Microneedles have emerged as an effective strategy to bypass the stratum corneum by creating microchannels in the skin, allowing for enhanced drug permeation with minimal invasiveness. Pectin, a natural polysaccharide, when modified with thiol (-SH) groups, exhibits redox-sensitive behavior, making it responsive to reducing agents such as glutathione and dithiothreitol (DTT). The objective of this study was to investigate the transdermal delivery efficacy of redox-responsive microneedle-containing thiolated pectin. Various molar ratios of thiolated pectin were synthesized through the ring-opening reaction followed by nucleophilic substitution of propylene sulfide with pectin. MN matrices were formulated with thiolated pectin at various molar ratios, hyaluronic acid, collagen, and trehalose. The stiffness and mechanical strength of the microneedles increased with higher thiol-containing pectin molecules. The in vitro skin permeation release showed a large amount of FITC release when no thiol group was conjugated to pectin. MN-thio: pectin (20:10) with H₂O₂ showed greater release at higher DTT concentrations, and in the absence of DTT, the release was 50 times less than without thiol. In summary, the redox-responsive microneedle containing thiolated pectin may be a promising vehicle for transdermal drug delivery by harvesting the reducing agents in the human body.