Inflammation of the mammary gland is the most costly disease affecting the US dairy sector resulting in costs around $2 billion/year. Histone deacetylase (HDAC) inhibition has anti-inflammatory properties in rodents, and short-chain fatty acids (SCFA) are effective in inhibiting HDACs. We hypothesized that SCFA would inhibit inflammation via HDAC-dependent regulation of gene expression. We aimed to evaluate sodium propionate (SP) and sodium butyrate (SB) effects on HDAC activity, histone H3 acetylation, and inflammatory gene expression. Bovine mammary epithelial (MAC-T) cells were used as the lipopolysaccharide (LPS)-induced inflammatory model. Cells were cultured in basal medium and cell lysates were incubated with increasing doses of SP or SB (0 to 5 mM) for 2 h before incubation with HDAC substrates (2 h). HDAC activity was determined by fluorescence detection. Cells were also pretreated with SP or SB (0 to 3mM) for 2 h, stimulated with LPS (1 µg/mL) for 2 h, and assessed for histone H3 acetylation by immunoblotting. Next, cells were pretreated with SP or SB (1mM) for 24 h, stimulated with LPS (1 µg/mL) for 2 h, and RNA was isolated. PCR array was used to examine the expression of 83 inflammatory genes and quantitative real-time PCR was used for gene validation. One-way ANOVA with Tukey post hoc analysis was used to assess significance (P ≤ 0.05). SP and SB dose-dependently and selectively inhibited class I HDAC activity, which differed between the SCFAs, where SB inhibited HDACs 2, 3, and 8, while SP inhibited HDACs 2 and 8. SP and SB dose-dependently increased histone H3 acetylation, differing between the SCFAs, where SB increased H3K9/14, H3K18 and H3K27 acetylation, while SP increased H3K9/14 and H3K18 acetylation. SCFAs increased the overall inflammatory gene expression in MAC-T cells. Under our experimental conditions, the findings suggest that SCFAs regulate epigenetic marks on nucleosomal DNA in addition to regulation of inflammatory gene events independent of HDAC activity in MAC-T cells.