药学学报

C	δ_H (Hz)	δ_C
1	0.87-0.91 (m)	50.5
	1.91-1.94 (m)
2	3.72-3.76 (m)	69.9
3	3.28-3.30 (m)	78.7
4	-	44.9
5	1.29 (br. s)	49.0
6	4.38 (br. s)	68.6
7	1.52 (dd, J = 14.3, 2.5)	41.4
	1.80 (dd, J = 14.3, 3.6)
8	-	44.1
9	1.62-1.71 (m)	49.7
10	-	38.6
11	1.98-2.05 (m)	24.7
	2.08-2.13 (m)
12	5.32-5.33 (m)	127.8
13	-	138.5
14	-	40.3
15	1.14-1.17 (m)	29.4
	1.98-2.05 (m)
16	1.85-1.90 (m)	25.4
	2.27-2.30 (m)
17	-	49.6
18	2.30-2.32 (m)	56.6
19	2.42-2.45 (m)	38.7
20	-	154.3
21	2.21-2.24 (m)	33.4
	2.32-2.35 (m)
22	1.62-1.71 (m)	38.6
	1.91-1.94 (m)
23	3.43 (d, J = 11.2)	66.0
	3.58 (d, J = 11.2)
24	1.06 (s)	15.4
25	1.40 (s)	19.4
26	1.09 (s)	19.3
27	1.16 (s)	24.1
28	-	177.2
29	1.03 (d, J = 6.5)	16.9
30	4.69 (br, s), 4.64 (br, s)	105.5
1'	5.34 (d, J = 8.2)	96.0
2'	3.31-3.32 (m)	74.0
3'	3.38-3.41 (m)	78.3
4'	3.35-3.36 (m)	71.4
5'	3.32-3.33 (m)	78.4
6'	3.79 (dd, J = 12.0, 2.9)	62.6
	3.67 (dd, J = 12.0, 4.7)

δ_H (Hz)

δ_C

0.87-0.91 (m)

50.5

1.91-1.94 (m)

3.72-3.76 (m)

69.9

3.28-3.30 (m)

78.7

44.9

1.29 (br. s)

49.0

4.38 (br. s)

68.6

1.52 (dd, J = 14.3, 2.5)

41.4

1.80 (dd, J = 14.3, 3.6)

44.1

1.62-1.71 (m)

49.7

38.6

1.98-2.05 (m)

24.7

2.08-2.13 (m)

5.32-5.33 (m)

127.8

138.5

40.3

1.14-1.17 (m)

29.4

1.98-2.05 (m)

1.85-1.90 (m)

25.4

2.27-2.30 (m)

49.6

2.30-2.32 (m)

56.6

2.42-2.45 (m)

38.7

154.3

2.21-2.24 (m)

33.4

2.32-2.35 (m)

1.62-1.71 (m)

38.6

1.91-1.94 (m)

3.43 (d, J = 11.2)

66.0

3.58 (d, J = 11.2)

1.06 (s)

15.4

1.40 (s)

19.4

1.09 (s)

19.3

1.16 (s)

24.1

177.2

1.03 (d, J = 6.5)

16.9

4.69 (br, s), 4.64 (br, s)

105.5

5.34 (d, J = 8.2)

96.0

3.31-3.32 (m)

74.0

3.38-3.41 (m)

78.3

3.35-3.36 (m)

71.4

3.32-3.33 (m)

78.4

3.79 (dd, J = 12.0, 2.9)

62.6

3.67 (dd, J = 12.0, 4.7)

C	δ_H (Hz)	δ_C
1	0.87-0.91 (m)	50.5
	1.91-1.94 (m)
2	3.72-3.76 (m)	69.9
3	3.28-3.30 (m)	78.7
4	-	44.9
5	1.29 (br. s)	49.0
6	4.38 (br. s)	68.6
7	1.52 (dd, J = 14.3, 2.5)	41.4
	1.80 (dd, J = 14.3, 3.6)
8	-	44.1
9	1.62-1.71 (m)	49.7
10	-	38.6
11	1.98-2.05 (m)	24.7
	2.08-2.13 (m)
12	5.32-5.33 (m)	127.8
13	-	138.5
14	-	40.3
15	1.14-1.17 (m)	29.4
	1.98-2.05 (m)
16	1.85-1.90 (m)	25.4
	2.27-2.30 (m)
17	-	49.6
18	2.30-2.32 (m)	56.6
19	2.42-2.45 (m)	38.7
20	-	154.3
21	2.21-2.24 (m)	33.4
	2.32-2.35 (m)
22	1.62-1.71 (m)	38.6
	1.91-1.94 (m)
23	3.43 (d, J = 11.2)	66.0
	3.58 (d, J = 11.2)
24	1.06 (s)	15.4
25	1.40 (s)	19.4
26	1.09 (s)	19.3
27	1.16 (s)	24.1
28	-	177.2
29	1.03 (d, J = 6.5)	16.9
30	4.69 (br, s), 4.64 (br, s)	105.5
1'	5.34 (d, J = 8.2)	96.0
2'	3.31-3.32 (m)	74.0
3'	3.38-3.41 (m)	78.3
4'	3.35-3.36 (m)	71.4
5'	3.32-3.33 (m)	78.4
6'	3.79 (dd, J = 12.0, 2.9)	62.6
	3.67 (dd, J = 12.0, 4.7)

δ_H (Hz)

δ_C

0.87-0.91 (m)

50.5

1.91-1.94 (m)

3.72-3.76 (m)

69.9

3.28-3.30 (m)

78.7

44.9

1.29 (br. s)

49.0

4.38 (br. s)

68.6

1.52 (dd, J = 14.3, 2.5)

41.4

1.80 (dd, J = 14.3, 3.6)

44.1

1.62-1.71 (m)

49.7

38.6

1.98-2.05 (m)

24.7

2.08-2.13 (m)

5.32-5.33 (m)

127.8

138.5

40.3

1.14-1.17 (m)

29.4

1.98-2.05 (m)

1.85-1.90 (m)

25.4

2.27-2.30 (m)

49.6

2.30-2.32 (m)

56.6

2.42-2.45 (m)

38.7

154.3

2.21-2.24 (m)

33.4

2.32-2.35 (m)

1.62-1.71 (m)

38.6

1.91-1.94 (m)

3.43 (d, J = 11.2)

66.0

3.58 (d, J = 11.2)

1.06 (s)

15.4

1.40 (s)

19.4

1.09 (s)

19.3

1.16 (s)

24.1

177.2

1.03 (d, J = 6.5)

16.9

4.69 (br, s), 4.64 (br, s)

105.5

5.34 (d, J = 8.2)

96.0

3.31-3.32 (m)

74.0

3.38-3.41 (m)

78.3

3.35-3.36 (m)

71.4

3.32-3.33 (m)

78.4

3.79 (dd, J = 12.0, 2.9)

62.6

3.67 (dd, J = 12.0, 4.7)

No.	2	3	4	5	6	7
1	51.0	51.0	48.1	50.5	50.3	48.4
2	69.9	69.9	69.9	69.8	69.9	69.9
3	78.3	78.3	78.4	78.3	78.4	78.4
4	44.9	44.9	44.3	44.9	45.0	44.3
5	48.7	48.7	48.7	49.6	49.1	48.0
6	68.5	19.0	19.4	68.6	68.9	19.4
7	42.6	35.3	33.7	41.4	40.6	33.7
8	39.5	40.5	41.4	40.6	41.5	41.2
9	49.7	48.6	48.3	48.8	49.1	48.7
10	38.6	39.1	39.1	38.6	38.4	39.3
11	24.5	24.5	24.9	25.4	24.9	24.9
12	128.0	127.7	129.6	127.5	130.0	129.8
13	139.3	139.8	139.9	138.7	139.2	140.4
14	46.3	46.0	42.9	44.0	43.4	43.2
15	29.9	29.8	29.8	29.4	29.8	29.8
16	36.0	36.0	27.4	24.7	26.7	26.8
17	50.8	49.7	49.5	49.4	49.6	49.7
18	134.3	134.2	55.1	54.4	55.1	55.3
19	137.8	137.8	73.8	40.4	73.8	73.8
20	35.8	35.8	42.9	40.3	43.1	42.9
21	27.6	27.6	26.6	31.9	27.3	27.2
22	31.8	31.7	38.5	37.6	38.6	39.2
23	66.0	66.4	66.5	66.0	66.1	66.5
24	15.4	14.2	14.0	15.4	15.4	14.0
25	19.9	19.1	17.8	19.4	19.2	17.8
26	19.9	18.9	17.8	19.4	18.9	17.7
27	20.6	19.9	24.9	24.2	24.9	25.0
28	176.8	176.7	178.7	178.1	178.6	180.9
29	22.6	22.5	27.2	17.8	27.2	27.5
30	19.1	18.5	16.8	21.7	16.8	16.8
1'	95.9	95.9	95.9	95.9	96.0
2'	74.2	74.2	74.0	74.1	74.0
3'	78.8	78.8	78.7	78.7	78.4
4'	71.4	71.3	71.2	71.4	71.3
5'	78.5	78.5	78.4	78.4	78.3
6'	62.7	62.7	62.6	62.1	62.5

No.

51.0

48.1

50.5

50.3

48.4

69.9

69.8

69.9

78.3

78.4

78.3

78.4

44.9

44.3

44.9

45.0

44.3

48.7

49.6

49.1

48.0

68.5

19.0

19.4

68.6

68.9

19.4

42.6

35.3

33.7

41.4

40.6

33.7

39.5

40.5

41.4

40.6

41.5

41.2

49.7

48.6

48.3

48.8

49.1

48.7

38.6

39.1

38.6

38.4

39.3

24.5

24.9

25.4

24.9

128.0

127.7

129.6

127.5

130.0

129.8

139.3

139.8

139.9

138.7

139.2

140.4

46.3

46.0

42.9

44.0

43.4

43.2

29.9

29.8

29.4

29.8

36.0

27.4

24.7

26.7

26.8

50.8

49.7

49.5

49.4

49.6

49.7

134.3

134.2

55.1

54.4

55.1

55.3

137.8

73.8

40.4

73.8

35.8

42.9

40.3

43.1

42.9

27.6

26.6

31.9

27.3

27.2

31.8

31.7

38.5

37.6

38.6

39.2

66.0

66.4

66.5

66.0

66.1

66.5

15.4

14.2

14.0

15.4

14.0

19.9

19.1

17.8

19.4

19.2

17.8

19.9

18.9

17.8

19.4

18.9

17.7

20.6

19.9

24.9

24.2

24.9

25.0

176.8

176.7

178.7

178.1

178.6

180.9

22.6

22.5

27.2

17.8

27.2

27.5

19.1

18.5

16.8

21.7

16.8

95.9

96.0

74.2

74.0

74.1

74.0

78.8

78.7

78.4

71.4

71.3

71.2

71.4

71.3

78.5

78.4

78.3

62.7

62.6

62.1

62.5

No.	2	3	4	5	6	7
1	51.0	51.0	48.1	50.5	50.3	48.4
2	69.9	69.9	69.9	69.8	69.9	69.9
3	78.3	78.3	78.4	78.3	78.4	78.4
4	44.9	44.9	44.3	44.9	45.0	44.3
5	48.7	48.7	48.7	49.6	49.1	48.0
6	68.5	19.0	19.4	68.6	68.9	19.4
7	42.6	35.3	33.7	41.4	40.6	33.7
8	39.5	40.5	41.4	40.6	41.5	41.2
9	49.7	48.6	48.3	48.8	49.1	48.7
10	38.6	39.1	39.1	38.6	38.4	39.3
11	24.5	24.5	24.9	25.4	24.9	24.9
12	128.0	127.7	129.6	127.5	130.0	129.8
13	139.3	139.8	139.9	138.7	139.2	140.4
14	46.3	46.0	42.9	44.0	43.4	43.2
15	29.9	29.8	29.8	29.4	29.8	29.8
16	36.0	36.0	27.4	24.7	26.7	26.8
17	50.8	49.7	49.5	49.4	49.6	49.7
18	134.3	134.2	55.1	54.4	55.1	55.3
19	137.8	137.8	73.8	40.4	73.8	73.8
20	35.8	35.8	42.9	40.3	43.1	42.9
21	27.6	27.6	26.6	31.9	27.3	27.2
22	31.8	31.7	38.5	37.6	38.6	39.2
23	66.0	66.4	66.5	66.0	66.1	66.5
24	15.4	14.2	14.0	15.4	15.4	14.0
25	19.9	19.1	17.8	19.4	19.2	17.8
26	19.9	18.9	17.8	19.4	18.9	17.7
27	20.6	19.9	24.9	24.2	24.9	25.0
28	176.8	176.7	178.7	178.1	178.6	180.9
29	22.6	22.5	27.2	17.8	27.2	27.5
30	19.1	18.5	16.8	21.7	16.8	16.8
1'	95.9	95.9	95.9	95.9	96.0
2'	74.2	74.2	74.0	74.1	74.0
3'	78.8	78.8	78.7	78.7	78.4
4'	71.4	71.3	71.2	71.4	71.3
5'	78.5	78.5	78.4	78.4	78.3
6'	62.7	62.7	62.6	62.1	62.5

No.

51.0

48.1

50.5

50.3

48.4

69.9

69.8

69.9

78.3

78.4

78.3

78.4

44.9

44.3

44.9

45.0

44.3

48.7

49.6

49.1

48.0

68.5

19.0

19.4

68.6

68.9

19.4

42.6

35.3

33.7

41.4

40.6

33.7

39.5

40.5

41.4

40.6

41.5

41.2

49.7

48.6

48.3

48.8

49.1

48.7

38.6

39.1

38.6

38.4

39.3

24.5

24.9

25.4

24.9

128.0

127.7

129.6

127.5

130.0

129.8

139.3

139.8

139.9

138.7

139.2

140.4

46.3

46.0

42.9

44.0

43.4

43.2

29.9

29.8

29.4

29.8

36.0

27.4

24.7

26.7

26.8

50.8

49.7

49.5

49.4

49.6

49.7

134.3

134.2

55.1

54.4

55.1

55.3

137.8

73.8

40.4

73.8

35.8

42.9

40.3

43.1

42.9

27.6

26.6

31.9

27.3

27.2

31.8

31.7

38.5

37.6

38.6

39.2

66.0

66.4

66.5

66.0

66.1

66.5

15.4

14.2

14.0

15.4

14.0

19.9

19.1

17.8

19.4

19.2

17.8

19.9

18.9

17.8

19.4

18.9

17.7

20.6

19.9

24.9

24.2

24.9

25.0

176.8

176.7

178.7

178.1

178.6

180.9

22.6

22.5

27.2

17.8

27.2

27.5

19.1

18.5

16.8

21.7

16.8

95.9

96.0

74.2

74.0

74.1

74.0

78.8

78.7

78.4

71.4

71.3

71.2

71.4

71.3

78.5

78.4

78.3

62.7

62.6

62.1

62.5

番石榴根中一个新的三萜类成分

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彭财英 , 黄应正 , 刘建群 , 黄慧莲 , 舒积成 ^*

药学学报 | 研究论文 2017,52(11): 1731-1736

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药学学报 | 研究论文 2017, 52(11): 1731-1736

番石榴根中一个新的三萜类成分

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彭财英, 黄应正, 刘建群, 黄慧莲, 舒积成^*

作者信息

江西中医药大学现代中药制剂教育部重点实验室, 江西南昌 330004

通讯作者:

* 舒积成, Tel: 86-791-87118658, E-mail: shujc210@163.com

A new triterpenoid from roots of Psidium guajava

Cai-ying PENG, Ying-zheng HUANG, Jian-qun LIU, Hui-lian HUANG, Ji-cheng SHU^*

Affiliations

Key Laboratory of Modern Preparation of Traditional Chinese Medicines, Ministry of Education, Jiangxi University of Traditional Chinese Medicines, Nanchang 330004, China

出版时间: 2017-11-12 doi: 10.16438/j.0513-4870.2017-0565

文章导航

摘要

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为了研究番石榴Psidium guajava根的化学成分，采用柱色谱技术分离纯化，通过波谱数据鉴定了7个乌苏烷型三萜类化合物：2α，3β，6β，23-四羟基乌苏酸-12，20（30）-双烯-28-O-β-D-葡萄糖苷（1）、2α，3β，6β，23-四羟基乌苏酸-12，18-双烯-28-O-β-D-葡萄糖苷（2）、2α，3β，23-三羟基乌苏酸-12，18-双烯-28-O-β-D-葡萄糖苷（3）、nigaichigoside F1（4）、积雪草苷C（5）、2α，3β，6β，19α，23-五羟基乌苏酸-12，18-双烯-28-O-β-D-葡萄糖苷（6）和2α，3β，19α，23-四羟基乌苏酸（7）。化合物1为新化合物，化合物2~6均为首次在此植物中分离得到。对新化合物进行了体外细胞毒活性测定，化合物1浓度为25 μmol·L^-1对人肝瘤细胞Bel 7402抑制率为52.5%，显示具有较好的体外抑制肿瘤细胞活性。

关键词

番石榴 / 化学成分 / 乌苏烷型 / 三萜 / 细胞毒

Abstract

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To investigated the chemical constituents of the roots of Psidium guajava, we isolated seven compounds by silica gel column chromatography.These include oleanane derivatives, 2 α, 3β, 6β, 23-tetrahydroxylurs-12, 20(30)-dien-28-oic acid β-D-glucopyranoside (1), 2α, 3β, 6β, 23-tetrahydroxylurs-12, 18-dien-28-oic acid β-D-glucopyranoside (2), 2α, 3β, 23-trihydroxylurs-12, 18-dien-28-oic acid β-D-glucopyranoside (3), nigaichigoside F1(4), asiaticoside C (5), 2α, 3β, 6β, 19α, 23-pentahydroxylurs-12, 18-dien-28-oic acid β-Dglucopyranoside (6) and 2α, 3β, 19α, 23-tetrahydroxylurs-12-en-28-oic acid (7).Their structures were elucidated on the basis of spectral analysis with reference to the published data.Compound 1 is brand new, compounds 2-6 were first isolated from this plant.The new compound was evaluated for their cytotoxic activity against human hepatoma Bel 7402 in vitro.The results were expressed as the ratio of inhibiting Bel 7402 cells growth by comparing to untreated cells.The new compound (concentration:25 μmol·L^-1) showed the ratio values of 52.5%.

Key words

Psidium guajava / chemical constituent / ursane / triterpenoid / cytotoxic activity

引用本文

彭财英, 黄应正, 刘建群, 黄慧莲, 舒积成. 番石榴根中一个新的三萜类成分. 药学学报, 2017 , 52 (11) : 1731 -1736 . DOI: 10.16438/j.0513-4870.2017-0565

Cai-ying PENG, Ying-zheng HUANG, Jian-qun LIU, Hui-lian HUANG, Ji-cheng SHU. A new triterpenoid from roots of Psidium guajava[J]. Acta Pharmaceutica Sinica, 2017 , 52 (11) : 1731 -1736 . DOI: 10.16438/j.0513-4870.2017-0565

正文

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番石榴Psidium guajava L.为桃金娘科Myrtaceae番石榴属Psidium植物。因其为重要的粮食作物和具有良好的药用价值^[1], 故在热带和亚热带地区均有种植。此植物不同部位具有不同的药物用途, 如根和不成熟果实可治疗胃肠炎和腹泻^[2]; 成熟果实可通便和除口臭^[3]; 叶水提取液可作退热剂和解痉剂^[4]; 树皮具止血功效^[5]; 花蕊具有止咳作用等^[6]; 种子具有较好抗肿瘤作用^[7]。近年来, 国内外学者主要对番石榴叶及果实的化学成分进行了较深入的研究, 并报道了大量的黄酮、萜类及有机酸类等成分^[8-10], 而有关其根部位的化学成分研究较少。作为从番石榴植物各个部位(包括果实、叶、皮、花及根)分离活性成分研究的一部分, 作者对番石榴根进行了系统的化学成分研究。此次报道从乙醇提取物的乙酸乙酯部分分离到7个乌苏烷型三萜化合物, 包括1个新化合物、5个首次在此植物中分离得到的化合物, 结构如图 1。并对新化合物进行了体外细胞毒活性测定, 结果显示较好的体外抑制人肝瘤细胞Bel 7402活性。

化合物1 白色无定形粉末(甲醇), [α]_D²⁵ -35.8 (c 0.026, MeOH); HR-ESI-MS准分子离子峰m/z: 665.389 6 [M+H]⁺ (计算值665.390 1, C₃₆H₅₆O₁₁); ¹H NMR和¹³C NMR数据见表 1。

化合物1 10%硫酸乙醇显色为粉色, Liebermann-Burchard阳性反应, 提示化合物为三萜类化合物。HR-ESI-MS给出准分子离子峰m/z: 665.389 6 [M+ H]⁺ (计算值665.390 1, C₃₆H₅₆O₁₁), 给出分子式为C₃₆H₅₆O₁₁。¹H NMR (表 1)显示, 4个单峰甲基信号: δ_H 1.06 (s, 3H, H-24)、1.40 (s, 3H, H-25)、1.09 (s, 3H, H-26)和1.16 (s, 3H, H-27); 1个双峰甲基信号: δ_H 1.03 (d, 3H, J = 6.5 Hz, H-29); 3个烯质子信号, 其中包括一对环外双键质子: δ_H 5.32~5.33 (m, 1H, H-12)、4.64 (br.s, 1H, H-30)和4.69 (br.s, 1H, H-30)。¹³C NMR (表 1)显示, 两组双键碳信号, 其中包括一对末端双键: δ_C 127.8 (C-12)、138.5 (C-13)、105.5 (C-20)和154.3 (C-30)。以上信息提示化合物1母核结构为具末端双键的12-烯-28-酸-乌苏烷型三萜^{[11, 12]}。

对¹H NMR和¹³C NMR分析, 化合物1具有一分子葡萄糖的结构单元^[13]:糖端基质子信号δ_H 5.34 (d, 1H, J = 8.2 Hz, H-1'); 6个连氧碳信号δ_C 96.0、78.4、78.3、74.0、71.4和62.6。根据葡萄糖端基质子的偶合常数为8.2 Hz, 提示所连接的糖单元为β-葡萄糖。化合物1经酸水解, 气相色谱检测, 保留时间与β-D-葡萄糖对照品结果一致, 提示化合物所连接的糖为β-D-葡萄糖^[14]。另外, 在¹H NMR和¹³C NMR谱中, 除上述葡萄糖质子和葡萄糖碳信号外, ¹H NMR还显示5个连氧质子: δ_H 3.72~3.76 (m, 1H, H-2)、3.29~3.40 (m, 1H, H-3)、4.38 (br.s, 1H, H-6)、3.43 (d, 1H, J = 11.2 Hz, H-23)和3.58 (d, J = 11.2 Hz, H-23);结合HSQC谱分析, ¹³C NMR还显示4个连氧碳信号: δ_C 69.9 (C-2)、78.7 (C-3)、68.6 (C-6)和66.0 (C-23)。综合上述分析, 提示化合物1是具四羟基取代的乌苏烷型三萜苷。

HMBC谱中(图 2), H-1 (δ_H 1.91~1.94)和H-3 (δ_H 3.29~3.40)与C-2 (δ_C 69.9)具有远程相关, 提示C-2位有OH取代; H-23 (δ_H 3.43, 3.58)与CH₃-24 (δ_H 1.06)和C-3 (δ_C 78.7)具有远程偶合, 提示C-3位有OH取代, 同时提示C-23位有OH取代; H-6 (δ_H 4.38)与C-5 (δ_C 49.0)、C-8 (δ_C 44.1)和C-10 (δ_C 38.6)有远程相关, 提示C-6位有OH取代; 两个末端双键质子δ_H 4.64 (br.s, 1H, H-30)和4.69 (br.s, 1H, H-30)均与C-19和C-21有远程偶合, 提示此双键在C-20和C-30位上; 糖端基质子H-1' (δ_H 5.34)与C-28 (δ_C 175.8)具有远程偶合, 提示葡萄糖连接在C-28位。NOESY谱中, H-2显示与H-24和H-25相关; H-3与H-5、H-6和H-23具有相关, 提示化合物的羟基取代的相对构型为2α-OH、3β-OH、6β-OH和23α。综上分析, 化合物1的结构鉴定为2α, 3β, 6β, 23-四羟基乌苏酸-12, 20(30)-双烯-28-O-β-D-葡萄糖苷。经Scifinder检索, 化合物1为新化合物。

实验部分

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AB Sciex 5600质谱仪; Bruker AVANCE Ⅲ HD 600 MHz型核磁共振波谱仪(瑞士布鲁克公司), TMS为内标; CD₃OD为溶剂。Finnigan Trace DSQ气质联用色谱仪(美国Thermo公司); Perkin Elmer 343型旋光测定仪(美国Perkin Elmer公司); Aglient 1100制备液相(美国安捷伦公司), YMC-Actus ODS-AC18 (250 mm × 10 mm, 5 µm)半制备柱(日本YMC公司)。柱色谱硅胶(200~300目), 薄层色谱硅胶(青岛海洋化工), 凝胶Sephadex LH-20 (日本三菱株式会社), 显色剂为5%浓硫酸乙醇溶液。常规分析纯化学试剂购自国药集团化学试剂有限公司。

番石榴根于2014年7月中旬采自广东省珠海市, 由江西中医药大学付小梅副教授鉴定, 标本(No. 201407003)现存于江西中医药大学现代中药制剂教育部重点实验室。

1 提取与分离

自然干燥的番石榴根(10.2 kg)粉碎, 经90%乙醇加热回流3次, 每次1 h, 提取液减压回收得流浸膏980 g, 所得流浸膏用适量的水分散, 然后依次用石油醚、乙酸乙酯萃取, 剩余部分旋干溶剂后, 用甲醇溶解。回收溶剂得石油醚部分86.0 g、乙酸乙酯部分146.2 g、甲醇部分596.5 g。

取乙酸乙酯萃取部位(145 g)进行硅胶柱色谱分离, 以二氯甲烷-甲醇(100:0, 50:1, 25:1, 10:1, 5:1, 1:1, 0:100)梯度洗脱得到Fr.1~9。Fr.3 (13.5 g)经硅胶柱(二氯甲烷-甲醇)分离, 得Fr.3-1~3-48。合并Fr.3-16~3-21, 再经Sephadex LH-20柱色谱(MeOH)分离, 得化合物7 (15 mg)。Fr.4 (21.9 g)经硅胶柱(二氯甲烷-甲醇)分离, 得Fr.4-1~4-86。合并Fr.4-18~3-23, 再经Sephadex LH-20柱(MeOH)分离, 得化合物3 (9 mg)。Fr.5 (17.9 g)经硅胶柱(二氯甲烷-甲醇)分离, 得Fr.5-1~5-52。合并Fr.2-15~2-20, 再经半制备柱色谱(甲醇-水=30:70, 3 mL·min^-1), 得到化合物1 (7 mg, t_R= 10.5 min)和化合物2 (8 mg, t_R= 11.7 min)。Fr.6 (25.8 g)经硅胶柱(二氯甲烷-甲醇)分离, 得Fr.6-1~6-71。合并Fr.6-16~6-24, 再经半制备柱色谱(甲醇-水=30:70, 3 mL·min^-1), 得到化合物4 (5 mg, t_R= 11.2 min)、化合物5 (8 mg, t_R= 12.6 min)和6 (9 mg, t_R= 13.8 min)。

2 结构鉴定

化合物1 白色无定形粉末(甲醇), [α]_D²⁵ -35.8 (c 0.026, MeOH); HR-ESI-MS给出准分子离子峰m/z: 665.389 6 [M+H]⁺ (计算值665.390 1, C₃₆H₅₆O₁₁); ¹H NMR和¹³C NMR数据见表 1。

化合物2 白色粉末(甲醇); ESI-MS m/z: 687.3 [M+Na]⁺, 结合核磁共振C、H谱, 提示分子式C₃₆H₅₆O₁₁。¹H NMR (600 MHz, CD₃OD) δ_H: 5.44 (1H, dd, J = 5.5, 2.3 Hz, H-12), 5.40 (1H, d, J = 8.2 Hz, glc-H-1), 4.37 (1H, s, H-6), 3.81 (1H, dd, J = 12.0, 1.8 Hz, glc-H-6a), 3.76 (1H, ddd, J = 11.4, 9.6, 4.6 Hz, H-2), 3.68 (1H, dd, J = 12.0, 4.2 Hz, glc-H-6b), 3.59 (1H, d, J = 11.2 Hz, H-23a), 3.44 (1H, d, J = 11.2 Hz, H-23b), 3.27~3.42 (4H, m, H-3, glc-H-2, 3, 4, 5), 1.76 (s, H-25), 1.43 (3H, s, H-29), 1.16 (3H, s, H-24), 1.11 (3H, d, J = 7.0 Hz, H-30), 1.06 (3H, s, H-26), 0.98 (3H, s, H-27)。¹³C NMR (150 MHz, CD₃OD) δ_C见表 2。以上¹H NMR、¹³C NMR数据与文献^[15]一致, 故鉴定化合物2为2α, 3β, 6β, 23-四羟基乌苏酸-12, 18-双烯-28-O-β-D-葡萄糖苷。

化合物3 白色粉末(甲醇); ESI-MS m/z: 649.3 [M+H]⁺, 结合核磁共振C、H谱, 提示分子式C₃₆H₅₆O₁₀。¹H NMR (600 MHz, CD₃OD) δ_H: 5.39~5.42 (1H, m, H-12), 5.39 (1H, d, J = 8.0 Hz, glc-H-1), 3.80 (1H, dd, J = 12.1, 1.9 Hz, glc-H-6a), 3.70~3.74 (1H, m, H-2), 3.67 (1H, dd, J = 12.1, 4.5 Hz, glc-H-6b), 3.50 (1H, d, J = 11.1, H-23a), 3.26~3.40 (6H, m, H-23b, H-3, glc-H-2, 3, 4, 5), 1.75 (3H, s, H-25), 1.10 (3H, d, J = 7.1 Hz, H-30), 1.09 (3H, s, H-29), 1.01 (3H, s, H-24), 0.93 (3H, s, H-26), 0.70 (3H, s, H-27)。¹³C NMR (150 MHz, CD₃OD) δ_C见表 2。以上¹H NMR、¹³C NMR数据与文献^[16]一致, 故鉴定化合物3为2α, 3β, 23-三羟基乌苏酸-12, 18-双烯-28-O-β-D-葡萄糖苷。

化合物4 白色粉末(甲醇); ESI-MS m/z: 689.3 [M+Na]⁺, 结合核磁共振C、H谱, 提示分子式C₃₆H₅₈O₁₁。¹H NMR (600 MHz, CD₃OD) δ_H: 5.31~5.34 (1H, m, H-12), 5.32 (1H, d, J = 8.4 Hz, glc-H-1), 3.80 (1H, dd, J = 12.0, 2.2 Hz, glc-6a), 3.68~3.72 (1H, m, H-2), 3.68 (1H, dd, J = 12.0, 4.7 Hz, glc-6b), 3.50 (1H, d, J = 11.0 Hz, H-23a), 3.35~3.42 (5H, m, H-3, glc-H-2, 3, 4, 5), 3.27 (1H, d, J = 11.0 Hz, H-23b), 1.34 (3H, s, H-27), 1.21 (3H, s, H-29), 1.04 (3H, s, H-25), 0.93 (3H, d, J = 6.5 Hz, H-30), 0.78 (3H, s, H-26), 0.70 (3H, s, H-24)。¹³C NMR (150 MHz, CD₃OD) δ_C见表 2。以上¹H NMR、¹³C NMR数据与文献^[17]一致, 故鉴定化合物4为nigaichigoside F1。

化合物5 白色粉末(甲醇); ESI-MS m/z: 689.3 [M+Na]⁺, 结合核磁共振C、H谱, 提示分子式C₃₆H₅₈O₁₁。¹H NMR (600 MHz, CD₃OD) δ_H: 5.34 (1H, d, J = 8.2 Hz, glc-H-1), 5.30 (1H, m, H-12), 4.37 (1H, s, H-6), 3.79 (1H, dd, J = 12.0, 2.0 Hz, glc-H-6a), 3.73~3.74 (1H, m, H-2), 3.68 (1H, d, J = 12.0, 4.5 Hz, H-glc-H-6b), 3.58 (1H, d, J = 11.4 Hz, H-23a), 3.43 (1H, d, J = 11.4 Hz, H-23b), 3.28~3.41 (5H, m, H-3, glc-H-2, 3, 4, 5), 1.40 (3H, s, H-25), 1.10 (3H, s, H-24), 1.09 (3H, s, H-26), 1.06 (3H, s, H-27), 0.96 (3H, s, H-29), 0.90 (3H, d, J = 6.5 Hz, H-30)。¹³C NMR (150 MHz, CD₃OD) δ_C见表 2。以上¹H NMR、¹³C NMR数据与文献^[18]一致, 故鉴定化合物5为积雪草苷C。

化合物6 白色粉末(甲醇); ESI-MS m/z: 705.4 [M+Na]⁺, 结合核磁共振C、H谱, 提示分子式C₃₆H₅₈O₁₂。¹H NMR (600 MHz, CD₃OD) δ_H: 5.36 (1H, t, J = 3.0 Hz, H-12), 5.31 (1H, d, J = 8.2 Hz, glc-H-1), 4.39 (1H, s, H-6), 3.80 (1H, dd, J = 12.0, 2.1 Hz, glc-H-6a), 3.73~3.77 (1H, m, H-2), 3.68 (1H, d, J = 12.0, 4.6 Hz, H-glc-H-6b), 3.59 (1H, d, J = 11.2 Hz, H-23a), 3.42 (1H, d, J = 11.2 Hz, H-23b), 3.23~3.39 (5H, m, H-3, glc-H-2, 3, 4, 5), 1.40 (3H, s, H-25), 1.33 (3H, s, H-27), 1.22 (3H, s, H-29), 1.07 (3H, s, H-24), 1.05 (3H, s, H-26), 0.94 (3H, d, J = 6.6 Hz, H-30)。¹³C NMR (150 MHz, CD₃OD) δ_C见表 2。以上¹H NMR、¹³C NMR数据与文献^[19]一致, 故鉴定化合物6为2α, 3β, 6β, 19α, 23-五羟基乌苏酸-12, 18-双烯-28-O-β-D-葡萄糖。

化合物7 白色粉末(甲醇); ESI-MS m/z: 511.3 [M+Na]⁺, 结合核磁共振C、H谱, 提示分子式C₃₀H₄₈O₅。¹H NMR (600 MHz, CD₃OD) δ_H: 5.30 (1H, like s, H-12), 3.70 (1H, m), 3.50 (1H, d, J = 11.2 Hz, H-23a), 3.36 (1H, m), 3.28 (1H, d, J = 11.2 Hz, H-23b), 2.52 (1H, s, H-18), 1.35 (3H, s), 1.20 (3H, s), 1.04 (3H, s), 0.93 (3H, d, J = 6.6 Hz, H-30), 0.81 (3H, s), 0.71 (3H, s)。¹³C NMR (150 MHz, CD₃OD) δ_C见表 2。以上¹H NMR、¹³C NMR数据与文献^[20]一致, 故鉴定化合物7为2α, 3β, 19α, 23-四羟基乌苏酸(2α, 3β, 19α, 23-tetrahydroxylurs-12-en-28-oic acid)。

3 化合物1酸水解

参照文献^[14], 取2 mg化合物1溶解于5 mL 3 mol·L^-1三氯乙酸中, 120 ℃加热2 h后减压浓缩至干, 干燥过夜。依次加入20 μL (2S)-1-氨基-2-丙醇-甲醇(1:8)混合液, 17 μL乙酸-甲醇(1:4)及13 μL 3%氰基硼氢化钠甲醇溶液。上述混合液放置65 ℃水浴加热2 h。冷却后缓慢加入3 mol·L^-1三氟乙酸至pH为1, 减压浓缩, 残余物真空干燥过夜, 再加入吡啶和乙酸酐各0.2 mL, 置100 ℃水浴加热1 h。冷却后, 加适量水, 用三氯甲烷萃取, 用0.5 mol·L^-1 Na₂CO₃和水各洗滤3次。三氯甲烷层加入无水硫酸钠干燥后过滤, 注入气质联用色谱仪。

4 化合物1抗肿瘤活性测定

采用MTT法, 参照文献^[21], 将处于对数生长期Bel 7402细胞按每孔100 μL接种于96孔培养板内, 每孔7.5×10³细胞, 培养24 h后, 加入以培养基稀释的化合物1工作液, 终浓度为25 μmol·L^-1。实验中每个浓度均为3个复孔, 另设空白对照孔。细胞在37 ℃、5% CO₂条件下分别孵育48 h后, 去除培养基, PBS洗1次后, 每孔加入新鲜培养基100 μL和MTT (5 mg·mL^-1, 用生理盐水配制) 20 μL; 继续培养4 h后, 去除孔中的培养基和MTT, 每孔加入二甲基亚砜(DMSO) 100 μL, 用酶标仪测OD₅₇₀_-₆₃₀值。样品对Bel 7402的抑制率按下述公式计算:抑制率(%) = [(OD₀-OD_S) / OD₀] × 100%, OD₀为对照组吸收值, OD_S为测试样品吸收值。

基金

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国家自然科学基金资助项目(81360630)
江西省青年科学家培养项目(20142BCB23023)
江西省自然科学基金资助项目(20161BAB205219)

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doi: 10.16438/j.0513-4870.2017-0565

接收时间：2017-06-13
首发时间：2026-01-14
出版时间：2017-11-12

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收稿日期：2017-06-13
修回日期：2017-07-04

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国家自然科学基金资助项目(81360630)

江西省青年科学家培养项目(20142BCB23023)

江西省自然科学基金资助项目(20161BAB205219)

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江西中医药大学现代中药制剂教育部重点实验室, 江西南昌 330004

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* 舒积成, Tel: 86-791-87118658, E-mail: shujc210@163.com

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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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C	δ_H (Hz)	δ_C
1	0.87-0.91 (m)	50.5
	1.91-1.94 (m)
2	3.72-3.76 (m)	69.9
3	3.28-3.30 (m)	78.7
4	-	44.9
5	1.29 (br. s)	49.0
6	4.38 (br. s)	68.6
7	1.52 (dd, J = 14.3, 2.5)	41.4
	1.80 (dd, J = 14.3, 3.6)
8	-	44.1
9	1.62-1.71 (m)	49.7
10	-	38.6
11	1.98-2.05 (m)	24.7
	2.08-2.13 (m)
12	5.32-5.33 (m)	127.8
13	-	138.5
14	-	40.3
15	1.14-1.17 (m)	29.4
	1.98-2.05 (m)
16	1.85-1.90 (m)	25.4
	2.27-2.30 (m)
17	-	49.6
18	2.30-2.32 (m)	56.6
19	2.42-2.45 (m)	38.7
20	-	154.3
21	2.21-2.24 (m)	33.4
	2.32-2.35 (m)
22	1.62-1.71 (m)	38.6
	1.91-1.94 (m)
23	3.43 (d, J = 11.2)	66.0
	3.58 (d, J = 11.2)
24	1.06 (s)	15.4
25	1.40 (s)	19.4
26	1.09 (s)	19.3
27	1.16 (s)	24.1
28	-	177.2
29	1.03 (d, J = 6.5)	16.9
30	4.69 (br, s), 4.64 (br, s)	105.5
1'	5.34 (d, J = 8.2)	96.0
2'	3.31-3.32 (m)	74.0
3'	3.38-3.41 (m)	78.3
4'	3.35-3.36 (m)	71.4
5'	3.32-3.33 (m)	78.4
6'	3.79 (dd, J = 12.0, 2.9)	62.6
	3.67 (dd, J = 12.0, 4.7)

δ_H (Hz)

δ_C

0.87-0.91 (m)

50.5

1.91-1.94 (m)

3.72-3.76 (m)

69.9

3.28-3.30 (m)

78.7

44.9

1.29 (br. s)

49.0

4.38 (br. s)

68.6

1.52 (dd, J = 14.3, 2.5)

41.4

1.80 (dd, J = 14.3, 3.6)

44.1

1.62-1.71 (m)

49.7

38.6

1.98-2.05 (m)

24.7

2.08-2.13 (m)

5.32-5.33 (m)

127.8

138.5

40.3

1.14-1.17 (m)

29.4

1.98-2.05 (m)

1.85-1.90 (m)

25.4

2.27-2.30 (m)

49.6

2.30-2.32 (m)

56.6

2.42-2.45 (m)

38.7

154.3

2.21-2.24 (m)

33.4

2.32-2.35 (m)

1.62-1.71 (m)

38.6

1.91-1.94 (m)

3.43 (d, J = 11.2)

66.0

3.58 (d, J = 11.2)

1.06 (s)

15.4

1.40 (s)

19.4

1.09 (s)

19.3

1.16 (s)

24.1

177.2

1.03 (d, J = 6.5)

16.9

4.69 (br, s), 4.64 (br, s)

105.5

5.34 (d, J = 8.2)

96.0

3.31-3.32 (m)

74.0

3.38-3.41 (m)

78.3

3.35-3.36 (m)

71.4

3.32-3.33 (m)

78.4

3.79 (dd, J = 12.0, 2.9)

62.6

3.67 (dd, J = 12.0, 4.7)

No.	2	3	4	5	6	7
1	51.0	51.0	48.1	50.5	50.3	48.4
2	69.9	69.9	69.9	69.8	69.9	69.9
3	78.3	78.3	78.4	78.3	78.4	78.4
4	44.9	44.9	44.3	44.9	45.0	44.3
5	48.7	48.7	48.7	49.6	49.1	48.0
6	68.5	19.0	19.4	68.6	68.9	19.4
7	42.6	35.3	33.7	41.4	40.6	33.7
8	39.5	40.5	41.4	40.6	41.5	41.2
9	49.7	48.6	48.3	48.8	49.1	48.7
10	38.6	39.1	39.1	38.6	38.4	39.3
11	24.5	24.5	24.9	25.4	24.9	24.9
12	128.0	127.7	129.6	127.5	130.0	129.8
13	139.3	139.8	139.9	138.7	139.2	140.4
14	46.3	46.0	42.9	44.0	43.4	43.2
15	29.9	29.8	29.8	29.4	29.8	29.8
16	36.0	36.0	27.4	24.7	26.7	26.8
17	50.8	49.7	49.5	49.4	49.6	49.7
18	134.3	134.2	55.1	54.4	55.1	55.3
19	137.8	137.8	73.8	40.4	73.8	73.8
20	35.8	35.8	42.9	40.3	43.1	42.9
21	27.6	27.6	26.6	31.9	27.3	27.2
22	31.8	31.7	38.5	37.6	38.6	39.2
23	66.0	66.4	66.5	66.0	66.1	66.5
24	15.4	14.2	14.0	15.4	15.4	14.0
25	19.9	19.1	17.8	19.4	19.2	17.8
26	19.9	18.9	17.8	19.4	18.9	17.7
27	20.6	19.9	24.9	24.2	24.9	25.0
28	176.8	176.7	178.7	178.1	178.6	180.9
29	22.6	22.5	27.2	17.8	27.2	27.5
30	19.1	18.5	16.8	21.7	16.8	16.8
1'	95.9	95.9	95.9	95.9	96.0
2'	74.2	74.2	74.0	74.1	74.0
3'	78.8	78.8	78.7	78.7	78.4
4'	71.4	71.3	71.2	71.4	71.3
5'	78.5	78.5	78.4	78.4	78.3
6'	62.7	62.7	62.6	62.1	62.5

No.

51.0

48.1

50.5

50.3

48.4

69.9

69.8

69.9

78.3

78.4

78.3

78.4

44.9

44.3

44.9

45.0

44.3

48.7

49.6

49.1

48.0

68.5

19.0

19.4

68.6

68.9

19.4

42.6

35.3

33.7

41.4

40.6

33.7

39.5

40.5

41.4

40.6

41.5

41.2

49.7

48.6

48.3

48.8

49.1

48.7

38.6

39.1

38.6

38.4

39.3

24.5

24.9

25.4

24.9

128.0

127.7

129.6

127.5

130.0

129.8

139.3

139.8

139.9

138.7

139.2

140.4

46.3

46.0

42.9

44.0

43.4

43.2

29.9

29.8

29.4

29.8

36.0

27.4

24.7

26.7

26.8

50.8

49.7

49.5

49.4

49.6

49.7

134.3

134.2

55.1

54.4

55.1

55.3

137.8

73.8

40.4

73.8

35.8

42.9

40.3

43.1

42.9

27.6

26.6

31.9

27.3

27.2

31.8

31.7

38.5

37.6

38.6

39.2

66.0

66.4

66.5

66.0

66.1

66.5

15.4

14.2

14.0

15.4

14.0

19.9

19.1

17.8

19.4

19.2

17.8

19.9

18.9

17.8

19.4

18.9

17.7

20.6

19.9

24.9

24.2

24.9

25.0

176.8

176.7

178.7

178.1

178.6

180.9

22.6

22.5

27.2

17.8

27.2

27.5

19.1

18.5

16.8

21.7

16.8

95.9

96.0

74.2

74.0

74.1

74.0

78.8

78.7

78.4

71.4

71.3

71.2

71.4

71.3

78.5

78.4

78.3

62.7

62.6

62.1

62.5