We previously reported that consumption of a resistant protein, sericin, reduces colon tumorigenesis, constipation, and serum TG in rodents. The present study was conducted to elucidate the effects of dietary sericin on the intestinal luminal environment in rats fed a high-fat (HF) diet. Rats were fed 300 or 50 g/kg of beef tallow with or without 40 g/kg sericin, a protein purified from cocoons of Bombix mori, for 3 wk. Intestinal luminal variables, including IgA (index of intestinal immune function), mucins (index of barrier function), organic acids, microflora, and secondary bile acids, were measured. Dietary sericin markedly elevated fecal IgA in the HF diet group (3-fold, P < 0.05) but not in the low-fat (LF) diet group. Fecal mucin levels were elevated by sericin intake in the HF diet group (P < 0.05). Cecal organic acids, including acetate, propionate, n-butyrate, and succinate, were significantly lower in the HF diet group compared with the LF diet group. Dietary sericin significantly elevated cecal acetate and n-butyrate in the HF diet group but not in the LF diet group. Compared with the LF diet, the HF diet significantly increased serum TG in the untreated group but not in those fed sericin. The HF diet increased lower density lipoprotein (VLDL + IDL + LDL) cholesterol and it was reduced by sericin intake (P < 0.05). There was an inverse correlation between serum TG and cecal acetate (Spearman rank correlation coefficient = -0.63; P < 0.001). The profile of microflora in cecal digesta and fecal secondary bile acids (a risk factor for colon cancer) did not differ between the HF diet and HF diet with sericin groups. These results suggest a novel and favorable effect of sericin on colon health by modulating intestinal immune and barrier functions and fermentation in rats fed a HF diet.