The therapeutic effects of iron, zinc and magnesium trace elements, as well as rifaximin were investigated and compared in HE rats. In this study, HE rats were treated with either ferrous sulfate (HE-Fe, 30 mg/kg/day), zinc sulfate (HE-Zn, 30 mg/kg/day), magnesium sulfate (HE-Mg, 50 mg/kg/day) or rifaximin (HE-Rf, 50 mg/kg/day), which was mixed with water and administered orally for 61 days. The Morris water maze (MWM) and open-field tests were used to evaluate cognitive and locomotor function. The blood ammonia levels before and after administration of the glutamine challenge test, manganese concentration and glutamine synthetase (GS) activity were measured. Significantly longer MWM escape latencies, less locomotor activity, higher blood ammonia levels, higher brain manganese concentrations and higher GS activity were observed in HE rats. However, HE-Mg and HE-Rf rats had significantly shorter MWM escape latencies, increased locomotor activity, lower blood ammonia, lower brain manganese concentrations and lower GS activity. Partial improvements were observed in HE-Fe and HE-Zn rats. The results indicated that oral administration of magnesium can significantly improve the cognitive and locomotor functions in HE rats by reducing the brain manganese concentration and regulating GS activity.Hepatic encephalopathy (HE) is a major complication of cirrhosis and (or) portal-systemic shunting 1 . Its pathogenesis is not completely clear. It has been suggested that nitrogen metabolism dysfunction may cause neurological symptoms 2 . HE significantly affects the quality of daily life and may ultimately lead to death; however, HE can be reversed with the appropriate treatment 1, 2 . HE patients are known to have higher levels of ammonia in their blood. Thus, current HE treatments focus on non-absorbable disaccharides (e.g., lactulose) or antibiotics (e.g., rifaximin) to reduce ammoniagenesis in the intestines and lower blood ammonia levels 3 . Recent studies showed that trace element concentrations varied in cirrhotic patients with HE 4 . Lower serum zinc, magnesium and iron but higher serum manganese and copper were reported in cirrhotic patients [5][6][7] . Zinc, magnesium, and iron are co-enzymes of a variety of metabolic enzymes. Deficiencies in zinc, magnesium and iron levels may lead to metabolic disorders and negatively impact behavior and cognition [8][9][10] . On the other hand, high manganese levels can be neurotoxic. Excessive levels of manganese lead to cognitive impairments and extrapyramidal symptoms 11 . Oral administration of zinc effectively improves HE symptoms 12 . However, whether oral administration of iron or magnesium is also beneficial is not known. Previous studies demonstrated that trace elements, such as iron and magnesium, can influence manganese absorption in the intestines 13,14 . We hypothesize that iron and magnesium supplementation may improve HE by reducing intestinal manganese absorption and subsequently
