Enhancing the nutritional values and quality of fruit crops is of utmost significance, necessitating the exploration and implementation of novel agricultural approaches. Therefore, the application of natural sources, such as biostimulants and the reduction of conventional fertilizers, are integral to sustainable agriculture, which aims to reduce environmental harm and maintain the long-term health of agricultural systems, meanwhile, maximizing productivity and profitability. The study’s objective was to assess the impact of applying arbuscular mycorrhizal fungi (AMF) and licorice root extract (LRE) on the growth and some biochemical traits of tomato fruit. For the AMF inoculation, 15 g/plant was applied to the soil, while the LRE was sprayed onto the canopy at varying doses of 0, 5, 10, and 15 g/L. A combination of AMF and LRE was also applied simultaneously. All treatment combinations had noticeable effects on plant growth and quality. The combination of AMF with LRE at a concentration of 15 g/L significantly influenced most of the growth, fruit, and physicochemical characteristics. AMF resulted in the lowest nitrate accumulation in the fruits. Furthermore, doses of 10 g/L LRE exhibited the highest levels of heavy metals, including zinc, manganese, and copper. This study suggests that the integrated use of AMF and LRE is a promising approach for the sustainable and economically viable production of tomato fruit under controlled or semi-controlled conditions.