Stability of Glutathione in Induced Sputum: Impact of Freezing

BACKGROUND: Oxidative stress has repeatedly been linked to the pathogenesis of pulmonary disorders like asthma and chronic obstructive pulmonary disease. Measuring glutathione (GSH) in induced sputum (IS) offers a noninvasive tool to study oxidative stress in airway diseases. OBJECTIVES: This study assessed the stability of GSH in sputum supernatant under varying conditions. METHODS: GSH in IS of 14 (7 healthy, 4 chronic obstructive pulmonary disease, 3 allergic rhinitis) nonsmoking subjects was quantified spectrophotometrically. The stability of GSH in supernatant was analyzed over 24 h under different ambient conditions (room temperature and cooling at 4 degrees C). Reproducibility of GSH measurements in immediately processed and frozen supernatant (+72 h) was expressed by intraclass correlation coefficient (R(i)) and coefficient of repeatability (CR). RESULTS: GSH recovery in supernatant decreased in a time- and temperature-dependent manner. Samples stored at 4 degrees C and room temperature showed a rapid decline of stability after 2 h. Mean GSH concentrations in IS after freezing (-20 degrees C) and thawing after 72 h were not significantly different from GSH values measured immediately after processing of the samples (immediate processing: 17.9 +/- 13.9 microM; 72 h freezing: 16.4 +/- 12.9 microM, p = 0.2). The reproducibility between immediately processed and frozen samples was excellent (R(i) = 0.97; CR = 5.7 microM). CONCLUSIONS: Storage of sputum supernatant at room temperature or 4 degrees C leads to a rapid decline of GSH recovery compared with baseline values. Immediate freezing of samples is a suitable and valid alternative to rapid processing and allows collection and shipment of samples for subsequent analysis.