在线超声提取96-孔盘技术高通量快速测定全叶马兰中果聚糖成分

摘要

An on-line method was developed for the extraction and determination of fructans in plant samples. Fructans was extracted with water by ultrasound assisted, and directly introduced into a well of 96-well format which was reacted with concentrated sulphuric acid and 5％ phenol for fast quantifying fructans. Response surface methodology (RSM) along with Box-Behnken design (BBD)was applied to optimize the critical variables of the extraction conditions of dynamic ultrasoundassisted extraction (DUAE) system for fructans yield from Kalimeris integrifolia. Three factors such as ultrasonic power, flow rate of solventand volume of solvent were investigated. The optimum extraction conditions were ultrasonic power 496 W, flow rate of solvent 0.63 mL/min, and volume of solvent 4.69 mL. Under these conditions, the maximum fructans yield was 17.60％. In addition, the single-factor-test was employed to optimize the major experimental parameters for the phenol-sulphuric acid procedure on 96-well microplate. The calibration curves of the proposed method under the optimised conditions corresponded very closely to a linear relationship (R≥0.99) over the concentration range from 20 to 100 mg/L for inulin. RSD of the method were 2.9％-4. 1％, respectively. In this study, only small amount of sample, i.e. 5 mg, can meet the demand of the extraction. All of the analysis steps were completed in less than 15 min, and 96 samples could be analysed in 1 h using a 96-well format.%采用在线动态超声提取装置对全叶马兰果聚糖成分进行提取,用效应面方法优化了在线提取条件.提取溶剂为纯水,待测样品为5 mg,提取条件为超声功率是496 W,溶剂流速 0.63 Ml/min,溶剂体积 4.69 Ml;用96-孔盘联合酚硫酸程序对果聚糖进行高通量快速检测,果聚糖的加标回收率为98.3%～105.7%,相对标准偏差为2.9%～4.1%,全叶马兰果聚糖含量为17.60%.15 min内可分析24个样品.试剂用量较常规比色分析减少了95%.本方法快速、准确,特别适合天然植物资源生态学研究中数以百计的大样本统计调查.