric lymph nodes (A), liver (B) and spleen (C).

Claims (9)

A pharmaceutical composition comprising cow colostrum or cow colostrum-derived product as an active ingredient, and for preventing or treating diseases associated with enterobacterial potential, endotoxemia and endotoxinemia due to intestinal wall injury. A pharmaceutical composition comprising an immunotrophic material, together with cow colostrum or colostrum-derived product, for preventing or treating a disease associated with enterobacterial bacterium, endotoxemia or endotoxemia due to intestinal wall injury. The pharmaceutical composition of claim 2, wherein the immunotrophic material is selected from the group consisting of glutamine, ω-3 fatty acids, arginine, nucleic acids, probiotics or prebiotics. The pharmaceutical composition according to claim 1 or 2, wherein the colostrum is selected from the group consisting of colostrum fraction, growth factor-rich colostrum and lactoferrin-rich colostrum. The gastrointestinal tract of claim 1, wherein the disease associated with endotoxemia comprises alcoholic liver disease, liver cirrhosis, liver cancer, pancreatitis, acute pancreatitis, acute sepsis, inflammatory bowel disease (IBD) and irritable bowel symptoms (IBS). Systemic diseases; Circulatory diseases including coronary artery disease, arteriosclerosis and cerebral infarction; Infectious diseases including sepsis, systemic inflammatory response syndrome (SIRS) and multi-organ dysfunction (MODS); Respiratory diseases including asthma, chronic obstructive pulmonary disease (COPD), colds and flu; Complications associated with intestinal bacterial potential and endotoxins, including complications after artificial hemodialysis or peritoneal dialysis, complications after organ transplant surgery, and mucositis after chemotherapy, and post-burn complications; A pharmaceutical composition selected from the group consisting of autoimmune diseases including atopic dermatitis, allergic rhinitis, allergic keratitis and rheumatoid arthritis, or congenital diseases such as AIDS, and gynecological diseases including infertility and miscarriage. The pharmaceutical composition according to claim 1 or 2 for the prevention of aging, treatment of chronic fatigue syndrome, relieving hangovers or promoting the growth of children. The pharmaceutical composition according to claim 1 or 2, wherein the daily dose of colostrum is 1 g to 50 g. The pharmaceutical composition according to claim 1 or 2, which is orally administered. The pharmaceutical composition of claim 8, wherein the oral dosage agent is selected from the group consisting of tablets, capsules, powders, pouches and solutions.