Nacystelyn, a novel lysine salt of N-acetylcysteine, to augment cellular antioxidant defence in vitro

Abstract

Nacystelyn (NAL), a recently-developed lysine salt of N-acetylcysteine (NAC), and NAG, both known to have excellent mucolytic capabilities, were tested for their ability to enhance cellular antioxidant defence mechanisms. To accomplish this, both drugs were tested in vitro for their capacity: (1) to inhibit O 2 − and H 2O 2 in cell-free assay systems; (2) to reduce O 2 − and H 2O 2 released by polymorphonuclear leukocytes (PMN); and (3) for their cellular glutathione (GSH) precursor effect. In comparison with GSH, NAL and NAC inhibited H 2O 2, but not O 2 −, in cell-free, in vitro test systems in a similar manner. The anti-H 2O 2 effect of these drugs was as potent as that of GSH, an important antioxidant in mammalian cells. To enhance cellular GSH levels, increasing concentrations (0–2 × 10 −4 mol 1 −1) of both substances were added to a transformed alveolar cell line (A549 cells). After NAC administration (2 × 10 −4 mol 1 −1), total intracellular GSH (GSH + 2GSSG) levels reached 4·5 ± 1·1 × 10 −6 mol per 10 6 cells, whereas NAL increased GSH to 8·3 ± 1·6 × 10 −6 mol per 10 6 cells. NAC and NAL administration also induced extracellular GSH secretion; about two-fold (NAC), and 1·5-fold (NAL), respectively. The GSH precursor potency of cystine was about two-fold higher than that of NAL and NAC, indicating that the deacetylation process of NAL and NAC slows the ability of both drugs to induce cellular glut production and secretion. Buthionine-sulphoximine, which is an inhibitor of GSH synthetase, blocked the cellular GSH precursor effect of all substances. In addition, these data demonstrate that NAC and NAL reduce H 2O 2 released by freshly-isolated cultured blood PMN from smokers with chronic obstructive pulmonary disease (COPD) ( n=10) in a similar manner (about 45% reduction of H 2O 2 activity by NAC or NAL at 4 × 10 −6 mol 1 −1). In accordance with the results obtained from cell-free, in vitro assays, O 2 − released by PMN was not affected. Ambroxol (concentrations: 10 −9−10 −3 mol 1 −1) did not reduce activity levels of H 2O 2 and O 2 − in vitro. Due to the basic effect of dissolved lysine, which separates easily in solution from NAL, the acidic function of the remaining NAC molecule is almost completely neutralized [at concentration 2 × 10 −4 M: pH 3·6 (NAC), pH 6·4 (NAL)]. Due to their function as H 2O 2 scavengers, and due to their ability to enhance cellular glutathione levels, NAL and NAC both have potent antioxidant capabilities in vitro. The advantage of NAL over NAC is two-fold; it enhances intracellular GSH levels twice as effectively, and it forms neutral pH solutions whereas NAC is acidic. Concluding from these in vitro results, NAL could be an interesting alternative to enhance the antioxidant capacity at the epithelial surface of the lung by aerosol administration.