WO2019041848A1

WO2019041848A1 - Use of butyric acid compound in preparation of drug for preventing and treating nec or nutrition preparation

Info

Publication number: WO2019041848A1
Application number: PCT/CN2018/084675
Authority: WO
Inventors: 文洁; 蔡威
Original assignee: 上海市儿科医学研究所; 上海交通大学医学院附属新华医院
Priority date: 2017-08-29
Filing date: 2018-04-26
Publication date: 2019-03-07

Abstract

Provided in the present invention is the use of a butyric acid compound in the preparation of a drug for preventing and treating necrotizing enterocolitis or a nutrition preparation, also provided is a nutrition preparation containing butyric acid which comprises a nutritional composition and a butyric acid compound, wherein the concentration of the butyric acid compound in the nutrition preparation is 0.5 to 200 mM (mmol/L). The butyric acid compound is selected from at least one of butyric acid, butyrate salts, and butyric acid derivatives. The present invention first finds that the butyric acid compound is very effective in preventing and treating necrotizing enterocolitis by adding it to a pharmaceutical or nutritional composition, including significantly reducing the incidence and mortality of necrotizing enterocolitis, shortening the duration of necrotizing enterocolitis, reducing its complications and/or reducing its severity. At the same time it also has the effects of promoting growth, immune system development and intestinal maturation.

Description

Application of butyric acid compounds in preparation of NEC drugs or nutritional preparations

Technical field

The invention relates to the field of medicine and food technology, in particular to a use of a butyric acid compound in the preparation of a medicament for preventing or treating necrotizing enterocolitis or a nutritional preparation.

Background technique

Necrotic enterocolitis (NEC) is an intestinal disease that endangers the life of children and is one of the important causes of neonatal death. There is no effective prevention and treatment method in clinical practice. With the advancement of perinatal medical technology, the birth rate and survival rate of premature infants continue to increase, especially with the application of in vitro fertilization combined with embryo transfer technology, the incidence of NEC has increased in recent years. The long-term outcome of children with NEC is not ideal, and the mortality rate is as high as 15%-50%. Even after successful surgical treatment, complications such as intestinal stenosis and short bowel syndrome often occur, which seriously affects the survival rate and quality of life of the children. Therefore, finding safe and effective prevention and treatment measures has become a problem that needs to be solved in clinical practice, and it is also a major difficulty in clinical practice.

Although necrotizing enterocolitis is similar in name to enteritis, it is a completely different disease from traditional enteritis and inflammatory bowel disease. The etiology and pathogenesis are different, and the prevention and treatment methods are completely different. It is widely believed that NEC is a multi-factor-caused disease, mainly occurring in premature infants, so immature gut is the basis of NEC. There are usually three factors in the small intestine of children with NEC: persistent intestinal ischemia and hypoxia damage, intestinal substrate (eg, enteral feeding), bacterial colonization. Intestinal ischemia and hypoxia damage can destroy the intestinal barrier, causing the intestinal tract to be susceptible to bacterial invasion. As the lesion progresses, it can lead to necrosis, perforation, peritonitis, sepsis and death of the entire intestinal wall. Compared with other intestinal diseases, NEC is more dangerous, progressing rapidly and endangering the lives of children. At present, the conservative treatment measures for NEC are mainly fasting, gastrointestinal decompression, the use of adequate antibiotics and intravenous nutritional support treatment. However, the use of antibiotics in large doses for a long time and repeated times is likely to cause damage to the whole body organs of the child, and problems such as antibiotic resistance and double infection may occur. Intravenous nutrition is extremely important or even the only nutritional support pathway for children with NEC risk or with NEC. However, long-term use of intravenous nutrition support, there will also be complications leading to the need to reduce or stop intravenous nutrition.

Due to the difficulties in the treatment of NEC, researchers at home and abroad have carried out a large number of experimental research and clinical research to explore ways to prevent and control NEC. Researchers have used different drugs or nutrients to intervene in various aspects of NEC pathogenesis, such as growth factors, arginine, glutamine, probiotics, prebiotics, IL-10, polyunsaturated fatty acids, etc. Although some drugs have achieved certain effects in animal experiments, their clinical application has been limited by the lack of safety and effectiveness. On the other hand, artificial feeding is more likely to occur in NEC than in breastfeeding because artificial feeding is another important risk factor for NEC. There are many protective factors in breast milk compared with formula, so many researchers at home and abroad are working to explore ways to prevent and control NEC by studying the dominant components in breast milk, but the effect is not significant.

Butyric acid is a natural component in mammalian milk, but it is low in content and is one of the metabolites of intestinal flora in the body. Butyric acid, as a kind of short-chain fatty acid, plays an important role in maintaining the normal physiological function of the intestine. In some intestinal diseases such as ulcerative colitis and Crohn's disease, butyric acid has the effect of alleviating the symptoms of the disease. However, long-term research has suggested that the role of butyric acid in preterm infants and neonatal intestines is not the case. Premature babies and newborns have special physiological characteristics, such as immature intestinal development, poor peristalsis, lack of lactase, and often accompanied by history of infection and asphyxia. Due to the lack of gastrointestinal motility, the food stays for a long time, which tends to cause bacterial growth. On the other hand, due to the relative lack of lactase in the small intestine, bacteria in the intestines will decompose a large amount of lactose into short-chain fatty acids, thereby producing excessive short-chain fatty acids. And accumulated in the intestinal lumen can not be removed in time. Intestinal development in premature infants and neonates is not yet mature, and the intestinal barrier function is not yet perfect. High concentrations of short-chain fatty acids can cause or aggravate intestinal mucosal damage. Numerous studies have confirmed that excessive production or accumulation of short-chain fatty acids such as butyric acid in premature infants and neonatal intestinal lumen is an important cause of NEC. For example, the study found that neonatal rats were administered enterally by short-chain fatty acids such as acetic acid and butyric acid, which caused NEC-like damage in the intestinal tract in a concentration-dependent manner (Lin J, et a1.Variable effects of short chain fatty acids and lactic acid in Inducing intestinal mucosal injury in newborn rats. J Pediatr Gastroenterol Nutr. 2002, 35(4): 545-550.). Peng et al. found that high concentrations of butyric acid may cause intestinal barrier damage by increasing the apoptotic rate of intestinal epithelial cells (Peng L, et a1.Effects of butyrate on intestinal barrier function in a Caco-2 cell monolayer model of intestinal barrier. Pediatr Res .2007, 61(1): 37-41.). Carbonaro et al. found that high concentrations of butyric acid caused an increase in permeability through damage to the intestinal mucosal barrier, which in turn led to bacterial translocation in the intestinal lumen (Carbonaro CA, et a1.A bacterial pathogenicity determinant associated with necrotizing enterocolitis. Microb Pathog. 1988, 5 ( 6): 427-436.). Therefore, based on these findings, short-chain fatty acid butyrate has long been considered to be an important factor in NEC.

In the prior art, the patent document with application number 200880120656.X discloses a preparation for improving the function of the gastrointestinal tract. This patent provides a protein-rich enteral nutrition formula supplemented with glutamine, docosahexaenoic acid (DHA) and butyric acid for improved gastrointestinal function. High protein in this formula can improve enteral nutrition and barrier function; glutamine is added to prevent intestinal inflammation and regulate stress-related apoptosis; add omega-3 fatty acids, such as DHA, to prevent the spread of intestinal inflammation And add short-chain fatty acid butyric acid to improve intestinal barrier function. The patent has been used to improve gastrointestinal function by adding several nutrients, and there is no evidence to prove the role of the formula in the prevention and treatment of necrotizing enterocolitis. And butyric acid is weak acid, volatile, and smelly. The patent adds butyric acid directly to the enteral nutrition formula, which not only brings into the unpleasant odor, but more importantly, the weakly acidic substance may not have much influence on the normal and mature intestinal tract, but directly adds weak acidity. Substances stimulate the immature intestines of newborn individuals, especially those at risk of necrotizing enterocolitis, which may accelerate intestinal damage.

Since necrotizing enterocolitis mainly occurs in infants and young children, the safety of control methods is the primary and most important consideration, and intervention with traditional drugs may cause certain side effects, especially when used for preventive purposes. Recognition of parents or medical staff of young subjects. Therefore, finding a safe and effective method is a major technical problem in the prevention and treatment of necrotizing enterocolitis.

Summary of the invention

The technical problem to be solved by the present invention is to provide an application of a butyric acid compound in the preparation of a medicament for preventing or treating necrotizing enterocolitis, and providing a nutrition containing a butyric acid compound, in view of the deficiencies of the prior art. The preparation can be used for preventing and treating necrotizing enterocolitis (NEC).

The present invention finds some problems by analyzing the research of the prior art. First, although bacterial overgrowth in the intestinal lumen of preterm infants results in the production of large amounts of short-chain fatty acids, there is no definitive clinical evidence to confirm the specific components of these short-chain fatty acids and their role in NEC. Secondly, studies have reported that butyric acid induces intestinal epithelial cell apoptosis and leads to intestinal barrier disruption (Peng L, et a1. Effects of butyrate on intestinal barrier function in a Caco-2 cell monolayer model of intestinal barrier. Pediatr Res. 2007, 61 (1): 37-41.) (Ruemmele FM, et a1. Buttyrate induced Caco-2cell apoptosisis mediated via the mitochondrial pathway. Gut. 2003, 52(1): 94-100.), but the intestines used in these studies The epithelial cells are Caco-2 cell lines derived from colon adenocarcinoma cells, whereas butyric acid has anti-cancer properties that inhibit cancer cell proliferation and promote cancer cell apoptosis (Mandal M1, et a1. Butyric acid induces) Apoptosis by up-regulating Bax expression via stimulation of the c-Jun N-terminal kinase/activation protein-1 pathway in human colon cancer cells. Gastroenterology. 2001 Jan; 120(1): 71-8.). Therefore, the conclusion that butyric acid increases the apoptosis of intestinal epithelial cells in these studies may not be established on normal intestinal epithelial cells. Finally, studies using high concentrations of butyric acid in animal experiments have resulted in damage to the intestinal mucosa and intestinal barrier (Lin J, et a1.Variable effects of short chain fatty acids and lactic acid in inducing intestinal mucosal injury in newborn rats. J Pediatr Gastroenterol Nutr. 2002, 35(4): 545-550.), but butyric acid is a weakly acidic substance. Direct administration of the butyric acid stock solution may cause chemical damage to the immature intestinal tract, thus masking its originality. Some physiological effects.

In the present invention, we innovatively provide a nutrient preparation containing butyric acid compound for preventing and treating necrotizing enterocolitis, overcoming the technical bias existing in the above existing research, and solving the technical problem of NEC prevention and treatment. NEC's control provides a safe, economical and effective method. Among them, butyric acid compounds include butyrate, butyric acid derivatives, and butyric acid. Butyrate and butyric acid derivatives are not acidic substances and therefore do not cause damage to the intestinal tract; in the present invention, when the butyric acid compound is butyric acid, it is added in the form of a coating, which not only prevents high concentration of butyric acid from directly contacting the intestinal tract. The mucous membrane causes damage and does not carry an unpleasant odor that affects the taste of the nutritional preparation. Moreover, the present invention takes into account the immature characteristics of premature infants and neonatal intestinal development, and does not adopt a direct administration method, but arranges butyrate, butyric acid derivative, and coated butyric acid in a nutritional combination such as a formula. On the one hand, avoid chemical damage to the intestine due to osmotic pressure, PH value, etc. On the other hand, adding butyric acid-containing compounds to the nutrient preparation can make it slowly play through the intestines with food to better play. Function, thereby effectively preventing NEC.

Second, because intravenous nutrition is extremely important or even the only nutritional support pathway for children with NEC risk or with NEC. In the present invention, we have innovatively added butyric acid compounds to parenteral nutrition preparations, providing a new and more reliable intervention route for the prevention and treatment of NEC.

The object of the invention is achieved by the following technical solutions:

The invention provides a use of a butyric acid compound in the preparation of a medicament for preventing and treating necrotizing enterocolitis.

Preferably, the butyric acid compound is selected from at least one of butyric acid or a pharmaceutically acceptable salt or derivative thereof.

Preferably, the pharmaceutically acceptable salt of butyric acid is selected from at least one of sodium butyrate, potassium butyrate, calcium butyrate, and magnesium butyrate.

Preferably, the pharmaceutically acceptable derivative of butyric acid is selected from the group consisting of glyceryl butyrate, mono-diglyceride butyrate, ethyl butyrate, methyl butyric acid, isoamyl butyrate, and butyric acid cyclodextrin. At least one of the substances.

Preferably, the medicament comprises a therapeutically effective amount of a butyric acid compound.

Preferably, the effective dose of the butyric acid compound is 0.05 to 20 mmol/kg/d.

Preferably, the medicament further comprises other drugs compatible with the butyric acid compound and a pharmaceutically acceptable carrier and/or adjuvant.

Preferably, the medicament is in a pharmaceutically acceptable dosage form.

Preferably, the dosage form is a powder, an injection, a capsule, a tablet or an oral solution.

Preferably, when the butyric acid compound is butyric acid, the butyric acid compound is added to the drug in the form of a coating or a coating.

The invention also provides a use of a butyric acid compound in the preparation of a nutritional preparation for preventing and treating necrotizing enterocolitis.

The present invention also provides a butyric acid-containing nutritional preparation comprising a nutritional composition and a butyric acid compound in a concentration of 0.5 to 200 mM (mmol/L) in the nutritional preparation.

Preferably, the butyric acid compound is at least one selected from the group consisting of butyric acid, butyrate, and butyric acid derivatives.

Preferably, the butyrate is selected from at least one of sodium butyrate, calcium butyrate, magnesium butyrate, and potassium butyrate.

Preferably, the butyric acid derivative is at least one selected from the group consisting of glyceryl butyrate, mono-diglyceride butyrate, ethyl butyrate, methyl butyric acid, isoamyl butyrate, and butyric acid cyclodextrin complex. Kind.

Preferably, the butyric acid compound is butyric acid and/or butyrate, and the concentration of butyric acid and/or butyrate in the nutritional preparation is from 1 to 200 mM.

More preferably, the butyric acid compound is butyric acid and/or butyrate, and the concentration of butyric acid and/or butyrate in the nutritional preparation is from 1 to 100 mM; most preferably, the concentration is from 50 to 100 mM. Nutritional preparations containing this concentration of butyric acid and/or butyrate are effective in preventing the occurrence of NEC, promoting growth, immune system development, and intestinal maturation.

More preferably, the butyric acid compound is butyric acid and/or butyrate, and the concentration of butyric acid and/or butyrate in the nutritional preparation is from 100 to 200 mM. A nutritional formulation containing this concentration of butyric acid and/or butyrate is effective in treating NEC.

Preferably, the butyric acid compound is a butyric acid derivative, and the concentration of the butyric acid derivative in the nutritional preparation is 0.5 to 200 mM.

More preferably, the butyric acid compound is a butyric acid derivative, and the concentration of the butyric acid derivative in the nutritional preparation is from 0.5 to 100 mM; most preferably, the concentration is from 50 to 100 mM. A nutrient preparation containing the concentration of the butyric acid derivative can effectively prevent NEC.

Preferably, the butyric acid compound is a butyric acid derivative, and the concentration of the butyric acid derivative in the nutritional preparation is 100 to 200 mM. A nutritional preparation containing this concentration of butyric acid derivative is effective for treating NEC.

Preferably, the nutritional composition is selected from at least one of a conventional formula, a special medical use formula, a parenteral nutrition preparation, and an enteral nutrition preparation.

Preferably, the conventional formula includes infant formula, infant formula, cereal milk powder, growing milk, pregnancy and lactating formula; and the special medical use formula includes premature/low birth weight infant formula, lactose-free formula Or low lactose formula, milk protein partially hydrolyzed formula, milk protein deep hydrolysis formula or amino acid formula, fortifiers or nutritional supplements for breast and infant formulas, for food intolerance, allergies, diseases or a dysfunctional formula; the parenteral nutrition preparation comprises a fat emulsion injection, an all-in-one nutrient solution, an intravenous injection; the enteral nutrition preparation comprises an amino acid enteral nutrition preparation, a short peptide type enteral nutrition preparation, Whole protein type enteral nutrition preparation, component type enteral nutrition preparation.

Food for Special Medical Purpose (FSMP) is a formula that is specially formulated to meet the special needs of people with limited food intake, digestive dysfunction, metabolic disorders or specific disease states. Such products must be consumed alone or in combination with other foods under the direction of a doctor or clinical nutritionist. Special medical use formulas are special dietary foods. When the target population cannot eat ordinary meals or can not meet their nutritional needs with daily diet, special medical use formulas can be used as a nutritional supplement, which plays an important nutritional support role in the treatment, rehabilitation and maintenance of body functions.

The invention also provides a preparation method of a butyric acid-containing nutritional preparation, the preparation method comprising: adding a butyric acid compound to the nutritional composition in proportion, and uniformly mixing; the butyric acid compound It is at least one selected from the group consisting of butyric acid, butyrate, and butyric acid derivatives.

Preferably, when the butyric acid compound is butyric acid, the butyric acid compound is added in the form of a coating or a coating.

It should be noted that, in the present invention, the butyric acid compound includes all forms of butyric acid, and the present invention is applicable not only to the above-mentioned respective butyric acid compounds but also to other butyric acid compounds. The butyric acid includes n-butyric acid and isobutyric acid.

It should be noted that the application form of butyric acid in the present invention is not important, and a butyric acid-containing substance such as butyric acid and/or butyrate and/or butyric acid derivative is added to a conventional formula, a special medical use formula, In the parenteral nutrition preparation and the enteral nutrition preparation, the effect of preventing and treating NEC can be achieved as long as an effective amount is applied.

As an alternative to formula application, the butyric acid, butyrate and butyric acid derivatives of the invention can be administered as a supplement rather than integrated into the formula. For example, butyric acid, butyrate and butyric acid derivatives can be ingested in the form of pills, tablets, capsules, caplets, powders, liquids or gels. For example, butyric acid, butyrate, and butyric acid derivatives can be combined with other nutritional supplements.

Compared with the prior art, the present invention has the following beneficial effects:

1. The present invention can effectively prevent necrotizing enterocolitis by adding an active ingredient, a butyric acid compound, to a nutritional preparation, including significantly reducing the incidence and mortality of necrotizing enterocolitis, and shortening the necrotizing small intestine colon The duration of inflammation, reducing its complications and/or reducing its severity. It also has the effects of promoting growth and development, immune system maturation and intestinal development.

The butyric acid compound added in the present invention has the following advantages:

First, as a direct source of energy for intestinal epithelial cells, it has the effect of promoting intestinal development;

Secondly, promote the development and improvement of the immune system of newborn individuals;

Finally, butyric acid compounds also have an important function that is easily overlooked, which promotes hemoglobin synthesis, thereby enhancing the body's oxygen supply capacity to resist intestinal damage caused by ischemia and hypoxia, and this is for the prevention and treatment of necrotizing small intestine colon Inflammation plays a vital role.

2. Butyric acid is a natural component found in mammalian milk and intestines. From a clinical application point of view, it is safer than other drugs for preventing and treating necrotizing enterocolitis. In this study, butyric acid compounds can be added in the form of a coating, which does not affect the application due to the acidity and odor of butyric acid.

3. Intravenous nutrition is extremely important or even the only nutritional support pathway for children with NEC risk or NEC. In the present invention, we have innovatively added butyric acid compounds to parenteral nutrition preparations, providing a new and more reliable intervention route for the prevention and treatment of NEC.

DRAWINGS

Other features, objects, and advantages of the present invention will become apparent from the Detailed Description of Description

Figure 1 is a schematic diagram of artificial feeding of oral intubation in neonatal rats;

Figure 2 is a general morphology diagram of the intestine of a newborn rat;

Figure 2a is a NEC model group;

Figure 2b shows the NEC model + butyric acid (100 mM, uncoated direct addition) intervention group;

Figure 2c is an NEC model + butyric acid (100 mM, added in the form of a capsule) intervention group;

Figure 2d shows the NEC model + sodium butyrate (100 mM) intervention group;

Figure 2e is a NEC model + butyrate (50 mM) intervention group;

Figure 2f is a NEC model + butyric acid (200 mM, uncoated direct addition) treatment group;

Figure 2g is a treatment group of NEC model + butyric acid (200 mM, added in the form of a capsule);

Figure 2h is a NEC model + sodium butyrate (200 mM) treatment group;

Figure 2i is a NEC model + butyrate (200 mM) treatment group;

Figure 3 is a diagram showing the histopathological changes of the intestine of newborn rats (200×);

Figure 3a is a NEC model group;

Figure 3b is a NEC model + butyric acid (100 mM, uncoated direct addition) intervention group;

Figure 3c is a NEC model + butyric acid (100 mM, added in the form of a capsule) intervention group;

Figure 3d shows the NEC model + sodium butyrate (100 mM) intervention group;

Figure 3e is a NEC model + butyrate (50 mM) intervention group;

Figure 3f is a NEC model + butyric acid (200 mM, uncoated direct addition) treatment group;

Figure 3g is a treatment group of NEC modeling + butyric acid (200 mM, added in the form of a capsule);

Figure 3h is a NEC model + sodium butyrate (200 mM) treatment group;

Figure 3i is a NEC model + butyrate (200 mM) treatment group.

Detailed ways

The invention will now be described in detail in connection with specific embodiments. The following examples are intended to further understand the invention, but are not intended to limit the invention in any way. It should be noted that a number of changes and modifications may be made by those skilled in the art without departing from the inventive concept. These are all within the scope of protection of the present invention.

The following Examples 1-6 provide a butyric acid-containing nutritional formulation comprising a nutritional composition and a butyric acid compound in a nutritional formulation at a concentration of from 0.5 to 200 mM (mmol/L).

The butyric acid compound is at least one selected from the group consisting of butyric acid, butyrate, and butyric acid derivatives.

The butyrate is selected from at least one of sodium butyrate, calcium butyrate, magnesium butyrate, and potassium butyrate.

The butyric acid derivative is at least one selected from the group consisting of glyceryl butyrate, mono-diglyceride butyrate, ethyl butyrate, methyl butyric acid, isoamyl butyrate, and butyric acid cyclodextrin.

The butyric acid compound is butyric acid and/or butyrate, and the concentration of butyric acid and/or butyrate in the nutritional preparation is from 1 to 200 mM.

When the concentration of the butyric acid and/or butyrate in the nutritional preparation is from 1 to 100 mM, especially at a concentration of from 50 to 100 mM, the nutritional preparation containing the concentration of butyric acid and/or butyrate can effectively prevent NEC. Occurs to promote growth, immune system development and intestinal maturation. When the concentration of the butyric acid and/or butyrate in the nutritional preparation is 100 to 200 mM, the nutritional preparation containing the concentration of butyric acid and/or butyrate is effective for treating NEC.

The butyric acid compound is a butyric acid derivative, and the concentration of the butyric acid derivative in the nutritional preparation is 0.5 to 200 mM.

When the concentration of the butyric acid derivative in the nutritional preparation is from 0.5 to 100 mM, particularly preferably from 50 to 100 mM, the nutritional preparation containing the butyric acid derivative of the concentration can effectively prevent NEC. When the concentration of the butyric acid derivative in the nutritional preparation is 100 to 200 mM, the nutritional preparation containing the butyric acid derivative of the concentration can effectively treat NEC.

The nutritional composition is selected from at least one of a conventional formula, a special medical use formula, a parenteral nutrition preparation, and an enteral nutrition preparation.

The conventional formula includes infant formula, infant formula, cereal milk powder, growing milk, pregnancy and lactating formula; and the special medical use formula includes premature/low birth weight infant formula, lactose-free formula or low lactose Formula, milk protein partially hydrolyzed formula, milk protein deep hydrolysis formula or amino acid formula, fortifiers or nutritional supplements for breast and infant formulas, for food intolerance, allergies, diseases or dysfunction a formula; the parenteral nutrition preparation comprises a fat emulsion injection, an all-in-one nutrient solution, an intravenous injection; the enteral nutrition preparation comprises an amino acid enteral nutrition preparation, a short peptide type enteral nutrition preparation, and a whole protein type Enteral nutrition preparation, component type enteral nutrition preparation.

Example 1

The present embodiment provides a nutrient preparation containing sodium butyrate and a preparation method thereof, which comprises: adding food grade or injection grade sodium butyrate to a nutrient preparation, and uniformly mixing, that is, obtaining. The concentration of sodium butyrate in the prepared nutritional preparation is from 1 to 100 mM.

Example 2

The present embodiment provides a nutrient preparation containing sodium butyrate and a preparation method thereof, which comprises: adding food grade or injection grade sodium butyrate to a nutrient preparation, and uniformly mixing, that is, obtaining. The concentration of sodium butyrate in the prepared nutritional preparation is 100 to 200 mM.

Example 3

The present embodiment provides a nutrient preparation containing glyceryl butyrate and a preparation method thereof, which comprises: adding food grade or injection grade butyric acid glyceride to a nutrient preparation, and uniformly mixing, that is, obtaining. The concentration of butyric acid glyceride in the prepared nutritional preparation is from 0.5 to 100 mM.

Example 4

The present embodiment provides a nutrient preparation containing glyceryl butyrate and a preparation method thereof, which comprises: adding food grade or injection grade butyric acid glyceride to a nutrient preparation, and uniformly mixing, that is, obtaining. The concentration of butyric acid glyceride in the prepared nutritional preparation is 100 to 200 mM.

Example 5

The present embodiment provides a nutrient preparation containing butyric acid and a preparation method thereof, which comprises: adding food grade butyric acid or injection grade butyric acid into a nutrient preparation in the form of a capsule, and uniformly mixing, that is, obtained. The concentration of butyric acid in the prepared nutritional preparation is from 1 to 100 mM.

Example 6

The present embodiment provides a nutrient preparation containing butyric acid and a preparation method thereof, which comprises: adding food grade butyric acid or injection grade butyric acid into a nutrient preparation in the form of a capsule, and uniformly mixing, that is, obtained. The prepared nutritional preparation has a concentration of butyric acid of 100 to 200 mM.

The butyric acid-containing nutrient preparation prepared in the above examples can effectively prevent and treat NEC, and has the functions of promoting body weight growth, immune system maturation and intestinal development.

Example 7

This embodiment provides an application of a butyric acid compound in the preparation of a medicament for preventing and treating necrotizing enterocolitis.

The butyric acid compound is selected from at least one of butyric acid or a pharmaceutically acceptable salt or derivative thereof.

The pharmaceutically acceptable salt of butyric acid is selected from at least one of sodium butyrate, potassium butyrate, calcium butyrate, and magnesium butyrate.

The pharmaceutically acceptable derivative of butyric acid is selected from the group consisting of glyceryl butyrate, mono-diglyceride butyrate, ethyl butyrate, methyl butyric acid, isoamyl butyrate, and butyric acid cyclodextrin complex. At least one.

The medicament includes a therapeutically effective amount of a butyric acid compound.

The effective dose of the butyric acid compound is 0.05 to 20 mmol/kg/d.

The medicament also includes other medicaments compatible with the butyrate compound, as well as pharmaceutically acceptable carriers and/or adjuvants.

The drug is in a pharmaceutically acceptable dosage form.

The dosage form is a powder, an injection, a capsule, a tablet or an oral solution.

When the butyric acid compound is butyric acid, the butyric acid compound is added to the drug in the form of a coating or a coating.

Verification of animal test results: effects of nutrient preparations containing butyric acid in the prevention and treatment of NEC

1. Experimental animals and grouping

Clean-grade newborn Sprague-Dawley (SD) rats, born on day 1, were male or female, weighing about 5-6 g, purchased from Shanghai Xipuer-Beikai Experimental Animal Co., Ltd. Raised at the Animal Experimental Center of Xinhua Hospital affiliated to Shanghai Jiao Tong University School of Medicine. Rat milk substitutes were prepared as reported by Nancy Auestad et al. and Gail Besner Laboratories.

2. Processing method

Artificial feeding control group: Newborn rats were separated from female rats and placed in an incubator. Artificial feeding of mouse milk substitutes was regularly fed at 0.15ml/6h/day;

NEC model group: the newborn rats were separated from the mother rats and placed in the incubator. The artificial milk feeding substitutes were 0.15ml/6h/day, and fed regularly. After 1 day of artificial feeding, hypoxia and cold stimulation were given twice a day. Lasts for three days;

NEC model + butyric acid (not coated directly) intervention group: newborn rats and mother rats were separated, placed in the incubator, artificial feeding of uncoated enveloped butyric acid substitute milk substitute 0.15ml/6h / day timing Feeding; after 1 day of artificial feeding, hypoxia and cold stimulation were given twice daily for three days;

NEC model + butyric acid (added in the form of capsule) intervention group: newborn rats were separated from the mother rats and placed in the incubator. Artificial feeding of rat milk substitute with coated butyric acid was applied at 0.15ml/6h/day. After 1 day of artificial feeding, hypoxia and cold stimulation were given twice daily for three days;

NEC model + sodium butyrate intervention group: the newborn rats were separated from the mother rats and placed in the incubator. The artificial milk supplemented with sodium butyrate was used for 0.15ml/6h/day feeding; after artificial feeding for 1 day, Hypoxia and cold stimulation twice daily for three days;

NEC model + butyric acid glyceride intervention group: newborn rats and mother rats were separated, placed in the incubator, artificial feeding of butyric acid glyceride substitute milk substitute 0.15ml/6h / day feeding; artificial feeding after 1 day Give hypoxia and cold stimulation twice daily for three days;

NEC model + butyric acid (not coated directly) treatment group: newborn rats and mother rats were separated, placed in the incubator, artificially fed mouse milk substitute 0.15ml/6h / day timing feeding; artificial feeding 1 day, Stimulate twice daily with hypoxia and cold for three days, and switch to a mouse milk substitute with uncoated butyric acid for treatment;

NEC model + butyric acid (added in the form of capsule) treatment group: newborn rats and mother rats were separated, placed in the incubator, artificially fed mouse milk substitute 0.15ml/6h / day timing feeding; 1 day after artificial feeding, Stimulate twice daily with hypoxia and cold for three days, and switch to a mouse milk substitute with coated butyric acid for treatment;

NEC model + sodium butyrate treatment group: newborn rats and mother rats were separated, placed in the incubator, artificially fed mouse milk substitute 0.15ml/6h / day timing feeding; artificial feeding 1 day, with hypoxia and cold stimulation Twice a day for three days, and switched to a rat milk substitute with sodium butyrate for treatment;

NEC model + butyric acid glyceride treatment group: newborn rats and mother rats were separated, placed in the incubator, artificially fed mouse milk substitute 0.15ml/6h / day timing feeding; artificial feeding 1 day, given hypoxia and cold Stimulate twice daily for three days and switch to a rat milk substitute with butyrate added for treatment;

The rats were fed with a clean PICC tube (Fig. 1), and the skin around the mouth of the newborn rats was cleaned with physiological saline before feeding. After feeding, the PICC was washed and disinfected with 75% alcohol.

3. Modeling method

Hypoxia and cold stimulation: Zero the oxygen meter and connect it to a closed anoxic chamber. Nitrogen was turned on and the flow rate was controlled to be about 15 L/min. The time was started when the oxygen concentration dropped to zero. After 90 seconds, the newborn rats were taken out and immediately placed in a refrigerator at 4 ° C for 10 minutes. Oxygen and cold stimulation were performed twice a day, and returned to the incubator after treatment.

After establishing the model, closely observe the feeding, defecation, abdominal condition and activity response of newborn rats. The body weight of the newborn rats was weighed before the experiment and after the modeling. Neonatal rats were sacrificed when clinical symptoms (severe bloating, bloody stools and cyanosis) appeared or on day 4 (96 hours) after modeling. The general morphology of the intestines was observed and intestinal and blood samples were collected.

4. Intestinal histopathology

The 2 cm intestine of the distal intestine was retrieved, and the cold saline was quickly washed, and the contents of the intestine were removed, and then fixed in the tissue fixing solution for 24-48 hours. After HE staining, the morphological changes of the intestinal tissue were observed under light microscope, and the pathological score was independently performed by the pathologist (double-blind method).

Refer to the scoring criteria used by Michael Caplan and Gail Besner:

0 points: The intestinal mucosa was intact and the tissue structure was normal.

1 point: slight submucosal and/or lamina propria separation.

2 points: moderate submucosal and/or lamina propria, or submucosal and muscular edema.

3 points: severe submucosa and / or lamina propria, or severe edema of the submucosa and muscle layer, local villi detachment.

4 points: Intestinal villi disappeared with full layer necrosis of the intestinal wall.

A score of ≧ 2 is considered as NEC.

5. Results

5.1 growth situation:

Control group: Rats had normal eating and bowel movements, and the activity was good. There were no symptoms such as bloating and bloody stools. During the experiment, the body weight increased and no death occurred.

NEC model group: After the model was established, the rats gradually showed different degrees of abdominal distension, decreased milk intake, slow response, decreased activity, slow movement, body blemishes, and dark redness. The weight was significantly reduced after modeling. The rat died within 72 hours after modeling;

NEC model + butyric acid (100 mM, uncoated directly added) intervention group: rats developed abdominal distension, slow movement, body blemishes, and dark red stools after modeling, and some rats died during modeling.

NEC model + butyric acid (100 mM, added in the form of capsule) intervention group: normal bowel movements in rats, good activity, ruddy body, no abdominal distension and blood in the stool, weight gain before modeling, and slight weight loss after modeling;

NEC model + sodium butyrate (100 mM) intervention group: rats with normal bowel movements, good activity, ruddy body, no abdominal distension and blood in the stool, weight gain before modeling, and slight weight loss after modeling;

NEC model + butyric acid glyceride (50 mM) intervention group: rats with normal bowel movements, good activity, ruddy body, no abdominal distension and blood in the stool, weight gain before modeling, and slight weight loss after modeling;

NEC model + butyric acid (200 mM, uncoated directly added) treatment group: rats developed abdominal distension, slow movement, body blemishes, and dark red stools after modeling, most rats died during modeling .

NEC model + butyric acid (200 mM, added in the form of a capsule) treatment group: the rats had better activity during the modeling period, the body was ruddy, no blood in the stool, and the body weight was slightly reduced after modeling;

NEC model + sodium butyrate (200 mM) treatment group: the rats had better activity during the modeling period, the body was ruddy, no blood in the stool, and the weight was slightly reduced after modeling.

NEC model + butyric acid glyceride (200 mM) treatment group: occasionally bloating during the model establishment, the activity was better, the body was ruddy, no blood in the stool, and the weight was slightly reduced after modeling.

5.2 intestinal tube situation (Figure 2):

Control group: the color of the intestine of the rat was rosy and the elasticity was good;

NEC model group: edema, hyperemia and necrosis of rat intestinal wall, reddish black, showing intestinal thinning, stenosis and even obstruction; as shown in Figure 2a;

NEC model + butyric acid (100 mM, uncoated directly added) intervention group: rat intestinal wall edema, congestion, necrosis, reddish black, intestinal thinning, stenosis and even obstruction; as shown in Figure 2b;

NEC model + butyric acid (100 mM, added in the form of capsule) intervention group: the color of the intestinal tube was normal, no congestive necrosis, no narrowing and narrowing, compared with the control group, the intestine was thicker and normal, as shown in Figure 2c. ;

NEC model + sodium butyrate (100 mM) intervention group: the color of the intestinal tube was normal, no congestive necrosis, no narrowing and narrowing. Compared with the control group, the intestinal tube was thicker and normal, as shown in Figure 2d.

NEC model + butyric acid glyceride (50 mM) intervention group; rat intestinal tube was thick, no narrowing, normal morphology, no congestive necrosis of the intestine, as shown in Figure 2e;

NEC model + butyric acid (200 mM, uncoated directly added) treatment group: rat intestinal wall edema, congestion, necrosis, reddish black, visible intestinal thinning, stenosis and even obstruction, as shown in Figure 2f;

NEC model + butyric acid (200 mM, added in the form of capsule) treatment group: normal color of the intestine, no congestion and necrosis, no narrowing and narrow, normal morphology, as shown in Figure 2g;

NEC model + sodium butyrate (200 mM) treatment group: normal color of the intestine, no congestion and necrosis, no narrowing and narrowing, normal morphology, thicker intestine, as shown in Figure 2h;

NEC model + butyric acid glyceride (200 mM) treatment group: normal bowel color, no congestion and necrosis, no narrowing, normal morphology, as shown in Figure 2i.

5.3 pathological conditions (Figure 3):

Control group: ileal villus intact, epithelial continuous, gland arrangement rules;

NEC model group: different degrees of mucosal and submucosal congestion and edema in the ileum, partial necrosis and necrosis, disordered glandular arrangement, mild to moderate separation of mucosal muscle layer, thinning or even rupture of muscle layer, inflammatory cells such as neutral Infiltration of granulocytes and lymphocytes, severe intestinal villus disappearance with intestinal necrosis, similar to the pathological changes in children with clinical NEC; as shown in Figure 3a;

NEC model + butyric acid (100 mM, uncoated directly added) intervention group: mucosal layer and muscle layer separation in the ileum, mucosal and submucosal congestion, edema, partial villus necrosis, glandular arrangement disorder, inflammatory cells such as Infiltration of neutrophils and lymphocytes, accompanied by disappearance of intestinal villi and intestinal necrosis, as shown in Figure 3b;

NEC model + butyric acid (100 mM, added in the form of capsule) intervention group: ileal villus intact structure, long villi, deep crypt, continuous epithelium, regular glandular arrangement, no expansion of lamina propria, no mucosal congestion, no Significant inflammatory cell infiltration, as shown in Figure 3c;

NEC model + sodium butyrate (100 mM) intervention group: ileal villus intact structure, long villus, deep crypt, continuous epithelium, regular arrangement of glands, no expansion of lamina propria, no mucosal congestion, no obvious inflammatory cell infiltration, As shown in Figure 3d;

NEC model + butyric acid glyceride (50 mM) intervention group: ileal villus intact structure, long villus, deep crypt, continuous epithelium, regular arrangement of glands, no expansion of lamina propria, no mucosal congestion, no obvious inflammatory cell infiltration , as shown in Figure 3e;

NEC model + butyric acid (200 mM, uncoated directly added) treatment group: mucosal layer and muscle layer separation in the ileum, mucosal layer and submucosa congestion, edema, partial villus necrosis, gland arrangement disorder, inflammatory cells such as Infiltration of granulocytes and lymphocytes, accompanied by disappearance of intestinal villi and intestinal necrosis, as shown in Figure 3f;

NEC model + butyric acid (200 mM, added in the form of capsule) treatment group: ileal villus intact structure, long villus, deep crypt, continuous epithelium, regular glandular arrangement, no expansion of lamina propria, no obvious mucosal congestion and Infiltration of inflammatory cells, as shown in Figure 3g;

NEC model + sodium butyrate (200 mM) treatment group: ileal villus intact structure, long villus, deep crypt, continuous epithelium, regular arrangement of glands, no expansion of lamina propria, no mucosal hyperemia and inflammatory cell infiltration, as shown 3h;

NEC model + butyric acid glyceride (200 mM) treatment group: ileal villus intact structure, long villus, deep crypt, continuous epithelium, regular arrangement of glands, no expansion of lamina propria, no obvious mucosal congestion and inflammatory cell infiltration, As shown in Figure 3i.

5.4 Development of the immune system:

Since the immune system of newborn rats has not yet matured, its ability to regulate the body's immune inflammatory response is poor. After intervention and treatment with sodium butyrate, butyrate and coated butyric acid, the maturation and proliferation of neonatal rat immune cells were promoted, and the ratio was closer to that of adult rats.

5.5 Incidence and mortality:

Control group: The incidence of NEC was 0% and the survival rate was 100%.

NEC model: NEC has a 100% incidence and a survival rate of less than 20%.

Morbidity and mortality in the intervention group:

NEC model + butyric acid (100 mM, uncoated directly added) intervention group: NEC incidence rate of 100%, survival rate of less than 5%.

NEC model + butyric acid (100 mM, added in the form of a capsule) intervention group: the incidence of NEC decreased to 15%, and the survival rate reached 90%.

NEC model + butyric acid (50 mM, added in the form of a capsule) intervention group: the incidence of NEC decreased to 20%, and the survival rate reached 85%.

NEC model + butyric acid (1 mM, added in the form of a capsule) intervention group: the incidence of NEC was 80%, and the survival rate was 35%.

NEC model + sodium butyrate (100 mM) intervention group: the incidence of NEC decreased to 10%, and the survival rate reached 95%.

NEC model + sodium butyrate (50 mM) intervention group: the incidence of NEC decreased to 15%, and the survival rate reached 90%.

NEC model + sodium butyrate (1 mM) intervention group: the incidence of NEC decreased slightly to 85%, and the survival rate was 35%.

NEC model + butyric acid glyceride (100 mM) intervention group: the incidence of NEC decreased to 15%, and the survival rate reached 95%.

NEC model + butyric acid glyceride (50 mM) intervention group: the incidence of NEC decreased to 20%, and the survival rate reached 90%.

NEC model + butyrate (0.5 mM) intervention group: the incidence of NEC was 90%, and the survival rate was 30%.

Treatment group morbidity and mortality:

NEC model + butyric acid (200 mM, uncoated directly added) treatment group: NEC incidence rate of 100%, survival rate of less than 10%.

NEC model + butyric acid (100 mM, added in the form of a capsule) treatment group: the incidence of NEC decreased to 20%, and the survival rate reached 80%.

NEC model + butyric acid (200 mM, added in the form of a capsule) treatment group: the incidence of NEC decreased to 15%, and the survival rate reached 85%.

NEC model + butyric acid (300 mM, added in the form of a capsule) treatment group: the incidence of NEC was 25%, and the survival rate was 80%.

NEC model + sodium butyrate (100 mM) treatment group: the incidence of NEC decreased to about 20%, and the survival rate reached 85%.

NEC model + sodium butyrate (200 mM) treatment group: the incidence of NEC decreased to about 15%, and the survival rate reached 90%.

NEC model + sodium butyrate (300 mM) treatment group: the incidence of NEC was 20%, and the survival rate was 85%;

NEC model + butyric acid glyceride (100 mM) treatment group: the incidence of NEC decreased to 20%, and the survival rate reached 85%.

NEC model + butyric acid glyceride (200 mM) treatment group: the incidence of NEC decreased to 15%, and the survival rate reached 90%.

NEC model + butyric acid glyceride (300 mM) treatment group: the incidence of NEC was 20%, and the survival rate reached 85%.

In summary, the NEC model was constructed by artificial feeding combined with hypoxia and cold stimulation, and intervention or treatment was performed using uncoated butyric acid, coated butyric acid, butyrate or butyric acid derivatives. The results showed that the direct addition of uncoated butyric acid for intervention, because of its weak acidity, increased the condition of NEC, and the mortality increased; while the intervention and treatment with enveloped butyric acid effectively reduced the morbidity and mortality of NEC. The addition of butyrate and butyric acid derivatives is also very effective in the prevention and treatment of NEC. The incidence rate of the NEC model group was 100%, and the survival rate was less than 20%. After a prophylactic intervention with an effective dose of butyrate, the incidence of NEC can be reduced to a minimum of 10% and the survival rate can reach up to 95%. After treatment with an effective dose of butyrate, the incidence of NEC can be reduced to a minimum of 15% and the survival rate can be as high as 90%. In the NEC model group, intestinal wall edema, hyperemia and necrosis appeared in red-black color, while NEC model + butyrate intervention or treatment group had normal intestinal color, no congestion and necrosis, and no obvious lesions. The pathological results showed that the ileum of the NEC model group showed hyperemia, edema, inflammatory cell infiltration, partial necrosis of nevus, disordered gland arrangement, mild to moderate separation of mucosal muscle layer, thinning or even fracture of muscle layer; In the intervention or treatment group of NEC model + butyric acid compound, the ileum was intact, the villus was long, the crypt was deep, the epithelium was continuous, the gland was arranged regularly, the lamina propria was not dilated, and there was no obvious mucosal hyperemia and inflammatory cell infiltration. Therefore, our research has subverted the long-term role of short-chain fatty acid-butyric acid in the pathogenesis of NEC, confirming that the effect of butyric acid compounds in the prevention and treatment of NEC is significantly better than other drugs reported at present, while butyric acid The compounds also have the effect of promoting growth of newborn rats, development of the immune system, and development of the gut. Moreover, butyric acid is a natural component in mammalian milk and is safer than other drugs from a clinical application point of view. Therefore, the addition of butyric acid compounds to nutritional preparations as a safe, economical and effective method is of great significance for the prevention and treatment of NEC.

The specific embodiments of the present invention have been described above. It is to be understood that the invention is not limited to the specific embodiments described above, and various changes or modifications may be made by those skilled in the art without departing from the scope of the invention. The features of the embodiments and the embodiments of the present application may be combined with each other arbitrarily without conflict.

Claims

A use of a butyric acid compound for the preparation of a medicament for preventing and treating necrotizing enterocolitis.
The use according to claim 1, wherein the butyric acid compound is at least one selected from the group consisting of butyric acid or a pharmaceutically acceptable salt or derivative thereof.
The use according to claim 2, wherein the pharmaceutically acceptable salt of butyric acid is at least one selected from the group consisting of sodium butyrate, potassium butyrate, calcium butyrate, and magnesium butyrate.
The use according to claim 2, wherein the pharmaceutically acceptable derivative of butyric acid is selected from the group consisting of glyceryl butyrate, mono-diglyceride butyrate, ethyl butyrate, methyl butyric acid, butyric acid. At least one of isoamyl ester and butyric acid cyclodextrin complex.
The use according to claim 2 wherein the medicament comprises a therapeutically effective amount of a butyric acid compound.
The use according to claim 5, characterized in that the effective dose of the butyric acid compound is 0.05 to 20 mmol/kg/d.
The use according to claim 1, wherein the medicament further comprises other drugs compatible with the butyric acid compound and a pharmaceutically acceptable carrier and/or adjuvant.
The use according to claim 7, wherein the medicament is in a pharmaceutically acceptable dosage form.
An use according to claim 8, characterized in that the dosage form is a powder, an injection, a capsule, a tablet or an oral solution.
A use of a butyric acid compound in the preparation of a nutritional preparation for preventing and treating necrotizing enterocolitis.
A nutrient preparation containing butyric acid, comprising a nutritional composition and a butyric acid compound, wherein the concentration of the butyric acid compound in the nutritional preparation is from 0.5 to 200 mM.
The butyric acid-containing nutrient preparation according to claim 11, wherein the butyric acid compound is at least one selected from the group consisting of butyric acid, butyrate, and butyric acid derivatives.
The butyric acid-containing nutritional preparation according to claim 12, wherein the butyrate is at least one selected from the group consisting of sodium butyrate, potassium butyrate, calcium butyrate, and magnesium butyrate.
The butyric acid-containing nutritional preparation according to claim 12, wherein the butyric acid derivative is selected from the group consisting of glyceryl butyrate, mono-diglyceride butyrate, ethyl butyrate, methyl butyric acid, butyric acid. At least one of isoamyl ester and butyric acid cyclodextrin complex.
The butyric acid-containing nutritional preparation according to claim 12, wherein the butyric acid compound is butyric acid and/or butyrate, and the concentration of butyric acid and/or butyrate in the nutritional preparation is 1 ~200 mM.
The butyric acid-containing nutritional preparation according to claim 12 or 14, wherein the butyric acid compound is a butyric acid derivative, and the concentration of the butyric acid derivative in the nutritional preparation is from 0.5 to 200 mM.
The butyric acid-containing nutritional preparation according to claim 11, wherein the nutritional composition is at least one selected from the group consisting of a conventional formula, a special medical use formula, a parenteral nutrition preparation, and an enteral nutrition preparation.
The butyric acid-containing nutritional preparation according to claim 17, wherein the conventional formula comprises infant formula, infant formula, cereal milk powder, growing milk, pregnancy and lactating formula; said special medical use Formulated foods include preterm/low birth weight infant formula, lactose-free or low-lactose formula, milk protein partially hydrolyzed formula, milk protein deep hydrolysis formula or amino acid formula, fortifiers or nutritional supplements for breast and infant formulas Medicament, a formula suitable for food intolerance, allergy, disease or dysfunction; the parenteral nutrition preparation comprises a fat emulsion injection, an all-in-one nutrient solution, an intravenous injection; the enteral nutrition preparation comprises Amino acid type enteral nutrition preparation, short peptide type enteral nutrition preparation, whole protein type enteral nutrition preparation, and component type enteral nutrition preparation.
A method for preparing a butyric acid-containing nutritional preparation according to claim 11, wherein the preparation method comprises: adding a butyric acid compound to the nutritional composition in proportion, and mixing uniformly; The butyric acid compound is at least one selected from the group consisting of butyric acid, butyrate, and butyric acid derivatives.
The method for producing a butyric acid-containing nutritional preparation according to claim 19, wherein when the butyric acid compound is butyric acid, the butyric acid compound is added in the form of a coating or a coating.