Enhancing the Oral Bioavailability of Glutathione Using Innovative Analogue Approaches

<b>Background:</b> Glutathione (GSH) is an essential antioxidant that protects against oxidative stress, but its oral bioavailability is below 1% due to enzymatic degradation and poor gastrointestinal absorption. Improving the oral bioavailability of GSH could significantly enhance its therapeutic efficacy. <b>Methods:</b> This study synthesised GSH analogues with chemical modifications to improve bioavailability. Seven GSH derivatives were designed: three analogues with altered stereochemistry (<b>1.62</b>, <b>1.63</b>, and <b>1.64</b>) and three <i>N</i>-methylated derivatives (<b>1.65</b>, <b>1.70</b>, and <b>1.71</b>), alongside a native GSH (<b>1.61</b>). The analogues were synthesised via Fmoc-solid-phase peptide synthesis, and they were characterised using reverse-phase high-performance liquid chromatography (RP-HPLC), electrospray ionisation mass spectrometry (ESI-MS), Fourier-transform infrared spectroscopy (FTIR), and nuclear magnetic resonance (NMR) spectroscopy. Their toxicity was assessed on Caco-2 cells for viability, and their antioxidant activity was assessed on UVA-irradiated fibroblast cells, enzymatic resistance, and interactions with GSH-metabolising enzymes. <b>Results</b>: Among the tested analogues, the <i>N-</i>methylated cysteine Compound (1.70) emerged as the most promising candidate. Compound <b>1.70</b> demonstrated superior resistance to enzymatic degradation, as well as showing enhanced cell viability and improved antioxidant activity. In vivo studies revealed a 16.8-fold increase in plasma half-life (t½) and a 16.1-fold increase in oral bioavailability compared to native GSH. <b>Conclusions</b>: Chemical modification strategies, particularly the <i>N-</i>methylation of GSH, present a viable approach to enhancing oral bioavailability. Compound <b>1.70</b> showed significant potential for therapeutic applications, warranting further investigation and development in clinical settings.