Improved measurements of Na+ fluxes in plants using calixarene-based microelectrodes.
Ion-selective microelectrodes are a powerful tool in studying adaptive responses of plant cells and tissues to various abiotic stresses. However, application of this technique in Na(+) flux measurements was limited due to poor selectivity for Na(+) ions of commercially available Na(+) cocktails. Often, these cocktails cannot discriminate between Na(+) and other interfering ions such as K(+) and Ca(2+), leading to inaccurate measurements of Na(+) concentration and, consequently, inaccurate Na(+) flux calculations. To overcome this problem, three Na(+)-selective cocktail mixtures were prepared using tetramethoxyethyl ester derivative of p-t-butyl calix[4]arene. These cocktail mixtures were compared with commercially available ETH 227-based Na(+) cocktail for selectivity for Na(+) ions over other ions (particularly K(+) and Ca(2+)). Among the three calixarene-based Na(+) cocktails tested, cocktail 2 [in % w/w: Na(+) ionophore (4-tert-butylcalix[4]arene-tetra acetic acid tetraethyl ester) 3.5, the plasticizer (2-nitrophenyl octyl ether) 95.9 and lipophilic anion (potassium tetrakis (4-chlorophenyl) borate) 0.6] showed the best selectivity for Na(+) ions over K(+) and Ca(2+) ions and was highly stable over time (up to 10h). Na(+) flux measurements under a wide range of NaCl concentrations (25-150 mM) using Na(+) cocktail 2 established a clear dose-response relationship between severity of salt stress and magnitude of Na(+) influx at the distal elongation and mature zones of Arabidopsis thaliana roots. Furthermore, Na(+) cocktail 2 was compared with commercially available ETH 227-based Na(+) cocktail by measuring Na(+) fluxes at the two Arabidopsis root zones in response to 100mM NaCl treatment. With calixarene-based Na(+) cocktail 2, a large decreasing Na(+) influx (0-15 min) followed by small Na(+) influx (15-45 min) was measured, whereas with ETH-based Na(+) cocktail Na(+) influx was short-lived (1-3 min) and was followed by Na(+) efflux (3-45 min) that might have been due to K(+) and Ca(2+) efflux measured together with Na(+) influx. In conclusion, Na(+)-selective calixarene-based microelectrodes have excellent potential to be used in real-time Na(+) flux measurements in plants.
School of Earth and Environment, The University of Western Australia, M087, Crawley, WA 6009, Perth, Australia. j.maheswari@gmail.com
Maheswari
M
Jayakannan
Olga
O
Babourina
Zed
Z
Rengel
D000081:Acetamides; D017360:Arabidopsis / Q000737:chemistry* / Q000502:physiology; D002118:Calcium / Q000032:analysis / Q000378:metabolism; D002138:Calibration; D047250:Calixarenes; D017136:Ion Transport; D017736:Ion-Selective Electrodes; D007476:Ionophores; D008839:Microelectrodes; D018517:Plant Roots / Q000737:chemistry / Q000502:physiology; D011188:Potassium / Q000032:analysis / Q000378:metabolism; D036226:Seedlings / Q000737:chemistry / Q000502:physiology; D012964:Sodium / Q000032:analysis* / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D013997:Time Factors
D000081:Acetamides; D007476:Ionophores; D047250:Calixarenes; D012965:Sodium Chloride; C050214:ETH 227; D012964:Sodium; D011188:Potassium; D002118:Calcium