<p>As a sustainable solution, ureolytic-pathway-based microbial self-healing concrete has gained increasing recognition as a promising strategy for repairing cracks. However, challenges remain in achieving effective deep crack repair, as the repair product tends to concentrate in the superficial regions. As fundamental components governing the mineralization reactions, the heterogeneous distribution of precipitation precursors concentration and pH within cracks may hold the key to addressing this issue. This study explored the <i>in-situ</i> distribution of main physicochemical characteristics (pH, Ca²⁺, urea, and nutrient) along the depth direction of microcracks in microbial self-healing concrete, and investigated the urease activity revival of spores in both shallow and deep regions within the cracks. The results showed that the bio-agent concentrations and pH exhibited a gradient distribution along the depth direction within the crack. From the crack mouth (0 mm) to bottom (40 mm), pH increased from approximately 10.5 to 12.7, Ca²⁺ concentration rose from around 3 to 50 mmol/L, urea concentration increased from about 50 to 600 mmol/L, and nutrient broth concentration grew from approximately 0.5 to 4.5 g/L. For crack regions with depths less than 5 mm, spores effectively germinated into active cells and revived sufficient urease activity within 15 d. In crack regions deeper than 5 mm, though higher urea concentrations (up to 769.90 mmol/L) and Ca²⁺ concentration (43–55 mmol/L) prolonged the time required for spore germination and revival of urease activity. However, as long as the pH remained below 12.0, spores could still revive urease activity and completely decompose urea. Excessively high environmental pH was the direct cause of the inhibition of spore germination and urease activity revival.</p>