具体实施方式
现结合实例,对本发明做进一步说明。实例仅限于说明本发明,而非对本发明的限定。
实施例1:皂苷提取物第一维馏分制备
三七叶水提物经制备型HPLC分离(C18 TDE柱;水(A)和乙腈(B)流动相体系;洗脱梯度为0-5min,体积浓度20%→32%B;5-45min,体积浓度32%→68%B;45-50min,体积浓度68%→100%B;50-55min,100%B);分离过程中采用紫外检测器检测,检测波长203nm。1-55min,每分钟收集一份,共计54个组分。分别为Fractions 1-54,皂苷主要集中在Fractions 6-30。
实施例2:化合物P1和P2的制备
选取Fraction 9采用亲水色谱模式进行第二维制备型HPLC分离(色谱柱为酰胺柱;水(A)和乙腈(B)流动相体系;体积浓度76%B等度洗脱)流速为20mL/min;检测波长为203nm;进样量为1mL。收集各个色谱峰,分别回收溶剂,经核磁实验确定,获得P1和P2两个化合物。HPLC检测纯度大于95%,经理化测定,数据如下:P1,白色粉末,HR-ESI-MS:[M+H]+(m/z):1211.6409.1H NMR(600MHz,pyridine-d5)δ:0.77(1H,s,H-19),0.92(2H,s,H-18,30),1.08(1H,s,H-29),1.25(1H,s,H-28),1.60(1H,s,H-21),1.62(1H,s,H-27),1.64(1H,s,H-26),3.28(1H,dd,J=4.2,11.4,H-3),5.29(1H,t,J=7.2,H-24),4.92(1H,d,J=7.8),5.50(1H,d,J=7.8),5.40(1H,d,J=6.6),5.13(1H,d,J=7.8),4.85(1H,brs).13C NMR:见表1.P2,白色粉末,HR-ESI-MS:[M+H]+(m/z):1241.6498.1H NMR(600MHz,pyridine-d5)δ:0.78(1H,s,H-19),0.93(1H,s,H-30),0.94(1H,s,H-18),1.08(1H,s,H-29),1.25(1H,s,H-28),1.58(1H,s,H-21),1.64(2H,s,H-26,27),3.28(1H,dd,J=4.2,11.4,H-3),5.29(1H,t,J=6.6,H-24),4.91(1H,d,J=7.8),5.49(1H,d,J=7.2),5.39(1H,d,J=7.2),5.12(1H,d,J=7.8),5.09(1H,d,J=7.8).13C NMR:见表1.
实施例3:化合物P3-P6的制备
选取Fraction 11采用亲水色谱模式进行第二维制备型HPLC分离(色谱柱为酰胺柱;水(A)和乙腈(B)流动相体系;体积浓度80%B等度洗脱)流速为20mL/min;检测波长为203nm;进样量为1mL。收集各个色谱峰,分别回收溶剂,经核磁实验确定,获得P3-P6四个化合物。HPLC检测纯度大于95%,经理化测定,数据如下:P3,白色粉末,HR-ESI-MS:[M+H]+(m/z):1079.5989.1H NMR(600MHz,pyridine-d5)δ:0.78(1H,s,H-19),0.92(1H,s,H-30),0.93(1H,s,H-18),1.08(1H,s,H-29),1.26(1H,s,H-28),1.60(1H,s,H-21),1.62(1H,s,H-27),1.64(1H,s,H-26),3.24(1H,dd,J=4.2,11.4,H-3),5.29(1H,t,J=6.6,H-24),4.90(1H,d,J=7.8),5.35(1H,d,J=7.8),5.12(1H,d,J=7.8),4.85(1H,brs).13C NMR:见表1.P4,白色粉末,HR-ESI-MS:[M+H]+(m/z):1109.6064.1H NMR(600MHz,pyridine-d5)δ:0.80(1H,s,H-19),0.94(2H,s,H-18,30),1.09(1H,s,H-29),1.27(1H,s,H-28),1.59(1H,s,H-21),1.64(2H,s,H-26,27),3.26(1H,dd,J=4.2, 11.4,H-3),5.30(1H,t,J=6.6,H-24),4.91(1H,d,J=7.8),5.36(1H,d,J=7.8),5.12(1H,d,J=7.8),5.09(1H,d,J=7.8).13C NMR:见表1.P5,白色粉末,HR-ESI-MS:[M+H]+(m/z):1211.6390.1H NMR(600MHz,pyridine-d5)δ:0.78(1H,s,H-19),0.92(1H,s,H-30),0.94(1H,s,H-18),1.09(1H,s,H-29),1.25(1H,s,H-28),1.59(1H,s,H-21),1.63(2H,s,H-26,27),3.28(1H,dd,J=4.2,12,H-3),5.29(1H,t,J=7.2,H-24),4.91(1H,d,J=7.8),5.50(1H,d,J=7.8),5.40(1H,d,J=7.2),5.12(1H,d,J=7.8),4.97(1H,d,J=7.2).13C NMR:见表1.P6,白色粉末,HR-ESI-MS:[M+H]+(m/z):1343.6930.1H NMR (600MHz,pyridine-d5)δ:0.78(1H,s,H-19),0.92(1H,s,H-30),0.95(1H,s,H-18),1.09(1H,s,H-29),1.25(1H,s,H-28),1.59(1H,s,H-21),1.62(1H,s,H-27),1.63(1H,s,H-26),3.28(1H,dd,J=4.2,11.4,H-3),5.28(1H,t,J=6.6,H-24),4.92(1H,d,J=6.0),5.50(1H,d,J=7.8),5.40(1H,d,J=6.6),5.10(1H,d,J=7.8),5.50(1H,d,J=7.8).13C NMR:见表1.
实施例4:化合物P7和P8的制备
选取Fraction 13采用亲水色谱模式进行第二维制备型HPLC分离(色谱柱为酰胺柱XAmide,250×20mm,i.d.,10μm);水(A)和乙腈(B)流动相体系;体积浓度80%B等度洗脱)流速为20mL/min;检测波长为203nm;进样量为1mL。收集各个色谱峰,分别回收溶剂,经核磁实验确定,获得P7和P8两个化合物。HPLC检测纯度大于95%,经理化测定,数据如下:P7,白色粉末,HR-ESI-MS:[M-H]-(m/z):945.5468.1H NMR(600MHz,pyridine-d5)δ:0.79(1H,s,H-19),0.93(1H,s,H-30),0.94(1H,s,H-18),1.09(1H,s,H-29),1.26(1H,s,H-28),1.57(2H,s,H-21,27),1.61(1H,s,H-26),3.25(1H,dd,J=4.2,12,H-3),5.22(1H,t,J=6.6,H-24),4.91(1H,d,J=7.8),5.35(1H,d,J=7.8),5.18(1H,d,J=7.2).13C NMR:见表1.P8,白色粉末,HR-ESI-MS:[M+H]+(m/z):1079.5948.1H NMR(600MHz,pyridine-d5)δ:0.78(1H,s,H-19),0.93(1H,s,H-30),0.95(1H,s,H-18),1.09(1H,s,H-29),1.26(1H,s,H-28),1.59(1H,s,H-21),1.63(2H,s,H-26,27),3.26(1H,dd,J=4.2,12,H-3),5.29(1H,t,J=6.6,H-24),4.90(1H,d,J=7.2),5.35(1H,d,J=7.8),5.11(1H,d,J=7.8),4.97(1H,d,J=7.8).13C NMR:见表1.
表1化合物P1-P8的13C NMR数据(150MHz,氘代吡啶)(The 13CNMR (150MHz)spectral data of the compounds P1-P8(in pyridine-d5))
glu:β-D-吡喃型葡萄糖基,xyl:β-D-吡喃型木糖基,ara:α-L-呋喃型阿拉伯糖基或α-L-吡喃型阿拉伯糖基(glu:β-D-glucopyranosyl,xyl:β-D-xylopyranosyl,ara:α-L-arabinofuranosyl or α-L-arabinopyranosyl)