Gemfibrozil-piperine nanostructure lipid carrier as well as preparation method and application thereof

Abstract

The invention discloses a gemfibrozil-piperine nanostructure lipid carrier as well as a preparation method and application thereof, and belongs to the technical field of medicines. According to the gemfibrozil-piperine nanostructure lipid carrier as well as the preparation method and the application thereof disclosed by the invention, the gemfibrozil and the piperine are entrapped in the nanostructure lipid carrier for the first time, so that the solubility of insoluble drugs is improved, the lipid-lowering effect of the gemfibrozil is enhanced, and the occurrence of adverse reactions of the gemfibrozil is reduced; meanwhile, the preparation process is controlled to realize the slow release effect of the medicine, the administration frequency is reduced, the compliance of a patient is improved, and a reference is provided for development and clinical application of a lipid-lowering medicine preparation.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and more particularly to a gemfibrozil-piperine nanostructured lipid carrier and a preparation method and application thereof. Background Art

[0002] Hyperlipidemia is a systemic disease caused by abnormal lipid metabolism or transport, also known as dyslipidemia. Common symptoms include dizziness, fatigue, headache, memory loss, and limb numbness. Hyperlipidemia is not only a standalone condition but also a contributing factor to numerous other conditions, including cardiovascular disease, atherosclerosis, diabetes, and obesity. With improvements in living standards and changes in lifestyle, the incidence of hyperlipidemia is increasing annually, and the age of onset is becoming younger and younger.

[0003] Currently, common clinical therapeutic agents include statins, fibrates, and niacin. However, while these drugs can lower blood lipids, they can also cause a variety of adverse reactions, making them unsuitable for long-term lipid regulation. In the search for more effective measures to prevent and treat hyperlipidemia, combination therapy is undoubtedly a promising lipid-lowering treatment approach. In recent years, numerous studies have demonstrated that dual-drug combinations offer superior lipid-lowering efficacy compared to either drug alone, leading to widespread clinical application and widespread acceptance.

[0004] Gemfibrozil belongs to the fibrate class of lipid-lowering drugs. This class of drugs activates peroxisome proliferator-activated receptor α, reducing ApoC3, which in turn activates LPL and ApoA5, lowering triglycerides (TG), triglycerides (TC), and LDL-C levels while increasing HDL-C levels, ultimately achieving the effect of lowering blood lipids. As a lipid regulator, gemfibrozil's lipid-regulating mechanism primarily inhibits the synthesis of very low-density lipoprotein (VLDL) in the liver, increases lipoprotein lipase activity, and accelerates the catabolism of VLDL, thereby inhibiting the synthesis of TG and LDL.

[0005] Piperine, derived from the natural product pepper, has a favorable safety profile and can inhibit the activity of various drug metabolism and transport enzymes, thereby improving the bioavailability of some therapeutic drugs and phytochemicals. Studies have shown that piperine exhibits numerous pharmacological effects, including antioxidant, anti-inflammatory, anticonvulsant, lipid-lowering, and anti-tumor activities. It has been reported that administration of piperine can improve metabolic abnormalities induced by a high-fat diet, such as lowering serum triglycerides, cholesterol, and low-density lipemia, as well as hepatic steatosis and insulin resistance. Piperine is also a well-established glucuronyltransferase inhibitor, effectively inhibiting the binding of glucuronic acid to drugs, thereby reducing their renal clearance and prolonging their duration of action.

[0006] Nanostructured lipid carriers are a new generation of colloidal drug delivery systems developed in the late 20th century based on solid lipid nanoparticles. They offer excellent physiological compatibility, high drug stability, and low irritation. They exhibit sustained-release, targeted, and long-lasting properties, and have a high encapsulation efficiency for lipophilic drugs. During the preparation process, liquid lipids with significantly different physical and chemical properties from the solid lipids are added to disrupt the perfect crystal lattice, thereby achieving higher drug encapsulation efficiency and improved stability. The solid-liquid lipid ratio is controlled, resulting in nanostructured lipid carriers with enhanced sustained-release and controlled-release properties. These nanostructured lipid carriers can be used for various routes of administration, including oral, transdermal, pulmonary, and intravenous administration. The oil phase in nanostructured lipid carriers promotes bile secretion, alters gastrointestinal motility, and stimulates the secretion of bile salts and phospholipids. Bile salts are endogenous surfactants present in the human intestine and can mix with phospholipids to form mixed micelles, increasing intestinal mucosal permeability and promoting intestinal lymphatic transport. The secretion of endogenous solubilizing components such as bile salts and the intake of exogenous components (oil phase) can enhance the solubility of drugs in the gastrointestinal tract, keep the drugs dissolved in the gastrointestinal tract, improve the absorption of drugs in the gastrointestinal tract, and achieve the purpose of improving bioavailability. It is a good new dosage form for oral administration with broad development prospects.

[0007] Therefore, providing a gemfibrozil-piperine nanostructured lipid carrier and a preparation method and application thereof is a problem that needs to be solved urgently by those skilled in the art. Summary of the Invention

[0008] In view of this, the present invention provides a gemfibrozil-piperine nanostructured lipid carrier, its preparation method, and application. This novel pharmaceutical preparation has a good lipid-lowering effect and can effectively alleviate gemfibrozil-induced muscle damage. The preparation method is simple and the preparation is stable.

[0009] In order to achieve the above object, the present invention adopts the following technical solutions:

[0010] The preparation method of gemfibrozil-piperine nanostructured lipid carrier comprises the following steps:

[0011] (1) Weigh 0.17 g of gemfibrozil, 0.08 g of piperine, 4.00 g of glyceryl behenate, and 1.00 g of isopropyl myristate, mix well, and heat in a 75°C water bath until completely melted to form the oil phase;

[0012] (2) Poloxamer 188 and Tween 80 were mixed in a mass ratio of 1:1 to obtain an emulsifier; 2.00 g of the emulsifier was dissolved in distilled water to a concentration of 1.91% as the aqueous phase;

[0013] (3) The 75°C aqueous phase was added dropwise to the oil phase at 800 r / min and magnetically stirred for 1.52 h to obtain colostrum; ultrasonic dispersion was performed at 350 W power for 10 min, and after cooling, the mixture was filtered through a 0.45 μm filter membrane to obtain the gemfibrozil-piperine nanostructured lipid carrier.

[0014] Furthermore, the method described above prepares a gemfibrozil-piperine nanostructured lipid carrier.

[0015] Furthermore, the gemfibrozil-piperine nanostructured lipid carrier is used in the preparation of lipid-lowering drugs.

[0016] Furthermore, the gemfibrozil-piperine nanostructured lipid carrier is used in the preparation of drugs that enhance the lipid-lowering effect.

[0017] Furthermore, the gemfibrozil-piperine nanostructured lipid carrier is used to improve the solubility of poorly soluble drugs.

[0018] Furthermore, the gemfibrozil-piperine nanostructured lipid carrier is used in drug sustained release.

[0019] Through the above technical solution, it can be known that compared with the prior art, the present invention discloses a gemfibrozil-piperine nanostructured lipid carrier and its preparation method and application. Hyperlipidemia is a systemic chronic disease and patients need long-term medication. The first-line lipid-lowering drug fibrates are often accompanied by side effects. While improving blood lipid levels, it is extremely important to pay attention to and improve the reduction of drug side effects. In view of the long-term medication characteristics of hyperlipidemia and the shortcomings of gemfibrozil and piperine being poorly water-soluble, the present invention encapsulates gemfibrozil and piperine in a nanostructured lipid carrier for the first time. While improving the solubility of poorly soluble drugs, the lipid-lowering effect of gemfibrozil is enhanced and the occurrence of its adverse reactions is reduced. At the same time, the preparation process is controlled to achieve a sustained-release effect of the drug, reduce the number of administrations, and improve the patient's compliance, providing a reference for the development and clinical application of lipid-lowering drug preparations. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are merely embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying any creative work.

[0021] Figure 1 The transmission electron micrograph (A) and sample image (B) of the gemfibrozil-piperine nanostructured lipid carrier of the present invention are shown;

[0022] Figure 2The particle size distribution diagram (A) and zeta potential distribution diagram (B) of the gemfibrozil-piperine nanostructured lipid carrier of the present invention;

[0023] Figure 3 In vitro release curves of gemfibrozil (Free-GEM), piperine (Free-PIP) and gemfibrozil-piperine nanostructured lipid carrier (GEM-PIP-NLC) of the present invention in (a) pH 1.2 and (b) pH 6.8 release media (n=3);

[0024] Figure 4 The in vivo drug-dose curves of the gemfibrozil bulk drug (Free GEM) and the gemfibrozil-loaded nanoformulation (GEM-loaded NLC) of the present invention were orally administered to rats (n=6);

[0025] Figure 5 The in vivo drug-dose curves of the piperine bulk drug (Free PIP) and the piperine-loaded nanoformulation (PIP-loaded NLC) of the present invention were orally administered to rats (n=6);

[0026] Figure 6 The serum biochemical indexes of the present invention were detected and analyzed; compared with the NC group, # P<0.05, ## P<0.01; compared with the MC group, * P<0.05, ** P<0.01, *** P < 0.0001;

[0027] Figure 7 The liver enzyme and antioxidant index detection of the present invention; compared with the NC group, # P<0.05, ## P<0.01; compared with the MC group, * P<0.05, ** P < 0.01;

[0028] Figure 8 This is a microscopic photograph of the pathological section of the mouse liver of the present invention (×200);

[0029] Figure 9 This is a microscopic photograph of the epididymal fat pathological section of the mouse of the present invention (×200);

[0030] Figure 10 This is the pathological change of gastrocnemius muscle tissue in the present invention (×200). DETAILED DESCRIPTION

[0031] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0032] Gemfibrozil (purity > 98%) and piperine (purity > 98%) were purchased from Shanghai MacLean Biochemical Technology Co., Ltd.; glyceryl behenate, poloxamer 188, and soybean lecithin were purchased from Sinopharm Chemical Reagent Co., Ltd.; isopropyl myristate was purchased from Aladdin Chemical Reagent Co., Ltd.; DF101-S heat-collecting constant temperature magnetic stirrer was purchased from Shanghai Lichen Instrument Technology Co., Ltd.; and JY92-2D ultrasonic cell disruptor was purchased from Ningbo Xinzhi Biotechnology Co., Ltd.

[0033] Example 1 GEM-PIP-NLC Preparation Method

[0034] Gemfibrozil-piperine nanostructured lipid carrier (GEM-PIP-NLC) was prepared by melt emulsification-ultrasonic dispersion. Gemfibrozil (0.17 g), piperine (0.08 g), solid lipid (glyceryl behenate) (4.00 g), and liquid lipid (isopropyl myristate) (1.00 g) were weighed and mixed. The mixture was heated in a 75°C water bath until completely melted, which served as the oil phase. The drug (gemfibrozil and piperine) to lipid (solid lipid and liquid lipid) ratio was 1:20. Take 2.00 g of the prescribed amount of emulsifier (Poloxamer 188 and Tween 80, mass ratio 1:1) and dissolve it in distilled water to a concentration of 1.91% as the aqueous phase; add the 75°C aqueous phase dropwise to the oil phase at 800 r / min and magnetically stir for 1.52 h to obtain colostrum; ultrasonically disperse at 350 W power for 10 min, cool and filter through a 0.45 μm filter membrane to obtain the GEM-PIP-NLC solution.

[0035] See the finished product Figure 1 Transmission electron microscopy images of gemfibrozil-piperine nanostructured lipid carriers ( Figure 1 A) and sample diagram ( Figure 1 B) Transmission electron microscopy revealed that the GEM-PIP-NLC particles were spherical, with a uniform particle size distribution and no obvious particle aggregation. The sample was homogeneous, transparent, and had a light blue opalescence.

[0036] Example 2 Characterization of Gemfibrozil-Piperine Nanostructured Lipid Carrier

[0037] Take 2 mL of the newly prepared Gemfibrozil-piperine nanostructured lipid carrier (GEM-PIP-NLC) sample and dissolve it in 20 mL of distilled water. Use a nanoparticle size analyzer to measure its particle size and Zeta potential. The results are shown in Figure 2 The particle size of GEM-PIP-NLC was 137.4±2.13 nm, its PDI result was 0.155±0.06, and its Zeta potential was -28.6±0.08 mV, indicating that the GEM-PIP-NLC prepared by the optimized formulation had a smaller particle size, more uniform distribution, and good stability.

[0038] The drug release characteristics were investigated by dynamic dialysis. Before the experiment, the cellulose dialysis bag (MW = 3500Da) was activated and then immersed in the release medium [artificial gastric juice (pH 1.2) and artificial intestinal juice (pH 7.8)] overnight.

[0039] Preparation method of artificial gastric juice (pH 1.2): According to the Chinese Pharmacopoeia: take 16.4 mL of 10% dilute hydrochloric acid, add 800 mL of water and 10 g of pepsin (37°C, 3.000-3.500 NFU / mg), shake well, and dilute with water to 1000 mL.

[0040] Preparation method of artificial intestinal fluid (pH 7.8): Take 6.8 g of potassium dihydrogen phosphate, add 500 mL of water to dissolve it, adjust the pH to 7.8 with 0.1 mol / L sodium hydroxide solution, add 10 g of pancreatic enzyme (37°C, 7000 U / mg), and dilute with water to 1000 mL.

[0041] 5 mL of gemfibrozil (GEM) solution (0.36 g / mL), piperine (PIP) solution (0.18 g / mL), and GEM-PIP-NLC solution (0.8 g / mL) were each accurately pipetted into a pre-treated dialysis bag and clamped. The three solutions were then placed in artificial gastric fluid (pH 1.2) and artificial intestinal fluid (pH 6.8), respectively. These solutions were then placed in a thermostatic oscillator at 37 ± 0.5°C and 100 rpm for in vitro release experiments. At the start of the experiment, 5 mL of release medium was pipetted at different time points, and the same volume of release medium was added simultaneously at the same temperature.

[0042] The sampling times for sample release in artificial gastric fluid (pH 1.2) were: 0.5, 1, 2, 4, 6, 8, 10, 12, and 24 h;

[0043] The sampling times for sample release in artificial intestinal fluid (pH 7.8) were: 0.5, 1, 2, 4, 6, 8, 10, 12, 24, 36, 48, and 72 h.

[0044] The samples were filtered through a 0.22 μm microporous membrane, and the contents of the raw drug GEM (Free-GEM) and PIP (Free-PIP) in the release medium, as well as the GEM (GEM-NLC) and PIP (PIP-NLC) in the nanostructured lipid carriers were determined. The cumulative release rates of GEM and PIP were calculated. Each experiment was repeated three times. The in vitro drug release curves were plotted with the cumulative release percentage F% as the ordinate and time t as the abscissa. Figure 3 . In artificial gastric fluid, the cumulative release of GEM and PIP from GEM-PIP-NLC was 5.68% and 7.05% after 24 hours, with almost no drug release, indicating that GEM-PIP-NLC is stable under artificial gastric fluid conditions. In artificial intestinal fluid, the cumulative release of GEM and PIP from GEM-PIP-NLC was 40.45% and 44.08% after 12 hours, with no sudden release. At 24 hours, the cumulative release of the two drugs exceeded 50%, and was almost completely released at 48 hours. At 72 hours, the cumulative release of GEM and PIP reached 72.67% and 76.88%, respectively, indicating that GEM-PIP-NLC has a sustained-release effect.

[0045] Example 3 Pharmacokinetic Study of Gemfibrozil-Piperine Nanostructured Lipid Carrier in Rats

[0046] Example 1: Approximately 104 mL of a GEM-PIP-NLC solution was prepared in a single batch. 1% mannitol was added as an excipient and preservative, and freeze-dried (pre-freezing: -50°C, 2°C / min cooling, 4 h; primary drying: -20°C, 30 Pa, 24 h; secondary drying: 25°C, 5 Pa, 8 h). Approximately 3.5 g of a white, fluffy cake was obtained, which was ground into a lyophilized powder. Prior to administration, the GEM-PIP-NLC lyophilized powder was weighed according to the rat's weight and dissolved in physiological saline for oral administration.

[0047] Twelve healthy male Sprague-Dawley rats were randomly divided into two groups (raw material drug group and formulation group), with 6 rats in each group. Both groups received a single oral gavage. Rats were fasted for 12 hours before dosing but had free access to water. The raw material drug group received a suspension of gemfibrozil (GEM-PIP) (54 mg / kg GEM and 27 mg / kg PIP) in 0.5% sodium carboxymethylcellulose (CMC-Na). The formulation group received a suspension of 1.2 g / kg lyophilized GEM-PIP-NLC (54 mg / kg GEM and 27 mg / kg PIP) dissolved in 2 mL of normal saline. Approximately 0.6 mL of blood was collected from the retroorbital venous ...

[0048] After oral administration of GEM-PIP-NLC and suspension, the drug concentration-time curves in serum were as follows: Figure 4 and Figure 5 The drug-drug curves of GEM-PIP-NLC and GEM-PIP suspension in rats were analyzed using DAS2.0. The results are shown in Tables 1 and 2. The relevant pharmacokinetic parameters were calculated using the non-compartmental model. After oral administration of GEM-PIP API, the plasma concentration increased continuously and reached the maximum plasma concentration C at 2 h. max , respectively 32.58 μg·mL -1 and 1044.60 ng·mL -1 After 2 hours, the blood drug concentration decreased rapidly. After 12 hours, the downward trend became gentle. After that, the blood drug concentration decreased rapidly, and the data was almost undetectable at 24 hours. This shows that the raw material drug is poorly absorbed and metabolized rapidly in the body after oral administration in rats. The elimination half-life of GEM and PIP was extended by 2.12 times and 3.28 times, respectively, significantly increasing the drug circulation time. max The MRTs were 1.82 times and 2.59 times of the raw materials, respectively. The area under the drug-time curve of GEM-PIP-NLC was also significantly improved within 48 hours, with the AUC increased by 3.07 times and 4.78 times, respectively. The results showed that GEM-PIP-NLC can significantly improve the oral bioavailability of GEM and PIP raw materials.

[0049] Table 1 Pharmacokinetic parameters of gemfibrozil and nanoformulation in rats

[0050]

[0051] Table 2 Pharmacokinetic parameters of piperine and nanoformulation in rats

[0052]

[0053]

[0054] Example 4 Pharmacodynamic Study of Gemfibrozil-Piperine Nanostructured Lipid Carrier

[0055] Animal modeling and drug administration

[0056] Preparation of high-fat emulsion: Weigh 40g of homemade lard and heat to melt (temperature 60-70°C), take 20g of cholesterol, 4g of sodium cholate, and 2g of propylthiouracil, grind them into fine powder in a mortar, add them to the melted lard, and stir quickly to dissolve evenly. Add 1mL of Tween-80 and stir evenly to fully emulsify the two. Add water in small amounts and multiple times to make up to 200mL to obtain a stable high-fat emulsion.

[0057] After adaptive feeding for one week, 80 KM mice were randomly divided into eight groups (n=10): normal group (NC), hyperlipidemia model group (MC), positive control group (PC), single-drug API group (GEM), dual-drug API group (GEM-PIP), high-dose dual-drug preparation group (H-GPN), medium-dose dual-drug preparation group (M-GPN), and low-dose dual-drug preparation group (L-GPN).

[0058] Models were established at a fixed time daily at 9:00 AM. The blank group received a gavage of normal saline (0.2 mL / 10 g), while the other groups received an equivalent dose of high-fat emulsion. After 4 weeks, the mice were fasted but not watered. Blood was collected from the NC and MC groups by tail-chopping. Plasma TC and TG levels were measured using an ELASA kit and statistically analyzed using statistical software. Successful model establishment was confirmed if P < 0.05 or P < 0.01 was found between the NC and MC groups.

[0059] Based on the clinical dosage of GEM, the dosage for a 70 kg adult is approximately 600 mg / day. Using the equivalent dose coefficient conversion method, the mouse dosage is approximately 80 mg / kg / day. Drug treatment was initiated after successful model establishment. The GEM group was given GEM suspension at 80 mg / kg / d, and the GEM-PIP group was given an equal amount of GEM and PIP suspension at 80 mg / kg / d (equivalent to the main drugs GEM 53.3 mg / kg / d and PIP 27.7 mg / kg / d). The oral doses of the GEM-PIP-NLC high, medium, and low treatment groups were 760 mg / kg / 2d, 380 mg / kg / 2d, and 190 mg / kg / 2d (equivalent to the main drug GEM 106.6 mg / kg / 2d, 53.3 mg / kg / 2d, and 26.65 mg / kg / 2d, calculated based on drug loading). The PC group was given a powdered simvastatin 0.1% CMC-Na suspension at a dose of (10 mg / kg / d). The NC and MC groups were gavaged with an equal dose of normal saline 0.2 mL / 10 g / d.

[0060] After the last administration, mice in each group were fasted for 48 hours. Blood was collected from the mouse eyeballs and placed in a centrifuge tube. The whole blood was allowed to stand for 30 minutes and centrifuged at 10,000 rpm for 10 minutes. The levels of TC, TG, HDL-C, and LDL-C in mouse plasma were measured using an ELASA kit according to the kit instructions and a microplate reader. The results are shown in the table. Figure 6 The four blood lipid levels of mice in each treatment group were improved. Among them, the combined use of GEM and PIP had a better therapeutic effect on hyperlipidemia than GEM alone, and the therapeutic effect of the preparation group was more significant.

[0061] Weigh 0.5 g of the right lobe of mouse liver, flush the surface blood with 4°C saline, absorb it with filter paper, add saline and prepare a homogenate in an ice bath (mass: volume = 1:9), centrifuge the homogenate at 4000 r / min for 15 min in a low-temperature centrifuge, and aspirate the supernatant to obtain a liver homogenate with a concentration of 10%.

[0062] like Figure 7 The levels of TC, TG, ALT, AST, MDA, and T-SOD in liver tissue homogenates were measured. Compared with the MC group, the TC content in the H-GPN group was significantly reduced by 44.74% (P<0.05). The high and medium dose groups significantly reduced TG levels (P<0.01). Compared with the NC group, the ALT and AST activities in the MC group were significantly higher than those in the NC group (P<0.01), MDA was significantly increased relative to the NC group, and T-SOD was significantly decreased (P<0.01), indicating that the modeling caused liver damage in mice and oxidative damage in vivo. Compared with the MC group, the AST levels in the PC and H-GPN groups were significantly reduced, by 13.32% and 27.47%, respectively, with the H-GPN group showing a greater effect. GEM-PIP-NLC has a certain alleviating effect on liver damage caused by hyperlipidemia. Compared with the MC group, the T-SOD levels in the GEM-PIP group and the H-GPN group increased by 16.22% and 24.25%, respectively, and the treatment effect of the H-GPN group was significant (P<0.01), indicating that GEM-PIP-NLC had a better effect on improving T-SOD and increased catalase activity.

[0063] Example 5 Histopathological results of Gemfibrozil-piperine nanostructured lipid carriers

[0064] Pathological observation of liver tissue: The liver was quickly removed and the liver surface was washed with 4°C physiological saline. A portion of liver tissue at the same position of the left lobe of the liver was obtained at the largest possible cross-section. The liver tissue was fixed in 4% paraformaldehyde solution. The tissue was dehydrated, embedded, and sectioned. HE staining was performed and the morphological changes of the liver pathological sections were observed using an electron microscope. Figure 8 , HE staining of liver sections observed under a 200x microscope. HE staining of liver sections in the NC group showed intact hepatic lobule structure, with round and plump hepatocytes; no obvious abnormalities in the portal tracts between adjacent hepatic lobules; and no obvious inflammatory changes. Compared with the NC group, the MC group showed disordered lobule structure, amorphous and irregularly arranged hepatic epithelial cells, vacuoles in the hepatocytes, and fatty degeneration. Compared with the MC group, all drug intervention groups showed some degree of improvement. Among them, the improvement effect of the dual-drug group was significantly stronger than that of the single-drug group, and the therapeutic effect of H-GPN in the preparation group was more obvious, higher than that of the other treatment groups.

[0065] Pathological observation of epididymal adipose tissue: The epididymal adipose tissue obtained by dissection was stored in paraformaldehyde fixative, embedded and sliced for HE staining, and the prepared tissue slices were observed under a microscope for morphological changes. Figure 9 , a high-fat diet can cause fat accumulation, and white adipose tissue is usually stored in the epididymis, which leads to an increase in tissue cells. The fat cells in the NC group were neatly arranged and dense, with regular morphology and clear boundaries; the fat cells in the MC group were irregular in morphology, with significantly increased cell size, lipid droplets appearing in the fat cells, and some cells swollen or differentiated. The adipose tissues in the PC group, GEM group, GEM-PIP group and preparation group were more neat and uniform compared to the MC group, with gradually reduced cell volume and significantly reduced lipid droplets. The adipocytes in the H-GPN group and M-GPN group were closer to the NC group. It can be seen that the combination of GEM-PIP can improve the body's lipid accumulation and alleviate the enlargement of adipose tissue cells caused by a high-fat diet, but the improvement in the GEM-PIP-NLC group was more obvious. In terms of morphology, the preparation group showed better effects than the PC group.

[0066] Pathological observation of gastrocnemius muscle: The middle part of gastrocnemius muscle was cut into sections, fixed with paraformaldehyde, embedded and sliced, and HE staining was performed to observe the morphological changes of muscle cells under a microscope. Figure 10 As shown, in the sections of the NC group, the gastrocnemius muscle cells were closely arranged and uniform in size, without shrinkage, swelling, or rupture, and the intracellular muscle fibers were regular and without deformation. The cells in the GEM group were significantly different from those in the NC group. The gastrocnemius muscle cells were more rounded in morphology, with wider intercellular spaces and loose and irregular arrangement. A large number of muscle fiber ruptures, myocyte ruptures, sarcoplasm outflow, and inflammatory infiltration were observed inside the muscle cells. After treatment with GEM-PIP-NLC, the muscle fibers in the GEM-PIP-NLC group were intact and without obvious ruptures compared to the injury model group. The cells were more closely arranged than those in the GEM group, and the degree of cell rounding was reduced. This confirms that GEM-PIP-NLC has the effect of alleviating GEM muscle damage.

[0067] The above description of the disclosed embodiments is intended to enable one skilled in the art to implement or use the present invention. Various modifications to these embodiments will be readily apparent to one skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention is not limited to the embodiments shown herein but is intended to conform to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for preparing a gemfibrozil-piperine nanostructured lipid carrier, characterized in that: The following steps are involved: (1) Weigh 0.17 g of gemfibrozil, 0.08 g of piperine, 4.00 g of glyceryl behenate, and 1.00 g of isopropyl myristate, mix well, and heat in a 75°C water bath until completely melted to form the oil phase; (2) Poloxamer 188 and Tween 80 were mixed in a mass ratio of 1:1 to obtain an emulsifier; 2.00 g of the emulsifier was dissolved in distilled water to a concentration of 1.91% as the aqueous phase; (3) The 75°C aqueous phase was added dropwise to the oil phase at 800 r / min and magnetically stirred for 1.52 h to obtain colostrum; ultrasonic dispersion was performed at 350 W power for 10 min, and after cooling, the mixture was filtered through a 0.45 μm filter membrane to obtain the gemfibrozil-piperine nanostructured lipid carrier.

2. The gemfibrozil-piperine nanostructured lipid carrier prepared by the method according to claim 1.

3. Use of the gemfibrozil-piperine nanostructured lipid carrier according to claim 2 in the preparation of lipid-lowering drugs.

4. Use of the gemfibrozil-piperine nanostructured lipid carrier according to claim 2 in the preparation of a drug for enhancing the effect of lipid-lowering.

5. Use of the gemfibrozil-piperine nanostructured lipid carrier according to claim 2 in improving the solubility of poorly soluble drugs.

6. Use of the gemfibrozil-piperine nanostructured lipid carrier according to claim 2 in drug sustained release.

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