Evaluation of [Co(gly)3]- as a 35Cl- NMR shift reagent for cellular studies.
We studied the efficacy of the tris-glycinatocobaltate(II) complex ([Co(gly)(3)](-)) as a shift reagent (SR) for chloride by (35)Cl NMR spectroscopy and compared to that of Co(2+)((aq)). Due to the relatively low thermodynamic stability of [Co(gly)(3)](-), a 1:3 Co(II)/gly stoichiometric solution at physiological pH is approximately a 2:1 mixture of [Co(gly)(2)(H(2)O)(2)] and [Co(gly)(H(2)O)(4)](+). This SR was found to be stable up to higher pH values than Co(2+)((aq)), better preventing Co(OH)(2) formation at alkaline pH. No significant differences in the (35)Cl(-) NMR chemical shift induced by Co(II)/gly or Co(2+)((aq)) were observed in the presence of physiological concentrations of either Ca(2+) or Mg(2+), or of either Na(+) or K(+). Although Co(2+)((aq)) was almost twice as effective as Co(II)/gly in shifting the (35)Cl(-) NMR resonance at the same high rho ([SR]/[Cl(-)]) value and low ionic strength, Co(2+)((aq)) showed a significant decrease (p < 0.05) in the (35)Cl(-) chemical shift at higher ionic strength. Line widths at half-height were significantly (p < 0.05) less for Co(II)/gly than for Co(2+)((aq)) at rho values in the range 0.066-0.40. Intracellular chloride was clearly detectable by (35)Cl NMR spectroscopy in human skin fibroblast cells suspended in medium containing 40 mM Co(II)/gly SR. We determined that, although Co(2+)((aq)) provides a larger shift than Co(II)/gly at the same rho value, there are significant advantages for using Co(II)/gly, such as pH stability, ionic strength independent chemical shifts, narrow (35)Cl(-) NMR resonances, and reduced cellular toxicity, as a SR in biological systems.
Diven
Conrad F
CF
Department of Chemistry, Loyola University Chicago, 6525 North Sheridan Road, Chicago, Illinois 60626, USA.
Wang
Fei
F
Abukhdeir
Abde M
AM
Salah
Wajeeh
W
Layden
Brian T
BT
Geraldes
Carlos F G C
CF
Mota de Freitas
Duarte
D
D002478:Cells, Cultured / Q000187:drug effects; D002712:Chlorides / Q000737:chemistry*; D003035:Cobalt / Q000737:chemistry*; D005347:Fibroblasts / Q000187:drug effects; D005998:Glycine / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents / Q000737:chemistry; D007554:Isotopes / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D013816:Thermodynamics
D002712:Chlorides; D007202:Indicators and Reagents; D007554:Isotopes; D003035:Cobalt; D005998:Glycine