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No-till (NT) cropping systems have the potential to enhance soil aggregation, providing physical protection and soil C sequestration. The existence of discrepancies in the impact of tillage on soil aggregation and soil C sequestration warrants further studies, particularly for different crop rotations. We hypothesized the following: a) NT biannual crop rotations tend to be more effective in restoring large macroaggregation and the concentrations of soil organic C (SOC), total N and permanganate oxidizable C (POXC) associated with macroaggregates than NT systems with a one-year frequency pattern and conventional tillage (CT); b) the continuous biomass-C inputs via crop residues in large macroaggregates under NT tend to increase the proportion of aliphatic C than those under CT. Therefore, the objectives of this study were: (i) to assess changes in the aggregate size distribution and levels of aggregate-associated total SOC, total N and POXC and (ii) to characterize humic acid (HA) using 13C CP-MAS nuclear magnetic resonance (NMR) spectra of 8- to 19-mm soil aggregate size class in a reference vegetation (RV) and in rice-, soybean- and cassava-based cropping systems (RcCS, SbCS and CsCS, respectively) in a clayed Oxisol after tillage and crop rotation management. We evaluated four treatments in each cropping system: 1) CT, and 2) three NT systems in a randomized complete block design with three replicates. Soil aggregate samples were collected at depths of 0–5, 5–10 and 10–20 cm. The conversion of RV to agricultural land influenced the distribution of aggregate size classes, soil aggregation indices and aggregate-associated SOC, total N and POXC in the two surface layers. The formation of large macroaggregates (8–19 mm) dominated the aggregate size distribution with a relatively higher proportion under RV and NT than under CT. Across all soil depths, the proportions of the 8- to 19-mm aggregate size fraction were 59% (NV), 43% and 47% (RcCS), 45% and 53% (SbCS) and 34% and 37% (CsCS) for the CT and NT systems, respectively. Among the three NT systems, the biannual crop rotations in the three cropping systems (NT2-Rice, NT2-Soybean, NT2-Cassava; NT3-Rice, NT3-Soybean and NT3-Cassava) indicated better performance than the one-year frequency pattern in restoring large macroaggregation and the concentrations of SOC, total N and POXC associated with large macroaggregates. Additionally, in the surface (0–5 cm) and subsurface (10–20 cm) soil layers, the SbCS with a high rate (7.32 Mg C ha−1 year−1) and diversity [Pennisetum typhoides) (Pearl millet)/maize + Brachiaria ruziziensis (Brz), Stylosanthes guianensis (St)] of biomass-C inputs reached the highest levels of lability of SOC and POXC in the macroaggregate size classes of 0.25–0.5 and 8–19 mm, respectively. The CP-MAS 13C NMR measurement suggests that the continuous and high biomass-C inputs with diverse crop residues under NT, such as millet, maize, Brz, St and Crotalaria juncea, tended to increase the proportion of aliphatic C than under CT; an opposite trend was observed for aromatic C. 13C NMR revealed an advance caused by the association between the quantity and quality of C addition via cultural residues in the discrimination of the composition of C in the macroaggregation in the tropical region.