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Microcrystalline cellulose (MCC) has several applications in food and pharmaceutical industries but the nutritional value for dairy cows and effects on in vivo digestion are not known. A feeding experiment was conducted using 24 dairy cows. The cows were offered MCC originating from unbleached softwood kraft pulp at 0, 10 or 100 g per kg diet dry matter (DM) to replace barley grain in the diet. The total daily DM intake was on average 25.6 kg and not significantly affected by the diet. Positive effects on rumen fermentation could not be demonstrated in a feeding situation where total mixed ration and a concentrate proportion of 0.50 on DM basis was used. Diet organic matter digestibility was not affected by MCC inclusion but fibre digestibility improved and the additional MCC fibre was virtually completely digested. The production potential of MCC was lower than that of barley grain as daily yields of energy-corrected milk, milk fat and protein, and milk protein concentration decreased when MCC replaced barley grain in the diet. Based on these results, MCC is not recommended as a dietary component for high-yielding dairy cows.