Application of ginger active composite component in related products for improving energy metabolism

Abstract

According to the application of the active compound components of the fresh ginger in related products for improving energy metabolism, main active components are extracted from the fresh ginger, and the biological activity of fresh ginger extracts with different concentrations is evaluated; screening a ginger component which is combined with a CaV1.2 calcium channel and activates the channel by utilizing molecular docking simulation; identifying the components of the ginger extract through high performance liquid chromatography; whether the screened ginger component can activate a CaV1.2 channel or not is verified, calcium ion inflow of the CaV1.2 channel is enhanced, 6-gingerol, 6-zingiberone phenol and 6-shogaol are compounded, and the optimal active component formula is screened out. The invention proves that the active components of ginger can promote the flow of calcium ions in cells through a targeted CaV1.2 channel, and further activate an AMPK signal channel, so that the energy metabolism level is remarkably improved, the bottleneck that a calcium ion channel and energy metabolism are difficult to accurately regulate and control by a traditional method is solved, and an accurate and effective natural product application scheme is provided.

Description

Technical Field

[0001] The present invention belongs to the field of extraction and application of natural products, and particularly relates to the use of active composite components of ginger in products related to enhancing energy metabolism. Background Art

[0002] Energy metabolism plays a crucial role in human health, cell function, and organ physiology, especially in controlling the energy supply and consumption of cells, which has a fundamental role.

[0003] Currently, in terms of energy metabolism regulation, traditional technologies mainly rely on drug molecules (such as AMPK activators, mitochondrial function promoters, etc.) to intervene through intracellular signaling pathways to promote metabolic activities such as fatty acid oxidation and glucose metabolism. However, most of the existing drugs are chemically synthesized compounds, which have certain side effects and are difficult to achieve long-term and precise energy regulation. In addition, traditional intervention methods mostly rely on the action on specific intracellular signaling pathways (such as AMPK, mTOR, etc.). Although these pathways have a significant impact on energy metabolism, they are often interfered by external factors (such as the bioavailability of drugs, cell status, etc.) and are difficult to achieve precise control. Summary of the Invention

[0004] Technical problems to be solved: Aiming at the above problems, the purpose of the present invention is to provide the use of active composite components of ginger in products related to enhancing energy metabolism. First, the main active substances of ginger are extracted, and the biological activities of different concentrations of ginger extracts are evaluated. Secondly, molecular docking simulation is used to screen out the ginger components that can bind to and activate the CaV1.2 calcium channel. Then, the components of the ginger extract are identified by high-performance liquid chromatography. Finally, it is verified whether the screened ginger components can activate the CaV1.2 channel, enhance the calcium influx of the CaV1.2 channel, and compound 6-gingerol, 6-paradol, and 6-shogaol according to different mass ratios to screen out the optimal active ingredient formulation. The active components screened by the present invention include 6-gingerol, 6-paradol, and 6-shogaol, which can highly selectively open its channel with CaV1.2, activate the key energy metabolism enzyme AMPK, and improve the body's energy metabolism level, which is beneficial to providing reference for functional foods and drugs targeting CaV1.2.

[0005] Technical solution: The use of active composite components of ginger in products related to enhancing energy metabolism, wherein the active composite components of ginger are a complex of 6-gingerol, 6-paradol, and 6-shogaol.

[0006] Further, the mass ratio of 6-gingerol, 6-paradol, and 6-shogaol is 1:(1 - 3):(1 - 3).

[0007] Furthermore, the screening of the ginger active composite component includes the following steps: S1. Prepare a ginger extract, dilute the prepared extract, and evaluate the biological activities of ginger extracts at different concentrations; S2. Use molecular docking simulation to screen out the ginger components that can bind to and activate the CaV1.2 calcium channel; S3. Identify the components of the ginger extract by high-performance liquid chromatography; S4. Verify whether the screened ginger components can activate the CaV1.2 channel and enhance the calcium influx of the CaV1.2 channel. Compound 6-gingerol, 6-paradol, and 6-shogaol according to different mass ratios, and screen out the active ingredient formulation with the best effect.

[0008] Furthermore, the preparation method of the ginger extract in step S1 is as follows: Take 10 g of ginger powder, dissolve it in 80 - 100 mL of ethyl acetate, perform ultrasonic-assisted extraction at 450 - 480 W for 50 - 60 min, then filter the extract, collect the filtrate, concentrate it by rotary evaporation, and obtain the ginger extract after freeze-drying.

[0009] Furthermore, the concentrations of the ginger extract in step S1 are 10 μg / mL, 50 μg / mL, and 100 μg / mL respectively.

[0010] Furthermore, the conditions of the high-performance liquid chromatography in step S3 are as follows: The chromatographic column is an ASB-C18 chromatographic column, 250×4.6 mm, 5 μm; binary gradient elution is adopted, mobile phase A is pure water, B is chromatographic-grade acetonitrile, and the elution program is: 0 - 5 min, 0 - 20% B; 5 - 45 min, 20 - 90% B; 45 - 70 min, 100% B; the injection volume is 20 μL, the flow rate is 1 mL / min, the column temperature is 30°C, and the detection wavelength is 280 nm.

[0011] Furthermore, the product includes drugs, health products, or foods for the treatment or prevention of metabolic syndrome.

[0012] Beneficial effects

[0013] The present invention is a novel mechanism for enhancing energy metabolism by targeting the calcium channel CaV1.2. Ginger bioactive substances can highly selectively activate the CaV1.2 channel, thereby promoting cellular energy metabolism. Ginger extract, especially at medium concentrations, significantly enhances the activity of AMPK, improves the intracellular energy metabolism level, and further promotes cell growth and proliferation. Low-concentration ginger extract has a slight anti-apoptotic effect, while excessive concentrations may induce apoptosis due to possible cytotoxicity. Ginger extract can significantly regulate the energy metabolism pathway within a certain concentration range. In particular, its active ingredients 6-gingerol, 6-paradol, and 6-shogaol can effectively enhance the activation of AMPK, thereby enhancing the energy metabolism ability of cells.

[0014] Through molecular docking simulation experiments, the present invention screened out ginger bioactive ingredients that can bind to the CaV1.2 calcium channel, including 6-gingerol, 6-paradol, and 6-shogaol. These ingredients can form stable complexes with the binding sites of CaV1.2, showing low binding energy and thus having strong binding ability, providing a strong basis for further screening CaV1.2 channel activators. 6-Gingerol, 6-paradol, and 6-shogaol can activate the CaV1.2 channel, induce calcium ions to flow into cells, increase the intracellular calcium ion concentration, enhance the current response of the channel and be accompanied by an increase in the Fluo-4 fluorescence signal, effectively promoting energy metabolism, further supporting the application potential of 6-gingerol, 6-paradol, and 6-shogaol as potential CaV1.2 channel activators in regulating energy metabolism.

[0015] The present invention not only clarifies the biological activity mechanism of ginger but also provides valuable theoretical basis for the development of functional foods and drugs based on the CaV1.2 channel. It further confirms that ginger ingredients such as 6-gingerol, 6-shogaol, and 6-paradol can effectively activate the CaV1.2 channel and increase the influx of calcium ions in different proportion combinations, thereby improving the energy metabolism level. Through detailed analysis of the binding force and activation effect of different combinations of ginger active formulas with the CaV1.2 channel, more precise optimized application plans can be formulated to improve the efficacy and safety of ginger ingredients in clinical applications. Brief Description of the Drawings

[0016] Figure 1 It is a flow chart for the extraction, screening of ginger ingredients and their binding to the CaV1.2 channel; Figure 2 It is a high-performance liquid chromatography (HPLC) chart of the standard polyphenol substances in ginger extract (the marked 1-4 are 6-gingerol, 6-paradol, gingerone, and 6-shogaol in sequence); Figure 3 It is a high-performance liquid chromatography (HPLC) chart of a 50 μg / mL ginger extract sample. Detailed Embodiments

[0017] The present invention provides the use of ginger active composite components in products for enhancing energy metabolism. To make the objectives, technical solutions and effects of the present invention clearer and more definite, the following will further elaborate on the present invention in conjunction with embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0018] Example 1

[0019] An in vitro experiment was conducted to detect the effect of ginger extract on the human muscle cell line C2C12, including the following steps: S1. Cell culture The human muscle cell line C2C12 was selected and cultured to the logarithmic growth phase. The cells were cultured with DMEM medium and maintained in an environment of 37°C and 5% CO 2 . S2. Preparation of ginger extract 10 g of ginger powder was taken and dissolved in ethyl acetate at a solid-liquid ratio of 1:10 (g / mL). After ultrasonic-assisted extraction at 480 W for 60 min, the extract was filtered by suction. The filtrate was collected, concentrated by rotary evaporation, and then freeze-dried to obtain ginger extract. S3. Cell viability assessment The prepared extract was diluted to 10 μg / mL, 50 μg / mL, and 100 μg / mL respectively and added to the cell culture medium. The cell proliferation, apoptosis rate, and expression of energy metabolism-related enzymes such as AMPK and ACC were evaluated after treatment for 24 h, 48 h, and 72 h respectively.

[0020] Comparative Example 1

[0021] The difference between this comparative example and Example 1 is that no ginger extract was added. Specifically as follows: An in vitro experiment was conducted to detect the effect of not adding ginger extract on the human muscle cell line C2C12, including the following steps: S1. Cell culture The human muscle cell line C2C12 was selected and cultured to the logarithmic growth phase. The cells were cultured according to the conventional method and maintained in an environment of 37°C and 5% CO 2 . S2. Cell viability assessment The cell proliferation, apoptosis rate, and expression of energy metabolism-related enzymes such as AMPK and ACC were evaluated after treatment for 24 h, 48 h, and 72 h.

[0022] Comparative Example 2

[0023] The difference between this comparative example and Example 1 is that no ginger extract was added, and an AMPK agonist was added. Specifically as follows: In vitro experiments were conducted to detect the effects of AMPK agonists on the human muscle cell line C2C12, including the following steps: S1. Cell culture Select the human muscle cell line C2C12 and culture it until the logarithmic growth phase. Culture the cells in DMEM medium and maintain them at 37°C in an environment of 5% CO 2 . S2. Cell activity assessment Add 1 mM AMPK agonist to the cell culture medium and evaluate cell proliferation, apoptosis rate, and the expression of AMPK and ACC energy metabolism-related enzymes after treatment for 24 h, 48 h, and 72 h, respectively.

[0024] Performance test (1) Cell activity assessment Use the MTT method to detect cell proliferation, flow cytometry to evaluate the apoptosis rate, and Western blot to detect the expression of AMPK and ACC energy metabolism-related enzymes.

[0025] Table 1 Effects of Example 1 and Comparative Examples 1-2 on cell activity

[0026] Concentration-effect curves were plotted using MTT absorbance data to evaluate the effect of ginger extract on cell proliferation, and cell apoptosis rate curves were plotted using flow cytometry data to evaluate the anti-apoptotic effect of ginger extract. Western blot analysis was used to evaluate the phosphorylation levels of AMPK and ACC to assess the activation effect of ginger extract on the energy metabolism pathway. In the above in vitro experiments, the regulatory effects of ginger extract (especially its active ingredients) on cell proliferation, apoptosis, and energy metabolism were investigated. In vitro experiments showed that 10 μg / mL and 50 μg / mL could significantly promote cell proliferation, while 100 μg / mL inhibited cell proliferation, indicating that ginger extract has a promoting effect on cell growth within a certain concentration range, but high concentrations have an inhibitory effect. At low concentrations, ginger extract has a slight anti-apoptotic effect; the inhibitory effect on apoptosis is best at 50 μg / mL, while 100 μg / mL may induce more cell apoptosis due to cytotoxicity. Ginger extract can significantly activate the AMPK pathway and promote energy metabolism, especially most significantly at 50 μg / mL. At 100 μg / mL, although the activation effect of AMPK is enhanced, the inhibitory effect on ACC is prominent, possibly due to the negative feedback mechanism of AMPK. Ginger extract, especially at 50 μg / mL, can significantly enhance the energy metabolism level of cells by activating the AMPK signaling pathway, thereby promoting cell proliferation and growth. Excessive concentrations may trigger cell apoptosis and have a negative impact on energy metabolism.

[0027] Example 2 Through molecular docking simulation, the ginger components that form stable complexes with the binding sites of the CaV1.2 channel were screened out, and the specific steps are as follows: S1. Download and construct the molecular structures of gingerols, shogaols, gingerones, and paradols through PubChem (https: / / pubchem.ncbi.nlm.nih.gov / ). S2. Optimize in Pymol software and then open the file in Autodock Tools software to adjust the charge number. S3. The CaV1.2 protein structure was downloaded from the PDB database (http: / / www.rcsb.org / ), and the receptor structure for docking was obtained through preprocessing such as removing water, adding hydrogen, and calculating charges. S4. Use the molecular docking software AutoDock Tools to dock the ginger components with adjusted charge numbers in S2 and the receptor structure in S3 to obtain the receptor-ligand conformation in the docking state, and calculate the docking activation energy.

[0028] Table 2 Docking binding energy of ginger components and CaV1.2

[0029] The binding free energy in the docking results, which is an index to measure the binding stability of molecules. A lower binding free energy value usually indicates more stable binding between molecules. As can be seen from Table 2, the order of binding energy is 6-gingerol < 6-shogaol < 6-paradol < 8-shogaol < 10-gingerol < gingerone < 12-gingerol < 10-shogaol < 8-gingerol. Therefore, according to the molecular docking experiment, the binding stability order of ginger active compounds and the CaV1.2 receptor is: 6-gingerol > 6-shogaol > 6-paradol > 8-shogaol > 10-gingerol > gingerone > 12-gingerol > 10-shogaol > 8-gingerol.

[0030] Example 3

[0031] Identify the components of the ginger extract by high performance liquid chromatography, including the following steps: S1. Load the ginger extract after passing through a 0.45 μm microporous organic filter membrane. S2. Identify the components of the filtered ginger extract by high performance liquid chromatography.

[0032] Performance Test

[0033] (1) High Performance Liquid Chromatography The chromatographic column was an ASB-C18 chromatographic column, 250×4.6 mm, 5 μm; binary gradient elution was adopted, mobile phase A was pure water, B was chromatographic grade acetonitrile, and the elution program was: 0 - 5 min, 0 - 20% B; 5 - 45 min, 20 - 90% B; 45 - 70 min, 100% B. The injection volume was 20 μL, the flow rate was 1 mL / min, the column temperature was 30°C, and the detection wavelength was 280 nm.

[0034] Figure 2 It is the high-performance liquid chromatogram of the polyphenol standard in ginger extract (the marked 1 - 4 are 6-gingerol, 6-paradol, gingerone, and 6-shogaol in sequence). Figure 3 It is the high-performance liquid chromatogram of the 50 μg / mL ginger extract sample. Through data processing and analysis, the regression equations of the four standards of 6-gingerol, 6-paradol, gingerone, and 6-shogaol are shown in Table 3. y represents the peak area, x represents the content. After comparing with the retention time chromatogram of the reference standard and calculating the integrated peak area, the relative contents of the four polyphenol compounds in ginger are obtained. The contents of 6-gingerol, 6-paradol, gingerone, and 6-shogaol are 9.14 mg / g, 1.23 mg / g, 0.34 mg / g, and 1.61 mg / g respectively. Among them, the content of 6-gingerol is the highest, and the content of gingerone is the lowest. The ginger extract contains 6-gingerol, 6-shogaol, and 6-paradol with strong binding stability to the CaV1.2 receptor.

[0035] Table 3 Linear regression equations of 6-gingerol, 6-paradol, gingerone, and 6-shogaol standards

[0036] Example 4

[0037] Verify whether 10 μg / mL 6-gingerol can activate the CaV1.2 channel, which includes the following steps in parts by weight: S1. Add the Fluo-4 calcium ion probe to C2C12 cells. Inoculate the C2C12 cells at 1×10 4 / mL into a 24-well plate and culture them routinely with DMEM complete culture medium. After the cells adhere for 12 h, change to DMEM complete culture medium containing different ginger polyphenol compounds and continue to culture. When the probe binds to the calcium ions in the cells, it emits fluorescence. Add 10 μg / mL 6-gingerol to the cells and monitor the influx of calcium ions through the change of fluorescence signal. S2. Confirm whether 6-gingerol can highly selectively open the CaV1.2 channel through electrophysiological techniques (whole-cell patch clamp).

[0038] Example 5

[0039] Verify whether 10 μg / mL 6-gingerenol can activate the CaV1.2 channel. By weight, it includes the following steps: S1. Add the Fluo-4 calcium ion probe to C2C12 cells. Seed the C2C12 cells at a density of 1×10 4 / mL into a 24-well plate and culture them routinely with DMEM complete culture medium. After the cells adhere for 12 h, change to DMEM complete culture medium containing different ginger polyphenolic compounds and continue culturing. When the probe binds to calcium ions inside the cells, it emits fluorescence. Add 10 μg / mL 6-gingerenol to the cells and monitor the influx of calcium ions through the change in fluorescence signal. S2. Confirm whether 6-gingerenol can highly selectively open the CaV1.2 channel through electrophysiological techniques (whole-cell patch clamp).

[0040] Example 6

[0041] Verify whether 10 μg / mL 6-paradol can activate the CaV1.2 channel. By weight, it includes the following steps: S1. Add the Fluo-4 calcium ion probe to C2C12 cells. Seed the C2C12 cells at a density of 1×10 4 / mL into a 24-well plate and culture them routinely with DMEM complete culture medium. After the cells adhere for 12 h, change to DMEM complete culture medium containing different ginger polyphenolic compounds and continue culturing. When the probe binds to calcium ions inside the cells, it emits fluorescence. Add 10 μg / mL 6-paradol to the cells and monitor the influx of calcium ions through the change in fluorescence signal. S2. Confirm whether 6-paradol can highly selectively open the CaV1.2 channel through electrophysiological techniques (whole-cell patch clamp).

[0042] Comparative Example 3

[0043] The difference between this comparative example and Example 4 is that no ginger active ingredient is added.

[0044] Verify whether the absence of any ginger active ingredient can activate the CaV1.2 channel. By weight, it includes the following steps: S1. Add the Fluo-4 calcium ion probe to C2C12 cells. Seed the C2C12 cells at a density of 1×10 4 / mL into a 24-well plate and culture them routinely with DMEM complete culture medium. After the cells adhere for 12 h, change to DMEM complete culture medium containing different ginger polyphenolic compounds and continue culturing. When the probe binds to calcium ions inside the cells, it emits fluorescence. Monitor the influx of calcium ions through the change in fluorescence signal. S2. Confirm whether the CaV1.2 channels can be highly selectively opened without adding any ginger active ingredients through electrophysiological techniques (whole-cell patch clamp).

[0045] Comparative Example 4

[0046] The difference between this comparative example and Example 4 is the use of Bay K8644.

[0047] Verify whether adding Bay K8644 can activate the CaV1.2 channels. By weight, it includes the following steps: S1. Add the Fluo-4 calcium ion probe to C2C12 cells. Seed the C2C12 cells at a density of 1×10 4 / mL into a 24-well plate and culture them routinely with DMEM complete culture medium. After the cells adhere for 12 h, change to DMEM complete culture medium containing different ginger polyphenol compounds and continue culturing. When the probe binds to calcium ions inside the cells, it emits fluorescence. Add 10 μM BayK8644 to the cells and monitor the influx of calcium ions through the change in fluorescence signal. S2. Confirm whether the CaV1.2 channels can be highly selectively opened without adding anything through electrophysiological techniques (whole-cell patch clamp).

[0048] Performance Test (1) Detection of intracellular calcium ion concentration Rinse the cells 2 - 3 times with PBS buffer to wash away impurities on the cell surface. Add Fluo-4 working solution (400 μL) and Pluronic F-127 (2.5 μL) to the 24-well plate to completely cover the cells and gently shake slowly. Wrap the cell plate with tin foil and incubate it in the dark at 37 °C for 30 minutes. Initially check under a fluorescence microscope whether there is green fluorescence inside the cells. After confirming the labeling is complete (when the green fluorescence of the cells is higher than the fluorescence of the solution background), wash away the non-specifically bound fluorescence outside the cells with PBS solution. The time should be short to avoid the probe being pumped out of the cells. Measure the intracellular Ca²⁺ concentration, observe the fluorescence with an excitation light wavelength of 528 nm (excitation: 490 - 500 nm) and save the experimental results.

[0049] (2) Whole-cell patch clamp

[0050] All the liquid preparations in the patch clamp experiment were carried out strictly in accordance with the requirements of electrophysiological experiments. The narrow-mouth bottles for storing the liquid and the bacterial filter membranes were sterilized before use. The inner / outer liquid components were dissolved in 900 mL of water, the pH was adjusted to 7.4 with NaOH, and then transferred to a volumetric flask and made up to 1 L. It was filtered through a bacterial filter membrane into a narrow-mouth bottle and stored in a 4 °C refrigerator. Since in the inside-out mode, 1 μM Ca²⁺ can saturate the current value of TMEM16A, the calcium-containing liquid was selected at a concentration of 1 μM. The inside-out mode was mainly used in the experiment. This mode can exclude the interaction between other substances and structures inside the cell and the natural compounds, and detect the direct effect of ginger polyphenol compounds on the calcium-activated chloride channel (CaCCs). After being polished, the glass microelectrode had an access resistance maintained between 0.7 and 1.5 MΩ, and the seal resistance was above 1.5 GΩ. The patch clamp amplifier used the EPC 10 series from HEKA Company in Germany. Pulse software was used for data acquisition. The itx format files could be converted into ABF type files by using mini analysis software, and then could be recognized and read by Clampfit 9.0 software. Clampfit 9.0 has functions such as removing baseline current, obtaining current peak values, current average values, and fitting inactivation and activation curves.

[0051] Table 4 Detection of intracellular calcium ion concentration and calcium signal results

[0052] As can be seen from Table 4, the Ca²⁺ concentration in Examples 4-6 increased significantly. More Ca²⁺ would bind to the binding site of the CaV1.2 channel, manifested as an increase in apparent binding energy. At the same time, the fluorescence signals in Examples 4-6 were significantly higher than those in Comparative Example 3 and similar to those in Comparative Example 4, indicating that they activated the CaV1.2 channel and increased calcium ion influx. Examples 4-6 significantly enhanced the current of the CaV1.2 channel, and the current characteristics were similar to those in Comparative Example 4, indicating that they could highly selectively activate the CaV1.2 channel.

[0053] Example 7 To screen the optimal active ingredient formulation, by weight, the following steps are included: S1. In cells expressing the CaV1.2 channel, add 1 part of 10 μg / mL 6-gingerol, 1 part of 10 μg / mL 6-paradol, and 1 part of 10 μg / mL 6-shogaol. Using the Fluo-4 calcium ion probe method, monitor the change in intracellular calcium ion concentration, record the fluorescence intensity, and analyze the change in calcium ion influx; S2. Fix the cells expressing the CaV1.2 channel in the patch clamp experimental system, add 1 part of 10 μg / mL 6-gingerol, 1 part of 10 μg / mL 6-paradol, and 1 part of 10 μg / mL 6-shogaol, record the changes in intracellular current, evaluate the opening and closing of the CaV1.2 channel, and analyze its effect on the CaV1.2 channel current.

[0054] S3. Effects on mouse body temperature and respiratory metabolism In the 4th week of the animal experiment, transfer the mice in different treatment groups to the small animal respiratory metabolism monitoring system. Supply feed and drinking water as usual. After the mice are adapted for 24 h, start collecting metabolic data. The monitoring indicators include: oxygen consumption rate ( ), energy consumption.

[0055] On the day when the experiment ends, use a digital medical infrared camera to take pictures of the mice in different treatment groups respectively, and analyze the epidermal temperature at the scapula with software.

[0056] Example 8

[0057] To screen the optimal active ingredient formulation, calculated by weight, the following steps are included: S1. In the cells expressing the CaV1.2 channel, add 1 part of 10 μg / mL 6-gingerol, 2 parts of 10 μg / mL 6-paradol, and 1 part of 10 μg / mL 6-shogaol. Use the Fluo-4 calcium ion probe method to monitor the changes in intracellular calcium ion concentration, record the fluorescence intensity, and analyze the changes in calcium ion influx; S2. Fix the cells expressing the CaV1.2 channel in the patch clamp experimental system, add 1 part of 10 μg / mL 6-gingerol, 2 parts of 10 μg / mL 6-paradol, and 1 part of 10 μg / mL 6-shogaol, record the changes in intracellular current, evaluate the opening and closing of the CaV1.2 channel, and analyze its effect on the CaV1.2 channel current.

[0058] S3. Effects on mouse body temperature and respiratory metabolism In the 4th week of the animal experiment, transfer the mice in different treatment groups to the small animal respiratory metabolism monitoring system. Supply feed and drinking water as usual. After the mice are adapted for 24 h, start collecting metabolic data. The monitoring indicators include: oxygen consumption rate ( ), energy consumption.

[0059] On the day when the experiment ends, use a digital medical infrared camera to take pictures of the mice in different treatment groups respectively, and analyze the epidermal temperature at the scapula with software.

[0060] Example 9

[0061] To screen the optimal active ingredient formulation, by weight, the following steps are included: S1. In cells expressing the CaV1.2 channel, add 1 part of 10 μg / mL 6-gingerol, 3 parts of 10 μg / mL 6-paradol, and 1 part of 10 μg / mL 6-shogaol. Using the Fluo-4 calcium ion probe method, monitor the change in intracellular calcium ion concentration, record the fluorescence intensity, and analyze the change in calcium ion influx; S2. Fix the cells expressing the CaV1.2 channel in the patch clamp experimental system, add 1 part of 10 μg / mL 6-gingerol, 3 parts of 10 μg / mL 6-paradol, and 1 part of 10 μg / mL 6-shogaol, record the change in intracellular current, evaluate the opening and closing of the CaV1.2 channel, and analyze its effect on the CaV1.2 channel current.

[0062] S3. Effects on mouse body temperature and respiratory metabolism In the 4th week of the animal experiment, transfer the mice in different treatment groups to the small animal respiratory metabolism monitoring system. Feed and drinking water are supplied as usual. After the mice are adapted for 24 h, start collecting metabolic data. The monitoring indicators include: oxygen consumption rate ( ), energy consumption.

[0063] On the day when the experiment ends, use a digital medical infrared camera to take pictures of the mice in different treatment groups respectively, and use software to analyze the epidermal temperature at the scapula.

[0064] Example 10

[0065] To screen the optimal active ingredient formulation, by weight, the following steps are included: S1. In cells expressing the CaV1.2 channel, add 1 part of 10 μg / mL 6-gingerol, 2 parts of 10 μg / mL 6-paradol, and 2 parts of 10 μg / mL 6-shogaol. Using the Fluo-4 calcium ion probe method, monitor the change in intracellular calcium ion concentration, record the fluorescence intensity, and analyze the change in calcium ion influx; S2. Fix the cells expressing the CaV1.2 channel in the patch clamp experimental system, add 1 part of 10 μg / mL 6-gingerol, 2 parts of 10 μg / mL 6-paradol, and 2 parts of 10 μg / mL 6-shogaol, record the change in intracellular current, evaluate the opening and closing of the CaV1.2 channel, and analyze its effect on the CaV1.2 channel current.

[0066] S3. Effects on mouse body temperature and respiratory metabolism In the 4th week of the animal experiment, mice in different treatment groups were transferred to a small animal respiratory metabolism monitoring system. Feed and drinking water were supplied as usual. After the mice were allowed to adapt for 24 h, metabolic data collection began. The monitoring indicators included: oxygen consumption rate ( ), and energy consumption.

[0067] On the day when the experiment ended, digital medical infrared cameras were used to take pictures of the mice in different treatment groups respectively, and software was used to analyze the epidermal temperature at the scapula.

[0068] Example 11

[0069] To screen the optimal active ingredient formulation, calculated by weight parts, the following steps are included: S1. In cells expressing the CaV1.2 channel, add 1 part of 10 μg / mL 6-gingerol, 2 parts of 10 μg / mL 6-paradol, and 3 parts of 10 μg / mL 6-shogaol. Using the Fluo-4 calcium ion probe method, monitor the change in intracellular calcium ion concentration, record the fluorescence intensity, and analyze the change in calcium ion influx; S2. Fix the cells expressing the CaV1.2 channel in the patch clamp experiment system, add 1 part of 10 μg / mL 6-gingerol, 2 parts of 10 μg / mL 6-paradol, and 3 parts of 10 μg / mL 6-shogaol, record the change in intracellular current, evaluate the opening and closing of the CaV1.2 channel, and analyze its effect on the CaV1.2 channel current.

[0070] Effect on mouse body temperature and respiratory metabolism In the 4th week of the animal experiment, mice in different treatment groups were transferred to a small animal respiratory metabolism monitoring system. Feed and drinking water were supplied as usual. After the mice were allowed to adapt for 24 h, metabolic data collection began. The monitoring indicators included: oxygen consumption rate ( ), and energy consumption.

[0071] On the day when the experiment ended, digital medical infrared cameras were used to take pictures of the mice in different treatment groups respectively, and software was used to analyze the epidermal temperature at the scapula.

[0072] Performance test (1) Energy consumption The calculation formula for energy consumption is as follows: Where represents the oxygen consumption rate, L / kg / h; 4.825 is the thermal equivalent of oxygen, kcal / L.

[0073] Table 5 Results of calcium flow experiments and electrophysiological techniques for Examples 7 - 11

[0074] As can be seen from Table 5, the calcium ion concentration gradually increases in Examples 7-11, and the corresponding fluorescence intensity gradually increases and the absolute value of the current intensity increases, indicating that ginger polyphenol compounds in different proportions can effectively enhance the intracellular calcium ion concentration; in the mouse experiment, Examples 7-11 show that under the intervention of different combinations of ginger polyphenol compounds, the body temperature and energy consumption increase to varying degrees, indicating the application prospect of the active composite components of ginger (6-gingerol, 6-paradol, and 6-shogaol) in enhancing energy metabolism.

[0075] The above are only the preferred embodiments of the present invention, and do not impose any form of limitation on the present invention. Any person skilled in the art can make many possible changes and modifications to the technical solution of the present invention by using the disclosed methods and technical contents without departing from the spirit and technical solution of the present invention, or modify it into equivalent embodiments with equivalent changes. Therefore, any simple modification, equivalent replacement, equivalent change and modification made to the above embodiments based on the technical essence of the present invention without departing from the technical solution of the present invention still fall within the scope of protection of the technical solution of the present invention.

Claims

1. The use of ginger active compound ingredients in products related to improving energy metabolism, characterized in that: The active composite component of ginger is a composite of 6-gingerol, 6-gingerone phenol and 6-shogaol.

2. The use of the ginger active compound component according to claim 1 in products related to improving energy metabolism, characterized in that: The mass ratio of 6-gingerol, 6-gingerone phenol and 6-shogaol is 1:(1-3):(1-3).

3. The use of the active compound component of ginger according to claim 1 in products related to improving energy metabolism, characterized in that: The screening of the active composite ingredients of ginger comprises the following steps: S1. preparing ginger extract, diluting the prepared extract, and evaluating the biological activity of ginger extract at different concentrations; S2. Molecular docking simulation was used to screen out ginger components that can bind to and activate CaV1.2 calcium channels; S3. Identification of ginger extract components by high performance liquid chromatography; S4. Verify whether the selected ginger ingredients can activate CaV1.2 channels and enhance the calcium ion influx of CaV1.2 channels, compound 6-gingerol, 6-gingerone phenol, and 6-shogaol according to different mass ratios, and screen out the active ingredient formula with the best effect.

4. The use of the active compound component of ginger according to claim 3 in products related to improving energy metabolism, characterized in that: The preparation method of the ginger extract in step S1 is as follows: Take 10 g of ginger powder, dissolve it in 80-100 mL of ethyl acetate, perform ultrasonic-assisted extraction at 450-480 W for 50-60 min, filter the extract, collect the filtrate, concentrate it by rotary evaporation, and then freeze-dry it to obtain the ginger extract.

5. The use of the active compound component of ginger according to claim 3 in products related to improving energy metabolism, characterized in that: The concentrations of the ginger extract in step S1 are 10 μg / mL, 50 μg / mL and 100 μg / mL respectively.

6. The use of the active compound component of ginger in products related to improving energy metabolism according to claim 3, characterized in that: The conditions of high performance liquid chromatography in step S3 are: The chromatographic column was an ASB-C18 column, 250×4.6 mm, 5 μm. Binary gradient elution was adopted, the mobile phase A was pure water, B was chromatographic grade acetonitrile, and the elution program was: 0-5 min, 0-20% B; 5-45 min, 20-90% B; 45-70 min, 100% B; the injection volume was 20 μL, the flow rate was 1 mL / min, the column temperature was 30°C, and the detection wavelength was 280 nm.

7. The use of the active compound component of ginger according to claim 1 in products related to improving energy metabolism, characterized in that: The products include medicines, health products or foods for treating or preventing metabolic syndrome.

Images