A folic acid derivative, its preparation method, and its application in preventing ferroptosis.

Abstract

This invention belongs to the field of pharmaceutical application technology, specifically relating to a folic acid derivative, its preparation method, and its application in preventing and treating ferroptosis. The folic acid derivative of this invention comprises compounds represented by general formula (I) and their pharmaceutically acceptable salts. The folic acid compounds provided by this invention, by introducing a lipophilic group or a specific group at the R position, yield compounds with good ferroptosis inhibitory activity, which can be used to treat non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, atherosclerosis, acute kidney injury, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and epilepsy.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical application technology, specifically relating to a folic acid derivative, its preparation method, and its application in preventing ferroptosis. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Ferroptosis is a recently discovered programmed cell death mechanism distinct from apoptosis, necrosis, and autophagy in morphology, biochemistry, and genetics. It is named ferroptosis because it is an oxidative, non-apoptotic programmed cell death process dependent on the presence of iron ions. Its mechanism involves an imbalance between the generation and degradation of reactive oxygen species (ROS) in cell membrane lipids; typical characteristics include smaller mitochondria, increased bilayer membrane density, and an increase in ROS free radicals in cell membrane lipids. Recent data also indicate that ferroptosis is closely related to some normal physiological conditions, such as tumor suppression, immunity, and development. For example, CD8+ T lymphocytes exert tumor immune function by inducing ferroptosis in cancer cells. Notably, lipid peroxidation-driven ferroptosis has attracted considerable attention, primarily because it is involved in numerous human pathological conditions, such as organ injury, neurodegeneration, and organ ischemia-reperfusion injury. Importantly, in various preclinical animal models, pharmacological modulation of ferroptosis has proven to be a promising therapeutic approach for these diseases.

[0004] Ferrostatin-1 and Liproxstatin-1 are first-generation small-molecule ferroptosis inhibitors that function by scavenging lipid free radical damage to cell membranes and blocking ferroptosis. These inhibitors have well-defined mechanisms of action and structure-activity relationships. However, both compounds suffer from drawbacks such as short half-lives, high toxicity, and difficulty in storage. Summary of the Invention

[0005] To address the shortcomings of existing technologies, the present invention aims to provide a folic acid derivative, its preparation method, and its applications. This invention provides a folic acid derivative comprising folic acid diester and folic acid diamide, and its preparation method. Based on research into the mechanism of ferroptosis, through cell membrane-level lipid free radical inhibition experiments, cell activity tests, and whole-animal-level pharmacological activity assays, the folic acid derivative has been discovered and verified as an inhibitor of ferroptosis. This invention offers novel applications in the treatment of non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, acute kidney injury, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and epilepsy.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solution:

[0007] In a first aspect, the present invention provides a folic acid derivative comprising a compound of general formula (I) and a pharmaceutically acceptable salt thereof;

[0008] General formula (I);

[0009] Wherein, R is one or more of alkyl, cycloalkyl, and aryl groups.

[0010] Secondly, the present invention provides a method for preparing the folic acid derivative described in the first aspect, comprising the following steps:

[0011] Folic acid and p-toluenesulfonic acid are dissolved in alcohol or amine and heated under reflux. After the reaction is complete, the reaction system is cooled to room temperature, the solvent is concentrated under reduced pressure, filtered, and the filter cake is washed with cold alcohol. The filter cake is dried under vacuum to obtain a crude product. The crude product is recrystallized to obtain a folic acid derivative.

[0012] Thirdly, the present invention provides the application of the folic acid derivative described in the first aspect in the preparation of a drug that inhibits cell ferroptosis.

[0013] Fourthly, the present invention provides the application of the folic acid derivative described in the first aspect in the preparation of a drug for preventing and treating ferroptosis-related diseases, wherein the ferroptosis-related diseases include one or more of non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, acute kidney injury, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases, wherein the neurodegenerative diseases include one or more of Alzheimer's disease, Parkinson's disease, and epilepsy.

[0014] Fifthly, the present invention provides the use of the folic acid derivative described in the first aspect in the preparation of a drug for preventing and treating diseases related to elevated homocysteine, said diseases including one or more of hypertension, fatty liver, osteoporosis, and preeclampsia with elevated homocysteine.

[0015] The beneficial effects achieved by one or more technical solutions of the present invention are as follows:

[0016] This invention successfully synthesizes folic acid derivatives of different substituents, such as folic acid esters or folic acid amides. These folic acid derivatives exhibit high activity and stability, and can block the chain reaction of membrane lipid free radicals, reduce the damaging effects of free radicals on cell membranes, and prevent cell ferroptosis.

[0017] This invention demonstrates through cytotoxicity experiments that folic acid derivatives at therapeutic doses are effective in treating non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, acute kidney injury, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and epilepsy without significant toxicity.

[0018] This invention provides treatment plans and theoretical basis for the treatment of non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, acute kidney injury, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and epilepsy, targeting ferroptosis. Attached Figure Description

[0019] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0020] Figure 1 This is a graph showing the inhibitory effects of folic acid derivatives A2 and B1 on inducer-induced ferroptosis in Example 4.

[0021] Figure 2 The graph shows the data on the prevention of lipid peroxidation by folic acid derivatives A1, A2, A3, A4, B1, and B2 in Example 4, where Ctrl represents the group that only added RSL3;

[0022] Figure 3 To assess kidney damage in mice treated with folic acid derivative A2 in Example 5, creatinine and urea were used as biochemical indicators.

[0023] Figure 4 HE staining image of kidney damage in mice treated with folic acid derivative A2 in Example 5;

[0024] Figure 5 To assess liver damage in mice treated with folic acid derivative A2 in Example 6, aspartate aminotransferase (ALT) and alanine aminotransferase (AST) were used as biochemical indicators.

[0025] Figure 6 HE staining image of liver damage in mice treated with folic acid derivative A2 in Example 6. Detailed Implementation

[0026] A first typical embodiment of the present invention is a folic acid derivative comprising a compound of general formula (I) and a pharmaceutically acceptable salt thereof;

[0027] General formula (I);

[0028] Wherein, R is one or more of alkyl, cycloalkyl, and aryl groups.

[0029] In one or more embodiments of this implementation, the alkyl group is selected from C1 to C20 alkyl groups, the cycloalkyl group is selected from an aliphatic cyclic structure having 6 atoms, and the aryl group is selected from alkylphenyl groups.

[0030] In one or more embodiments of this implementation, the pharmaceutically acceptable salt includes one or more of hydrochloride, sulfate, and phosphate.

[0031] A second typical embodiment of the present invention, a method for preparing the folic acid derivative described in the first typical embodiment, characterized by comprising the following steps:

[0032] Folic acid and paracatalytic acid are dissolved in alcohol or amine and heated under reflux. After the reaction is complete, the reaction system is cooled to room temperature, the solvent is concentrated under reduced pressure, filtered, and the filter cake is washed with cold alcohol. The filter cake is dried under vacuum to obtain a crude product. The crude product is recrystallized to obtain a folic acid derivative.

[0033] In one or more embodiments of this implementation, the alcohol includes one or more of alkyl alcohols, cycloalkyl alcohols, and aryl alcohols;

[0034] Alternatively, the amine may include one or more of alkylamines, cycloalkylamines, and arylamines;

[0035] Preferably, the alkyl group of the alkyl alcohol is selected from C1 to C20 alkyl groups, the cycloalkyl group of the cycloalkyl alcohol is selected from an aliphatic cyclic structure having 6 atoms, and the aryl group of the aryl alcohol is selected from alkylphenyl groups;

[0036] Preferably, the alkyl group of the alkylamine is selected from C1 to C20 alkyl groups, the cycloalkyl group of the cycloalkylamine is selected from an aliphatic cyclic structure having 6 atoms, and the aryl group of the arylamine is selected from alkylphenyl groups.

[0037] In one or more embodiments of this implementation, the catalytic acid is p-toluenesulfonic acid or sulfuric acid; the molar ratio of folic acid to the catalytic acid is 1:0.99-1.01.

[0038] In one or more embodiments of this implementation, the reflux temperature is 79-81 °C and the reaction time is 11-13 h.

[0039] The third typical embodiment of the present invention is the application of the folic acid derivative described in the first typical embodiment in the preparation of a drug that inhibits cell ferroptosis.

[0040] The fourth typical embodiment of the present invention is the application of the folic acid derivative described in the first typical embodiment in the preparation of a drug for preventing and treating ferroptosis-related diseases. The ferroptosis-related diseases include one or more of non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, acute kidney injury, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases. The neurodegenerative diseases include one or more of Alzheimer's disease, Parkinson's disease, and epilepsy.

[0041] The fifth typical embodiment of the present invention is the use of the folic acid derivative described in the first typical embodiment in the preparation of a drug for preventing and treating diseases related to elevated homocysteine, wherein the diseases include one or more of the following: hypertension, fatty liver, osteoporosis, and preeclampsia with elevated homocysteine.

[0042] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments and comparative examples.

[0043] Example 1

[0044] Preparation of target compounds A1-A31

[0045] Folic acid (440 mg, 1 mmol) and a catalytic acid (1 mmol) were added to a round-bottom flask, along with a large excess of alcohol as a solvent. The mixture was refluxed at 80 °C for 12 h. After the reaction was complete, the reactants were cooled to room temperature, the solvent was concentrated under reduced pressure, filtered, and the filter cake was washed three times with the corresponding cold alcohol. The filter cake was then dried under vacuum to obtain a crude product. The dried crude product was recrystallized to obtain the target compound. Different target compounds, A1-A31, were obtained by changing the types of alcohol and catalytic acid.

[0046] When methanol was used as the alcohol and p-benzylbenzenesulfonic acid was used as the catalytic acid, a yellow solid, target compound A1, was obtained in 80% yield. The chemical formula of A1 is... 1 The H NMR data are as follows:

[0047] ;

[0048] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 3.61 (s, 3H), 3.57 (s, 3H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H).

[0049] When the alcohol is ethanol and the catalytic acid is p-benzylbenzenesulfonic acid, a yellow solid, target compound A2, is obtained in 75% yield. The chemical formula of A2 is... 1 The H NMR data are as follows:

[0050] ;

[0051] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.26 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.8 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J = 8.8 Hz, 2H),4.49 (s, 2H), 4.38 (ddd, J = 9.6, 7.4, 5.3 Hz, 1H), 4.11 – 3.87 (m, 4H), 2.42(t, J = 7.6 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.56 (tq, J = 14.0, 7.1 Hz, 4H), 0.98 – 0.82 (m, 6H).

[0052] When the alcohol is n-propanol and the catalytic acid is p-benzylbenzenesulfonic acid, a yellow solid, target compound A3, is given in 55% yield. The chemical formula of A3 is... 1The H NMR data are as follows:

[0053] ;

[0054] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.26 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.8 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J = 8.8 Hz, 2H),4.49 (s, 2H), 4.38 (ddd, J = 9.6, 7.4, 5.3 Hz, 1H), 4.11 – 3.87 (m, 4H), 2.42(t, J = 7.6 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.56 (tq, J = 14.0, 7.1 Hz, 4H), 0.98 – 0.82 (m, 6H).

[0055] When isopropanol is used as the alcohol and sulfuric acid is used as the catalytic acid, a yellow solid, target compound A4, is given in 60% yield. The chemical formula of A4 is... 1 The H NMR data are as follows:

[0056] ;

[0057] 1 H NMR(400 MHz, DMSO) δ 11.43 (s, 1H), 8.65 (s, 1H), 8.21 (d, J = 7.4Hz, 1H), 7.64 (d, J = 8.7 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 4.88 (dq, J = 12.1, 6.0 Hz, 2H), 4.49 (d, J = 3.8 Hz, 2H), 4.36 – 4.28 (m,1H), 2.35 (dd, J = 13.8, 6.2 Hz, 2H), 2.06 – 1.91 (m, 2H), 1.17 (dd, J =11.2, 6.3 Hz, 12H).

[0058] When the alcohol is n-butanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid, target compound A5, is given in 60% yield. The chemical formula of A5 is... 1 The H NMR data are as follows:

[0059] ;

[0060] 1 H NMR(400 MHz, DMSO) δ 11.48 (s, 1H), 8.65 (s, 1H), 8.25 (d, J = 7.4Hz, 1H), 7.64 (d, J = 8.5 Hz, 2H), 6.96 (s, 3H), 6.64 (d, J = 8.6 Hz, 2H), 4.49 (s, 2H), 4.36 (dd, J = 13.5, 8.3 Hz, 1H), 4.02 (dt, J = 13.3, 6.3 Hz, 4H), 2.41 (t, J = 7.4 Hz, 2H), 2.10 – 1.93 (m, 2H), 1.52 (dd, J = 14.0, 6.3Hz, 4H), 1.36 – 1.26 (m, 4H), 0.86 (t, J = 7.3 Hz, 6H).

[0061] When isobutanol is used as the alcohol and sulfuric acid is used as the catalytic acid, a yellow solid, target compound A6, is given in 60% yield. The chemical formula of A6 is... 1 The H NMR data are as follows:

[0062] ;

[0063] 1 H NMR(400 MHz, DMSO) δ 11.42 (s, 1H), 8.65 (s, 1H), 8.27 (d, J = 7.5Hz, 1H), 7.64 (d, J = 8.7 Hz, 2H), 6.97 (t, J = 5.7 Hz, 3H), 6.64 (d, J = 8.8Hz, 2H), 4.48 (d, J = 5.5 Hz, 2H), 4.43 – 4.35 (m, 1H), 3.83 (ddd, J = 19.3,10.7, 6.6 Hz, 4H), 2.44 (t, J = 7.5 Hz, 2H), 2.13 – 1.95 (m, 2H), 1.91 – 1.79 (m, 2H), 0.84 (t, J = 10.1 Hz, 12H).

[0064] When the alcohol is sec-butanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid, target compound A7, is given in 70% yield. The chemical formula of A7 is... 1 The H NMR data are as follows:

[0065] ;

[0066] 1 H NMR (400 MHz, DMSO) δ 11.52 (s, 1H), 8.66 (s, 1H), 8.23 ​​(d, J = 5.4Hz, 1H), 7.65 (d, J = 8.3 Hz, 2H), 6.96 (s, 3H), 6.64 (d, J = 8.5 Hz, 2H), 4.73 (d, J = 5.9 Hz, 2H), 4.49 (s, 2H), 4.34 (s, 1H), 2.39 (s, 2H), 2.03 (dd,J = 24.7, 18.5 Hz, 2H), 1.57 – 1.42 (m, 4H), 1.14 (dd, J = 9.2, 6.7 Hz, 6H),0.82 (dd, J = 15.7, 7.6 Hz, 6H).

[0067] When the alcohol is tert-butanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid, target compound A8, is given in 78% yield. The chemical formula of A8 is... 1 The H NMR data are as follows:

[0068] ;

[0069] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.25 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J =7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.42(s, 18H).

[0070] When the alcohol is n-pentanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid, target compound A9, is given in 85% yield. The chemical formula of A9 is... 1 The H NMR data are as follows:

[0071] ;

[0072] 1 H NMR(400 MHz, DMSO) δ 11.48 (s, 1H), 8.65 (s, 1H), 8.25 (d, J = 7.4Hz, 1H), 7.64 (d, J = 8.5 Hz, 2H), 6.96 (s, 3H), 6.64 (d, J = 8.6 Hz, 2H), 4.49 (s, 2H), 4.36 (dd, J = 13.5, 8.3 Hz, 1H), 4.02 (dt, 4H), 2.41 (t, J =7.4 Hz, 2H), 2.10 – 1.93 (m, 2H), 1.60 (m, 4H), 1.36 (m, 8H), 0.86 (t, 6H).

[0073] When the alcohol is 2-pentanol and the catalytic acid is p-toluenesulfonic acid, the target compound A10, a yellow solid, is given in 68% yield. The chemical formula of A10 is... 1 The H NMR data are as follows:

[0074] ;

[0075] 1 H NMR (400 MHz, DMSO) δ 11.52 (s, 1H), 8.66 (s, 1H), 8.23 ​​(d, J = 5.4Hz, 1H), 7.65 (d, J = 8.3 Hz, 2H), 6.96 (s, 3H), 6.64 (d, J = 8.5 Hz, 2H),4.73 (d, 2H), 4.49 (s, 2H), 4.34 (s, 1H), 2.39 (s, 2H), 2.03 (dd, J = 24.7,18.5 Hz, 2H), 1.50 – 1.42 (m, 4H), 1.30 (m, 4H), 1.14 (d, 6H), 0.82 (dt, 6H).

[0076] When the alcohol is 3-pentanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid target compound A11 is given in 79% yield. The chemical formula of A11 is... 1 The H NMR data are as follows:

[0077] ;

[0078] 1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H), 8.66 (s, 1H), 8.23 ​​(d, J = 5.4Hz, 1H), 7.65 (d, J = 8.3 Hz, 2H), 6.96 (s, 3H), 6.64 (d, J = 8.5 Hz, 2H),4.67 (m, 1H), 4.57 (m, 1H), 4.49 (s, 2H), 4.34 (s, 1H), 2.39 (s, 2H), 2.03(dd, J = 24.7, 18.5 Hz, 2H), 1.57 – 1.42 (m, 8H), 0.89 (dt, 12H).

[0079] When the alcohol is 2-methyl-1-butanol and the catalytic acid is p-toluenesulfonic acid, the target compound A12, a yellow solid, is given in 73% yield. The chemical formula of A12 is... 1 The H NMR data are as follows:

[0080] ;

[0081] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H),4.18(dd, 4H),2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.55(m, 4H), 0.93(dt, 6H), 0.85(d, 6H).

[0082] When the alcohol is 2-methyl-2-butanol and the catalytic acid is p-toluenesulfonic acid, the target compound A13, a yellow solid, is given in 79% yield. The chemical formula of A13 is... 1 The H NMR data are as follows:

[0083] ;

[0084] 1H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J =7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.68(q, 2H), 1.48(q, 2H), 1.33(m, 12H), 0.90(dt, 6H).

[0085] When the alcohol is 3-methyl-1-butanol and the catalytic acid is p-toluenesulfonic acid, the target compound A14, a yellow solid, is given in 65% yield. The chemical formula of A14 is... 1 The H NMR data are as follows:

[0086] ;

[0087] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.10(dt, 4H),1.52(dt, 4H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 0.97 (s, 12H).

[0088] When the alcohol is 3-methyl-2-butanol and the catalytic acid is p-toluenesulfonic acid, the target compound A15, a yellow solid, is given in 78% yield. The chemical formula of A15 is... 1 The H NMR data are as follows:

[0089] ;

[0090] 1H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.34(m, 2H),2.51(m, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.32(s, 6H), 0.98 (s, 12H).

[0091] When the alcohol is 2,2-dimethyl-1-propanol and the catalytic acid is p-toluenesulfonic acid, the target compound A16, a yellow solid, is given in 78% yield. The chemical formula of A16 is... 1 The H NMR data are as follows:

[0092] ;

[0093] 1 H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H),3.97(s, 2H),3.81(s, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.25(s, 18H).

[0094] When the alcohol is cyclohexanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid, target compound A17, is given in 84% yield. The chemical formula of A17 is... 1 The H NMR data are as follows:

[0095] ;

[0096] 1H NMR (400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 4.60 – 4.53 (m, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.63(m,8H), 1.53(m,8H), 1.46 (m, 4H).

[0097] When the alcohol is cyclopentylethanol and the catalytic acid is p-toluenesulfonic acid, the target compound A18, a yellow solid, is given in 77% yield. The chemical formula of A18 is... 1 The H NMR data are as follows:

[0098] ;

[0099] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.29(d, 2H),3.85(d, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H),1.76(m, 8H), 1.68– 1.47 (m, 10H).

[0100] When the alcohol is 2-methylcyclopentanol and the catalytic acid is p-toluenesulfonic acid, the target compound A19, a yellow solid, is given in 84% yield. The chemical formula of A19 is... 1 The H NMR data are as follows:

[0101] ;

[0102] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.94(m, 1H), 4.58(m, 1H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4,5.4 Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.76 – 1.62(m,13H), 1.39(m, 1H), 0.90(dd, 6H).

[0103] When the alcohol is 3-methylcyclopentanol and the catalytic acid is p-toluenesulfonic acid, the target compound A20, a yellow solid, is given in 82% yield. The chemical formula of A20 is... 1 The H NMR data are as follows:

[0104] ;

[0105] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),5.15(m, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H),2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.85 – 1.65(m, 14H), 0.86 (s, 6H).

[0106] When the alcohol is cyclobutylethanol and the catalytic acid is p-toluenesulfonic acid, a yellow solid, target compound A21, is given in 85% yield. The chemical formula of A21 is... 1 The H NMR data are as follows:

[0107] ;

[0108] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.13 – 4.06(m, 4H), 2.42 (t, J = 7.4 Hz, 2H), 2.14 – 1.77 (m, 16H), 1.52(m, 4H).

[0109] When the alcohol is (2-methylcyclobutyl)methanol and the catalytic acid is p-toluenesulfonic acid, the target compound A22, a yellow solid, is given in 90% yield. The chemical formula of A22 is... 1 The H NMR data are as follows:

[0110] ;

[0111] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.24(dd, 4H),2.63(m, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 4H), 1.72 – 1.47(m,8H), 0.88 (dd, 6H).

[0112] When the alcohol is (3-methylcyclobutyl)methanol and the catalytic acid is p-toluenesulfonic acid, the target compound A23, a yellow solid, is given in 80% yield. The chemical formula of A23 is... 1 The H NMR data are as follows:

[0113] ;

[0114] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.29(d, 2H),3.84(m, 2H), 2.73(m, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.24(m, 2H), 2.12 – 1.93(m, 2H), 1.66 – 1.41(m, 8H), 0.86(d, 6H).

[0115] When the alcohol is 2,3-dimethylcyclobutanol and the catalytic acid is p-toluenesulfonic acid, the target compound A24, a yellow solid, is given in 89% yield. The chemical formula of A24 is... 1 The H NMR data are as follows:

[0116] ;

[0117] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.23(m, 2H),2.88(m, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.29(m, 2H), 2.12 – 1.86 (m, 6H), 0.88 (d, 12H).

[0118] When the alcohol is 2,4-dimethylcyclobutanol and the catalytic acid is p-toluenesulfonic acid, the target compound A25, a yellow solid, is given in 83% yield. The chemical formula of A25 is... 1 The H NMR data are as follows:

[0119] ;

[0120] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H),4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.23(s, 2H),2.98(m, 4H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.80 – 1.55(m,4H), 0.88 (d, 12H).

[0121] When benzyl alcohol was used as the alcohol and p-toluenesulfonic acid was used as the catalytic acid, the target compound A26, a yellow solid, was obtained in 67% yield. The chemical formula of A26 is... 1 The H NMR data are as follows:

[0122] ;

[0123] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.33(d, 10H), 6.98 (s, 3H), 6.64 (d, J =8.7 Hz, 2H), 5.34(s, 2H), 5.20(s, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J= 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H).

[0124] When the alcohol is 1-phenylethanol and the catalytic acid is p-toluenesulfonic acid, the target compound A27, a yellow solid, is given in 73% yield. The chemical formula of A27 is... 1 The H NMR data are as follows:

[0125] ;

[0126] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.32 – 7.25(m, 10H), 6.98 (s, 3H), 6.64(d, J = 8.7 Hz, 2H), 6.05(m, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J =9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.68(dd, 6H).

[0127] When the alcohol is 2-phenylethanol and the catalytic acid is p-toluenesulfonic acid, the target compound A28, a yellow solid, is given in 66% yield. The chemical formula of A28 is... 1 The H NMR data are as follows:

[0128] ;

[0129] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.23 – 7.19(m, 10H), 6.98 (s, 3H), 6.64(d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.41(dt, 4H), 4.38 (ddd, J =9.5, 7.4, 5.4 Hz, 1H), 2.96(dt, 4H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H).

[0130] When the alcohol is 2-methylbenzyl alcohol and the catalytic acid is p-toluenesulfonic acid, the target compound A29, a yellow solid, is given in 87% yield. The chemical formula of A29 is... 1 The H NMR data are as follows:

[0131] ;

[0132] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.32(d, 2H), 7.23(m, 4H), 7.12(m, 2H),6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 5.34(s, 2H), 5.05(s, 2H), 4.49 (d, J= 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.29 (s, 6H), 2.12 – 1.93 (m, 2H).

[0133] When the alcohol is 2-methylbenzyl alcohol and the catalytic acid is p-toluenesulfonic acid, the target compound A30, a yellow solid, is given in 66% yield. The chemical formula of A30 is... 1 The H NMR data are as follows:

[0134] ;

[0135] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.51(dt, 2H), 7.37(dd, 2H), 7.19(s, 2H),7.05(dd, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 5.34(s, 2H), 5.20(s,2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t,J = 7.4 Hz, 2H), 2.31(s, 6H), 2.12 – 1.93 (m, 2H).

[0136] When the alcohol is 4-methylbenzyl alcohol and the catalytic acid is p-toluenesulfonic acid, the target compound A31, a yellow solid, is given in 65% yield. The chemical formula of A31 is... 1 The H NMR data are as follows:

[0137] ;

[0138] 1 H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.13(t, 4H), 7.09(t, 4H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz, 2H), 5.34 (s, 2H), 5.20 (s, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.19(s, 6H), 2.12 – 1.93 (m, 2H).

[0139] Example 2

[0140] Folic acid diester (1 mmol) was added to a round-bottom flask, followed by a large excess of the corresponding amine as a solvent. The mixture was refluxed at 80 °C for 12 h. After the reaction was complete, the reactants were cooled to room temperature, the solvent was concentrated under reduced pressure, filtered, and the filter cake was washed three times with the cold corresponding amine. The filter cake was then dried under vacuum to obtain a crude product. The dried crude product was recrystallized to obtain a yellow solid. The target compounds B1-B10 were prepared by using different substituted amines.

[0141] When the amine used was an aqueous solution of ethylamine, the product was a yellow solid, target compound B1, in 75% yield. The chemical formula and 1H NMR data of B1 are as follows:

[0142] ;

[0143] 1H NMR(400 MHz, DMSO) δ 11.45 (s, 1H), 8.65 (s, 1H), 8.25 (d, J = 7.5Hz, 1H), 8.01(s, 2H), 7.65 (d, J = 8.7 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J =8.7 Hz, 2H), 4.49 (s, 2H), 4.36 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 3.24 – 3.11(m, 4H), 2.40 (t, J = 7.5 Hz, 2H), 2.11 – 1.90 (m, 2H), 0.99 (q, 6H).

[0144] When isopropylamine was used as the amine, the product was a yellow solid, target compound B2, with a yield of 83%. The chemical formula and 1H NMR data of B2 are as follows:

[0145] ;

[0146] 1H NMR(400 MHz, DMSO) δ 11.43 (s, 1H), 8.65 (s, 1H), 8.21 (d, J = 7.4Hz, 1H), 8.14(s, 2H), 7.64 (d, J = 8.7 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J =8.7 Hz, 2H), 3.81 (dq, 2H), 4.49 (d, J = 3.8 Hz, 2H), 4.36 – 4.28 (m, 1H), 2.35 (dd, J = 13.8, 6.2 Hz, 2H), 2.06 – 1.91 (m, 2H), 1.00 (dd, 12H).

[0147] When n-propylamine is used as the amine, the product is a yellow solid B3 with a yield of 90%. The chemical formula of B3 is [formula missing]. 1 The H NMR data are as follows:

[0148] ;

[0149] 1H NMR(400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.26 (d, J = 7.5Hz, 1H), 8.01(s, 1H), 7.75(s, 1H), 7.65 (d, J = 8.8 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J = 8.8 Hz, 2H), 4.49 (s, 2H), 4.38 (ddd, J = 9.6, 7.4, 5.3 Hz, 1H), 3.42 – 3.18 (m, 4H), 2.42 (t, J = 7.6 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.56 (tq, 4H), 0.98 – 0.82 (m, 6H).

[0150] When 3-pentylamine is used, the product is a yellow solid B4, with a yield of 75%. The chemical formula of B4 is [formula missing]. 1 The H NMR data are as follows:

[0151] ;

[0152] 1H NMR(400 MHz, DMSO) δ 11.52 (s, 1H), 8.66 (s, 1H), 8.23 ​​(d, J = 5.4Hz, 1H), 8.14(s, 2H), 7.65 (d, J = 8.3 Hz, 2H), 6.96 (s, 3H), 6.64 (d, J =8.5 Hz, 2H), 4.49 (s, 2H), 4.34 (ddd, 1H), 3.40(m, 2H), 2.39 (s, 2H), 2.03(dd, J = 24.7, 18.5 Hz, 2H), 1.57 – 1.42 (m, 8H), 0.89 (dt, 12H).

[0153] When tert-butylamine is used, the product is a yellow solid B5, with a yield of 65%. The chemical formula of B5 is [formula missing]. 1 The H NMR data are as follows:

[0154] ;

[0155] 1H NMR(400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.25 (d, J = 7.5Hz, 1H), 8.18(s, 2H), 7.65 (d, J = 8.7 Hz, 2H), 6.97 (s, 3H), 6.64 (d, J =8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.37(s, 18H).

[0156] When benzylamine is used, the product is a yellow solid B6, with a yield of 73%. The chemical formula of B6 is [formula missing]. 1 The H NMR data are as follows:

[0157] ;

[0158] 1H NMR (400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.31 – 7.23(m, 10H), 6.98 (s, 3H), 6.64(d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.44(s, 2H), 4.38 (ddd, J =9.5, 7.4, 5.4 Hz, 1H), 4.24(s, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m,2H).

[0159] When cyclopentylmethylamine was used as the amine, the product was a yellow solid, B7, with a yield of 78%. The chemical formula of B7 is [formula missing]. 1 The H NMR data are as follows:

[0160] ;

[0161] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 8.09(s, 2H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J =8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 3.12(dd, 4H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 1.76(m, 8H), 1.68 – 1.47 (m, 10H).

[0162] When neopentyl amine is used, the product is a yellow solid B8, with a yield of 82%. The chemical formula of B8 is [formula missing]. 1 The H NMR data are as follows:

[0163] ;

[0164] 1H NMR(400 MHz, DMSO) δ 11.44 (s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 8.01(s, 2H), 7.65 (d, J = 8.7 Hz, 2H), 6.98 (s, 3H), 6.64 (d, J =8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 3.06(s, 4H), 2.42 (t, J = 7.4 Hz, 2H), 2.12 – 1.93 (m, 2H), 0.94 (s, 18H).

[0165] When 3-methylbenzylamine is used as the amine, the product is a yellow solid, B9, with a yield of 90%. The chemical formula of B9 is [formula missing]. 1 The HNMR data are as follows:

[0166] ;

[0167] 1H NMR (400 MHz, DMSO) δ 11.44(s, 1H), 8.87(s, 1H), 8.65 (s, 1H), 8.29(d, J = 7.5 Hz, 1H), 8.18(s, 1H), 7.65 (d, J = 8.7 Hz, 2H), 7.51(dt, 2H),7.37(dd, 2H), 7.19(s, 2H), 7.05(dd, 2H), 6.98 (s, 3H), 6.64 (d, J = 8.7 Hz,2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5, 7.4, 5.4 Hz, 1H), 4.34(s,2H), 4.24(s, 2H), 2.42 (t, J = 7.4 Hz, 2H), 2.31(s, 6H), 2.12 – 1.93 (m, 2H).

[0168] The amine used was 2-phenylethylamine, and the product was a yellow solid, B10, with a yield of 81%. The chemical formula of B10 is [formula missing]. 1 The H NMR data are as follows:

[0169] ;

[0170] 1H NMR(400 MHz, DMSO) δ 11.44(s, 1H), 8.65 (s, 1H), 8.29 (d, J = 7.5Hz, 1H), 8.01(s, 2H), 7.65 (d, J = 8.7 Hz, 2H), 7.23 – 7.19(m, 10H), 6.98 (s,3H), 6.64 (d, J = 8.7 Hz, 2H), 4.49 (d, J = 3.9 Hz, 2H), 4.38 (ddd, J = 9.5,7.4, 5.4 Hz, 1H), 3.37(d, 2H), 3.21(d, 2H), 2.74(dt, 4H), 2.42 (t, J = 7.4Hz, 2H), 2.12 – 1.93 (m, 2H).

[0171] Example 3

[0172] Ferrocyte inhibition by folic acid derivatives

[0173] Ferroplasmosis inducer: RSL3; Ferroplasmosis inhibitor (control): Ferrostatin-1 (Fer-1), purchased from Sigma-Aldrich. Folic acid derivatives A1-A32, B1-B10. All reagents were prepared as 10 mM stock solutions using dimethyl sulfoxide (DMSO) and then diluted to the required concentrations using cell culture media as needed.

[0174] Culture conditions for human chorionic trophoblast cells HTR-8 / SVneo: DMEM high-glucose medium (GIBCO) containing 10% FBS (GIBCO), 37℃, 5% CO2 saturated humidity incubator. After cell attachment, DMSO, RSL3 (3 μM, final concentration), RSL3 (3 μM) + folic acid derivative gradient concentrations (1000, 500, 250, 125, 62.5 nM), and RSL3 (3 μM) + Ferrostatin-1 gradient concentrations (1000, 500, 250, 125, 62.5 nM) were added to the medium. HT-1080 cells treated with these drugs for 24 hours were viable using the CCK8 colorimetric assay. The CCK8 kit was purchased from Bebo Biotechnology. The half-maximal effective concentration (EC50) was calculated. 50 (value), the experiment was repeated 3 times.

[0175] The results are shown in Table 1. All folic acid derivatives significantly inhibited ferroptosis, and their inhibitory activity was comparable to that of the positive control drug Fer-1. Among them, the half-maximal effective concentration (IC50) of A2 folic acid diethyl ester was 57.7 nM, which was significantly better than that of Fer-1.

[0176] Table 1. EC5 values ​​of the inhibitory effect of folic acid derivatives on RSL3-induced ferroptosis 50 value

[0177]

[0178] Example 4

[0179] Inhibitory effects of folic acid derivatives on other ferroptosis inducers and their mechanisms of action

[0180] Ferrocyte inducing agents: FIN56 and Sulfasalazine were purchased from Aladdin Reagents, FINO2 from Selleck, ML162, ML210, and DL-Buthionine-(S,R)-sulfoximine (BSO) were purchased from MCE, and Erastin was purchased from Adamas. All reagents and folic acid derivatives were prepared as 10 mM stock solutions with dimethyl sulfoxide (DMSO) and then diluted to the required concentrations using cell culture media as needed.

[0181] Culture conditions for human chorionic trophoblast cells HTR-8 / SVneo: DMEM high-glucose medium (GIBCO) containing 10% FBS (GIBCO), 37 ℃, 5% CO2 saturated humidity incubator.

[0182] HTR-8 / SVneo cells were cultured under the above conditions. After cell attachment, DMSO, an inducer, an inducer + folic acid derivative A2 (300 nM), and an inducer + folic acid derivative B1 (300 nM) were added to the culture medium, respectively. The inducer concentration was determined based on a cell viability of 30-40%. The inducer concentrations selected in this experiment were: 8 μM for FIN56; 250 μM for Sulfasalazine; 20 μM for FINO2; 10 μM for ML162; 90 μM for ML210; 800 μM for DL-Buthionine-(S,R)-sulfoximine (BSO); and 10 μM for Erastin. HT-1080 cells treated with these drugs for 24 hours were viable using the CCK8 colorimetric assay. The CCK8 kit was purchased from Bebo Biotechnology, and the experiment was repeated three times. Figure 1 As shown, folic acid derivatives can effectively inhibit ferroptosis induced by different inducers and improve cell survival.

[0183] HTR-8 / SVneo cells were cultured under the above conditions. After cell adhesion, DMSO, RSL3, and RSL3+ folic acid derivatives (300 nM) were added to the culture medium. After 6 hours of treatment, HTR cells were digested into single-cell suspensions using trypsin. Folic acid derivatives A1, A2, A3, A4, B1, and B2 were selected. Cells were incubated with C11-BODIPY (10 μM) at room temperature in the dark for 30 minutes, washed three times with PBS, and the level of lipid peroxidation in the cell membrane was detected by flow cytometry. Figure 2 As shown, folic acid derivatives can effectively reduce the level of cell membrane lipid peroxidation, block the chain reaction of membrane lipid free radicals, reduce the damaging effect of free radicals on cell membranes, and prevent cell ferroptosis.

[0184] Example 5

[0185] Protective effect of folic acid derivatives against renal ischemia-reperfusion injury

[0186] Male C57BL / 6N mice aged 8 to 12 weeks were fed a standard diet and subjected to renal ischemia-reperfusion injury to establish a mouse model of renal ischemia-reperfusion injury. Bilateral renal stalk clamping was performed via a midline abdominal incision for 36 minutes. Body temperature was maintained between 36 and 37°C throughout the procedure. After clamp removal, the abdomen was closed, allowing restoration of blood flow. Sham-operated mice underwent the same procedure except for the clamping of the kidneys to prevent obstruction. All mice were euthanized 48 hours after reperfusion. All ischemia-reperfusion experiments were performed in a double-blind manner. Folic acid derivative A2 (10 mg / kg) was administered intraperitoneally 1 hour before the onset of ischemia. -1 (or medication). Serum creatinine and urea were measured at the Institute of Clinical Chemistry. Tissue was stained with hematoxylin and eosin (HE), and the morphological features of renal failure were determined by light microscopy.

[0187] Figure 3 , 4 The results showed that, compared with the renal ischemia-reperfusion model treatment group, the necrotic area in the A2 treatment group was significantly reduced, and the serum creatinine and urea nitrogen levels in the A2 treatment group were significantly lower. Figure 3 Therefore, A2 has a significant alleviating effect on acute renal ischemia-reperfusion injury in mice. Figure 4 ).

[0188] Example 6

[0189] Protective effect of diethyl folic acid against non-alcoholic fatty liver disease.

[0190] Forty animals were randomly divided into four groups using a random number table: a control group (n=10) fed a normal diet, an A2 group (n=10) administered A2 (2 mL / kg) by gavage every morning, a high-fat group (n=10) fed a high-fat diet, and a treatment group (n=10) fed a high-fat diet and administered A2 (2 mL / kg) by gavage every morning. All animals underwent liver blood biochemistry and pathological examinations after 8 weeks of feeding.

[0191] Figure 5 , 6 The results showed that, compared with the model group, the necrotic area in the A2 treatment group was significantly reduced, and the serum ALT and AST levels in the A2 treatment group were significantly decreased. Figure 5 Therefore, A2 has a mitigating effect on acute drug-induced liver injury in high-fat rats. Figure 6 ).

[0192] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A folic acid derivative, characterized in that, The folic acid derivative is one or more of the following compounds: 。 2. The pharmaceutically acceptable salt of the folic acid derivative as described in claim 1, characterized in that, The pharmaceutically acceptable salt is one or more of the following: hydrochloride, sulfate, and phosphate.

3. A method for preparing a folic acid derivative according to any one of claims 1-2, characterized in that, Includes the following steps: Folic acid and catalytic acid are dissolved in alcohol or amine and heated under reflux. After the reaction is complete, the reaction system is cooled to room temperature, the solvent is concentrated under reduced pressure, filtered, and the filter cake is washed with cold alcohol or amine. The filter cake is dried under vacuum to obtain crude product. The crude product is recrystallized to obtain folic acid derivatives.

4. The preparation method according to claim 3, characterized in that, The alcohol is one or more of alkyl alcohols, cycloalkyl alcohols, and aryl alcohols.

5. The preparation method according to claim 3, characterized in that, The amine is one or more of alkylamines, cycloalkylamines, and arylamines.

6. The preparation method according to claim 4, characterized in that, The alkyl group of the alkyl alcohol is selected from C1 to C20 alkyl groups, the cycloalkyl group of the cycloalkyl alcohol is selected from an aliphatic cyclic structure with 6 atoms, and the aryl group of the aryl alcohol is selected from alkylphenyl groups.

7. The preparation method according to claim 5, characterized in that, The alkyl group of the alkylamine is selected from C1 to C20 alkyl groups, the cycloalkyl group of the cycloalkylamine is selected from an aliphatic cyclic structure with 6 atoms, and the aryl group of the arylamine is selected from alkylphenyl groups.

8. The preparation method according to claim 3, characterized in that, The catalytic acid is p-benzylbenzenesulfonic acid or sulfuric acid; the molar ratio of folic acid to the catalytic acid is 1:0.99-1.

01.

9. The preparation method according to claim 3, characterized in that, The reflux temperature was 79-81 °C, and the reaction time was 11-13 h.

10. The use of a folic acid derivative in the preparation of a drug that inhibits ferroptosis; wherein the folic acid derivative is selected from the folic acid derivatives described in any one of claims 1-2.

11. The application of a folic acid derivative in the preparation of drugs for preventing and treating diseases related to ferroptosis, characterized in that, The cell ferroptosis-related diseases include one or more of the following: non-alcoholic steatohepatitis, acute liver injury, cardiomyopathy, drug-induced cardiotoxicity, acute kidney injury, atherosclerosis, renal fibrosis, pneumonia, preeclampsia, osteoporosis, osteoarthritis, traumatic brain injury, and neurodegenerative diseases. The neurodegenerative diseases include one or more of the following: Alzheimer's disease, Parkinson's disease, and epilepsy. The folic acid derivative is selected from the folic acid derivatives according to any one of claims 1-2.

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