Dietary butyrylated high-amylose starch reduces azoxymethane-induced colonic O6-methylguanine adducts in rats as measured by immunohistochemistry and high-pressure liquid chromatography

O6-methyl guanine (O6MeG) adducts are major toxic, promutagenic, and procarcinogenic adducts involved in colorectal carcinogenesis. Resistant starch and its colonic metabolite butyrate are known to protect against oncogenesis in the colon. In this study, we hypothesized that a dietary intervention that specifically delivers butyrate to the large bowel (notably butyrylated high-amylose maize starch [HAMSB]) would reduce colonic levels of O6MeG in rats shortly after exposure to the deoxyribonucleic acid (DNA) alkylating agent azoxymethane (AOM) when compared with a low-amylose maize starch (LAMS). A further objective was to validate an immunohistochemistry (IHC) method for quantifying O6MeG against a high-performance liquid chromatography method using fluorescence and diode array detection. Rats were fed either LAMS or HAMSB diets for 4 weeks followed by a single injection of AOM or saline and killed 6 hours later. After AOM exposure, both IHC and high-performance liquid chromatography method using fluorescence and diode array detection measured a substantially increased quantity of DNA adducts in the colon (P < .001). Both techniques demonstrated equally that consumption of HAMSB provided a protective effect by reducing colonic adduct load compared with the LAMS diet (P < .05). In addition, IHC allowed visualization of the O6MeG distribution, where adduct load was reduced in the lower third of the crypt compartment in HAMSB-fed rats (P = .036). The apoptotic response to AOM was higher in the HAMSB-fed rats (P = .002). In conclusion, the reduction in O6MeG levels and enhancement of the apoptotic response to DNA damage in the colonic epithelium through consumption of HAMSB provide mechanistic insights into how HAMSB protects against colorectal tumorigenesis.