Atypical tetracycline antibiotic derivatives, processes for their preparation and use thereof

Abstract

This invention provides an atypical tetracycline antibiotic derivative, its preparation method, and its application. The atypical tetracycline antibiotic derivative has the compound structure shown in Formula II: wherein, R 1’ Represents any one of H, amino group and its substituents or their hydrochloride salts, amine group, amide group, sulfonamide group, hydroxyl group and its substituents, and ester group; R 2’ Represents any one of H, alkyl, aryl, acyl, nitroso, amino, and amide. Based on a series of intermediates with structures shown in Formula I, this invention further synthesizes atypical tetracycline derivative structures with multiple site modifications. Cell activity tests show that they have varying degrees of inhibitory effects on *Escherichia coli* and *Staphylococcus aureus*, demonstrating potential for development into antibacterial therapies.

Description

Technical Field

[0001] This invention relates to the fields of organic synthesis and medicinal chemistry, specifically to a method for structural modification and chemical modification of a class of novel atypical tetracyclines, and particularly to an atypical tetracycline antibiotic derivative, its preparation method, and its application. Background Technology

[0002] Pathogenic bacteria are widely distributed across the globe, posing a significant threat to public health. Despite the abundance and widespread use of antibiotics, antibiotic resistance in bacteria is rapidly increasing, potentially leading to serious consequences, compared to infectious diseases caused by viruses and parasites. Therefore, developing novel antibiotics to combat drug-resistant bacterial infections has become an urgent task. Modifying existing antibiotics using chemical methods to give them different mechanisms of action is one of the effective strategies for treating current drug-resistant bacterial infections.

[0003]

[0004] Chelocardin (CHD) is an atypical tetracycline compound with a unique molecular structure. It features acetyl substitution at the C2 position and aromatization of the C ring, exhibiting an overall aromatic character. This molecule was characterized by single-crystal diffraction in 2023 (structure shown above). In the 1970s, CHD was used in a small phase II clinical trial to study urinary tract infections, showing strong inhibitory activity, but the trial was ultimately terminated due to excessive cytotoxicity. Subsequently, as a genetically engineered fermentation derivative of CHD, CDHD (structure shown above) introduced an amide structure at the C2 position, exhibiting a broader spectrum of antibacterial activity. The antibacterial mechanism of CHD differs from that of classic tetracyclines; this atypical tetracycline primarily acts on the cell membrane, causing bacterial cell membrane rupture, thereby achieving its antibacterial effect. Furthermore, CDHD also exhibits the ability to overcome bacterial tetracycline resistance, which may be related to its unique mechanism of action. Therefore, the semi-synthetic modification of CDHD, serving as a lead compound skeleton, provides a new approach to combating tetracycline-resistant bacterial infections. Summary of the Invention

[0005] To address the aforementioned problems, the present invention aims to provide an atypical tetracycline antibiotic derivative and its preparation method, specifically a method for preparing novel antibacterial tetracycline lead compounds from CDCHD skeleton analogs, providing new applications for this type of derivative in the field of antibacterial therapy, and offering certain reference value for the development of atypical tetracyclines with antibacterial activity.

[0006] The objective of this invention is achieved through the following technical solution:

[0007] In a first aspect, the present invention provides an intermediate for an atypical tetracycline antibiotic derivative, the intermediate having a compound structure as shown in Formula I:

[0008]

[0009] Among them, R 1 Represents any one of H, amino group and its substituents, amine group, amide group, sulfonamide group, hydroxyl group and its substituents, and ester group; R 2 It represents any one of H, alkyl, alkenyl, alkynyl, aryl, and acyl groups.

[0010] As a preferred option, in Equation I, R 1 It can be either S-type or R-type.

[0011] As a preferred option, in Equation I, R 1 It represents any one of H, amino and its substituents, hydroxyl and its substituents.

[0012] As a preferred embodiment, the atypical tetracycline antibiotic derivative intermediate comprises the following compound structures:

[0013]

[0014] In Equations I-1 and I-2, R 3 Represents any one of H, Me, Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, or Ms, R 4 Represents H or Me; in Equation I-3, R 2 It represents any one of H, alkenyl, alkynyl, or aryl.

[0015] As a preferred option, in equations I-1 and I-2, NR 3 R 4 OR 3 It can be either S-type or R-type.

[0016] As a preferred embodiment, the R 2 In this compound, the alkenyl group includes any one of vinyl, 1-butenyl, isopropenyl, and cyclohexenyl; the alkynyl group includes any one of ethynyl and 1-butynyl; and the aryl group includes any one of phenyl, p-methylphenyl, p-methoxyphenyl, p-nitrophenyl, p-acetylphenyl, o-methoxyphenyl, o-nitrophenyl, and furan.

[0017] Secondly, this invention provides a method for preparing an intermediate of an atypical tetracycline antibiotic derivative, wherein in the compounds represented by formulas I-1 and I-2, R 3 R 4 When all atoms are H, the preparation method includes the following steps:

[0018] A1. Tetracycline hydrochloride was oxidized by NCS under acidic conditions to obtain a mixture of hemiketals;

[0019] A2. A mixture of hemiketals is reacted with potassium bicarbonate and hydroxylamine hydrochloride to form an oxime, which is then reduced with a reducing agent to obtain R. 3 R 4 All are compounds of formula I-1 with H; or R is obtained by reducing a mixture of hemiketals under conditions of sodium borohydride and cerium chloride heptahydrate. 3 The compound represented by formula I-2 is H;

[0020] In the compounds represented by formulas I-1 and I-2, R 3 For Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, or Ms, and R 4 When the H content is used, the preparation method includes the following steps:

[0021] A3. The R prepared in steps A1-A2 above... 3 R 4 All are compounds of formula I-1 represented by H or R 3 The compound represented by formula 1-2 with H undergoes an acylation reaction with an acylation reagent under basic conditions to give R. 3 For Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, or Ms and R 4 Compounds of formula I-1 and I-2 for H;

[0022] In the compound represented by formula I-1, R 3 R 4 When all components are Me, the preparation method includes the following steps:

[0023] A4. The R prepared in steps A1-A2 above... 3 R 4 The compound represented by formula I-1, which contains H, reacts with paraformaldehyde via reductive amination to obtain R. 3 R 4 All are compounds of formula I-1 represented by Me;

[0024] In the compounds represented by Formula I-3, when R 2 When the H content is used, the preparation method includes the following steps:

[0025] A5. Tetracycline was reacted in the dark with iodomethane and tetrahydrofuran. The resulting intermediate was reduced with a reducing agent to obtain R. 2 The compound represented by formula I-3 is H;

[0026] In the compounds represented by Formula I-3, when R2 When the group is alkenyl, alkynyl, or aryl, the preparation method includes the following steps:

[0027] A6. The R prepared in step A5 above... 2 The compound of formula I-3 with H undergoes a C9 iodination reaction, and the resulting iodinated derivative then undergoes a Stille coupling reaction with a tin reagent to obtain R. 2 Compounds of formula I-3 that are alkenyl, alkynyl or aryl.

[0028] As a preferred embodiment, in step A3, the acylation reagent is selected from at least one of acetyl chloride, ethyl chloroformate, benzyl chloroformate, o-methoxybenzoyl chloride, benzoyl chloride, o-acetoxybenzoyl chloride, and acetic anhydride;

[0029] The alkaline condition is achieved by selecting at least one of potassium carbonate, sodium bicarbonate, pyridine, TMEDA, DBU, and triethylamine.

[0030] In steps A2 and A5, the reducing agent is selected from zinc powder;

[0031] In step A6, the tin reagent is a tin reagent containing aryl, alkynyl, or alkenyl groups; further, the tin reagent is selected from at least one of vinyltri-n-butyltin, 1-butenyltri-n-butyltin, isopropenyltri-n-butyltin, tri-n-butyltin-cyclohexene, ethynyltri-n-butyltin, 1-butynyltri-n-butyltin, phenyltri-n-butyltin, p-methylphenyltri-n-butyltin, p-methoxyphenyltri-n-butyltin, p-nitrophenyltri-n-butyltin, p-acetylphenyltri-n-butyltin, o-methoxyphenyltri-n-butyltin, o-nitrophenyltri-n-butyltin, and furantri-n-butyltin.

[0032] As a preferred embodiment, in step A3, the reaction is carried out under solvent conditions, wherein the solvent is selected from one or a mixture of two of the following solvents: THF, DMF, 1,4-dioxane, dichloromethane, dichloroethane, and water.

[0033] Thirdly, the present invention provides the use of an intermediate of atypical tetracycline antibiotic derivatives in the preparation of atypical tetracycline antibiotic derivatives.

[0034] Fourthly, the present invention provides an atypical tetracycline antibiotic derivative having the compound structure shown in Formula II:

[0035]

[0036] Among them, R 1’ Represents any one of H, amino group and its substituents or their hydrochloride salts, amine group, amide group, sulfonamide group, hydroxyl group and its substituents, and ester group; R 2’It represents any one of H, alkyl, aryl, acyl, nitroso, amino, and amide.

[0037] As a preferred option, in Equation II, R 1’ It can be either S-type or R-type.

[0038] As a preferred embodiment, the atypical tetracycline antibiotic derivative comprises the following compound structures:

[0039]

[0040] In Equations II-2 and II-5, R 3 Represents any one of H, Me, Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, Ms, R 4 Represents H or Me;

[0041] In Equation II-6, R 2’ Represents any one of H, alkyl, aryl, acyl, nitroso, amino, and amide; wherein the amide is selected from acetamido, butyramido, benzamide, o-benzyloxybenzamide, p-nitrobenzamide, maleimide, phthalimide, etc. Any one of them.

[0042] As a preferred embodiment, in formulas II-1, II-2, and II-5, NH2·HCl and NR 3 R 4 OR 3 It can be either S-type or R-type.

[0043] As a preferred option, R in Equation II-6 2’ In this compound, the alkyl group includes any one of ethyl, n-butyl, isopropyl, and cyclohexyl; the aryl group includes any one of phenyl, p-methylphenyl, p-methoxyphenyl, p-nitrophenyl, p-acetylphenyl, o-methoxyphenyl, o-nitrophenyl, and furan; the acyl group includes any one of acetyl and butyryl; and the amino group includes any one of n-butylamino, dimethylamino, n-octylamino, and phenethylamino.

[0044] Fifthly, the present invention provides a method for preparing an atypical tetracycline antibiotic derivative, wherein the method for preparing the compound represented by Formula II-1 includes the following steps:

[0045] B1, R 3 R 4 The compound represented by formula I-1, which contains H, is reacted under concentrated hydrochloric acid conditions to obtain the compound represented by formula II-1.

[0046] In the compound shown in II-2, R 3 R 4When both are H or Me, the preparation method includes the following steps:

[0047] B2. The compound of formula II-1 prepared in step B1 is added to a base for neutralization or reacted with paraformaldehyde via a reducing amination reaction to obtain R. 3 R 4 All are compounds represented by II-2 of H or Me;

[0048] In the compound shown in II-2, R 3 For Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz or CO2Et and R 4 When the H content is used, the preparation method includes the following steps:

[0049] B3, R 3 The compound of formula I-1, consisting of Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, or CO2Et, is reacted with p-toluenesulfonic acid monohydrate or concentrated hydrochloric acid to obtain R. 3 For Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz or CO2Et and R 4 The compound represented by formula II-2 is H;

[0050] In the compound shown in II-2, R 3 For Ac or Ms and R 4 When the H content is used, the preparation method includes the following steps:

[0051] B4. The compound of formula II-1 prepared in step B1 is subjected to an acylation reaction with methanesulfonyl chloride or acetic acid to obtain R. 3 For Ac or Ms and R 4 The compound is shown as II-2 of H;

[0052] The preparation method of the compounds represented by formulas II-3 and II-4 includes the following steps:

[0053] B5. The compound of formula II-1 prepared in step B1 is subjected to an acylation reaction with N-phthalimide or maleic anhydride to obtain the compounds of formula II-3 and II-4.

[0054] The preparation method of the compound shown in II-5 includes the following steps:

[0055] B6. Add the compound shown in formula I-2 to p-toluenesulfonic acid monohydrate or concentrated hydrochloric acid to react and obtain the compound shown in II-5.

[0056] In the compound shown in II-6, R 2’ When the group is H or aryl, the preparation method includes the following steps:

[0057] B7, R 2 The compound of formula I-3 with H or aryl groups is reacted with p-toluenesulfonic acid monohydrate to obtain R. 2’ Compounds represented by II-6 with H or aryl groups;

[0058] In the compound shown in II-6, R 2’ When the alkyl group is used, the preparation method includes the following steps:

[0059] B8, R 2 Hydrogenation of the compound represented by formula I-3 with an alkynyl group yields R. 2’ Compounds shown in II-6 are alkyl groups;

[0060] In the compound shown in II-6, R 2’ When the group is acyl, the preparation method includes the following steps:

[0061] B9, R 2 The compound of formula I-3 with an alkynyl group is reacted with p-toluenesulfonic acid monohydrate to obtain R. 2’ Compounds represented by acyl group II-6;

[0062] In the compound shown in II-6, R 2’ When the group is nitrosyl, the preparation method includes the following steps:

[0063] B10. The R prepared in step B7 above... 2’ The compound shown in II-6 of H is reacted with nitrite to obtain R. 2’ The compound represented by II-6 is nitroso;

[0064] In the compound shown in II-6, R 2’ When the amide is used, the preparation method includes the following steps:

[0065] B11, the R prepared in step B10 above... 2’ The compound shown as II-6, which is nitroso, is reduced with sodium hydrosulfite. After the conversion of the raw materials is complete, a base and an acylation reagent are directly added to react and yield R. 2’ The compound is shown as II-6 of the amide group.

[0066] In the compound shown in II-6, R 2’ When the group is amine, the preparation method includes the following steps:

[0067] B12, the R prepared in step B10 above 2’ The compound shown as II-6, which is nitroso, is reduced with sodium hydrosulfite. After the raw material is completely converted, it is directly reacted with an aldehyde group via a reductive amination reaction to obtain R.2’ The compound shown is II-6, which is an amino group.

[0068] As a preferred embodiment, in step B2, the alkali is selected from either NaOH or MeOH;

[0069] In step B11, the acylation reagent is selected from acetyl chloride, butyryl chloride, benzoyl chloride, o-benzyloxybenzoyl chloride, p-nitrobenzoyl chloride, maleic anhydride, phthalic anhydride, etc. any one of them;

[0070] In step B12, the aldehyde is selected from any one of butyraldehyde, paraformaldehyde, octanaldehyde, and phenylacetaldehyde.

[0071] As a preferred option, the solvent used in each step is selected from one or a mixture of dichloromethane, dichloroethane, tetrahydrofuran, or 1,4-dioxane; when alkaline conditions are required in each step, the base used is potassium carbonate, sodium bicarbonate, pyridine, TMEDA, DBU, triethylamine or other bases.

[0072] Sixthly, the present invention provides the use of an atypical tetracycline antibiotic derivative in the preparation of antibacterial products.

[0073] As a preferred embodiment, the antibacterial product includes at least one of products that inhibit Escherichia coli and products that inhibit Staphylococcus aureus.

[0074] Compared with the prior art, the present invention has the following beneficial effects:

[0075] Based on a series of intermediates with structures shown in Formula I, this invention further synthesizes atypical tetracycline derivative structures with multiple site modifications. Cell activity tests show that they have different degrees of inhibitory effects on Escherichia coli and Staphylococcus aureus, and have the potential to be developed for antibacterial therapy. Detailed Implementation

[0076] The present invention will now be described in detail with reference to specific embodiments. These embodiments will help those skilled in the art to further understand the present invention, but do not limit the invention in any way. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention. These all fall within the scope of protection of the present invention.

[0077] Example 1: Preparation of Compound 3

[0078]

[0079] 10 mL of concentrated hydrochloric acid was added to an open flask containing 1000 mL of water and stirred for 5 minutes. Tetracycline hydrochloride (compound 1, 50 g, 104 mmol) and NCS (35 g, 260 mmol) were added. The reaction mixture was stirred at room temperature for 40 minutes and then filtered to obtain a yellow filter cake. The filter cake was washed with water to obtain a mixture of two hemiketals - compounds 2a / 2b (33.2 g, 78%), which were used directly in the next step of the reaction. The obtained hemiketal mixture was dissolved in 130 mL of methanol, and potassium bicarbonate (63.58 g, 635.92 mmol) and NH2OH·HCl (13.8 g, 198.7 mmol) were added. After stirring for 0.5 h, the mixture was filtered. The filter cake was washed with ethyl acetate. The combined organic phases were washed successively with 3.0 g hydrochloric acid, saturated NH4Cl solution and saturated NaCl solution to obtain a brown crude product. 450 mL of PE / MeOH (v / v = 10:1) was added and the mixture was slurried. After filtration, a yellow powder - compound 3 (27.0 g, 79%) was obtained.

[0080] Compound 3: 1 H NMR (400MHz, DMSO-d6) δ18.21(s,1H),15.22(s,1H),12.98(s,1H),11.85(s,1H),8.85(s,2H),7.52(t,J=8.0Hz,1H),7.08(d,J=7.7Hz,1H), 6.98-6.86(m,2H),4.96(s,1H),3.76-3.68(m,1H),2.83(dd,J=11.4,5.2Hz,1H),2.00-1.96(m,1H),1.67(q,J=12.7Hz,1H),1.47(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ193.2,178.2,176.4,173.6,161.6,148.1,136.6,117.1,115.3,114.7,106.7,98.7,74.8,68.1,42.0,37.5,23.3,22.5ppm.

[0081] Example 2 Preparation of compounds 4 and 4'

[0082]

[0083] Compound 3 (26 g, 60.4 mmol) and zinc powder (10.2 g, 15.6 mmol) were added to an open flask containing 250 mL of acetic acid / water (v / v 1:1). The mixture was stirred at room temperature for 5 h, filtered through a diatomaceous earth liner, and concentrated. The crude product was subjected to C18 reversed-phase column chromatography (water / acetonitrile = 9:1) (the aqueous phase contained 0.1% acetic acid) to give the reduced product – compound 4 (14.5 g, 58%) and compound 4' (0.26 g, 1%, LC purity 90%). Compound 4: 1 H NMR (400MHz, DMSO-d6) δ12.12(s,1H),8.81(d,J=74.0Hz,2H),7.49(t,J=8.0Hz,1H),7.07(d,J=7.8Hz,1H),6.87(d,J=8.2Hz,1H),6.43(br,1H),4. 88(s,1H),4.23(d,J=4.6Hz,1H),2.62-2.57(m,1H),2.42(dd,J=14.1,4. 9Hz, 1H), 2.10 (d, J = 12.9Hz, 1H), 1.60 (d, J = 11.9Hz, 1H), 1.49 (s, 3H) ppm. 13 C NMR(101MHz,DMSO-d6)δ189.3,179.3,170.9,170.5,161.6,148.1,136.0,11 6.9,115.2,114.9,105.0,99.3,81.8,75.3,68.2,52.7,41.9,22.7,19.2ppm.

[0084] Compound 4': 1 H NMR (400MHz, DMSO-d6) δ15.17(s,1H),11.80(s,1H),9.40(d,J=67.3Hz,1H),8.38(s,3H),7.56(t,J=8.0Hz,1H),7.14(d,J=7.6Hz,1H),6.9 3(d,J=8.3Hz,1H),5.06(s,1H),3.97(s,1H),2.86-2.82(m,1H),2.21-2.17(m,1H),2.03-1.97(m,1H),1.80(s,1H),1.26-1.22(m,3H)ppm.

[0085] Example 3 Preparation of Compound 5

[0086]

[0087] Compound 4 (5.4 g, 12.9 mmol) was dissolved in 20 mL of tetrahydrofuran, 4 mL of concentrated hydrochloric acid was added, and the mixture was stirred at room temperature for 6 hours. The mixture was extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and subjected to reversed-phase column chromatography (water / acetonitrile = 4:1-3:1) to give aromatic compound 5 (2.70 g, 48%).

[0088] Compound 5: 1 H NMR (400MHz, DMSO-d6) δ16.57(br,1H),15.26(br,1H),10.06(s,1H),9.58(s,1H),9.35(s,1H),8.87(s,2H),7.60(t,J=8.2Hz,1H),7.43( d,J=8.4Hz,1H),6.90(d,J=7.8Hz,1H),4.70(s,1H),3.51-3.47(m,2H),3.24-3.18(m,1H),2.74(dd,J=17.8,12.7Hz,1H),2.36(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ189.3,172.5,138.8,132.8,130.6,114.6,112.6,110.8,108.7,96.9,76.9,52.3,40.4,25.6,14.1ppm.

[0089] Example 4 Preparation of compound 5a

[0090]

[0091] Compound 5 (50 mg, 0.12 mmol) was dissolved in 0.5 mL of tetrahydrofuran, and 0.2 mL of 1.0 M NaOH solution was added. The mixture was stirred at room temperature for 1.5 hours, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and subjected to reversed-phase column chromatography (water / acetonitrile = 4:1-3:1) to give aromatic compound 5a (21.3 mg, 47%).

[0092] Compound 5a: 1 H NMR (500MHz, DMSO-d6) δ17.41(br,1H),9.25(s,1H),7.21(t,J=8.1Hz,1H),6.86(d,J=8.4Hz,1H),6.31(d,J=7.8Hz,1H ),6.27(br,1H),5.70(br,1H),4.06(s,1H),3.19-3.17(m,1H),2.45-2.38(m,1H),2.24-2.17(m,1H),2.11(s,3H)ppm.13 CNMR(101MHz,DMSO-d6)δ195.6,170.9,162.6,145.4,138.2,120.0,115.8,115 .0,113.1,87.5,79.8,79.4,79.1,68.5,62.4,52.0,41.9,23.0,22.0,17.0ppm.

[0093] Example 5 Preparation of compound 6a

[0094]

[0095] Compound 4 (100 mg, 0.24 mmol) and sodium bicarbonate (30 mg, 0.36 mmol) were dissolved in a mixed solvent of THF / H2O (4:1). Ethyl chloroformate (34 μL, 0.36 mmol) was added dropwise at 0 °C. After stirring for 30 minutes, the mixture was quenched with water, extracted with ethyl acetate, washed successively with saturated ammonium chloride and saturated brine, dried, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 7:3) to give compound 6a (56.9 mg, 48%).

[0096] Compound 6a: 1 H NMR(400MHz,DMSO-d6)δ17.91(s,1H),15.31(s,1H),11.86(s,1H),9.51–9.22(m,1H),9 .12(s,1H),7.64(s,1H),7.52(t,J=8.0Hz,1H),7.09(d,J=7.7Hz,1H),6.90(d,J=8.4Hz ,1H),5.18–5.04(m,1H),4.90(s,1H),4.11–4.02(m,2H),2.67(dd,J=11.3,5.1Hz,1H), 2.32(d,J=11.9Hz,1H),2.21(d,J=13.4Hz,1H),1.50(s,3H),1.21(t,J=7.2Hz,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ174.0,161.9,157.5,148.7,136.9,117.4,115.8,115.0,96.2,68.6,60.7,42.0,23.0,20.0,15.1ppm.

[0097] Example 6 Preparation of compound 6b

[0098]

[0099] Compound 4 (100 mg, 0.24 mmol), CbzCl (40 μL, 0.27 mmol), and sodium bicarbonate (30 mg, 0.35 mmol) were dissolved in 5 mL of THF / H2O (v / v 4:1) mixed solvent. CbzCl (40 μL, 0.27 mmol) was added dropwise at 0 °C. After stirring for 30 min, the mixture was quenched with water, extracted with ethyl acetate, washed successively with saturated ammonium chloride and saturated brine, dried, filtered, and concentrated. The crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give compound 6b (55.2 mg, 42%).

[0100] Compound 6b: 1 H NMR(400MHz,DMSO-d6)δ17.85(br,1H),15.30(br,1H),11.87(s,1H),9.35(s,1H ),9.11(s,1H),7.77(s,1H),7.52(t,J=8.0Hz,1H),7.42-7.32(m,5H),7.08(d,J =7.7Hz,1H),6.90(d,J=8.3Hz,1H),5.15-5.03(m,3H),4.89(s,1H),2.71-2.65( m,1H),2.39-2.32(m,1H),2.23-2.20(m,1H),1.49(s,3H),1.45-1.41(m,1H)ppm. 13 C NMR(101MHz,DMSO-d6)δ193.0,173.5,161.5,156.9,148.3,136.9,136.4,128.4, 128.1,128.0,116.9,115.3,114.6,95.7,68.1,65.9,42.6,41.6,22.5,19.6ppm.

[0101] Example 7 Preparation of compound 6c

[0102]

[0103] Compound 4 (100 mg, 0.24 mmol) was dissolved in 3 mL of DCM. TMEDA (380 μL, 2.40 mmol) and B2Cl (56 μL, 0.48 mmol) were added at 0 °C. After stirring for half an hour, the mixture was quenched with water, extracted twice with ethyl acetate, and the organic phases were combined. The mixture was washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give compound 6c (54.3 mg, 43%).

[0104] Compound 6c: 1H NMR(400MHz,DMSO-d6)δ18.07(br,1H),15.32(s,1H),11.87(s,1H),9.46-9.14(m,2H) ,8.76(br,1H),8.00-7.95(m,2H),7.59-7.54(m,1H),7.54-7.48(m,3H),7.08(d,J=7. 7Hz,1H),6.90(d,J=8.3Hz,1H),5.74(br,1H),4.92(s,1H),2.69(dd,J=11.3,5.1Hz,1 H),2.48-2.42(m,1H),2.30(d,J=14.0Hz,1H),1.61(q,J=13.5Hz,1H),1.53(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ194.2,173.6,167.0,161.5,148.3,136.4,134.1,131 .5,128.3,127.7,116.9,115.3,114.5,96.1,68.1,42.4,41.6,22.6,20.0ppm.

[0105] Example 8 Preparation of compound 6d

[0106]

[0107] Compound 4 (150 mg, 0.36 mmol) and sodium bicarbonate (45 mg, 0.54 mmol) were dissolved in 5 mL of a mixed solvent of THF / H2O (4:1). o-methoxybenzoyl chloride (61 μL, 0.43 mmol) was added dropwise under an ice-water bath. The other steps were the same as those for the preparation of compound 6a, to obtain a yellow solid compound 6d (92.0 mg, 46%).

[0108] Compound 6d: 1H NMR(400MHz,DMSO-d6)δ17.84(s,1H),15.33(s,1H),11.86(s,1H),9.38(s,1H),9.22(s,1H),8.65(d, J=7.7Hz,1H),7.82(d,J=7.6Hz,1H),7.52(t,J=7.9Hz,2H),7.20(d,J=8.4Hz,1H),7.07(dt,J=7.5,3.7 Hz,2H),6.98(br,1H),6.90(d,J=8.4Hz,1H),5.56(br,1H),4.96(s,1H),3.94(s,3H),2.70(dd,J=11.4 ,5.2Hz,1H),2.65-2.62(m,1H),2.06(d,J=12.5Hz,1H),1.60(q,J=12.5,12.1Hz,1H),1.49(s,3H)ppm.

[0109] Example 9 Preparation of compound 6e

[0110]

[0111] Compound 4 (200 mg, 0.48 mmol) and paraformaldehyde (288 mg, 9.6 mmol) were dissolved in a mixed solvent of MeOH (2 mL). Sodium cyanoborohydride (605 mg, 9.6 mmol) was added to the mixture at room temperature. After 2 h, the mixture was directly subjected to reversed-phase column chromatography (water / acetonitrile = 4:1) to give a yellow solid compound 6e (64.8 mg, 30%).

[0112] Compound 6e: 1 H NMR (400MHz, DMSO-d6) δ11.89(s,1H),9.37(br,2H),7.53(t,J=8.0Hz,1H),7.09(d,J=7.8Hz,1H),6.91(d,J =8.3Hz,1H),6.71(br,1H),4.95(s,1H),4.02(s,1H),2.85-2.72(m,1H),2.76(s,6H),1.58-1.41(m,4H)ppm. 13 C NMR(101MHz,DMSO-d6)δ193.0,174.6,172.0,161.6,149.3,135.7,117.2,11 5.4,113.6,106.0,97.4,80.1,75.8,69.3,36.5,45.5,45.3,22.1,22.0ppm.

[0113] Example 10 Preparation of compound 6a'

[0114]

[0115] A mixture of compounds 4 and 4' (130 mg, 0.31 mmol, ratio of compounds 4 and 4': 1:1) and sodium bicarbonate (39.4 mg, 0.46 mmol) was dissolved in a THF / H2O (4:1) mixed solvent. Ethyl chloroformate (36 μL, 0.37 mmol) was added dropwise under an ice-water bath. The other steps were the same as those for the preparation of compound 6a, to obtain a yellow solid compound 6a' (19.9 mg, 13%).

[0116] Compound 6a': 1 H NMR(400MHz,DMSO-d6)δ18.29(br,1H),15.21(br,1H),11.86(s,1H),9.09(s,2H),7.53( t,J=8.0Hz,1H),7.25(d,J=29.5Hz,1H),7.10(d,J=7.8Hz,1H),6.90(d,J=8.5Hz,1H),6.7 8(br,1H),4.93(s,1H),4.23(s,1H),4.01(q,J=7.1Hz,2H),2.84(t,J=8.2Hz,1H),2.42- 2.32(m,1H)2.03-1.97(m,1H),1.51(s,3H),1.48-1.41(m,1H),1.16(t,J=7.1Hz,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ193.3,173.9,161.9,156.1,148.6,137.0,117.4,115.9,114.9,68.6,60.9,49.1,23.1,15.0ppm.

[0117] Example 11 Preparation of compound 7a

[0118]

[0119] Compound 6a (35 mg, 0.07 mmol) was dissolved in a mixed solution of 1.0 mL tetrahydrofuran and 0.2 mL concentrated hydrochloric acid. After stirring at room temperature for 8 hours, the reaction solution was concentrated. The crude product was eluted by reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 7a (29.7 mg, 88%).

[0120] Compound 7a: 1H NMR(400MHz,DMSO-d6)δ17.66(br,1H),15.33(br,1H),10.07(br,1H),9.21(br,2 H),7.77(d,J=9.6Hz,1H),7.57(t,J=8.1Hz,1H),7.39(d,J=8.5Hz,1H),7.18(br,1 H),6.85(d,J=7.8Hz,1H),5.21(s,1H),4.08(qt,J=7.2,3.6Hz,2H),3.47-3.39(m, 1H),2.83-2.77(m,1H),2.67-2.59(m,1H),2.35(s,3H),1.23(t,J=7.1Hz,3H)ppm. 13 CNMR(101MHz,DMSO-d6)δ173.1,157.2,138.8,132.7,131.6,110.9,109.0,97.5,79.3,79.0,60.3,41.9,26.1,22.6,14.7,14.1ppm.

[0121] Example 12 Preparation of compound 7b

[0122]

[0123] Using compound 6b (40.1 mg, 0.072 mmol) as a starting material, and following the preparation method of compound 7a, a yellow solid compound 7b (32.1 mg, 84%) was obtained.

[0124] Compound 7b: 1 H NMR (400MHz, DMSO-d6) δ17.68(br,1H),9.20(s,2H),7.93(d,J=10.0Hz,1H),7.55(t,J=7.8Hz,1H),7.43-7.33(m,6H),7.14(br, 1H),6.82(d,J=7.7Hz,1H),5.22(br,1H),5.17-5.07(m,2H),3.43(s,1H),2.85-2.77(m,1H),2.68-2.58(m,1H),2.32(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ173.1,157.1,138.9,136.9,132.8,131.5,128.5,128.1,128.1,112.2,111.0,109.0,97.5,66.0,42.0,26.1,14.1ppm.

[0125] Example 13 Preparation of compound 7c

[0126]

[0127] Compound 6c (30 mg, 0.057 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (44 mg, 0.231 mmol) was added. After stirring at room temperature for 30 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 7c (18 mg, 62%).

[0128] Compound 7c: 1 H NMR (400MHz, DMSO-d6) δ17.70(br,1H),9.19(s,1H),8.89(d,J=8.7Hz,1H),7.97(d,J=7.5Hz,2H),7.60-7.50(m,4H),7 .35(br,1H),7.12(br,1H),6.81(d,J=7.4Hz,1H),5.78(s,1H),3.46-3.42(m,1H),2.88-2.77(m,2H),2.35(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ173.1,167.1,138.7,134.1,132.7,131.6,131.5,128 .3,127.6,127.2,114.9,112.1,110.8,97.8,79.3,78.9,41.6,26.4,14.1ppm.

[0129] Example 14 Preparation of compound 7d

[0130]

[0131] Compound 6d (52.0 mg, 0.094 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (71.9 mg, 0.378 mmol) was added. The mixture was stirred at 40 °C for 14 hours. The reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 7d (40.5 mg, 81%).

[0132] Compound 7d: 1H NMR(400MHz, DMSO-d6)δ8.71(d,J=7.9Hz,1H),7.80(dd,J=7.6,1.8Hz,1H),7.55-7.51(m,2H),7.31(br,1H),7.21(d,J=8.4Hz,1H), 7.08(t,J=7.4Hz,1H),6.79(br,1H),5.63(br,1H),3.97(s,3H),3.28(m,1H),3.08-2.98(m,1H),2.86-2.82(m,1H),2.31(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ173.2,171.6,165.6,157.2,138.8,132.7,131.5,130.6, 122.8,120.7,114.7,112.3,110.8,109.0,97.8,56.2,41.1,25.7,22.5,14.0ppm.

[0133] Example 15 Preparation of compound 7a'

[0134]

[0135] Using a mixture of compounds 6a and 6a' (45.0 mg, liquid ratio: 1:7, 0.092 mmol) as raw materials, and following the preparation method of compound 7a, a yellow solid compound 7a' (16.1 mg, 37%) was obtained.

[0136] Compound 7a': 1 H NMR(400MHz,DMSO-d6)δ18.09(br,1H),14.83(br,1H),10.10(br,1H),9.41(s,1H),9.19(s,1H),7.60-7.56(m,1H),7.45(br,2H),6.86(br,1H), 4.15-4.00(m,1H),3.96-3.92(m,2H),3.23-3.16(m,1H),3.05(d,J=15. 9Hz, 1H), 2.84 (d, J = 12.3Hz, 1H), 2.33 (s, 3H), 1.14 (t, J = 7.1Hz, 3H) ppm. 13 C NMR (101MHz, DMSO-d6) δ173.1,171.6,156.6,139.3,132.8,130.9,118.2,111.0,108.4,101.0,76.5,60.2,42.0,24.1,22.6,14.6,14.1ppm.

[0137] Example 16 Preparation of compound 7e

[0138]

[0139] Compound 5 (100 mg, 0.20 mmol) was dissolved in 0.5 mL of tetrahydrofuran, and 0.5 mL of DIPEA and ethyl N-phthalimide (64.5 mg, 0.30 mmol) were added. After stirring at room temperature for 16 hours, the reaction solution was quenched with water, extracted three times with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give the yellow solid product compound 7e (34.4 mg, 32%).

[0140] Compound 7e: 1 H NMR(500MHz,DMSO-d6)δ17.12(br,1H),15.31(br,1H),10.23(br,1H),9.39(s,1H),9.24(br,1H),7.99(q,J=3.6Hz,2H),7.9 6-7.89(m,2H),7.55(t,J=8.1Hz,1H),7.45(br,1H),7.37(d,J=8.5Hz,1H),6.84(d,J=7.7Hz,1H),5.76(s,1H),3.43-3.41(m overlapped with water,1H),3.17-3.01(m,2H),2.24(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ190.0,172.9,168.2,167.7,163.1,138.8,135.0,134.8,1 32.7,131.8,131.6,131.3,123.5,114.8,110.8,108.9,97.5,42.1,27.2,14.0ppm.

[0141] Example 17 Preparation of compound 7f

[0142]

[0143] Compound 5 (30 mg, 0.069 mmol) was dissolved in 2.0 mL of acetic acid, and maleic anhydride (20.3 mg, 0.20 mmol) was added at room temperature. The reaction mixture was stirred at 100 °C for 10 hours, quenched with water, extracted with ethyl acetate, and the organic phase was washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 7f (15.1 mg, 45%).

[0144] Compound 7f: 1 H NMR (400MHz, DMSO-d6) δ17.43(br,1H),9.31-9.21(m,3H),7.57(t,J=8.1Hz,1H),7.39(d,J=8.3Hz,1H),7.19(s,1H),6.85( d,J=7.8Hz,1H),6.57(d,J=12.3Hz,1H),6.32(d,J=12.2Hz,1H),5.54(dd,J=8.9,4.4Hz,1H),3.44-3.41(m,1H,overlapped with water), 2.89 (dt, J=12.7, 4.9Hz, 1H), 2.69 (dd, J=17.7, 12.6Hz, 1H), 2.34 (s, 3H), 1.26-1.20 (m, 1H)ppm. 13 C NMR(101MHz,DMSO-d6)δ173.0,166.3,165.4,158.1,138.8,132.7,131.4, 131.3,114.8,112.3,110.8,108.9,97.7,77.8,51.3,41.2,25.9,14.0ppm.

[0145] Example 18 Preparation of 7g of compound

[0146]

[0147] Compound 5 (200 mg, 0.46 mmol) and sodium bicarbonate (270 mg, 3.2 mmol) were dissolved in a mixed solvent of THF / H2O (6.0 mL, v / v 4:1), followed by the addition of methanesulfonyl chloride (130 μL, 1.68 mmol). The mixture was stirred at room temperature for 10 hours, and the remaining steps were performed as described for compound 6a. The mixture was then subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give 7 g (25.2 mg, 11%) of a yellow solid.

[0148] Compound 7g: 1H NMR (500MHz, DMSO-d6) δ17.41(br,1H),15.29(br,1H),10.05(br,1H),9.30(br,2H),8.02(d,J=9.3Hz,1H),7.58(t,J=8.1Hz,1H),7.40(d,J=8.5Hz,1H), 7.25(br,1H),6.86(d,J=7.8Hz,1H),4.92(dd,J=9.5,4.4Hz,1H),3.45-3.42 (m,1H),3.10(s,3H),2.88-2.83(m,1H),2.66-2.59(m,1H),2.35(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ173.0,157.9,138.8,132.8,131.4,114.9,110.9,109.0,97.5,78.1,43.3,42.1,25.8,22.5,14.0ppm.

[0149] Example 19 Preparation of Compound 7h

[0150]

[0151] Compound 5 (400 mg, 0.92 mmol), acetic acid (105 μL, 1.84 mmol), HATU (700 mg, 1.84 mmol), and HOBt (124 mg, 0.92 mmol) were dissolved in 10 mL of tetrahydrofuran. The mixture was stirred at room temperature for 5 hours. After conversion, the pH of the reaction solution was adjusted to 3-4, and the reaction solution was extracted twice with ethyl acetate. The organic phases were combined, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to reversed-phase column chromatography (water / acetonitrile = 1:1) to give a yellow solid compound 7h (232 mg, 57%). Compound 7h: 1 H NMR (400MHz, DMSO-d6) δ17.72(s,1H),15.35(s,1H),10.07(s,1H),9.19(s,2H),8.42(d,J=9.2Hz,1H),7.58(t,J=8.0Hz,1H),7. 41(d,J=8.4Hz,1H),7.15(s,1H),6.86(d,J=7.9Hz,1H),5.52(d,J=8.4Hz,1H),2.80-2.62(m,3H),2.36(s,3H),1.99(s,3H)ppm. 13C NMR (101MHz, DMSO-d6) δ173.2,169.9,157.9,138.8,132.7,131.5,115.3,110.9,108.9,97.8,41.6,26.0,22.7,14.1ppm.

[0152] Example 20 Preparation of compound 7i

[0153]

[0154] Compound 5 (100 mg, 0.23 mmol) and sodium cyanoborohydride (288 mg, 4.6 mmol) were dissolved in 20 mL of methanol, and paraformaldehyde (320 μL, 4.6 mmol) was added. The mixture was stirred at room temperature for 5 hours. After the conversion was complete, the reaction system was diluted with water, and the reaction solution was extracted twice with ethyl acetate. The organic phases were combined, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to reversed-phase column chromatography (water / acetonitrile = 2:3) to give a yellow solid compound 7i (56.9 mg, 58%).

[0155] Compound 7i: 1 H NMR (400MHz, DMSO-d6) δ9.04(s,1H),8.74(s,1H),7.47(t,J=8.0Hz,1H),7.25(d,J=8.5Hz,1H),6.89(s,1H), 6.70(d,J=7.8Hz,1H),4.48(s,1H),3.08–3.03(m,1H),2.81(s,6H),2.39(d,J=11.7Hz,2H),2.26(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ188.2,172.6,158.0,139.1,138.9,132.9,130.7,115.0,112.2,111.1,108.8,76.9,66.2,41.8,34.1,26.4,14.3ppm.

[0156] Example 21 Preparation of compounds 8 and 8'

[0157]

[0158] Tetracycline hydrochloride (compound 1) was used as the starting material and oxidized by NCS to obtain a mixture of hemiacetals (the preparation method is the same as in Example 1). Subsequently, the hemiacetals were reduced by the reported method (Tetrahedron Lett. 2023, 133, 154829.) to obtain 4-OH type tetracycline derivatives - compounds 8 (20%) and 8' (21%).

[0159] Compound 8: 1 H NMR(400MHz,DMSO-d6)δ18.29(br,1H),15.34(s,1H),11.84(s,1H),9.05(br, 2H),7.53(t,J=8.0Hz,1H),7.09(d,J=7.6Hz,1H),6.91(d,J=8.0Hz,1H),6.76 (br,1H),5.88(br,1H),4.92(s,1H),4.73(br,1H),2.69(dd,J=11.2,5.0Hz,1 H),2.46-2.41(m,1H),2.18-2.14(m,1H),1.51-1.35(m,1H),1.48(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ173.7,161.5,148.2,136.5,117.0,115.3,114.6,95.4,68.2,43.4,41.7,22.5,18.5.

[0160] Compound 8': 1 H NMR(400MHz,DMSO-d6)δ18.29(br,1H),15.17(s,1H),11.82(s,1H),8.98(br,2 H),7.53(t,J=8.0Hz,1H),7.11(d,J=7.6Hz,1H),6.91(d,J=8.3Hz,1H),6.75(s ,1H),5.75(d,J=7.0Hz,1H),4.96(s,1H),4.06(br,1H),2.85(dd,J=9.5,6.7Hz ,1H),2.40-2.34(m,1H),2.06-1.99(m,1H),1.80-1.70(m,1H),1.51(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ193.7,192.8,173.6,161.4,148.1,136.5,117.0,115.5,114.5,106.1,68.2,42.3,41.0,22.8,22.1ppm.

[0161] Example 22 Preparation of compounds 9 and 9'

[0162]

[0163] Compound 8 (5.0 g, 12.0 mmol) was dissolved in 15 mL of tetrahydrofuran, and p-TsOH·H2O (11.4 g, 60.0 mmol) was added. The mixture was stirred at 40 °C for 12 hours, concentrated under reduced pressure, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 2:3) to give compound 9 (4.3 g, 90%) as a yellow solid.

[0164] Compound 9: 1 H NMR (400MHz, DMSO-d6) δ18.01(br,1H),15.38(br,1H),10.02(br,1H),9.09(br,2H),7.58(t,J=8.2Hz,1H),7.42(d,J=8.4Hz,1H), 7.01(br,1H),6.86(d,J=7.8Hz,1H),5.99(s,1H),4.77(s,1H),2.95-2.88(m,1H),2.62(dd,J=18.2,12.0Hz,2H),2.32(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ201.2,196.9,190.5,173.3,162.8,157.8,138.7,132.7, 131.9,121.5,115.0,112.1,110.8,109.1,97.4,78.3,67.9,42.8,25.0,14.0ppm.

[0165] Compound 8' (736 mg, 1.8 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and p-TsOH·H2O (1.70 g, 9.0 mmol) was added. The mixture was stirred at 40 °C for 12 hours, concentrated under reduced pressure, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 2:3) to give a yellow solid compound 9' (203 mg, 29%).

[0166] Compound 9': 1 H NMR (400MHz, DMSO-d6) δ18.23(br,1H),14.87(br,1H),10.04(br,1H),9.39(s,1H),9.15(s,1H),7.58(t,J=8.2Hz,1H),7.42(d,J=8.4Hz,1H),6.90(br ,1H),6.86(d,J=7.7Hz,1H),5.90(br,1H),4.00(d,J=11.5Hz,1H),3.38-3. 32(m,1H),3.20(dd,J=17.2,5.1Hz,1H),2.70-2.65(m,1H),2.39(s,3H)ppm. 13C NMR(176MHz,DMSO-d6)δ198.7,173.2,164.9,158.0,139.2,132.7,131.1,129 .8,121.8,114.9,112.2,110.9,108.7,97.7,76.4,68.1,44.6,23.4,14.2ppm.

[0167] Example 23 Preparation of compound 10a

[0168]

[0169] Using compound 8 (200 mg, 0.48 mmol) and acetyl chloride as raw materials, compound 10a (103.7 mg, 47%) was obtained as a yellow solid by the method described below for preparing compound 10b.

[0170] Compound 10a: 1 H NMR (400MHz, DMSO-d6) δ17.50(s,1H),15.31(s,1H),11.81(s,1H),9.35(s,1H),9.28(s,1H),7.53(t,J=8.0Hz,1H),7.10(d,J=7.7Hz,2H),6.92(d, J=8.3Hz,1H),6.01(br,1H),4.96(s,1H),2.76(dd,J=11.3,5.3Hz,1H),2 .57(ddd,J=13.9,5.2,2.7Hz,1H),2.19(s,3H),2.13-2.06(m,1H),1.64(q like,J=11.9,11.0Hz,1H),1.49(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ193.2,173.4,169.7,161.5,148.1,136.6,117.1,115.4,114.6,106.7,95.2,68.1,41.4,41.3,22.5,20.7,19.4ppm.

[0171] Example 24 Preparation of compound 10b

[0172]

[0173] Compound 8 (200 mg, 0.48 mmol) was dissolved in 2.0 mL of DCM. Pyridine (190 μL, 2.4 mmol) was added at 0 °C and stirred for 5 minutes. Then, ethyl chloroformate (114 μL, 1.2 mmol) was added dropwise. After stirring at room temperature for 24 hours, the mixture was quenched with water, extracted with ethyl acetate, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 10b (72.6 mg, 31%).

[0174] Compound 10b: 1 H NMR (400MHz, DMSO-d6) δ17.35(br,1H),15.31(br,1H),11.80(s,1H),9.37(s,1H),9.34(br,1H),7.53(t,J=8.0Hz,1H),7.10(d,J=7.6Hz,2H),6. 92(d,J=8.3Hz,1H),5.79(br,1H),4.97(s,1H),4.22(q,J=7.0Hz,2H),2. 78(dd,J=11.3,5.3Hz,1H),2.70-2.64(m,1H),2.06-2.00(m,1H),1.63(q like,J=12.4,12.0Hz,1H),1.48(s,3H),1.28(t,J=7.1Hz,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ193.3,173.3,161.5,154.3,148.1,136.6,117.1,11 5.3,114.6,106.8,94.9,74.4,73.1,68.1,64.2,41.3,22.5,19.3,14.1ppm.

[0175] Example 25 Preparation of compound 10c

[0176]

[0177] Using compound 8 (200 mg, 0.48 mmol) and o-methoxybenzoyl chloride as raw materials, and following the preparation method of compound 10b, a yellow solid compound 10c (121.3 mg, 46%) was obtained.

[0178] Compound 10c: 1H NMR(400MHz,DMSO-d6)δ17.49(br,1H),15.33(br,1H),11.82(s,1H),9.36(s,1H),9.27(br,1H),7.84( dd,J=7.8,1.8Hz,1H),7.62-7.58(m,1H),7.53(t,J=8.0Hz,1H),7.19(d,J=8.4Hz,1H),7.11-7.06(m,3 H),6.91(d,J=8.3Hz,1H),6.22(br,1H),5.00(s,1H),3.84(s,3H),2.81(dd,J=11.3,5.3Hz,1H),δ2.67 (ddd,J=13.7,5.3,2.7Hz,1H),2.26(ddd,J=13.2,5.4,2.8Hz,1H),1.72-1.66(m,1H),1.52(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ193.2,164.8,161.5,158.9,148.1,136.6,134.4,131 .6,120.3,117.1,115.4,114.6,112.9,68.1,55.9,41.6,41.4,22.6,19.6ppm.

[0179] Example 26 Preparation of compound 10d

[0180]

[0181] Using compound 8 (200 mg, 0.48 mmol) and benzoyl chloride as raw materials, and following the preparation method of compound 10b, a yellow solid compound 10d (121.3 mg, 43%) was obtained.

[0182] Compound 10d: 1 H NMR(400MHz,DMSO-d6)δ17.49(br,1H),15.34(s,1H),11.83(s,1H),9.38(s,1H),9. 33(s,1H),8.08(d,J=23.4Hz,2H),7.72(t,J=7.4Hz,1H),7.61-7.52(m,4H),7.11(d, J=7.7Hz,1H),6.92(d,J=8.3Hz,1H),6.29(br,1H),4.99(br,1H),2.82(dd,J=11.3,5 .3Hz,1H),2.78-2.73(m,1H),2.32-2.27(m,1H),1.83-1.73(m,1H),1.56(s,3H)ppm.13 C NMR(101MHz,DMSO-d6)δ193.2,173.3,165.0,161.5,148.1,136.6,133.8,1 29.6,129.3,129.0,117.1,115.4,114.6,95.1,68.1,41.4,22.6,19.6ppm.

[0183] Example 27 Preparation of compound 10e

[0184]

[0185] Compound 8 (5.0 g, 12 mmol) was dissolved in 70 mL of DCM. Pyridine (19 mL, 240 mmol) was added at 0 °C and stirred for 5 minutes. Then o-acetoxybenzoyl chloride (4.70 g, 24.0 mmol) was added dropwise. After stirring at 40 °C for 2.5 hours, the mixture was quenched with water, extracted with ethyl acetate, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 10e (2.7 g, 39%).

[0186] Compound 10e: 1 H NMR(400MHz,DMSO-d6)δ17.48(br,1H),15.36(s,1H),11.85(s,1H),9.41(br,1H),9.38(br,1H),8.06(d ,J=6.8Hz,1H),7.80-7.71(m,1H),7.55(t,J=8.0Hz,1H),7.50(t,J=7.6Hz,1H),7.32(d,J=8.0Hz,1H),7. 17(s,1H),7.12(d,J=7.7Hz,1H),6.94(d,J=8.3Hz,1H),6.25(br,1H),5.02(s,1H),2.81(dd,J=11.2,5.2 Hz,1H),2.72(d,J=11.5Hz,1H),2.27(s,3H),2.25-2.18(m,1H),1.76(q,J=13.0Hz,1H),1.54(s,3H)ppm. 13CNMR(101MHz,DMSO-d6)δ193.2,173.3,169.1,163.4,161.5,150.1,148.0,136.6,134.5,131.5,12 6.5,124.3,123.0,117.1,115.3,114.5,106.7,99.5,95.0,74.4,68.1,41.3,22.5,20.8,19.4ppm.

[0187] Example 28 Preparation of compound 10a'

[0188]

[0189] Compound 8' (200 mg, 0.48 mmol) was dissolved in 2.0 mL of dichloromethane. Pyridine (0.2 mL, 2.4 mmol) and acetic anhydride (90 μL, 0.96 mmol) were added at 0 °C. After stirring at room temperature for 2.5 hours, the mixture was quenched with water, extracted with ethyl acetate, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 10a' (30.7 mg, 14%).

[0190] Compound 10a': 1 H NMR (400MHz, DMSO-d6) δ15.19(br,1H),11.83(br,1H),9.18(br,2H),7.53(t,J=8.0Hz,1H),7.10(d,J=7.7Hz,1H),6.91(d,J=8.3Hz,1H),6.64(br, 1H),5.19(s,1H),4.97(s,1H),2.81(dd,J=9.9,6.3Hz,1H),2.47-2.43(m ,1H),2.15-2.08(m,1H),2.07(s,3H),1.79-1.70(m,1H),1.52(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ192.9,173.2,169.9,161.5,148.2,136.7,117.1,115.6,114.6,106.3,68.2,41.3,22.8,22.6,20.9ppm.

[0191] Example 29 Preparation of compound 10b'

[0192]

[0193] After stirring the mixture of compound 8' (50 mg, 0.12 mmol) in 50 μL (0.6 mmol) for 5 minutes, ethyl chloroformate (30 μL, 0.3 mmol) was added dropwise. The mixture was stirred at room temperature for 5 hours, then quenched with water, extracted with ethyl acetate, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 10b' (24.4 mg, 42%).

[0194] Compound 10b': 1 H NMR(400MHz,DMSO-d6)δ17.64(br,1H),15.17(br,1H),11.83(s,1H),9.21(br,2H), 7.54(t,J=8.0Hz,1H),7.11(d,J=7.7Hz,1H),6.91(d,J=8.3Hz,1H),6.69(br,1H),5. 06(br,1H),4.98(s,1H),4.18-4.13(m,2H),2.85(dd,J=9.4,6.6Hz,1H),2.59-2.53( m,1H),2.14-2.07(m,1H),1.84-1.75(m,1H),1.53(s,3H),1.23(t,J=7.1Hz,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ192.7,188.6,173.0,161.4,154.3,148.1,136.5, 117.0,115.5,114.5,106.1,74.1,68.1,64.0,41.0,40.48,22.8,14.1ppm.

[0195] Example 30 Preparation of compound 10c'

[0196]

[0197] Compound 8' (200 mg, 0.48 mmol) was dissolved in 2.0 mL of dry dichloromethane. Pyridine (0.38 mL, 4.8 mmol) was added under an ice-water bath. After stirring for 5 minutes, 2-methoxybenzoyl chloride (102 μL, 0.72 mmol) was added. The mixture was stirred at room temperature for 8 hours, quenched with water, extracted with ethyl acetate, washed successively with saturated ammonium chloride and saturated brine, dried, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 10c' (97 mg, 37%).

[0198] Compound 10c': 1H NMR(400MHz,DMSO-d6)δ15.23(br,1H),11.89(br,1H),9.21(br,2H),7.75(d, J=7.7Hz,1H),7.54(dt,J=14.2,8.1Hz,2H),7.16(d,J=8.4Hz,1H),7.10(d,J= 7.6Hz,1H),7.02(t,J=7.6Hz,1H),6.90(d,J=8.3Hz,1H),6.63(br,1H),5.36( br,1H),4.95(s,1H),3.82(s,3H),2.86(t,J=8.3Hz,1H),2.54(m,overlapped with DMSO),2.19-2.13(m,1H),1.91-1.83(m,1H),1.54(s,3H)ppm. 13 CNMR(101MHz,DMSO-d6)δ192.8,173.2,164.0,161.5,161.4,158.8,158.7,148.1,148.0,136.5, 134.1,131.2,120.0,119.3,117.0,115.5,114.5,112.7,106.3,68.1,55.8,41.4,40.4,22.7ppm.

[0199] Example 31 Preparation of compound 10d'

[0200]

[0201] Compound 8' (950 mg, 2.8 mmol, 1.0 equiv.) was dissolved in 10 mL of DCE. Pyridine (3.3 mL, 2.3 mmol) was added at 0 °C and stirred for 5 minutes. Benzoyl chloride (490 μL, 4.2 mmol) was added dropwise, and the mixture was stirred at 40 °C for 0.5 hours. The mixture was then quenched with water, extracted with ethyl acetate, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:2) to give a yellow solid compound 10d' (261 mg, 22%).

[0202] Compound 10d': 1H NMR(400MHz,DMSO-d6)δ17.77(br,1H),15.20(br,1H),11.84(s,1H),9.24(b r,2H),8.00(d,J=7.3Hz,2H),7.71-7.66(m,1H),7.57-7.51(m,3H),7.10(d, J=7.7Hz,1H),6.91(d,J=8.3Hz,1H),6.76(br,1H),5.50(br,1H),4.98(s,1H ),2.86(dd,J=10.0,6.2Hz,1H),2.62-2.59(m,1H),2.23-2.16(m,1H),2.83(q like,J=12.0Hz,1H),1.53(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ192.8,173.2,165.1,161.4,148.1,136.5,133.6,129.5,128.8,117.1,115.4,114.5,106.3,68.1,22.7ppm.

[0203] Example 32 Preparation of compound 11a

[0204]

[0205] Compound 10a (73 mg, 0.16 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and 0.4 mL of concentrated hydrochloric acid was added at 0 °C. The mixture was stirred at room temperature for 3 hours, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 2:3) to give a yellow solid compound 11a (42.4 mg, 60%).

[0206] Compound 11a: 1 H NMR (400MHz, DMSO-d6) δ17.97(br,1H),14.41(s,1H),9.10(br,2H),7.93(d,J=8.6Hz,1H),7.72(t,J=8.1Hz,1H),7.18(d,J=7.5Hz,1 H),6.99(br,1H),6.00(br,1H),4.76(s,1H),3.42-3.40(m,1H),2.96-2.91(m,1H),2.71-2.64(m,1H),2.39(s,3H),2.36(s,3H)ppm. 13C NMR (101MHz, DMSO-d6) δ173.1,169.9,160.6,149.0,138.6,133.1,131.0,122.4,121.6,119.6,116.6,110.7,42.4,25.3,21.1,14.1ppm.

[0207] Example 33 Preparation of compound 11b

[0208]

[0209] Using compound 10b (60 mg, 0.12 mmol) as a starting material, and following the preparation method of compound 11a, a yellow solid compound 11b (29.1 mg, 50%) was obtained.

[0210] Compound 11b: 1 H NMR (400MHz, DMSO-d6) δ17.97(br,1H),14.41(s,1H),9.10(br,2H),7.93(d,J=8.6Hz,1H),7.72(t,J=8.1Hz,1H),7.18(d,J=7.5Hz,1 H),6.99(br,1H),6.00(br,1H),4.76(s,1H),3.42-3.40(m,1H),2.96-2.91(m,1H),2.71-2.64(m,1H),2.39(s,3H),2.36(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ173.1,169.9,160.6,149.0,138.6,133.1,131.0,122.4,121.6,119.6,116.6,110.7,42.4,25.3,21.1,14.1ppm.

[0211] Example 34 Preparation of compound 11c

[0212]

[0213] Compound 10c (103.2 mg, 0.187 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (217.0 mg, 1.14 mmol) was added. After stirring at 40 °C for 30 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 11c (44.7 mg, 45%).

[0214] Compound 11c: 1H NMR (400MHz, DMSO-d6) δ17.16(br,1H),9.33-9.28(m,2H),7.84(s,1H),7.61-7.54(m,2H),7.38(s,1H),7.18(d,J=8.5Hz,1H),7.06(t,J =7.6Hz,1H),6.81(br,1H),6.29(d,J=4.6Hz,1H),3.86(s,3H),3.46-3.42(m,1H),3.17-3.14(m,1H),2.96-2.89(m,1H),2.33(s,3H)ppm. 13 C NMR (176MHz, DMSO-d6) δ173.0,164.6,158.9,138.8,134.4,132.7,131.6,120.2,112.8,56.0,25.6,22.6,14.0ppm.

[0215] Preparation of compound 11d in Example 35

[0216]

[0217] Compound 10d (1.4 g, 2.68 mmol) was dissolved in 15 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (2.36 g, 12.4 mmol) was added. After stirring at 40 °C for 8 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid 11d (1.2 g, 89%).

[0218] Compound 11d: 1 H NMR (400MHz, DMSO-d6) δ17.05(s,1H),9.38(s,1H),9.30(s,1H),8.10-7.99(m,2H),7.69(t,J=7.5Hz,1H),7.53-7.60(m,3H),7.43-7.31( m,2H),6.83(d,J=7.8Hz,1H),6.34(d,J=4.8Hz,1H),3.53-3.49(m,1H),3.25-3.21(m,1H),2.99(dd,J=17.4,11.8Hz,1H),2.35(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ173.0,165.0,162.9,157.8,138.7,133.8,132.8,131.0,129 .5,128.9,121.8,115.0,112.1,111.0,108.9,97.3,59.8,40.8,25.3,20.8,14.1ppm.

[0219] Example 36 Preparation of compound 11e

[0220]

[0221] Compound 10e (0.6 g, 1.03 mmol) was dissolved in 10 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (1.03 g, 5.15 mmol) was added. After stirring at 40 °C for 30 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give compound 11e (432 mg, 74%).

[0222] Compound 11e: 1 H NMR(400MHz,DMSO-d6)δ17.05(br,1H),15.27(br,1H),10.12(br,1H),δ9.38(s,1H) ,9.31(s,1H),8.01(br,1H),7.70-7.67(m,1H),7.56(t,J=8.1Hz,1H),7.48-7.38(m, 2H),7.34-7.25(m,2H),δ6.84(d,J=7.8Hz,1H).6.27(d,J=4.7Hz,1H),3.43-3.40(m, 1H),3.22-3.15(m,1H),2.95(dd,J=17.6,11.5Hz,1H),2.34(s,3H),2.25(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ173.0,169.2,163.2,150.2,138.7,134.6,132.7,131.4,131.0, 126.4,124.2,122.1,114.8,112.1,108.9,97.4,79.0,71.1,40.7,25.2,20.7,14.0ppm.

[0223] Example 37 Preparation of compound 11a'

[0224]

[0225] Compound 10a' (20 mg, 0.043 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (41.4 mg, 1.14 mmol) was added. After stirring at room temperature for 10 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 11a' (8.7 mg, 45%).

[0226] Compound 11a': 1H NMR (400MHz, DMSO-d6) δ17.70(br,1H),14.82(br,1H),10.10(br,1H),9.41(s,1H),9.34(br,1H),7.57(t,J=8.0Hz,1H),7.48-7. 33(m,1H),7.06(br,1H),6.84(s,1H),5.31(d,J=11.7Hz,1H),3.27-3.21(m,1H),3.05-2.94(m,2H),2.33(s,3H),2.12(s,3H)ppm. 13 C NMR (176MHz, DMSO-d6) δ179.5,173.0,169.7,158.1,157.9,139.3,129.7,118.1,116.4,45.7,40.0,29.6,20.3,14.1ppm.

[0227] Example 38 Preparation of compound 11b'

[0228]

[0229] Using compound 10b' (60 mg, 0.12 mmol) as a starting material, and following the preparation method of compound 11a, a yellow solid compound 11b' (23.1 mg, 40%) was obtained.

[0230] Compound 11b': 1 H NMR (400MHz, DMSO-d6) δ17.59(br,1H),14.83(br,1H),10.07(s,1H),9.44(br,2H),7.61(t,J=8.1Hz,1H),7.47(d,J=8.5Hz,1H),7.21(br,1H),6.90(d ,J=7.7Hz,1H),5.10(d,J=11.7Hz,1H),4.11(q,J=7.1Hz,2H),3.29(dd,J=1 7.3, 4.4Hz, 1H), 3.10-3.00 (m, 2H), 2.35 (s, 3H), 1.19 (t, J = 7.10Hz, 3H) ppm. 13 C NMR (101MHz, DMSO-d6) δ172.9,164.3,158.0,154.2,139.2,133.0,130.0,115.2,112.2,111.3,97.4,76.0,64.4,48.2,41.8,23.5,14.13,14.0ppm.

[0231] Example 39 Preparation of compound 11c'

[0232]

[0233] Compound 10c' (44 mg, 0.08 mmol) was dissolved in 2.0 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (61.1 mg, 0.32 mmol) was added. After stirring at 40 °C for 60 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 11c' (16.3 mg, 38%).

[0234] Compound 11c': 1 H NMR(400MHz,DMSO-d6)δ17.69(br,1H),14.88(br,1H),10.07(br,1H),9.46(s,1H) ,9.40(br,1H),7.68(dd,J=7.7,1.8Hz,1H),7.62-7.57(m,2H),7.46(d,J=8.5Hz,1H ),7.19(d,J=8.5Hz,2H),7.05(t,J=7.5Hz,1H),6.90(d,J=8.0Hz,1H),5.53(d,J=11 .7Hz,1H),3.84(s,3H),3.34-3.30(m,1H),3.16(d,J=15.7Hz,2H),2.32(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ196.4,173.0,164.6,158.3,139.2,134.1,133.0,1 30.8,130.2,120.2,119.1,115.1,112.7,76.2,55.9,47.1,42.0,13.9ppm.

[0235] Example 40 Preparation of compound 11d'

[0236]

[0237] Compound 10d' (240 mg, 0.46 mmol) was dissolved in 5.0 mL of tetrahydrofuran, and p-toluenesulfonic acid monohydrate (437 mg, 2.3 mmol) was added. After stirring at 40 °C for 12 hours, the reaction solution was concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid 11d' (184 mg, 79%).

[0238] Compound 11d': 1H NMR(400MHz,DMSO-d6)δ17.77(br,1H),15.20(br,1H),11.84(s,1H),9.24(b r,2H),8.00(d,J=7.3Hz,2H),7.71-7.66(m,1H),7.57-7.51(m,3H),7.10(d, J=7.7Hz,1H),6.91(d,J=8.3Hz,1H),6.76(br,1H),5.50(br,1H),4.98(s,1H ),2.86(dd,J=10.0,6.2Hz,1H),2.62-2.59(m,1H),2.23-2.16(m,1H),2.83(q like,J=12.0Hz,1H),1.53(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ192.8,173.2,165.1,161.4,148.1,136.5,133.6,129.5,128.8 117.0,115.4,114.5,106.3,68.1,22.7ppm.

[0239] Example 41 Preparation of Compound 13

[0240]

[0241] Tetracycline (1) (30g, 68mmol) was dissolved in THF (600mL), and MeI (72mL, 1.156mol) was added. The mixture was stirred at room temperature in the dark for 5 days. After the raw material was completely converted, it was directly filtered to obtain a yellow solid compound 12 (36g, 90%).

[0242] Compound 12: 1 H NMR(400MHz,DMSO-d6)δ16.95(s,1H),15.22(s,1H),11.74(s,1H),9.63(s,1H),9.47 (s,1H),7.73(s,1H),7.55(t,J=8.0Hz,1H),7.11(d,J=7.7Hz,1H),6.93(d,J=8.4Hz,1 H),5.04(d,J=25.7Hz,1H),4.46(s,1H),3.61-3.57(m,1H),3.39(s,10H),3.17-3.10( m,1H),2.91(dd,J=11.2,5.1Hz,1H),2.32-2.25(m,1H),1.71(s,1H),1.51(s,3H)ppm. 13C NMR(101MHz,DMSO-d6)δ193.1,192.3,186.3,174.9,173.0,161.5,147.9,136.8,117.2,1 15.4,114.5,106.7,97.4,75.5,72.2,68.0,67.1,54.0,41.9,36.1,26.9,25.2,22.6ppm.

[0243] Acetic acid (21 mL) and H2O (21 mL) were added to a 100 mL round-bottom flask containing quaternary ammonium salt intermediate compound 12 (7 g, 11.92 mmol). Zn powder was added while stirring at 0 °C. After the addition was complete, the reaction was continued for 0.5 hours. The mixture was then filtered, and the filtrate was extracted three times with ethyl acetate (300 mL). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and evaporated under reduced pressure at 27 °C to obtain the dedimethylamine product - compound 13 (3.2 g, 67%).

[0244] Compound 13: 1 H NMR(400MHz,DMSO-d6)δ18.43(s,1H),15.30(s,1H),11.88(s,1H),8.97(s,1H) ,8.74(s,1H),7.52(t,J=8.0Hz,1H),7.09(d,J=7.7Hz,1H),6.91(d,J=8.3Hz,1 H),6.63(s,1H),4.89(s,1H),3.19-3.15(m,1H),2.76(dd,J=11.1,5.4Hz,1H), 2.43-2.33(m,2H),1.98-1.94(m,1H),1.78(q,J=12.7Hz,1H),1.48(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ194.9,192.9,191.8,178.1,173.6,161.4,148.1,136.4 ,117.0,115.3,114.6,106.4,97.4,74.8,68.0,41.9,35.4,34.7,24.9,22.5ppm.

[0245] Example 42 Preparation of Compound 16

[0246]

[0247] To a round-bottom flask containing HFIP (4 mL), the demethylamine intermediate compound 13 (400 mg, 1 mmol), NIS (450 mg, 2 mmol), and AcOH (228 μL, 4 mmol) were added sequentially. After reacting at room temperature for 8 hours, saturated Na₂S₂O₃ solution (2 mL) was added, and the mixture was extracted three times with EtOAc (10 mL). The organic phases were combined, dried over anhydrous Na₂SO₄, filtered, and evaporated to dryness under reduced pressure. The residue was purified by reversed-phase column chromatography (water / acetonitrile = 3:2) to give a brown solid compound 14 (323 mg, 61%).

[0248] Compound 14: 1 H NMR (400MHz, DMSO-d6) δ18.41(s,1H),15.14(s,1H),12.79(s,1H),8.84(d,J=87.7Hz,2H),8.00(d,J=8.1Hz,1H),6.92(d,J=8.2Hz,1H),6.66(s, 1H),4.96(s,1H),3.16(d,J=16.4Hz,1H),2.77(dd,J=11.2,5.4Hz,1H),2 .43-2.33(m,2H),1.99-1.94(m,1H),1.82-1.75(m,1H),1.46(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ195.0,192.4,178.8,173.6,160.0,148.3,145.0,11 7.4,114.9,105.8,97.1,85.5,74.7,67.9,41.7,35.5,34.7,24.8,22.4ppm.

[0249] Compound 14 (200 mg, 0.38 mmol), bis(triphenylphosphine)palladium dichloride (26.6 mg, 0.038 mmol), triphenylarsine (11.6 mg, 0.038 mmol), and cuprous iodide (7.2 mg, 0.038 mmol) were dissolved in 6 mL of dry toluene. The mixture was purged with nitrogen three times. Then, phenyltri-n-butyltin reagent (180 μL, 0.56 mmol) was added, and the mixture was reacted at 90 °C for 12 hours. The reaction solution was directly concentrated, and the residue was purified by reversed-phase column chromatography (water / acetonitrile = 2:3) to obtain a yellow solid compound 15 (68.3 mg, 34%).

[0250] Compound 15: 1H NMR(400MHz,DMSO-d6)δ18.42(s,1H),15.21(s,1H),12.56(s,1H),8.98(br,1H),8. 73(br,1H),7.60-7.54(m,3H),7.44(t,J=7.6Hz,2H),7.38-7.34(m,1H),7.17(d,J= 8.0Hz,1H),6.66(br,1H),4.95(s,1H),3.21-3.16(m,1H),2.81(dd,J=11.3,5.4Hz, 1H),2.45-2.36(m,2H),1.98-1.96(m,1H),1.80(q,J=12.1Hz,1H),1.52(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ195.0,193.5,191.9,178.1,173.7,158.8,147.4,136.8,136.7,129.2,12 9.0,128.2,127.4,115.4,114.8,106.7,97.5,74.9,68.0,42.0,35.4,34.8,24.9,22.5,13.7ppm.

[0251] Compound 15 (50 mg, 0.1 mmol) and p-toluenesulfonic acid monohydrate (100 mg, 0.52 mmol) were dissolved in 1.5 mL of tetrahydrofuran. The mixture was stirred at room temperature for 60 hours. The reaction solution was extracted with ethyl acetate, washed with water, washed with saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give the yellow solid product compound 16 (21.3 mg, 44%). Compound 16: 1 H NMR(400MHz,DMSO-d6)δ18.32(br,1H),15.74(br,1H),10.61(br,1H),8.87(br,2H ),7.68(d,J=8.7Hz,1H),7.63(d,J=7.6Hz,2H),7.52(d,J=8.8Hz,1H),7.45(t,J=7. 6Hz,2H),7.34(t,J=7.4Hz,1H),6.93(br,1H),3.61-3.58(m,1H),3.34(d,J=8.9Hz, 2H),3.03(d,J=17.1Hz,1H),2.93-2.80(m,2H),2.57-2.51(m,1H),2.37(s,3H)ppm. 13C NMR (176MHz, DMSO-d6) δ173.3,138.3,129.4,128.1,118.2,79.2,79.1,78.9,65.2,35.6,29.6,22.6,13.9ppm.

[0252] Example 43 Preparation of Compound 18

[0253]

[0254] Using compound 14 as a starting material, and referring to the preparation method of compound 15, the phenyltri-n-butyltin reagent was replaced with ethynyltri-n-butyltin, and the reaction temperature was changed to 80℃, to obtain 9-ethynyltetracycline derivative compound 17 (54.3 mg, 67%).

[0255] Compound 17: 1 H NMR(400MHz,DMSO-d6)δ18.40(s,1H),15.17(s,1H),12.46(s,1H),8.95(s, 1H),8.72(s,1H),7.65(d,J=7.9Hz,1H),7.08(d,J=8.1Hz,1H),6.67(br,1H) ,4.99(br,1H),4.33(s,1H),3.19-3.14(m,1H),2.77(dd,J=11.1,5.4Hz,1H ),2.43-2.34(m,2H),1.98-1.93(m,1H),1.82-1.72(m,1H),1.47(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ195.0,192.6,191.7,178.7,173.6,162.5,148.7,139.5,131.4 ,130.2,115.3,114.7,110.6,106.4,85.6,78.8,68.0,41.7,35.3,34.7,24.8,22.4ppm.

[0256] Compound 17 (54 mg, 0.127 mmol) and 20% Pd(OH)₂ / C (23 mg) were dissolved in 3.0 mL of methanol, purged three times with hydrogen, stirred at 35 °C for 6 hours, filtered through diatomaceous earth, concentrated, and the crude product was dissolved in 2.0 mL of a mixed solvent of HCl (con.) / THF (v / v 1:4), stirred at 35 °C for 9 hours, concentrated, and subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give 9-ethyl derivative-compound 18 (28.3 mg, 54%). Compound 18: 1H NMR(400MHz,DMSO-d6)δ18.41(s,1H),15.18(s,1H),12.51(s,1H),8.96(s,1H),8.73(s,1H), 7.77(d,J=8.0Hz,1H),7.09(d,J=8.1Hz,1H),6.96(dd,J=17.8,11.3Hz,1H),6.63(s,1H),5.8 8(dd,J=17.8,1.4Hz,1H),5.34(d,J=12.0Hz,1H),4.90(s,1H),3.19-3.14(m,1H),2.76(dd,J =11.1,5.3Hz,1H),2.43-2.33(m,2H),1.99-1.93(m,1H),1.82-1.76(m,1H),1.48(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ195.0,193.3,191.8,178.2,173.7,158.8,147.6,132.7,129.8 ,125.1,115.8,115.2,114.6,106.5,97.4,74.9,67.9,41.9,35.3,34.7,24.9,22.5ppm.

[0257] Example 44 Preparation of Compound 19

[0258]

[0259] Compound 17 (60 mg, 0.14 mmol) and p-toluenesulfonic acid monohydrate (134 mg, 0.70 mmol) were dissolved in 3 mL of tetrahydrofuran and stirred at room temperature for 40 hours. The reaction mixture was extracted with ethyl acetate, washed with water, washed with saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was subjected to reversed-phase column chromatography (water / acetonitrile = 3:7) to give a yellow solid compound 19 (19.3 mg, 32%).

[0260] Compound 19: 1 H NMR (400MHz, DMSO-d6) δ18.32(br,1H),14.63(br,1H),8.87(br,2H),8.00(d,J=9.1Hz,1H),7.36(d,J=9.2Hz,1H),6.85(br,1H),3.33-3.31(m,overlappedwith water,1H),3.04-3.00(m,1H),2.93-2.86(m,1H),2.84-2.77(m,1H),2.56-2.52(m,1H),2.32(s,3H)ppm.13 C NMR (176MHz, DMSO-d6) δ195.8,173.2,171.6,165.8,142.3,137.9,131.6,121.1 ,114.2,114.0,113.1,110.5,99.4,77.7,35.2,34.7,29.4,27.9,22.6,13.9ppm.

[0261] Example 45 Preparation of Compound 20

[0262]

[0263] Compound 13 (0.6 g, 1.49 mmol) was dissolved in 1.6 mL of tetrahydrofuran. 0.4 mL of concentrated hydrochloric acid was slowly added at 0 °C. After stirring for 5 minutes, the mixture was brought to room temperature and stirred for 8 hours. The mixture was extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and the crude product was slurried with 30 mL of petroleum ether-ethyl acetate (5:1) to give the yellow solid product compound 20 (0.5 g, 87%).

[0264] Compound 20: 1 H NMR (400MHz, DMSO-d6) δ18.32(br,1H),15.32(s,1H),9.96(s,1H),8.86(br,2H),7.58(t,J=8.1Hz,1H),7.42(d,J=8.4Hz,1H), 6.86(d,J=7.7Hz,1H),3.31(d,J=4.5Hz,1H),3.02(d,J=18.6Hz,1H),2.91-2.79333(m,2H),2.56-2.54(m,1H),2.33(s,3H)ppm. 13 C NMR(101MHz,DMSO-d6)δ201.7,192.3,173.6,163.7,158.3,139.3,133.2,1 32.2,122.1,115.4,112.5,111.4,109.3,99.7,78.0,36.0,29.3,14.5ppm.

[0265] Example 46 Preparation of Compound 21

[0266]

[0267] Compound 20 (1.20 g, 3.13 mmol) was dissolved in 36 mL of tetrahydrofuran. Sodium nitrite (0.64 g, 9.36 mmol) was added at 0 °C, followed by the slow addition of 6.2 mL of dilute hydrochloric acid (1.0 N) and 30 mL of water. After stirring at the same temperature for 1.5 hours, the reaction solution was directly subjected to reversed-phase column chromatography (water / methanol = 1:9-9:1) to give the brownish-red solid product compound 21 (1.01 g, 82%).

[0268] Compound 21: 1 H NMR (400MHz, DMSO-d6) δ18.30(br,1H),14.37(s,1H),8.96(s,1H),8.81(s,1H),7.34(d,J=10.5Hz,1H),7.20( d,J=10.5Hz,1H),6.55(br,1H),3.33–3.26(m,2H),2.97(d,J=18.0Hz,1H),2.86–2.75(m,2H),2.25(s,3H)ppm. 13 C NMR (101MHz, DMSO-d6) δ185.9,173.1,163.2,151.5,138.1,126.0,125.8,119.9,117.5,114.0,99.6,78.7,78.0,35.3,30.8,13.5ppm.

[0269] Example 47 Preparation of Compound 23

[0270]

[0271] Compound 21 (164 mg, 0.40 mmol) and sodium hydrosulfite (140 mg, 0.8 mmol) were dissolved in 8 mL of a tetrahydrofuran-water mixture (v / v 1:1). After stirring for 10 minutes, the reaction was stopped. Sodium bicarbonate (50.5 mg, 0.6 mmol) and acetyl chloride (57 μL, 0.8 mmol) were added at 0 °C. After stirring for 30 minutes at the same temperature, the reaction was stopped, quenched with water, and extracted twice with ethyl acetate. The organic phases were combined, washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to reversed-phase column chromatography (water / acetonitrile = 1:1) to give a yellow solid compound 23 (74.3 mg, 42%).

[0272] Compound 23: 1H NMR (400MHz, DMSO-d6) δ18.34(s,1H),10.13(s,1H),9.41(s,1H),8.88(s,2H),8.16(d,J=9.0Hz,1H),7.40(d,J=9.2Hz, 1H),6.91(s,1H),3.49(br,1H),3.33–3.28(m,1H),3.10–2.95(m,1H),2.89–2.77(m,2H),2.31(s,3H),2.14(s,3H)ppm. 13 CNMR(101MHz,DMSO-d6)δ195.9,173.2,168.8,162.9,147.1,135.0,131.7,130.6,12 8.8,128.0,121.8,114.5,111.9,108.9,99.3,77.6,52.7,35.6,28.8,23.7,13.8ppm.

[0273] Example 49 Preparation of Compound 24

[0274]

[0275] Compound 21 (164 mg, 0.40 mmol) and sodium hydrosulfite (140 mg, 0.8 mmol) were dissolved in 8 mL of a tetrahydrofuran-water mixture (v / v 1:1). After stirring for 10 minutes, the reaction was stopped. Sodium cyanoborohydride (250 mg, 4.0 mmol) and butyraldehyde (70 μL, 0.8 mmol) were added at 0 °C. After stirring at room temperature for 2 hours, the reaction was stopped, quenched with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated ammonium chloride and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and the residue was subjected to reversed-phase column chromatography (water / acetonitrile = 2:3) to give a yellow solid compound 24 (91.4 mg, 50%).

[0276] Compound 24: 1 H NMR(400MHz,DMSO-d6)δ18.31(br,1H),8.91(s,1H),8.83(s,1H),7.74–7.64(m,1H),7.40–7.10(m,2H),6.77(br,1H),3.25-3.13(m,2 H),3.06-2.91(mi,2H),2.85-2.70(m,2H),2.36(br,1H),2.28(s,3H),1.67-1.55(m,2H),1.41-1.32(m,2H),0.91(t,J=7.3Hz,3H)ppm. 13C NMR (101MHz, DMSO-d6) δ173.1,167.0,131.7,131.5,128.7,114.3,99.2,91.7,65.0,35.6,30.0,29.1,19.7,19.0,18.7,13.6ppm.

[0277] Antibacterial activity test

[0278] The antibacterial effect was tested using in vitro cell experiments with representative derivatives (evaluated by MIC μg / mL value).

[0279] The test strains were Escherichia coli ATCC25922 and Staphylococcus aureus ATCC25923, and the two control groups were tetracycline hydrochloride and minocycline hydrochloride, respectively.

[0280] Antibacterial activity testing procedure:

[0281] 1. Dissolve each compound shown in Table 1 in DMSO to prepare a stock solution of each compound at a concentration of 51.2 mg / mL;

[0282] 2. Add 190 μL of MH medium to wells A1-H1 of a 96-well plate, and add 100 μL of MH medium to the remaining wells;

[0283] 3. Add 10 μL of the 51.2 mg / mL compound stock solution to well A1-H1 of a 96-well plate, then pipette 100 μL from well A1-H1 and add it to well A2-H2 and mix well. Continue in this manner until well A11-H11 is reached. At this point, the concentrations from left to right are 256 μg / mL and 0.25 μg / mL, respectively.

[0284] 4. Prepare a new 96-well plate and transfer the above sample solution into the new 96-well plate at a rate of 20 μL / well.

[0285] 5. The cultured bacterial solution is equivalent to McFarland turbidity 0.5 (OD625 0.08-0.13). The actual OD625 is 0.110. Then, the bacterial solution is diluted 1000 times with MH medium and 180 μL / well is added to a 96-well plate. The concentration of the compound in the well is recorded at this time.

[0286] 6. Place in a 35℃ incubator for 16-20 hours (actually 20 hours), and observe. The MIC value (μg / mL) for this group is defined as the absence of obvious colonies and the sample is clear.

[0287] Based on this testing procedure, the MIC values ​​(μg / mL) of representative derivatives are summarized in Table 1 below:

[0288] Table 1

[0289]

[0290]

[0291] As shown in Table 1, compounds 7b, 7c, 7e, 11e, 11d', and 20 showed the best antibacterial effect against Staphylococcus aureus (MIC value ≤ 0.25 μg / mL), comparable to or even better than tetracycline or minocycline; compounds 5, 5a, 7a, 7d, 7a', 9, 9', 11b, 11d, 11a', 11b', 11c', 16, 18, and 19 showed the second best effect (0.5 μg / mL ≤ MIC value ≤ 8 μg / mL); followed by compounds 7g, 7h, 7f, 7i, 11a, 11c, 21, 23, and 24 (12 μg / mL ≤ MIC value ≤ 128 μg / mL).

[0292] For Escherichia coli, compounds 9, 9', and 20 showed the best antibacterial effect (MIC value ≤ 8 μg / mL); compounds 5, 5a, 7a, 7e, 11a, 11b, 11b', 23, and 24 showed the next best effect (16 μg / mL ≤ MIC value ≤ 128 μg / mL); while other compounds showed poor antibacterial effect (MIC value > 128 μg / mL).

[0293] Analysis of experimental results: The compounds obtained in this invention generally showed better inhibitory effects against Staphylococcus aureus than against Escherichia coli, but compared with tetracycline or minocycline, their overall antibacterial effects were slightly weaker. It is worth noting that the C-ring aromatic molecular skeleton synthesized in this invention is considered an atypical tetracycline, and the mechanism of action of similar skeletons with CDHD differs from that of traditional tetracyclines acting on the 30S subunit of bacterial ribosomes, possibly involving inhibition of bacterial proliferation through binding and cleavage of the bacterial cell membrane. Due to this difference in mechanism, some compounds obtained in this invention may have potential inhibitory effects against infections caused by tetracycline-resistant bacteria. Related literature mentions that the C-ring aromatization of the molecular skeleton may reduce its binding ability to bacteria, which may explain why most of our modified compounds at the C4 and C9 positions showed lower antibacterial effects than tetracycline or minocycline.

[0294] The above description of the embodiments is provided to enable those skilled in the art to understand and use the invention. It will be apparent to those skilled in the art that various modifications can be made to these embodiments, and the general principles described herein can be applied to other embodiments without inventive effort. Therefore, the present invention is not limited to the above embodiments, and any improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the invention should be within the protection scope of the present invention.

Claims

1. An intermediate for an atypical tetracycline antibiotic derivative, characterized in that, The atypical tetracycline antibiotic derivative intermediate has the compound structure shown in Formula I: Among them, R 1 Represents any one of H, amino group and its substituents, amine group, amide group, sulfonamide group, hydroxyl group and its substituents, and ester group; R 2 It represents any one of H, alkyl, alkenyl, alkynyl, aryl, and acyl groups.

2. The atypical tetracycline antibiotic derivative intermediate according to claim 1, characterized in that, The atypical tetracycline antibiotic derivative intermediates include the following compound structures: In Equations I-1 and I-2, R 3 Represents any one of H, Me, Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, or Ms, R 4 Represents H or Me; in Equation I-3, R 2 It represents any one of H, alkenyl, alkynyl, or aryl.

3. A method for preparing an intermediate of an atypical tetracycline antibiotic derivative according to claim 1 or 2, characterized in that, In the compounds represented by formulas I-1 and I-2, R 3 R 4 When all atoms are H, the preparation method includes the following steps: A1. Tetracycline hydrochloride was oxidized by NCS under acidic conditions to obtain a mixture of hemiketals; A2. A mixture of hemiketals is reacted with potassium bicarbonate and hydroxylamine hydrochloride to form an oxime, which is then reduced with a reducing agent to obtain R. 3 R 4 All are compounds of formula I-1 with H; or R is obtained by reducing a mixture of hemiketals under conditions of sodium borohydride and cerium chloride heptahydrate. 3 The compound represented by formula I-2 is H; In the compounds represented by formulas I-1 and I-2, R 3 For Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, or Ms, and R 4 When the H content is used, the preparation method includes the following steps: A3. The R prepared in steps A1-A2 above... 3 R 4 All are compounds of formula I-1 represented by H or R 3 The compound represented by formula 1-2 with H undergoes an acylation reaction with an acylation reagent under basic conditions to give R. 3 For Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, or Ms and R 4 Compounds of formula I-1 and I-2 for H; In the compound represented by formula I-1, R 3 R 4 When all components are Me, the preparation method includes the following steps: A4. The R prepared in steps A1-A2 above... 3 R 4 The compound represented by formula I-1, which contains H, reacts with paraformaldehyde via a reductive amination reaction to obtain R. 3 R 4 All are compounds of formula I-1 represented by Me; In the compounds represented by Formula I-3, when R 2 When the H content is used, the preparation method includes the following steps: A5. Tetracycline was reacted in the dark with iodomethane and tetrahydrofuran. The resulting intermediate was reduced with a reducing agent to obtain R. 2 The compound represented by formula I-3 is H; In the compounds represented by Formula I-3, when R 2 When the group is alkenyl, alkynyl, or aryl, the preparation method includes the following steps: A6. The R prepared in step A5 above... 2 The compound of formula I-3 with H undergoes a C9 iodination reaction, and the resulting iodinated derivative then undergoes a Stille coupling reaction with a tin reagent to obtain R. 2 Compounds of formula I-3 that are alkenyl, alkynyl or aryl.

4. The method for preparing the intermediate of atypical tetracycline antibiotic derivatives according to claim 3, characterized in that, In step A3, the acylation reagent is selected from at least one of acetyl chloride, ethyl chloroformate, benzyl chloroformate, o-methoxybenzoyl chloride, benzoyl chloride, o-acetoxybenzoyl chloride, and acetic anhydride; The alkaline condition is achieved by selecting at least one of potassium carbonate, sodium bicarbonate, pyridine, TMEDA, DBU, and triethylamine. In steps A2 and A5, the reducing agent is selected from zinc powder; In step A6, the tin reagent is a tin reagent containing aryl, alkynyl, or alkenyl groups.

5. Use of an intermediate for an atypical tetracycline antibiotic derivative according to claim 1 or 2 in the preparation of atypical tetracycline antibiotic derivatives.

6. An atypical tetracycline antibiotic derivative, characterized in that, The atypical tetracycline antibiotic derivative has the compound structure shown in Formula II: Among them, R 1’ Represents any one of H, amino group and its substituents or their hydrochloride salts, amine group, amide group, sulfonamide group, hydroxyl group and its substituents, and ester group; R 2’ It represents any one of H, alkyl, aryl, acyl, nitroso, amino, and amide.

7. The atypical tetracycline antibiotic derivative according to claim 6, characterized in that, The atypical tetracycline antibiotic derivatives include the following compound structures: In Equations II-2 and II-5, R 3 Represents any one of H, Me, Ac, Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, CO2Et, Ms, R 4 Represents H or Me; In Equation II-6, R 2’ Represents any one of H, alkyl, aryl, acyl, nitroso, amino, and amide; wherein the amide is selected from acetamido, butyramido, benzamide, o-benzyloxybenzamide, p-nitrobenzamide, maleimide, phthalimide, etc. Any one of them.

8. A method for preparing an atypical tetracycline antibiotic derivative according to claim 6 or 7, characterized in that, The method for preparing the compound represented by Formula II-1 includes the following steps: B1, R 3 R 4 The compound represented by formula I-1, which contains H, is reacted under concentrated hydrochloric acid conditions to obtain the compound represented by formula II-1. In the compound shown in II-2, R 3 R 4 When both are H or Me, the preparation method includes the following steps: B2. The compound of formula II-1 prepared in step B1 is added to a base for neutralization or reacted with paraformaldehyde via a reducing amination reaction to obtain R. 3 R 4 All are compounds represented by II-2 of H or Me; In the compound shown in II-2, R 3 For Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz or CO2Et and R 4 When the H content is used, the preparation method includes the following steps: B3, R 3 The compound of formula I-1, consisting of Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz, or CO2Et, is reacted with p-toluenesulfonic acid monohydrate or concentrated hydrochloric acid to obtain R. 3 For Cbz, Bz, (2-OMe)Bz, (2-OAc)Bz or CO2Et and R 4 The compound represented by formula II-2 is H; In the compound shown in II-2, R 3 For Ac or Ms and R 4 When the H content is used, the preparation method includes the following steps: B4. The compound of formula II-1 prepared in step B1 is subjected to an acylation reaction with methanesulfonyl chloride or acetic acid to obtain R. 3 For Ac or Ms and R 4 The compound is shown as II-2 of H; The preparation method of the compounds represented by formulas II-3 and II-4 includes the following steps: B5. The compound of formula II-1 prepared in step B1 is subjected to an acylation reaction with N-phthalimide or maleic anhydride to obtain the compounds of formula II-3 and II-4. The preparation method of the compound shown in II-5 includes the following steps: B6. Add the compound shown in formula I-2 to p-toluenesulfonic acid monohydrate or concentrated hydrochloric acid to react and obtain the compound shown in II-5. In the compound shown in II-6, R 2’ When the group is H or aryl, the preparation method includes the following steps: B7, R 2 The compound of formula I-3 with H or aryl groups is reacted with p-toluenesulfonic acid monohydrate to obtain R. 2’ Compounds represented by II-6 with H or aryl groups; In the compound shown in II-6, R 2’ When the alkyl group is used, the preparation method includes the following steps: B8, R 2 Hydrogenation of the compound represented by formula I-3 with an alkynyl group yields R. 2’ Compounds shown in II-6 are alkyl groups; In the compound shown in II-6, R 2’ When the group is acyl, the preparation method includes the following steps: B9, R 2 The compound represented by alkynyl formula I-3 is reacted with p-toluenesulfonic acid monohydrate to obtain R. 2’ Compounds represented by acyl group II-6; In the compound shown in II-6, R 2’ When the group is nitrosyl, the preparation method includes the following steps: B10. The R prepared in step B7 above... 2’ The compound shown in II-6 of H is reacted with nitrite to obtain R. 2’ The compound represented by II-6 is nitroso; In the compound shown in II-6, R 2’ When the amide is used, the preparation method includes the following steps: B11, the R prepared in step B10 above... 2’ The compound shown as II-6, which is nitroso, is reduced with sodium hydrosulfite. After the conversion of the raw materials is complete, a base and an acylation reagent are directly added to react and yield R. 2’ The compound is shown as II-6 of the amide group. In the compound shown in II-6, R 2’ When the group is amine, the preparation method includes the following steps: B12, the R prepared in step B10 above 2’ The compound represented by II-6, which is nitroso, is reduced with sodium hydrosulfite. After the conversion of the raw material is complete, it is directly reacted with an aldehyde via a reducing amination reaction to obtain R. 2’ The compound shown is II-6, which is an amino group.

9. The method for preparing atypical tetracycline antibiotic derivatives according to claim 8, characterized in that, In step B2, the alkali is selected from any one of NaOH, NaHCO3, and Na2CO3; In step B11, the acylation reagent is selected from acetyl chloride, butyryl chloride, benzoyl chloride, o-benzyloxybenzoyl chloride, p-nitrobenzoyl chloride, maleic anhydride, phthalic anhydride, etc. any one of them; In step B12, the aldehyde is selected from any one of butyraldehyde, paraformaldehyde, octanaldehyde, and phenylacetaldehyde.

10. Use of an atypical tetracycline antibiotic derivative according to any one of claims 6-7 in the preparation of an antibacterial product.

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