Chitosan was evaluated as a feed additive to mitigate in vivo CH4 emissions in beef cattle. Twenty-four crossbred heifers (BW = 318 ± 35 kg) were used in a randomized block design replicated in 2 periods. The design included a 2 × 3 factorial arrangement of treatments, which included diet (high concentrate [HC] or low concentrate [LC]) and 0.0, 0.5, or 1.0% of chitosan inclusion (DM basis). Diets were offered ad libitum and individual intake was recorded. An in vitro experiment to analyze chitosan’s effect on fermentation parameters and gas production kinetics was performed. A diet effect (P < 0.01) was observed for CH4 emissions expressed as grams/day, grams/kilogram of BW0.75, and grams/kilogram of DMI. Heifers consuming the LC diet produced 130 g of CH4/d vs. 45 g of CH4/d in those consuming the HC diet. Incubation fluid pH increased linearly (P < 0.05) when chitosan was included in HC substrates. In vitro CH4 production was not affected (P > 0.10) by chitosan in HC substrate; however, when incubated with the LC substrate, CH4 production increased quadratically (P < 0.01) as chitosan inclusion increased. A digestibility marker × diet interaction occurred (P < 0.05) for DM, OM, CP, NDF, and ADF digestibility. Diet × chitosan interactions (P < 0.05) occurred for DM, OM, NDF, and ADF digestibility when Cr2O3 was used. When TiO2 was used, diet × chitosan interactions (P < 0.05) were observed for NDF and ADF. However, using indigestible NDF as an internal marker, DM and OM digestibility were improved (P < 0.05) by 21 and 19%, respectively, when chitosan was included in LC diets. In conclusion, feeding up to 1% of chitosan (DM basis) to heifers consuming a LC diet increased apparent total tract digestibility of nutrients. Enteric CH4 emissions were not affected by chitosan feeding, regardless of type of diet, and heifers consuming a 36% concentrate diet produced 2.6 times more methane per day than those consuming an 85% concentrate diet.

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