Background: Fasciola spp. is endemic in Iran, and even in recent years, some human epidemics of this parasite have been reported in the northern parts of Iran. Biliary fluid is one of the most important substances that is altered by invasion and replacement of the parasite, and it seems that biliary fluid can be used to determine the progression of the disease and the degree of the damage to the liver cells. The aim of this study was to investigate whether there was a relationship between the amount of bilirubin, phosphorus, calcium and cholesterol in the biliary fluid of healthy livers and those in livers infected with Fasciola spp. Methods: A total of 50 samples of biliary fluid from the infected livers and 50 samples of biliary fluid from healthy livestock in terms of presence or absence of contamination were collected. Results: Direct bilirubin and cholesterol in infected livers is lower than those in healthy livers. Also, the level of calcium in the liver contaminated with the parasite was higher than that of the healthy liver; the difference was statistically significant. The level of phosphorus in the infected livers was higher than that in the healthy liver, however, the difference was not statistically significant. Conclusions: This study showed that bilirubin, cholesterol, calcium, and phosphorus in the liver consistent with hepatic fascioliasis were altered significantly, indicating valuable effects of the parasites on hepatic cells.