药学学报

No.	δ_C	δ_H, mult		No.	δ_C	δ_H, mult
1	84.8	-		18	25.9	1.70, s
2	203.1	-	19	31.1	1.48, d, J = 13.0 Hz
					2.97, dd, J = 13.0, 5.5 Hz
3	77.7	-	20	82.0	4.29, d, J = 5.5 Hz
4	111.1	-	21	84.3	-
5	61.6	-	22	28.0	1.30, s
6	37.2	1.57, t, J = 14.0 Hz	23	21.9	1.37, s
		2.54, dd, J = 14.0, 4.0 Hz
7	46.0	1.34, m	24	27.9	1.68, m
					2.12, m
8	54.1	-	25	122.5	4.99, t, J = 7.0 Hz
9	206.6	-	26	133.4	-
10	209.3	-	27	18.3	1.70, s
11	41.2	2.69, sept, J = 6.5 Hz	28	18.2	1.57, s
12	21.7	0.97, d, J = 6.5 Hz	29	13.9	1.05, s
13	20.3	1.08, d, J = 6.5 Hz	30	37.0	1.16, m
					2.05, m
14	31.9	2.61, dd, J = 14.0, 7.5 Hz	31	25.6	1.88, m
		2.91, dd, J = 14.0, 7.5 Hz			2.16, m
15	118.2	5.36, t, J = 7.5 Hz	32	124.7	5.07, t, J = 6.5 Hz
16	136.4	-	33	131.5	-
17	26.4	1.73, s	34	25.9	1.66, s
			35	17.9	1.60, s

No.	δ_C	δ_H, mult		No.	δ_C	δ_H, mult
1	84.8	-		18	25.9	1.70, s
2	203.1	-	19	31.1	1.48, d, J = 13.0 Hz
					2.97, dd, J = 13.0, 5.5 Hz
3	77.7	-	20	82.0	4.29, d, J = 5.5 Hz
4	111.1	-	21	84.3	-
5	61.6	-	22	28.0	1.30, s
6	37.2	1.57, t, J = 14.0 Hz	23	21.9	1.37, s
		2.54, dd, J = 14.0, 4.0 Hz
7	46.0	1.34, m	24	27.9	1.68, m
					2.12, m
8	54.1	-	25	122.5	4.99, t, J = 7.0 Hz
9	206.6	-	26	133.4	-
10	209.3	-	27	18.3	1.70, s
11	41.2	2.69, sept, J = 6.5 Hz	28	18.2	1.57, s
12	21.7	0.97, d, J = 6.5 Hz	29	13.9	1.05, s
13	20.3	1.08, d, J = 6.5 Hz	30	37.0	1.16, m
					2.05, m
14	31.9	2.61, dd, J = 14.0, 7.5 Hz	31	25.6	1.88, m
		2.91, dd, J = 14.0, 7.5 Hz			2.16, m
15	118.2	5.36, t, J = 7.5 Hz	32	124.7	5.07, t, J = 6.5 Hz
16	136.4	-	33	131.5	-
17	26.4	1.73, s	34	25.9	1.66, s
			35	17.9	1.60, s

Compound	NO inhibition rate	Cell proliferation inhibition rate
Dex	94.88%	8.26%
1	57.99%	7.63%
2	59.42%	26.18%
3	54.92%	11.97%
4	48.15%	33.37%
5	65.99%	6.67%
6	53.23%	26.29%
7	55.43%	11.54%
8	51.34%	37.64%

Compound	NO inhibition rate	Cell proliferation inhibition rate
Dex	94.88%	8.26%
1	57.99%	7.63%
2	59.42%	26.18%
3	54.92%	11.97%
4	48.15%	33.37%
5	65.99%	6.67%
6	53.23%	26.29%
7	55.43%	11.54%
8	51.34%	37.64%

北栽秧花中一个新的间苯三酚类化合物

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马永慧 ¹^,² , 孙浩然 ² , 李进 ¹^,^* , 吉腾飞 ²^,³^,^*

药学学报 | 研究论文 2022,57(12): 3596-3602

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药学学报 | 研究论文 2022, 57(12): 3596-3602

北栽秧花中一个新的间苯三酚类化合物

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马永慧¹^,², 孙浩然², 李进¹^,^*, 吉腾飞²^,³^,^*

作者信息

1.新疆师范大学生命科学学院, 新疆特殊环境物种保护与调控生物学实验室, 干旱区植物逆境生物学实验室, 新疆乌鲁木齐 830054

2.中国医学科学院、北京协和医学院药物研究所, 天然药物活性物质与功能国家重点实验室, 北京 100050

3.中国科学院西北高原生物研究所, 青海省藏药研究重点实验室, 藏药研究重点实验室, 青海西宁 810001

通讯作者:

*李进, Tel: 86-991-4332325, E-mail: xjcjlj4@xjnu.edu.cn;

吉腾飞 , Tel: 86-10-63165226, E-mail: jitf@imm.ac.cn

A new polycyclic polyprenylated acylphloroglucinol from Hypericum pseudohenryi

Yong-hui MA¹^,², Hao-ran SUN², Jin LI¹^,^*, Teng-fei JI²^,³^,^*

Affiliations

1. Xinjiang Key Laboratory of Special Species Conservation and Regulatory Biology, Key Laboratory of Plant Stress Biology in Arid Land, College of Life Science, Xinjiang Normal University, Urumqi 830054, China

2. State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China

3. Qinghai Provincial Key Laboratory of Tibetan Medicine Research, Key Laboratory of Tibetan Medicine Research, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, Xining 810001, China

出版时间: 2022-12-12 doi: 10.16438/j.0513-4870.2022-0654

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摘要

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采用MCI柱色谱、硅胶柱色谱、制备高效液相色谱等对北栽秧花(Hypericum pseudohenryi N. Robson) 95%乙醇提取物中间苯三酚类成分进行分离纯化, 从中分离得到8个多环多异戊烯基酰基化间苯三酚化合物, 分别鉴定为hypseudohenrin L (1)、hyperbeanin P (2)、furohyperforin (3)、furoadhyperforin (4)、ascyronone F (5)、attenuatumione E (6)、hyphenrone T (7) 和hyperforatone N (8), 其中化合物1为新间苯三酚类成分。利用MTT法探究8个化合物对脂多糖(LPS) 诱导的原代小鼠腹腔巨噬细胞一氧化氮(NO) 生成的影响, 结果表明, 化合物1、3、5和7对LPS诱导巨噬细胞NO生成具有显著的抑制作用, 说明其具有潜在的抗炎活性。

关键词

北栽秧花 / 间苯三酚 / 一氧化氮 / 抗炎活性

Abstract

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Eight polycyclic polyprenylated acylphloroglucinols (PPAPs) were isolated from the 95% ethanol extract of Hypericum pseudohenryi by MCI column, silica gel column and preparative HPLC. These compounds were defined as hypseudohenrin L (1), hyperbeanin P (2), furohyperforin (3), furoadhyperforin (4), ascyronone F (5), attenuatumione E (6), hyphenrone T (7) and hyperforatone N (8), compound 1 is a new compound. Compounds 1, 3, 5, 7 showed an inhibitory effect on lipopolysaccharide (LPS)-induced nitric oxide (NO) production in macrophage cells.

Key words

Hypericum pseudohenryi / polycyclic polyprenylated acylphloroglucinol / nitric oxide / anti-inflammatoy activity

引用本文

马永慧, 孙浩然, 李进, 吉腾飞. 北栽秧花中一个新的间苯三酚类化合物. 药学学报, 2022 , 57 (12) : 3596 -3602 . DOI: 10.16438/j.0513-4870.2022-0654

Yong-hui MA, Hao-ran SUN, Jin LI, Teng-fei JI. A new polycyclic polyprenylated acylphloroglucinol from Hypericum pseudohenryi[J]. Acta Pharmaceutica Sinica, 2022 , 57 (12) : 3596 -3602 . DOI: 10.16438/j.0513-4870.2022-0654

正文

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北栽秧花(Hypericum pseudohenryi N. Robson) 为藤黄科(Guttiferae) 金丝桃属植物, 该属植物在全球约有400种, 其主要特征性成分为多环多异戊烯基取代酰基间苯三酚类化合物(polycyclic polyprenylated acylphoroglucinols, PPAPs)^[1], 是一类具有异戊烯基、香叶烯基及酰基侧链取代, 高度氧化环合结构特点的化合物。金丝桃属代表性植物贯叶金丝桃已有2 400余年的药用历史^[2], 北栽秧花用于治疗急慢性黄疸型肝炎、尿道感染、结石、风湿疼痛、跌打损伤^[3]等。现代药理学研究表明, 金丝桃属植物具有抗炎、抗肿瘤及抗神经退行性疾病等活性, 而炎症为疾病基本病理过程, 与多种疾病的进程及转归相关, 因此对天然药物的抗炎药效物质基础进行研究, 发现先导化合物是切实可行的。近年发现一系列抗炎活性良好的天然产物, 包括黄酮类化合物桑色素^[4]、倍半萜^[5]、西松烷型二萜^[6]和多糖^[7]等。

本研究在已有间苯三酚类成分的工作基础上^[8], 开展了北栽秧花间苯三酚类成分的研究。分得8个间苯三酚类化合物, 化合物1为新化合物。结合其民间用于治疗尿路感染、风湿疼痛等炎症相关疾病, 以一氧化氮(NO) 释放抑制作用为切入点, 筛选北栽秧花中抗炎活性成分, 采用MTT法探究8个化合物对脂多糖(LPS) 诱导的原代小鼠腹腔巨噬细胞NO生成的影响, 结果表明, 化合物1、3、5和7对LPS诱导巨噬细胞NO生成显著的抑制作用, 说明其具有潜在的抗炎活性。

结果与讨论

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1 结构鉴定

化合物1 无色油状物, [α]$ {}_{\mathrm{D}}^{20} $ +22.4 (c 0.71, MeOH)。由高分辨质谱数据可知其分子式为C₃₅H₅₂O₆ (m/z 569.383 0 [M + H]⁺, 计算值569.383 7), 不饱和度为10。在¹H NMR (500 MHz, CDCl₃) 图谱中, 未观察到苯环上的氢信号, 但存在一个次甲基和两个明显的双峰甲基信号, 且偶合常数相同(δ_H 0.97, 3H, d, J = 6.5 Hz; 1.08, 3H, d, J = 6.5 Hz; 2.69, 1H, sept, J = 6.5 Hz), 提示该化合物中可能存在异丁酰基, 高场区显示9个单峰甲基信号δ_H 1.73 (3H, s)、1.70 (3H, s)、1.70 (3H, s)、1.66 (3H, s)、1.60 (3H, s)、1.57 (3H, s)、1.37 (3H, s)、1.30 (3H, s)、1.05 (3H, s) 和3个烯氢信号δ_H 5.36 (1H, t, J = 7.5 Hz)、5.07 (1H, t, J = 6.5 Hz)、4.99 (1H, t, J = 7.0 Hz)。结合¹³C NMR (125 MHz, CDCl₃), 该化合物共有35个碳信号, 可见3个非共轭的羰基信号(δ_C 209.3、206.6、203.1) 和1个双连氧的碳信号(δ_C 111.1), 上述信号包含6个不饱和度, 因此, 化合物1中存在4个环系。

在HMBC谱中, H-14 (δ_H 2.61) 与3个季碳相关(C-2, δ_C 203.1; C-3, δ_C 77.7; C-4, δ_C 111.1), H-24 (δ_H 2.12) 与C-6 (δ_C 37.2)、C-7 (δ_C 46.0)、C-8 (δ_C 54.1)、CH₃-29 (δ_C 13.9) 相关, H-30 (δ_H 2.05) 与C-8、C-1 (δ_C 84.8) 相关, 明确了3个异戊烯基与母核的连接位置, 且CH₃-29不与C-3相关, 故化合物1应为Type A双环多异戊烯基取代的酰基间苯三酚型(bicycle polyprenylated acylphloroglucinols, BPAPs) 化合物。H-19 (δ_H 1.48) 与C-4 (δ_C 111.1)、C-5 (δ_C 61.6) 相关, 而C-4又为双连氧碳, 提示C₄-C₅位置可能稠合了四氢呋喃环。偕二甲基(H-22, δ_H 1.30; H-23, 1.37) 与两个连氧碳(C-20, δ_C 82.0; C-21, δ_C 84.3) 相关, 确定了四氢呋喃环以及2-羟基丙基的存在。C-4位在形成一个四氢呋喃环之外, 另一个氧原子也需要成环, 因此在C₃-C₄之间存在一个环氧结构。化合物1的平面结构如图 1所示, 其波谱数据归属见表 1。

化合物1的相对构型由ROESY实验确定。H-20 (δ_H 4.29, d, J = 5.5 Hz) 仅有一个偶合常数, 表明在具有四氢呋喃环的BPAP结构中该氢原子处于β-取向, 因为其与H-19α (δ_H 1.48, d, J = 13.0 Hz) 的二面角几乎为90度, 根据Karplus规则^[9]可知, 此时两个氢的偶合常数几乎等于零; 若是H-20处于α-取向, 则其峰形为dd峰。C-7的化学位移为46.0, 且H-6α (δ_H 2.54) 与H-6β (δ_H 1.57) 的化学位移差值为0.97, 故C-7处的异戊烯基处于β-取向。在ROESY谱中, 可以观察到CH₃-29 (δ_H 1.05) 与H-6β的相关信号, 故CH₃-29也处于β-取向; 同时可以观察到CH₂ -14 (δ_H 2.61) 与H-7 (δ_H 1.34) 的相关信号, 确定3, 4位环氧处于β-取向; 因此化合物1的相对构型确定如图 2所示。

化合物1的绝对构型是根据其ECD图谱与化合物kiiacylphnol A^[10]的相关数据进行对比确定的(图 3)。化合物1平面结构与化合物kiiacylphnol A类似, 区别在于化合物1的C-4, 5骈合的是四氢呋喃环; 而化合物kiiacylphnol A则是四氢吡喃环, 二者的母核与相对构型也一致。化合物1的实验ECD图谱曲线趋势与化合物kiiacylphnol A实验和计算ECD图谱曲线趋势一致, 确定其绝对构型为1S, 3S, 4R, 5R, 7S, 8R, 23R, 将化合物1命名为hypseudohenrin L。

2 化合物活性测试

利用MTT法对8个间苯三酚类化合物进行细菌LPS诱导原代小鼠腹腔巨噬细胞产生NO生成的影响, 并以地塞米松(Dex) 为阳性对照进行评价。结果表明化合物1、3、5和7对炎性因子NO的产生具有显著的抑制作用(表 2)。

实验部分

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V-650紫外可见分光光度计、J-815圆二色性偏光光度计(日本JASCO公司); AUTOPOL V自动旋光仪(美国Rudolph Research Analytical公司); Nicolet 5700傅立叶变换红外光谱仪、Nicolet is50傅立叶变换红外光谱仪、Q Exactive液相色谱-质谱联用系统(美国Thermo Fisher Scientific公司); ECZ-400S (日本JEOL公司); WNMR-I 500 (Q.One Instruments Ltd.); Bruker Av III 500/600型核磁共振波谱仪(德国Bruker公司); 硅胶H、薄层色谱硅胶(青岛海洋化工厂); LC-6AD与SPD-20A制备型高效液相色谱仪系统(日本Shimadzu公司); Agilent 1200 (分析型; 美国Agilent公司); UG80 5 μm, 250 mm × 20 mm (日本Shiseido公司); 乙腈、甲醇(色谱纯, 美国Fisher公司); 其余试剂均为国产分析纯。

北栽秧花于2017年6月采自云南省丽江市, 并由中国科学院昆明植物研究所张君老师鉴定为藤黄科金丝桃属植物北栽秧花(H. pseudohenryi), 标本现存于中国医学科学院药物研究所植物标本室, 标本号为No. ID-24705。

1 提取与分离

取干燥的北栽秧花地上部分25.4 kg, 粉碎后用95%乙醇加热回流提取3次, 每次3 h, 提取液减压回收溶剂后得浸膏1.7 kg, 将此浸膏混悬于5 L水中, 用等体积石油醚萃取3次; 再将水层用等体积乙酸乙酯萃取3次, 合并减压回收溶剂得浸膏1.2 kg。将该浸膏经MCI柱色谱, 60%~95%乙醇梯度洗脱, 合并相同极性部分共得到9个组分Fr. I~Fr. IX。流分Fr. IV (232 g) 经MCI柱色谱, 45%~95%乙醇梯度洗脱, 共得到9个组分(Fr. IV.1~IV.9)。Fr. IV.7 (115.9 g) 经DIOL柱色谱, 石油醚-乙酸乙酯(1∶0~0∶1) 梯度洗脱, 合并得到15个组分(Fr. IV.7.1~IV.7.15)。Fr. IV.7.8 (10.3 g) 经硅胶(200~300目) 柱色谱, 石油醚-乙酸乙酯(1∶0~0∶1) 梯度洗脱, 得到8个组分(Fr. IV.7.8.1~IV.7.8.8)。流分Fr. IV.7.8.2 (295 mg) 经制备HPLC (90% MeOH-H₂O) 分离纯化, 得化合物1 (15 mg); 流分Fr. IV.7.8.3 (1 651 mg) 经制备HPLC (93% MeOH-H₂O) 分离纯化, 得化合物2 (7 mg)、4 (32 mg)、6 (22 mg)、7 (28 mg)、8 (108 mg); 流分Fr. IV.7.8.4 (814 mg) 经制备HPLC (95% CH₃CN-H₂O) 分离纯化, 得化合物3 (3 mg)、5 (35 mg)。

2 结构鉴定

化合物1 无色油状物; [α]$ {}_{\mathrm{D}}^{20} $ +22.4 (c 0.71, MeOH); UV (MeOH) λ_max (log ε) 204.0 (4.12) nm; ECD (MeOH) λ_max (Δε) 300.0 (+1.74), 336.0 (+0.38), 366.5 (-0.42), 388.0 (+0.16) nm; IR ν_max 2 974, 2 929, 2 872, 1 734, 1 722, 1 697, 1 448, 1 379, 1 057, 987 cm^-1; ¹H NMR (CDCl₃, 500 MHz) 和¹³C NMR (CDCl₃, 125 MHz) 数据见表 1; HR-ESI-MS m/z 569.383 0 [M+H]⁺ (计算值569.383 7, C₃₅H₅₃O₆)。

化合物2 无色油状物; HR-ESI-MS m/z 585.357 5, [M+H]⁺ (计算值585.357 5, C₃₈H₄₉O₅); ¹H NMR (CDCl₃, 400 MHz) δ_H: 7.78 (1H, d, J = 6.0 Hz, H-12), 7.78 (1H, d, J = 6.4 Hz, H-16), 7.45 (1H, t, J = 6.4 Hz, H-14), 7.31 (1H, t, J = 6.4 Hz, H-13), 7.31 (1H, t, J = 6.4 Hz, H-15), 5.93 (1H, d, J = 7.6 Hz, H-33), 5.11 (1H, t, J =6.4 Hz, H-18), 4.90 (1H, t, J = 5.6 Hz, H-28), 4.72 (1H, t, J = 7.2 Hz, H-23), 4.42 (1H, d, J =9.6 Hz, H-35), 3.66 (1H, t, J = 9.2 Hz, H-34), 3.16~3.27 (1H, m, H-17), 2.72 (1H, dd, J = 10.4, 7.2 Hz, H-22a), 2.59 (1H, m, H-7), 2.37 (1H, d, J = 5.6 Hz, H-6a), 2.37 (1H, dd, J = 10.4, 7.2 Hz, H-22b), 1.97~2.12 (2H, m, H-27), 1.72 (3H, s, H-20), 1.69 (3H, s, H-30), 1.66 (3H, s, H-21), 1.64 (3H, s, H-32), 1.58 (3H, s, H-31), 1.53 (3H, s, H-25), 1.42 (1H, t, J = 10.4 Hz, H-6b), 1.38 (3H, s, H-26), 1.34 (3H, s, H-37), 1.31 (3H, s, H-38); ¹³C NMR (CDCl₃, 100 MHz) δ_C: 74.6 (C-1), 166.1 (C-2), 125.0 (C-3), 199.7 (C-4), 62.8 (C-5), 43.4 (C-6), 39.0 (C-7), 145.8 (C-8), 204.7 (C-9), 195.5 (C-10), 136.8 (C-11), 128.4 (C-12), 128.6 (C-13), 132.8 (C-14), 128.6 (C-15), 128.4 (C-16), 22.6 (C-17), 120.7 (C-18), 133.2 (C-19), 26.1 (C-20), 18.1 (C-21), 40.0 (C-22), 118.2 (C-23), 136.1 (C-24), 26.2 (C-25), 17.7 (C-26), 32.9 (C-27), 122.0 (C-28), 133.6 (C-29), 25.9 (C-30), 18.3 (C-31), 19.3 (C-32), 134.6 (C-33), 51.5 (C-34), 73.2 (C-35), 90.8 (C-36), 28.5 (C-37), 19.9 (C-38)。以上数据与文献^[11]报道基本一致, 故鉴定化合物为hyperbeanin P。

化合物3 无色油状物; HR-ESI-MS m/z 553.388 7, [M+H]⁺ (计算值585.388 7, C₃₅H₅₃O₅); ¹H NMR (400 MHz, CDCl₃) δ_H: 5.08 (1H, br dd, J = 6.8, 5.2 Hz, H-27), 5.04 (1H, m, H-17), 4.94 (1H, m, H-22), 4.63 (1H, dd, J = 10.8, 10.8 Hz, H-32), 4.55 (1H, dd, J = 10.8, 10.8 Hz, H-32), 3.11 (1H, br dd, J = 14.4, 14.4 Hz, H-26a), 3.00 (1H, br dd, J = 14.4, 14.4 Hz, H-26b), 2.64 (1H, dd, J = 13.2, 13.2 Hz, H-31a), 2.14 (1H, m, H-16a), 2.05 (1H, m, H-15a), 2.01 (1H, br dd, J = 8.0, 8.0 Hz, H-5a), 1.99 (1H, qq, J = 8.0, 8.0 Hz, H-11), 1.95 (1H, m, H-16b), 1.80 (2H, m, H-21), 1.77 (1H, dd, J = 12.0, 12.0 Hz, H-31b), 1.70 (3H, s, H-24), 1.70 (3H, s, H-30), 1.68 (3H, br s, H-19), 1.68 (3H, br s, H-29), 1.63 (3H, br s, H-20), 1.62 (1H, m, H-4), 1.58 (3H, s, H-25), 1.51 (1H, dd, J = 2.4, 2.8 Hz, H-5b), 1.36 (3H, s, H-34), 1.30 (1H, m, H-15b), 1.20 (3H, s, H-35), 1.08 (3H, d, J = 8.0 Hz, H-12), 1.03 (3H, s, H-14), 1.00 (3H, d, J = 8.0 Hz, H-13); ¹³C NMR (100 MHz, CDCl₃) δ_C: 204.7 (C-1), 83.4 (C-2), 48.4 (C-3), 43.4 (C-4), 38.2 (C-5), 59.6 (C-6), 173.1 (C-7), 116.8 (C-8), 192.9 (C-9), 209.6 (C-10), 42.1 (C-11), 20.5 (C-12), 21.5 (C-13), 13.6 (C-14), 36.5 (C-15), 25.3 (C-16), 124.9 (C-17), 131.2 (C-18), 25.8 (C-19), 17.8 (C-20), 27.0 (C-21), 122.4 (C-22), 133.7 (C-23), 25.8 (C-24), 18.0 (C-25), 22.3 (C-26), 121.3 (C-27), 132.7 (C-28), 25.8 (C-29), 17.8 (C-30), 30.3 (C-31), 90.3 (C-32), 71.0 (C-33), 27.2 (C-34), 24.2 (C-35)。以上数据与文献^[12]报道基本一致, 故鉴定化合物为furohyperforin。

化合物4 无色油状物; HR-ESI-MS m/z 567.405 1, [M+H]⁺ (计算值567.404 4, C₃₆H₅₅O₅); ¹H NMR (400 MHz, CD₃OD) δ_H: 5.11 (1H, br t, J = 4.8 Hz, H-17), 5.11 (1H, br t, J = 4.8 Hz, H-27), 4.88 (1H, t, J = 4.4 Hz, H-22), 4.61 (1H, dd, J = 3.2, 7.2 Hz, H-32), 3.09 (1H, dd, J = 4.0, 10.4 Hz, H-26a), 2.97 (1H, dd, J = 5.2, 10.4 Hz, H-26b), 2.61 (1H, dd, J = 7.2, 9.6 Hz, H-31a), 2.15 (2H, m, H-21), 2.13 (1H, m, H-16a), 2.01 (1H, dd, J = 2.8, 10.0 Hz, H-5a), 1.93 (1H, m, H-15a), 1.92 (1H, m, H-16b), 1.84 (1H, dd, J = 3.6, 9.6 Hz, H-31b), 1.72 (1H, m, H-11), 1.70 (3H, s, H-24), 1.70 (3H, s, H-30), 1.69 (2H, m, H-13b), 1.67 (3H, s, H-29), 1.64 (3H, s, H-19), 1.62 (1H, m, H-4), 1.59 (3H, s, H-20), 1.57 (3H, s, H-25), 1.54 (1H, t, J = 9.6 Hz, H-5b), 1.30 (1H, m, H-15b), 1.30 (3H, s, H-34), 1.22 (1H, m, H-13a), 1.18 (1H, s, H-35), 1.03 (3H, d, J = 3.6 Hz, H-12), 1.01 (3H, s, H-14), 0.74 (13-OCH₃); ¹³C NMR (100 MHz, CD₃OD) δ_C: 204.6 (C-1), 83.3 (C-2), 48.5 (C-3), 43.0 (C-4), 37.8 (C-5), 59.6 (C-6), 174.7 (C-7), 116.2 (C-8), 193.4 (C-9), 209.7 (C-10), 48.5 (C-11), 16.7 (C-12), 27.2 (C-13), 13.1 (C-14), 36.5 (C-15), 24.8 (C-16), 124.6 (C-17), 130.7 (C-18), 24.8 (C-19), 16.7 (C-20), 27.2 (C-21), 122.2 (C-22), 133.2 (C-23), 26.8 (C-24), 16.7 (C-25), 21.7 (C-26), 121.4 (C-27), 131.8 (C-28), 24.8 (C-29), 16.7 (C-30), 29.5 (C-31), 90.8 (C-32), 70.0 (C-33), 26.8 (C-34), 24.2 (C-35), 10.6 (13-OCH₃)。以上数据与文献^[13]报道基本一致, 故鉴定化合物为furoadhyperforin。

化合物5 无色油状物; HR-ESI-MS m/z 587.372 8, [M+H]⁺ (计算值587.373 1, C₃₈H₅₁O₅); ¹H NMR (500 MHz, CDCl₃) δ_H: 7.56 (1H, d, J = 10.0 Hz, H-12), 7.56 (1H, d, J = 10.0 Hz, H-16), 7.48 (1H, t, J = 9.5 Hz, H-14), 7.34 (1H, dd, J = 8.8, 7.2 Hz, H-13), 7.34 (1H, dd, J = 10.0, 9.5 Hz, H-15), 5.02 (1H, m, H-19), 4.99 (1H, m, H-25), 4.95 (1H, m, H-30), 4.59 (1H, t, J = 10.0 Hz, H-35), 2.94 (1H, dd, J = 10.0, 4.5 Hz, H-34), 2.56 (1H, m, H-29b), 2.47 (1H, m, H-29a), 2.11 (1H, m, H-24), 2.09 (1H, m, H-18), 2.01 (1H, m, H-9a), 1.80 (1H, m, H-8), 1.69 (1H, m, H-17), 1.67 (3H, s, H-27), 1.67 (3H, s, H-32), 1.66 (3H, s, H-33), 1.64 (3H, s, H-21), 1.59 (3H, s, H-22), 1.57 (3H, s, H-28), 1.44 (1H, m, H-9b), 1.25 (3H, s, H-23), 0.90 (3H, s, H-37), 0.86 (3H, s, H-38); ¹³C NMR (125 MHz, CDCl₃) δ_C: 70.7 (C-1), 207.0 (C-2), 65.1 (C-3), 190.6 (C-4), 171.8 (C-5), 118.9 (C-6), 49.5 (C-7), 41.6 (C-8), 41.1 (C-9), 193.4 (C-10), 137.8 (C-11), 128.6 (C-12), 128.3 (C-13), 132.8 (C-14), 128.3 (C-15), 128.6 (C-16), 38.0 (C-17), 25.8 (C-18), 124.4 (C-19), 131.9 (C-20), 25.8 (C-21), 17.9 (C-22), 15.0 (C-23), 28.3 (C-24), 122.3 (C-25), 133.8 (C-26), 25.8 (C-27), 17.9 (C-28), 29.5 (C-29), 119.6 (C-30), 134.9 (C-31), 18.2 (C-32), 26.0 (C-33), 26.2 (C-34), 93.7 (C-35), 70.7 (C-36), 24.0 (C-37), 26.5 (C-38)。以上数据与文献^[14]报道基本一致, 故鉴定化合物为ascyronone F。

化合物6 无色油状物; HR-ESI-MS m/z 553.388 7, [M+H]⁺ (计算值553.388 7, C₃₅H₅₃O₅); ¹H NMR (400 MHz, CDCl₃) δ_H: 5.00 (1H, dd, J = 9.2, 5.6 Hz, H-20), 5.00 (1H, dd, J = 9.2, 5.6 Hz, H-32), 4.94 (1H, t, J = 6.0 Hz, H-25), 4.77 (1H, t, J = 8.0 Hz, H-11), 2.95 (2H, d, J = 8.0 Hz, H-10), 2.55 (1H, dd, J = 8.8, 5.2 Hz, H-16), 2.44 (2H, dd, J = 10.4, 5.6 Hz, H-19), 2.07 (1H, dd, J = 6.0, 4.8 Hz, H-24a), 2.00 (1H, dd, J = 8.8, 5.2 Hz, H-31a), 1.94 (1H, dd, J = 8.4, 5.2 Hz, H-31b), 1.78 (2H, dd, J = 9.2, 2.8 Hz, H-6), 1.70 (1H, m, H-24b), 1.68 (1H, s, H-7), 1.67 (3H, s, H-27), 1.67 (2H, s, H-30), 1.64 (3H, s, H-23), 1.63 (3H, s, H-33), 1.57 (3H, s, H-35), 1.55 (3H, s, H-28), 1.42 (3H, m, H-13), 1.22 (3H, s, H-14), 1.20 (3H, s, H-18), 1.18 (3H, s, H-17), 1.12 (3H, s, H-29); ¹³C NMR (100 MHz, CDCl₃) δ_C: 74.2 (C-1), 171.6 (C-2), 120.8 (C-3), 190.8 (C-4), 64.0 (C-5), 39.6 (C-6), 42.4 (C-7), 47.7 (C-8), 205.8 (C-9), 26.9 (C-10), 94.0 (C-11), 71.2 (C-12), 27.1 (C-13), 25.4 (C-14), 208.9 (C-15), 41.0 (C-16), 21.2 (C-17), 21.2 (C-18), 29.4 (C-19), 119.7 (C-20), 134.4 (C-21), 26.0 (C-22), 18.2 (C-23), 28.3 (C-24), 122.4 (C-25), 133.6 (C-26), 26.1 (C-27), 17.9 (C-28), 14.6 (C-29), 38.2 (C-30), 24.4 (C-31), 124.5 (C-32), 131.8 (C-33), 25.8 (C-34), 18.2 (C-35)。以上数据与文献^[15]报道基本一致, 故鉴定化合物为attenuatumione E。

化合物7 无色油状物; HR-ESI-MS m/z 569.383 5, [M+H]⁺ (计算值569.383 6, C₃₅H₅₃O₆); ¹H NMR (400 MHz, CD₃OD) δ_H: 5.09 (1H, t, J = 6.4 Hz, H-18), 5.01 (1H, t, J = 6.0 Hz, H-35), 4.87 (1H, t, J = 6.0 Hz, H-28), 3.79 (1H, m, H-23), 2.91 (1H, sept, J = 4.8 Hz, H-11), 2.69 (1H, dd, J = 8.0, 6.4 Hz, H-17a), 2.55 (1H, dd, J = 8.0, 6.4 Hz, H-17b), 2.15 (2H, t, J = 8.8 Hz, H-6), 2.06 (1H, m, H-34a), 2.05 (2H, m, H-22), 1.87 (1H, m, H-34b), 1.75 (1H, m, H-33a), 1.68 (2H, t, J = 4.0 Hz, H-27), 1.65 (3H, s, H-30), 1.65 (3H, s, H-20), 1.64 (3H, s, H-21), 1.62 (3H, s, H-37), 1.55 (3H, s, H-38), 1.52 (3H, s, H-31), 1.43 (3H, s, H-25), 1.33 (3H, s, H-26), 1.27 (3H, s, H-32), 1.24 (1H, m, H-33b), 1.09 (3H, d, J = 4.8 Hz, H-13), 0.92 (3H, d, J = 4.8 Hz, H-12), 0.75 (1H, m, H-7); ¹³C NMR (100 MHz, CD₃OD) δ_C: 77.4 (C-1), 206.9 (C-2), 97.5 (C-3), 210.0 (C-4), 46.7 (C-5), 35.2 (C-6), 46.7 (C-7), 45.2 (C-8), 110.0 (C-9), 210.0 (C-10), 44.3 (C-11), 21.0 (C-12), 19.6 (C-13), 24.2 (C-14), 117.1 (C-15), 137.8 (C-16), 26.0 (C-17), 18.0 (C-18), 31.4 (C-19), 69.1 (C-20), 80.5 (C-21), 28.0 (C-22), 25.9 (C-23), 28.9 (C-24), 123.9 (C-25), 133.9 (C-26), 26.1 (C-27), 18.2 (C-28), 13.8 (C-29), 39.0 (C-30), 26.6 (C-31), 125.9 (C-32), 132.0 (C-33), 26.1 (C-34), 17.8 (C-35)。以上数据与文献^[16]报道基本一致, 故鉴定化合物为hyphenrone T。

化合物8 无色油状物; HR-ESI-MS m/z 639.386 5, [M+Na]⁺ (计算值639.386 7, C₃₆H₄₉O₈Na); ¹H NMR (400 MHz, CDCl₃) δ_H: 5.08 (1H, t, J = 7.6 Hz, 1H-16), 4.89 (1H, t, J = 6.8 Hz, H-26), 3.79 (1H, t, J = 4.4 Hz, H-21), 3.26 (1H, overlapped, H-33), 3.18 (34-OCH₃), 2.97 (1H, sept, J = 6.4 Hz, H-11), 2.66 (1H, dd, J = 15.2, 7.2 Hz, H-15b), 2.56 (1H, dd, J = 15.2, 7.8 Hz, H-15a), 2.29 (2H, overlapped, H-6), 2.12 (2H, m, H-25), 2.08 (2H, dd, J = 14.8, 4.2 Hz, H-20), 1.67 (3H, s, H-18), 1.67 (3H, s, H-28), 1.64 (3H, s, H-19), 1.53 (3H, s, H-29), 1.43 (3H, s, H-23), 1.32 (3H, s, H-24), 1.26 (3H, s, H-30), 1.82 (2H, m, H-31), 1.22 (2H, m, H-32), 1.09 (3H, d, J = 6.4 Hz, H-13), 1.09 (3H, s, H-35), 1.05 (3H, s, H-36), 0.92 (3H, d, J = 6.4 Hz, H-12), 0.78 (1H, m, H-7); ¹³C NMR (100 MHz, CDCl₃) δ_C: 76.9 (C-1), 206.5 (C-2), 97.0 (C-3), 209.3 (C-4), 49.2 (C-5), 34.7 (C-6), 46.5 (C-7), 44.8 (C-8), 109.6 (C-9), 209.3 (C-10), 44.8 (C-11), 21.0 (C-12), 19.7 (C-13), 23.9 (C-14), 117.0 (C-15), 137.2 (C-16), 26.1 (C-17), 18.2 (C-18), 30.5 (C-19), 68.7 (C-20), 80.0 (C-21), 27.8 (C-22), 26.1 (C-23), 28.4 (C-24), 124.0 (C-25), 133.2 (C-26), 26.1 (C-27), 18.1 (C-28), 13.8 (C-29), 36.2 (C-30), 29.3 (C-31), 77.9 (C-32), 78.0 (C-33), 21.0 (C-34), 20.9 (C-35), 49.2 (34-OCH₃)。以上数据与文献^[17]报道基本一致, 故鉴定化合物为hyperforatone N。

3 抗炎活性筛选

实验选用的细胞株为原代小鼠腹腔巨噬细胞, 其可在细菌脂多糖LPS诱导下产生NO等炎性因子, 参与并介导炎症反应, 在多种炎症免疫过程初期与病理发展过程中均有较高的水平。通过检测原代培养的小鼠巨噬细胞NO生成量, 可作为体外初步观察和筛选有一定抗炎活性的组分或化合物的指标。取原代小鼠腹腔巨噬细胞接种于96孔板中, 加入待测化合物(10 μmol·L^-1) 和同等浓度的阳性对照药Dex预保护1 h; 然后, 加入1 μg·mL^-1的LPS于37 ℃、5% CO₂培养箱中培养24 h后, 收集上清液, 采用Griess法测定NO的含量(NO抑制率%), 同时用MTT法测定细胞增殖抑制率。该实验方案下, 表 2中的化合物1、3、5和7在10 μmol·L^-1对LPS诱导原代小鼠腹腔巨噬细胞NO生成具有显著的抑制活性, 结果表明具有潜在抗炎活性。

致谢中国医学科学院药物研究所药理学研究室林明宝老师提供平台并进行抗炎活性实验。

作者贡献: 马永慧负责实验、结构鉴定、论文撰写; 孙浩然负责进行分离实验和部分化合物结构鉴定; 吉腾飞负责研究选题、方案设计及论文修改; 李进负责方案设计及论文修改。

利益冲突: 本文不存在任何利益冲突。

基金

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新疆维吾尔自治区高校科研计划项目(XJEDU2019I022)
中国医学科学院医学与健康科技创新工程项目(2021-I2M-1-028)
北京高校卓越青年科学家计划项目(BJJWZYJH01201910023028)
青海省重点研发与转化计划-国际合作专项(2020-HZ-803)

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2022年第57卷第12期

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doi: 10.16438/j.0513-4870.2022-0654

接收时间：2022-05-27
首发时间：2025-12-24
出版时间：2022-12-12

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收稿日期：2022-05-27
修回日期：2022-06-10

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新疆维吾尔自治区高校科研计划项目(XJEDU2019I022)

中国医学科学院医学与健康科技创新工程项目(2021-I2M-1-028)

北京高校卓越青年科学家计划项目(BJJWZYJH01201910023028)

青海省重点研发与转化计划-国际合作专项(2020-HZ-803)

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1.新疆师范大学生命科学学院, 新疆特殊环境物种保护与调控生物学实验室, 干旱区植物逆境生物学实验室, 新疆乌鲁木齐 830054

2.中国医学科学院、北京协和医学院药物研究所, 天然药物活性物质与功能国家重点实验室, 北京 100050

3.中国科学院西北高原生物研究所, 青海省藏药研究重点实验室, 藏药研究重点实验室, 青海西宁 810001

通讯作者:

*李进, Tel: 86-991-4332325, E-mail: xjcjlj4@xjnu.edu.cn;

吉腾飞 , Tel: 86-10-63165226, E-mail: jitf@imm.ac.cn

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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