Hydrazone-containing small molecule gelator, its synthesis method and application in preparing ice flake assembly

Abstract

The application discloses a small-molecule gelling agent containing a hydrazone group, a synthesis method of the small-molecule gelling agent and application of the small-molecule gelling agent in preparation of borneol assembly. The small-molecule gelling agent is prepared through Williamson etherification reaction of a hydroxybenzaldehyde compound and a bromoalkane and then condensation reaction with a hydrazine compound, and the structure of the small-molecule gelling agent contains functional groups such as a hydrazone group, an alkyl chain and a phenolic hydroxyl group, and the small-molecule gelling agent can self-assemble into a three-dimensional network gel structure through non-covalent interactions such as hydrogen bonds and pi-pi stacking. The gelling agent can be used for clathration of volatile drugs borneol, and preparation of borneol-small-molecule gelling agent xerogel, thereby significantly reducing the volatile loss of borneol, and improving the thermal stability, light stability and wet stability of the borneol. The assembly has lower toxicity than free borneol. When the assembly is applied to compound Danshen tablets, the pharmaceutical stability of borneol can be effectively improved, and the anti-myocardial ischemia efficacy of the preparation can be ensured. The application provides a novel, efficient and low-toxicity solution for the stabilization of volatile traditional Chinese medicine components.

Description

Technical Field

[0001] This invention belongs to the field of small molecule gelling agent technology, specifically relating to a class of hydrazone-containing small molecule gelling agents and their synthesis methods, as well as the application of the small molecule gelling agent as a carrier in the preparation of assemblies for stabilizing borneol. Background Technology

[0002] Borneol, a volatile component of traditional Chinese medicine, is chemically known as 2-borneol. It is used to clear the mind, relieve pain, and has the effects of invigorating the spirit and clearing heat. It is widely used in over 170 kinds of traditional Chinese medicine preparations, including Compound Danshen Tablets. However, borneol is prone to sublimation at room temperature and is unstable during preparation and storage, leading to a decrease in its content and affecting the efficacy of the drug. According to literature reports, the actual borneol content in Compound Danshen Tablets is only 1.88% to 33.75% of the theoretical dosage in the pharmacopoeia, which is closely related to the volatility of borneol itself.

[0003] Currently, methods to improve the stability of borneol mainly include β-cyclodextrin inclusion technology, microencapsulation technology, and solid dispersion technology. However, these methods have problems such as large excipient usage, long preparation time, and drug burst release. For example, when using β-cyclodextrin to encapsulate borneol, a 1:6 feed ratio is required, resulting in a large amount of excipients; when using hollow mesoporous silica spheres as a carrier, the preparation time is long, which is not conducive to industrial production; although physical adsorption is simple, it is prone to causing initial burst release of the drug, making it difficult to achieve sustained release or targeted delivery.

[0004] Small molecule organic gelling agents can self-assemble into three-dimensional network structures through non-covalent interactions such as hydrogen bonding, π-π stacking, and van der Waals forces, immobilizing solvent molecules or drugs within them, providing a new approach for the stabilization of volatile drugs. Hydrazones, with their simple synthesis, tunable structure, and ease of hydrogen bonding, are ideal structural units for constructing small molecule gelling agents. Therefore, developing a novel small molecule gelling agent that is simple to prepare, low in cost, has strong gelling ability, effectively stabilizes borneol, and exhibits good biocompatibility is of significant practical importance. Summary of the Invention

[0005] The present invention aims to provide a class of small molecule gelling agents containing hydrazone groups. This compound has good gelling ability and can self-assemble with borneol to form stable assemblies, thereby significantly reducing the volatility of borneol and improving its pharmaceutical stability.

[0006] The present invention also provides a method for synthesizing the hydrazone-containing small molecule gelling agent and its application in the preparation of borneol assemblies.

[0007] The hydrazone-containing small molecule gelling agent of the present invention has the following structural formula:

[0008]

[0009] Where R1 represents C4~C 18 A straight-chain alkyl group; R2 represents hydrogen or hydroxyl; R3 is selected from hydrogen, , , , , , , , Any one of them.

[0010] Furthermore, R3 is selected from... , , , , , Any one of them.

[0011] The synthesis method of the above-mentioned hydrazone-containing small molecule gelling agent includes the following steps:

[0012] Step 1: React the p-hydroxybenzaldehyde compound shown in Formula I with the bromoalkane shown in Formula II under alkaline conditions to generate the alkoxybenzaldehyde intermediate shown in Formula III.

[0013] Step 2: The alkoxybenzaldehyde intermediate obtained in Step 1 is reacted with the hydrazine compound shown in Formula IV in a high-boiling-point polar solvent by heating. After cooling, washing, and pulping, a small molecule gelling agent containing a hydrazone group is obtained. The synthetic route is as follows:

[0014]

[0015] Furthermore, the alkaline conditions described in step 1 are provided by anhydrous potassium carbonate; the reaction temperature is 50–70°C, and the reaction time is 5–24 h.

[0016] Furthermore, the high-boiling-point polar solvent in step 2 is diethylene glycol; the reaction temperature is 120–135°C.

[0017] This invention also provides a small molecule gelling agent-borneol assembly, which is obtained by self-assembling the above-mentioned hydrazone-containing small molecule gelling agent and borneol in an organic solvent to form a gel, followed by freeze-drying. The organic solvent is any one of petroleum ether, cyclohexane, isopropanol, n-butanol, acetonitrile, and DMSO.

[0018] Further, the preparation method of the assembly is as follows: the hydrazone-containing small molecule gelling agent and borneol are added to an organic solvent at a mass ratio of 1:2 to 4, heated to 60 to 90°C under sealed conditions until completely dissolved, and then allowed to stand at 0 to 5°C for 10 to 30 minutes to form a gel. The mixture is then pre-frozen at -25 to -15°C for 4 to 6 hours, and subsequently freeze-dried under vacuum at -80 to -60°C for 20 to 24 hours to obtain the small molecule gelling agent-borneol assembly.

[0019] Preferably, the mass of the hydrazone-containing small molecule gelling agent added to each milliliter of organic solvent is 50-70 mg.

[0020] Preferably, the organic solvent is petroleum ether or ethanol.

[0021] The present invention further provides the application of the above-mentioned hydrazone-containing small molecule gelling agent or the above-mentioned small molecule gelling agent-borneol assembly in the preparation of pharmaceutical formulations that improve the stability of borneol and reduce the toxicity of borneol.

[0022] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0023] 1. This invention synthesizes a novel class of hydrazone-containing small molecule gelling agents that exhibit excellent gelling ability in a variety of organic solvents, can rapidly form stable gels, and have a high gel-sol phase transition temperature. Through non-covalent interactions such as hydrogen bonding and π-π stacking, they self-assemble into a three-dimensional network structure, laying the foundation for their use as drug carriers. The synthesis route is simple, the conditions are mild, and the yield is high, which is beneficial for industrial production. The purity is all above 98%, meeting the requirements for pharmaceutical excipients.

[0024] 2. This invention prepares a borneol-small molecule gelling agent dry gel by co-assembling a small molecule gelling agent with borneol. This assembly forms a three-dimensional network structure through multiple interactions, including hydrogen bonding, significantly reducing the volatility of borneol and improving its tolerance to environmental factors such as high temperature, light, and high humidity. Acute toxicity experiments in zebrafish and organ pathological observations confirm that the small molecule gelling agent of this invention has good safety within the effective concentration range. After forming an assembly with borneol, its toxicity is lower than that of the same dose of free borneol, and it does not cause significant damage to the heart or liver, demonstrating good biocompatibility. In vivo pharmacokinetic studies in rats show that this assembly has the characteristic of delaying borneol release, which is expected to improve the pharmacokinetic behavior of borneol in vivo and achieve a long-lasting effect. When the assembly is applied to compound Danshen tablets, it shows efficacy comparable to ordinary compound Danshen tablets in rats with acute myocardial ischemia, indicating that the introduction of the gelling agent does not interfere with the original therapeutic effect of the formulation and has good application prospects. Attached Figure Description

[0025] Figure 1 These are scanning electron microscope (SEM) images of the dry gels, where A is borneol, B is D2-3 dry gel, and C, D, and E are dry gels with borneol-D2-3 gelling agent mass ratios of 2:1, 3:1, and 4:1, respectively.

[0026] Figure 2 This is a differential scanning calorimetry (DSC) analysis chromatogram of the dry gel.

[0027] Figure 3This is a comparison of the Fourier Transform Infrared (FT-IR) spectra of borneol and dry gel.

[0028] Figure 4 This is a comparison chart of the loss rates of borneol and borneol-D2-3 gelling agent dry gel at 50℃.

[0029] Figure 5 This is a graph showing the mortality rate of zebrafish at different concentrations in each group.

[0030] Figure 6 This is a comparison chart of the cumulative mortality of zebrafish within 72 hours between borneol and borneol-D2-3 gelling agent dry gel. The left chart is the borneol group, and the right chart is the assembly group.

[0031] Figure 7 These are pathological morphological changes in the myocardial tissue of each group of zebrafish (200x).

[0032] Figure 8 These are pathological morphological changes in the liver tissue of zebrafish from different groups (400x).

[0033] Figure 9 This is a comparison chart of the infarct area in a rat model of acute myocardial ischemia.

[0034] Figure 10 This is a diagram showing the pathological morphological changes in rat myocardial tissue. Detailed Implementation

[0035] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments, but the scope of protection of the present invention is not limited to these embodiments.

[0036] Example 1

[0037] Step 1: Under nitrogen protection, p-hydroxybenzaldehyde (10.00 g, 81.89 mmol), 1-bromohexadecane (26.23 g, 85.98 mmol), sodium iodide (2.45 g, 16.35 mmol), anhydrous potassium carbonate (11.31 g, 81.89 mmol), and 80 mL of acetone were placed in a three-necked flask equipped with a stirrer and a condenser. The mixture was refluxed at 60 °C for 24 h. The reaction was confirmed to be complete by TLC, and the reaction was quenched with water. The mixture was extracted three times with ethyl acetate, and the organic phases were combined and separated by silica gel column chromatography. The eluent was petroleum ether:ethyl acetate (v / v) = 16:1 to give compound C-1, a white solid with a yield of 62% and a melting point of 52–53 °C.

[0038] The reaction was carried out with equimolar amounts of 2,4-dihydroxybenzaldehyde instead of p-hydroxybenzaldehyde and 1-bromohexadecane, under otherwise unchanged conditions, to give compound C-2 (yield 29%).

[0039]

[0040] C-1 C-2

[0041] The structural characterization data of compound C-1 are as follows: EI-MS (m / z): 275.070 [M-C5H 11 ] + ; 1 H NMR (400MHz, Chloroform-d) δ 9.87 (s, 1H,), 7.90 – 7.66 (m, 2H,), 7.07 – 6.85 (m,2H,), 4.03 (t, J = 6.5 Hz, 2H), 1.80 (dt, J = 14.6, 6.6 Hz, 2H), 1.50 – 1.38 (m, 2H), 1.25 (s, 24H), 0.89 – 0.84 (m, 3H); 13 C NMR (101 MHz, CDCl3) δ191.04, 190.87, 164.37, 132.12, 132.07, 129.79, 115.04, 114.92, 114.71,114.59, 77.46, 77.14, 76.83, 68.51, 32.03, 29.80, 29.71, 29.47, 29.15, 26.06,22.81, 14.24; IR (KBr), ν, cm -1 : 2920, 2848, 1695, 1600, 1469, 834.

[0042] The structural characterization data of compound C-2 are as follows: 1 H NMR (400 MHz, Chloroform-d) δ 11.47 (s,1H), 9.70 (s, 1H), 7.41 (d, J = 8.7 Hz, 1H), 6.52 (dd, J = 8.7, 2.3 Hz, 1H), 6.41 (d, J = 2.3 Hz, 1H), 4.00 (t, J = 6.6 Hz, 2H), 1.79 (dt, J = 14.6, 6.7Hz, 2H), 1.44 (p, J = 6.8 Hz, 2H), 1.26 (s, 24H), 0.90 – 0.85 (m, 3H); 13C NMR(101 MHz, CDCl3) δ 194.38, 166.58, 164.63, 135.28, 115.11, 108.88, 101.16,68.70, 32.00, 29.77, 29.74, 29.65, 29.60, 29.44, 29.38, 29.00, 25.99, 22.77,14.19; IR (KBr), ν, cm -1 : 3168, 2918, 2850, 1672, 1627, 1210, 827.

[0043] Step 2: Compound C-1 (5 g, 14.45 mmol), 4-hydroxybenzoyl hydrazide (6.59 g, 43.35 mmol), and 60 mL of diethylene glycol were added to a reaction flask and reacted at 135 °C for 3 h. The reaction was confirmed to be complete by TLC. The mixture was washed with water, methanol, and petroleum ether in sequence, and then slurried to obtain compound D2-3, a white solid with a yield of 47% and a liquid phase purity of 99%.

[0044] By replacing the above-mentioned 4-hydroxybenzoyl hydrazide with equimolar amounts of benzoyl hydrazide, 2-naphthyl hydrazide hydrochloride, phenylhydrazide, and aminothiourea, respectively, and under the same conditions as the synthesis of D2-3, compounds D2-1 (white solid, yield 42%), D2-2 (white solid, yield 55%), D2-4 (pale yellow solid, yield 53%), and D2-5 (brownish-yellow solid, yield 35%) were obtained.

[0045] By replacing compound C-1 with equimolar compound C-2 and equimolar benzoyl hydrazide with 4-hydroxybenzoyl hydrazide, and under the same conditions as the synthesis of D2-3, compound D2-6 (white solid, yield 23%) was obtained.

[0046] By replacing compound C-1 with equimolar compound C-2 and equimolar phenylhydrazine with 4-hydroxybenzoylhydrazine, and under the same conditions as the synthesis of D2-3, compound D2-7 (yellow solid, yield 16%) was obtained.

[0047] By replacing the above-mentioned 4-hydroxybenzoylhydrazide with an equimolar allyl benzoylhydrazide, and under the same conditions as the synthesis of D2-3, compound D2-8 (white solid, yield 32%) was obtained.

[0048] The above-mentioned 4-hydroxybenzoyl hydrazine was replaced with an equimolar amount of 4-hydroxy-3-methoxybenzoyl hydrazine (prepared by reacting methyl 4-hydroxy-3-methoxybenzoate with hydrazine hydrate), and the other conditions were the same as those for the synthesis of D2-3, to obtain compound D2-9 (white solid, yield 32%).

[0049]

[0050] D2-1 D2-2

[0051]

[0052] D2-3 D2-4

[0053]

[0054] D2-5 D2-6

[0055]

[0056] D2-7 D2-8

[0057]

[0058] D2-9

[0059] The structures of the above D2 series compounds were determined by 1 H NMR, 13 C NMR and FT-IR confirmed that the structure was consistent with the target structure.

[0060] The structural characterization data of compound D2-1 are as follows: 1 H NMR (600 MHz, Chloroform-d) δ 9.10 (s,1H), 7.74 (s, 1H), 7.61 – 7.55 (m, 2H), 7.18 (s, 1H), 6.91 (dd, J = 8.6, 3.3Hz, 2H), 6.25 (s, 1H), 3.99 (q, J = 5.7 Hz, 2H), 1.83 – 1.76 (m, 2H), 1.45 (d, J = 9.9 Hz, 2H), 1.39 – 1.22 (m, 24H), 0.88 (t, J = 8.6 Hz, 3H). 13 C NMR(151 MHz, CDCl3) δ 178.40, 161.50, 143.67, 129.12, 125.12, 114.91, 68.24,31.93, 29.70, 29.68, 29.66, 29.59, 29.56, 29.37, 29.14, 25.99, 22.70, 14.13.IR (KBr), ν, cm -1: 3408, 3254, 3152, 2925, 2851, 1600, 1531, 1472, 1297, 1241,833.

[0061] The structural characterization data of compound D2-2 are as follows: 1 H NMR (600 MHz, Chloroform-d) δ 9.11 (s,1H), 8.23 ​​(s, 1H), 7.85 (s, 2H), 7.71 (s, 2H), 7.54 (t, J = 5.7 Hz, 1H), 7.50– 7.44 (m, 2H), 6.91 (t, J = 5.7 Hz, 2H), 3.98 (q, J = 5.8 Hz, 2H), 1.83 –1.75 (m, 2H), 1.45 (s, 2H), 1.36 – 1.22 (m, 24H), 0.88 (t, J = 8.8 Hz, 3H). 13 C NMR (151 MHz, CDCl3) δ 159.65, 147.40, 130.97, 128.40, 127.80, 126.12,124.83, 113.69, 67.15, 30.91, 28.67, 28.66, 28.64, 28.58, 28.55, 28.36,28.34, 28.16, 24.99, 21.67, 13.10. IR (KBr), ν, cm -1 : 3250, 2920, 2853, 1653,1608, 1509, 1255.

[0062] The structural characterization data of compound D2-3 are as follows: 1 H NMR (600 MHz, DMSO-d6) δ 11.49 (s, 1H),10.08 (s, 1H), 8.36 (s, 1H), 7.79 (d, J = 6.9 Hz, 2H), 7.63 (d, J = 8.2 Hz,2H), 6.99 (d, J = 8.2 Hz, 2H), 6.85 (d, J = 8.2 Hz, 2H), 4.00 (q, J = 5.6 Hz,2H), 1.76 – 1.68 (m, 2H), 1.41 (t, J = 7.5 Hz, 2H), 1.34 – 1.17 (m, 24H),0.85 (t, J = 8.2 Hz, 3H).13 C NMR (151 MHz, DMSO) δ 163.02, 161.10, 160.61,147.21, 130.04, 128.97, 127.43, 124.54, 115.44, 115.21, 68.07, 31.77, 29.52,29.51, 29.48, 29.44, 29.22, 29.18, 29.09, 25.94, 22.56, 14.41. IR (KBr), ν,cm -1 : 3382, 3239, 2922, 2855, 1650, 1609, 1509, 1301, 1252, 837.

[0063] The structural characterization data of compound D2-4 are as follows: 1 H NMR (400 MHz, Chloroform-d) δ 7.65 (s,1H), 7.61 – 7.56 (m, 2H), 7.30 – 7.23 (m, 2H), 7.10 (d, J = 8.0 Hz, 2H), 6.92– 6.87 (m, 2H), 6.85 (t, J = 7.3 Hz, 1H), 3.98 (t, J = 6.6 Hz, 2H), 1.85 –1.74 (m, 2H), 1.46 (dd, J = 23.2, 6.0 Hz, 2H), 1.27 (s, 24H), 0.93 – 0.85 (m,3H). 13 C NMR (151 MHz, CDCl3) δ 159.65, 145.00, 137.57, 129.26, 127.56,119.77, 114.68, 112.66, 68.12, 31.93, 29.70, 29.67, 29.61, 29.58, 29.41,29.37, 29.26, 29.12, 29.06, 26.05, 22.70, 14.12. IR (KBr), ν, cm -1 : 3297.32,2920, 2854, 1601, 1503, 1254, 1118, 827.

[0064] The structural characterization data of compound D2-5 are as follows: 1H NMR (600 MHz, Chloroform-d) δ 7.75 (s,1H), 7.73 (s, 3H), 7.64 (t, J = 6.4 Hz, 2H), 7.46 (d, J = 4.4 Hz, 1H), 7.40(d, J = 6.5 Hz, 1H), 7.32 – 7.27 (m, 2H), 6.93 (t, J = 6.7 Hz, 2H), 3.99 (q,J = 6.1 Hz, 2H), 1.80 (q, J = 6.8 Hz, 2H), 1.47 (d, J = 7.2 Hz, 2H), 1.38 –1.23 (m, 24H), 0.88 (q, J = 6.4 Hz, 3H). 13 C NMR (151 MHz, CDCl3) δ 159.80,142.58, 138.11, 134.86, 129.16, 128.87, 127.76, 127.70, 126.49, 126.46,122.80, 115.46, 114.74, 106.69, 68.15, 31.94, 29.71, 29.67, 29.61, 29.59,29.42, 29.37, 29.27, 26.06, 22.70, 14.12. IR (KBr), ν, cm -1 : 3288, 2918, 2852,1601, 1504, 1469, 1293, 1254, 842.

[0065] The structural characterization data of compound D2-6 are as follows: 1 H NMR (600 MHz, Chloroform-d) δ 9.59 (s,1H), 8.50 (s, 1H), 7.87 (s, 2H), 7.55 (d, J = 7.4 Hz, 1H), 7.48 (d, J = 7.6Hz, 2H), 7.08 (d, J = 8.0 Hz, 1H), 6.55 – 6.40 (m, 2H), 3.92 (t, J = 7.1 Hz,2H), 1.75 (t, J = 7.4 Hz, 2H), 1.43 (q, J = 7.6 Hz, 2H), 1.39 – 1.20 (m,24H), 0.88 (t, J = 7.3 Hz, 3H). 13C NMR (151 MHz, CDCl3) δ 164.55, 160.54,159.83, 152.14, 135.21, 132.22, 128.90, 127.19, 110.48, 107.69, 101.94,77.23, 77.02, IR (KBr), ν,cm -1 : 3430,3269, 2921, 2855, 1639, 1522,1295, 1161, 701.

[0066] The structural characterization data of compound D2-7 are as follows: 1 H NMR (600 MHz, Chloroform-d) δ 11.07 (s,1H), 7.81 (s, 1H), 7.43 (d, J = 5.7 Hz, 2H), 7.38 (d, J = 6.7 Hz, 2H), 7.32(d, J = 7.0 Hz, 1H), 7.02 (t, J = 6.6 Hz, 1H), 6.93 (d, J = 5.6 Hz, 4H), 6.51(s, 1H), 6.45 (t, J = 6.7 Hz, 1H), 5.03 (d, J = 4.7 Hz, 2H), 3.96 (t, J = 6.4Hz, 2H), 1.78 (p, J = 6.6 Hz, 2H), 1.45 (q, J = 7.2 Hz, 2H), 1.38 – 1.22 (m, 24H), 0.88 (t, J = 6.7 Hz, 3H). 13C NMR (151 MHz, CDCl3) δ 161.25, 158.63,145.86, 141.44, 137.28, 130.20, 128.56, 127.91, 127.51, 116.15, 114.03,106.97, 101.94, 99.72, 70.71, 68.16, 31.94, 29.71, 29.69, 29.67, 29.61,29.58, 29.39, 29.37, 29.18, 26.03, 22.70, 14.12.IR (KBr), ν, cm -1 : 3441,3318, 2920, 2850, 1634, 1522, 1467, 1291, 1244, 807.

[0067] The structural characterization data of compound D2-8 are as follows: 1 H NMR (400 MHz, Chloroform-d) δ 9.19 (s,1H), 8.20 (s, 1H), 7.83 (s, 1H), 7.67 (s, 1H), 6.99 – 6.84 (m, 4H), 6.04 (t,J = 11.8 Hz, 1H), 5.43 (d, J = 17.6 Hz, 1H), 5.32 (dt, J = 10.4, 1.5 Hz, 1H), 4.58 (s, 2H), 3.97 (t, J = 6.5 Hz, 2H), 1.79 (q, J = 7.1 Hz, 2H), 1.43 (q, J= 7.3 Hz, 2H), 1.25 (s, 24H), 0.87 (t, J = 6.7 Hz, 3H). 13 C NMR (151 MHz, CDCl3) δ 163.21, 161.52, 161.14, 147.96, 132.59, 129.30, 129.04, 126.03,118.15, 114.68, 68.88, 68.15, 31.93, 29.70, 29.68, 29.67, 29.61, 29.58,29.40, 29.37, 29.19, 26.02, 22.70, 14.13. IR (KBr), ν, cm -1 : 3293, 2926, 2853,1651, 1609, 1551, 1514, 1252, 840.

[0068] The structural characterization data of compound D2-9 are as follows: 1 H NMR (600 MHz, DMSO-d6) δ 11.47 (s, 1H),9.68 (s, 1H), 8.38 (s, 1H), 7.64 (d, J = 8.3 Hz, 2H), 7.47 (d, J = 2.1 Hz,1H), 7.43 (dd, J = 8.3, 1.41 (p, J = 7.0 Hz, 2H), 1.33 – 1.21 (m, 24H), 0.85 (t, J = 6.9 Hz, 3H). 13 C NMR (151 MHz, DMSO) δ 162.97, 160.63, 150.24, 147.74, 147.31, 129.00,127.39, 124.80, 121.68, 115.40, 115.24, 112.06, 68.07, 56.20, 40.42, 40.28,40.14, 40.00, 39.87, 39.73, 39.59, 31.76, 29.52, 29.50, 29.47, 29.44, 29.43,29.20, 29.17, 29.08, 25.93, 22.56, 14.42. IR (KBr), ν, cm -1 : 3451, 3226, 2921,2852, 1602, 1646, 1511, 1303, 1249, 1031, 833.

[0069] The gelling ability of the D2 series compounds was evaluated using the inverted vial method. Accurately weighed amounts of the compound and a measured amount of solvent were added to a stoppered test tube, and the mixture was heated until the solid completely dissolved. The test tube was then cooled to room temperature, and the state of the system was observed. If a homogeneous gel formed after cooling and no liquid flowed in the inverted test tube, it was recorded as a gel (G); if partial gelation occurred, it was recorded as a hemigel (PG); if crystals precipitated, it was recorded as crystals (P); if it remained a solution, it was recorded as a solution (S); if it could not completely dissolve during heating, it was recorded as insoluble (I). The time required for gel formation was recorded, and the results are shown in Table 1.

[0070] Table 1. Gel state of D2 series compounds in common solvents

[0071]

[0072] Table 1 shows that D2-3 can form gels in 5 wt% and 2 wt% petroleum ether, cyclohexane, isopropanol, n-butanol, acetonitrile, and DMSO, exhibiting excellent gelling ability. In rapeseed oil, D2-3 can form a gel at 5 wt%, with a sol temperature greater than 110℃, demonstrating good thermal stability. Considering its gelling ability, solubility, and stability, D2-3 was selected as the representative compound for subsequent research.

[0073] Example 2

[0074] Accurately weigh 30 mg of D2-3 and 60 mg, 90 mg, and 120 mg of borneol (mass ratios of 1:2, 1:3, and 1:4, respectively), add 0.5 mL of petroleum ether, seal, heat at 65 °C until completely dissolved, cool to form a gel, and freeze-dry in the same way to obtain borneol-D2-3 gelling agent dry gel, i.e., borneol-D2-3 assembly.

[0075] Simultaneously, 30 mg of D2-3 was accurately weighed, added to 0.5 mL of petroleum ether, heated to dissolve, and then cooled to form a gel. The gel was pre-frozen at -20°C for 12 h and then freeze-dried under vacuum at -80°C for 24 h to obtain a D2-3 dry gel. A comparative experiment was conducted with the above-mentioned borneol-D2-3 gelling agent dry gel.

[0076] To observe the changes in microstructure after assembly formation, field emission scanning electron microscopy was used to observe the microstructure of borneol, D2-3 dry gel, and assemblies with different proportions of borneol-D2-3. The results are as follows: Figure 1 As shown. Figure 1 A shows that the borneol raw material is in the form of irregular blocky crystals. Figure 1 B shows that the D2-3 dry gel has a long strip structure of about 3.00 μm, with fibers interwoven to form a three-dimensional network. Figure 1 The microstructure of C (1:2 assembly) tends to resemble that of D2-3 dry gel. As the proportion of borneol increases, Figure 1 D (1:3 assembly) exhibits a stacked sheet-like structure with a size of approximately 2 μm; Figure 1 The E (1:4 assembly) sheet-like structure is more dense, with a size of approximately 1 μm. The results indicate that D2-3 and borneol successfully assemble to form a new microstructure, and the assembly size gradually decreases and the network structure becomes more dense as the proportion of borneol increases.

[0077] To investigate whether new phases are formed during assembly formation, differential scanning calorimetry (DSC) was used to perform thermal analysis on borneol, D2-3 powder, D2-3 dry gel, and assemblies of different borneol-D2-3 ratios. The results are as follows: Figure 2 As shown. Figure 2 The results showed that borneol exhibited an endothermic peak at 204.15℃; the endothermic peaks of D2-3 powder and D2-3 dry gel appeared at 186.42℃ and 185.89℃, respectively, with the difference possibly related to the increased specific surface area and microstructure changes of the dry gel. When borneol and D2-3 formed assemblies, the 1:2, 1:3, and 1:4 assemblies showed new endothermic peaks at 134.94℃, 135.97℃, and 134.54℃, respectively, which were significantly different from the endothermic peaks of the raw materials, proving that a new phase was formed rather than a simple physical mixture.

[0078] To investigate the intermolecular forces involved in the formation of assemblies, Fourier transform infrared spectroscopy was used to characterize D2-3 powder, D2-3 dry gel, and assemblies of different proportions of borneol-D2-3. The results are as follows: Figure 3 As shown. Figure 3 The results show that the hydroxyl stretching vibration peak of borneol is at 3630 cm⁻¹. -1 The hydroxyl stretching vibration peak of the D2-3 dry gel is located at 3381 cm⁻¹. -1 Compared to D2-3 powder (3382cm) -1 There was a slight red shift. After assembly, the hydroxyl stretching vibration peaks of the 1:2, 1:3, and 1:4 assemblies shifted to 3351, 3356, and 3361 cm⁻¹, respectively. -1 All of these shifted towards lower wavenumbers. These results indicate that hydrogen bonding exists between borneol and D2-3 during self-assembly, and that hydrogen bonding is a crucial driving force for assembly formation.

[0079] To evaluate the inhibitory effect of the assembly on the volatility of borneol, borneol alone, a physical mixture of borneol and D2-3 (mass ratio 3:1), and borneol-D2-3 assemblies with different proportions were placed in a 50℃ constant temperature drying oven. Samples were taken and weighed at 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 hours, and the cumulative loss rate of borneol was calculated. The results are as follows: Figure 4 As shown. Figure 4 The results showed that pure borneol evaporated rapidly, with a loss rate of 39.34% after 5 hours and complete evaporation after 10 hours. The loss rate of the physically mixed samples after 10 hours was 55.48%, lower than that of pure borneol but higher than that of the assembled products. The loss rates of the 1:2, 1:3, and 1:4 assemblies after 10 hours were 14.14%, 10.45%, and 10.64%, respectively, significantly lower than those of the control group.

[0080] To further investigate the stability of the assembly under different environmental conditions, individual borneol and the borneol-D2-3 assembly with a mass ratio of 1:3 were placed under high temperature (50℃), light (4500 lx), and high humidity (25℃, relative humidity 90%±5%) conditions for 30 days. Samples were taken on days 0, 10, 20, and 30 to observe the appearance changes and determine the borneol retention rate. The results are shown in Table 2.

[0081] Table 2 Effects of high temperature, light, and high humidity on borneol and 1:3 assemblies (n=3)

[0082]

[0083] Table 2 shows that under high temperature conditions, borneol completely evaporates within 10 days, while the 1:3 assembly retains 8.11±2.11% after 30 days; under light conditions, the borneol retention rate is 96.99±0.94% after 30 days, and the assembly's is 98.01±0.88%; under high humidity conditions, the borneol retention rate is 99.31±0.31% after 30 days, and the assembly's is 99.11±0.67%. The assembly significantly improves the stability of borneol under high temperatures and also exhibits good tolerance to light and humidity.

[0084] The acute toxicity of D2-3 gelling agent and borneol-D2-3 assembly was evaluated using a zebrafish model. Wild-type AB strain zebrafish aged 4–5 months were acclimatized for one week and randomly divided into four groups: a blank control group, a solvent control group (0.1% DMSO), a borneol group, a D2-3 gelling agent group, and a borneol-D2-3 dry gel group (1:3 assembly), with 10 fish in each group. The borneol groups were administered at doses of 300, 600, 900, 1200, and 1500 μM; the D2-3 groups at doses of 100, 300, 600, and 900 μM; and the assembly groups at borneol concentrations of 300:100, 600:200, 900:300, 1200:400, and 1500:500 μM. The zebrafish's condition, poisoning symptoms, and number of deaths were observed at 1, 2, 3, 4, 6, 12, 24, 48, and 72 hours. The mortality rate and median lethal concentration (LC50) were calculated. Heart and liver tissues were collected for HE staining to observe pathological changes. The results are as follows: Figures 5-8 As shown.

[0085] Figure 5 The results showed that the LC50 of borneol was 855.1 μM; no death was observed in D2-3 gelling agent at concentrations of 0–900 μM, and the LC50 was not detected; the LC50 of the assembly (based on borneol) was 976.8 μM, which was higher than that of pure borneol, indicating that the toxicity was reduced after the assembly was formed. Figure 6 The cumulative mortality rate within 72 hours was compared between the borneol group (900 μM) and the assembly group (900:300 μM, left), and between the borneol group (1200 μM) and the assembly group (1200:400 μM, right). In the borneol 900 μM group, the mortality rate was 10% within 1 hour and reached 70% cumulatively within 3 hours; while in the assembly group (900:300 μM), there were no deaths within 4 hours, but the mortality rate slowly increased to 20% from 6 to 72 hours. In the borneol 1200 μM group, the mortality rate was 10% within 1 hour and all patients died within 4 hours; in the assembly group (1200:400 μM), there were no deaths within 2 hours, the mortality rate was 30% within 3 hours, and all patients died within 6 hours. These results indicate that the assembly delayed the toxicity expression process of borneol, exhibiting sustained-release characteristics.

[0086] Figure 7 The results showed that at concentrations above 900 μM, the heart showed varying degrees of inflammatory cell infiltration, fibrosis, and myocardial fiber degeneration and necrosis; while the corresponding organs in the assembled group showed milder pathological damage and no obvious pathological changes. Figure 8 The results showed that at concentrations above 900 μM, a small amount of hepatocyte necrosis was observed in the liver in the borneol group; only at high doses (1200:400, 1500:500 μM), slight sinusoidal congestion was observed in the assembly group, with no other significant pathological changes. These results indicate that D2-3 gelling agent is safe at concentrations ranging from 0 to 900 μM, and that forming assemblies with borneol can reduce the peak acute toxicity of borneol and alleviate organ pathological damage.

[0087] The efficacy of compound Danshen tablets containing borneol-D2-3 assemblies was evaluated using a rat model of acute myocardial ischemia. The effects of the assemblies on the anti-myocardial ischemia efficacy were investigated by detecting myocardial enzyme activity, myocardial infarction area, and observing myocardial tissue pathological changes. Specific experiments and results are as follows:

[0088] 1. Sample preparation

[0089] Weigh 225g of Salvia miltiorrhiza, crush it, add 5 times the amount of anhydrous ethanol, heat under reflux for 1.5h, filter the extract, recover the ethanol from the filtrate and concentrate it for later use; add 5 times the amount of 50% ethanol to the residue, heat under reflux for 1.5h, filter the extract, recover the ethanol from the filtrate and concentrate it to an appropriate amount for later use; add 8 times the amount of water to the residue and decoct for 2h, filter the decoction, and concentrate the filtrate to an appropriate amount for later use. Combine the concentrates, vacuum dry to obtain 98g of dry powder, with an extraction rate of 43.56% for Salvia miltiorrhiza. Weigh 70.5g of Panax notoginseng, pulverize it into a fine powder for later use. Grind 4g of borneol into a fine powder for later use. Prepare borneol-D2-3 dry gel (1:3 assembly) at a mass ratio of 3:1 (borneol to D2-3), freeze dry for later use.

[0090] 2. Animal grouping and administration

[0091] According to the 2025 edition of the Chinese Pharmacopoeia, the recommended daily dose for Compound Danshen Tablets for adults is 9 tablets (containing 4.05g of Danshen, 1.27g of Panax notoginseng, and 0.072g of borneol). For experimental rats, the dosage was calculated at twice the clinical dose: 0.064g / rat / day of Danshen extract, 0.046g / rat / day of Panax notoginseng, and 0.0026g / rat / day of borneol. The pharmacopoeia stipulates that each tablet of Compound Danshen Tablets contains 8 mg of borneol, and the dosage for the high-dose group (CDT-H) is 0.563 g / kg; when each tablet contains 0.15 mg of borneol, the dosage for the low-dose group (CDT-L) is 0.550 g / kg; each tablet of Compound Danshen Tablets containing the assembly contains 8 mg of borneol, and the dosage is 0.563 g / kg, with a gelling agent dosage of 0.0216 g / kg (CDT-H-LMWGs group); the dosage for the D2-3 gelling agent-only group (LMWGs group) is 0.0216 g / kg.

[0092] Sixty male SD rats were acclimatized for one week and then randomly divided into six groups: control group (Con), model group (ISO), CDT-H group, CDT-L group, CDT-H-LMWGs group, and LMWGs group, with 10 rats in each group. The drugs were administered by gavage once daily for 7 days. On day 6, except for the Con group, all other groups were injected subcutaneously with isoproterenol (85 mg / kg) one hour after drug administration to establish an acute myocardial ischemia model, with injections continuing for two consecutive days. The Con group received an equal volume of physiological saline subcutaneously.

[0093] 3. Sample collection and testing

[0094] Following the last subcutaneous injection of ISO, the patient fasted for 12 hours, then received an intraperitoneal injection of 20% urethane (5 mL / kg) for anesthesia. Blood was drawn from the abdominal aorta, allowed to stand at room temperature for 1 hour, and centrifuged at 3500 rpm for 15 minutes. The supernatant was collected as serum and stored at -80℃ for later use. The heart was removed, rinsed with physiological saline, and divided into three portions: one for TTC staining to determine the myocardial infarction area, one fixed with 4% paraformaldehyde for pathological sections, and one for biochemical index detection.

[0095] 4. Heart Mass Index

[0096] Weigh the whole heart and calculate the cardiac index = whole heart weight / body weight (mg / g). The result is as follows. Figure 9 As shown. Compared with the Con group, the cardiac index of the ISO group was significantly increased (P<0.01); compared with the ISO group, the cardiac index of the CDT-L group was decreased (P<0.05), and the cardiac index of the CDT-H and CDT-H-LMWGs groups was significantly decreased (P<0.01), while there was no significant difference in the LMWGs group (P>0.05).

[0097] 5. Measurement of myocardial infarction area

[0098] After being frozen at -20°C for 30 minutes, the heart was cut into 1-2 mm thin sections and stained with 2% TTC in the dark for 20 minutes. Normal myocardium stained red, while infarcted myocardium stained white. The infarct area was analyzed using ImageJ software, and the results are as follows: Figure 9 As shown in the figure. Compared with the Con group, the myocardial infarction area in the ISO group was significantly increased (P<0.01); compared with the ISO group, the myocardial infarction area in the LMWGs group, CDT-L group, CDT-H group and CDT-H-LMWGs group was significantly reduced (P<0.01), among which the CDT-H-LMWGs group had the same effect as the CDT-H group.

[0099] 6. Detection of myocardial enzymes and inflammatory factors

[0100] Serum CK, LDH, CK-MB activities and cTnI concentrations, as well as the levels of inflammatory factors TNF-α, IL-1β, and IL-6, were measured according to the kit instructions. Myocardial SOD activity and MDA levels were also measured. Results showed that, compared with the ISO group, the CDT-H-LMWGs group significantly reduced myocardial enzyme activity (P<0.01), decreased inflammatory factor levels (P<0.05 or P<0.01), increased SOD activity, and decreased MDA levels (P<0.05), with effects comparable to the CDT-H group. The LMWGs group, however, showed no significant therapeutic effect.

[0101] 7. Pathological observation of myocardial tissue

[0102] HE staining was used to observe pathological changes in myocardial tissue, and the results were as follows: Figure 10 As shown in the diagram, the Con group showed normal cardiomyocyte structure and tightly and neatly arranged myocardial fibers. The ISO group showed loose and disordered myocardial fiber arrangement, interstitial edema, extensive myocardial cell degeneration and necrosis, and extensive inflammatory cell infiltration. The LMWGs group showed a small amount of myocardial cell degeneration and necrosis, and a small amount of inflammatory cell infiltration. The CDT-L group showed loosely arranged myocardial fibers and a small amount of myocardial degeneration and edema. The CDT-H group and the CDT-H-LMWGs group showed significantly reduced myocardial tissue structure disorder, significantly reduced inflammatory cell infiltration and myocardial cell degeneration and necrosis, and significantly reduced overall pathological changes.

[0103] The results showed that the compound Danshen tablets containing the assembly could effectively improve acute myocardial ischemia. The introduction of the gelling agent did not affect the anti-myocardial ischemia efficacy of the formulation. D2-3, as a drug delivery excipient, did not participate in the therapeutic effect itself.

Claims

1. A small molecule gelling agent containing hydrazone, characterized in that: The structural formula of the gelling agent is shown below: Where R1 represents C4~C 18 A straight-chain alkyl group; R2 represents hydrogen or hydroxyl; R3 is selected from hydrogen, , , , , , , , Any one of them.

2. The hydrazone-containing small molecule gelling agent according to claim 1, characterized in that: R3 is selected from , , , , , Any one of them.

3. A method for synthesizing a small molecule gelling agent containing hydrazone as described in claim 1 or 2, characterized in that... Includes the following steps: Step 1: React the p-hydroxybenzaldehyde compound shown in Formula I with the bromoalkane shown in Formula II under alkaline conditions to generate the alkoxybenzaldehyde intermediate shown in Formula III. Step 2: The alkoxybenzaldehyde intermediate obtained in Step 1 is reacted with the hydrazine compound shown in Formula IV in a high-boiling-point polar solvent by heating. After cooling, washing, and pulping, a small molecule gelling agent containing a hydrazone group as shown in Formula V is obtained. The synthetic route is as follows: 。 4. The method for synthesizing the hydrazone-containing small molecule gelling agent according to claim 3, characterized in that, In step 1, the molar ratio of the p-hydroxybenzaldehyde compound to the bromoalkanes is 1:1 to 1.2, the alkaline conditions are provided by anhydrous potassium carbonate, the reaction temperature is 50 to 70°C, and the reaction time is 5 to 24 hours.

5. The method for synthesizing the hydrazone-containing small molecule gelling agent according to claim 3, characterized in that, The high-boiling-point polar solvent mentioned in step 2 is diethylene glycol; the reaction temperature is 120–135°C.

6. A small molecule gelling agent-borneol assembly, characterized in that: The hydrazone-containing small molecule gelling agent of claim 1 is self-assembled with borneol in an organic solvent to form a gel, and then the assembly is obtained by freeze-drying; the organic solvent is any one of petroleum ether, cyclohexane, isopropanol, n-butanol, acetonitrile, and DMSO.

7. The small molecule gelling agent-borneol assembly according to claim 6, characterized in that: The hydrazone-containing small molecule gelling agent and borneol were added to an organic solvent at a mass ratio of 1:2 to 4. The mixture was heated to 60 to 90°C under sealed conditions until completely dissolved, and then allowed to stand at 0 to 5°C for 10 to 30 minutes to form a gel. The mixture was then pre-frozen at -25 to -15°C for 4 to 6 hours, and subsequently freeze-dried under vacuum at -80 to -60°C for 20 to 24 hours to obtain the small molecule gelling agent-borneol assembly.

8. The small molecule gelling agent-borneol assembly according to claim 7, characterized in that: The mass of the small molecule gelling agent containing hydrazone added per milliliter of organic solvent is 50–70 mg.

9. The small molecule gelling agent-borneol assembly according to claim 6, characterized in that: The organic solvent is petroleum ether or ethanol.

10. The use of the hydrazone-containing small molecule gelling agent of claim 1 or the small molecule gelling agent-borneol assembly of any one of claims 6 to 9 in the preparation of pharmaceutical formulations that improve the stability of borneol and reduce its toxicity.

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