Application of piperine in preparation of product for improving heat loss

Abstract

The invention belongs to the technical field of biological medicine. More specifically, the invention provides an application of piperine in preparation of a product for improving heat loss. By constructing a mouse heat loss model, the research finds that the piperine can obviously increase the quantity and ridge number of mitochondria in mouse scapular brown adipose tissue, so that the mitochondrial dysfunction is improved, and the energy metabolism efficiency is improved. The effect is beneficial to promoting lipolysis and heat energy release, so that the adaptive body temperature regulation ability of the mouse is enhanced, and more heat support is provided for the body. As a natural plant component, the piperine is nontoxic to human bodies and has very high safety. The research finds that piperine has multiple effects of enhancing mitochondrial function, activating fat heat production and improving body temperature regulation capability, can be widely applied to preparation of related products for improving heat loss, hypothermia, heat production disorder and the like, and has good application potential and prospect.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical technology. More specifically, it relates to the application of piperine in the preparation of products for improving hypothermia. Background Art

[0002] Hypothermia is a pathological state caused by the core body temperature dropping below the normal range (usually below 35°C), which commonly occurs under cold exposure or metabolic stress conditions. The occurrence mechanism of hypothermia is mainly related to the body's heat loss being greater than heat supply, resulting in damage to the core organs (such as the brain, heart, lungs, etc.) that maintain life. According to the severity, hypothermia can be divided into mild, moderate, and severe. Mild hypothermia is manifested by symptoms such as shivering, coldness, and muscle spasms; moderate hypothermia may be accompanied by bradycardia, decreased blood pressure, confusion, etc.; while patients with severe hypothermia may have weak pulse and respiration, whole body muscle stiffness, and even loss of consciousness, which may lead to death if not treated in time.

[0003] Currently, the clinical treatment methods for hypothermia mainly rely on physical rewarming (such as using heating blankets, hot drinks, electric heating devices, etc.) and supportive measures such as environmental control. However, these methods can only relieve symptoms and cannot fundamentally regulate the body's metabolic pathways to restore the body temperature regulation ability under extremely cold environments or metabolic disorder states. There is an urgent need to develop new drugs that can enhance the heat production function, improve mitochondrial function disorders, and restore body temperature homeostasis. Summary of the Invention

[0004] The purpose of the present invention is to provide the application of piperine in the preparation of products for improving hypothermia, cryopathy, heat production disorders, etc.

[0005] The above object of the present invention is achieved by the following technical solutions:

[0006] The research of the present invention shows that piperine can enhance mitochondrial function, activate fat heat production, and improve the body temperature regulation ability. Therefore, the present invention claims the following application solutions:

[0007] The application of piperine in the preparation of products for improving hypothermia.

[0008] The application of piperine in the preparation of products for improving cryopathy.

[0009] The application of piperine in the preparation of products for activating fat heat production.

[0010] The application of piperine in the preparation of products for improving mitochondrial function disorders.

[0011] The application of piperine in the preparation of products for enhancing the body temperature regulation ability.

[0012] The present invention provides a drug for improving hypothermia or low temperature disease, which contains piperine and a pharmaceutically acceptable pharmaceutical excipient.

[0013] The present invention provides a drug for improving mitochondrial disease, which contains piperine and a pharmaceutically acceptable pharmaceutical excipient.

[0014] As an alternative embodiment, the pharmaceutical excipient includes any one or a combination of at least two of a carrier, a diluent, an excipient, a filler, a binder, a wetting agent, an emulsifier, a solubilizer, a surfactant or a buffer.

[0015] As an alternative embodiment, the pharmaceutical excipient further includes any one or a combination of at least two of a colorant, a pH regulator, an antioxidant, an antibacterial agent.

[0016] As an alternative embodiment, the dosage form of the drug is a tablet, a capsule, an aerosol, a pill, a powder, a solution, an emulsion or a granule.

[0017] The present invention has the following beneficial effects:

[0018] By constructing a mouse hypothermia model, the present invention finds through research that piperine (Piperine, PIP) has a significant effect in improving hypothermia, low temperature disease and related body temperature regulation disorders. Experiments prove that piperine can significantly increase the number and cristae number of mitochondria in the mouse scapular brown adipose tissue, thereby improving mitochondrial function disorders and enhancing the energy metabolism efficiency. This effect helps to promote fat decomposition and heat release, thereby enhancing the adaptive body temperature regulation ability of mice and providing more heat support for the body.

[0019] As a natural plant component, piperine is non-toxic to the human body and has high safety. The present invention finds through research that piperine has multiple functions of enhancing mitochondrial function, activating fat thermogenesis and improving body temperature regulation ability, and can be widely used in the preparation of related products for improving hypothermia, low temperature disease, thermogenesis disorders, etc., and has good application potential and prospects. Description of the Drawings

[0020] Figure 1 For the determination results of the thermogenic function of mice in each group (Figure A is the infrared thermal imaging diagram of the back temperature of mice in each group; Figure B is the temperature difference result of the back of mice in each group during the cold exposure experiment).

[0021] Figure 2 For the transmission electron microscope results and quantitative analysis results of mitochondria in the scapular brown adipose tissue of mice in each group (Figure A is the transmission electron microscope result; Figure B is the analysis result of the number of mitochondria; Figure C is the analysis result of the cristae number of mitochondria).

[0022] Figure 3 For the determination results of the energy metabolism gene expression levels in the scapular brown adipose tissue of mice in each group. Detailed implementation manners

[0023] The present invention will be further described below in conjunction with the accompanying drawings of the specification and specific embodiments. However, the embodiments do not impose any form of limitation on the present invention. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the technical field.

[0024] Unless otherwise specified, the reagents and materials used in the following examples are all commercially available.

[0025] Piperine used in the examples was purchased from Xi'an Ruilin Biotechnology Co., Ltd., and the CAS number is 94-62-2.

[0026] The manufacturer of the standard diet feed in the examples is: Jiangsu Xietong Pharmaceutical Biotechnology Co., Ltd., and the product number is 1010088.

[0027] The manufacturer of the high-sugar and high-fat diet feed in the examples is: Jiangsu Xietong Pharmaceutical Biotechnology Co., Ltd., and the product number is XT310.

[0028] The preparation method of the mouse hypothermia model is: feeding the mice with a high-sugar and high-fat diet for 6-10 weeks, placing the mice at 2-8 °C for 10-20 min, and then placing the mice back into the room temperature environment to obtain the hypothermia model mice.

[0029] As an alternative implementation manner, the preparation method of the mouse hypothermia model is: feeding the mice with a high-sugar and high-fat diet for 8 weeks, placing the mice at 4 °C for 15 min, and then placing the mice back into the room temperature environment to obtain the hypothermia model mice.

[0030] As an alternative implementation manner, after the mice are adapted to the light cycle for 1 week, the hypothermia model is prepared.

[0031] Example 1 Influence of piperine on the mouse hypothermia model

[0032] I. Experimental method

[0033] (1) Animal feeding and grouping

[0034] C57BL / 6 mice were selected. All mice were housed in a self - regulating environment (temperature: 25 ± 2 °C; relative humidity: 65 ± 5%). They were in a 12 - hour light - dark cycle (zeitgeber time (ZT), ZT0 = 8:00 am, lights on; ZT12 = 8:00 pm, lights off; ZT6 = 2:00 pm, ZT18 = 2:00 am). After a 1 - week acclimation period, all mice were randomly divided into three groups (12 mice in each group): mice in the blank control (CONT) group were fed a standard diet; mice in the hypothermia model (ACE, acute cold exposure) group were fed a high - sugar and high - fat diet; mice in the piperine intervention group (PIP) were given piperine intervention. Piperine was administered by gavage at a dose of 0.03 mg / g / animal.

[0035] (2) Detection of thermogenic function in mice

[0036] After 8 weeks of corresponding treatment for the three groups of mice, a cold exposure experiment was conducted on each group of mice. An infrared imaging instrument was used to evaluate the ability of mice to activate adipose tissue thermogenesis and restore their core body temperature. The specific experimental steps were as follows: The core body temperature of the mice was measured after they were placed at 4 °C for 15 minutes, and then they were placed at 30 °C room temperature, and the change in the back temperature of the mice was continuously measured within ten minutes.

[0037] (3) Mouse dissection

[0038] Each group of mice was fasted for 8 hours before sacrifice and sacrificed at four time points: ZT0, ZT6, ZT12, and ZT18. After dissecting the mice, the interscapular brown adipose tissue was removed and weighed. Then, it was placed in physiological saline to wash away impurities, and the same part of the interscapular brown adipose tissue was precisely cut and weighed, and stored in a - 80 °C refrigerator for subsequent determination of indicators.

[0039] (4) Analysis of mitochondrial changes in mouse adipose tissue

[0040] The interscapular brown adipose tissue was washed with phosphate - buffered saline and fixed in 4% paraformaldehyde, followed by trimming, dehydration, embedding, sectioning, staining, and electron microscopy observation. The number of mitochondria and the number of cristae were analyzed and measured by transmission electron microscopy at magnifications of 8k and 15k respectively.

[0041] II. Experimental results

[0042] The results of the thermogenic function determination of each group of mice were as Figure 1 shown. The results indicated that the adaptive thermoregulatory ability of the mice in the ACE group was significantly lower than that of normal mice, suggesting that the thermogenic function of the mice was impaired. After treatment with piperine, the thermoregulatory ability of the mice in the ACE group was significantly improved. In addition, the infrared imaging temperatures of normal mice and mice after piperine intervention were higher than those of the ACE group.

[0043] The transmission electron microscopy results and quantitative analysis results of mitochondria in the scapular brown fat of each group of mice are as follows Figure 2 shown. Compared with the blank group, the number of mitochondria and the number of cristae in the brown adipose tissue of mice in the ACE group decreased significantly. However, after intervention with piperine, the number of some mitochondria increased and the number of cristae of some mitochondria was restored. This indicates that piperine increases energy consumption by increasing the number of mitochondria and improving mitochondrial dysfunction.

[0044] Example 2 Temperature of the hypothermic mouse model by piperine

[0045] I. RNA extraction

[0046] Take the fat of the mice in Example 1 for RNA extraction. The specific method is as follows: Take the fat stored at -80 °C on ice, quickly weigh 90 - 100 mg of fat and put it into a centrifuge tube containing 1 mL of Trizol, homogenize it with a homogenizer, and pipette it repeatedly. Let it stand at room temperature for 5 min, add 0.2 mL of chloroform, vortex it vigorously to mix evenly, and incubate at room temperature for 3 min. After incubation, centrifuge at 10000 g at 4 °C for 15 min, transfer the colorless upper aqueous phase to a new centrifuge tube, add 0.5 mL of isopropanol, mix well, and incubate at room temperature for 10 min. After standing, centrifuge at 10000 g at 4 °C for 10 min, carefully discard the supernatant to obtain a gelatinous precipitate. Add 1 mL of 75% ethanol to wash the precipitate, vortex it vigorously, centrifuge at 7500 g at 4 °C for 5 min, discard the supernatant, and dry the precipitate at room temperature. The RNA precipitate is dissolved in 100 μL of RNA dissolution solution. Use a ultra-micro spectrophotometer to measure the concentration and purity of RNA. When OD 260 / OD 280 is between 1.8 - 2.0, it indicates that the purity and integrity of the total RNA are good. The total RNA is stored in a -80 °C ultra-low temperature freezer for subsequent experiments.

[0047] II. RT-PCR detection of changes in mouse fat expression

[0048] PCR synthesis of cDNA: Dilute the total fat RNA to 100 ng / μL, load the sample, and react the system on a PCR instrument at 42 °C for 15 min, 85 °C for 5 s to synthesize the cDNA strand, and store it at -20 °C for use.

[0049] RT-PCR experiment: After loading the sample into a 96-well plate according to the qPCR kit, detect it on a qPCR instrument. Set the qPCR program as follows: pre-denaturation at 94 °C for 30 s; denaturation at 94 °C for 5 s, annealing at 60 °C for 30 s, repeat 40 cycles. The Ct value is normalized to GAPDH, and 2 -△△CtThe relative expression levels of genes related to energy metabolism were calculated by the method (the related genes include Cox7a1 gene, Cpt-1b gene, Pgc-1 gene, Ppar-y gene, and Ucp1 gene).

[0050] III. Experimental Results

[0051] The measurement results of the expression levels of genes related to energy metabolism in each group of mice are as Figure 3 shown. Compared with the blank control group, the genes related to energy metabolism in the ACE group of mice were significantly down-regulated. After intervention with piperine, the energy metabolism genes in the mice were significantly up-regulated and recovered, indicating that piperine can significantly increase the expression of energy metabolism genes and improve the symptoms of hypothermia.

[0052] The above embodiments are the preferred embodiments of the present invention, but the embodiments of the present invention are not limited by the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principle of the present invention shall be equivalent replacement methods and are all included in the protection scope of the present invention.

Claims

1. Application of piperine in the preparation of products for improving hypothermia.

2. Application of piperine in the preparation of products for improving hypothermia.

3. Application of piperine in the preparation of products for improving mitochondrial dysfunction.

4. Application of piperine in the preparation of products for enhancing the ability to regulate body temperature.

5. A drug for improving hypothermia or hypothermia, characterized in that: The invention contains piperine and pharmaceutically acceptable auxiliary materials.

6. A drug for improving mitochondrial disease, characterized in that: The invention contains piperine and pharmaceutically acceptable auxiliary materials.

7. The drug according to claim 5 or 6, characterized in that: Pharmaceutical excipients include any one or a combination of at least two of carriers, diluents, excipients, fillers, binders, wetting agents, emulsifiers, solubilizers, surfactants or buffers.

8. The drug according to claim 7, characterized in that: Pharmaceutical excipients also include any one of colorants, pH regulators, antioxidants, and antibacterial agents, or a combination of at least two of them.

9. The drug according to claim 7, characterized in that: The carrier includes liposomes, micelles, dendrimers, microspheres or microcapsules.

10. The drug according to any one of claims 6 to 9, characterized in that: The dosage form of the drug is tablet, capsule, aerosol, pill, powder, solution, emulsion or granule.

Images