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Diabetic foot infections (DFIs) are frequently linked to diabetic-related morbidity and death because of the ineffectiveness of conventional antibiotics against multidrug-resistant bacteria. Pexiganan and nisin A are antimicrobial peptides (AMPs), and their application may complement conventional antibiotics in DFI treatment. A collagen 3D model, previously established to mimic a soft-tissue collagen matrix, was used to evaluate the antibacterial efficacy of a guar gum gel containing pexiganan and nisin alone and combined with three antimicrobials toward the biofilms of <i>Staphylococcus aureus</i> and <i>Pseudomonas aeruginosa</i> isolated from infected foot ulcers. Antimicrobials and bacterial diffusion were confirmed by spot-on-lawn and bacterial growth by bacterial count (cfu/mL). Our main conclusion was that the dual-AMP biogel combined with gentamicin, clindamycin, or vancomycin was not able to significantly reduce bacterial growth or eradicate <i>S. aureus</i> and <i>P. aeruginosa</i> DFI isolates. We further reported an antagonism between dual-AMP and dual-AMP combined with antibiotics against <i>S. aureus</i>.