Suspension culture establishment and alkaloids analysis of tubercles of Pinellia ternata | 半夏小块茎悬浮培养及其生物碱类化合物的测定

Chinese. 筛选半夏小块茎悬浮培养过程中诱导和继代的最佳培养基，并对小块茎中生物碱类化合物的含量进行测定，为半夏生物碱的生产及其次生代谢物的调控提供参考。在添加不同激素的MS培养基上诱导、增殖半夏小块茎，并建立未分化小块茎的液体悬浮培养体系，采用酸性染料比色法测定小块茎的总生物碱含量，高效液相色谱法测定小块茎中鸟苷、肌苷和葫芦巴碱的含量。以块茎、叶片叶柄为外植体诱导半夏小块茎的最佳培养基分别为 MS + 0.5 mg/L NAA + 1.0 mg/L 6-BA 和MS + 0.2 mg/L NAA + 1.0 mg/L 6-BA；小块茎最适继代培养基为固体1/2 MS + 0.6 mg/L ABA;小块茎的液体悬浮培养基为液体1/2 MS + 0.6 mg/L ABA。与栽培块茎相比，以块茎及叶片叶柄为外植体诱导的半夏小块茎总生物碱含量分别提高了4.46倍和2.07倍。以块茎为外植体诱导的半夏小块茎中，鸟苷和葫芦巴碱的含量分别是栽培块茎的10.6和2.5倍；而肌苷含量降低，约是栽培块茎的1/3。以叶片叶柄为外植体诱导的半夏小块茎中，鸟苷和葫芦巴碱的含量分别比栽培块茎增加了1.1和1.9倍，肌苷含量约为栽培块茎的1/4。半夏组培小块茎可作为一种模式材料，用于半夏生物碱的生产以及鸟苷、肌苷和葫芦巴碱的代谢调控。

English. The induction and subculture media to establish suspension cultures of Pinellia tubercles were selected. Alkaloids in tubercles were examined. The objective was to provide evidence for Pinellia tubercles on alkaloids production or metabolism regulation. The induction, propagation and suspension culture establishment of tubercles were investigated on MS medium supplemented with various hormones. Total alkaloids were determined using the method of uvvis spectrophotometry. The contents of guanosine, inosine and trigonelline in tubercles and field grown tuber were determined by HPLC. The suitable media to induce tubercles from tuber as well as leaf and petiole explants were MS medium containing 0.5 mg/L NAA and 1.0 mg/L 6-BA and MS medium containing 0.2 mg/L NAA and 1.0 mg/L 6-BA, respectively. The suitable medium to subculture tubercles was solid one-half strength MS medium containing 0.6 mg/L ABA. Suspension culture of tubercles was established in liquid one-half strength MS medium containing 0.6 mg/L ABA. Total alkaloids contents in tubercles induced from tuber explant, tubercles induced from leaf and petiole explant were 4.46 and 2.07 times higher than that in field-grown tubers respectively. The contents of guanosine and trigonelline in tuber derived tubercles were 10.6 and 2.5 times higher respectively than that in field-grown tubers, while inosine content was about one third of that in field-grown tubers. The contents of guanosine and trigonelline in leaf and petiole derived tubercles were 1.1 and 1.9 times higher respectively than that in field-grown tubers, while inosine content was about one fourth of that in field-grown tubers. Pinellia tubercles not only can serve as a novel material for alkaloids production, but also for guanosine, inosine and trigonelline metabolism regulation.