KR102537907B1 - Peptide(PEPTIDE 15) for anti-obesity and use thereof - Google Patents

Abstract

The present invention relates to a peptide exhibiting anti-obesity efficacy or an anti-obesity composition containing the same. According to the present invention, since the composition exhibited a significant differentiation inhibitory effect on adipocytes, it is expected that a more fundamental approach to target treatment in the prevention or treatment of obesity will be possible.

Description

Peptide (Peptide 15) having anti-obesity effect and its use {Peptide (PEPTIDE 15) for anti-obesity and use thereof}

The present invention relates to peptides exhibiting anti-obesity efficacy and uses thereof.

Obesity is caused by abnormal hypertrophy of subcutaneous fat tissue caused by accumulation of excess energy in the body as fat when an imbalance in metabolic processes occurs due to endocrine factors, genetic factors, social and environmental factors, and the like. Adipose tissue hypertrophy is a phenomenon in which adipocytes increase in size (adipocyte hypertrophy) or increase in number (adipocyte hyperplasia), which also affects the stasis of the local venous-lymphatic system, resulting in dermal-subcutaneous vascular tissue disease. can cause Therefore, obesity is recognized as an independent disease in itself, and the World Health Organization treats obesity as a global nutritional problem and recognizes it as a disease to be treated rather than a simple risk factor that harms health.

Current anti-obesity drugs can be classified into drugs that affect appetite by acting on the central nervous system and drugs that inhibit absorption by acting on the gastrointestinal tract according to their mechanism of action. Drugs that act on the central nervous system include fenfluramine and dexfenfluramine, which inhibit serotonin (5-HT) reuptake, ephedrine and caffeine, which inhibit noradrenaline reuptake, and recently, sibutramine, which simultaneously acts on serotonin and noradrenaline nervous systems. there is. As a drug that inhibits obesity by acting on the gastrointestinal tract, there is orlistat, which is approved as a treatment for obesity by inhibiting lipase produced in the pancreas to reduce fat absorption. However, among previously used drugs, fenfluramine, an appetite suppressant, was not only classified as a narcotic component, but also banned because of side effects such as primary pulmonary hypertension or heart valve lesions. Its use is limited in patients with heart failure and renal disease. Orlistat, a lipolysis inhibitor, also causes side effects in the gastrointestinal tract, causing discomfort and hindering the absorption of fat-soluble vitamins. this was banned

One of the alternatives for reducing the side effects of the above drugs and preventing and suppressing more fundamental obesity is an adipocyte differentiation inhibitor. Currently known secreted substances include TNF-α, IL-6, IL-8, plasminogen activator inhibitor-1, angiotensin-II, leptin, and adiponectin, but these are known to cause various metabolic diseases and cardiovascular diseases. known to cause disease. Energy storage and secretion of endocrine substances in adipose tissue are closely related to the differentiation and growth of adipocytes and occur simultaneously with the adipogenesis process. The growth of adipocytes is a process in which hypertrophic adipocytes are formed by accumulation of neutral fat in differentiated adipocytes without a change in the number of adipocytes. possible.

Under this technical background, research on new anti-obesity substances is being actively conducted (Korean Registered Patent No. 10-1662062), but it is still incomplete.

An object of the present invention is to provide a peptide exhibiting anti-obesity efficacy and a composition for preventing or treating obesity containing the same as an active ingredient.

In addition, another object of the present invention is to provide a composition for external application for skin for preventing or improving obesity, comprising the peptide as an active ingredient.

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems not mentioned will be clearly understood by those skilled in the art from the following description.

The present invention was made to solve the above problems, and the present inventors synthesized peptides having no cytotoxicity and exhibiting anti-obesity efficacy and confirming their significant adipocyte differentiation inhibitory effect, that is, the anti-obesity effect. has completed

Accordingly, the present invention provides a peptide exhibiting anti-obesity efficacy.

As used herein, the term “obesity” means excessive accumulation of body fat in the body.

Specifically, the present invention may be a peptide consisting of one of the amino acid sequences represented by SEQ ID NO: 1 to SEQ ID NO: 23 showing anti-obesity activity.

As used herein, the term "peptide" refers to a linear molecule formed by binding amino acid residues to each other by a peptide bond. The peptides of the present invention can be synthesized using chemical synthesis methods known in the art, particularly solid-phase synthesis techniques; Merrifield, J. Amer. Chem. Soc. 85:2149-54 (1963); Stewart, et al., Solid Phase Peptide Synthesis, 2nd. ed., Pierce Chem. Co.: Rockford, 111 (1984)) or liquid synthesis technology (US Patent No. 5,516,891), but is not limited thereto. Peptide synthesis methods can be used.

The peptide of the present invention may select a portion of the amino acid sequence and induce modification at the N-terminus or C-terminus to increase its activity. Through this modification, the peptide of the present invention can have a high half-life that increases the half-life upon in vivo administration.

In addition, the C-terminus of the peptide of the present invention is modified with a hydroxy group (-OH), amino group (-NH ₂ ), azide (-NHNH ₂ ), etc., and the N-terminus of the peptide is acetyl group, fluorenyl methyl A protecting group selected from the group consisting of oxycarbonyl group, formyl group, palmitoyl group, myristyl group, stearyl group, and polyethylene glycol (PEG) may be bound.

Modification of the amino acids described above serves to greatly improve the stability of the peptides of the present invention. In this specification, the term "stability" means not only in vivo stability, but also storage stability (eg, storage stability at room temperature). The above-described protecting group serves to protect the peptide of the present invention from attack by proteolytic enzymes in vivo.

In addition, the present invention provides a pharmaceutical composition for preventing or treating obesity comprising the peptide as an active ingredient.

In addition, the present invention provides a composition for external application for skin for preventing or improving obesity, comprising the peptide as an active ingredient.

Since the contents related to the peptides in the composition are the same as those described above, the description of duplicate contents will be omitted.

The composition can be applied directly to local areas with a lot of subcutaneous fat, such as the face, abdomen, thighs, buttocks, arms, etc., to locally reduce fat in these areas, and is formulated in the form of an external skin preparation suitable for this purpose. However, the formulation is not particularly limited. In addition, in the composition of one aspect of the present invention, the composition can reduce fat in the body by drinking or taking for oral administration, and may be in a liquid dosage form for oral administration to be suitable for this purpose. The composition of the present invention, in addition to the above components, is appropriately blended with auxiliary components commonly used in the field, and is formulated into tablets, capsules, soft capsules, pills, granules, and drinks, as health supplements or pharmaceuticals. can be used

The present invention relates to a peptide exhibiting anti-obesity efficacy and an anti-obesity composition comprising the same as an active ingredient. According to the present invention, since the peptide and the composition containing the peptide showed a significant differentiation inhibitory effect on adipocytes, it is expected that a more fundamental approach to target treatment in the prevention or treatment of obesity will be possible.

1 shows the result of confirming the change in the differentiation rate and fat accumulation from 3T3-L1 preadipocytes to adipocytes according to the treatment of peptides.
Figure 2 shows the results of a cytotoxicity test by measuring the survival rate of 3T3-L1 preadipocytes according to the treatment of peptides.

Hereinafter, a preferred embodiment is presented to aid understanding of the present invention. However, the following examples are provided to more easily understand the present invention, and the content of the present invention is not limited by the following examples.

[Experimental example]

Experimental Example 1. Experimental Materials and Experimental Methods

Synthesis of 1-1 Peptides

23 types of peptide samples were obtained by requesting the synthesis of peptides to Peptron Co., Ltd. A specific peptide synthesis method is as follows. Peptide synthesis was performed using the solid phase Fmoc Chemistry method through continuous flow conditions using an organic chemical reaction. This is a method of synthesizing each amino acid in resin one by one in a predetermined order. Peptron Co., Ltd. synthesized 48 different peptides at the same time using an automatic synthesizer developed by itself. The synthesized crude peptide was purified using reverse phase HPLC and dried using a lyophilizer (Table 1).

sequence number Amino acid sequence information chain length molecular composition Molecular Weight
(Da) One KLPFQR
(Lys-Leu-Pro-Phe-Gln-Arg) 6 C37H61N11O8 787 2 STELLIR (Ser-Thr-Glu-Leu-Lue-Ile-Arg) 7 C36H66N10O12 830 3 EIAQDFK (Glu-Ile-Ala-Gln-Asp-Phe-Lys) 7 C38H59N9O13 849 4 HLQLAIR (His-Leu-Gln-Leu-Ala-Ile-Arg) 7 C38H67N13O9 849 5 AVQGLLK (Ala-Val-Gln-Gly-Leu-Leu-Lys) 7 C33H61N9O9 727 6 IAQGGVLP (Ile-Ala-Gln-Gly-Gly-Val-Leu-Pro) 8 C34H59N9O10 753 7 NDEELNKLL (Asn-Asp-Glu-Glu-Leu-Asn-Lys-Leu-Leu) 9 C46H78N12O18 1,086 8 AGLQFPVGR (Ala-Gly-Leu-Gln-Phe-Pro-Val-Gly-Arg) 9 C43H69N13O11 943 9 YRPGTVALR (Tyr-Arg-Pro-Gly-Thr-Val-Ala-Leu-Arg) 9 C46H77N15O12 1,031 10 KSTGGKAPR (Lys-Ser-Thr-Gly-Gly-Lys-Ala-Pro-Arg) 9 C37H68N14O12 900 11 KQLATKAAR (Lys-Gln-Leu-Ala-Thr-Lys-Ala-Ala-Arg) 9 C42H79N15O12 985 12 RFQSSAVMA (Arg-Phe-Gln-Ser-Ser-Ala-Val-Met-Ala) 9 C42H69N13O13S1 995 13 TLSDYNIQK (Thr-Leu-Ser-Asp-Tyr-Asn-Ile-Gln-Lys) 9 C47H76N12O17 1,080 14 NKLLSGVTIA (Asn-Lys-Leu-Leu-Ser-Gly-Val-Thr-Ile-Ala) 10 C45H82N12O14 1,014 15 EIAQDFKTDL (Glu-Ile-Ala-Gln-Asp-Phe-Lys-Thr-Asp-Leu) 10 C52H82N12O19 1,178 16 ALNQAWAFLK
(Ala-Leu-Asn-Gln-Ala-Trp-Ala-Phe-Leu-Lys) 10 C56H84N14O13 1,160 17 AERVGAGAPVY Ala-Glu-Arg-Val-Gly-Ala-Gly-Ala-Pro-Val-Tyr 11 C48H76N14O15 1,088 18 RIVDFHMLESR Arg-Ile-Val-Asp-Phe-His-Met-Leu-Glu-Ser-Arg 11 C61H99N19O17S1 1,401 19 SGVTIAQGGVLP
Ser-Gly-Val-Thr-Ile-Ala-Gln-Gly-Gly-Val-Leu-Pro 12 C48H83N13O16 1,097 20 APRKQLATKAAR Ala-Pro-Arg-Lys-Gln-Leu-Ala-Thr-Lys-Ala-Ala-Arg 12 C56H103N21O15 1,309 21 QNIIPASTGAAK
Gln-Asn-Ile-Ile-Pro-Ala-Ser-Thr-Gly-Ala-Ala-Lys 12 C50H87N15O17 1,169 22 HLQLAIRNDEELNK (His-Leu-Gln-Leu-Ala-Ile-Arg-Asn-Asp-Glu-Glu-Leu-Asn-Lys) 14 C72H121N23O24 1,691 23 VTIAQGGVLPNIQA (Val-Thr-Ile-Ala-Gln-Gly-Gly-Val-Leu-Pro-Asn-Ile-Gln-Ala) 14 C61H105N17O19 1,379

1-2 Cell culture and fat measurement method

The 3T3-L1 cell line, a mast cell, was purchased from ATCC and used. Penicillin (100 U/mL) and streptomycin (100 μg/mL) were added to DMEM containing 10% calf serum at 37˚C, 5% Cultured and maintained under CO ₂ . For the differentiation of 3T3-L1 pre-adipocytes into adipocytes, 5×10 ⁵ cells/well of cells were dispensed in a 6-well plate, and the cells were cultured in sufficient density. After 2 days, differentiation was initiated by culturing for 2 days in DMEM containing 0.5 mM isobutylmethylxanthine (IBMX), 1 μM dexamethasone, 10 μg/mL insulin (MDI solution), and 10% FBS, followed by 10 μg/mL insulin and Differentiation was performed for 3 days by replacing the cells with DMEM containing 10% fetal bovine serum (FBS). After that, by culturing in DMEM containing only 10% FBS, the cells were differentiated into adipocytes that form lipid droplets, and the amount of lipid droplets was confirmed using Oil Red O. Specifically, after differentiation, the medium was removed and the cells were fixed with a 10% formaldehyde solution. After fixing for 30 minutes at room temperature, the solution was removed, washed twice with PBS and twice with 70% alcohol, and then stained using Oil Red O solution. Stained cells were observed under a microscope, dissolved using a solution containing 4% NP-40 in isopropyl alcohol, and absorbance was measured at 510 nm.

1-3 Cytotoxicity measurement method

3T3-L1 cells (5×10 5 cells) were dispensed into each well of a 96-well microplate, and a peptide sample was treated at a concentration of 62.5 μg/mL in each well containing 3T3-L1 cells, and the cells were cultured for 24 hours. Then, each well was treated with 50 μL of MTT (1.0 mg/ml) solution and incubated for 4 more hours. The cultured cells were centrifuged (3000 rpm, 10 min, 20 ℃) to remove the supernatant, treated with 200 μL of dimethylsulfoxide (DMSO) solution, and dissolved by pipetting the formazan crystals, and after 10 minutes, a 96-well microplate reader was used. Absorbance was measured at 540 nm using At this time, the result analysis was calculated as a percentage of the average absorbance of the control group by obtaining the average absorbance of the parasitic sample group.

Animal test evaluation of 1-4 peptide anti-obesity efficacy (high-fat diet mouse feeding experiment)

4-week-old C57BL/6 mice were purchased from the Nara Bio Animal Center (Nara Biotech, Seoul, Korea) and reared in disease-free plastic cages for experiments. The animal room environment was controlled with a 12-hour dark light cycle at 20-21°C and a relative humidity of 40-45%. All experiments were approved by the Animal Care and Use Committee of Konkuk University.

After 1 week of adaptation, the mice were randomly divided into two groups, a control group and an experimental group. The control group was fed a normal diet with 8 animals (control, 10% kcal fat content), and the experimental group was fed a high-fat diet with 30 animals (HFD, 60 % kcal fat content). The food intake of the rats was recorded daily, and the rats' body weight was measured twice weekly. After 5 weeks, mice whose body weight was 20% higher than the control group were selected and randomly divided into three groups (n = 8/group). HFD (high fat diet) mice treated with saline, treated with HFD + peptide (SEQ ID NO: 8 or 15, 100 mg/kg), HFD + Orlistat (60 mg/kg), and orlistat was used as a positive control. After 12 weeks, the animals were slaughtered and examined after fasting for one day.

Blood samples were taken for further analysis. Adipose tissue (cortical, subcutaneous, visceral, hepatic) was weighed, extracutaneous adipose tissue and liver tissue were collected, frozen in liquid nitrogen, and stored at -80 °C until further analysis. For histological analysis, sections of percutaneous lipid and liver tissue were fixed in 10% formaldehyde.

1-5 Body fat composition analysis

In order to compare body fat, dual-energy X-ray absorptiometry (DXA) was performed, and in the 4th week of the experiment, bone and tissue were measured using a whole body scanner once at low energy and once at high energy. Fat and lean body weights were calculated.

1-6 Histological analysis

Liver and epidural adipose tissue was dissected, fixed in 10% natural buffered formalin, and embedded in paraffin for histological examination. Formalin-fixed and paraffin-embedded tissues were cut at 4 μm thickness and stained with hematoxylin and eosin (H&E). The frozen liver was fixed with 10% formaldehyde solution for 10 minutes and then rinsed with running water. After washing with distilled water for a few seconds, immersed in an isopropanol solution for 20 to 30 seconds, stained with oil red O reagent for 15 to 20 minutes, and rinsed with distilled water. Finally, after staining with hematoxylin for 40 seconds, they were immersed in tap water for 5 minutes. Liver tissue was photographed at 200x or less, and adipocytes were analyzed using a microscope.

1-7 Biological Analysis of Blood

Blood samples were obtained from the hearts of mice, and serum was separated by centrifugation (3000 rpm per 20 min) and stored at -80 °C until further analysis. Serum levels of blood urea nitrogen, alanine aminotransferase (ALT), alkaline phosphate, aspartate aminotransferase (AST), total protein, serum albumin, and electrolytes were measured automatically (Abaxis VETSVAN VS2 Chemical Analyzer, CA, USA) was measured with Total cholesterol, low-density lipoprotein cholesterol (LDL), high-density lipoprotein cholesterol (HDL), and TG were measured using a high-speed blood lipid analyzer (OSAN Healthcare Lipid Pro, Anyang-si, Korea). Adiponectin was analyzed by western blotting, and leptin was quantified using the leptin enzyme-linked immuno-oncology (ELISA) kit (MERCK, DHB, Germany).

[Example]

Example 1. Confirmation of anti-obesity efficacy (in vitro)

In order to evaluate the anti-obesity effect of the peptides according to the present invention, a total of 23 peptide samples disclosed in Experimental Example 1-1 were treated with 3-L1 adipocytes, and the resulting lipid droplet formation was tested using Oil red O test method confirmed through. On the other hand, as a control group, separate peptide treatment was not performed (hereinafter, the control group is referred to as an untreated group).

As a result, the experimental results of Table 2 or FIG. 1 were confirmed.

Adipocyte differentiation rate (%) CON. 99.8 ± 0.36 One 57.9 ± 0.60 2 59.0 ± 0.60 3 52.2 ± 0.72 4 63.3 ± 0.12 5 48.4 ± 0.36 6 38.0 ± 0.72 7 39.4 ± 0.36 8 33.7 ± 0.84 9 40.6 ± 0.60 10 48.4 ± 0.36 11 94.3 ± 0.24 12 57.5 ± 0.60 13 68.0 ± 0.60 14 77.8 ± 0.24 15 30.2 ± 1.44 16 57.5 ± 0.60 17 35.1 ± 0.84 18 41.9 ± 0.24 19 68.0 ± 0.60 20 94.8 ± 0.60 21 44.4 ± 1.32 22 83.1 ± 1.80 23 74.4±0.96

Sample Concentration The growth of preadipocytes was inhibited in most of the 23 samples in the experiment treated with a concentration of 62.5 μg/mL. In the control group, 99.8% of adipocytes were differentiated, whereas 15 (30.2%), 8 (33.7%), 17 (35.1%), 6 (38%), and 7 (39.4%) were 40%. Adipocytes were differentiated below, showing excellent ability to inhibit differentiation of adipocytes. No. 3 (52.2%), No. 12 (57.5%), No. 16 (57.5%), No. 12 (57.5%), No. 1 (57.9%), No. 2 (59%) are the fat cells in the 40-60% range showed anger. Others No. 4 (63.3%), No. 13 (68%), No. 19 (68%), No. 23 (74.4%), No. 14 (77.8%), No. 22 (83.1%), No. 11 (94.3%) , No. 20 (94.8%) showed a similar or slightly superior ability to inhibit differentiation of adipocytes to that of the control group.

Example 2. Confirmation of cytotoxicity of peptide samples

In order to evaluate the anti-obesity effect of the peptides according to the present invention, 3-L1 adipocytes were treated with a total of 23 peptide samples disclosed in Experimental Example 1-1, and the survival rate of pre-adipocytes was measured accordingly. On the other hand, as a comparative group, no separate peptide treatment was performed (hereinafter, the comparative group is referred to as an untreated group.)

As a result, the experimental results of Table 3 or FIG. 2 were confirmed.

sample Cell viability (%) 24 hours 48 hours untreated 100.0 ± 0.01 100.0 ± 0.01 1 (KLPFQR) 90.2 ± 0.95 81.5 ± 0.84 2 (STELLIR) 95.6 ± 0.35 81.3 ± 0.92 3 (EIAQDFK) 94.1 ± 0.94 84.7 ± 0.78 4 (HLQLAIR) 97.1 ± 1.38 91.0 ± 1.20 5 (AVQGLLK) 97.4 ± 0.93 89.9±1.67 6 (IAQGGVLP) 96.0 ± 1.23 86.1 ± 1.60 7 (NDEELNKLL) 89.1±0.69 82.6 ± 0.77 8 (AGLQFPVGR) 97.8 ± 0.49 87.6 ± 0.93 9 (YRPGTVALR) 91.7 ± 0.34 86.5 ± 0.19 10 (KSTGGKAPR) 90.6 ± 0.76 79.8 ± 1.43 11 (KQLATKAAR) 95.7 ± 0.67 86.7 ± 0.40 12 (RFQSSAVMA) 90.7 ± 1.14 74.7 ± 0.64 13 (TLSDYNIQK) 86.8 ± 0.15 80.3 ± 0.97 14 (NKLLSGVTIA) 93.8 ± 0.45 87.4 ± 1.12 15 (EIAQDFKTDL) 96.6 ± 0.26 82.7 ± 0.60 16 (ALNQAWFLK) 109.6 ± 0.52 90.0 ± 0.42 17 (AERVGAGAPVY) 105.4 ± 0.59 88.0±1.95 18 (RIVDFHMLESR) 89.7 ± 0.40 88.6 ± 0.53 19 (SGVTIAQGGVLP) 91.0 ± 1.99 83.1 ± 0.66 20 (APRKQLATKAAR) 95.0 ± 1.24 85.4 ± 0.91 21 (QNIIPASTGAAK) 91.7 ± 0.94 76.9 ± 0.94 22 (HLQLAIRNDEELNK) 94.7 ± 0.79 86.2 ± 0.70 23 (VTIAQGGVLPNIQA) 88.7 ± 0.30 76.5 ± 0.76

At a sample concentration of 62.5 μg/mL, 3T3-L1 preadipocytes used in the anti-obesity activity assay were treated to conduct a toxicity test on the cells. As a result of the experiment, the cell viability was 85% or more until 24 hours after treatment, and the toxicity of the sample to cells was insignificant, as it was more than 80% even after 48 hours. Since they were able to effectively inhibit the differentiation of adipocytes, they could be used for prevention, improvement, or treatment of obesity.

Example 3. Changes in body composition in mice fed high-fat nutrition

Changes in body weight and fat composition of high-fat diet mice according to peptide feeding treatment body weight (g/mouse) fat weight
(g/mouse) before experiment Week 5 increment Normal diet group (control) 22.37 28.82 6.45 4.0 high fat diet 22.95 43.16 20.21 ^## 18.2 ^## high fat diet +
Orlistat (positive control) 23.46 36.08 12.62 ^** 13.7 ^** High-fat diet + peptide (SEQ ID NO: 8) 23.85 35.15 11.30 ^** 12.7 ^** High-fat diet + peptide (SEQ ID NO: 15) 25.06 36.97 11.91 ^** 14.5 ^**

Significant differences at ^## p < 0.01 compared with the Con group. Significant differences at ^* p < 0.05, ^** p < 0.01 compared with the HFD group.

As shown in Table 4, the weight gain rate of the rats on the normal diet increased by 6.45 g for 5 weeks, while the rats fed the high-fat diet increased by 20.21 g. The weight gain inhibition rate was 37.6%. Rats that consumed the high-fat diet and peptide (SEQ ID NO: 8) at the same time showed a 44.1% weight gain inhibition rate with an increase of 11.3 g, and the rats that consumed the peptide (SEQ ID NO: 15) at the same time showed a 41.1% weight gain inhibition rate with an increase of 11.91 g. showed that the anti-obesity effect was excellent in the group that consumed the peptide at the same time.

Mouse body fat composition under high-fat diet (double energy x-ray absorptiometry) treatment Body composition (%) province Lean fat Normal diet group (control) 13.9 80.4 high fat diet 41.7 53.6 high fat diet +
Orlistat (positive control) 36.9 59.0 High-fat diet + peptide (SEQ ID NO: 8) 35.6 59.9 High-fat diet + peptide (SEQ ID NO: 15) 38.5 57.3

As shown in Table 5, as a result of analysis of the body fat composition of the mice measured by dual energy x-ray absorptiometry, the fat composition was 13.9% in the normal diet rats, but very high at 41.7% in the high-fat diet rats. The rats that consumed Orlistat, a positive control group, and Orlistat at the same time showed an effect of lowering the fat composition to 11.5% compared to the high-fat diet by 36.9%. Rats consuming the high-fat diet and peptide (SEQ ID NO: 8) at the same time showed an effect of lowering the fat composition by 14.6% compared to the high-fat diet, with a fat composition of 35.6%. showed the effect of lowering the fat composition by 7.7% compared to the high-fat diet, and the peptide showed the anti-obesity effect of suppressing the increase in fat composition.

Example 4. Blood Biological Analysis

Comparison of blood components in mice fed a peptide-fed high-fat diet treatment changes in blood composition TG ^a
(mg/dL) TC ^b
(mg/dL) HDL ^-c
(mg/dL) LDL/VLDL ^d
(mg/dL)
Normal diet group (control) 108.3 96.1 79.8 16.3
high fat diet 232.6 ^## 119.7 ^## 47.4 ^## 72.3 ^##
high fat diet +
Orlistat (positive control) 117.9 ^** 119.3 ^** 77.8 ^** 44.2 ^**
high fat diet +
peptide (SEQ ID NO: 8) 138.8 ^** 120.5 ^** 75.1 ^** 42.7 ^**
high fat diet +
peptide (SEQ ID NO: 15) 140.3 ^** 107.5 ^** 74.5 ^** 33.0 ^**

Significant differences at ^## p < 0.01 compared with the Con group. Significant differences at ^* p < 0.05, ^** p < 0.01 compared with the HFD group.

^a triglyceride (TG), ^b total cholesterol (TC), ^c high density lipoprotein (HDL), ^d low density lipoprotein (LDL)

As shown in Table 6, the triglyceride (TG) of mice raised on a high-fat diet was 232.6 mg/dL, total cholesterol (TC) was 119.7 mg/dL, high-density lipoprotein (HDL) was 47.4 mg/dL, and density Lipoprotein (LDL) was 72.3 mg/dL. Triglyceride (TG), total cholesterol (TC), and high-density lipoprotein (HDL) were 75.1 mg/dL, 138.8 mg/dL, and 75.1 mg/dL, respectively. Lipoprotein (LDL) was 42.7mg/dL, which reduced neutral fat and low-density lipoprotein according to peptide intake, and showed an effect of increasing HDL, which reduces vascular diseases such as arteriosclerosis.

In the case of mice simultaneously ingesting a high-fat diet and peptide (SEQ ID NO: 15), triglyceride (TG) was 140.3mg/dL, total cholesterol (TC) was 107.5mg/dL, high-density lipoprotein (HDL) was 74.5mg/dL, Low-density lipoprotein (LDL) was 33.0 mg/dL, which reduced triglyceride, total cholesterol, and low-density lipoprotein according to peptide intake, and showed an effect of increasing HDL.

Example 5. Comparison of histological characteristics of mice fed high fat nutrition

<peptide (SEQ ID NO: 8)>

In mice fed a high-fat diet, compared to the normal diet group, the liver size increased (Fig. 3A), the amount of white adipose tissue increased (Fig. 3B), and the amount of red adipose tissue increased when stained with oil red o (Fig. 3C). , It was found that the size of fat was also significantly larger than that of normal mice (FIG. 3D). Mice fed the high-fat diet and the positive control orlistat at the same time had a significantly smaller liver size and a smaller amount of white adipose tissue compared to the high-fat diet group. The size was also found to be relatively small. Similar to the orilistat intake group, the peptide 8 group was similar to the high-fat diet group, and the shape and size of the liver were close to normal, and the number and size of white adipose tissue were also close to normal. showed

<peptide (SEQ ID NO: 15)>

Compared to the high-fat diet group, the group fed the high-fat diet and the peptide (SEQ ID NO: 15) simultaneously showed that the shape and size of the liver were close to normal, and the number and size of white adipose tissue were also close to normal, suppressing fat gain. showed an effect (FIG. 4).

<110> Republic of Korea <120> Peptide (PEPTIDE 15) for anti-obesity and use thereof <130> P20G19C2080 <150> KR 10-2019-0136552 <151> 2019-10-30 <160> 23 <170> KoPatentIn 3.0 <210> 1 <211> 6 <212> PRT <213> artificial sequence <220> <223> peptide 1 <400> 1 Lys Leu Pro Phe Gln Arg 1 5 <210> 2 <211> 7 <212> PRT <213> artificial sequence <220> <223> peptide 2 <400> 2 Ser Thr Glu Leu Leu Ile Arg 1 5 <210> 3 <211> 7 <212> PRT <213> artificial sequence <220> <223> peptide 3 <400> 3 Glu Ile Ala Gln Asp Phe Lys 1 5 <210> 4 <211> 7 <212> PRT <213> artificial sequence <220> <223> peptide 4 <400> 4 His Leu Gln Leu Ala Ile Arg 1 5 <210> 5 <211> 7 <212> PRT <213> artificial sequence <220> <223> peptide 5 <400> 5 Ala Val Gln Gly Leu Leu Lys 1 5 <210> 6 <211> 8 <212> PRT <213> artificial sequence <220> <223> peptide 6 <400> 6 Ile Ala Gln Gly Gly Val Leu Pro 1 5 <210> 7 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 7 <400> 7 Asn Asp Glu Glu Leu Asn Lys Leu Leu 1 5 <210> 8 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 8 <400> 8 Ala Gly Leu Gln Phe Pro Val Gly Arg 1 5 <210> 9 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 9 <400> 9 Tyr Arg Pro Gly Thr Val Ala Leu Arg 1 5 <210> 10 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 10 <400> 10 Lys Ser Thr Gly Gly Lys Ala Pro Arg 1 5 <210> 11 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 11 <400> 11 Lys Gln Leu Ala Thr Lys Ala Ala Arg 1 5 <210> 12 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 12 <400> 12 Arg Phe Gln Ser Ser Ala Val Met Ala 1 5 <210> 13 <211> 9 <212> PRT <213> artificial sequence <220> <223> peptide 13 <400> 13 Thr Leu Ser Asp Tyr Asn Ile Gln Lys 1 5 <210> 14 <211> 10 <212> PRT <213> artificial sequence <220> <223> peptide 14 <400> 14 Asn Lys Leu Leu Ser Gly Val Thr Ile Ala 1 5 10 <210> 15 <211> 10 <212> PRT <213> artificial sequence <220> <223> peptide 15 <400> 15 Glu Ile Ala Gln Asp Phe Lys Thr Asp Leu 1 5 10 <210> 16 <211> 10 <212> PRT <213> artificial sequence <220> <223> peptide 16 <400> 16 Ala Leu Asn Gln Ala Trp Ala Phe Leu Lys 1 5 10 <210> 17 <211> 11 <212> PRT <213> artificial sequence <220> <223> peptide 17 <400> 17 Ala Glu Arg Val Gly Ala Gly Ala Pro Val Tyr 1 5 10 <210> 18 <211> 11 <212> PRT <213> artificial sequence <220> <223> peptide 18 <400> 18 Arg Ile Val Asp Phe His Met Leu Glu Ser Arg 1 5 10 <210> 19 <211> 12 <212> PRT <213> artificial sequence <220> <223> peptide 19 <400> 19 Ser Gly Val Thr Ile Ala Gln Gly Gly Val Leu Pro 1 5 10 <210> 20 <211> 12 <212> PRT <213> artificial sequence <220> <223> peptide 20 <400> 20 Ala Pro Arg Lys Gln Leu Ala Thr Lys Ala Ala Arg 1 5 10 <210> 21 <211> 12 <212> PRT <213> artificial sequence <220> <223> peptide 21 <400> 21 Gln Asn Ile Ile Pro Ala Ser Thr Gly Ala Ala Lys 1 5 10 <210> 22 <211> 14 <212> PRT <213> artificial sequence <220> <223> peptide 22 <400> 22 His Leu Gln Leu Ala Ile Arg Asn Asp Glu Glu Leu Asn Lys 1 5 10 <210> 23 <211> 14 <212> PRT <213> artificial sequence <220> <223> peptide 23 <400> 23 Val Thr Ile Ala Gln Gly Gly Val Leu Pro Asn Ile Gln Ala 1 5 10

Claims (7)

A peptide showing anti-obesity activity consisting of the amino acid sequence represented by SEQ ID NO: 15. A pharmaceutical composition for preventing or treating obesity comprising the peptide of claim 1. delete A composition for external application for skin for preventing or improving obesity, comprising the peptide of claim 1. delete A food composition for preventing or improving obesity comprising the peptide of claim 1. delete

Images