KR101997518B1 - Synthetic method for nocarbenzoxazole G and derivatives of nocarbenzoxaoles - Google Patents

Abstract

An object of the present invention is to provide an anti-inflammation agent having excellent efficacy and minimized side effects by solving side effects of existing anti-inflammation agents, and to provide a novel nocarbenzoxazole derivative and a synthesis method thereof. By using a POCl_3-mediated ring dehydration reaction using commercially available inexpensive raw materials and selective and/or complete demethylation and a reduction reaction as main reactions, nocarbenzoxazole G and nocarbenzoxazole derivatives are easily synthesized in a 15 to 49% of yield and it is confirmed by testing anti-inflammatory activity of compounds.

Description

Synthetic method for nocarbenzoxazole G and derivatives of nocarbenzoxaoles}

The present invention relates to a method for synthesizing nocabenzoxazole G and nocabenzoxazole derivatives from a commercially available economical raw material in an easy process and an anti-inflammatory pharmaceutical composition containing the compound.

Inflammation is the body's local defense response to endogenous and exogenous injuries such as pathogens, infections or diseases. Inflammation It can be divided into acute or chronic inflammation. Although this reaction can cause the body to limit invasion and damage after trauma, long-term inflammation is associated with the induction of vaginal diseases such as cardiovascular disease, rheumatoid arthritis, cancer, bronchitis, diabetes and neurodegenerative diseases. The inflammatory process involves the release of various chemical mediators such as cytokines (interleukin-1β, interleukin-6 and tumor necrosis factor α), chemokines, histamine, prostaglandins, complement fragments, and active molecules such as nitric oxide released from white blood cells. . Of these, nitric oxide (NO) is a small, temporary free radical produced by three kinds of similar nitric oxide synthase (NOS): endothelial NOS, neuronal NOS and induced NOS, and plays an important role in the development of inflammation. This acts as a "friend and enemy" face depending on concentration. Nitric oxide has an anti-inflammatory effect under normal physiological concentrations. However, nitric oxide production by induced NOS is involved in several pathophysiological processes. Thus, pharmaceutical intervention in nitric oxide production will have a strong impact on the treatment of inflammation based diseases.

Non-steroidal anti-inflammatory drugs (NSAIDs), including over-the-counter drugs such as aspirin and ibuprofen, corticosteroids, selective COX-2 inhibitors (COXIBs), and antihistamines, are used for pain, fever, and inflammation. It is usually prescribed for treatment. They work by blocking the synthesis or activity of mediators that cause the host's response to injury. These small molecule inhibitors provide major prescriptions for various inflammatory diseases, and their use is limited due to common side effects such as gastric ulcers, joint destruction, kidney toxicity, and cardiovascular disease. As a result, in order to solve these problems, new drugs need to be developed to improve the effects and minimize side effects.

Benzoxazole is an oxazole ring structure fused with benzene and is an important heterocycle. Numerous benzoxazole derivatives are used as building blocks in the biochemical and pharmaceutical fields, as well as in other industries such as pesticides, pigments, textile auxiliaries and plastics. Substituted benzoxazoles exhibit a broad spectrum of bioactivity such as antibiotics, anticancer agents, antimicrobial agents, antifungal agents, Parkinson's disease agents, anti-inflammatory agents, anticonvulsants, immunosuppressants, antiviral agents and antiparasitic agents (8, 9). In addition, some benzoxazoles exhibit enzymatic inhibitory activity (10). In addition, some benzoxazoles are recognized as important structures in fluorescent probes such as anion and metal cation sensors (11).

Republic of Korea Patent No. 10-1792743 "Novel benzobenzoxazole derivatives, preparation method thereof and uses thereof"

A. Ortega-Gomez, M. Perretti O. Soehnlein, EMBO Mol. Med. 2013, 5, 661. P. Libby, Nutr. Rev. 2007, 65, 140. D. W. Gilroy, T. Lawrence, M. Perretti, A. G. Rossi, Nat. Rev. Drug Discov. 2004, 3, 401. J. A. McCleverty, Chem. Rev. 2004, 104, 403. J. Lei, Y. Vodovotz, E. Tzeng, T.R. Billiar, Nitric Oxide 2013, 35, 175. R. J. Flower, Nature Rev. Drug Discov. 2003, 2, 179. I. L. Meek, M. A. F. J. van de Laar, H. E. Vonkeman, Pharmaceuticals 2010, 3, 2146. B. Maleki, M. Baghayeri, S. M. Vahdat, A. Mohammadzadeh, S. Akhoondic, RSC Adv. 2015, 5, 46545. S. Rajasekhar, B. Maiti, K. Chanda, Synlett 2017, 28, 521. (a) P. Malapati, V.S. Krishna, R. Nallangi, R. R. Srilakshmi, D. Sriram, Eur. J. Med. Chem. 2018, 145, 23; (b) M. -H. Nam, M. Park, H. Park, Y. Kim, S. Yoon, V. S. Sawant, J. W. Choi, J. -H. Park, K. D. Park, S.-J. Min, C. J. Lee, H. Choo, ACS Chem. Neurosci, 2017, 8 1519. M. Taki, J. L. Wolford, T. V. O'Halloran, J. Am. Chem. Soc. 2004, 126, 712. M. Sun, X. Zhang, H. Hao, W. Li, C. Lu, J. Nat. Prod. 2015, 78, 2123. (a) K. Damodar, J.-K. Kim, J.-G. Jun, Tetrahedron Lett. 2017, 58, 50; (b) H. Y. Jang, H. J. Park, K. Damodar, J.-K. Kim, J.-G. Jun, Bioorg. Med. Chem. Lett., 2016, @ 26, 5438. (a) H. Ramser, E. Wiberg, Chem. Ber. 1930, 63, 1136; (b) W. Gerrard, M. F. Lappert, J. Chem. Soc. 1952, 1486. (a) D. Tsikas, J. Chromatogr. B, 2007, 851, 51; (b) E. M. Hetrick, M. H. Schoenfisch, Annu. Rev. Anal. Chem. 2009, 2, 409. (a) J. D. Laskin, D. E. Heck, D. L. Laskin, Trends Endocrinol. Metab. 1994, 5, 377; (b) D. Schade, J. Kotthaus, B. Clement, Pharmacol. Ther. 2010, 126, 279; (c) C. S. Kim, L. Subedi, J. Oh, S. Y. Kim, S. U. Choi, K. R. Lee, J. Nat. Prod. 2017, 80, 1134. J. Carmichael, A. F. Gazdar, Cancer Res. 1987, 47, 943.

An object of the present invention is to provide an anti-inflammatory agent that is excellent in efficacy and minimized side effects by solving the side effects of the conventional anti-inflammatory agent.

It is another object of the present invention to provide a method for synthesizing new nocabenzoxazoles and derivatives.

The present inventors easily synthesized nocabenzoxazole G (compound 5) and nocabenzoxazole derivatives (compounds 4 and 6) in 15-49% yield using commercially available inexpensive raw materials.

POCl ₃ -mediated toroidal dehydration, selective and / or complete demethylation and reduction are key steps in the success of this synthetic route. In addition, these benzoxazole compounds showed mild nitric oxide production inhibitory activity in a concentration-dependent manner without noticeable cytotoxicity in LPS-induced RAW264.7 cells.

The nocabenzoxazole G and nocabenzoxazole derivatives of the present invention are shown in FIG. 1. Compound 5 (nocarbenzoxazoles G) is a natural product (12) isolated from the basophilic Nocardiopsis lucentensis DSM 44048, while compounds 4 and 6 are synthetic derivatives. There was no report of anti-inflammatory activity, and no method of synthesizing compounds 4 and 6 was reported.

The present invention

A) Ethyl 3-amino-4-hydroxybenzoate was added to anhydrous 1,4-dioxane, and the compound 16 represented by the formula (16) was added to the stirred solution under normal temperature and nitrogen atmosphere, warmed to 90 ° C and stirred, and then POCl ₃ dropwise addition and stirring to obtain ethyl 2- (2,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate;

B) Ethyl 2- (2,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate was added to anhydrous CH ₂ Cl ₂ and stirred in a solution of BCl ₃ dropwise at 0 ° C. under nitrogen atmosphere. After addition, warming to room temperature and stirring to obtain ethyl 2- (2-hydroxy-4-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate; And

C) Add ethyl 2- (2,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate to anhydrous CH ₂ Cl ₂ , add AlCl ₃ to a stirred solution under nitrogen atmosphere, and stir at room temperature. After completion of the reaction, H ₂ O was slowly added and stirred to obtain ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate;

D) Ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate was added to anhydrous CH ₂ Cl ₂ , and stirred in a solution of BCl ₃ at 0 ° C. under a nitrogen atmosphere. After dropwise addition, warming to room temperature and stirring to obtain ethyl 2- (2,4-dihydroxyphenyl) benzo [ d ] oxazole-5-carboxylate; And

E) ethyl 2- (2-hydroxy-4-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate or ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole To a solution of -5-carboxylate or ethyl 2- (2,4-dihydroxyphenyl) benzo [ d ] oxazole-5-carboxylate in anhydrous THF, stirred LiAlH ₄ solution under 0 ° C. and nitrogen atmosphere. After dropwise addition, the mixture was warmed to room temperature, stirred to complete the reaction, cooled to 0 ° C., and saturated Na ₂ CO ₃ aqueous solution was slowly added to quench excess LiAlH ₄ and stirred, respectively, to correspond to 2- (5- (hydroxy). Methyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol, 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -3-methoxyphenol or 4- It relates to a method for synthesizing nocabenzoxazole or a derivative thereof, comprising the step of obtaining (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol.

<Formula 16>

The present invention also relates to novel 2- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol.

The present invention also relates to novel 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol.

The present invention also provides 2- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol and 4- (5- (hydroxymethyl) benzo [ d ] oxazole- Anti-inflammatory containing one or more selected from 2-yl) -3-methoxyphenol and 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol It relates to a pharmaceutical composition.

The pharmaceutical compositions of the present invention may be formulated in oral or injectable form by conventional methods in combination with a carrier which is commonly acceptable in the pharmaceutical art. Oral compositions include, for example, tablets and gelatin capsules, which, in addition to the active ingredient, may contain diluents (e.g. lactose, dextrose, sucrose, mannitol, sorbitol, cellulose and / or glycine), lubricants (e.g. silica, talc) , Stearic acid and its magnesium or calcium salts and / or polyethylene glycols, the tablets also contain binders (e.g. magnesium aluminum silicate, starch paste, gelatin, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidone). ), Optionally with disintegrants (e.g., starches, agar, alginic acid or sodium salts thereof) or boiling mixtures and / or absorbents, colorants, flavors and sweeteners. Injectable compositions are preferably aqueous isotonic solutions or suspensions, and the compositions mentioned are sterile and / or contain auxiliaries (eg, preservatives, stabilizers, wetting or emulsifier solution promoters, salts / or buffers to control osmotic pressure). In addition, they may contain other therapeutically valuable substances.

The pharmaceutical preparations thus prepared can be administered orally as desired, or parenterally, ie, intravenously, subcutaneously, intraperitoneally, or topically. The dose may be administered by dividing the daily dose between 0.0001 and 100 mg / kg in one to several times. Dosage levels for a particular patient may vary depending on the patient's weight, age, sex, health condition, time of administration, method of administration, rate of excretion, severity of the disease, and the like.

Furthermore, the present invention provides the above 2- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol, 4- (5- (hydroxymethyl) benzo [ d ] oxazole At least one selected from 2-yl) -3-methoxyphenol and 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol as an active ingredient And it provides a pharmaceutical composition useful for the prevention and treatment of inflammatory diseases, including dermatitis, such as atopic dermatitis, characterized in that it comprises a pharmaceutically acceptable carrier.

Inflammatory diseases as defined in the present invention are dermatitis diseases including atopic dermatitis, neuronal inflammatory diseases such as glioma cells, spondylitis, urethritis, cystitis, nephritis, pyelonephritis, vasculitis, rhinitis, sore throat, tonsillitis, acute pain or inflammatory bowel Diseases, and the like, and are preferably dermatitis diseases, urethritis, cystitis, nephritis, pyelonephritis, rhinitis, sore throat, tonsillitis, or inflammatory bowel disease.

According to the present invention, it is possible to easily synthesize nocabenzoxazole G (compound 5) and nocabenzoxazole derivatives (compounds 4 and 6) in 15-49% yield using commercially available inexpensive raw materials.

In addition, according to the present invention, it is possible to provide an anti-inflammatory pharmaceutical composition comprising at least one of the synthesized nocabenzoxazole G and derivatives.

Figure 1 shows the chemical structures of nocabenzoxazole F, G and its derivatives.
Figure 2 shows a method for synthesizing nocabenzoxazole F, G and its derivatives.

Hereinafter, the configuration of the present invention will be described in more detail with reference to specific embodiments. However, it is obvious to those skilled in the art that the scope of the present invention is not limited only to the description of the embodiments.

All chemicals were used without purification as purchased unless otherwise noted. All solvents used for the reaction were distilled from an appropriate dehydrating agent under a nitrogen atmosphere. All solvents used for chromatography were purchased and used directly without further purification.

All reactions were monitored by thin layer chromatography (TLC) using a plate coated with silica gel on a DC-Plastikfolien 60, F ₂₅₄ (Merck, 0.2 mm layer) plastic plate and observed using UV (254 nm) or p -Observed by staining with anisaldehyde and phosphomolybdic acid (PMA). Silica gel 60 (230-400 mesh, Merck, Darmstadt, Germany) was used to purify the compound obtained by column chromatography. ¹ H-NMR spectra were recorded at 75 MHz for Varian Mercury-300 MHz FT-NMR (Palo alto, CA, USA) and ¹³ C, and chemical shifts (δ) are expressed in parts per million (ppm) for TMS. Coupling constants ( J ) are quoted in Hz. Peak division patterns are abbreviated as s (singlet), d (doublet), t (triplet), q (quartet), dd (doublet of doublet) and m (multiplet), and CDCl ₃ / CD ₃ OD / acetone d ₆ / DMSO- d ₆ was used as solvent and internal standard. Low resolution mass spectra (electron ionization, EI) were recorded using an Agilent-5977E spectrometer, and high resolution mass spectra were recorded using JMS-700 (JEOL, Japan). Melting points were measured on a MEL-TEMP II instrument and were not corrected.

Ethyl 3-amino-4-hydroxybenzoate (Ethyl 3-amino-4- hydroxybenzoate) ( Compound 10): 3-amino-4-hydroxy-benzoic acid (Compound 9) (1.0 g, 6.53 mmol ) EtOH (10 mL) was added to the stirred solution and concentrated H ₂ SO ₄ (3.0 mL) was added at 0 ° C. under a nitrogen atmosphere. After the reaction was completed, the mixture was warmed and refluxed for 12 hours. After the reaction was completed, the mixture was cooled to room temperature, poured into crushed ice, and saturated with NaHCO _3. Neutralized with aqueous solution. The reaction mixture was extracted with EtOAc (3 x 45 mL). The mixed organic solvent layer was washed with H ₂ O (2 × 20 mL), brine (20 mL), dried over anhydrous Na ₂ SO ₄ , and concentrated in vacuo. The crude compound was purified by column chromatography (EtOAc: Hexane = 1: 2) to give a pale orange solid compound 10 (1.01 g, 85%). Melting point 96-98 ° C .; R _f = 0.33 (EtOAc / hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 7.44 (1H, d, J = 1.8 Hz), 7.41 (1H, dd, J = 7.8, 1.8 Hz), 6.75 (1H, d, J = 7.8 Hz), 4.32 (2H, q, J = 7.2 Hz) 1.37 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 167.0, 148.6, 134.1, 123.3, 122.2, 118.2, 114.6, 61.0, 14.7.

Substituted  Benzoic Acid (Compound 11- From 13)  Substituted Benzoyl  General procedure for the synthesis of chlorides (compounds 14, 16 and 17):

SOCl ₂ (2 mL) was added to substituted benzoic acid (0.55 mmol) at 0 ° C. under a nitrogen atmosphere. The mixture was stirred at 60 ° C. for 2 hours. After the reaction was completed, the reaction mixture was cooled to room temperature, and excess SOCl ₂ was removed under reduced pressure. The resulting benzoyl chloride was dried under vacuum for 30 minutes and used in the next step without further purification.

2-aryl Benzo [d] oxazole -5- Carboxylate  (2-aryl benzo [ d ] oxazole -5-carboxylate) POCl ₃  General procedure for mediated synthesis:

Compound 10 (0.55 mmol) was added to anhydrous 1,4-dioxane (3 mL), and substituted benzoyl chloride (1.1 mmol, 2.0 equiv) was added to the stirred solution under normal temperature and nitrogen atmosphere. The mixture was warmed to 90 ° C. and stirred for 3 hours. In this step, POCl ₃ (0.16 mL, 1.65 mmol, 3.0 equiv) was added dropwise and stirred for a further 12 h. After the reaction was completed, the reaction mixture was cooled to room temperature, and the solvent and POCl ₃ were removed under reduced pressure. EtOAc (25 mL), H ₂ O (10 mL) were added to the crude compound and the two layers were separated. The aqueous layer was extracted with EtOAc (2 x 20 mL). The mixed organic solvent layer was washed with H ₂ O (2 × 15 mL) and brine (15 mL), dried over anhydrous Na ₂ SO ₄ , and concentrated in vacuo. The crude compound was purified by column chromatography (EtOAc: Hexane = 1: 5-1: 2) to give pure compound.

Ethyl 2- (4-methoxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  {Ethyl 2- (4-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate} (Compound 18):

Yield: 55%; Off-white solid; Melting point 156-158 ° C .; R _f = 0.73 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.38 (1H, s), 8.14 (2H, d, J = 8.7 Hz), 8.04 (1H, d, J = 8.4 Hz), 7.53 (1H, d, J = 8.4 Hz), 6.99 (2H, d, J = 8.7 Hz), 4.40 (2H, q, J = 7.2 Hz), 3.86 (3H, s), 1.42 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 166.3, 164.3, 162.6, 153.5, 142.4, 129.6, 127.2, 126.6, 121.5, 119.1, 114.5, 110.1, 61.3, 55.6, 14.6.

Ethyl 2- Phenylbenzo [ d ] Oxazole -5- Carboxylate  (Ethyl 2-phenylbenzo [ d ] oxazole-5-carboxylate) (compound 19):

Yield: 60%; Off-white solid; Melting point 102-104 ° C; R _f = 0.54 (EtOAc / Hexane = 1/3); ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.42 (1H, d, J = 0.9 Hz), 8.22-8.18 (2H, m), 8.06 (1H, dd, J = 8.7, 1.8 Hz), 7.56- 7.48 ( 4H, m), 4.40 (2H, q, J = 7.2 Hz), 1.42 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 166.1, 164.2, 153.6, 142.2, 131.9, 129.0, 127.8, 127.4, 127.0, 126.7, 122.0, 110.3, 61.3, 14.6.

Ethyl 2- (2,4-dimethoxyphenyl) benzo [d] oxazole-5-carboxylate (Ethyl 2- (2,4-dimethoxyphenyl) benzo [d] oxazole-5-carboxylate) (compound 20): Yield: 48%; Off-white solid; Melting point 148-150 ° C .; R _f = 0.61 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.45 (1H, s), 8.08 (1H, J = 9.3 Hz), 8.03 (1H, d, J = 9.3 Hz), 7.54 (1H, d, J = 8.7 Hz ), 6.60 (1H, d, J = 8.7 Hz), 6.58 (1H, s), 4.40 (2H, q, J = 6.9 Hz), 4.00 (3H, s), 3.87 (3H, s), 1.42 (3H , t, J = 6.9 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 166.2, 163.7, 162.8, 159.9, 152.9, 142.2, 132.5, 126.8, 126.2, 121.6, 109.8, 108.4, 105.3, 99.0, 61.0, 56.1, 55.5, 14.4.

Ethyl 2- (3,4-dimethoxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  (Ethyl 2- (3,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate) (Compound 21):

Yield: 53%; Off-white solid; Melting point 146-148 ° C .; R _f = 0.47 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.36 (1H, d, J = 1.8 Hz), 8.03 (1H, dd, J = 8.7, 1.8 Hz), 7.78 (1H, dd, J = 8.4, 2.1 Hz) , 7.67 (1H, d, J = 2.1 Hz), 7.50 (1H, d, J = 8.7 Hz), 6.92 (1H, d, J = 8.4 Hz), 4.39 (2H, q, J = 7.2 Hz), 3.98 (3H, s), 3.94 (3H, s), 1.42 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 166.2, 164.3, 153.5, 152.3, 149.3, 142.3, 127.3, 126.6, 121.5, 121.4, 119.2, 111.1, 110.2, 110.1, 61.3, 56.3, 56.2, 14.6.

BBr ₃ To  General Procedure of Demethylation Using:

Ethyl 2-aryl benzo [ d ] oxazole-5-carboxylate (0.2 mmol) was added to anhydrous CH ₂ Cl ₂ (4 mL) and stirred in BBr ₃ (1.0 M in CH ₂ Cl ₂ , 0.3 mL, 1.5). Equivalent) was added dropwise at 0 ° C. under a nitrogen atmosphere. The mixture was warmed to room temperature and stirred for 3 hours. After the reaction was completed, the reaction mixture was cooled to 0 ° C, 1.0 N HCl (1 mL) was slowly added thereto, and stirred for 15 minutes. CH ₂ Cl ₂ Extracted with (2 x 20 mL). The mixed organic solvent layer was washed with H ₂ O (2 × 10 mL) and brine (10 mL), dried over anhydrous Na ₂ SO ₄ , and concentrated in vacuo. The crude compound was purified by column chromatography (EtOAc: Hexane = 1: 2-1: 1) to obtain a pure compound. [Note: In the case of compound 21, 3.0 equivalents of BBr ₃ were used].

Ethyl 2- (4-hydroxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  (Ethyl 2- (4-hydroxyphenyl) benzo [ d ] oxazole-5-carboxylate) (compound 22):

Yield: 71%; Off-white solid; Melting point 210-212 ° C; R _f = 0.55 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, DMSO-d6) δ 8.26 (1H, d, J = 1.5 Hz), 8.08-8.03 (3H, m), 7.68 (1H, d, J = 8.7 Hz), 6.95 (2H, d , J = 8.4 Hz), 4.38 (2H, q, J = 7.2 Hz), 1.43 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, DMSO-d6) δ 167.5, 166.6, 163.1, 154.8, 143.3, 131.0, 128.6, 127.7, 121.6, 118.4, 117.2, 111.6, 62.6, 14.9.

Ethyl 2- (3,4-dihydroxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  (Ethyl 2- (3,4-dihydroxyphenyl) benzo [ d ] oxazole-5-carboxylate) (Compound 23):

Yield: 75%; Off-white solid; Melting point 214-216 ° C .; R _f = 0.35 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, DMSO-d6) δ 8.21 (1H, d, J = 1.8 Hz), 7.96 (1H, d, J = 8.4 Hz), 7.81 (1H, dd, J = 8.4, 1.8 Hz), 7.58 (1H, d, J = 2.1 Hz), 7.54 (1H, dd, J = 8.4, 2.1 Hz), 6.92 (1H, d, J = 8.4 Hz), 4.34 (2H, q, J = 7.2 Hz), 1.35 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, DMSO-d6) δ 165.2, 164.1, 152.8, 149.8, 145.6, 141.8, 126.5, 125.9, 120.0, 119.9, 116.6, 116.0, 114.3, 110.6, 60.8, 14.2.

Ethyl 2- (2-hydroxy-4-methoxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  {Ethyl 2- (2-hydroxy-4-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate} (Compound 24):

Compound 20 (0.09 g, 0.27 mmol) was added to anhydrous CH ₂ Cl ₂ (4 mL), and BCl ₃ (1.0 M in CH ₂ Cl ₂ , 0.28 mL) was added to the stirred solution under 0 ° C. under a nitrogen atmosphere. The reaction mixture was allowed to warm to room temperature and stirred for 2 hours. After the reaction was completed, the reaction mixture was cooled to 0 ° C, 1.0 N HCl (1 mL) was added slowly, and stirred for 15 minutes. CH ₂ Cl ₂ Extracted with (2 x 20 mL). The mixed organic solvent layer was washed with H ₂ O (2 × 10 mL) and brine (10 mL), dried over anhydrous Na ₂ SO ₄ , and concentrated in vacuo. The crude compound was purified by column chromatography (EtOAc: Hexane = 1: 2) to give pure off-white solid compound 24 (0.08 g, 89%). Melting point 154-156 ° C .; R _f = 0.70 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 11.33 (1H, s), 8.31 (1H, d, J = 1.8 Hz), 8.05 (1H, dd, J = 8.4, 1.8 Hz), 7.84 (1H, d, J = 9.0 Hz), 7.54 (1H, d, J = 8.4 Hz), 6.57 (1H, d, J = 2.1 Hz), 6.54 (1H, dd, J = 9.0, 2.1 Hz), 4.41 (2H, q, J = 7.2 Hz), 3.85 (3H, s), 1.44 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 165.8, 164.4, 164.1, 160.8, 151.6, 140.2, 128.2, 127.5, 126.6, 120.4, 110.0, 107.8, 103.1, 101.2, 61.2, 55.5, 14.4.

Ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  {Ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate} (Compound 25):

Compound 20 (0.09 g, 0.27 mmol) was added to anhydrous CH ₂ Cl ₂ (4 mL) and AlCl ₃ (0.12 g, 0.92 mmol) was added to the stirred solution under a nitrogen atmosphere. The reaction mixture was stirred at room temperature for 48 hours. After completion of the reaction, H ₂ O (1 mL) was slowly added and stirred for 15 minutes. CH ₂ Cl ₂ Extracted with (2 x 20 mL). The mixed organic solvent layer was washed with H ₂ O (2 × 10 mL) and brine (10 mL), dried over anhydrous Na ₂ SO ₄ , and concentrated in vacuo. The crude compound was purified by column chromatography (EtOAc: Hexane = 1: 2) to give compound 25 (0.04 g, 42%) as a pure off-white solid. Melting point 204-206 ° C; R _f = 0.22 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CD ₃ COCD ₃ ) δ 9.48 (1H, s), 8.29 (1H, s), 8.04 (1H, d, J = 8.4 Hz), 8.00 (1H, d, J = 8.4 Hz) , 7.71 (1H, d, J = 8.7 Hz), 6.68 (1H, s), 6.63 (1H, d, J = 8.4 Hz), 4.38 (2H, q, J = 7.2 Hz), 3.94 (3H, s) , 1.40 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CD ₃ COCD ₃ ) δ 166.3, 163.2, 161.3, 154.1, 143.1, 133.6, 127.8, 126.8, 121.4, 111.0, 108.8, 106.9, 100.5, 99.7, 61.6, 56.2, 14.7.

Ethyl 2- (2,4-dihydroxyphenyl) benzo [ d ] Oxazole -5- Carboxylate  {Ethyl 2- (2,4-dihydroxyphenyl) benzo [ d ] oxazole-5-carboxylate} (Compound 26):

According to the procedure used for preparing compound 24, compound 25 (0.09 g, 0.29 mmol) was treated with BCl ₃ (1.0 M in CH ₂ Cl ₂ , 0.58 mL) to give an off-white solid compound 26 (0.08 g, 97%). . Melting point 218-220 ° C .; R _f = 0.12 (EtOAc / Hexane = 1/1); ¹ H NMR (300 MHz, CD ₃ COCD ₃ ) δ 11.19 (1H, s), 9.53 (1H, s), 8.27 (1H, d, J = 1.2 Hz), 8.06 (1H, dd, J = 8.4, 1.8 Hz), 7.85 (1H, d, J = 9.0 Hz), 7.76 (1H, d, J = 8.4 Hz), 6.59 (1H, dd, J = 9.0, 1.8 Hz), 6.52 (1H, d, J = 1.8 Hz), 4.37 (2H, q, J = 7.2 Hz), 1.40 (3H, t, J = 7.2 Hz); ¹³ C NMR (75 MHz, CD ₃ COCD ₃ ) δ 166.0, 165.4, 163.9, 161.7, 152.4, 141.0, 129.6, 128.5, 127.3, 120.5, 111.2, 109.5, 103.8, 102.9, 61.8, 14.6.

Ethyl 2-aryl Benzo [ d ]] Oxazole -5- Carboxylate  (ethyl 2-aryl benzo [ d general procedure for the reduction of oxazole-5-carboxylate):

To a stirred solution of ethyl 2-aryl benzo [ d ] oxazole-5-carboxylate (0.17 mmol) in anhydrous THF (3 mL) was added LiAlH ₄ solution (1.0 M in THF, 0.34 mL, 0.34 mmol, 2.0 equiv). .) Was added dropwise at 0 ° C. under a nitrogen atmosphere. The mixture was warmed to room temperature and stirred for 30 minutes. After the reaction was completed, the reaction mixture was cooled to 0 ° C, and saturated Na ₂ CO ₃ aqueous solution (1 mL) was slowly added to quench excess LiAlH ₄ and stirred for 20 minutes. The resulting suspension was filtered, washed with ethyl acetate (2 Х 10 mL), and the mixed organic solvent layer was dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure. The crude compound was purified by column chromatography (MeOH / CH ₂ Cl ₂ = 1/10) to obtain pure compound. [Note: For purification of compounds 2, 3 and 7, EtOAc: Hexane = 1: 2-1: 1 was used. ].

4- (5- (Hydroxymethyl) benzo [ d ] Oxazole -2-yl) phenol ( Nocabenzoxazole  F) (4- (5- (Hydroxymethyl) benzo [ d ] oxazol-2-yl) phenol ( nocarbenzoxazole  F)) (Compound 1):

Yield: 95%; Pale yellow solid; Melting point 230-232 ° C .; R _f = 0.25 (MeOH / CH ₂ Cl ₂ = 1/10); ¹ H NMR (300 MHz, DMSO-d6) δ 10.36 (1H, s), 8.01 (2H, d, J = 7.5 Hz), 7.64 (1H, d, J = 8.1 Hz), 7.62 (1H, s), 7.29 (1H, d, J = 8.1 Hz), 6.94 (2H, d, J = 7.5 Hz), 5.35 (1H, br s), 4.59 (2H, s); ¹³ C NMR (75 MHz, DMSO-d6) δ 162.8, 160.7, 148.9, 141.6, 139.2, 129.2, 123.3, 117.2, 116.9, 116.0, 109.9, 62.8; EI-MS m / z 241 (M ⁺ ), 207 (base), 148, 119; HRMS: Calcd for C ₁₄ H ₁₁ NO ₃ (M ⁺ ): 241.0739, found: 241.0744.

2- Phenylbenzo [ d ] Oxazole -5-yl) methanol {(2- Phenylbenzo [ d ] oxazol -5-yl) methanol} (Compound 2):

Yield: 97%; Pale yellow solid; Melting point 146-148 ° C .; R _f = 0.31 (MeOH / CH ₂ Cl ₂ = 1/20); ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.24-8.21 (2H, m), 7.73 (1H, s), 7.55-7.50 (4H, m), 7.35 (1H, d, J = 8.4 Hz), 4.80 ( 2H, s), 2.41 (1H, broad singlet); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 163.6, 150.3, 142.3, 137.9, 131.7, 129.0, 127.8, 127.2, 124.5, 118.6, 110.6, 65.5; EI-MS m / z 225 (M ⁺ , base), 224 (MH), 208, 194; HRMS: Calcd for C ₁₄ H ₁₁ NO ₂ (M ⁺ ): 225.0790, found: 225.0783.

2- (2,4-dimethoxyphenyl) benzo [ d ] Oxazole -5-yl) methanol {(2- (2,4-Dimethoxyphenyl) benzo [ d ] oxazol-5-yl) methanol} (Compound 3):

Yield: 92%; White solid; Melting point 134-136 ° C .; R _f = 0.3 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.05 (1H, d, J = 8.4 Hz), 7.70 (1H, s), 7.45 (1H, d, J = 8.4 Hz), 7.25 (1H, d, J = 8.7 Hz), 6.60 (1H, dd, J = 8.7, 2.1 Hz), 6.56 (1H, d, J = 2.1 Hz), 4.75 (2H, s), 3.99 (3H, s), 3.87 (3H, s) , 2.62 (1H, broad singlet); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 163.6, 162.2, 160.0, 149.6, 142.3, 137.6, 132.5, 123.9, 118.4, 110.2, 109.0, 105.5, 99.2, 65.5, 56.3, 55.8; EI-MS m / z 285 (M ⁺ , base), 284 (MH), 254, 148; HRMS: Calcd for C ₁₆ H ₁₅ NO ₄ (M ⁺ ): 285.1001, found: 285.1011.

2- (5- (Hydroxymethyl) benzo [ d ] Oxazole 2-yl) -5- Methoxyphenol  {2- (5- (Hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol} (Compound 4):

Yield: 93%; Pale yellow solid; Melting point 170-172 ° C .; R _f = 0.30 (MeOH / CH ₂ Cl ₂ = 1/15); ¹ H NMR (300 MHz, CDCl ₃ ) δ 7.90 (1H, d, J = 8.4 Hz), 7.68 (1H, d, J = 1.5 Hz), 7.55 (1H, d, J = 8.7 Hz), 7.36 (1H , d, J = 8.4, 1.5 Hz), 6.63 (1H, d, J = 2.4 Hz), 6.59 (1H, dd, J = 8.7, 2.4 Hz), 4.83 (2H, s), 3.88 (3H, s) ; ¹³ C NMR (75 MHz, CDCl ₃ ) δ 164.2, 163.7, 160.8, 148.6, 140.5, 138.1, 128.4, 124.1, 117.4, 110.5, 107.9, 103.8, 101.4, 65.5, 55.8; EI-MS m / z 271 (M ⁺ , base), 254, 240, 149; HRMS: Calcd for C ₁₅ H ₁₃ NO ₄ (M ⁺ ): 271.0845, found: 271.0851.

4- (5- (Hydroxymethyl) benzo [ d ] Oxazole -2-yl) -3- Methoxyphenol  {4- (5- (Hydroxymethyl) benzo [ d ] oxazol-2-yl) -3-methoxyphenol} (Compound 5):

Yield: 94%; Pale yellow solid; Melting point 214-216 ° C .; R _f = 0.18 (MeOH / CH ₂ Cl ₂ = 1/10); ¹ H NMR (300 MHz, CD ₃ OD) δ 7.93 (1H, d, J = 8.7 Hz), 7.68 (1H, s), 7.56 (1H, d, J = 8.7 Hz), 7.35 (1H, d, J = 8.4 Hz), 6.60 (1H, d, J = 1.8 Hz), 6.54 (1H, dd, J = 8.4, 1.8 Hz), 4.72 (2H, s), 3.95 (3H, s); ¹³ C NMR (75 MHz, CD ₃ OD) δ 164.1, 161.9, 143.0, 139.8, 133.3, 125.2, 118.3, 111.1, 109.2, 107.8, 107.6, 100.6, 99.7, 65.3, 56.3; EI-MS m / z 271 (M ⁺ , base), 254, 240, 149; HRMS: Calcd for C ₁₅ H ₁₃ NO ₄ (M ⁺ ): 271.0845, found: 271.0834.

4- (5- (Hydroxymethyl) benzo [ d ] Oxazole -2-yl) benzene-1,3- Dior  {4- (5- (Hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol} (Compound 6):

Yield: 92%; Pale yellow solid; Melting point 224-226 ° C .; R _f = 0.21 (MeOH / CH ₂ Cl ₂ = 1/10); ¹ H NMR (300 MHz, CD ₃ OD) δ 11.28 (1H, s), 10.45 (1H, s), 7.81 (1H, dd, J = 8.7, 2.1 Hz), 7.70 (1H, dd, J = 8.4, 1.8 Hz), 7.65 (1H, s), 7.33 (1H, d, J = 8.4 Hz), 6.50 (1H, d, J = 8.7 Hz), 6.45 (1H, s), 5.38 (1H, t, J = 4.5 Hz), 4.60 (2H, d, J = 4.5 Hz); ¹³ C NMR (75 MHz, CD ₃ OD) δ 162.9, 162.6, 159.6, 147.3, 139.8, 139.4, 128.7, 123.5, 116.1, 110.1, 108.7, 102.8, 102.0, 62.7; EI-MS m / z 257 (M ⁺ , base), 256 (MH), 240, 228; HRMS: Calcd for C ₁₄ H ₁₁ NO ₄ (M ⁺ ): 257.0688, found: 257.0692.

(2- (2,4-dimethoxyphenyl) benzo [ d ] Oxazole -5-yl) methanol {(2- (2,4-Dimethoxyphenyl) benzo [ d ] oxazol-5-yl) methanol} (Compound 7):

Yield: 95%; White solid; Melting point 130-132 ° C; R _f = 0.25 (EtOAc / Hexane = 1/2); ¹ H NMR (300 MHz, CDCl ₃ ) δ 7.86 (1H, dd, J = 8.1, 1.5 Hz), 7.75 (1H, d, J = 1.5 Hz), 7.73 (1H, s), 7.54 (1H, d, J = 8.4 Hz), 7.35 (1H, d, J = 8.1 Hz), 7.00 (1H, d, J = 8.4 Hz), 4.82 (2H, d, J = 6.0 Hz), 4.03 (3H, s), 3.99 (3H, s), 1.88 (1H, broad singlet, J = 6.0 Hz); ¹³ C NMR (75 MHz, CDCl ₃ ) δ 162.9, 152.2, 150.4, 149.4, 142.6, 137.8, 124.1, 121.4, 119.9, 118.3, 111.2, 110.5, 110.3, 65.7, 56.4, 56.3; EI-MS m / z 285 (M ⁺ , base), 284 (MH), 268, 163; HRMS: Calcd for C ₁₆ H ₁₅ NO ₄ (M ⁺ ): 285.1001, found: 285.0983.

4- (5- (Hydroxymethyl) benzo [ d ] Oxazole -2-yl) benzene-1,2- Dior  {4- (5- (Hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,2-diol} (Compound 8):

Yield: 91%; Pale yellow solid; Melting point 222-224 ° C .; R _f = 0.18 R _f = 0.18 (MeOH / CH ₂ Cl ₂ = 1/10); ¹ H NMR (300 MHz, DMSO-d6) δ 9.80 (1H, s), 9.53 (1H, s), 7.63 (H, d, J = 8.7 Hz), 7.62 (1H, s), 7.56 (1H, d , J = 2.1 Hz), 7.51 (1H, dd, J = 8.4, 2.1 Hz), 7.29 (1H, d, J = 8.7 Hz), 6.90 (1H, d, J = 8.4 Hz), 5.30 (1H, t , J = 4.2 Hz), 4.60 (2H, d, J = 4.2 Hz); ¹³ C NMR (75 MHz, DMSO-d6) δ 162.9, 149.2, 148.8, 145.6, 141.6, 139.1, 123.2, 119.4, 117.3, 116.8, 116.0, 114.1, 109.8, 62.8; EI-MS m / z 257 (M ⁺ , base), 256 (MH), 240, 228; HRMS: Calcd for C ₁₄ H ₁₁ NO ₄ (M ⁺ ): 257.0688, found: 257.0681.

result

The synthesis of the present invention begins with the esterification of commercially available 3-amino-4-hydroxybenzoic acid (Compound 9) (FIG. 1). Treatment of compound 9 with ethanol in the presence of H ₂ SO ₄ at reflux gave a substituted benzoic acid compound 10 in 85% yield. Subsequently, the reaction of substituted benzoic acid compounds 11-13 with SOCl ₂ at 60 ° C. yields the corresponding acid chlorides 14, 16 and 17, respectively. It is used directly in the next step without purification. Next, the main cyclodehydration reaction was studied using the main fragments in hand and using compounds 10 and 14 as model substrates (Table 1). POCl ₃ , PCl ₅ , H ₂ SO ₄ , P ₂ O ₅ , SOCl ₂ , CH ₃ SO ₃ H and CF ₃ SO ₃ H are dehydrated in other solvents such as DMF, acetic anhydride, 1,4-dioxane and pyridine Screened topically. All attempts to proceed to ring dehydration either failed (3, 4 and 7 in Table 1) or yielded compound 18 in low yield (1, 2, 5, 6 and 8-11 in Table 1). Fortunately, POCl ₃ was able to obtain compound 18 in 55% yield via mediated ring dehydration in 1,4-dioxane (12 in Table 1). Reducing or increasing the reaction time did not improve product yields (13 and 14 in Table 1). According to these optimized conditions, benzoxazole compound 19-21 was obtained by reaction of compound 10 with substituted benzoyl chloride compound 15-17, respectively. Table 1 shows the results of testing the optimum conditions for the ring dehydration reaction between compound 10 and compound 14.

Entry Reagent Solvent Time (h) Yield ^b  ( % ) One POCl ₃ DMF 0.5 15 2 POCl ₃ 1,4-dioxane 0.5 15 3 POCl ₃ Pyridine 0.5 N.R 4 H ₂ SO ₄ Ac ₂ O One N.R 5 POCl ₃ DMF 3 16 6 POCl ₃ 1,4-dioxane 3 29 7 PCl ₅ 1,4-dioxane 3 N.R 8 P ₂ O ₅ 1,4-dioxane 3 24 9 SOCl ₂ 1,4-dioxane 3 17 10 CH ₃ SO ₃ H 1,4-dioxane 3 25 11 CF ₃ SO ₃ H 1,4-dioxane 3 20 12 POCl ₃ 1,4- dioxane 15 55 13 POCl ₃ 1,4-dioxane 12 45 14 POCl ₃ 1,4-dioxane 24 35

Next, Compounds 18 and 21 were treated with BBr ₃ (boron tribromide) solution (1.0 M in CH ₂ Cl ₂ ) to yield Compounds 22 and 23, respectively. Compound 20 was selectively demethylated with BCl ₃ solution (1.0M in CH ₂ Cl ₂ ) and AlCl ₃ to yield compounds 24 and 25, respectively. Due to chelation with nitrogen or oxygen atoms of the benzoxazole ring, BCl ₃ is significantly more effective at demethylation at the 2 'position than at the 4' position. Since BCl ₃ does not selectively remove the separated arylmethyl groups, the inventors have some success by affecting the demethylation of the 4 ′ position with AlCl ₃ . Compound 25 was further methylated with BCl ₃ to form benzoxazole compound 26. Finally, Compounds 22, 19, 20, 24-26, 21 and 23 obtained target benzoxazole compounds 1-8 in high yield, respectively, using LiAlH ₄ (91-97%). . All target compounds 1-8 were determined by NMR ( ¹ H- and ¹³ C-) and MS data.

Nitric Oxide Production Inhibition

The biological activity of the synthesized benzoxazole compound 1-8 as an inhibitor of nitric oxide production was evaluated by measuring the amount of nitric oxide as an indicator of anti-inflammatory activity in RAW 264.7 cells stimulated with lipopolysaccharide (LPS). Due to the very short physiological half-life of nitric oxide and the presence of other waste-removing molecules, nitric oxide levels were indirectly assessed by its spectroscopic method using the Griess reaction, its stable metabolites, nitrite and nitrate concentrations. L-NMMA ( N ^G -Monomethyl-L-arginine acetate) was used as a positive control. Table 2 shows the inhibitory effect of nitric oxide production of nocabenzoxazole F (compound 1) and its derivatives (compounds 2-4 and 6-8).

Compound NO Production (% inhibition) ^{a, b} Proliferation ^a IC ₅₀
(μM) 1 μM 10 μM 1 μM 10 μM LPS 100.0 ± 1.8 (0.0) 100.0 ± 1.8 (0.0) One 120.6 ± 1.1 (-20.6) ** 97.2 ± 3.9 (2.8) * 111.9 ± 2.1 * 91.8 ± 4.8 26.24 2 107.7 ± 2.5 (-7.7) 92.2 ± 2.7 (7.8) 90.2 ± 3.1 84.6 ± 1.4 ** 29.11 3 117.7 ± 0.7 (-17.7) ** 92.6 ± 1.7 (7.4) 92.7 ± 3.8 103.1 ± 1.1 27.31 4 117.0 ± 2.3 (-17.0) ** 96.6 ± 1.5 (3.4) 99.2 ± 5.1 98.9 ± 4.1 21.01 6 115.0 ± 1.3 (-15.0) * 87.3 ± 0.7 (12.7) 104.4 ± 4.1 98.1 ± 5.1 28.26 7 114.6 ± 4.4 (-14.6) 110.4 ± 2.5 (-10.4) 97.1 ± 2.0 92.2 ± 4.0 39.91 8 90.6 ± 25.4 (9.4) 46.5 ± 3.3 (53.5) *** 99.9 ± 3.9 90.9 ± 6.2 8.17 L- NMMA 95.2 ± 1.1 (4.8) 80.5 ± 2.9 (19.5) ** 102.2 ± 2.5 100.7 ± 1.8 18.77

^The results are expressed as three means ± standard deviation. Statistical significance was based on differences from LPS treated groups (* P <0.05, ** P <0.01, *** P <0.001).

^b % Inhibition is shown in parentheses.

The effect of compound 1-8 on nitric oxide inhibition by induced macrophages was monitored (Table 2). Nocabenzoxazole G (Compound 5) was very unstable and readily degraded within 24 hours. LPS-induced nitric oxide production in macrophages was inhibited concentration-dependently by benzoxazole. All compounds reduced nitric oxide production at the lowest concentration (1 μM). However, compounds 6 and 8 showed the best inhibitory activity at the highest concentration (10 μM) compared to the control L-NMMA. In addition, cell viability was determined by MTT {3- [4,5-dimethylthiazol-2-yl] -2,5-diphenyltetrazolium bromide} to determine whether the inhibitory effect on nitric oxide production was due to the cytotoxicity of the test compound. The evaluation was based on the reduction of the tetrazolium salt to formazan crystals. As a result, Compounds 1-4 and 6-8 did not show significant cytotoxicity in RAW 264.7 cells at the concentrations used for evaluation. The concentrations required to inhibit nitric oxide production 50%, ie IC ₅₀ values, were evaluated with GraphPad Prism 4.0 software and were 26.24, 29.11, 27.31, 21.01, 28.26, 39.91 and 8.17 μM, respectively. Taken together, the compound 8 with catechol at position 2 of the benzoxazole structure showed better nitric oxide inhibitory effect than the positive control L-NMMA.

Claims (4)

A) Ethyl 3-amino-4-hydroxybenzoate was added to anhydrous 1,4-dioxane, and the compound 16 represented by the formula (16) was added to the stirred solution under normal temperature and nitrogen atmosphere, warmed to 90 ° C and stirred, and then POCl ₃ dropwise addition and stirring to obtain ethyl 2- (2,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate;
B) Ethyl 2- (2,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate was added to anhydrous CH ₂ Cl ₂ and stirred in a solution of BCl ₃ dropwise at 0 ° C. under nitrogen atmosphere. After addition, warming to room temperature and stirring to obtain ethyl 2- (2-hydroxy-4-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate; And
C) Add ethyl 2- (2,4-dimethoxyphenyl) benzo [ d ] oxazole-5-carboxylate to anhydrous CH ₂ Cl ₂ , add AlCl ₃ to a stirred solution under nitrogen atmosphere, and stir at room temperature. After completion of the reaction, H ₂ O was slowly added and stirred to obtain ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate;
D) Ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate was added to anhydrous CH ₂ Cl ₂ , and stirred in a solution of BCl ₃ at 0 ° C. under a nitrogen atmosphere. After dropwise addition, warming to room temperature and stirring to obtain ethyl 2- (2,4-dihydroxyphenyl) benzo [ d ] oxazole-5-carboxylate; And
E) ethyl 2- (2-hydroxy-4-methoxyphenyl) benzo [ d ] oxazole-5-carboxylate or ethyl 2- (4-hydroxy-2-methoxyphenyl) benzo [ d ] oxazole To a solution of -5-carboxylate or ethyl 2- (2,4-dihydroxyphenyl) benzo [ d ] oxazole-5-carboxylate in anhydrous THF (tetrahydrofuran), add LiAlH ₄ solution to 0 ° C and nitrogen. After dropwise addition in an atmosphere, the mixture was warmed to room temperature, stirred, and cooled to 0 ° C after completion of the reaction, and saturated Na ₂ CO ₃ aqueous solution was slowly added to quench excess LiAlH ₄ and stirred, respectively, corresponding to 2- (5-). (Hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol, 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -3-methoxyphenol Or 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol; obtaining nocabenzoxazole or a derivative thereof
<Formula 16>

Novel 2- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol
Novel 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol
2- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) -5-methoxyphenol, 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl)- Anti-inflammatory pharmaceutical composition containing at least one selected from 3-methoxyphenol and 4- (5- (hydroxymethyl) benzo [ d ] oxazol-2-yl) benzene-1,3-diol

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