Katsu T, Nakagawa H, Kanamori T, Kamo N, Tsuchiya T
Faculty of Pharmaceutical Sciences, Okayama University, Tsushima, Okayama 700-8530, Japan.
Anal Chem. 2001 Apr 15;73(8):1849-54. doi: 10.1021/ac001090t.
A triethylammonium-sensitive electrode was constructed using sodium tetrakis[3,5-bis(2-methoxyhexafluoro-2-propyl)phenyl]borate as an ion-exchanger and benzyl 2-nitrophenyl ether as a solvent mediator in a poly(vinylchloride) membrane matrix and was used to determine the pH difference across a cell membrane. The method is based on monitoring of the pH gradient-induced uptake of triethylammonium in situ. The triethylammonium electrode exhibited a near-Nernstian response to triethylammonium in the concentration range of 5 x 10(-6)-1 x 10(-2) M with a slope of 58.5 mV per concentration decade in a buffer solution composed of 150 mM NaCl and 10 mM NaH2PO4/Na2HPO4 (pH 7.5). The limit of detection was 1 microM. In experiments using liposomes, the uptake of triethylammonium into liposomes was quantitatively induced according to the pH difference across the liposomal membrane. The transmembrane pH differences in Escherichia coli cells and the light-induced pH differences across the envelope vesicles of Halobacterium halobium were successfully determined by the present method.
以四[3,5-双(2-甲氧基六氟-2-丙基)苯基]硼酸钠作为离子交换剂、苄基2-硝基苯基醚作为溶剂介导剂，在聚氯乙烯膜基质中构建了一种对三乙铵敏感的电极，并用于测定细胞膜两侧的pH差值。该方法基于对pH梯度原位诱导的三乙铵摄取的监测。在由150 mM氯化钠和10 mM磷酸二氢钠/磷酸氢二钠（pH 7.5）组成的缓冲溶液中，三乙铵电极在5×10⁻⁶ - 1×10⁻² M的浓度范围内对三乙铵表现出近能斯特响应，每浓度 decade 的斜率为58.5 mV。检测限为1 μM。在使用脂质体的实验中，三乙铵进入脂质体的摄取根据脂质体膜两侧的pH差值被定量诱导。通过本方法成功测定了大肠杆菌细胞中的跨膜pH差值以及嗜盐菌包膜囊泡中的光诱导pH差值。
