The study evaluated the effects of a polyol blend (equal parts xylitol, arabinitol, and sorbitol) added to beef cattle total mixed rations (TMR) on in vitro rumen fermentation, total gas production, digestibility, and organic acid profiles. Polyols were supplemented at 0, 2, 4, 8, and 12% of dry matter. Results showed no significant differences in total gas production, metabolic energy, lactation net energy, or organic matter digestibility (OMd) between polyol-supplemented groups and the control (P < 0.05). However, the addition of 2, 4 and 8% polyols increased in vitro rumen pH (P < 0.05). All polyol levels reduced the proportions of acetic acid (AA), butyric acid (BA), propionic acid (PA), iso-butyric acid (IBA), valeric acid (VA), iso-valeric acid (IVA), and branched short chain fatty acids (BSCFA) in total short-chain fatty acids (T-SCFA), while only 12% supplementation elevated straight-chain SCFA concentrations (P < 0.05). Overall, polyol addition at 2, 4, 8, and 12% did not significantly alter in vitro rumen fermentation parameters (total gas production, OMd, energy values and SCFAs) in beef cattle TMR. However, the increase in rumen fluid pH observed with moderate polyol doses (2–8%) suggests a potential buffering effect, which may be beneficial in starch-rich diets used during intensive fattening or early lactation in dairy cattle.