CN114940693B - 硫代苯甲酸酯糖苷类化合物及其制备方法和应用 - Google Patents
硫代苯甲酸酯糖苷类化合物及其制备方法和应用 Download PDFInfo
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Abstract
本发明公开了一类结构新颖的硫代苯甲酸酯糖苷类化合物,是可通过大米固体培养基发酵培养,而后提纯,最终得到的天然产物。研究表明,本发明的硫代苯甲酸酯糖苷类化合物可以用于制备抗前列腺癌、抗乳腺癌和抗结肠癌药物。本发明增加了硫代苯甲酸酯糖苷类似物种类和活性的多样性。
Description
技术领域
本发明涉及放线菌次级代谢产物制备活性化合物的领域,具体涉及一类Suncheonosides类化合物及其制备方法和应用。
背景技术
Suncheonosides(结构式如下式(1))属于六取代的硫代苯甲酸酯糖苷类化合物,最早于2015年,由韩国首尔国立大学药学院天然产物研究所从韩国顺天湾的沉积物中得到的海洋链霉菌菌株SSC21 中分离得到该化合物。
研究表明,Suncheonosides可以提高脂联素的产生,从而具有成为增加胰岛素敏感性的抗糖尿病药物的潜力。
自然界中,含有硫代苯酸甲酯的化合物并不常见,并且大部分的化合物来自于放线菌。例如:
Resorthiomycin I(结构式如下式(6))来自于Streptomyces collinus,具有抗肿瘤和抗生素活性(Tahara M,Okabe T,Furihata K,et al. Revised structure ofresorthiomycin[J].Journal of Antibiotics,1991, 44(2):255);
Calicheamicins(结构式如下式(7)),来自于棘孢小单胞菌,是烯二炔部分与硫代苯甲酸酯结合而成的化合物,具有抗癌活性(MD Lee,Manning J K,Williams D R,etal.CALICHEAMICINS,ANOVEL FAMILY OF ANTITUMOR ANTIBIOTICS[J].Journal ofAntibiotics, 1989,42(7):1070-87);
2012年马来西亚博特拉大学从海洋来源的放线菌中分出来了 Suncheonosides苷元(结构式如下式(5)),没有做生物学活性评价 (Mahyudin N A,Blunt J W,Cole AL J,etal.The Isolation of a New S-Methyl Benzothioate Compound from a Marine-Derived Streptomyces sp.[J].Journal of Biomedicine and Biotechnology,2012,2012:1-4);
最近,2015年韩国首尔国立大学药学院天然产物研究所从韩国顺天湾的沉积物中得到的海洋链霉菌菌株SSC21中分离得到 Suncheonosides A-E(结构式分别如下式(1)-(5)),研究表明可能具有潜在的抗糖尿病活性(Shin B,Ahn S,Noh M,et al.Suncheonosides A-D,Benzothioate Glycosides from a Marine-Derived Streptomycessp.[J].Journal of Natural Products,2015,78(6):1390-6)。
六取代的硫代苯甲酸酯很少从放线菌以外的样品中发现,目前只有mortivinactin B(结构式如下式(8))是来自真菌Mortierellavinacea 的,具有抗细菌和抗真菌活性(Soman AG,Gloer J B,Wicklow D T. Antifungal and antibacterialmetabolites from a sclerotium-colonizing isolate of Mortierellavinacea.[J].Journal of Natural Products,1999, 62(2):386-388);另一个是polycarpamine B(结构式如下式(9))来自海洋海鞘Polycarpaauzata,其在体外对酿酒酵母和白色念珠菌具有显着的抗真菌活性(Lindquist N,Fenical W.Polycarpamines A-E, antifungaldisulfides from the marine ascidian polycarpaauzata.[J]. Tetrahedron Letters,1990,31(17):2389-2392)。
现阶段存在的Suncheonosides类似物只有四种而且只测得了抗糖尿病的活性。
为了增加化合物的多样性,及其活性的多样化,本发明分离得到 13个新的Suncheonosides类似物,并测得其具有良好的抗肿瘤活性。
发明内容
本发明提供了一类共十三种结构新颖的Suncheonosides类化合物,通过大米固体培养基发酵培养,而后提纯,最终得到的天然产物。
一类Suncheonosides类化合物,结构式分别如下式(1)~(13) 所示:
本发明还提供了上述Suncheonosides类化合物的制备方法,包括步骤:
1)将放线菌菌株解冻活化后接种高氏一号液体培养基中,摇床培养,获得种子液,取适量种子液加入到大米固体培养基中,静置培养得到发酵液;
所述放线菌,采用中国工业微生物菌种保藏管理中心出售的链霉菌Streptomycessp.CICC 11011;
2)上述发酵液经过有机溶剂萃取后得到萃取液;
3)上述萃取液经浓缩后进行分离纯化,最终获得结构式如式(1) ~(13)所示的Suncheonosides类化合物。
在一优选例中,步骤1)中,放线菌的活化过程包括:将放线菌接种于高氏一号固体培养基中,在26~30℃条件下活化培养3~4天。
在一优选例中,步骤1)中:
高氏一号液体培养基的配制方法为:以培养基1L计,将可溶性淀粉20g、KNO3 1g、K2HPO4 0.5g、MgSO4·7H2O 0.5g、NaCl 0.5g、 FeSO4·7H2O 0.01g、海盐25g混合后加水至1L,再调节pH值至7.2 ~7.4;
大米固体培养基的配制方法为:称取25g海盐于1L水中混合均匀,将pH值调至7.2~7.4,配制成海盐水;每个500mL三角瓶中加入40g大米以及60mL海盐水,搅拌均匀,灭菌备用;
高氏一号液体培养基的条件为:28℃下,180rpm摇床中培养4 天;
种子液接种量为:5mL/瓶;
大米固体培养基培养条件为:28℃,静置40天。
在一优选例中,步骤2)中,所述的有机溶剂为乙酸乙酯。
在一优选例中,步骤3)中:
所述浓缩操作,具体是:所述萃取液经真空干燥除去溶剂;
所述分离纯化操作,具体是:
(a)浓缩后的萃取液采用硅胶柱层析的分离方法,通过体积比为10:1至5:1石油醚-乙酸乙酯体系和40:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得12个馏分,分别记为Fr.A-Fr.L;
(b)将步骤(a)所得馏分采用反相高效液相色谱法进行分离,得到结构式如式(1)~(13)所示的Suncheonosides类化合物。
在一优选例中,步骤(b)中:
Fr.B采用甲醇体积百分数为60%~90%的甲醇-水体系,以10 mL/min进行梯度洗脱40min,收集17~18min的洗脱液得到如式(12) 结构的化合物;
Fr.C采用甲醇体积百分数为60%~90%的甲醇-水体系,以10 mL/min进行梯度洗脱40min,收集26~27min的洗脱液得到如式(10) 结构的化合物,收集17~18min的洗脱液得到如式(13)结构的化合物,收集20~21min的洗脱液得到如式(11)结构的化合物;
Fr.J通过体积比为60:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得13个馏分(Fr.J 1–Fr.J 13);
Fr.J 9采用甲醇体积百分数为10%~100%的甲醇-水体系,以10 mL/min进行梯度洗脱40min,收集33~34min的洗脱液得到如式(8) 结构的化合物,收集35~36min的洗脱液得到如式(7)结构的化合物,收集40~41min的洗脱液得到如式(9)结构的化合物;
Fr.K通过体积比为40:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得10个馏分(Fr.K 1–Fr.K 10);
Fr.K 9采用甲醇体积百分数为20%~90%的甲醇-水体系,以10 mL/min进行梯度洗脱50min,收集32~33min的洗脱液得到如式(1) 结构的化合物;
Fr.K 8采用甲醇体积百分数为10%~100%的甲醇-水体系,以10 mL/min进行中压制备60min,每15min为一个梯度,总共得到6个梯度(Fr.K 8-1–Fr.K 8-6);
Fr.K 8-5采用甲醇体积百分数为55%~90%的甲醇-水体系,以 10mL/min进行梯度洗脱40min,收集13~14min的洗脱液得到如式 (6)结构的化合物,收集19~20min的洗脱液得到如式(3)结构的化合物,收集20~21min的洗脱液得到如式(5)结构的化合物;
Fr.K 8-6采用甲醇体积百分数为55%~85%的甲醇-水体系,以 10mL/min进行梯度洗脱40min,收集17~18min的洗脱液得到如式 (2)结构的化合物,收集27~28min的洗脱液得到如式(4)结构的化合物。
本发明还提供了所述的Suncheonosides类化合物在制备抗肿瘤药物中的应用。
优选的,所述抗肿瘤药物为抗前列腺癌药物、抗乳腺癌药物或抗结肠癌药物。
采用人前列腺癌细胞株(PC-3)、人乳腺癌细胞株(MDA-MB-231) 和人结肠癌细胞株(HCT-116)对本发明分离得到的Suncheonosides 类化合物进行了抗肿瘤活性评价试验。结果表明,式(6)所示的化合物具有较好的PC-3细胞毒活性,为4.1±0.1μM;式(8)~式(10)具有较好的抗MDA-MB-231细胞株的活性,为10.9±0.0~35.7±5.6 μM式(12)所示的化合物具有较好的HCT-116细胞毒活性,为7.3± 0.4μM。
本发明与现有技术相比,主要优点包括:
本发明公开了多种结构新颖的Suncheonosides类化合物,均为放线菌通过发酵培养及提纯后得到。其中,具有式(1)~式(13)对应的六个化合物具有不同于Suncheonosides的母核结构,活性测试发现式(6)具有很好的抗PC-3细胞株的活性,式(8)~式(10)具有较好的抗MDA-MB-231细胞株的活性,式(12)具有很好的抗HCT-116 细胞株的活性。研究表明,本发明的Suncheonosides类化合物可以用于制备抗前列腺癌、抗乳腺癌和抗结肠癌药物。本发明增加了 Suncheonosides类似物种类和活性的多样性。
具体实施方式
菌种来源
本发明放线菌采用中国工业微生物菌种保藏管理中心出售的链霉菌Streptomyces sp.CICC 11011。
培养基
高氏一号平板固体培养基:以培养基1L计,将可溶性淀粉20g、 KNO3 1g、K2HPO40.5g、MgSO4·7H2O 0.5g、NaCl 0.5g、FeSO4·7H2O 0.01g、琼脂20g、海盐25g混合后加水至1L,再调节pH值至7.2 ~7.4。
高氏一号液体培养基:以培养基1L计,将可溶性淀粉20g、KNO3 1g、K2HPO4 0.5g、MgSO4·7H2O 0.5g、NaCl 0.5g、FeSO4·7H2O 0.01 g、海盐25g混合后加水至1L,再调节pH值至7.2~7.4。
大米固体培养基:称取25g海盐于1L纯净水中混合均匀,将 pH值调至7.2~7.4,配制成海盐水,500mL三角瓶中称取40g大米,并加入60mL海盐水。
实施例1
一、化合物的发酵
1)取放线菌适量,接种到高氏一号平板固体培养基上,于28℃培养箱中静置,活化培养3~4天。
2)将步骤1)中己经活化的放线菌的单菌落接种于含有250mL 液体培养基的500mL锥形瓶中,每瓶含250mL高氏一号液体培养基,在摇床中28℃下以180rpm振荡培养4天,获得种子液。
3)取步骤2)中的种子液5mL于含有40g大米固体培养基的 500mL锥形瓶中,28℃静置培养40天,得到发酵液。
二、化合物的制备
(a)将所述的发酵液采用250mL乙酸乙酯萃取3次,所得萃取液经真空干燥浓缩除去乙酸乙酯溶剂。将所得乙酸乙酯部分进行硅胶柱层析,用体积比为10:1至5:1的石油醚-乙酸乙酯体系和40:1至0:1 的二氯甲烷-甲醇体系梯度洗脱,薄层检识含有新化合物的馏分,合并。
(b)所得含有新化合物的馏分Fr.B用反相高效液相色谱分离 (Agilent PursuitC-18(10μm,21.2×250mm))色谱柱,检测波长210 nm,采用甲醇体积百分数为60%~90%的甲醇-水体系,以10mL/min 进行梯度洗脱40min,收集17~18min的洗脱液。将分离纯化得到的化合物进行结构鉴定,分子式根据高分辨质谱HR-ESI-MS计算为 C13H18O4([M+H]+m/z239.1270,计算值为239.1278),进一步依据核磁共振数据(表1),其具体结构如下所示,记为 2,4-dihydroxy-6-isopropyl-3,5-dimethylbenzoate:
该化合物的核磁共振数据列于下表1中,核磁共振参数1H 600 MHz,13C 150MHz,溶剂CD3OD-d4。
表1化合物12的1H和13C数据
发酵液的获得与实施例1中相同,区别仅在于,化合物的制备步骤(b)中,所得含有新化合物的馏分Fr.C用反相高效液相色谱分离 (Agilent Pursuit C-18(10μm,21.2×250mm)色谱柱,检测波长为210 nm,采用甲醇体积百分数为60%~90%的甲醇-水体系,以10 mL/min进行梯度洗脱40min,分别收集17~18min、20~21min和 26~27min的洗脱液。将分离纯化得到的化合物进行结构鉴定,分子式根据高分辨质谱HR-ESI-MS计算,26~27min化合物为C14H20SO3 ([M+Na]+m/z 291.1028,计算值为291.1026),进一步依据核磁共振数据(表2),其具体结构如下所示,记为S-methyl 4-hydroxy-6-isopropyl-2-methoxy-3,5-dimethylbenzothioate:
该化合物的核磁共振数据列于下表2中。
表2化合物10的1H和13C数据
20~21min化合物为C12H16O3S([M+Na]+m/z 263.0714,计算值为263.0713),进一步依据核磁共振数据(表3),其具体结构如下所示,记为S-methyl 6-ethyl-2,4-dihydroxy-3,5-dimethylbenzothioate:
该化合物的核磁共振数据列于下表3中。
表3化合物11的1H和13C数据
17~18min化合物为C11H16O2([M+Na]+m/z 203.1053,计算值为203.1043),进一步依据核磁共振数据(表4),其具体结构如下所示,记为5-isopropyl-2,4-dimethylbenzene-1,3-diol:
该化合物的核磁共振数据列于下表4中。
表4化合物13的1H和13C数据
Fr.J通过体积比为60:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得13个馏分(Fr.J 1–J 13);
所得含有新化合物的馏分Fr.J 9用反相高效液相色谱分离(Agilent Pursuit C-18(10μm,21.2×250mm)色谱柱,检测波长为210 nm,采用甲醇体积百分数为10%~100%的甲醇-水体系,以10mL/min 进行梯度洗脱40min,收集33~34min、35~36min和40~41min的洗脱液。将分离纯化得到的化合物分别进行结构鉴定,分子式根据高分辨质谱HR-ESI-MS计算,33~34min化合物为C20H28O9S ([M+Na]+m/z 467.1346,计算值为467.1347),进一步依据核磁共振数据(表5),其具体结构如下所示,记为Suncheonosides L:
该化合物的核磁共振数据列于下表5中。
表5化合物8的1H和13C数据
35~36min化合物分子式根据高分辨质谱HR-ESI-MS计算为 C21H30O9S([M+Na]+m/z481.1505,计算值为481.1503),进一步依据核磁共振数据(表6),其具体结构如下所示,记为Suncheonosides K:
该化合物的核磁共振数据列于下表6中。
表6化合物7的1H和13C数据
40~41min化合物分子式根据高分辨质谱HR-ESI-MS计算为 C31H42O11S([M+Na]+m/z 645.2339,计算值为645.2341),进一步依据核磁共振数据(表7),其具体结构如下所示,记为Suncheonosides M:
该化合物的核磁共振数据列于下表7中。
表7化合物9的1H和13C数据
*数据通过BC谱获得。
Fr.K通过体积比为40:0至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得10个馏分(Fr.K1–K10);
所得含有新化合物的馏分Fr.K 9用反相高效液相色谱分离(Agilent Pursuit C-18(10μm,21.2×250mm)色谱柱,检测波长为210 nm,采用甲醇体积百分数为20%~90%的甲醇-水体系,以10mL/min 进行梯度洗脱50min,收集32~33min的洗脱液,将分离纯化得到的化合物分别进行结构鉴定,分子式根据高分辨质谱HR-ESI-MS计算为C19H28O8S([M+Na]+m/z 439.1403,计算值为439.1398),进一步依据核磁共振数据(表8),其具体结构如下所示,记为 Suncheonosides E:
该化合物的核磁共振数据列于下表8中。
表8化合物1的1H和13C数据
Fr.K 8采用甲醇体积百分数为10%~100%的甲醇-水体系,以10 mL/min进行中压制备60min,每15min为一个梯度,总共得到6个梯度(K 8-1-K 8-6);
所得含有新化合物的馏分Fr.K 8-5用反相高效液相色谱分离 (Agilent PursuitC-18(10μm,21.2×250mm)色谱柱,检测波长为210 nm,采用甲醇体积百分数为55%~90%的甲醇-水体系,以10mL/min 进行梯度洗脱40min,分别收集13~14min、19~20min和20~21 min的洗脱液。将分离纯化得到的化合物分别进行结构鉴定,分子式根据高分辨质谱HR-ESI-MS计算,13~14min化合物为C17H26O8 ([M+Na]+m/z 381.1517,计算值为381.1520),进一步依据核磁共振数据(表9),其具体结构如下所示,记为Suncheonosides J:
该化合物的核磁共振数据列于下表9中。
表9化合物6的1H和13C数据
19~20min化合物为C17H24O8S([M+Na]+m/z 411.1079,计算值为411.1085),进一步依据核磁共振数据(表10),其具体结构如下所示,记为Suncheonosides G:
该化合物的核磁共振数据列于下表10中。
表10化合物3的1H和13C数据
20~21min化合物为C18H26O8S([M+Na]+m/z 425.1243,计算值为425.1241),进一步依据核磁共振数据(表11),其具体结构如下所示,记为Suncheonosides I:
该化合物的核磁共振数据列于下表11中。
表11化合物5的1H和13C数据
所得含有新化合物的馏分Fr.K 8-6用反相高效液相色谱分离 (Agilent PursuitC-18(10μm,21.2×250mm)色谱柱,检测波长为210 nm,采用甲醇体积百分数为55%~85%的甲醇-水体系,以10mL/min 进行梯度洗脱40min,分别收集17~18min和27~28min的洗脱液。将分离纯化得到的化合物分别进行结构鉴定,分子式根据高分辨质谱HR-ESI-MS计算,17~18min化合物为C18H26O8S([M+Na]+ 425.1248,计算值为425.1241),进一步依据核磁共振数据(表12),其具体结构如下所示,记为Suncheonosides F:
该化合物的核磁共振数据列于下表12中。
表12化合物2的1H和13C数据
27~28min化合物为C20H30O8S([M+Na]+m/z 453.1554,计算值为453.1554),进一步依据核磁共振数据(表13),其具体结构如下所示,记为Suncheonosides H:
该化合物的核磁共振数据列于下表13中。
表13化合物4的1H和13C数据
*数据通过BC谱获得。
活性测试:
抗肿瘤活性实验
人前列腺癌细胞PC-3、人乳腺癌细胞MDA-MB-231和人结肠癌细胞HCT-116抑制率实验。测定不同浓度实施例1~13分别纯化得到的化合物的抑制活性。以阿霉素作为阳性药对照。取正常培养的处于对数期的细胞,3×104个/mL铺96孔板,加入药物培养24h后CCK8检测细胞活力。细胞存活率(%)=实验组OD值/不加药组 OD值×100%。最终根据各浓度抑制率计算半数抑制浓度IC50,如表14化合物1~13细胞毒性检测结果所示。
表14化合物1~13细胞毒性检测结果(IC50/μM)
No. | PC-3 | MDA-MB-231 | HCT-116 |
1 | >40 | >40 | >40 |
2 | >40 | >40 | >40 |
3 | >40 | >40 | >40 |
4 | >40 | >40 | >40 |
5 | >40 | >40 | >40 |
6 | 4.1±0.1 | >40 | >40 |
7 | 未测 | >40 | >40 |
8 | 未测 | 15.9±1.1 | >40 |
9 | 未测 | 10.9±0.0 | >40 |
10 | >40 | 35.7±5.6 | >40 |
11 | >40 | >40 | >40 |
12 | >40 | >40 | 7.3±0.44 |
13 | >40 | >40 | >40 |
阿霉素 | 0.4±0.2 | 0.4±0.0 | 0.07±0.0 |
此外应理解,在阅读了本发明的上述描述内容之后,本领域技术人员可以对本发明作各种改动或修改,这些等价形式同样落于本申请所附权利要求书所限定的范围。
Claims (3)
1.一类Suncheonosides类化合物,其特征在于,结构式分别如下式(1)~(13)所示:
。
2.根据权利要求1所述的Suncheonosides类化合物的制备方法,其特征在于,包括步骤:
1)将放线菌菌株解冻活化后接种高氏一号液体培养基中,28℃下,180 rpm摇床中培养4天,获得种子液,取适量种子液加入到大米固体培养基中,28℃静置培养40天得到发酵液;
所述放线菌,采用中国工业微生物菌种保藏管理中心出售的链霉菌Streptomyces sp. CICC 11011;
放线菌的活化过程包括:将放线菌接种于高氏一号固体培养基中,在26 ~ 30℃条件下活化培养3 ~ 4天;
高氏一号液体培养基的配制方法为:以培养基1 L计,将可溶性淀粉20 g、KNO3 1 g、K2HPO4 0.5 g、MgSO4•7H2O 0.5 g、NaCl 0.5 g、FeSO4•7H2O 0.01 g、海盐25 g混合后加水至1 L,再调节pH 值至7.2 ~ 7.4;
大米固体培养基的配制方法为:称取25 g海盐于1 L水中混合均匀,将pH值调至7.2 ~7.4,配制成海盐水;每个500 mL三角瓶中加入40 g大米以及60 mL海盐水,搅拌均匀,灭菌备用;
种子液接种量为:5 mL/瓶;
2)上述发酵液经过有机溶剂乙酸乙酯萃取后得到萃取液;
3)上述萃取液经真空干燥除去溶剂浓缩后进行分离纯化,最终获得结构式如式(1)~(13)所示的Suncheonosides类化合物;
分离纯化操作具体是:
(a)浓缩后的萃取液采用硅胶柱层析的分离方法,通过体积比为10:1至5:1石油醚-乙酸乙酯体系和40:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得 12 个馏分,分别记为Fr. A-Fr. L;
(b)将步骤(a)所得馏分采用反相高效液相色谱法进行分离,得到结构式如式(1)~(13)所示的Suncheonosides类化合物;
步骤(b)中:
Fr.B采用甲醇体积百分数为60% ~ 90%的甲醇-水体系,以10 mL/min进行梯度洗脱40min,收集17 ~ 18 min的洗脱液得到如式(12)结构的化合物;
Fr.C采用甲醇体积百分数为60% ~ 90%的甲醇-水体系,以10 mL/min进行梯度洗脱40min,收集26 ~ 27 min的洗脱液得到如式(10)结构的化合物,收集17 ~ 18 min的洗脱液得到如式(13)结构的化合物,收集20 ~ 21 min的洗脱液得到如式(11)结构的化合物;
Fr.J通过体积比为60:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得 13 个馏分,分别记为Fr. J 1–Fr. J 13;
Fr.J 9采用甲醇体积百分数为10% ~ 100%的甲醇-水体系,以10 mL/min进行梯度洗脱40 min,收集33 ~ 34 min的洗脱液得到如式(8)结构的化合物,收集35 ~ 36 min的洗脱液得到如式(7)结构的化合物,收集40 ~ 41 min的洗脱液得到如式(9)结构的化合物;
Fr.K通过体积比为40:1至0:1的二氯甲烷-甲醇体系进行梯度洗脱,收集包含目标化合物的馏分,合并得 10 个馏分,分别记为Fr. K 1–Fr.K 10;
Fr.K 9采用甲醇体积百分数为20% ~ 90%的甲醇-水体系,以10 mL/min进行梯度洗脱50 min,收集32 ~ 33 min的洗脱液得到如式(1)结构的化合物;
Fr.K 8 采用甲醇体积百分数为10% ~ 100%的甲醇-水体系,以10 mL/min进行中压制备60 min,每15 min为一个梯度,总共得到6个梯度,分别记为Fr.K 8-1–Fr.K 8-6;
Fr.K 8-5采用甲醇体积百分数为55% ~ 90%的甲醇-水体系,以10 mL/min进行梯度洗脱40 min,收集13 ~ 14 min的洗脱液得到如式(6)结构的化合物,收集19 ~ 20 min的洗脱液得到如式(3)结构的化合物,收集20 ~ 21 min的洗脱液得到如式(5)结构的化合物;
Fr.K 8-6采用甲醇体积百分数为55% ~ 85%的甲醇-水体系,以10 mL/min进行梯度洗脱40 min,收集17 ~ 18 min的洗脱液得到如式(2)结构的化合物,收集27 ~ 28 min的洗脱液得到如式(4)结构的化合物。
3.根据权利要求1所述的Suncheonosides类化合物在制备抗肿瘤药物中的应用,其特征在于,所述抗肿瘤药物为抗前列腺癌药物、抗乳腺癌药物或抗结肠癌药物。
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