Integrated safety and microbiota profiling of fulvic acid formulations across in vitro and in vivo models

Fulvic acid, a naturally occurring organic compound derived from humic substances, has recently emerged as a multifunctional bioactive agent with potential biological relevance. Here, we evaluated the in vitro bioactivity of various fulvic acid-containing formulations, focusing on cytocompatibility, genotoxicity, anti-inflammatory effects, regenerative potential, and probiotic-stimulating potential microbiome-related activity. Two fossil-based fulvic acid formulations (MLG-50 and alkaline MLG-A50) were assessed in eukaryotic cell models (L929 fibroblasts, LoVo epithelial cells, HepG2 hepatocytes) as well as in vitro and in vivo microbial systems. Both formulations demonstrated high cytocompatibility across a broad concentration range, with genotoxicity levels significantly lower than those induced by reference dietary compounds and no relevant nuclear abnormalities detected. Cell proliferation assays revealed enhanced intestinal epithelial cell growth at sub-toxic concentrations. Wound healing assays indicated a pro-regenerative effect of MLG-A50 in LoVo cells, as reflected by an increased wound closure rate compared with the control. Anti-inflammatory activity was evidenced by the suppression of NF-κB activation in LPS-stimulated monocytes via LPS neutralization and receptor-level inhibition, accompanied by reduced TNF and IL-6 secretion without impairment of IL-10 production. In vitro and in vivo microbiome profiling revealed stimulation of beneficial bacterial taxa, including Lactobacillus and Clostridia spp., alongside reduced growth of pathogenic strains. In vivo supplementation with MLG-50 and MLG-A50 further increased microbial diversity and abundance of health-associated taxa. Overall, fulvic acid-based formulations exhibited a favorable safety profile and measurable bioactivity in cellular and microbiome models. These results support further investigation of fulvic acid as a multifunctional bioactive component, particularly in the context of immunonutrition, microbiome modulation, and regenerative-supportive strategies, while acknowledging the need for future mechanistic and translational studies.