Phytochemical extracts from the leaves and stem of red sorghum (Sorghum bicolor, L. Moench) potentially improve in vitro fermentation by modulating rumen protozoa

Abstract

Plants enriched with phytochemicals potentially improve nitrogen utilization efficiency (NUE) in dairy cows. We hypothesized that phytochemicals extracted from the leaves and stem of red sorghum (Sorghum bicolor, L. Moench; RSLS) and its fractions could regulate rumen protozoa and nitrogen utilization. The Box-Behnken design was used to optimize ethanol concentrations (60%, 80%, and 100%), extraction time (2, 3, and 4 h), and extraction repetitions (2, 3, and 4 times) to maximize extraction yields, total phenolic content (TPC), and total flavonoid content (TFC) from RSLS, and then evaluate the antiprotozoal activities in vitro (n = 6). Crude extracts (CE) produced using the optimum extraction method were assessed for the effects on in situ DM degradation (in situ DMD) after 12, 18, 24, 36, and 48 h of in situ incubation. The CE was further fractionated by petroleum ether, ethyl acetate, n-butanol, and aqueous to assess antiprotozoal activity. Data were analyzed using one-way ANOVA, and differences among means were assessed using Bonferroni-corrected pairwise t-tests in R (R Foundation for Statistical Computing; version 1.4.1717). The optimized extraction conditions, 80% ethanol, 6-h extraction, repeated 3 times, achieved yields of 48% from RSLS, with TPC levels of 400 mg tannic acid/g, and TFC at 120 mg quercetin/g. Compared with the control, the CE decreased total protozoa, Entodinium, and ammonia-nitrogen (NH3-N) by 47.0%, 49.0%, and 31.2%, while increasing microbial CP and in vitro DMD by 66.8% and 24.3%, respectively. The CE increased in situ DMD by 24.2% and 20.6% after 24 and 36 h of fermentation. After purification, all fractions, particularly the aqueous fraction, reduced total protozoa, Entodinium, and NH3-N compared with the control group. Therefore, ethanol-extracted phytochemicals from RSLS potentially improve NUE by modulating protozoa and in vitro fermentation. Future studies should elucidate the chemical structure of the compounds responsible for these findings to exploit their promising therapeutic properties.