Abstract
β2-Agonists are the most effective bronchodilators for the rapid relief of asthma symptoms, but for unclear reasons, their effectiveness may be decreased during severe exacerbations. Because peroxidase activity and nitrogen oxides are increased in the asthmatic airway, we examined whether salbutamol, a clinically important β2-agonist, is subject to potentially inactivating nitration. When salbutamol was exposed to myeloperoxidase, eosinophil peroxidase or lactoperoxidase in the presence of hydrogen peroxide (H2O2) and nitrite (NO2−), both absorption spectroscopy and mass spectrometry indicated formation of a new metabolite with features expected for the nitrated drug. The new metabolites showed an absorption maximum at 410 nm and pKa of 6.6 of the phenolic hydroxyl group. In addition to nitrosalbutamol (m/z 285.14), a salbutamol-derived nitrophenol, formed by elimination of the formaldehyde group, was also detected (m/z 255.13) by mass spectrometry. Notably, the latter metabolite was detected in exhaled breath condensates of asthma patients receiving salbutamol but not in unexposed control subjects, indicating potential for β2-agonist nitration to occur in the inflamed airway in vivo. Salbutamol nitration was inhibited in vitro by ascorbate, thiocyanate, and by the pharmacological agents methimazole and dapsone. The efficacy of inhibition depended upon the nitrating system, with the lactoperoxidase/H2O2/NO2− being the most affected. Functionally, nitrated salbutamol showed decreased affinity for β2-adrenergic receptors and impaired cAMP synthesis in airway smooth muscle cells when compared to the native drug. These results suggest that under inflammatory conditions associated with asthma, phenolic β2-agonists may be subject to peroxidase-catalyzed nitration that could potentially diminish their therapeutic efficacy.
- Received May 5, 2010.
- Revision received October 19, 2010.
- Accepted October 19, 2010.
- The American Society for Pharmacology and Experimental Therapeutics