Kolb N, Herrera J L, Ferreyra D J, Uliana R F
Facultad de Ciencias Exactas, Químicas y Naturales, Universidad Nacional de Misiones, Felix de Azara 1552, 3300 Posadas (Misiones), Argentina.
J Agric Food Chem. 2001 Oct;49(10):4538-41. doi: 10.1021/jf010475p.
An improved analytical method was developed which may be applied to quality control of stevioside and rebaudioside A contents in dried leaves of Stevia rebaudiana before processing; in a selective sampling program searching for plants of higher yield in diterpene glycosides content; or when a large number of samples are sent to the laboratory for analysis. The procedure developed involves two steps: solvent extraction followed by an isocratic HPLC analysis. The sample, 1 g of dried leaves of S. rebaudiana, is ground and solvent-extracted with EtOH 70% (w/w) in Erlenmeyer flasks by shaking for 30 min in a 70 degrees C water bath. After the extract was cooled, it was filtered and analyzed by HPLC using an NH(2) column (250 x 4.6 mm) and a mixture of acetonitrile/water (80:20, v/v) as mobile phase, pH 5 adjusted with acetic acid. The detection was in the UV range at 210 nm (0.04 AUFS). Quantitation was performed by means of an external standard calibration curve for each analyte which had been obtained from standard solutions of pure stevioside and rebaudioside A. Working under these conditions there were no observed interference effects. The method saves time in sample preparation, and reduces sample handling and chromatographic analysis time, while having little loss of precision [coefficient of variation (CV%) between 1.8% and 3.0%] and recovery [between 98.5% and 100.5%]. The method was applied to 30 samples of S. rebaudiana from Misiones (Northeastern Argentina), and the stevioside content found ranged between 3.78 and 9.75% (weight) whereas Rebaudioside A content ranged between 1.62 and 7.27% (weight).
开发了一种改进的分析方法，该方法可应用于甜叶菊干叶加工前甜菊糖苷和莱鲍迪苷A含量的质量控制；用于在选择性采样程序中寻找二萜糖苷含量较高的植物；或者当大量样品被送到实验室进行分析时。所开发的程序包括两个步骤：溶剂萃取，然后进行等度高效液相色谱分析。取1克甜叶菊干叶样品研磨后，在锥形瓶中用70%（w/w）乙醇在70℃水浴中振荡萃取30分钟。萃取液冷却后过滤，采用氨基柱（250×4.6毫米），以乙腈/水（80:20，v/v）混合物为流动相，用乙酸调节pH值至5，通过高效液相色谱进行分析。检测在210纳米的紫外范围内（0.04 AUFS）。通过从纯甜菊糖苷和莱鲍迪苷A标准溶液获得的每种分析物的外标校准曲线进行定量。在这些条件下工作，未观察到干扰效应。该方法节省了样品制备时间，减少了样品处理和色谱分析时间，同时精度损失很小[变异系数（CV%）在1.8%至3.0%之间]，回收率在98.5%至100.5%之间。该方法应用于来自米西奥内斯（阿根廷东北部）的30个甜叶菊样品，发现甜菊糖苷含量在3.78%至9.75%（重量）之间，而莱鲍迪苷A含量在1.62%至7.27%（重量）之间。
