CN1322729A - 薯蓣皂甙的酰基化衍生物、制备方法及用途 - Google Patents

薯蓣皂甙的酰基化衍生物、制备方法及用途 Download PDF

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CN1322729A
CN1322729A CN 01112870 CN01112870A CN1322729A CN 1322729 A CN1322729 A CN 1322729A CN 01112870 CN01112870 CN 01112870 CN 01112870 A CN01112870 A CN 01112870A CN 1322729 A CN1322729 A CN 1322729A
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俞飚
邢国文
惠永正
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Shanghai Institute of Organic Chemistry of CAS
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Abstract

本发明涉及一种单酰基化与双酰基化薯蓣皂甙衍生物,系在有机溶剂中利用脂肪酶催化薯蓣皂甙衍生物和酯制得,并用质谱、核磁共振波谱和元素分析等对其结构进行了鉴定,该衍生物具有生理活性,可用于抗肿瘤的药物。

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薯蓣皂甙的酰基化衍生物、制备方法及用途
本发明涉及薯蓣皂甙衍生物。具体的说是一种单酰基化与双酰基化薯蓣皂甙衍生物，该衍生物是在有机溶剂中利用脂肪酶催化制备这类化合物，并用质谱、核磁共振波谱和元素分析等对其结构进行了鉴定。该衍生物具有生理活性，可用于抗肿瘤的药物。
皂甙是一种具有多种生理活性的糖缀合物，广泛地存在于植物的根、茎、叶中。我国传统的中草药如人参、远志、黄芪等中都含有大量的皂甙。根据其分子中甙元的不同，皂甙通常被分为三萜皂甙和甾体皂甙两大类。在自然界中，许多三萜皂甙糖链上的自由羟基被酰基官能团所修饰，如：乙酰基，丁酰基，苯甲酰基等。但是很少有报道甾体皂甙中自由羟基被酰基官能团修饰的情况。为了深入研究天然的甾体皂甙经过酰基修饰后其生物活性的变化，为药物筛选提供具有生理活性的化合物，我们对这类化合物的合成进行了研究。由于皂甙分子中糖链部分存在多个反应活性接近的羟基官能团，且糖链的构成复杂多变，用传统的化学合成方法制备单取代或双取代的皂甙酰基化衍生物具有合成步骤多，合成周期长等不足。酶催化反应具有立体选择性高，区域选择性强以及反应条件温和等优点，在不对称合成、光学活性化合物拆分及天然产物的制备等领域得到了许多应用。近年来，Riva等人报道了在叔戊醇中利用脂肪酶催化合成了一种三萜皂甙--人参皂甙的酰基化衍生物(Danieli,B.；Luisetti,M；Riva,S.；Bertinotti,A.；Ragg,E.；Scaglioni,L.；Bombardelli,E.J.Org.Chem.,1995,60,3637-3642.)。但是到目前为止还没有使用脂肪酶合成甾体皂甙--薯蓣皂甙酰基化衍生物的报道。因此，提供单酰基化与双酰基化薯蓣皂甙衍生物和使用酶法方便地制备薯蓣皂甙的各种酰基化衍生物是很有意义的。
本发明的目的是在发明人对一系列薯蓣皂甙的化学合成进行了系统的研究基础上，提供获得的一系列不同单酰基化与双酰基化薯蓣皂甙衍生物，即单酰基化或双酰基化的二糖和三糖薯蓣皂甙衍生物。
本发明的另一目的是提供一种制备酰基化薯蓣皂甙衍生物的酶催化合成方法。
本发明的目的还提供一种上述酰基化薯蓣皂甙衍生物的用途，即为药物筛选提供具有生理活性的化合物。
本发明提供的一种单酰基化与双酰基化薯蓣皂甙衍生物结构式如下：结构式中，R=C1-8的酰基或烯酰基。如乙酰基(Ac)，丁酰基[CH3(CH2)2C(O)]，己酰基[CH3(CH2)4C(O)]或乙烯基己二酰基[CH2=CHOC(O)(CH2)4C(O)]等；r,p,q=0、1或2，且，0＜r+p+q≤2；M=葡萄糖基
Figure A0111287000061
N,U=鼠李糖基
Figure A0111287000062
或阿拉伯糖基 n或u=0或1。
本发明的上述单酰基化与双酰基化薯蓣皂甙衍生物可以是下述结构式中的化合物：
Figure A0111287000064
上述结构式中：R1、R2或R3=H或R,R=C1-8的酰基或烯酰基，其中R1、R2或R3不能同时等于H。
本发明的合成方法系采用脂肪酶催化的方法，以不同的薯蓣皂甙为原料经过一步酶催化反应，合成本发明的单酰基化或双酰基化的二糖或三糖的薯蓣皂甙衍生物，并用质谱，一维核磁共振及二维核磁共振波谱，元素分析等对其结构进行了鉴定。这些核磁共振波谱包括1H-NMR(核磁共振氢谱)，13C-NMR(核磁共振碳谱)，1H-1HCOSY(同核化学位移相关谱)，TOCSY(全相关谱)，HSQC(异核单量子相关谱)等。该类化合物的结构可以如上所述。
本发明方法采用薯蓣皂甙为原料，其结构式如下：结构式中，M=葡萄糖基
Figure A0111287000072
N,U=鼠李糖基 或阿拉伯糖基
Figure A0111287000074
n,u=0或1。
本发明的方法中可以是薯蓣甙元-β-D-葡萄糖甙(1)、薯蓣甙元-α-L-鼠李糖基-(1→2)-β-D-葡萄糖甙(2)、薯蓣甙元-α-L-鼠李糖基-(1→4)-β-D-葡萄糖甙(3)、薯蓣甙元-α-L-鼠李糖基-(1→2)-[α-L-鼠李糖基-(1→4)]-β-D-葡萄糖甙(4)、薯蓣甙元-α-L-鼠李糖基-(1→2)-[α-L-阿拉伯糖基-(1→4)]-β-D-葡萄糖甙(5)或薯蓣甙元-α-L-鼠李糖基-(1→6)-β-D-葡萄糖甙(6)等薯蓣皂甙为原料，在有机溶剂中和脂肪酶为催化剂，与不同酯的转酯反应，得到了各种单酰基化或双酰基化的二糖和三糖薯蓣皂甙衍生物。
本发明的方法还可进一步描述如下：
酰基给体ROR’和上述的酰基受体薯蓣皂甙在极性有机溶剂中，并在脂肪酶的催化下发生转酯反应，在酶促反应中，所述酰基受体、酰基给体与脂肪酶的摩尔比依次为1-7∶100-210∶0.02-0.4。上述化合物在极性溶剂中，反应温度为室温-100℃，反应1小时-120小时得到单酰基化或双酰基化的薯蓣皂甙衍生物。所述酰基给体ROR’中，R为C1-8的酰基或烯酰基。如：乙酰基，丁酰基，己酰基，苯甲酰基，丁烯酰基，乙烯基己二酰基等。R’为乙烯基或三氟乙基。所述极性溶剂为四氢呋喃(THF)，三氯甲烷，N,N-二甲基甲酰胺(DMF)，二氧六环或水等。所用催化剂脂肪酶为下述各种脂肪酶中的任意一种：脂肪酶CRL(lipase from Candida rugosa)，脂肪酶PPL(lipase from Porcine Pancreas)，脂肪酶MJL(lipase from Mucor javanicus)，脂肪酶WGL(lipase from Wheat Germ)，脂肪酶PFL(Amano lipase from Pseudomonasfluorescens)，脂肪酶Novozyme 435(Lipase B from Candida antar ctica immobilized onacrylic resin)等。反应产物单酰基化或双酰基化的薯蓣皂甙衍生物可经硅胶柱层析分离纯化。
以脂肪酶催化制备6-O-乙酰基延龄草甙1a和4,6-O-二乙酰基延龄草甙1b为例，本发明的方法可以用下述反应式表示：
Figure A0111287000075
本发明的单酰基化或双酰基化的薯蓣皂甙衍生物和皂甙库具有抗肿瘤等生理活性。可以用于抗肿瘤的药物。
本发明首次提供和制备了一系列具有生理活性的单酰基化或双酰基化的薯蓣皂甙衍生物。利用酶促反应制备酰基化薯蓣皂甙衍生物，区域选择性高，方法简便，大大提高了合成此类化合物的效率，为发现和筛选新的皂甙化合物提供了一种高效、方便的方法。与化学法合成相比，酶法制备酰基化薯蓣皂甙衍生物具有工业化前景。
通过下述实施例将有助于理解本发明，但不能限制本发明的内容：
                        实施例1 实验试剂与分析仪器
各种脂肪酶购自Sigma或Aldrich。各种乙烯酯与三氟乙酯购自东京化成或自制。反应溶剂THF,DMF，二氧六环为分析纯试剂。柱层析硅胶H(10-40μ)为国产硅胶。比旋光值经Pekin-Elmer 241MC旋光仪测得，测试温度未经特别注明一般为17℃。各种一维和二维核磁共振(NMR)波谱用Bruker AM300,Bruker DPX300或Bruker DPX400核磁共振波谱仪测得，测试用溶剂为氘代吡啶。电喷雾质谱(ESI-MS)所用质谱仪型号为PE Mariner API-TOF。元素分析仪型号为Elementar Vario EL。
                        实施例2 脂肪酶催化反应的一般步骤
取薯蓣皂甙30mg于1-5mlTHF中，加入乙烯酯或三氟乙酯0.1-2ml以及脂肪酶10-200mg。于40℃振荡反应1-3天后，减压蒸除溶剂，残余物用粗硅胶拌样上柱，用硅胶H柱层析纯化。用CH2Cl2→CH2Cl2-CH3OH(30∶1)→CH2Cl2-CH3OH(20∶1)→CH2Cl2-CH3OH(10∶1)梯度洗脱，用TLC检测收集的洗脱液，将含产品的部分减压浓缩得白色固体。
                        实施例3 实验结果
     表1  反应条件及各种乙酰化薯蓣皂甙衍生物的产率皂甙       脂肪酶              反应溶剂      反应时间      产物    产率％1          PPL,MJL or WGL      THF           1-2           1a      101a        CRL                 THF           4             1a      171b        CRL                 DMF           2             1a      151c        PFL                 THF           4             1a      431          Novozyme 435        THF           1             1a,1b   75,171          Novozyme 435        THF           2             1a,1b   64,351          Novozyme 435        二氧六环      3             1a,1b   62,322          Novozyme 435        THF           2             2a,2b   53,313          Novozyme 435        THF           2             3a      914          Novozyme 435        THF           2             4a      865          Novozyme 435        THF           3             5a,5b   55,266          Novozyme 435        THF           2             6a      70ea，加入磷酸盐缓冲液(pH7.4)于反应体系中，使THF的含水量为0.8％(V/V)。b，加入磷酸盐缓冲液(pH7.4)于反应体系中，使DMF的含水量为10.0％(V/V)。c，加入磷酸盐缓冲液(pH8.0)于反应体系中，使THF的含水量为0.8％(V/V)。
                        实施例4实验结果
表2利用Novozyme 435在THF中催化制备各种酰基化皂甙4a-4d的反应产率酰基给体                          酰基受体    产物      产率％AcOCH=CH2                        4           4a        86AcOCH2CF3                       4           4a        76CH3(CH2)2C(O)OCH=CH2         4           4b        63CH3(CH2)2C(O)OCH2CF3        4           4b        60CH3(CH2)4C(O)OCH=CH2         4           4c        40CH3CH=CHC(O)OCH=CH2             4           /         10PhC(O)OCH=CH2                    4           /         5CH2=CHOC(O)(CH2)4C(O)OCH=CH2 4           4d        72
实施例5各种酰基化薯蓣皂甙衍生物的物理常数及波谱数据薯蓣甙元-6-O-乙酰基-β-D-葡萄糖甙(Diosgenyl 6-O-acetyl-β-D-glucopyranoside)1aRf:0.52[CH2Cl2-CH3OH(10∶1)]；[α]D=-94.6°(c0.56,THF)；ESI-MS:641.6(M+Na),657.6(M+K)；1H-NMR(400MHz)5.26(d,1H,J=4.8Hz),4.94(d,1H,J=7.6Hz),4.87(d,1H,J=10.0Hz),4.75(dd,1H,J=11.6,5.6Hz),4.48(m,1H),4.20(t,1H,J=8.8Hz),4.05(t,1H,J=9.2Hz),3.98(m,2H),3.86(m,1H),3.54-3.41(m,2H),1.90(s,3H),1.07(d,3H,J=6.8Hz),0.84(s,3H),0.77(s,3H),0.62(d,3H,J=5.2Hz)；13C-NMR(100MHz):171.0(C=O),141.1,121.9,109.4,103.0,81.3,78.8,78.5,75.4,75.3,71.6,67.0,65.0,63.1,56.8,50.4,42.1,40.6,40.0,39.5,37.7,37.2,32.4(2×C),32.0,31.8,30.8,30.4,29.4,21.3,21.0,19.6,17.5,16.6,15.2.Anal.Calcd.for C35H54O9·H2O:C,66.01；H,8.86.Found:C,66.41；H8.87。薯蓣甙元-4,6-O-二乙酰基-β-D-葡萄糖甙(Diosgenyl 4,6-O-di-acetyl-β-D-glucopyranoside)1bRf:0.72[CH2Cl2-CH3OH(10∶1)]；[α]D=-64.1°(c,0.30,THF)；ESI-MS:661.6(M+1),683.6(M+Na),699.6(M+K)；1H-NMR(400MHz)5.80(t,1H,J=9.4Hz),5.23(d,1H,J=4.8Hz),4.94(d,1H,J=7.6Hz),4.84(dd,1H,J=11.8,1.8Hz),4.72(dd,1H,J=11.8,5.4Hz),4.48(m,1H),4.09(t,1H,J=9.4Hz),4.03-3.97(m,2H),3.82(m,1H),3.54-3.40(m,2H),1.93(s,3H),1.07(d,3H,J=6.8Hz),0.80(s,3H),0.76(s,3H),0.62(d,3H,J=5.6Hz)；13C-NMR(100MHz):171.0(2×C=O),141.0,122.0,109.4,102.7,81.3,79.2,78.9,75.0,73.2,69.7,67.0,64.5,63.1,56.8,50.4,42.1,40.6,40.0,39.3,37.6,37.2,32.4(2×C),32.0,31.8,30.8,30.3,29.4,21.4,21.3,20.9,19.5,17.5,16.5,15.2.Anal.Calcd.for C37H56O10·H2O:C,65.46；H,8.61.Found:C,65.82；H,8.32.薯蓣甙元-α-L-鼠李糖基-(1→2)-6-O-乙酰基-β-D-葡萄糖甙(Diosgenyl α-L-rhamnopyranosyl-(1→2)-6-O-acetyl-β-D-glucopyranoside)2aRf:0.15[CH2Cl2-CH3OH(8∶1)]；[α]D=-99.8°(c,0.29,THF)；ESI-MS:765.7(M+1),787.7(M+Na)；1H-NMR(400MHz):6.37(s,1H),5.25(d,1H,J=4.8Hz),4.96-4.91(m,2H),4.81(d,1H,J=10.8Hz),4.75-4.68(m,2H),4.57(dd,1H,J=9.2,3.2Hz),4.49(m,1H),4.30(t,1H,J=9.5Hz),4.25-4.17(m,2H),3.97-3.89(m,3H),3.54-3.41(m,2H),1.88(s,3H),1.71(d,3H,J=6.4Hz),1.07(d,3H,6.8Hz),0.98(s,3H),0.76(s,3H),0.63(d,3H,J=5.2Hz)；13C-NMR(100MHz):171.0(C=O),141.0,121.9,109.4,102.3,100.8,81.3,79.5,78.7,77.8,75.0,74.3,73.0,72.7,71.7,69.8,67.0,64.7,63.1,56.8,50.4,42.1,40.6,40.0,39.2,37.7,37.3,32.5,32.4,32.0,31.9,30.8,30.4,29.4,21.3,20.9,19.6,18.9,17.5,16.5,15.2.Anal.Calcd.for C41H64O13·2H2O:C,61.48；H,8.56；.Found:C,61.20；H,8.48.薯蓣甙元-4-O-乙酰基-α-L-鼠李糖基-(1→2)-6-O-乙酰基-β-D-葡萄糖甙(Diosgenyl 4-O-acetyl-α-L-rhamnopyranosyl-(1→2)-6-O-acetyl-β-D-glucopyranoside)2bRf:0.37[CH2Cl2-CH3OH(8:1)]；[α]D=-80.4°(c0.28,THF)；ESI-MS:807.7(M+1),829.7(M+Na),845.7(M+K)；1H-NMR(400MHz)6.36(s,1H),5.83(t,1H,J=9.8Hz),5.33(d,1H,J=4.8Hz),5.01-4.94(m,2H),4.81(d,1H,J=10.4Hz),4.72-4.68(m,2H),4.61(dd,1H,J=9.6,3.2Hz),4.51(m,1H),4.22-4.15(m,2H),3.97-3.88(m,3H),3.54-3.41(m,2H),2.04(s,3H),1.86(s,3H),1.44(d,3H,J=6.0Hz),1.09(d,3H,6.8Hz),1.04(s,3H),0.81(s,3H),0.63(d,3H,J=5.2Hz)；13C-NMR(100MHz):171.0(C=O),170.9(C=O),140.9,122.2,109.5,101.7,100.2,81.3,79.4,78.2,76.9,76.3,75.1,72.6,71.6,70.4,67.0(2×C),64.6,63.1,56.8,50.5,42.1,40.7,40.0,39.2,37.6,37.3,32.5,32.4,32.0,31.9,30.8,30.3,29.4,21.4,21.3,20.9,19.6,18.2,17.5,16.6,15.2.Anal.Calcd.for C43H66O14:C,64.00；H,8.24.Found:C,64.06；H,8.54.薯蓣甙元-4-O-乙酰基-α-L-鼠李糖基-(1→4)-β-D-葡萄糖甙(Diosgenyl 4-O-acetyl-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranoside)3aRf:0.27[CH2Cl2-CH3OH(10:1)]；[α]D=-95.8°(c,0.16,THF)；ESI-MS:765.7(M+1),787.7(M+Na),803.6(M+K)；1H-NMR(400MHz):5.90(s,1H),5.80(t,1H,J=9.6Hz),5.25(d,1H,J=5.2Hz),5.07(m,1H),4.92(d,1H,J=7.6Hz),4.63(brs,1H),4.56(dd,1H,J=9.6,3.2Hz),4.50-4.44(m,2H),4.23-4.18(m,2H),4.06(brd,1H),3.92(t,1H,J=8.4Hz),3.82(m,1H),3.66(brd,1H),3.53-3.40(m,2H),1.96(s,3H),1.40(d,3H,J=6.0Hz),1.07(d,3H,J=6.8Hz),0.84(s,3H),0.76(s,3H),0.62(d,3H,J=5.2Hz)；13C-NMR(100MHz):170.9(C=O),141.0,122.0,109.4,102.7,102.3,81.3,78.4,77.6,77.4,76.8,76.2,75.9,72.7,70.5,67.6,67.0,63.0,61.5,56.8,50.4,42.1,40.6,40.0,39.5,37.6,37.2,32.4(2×C),32.0,31.8,30.8,30.4,29.4,21.3(2×C),19.6,18.1,17.5.16.5,15.2.Anal.Calcd.for C41H64O13:C,64.38；H,8.43.Found:C,64.60；H,8.83。薯蓣甙元-α-L-鼠李糖基-(1→2)-[4-O-乙酰基-α-L-鼠李糖基-(1→4)]-β-D-葡萄糖甙(Diosgenyl α-L-rhamnopyranosyl-(1→2)-[4-O-acetyl-α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside)4aRf:0.48[CH2Cl2-CH3OH(6∶1)]；[α]D=-100.8°(c,0.39,THF)；ESI-MS:911.9(M+1),933.8(M+Na),949.8(M+K)；1H-NMR(400MHz):6.40(s,1H),5.87(s,1H),5.79(t,1H,J=9.8Hz),5.24(d,1H,J=4.4Hz),4.98-4.90(m,3H),4.80(brs,1H),4.62-4.47(m,4H),4.40(t,1H,J=9.0Hz),4.33(t,1H,J=9.4Hz),4.21-4.17(m,3H),4.02(brd,1H),3.82(m,1H),3.58(brd,1H),3.53-3.43(m,2H),1.96(s,3H),1.71(d,3H,J=6.4Hz),1.31(d,3H,J=6.0Hz),1.07(d,3H,J=6.8Hz),0.98(s,3H),0.75(s,3H),0.62(d,3H,J=4.8Hz)；13C-NMR(100MHz):170.9(C=O),140.9,122.0,109.4,102.5,102.2,100.5,81.3,78.2,78.1(3×C),77.2,76.0,74.3,73.0,72.8,72.6,70.4,69.7,67.7,67.0,63.0,61.3,56.8,50.4,42.1,40.6,40.0,39.1,37.7,37.3,32.5,32.4,32.0,31.9,30.8,30.3,29.4,21.3(2×C),19.6,18.8,18.0,17.5,16.5,15.3.Anal.Calcd.for C47H74O17:C,61.96；H,8.19.Found:C,61.68；H,8.50.薯蓣甙元-α-L-鼠李糖基-(1→2)-[4-O-丁酰基-α-L-鼠李糖基-(1→4)]-β-D-葡萄糖甙(Diosgenylα-L-rhamnopyranosyl-(1→2)-[4-O-butyryl-α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside)4bRf:0.27[CH2Cl2-CH3OH(8∶1)]；[α]D=102.0°(c,0.36,THF)；ESI-MS:961.8(M+Na),978.1(M+K)；1H-NMR(400MHz):6.37(s,1H),5.84(s,1H),5.79(t,1H,J=9.7Hz),5.25(d,1H,J=4.3Hz),4.95-4.89(m,3H),4.76(d,1H,J=1.1Hz),4.60-4.45(m,4H),4.38(t,1H,J=8.8Hz),4.30(t-like,1H,J=9.9,9.0Hz),4.20-4.14(m,3H),4.02(brd,1H),3.82(m,1H),3.58(brd,1H),3.53-3.41(m,2H),2.28(t,2H,J=7.4Hz),1.70(d,3H,J=5.9Hz),1.32(d,3H,J=6.1Hz),1.07(d,3H,J=6.9Hz),0.99(s,3H),0.78(t,3H,J=7.6Hz),0.77(s,3H),0.63(d,3H,J=4.7Hz).Anal.Calcd.for C49H78O17·1.5H2O:C,60.91；H,8.45；.Found:C,60.77；H,8.52。薯蓣甙元-α-L-鼠李糖基-(1→2)-[4-O-己酰基-α-L-鼠李糖基-(1→4)]-β-D-葡萄糖甙(Diosgenyl α-L-rhamnopyranosyl-(1→2)-[4-O-caproyl-α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside)4cRf:0.29[CH2Cl2-CH3OH(8∶1)]；[α]D 21=-92.8。(c,0.40,THF)；ESl-MS:989.9(M+Na)；1H-NMR(400MHz):6.37(s,1H),5.84(s,1H),5.80(t,1H,J=9.9Hz),5.26(d,1H,J=4.0Hz),4.97-4.89(m,3H),4.76(m,1H),4.60-4.45(m,4H),4.38(t,1H,J=8.9Hz),4.30(t,1H,J=9.4Hz),4.20-4.14(m,3H),4.02(brd,1H),3.82(m,1H),3.58(brd,1H),3.54-3.41(m,2H),2.32(m,2H),1.70(d,3H,J=6.0Hz),1.34(d,3H,J=6.2Hz),1.07(d,3H,J=6.9Hz),0.99(s,3H),0.77(s,3H),0.69(t,3H,J=7.0Hz),0.63(d,3H,J=4.7Hz)Anal.Calcd.for C51H82O17:C,63.33；H,8.55.Found:C,63.08；H,8.63.薯蓣甙元-α-L-鼠李糖基-(1→2)-[4-O-乙烯基己二酰基-α-L-鼠李糖基-(1-4)]-β-D-葡萄糖甙(Diosgenyl α-L-rhamnopyranosyl-(1→2)-[4-O-vinyladipyl-α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside)4dRf:0.40[CH2Cl2-CH3OH(8∶1)]；[α]D 21=-99.7°(c,0.19,THF)；ESI-MS:1045.7(M+Na)；1H-NMR(400MHz):7.36(dd,1H,J=14.0,6.3Hz),6.36(s,1H),5.84(s,1H),5.78(t,1H,J=9.8Hz),5.26(d,1H,J=5.1Hz),4.95-4.88(m,3H),4.84(dd,1H,J=14.0,1.5Hz),4.76(m,1H),4.60-4.46(m,5H),4.37(t,1H,J=9.0Hz),4.30(t,1H,J=9.4Hz),4.20-4.14(m,2H),4.02(dd,1H,J=12.2,3.0Hz),3.82(m,1H),3.58(brd,1H),3.54-3.41(m,2H),2.34-2.26(m,4H),1.70(d,3H,J=6.2Hz),1.31(d,3H,J=6.2Hz),1.07(d,3H,J=7.0Hz),0.99(s,3H),0.77(s,3H),0.63(t,3H,J=5.6Hz).Anal.Calcd.forC53H82O19·2H2O:C,60.10；H,8.18.Found:C,60.24；H,7.86.薯蓣甙元-α-L-鼠李糖基-(1→2)-[3,5-O-二乙酰基-α-L-阿拉伯糖基-(1→4)]-β-D-葡萄糖甙(Diosgenyl α-L-rhamnopyranosyl-(1→2)-[3,5-O-di-acetyl-α-L-arabinofuranosyl-(1→4)]-β-D-glucopyranoside)5aRf:0.28[CH2Cl2-CH3OH(10∶1)]；[α]D=-92.7°(c,0.19,THF)；ESI-MS:961.8(M+Na),977.8(M+K)；1H-NMR(400MHz):6.30(s,1H),6.07(s,1H),5.45(dd,1H,J=5.0,1.8Hz),5.24(d,1H,J=4.8Hz),4.98(m,1H),4.93-4.86(m,3H),4.73(brd,1H),4.69(dd,1H,J=11.6,4.0Hz),4.57-4.47(m,3H),4.35-4.12(m,6H),3.81(m,1H),3.65(brd,1H),3.53-3.41(m,2H),1.89(s,3H),1.81(s,3H),1.71(d,3H,J=6.0Hz),1.07(d,3H,J=6.8Hz),0.99(s,3H),0.76(s,3H),0.62(d,3H,5.2Hz)；13C-NMR(100MHz):170.8(2×C=O),140.9,122.0,109.8,109.4,102.2,100.4,81.4,81.3(2×C),80.4,78.2,77.9(2×C),76.8(2×C),74.3,73.0,72.7,69.7,67.0,64.7,63.1,61.2,56.8,50.5,42.1,40.6,40.0,39.1,37.7,37.3,32.5,32.4,32.0,31.8,30.8,30.3,29.4,21.3,20.9,20.8,19.6,18.8,17.5,16.5,15.2.Anal.Calcd.for C48H74O18:C,61.39；H,7.94.Found:C,61.18；H,7.97.薯蓣甙元-α-L-鼠李糖基-(1→2)-[2,5-O-二乙酰基-α-L-阿拉伯糖基-(1→4)]-β-D-葡萄糖甙(Diosgenyl α-L-rhamnopyranosyl-(1→2)-[2,5-O-di-acetyl-α-L-arabinofuranosyl-(1→4)]-β-D-glucopyranoside)5bRf:0.20[CH2Cl2-CH3OH(10∶1)]；[α]D=-90.7°(c,0.18,THF)；ESI-MS:961.8(M+Na),977.8(M+K)；1H-NMR(400MHz):6.32(s,1H),5.88(s,1H),5.60(d,1H,J=2.4Hz),5.23(d,1H,J=4.4Hz),5.09(m,1H),4.93-4.86(m,2H),4.75(brs,1H),4.64(dd,1H,J=12.0,3.6Hz),4.57-4.54(m,2H),4.49-4.39(m,3H),4.33-4.25(m,3H),4.17-4.15(m,2H),3.80(m,1H),3.63(brd,1H),3.53-3.41(m,2H),1.87(s,3H),1.80(s,3H),1.70(d,3H,J=6.4Hz),1.07(d,3H,J=6.8Hz),0.98(s,3H),0.76(s,3H),0.62(d,3H,5.2Hz)；13C-NMR(100MHz):170.8(C=O),170.6(C=O),140.9,122.0,109.4,106.6,102.1,100.5,85.8,82.5,81.3,78.2,77.7(2×C),76.9,76.1(2×C),74.3,73.0,72.7,69.7,67.0,64.3,63.0,61.2,56.8,50.4,42.1,40.6,40.0,39.1,37.6,37.3,32.5,32.4,32.0,31.8,30.8,30.2,29.4,23.1,20.8(2×C),19.6,18.8,17.5,16.5,15.2.薯蓣甙元-4-O-乙酰基-α-L-鼠李糖基-(1→6)-β-D-葡萄糖甙(Diosgenyl 4-O-acetyl-α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside)6aRf:0.51[CH2Cl2-CH3OH(5∶1)]；[α]D=-75.3°(c,0.24,THF)；ESI-MS:765.7(M+1),787.6(M+Na),803.6(M+K)；1H-NMR(300MHz):5.67(t,1H,J=9.5Hz),5.41(s,1H),5.24(d,1H,J=4.9Hz),4.92(d,1H,J=7.7Hz),4.53-4.43(m,4H),4.20-3.84(m,6H),3.52-3.42(m,2H),1.95(s,3H),1.28(d,3H,J=6.3Hz),1.06(d,3H,J=6.9Hz),0.84(s,3H),0.75(s,3H),0.62(d,3H,J=5.5Hz)；13C-NMR(100MHz):170.9(C=O),141.2,121.8,109.5,103.2,102.5,81.3,79.1,78.7,76.9,75.9,75.4,72.4,71.9,70.4,68.5,67.2,67.0,64.6(CH3OH),63.0,56.8,50.4,42.1,40.6,40.0,39.7,37.7,37.2,32.4,32.3,32.0,31.8,30.8,30.6,29.4,21.3(2×C),19.6,18.3,17.5,16.5,15.2.Anal.Calcd.forC41H64O13·H2O:C,62.90；H,8.50.Found:C,63.16；H,8.82.
                        实施例6表3 乙酰化薯蓣皂甙衍生物中糖基部分13C的化学位移(δ)a
   1a      1b       2a       2b       3a       4a       5a       5bC-1’  103.0    102.7    100.8    100.2    102.3    100.5    100.4    100.5C-2’  75.3     75.0     79.5     79.4     75.9     78.1     77.9     77.7C-3’  78.8     69.7     77.8     76.9     76.8     78.1     77.9     77.7C-4’  71.6     79.2     71.7     71.6     77.6     78.1     76.8     76.0C-5’  75.4     73.2     75.0     75.1     77.4     77.2     76.8     76.9C-6’  65.0     64.5     64.7     64.6     61.5     61.3     61.2     61.2C-1r                   102.3    101.7    102.7    102.2    102.2    102.1C-2r                   72.7     72.6     72.7     72.8     72.7     72.7C-3r                   73.0     70.4     70.5     73.0     73.0     73.0C-4r                   74.3     76.3     76.2     74.3     74.3     74.3C-5r                   69.8     67.0     67.6     69.7     69.7     69.7C-6r                   18.9     18.2     18.1     18.8     18.8     18.8C-                                                102.5    109.8    106.61r’(1a)C-                                                72.6     80.4     85.82r’(2a)C-                                                70.4     81.4     76.93r’(3a)C-                                                76.0     81.3     82.54r’(4a)C-                                                67.0     64.7     64.35r’(5a)C-6r’                                            18.1a，此化学位移的归属是经1H-NMR,13C-NMR,1H-1HCOSY,TOCSY,HSQC谱综合分析的结果
       实施例7  抗肿瘤生物活性体外筛选实验
结果评定；无效：10-5mol/1＜85％
          弱效：10-5mol/l≥85％或10-6mol/l≥50％
          强效：10-6mol/l≥85％或10-7mol/l＞50％
                        表4筛选方法：磺酰罗丹明B(sulforhodamine B,SRB)蛋白染色法细胞株：A-549人肺腺癌    作用时间：72h
           对肿瘤细胞生长的抑制率％
皂甙     A-549 评价
 10-4  10-5 10-6  10-7  10-8
 2a  100  3.6  8.1  4.3  4.2 无效
 2b  100  93.0  7.1  8.7  4.4 弱效
3a  100  14.8  9.1  7.4  3.4 无效
4a  100  99.0  4.7  8.3  7.8 弱效
5a  100  100  26.4  5.2  5.0 弱效
5b  99.8  100  5.9  1.3  0 弱效
                        表5筛选方法：四氮唑盐(microculture tetrozolium,MTT)还原法细胞株：P388小鼠白血病    作用时间：48h
             对肿瘤细胞生长的抑制率％
皂甙     P-388 评价
 10-4  15-5  10-6  10-7  10-8
 2a  99.4  9.2  13.7  0  0 无效
 2b  99.2  37.5  2.3  0  0 无效
 3a  97.6  0  0  0  0 无效
 4a  99.3  97.8  0  0  0 弱效
 5a  98.7  98.6  24.9  11.0  10.7 弱效
 5b  99.3  98.2  0  0  0 弱效

Claims (10)

1、一种薯蓣皂甙的酰基化衍生物，其特征是其结构式如下：，结构式中，R=C1-8的酰基或烯酰基；r,p,q=0、1或2，且，0＜r+p+q≤2；M=葡萄糖基(
Figure A0111287000022
)；N,U=鼠李糖基(
Figure A0111287000023
)或阿拉伯糖基(
Figure A0111287000024
)；n,u=0或1。＜
2、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物，其特征是所述的结构式如下：
Figure A0111287000025
其中R1或R2=H或R,R1和R2不能同时等于H,R如权利要求1所述。
3、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物，其特征是所述的结构式如下：其中R1或R2=H或R,R1和R2不能同时等于H,R如权利要求1所述。
4、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物，其特征是所述的结构式如下：
Figure A0111287000027
其中R如权利要求1所述。
5、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物，其特征是所述的结构式如下：，其中R1、R2或R3=H或R,R1、R2和R3不能同时等于H,R如权利要求1所述。
6、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物，其特征是所述的结构式如下：
Figure A0111287000032
其中R1、R2或R3=H或R,R1、R2和R3不能同时等于H,R如权利要求1所述。
7、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物，其特征是所述的结构式如下：其中R如权利要求1所述。
8、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物的酶催化合成方法，其特征是在极性溶剂中和反应温度为室温-100℃时，酰基受体、酰基给体与脂肪酶的摩尔比依次为1-7∶100-210∶0.02-0.4，反应1小时-120小时，所述的酰基受体薯蓣皂甙结构式如下：
Figure A0111287000034
结构式中，M=葡萄糖基
Figure A0111287000035
N,U=鼠李糖基 或阿拉伯糖基 n,u=0或1，所述的酰基给体分子式为ROR’，其中R=C1-8的酰基或烯酰基，R’为乙烯基或三氟乙基，所述的脂肪酶是脂肪酶CRL、脂肪酶PPL、脂肪酶MJL、脂肪酶WGL、脂肪酶PFL或脂肪酶Novozyme435。
9、如权利要求3所述的一种薯蓣皂甙的酰基化衍生物的酶催化合成方法，其特征是所述的酰基受体薯蓣皂甙是薯蓣甙元-β-D-葡萄糖甙、薯蓣甙元-α-L-鼠李糖基-(1→2)-β-D-葡萄糖甙、薯蓣甙元-α-L-鼠李糖基-(1→4)-β-D-葡萄糖甙、薯蓣甙元-α-L-鼠李糖基-(1→2)-[α-L-鼠李糖基-(1→4)]-β-D-葡萄糖甙、薯蓣甙元-α-L-鼠李糖基-(1→2)-[α-L-阿拉伯糖基-(1→4)]-β-D-葡萄糖甙或薯蓣甙元-α-L-鼠李糖基-(1→6)-β-D-葡萄糖甙。
10、如权利要求1所述的一种薯蓣皂甙的酰基化衍生物的用途，其特征是用于抗肿瘤的药物。
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CN101787069A (zh) * 2010-03-22 2010-07-28 四川大学 薯蓣皂苷元哌嗪类衍生物及其制备方法
WO2013185613A1 (zh) 2012-06-13 2013-12-19 杭州本生药业有限公司 重楼皂苷i的酰化衍生物、及其制备方法和应用
CN104334571A (zh) * 2012-06-13 2015-02-04 杭州本生药业有限公司 重楼皂苷i的酰化衍生物、及其制备方法和应用
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Publication number Priority date Publication date Assignee Title
CN101787069A (zh) * 2010-03-22 2010-07-28 四川大学 薯蓣皂苷元哌嗪类衍生物及其制备方法
WO2013185613A1 (zh) 2012-06-13 2013-12-19 杭州本生药业有限公司 重楼皂苷i的酰化衍生物、及其制备方法和应用
CN104334571A (zh) * 2012-06-13 2015-02-04 杭州本生药业有限公司 重楼皂苷i的酰化衍生物、及其制备方法和应用
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CN104334571B (zh) * 2012-06-13 2016-08-31 杭州本生药业有限公司 重楼皂苷i的酰化衍生物、及其制备方法和应用
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