The potential of pharmacologically active secondary plant metabolites is limited by the low yield from often rare plants, and the lack of economically feasible chemical synthesis of these complex compounds. Plant cell fermentation offers an alternative strategy to overcome these constraints. However, the efficiency of this approach is limited by intracellular sequestration of the products, such that continuous bioprocessing is not possible. As a precondition for such a, more attractive, continuous process, it is of great importance to stimulate the export of the product into the medium without impairing viability and, thus, the productivity of the cells. Using nicotine alkaloids of tobacco as a case study, an alternative strategy is explored, where nanosecond pulsed electric fields (nsPEFs) are applied for the efficient downstream recovery of the products. To maintain cell viability and allow for the further use of biomass, cells were exposed to strong (1–20 kV·cm−1), but very short (10–100 ns) electric pulses, which leads to a temporary permeabilisation of cell membranes. Using two transgenic cell lines, where two key genes involved in the metabolism of the anti-Alzheimer compound nornicotine were overexpressed, we could show that this nsPEF treatment improved the partitioning of some nicotine alkaloids to the culture medium without impairing viability, nor the synthesis of alkaloids. However, this release was only partial and did not work for nornicotine. Thus, nsPEFs produced a fractionation of alkaloids. We explain this electrofractionation by a working model considering the differential intracellular compartmentalization of nicotineic alkaloids.

中文翻译：

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纳秒脉冲电场提高植物细胞发酵中的产品回收率

具有药理活性的次生植物代谢物的潜力受到通常稀有植物的低产量以及这些复杂化合物缺乏经济可行的化学合成的限制。植物细胞发酵提供了一种替代策略来克服这些限制。然而，这种方法的效率受到产品的细胞内隔离的限制，因此无法进行连续的生物加工。作为这种更具吸引力的连续过程的先决条件，在不损害细胞活力的情况下刺激产品输出到培养基中是非常重要的，因此也不会损害细胞的生产力。使用烟草的尼古丁生物碱作为案例研究，探索了一种替代策略，其中应用纳秒脉冲电场 (nsPEF) 以实现产品的高效下游回收。为了保持细胞活力并允许进一步利用生物质，细胞暴露于强（1-20 kV·cm-1）但非常短（10-100 ns）的电脉冲，这会导致细胞膜的暂时透化. 使用两种转基因细胞系，其中参与抗阿尔茨海默病化合物降烟碱代谢的两个关键基因被过表达，我们可以证明这种 nsPEF 处理改善了一些尼古丁生物碱在培养基中的分配，而不会损害生存能力，也不会影响生物碱。然而，这个版本只是部分的，对去甲尼古丁不起作用。因此，nsPEFs 产生了生物碱的分馏。