JPWO2011052759A1 - Novel nitroxyl radical compound and method for producing the same - Google Patents

Abstract

The present invention relates to a method for producing a novel nitroxyl radical and a compound thereof. Furthermore, the nitroxyl radical obtained by the production method of the present application has a therapeutic effect on gastric ulcer and hypertension. [Selection figure] None

Description

  This application claims priority to US Provisional Application No. 61 / 256,184, filed Oct. 29, 2009, the entire disclosure of which is incorporated herein. is there.

  The present invention relates to a 2,6-substituted piperidine-based nitroxyl radical and a novel synthesis method thereof.

  Nitroxyl radicals are known as relatively stable radicals, but they react sensitively to various active oxygen and redox substances, so that the effects of nitroxyl radicals on various diseases involving free radicals are reduced. Expected.

Diseases involving free radicals are different in free radical species that cause the disease, and therefore, effective radical scavenging species that specifically act on specific diseases are desired. It is known that the reactivity differs depending on the basic skeleton and side chain of the nitroxyl radical according to previous studies (F. Vianello, et al., Kinetics of nitroxide spin label removal in biological systems: An in vitro and in vivo ESR study.Magn Reson Imag, 1995.13 (2): 219-226.; WR Couette, et al., Influencing of chemical structure of the brain. 1985.41 (7): 1165-1172; . Eshita, et al, Kinetic study on ESR signal decay of nitroxyl radicals, potent redox probes for in vivo ESR spectroscopy, caused by reactive oxygen species.Biochim Biophys Acta, 2002.1573 (2): 156-64).. For example, nitroxyl radicals have been reported to react with ascorbic acid, thiols (metal-catalyzed reactions), mitochondrial respiratory chain, NADPH, cytochrome P450, etc. in cells, and induce apoptosis or via cytochrome P450. It has been reported that production of H ₂ O ₂ and the like cause cytotoxicity. On the other hand, nitroxyl radicals having a tetraethyl group have been reported to have a significantly reduced reactivity with ascorbic acid.

  In recent years, by replacing the α-position substituent of the nitroxyl radical of the isoindoline skeleton with an ethyl group, compared with TEMPO, p-phenylenediamine (N. Kocherginsky and HM Swartz, Chemical reactivity of nitrogens, in Nitroxide Spin Labels: Reactions in Biology and Chemistry, N. Kocherginsky and HM Swartz, Editors. 1995, CRC Press: Boca Raton. P. 27-66. comparative study of the reduction by associate of 1,1,3,3-tetraethylindolinin-2-yloxyl and of 1,1,3,3-tetramethylindolinin-2-yloxyl. J Chem Soc, Perkin Trans 1,2000: 1181.) It is reported to increase. In addition, it has been reported that a substance in which the α-position carbon of the nitroxyl radical of the imidazole skeleton is substituted with an ethyl group exhibits reduction resistance to ascorbic acid (AA Bobko, et al., Reverseable). reduction of nitroxides to hydroxylamines: roles for ascorbate and glutathione. Free Radic Biol Med, 2007.42 (3): 404-12.). Thus, it is thought that the reactivity with respect to various substances can be controlled by changing the substituent at the α-position of the nitroxyl radical.

  A conventional method for synthesizing 2,6-substituted nitroxyl radicals is shown below.

  As shown in the figure, the conventional method has many reaction steps and is complicated, so that the yield of the 2,6-position substituted nitroxy radical is very low. In order to solve this problem, development of a method for synthesizing a nitroxy radical in which various substituents are introduced into the 2,6-positions in a simple and high yield is desired.

  In addition, a method for producing a nitroxy radical developed by the inventor is disclosed in International Patent Publication No. WO2008 / 093881, and this patent application is incorporated herein by this reference.

  The inventors have conducted various studies to solve the above problems, and as a result, have developed a simple and high-yield synthesis method for 2,6-position substituted piperidine-based nitroxyl radicals. Furthermore, when the effectiveness of the nitroxyl radical obtained by the production method was examined, it was revealed that it has a gastric mucosal damage suppressing effect and a blood pressure lowering effect in an adjuvant arthritis model rat.

  According to the method of the present invention, when 1,2,2,6,6-pentaalkylpiperidin-4-one is used as a starting material and a carbonyl compound is reacted in the presence of an ammonium salt, piperidone substituted at the 2- and 6-positions Further, by oxidizing this compound, a nitroxy radical substituted at the 2- and 6-positions can be obtained.

  A feature of the present invention is that a 2,6-substituted piperidine having an alkyl group such as a methyl group on a nitrogen atom is used. When 1,2,2,6,6-pentamethylpiperidin-4-one having a methyl group on nitrogen is used, it is remarkable compared to 2,2,6,6-tetramethylpiperidin-4-one An increase in yield and a reaction promoting effect were observed. Therefore, in this reaction, it is most important to use a piperidine compound in which an alkyl group such as a methyl group is substituted on nitrogen as a starting material.

  Examples of the ketone used in the production method of the present application include cyclic / chain carbonyl compounds such as cyclohexanone, pyran ring, thiopyran ring, acetaminophen, and benzaldehyde.

  Specific carbonyl compounds and corresponding products (piperidone) are shown in Table 2.

  As apparent from the above table, the carbonyl compound having a cyclic structure and the linear carbonyl compound have different selectivity for the pyrrolidone product. That is, when a carbonyl compound having a ring structure is used as a raw material, a disubstituted product is obtained as a main product, whereas when a linear carbonyl compound is used as a raw material, a monosubstituted product is mainly used. Obtained as product. In the case of a carbonyl compound having a phenyl group, a mono-substituted product is selectively obtained. Furthermore, when 2-adamantanone is used as the carbonyl compound, both mono- and di-substituted products are obtained.

As described above, the selectivity of mono- and di-substituents with respect to the piperidone derivative product varies depending on the steric structure of the ketone, and TEMPONE is changed from NH ₄ Cl to ¹⁵ NH ₄ Cl. When the invention is carried out using the above, the reaction mechanism shown in the following scheme is presumed because [ ¹⁵ N] TEMPONE is obtained.

  In the production method of the present invention, ammonium chloride is used as the base used in the production step of the piperidone derivative, but is not limited thereto.

  In addition, when cesium hydroxide (CsOH) or benzyltrimethylammonium hydroxide (Triton B) is used together with ammonium chloride, an improvement in yield is observed.

  In general, oxidation of a piperidone derivative to a nitroxy radical derivative uses a tungsten catalyst and hydrogen peroxide, but is not limited thereto. MCPBA is used for oxidation to the 2,6-positioned adamantane-substituted nitroxyl radical.

  The nitroxy radical derivative obtained by the production method of the present invention was dissolved in 10 mM PB (pH 7.4) (50 μM) and measured at room temperature to calculate the ESR parameter. However, 12 contains 0.5% EtOH from the viewpoint of solubility. The results are shown in the table below.

式中、
Ｒ^１〜Ｒ^４は、独立して、Ｃ_１〜Ｃ_１０のアルキルであるか、或いは、Ｒ^１及びＲ^２、又はＲ^３及びＲ^４が共に結合して、（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＯ（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＳＯ_２（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＮＸ（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＣＹ（ＣＨ_２）_ｎ、
Ｘは、ＣＨ_３ＣＯ、ＣＦ_３ＣＯ、又はＲ^１ＣＯであり、
Ｙは、酸素（＝Ｏ）、（ＯＣＨ_２）_ｎであり、
ｎは２〜４の整数であり、
当該方法は、
化学式２で示される化合物であって、

According to one embodiment of the present invention, the present invention is a method for producing a nitroxy radical compound represented by Formula 1,

Where
R ^{1 to} R ⁴ are independently C _{1 to} C ₁₀ alkyl, or R ¹ and R ² , or R ³ and R ⁴ are bonded together to form (CH ₂ ) _n , (CH _{2) n O (CH 2)} n, (CH 2) n SO 2 (CH 2) n, (CH 2) n NX (CH 2) n, (CH 2) n CY (CH 2) n,
X is CH ₃ CO, CF ₃ CO, or R ¹ CO;
Y is oxygen (= O), a _{(OCH 2) n,}
n is an integer of 2 to 4,
The method is
A compound represented by Formula 2,

Where
R ^{5 to} R ⁸ are independently a C _{1 to} C ₂₀ alkyl compound represented by Formula 2,
A compound represented by Formula 3,

Where
R ^{1 to} R ⁴ are independently C _{1 to} C ₁₀ alkyl, R ¹ and R ² , or R ³ and R ⁴ are bonded together to form (CH ₂ ) _n , (CH ₂ ) _n O ( _{CH 2) n, (CH 2} ) n S (CH 2) n, (CH 2) n NX (CH 2) n, (CH 2) n CY (CH 2) n,
X is CH ₃ CO, CF ₃ CO, or R ¹ CO;
Y is oxygen (= O), a _{(OCH 2) n,}
n is an integer of 2 to 4, and a compound represented by Formula 3 is reacted in the presence of an ammonium salt to produce a 2,6-substituted-4-piperidone derivative;
Oxidizing the 2,6-substituted-4-piperidone derivative;
A process for producing the nitroxy radical compound is provided.

  According to another embodiment of the present invention, a compound produced by the above production method is provided. In this case, the compound can be used for the treatment of gastric mucosal damage (gastric ulcer) or hypertension.

  Furthermore, according to another embodiment of the present invention, there is provided a pharmaceutical composition comprising the above compound and a pharmaceutically acceptable carrier.

  Moreover, according to another embodiment of the present invention, there is provided a method for treating gastric mucosal damage (gastric ulcer), the method comprising the step of administering the compound to a subject, and treating hypertension. A method is provided comprising the step of administering the compound to a subject. In this case, in the two methods, the compound is preferably administered in an amount of about 0.0001-4 g / day. Further, in treating gastric mucosal damage (gastric ulcer), when the compound is 18 mM, it can be administered at a dose of 90 μmol / kg.

FIG. 1 is a graph showing the evaluation of cytotoxicity of a nitroxyl radical compound according to an embodiment of the present invention. FIG. 2 is a graph showing the effect of the nitroxyl radical compound according to one embodiment of the present invention on the gastric mucosa. FIG. 3 is a graph showing the effect of a nitroxyl radical compound according to an embodiment of the present invention on blood pressure fluctuations.

  Hereinafter, the present invention will be described with reference to examples, but it is understood that the present invention is not limited thereto.

Reagents were purchased from Sigma-Aldrich, Wako Pure Chemical Industries, Ltd. and used without purification. The melting point was measured using a micro melting point measuring apparatus MP-500P (Yanaco). Mass spectrometry was measured with JMS-600H (JEOL), and 3-nitrobenzyl alcohol was used as a matrix. The IR spectrum was measured by a solution method or a total reflection method using FT / IR-4200 (JASCO). ¹ H-NMR was measured using Unity INOVA 400 (Varian), and tetramethylsilane (TMS) was used as a standard substance. The chemical shift (δ) was based on TMS (0 ppm), and the coupling constant was expressed in Hz. Elemental analysis was requested from Kyushu University Faculty of Science, Central Elemental Analysis Laboratory.

7-Azadispiro [5.1.5.3] hexadecan-15-one (3a)
Method A
1.55 g (10 mmol) of 2,2,6,6-tetramethylpiperidin-4-one (1) and 2.94 g (30 mmol) of cyclohexanone (a) were dissolved in 15 ml of DMSO, and 3.21 g (60 mmol) of NH ₄ Cl. ) Was added at room temperature. The solution was stirred at 60 ° C. for 20 hours. The reaction mixture was diluted by adding 40 mL of water and acidified with a 7% aqueous hydrochloric acid solution. This was washed with ether and the aqueous layer was made alkaline with 10% K ₂ CO ₃ aqueous solution. Further, this was extracted with ethyl acetate, washed with water, the organic layer was dried over sodium sulfate, and the solvent was completely distilled off. Thereafter, the residue was purified by silica gel column chromatography (hexane, ethyl acetate) and recrystallized to obtain 376 mg (yield: 16%) of white crystals.

Method B
1.69 g (10 mmol) of 1,2,2,6,6-pentamethylpiperidin-4-one (2) and 2.94 g (30 mmol) of cyclohexanone (a) were dissolved in 15 ml of DMSO, and 3.21 g (60 mmol) of NH ₄ Cl. ) Was added at room temperature. The solution was stirred at 60 ° C. for 5 hours. The reaction mixture was diluted by adding 40 mL of water and acidified with a 7% aqueous hydrochloric acid solution. This was washed with ether and the aqueous layer was made alkaline with 10% K ₂ CO ₃ aqueous solution. Further, this was extracted with ethyl acetate, washed with water, the organic layer was dried over sodium sulfate, and the solvent was completely distilled off. Thereafter, the product was purified by silica gel column chromatography (hexane, ethyl acetate) and recrystallized to obtain 799 mg of white crystals (yield: 34%).
mp: 103 ° C. (hexane-EtOAc, lit. mp 101-103 ° C. [19]);
MS (FAB ⁺ ): 236.3 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1689 (C═O);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.36 to 1.46 (8H, m), 1.47 to 1.56 (8H, m), 1.68 to 1.71 (4H) , M), 2.31 (4H, s);
Found: C, 76.51; H, 10.68; N, 5.93%. Calc. forC ₁₅ H ₂₅ NO: C, 76.55; H, 10.71; N, 5.95%.

7-Aza-3,11-dioxadaspiro [5.1.5.3] hexadecan-15-one (3b)
According to Method B, cyclohexanone (a) was changed to 3.00 g (30 mmol) of tetrahydro-4H-pyran-4-one (b). The obtained crystals were recrystallized from Hexane-EtOAc.
Yield: 32%;
mp: 167 ° C;
MS (FAB ⁺ ): 240.1 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1692 (C═O);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.64 (8H, t, J = 5.6 Hz), 2.40 (4H, s), 3.54 to 3.60 (4H, m ), 3.81-3.87 (4H, m);
Found: C, 65.17; H, 8.85; N, 5.86%. Calc. _{for C 13 H 21 NO 3:} C, 65.25; H, 8.84; N, 5.85%.

7-Aza-3,11-dithiadispiro [5.1.5.3] hexadecan-15-one (3c)
According to Method B, cyclohexanone (a) was changed to 3.49 g (30 mmol) of tetrahydro-4H-thiopyran-4-one (c).
Yield: 30%;
mp: 155-157 ° C. (EtOAc);
MS (FAB ⁺ ): 272.2 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1697 (C═O);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 0.8 (1 s, br s), 1.75-1.90 (8 H, m), 2.27 (4 H, s), 2.43 -2.50 (4H, m), 2.88-2.95 (4H, m);
Found: C, 57.54; H, 7.78; N, 5.11%. Calc. _{for C 13 H 21 NOS 2:} C, 57.52; H, 7.80; N, 5.16%.

3,11-Diacetyl-3,7,11-trithiadispiro [5.1.5.3] hexadecan-15-one (3d)
9-Acetyl-2,2-dimethyl-1,9-diazaspiro [5.5] undecan-4-one (3d ′)
1,2,2,6,6-pentamethylpiperidin-4-one (2) 0.845 g (5 mmol), 1-acetyl-4-piperidone (d) 2.12 g (15 mmol) as in Method B, A mixture of 1.61 g (30 mmol) of NH ₄ Cl and 12 ml of DMSO is heated and stirred at 58 to 62 ° C. for 8 hours. After diluting with 40 mL of water and acidifying with 7% aqueous hydrochloric acid, the neutral part was extracted and removed with ether. The aqueous layer was adjusted to pH 9-10 with 10% aqueous K ₂ CO ₃ solution, extracted with chloroform, and dried over sodium sulfate. The oily substance obtained by evaporating the solvent under reduced pressure was purified by silica gel column chromatography (1-5% MeOH / CHCl ₃ ), recrystallized from hexane-EtOAc, and 3d 328 mg, 3d '556 mg were obtained. Obtained.
3d;
Yield: 21%;
mp: 164.2 ° C;
MS (FAB ⁺ ): 322.3 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1687 (C═O), 1634 (N- Ac );
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.58-1.66 (8H, m), 2.08 (6H, s), 2.38 (4H, s), 3.37- 3.71 (8H, m);
Found: C, 62.92; H, 8.62; N, 12.12%. Calc. _{for C 17 H 27 N 3 O} 3: C, 63.53; H, 8.47; N, 13.07%.
3d ′;
Yield: 47%;
mp: 80.4 [deg.] C (hexane-EtOAc);
MS (FAB ⁺ ): 239.2 (M ⁺⁺ 1);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.21 (6H, d, J = 13.6 Hz), 1.50-1.69 (4H, m), 2.05 (3H, s ), 2.28 (4H, d, J = 9.2 Hz), 3.34-3.78 (4H, m);
Found: C, 64.86; H, 9.28; N, 11.67%. Calc. _{for C 13 H 22 N 2 O} 2: C, 65.51; H, 9.30; N, 11.75%.

1,4,14,17-Tetraoxa-9-azetrastropiro [4.2.1.2.4.2.3.2] tetracosan-21-one (3e)
According to Method B, cyclohexanone (a) was changed to 4.69 g (30 mmol) of 1,4-cyclohexanedione monoethylene acetal (e) and recrystallized from Hexane-EtOAc to obtain 3e 1.12 g.
Yield: 32%;
mp: 190.6 ° C;
MS (FAB ^<+> ): 352.4 (M ⁺⁺ 1).
ν _max / cm ⁻¹ : 1699 (C═O), 1083 (—O—CH ₂ —CH ₂ —O—);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.56-1.74 (12H, m), 1.85-1.92 (4H, m), 2.32 (4H, s), 3.93 (8H, t, J = 2.8 Hz);
Found: C, 64.95; H, 8.30; N, 4.05%. Calc. _{for C 19 H 29 NO 5:} C, 64.93; H, 8.32; N, 3.99%.

2,2,6-Trimethyl-6-phenylpiperidin-4-one (3f)
According to Method B, cyclohexanone (a) was changed to 3.60 g (30 mmol) of acetophenone (f).
Yield: 16%;
mp: 109.5 ° C. (EtOAc);
MS (FAB ^<+> ): 218.2 (M ⁺⁺ 1).
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.20 (3H, s), 1.31 (3H, s), 2.23 to 2.50 (4H, m), 2.34 ( 3H, s), 4.17-4.20 (1H, m), 7.11-7.33 (4H, m);
Found: C, 77.92; H, 8.78; N, 6.44%. Calc. _{for C 19 H 29 NO 5:} C, 77.38; H, 8.81; N, 6.45%.

Methyl 3- (2,6,6-trimethyl-4-oxopiperidin-2-yl) propanoate (3g)
According to Method B, cyclohexanone was changed to methyl levulinate (g). This compound has been obtained as a mixture with a disubstituted product and has not yet been isolated. The structure was identified by methyl 3- (2,6,6-trimethyl-4-oxoperidin-1-oxyl-2-yl) propanoate (4 g) obtained by oxidizing 3 g.

2,2-Dimethyl-6-p-tolypiperidin-4-one (3h)
According to Method B, cyclohexanone (a) was changed to 3.60 g (30 mmol) of p-tolualdehyde (h).
Yield: 10%;
mp: 109.5 ° C (hexane);
MS (FAB ⁺ ): 218.2 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1693 (C═O);
¹ H-NMR (500 MHz; CDCl ₃ ) δ (ppm): 1.20 (3H, s), 1.31 (3H, s), 1.53 (1H, brs), 2.25 to 2.50 ( 4H, m), 2.34 (3H, s), 4.18 (1H, dd, J ₁ = 11.5 Hz, J ₂ = 3.5 Hz), 7.16 (2H, d, J = 8.0 Hz) ), 7.29 (2H, d, J = 8.0 Hz);
Found: C, 77.55; H, 8.82; N, 6.53%. Calc. _{for C 14 H 19 NO: C} , 77.38; H, 8.81; N, 6.45%.

3,11-Trifluoroacetyl-3,7,11-trithiadispiro [5.1.5.3] hexadecan-15-one (3i)
9-Trifluoroacetyl-2,2-dimethyl-1,9-diazaspiro [5.5] undecan-4-one (3i ′)
According to Method B, cyclohexanone (a) was changed to 5.85 g (30 mmol) of 1-trifluoroacetyl-4-piperidone (i). The neutral part was removed by extraction with EtOAc, adjusted to pH 9-10, and extracted with CHCl ₃ . Recrystallization from Hexane-EtOAc gave 3i 192 mg, 3i ′ 126 mg.
3i;
Yield: 4%;
mp: 172.6-173.2 ° C;
MS (FAB ⁺ ): 430.2 (M ⁺⁺ 1);
^{_{ν max / cm -1: 1688 (}} - N-CO -CF 3);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.64-1.77 (8H, m), 2.42 (4H, s), 3.57-3.82 (8H, m);
Found: C, 47.62; H, 5.01; N, 9.79%. Calc. _{for C 17 H 21 F 6 N} 3 O 3: C, 47.56; H, 4.93; N, 9.79%.
3i ';
Yield: 4%;
mp: 85.8-86.4 ° C. (hexane-EtOAc);
MS (FAB ⁺ ): 293.1 (M ⁺⁺ 1);
^{_{ν max / cm -1: 1692 (}} - N-CO -CF 3);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.23 (3H, s), 1.25 (3H, s), 1.62-1.80 (4H, m), 2.30 ( 2H, s), 2.33 (2H, s), 3.54-3.96 (4H, m);
Found: C, 53.30; H, 6.44; N, 9.58%. Calc. _{for C 13 H 19 F 3 N} 2 O 2: C, 53.42; H, 6.55; N, 9.58%.

Method C
7-Aza-15-oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4a)
7-Azadispiro [5.1.5.3] hexadecan- 15-one and (3a) 235mg (0.99mmol) was dissolved in ethanol _{10mL, Na 2 WO 4 · 2H} 2 O 50mg The (0.15 mmol) was added, To this solution was slowly added 2 mL of H ₂ O ₂ (30%). This was stirred at room temperature for 24 hours. _Then quenched added K 2 CO _3, and extracted with chloroform. Subsequently, the solvent was completely distilled off and purified by recrystallization (hexane) to obtain 208 mg (yield: 83%) of slightly yellow scale crystals.
mp: 114.2 ° C (lit. mp 114-116 ° C);
MS (FAB ^<+> ): 250.3 (M ^<+> );
ν _max / cm ⁻¹ : 1717 (C═O);
Found: C, 71.86; H, 9.62; N, 5.57%. Calc. _{for C 15 H 24 NO 2:} C, 71.96; H, 9.66; N, 5.59%.

7-Aza-3,11-dioxa-15-oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4b)
According to Method C, 7-Aza-3,11-dioxadaspiro [5.1.5.3] hexadecan-15-one (3b) 130 mg (0.54 mmol) was used as a starting material. Recrystallization from EtOAc gave 4 mg 110 mg (0.43 mmol).
Yield: 80%;
mp: 149.5 ° C;
MS (FAB ^<+> ): 255.2 (M ^< + ^{> +} 2);
ν _max / cm ⁻¹ : 1717 (C═O);
Found: C, 61.23; H, 7.97; N, 5.40%. Calc. _{for C 13 H 20 NO 4:} C, 61.40; H, 7.93; N, 5.51%.

7-Aza-3,11-dithia-3,3,11,11,15-pentaoxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4c)
7-Aza-3,11-dithiadispiro [5.1.5.3] hexadecan-15-one (3c) 210 mg (0.78 mmol), Na ₂ WO ₄ .2H ₂ O 118 mg (0.36 mmol), MeOH 3 ml And 1 mL of H ₂ O ₂ (30%) were mixed and stirred at room temperature for 48 hours. K ₂ CO ₃ was added, extraction was performed with a mixed solvent of CHCl ₃ : MeOH = 3: 1, and the solvent was distilled off. This was recrystallized from water to obtain 80 mg (0.23 mmol) of 4c.
Yield: 29%;
mp: 212.5-214.2 ° C;
MS (FAB ^<+> ): 351.0 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1712 (C═O), 1287 (SO ₂ ), 1122 (SO ₂ );
Found: C, 44.38; H, 5.79; N, 3.93%. Calc. _{for C 13 H 21 NO 5 S} 2: C, 44.56; H, 5.75; N, 4.00%.

3,11-Diacetyl-15-oxo-3,7,11-trithiadispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4d)
According to Method C, 3,11-Diacetyl-3,7,11-trithiadispiro [5.1.5.3] hexadecan-15-one (3d) was used as a starting material.
mp: 162-163 ° C;
MS (FAB ⁺ ): 337.3 (M ⁺⁺ 1);
^{_{ν max / cm -1: 1720 (}} C = O), 1637 (- N-CO -CH 3);

1,4,14,17-Tetraoxa-9-aza-21-oxotetraspiro [4.2.1.2.4.2.3.2.2] tetracos-9-yl-9-oxyl (4e)
According to Method C, 1,4,14,17-Tetraoxa-9-azetratraspiro [4.2.1.2.4.2.3.2] tetracosan-21-one (3e) was used as a starting material.
mp: 183.2-184.2 ° C;
MS (FAB ⁺ ): 367.3 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1713 (C═O);
Found: C, 62.18; H, 7.38; N, 3.88%. Calc. _{for C 19 H 28 NO 6:} C, 62.28; H, 7.70; N, 3.82%.

7-Aza-3,11,15-trioxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4e ')
1,4,14,17-Tetraoxa-9-azetratraspiro [4.2.1.2.4.2.3.2.2] tetracosan-21-one (3e) (536 mg, 1.53 mmol) in water (2 mL) and acetic acid (5 mL) 10% HCl was added dropwise and stirred at 70 ° C. The solution was diluted with water, the pH was adjusted to 7, and extracted with ethyl acetate. The organic layer was dried over sodium sulfate and the solvent was completely distilled off. Thereafter, the resultant was separated and purified by silica gel column chromatography hexane / EtOAc, 1: 4), and recrystallized (hexane, ethyl acetate) from 7-Aza-3,11,15-trioxodispiro [5.1.5.3] hexadecane. 330 mg (1.25 mmol) of white solid was obtained.
Yield: 82%
mp: 99.7 ° C;
MS (FAB ⁺ ): 264.2 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1705 (C═O);
¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.24 (1H, brs), 1.87-2.05 (8H, m), 2.29-2.67 (8H, m), 2.49 (4H, s);
Found: C, 76.51; H, 10.68; N, 5.93%. Calc. _{for C 15 H 25 NO: C} , 76.55; H, 10.71; N, 5.95%.
Subsequently, 215 mg (0.82 mmol) of 7-Aza-3,11,15-trioxodispiro [5.1.5.3] hexadecane obtained was oxidized according to Method C to obtain 4e′170 mg (0.61 mmol). It was.
Yield: 75%
mp: 161.5 ° C;
MS (FAB ⁺ ): 279.3 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1708 (C═O);
Found: C, 64.71; H, 7.25; N, 5.06%. Calc. _{for C 15 H 20 NO 4:} C, 64.73; H, 7.24; N, 5.03%.

Methyl 3- (2,6,6-trimethyl-4-oxoperidin-1-oxyl-2-yl) propanoate (4 g)
According to Method C, 147 mg of a mixture of methyl 3- (2,6,6-trimethyl-4-oxoperidin-2-yl) propanoate (3 g) and a disubstituted product was used as a raw material to obtain a red oily substance, 37 mg (0.15 mmol ).
MS (FAB ^<+> ): 242.2 (M ^<+> );
ν _max / cm ⁻¹ : 1720 (C═O);
Found: C, 58.93; H, 8.27; N, 5.65%. Calc. _{for C 12 H 20 NO 4:} C, 59.49; H, 8.32; N, 5.78%.

3,11-Trifluoroacetyl-3,7,11-trithia-15-oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4i)
According to Method C, 3,11-Trifluoroacetyl-3,7,11-trithiadispiro [5.1.5.3] hexadecan-15-one (3i) 130 mg (0.30 mmol) was used as starting material, and 4i 57 mg (0.13 mmol) )
Yield: 42%
MS (FAB ⁺ ): 445.1 (M ⁺⁺ 1);
Found: C, 44.90; H, 4.46; N, 9.25%. Calc. _{for C 17 H 20 F 6 N} 3 O 4: C, 45.95; H, 4.54; N, 9.46%.

^{7- [15 N] -Aza-15} -oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (5a)
NH ₄ Cl was changed to ¹⁵ NH ₄ Cl, and the same procedure as in 7-Aza-15-oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4a) was performed.
mp: 115.4-117.4 ° C. (hexane);
MS (FAB ⁺ ): 251.3 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1717 (C═O);
Found: C, 71.67; H, 9.79; N, 5.67%. Calc. _{^{for C 15 H 24 15 NO 2}} : C, 71.68; H, 9.62; N, 5.57%.

^{7- [15 N] -Aza-3,11} -Dioxa-15-oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (5b)
NH ₄ Cl is changed to ¹⁵ NH ₄ Cl, and the same operation as 7-Aza-3,11-Dioxa-15-oxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4b) is performed. It was.
mp: 171.7-172.0 ° C. (EtOAc);
MS (FAB ^<+> ): 256.23 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1719 (C═O);
Found: C, 61.18; H, 7.90; N, 5.52%. Calc. _{^{for C 13 H 20 15 NO 4}} : C, 61.16; H, 7.90; N, 5.48%.

^{7- [15 N] -Aza-3,11,15} -trioxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (5c)
NH ₄ Cl was changed to ¹⁵ NH ₄ Cl, and the same procedure as 7-Aza-3,11,15-trioxodispiro [5.1.5.3] hexadec-7-yl-7-oxyl (4e ′) was performed. .
mp: 165.8 ° C. (Hexane-EtOAc);
MS (FAB ⁺ ): 280.22 (M ⁺⁺ 1);
ν _max / cm ⁻¹ : 1707 (C═O);
Found: C, 64.47; H, 7.22; N, 5.01%. Calc. _{^{for C 15 H 20 15 NO 4}} : C, 64.50; H, 7.22; N, 5.01%.

2,2-diethyl-6,6-dimethylpiperidin-4-one (1)
1.69 g (10 mmol) of 1,2,2,6,6-pentamethylpiperidin-4-one and 5.16 g (60 mmol) of 3-pentanone were dissolved in 28 ml of DMSO, and 1.50 g (10 mmol) of CsOH, NH ₄ Cl 3.22 g (60 mmol) was added at room temperature. The solution was stirred at 80 ° C. for 4 hours. An appropriate amount of water was added for dilution, and the mixture was acidified with a 7-8% aqueous hydrochloric acid solution. This was washed with ether and the aqueous layer was made alkaline with aqueous K ₂ CO ₃ solution. Further, this was extracted with ethyl acetate, the organic layer was dried over sodium sulfate, and the solvent was completely distilled off. Thereafter, the residue was purified by silica gel column chromatography (hexane, ethyl acetate) to obtain 25 mg (yield: 1%) of a pale yellow oily substance.

Method B
2,2-dimethyl-6,6-dipropylpiperidin-4-one (3)
1.69 g (10 mmol) of 1,2,2,6,6-pentamethylpiperidin-4-one and 3.43 g (30 mmol) of 4-heptanone were dissolved in 16 ml of DMSO, 2 ml (4.40 mmol) of TritonB, NH ₄ 2.68 g (50 mmol) of Cl was added at room temperature. The solution was stirred at 50 ° C. for 4.5 hours. An appropriate amount of water was added for dilution, and the mixture was acidified with a 7-8% aqueous hydrochloric acid solution. This was washed with ether and the aqueous layer was made alkaline with aqueous K ₂ CO ₃ solution. Further, this was extracted with ethyl acetate, the organic layer was dried over sodium sulfate, and the solvent was completely distilled off. Then, it refine | purified by silica gel column chromatography (hexane, ethyl acetate), and the obtained crystal was recrystallized from hexane to obtain 368 mg (yield: 17%) of pale yellow needle-like crystals. mp: 53.4-55.0 ° C .; MS (FAB ⁺ ): 212.2 (M ⁺ +1); v _max / cm ⁻¹ : 1690 (C═O); ¹ H-NMR (300 MHz; CDCl ₃ ) δ (ppm): 0.88 (6H, t, J = 7.05 Hz), 1.21 (6H, s), 1.26-1.45 (8H, m), 2.23 (2H, s) 2.26 (2H, s); Found: C, 73.74; H, 11.88; N, 6.67%. Calc. _{for C 13 H 25 NO: C} , 73.88; H, 11.92; N, 6.63%.

2,2-dimethyl-6,6-dipropyl-4-oxo-piperidin-1-yl 1-oxyl (4)
According to Method A, 2,2-dimethyl-6,6-dipropylpiperidin-4-one (3) 59 mg (0.28 mmol) was used as a starting material. The reaction solution was stirred overnight at room temperature. Moreover, the progress of the reaction was confirmed by TLC, and H ₂ O ₂ and Na ₂ WO ₄ · 2H ₂ O were appropriately added. After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate) to obtain 38 mg (yield: 60%) of an orange oily substance. MS (FAB ^<+> ): 226.3 (M ^<+> ); _vmax / cm ^{< -1} >: 1720 (C = O); Found: C, 68.95; H, 10.76; N, 6.28%. Calc. _{for C 13 H 24 NO 2:} C, 68.99; H, 10.69; N, 6.19%.

2,2-dibutyl-6,6-dimethylpiperidin-4-one (5)
According to Method B, 4-heptanone was changed to 4.27 g (30 mmol) of 5-decanone. The reaction solution was stirred in a 60 ° C. water bath for 4 hours. Purification by silica gel column chromatography (hexane / ethyl acetate, 85:15) gave 106 mg (yield: 4%) of a pale yellow oily substance. MS (FAB ⁺ ): 240.3 (M ⁺ +1); v _max / cm ⁻¹ : 1705 (C═O); ¹ H-NMR (300 MHz; CDCl ₃ ) δ (ppm): 0.89 (6H, t, J = 6.7 Hz), 1.25 (6H, s), 1.24-1.56 (12H, m), 2.23 (2H, s), 2.26 (2H, s); : C, 74.83; H, 11.96; N, 5.97%. Calc. _{for C 15 H 29 NO: C} , 75.26; H, 12.21; N, 5.85%.

2,2-dibutyl-6,6-dimethyl-4-oxo-piperidin-1-yl 1-oxyl (6)
According to Method A, 2,2-dibutyl-6,6-dimethylpiperidin-4-one (5) 100 mg (0.42 mmol) was used as a starting material. The reaction solution was stirred overnight at room temperature. Moreover, the progress of the reaction was confirmed by TLC, and H ₂ O ₂ and Na ₂ WO ₄ · 2H ₂ O were appropriately added. After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate) to obtain 53 mg (yield: 50%) of an orange oily substance. ^{_{MS (FAB +): 254.3 (}} M +); v max / cm -1: 1720 (C = O); Found: C, 70.55; H, 11.05; N, 5.24%. Calc. _{for C 15 H 28 NO 2:} C, 70.82; H, 11.09; N, 5.51%.

2-butyl-2,6,6-trimethylpiperidin-4-one (7)
2,6-dibutyl-2,6-dimethylpiperidin-4-one (9)
According to Method B, 4-heptanone was changed to 3.00 g (30 mmol) of 2-hexanone. The reaction solution was stirred in a 50 ° C. water bath for 2.5 hours. After extraction, the product was purified by silica gel column chromatography (hexane / ethyl acetate, 9: 1) to obtain 239 mg (yield: 12%) of a pale yellow oily substance (7) and 98 mg (yield: 4) of a pale yellow oily substance (9). %). (7): MS (FAB ⁺ ): 198.3 (M ⁺ +1); v _max / cm ⁻¹ : 1703 (C═O); ¹ H-NMR (300 MHz; CDCl ₃ ) δ (ppm): 0. 90 (3H, t, J = 7.1 Hz), 1.16 (3H, s), 1.23 (6H, s), 1.29-1.33 (6H, m), 2.22-2. 26 (4H, m); Found: C, 72.83; H, 11.63; N, 7.02%. Calc. _{for C 12 H 23 NO: C} , 73.04; H, 11.75; N, 7.10%. (9): MS (FAB ⁺ ): 240.4 (M ⁺ +1); v _max / cm ⁻¹ : 1706 (C═O); ¹ H-NMR (300 MHz; CDCl ₃ ) δ (ppm): 0. 91 (6H, t, J = 3.3 Hz), 1.14 (6H, s), 1.28-1.46 (12H, m), 2.18-2.31 (4H, m); C, 75.15; H, 12.11; N, 5.75%. Calc. _{for C 15 H 29 NO: C} , 75.26; H, 12.21; N, 5.85%.

2-butyl-2,6,6-trimethyl-4-oxo-piperidin-1-yl 1-oxyl (8)
According to Method A, 2-butyl-2,6,6-trimethylpiperidin-4-one (7) (199 mg, 1.01 mmol) was used as a starting material. The reaction solution was stirred overnight at room temperature. Moreover, the progress of the reaction was confirmed by TLC, and H ₂ O ₂ and Na ₂ WO ₄ · 2H ₂ O were appropriately added. After extraction, the residue was purified by silica gel column chromatography (hexane / ethyl acetate, 95: 5) to obtain 68 mg (yield: 32%) of an orange oily substance. ^{_{MS (FAB +): 212.3 (}} M +); v max / cm -1: 1720 (C = O); Found: C, 68.01; H, 10.44; N, 6.53%. Calc. _{for C 12 H 22 NO 2:} C, 67.89; H, 10.44; N, 6.60%.

2,6-dibutyl-2,6-dimethyl-4-oxo-piperidin-1-yl-oxyl (10)
According to Method A, 77 mg (0.32 mmol) of 2,6-dibutyl-2,6-dimethylpiperidin-4-one (9) was used as a starting material. The reaction solution was stirred overnight at room temperature. Moreover, the progress of the reaction was confirmed by TLC, and H ₂ O ₂ and Na ₂ WO ₄ · 2H ₂ O were appropriately added. After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate) to obtain 25 mg (yield: 31%) of an orange oily substance. ^{_{MS (FAB +): 254.3 (}} M +); v max / cm -1: 1720 (C = O); Found: C, 70.94; H, 10.98; N, 5.18%. Calc. _{for C 15 H 28 NO 2:} C, 70.82; H, 11.09; N, 5.51%.

2,2,6-trimethyl-6-octipeliperidin-4-one (11)
According to Method B, 4-heptanone was changed to 4.69 g (30 mmol) of 2-decanone. The reaction solution was stirred in a 50 ° C. water bath for 4 hours. After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate) to obtain 137 mg (yield: 5%) of a pale yellow oily substance. MS (FAB ⁺ ): 254.3 (M ⁺ +1); v _max / cm ⁻¹ : 1708 (C═O); ¹ H-NMR (300 MHz; CDCl ₃ ) δ (ppm): 0.87 (3H, t, J = 5.7 Hz), 1.16 (3H, s), 1.22 (6H, s), 2.16-2.29 (4H, m); Found: C, 75.57; H, 12.15; N, 5.09%. Calc. for C ₁₆ H ₃₁ NO: C, 75.83; H, 12.33; N, 5.53%.

2,2,6-trimethyl-6-octyl-4-oxo-piperidin-1-yl 1-oxyl (12)
According to Method A, 2,2,6-trimethyl-6-octylpiperidin-4-one (11) 100 mg (0.39 mmol) was used as a starting material. The reaction solution was stirred overnight at room temperature. Moreover, the progress of the reaction was confirmed by TLC, and H ₂ O ₂ and Na ₂ WO ₄ · 2H ₂ O were appropriately added. After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate) to obtain 78 mg (yield: 74%) of an orange oily substance. ^{_{MS (FAB +): 268.3 (}} M +); v max / cm -1: 1721 (C = O); Found: C, 71.45; H, 11.13; N, 5.10%. Calc. _{for C 16 H 30 NO 2:} C, 71.59; H, 11.27; N, 5.22%.

2,2-dimethyl-6-spiro-2'-tricyclo [3.3.1.1 ^{3 ', 7'} ] decane piperidine-4-one (13)
2,6-dispiro-2 ′, 2′-diisticcyclo [3.3.1.1 ^{3 ′, 7 ′} ] decane piperidine-4-one (16)
According to Method B, 4-heptanone was changed to 4.51 g (30 mmol) of 2-adamantanone. The reaction conditions are shown in Table. After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate), and the resulting crystals were recrystallized from hexane to obtain white needle crystals (13) and pale yellow scale crystals (16). (13): Yield: (crude crystals); mp: 92.5-92.6 ° C .; MS (FAB ⁺ ): 248.3 (M ⁺ +1); v _max / cm ⁻¹ : 1699 (C═O ¹ H-NMR (300 MHz; CDCl ₃ ) δ (ppm): 1.23 (6H, s), 1.50-1.95 (14 H, m), 2.27 (2H, s), 2. 53 (2H, s); Found: C, 77.65; H, 10.12; N, 5.60%. Calc. for C ₁₆ H ₂₅ NO: C, 77.68; H, 10.19; N, 5.66%. (16): Yield: 0.62% (2.5 hours at 50 ° C.) (crude crystals); mp: 131.2-131.5 ° C .; MS (FAB ⁺ ): 340.4 (M ⁺ +1); v _max / cm ⁻¹ : 1709 (C═O); ¹ H-NMR (400 MHz; CDCl ₃ ) δ (ppm): 1.25 (brs), 1.56-1.69 (m), 1.79 -1.92 (m), 2.33 (brs), 2.36 (brs), 2.61 (4H, s); Found: C, 81.30; H, 9.77; N, 4.07 %. Calc. _{for C 23 H 33 NO: C} , 81.37; H, 9.80; N, 4.13%.

Method C
2,2-dimethyl-6-spiro-2′-tricyclo [3.3.1.1 ^{3 ′, 7 ′} ] decane-4-oxo-piperidin-1-yl 1oxyl (14)
2,2-dimethyl-6-spiro-2′-tricyclo [3.3.1.1 ^{3 ′, 7 ′} ] decane piperidine-4-one (13) 410 mg (1.66 mmol) was dissolved in chloroform 30 ml, ice Stir in the bath. 371 mg (1.66 mmol) of MCPBA was added, and after 15 minutes, the temperature was returned to room temperature, and stirring was further continued for 2 hours. Extraction was performed with chloroform, the organic layer was dried over sodium sulfate, and the solvent was completely distilled off. Then, it refine | purified by silica gel column chromatography (hexane, ethyl acetate), and the obtained crystal | crystallization was recrystallized from diisopropyl ether, and 172 mg (yield: 40%) of the orange scale-like crystal | crystallization was obtained. ^{mp: 134.7-135.3 ℃; MS (FAB} +): 262.3 (M +); v max / cm -1: 1709 (C = O); Found: C, 73.13; H, 9 .14; N, 5.29%. Calc. _{for C 16 H 24 NO 2:} C, 73.25; H, 9.22; N, 5.34%.

2,2-dimethyl-6-spiro-2'-tricyclo [3.3.1.1 ^{3 ', 7'} ] decane-4-hydroxy-piperidin-1-yl 1oxyl (15)
2,2-dimethyl-6-spiro-2′-tricyclo [3.3.1.1 ^{3 ′, 7 ′} ] decane-4-oxo-piperidin-1-yl 1oxyl (14) 180 mg (0.73 mmol) Dissolved in 5 ml of ethanol and stirred in a water bath. NaBH ₄ 34 mg (0.91 mmol) was added, the temperature was returned to room temperature, and stirring was continued overnight. Further, the progress of the reaction was confirmed by TLC, and NaBH ₄ was appropriately added. This was extracted with chloroform, the organic layer was dried over sodium sulfate, and the solvent was completely distilled off. Thereafter, the product was purified by silica gel column chromatography (chloroform / methanol, 99: 1), and the obtained crystal was recrystallized from ethyl acetate to obtain 89 mg (yield: 46%) of reddish purple scaly crystals. mp: 132.0-133.6 ° C .; MS (FAB ⁺ ): 264.3 (M ⁺ ); v _max / cm ⁻¹ : 3407 (br, OH); Found: C, 72.55; H, 9 .89; N, 5.36%. Calc. _{for C 16 H 24 NO 2:} C, 72.69; H, 9.91; N, 5.30%.

2,6-dispiro-2 ', 2'-dicycliccyclo [3.3.1.1 ^{3', 7 '} ] decane piperidine-4-oxo-piperidin-1-yl 1-oxyl (17)
According to Method C, 2,6-dispiro-2 ′, 2′-diisticcyclo [3.3.1.1 ^{3 ′, 7 ′} ] decane piperidine-4-one (16) 149 mg (0.439 mmol) was used as a starting material. . After extraction, the product was purified by silica gel column chromatography (hexane, ethyl acetate), and the obtained crystals were recrystallized from hexane to obtain 18 mg of purple spherical crystals (yield: 12%). mp: 154.6-160.9 ° C .; MS (FAB ⁺ ): 354.3 (M ⁺ ); v _max / cm ⁻¹ : 1705 (C═O); Found: C, 77.67; H, 9 .03; N, 3.76%. Calc. _{for C 23 H 32 NO 2:} C, 77.92; H, 9.10; N, 3.95%.

Cytotoxic nitroxyl radicals have been reported to react with ascorbic acid, thiols (metal-catalyzed reactions), mitochondrial respiratory chain, NADPH, cytochrome P450, etc. in cells, and induce apoptosis and mediated through cytochrome P450 It has been reported that production of H ₂ O ₂ and the like cause cytotoxicity. On the other hand, the present inventors have reported that the reactivity with the ascorbic acid of the nitroxyl radical having a tetraethyl group is significantly reduced. Therefore, the cytotoxicity of the synthesized nitroxyl radical compound was evaluated.

Method HUVEC (human umbilical vein endothelial cells) were cultured in a CO ₂ incubator at 37 ° C. under 5% CO ₂ using Brett Kit EGM as a medium. Moreover, in order to maintain a logarithmic growth phase, HUVEC was subcultured at regular intervals, and used between 2 and 5 subcultures for experiments. HUVEC (5000 cells / well) was incubated for 24 hours after adding nitroxyl radicals at various concentrations using a collagen I-coated transparent flat bottom plate, and the cell viability was analyzed by the WST-1 method. After 24 hours of incubation, 10 μl of WST-1 reagent was added to each well, and after 3 hours of incubation, the absorbance at 450 nm was measured with a plate reader. 2% Tween 20 was used as a positive control, PBS was used as a negative control, and data was expressed as a relative value (%) to the control.

Results A nitroxyl radical compound was added to HUVEC cells, and the cell growth ability after 24 hours was measured. As a result, Oxo-TEMPO and Tempol showed a concentration-dependent cell growth ability inhibitory effect. On the other hand, a compound having a tetraethyl group introduced showed no inhibitory effect on cell growth ability up to a concentration of 1 mM, indicating that it was low in toxicity despite its high fat solubility (see FIG. 1).

Evaluation of drug efficacy using gastric ulcer model animals Creation of adjuvant arthritic rats It was investigated whether nitroxyl radical suppresses indomethacin-induced gastric ulcers in adjuvant arthritic rats (AA rats).

Method Mycobacterium tuberculosis H37 RA was ground in an agate mortar, and heavy minor oil was added dropwise thereto, and further ground to 10 mg / ml. This tuberculosis killed oil-oil suspension was sonicated for 3 minutes and then taken into a 1 ml syringe, and 100 μL was administered into the left footpad of a 5-week-old DA rat under isoflurane anesthesia to induce adjuvant arthritis. In the negative control group, only heavy mineral oil was administered into the left footpad.

DA rats were fasted in the breeding cage on day 13 after administration of the adjuvant. However, drinking water was free drinking. After 24 hours from the start of fasting, indomethacin (20 mg / kg) was orally administered to rats to create gastric ulcers. As a control, the same amount of 5% NaHCO ₃ was orally administered as a vehicle. The nitroxyl radical compound was orally administered 5 minutes before administration of indomethacin.

Measurement of gastric mucosal damage area 4 hours after induction of gastric ulcer by indomethacin, the rat was treated by cervical dislocation under overanaesthesia, and the stomach was immediately cut and excised leaving about 1 cm each of the esophagus and the small intestine. The pylorus part side was fixed with forceps, 10 ml of 1% formalin solution was injected, and the cardia part side was fixed with forceps. The stomach wall was fixed from both sides by continuing soaking for 10-20 minutes. After fixation, the stomach was incised along the Great Bay, and the gastric lumen was photographed with a digital camera. Using the image processing software Image J, the damaged area generated in the gastric floor mucosa was measured.

Results The degree of gastric ulcer induced by indomethacin was evaluated by the damage area of the gastric fundus mucosa of AA rats to which indomethacin was orally administered. That is, i.e., inflammation of the gastric floor mucosa was confirmed. On the other hand, the AA rats administered with the nitroxyl radical compound showed a significantly reduced damage area compared to the AA rats not administered with the nitroxyl radical compound. It showed that mucosal damage was significantly suppressed.

  Tempol (see FIG. 2), a known nitroxyl radical compound, shows an inhibitory action at a concentration of 900 μmol / kg, but the nitroxyl radical compound synthesized according to the present invention (compounds A and B in FIG. 2). ) Was also shown to have significant inhibitory action on gastric mucosal damage at a concentration of up to 1/10 compared to Tempol, and it became clear that it showed an inhibitory effect at a lower concentration than known compounds .

TEMPO-based nitroxyl radical compounds such as Tempol, a blood pressure lowering action of nitroxyl radical, have a blood pressure lowering action and are widely used in hypertension research. Although the cause of the blood pressure lowering action has not yet been clarified, it is considered that it is due to the ability to scavenge active oxygen because it has a correlation with the SOD-like action. Therefore, we clarify the blood pressure lowering action of the newly synthesized nitroxyl radical compound.

Method All experiments were performed between 9:00 and 16:00 in order to avoid effects on the diurnal rhythm. Spontaneously hypertensive model rats (SHR; ♂, 208-330 g) were anesthetized with isoflurane. Mean blood pressure (MAP) cannulated polyethylene tube (PE-50) in femoral artery, then transducer (MLT0670 BP Transducer, ADinstruments, Sydney, Australia) and amplifier (ML117 BP Amp, Australia) Measured at. Nitroxyl radical solution (60 μmol / kg) was administered from the femoral vein at a rate of 1 ml / min, and blood pressure and heart rate were measured for 20 minutes after administration.

Results FIG. 3 shows changes in blood pressure over time after administration of the nitroxyl radical compound. The blood pressure lowering action exhibited by the newly developed nitroxyl radical compound (compound C) was significantly higher than that of Tempol, which is a known nitroxyl radical compound. Moreover, it was also shown that the duration of the blood pressure lowering action lasts for a long time when the novel nitroxyl compound (Compound C) is compared with Tempol.

  The above-described embodiment is an example of a preferred embodiment of the present invention, but the scope thereof is not limited, and various modifications can be made without departing from the scope of the present invention.

References relating to the present invention are set forth below and are fully incorporated herein by this reference.
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[0006]
[Table 2-1]

［００１８］
上記表から明らかなように、環状構造を有するカルボニル化合物と、直鎖状カルボニル化合物では、ピロリドン生成物に対して異なる選択性が見られる。つまり、環構造を有するカルボニル化合物を原料として用いた場合、二[0007]
[0017]
[Table 2-2]

[0018]
As apparent from the above table, the carbonyl compound having a cyclic structure and the linear carbonyl compound have different selectivity for the pyrrolidone product. That is, when a carbonyl compound having a ring structure is used as a raw material,

［００２６］[0010]
[Table 3-1]

[0026]

［００２８］
本発明の一実施形態によると、当該発明は、化学式１で示されるニトロキシラジカル化合物の製造方法であって、
［化３］

式中、
Ｒ^１〜Ｒ^４は、独立して、Ｃ_１〜Ｃ_１０のアルキルであるか、或いは、Ｒ^１及びＲ^２、又はＲ^３及びＲ^４が共に結合して、（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＯ（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＳＯ_２（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＮＸ（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＣＹ（ＣＨ_２）_ｎ、[0012]
[Table 3-3]

[0028]
According to one embodiment of the present invention, the present invention is a method for producing a nitroxy radical compound represented by Formula 1,
[Chemical formula 3]

Where
R ^{1 to} R ⁴ are independently C _{1 to} C ₁₀ alkyl, or R ¹ and R ² , or R ³ and R ⁴ are bonded together to form (CH ₂ ) _n , (CH _{2) n O (CH 2)} n, (CH 2) n SO 2 (CH 2) n, (CH 2) n NX (CH 2) n, (CH 2) n CY (CH 2) n,

［００２９］
式中、
Ｒ^５〜Ｒ^８は、独立して、Ｃ１〜Ｃ２０のアルキルである、化学式２で示される化合物と、
化学式３で示される化合物であって、
［化５］

［００３０］
式中、
Ｒ^１〜Ｒ^４は、独立して、Ｃ_１〜Ｃ_１０のアルキル、Ｒ^１及びＲ^２、又はＲ^３及びＲ^４が共に結合して、（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＯ（ＣＨ_２）_ｎ、（ＣＨ_２）_ｎＳ[0013]
X is CH ₃ CO, CF ₃ CO, or R ¹ CO;
Y is oxygen (= O), a _{(OCH 2) n,}
n is an integer of 2 to 4,
The method is
A compound represented by Formula 2,
[Chemical formula 4]

[0029]
Where
R ^{5 to} R ⁸ are independently a compound represented by Chemical Formula 2, which is C1-C20 alkyl;
A compound represented by Formula 3,
[Chemical formula 5]

[0030]
Where
R ^{1 to} R ⁴ are independently C _{1 to} C ₁₀ alkyl, R ¹ and R ² , or R ³ and R ⁴ are bonded together to form (CH ₂ ) _n , (CH ₂ ) _n O ( _{CH 2) n, (CH 2} ) n S

Claims (13)

A method for producing a nitroxy radical compound represented by Chemical Formula 1,

Where
R ^{1 to} R ⁴ are independently C _{1 to} C ₁₀ alkyl, or R ¹ and R ² , or R ³ and R ⁴ are bonded together to form (CH ₂ ) _n , (CH _{2) n O (CH 2)} n, (CH 2) n SO 2 (CH 2) n, (CH 2) n NX (CH 2) n, (CH 2) n CY (CH 2) n,
X is CH ₃ CO, CF ₃ CO, or R ¹ CO;
Y is oxygen (= O), a _{(OCH 2) n,}
n is an integer of 2 to 4,
The method is
A compound represented by Formula 2,

Where
R ^{5 to} R ⁸ are independently a C _{1 to} C ₂₀ alkyl compound represented by Formula 2,
A compound represented by Formula 3,

Where
R ^{1 to} R ⁴ are independently C _{1 to} C ₁₀ alkyl, R ¹ and R ² , or R ³ and R ⁴ are bonded together to form (CH ₂ ) _n , (CH ₂ ) _n O ( _{CH 2) n, (CH 2} ) n S (CH 2) n, (CH 2) n NX (CH 2) n, (CH 2) n CY (CH 2) n,
X is CH ₃ CO, CF ₃ CO, or R ¹ CO;
Y is oxygen (= O), a _{(OCH 2) n,}
n is an integer of 2 to 4, and a compound represented by Formula 3 is reacted in the presence of an ammonium salt to produce a 2,6-substituted-4-piperidone derivative;
Oxidizing the 2,6-substituted-4-piperidone derivative;
A process for producing the nitroxy radical compound, comprising:   The method of claim 1, wherein the ammonium salt is ammonium chloride.   2. The method of claim 1, wherein the step of producing the 2,6-substituted-4-piperidone derivative comprises cesium hydroxide.   The method according to claim 1, wherein the step of producing the 2,6-substituted-4-piperidone derivative comprises benzyltrimethylammonium hydroxide (Triton B). A compound represented by Formula 2,

Where
R ^{5 to} R ⁸ are C _{1 to} C ₂₀ alkyl, the compound represented by Formula 2,
A compound represented by Formula 3,

Where
R ^{9 to} R ¹⁰ are C _{1 to} C ₁₀ alkyl, R ⁹ and R ¹⁰ are bonded together, and (CH ₂ ) _n , (CH ₂ ) _n O (CH ₂ ) _n , (CH ₂ ) _n S (CH ₂ ) _n , (CH ₂ ) _n NX (CH ₂ ) _n , (CH ₂ ) _n CY (CH ₂ ) _n ,
X is CH ₃ CO, CF ₃ CO, or R ¹ CO;
Y is oxygen (= O), a _{(OCH 2) n,}
n is an integer of 2 to 4, and a compound represented by Formula 3 is reacted in the presence of an ammonium salt to produce a 2,6-substituted-4-piperidone derivative;
Oxidizing the 2,6-substituted-4-piperidone derivative;
Chemical formula 1 produced by a method comprising

A compound represented by 6. The compound of claim 5, wherein the compound has the following chemical formula:

A nitroxy radical compound represented by   6. A compound according to claim 5, wherein the compound is used for the treatment of gastric mucosal damage.   6. A compound according to claim 5, wherein the compound is used for the treatment of hypertension.   A pharmaceutical composition comprising the compound according to claim 5 and a pharmaceutically acceptable carrier.   A method of treating gastric mucosal damage, comprising the step of administering to a subject a compound according to claim 5.   12. The method of claim 10, wherein the compound is to be administered in an amount of about 0.0001-4 g / day.   A method of treating hypertension, the method comprising administering to a subject a compound according to claim 5.   13. The method of claim 12, wherein the compound is to be administered in an amount of about 0.0001-4 g / day.

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