Thyroid benign nodules (TBNs) are the most common lesions of this endocrine gland and are prevalent diseases around the world. Among TBNs the colloid goiter (CG) and thyroid adenoma (TA) are very frequent diseases. An evaluation of the variant of TBNs is clinically important for subsequent therapeutic interventions, as well as for more clear understanding the etiology of these disorders. The aim of this exploratory study was to examine differences in the content of silver (Ag), bromine (Br), calcium (Ca), chlorine (Cl), cobalt (Co), chromium (Cr), cooper (Cu), iron (Fe), mercury (Hg), iodine (I), potassium (K), magnesium (Mg), manganese (Mn), sodium (Na), rubidium (Rb), ammonium (Sb), scandium (Sc), selenium (Se), strontium (Sr), and zinc (Zn) in tissues of CG, TA, and T. Thyroid tissue levels of twenty chemical elements (ChE) were prospectively evaluated in 46 patients with CG, 19 patients with TA, and 12 patients with T. Measurements were performed using non-destructive energy-dispersive X-Ray fluorescent analysis combined with instrumental neutron activation analysis with high resolution spectrometry of short- and long-lived radionuclides. Tissue samples were divided into two portions. One was used for morphological study while the other was intended for ChE analysis. It was observed that in CG, TA and T tissues content of Ag, Br, and Hg was significantly higher, while level of I was lower than in normal thyroid tissue. In addition to these ChE, in CG and TA samples content of Cl, Cr, and Na was higher than in normal gland. ChE composition of T tissue differed from CG and TA tissues by elevated Rb level and reduced Se and Zn levels. The abnormal increase in Ag, Br, Hg level and decrease in I level in all TBNs might demonstrate an involvement of these ChE in etiology and pathogenesis of TBNs. In addition, elevated levels of such ChE as Cl, Cr, and Na can be also important for goitrous and adenomatous transformation of thyroid. It was supposed that elevated level of Rb and reduced levels of Se and Zn content in thyroid with T could possibly be explored for differencial diagnosis of T among other TBNs.