Quantitative electron paramagnetic resonance and spectrophotometric determination of the free radical 4-hydroxy-2,2,6,6-tetramethylpiperidinyloxy.

The first quantitative estimation of the stable free radical 4-hydroxy-2,2,6,6-tetramethylpiperidinyloxy (TEMPOL) simultaneously performed by electron paramagnetic resonance (EPR) and visible spectrophotometric titration is reported. The obtained molar absorptivity of TEMPOL 13.2 +/- 0.1 l(-1) m(-1) cm(-1) at 22,900 cm(-1) may be used as the future criterium about the purity of the EPR active substance. The studied compound may be considered as the second (after 1,1-diphenyl-2-pikrilhydrazil) material calibrated as the primary standard for quantitative EPR spectrometry.

Stability-indicating colorimetric assay for indicine N-oxide using TLC.

Aliquots of aqueous solutions in which indicine N-oxide may be degraded were mixed with 0.5 M formic acid (1:3) to adjust the pH to approximately 2-4 to quench the reaction and to ensure adequate TLC resolution. Silica-coated aluminum sheets were used to isolate indicine N-oxide by cutting the appropriate region from the chromatogram. By a modification of a known procedure, the silica gel then was treated with an acetic anhydride-diglyme mixture, and the mixture was heated to convert the drug to a pyrrole, which was then coupled with 4-dimethylaminobenzaldehyde to produce a color. The absorbance of the resulting solution was determined at 566 nm, and the apparent molar absorptivity, epsilon, based on the final indicine N-oxide concentration was 6.13 x 10(4). The recovery was approximately 92%, and the assays were readily reproducible with a coefficient of variation of 4.4%.