Accumulating above-ground biomass (AGB) and conservation of biodiversity are two major services provided by any ecosystem and homegardens are no exceptions. The permanent structure of a homegarden is owing to its perennial tree-community which steadily builds up and maintains the tree diversity and AGB accumulation. Being a manmade agroforestry system, humans play a vital role in mediating each component of homegarden ecosystem including its carbon stock and biodiversity through management of the tree community. The objectives of this study were to compute the tree diversity and AGB and human interventions in homegarden ecosystem in Jaffna peninsula and to identify the most prominent factors affecting the AGB. A primary survey was conducted capturing 135 homegardens from two agro-ecological regions (DL3 and DL4) in Jaffna peninsula (Northern province, Sri Lanka), where homegardens are observed as the primary tree-based vegetation. Data on floral composition, livestock and, household-interventions in homegardens were gathered through a structured questionnaire. Tree diversity and AGB were computed using Shannon-Wiener index (SWI) and carbon stock respectively. A multiple linear regression model was estimated to determine the effects of pre-defined interventions on AGB of homegardens. The study sample consisted of 122 small-scale (<0.2 ha), 11 medium-scale (0.2–0.8 ha) and 02 large-scale (>0.8 ha) homegardens. The mean tree diversity as estimated by SWI was 1.1 ± 0.6 with 38 perennial species and 1286 trees; 32 genera and 17 families. An average homegarden is estimated to store 23.8 ± 20.9 Mg C/ha as AGB. Homegardens which reared livestock animals, had a significantly lower SWI value, yet, a higher AGB stock, compared to the opposed group (p < 0.05). The SWI, time allocation (hours) for homegarden/week/ha, technical knowledge on farming and non-timber usage within homegarden have significantly (p < 0.05) contributed to the AGB of homegardens, with regression-coefficients of 6.12, 0.02, 1.38 and 7.91, respectively. The results indicate that up-keeping the commitment of households in managing their homegardens contribute to environmental sustainability. The results revealed a significant contribution of human interactions on the size of the AGB of homegardens in Jaffna peninsula. This contribution was not adequately discussed in previously literature and the findings suggest the need to manage human interactions in homegarden agroforestry systems to optimize carbon stock in traditional homegardens. Display omitted •Humans play a vital role in mediating each element of homegardens, including the above-ground biomass (AGB).•The study aimed to assess the relations of AGB vs human interventions, in modulating homegardens as AGB carbon storages.•Tree diversity, time spent on homegarden, technical awareness and non-timber usage, significantly affected the AGB.•The model suggests that effective mediation of humans could optimize traditional homegardens at storing AGB carbon.•The impact of human intervention on homegardens by means of a promising carbon storage is hardly discussed in literature.