The aim of the study was to investigate the effect of different levels of protein in a diet on the incorporation of endogenous urea nitrogen (EUN) into individual amino acids (AA) of the ruminal bacteria of goats fed a low- (LP), medium- (MP), or high-protein diet (HP) in a 3 × 3 Latin square design. Three Alpine goats of about 35 kg body weight fitted with cannula into the rumen and catheter into the jugular vein were fed three isoenergetic diets containing 11% (LP), 13% (MP), or 16% (HP) crude protein in dry matter. The goats were infused for 6 days continuously with an ¹⁵N urea solution into the jugular vein. Ruminal bacteria were hydrolysed with 6M HCl. Next, butyl derivatives of free bacterial AA were obtained using HCl in butanol, then N-acylated using trifluoroacetic acid anhydride and analysed by gas chromatography using a mass-selective detector. The concentration of urea in plasma was 178, 356 and 667 mg · l^–1 in goats from groups LP, MP and HP, respectively. ¹⁵N-excess during the infusion of labelled urea was significantly higher (P < 0.05) in the vast majority of AA of ruminal bacteria from goats fed the LP diet in comparison with goats fed the HP diet. Therefore, the level of protein in the diets affects the incorporation of EUN into bacterial AA. With the LP diet, EUN was incorporated mostly into glutamic acid, isoleucine and arginine, while in the case of the HP diet, into glutamic acid and arginine, as well as methionine. Regardless of the level of nitrogen in the diets, the incorporation of ¹⁵N into proline was very low. Irrespective of the dietary nitrogen level, EUN appears to be predominantly used for synthesis of glutamic acid in ruminal bacteria.