Compositions and methods for increasing bioavailable rate of active ingredients

Abstract

A composition comprising an oily medium system and an active ingredient, and a method of increasing the bioavailable rate of an active ingredient, where the method comprises incorporating the active ingredient into an oily medium system, and where the oily medium system contains a vegetable oil and / or a component of a vegetable oil, the liquid does not contain the following substances: a phosphate buffer solution, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxystearic acid hydroxyethyl ester and tocopherol polyethylene glycol succinate; the active ingredient is selected from the group consisting of compounds of formula (I) wherein T1 is a C6 cyclic hydrocarbon and R1 is a C1-C8 aliphatic hydrocarbon group, pharmaceutically acceptable salts thereof, and combinations thereof.

Description

Compositions and methods for increasing the bioavailability of active ingredients Technical Field

[0001] The present invention relates to a method for improving the bioavailability (BA) of an active ingredient, which comprises incorporating the active ingredient into an oily medium system. The present invention also relates to a composition comprising the oily medium system and the active ingredient. In the aforementioned method and composition, the oily medium system contains vegetable oil and / or vegetable oil components, but does not contain the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxystearate hydroxyethyl ester, and tocopheryl polyethylene glycol succinate; in addition, the active ingredient is selected from the following groups: a compound of formula (I), a pharmaceutically acceptable salt thereof, and a combination thereof,

[0002] wherein T1 is a C6 cyclic hydrocarbon, and R1 is a C1-C8 aliphatic hydrocarbon group. Background Art

[0003] Both brain cancer and pancreatic cancer are highly invasive, have high recurrence rates, high mortality rates, and poor prognoses. Brain cancer can occur in all age groups. For common malignant brain tumors such as anaplastic astrocytomas and glioblastoma multiforme (GBM), traditional surgical treatments can remove most tumors but cannot effectively eradicate all cancer cells and must be supplemented with other treatments (such as chemotherapy and radiotherapy). As for pancreatic cancer, because pancreatic cancer cells often migrate, invade, and metastasize to surrounding nerves or vascular tissues, most pancreatic cancer patients are unable to undergo surgical treatment by the time they are discovered and must rely on chemotherapy as the primary treatment.

[0004] However, the current clinical use of chemical drugs for brain cancer or pancreatic cancer is still quite limited, and the therapeutic effects are mostly low. Therefore, it is still necessary to continue to develop drugs or methods that can effectively treat brain cancer or pancreatic cancer.

[0005] Research has shown that butylidenephthalide can not only be used to delay or treat neurological diseases such as amyotrophic lateral sclerosis (ALS), spinocerebellar ataxia, and Alzheimer's disease, but can also be used to treat cerebrovascular diseases such as stroke, improve diabetes, and combat liver and pulmonary fibrosis. It also has excellent anti-cancer effects on brain and pancreatic cancer cells, making it suitable for cancer treatment.

[0006] Despite its aforementioned efficacy, butylidenephthalide still faces limitations in its application. This is primarily due to its reduced bioavailability due to hepatic metabolism and elimination, often requiring higher doses to achieve the desired therapeutic effect. This increases the risk of systemic side effects and burdens patients. This issue, caused by the increased dosage required due to bioavailability, is particularly acute with oral administration. Therefore, effectively increasing the bioavailability of the active ingredient would avoid unnecessary dosage increases, address these limitations, and provide patients with more effective treatment options.

[0007] Summary of the Invention

[0008] The inventors of this application have discovered that the active ingredient of the present invention (i.e., the compound of formula (I) and / or its pharmaceutically acceptable salt) has good solubility in the oily medium system of the present invention and can be stably present in the oily medium system for a long period of time (e.g., up to two years when stored at room temperature). Further research has revealed that when the active ingredient of the present invention is administered in a form incorporated into the oily medium system, its bioavailability can be increased, thereby avoiding the problem of requiring incremental dosage to achieve the desired therapeutic benefit and mitigating or eliminating unwanted side effects.

[0009] Therefore, an object of the present invention is to provide a composition comprising:

[0010] (1) an oily medium system containing vegetable oil and / or vegetable oil components, but not containing the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxyethyl hydroxystearate, and tocopheryl polyethylene glycol succinate; and

[0011] (2) an active ingredient selected from the group consisting of a compound of formula (I), a pharmaceutically acceptable salt thereof, and combinations thereof,

[0012] wherein T1 is a C6 cyclic hydrocarbon, and R1 is a C1-C8 aliphatic hydrocarbon group.

[0013] Preferably, the composition is used to increase the bioavailability of the active ingredient. The composition is preferably a pharmaceutical composition, a food composition, or a food additive composition. When the composition is a pharmaceutical composition, it is administered orally or sublingually; or when the composition is a food composition, it is a health food, dietary supplement, functional food, nutritional supplement, or special nutritional food.

[0014] Another object of the present invention is to provide a method for increasing the bioavailability of an active ingredient, comprising incorporating the active ingredient into an oily medium system, wherein:

[0015] (1) The oily medium system contains vegetable oil and / or vegetable oil components, but does not contain the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxyethyl hydroxystearate, and tocopheryl polyethylene glycol succinate; and

[0016] (2) The active ingredient is selected from the group consisting of: a compound of formula (I), a pharmaceutically acceptable salt thereof, and a combination thereof,

[0017] wherein T1 is a C6 cyclic hydrocarbon, and R1 is a C1-C8 aliphatic hydrocarbon group.

[0018] Preferably, the combined oily medium system and active ingredient are in the form of a pharmaceutical composition, a food composition, or a food additive composition. Preferably, when the combined oily medium system and active ingredient are in the form of a pharmaceutical composition, it is for oral or sublingual administration; when the combined oily medium system and active ingredient are in the form of a food composition, it is a health food, a dietary supplement, a functional food, a nutritional supplement, or a special nutritional food.

[0019] In the composition or method according to the present invention, the vegetable oil is preferably selected from the group consisting of sesame oil, perilla oil, inca inchi oil, brown rice oil, soybean oil, sea buckthorn oil, coconut oil, corn oil, palm oil, peanut oil, grapeseed oil, safflower oil, pumpkin seed oil, almond oil, cashew oil, hazelnut oil, walnut kernel oil, black soybean oil, pine nut oil, black cumin oil, evening primrose oil, sesame leaf essential oil, canola oil, cottonseed oil, olive oil, rapeseed oil, sunflower oil, pecan oil, pistachio oil, castor oil, and combinations thereof. More preferably, the vegetable oil is selected from the group consisting of sesame oil, soybean oil, canola oil, and combinations thereof.

[0020] In the above-mentioned composition or method according to the present invention, the active ingredient is preferably selected from the group consisting of: a compound of formula (II), a pharmaceutically acceptable salt thereof, a compound of formula (III), a pharmaceutically acceptable salt thereof, and combinations thereof.

[0021] Wherein, R1 is a C1-C6 aliphatic hydrocarbon group (preferably a C4 alkyl or alkenyl group).

[0022] In the composition or method according to the present invention, the concentration of the active ingredient is preferably 0.001 mg / mL to 1,500 mg / mL per mL of the oily medium system. In addition, the volume ratio of the oily medium system to the active ingredient is preferably 1:15 to 30:1. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] FIG1 is a photograph of the appearance of a composition according to different embodiments of the present invention after configuration.

[0024] Figure 2 is a photograph of the appearance of a soft capsule made from a composition according to one embodiment of the present invention, Figure 3A is an HPLC analysis of the contents of the soft capsule when it is made, Figure 3B is an HPLC analysis of the contents of the soft capsule after one year of storage, and Figure 3C is an HPLC analysis of ACN (i.e., blank group).

[0025] FIG4 is a photograph of the appearance of a composition according to different embodiments of the present invention after being stored for two years.

[0026] FIG5 is a line graph showing the concentration of the active ingredient in the serum extracted at different time points after the composition according to one embodiment of the present invention was administered to rats.

[0027] FIG6 is a line graph showing the concentration of the active ingredient in serum extracted at different time points after the composition according to one embodiment of the present invention is administered to a human subject.

[0028] FIG7 is a bar graph showing the relative survival rates of the brain cancer cell line 1XM (or pancreatic cancer cell line Mia-PaCa2) after different treatments compared to the blank control group of the brain cancer cell line 1XM (or the blank control group of the pancreatic cancer cell line Mia-PaCa2). “***” indicates that the p-value between the two groups is <0.001.

[0029] FIG8 is a bar graph showing the relative survival rates of the brain cancer cell line 1XM treated with different methods compared to the brain cancer cell line 1XM in the blank group, wherein “***” indicates that the p-value between the two groups is <0.001, “**” indicates that the p-value between the two groups is <0.01, and “*” indicates that the p-value between the two groups is <0.05. DETAILED DESCRIPTION

[0030] The following describes the detailed technical content and some specific implementation plans of the present invention so that those skilled in the art can understand the characteristics of the present invention. However, without departing from the spirit of the present invention, the present invention can be practiced in different forms, and the scope of protection of the present invention should not be interpreted as limited to what is specifically described in the specification.

[0031] Unless otherwise specified herein, the terms "a", "an", "the" and similar terms used in this specification (especially in the scope of the patent application mentioned below) should be understood to include both the singular and the plural forms; the numerical range used (for example, 5 to 100) should be understood to also include all rational numbers within the range and the range composed of any rational numbers within the range, and therefore include all possible combinations of numerical values ​​between the lowest and highest values ​​listed; "mg / kg body weight" refers to the dosage per kilogram of individual body weight; the so-called "individual" refers to mammals, including humans or non-human animals; bioavailability (BA) refers to an indicator of the rate (rate) and extent (extent) of the effective ingredient (also known as active ingredient) of a drug absorbed from a preparation into the systemic blood circulation or the site of action.

[0032] The bioavailability (BA) of a known active ingredient can be obtained by plotting the concentration of the active ingredient in the serum of an organism over time after administration of the active ingredient. In short, the active ingredient is first administered to the organism, and serum samples are continuously collected from the organism at specific time intervals after administration. The concentration of the active ingredient in the sample is then analyzed and a concentration-time curve is plotted. The area under the curve (AUC) is then calculated. The AUC value is positively correlated with the bioavailability. A higher AUC indicates a higher bioavailability of the active ingredient, and conversely, a lower AUC indicates a lower bioavailability of the active ingredient. For the above, see, for example, Drug Bioavailability (National Library of Medicine. Gary Price; Deven A. Patel. July 30, 2023), the entire text of which is incorporated herein by reference.

[0033] While the compounds of formula (I) of the present invention are known to have certain therapeutic benefits in neurological diseases, cerebrovascular diseases, and / or cancer, their application remains limited. This is primarily due to their relatively low bioavailability (AUC < 300 ng × time / ml), often requiring higher doses to achieve the desired therapeutic effect. This increases the risk of systemic side effects and burdens patients. The aforementioned problem of increasing the dosage required due to bioavailability is particularly severe in the case of oral administration. For more information, see, for example, Curr Drug Metab. 2012 Jun 1; 13(5): 524-34 and Drug Metab Dispos. 2008 Feb; 36(2): 400-8, the entire texts of which are incorporated herein by reference.

[0034] To address the aforementioned limitations in application, the inventors of this invention have discovered that the compound of formula (I) and / or its pharmaceutically acceptable salts exhibit excellent solubility in the oily medium system of the present invention and can remain stably in the oily medium system for extended periods of time (e.g., up to two years at room temperature). Further research has revealed that incorporating the active ingredient of the present invention into the oily medium system improves its bioavailability, thereby avoiding the need for incremental dosage to achieve the desired therapeutic benefit and mitigating or eliminating unwanted side effects.

[0035] Therefore, the present invention relates to a composition comprising:

[0036] (1) an oily medium system containing vegetable oil and / or vegetable oil components, but not containing the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxyethyl hydroxystearate, and tocopheryl polyethylene glycol succinate; and

[0037] (2) an active ingredient selected from the group consisting of a compound of formula (I), a pharmaceutically acceptable salt thereof, and combinations thereof,

[0038] wherein T1 is a C6 cyclic hydrocarbon, and R1 is a C1-C8 aliphatic hydrocarbon group.

[0039] In some embodiments according to the present invention, the composition is used to increase the bioavailability of the active ingredient.

[0040] The present invention also relates to a method for increasing the bioavailability of an active ingredient, comprising incorporating the active ingredient into an oily medium system, wherein:

[0041] (1) The oily medium system contains vegetable oil and / or vegetable oil components, but does not contain the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxyethyl hydroxystearate, and tocopheryl polyethylene glycol succinate; and

[0042] (2) The active ingredient is selected from the group consisting of: a compound of formula (I), a pharmaceutically acceptable salt thereof, and a combination thereof,

[0043] wherein T1 is a C6 cyclic hydrocarbon, and R1 is a C1-C8 aliphatic hydrocarbon group.

[0044] In the composition or method according to the present invention, the vegetable oil is preferably selected from the group consisting of: sesame oil, perilla oil, inca inchi oil, brown rice oil, soybean oil, sea buckthorn fruit oil, coconut oil, corn oil, palm oil, peanut oil, grape seed oil, safflower seed oil, pumpkin seed oil, almond oil, cashew oil, hazelnut oil, walnut kernel oil, black soybean oil, pine nut oil, black cumin oil, evening primrose oil, sesame leaf essential oil, canola oil, cottonseed oil, olive oil, rapeseed oil, sunflower oil, pecan oil, pistachio oil, castor oil, and combinations thereof; more preferably selected from the group consisting of: sesame oil, soybean oil, canola oil, and combinations thereof.

[0045] In some embodiments of the composition or method according to the present invention, the oily medium system consists of the vegetable oil.

[0046] In the composition or method according to the present invention, the active ingredient is preferably selected from the group consisting of a compound of formula (II), a pharmaceutically acceptable salt thereof, a compound of formula (III), a pharmaceutically acceptable salt thereof, and combinations thereof.

[0047] Wherein, R1 is a C1-C6 aliphatic hydrocarbon group (preferably a C4 alkyl or alkenyl group).

[0048] In the compositions or methods according to the present invention, the compounds of formula (I) and pharmaceutically acceptable salts thereof may be commercially available, prepared by synthetic methods known in the art, or isolated from natural extracts, but are not limited thereto.

[0049] In the composition or method according to the present invention, the concentration of the active ingredient in the oily medium system is such that no separation occurs after mixing the two. Preferably, the concentration of the active ingredient can be 50 mg / g to 500 mg / g per gram of the oily medium system; and the concentration of the active ingredient can be 0.001 mg / ml to 1,500 mg / ml per milliliter of the oily medium system. In some embodiments of the present invention, the volume ratio of the oily medium system to the active ingredient can be 1:15 to 30:1, for example, 1:13, 1:11.5, 1:10, 1:8, 1:6, 1:4, 1:2, 1:1, 2:1, 3:1, 4:1, 5:1, 8:1, 10:1, 13:1, 15:1, 17:1, 20:1, or 25:1. In some embodiments of the present invention, when the oily medium system contains sesame oil, the compound of formula (I) can still be stably present in the oily medium system when the concentration reaches 920 mg / ml per ml of the volume of the oily medium system.

[0050] The composition of the present invention can be prepared by simply mixing an oily medium system with the active ingredient. If the active ingredient is soluble in the oily medium system, the active ingredient can be directly added to the oily medium system to dissolve therein. If the active ingredient is insoluble in the oily medium system, the active ingredient can be added to the oily medium system and then subjected to a physical force (e.g., oscillation, magnetic stirring) to provide a stable suspension of the active ingredient in the form of micron or nanodroplets.

[0051] The composition provided according to the present invention can be a pharmaceutical composition, a food composition, or a food additive composition. Furthermore, in the methods according to the present invention, the combination of the active ingredient and the oily medium system can be in the form of a pharmaceutical composition, a food composition, or a food additive composition. In some embodiments of the present invention, the combination of the active ingredient and the oily medium system is provided as an orally administered pharmaceutical composition.

[0052] The pharmaceutical composition provided by the present invention can be used for systemic administration or local administration and can be delivered by various drug delivery systems (DDS). For example, but not limited to, the pharmaceutical composition can be delivered by an oral drug delivery system, a transmucosal drug delivery system, a transdermal drug delivery system, and / or an injectable drug delivery system. In addition, the pharmaceutical composition can be administered to an individual in need by one or more of the following modes of administration: oral, sublingual, subcutaneous, intramuscular, intraperitoneal, and intravenous (including drip infusion and rapid injection), nasal administration, transdermal administration, subcutaneous implantation, and interstitial implantation.

[0053] Depending on the form of use and application, any pharmaceutically acceptable excipient may be used to provide the pharmaceutical composition, as long as the excipient does not adversely affect the desired benefits of the compound of formula (I) and / or its pharmaceutically acceptable salt and its composition in the oily medium. The pharmaceutically acceptable excipient may be any known excipient in the art, including, but not limited to, diluents, surfactants, glidants, disintegrants, binders, buffers, colorants, flavorings, antioxidants, preservatives, film-forming agents, and the like.

[0054] The food composition provided according to the present invention can be a health food, a dietary supplement, a functional food, a nutritional supplement, or a special nutritional food. For example, but not limited to, the food composition can be provided in the form of an oil product, such as a capsule containing oil droplets.

[0055] Depending on the form of use and application, the food additive composition provided by the present invention can be in any suitable form, without particular limitation, as long as it is convenient for addition during the food manufacturing process. For example, but not limited to, the food additive composition can be provided in the form of an oily food additive, for example, in the form of an edible essential oil.

[0056] The present invention is further illustrated by the following examples. These examples are provided for illustration only and are not intended to limit the scope of the present invention. The scope of the present invention is as set forth in the claims.

[0057] Example

[0058] In the following examples, the materials and equipment used are as follows:

[0059] 1. Black sesame oil: purchased from Yimei Food Company, product name: Yimei 100% pure black sesame oil.

[0060] 2. White sesame oil: purchased from Seitenbacher Company, product name: Premium Sesame oil.

[0061] 3. Canola oil: purchased from TTET UNION.

[0062] 4. Soybean oil: purchased from TTET UNION.

[0063] 5. 2,6-Dibutyl-p-cresol, also known as 2,6-dibutylhydroxytoluene (Butylated hydroxytoluene, referred to as BHT): purchased from Alfa Aesar.

[0064] 6. n-Butylidenephthalide (BP): purchased from Penta Manufacturing; purity ≥ 97%.

[0065] 7. (Z)-n-butylidenephthalide (Z-BP): provided by Changhong Biotechnology Co., Ltd.; batch number: F212TR12001; purity: 99.8%.

[0066] 8. Ligustilide (LG): purchased from Sigma-Aldrich; purity ≥96%.

[0067] 9. Butylphthalide: purchased from Toronto Research Chemicals; purity ≥ 98%.

[0068] 10.1XM cells (a human glioblastoma cell line): obtained from the research team of Professor Han Hongzhi at Hualien Tzu Chi Hospital (website: hlm.tzuchi.com.tw / ).

[0069] 11. Mia-PaCa2 cells (a human pancreatic cancer cell line): obtained from the Bioresource Collection and Research Center (BCRC; website: www.bcrc.firdi.org.tw / ); accession number: BCRC 60139.

[0070] 12.1XM cell culture medium: DMEM culture medium (purchased from Hyclone) supplemented with 10% fetal bovine serum (FBS; purchased from Hyclone) and 1% penicillin and streptomycin (P / S; purchased from Hyclone).

[0071] 13. Mia-PaCa2 cell culture medium: DMEM / High Glucose culture medium (purchased from Hyclone) supplemented with 10% fetal bovine serum (FBS; purchased from Hyclone), 2.5% horse serum (purchased from Gibco), and 1% penicillin and streptomycin (P / S; purchased from Hyclone).

[0072] 14.MTT assay kit: purchased from Sigma-Aldrich.

[0073] 15. ELISA reader (immune enzyme analyzer): purchased from PerkinElmer (USA).

[0074] Example 1: Preparation of the composition of the present invention

[0075] 1-1. Preparation of composition

[0076] In a glass bottle, the ingredients were mixed in the proportions shown in Table 1 and rapidly stirred with a magnet for 20 minutes to form a homogeneous liquid. Compositions 1 to 4 were prepared. After removing the magnet, the glass bottle was tightly closed, and its appearance was observed and photographed. The results are shown in Figure 1.

[0077] Table 1: Volume percentage of compositions 1 to 4

[0078] As shown in Figure 1 , none of the compositions 1 to 4 showed delamination, indicating good solubility of BP in sesame oil. Furthermore, composition 3 was further prepared into soft capsules, a photograph of which is shown in Figure 2 .

[0079] 1-2. Component Analysis

[0080] The contents of the soft capsules provided in Example 1-1 were analyzed at various times, including upon production and after storage at room temperature in a sunlight-free environment for one year. Samples were prepared with acetonitrile (ACN) to a concentration of 1 mg / mL and analyzed by high-performance liquid chromatography (HPLC) under the conditions shown in Table 2. The results are shown in Figures 3A and 3B, with Figure 3C showing the HPLC analysis of ACN. As shown in Figures 3A and 3C, the soft capsules still contained Z-BP and E-BP even after storage at room temperature for one year, demonstrating the long-term stability of the active ingredients in the oily medium system of the present invention.

[0081] Table 2

[0082] Example 2: Stability of the composition of the present invention

[0083] 2-1. Preparation of composition

[0084] In a glass bottle, the ingredients were mixed in the proportions shown in Table 3 and rapidly stirred with a magnet for 20 minutes to form a homogeneous liquid, thereby preparing compositions 5 to 11. After removing the magnet, the glass bottle was tightly closed.

[0085] Table 3: Weight percentage of compositions 5 to 11

[0086] 2-2. Stability test

[0087] Compositions 5 to 10 provided in Example 2-1 were placed in a sunlight-free environment at room temperature for two years. Their appearances were observed and photographed. The results are shown in FIG4 .

[0088] As shown in Figure 4 , compositions 5 to 10 did not delaminate after being stored at room temperature for up to two years. These results further demonstrate that the compositions of the present invention exhibit excellent stability, maintaining the active ingredients contained therein in the oily medium even after prolonged storage.

[0089] Example 3: Bioavailability Test (Experimental Animals)

[0090] 3-1. Preparation of serum samples

[0091] After fasting for 12 hours, 0.3 ml of blood was drawn from the rat's tail vein and injected into a blood collection tube containing Potassium (K2) EDTA. The blood collection tube was then centrifuged for 10 minutes (2000×g, 2-8°C). The supernatant (i.e., blank group serum) was frozen at -70°C for subsequent analysis.

[0092] Thereafter, the rats were randomly divided into five groups (2 rats in each group) and treated according to the following conditions:

[0093] 1. F10900 group: The composition 8 provided in Example 2-1 was orally administered to rats at a dose of 300 mg (BP) / kg body weight.

[0094] 2. Group F10300: proceed as described for Group F10900, except that composition 8 was replaced by composition 9.

[0095] 3. Group F1900: proceed as described for Group F10900, except that composition 8 was replaced by composition 10.

[0096] 4. Group F1300: The same procedures as those for Group F10900 were followed, except that Composition 8 was replaced by Composition 11.

[0097] 5. Group D2018-2: proceed as described for Group F10900, except that composition 8 was replaced by composition 12.

[0098] At 1, 2, 4, 6, 8, and 24 hours after feeding, 0.3 ml of blood was drawn from the tail vein of each group of rats and injected into a blood collection tube containing potassium ethylenediaminetetraacetate (EDTA K2) and placed on ice. Each tube was then centrifuged for 10 minutes (2000 x g, 2-8°C), and the supernatant was collected. The resulting serum was divided into the F10300 group, the F1900 group, the F1300 group, the F1900 group, and the D2018-2 group. The serum from each group was frozen at -70°C until further analysis.

[0099] 3-2. Preparation of analytical standards

[0100] 50 μL of the standard solution (containing a BP concentration range of 0 to 0.5 mg / mL) was mixed with 150 μL of the blank serum provided in Example 3-1 and ultrasonically shaken for 3 minutes. Then, 400 μL of acetonitrile (ACN) was added, mixed, and ultrasonically shaken for another 3 minutes. The mixture was then placed in an environment below 0°C for 10 minutes, removed, and returned to room temperature for 5 minutes before being filtered through a 0.2 μm PTFE filter. The resulting filtrate was the analytical standard.

[0101] 3-3. Preparation of analytical samples

[0102] 150 μl of the serum from group F10300, group F1900, group F1300, group F1900, or group D2018-2 provided in Example 3-1 was mixed with 450 μl of ACN. The mixture was then ultrasonically shaken for 3 minutes. The mixture was then placed in an environment below 0°C for 10 minutes, removed from the oven, returned to room temperature for 5 minutes, and then filtered through a 0.2 μm PTFE membrane. The resulting filtrate was the analytical sample.

[0103] 3-4.HPLC and pharmacokinetic analysis

[0104] The analytical standards and analytical samples provided in Examples 3-2 and 3-3 were subjected to HPLC analysis under the conditions shown in Table 4 to quantify BP in serum samples at each sampling time point. The results are shown in FIG5 .

[0105] Table 4

[0106] The BP concentration values ​​at each sampling time point were input into Pksolver software and the NCA IV Bolus mode and linear log Trapezoidal method were used to calculate Tmax, Cmax, AUC, T 1 / 2 The results of parameters such as bioavailability (BA) are shown in Table 5.

[0107] Table 5

[0108] As shown in Figure 5 and Table 5, BP was detected in the serum of rats in the F10300, F1300, F10900, and F1900 groups within 24 hours after oral administration of the composition of the present invention. Serum BP levels in the F10300, F1300, F10900, F1900, and D2018-2 groups all reached a peak within 3 hours after oral administration of the composition of the present invention. Furthermore, the AUC and BA of the F10300 group were higher than those of the F10900 group, and the AUC of the F1300 group was higher than that of the F1900 group. These results demonstrate that the oral absorption rate and extent of the composition of the present invention in rats are excellent, and that the bioavailability of the active ingredient in the composition of the present invention increases with increasing the proportion of vegetable oil in the composition.

[0109] The results of this example show that incorporating the compound of formula (I) of the present invention into the oily medium system of the present invention and then administering it to an animal subject can increase the bioavailability of the compound of formula (I).

[0110] Example 4: Bioavailability Test (Human Body)

[0111] 4-1. Preparation of serum samples

[0112] 1. Blank Group: Before subjects (weight: 70 kg) orally administered the soft capsules provided in Example 1-1, 5 ml of blood was drawn and injected into a blood collection tube containing EDTA K2. The tube was then centrifuged for 10 minutes (1900 x g, 2-8°C). The supernatant (i.e., blank group serum) was frozen at -70°C for subsequent analysis.

[0113] 2. "Experimental (Sesame Oil)" Group: 1, 2, 4, 8, and 24 hours after the subjects orally administered the soft capsules provided in Example 1-1 (dosage: 4.285 mg / kg body weight, based on blood pressure), 5 ml of blood was drawn from each sample into EDTA K2-containing blood collection tubes placed on ice. Subsequently, each tube was centrifuged for 10 minutes (1900 x g, 2-8°C), and the supernatant (i.e., serum) was frozen at -70°C for subsequent analysis.

[0114] 4-2. Preparation of analytical standards

[0115] The same procedure was followed as in Example 3-2, except that the blank serum provided in Example 3-1 was replaced with the blank serum provided in Example 4-1 to provide analytical standards.

[0116] 4-3. Preparation of analytical samples

[0117] The same procedure was followed as in Example 3-3, except that the serum of group F10300 / group F1900 / group F1300 / group F1900 was replaced with the serum of group "experimental (sesame oil)" provided in Example 4-1 to provide analysis samples.

[0118] HPLC analysis

[0119] By comparing the method and conditions of Example 3-4, the analytical standards and analytical samples provided in Examples 4-2 and 4-3 were analyzed, and the BP in the serum samples at each sampling time point was quantified. The results are shown in FIG6 .

[0120] As shown in Figure 6 , BP was detectable in the serum of subjects for 24 hours after oral administration of the composition of the present invention, with serum BP levels reaching a peak approximately 2 hours after oral administration. These results demonstrate that the oral absorption rate and extent of the composition of the present invention in humans are also excellent.

[0121] Example 5: Cancer cell toxicity test I

[0122] In this example, the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay was used to investigate the cancer cell cytotoxicity of serum obtained after administration of the composition of the present invention.

[0123] The cell cultures in this example were all carried out in an environment of 95% relative humidity, 5% CO 2 , and 37° C.

[0124] 5-1. Cell Culture

[0125] Brain cancer cell line 1XM (or pancreatic cancer cell line Mia-PaCa2) were divided into groups and cultured in 6-well cell culture plates with the following culture media for 48 hours:

[0126] 1. Blank group: 1XM cell culture medium (or Mia-PaCa2 cell culture medium).

[0127] 2. “BP(50)” group or “BP(100)” group: 1XM cell culture medium (or Mia-PaCa2 cell culture medium) supplemented with BP (50 or 100 μg / ml).

[0128] 3. "Blank serum (low)" group, "Blank serum (medium)" group, and "Blank serum (high)" group: 1XM cell culture medium (or Mia-PaCa2 cell culture medium) supplemented with the blank serum provided in Example 4-1 (at a content of 6.25 volume%, 12.5 volume%, or 25 volume% in the culture medium).

[0129] 4. "Experimental serum (low)" group, "Experimental serum (medium)" group, and "Experimental serum (high)" group: 1XM cell culture medium (or Mia-PaCa2 cell culture medium) supplemented with serum from the experimental group taken 1, 2, 4, 8, or 24 hours after oral administration of the soft capsule of Example 4-1 (the content in the culture medium was 6.25 volume%, 12.5 volume%, or 25 volume%).

[0130] 5-2. MTT assay

[0131] Remove the culture medium from each well of the cell culture plate and add MTT reagent (so that the final concentration of MTT in the culture medium of each well is 0.5 mg / ml). Then place the cell culture plate in an incubator at 37°C and 5% CO2 for 2 to 4 hours to form a (formazan) purple needle crystals. Afterwards, the solution was aspirated and 500 μl of dimethyl sulfoxide (DMSO) was added to each well. Shake on an oscillator for 5 to 10 minutes to allow the DMSO to After the purple needle-shaped crystals completely dissolved, the absorbance at 570 or 595 nm was measured using an ELISA reader. Cell survival rates were calculated based on this absorbance. The relative survival rates of the remaining groups were calculated using the blank control group as a benchmark (i.e., the blank control group's survival rate was set at 100%). The results are shown in Figures 7 and 8. Figure 7 shows the data for serum from the experimental group collected 2 hours after oral softgel administration, while Figure 8 shows the data for serum from the experimental group collected 1, 4, 8, or 24 hours after oral softgel administration.

[0132] As shown in Figures 7 and 8, compared with the blank group, the cancer cell survival rates of the "experimental serum (low)" group, the "experimental serum (medium)" group, and the "experimental serum (high)" group were all lower, which were comparable to the "BP (50)" group or the "BP (100)" group; on the other hand, the degree of reduction in cancer cell survival rate was "experimental serum (low)" group < "experimental serum (medium)" group < "experimental serum (high)" group.

[0133] These results demonstrate that serum obtained after oral administration of soft capsules containing the composition of the present invention is toxic to brain and pancreatic cancer cells. This effect increases with increasing concentrations of the active ingredient in the serum and is comparable to direct treatment of cancer cells with the active ingredient. This also demonstrates that the active ingredient contained in the composition of the present invention has excellent bioavailability.

[0134] Example 6: Cancer cell toxicity test II

[0135] The cell cultures in this example were all carried out in an environment of 95% relative humidity, 5% CO 2 , and 37° C.

[0136] 6-1. Cell Culture

[0137] Pancreatic cancer cell line Mia-PaCa2 was divided into groups and cultured in 96-well cell culture plates with the following culture media for 24 hours:

[0138] 1. “Z-BP” group: Mia-PaCa2 cell culture medium supplemented with Z-BP.

[0139] 2. “Z-BP + sesame oil” group: “Z-BP + canola oil” group, “Z-BP + soybean oil” group: Mia-PaCa2 cell culture medium supplemented with a mixture of Z-BP and sesame oil, canola oil, or soybean oil in equal proportions.

[0140] 3. “LG+sesame oil” group: “LG+canola oil” group, “LG+soybean oil” group: Mia-PaCa2 cell culture medium supplemented with a mixture of LG and sesame oil, canola oil, or soybean oil in equal proportions.

[0141] MTT assay

[0142] MTT analysis was performed in the same manner as in Example 5-2, and the concentrations (IC 50 The results are shown in Table 6.

[0143] Table 6

[0144] The results in Table 6 show that incorporating the compound of formula (I) of the present invention into the oily medium system of the present invention improves the cancer cell-killing effect of the compound of formula (I).

[0145] The results of the above examples demonstrate that the active ingredient of the present invention (i.e., the compound of formula (I) and / or its pharmaceutically acceptable salt) exhibits good solubility in the oily medium system of the present invention and can be stably present in the oily medium system for an extended period of time. Furthermore, when the active ingredient of the present invention is administered in an oily medium system, its bioavailability can be increased, thereby avoiding the problem of requiring incremental dosage to achieve the desired therapeutic benefit and mitigating or eliminating unwanted side effects.

Claims

1. A composition, characterized in that It contains: (1) an oil medium system containing vegetable oil and / or vegetable oil components, but not containing the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxystearate hydroxyethyl ester, and tocopherol polyethylene glycol succinate; and (2) an active ingredient selected from the group consisting of a compound of formula (I), a pharmaceutically acceptable salt thereof, and a combination thereof, in, T1 is a C6 cyclic hydrocarbon; and R1 is a C1-C8 aliphatic hydrocarbon group.

2. The composition according to claim 1, characterized in that The vegetable oil is selected from the group consisting of: sesame oil, perilla oil, inca inchi oil, brown rice oil, soybean oil, sea buckthorn fruit oil, coconut oil, corn oil, palm oil, peanut oil, grape seed oil, safflower seed oil, pumpkin seed oil, almond oil, cashew oil, hazelnut oil, walnut kernel oil, black soybean oil, pine nut oil, black cumin oil, evening primrose oil, sesame leaf essential oil, canola oil, cottonseed oil, olive oil, rapeseed oil, sunflower oil, pecan oil, pistachio oil, castor oil, and combinations thereof.

3. The composition according to claim 1, characterized in that The vegetable oil is selected from the group consisting of sesame oil, soybean oil, canola oil, and combinations thereof.

4. The composition according to claim 1, characterized in that The active ingredient is selected from the group consisting of: a compound of formula (II), a pharmaceutically acceptable salt thereof, a compound of formula (III), a pharmaceutically acceptable salt thereof, and a combination thereof, Wherein, R1 is a C1-C6 aliphatic hydrocarbon group.

5. The composition according to claim 4, characterized in that R1 is a C4 alkyl or alkenyl group.

6. The composition according to any one of claims 1 to 5, characterized in that The concentration of the active ingredient is 0.001 mg / ml to 1,500 mg / ml per ml of the oil medium system.

7. The composition according to any one of claims 1 to 5, characterized in that The volume ratio of the oily medium system to the active ingredient is 1:15 to 30:

1.

8. The composition according to any one of claims 1 to 5, characterized in that It serves to increase the bioavailability of the active ingredient.

9. The composition according to claim 8, characterized in that It is a pharmaceutical composition, a food composition, or a food additive composition.

10. The composition according to claim 9, characterized in that It is a pharmaceutical composition in the form of oral or sublingual administration.

11. The composition according to claim 9, characterized in that The invention is a food composition, which is a health food, a dietary supplement, a functional food, a nutritional supplement food or a special nutritional food.

12. A method for increasing the bioavailability of an active ingredient, characterized in that: It comprises incorporating the active ingredient into an oily medium system, wherein: (1) The oil medium system contains vegetable oil and / or vegetable oil components, but does not contain the following substances: phosphate buffer, polyethylene glycol, dimethyl sulfoxide, ethanol, polypropylene glycol, polysorbate, ethyl acetate, 12-hydroxystearate hydroxyethyl ester, and tocopherol polyethylene glycol succinate; and (2) the active ingredient is selected from the group consisting of a compound of formula (I), a pharmaceutically acceptable salt thereof, and a combination thereof, in, T1 is a C6 cyclic hydrocarbon; and R1 is a C1-C8 aliphatic hydrocarbon group.

13. The method according to claim 12, characterized in that The vegetable oil is selected from the group consisting of: sesame oil, perilla oil, inca inchi oil, brown rice oil, soybean oil, sea buckthorn fruit oil, coconut oil, corn oil, palm oil, peanut oil, grape seed oil, safflower seed oil, pumpkin seed oil, almond oil, cashew oil, hazelnut oil, walnut kernel oil, black soybean oil, pine nut oil, black cumin oil, evening primrose oil, sesame leaf essential oil, canola oil, cottonseed oil, olive oil, rapeseed oil, sunflower oil, pecan oil, pistachio oil, castor oil, and combinations thereof.

14. The method of claim 12, wherein: The vegetable oil is selected from the group consisting of sesame oil, soybean oil, canola oil, and combinations thereof.

15. The method of claim 12, wherein: The active ingredient is selected from the group consisting of: a compound of formula (II), a pharmaceutically acceptable salt thereof, a compound of formula (III), a pharmaceutically acceptable salt thereof, and a combination thereof, Wherein, R1 is a C1-C6 aliphatic hydrocarbon group.

16. The method of claim 15, wherein: R1 is a C4 alkyl or alkenyl group.

17. The method according to any one of claims 12 to 16, characterized in that The concentration of the active ingredient is 0.001 mg / ml to 1,500 mg / ml per ml of the oil medium system.

18. The method according to any one of claims 12 to 16, characterized in that The volume ratio of the oily medium system to the active ingredient is 1:15 to 30:

1.

19. The method according to any one of claims 12 to 16, characterized in that The oily medium system and the active ingredient are in the form of a pharmaceutical composition, a food composition, or a food additive composition.

20. The method of claim 19, wherein: When the combined oily medium system and the active ingredient are in the form of a pharmaceutical composition, it is used for oral or sublingual administration.

21. The method of claim 19, wherein: When the combined oily medium system and the active ingredient are in the form of a food composition, the food composition is a health food, a dietary supplement, a functional food, a nutritional supplement food or a special nutritional food.

Images