Application of propyl gallate in preparation of anti-obesity drug or functional food

Abstract

The invention provides application of propyl gallate in preparation of anti-obesity drugs or functional food for weight management, and belongs to the field of biological medicine technology and food. The invention discloses that the propyl gallate has the health benefit of positively regulating basic metabolism for the first time, and the propyl gallate can obviously reduce the weight of obese mice induced by high fat diet, reduce body fat, effectively relieve obesity and improve metabolic conditions such as glucose tolerance and insulin resistance; and brown fat can be activated to produce heat, respiratory metabolism and energy consumption are enhanced, and a new way is opened up for preventing and / or treating obesity. Propyl gallate is an edible substance, is high in safety and oxidation resistance, and is widely used as a food antioxidant, so that propyl gallate can be used for preparing anti-obesity drugs or functional foods for weight management, and has a good application prospect.

Description

Technical Field

[0001] The invention relates to the fields of medical technology and food, and in particular to the application of propyl gallate in the preparation of anti-obesity drugs or functional foods for weight management. Background Art

[0002] Obesity was defined as a body mass index (BMI, weight divided by the square of height) ≥ 30 kg / m 2 , is a chronic disease caused by excessive fat accumulation. Obesity is a widespread global problem. As of 2022, the number of obese people in the world has exceeded 1 billion, bringing a huge burden to global public health.

[0003] Obesity increases the incidence of related complications including asthma, type 2 diabetes, hypertension, cardiovascular disease, cancer, obstructive sleep apnea, and osteoarthritis through associated pathophysiological and mechanical changes.

[0004] The root cause of obesity is the imbalance of energy metabolism, that is, the imbalance between excessive energy intake and insufficient energy consumption. Existing treatment methods mainly include dietary restriction, exercise, drug weight loss, and surgical weight loss. However, these methods have certain risks, side effects, or are difficult to adhere to. For example, existing drug therapies (such as orlistat, GLP-1 receptor agonists) are often accompanied by gastrointestinal reactions, cardiovascular risks and other side effects, while surgical interventions have the risk of complications such as infection and poor nutrient absorption. Therefore, finding a safe, effective, and easy-to-adhere weight loss method is an urgent problem to be solved.

[0005] Adipose tissue is one of the most plastic organs and is now widely accepted as an important player in maintaining whole-body energy homeostasis. Unlike white adipose tissue (WAT), which stores energy, brown adipose tissue (BAT) is rich in triglyceride droplets, which can be rapidly hydrolyzed and oxidized to fatty acids. It also contains a large number of mitochondria and expresses high levels of uncoupling protein 1 (UCP1). When UCP1 is activated, it can uncouple mitochondrial oxidative phosphorylation from ATP production, thereby dissipating energy in the form of heat. In addition, the creatine kinase B (CKB)-mediated creatine ineffective cycle runs in parallel with UCP1, jointly promoting BAT thermogenesis. Due to its excellent thermogenic energy-consuming function, BAT is considered a new target for the treatment and / or prevention of obesity and its metabolic complications.

[0006] Propyl gallate (PG) is one of the active ingredients of the commonly used Chinese medicine red peony root. It was first obtained by Chinese scholars through structural modification of gallic acid, the active ingredient of red peony root. It has stronger biological effects than gallic acid and is named red peony root 801. PG is an alkyl ester of the natural phenolic acid gallic acid and belongs to the phenolic acid derivatives. It is widely used as a food antioxidant due to its high safety and strong antioxidant properties. It is also used in the treatment of cardiovascular diseases as a tongmai ester. Phenolic acid compounds are a class of low-molecular natural plant polyphenols, which belong to plant secondary metabolites. They are widely present in plant foods and Chinese herbal medicines and have multiple physiological activities such as antioxidant, anti-inflammatory, and anti-obesity. In vivo, PG has higher stability and lipophilicity than phenolic acid, and is easier to enter various intracellular environments, which helps stimulate the absorption of oxygen by the mitochondrial and microsomal electron transport chains. However, there is no relevant research on the anti-obesity effect of propyl gallate. Summary of the invention

[0007] The purpose of the present invention is to provide a food-grade substance that has the function of regulating the basal metabolism of humans or animals, and to apply propyl gallate to the preparation of anti-obesity drugs or functional foods for weight management, thereby providing a new idea for the prevention and / or treatment of obesity and its metabolic complications.

[0008] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0009] The present invention provides the use of propyl gallate in the preparation of anti-obesity drugs or functional foods for weight management. The present invention found that propyl gallate has the health benefits of positively regulating basal metabolism in an obese mouse model induced by a high-fat diet, mainly including weight loss, lowering body fat, improving glucose tolerance and insulin resistance, activating brown fat thermogenesis, and enhancing energy consumption, thereby effectively alleviating obesity.

[0010] Therefore, propyl gallate can be used to prepare a composition for supporting weight management, promoting weight loss, preventing and / or treating obesity.

[0011] Specifically, the present invention provides an anti-obesity pharmaceutical composition, comprising an effective dose of propyl gallate and a pharmaceutically acceptable carrier.

[0012] The pharmaceutical composition is prepared by adding propyl gallate as the main active ingredient and a pharmaceutically acceptable carrier, and can be prepared according to the preparation method recorded in pharmacy. The preparation form of the drug can be, but is not limited to, an oral preparation. The specific dosage is adjusted according to the type of disease, the degree of disease, the age and the purpose of administration.

[0013] In the mouse model, the effective dose of propyl gallate is 50-100 mg / kg body weight / day, with no obvious toxic side effects.

[0014] The present invention provides the use of propyl gallate in the preparation of functional foods for improving energy metabolism, promoting heat production or assisting weight loss. Propyl gallate intervention can increase the heat production of brown fat, increase body temperature within a normal range, and increase energy consumption. Mechanism studies have shown that propyl gallate enhances the glucose uptake capacity of brown adipose tissue, promotes the expression of heat-related genes and proteins, activates brown fat activity, thereby enhancing the body's energy metabolism and heat production, and is used to assist weight management.

[0015] As an antioxidant food additive with high biosafety, propyl gallate is expected to be developed into an anti-obesity drug, or a functional food for improving energy metabolism, promoting heat production, and assisting weight management.

[0016] Specifically, the present invention provides a functional food for weight loss, comprising propyl gallate and a food science acceptable auxiliary material. The present invention uses propyl gallate as an active ingredient to prepare a corresponding functional food, which can also be added to food or beverages in the form of a food additive to promote weight loss of the user.

[0017] By adopting the above technical solution, the present invention has the following beneficial effects:

[0018] The present invention discloses for the first time that propyl gallate has the health benefits of positively regulating basal metabolism. For people or animals on a high-fat diet, propyl gallate can significantly reduce body weight and body fat, effectively alleviate obesity, and improve metabolic conditions such as glucose tolerance and insulin resistance; it can also activate brown fat thermogenesis, and enhance respiratory metabolism and energy consumption. Propyl gallate is an edible substance with high safety and strong antioxidant properties. It is widely used as a food antioxidant, so it can be used to prepare anti-obesity drugs or functional foods for weight management, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 The weight and adipose tissue changes of mice in Example 1, including (A) weight changes of mice; (B) adipose tissue of mice; (CD) organ indexes of iWAT and eWAT of mice; (E) HE staining of BAT, iWAT and eWAT of mice; in the figure, * indicates significant analysis compared with the DIO group, * represents p<0.05, ** represents p<0.01, and *** represents p<0.001.

[0020] Figure 2The glucose tolerance and insulin sensitivity of mice in Example 1, wherein (AB) glucose tolerance curve and area under the curve of mice; (CD) insulin tolerance curve and area under the curve of mice; in the figure, * indicates a significant difference compared with the DIO group, * represents p<0.05, ** represents p<0.01, and *** represents p<0.001.

[0021] Figure 3 The energy metabolism of the mice in Example 1, including (A) daily food intake of the mice; (B) daily average energy intake of the mice; (C) daily exercise volume of the mice; and (D) respiratory metabolic oxygen consumption of the mice.

[0022] Figure 4 The BAT activity of mice in Example 2, wherein (A) body temperature change of mice after cold stimulation; (B) infrared thermal imaging of mice; (C) PET-CT imaging of mice; (D) semi-quantitative results of PET-CT of mice; in the figure, * indicates a significant difference compared with the DIO group, * represents p<0.05, ** represents p<0.01, and *** represents p<0.001.

[0023] Figure 5 The expression of related genes and proteins in the BAT of mice in Example 2, wherein (AC) the relative expression of thermogenesis-related genes Ucp1, Pgc1α, and Ckb in the BAT of mice; (DG) the expression of thermogenesis-related proteins UCP1 and PGC-1α in the BAT of mice; in the figure, * indicates a significant difference with the DIO group, * represents p<0.05, ** represents p<0.01, and *** represents p<0.001.

[0024] Figure 6 The effect of PG on UCP1-Luciferase mice in Example 2, wherein (A) IVIS in vivo fluorescence imaging of UCP1-Luciferase gene knock-in mice treated with 100 mg / kg PG; (B) IVIS in vivo fluorescence quantification of UCP1-Luciferase gene knock-in mice; in the figure, * indicates a significant difference compared with the CHOW group, * represents p<0.05, ** represents p<0.01, and *** represents p<0.001.

[0025] All the above data are expressed as mean ± SEM. The experimental data were subjected to one-way ANOVA, and p < 0.05 was considered significant. DETAILED DESCRIPTION

[0026] The present invention provides the use of propyl gallate in the preparation of anti-obesity drugs or functional foods. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve it. It is particularly important to point out that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the method and application of this article without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0027] Terminology Note:

[0028] The term "comprising" is an open expression, that is, it includes the contents specified in the present invention but does not exclude other contents.

[0029] The terms "anti-", "prevent and / or treat" refer in some embodiments to ameliorating a disease or condition (i.e., slowing or preventing or alleviating the development of a disease or at least one clinical symptom thereof). In other embodiments, it refers to alleviating or improving at least one physical parameter, including physical parameters that may not be perceived by the subject. In other embodiments, it refers to regulating a disease or condition physically (e.g., stabilizing perceptible symptoms) or physiologically (e.g., stabilizing physical parameters), or both. In other embodiments, it refers to preventing or delaying the onset, onset, or worsening of a disease or condition.

[0030] The term "effective dose" refers to an amount of a compound that is sufficient to be effective in treating a disease when administered to a subject. The effective dose may vary with the severity of the disease, and the physical condition, age, weight, sex, etc. of the subject to be treated.

[0031] The term "pharmaceutically or edible acceptable carrier / excipient" refers to any preparation or carrier medium that can deliver an effective dose of the active substance of the present invention without interfering with the biological activity of the active substance and having no toxic side effects on the host or subject.

[0032] The test materials used in the present invention are all common commercial products and can be purchased in the market.

[0033] In the present invention, propyl gallate can significantly reduce the body weight and body fat of obese mice induced by a high-fat diet, improve metabolic conditions such as glucose tolerance and insulin resistance, and activate brown fat thermogenesis, enhance respiratory metabolism and energy consumption, thereby effectively alleviating obesity.

[0034] In the present invention, the concentration of propyl gallate in the drug is 10-20 mg / mL, and the dosage is 50-100 mg / kg / day.

[0035] In the present invention, the anti-obesity drug further includes a pharmaceutically acceptable carrier or excipient.

[0036] The present invention provides the use of propyl gallate in the preparation of brown fat activators or functional foods for improving energy metabolism, promoting heat production and assisting weight loss. Propyl gallate activates brown fat activity by enhancing the glucose uptake capacity of brown fat tissue, promoting the expression of heat-related genes and proteins.

[0037] In the present invention, the concentration of propyl gallate in the drug is 10-20 mg / mL, and the dosage is 50-100 mg / kg / day.

[0038] Propyl gallate used in the examples of the present invention was purchased from Shanghai MacLean Biochemical Technology Co., Ltd.

[0039] The purity of propyl gallate used in the examples of the present invention is 98%.

[0040] The C57BL6 / J mice used in the examples of the present invention were purchased from Sibeifu (Beijing) Biotechnology Co., Ltd.

[0041] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0042] Example 1 Effect of Propyl Gallate (PG) on Obese Mice Induced by High-Fat Diet

[0043] A total of 48 6-week-old C57BL6 / J male mice were selected and raised in an SPF-level mouse house with 12h light and dark alternation, temperature 25°C, and humidity 45%. After one week of adaptive feeding, they were randomly divided into cages according to body weight, with 4 mice in each cage, and divided into 4 groups, including a control group (CHOW), a high-fat group (DIO), a low-dose propyl gallate group (DIO+L-PG), and a high-dose propyl gallate group (DIO+H-PG). The CHOW group was fed with normal feed (mouse growth maintenance feed, purchased from Beijing Ke'ao Xieli Feed Co., Ltd.), and the remaining groups were fed with high-fat feed (mouse high-fat feed, purchased from Beijing Ke'ao Xieli Feed Co., Ltd.), and each group had free access to normal water. The DIO+L-PG group and the DIO+H-PG group were gavaged with propyl gallate solution at a dose of 0.1 mL / 10 g body weight (a solution prepared by heating 98% pure propyl gallate to 50°C to dissolve and then cooling); the dosage of the DIO+L-PG group was 50 mg / kg body weight / day, and the dosage of the DIO+H-PG group was 100 mg / kg body weight / day. The CON group and the DIO group were gavaged with the corresponding volume of purified water.

[0044] The drug was administered continuously for 13 weeks, and the weight of the mice was recorded every week. From the 14th week of the mouse experiment, glucose tolerance test, insulin tolerance test, cold stimulation body temperature detection, infrared thermal imaging, PET-CT scanning, mouse exercise and respiratory metabolism test were carried out. At the end of the experiment, the mice were fasted for 12 hours, blood was collected from the eyeballs and the mice were killed by dislocation of the neck. The blood was collected in a sodium heparin anticoagulant tube, centrifuged at 4℃3000rpm for 15min, and the supernatant was taken. The plasma was frozen at -80℃ for use; the liver, fat and other tissues of the mice were collected, weighed and tested for relevant indicators.

[0045] The results showed that PG could inhibit the weight gain of DIO mice in a dose-dependent manner (p<0.05). 50mg / kg and 100mg / kg of PG treatment reduced the body weight of DIO mice by 18.24% and 26.18%, respectively (Fig. Figure 1 A) From Figure 1 As can be seen in B, PG treatment significantly reduced the fat volume of DIO mice. The tissue and organ index showed that the PG treatment group, especially the DIO+H-PG group, could significantly reduce the weight of the subcutaneous adipose tissue (iWAT) and epididymal adipose tissue (eWAT) of the mice ( Figure 1 C, D) (*p<0.05, **p<0.01, ***p<0.001). Further HE staining of adipose tissue sections revealed that the volume of BAT, iWAT and eWAT cells in the PG-treated mice was significantly smaller than that in the high-fat group ( Figure 1 E). This indicates that PG has the effect of reducing obesity in DIO mice.

[0046] Figure 2 A and B show that compared with the DIO group, PG significantly enhanced the glucose tolerance of DIO mice (p<0.05); at the same time, the insulin sensitivity of mice was also significantly improved (p<0.05) ( Figure 2 C, D).

[0047] Figure 3 AC showed that there was no significant difference in the food intake and exercise volume of mice among the groups, indicating that PG treatment inhibited the weight gain of DIO mice not because of reduced food intake or increased exercise volume, but may be caused by increased energy consumption. Further results of mouse energy metabolism showed ( Figure 3 D), compared with the DIO group mice, the daytime basal metabolic rate and the oxygen consumption during the active period at night of the PG-treated mice were significantly increased, indicating that PG can significantly increase the body energy consumption of DIO mice, thereby effectively resisting obesity.

[0048] Example 2 Activation of BAT by Propyl Gallate

[0049] (I) BAT activity assay

[0050] BAT is a key organ for heat production and energy consumption in the body. Cold stimulation is an effective means to activate BAT. The body temperature of mice after cold stimulation was tested ( Figure 4 A) and infrared thermal imaging ( Figure 4 B) Detection found that PG significantly increased the body surface temperature of DIO mice. 18 F-FDG PET / CT test results ( Figure 4 C) shows BAT absorption in PG-treated mice 18 The F-FDG signal increased significantly, indicating that the BAT activity of mice was enhanced. The semi-quantitative results (SUV value) Figure 4 D) Consistent with the PET-CT images, PG treatment significantly increased the activity of BAT in DIO mice (*p<0.05, **p<0.01, ***p<0.001), indicating that PG treatment can activate BAT activity in DIO mice and enhance thermogenesis.

[0051] (II) Expression of thermogenesis-related genes and proteins in BAT

[0052] BAT tissues were obtained from mice to measure the expression of heat-related genes and proteins in BAT.

[0053] The total RNA of BAT was extracted by Trizol method, and chloroform was added for extraction. The supernatant was precipitated with an equal volume of isopropanol, and then the precipitate was washed with 75% ethanol. The RNA precipitate was dissolved in DEPC water for subsequent experiments. Reverse transcription was performed using a cDNA kit (purchased from Beijing Polymer Biotechnology Co., Ltd.). Primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd., and the primer sequences are shown in Table 1. The reaction system was prepared using a qPCR premix kit (purchased from Beijing Polymer Biotechnology Co., Ltd.), and the mRNA level was quantified using Cyclophilin A as the internal standard.

[0054] Table 1 Primer sequences

[0055]

[0056] Total BAT protein was extracted using RIPA lysis buffer containing PMSF, the protein concentration was adjusted to 1 μg / μL, and the protein was denatured by heating at 95°C for 5 min. SDS-PAGE 10% separation gel and 5% concentration gel were prepared, protein samples and markers were added, and electrophoresis was performed at 80V for 25 min and 120V for 75 min. Methanol activated the PVDP film, and the "wet transfer method" was used for 75 min at 110V. The PVDF membrane was removed and blocked with Biyuntian fast blocking solution for 10 min, incubated with primary antibody (1:1000) at 4°C overnight, washed with TBST, incubated with secondary antibody (1:5000) at room temperature for 1 h, and washed with TBST. ECL chemiluminescent solution was added for imaging under the developer.

[0057] The results showed that PG treatment significantly enhanced the expression of heat-related genes such as UCP1, PGC-1α, and CKB in BAT of DIO mice ( Figure 5 AC) and protein ( Figure 5 DG) (*p<0.05, **p<0.01, ***p<0.001). This indicates that PG can activate BAT activity in DIO mice and enhance the body's heat production and energy consumption.

[0058] (III) Activation effect of PG on UCP1 in BAT

[0059] In order to further verify the activation effect of PG on UCP1 in BAT in vivo, UCP1-Luciferase gene knock-in mice were treated with 100 mg / kg PG for 14 days and then subjected to in vivo imaging detection ( Figure 6 ) found that PG can significantly enhance the IVIS in vivo fluorescence expression of BAT in the scapula of mice (*p<0.05, **p<0.01, ***p<0.001). This shows that PG can be used as an effective activator of BAT, enhancing the body's heat production and energy consumption, thereby effectively improving high-fat diet-induced obesity in mice.

[0060] Based on the above embodiments and test examples, it can be seen that the use of propyl gallate described in the present invention can effectively inhibit the weight gain of obese mice induced by a high-fat diet, reduce the fat weight of mice, improve metabolic conditions such as glucose tolerance and insulin resistance, enhance respiratory metabolism and energy consumption, and promote the expression of heat-related genes and proteins by activating BAT activity, thereby increasing the body's heat production and energy consumption, thereby effectively improving obesity in mice.

[0061] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. The scope of protection of the present invention is based on the claims, and any equivalent substitutions or modifications based on the essence of the present invention fall within the scope of protection of the present invention.

[0062] The above are only preferred embodiments of the present invention. It should be pointed out that, for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. The use of propyl gallate in the preparation of anti-obesity drugs, characterized in that: Propyl gallate can significantly reduce weight gain induced by a high-fat diet, reduce body fat, and improve metabolic conditions such as glucose tolerance and insulin resistance; it can also activate brown fat thermogenesis, enhance respiratory metabolism and energy consumption, and thus effectively alleviate obesity; The concentration of propyl gallate in the drug is 10-20 mg / mL, and the dosage is 50-100 mg / kg body weight / day.

2. The use according to claim 1, characterized in that: The anti-obesity drug also includes a pharmaceutically acceptable carrier or auxiliary material.

3. The use of propyl gallate in the preparation of functional foods for improving energy metabolism, promoting heat production or assisting weight loss, characterized in that: Propyl gallate can be used to assist in weight management by enhancing the ability of brown adipose tissue to absorb glucose, promoting the expression of heat-related genes and proteins, activating brown fat activity, thereby enhancing the body's energy metabolism and heat production.

4. The use according to claim 3, characterized in that: The functional food also includes excipients that are acceptable in food science.

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