Sirtuins are NAD+-dependent deacetylases and ADP ribosyltransferases that are activated under stress conditions such as calorie restriction and starvation. Boron, which is mostly found in the form of boric acid (BA) or sodium borate (NaB), is known to bind NAD+. In this study, the effect of NaB on hepatoma cell line HEP3B was investigated by analyzing the proteins harvested from NaB-treated and serum-starved HEP3B cells. The NaB treatment (15 μg/mL) led to a decrease in the overall proteome acetylation and particularly in mitochondria, the synthesis rate of respiratory complexes, the amount of cellular reactive oxygen species, and the proliferation rate of HEP3B cells. On the other hand, the cellular ratio of NAD+/NADH and the deacetylase activity of mitochondrial sirtuin, SIRT3, were found to be elevated. The results of this study suggest a link between boron treatment and activation of SIRT3 by means of a NAD+-NaB interaction, which reprograms cellular metabolism. This study is expected to pave the way for new findings uncovering the metabolic changes in HEP3B cells related to SIRT3 activity upon NaB treatment.