[发明专利]中药特征指纹谱的建立及其应用

摘要

本发明涉及中药技术领域。具体涉及一种中药特征指纹谱的建立及其应用。用本发明方法建立的特征指纹谱不仅能用于含有相同有效成分的不同中药间区分和鉴别，并能用于品种的鉴定，还能为中成药所含药味的鉴定提供依据。

主权项

1.一种中药特征指纹谱的建立，其特征在于该指纹谱的建立包括下列步骤：(1)标准药材的选取，收集业经生药鉴定的该品种药材五个以上的批号，能有更多的批号则更佳，因为考虑到生物样品的个体差异，只有在相当数量的样品中，才能清楚地显现出它的特征，此时还需注意中药样品批号的含义，除样品的产地、采收时间和季节，以及药用部位外，气侯的影响，储运保管条件以及某些人为因素均应顾及，因此样品批号宜多不宜少；(2)规范化的操作条件和步骤：应从两方面的工作于以规范化，一是样品的预处理，生物样品都需经预处理后方能进行色谱分析，预处理的方法有多种，例如超声萃取，加热萃取，pH萃取等各种萃取技术，二是萃取样品量的大小和样品粉碎程度，以及萃取液是否再经处理等因素，总之所有样品处理过程都必须保持一致以保证样品液的分析具有良好的精密度、正确性、重现性和相互间的可比性；(3)最佳色谱条件的建立：色谱条件包括色谱柱、流动相、检测器等的选择，要建立良好色谱条件使中药样品中所含之成分尽可能地获得分离，也即分得的色谱峰愈多愈好，可以说是多多益善，使中药的内在特性都显现出来，为鉴定提供足够的信息，色谱柱的选择可根据中药所含的主要成分而定，例如含生物碱类化合物为主的中药，可选用离子交换柱；含多糖类化合物为主的中药可选用凝胶柱，含甾体类化合物为主的中药可选用c-18反相柱；......等，流动相的组成除应考虑化合物的性质外，因为中药所含成分多而杂，采用常规的等梯度洗脱难以使各种性质的成分获得分离，使色谱图中所出现的峰数较少，不能给出足够的鉴定所需的信息，在大多数的情况下以采用梯度洗脱为宜，在合适的梯度条件下，使性质差异较大的各种化合物均被分离，可充分提供鉴定所需的信息，分得的色谱峰越多越有利于鉴定，目前可供选择的检测器，在气相色谱中有氢火焰离子化检测器(FID)、热导检测器(TCD)、电子捕获检测器(ECD)、火焰光度检测器(FPD)等，其中以FID最为常用，凡以碳、氢、氧等元素组成的有机化合物均可被检出，对中药所含之挥发性成分的分析是十分有效的，在高效液相色谱中有可变紫外-可见光(UV-VIS)检测器、荧光检测器、电化学检测器、示差折射检测器(RID)、蒸发光散射光检测器(ELSD)等，其中紫外-可见光检测器用得较多，适宜于化合物中有共轭双键的成分分析，而大量的不含共轭双键的化合物不能被检出，近十年来，ELSD检测器的发展，使中药中甾体类、皂甙类、多糖类等化合物均可采用ELSD作检测；(4)特征指纹的确立：对上述五个以上批号的标准药材，在优化的提取条件下制备供试液，以及在最佳色谱条件下进行分析，每个样品同品种的中药五个以上的批号，分别进样三次，由此获得五个批号样品的十五张色谱图，并转化为十五张色谱指纹谱，现将这十五张色谱指纹谱中的相对保留值汇集并列成表，以便选择特征色谱峰来建立特征指纹。表I色谱指纹谱汇集表(面积归一化值)药材α0.0840.1000.1090.1280.1410.1680.1980.2220.2520.2840.4040.4460.49610.330.150.361.040.891.301.120.600.740.450.9620.450.380.381.060.821.261.080.490.730.401.0130.260.240.451.100.841.311.070.480.750.511.1540.330.290.890.780.981.010.710.690.4950.150.470.340.920.700.961.050.650.720.520.9960.400.260.900.680.990.990.730.700.510.9770.070.750.320.411.110.920.900.980.570.810.390.930.8980.030.680.280.371.090.880.940.960.640.780.430.880.9690.700.090.341.060.950.940.960.640.760.450.751.03100.890.210.220.840.761.151.180.700.770.571.08110.170.720.180.270.900.881.181.170.650.710.501.12120.040.770.160.180.920.801.201.170.660.730.520.920.96130.220.290.900.861.241.180.680.750.440.860.88140.010.300.480.960.871.241.190.670.790.420.941.00150.360.500.990.821.201.230.650.780.411.05共有样品数96151515151515151515614药材α0.5480.5760.6840.8040.8951.0001.1271.2891.3561.4191.5061.5731.6071.79010.320.281.0001.010.030.301.6720.540.211.0000.930.070.281.541.3130.400.251.0000.900.050.321.601.2841.940.201.0000.880.401.810.021.3551.880.241.0000.780.381.760.031.3062.000.271.0000.840.411.700.011.3270.401.560.301.0000.940.351.591.4480.321.660.261.0001.040.361.631.4190.381.660.251.0000.610.371.671.29100.771.0000.210.670.281.561.48110.230.701.0000.180.730.301.651.45120.270.751.0000.120.860.291.711.47130.261.0000.870.310.321.781.35140.201.0001.010.310.341.761.37150.221.0001.040.330.351.741.33共有样品数36314315315331515314药材α1.8201.8822.0022.0562.1662.2222.3002.3972.5602.6702.7352.9663.0213.1403.23810.661.5301.440.110.340.380.2420.630.541.5901.500.100.360.410.020.2830.650.551.5301.460.090.380.420.2240.770.740.601.4801.250.110.370.4550.750.730.581.410.080.400.4360.680.760.571.4501.420.090.350.4170.780.630.041.4301.480.100.390.3980.800.621.4001.490.080.110.330.410.0190.810.591.5501.510.100.340.410.02100.630.100.661.5701.430.080.380.35110.650.111.6001.450.070.350.38120.610.150.681.5801.460.100.400.39130.600.681.5001.211.510.090.160.330.401463.000.671.5101.151.530.080.130.370.430.06150.631.531.201.500.100.150.360.43共有样品数3143131143151153151534药材α3.2983.3843.4603.5263.5803.7423.7893.8223.9024.0684.1794.4364.5034.57910.671.100.580.500.0620.650.091.140.570.560.040.780.050.0330.680.111.190.600.530.030.7740.660.611.7400.7950.670.601.7000.270.800.9360.690.591.7600.760.8070.660.570.310.440.7880.650.560.290.7790.670.580.280.470.79100.600.760.01110.700.610.030.81120.690.600.010.79130.670.590.160.78140.710.600.180.76150.690.610.130.77共有样品数14231533254314311药材α4.6325.1755.5305.6505.76412.140.1222.180.0832.200.0942.080.6652.100.6762.140.6371.960.0781.980.0492.060.10100.042.10110.031.940.83120.011.990.88132.00142.09152.07共有样品数315533总峰数：61