In order to evaluate rumen microbial activity, the effects of two nitrogen sources (organic and inorganic) were analyzed in vitro by inoculating dairy cow rumen fluid and using citrus byproduct (CBP) and milk thistle (Silybum marianum, MT) as energy source substrates. The experimental design was 2 × 3 where we used three different nitrogen sources (N1: no nitrogen added, N2: ammonium bicarbonate, N3: sodium glutamate) with a basic concentration of nitrogen (100 mg/1 L). Culture pH, dry matter

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... gradability %, ammonia-N (NH 3 -N), total volatile fatty acids (TVFAs), individual volatile fatty acid proportions, and acetate propionate ratio (A:P) were measured after 24 h of fermentation. Culture pH values were in the optimal range with CBP and MT in all media. Supplementing nitrogen sources had similar tendencies to increase dry matter degradability (P < 0.01) and NH 3 -N production in N2 and N3 with MT and CBP. Glutamate significantly (P < 0.01) increased TVFAs, which were twice as high than in N1 and N2 with MT. These results show that N supplementation had minor effects on in vitro rumen microbiota activity compared to N1; however, it could be proposed to improve microbial biomass production in mixed rations of agricultural byproducts with local forages.