含1,2,4-三唑结构的黄酮衍生物的合成及抗氧化活性

引用本文

DUAN Zhi-fang, SHAO Ling, LI Sai-nan. Synthesis and antioxidative activity of flavone derivatives containing 1,2,4-triazole structure[J]. Acta Pharm Sin, 2015, 50(7): 861-867. 复制到剪切板

段志芳, 邵玲, 李赛男. 含1,2,4-三唑结构的黄酮衍生物的合成及抗氧化活性[J]. 药学学报, 2015, 50(7): 861-867. 复制到剪切板

含1,2,4-三唑结构的黄酮衍生物的合成及抗氧化活性

段志芳^1,2 , 邵玲¹, 李赛男¹

1. 肇庆学院生命科学学院, 广东肇庆 526061;
2. 肇庆学院生物医药工程中心, 广东肇庆 526061

收稿日期: 2015-01-28;修回日期: 2015-03-05.

基金项目：国家自然科学基金资助项目 (31201142); 肇庆市创新科技基金资助项目 (2012G25).

* 通讯作者：Tel/Fax: 86-758-2716359, E-mail: zhaoqing606@126.com

摘要：分别以7-羟基黄酮、水杨醛、肉桂酸和3-取代-4-氨基-1,2,4-三唑-5-硫酮等为起始原料, 根据活性基团拼合原则, 经多步反应得到3种类型共15个含1,2,4-三唑杂环结构的黄酮类衍生物。用红外、核磁共振氢谱、质谱及元素分析对化合物的结构进行了确证。测定了新化合物清除超氧自由基 (O₂^-· )、羟自由基 (·OH) 和2,2-二苯基-1-苦味酰基自由基 (DPPH·) 的活性及总还原能力。结果表明, 在0.5 mg·mL^-1浓度时, 多数化合物具有抗氧化活性, 其中化合物7-(3-间甲苯基-4-水杨醛亚氨基-1,2,4-三唑-5-硫)乙氧基黄酮 (5e) 对O₂^-· 抑制作用与对照维生素C相当, 其余化合物活性均弱于维生素C。

关键词： 1,2,4-三唑黄酮衍生物合成抗氧化

Synthesis and antioxidative activity of flavone derivatives containing 1,2,4-triazole structure

DUAN Zhi-fang^1,2 , SHAO Ling¹, LI Sai-nan¹

1. Zhaoqing University, College of Life Science, Zhaoqing 526061, China;
2. Zhaoqing University, Biopharmaceutical Research and Development Center, Zhaoqing 526061, China

Abstract: Using bioactive compounds 7-hydroxy flavone, salicylaldehyde, cinnamic acid and 4-amino-5-mercapto-1,2,4-triazoles as starting materials, three new types of flavone derivatives containing 1,2,4-triazole structure were synthesized via different step reactions. These new compounds were characterized by ¹H NMR, ESI-MS, IR and elemental analysis. Their scavenging effects on the superoxide radical (O₂^-·), hydroxyl radical (·OH), DPPH· radical and their total reduction activities were tested. The results showed that all of the compounds possessed some antioxidative activity at the concentration of 0.5 mg·mL^-1, but the scavenging ability of the target compounds was lower than that of the standard compound Vc.

Key words: 1,2,4-triazole flavone derivative synthesis antioxidation

抗氧化自由基 (anti-oxidant) 简称抗氧化, 人体内主要自由基包括超氧阴离子自由基 (O₂^－· )、羟自由基 (·OH) 和硝基自由基 (·ONOO) 等, 过量自由基对生物体具有潜在的氧化破坏作用, 可引起多种疾病, 如高血压、动脉硬化、心肌梗死、癌症、艾滋病等疾病的发生^{[1, 2]}。因此对自由基清除剂—抗氧化剂的研究便显得尤为重要。目前国内外测定抗氧化能力的方法主要包括体内实验和体外实验两大类, 而体外抗氧化实验被广泛的应用于抗氧化剂的筛选和研究上。

黄酮类化合物是一类广泛存在于植物中的常见天然化合物, 具有多种药用保健功能, 如抗氧化、抗衰老、抗心血管疾病、降血脂、降血压、降血糖以及抗癌等作用^[3], 而以黄酮类化合物为先导化合物, 对其进行结构改造和结构优化, 以期研发具有更强药理作用的黄酮类新药, 是该研究领域的一个重要课题^[4]。1,2,4-三唑类化合物具有广泛的药理活性, 如抗菌、抗肿瘤、抗过敏、抗抑郁、抗惊厥和抗炎等^{[5, 6]}, 作为药物分子设计与合成的基本结构一直备受关注。

本文以自制7-羟基黄酮为先导化合物, 与过量1,2-二溴乙烷反应得到7-溴乙氧基黄酮, 然后通过缩合反应将3种不同类型的1,2,4-三唑片段引入到黄酮结构中, 设计合成了15个新型的含1,2,4-三唑杂环的黄酮类化合物, 用红外、核磁共振氢谱、质谱及元素分析对这些化合物的结构进行了确证。初步测定了这些新化合物清除O₂^－· 、·OH和2,2-二苯基-1-苦味酰基自由基(DPPH·) 的活性及总还原能力, 以期研发具有更强药理作用的黄酮类新药, 为黄酮的广泛开发和应用提供实验数据。

中间体3-取代苯基-4-水杨醛亚氨基-1,2,4-三唑- 5-硫酮 (化合物1) 及3-取代苯基-5-巯基-1,2,4-三唑(化合物3) 的合成如合成路线1所示。目标化合物7-[2-(3-硫酮-5-取代-1,2,4-三唑-4-N-亚胺次甲基)]-苯氧乙氧基黄酮 (化合物4)、7-(3-取代苯基-4-水杨醛亚氨基-1,2,4-三唑-5-硫)乙氧基黄酮 (化合物5) 和7- (3-取代苯基-1,2,4-三唑-5-硫) 乙氧基黄酮 (化合物6) 的合成如合成路线2所示。

结果与讨论 1 目标化合物的合成

对黄酮天然产物进行结构修饰与改造是发现黄酮类新药的重要手段和途径。肉桂酸和水杨醛是两类非常重要的天然活性化合物, 其衍生物抗氧化性均较强^{[7, 8]}。本文原计划设计将肉桂酸和水杨醛结构通过1,2,4-三唑杂环引入黄酮中, 以期得到抗氧化活性强的黄酮衍生物。结果原料制备中没有得到预期的3-苯基-4-肉桂酰基-5-巯基-1,2,4-三唑, 而是得到其水解产物3-苯基-4-氢-5-巯基-1,2,4-三唑(化合物3)。7-溴乙氧基黄酮不是与三唑环中的NH缩合, 而是与SH发生缩合, 在制备目标化合物5和6时, 用NaOH/CH₃CH₂OH代替常用的K₂CO₃/CH₃COCH₃, 化合物5直接从反应体系中析出, 化合物6减压蒸除部分乙醇后析出, 后处理简化。目标化合物的结构经¹H NMR、IR、ESI-MS及元素分析确认, 理化和光谱数据见表 1～3。

2 抗氧化活性

抗氧化与许多疾病如肿瘤、炎症、衰老等密切相关, 而黄酮类抗氧化成分是人们研究的热点课题, 因此本文测定了目标新化合物的体外抗氧化能力, 包括O₂^－·、羟自由基 ·OH和2,2-二苯DPPH· 的活性及总还原能力。实验过程中为了便于比较, 选用了浓度为0.5 mg·mL⁻¹时的实验结果, 实验进行3次, 结果取平均值, 见表 4。从表中可以看出多数化合物具有抗氧化活性, 其中化合物7-(3-间甲苯基-4-水杨醛亚氨基- 1,2,4-三唑-5-硫) 乙氧基黄酮 (5e) 对O₂^－· 抑制作用与对照维生素C相当, 其余化合物均弱于维生素C。

Scheme 1 Synthetic routes of the intermediate compounds 1 and 3

Scheme 2 Synthetic routes of the target compounds 4, 5 and 6

3 小结

本论文合成了3种类型的1,2,4-三唑化合物: 3-取代-4-氨基-1,2,4-三唑-5-硫酮、3-取代苯基-4-水杨醛亚氨基-1,2,4-三唑-5-硫酮(化合物1) 及3-取代苯基- 4-氢-5-巯基-1,2,4-三唑(化合物3), 化合物1和3分别与7-溴乙氧基黄酮通过硫醚化反应合成了9个含1,2,4-三唑结构的硫乙氧基黄酮类衍生物, 化合物3-取代-4-氨基-1,2,4-三唑-5-硫酮与7-水杨醛乙氧基黄酮反应合成了6个含1,2,4-三唑结构的苯氧乙氧基黄酮类衍生物。测定了新化合物的清除O₂^－·、·OH和DPPH·的活性及总还原能力。抗氧化测试结果表明, 在浓度为0.5 mg·mL⁻¹时, 多数化合物具有抗氧化活性, 其中化合物7-(3-间甲苯基-4-水杨醛亚氨基-1,2,4-三唑-5-硫) 乙氧基黄酮 (5e) 对O₂^－· 抑制作用与对照维生素C相当, 值得进一步研究。

实验部分

X-4型显微熔点测定仪 (北京泰克公司, 温度计未校正); Vario EL cube型元素分析仪 (德国Elementar公司); FT-IR 6700型红外光谱仪 (美国Nicolet公司, KBr压片); Avance AV 500 MHz超导核磁共振仪 (德国Bruker公司, 溶剂为DMSO-d₆, 内标TMS); DSQ质谱仪 (美国)、UPLC SQD质谱仪 (Waters)。7-羟基黄酮参考文献^[9]制备; 3-取代-4-氨基-1,2,4-三唑-5-硫酮参考文献^{[10, 11]}制备; DPPH· 为Sigma产品; 其余试剂为国产化学纯或分析纯。

1 化合物的合成 1.1 3-苯基-4-水杨醛亚氨基-1,2,4-三唑-5-硫酮 (1a) 的合成

在100 mL圆底烧瓶中加入3 mmol水杨醛、3 mmol 3-取代苯基-4-氨基-1,2,4-三唑-5-硫酮和30 mL无水乙醇, 搅拌微热溶解后滴加2滴浓盐酸, 回流反应3 h。静置, 冷却, 抽滤, 蒸馏水洗涤得到白色固体, 用乙醇重结晶, 得3-苯基-4-水杨醛亚氨基- 1,2,4-三唑-5-硫酮(1a)。类似方法合成化合物1b～ 1e, 化合物理化与波谱数据见表 1。

Table 1 Physical property and spectra data of compounds 1, 2 and 3

Compd.	Yield/%	mp/℃	Physical property	¹H NMR (DMSO-d₆, 500 MHz) δ	IR (KBr) /cm^-1	ESI-MS m/z [M+H]⁺
1a	65.0	247−249	White solid	14.14 (s, 1H, NH), 10.41 (s, 1H, N=CH), 9.92 (s, 1H, OH), 7.83−7.88 (m, 2H, ArH), 7.82 (d, J = 1.5 Hz, 1H, ArH), 7.52−7.54 (m, 3H, ArH), 7.44 (t, J = 8.5 Hz, 1H, ArH), 6.99 (d, J = 8.0 Hz, 1H, ArH), 6.95 (t, J = 7.5 Hz, 1H, ArH)	3 108 (NH), 1 599 (N=C), 1 269 (C=S)	297.44
1b	67.2	224−226	White solid	14.08 (s, 1H, NH), 10.47 (s, 1H, N=CH), 9.88 (s, 1H, OH), 7.80−7.85 (m, 3H, ArH), 7.45 (t, J = 7.75 Hz, 1H, ArH), 7.08 (d, J = 8.5 Hz, 2H, ArH), 6.99 (d, J = 8.0 Hz, 1H, ArH), 6.95 (t, J = 7.5 Hz, 1H, ArH), 3.81 (s, 3H, CH₃O)	3 108 (NH), 1 610 (N=C), 1 297 (C=S)	327.23
1c	78.9	249−251	White solid	14.19 (s, 1H, NH), 10.41 (s, 1H, N=CH), 9.96 (s, 1H, OH), 7.88−7.90 (m, 2H, ArH), 7.82−7.84 (m, 1H, ArH), 7.60−7.63 (m, 2H, ArH), 7.43− 7.46 (m, 1H, ArH), 6.99 (d, J = 8.0 Hz, 1H, ArH), 6.95 (t, J = 7.0 Hz, 1H, ArH)	3 090 (NH), 1 601 (N=C), 1 275 (C=S)	331.27
1d	57.8	280−282	Yellow solid	14.17 (s, 1H, NH), 10.43 (s, 1H, N=CH), 9.96 (s, 1H, OH), 7.96 (d, J = 1.5 Hz, 2H, ArH), 7.87 (d, J = 1.5 Hz, 1H, ArH), 7.43−7.47 (m, 1H, ArH), 6.94−6.99 (m, 4H, ArH)	3 101 (NH), 1 606 (N=C), 1 274 (C=S)	341.41
1e	51.6	217−219	White solid	14.12 (s, 1H, NH), 10.40 (s, 1H, N=CH) , 9.91 (s, 1H, OH), 7.81−7.83 (dd, J = 1.5, 6.5 Hz, 1H, ArH), 7.70 (s, 1H, ArH), 7.65 (d, J = 7.5 Hz, 1H, ArH), 7.35−7.46 (m, 3H, ArH), 7.00 (d, J = 8.5 Hz, 1H, ArH), 6.95 (t, J = 7.5Hz, 1H, ArH), 2.35 (s, 3H, CH₃)	3 107 (NH), 1 604 (N=C), 1 270 (C=S)	311.17
2a	81.3	228−229	White solid	12.30 (s, 1H, NH), 11.69 (s, 1H, NH), 11.07 (s, 1H, NH) , 7.91−7.93 (m, 2H, ArH), 7.79 (d, J = 16.0 Hz, 1H, CH), 7.65−7.66 (m, 2H, ArH), 7.62−7.64 (m, 1H, ArH), 7.61 (d, J = 7.0 Hz, 2H, ArH), 7.48−7.54 (m, 3H, ArH), 7.07 (d, J = 16.0 Hz, 1H, CH)	3 157 (NH), 1 691, 1 652 (C=O), 1 634 (C=C), 1 145 (C=S)	326.13
2b	85.6	226−227	White solid	12.23 (s, 1H, NH), 11.68 (s, 1H, NH), 10.84 (s, 1H, NH), 7.78 (d, J = 16.0 Hz, 1H, CH), 7.64−7.66 (m, 2H, ArH), 7.54−7.56 (m, 1H, ArH), 7.48−7.50 (m, 3H, ArH), 7.38−7.42 (m, 1H, ArH), 7.27−7.30 (m, 2H, ArH), 7.05−7.08 (d, J = 15.5 Hz, 1H, CH), 2.43 (s, 3H, CH₃)	3 280 (NH), 1 696, 1 668 (C=O), 1 627 (C=C), 1 139 (C=S)	340.25
2c	61.1	208−209	Light yellow solid	12.31 (s, 1H, NH), 11.68 (s, 1H, NH), 11.01 (s, 1H, NH), 7.78 (d, J = 16.0 Hz, 1H, CH), 7.74 (s, 1H, ArH), 7.71 (d, J = 6.0 Hz, 1H, ArH), 7.64−7.66 (m, 2H, ArH), 7.47−7.49 (m, 3H, ArH), 7.41−7.42 (m, 2H, ArH), 7.06 (d, J = 16.0 Hz, 1H, CH), 2.39 (s, 3H, CH₃)	3 229 (NH), 1 687, 1 667 (C=O), 1 627 (C=C), 1 151 (C=S)	340.25
2d	89.5	230−231	White solid	12.31 (s, 1H, NH), 11.67 (s, 1H, NH), 10.99 (s, 1H, NH), 7.82 (d, J = 8.0 Hz, 2H, ArH), 7.78 (d, J = 16.0 Hz, 1H, CH), 7.64−7.66 (m, 2H, ArH), 7.47−7.49 (m, 3H, ArH), 7.33 (d, J = 8.0 Hz, 2H, ArH), 7.06 (d, J = 16.0 Hz, 1H, CH), 2.38 (s, 3H, CH₃)	3 178 (NH), 1 678, 1 645 (C=O), 1 628 (C=C), 1 156 (C=S)	340.25
3a	52.2	232−233	White solid	13.80 (s, 1H, NH), 13.64 (s, 1H, NH), 7.90−7.92 (m, 2H, ArH), 7.41− 7.52 (m, 3H, ArH)	3 233 (NH), 1 630 (C=N), 1 289 (C=S)	178.12
3b	62.4	220−221	White solid	13.64 (s, 1H, NH), 13.54 (s, 1H, NH), 7.60 (d, J = 7.5 Hz, 1H, ArH), 7.37−7.42 (m, 1H, ArH), 7.35 (d, J = 8.0 Hz, 1H, ArH), 7.31 (d, J = 7.5 Hz, 1H, ArH), 2.45 (s, 3H, CH₃)	3 232 (NH), 1 630 (C=N), 1 285 (C=S)	192.21
3c	63.7	238−240	White solid	13.74 (s, 1H, NH), 13.61 (s, 1H, NH), 7.75 (s, 1H, ArH), 7.70 (d, J = 8.0 Hz, 1H, ArH), 7.38−7.41 (m, 1H, ArH), 7.32 (d, J = 8.0 Hz, 1H, ArH), 2.36 (s, 3H, CH₃)	3 235 (NH), 1 629 (C=N), 1 287 (C=S)	192.21
3d	66.9	241−243	White solid	13.71 (s, 1H, NH), 13.57 (s, 1H, NH), 7.79 (d, J = 8.0 Hz, 2H, ArH), 7.32 (d, J = 8.0 Hz, 2H, ArH), 2.35 (s, 3H, CH₃)	3 233 (NH), 1 635 (C=N), 1 299 (C=S)	192.21

Table 1 Physical property and spectra data of compounds 1, 2 and 3

1.2 3-苯基-4-氢-5-巯基-1,2,4-三唑 (3a) 的合成

苯甲酰胺基肉桂酰基硫脲(2a) 参考文献^[12]制备。向15 mL的2 mmol该化合物的乙醇悬浮液中加入2 mL 5 mol·L⁻¹ NaOH水溶液, 加热回流2 h, 冷却, 过滤, 滤液用稀盐酸调节pH=5～6, 静置, 过滤, 蒸馏水洗涤, 干燥后乙醇重结晶, 得3-苯基-4-氢-5-巯基-1,2,4-三唑(3a)。类似方法合成化合物3b～3d, 化合物理化与波谱数据见表 1。

1.3 7-溴乙氧基黄酮的合成

参考文献^[13]方法合成。7-溴乙氧基黄酮为淡黄色颗粒状固体, 产率73.4%, mp 142～144 ℃; ESI-MS m/z: 345.4 ([M]⁺, 100), 347.3 ([M]⁺, 98, 同位素峰)。Elemental analysis (%) Calcd. for C₁₇H₁₃BrO₃: C 59.13, H 3.77; Found: C 59.46, H 3.64。

1.4 7-水杨醛乙氧基黄酮的合成

在250 mL圆底烧瓶中加入10 mmol水杨醛、100 mL无水乙醇和10 mmol氢氧化钠, 搅拌溶解后加入10 mmol的7-溴乙氧基黄酮, 加热回流反应16 h (TLC跟踪), 反应完毕后冷却至室温, 抽滤, 用蒸馏水洗涤固体, 无水乙醇重结晶, 得淡黄色固体, 产率79.2%, mp 193～19 ℃。¹H NMR (DMSO-d₆, 400 MHz) δ: 10.37 (s, 1H, CHO), 8.10 (dd, J = 2.2, 5.8 Hz, 2H, ArH), 7.97 (d, J = 8.8 Hz, 1H, 5-H), 7.67～7.71 (m, 2H, ArH), 7.57～7.62 (m, 3H, ArH), 7.44 (d, J = 2.3 Hz, 1H, 8-H), 7.34 (d, J = 8.2 Hz, 1H, ArH), 7.10～7.14 (m, 2H, ArH, 6-H), 6.97 (s, 1H, 3-H), 4.58～4.62 (m, 4H, CH₂CH₂); IR (KBr) cm^-1: 3 068, 3 036 (Ar-H), 2 948, 2 932 (-CH₂), 2 867, 2 767 (CH=O), 1 686 (CH=O), 1 639 (C=O), 1 598, 1 483, 1 453 (Ar), 1 373, 1 292, 1 245, 1 178 (C-O-C), 1 089, 1 058, 844, 775, 685, 658。ESI-MS m/z: 387.4 ([M+H] ⁺, 100)。Elemental analysis (%) Calcd. for C₂₄H₁₈O₅: C 74.61, H 4.66; Found: C 74.78, H 4.52。

1.5 7-[2-(3-硫酮-5-苯氧甲基-1,2,4-三唑-4-N-亚胺次甲基)]-苯氧乙氧基黄酮 (4a) 的合成

在50 mL圆底烧瓶中加入20 mL无水乙醇和1 mmol 7-水杨醛乙氧基黄酮, 搅拌微热溶解后, 加入1 mmol 3-硫酮-4-氨基-5-苯氧甲基-1,2,4-三唑, 滴加催化量冰醋酸, 缓慢升温至回流, TLC跟踪 (展开剂为V_乙酸乙酯∶V_石油醚= 1∶1), 待原料点7-水杨醛乙氧基黄酮消失后停止反应。静置, 冷却, 抽滤得到固体粗品, 用乙醇/N,N-二甲基甲酰胺重结晶, 得7-[2-(3-硫酮-5-苯氧甲基-1,2,4-三唑-4-N-亚胺次甲基)]-苯氧乙氧基黄酮 (4a)。类似方法合成化合物4b～4f, 化合物理化和波谱数据见表 2和表 3。

Table 2 Physical property of target compounds 4, 5 and 6

Compd.	Yield/%	mp/℃	Physical property	Compd.	Yield/%	mp/℃	Physical property
4a	62.7	223−225	Yellow solid	5c	49.1	184−186	White solid
4b	71.8	173−175	White solid	5d	44.1	173−175	Yellow solid
4c	62.0	210−212	White solid	5e	53.9	120−121	White solid
4d	81.1	245−247	Light yellow solid	6a	57.3	207−208	White solid
4e	67.1	225−227	Light yellow solid	6b	52.1	175−176	White solid
4f	78.5	256−258	White solid	6c	70.8	202−203	White solid
5a	45.4	114−116	White solid	6d	67.4	203−204	White solid
5b	57.0	168−170	White solid

Table 2 Physical property of target compounds 4, 5 and 6

Table 3 Spectra data of compounds 4, 5 and 6

Compd.	¹H NMR (DMSO-d₆, 500 MHz), IR, MS and elemental analysis
4a	¹H NMR δ: 13.99 (s, 1H, NH), 10.44 (s, 1H, CH=N), 8.07−8.09 (m, 2H, ArH), 7.86−7.94 (m, 2H, ArH, 5-H), 7.56−7.61 (m, 4H, ArH), 7.24− 7.37 (m, 4H, ArH, 8-H), 7.05−7.11 (m, 2H, ArH), 6.99−7.04 (m, 3H, ArH, 6-H), 6.97 (s, 1H, 3-H), 5.21 (s, 2H, CH₂), 4.60 (s, 2H, CH₂), 4.54 (s, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 243 (NH), 3 037 (ArH), 2 931, 2 850 (CH₂), 1 630 (C=O), 1 613 (C=N), 1 599, 1 587, 1 566, 1 492 (Ar), 1 447, 1 382, 1 361, 1 292; ESI-MS m/z: 591.3 ([M]⁺, 100); Elemental anal. (%) Calcd. for C₃₃H₂₆N₄O₅S: C 67.12, H 4.41, N 9.49; Found: C 67.14, H 4.36, N 9.47.
4b	¹H NMR δ: 11.73 (s, 1H, NH), 11.42 (s, 1H, CH=N), 8.07−8.11 (m, 2H, ArH, 5-H), 8.00−8.03 (m, 2H, ArH), 7.77−7.98 (m, 2H, ArH), 7.57− 7.60 (m, 4H, ArH), 7.36−7.50 (m, 4H, ArH, 8-H), 7.12−7.22 (m, 2H, ArH), 6.98−7.11 (m, 3H, ArH, 6-H), 6.96 (s, 1H, 3-H), 4.51−4.57 (m, 2H, CH₂), 4.41 (s, 2H, CH₂), 3.98 (s, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 242, 3 208 (NH), 3 062 (ArH), 2 948, 2 884 (CH₂), 1 672 (C=O), 1 655, 1 626 (C=N); ESI-MS m/z: 569.3 ([M−C₄H₇]⁺, 100); Elemental anal. (%) Calcd. for C₃₇H₂₈N₄O₄S: C 71.15, H 4.49, N 8.97; Found: C 71.09, H 4.42, N 8.79.
4c	¹H NMR δ: 14.02 (s, 1H, NH), 10.38 (s, 1H, CH=N), 8.36 (s, 1H, 5-H), 8.08−8.10 (m, 2H, ArH), 7.91−7.99 (m, 1H, ArH), 7.78−7.84 (m, 1H, ArH), 7.55−7.60 (m, 4H, ArH), 7.33−7.51 (m, 3H, ArH, 8-H), 7.20−7.30 (m, 1H, ArH), 6.99−7.15 (m, 2H, ArH, 6-H), 6.97 (s, 1H, 3-H), 5.34 (s, 2H, CH₂), 5.23 (s, 2H, CH₂), 4.53 (s, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 320 (NH), 3 062 (ArH), 2 941, 2 879, 2 850 (CH₂), 1 694 (C=O), 1 630 (C=N); ESI-MS m/z: 603.3 ([M−C₄H₇] ⁺, 100); Elemental anal. (%) Calcd. for C₃₃H₂₄Cl₂N₄O₅S: C 60.09, H 3.64, N 8.50; Found: C 60.12, H 3.49, N 8.57.
4d	¹H NMR δ: 14.14 (s, 1H, NH), 10.12 (s, 1H, CH=N), 8.04−8.08 (m, 2H, ArH, 5-H), 7.89−7.95 (m, 2H, ArH), 7.81−7.83 (m, 2H, ArH), 7.53− 7.66 (m, 6H, ArH, 8-H), 7.32−7.35 (m, 2H, ArH), 7.07−7.16 (m, 2H, ArH, 6-H), 6.91 (s, 1H, 3-H), 4.55 (s, 2H, CH₂), 4.60 (s, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 210 (NH), 3 064, 3 036 (ArH), 2 986, 2 883, 2 730 (CH₂), 1 629 (C=O), 1 613 (C=N); ESI-MS m/z: 595.2 ([M]⁺, 62.5); Elemental anal. (%) Calcd. for C₃₂H₂₃ClN₄O₄S: C 64.59, H 3.87, N 9.42; Found: C 64.63, H 3.91, N 9.53.
4e	¹H NMR δ: 13.97 (s, 1H, NH), 10.05 (s, 1H, CH=N), 8.04−8.06 (dd, J = 1.0, 6.5 Hz, 2H, ArH), 7.94−7.96 (dd, J = 1.5, 6.5 Hz, 1H, ArH), 7.89 (d, J = 9.0 Hz, 1H, ArH), 7.73−7.51 (dd, J = 2.0, 5.0 Hz, 2H, ArH), 7.55−7.63 (m, 4H, ArH), 7.34−7.35 (m, 1H, ArH), 7.32 (s, 1H, ArH), 7.14 (t, J = 7.5 Hz, 1H, ArH), 7.07−7.09 (dd, J = 2.5, 6.5 Hz, 1H, ArH), 7.02 (d, J = 2.0 Hz, 2H, ArH), 6.91 (s, 1H, 3-H), 4.53−4.56 (m, 4H, CH₂CH₂), 3.76 (s, 3H, CH₃O); IR (KBr) ν/cm⁻¹: 3 290 (NH), 3 052 (ArH), 2 919, 2 733 (CH₂), 1 630 (C=O), 1 613 (C=N); ESI-MS m/z: 338.56 (100), 360.51 (92), 591.29 (81), 697.81 (99); Elemental anal. (%) Calcd. for C₃₃H₂₆N₄O₅S: C 67.12, H 4.41, N 9.49; Found: C 67.02, H 4.23, N 9.57.
4f	¹H NMR δ: 13.91 (s, 1H, NH), 10.06 (s, 1H, CH=N), 8.06−8.08 (dd, J = 1.0, 6.5 Hz, 2H, ArH), 7.90−7.92 (m, 2H, ArH), 7.57−7.65 (m, 6H, ArH), 7.33−7.37 (m, 2H, ArH), 7.14 (t, J = 7.8 Hz, 1H, ArH), 7.08−7.10 (dd, J = 2.5, 6.5 Hz, 1H, ArH), 6.93 (s, 1H, 3-H), 6.84 (d, J = 9.0 Hz, 2H, ArH), 4.55−4.56 (m, 4H, CH₂CH₂); IR (KBr) ν/cm⁻¹: 3 298 (NH), 3 136 (ArH), 1 624 (C=O), 1 575, 1 508 (Ar); ESI-MS m/z: 338.50 (92), 360.51 (84), 577.18 ([M+H]⁺, 21); Elemental anal. (%) Calcd. for C₃₂H₂₄N₄O₅S: C 66.67, H 4.17, N 9.72; Found: C 66.93, H 4.19, N 9.47.
5a	¹H NMR δ: 10.49 (s, 1H, N=CH), 8.97 (s, 1H, OH), 8.10−8.11 (m, 2H, ArH), 7.91 (d, J = 8.5 Hz, 1H, ArH), 7.84−7.86 (dd, J = 2.0, 6.0 Hz, 2H, ArH), 7.80−7.82 (dd, J = 1.5, 6.5 Hz, 1H, ArH), 7.56−7.61 (m, 3H, ArH), 7.49−7.51 (m, 3H, ArH), 7.41−7.44 (m, 2H, ArH), 7.02−7.04 (dd, J = 2.0, 6.5 Hz, 1H, ArH), 6.97 (d, J = 8.0 Hz, 1H, ArH), 6.96 (s, 1H, 3-H), 6.93 (t, J = 9.0 Hz, 1H, ArH), 4.50 (t, J = 6.0 Hz, 2H, CH₂), 3.65 (t, J = 6.25 Hz, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 415 (NH), 3 068, 2 940, 2 736, 1 627 (C=O), 1 597 (C=N); ESI-MS m/z: 159.13 (30), 338.50 (81), 360.64 (43), 561.31 ([M+H]⁺, 100); Elemental anal. (%) Calcd. for C₃₂H₂₄N₄O₄S: C 68.57, H 4.29, N 10.00; Found: C 67.01, H 4.26, N 10.13.
5b	¹H NMR δ: 10.50 (s, 1H, N=CH), 8.97 (s, 1H, OH), 8.09−8.11 (m, 2H, ArH), 7.91 (d, J = 9.0 Hz, 1H, ArH), 7.78−7.82 (m, 3H, ArH), 7.56− 7.61 (m, 3H, ArH), 7.42−7.45 (m, 1H, ArH), 7.39 (d, J = 2.5 Hz, 1H, ArH), 7.06 (d, J = 9.0 Hz, 2H, ArH), 7.01−7.03 (dd, J = 2.5, 6.5 Hz, 1H, ArH), 6.98 (s, 1H, 3-H), 6.96 (s, 1H, ArH), 6.92 (t, J = 7.5 Hz, 1H, ArH), 4.49 (t, J = 6.3 Hz, 2H, CH₂), 3.79 (s, 3H, CH₃O), 3.62 (t, J = 6.0 Hz, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 425 (NH), 3 055, 2 840, 1 663 (C=O), 1 618 (C=N), 1 483, 1 443, 1 376; ESI-MS m/z: 159.19 (31), 453.43 (27), 566.95 (24), 591.49 ([M+H]⁺, 100); Elemental anal. (%) Calcd. for C₃₃H₂₆N₄O₅S: C 67.12, H 4.41, N 9.49; Found: C 66.97, H 4.52, N 9.64.
5c	¹H NMR δ: 10.56 (s, 1H, N=CH), 8.99 (s, 1H, OH), 8.09−8.11 (m, 2H, ArH), 7.82−7.89 (m, 3H, ArH), 7.81 (d, J = 1.5 Hz, 1H, ArH), 7.56− 7.61 (m, 5H, ArH), 7.43−7.46 (m, 1H, ArH), 7.39 (d, J = 2.0 Hz, 1H, ArH), 7.01−7.03 (dd, J = 2.5, 6.5 Hz, 1H, ArH), 6.97 (d, J = 8.3 Hz, 1H, ArH), 6.96 (s, 1H, 3-H), 6.92 (t, J = 7.8 Hz, 1H, ArH), 4.50 (t, J = 6.3 Hz, 2H, CH₂), 3.65 (t, J = 6.0 Hz, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 428 (NH), 3 067, 2 947, 2 733, 1 639 (C=O), 1 446, 1 365; ESI-MS m/z: 338.43 (89), 360.51 (82), 595.31 ([M+H]⁺, 76); Elemental anal. (%) Calcd. for C₃₂H₂₃ClN₄O₄S: C 64.59, H 3.87, N 9.42; Found: C 64.91, H 3.69, N 10.02.
5d	¹H NMR δ: 10.53 (s, 1H, N=CH), 8.99 (s, 1H, OH), 8.09−8.11 (m, 2H, ArH), 7.91−7.96 (m, 2H, ArH), 7.84 (d, J = 7.5 Hz, 1H, ArH), 7.66 (d, J = 1.5 Hz, 2H, ArH), 7.53−7.60 (m, 4H, ArH), 7.40−7.45 (m, 2H, ArH), 6.98−7.04 (m, 2H, ArH), 6.97 (s, 1H, 3-H), 6.94 (t, J = 9.0 Hz, 1H, ArH), 4.51 (t, J = 6.3 Hz, 2H, CH₂), 3.66 (t, J = 6.0 Hz, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 431 (NH), 3 069, 2 925, 2 861, 1 631 (C=O), 1 569 (C=N), 1 445, 1 365; ESI-MS m/z: 159.07 (61), 338.56 (100), 397.03 (93), 431.16 (64), 677.03 (64); Elemental anal. (%) Calcd. for C₃₂H₂₃N₅O₆S: C 63.47, H 3.80, N 11.57; Found: C 62.99, H 3.76, N 11.62
5e	¹H NMR δ: 10.49 (s, 1H, N=CH), 8.95 (s, 1H, OH), 8.10−8.11 (m, 2H, ArH), 7.91 (d, J = 9.0 Hz, 1H, ArH), 7.80−7.82 (dd, J = 1.5, 6.5 Hz, 1H, ArH), 7.68 (s, 1H, ArH), 7.56−7.61 (m, 4H, ArH), 7.41−7.45 (m, 2H, ArH), 7.38 (t, J = 7.8 Hz, 1H, ArH), 7.30 (d, J = 8.0 Hz, 1H, ArH), 7.01− 7.04 (dd, J = 2.5, 6.5 Hz, 1H, ArH), 6.97 (d, J = 8.0 Hz, 1H, ArH), 6.96 (s, 1H, 3-H), 6.92 (t, J = 7.5 Hz, 1H, ArH), 4.50 (t, J = 6.3 Hz, 2H, CH₂), 3.65 (t, J = 6.3 Hz, 2H, CH₂), 2.33 (s, 3H, CH₃); IR (KBr) ν/cm⁻¹: 3 416 (NH), 3 066, 2 946, 2 873, 2 738, 1 626 (C=O), 1 597 (C=N), 1 496, 1 450; ESI-MS m/z: 159.26 (37), 338.69 (100), 360.64 (64), 575.49 ([M+H]⁺, 84); Elemental anal. (%) Calcd. for C₃₃H₂₆N₄O₄S: C 68.99, H 4.53, N 9.76; Found: C 69.15, H 4.74, N 9.69.
6a	¹H NMR δ: 14.49 (s), 14.19 (s, 1H, NH), 8.05 (d, J = 7.0 Hz, 2H, ArH), 7.93−7.97 (m, 3H, ArH), 7.56−7.59 (m, 3H, ArH), 7.39−7.55 (m, 3H, ArH), 7.39 (s, 1H, ArH), 7.10 (d, J = 8.5 Hz, 1H, ArH), 6.94 (s, 1H, 3-H), 4.51 (t, J = 6.5 Hz, 2H, CH₂), 3.61 (s, 2H, CH₂); IR (KBr) ν/cm⁻¹: 3 442 (NH), 3 095 (Ar), 2 989, 2 875 (CH₂), 1 631 (C=O), 1 589, 1 454; ESI-MS m/z: 157.14 (100), 159.12 (62), 442.26 ([M+H]⁺, 72); Elemental anal. (%) Calcd. for C₂₅H₁₉N₃O₃S: C 68.03, H 4.31, N 9.52; Found: C 67.99, H 4.40, N 9.54.
6b	¹H NMR δ: 14.20 (s, 1H, NH), 8.06 (d, J = 7.0 Hz, 2H, ArH), 7.94 (d, J = 9.0 Hz, 1H, ArH), 7.54−7.61 (m, 4H, ArH), 7.33−7.38 (m, 4H, ArH), 7.10 (d, J = 8.0 Hz, 1H, ArH), 6.95 (s, 1H, 3-H), 4.51 (t, J = 6.5 Hz, 2H, CH₂), 3.60 (s, 2H, CH₂), 2.45 (s, 3H, CH₃); IR (KBr) ν/cm⁻¹: 3 448 (NH), 3 085 (Ar), 2 997, 2 918 (CH₂), 1 639 (C=O), 1 560, 1 517, 1 461; ESI-MS m/z: 157.14 (100), 179.14 (52), 456.22 ([M+H]⁺, 88); Elemental anal. (%) Calcd. for C₂₆H₂₁N₃O₃S: C 68.57, H 4.62, N 9.23; Found: C 68.61, H 4.58, N 9.22.
6c	¹H NMR δ: 14.38 (s), 14.09 (s, 1H, NH), 8.05 (d, J = 7.0 Hz, 2H, ArH), 7.94 (d, J = 8.5 Hz, 1H, ArH), 7.84 (d, J = 8.0 Hz, 2H, ArH), 7.53−7.61 (m, 3H, ArH), 7.39 (s, 1H, ArH), 7.29 (s, 2H, ArH), 7.11 (d, J = 8.5 Hz, 1H, ArH), 6.95 (s, 1H, 3-H), 4.51 (t, J = 6.5 Hz, 2H, CH₂), 3.58 (s, 2H, CH₂), 2.32 (s, 3H, CH₃); IR (KBr) ν/cm⁻¹: 3 442 (NH), 3 103 (Ar), 2 981 (CH₂), 1 632 (C=O), 1 504, 1 454; ESI-MS m/z: 157.14 (100), 179.14 (44), 456.29 ([M+H]⁺, 61); Elemental anal. (%) Calcd. for C₂₆H₂₁N₃O₃S: C 68.57, H 4.62, N 9.23; Found: C 68.67, H 4.59, N 9.23.
6d	¹H NMR δ: 14.44 (s), 14.13 (s, 1H, NH), 8.05 (d, J = 7.5 Hz, 2H, ArH), 7.94 (d, J = 9.0 Hz, 1H, ArH), 7.79 (s, 1H, ArH), 7.75 (d, J = 7.5 Hz, 1H, ArH), 7.53−7.61 (m, 3H, ArH), 7.39 (s, 1H, ArH), 7.29 (s, 2H, ArH), 7.11 (d, J = 8.0 Hz, 1H, ArH), 6.94 (s, 1H, 3-H), 4.51 (t, J = 6.5 Hz, 2H, CH₂), 3.59 (s, 2H, CH₂), 2.34 (s, 3H, CH₃); IR (KBr) ν/cm⁻¹: 3 464 (NH), 3 082 (Ar), 2 933 (CH₂), 1 624 (C=O), 1 512, 1 448; ESI-MS m/z: 157.14 (100), 179.14 (87), 456.22 ([M+H]⁺, 61); Elemental anal. (%) Calcd. for C₂₆H₂₁N₃O₃S: C 68.57, H 4.62, N 9.23; Found: C 68.62, H 4.61, N 9.30.

Table 3 Spectra data of compounds 4, 5 and 6

1.6 7-(3-苯基-4-水杨醛亚氨基-1,2,4-三唑-5-硫)乙氧基黄酮 (5a) 的合成

在50 mL圆底烧瓶中加入0.04 g (1 mmol) 氢氧化钠和20 mL无水乙醇, 溶解后加入1 mmol的3-苯基-4-水杨醛亚氨基-1,2,4-三唑- 5-硫酮 (化合物1a), 继续搅拌微热至溶解后, 加入0.345 g (1 mmol) 7-溴乙氧基黄酮, 缓慢升温至回流, TLC跟踪 (展开剂为V_乙酸乙酯∶V_石油醚=1∶1), 待原料点7-溴乙氧基黄酮消失后停止反应。静置, 冷却, 抽滤得到白色固体, 用乙醇/二甲亚砜重结晶, 得7-(3-苯基- 4-水杨醛亚氨基-1,2,4-三唑-5-硫)乙氧基黄酮5a。类似方法合成化合物5b～5e, 化合物理化和波谱数据见表 2和表 3。

Table 4 Antioxidative activity of target compounds

Compd.	The relative inhibition rate on free radicals/%			The relative reducibility/%
Compd.	DPPH	OH	O₂^－·	The relative reducibility/%
4a	48.73	11.95	0.45	15.53
4b	6.16	1.71	11.03	17.81
4c	17.08	14.39	0.89	15.41
4d	52.93	11.95	1.34	15.98
4e	21.56	19.51	13.46	17.69
4f	45.66	3.17	2.96	16.44
5a	2.52	20.00	8.07	15.75
5b	3.92	7.56	86.33	15.98
5c	0.56	10.24	20.46	15.53
5d	7.85	6.09	2.25	16.63
5e	0.24	23.63	95.54	17.12
6a	67.33	37.43	68.79	30.04
6b	69.01	41.05	74.45	31.73
6c	77.64	46.79	79.03	29.66
6d	47.83	34.67	85.09	34.23
Vc	100	100	100	100

Table 4 Antioxidative activity of target compounds

1.7 7-(3-苯基-1,2,4-三唑-5-硫)乙氧基黄酮 (6a) 的合成

在50 mL圆底烧瓶中加入0.04 g (1 mmol) 氢氧化钠和20 mL无水乙醇, 溶解后加入1 mmol的3-苯基-4-氢-5-巯基-1,2,4-三唑(化合物3a), 继续搅拌加入0.345 g (1 mmol) 7-溴乙氧基黄酮, 缓慢升温至回流, TLC跟踪 (展开剂为V_乙酸乙酯∶V_石油醚=1∶1), 待原料点7-溴乙氧基黄酮消失后停止反应。静置, 冷却, 减压蒸除部分乙醇, 抽滤得到白色固体, 用乙醇重结晶, 得7-(3-苯基-4-氢-1,2,4-三唑-5-硫)乙氧基黄酮6a。类似方法合成化合物6b～6d, 化合物理化和波谱数据见表 2和表 3。

2 体外抗氧化活性实验

目标化合物抗氧化活性实验参考文献^[14]方法进行, 对照选用常用抗氧化剂维生素C, 结果用相对清除率表示, 其中羟自由基由Fe²^＋-H₂O₂-亚甲蓝体系产生, 清除率 = [(A₀ − A_样) / A₀] × 100%, 相对清除率 = 样品的清除率 / Vc的清除率 × 100%; 超氧自由基采用碱性条件下邻苯三酚的自氧化产生, 清除率 = [(A₀ − A_样) / A₀]×100%, 相对清除率=样品的清除率 / Vc的清除率 × 100%; DPPH·自由基直接用无水乙醇配制使用, 清除率 = [1 − (A₁ − A₂) / A₀] ×100%, 相对清除率 = 样品的清除率 / Vc的清除率 ×100%; 还原能力测定采用铁氰化钾还原法, 还原能力∆A = A_样 − A₀, 相对还原能力 = 样品的还原能力 / Vc的还原能力×100%。试样浓度配制为0.5 mg·mL⁻¹的N,N-二甲基甲酰胺溶液。

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药学学报 2015, Vol. 50 Issue (7): 861-867	PDF