This in vitro study aims to evaluate of hydrogen peroxide (HP) diffusion into the pulp chamber, bleaching efficacy (BE), and pH stability (pH) of single-application high concentrated in-office bleaching gels. Eighty-eight healthy premolars were randomly into eleven groups (n = 8) according to the in-office dental bleaching: DSP White Clinic 35% calcium (DW), Nano White 35% (NW), Opalescence XTra Boost 40% (OB), Pola Office + 37.5% (PO), Potenza Bianco Pro SS 38% (PB), Total Blanc 35% (TB), Total Blanc One-Step 35% (TO), Whiteness Automixx 35% (WA), Whiteness Automixx Plus 35% (WP), and Whiteness HP Blue 35% (WB). A group not exposed to bleaching agents was the control group (CG). All bleaching agents were applied in one session with a single application. After the bleaching procedure, the concentration of HP diffusion (μg/mL) into the pulp chamber was assessed using UV-Vis spectrophotometry. The BE (ΔEab and ΔE00) was evaluated before and 1 week after the bleaching procedure using a digital spectrophotometer. The pH of each bleaching gel was evaluated by digital pHmeter. The one-way ANOVA and Tukey's was used for a statistical analysis (α = 0.05). The concentration of HP diffusion into the pulp chamber was higher in all in-office bleaching gels when compared to CG (p < 0.0000001). However, there are a significant difference between them (p = 0.0001). A significant BE was observed in all in-office bleaching gels (p < 0.0001 for ΔEab and ΔE00), with a significant difference between them (p < 0.0001). PO, OB, TB, WP, and WB showed a higher BE when compared to DW, PB, and WA (p < 0.0001). Most bleaching gels were slightly acidic or alkaline during the total application time, while DW, PB, TB, and WA showed a high acidic behavior after 30 min of application. A single application was able to produce a bleaching efficacy. However, usually, gels with slightly acidic or alkaline pH during the application time reduces the HP diffusion into the pulp chamber. The single-application of bleaching gels with slightly acidic or alkaline and stable pH decreased the penetration of hydrogen peroxide into the pulp chamber in in-office bleaching and maintained the bleaching efficacy.