Use of membrane spin label spectra to monitor rates of reaction of partitioning compounds: hydrolysis of a local anesthetic analog.
ESR spectra of membrane spin probes are conventionally used to obtain structural information. Here we show, for the first time, that when a membrane-soluble compound undergoes a chemical reaction, time-dependent changes in the ESR spectra of membrane spin probes can yield information about the kinetics of reaction. A benzoic acid ester, analog of the local anesthetic tetracaine, partitions between aqueous and membrane phases, causing changes in membrane structure as monitored by the ESR spectra of a probe. At alkaline pH, the lineshapes are time-dependent and the spectra go back to that in the absence of drug. The changes follow pseudo-first order kinetics. This effect is due to drug hydrolysis leading to water-soluble products, as confirmed by direct spectrophotometric measurements of the reaction. The pseudo-first order rate constants found by the latter method are in very good agreement with those calculated by ESR. The rate of hydrolysis decreases with increasing membrane concentration. This phenomenon accounts in part for the increased potency and toxicity of the more membrane-soluble local anesthetics.
Bianconi
M L
ML
Department of Biochemistry, Universidade de São Paulo, Brazil.
do Amaral
A T
AT
Schreier
S
S
D003497:Cyclic N-Oxides; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008051:Lipid Bilayers; D008081:Liposomes; D010713:Phosphatidylcholines; D013053:Spectrophotometry; D013113:Spin Labels; D013748:Tetracaine / Q000031:analogs & derivatives*
D003497:Cyclic N-Oxides; D008051:Lipid Bilayers; D008081:Liposomes; D010713:Phosphatidylcholines; D013113:Spin Labels; D013748:Tetracaine; C055584:chlorotetracaine; C055585:5-doxylstearic acid methyl ester