This study developed a novel mucoadhesive hydrogel via electrostatic complexation of sulfhydrl-modified low-methoxyl pectin (PEC-SH) and chitosan oligosaccharide (COS) for colon-specific delivery of Lactobacillus rhamnosus (LGG). Free thiol groups were successfully grafted onto the pectin backbone via a carbodiimide-mediated amidation reaction with L-cysteine. Under the optimal reaction conditions employing a 1:1 molar ratio of EDC/NHS at a concentration of 0.2 mg/mL and a cysteine concentration of 1.5 mol/L, the disulfide bond content of the thiol-modified pectin reached its peak value of 281.82 μmol/g. The polyelectrolyte hydrogel formed by electrostatic interaction between PEC-SH and COS (mass ratio 1:8) exhibited strong viscoelasticity (G'>350 Pa), a 260% pH-responsive swelling ratio, and high encapsulation efficiency for LGG (>92%). After 28 days at 4°C, probiotic viability in the PEC-SH/COS composite gel remained above 7.5 log CFU/mL. Simulated gastrointestinal digestion showed the composite gel effectively protected probiotics, maintaining viable counts above 10⁸ CFU/mL. Ex vivo porcine mucus adhesion tests revealed that the COS:PEC-SH (1:8, w/w) composite hydrogel exhibited an approximately 17-fold enhancement in mucoadhesive force compared to free probiotics. This superior adhesion was further corroborated by isothermal titration calorimetry (ITC), which indicated an exothermic interaction between the hydrogel and mucosal components, along with a high binding constant. The colonic perfusion assays demonstrated a mucoadhesion ratio of 16.61 ± 0.40% for the composite hydrogel. These collective results confirmed that the PEC-SH/COS composite hydrogel could serve as an effective colon-specific delivery system with considerable potential to enhance probiotic adhesion and colonization