Beta Glucans (β-glucans) are naturally occurring polysaccharides that have positive effects on healing in acute and chronic wounds. This study aimed to identify how β-glucans modulate macrophage polarisation and inflammation to aid the healing response. Flow cytometry was used to assess the effect of β-glucan on human monocytes during differentiation into M0, M1 and M2 macrophages. Subsequently, a murine full-thickness excisional wound healing model was conducted where wounds were treated with either β-glucan hydrogel or vehicle, at the time of wounding. The wounds were analysed to determine the rate of wound closure, the effect on inflammation, and matrix deposition. β-glucan promoted differentiation of monocytes to M0 macrophages but inhibited differentiation of M0 macrophages to pro-inflammatory M1 macrophages with no effect on M2 macrophage formation. In vivo, treatment of excisional wounds with β-glucan hydrogel accelerated healing with an earlier, more resolved inflammatory phase containing greater numbers of M2 macrophages and fewer neutrophils within the wound. No statistically significant effect on matrix deposition was observed. β-glucans modulate macrophage differentiation and accelerate healing in excisional wounds with no adverse effect on matrix formation. β-glucans are a potential therapeutic approach for treatment of hard-to-heal wounds in humans.