Controlled C–N configurations, i.e., pyrrolic-N, pyridinic-N, and graphitic-N, are promising strategies to tailor the carbon dots’ (CDs) optical properties into the first near infrared (NIR) window (650–900 nm), a responsive range for biomedical application. However, a deep understanding of the role of the C–N configuration in the CDs’ properties is still challenging and thought-provoking owing to their complex structure. Here, an underlying pyrrolic-N concentration and position effect on the pyrrolic-N-rich CDs’ absorption was comprehensively elucidated based on the integrated experimental and computational studies. The as-synthesized pyrrolic-N-rich CDs exhibit a first NIR window absorption centered at 650 nm with high photothermal conversion. Pyrrolic-N concentrations from 1.4% to 11.3% and positions (edge and mid-site) were systematically investigated. A mid-site pyrrolic-N was subsequently generated after the pyrrolic-N concentration more than 10%. Edge-site pyrrolic-N induces a frontier orbital hybridization, reducing bandgap energy, while mid-site pyrrolic-N plays a critical role in inducing a first NIR window absorption owing to their high charge transfer. Also, pyrrolic-N-rich CDs inherit a bowl-like topological feature, elevating the CDs’ layer thickness as much as 0.71 nm. This study shed light on the design and optimization of pyrrolic-N on CDs for the first NIR window responsive materials in any biomedical application.