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A prospective, unmatched case control study was performed to identify dietary and environmental risk factors for enterolithiasis in horses in California and to determine whether colonic ingesta analyses differed between horses with and without enteroliths. Forty-three horses with enterolithiasis were compared with 19 horses with surgical colic attributable to nonstrangulating obstruction of the colon without enteroliths. Colonic ingesta samples were collected at surgery from horses with enteroliths and control horses. Colonic pH and colonic concentrations of magnesium, phosphorus, sulfur, sodium, calcium, potassium, and nitrogen were measured. Questionnaires were distributed to owners to determine diet and management practices. Student's t-test and Mann-Whitney tests were used to evaluate differences in pH, dry matter content, percent nitrogen, and mineral content. Associations between dietary and management risk factors and enterolith occurrence were quantified by odds ratios. Mean pH of colonic contents from horses with enterolithiasis was significantly higher than for control horses. Horses with enterolithiasis had significantly lower percent dry matter in colonic fecal samples and higher mean mineral concentrations than controls. On the basis of reported feeding and management practices, horses with enterolithiasis were fed a significantly higher proportion of alfalfa in their diet and were less likely to have daily access to pasture grass than horses without enteroliths. Results suggest that decreasing alfalfa consumption and allowing daily access to pasture grazing might reduce the risk of enterolithiasis. Dietary modifications promoting acidification of colonic contents and dilution of minerals might be beneficial as preventive measures for enterolithiasis in horses.