Chemical investigation of the endophytic MEXU 26343, isolated from the medicinal plant Hintonia latiflora, yielded the known polyketide vermelhotin (1) and a new salicylic aldehyde derivative, namely, 9S,11R-(+)-ascosalitoxin (2). The structure and absolute configuration of the new compound were established through extensive NMR spectroscopy and molecular modeling calculations at the DFT B3LYP/DGDZVP level, which included the comparison between theoretical and experimental optical rotation values. In addition, chemical transformations of 2 yielded suitable derivatives for NOESY and (1)H-(1)H NMR coupling constant analyses, which reinforce the stereochemical assignment. The potential affinity of 1 and 2 with (Ca(2+))(4)-hCaM in solution was measured using the fluorescent biosensor hCaM M124C-mBBr. The results showed that 1 bound to the protein with a dissociation constant (K(d)) of 0.25 ± 0.04 μM, close to that of chlorpromazine (K(d) = 0.64 ± 0.03 μM), a classical CaM inhibitor. The stoichiometry ratio of 1 to (Ca(2+))(4)-hCaM was 1:4, similar to other well-known CaM ligands.