离子选择性微电极用于原位测量离子扩散系数

扩散系数是描述物质扩散过程的重要参数,而用膜池法、放射性或荧光示踪法、分子动力学模拟等现有方法无法原位进行生物体系中离子扩散系数的实时测量。 本文利用离子选择性微电极响应迅速、高选择性、高灵敏度、高空间分辨率、对样品无污染等优势,通过分析单个植物细胞原生质体在培养液中破裂时所形成的离子浓度脉冲信号,建立了相应的点源扩散模型,推导出了描述离子浓度随时间变化的理论公式,并通过该公式对实验测得的脉冲信号进行拟合,得到了离子的扩散系数,从而建立了一种用离子选择性微电极原位测定离子扩散系数的新方法,并将其应用于芦荟细胞原生质体破裂时离子扩散系数的测定,得到了Ca²⁺、Na⁺和K⁺的扩散系数分别为(6.51±0.12)×10^-6、(2.93±0.15)×10^-5和(3.03±0.35)×10^-5 cm²/s。 对比发现,拟合得到的Ca²⁺、Na⁺和K⁺扩散系数均略高于已报道的数值(纯水中),这一现象的产生可能是因为原生质体是在低渗液中吸水膨胀,细胞膜内压力升高产生内外压力差,该压力差会加速细胞破裂时离子的扩散。 这一方法对生物体系无干扰,较好地解决了生物体系中离子扩散系数原位实时测量的难题。

An Ion-Selective Microelectrode Method for In-situ Measurement of the Diffusion Coefficients of Ions

Diffusion coefficient is an important parameter describing the diffusion process of a substance. However, the existing methods such as the membrane pool method, radioactive or fluorescent tracer method, and molecular dynamics simulation cannot be used to measure the ion diffusion coefficient in the biological system in real time. The ion-selective microelectrode has the advantages of rapid response, high selectivity, high sensitivity, high spatial resolution, and no pollution to the sample. Using the advantages of microelectrodes, this paper established the corresponding point source diffusion model by analyzing the ion concentration pulse signal formed by the rupture of protoplasts of single plant cells in culture medium, and derived the theoretical formula describing the ion concentration change with time. By fitting the experimentally measured pulse signal to obtain the diffusion coefficient of ions, a new method for in situ determination of ion diffusion coefficient by ion-selective microelectrode was established and applied to aloe cell protoplasts. When the ion diffusion coefficient is measured during the body rupture, the diffusion coefficients of Ca²⁺, Na⁺ and K⁺ are (6.51±0.12)×10^-6 cm²/s, (2.93±0.15)×10^-5 cm²/s and (3.03±0.35)×10^-5 cm²/s, respectively. The results show that the Ca²⁺, Na⁺, and K⁺ diffusion coefficients obtained are slightly higher than those reported values (in pure water). This phenomenon might be caused by the increase of the intracellular pressure of the protoplasts in the hypotonic fluid. The increased pressure might have accelerated the diffusion of ions when the cell ruptured. This method does not interfere with the biological system, and better solves the problem of in-situ real-time measurement of ion diffusion coefficient in biological systems.