Objective: To investigate plasma-free carnitine (Fc), acylcarnitine (Ac), and total carnitine (Tc) levels in patients undergoing hemodialysis (HD), and to explore their clinical significance.Methods: A total of 20 subjects were in the normal control group and 133 patients undergoing HD were divided into medicated (received carnitine treatment) and non-medicated groups. The medicated group was further divided into three subgroups according to Fc level: Fc=80-199, 200-299, and300mol/L. We used non-derivative tandem mass spectrometry to determine carnitine levels, and clinical symptoms such as weakness, hypotension, and muscle cramps were recorded during dialysis.Results: Fc and Tc levels were significantly lower in the non-medicated group than in the control group, whereas Fc, Ac, and Tc levels were higher in the medicated than non-medicated group (p<.05). The medicated group had fewer symptoms during dialysis than the non-medicated group such as weakness, hypotension, and muscle cramps (p<.05). An additional comparison showed that the incidence rates of hypotension and muscle cramps in the Fc<80-199mol/L group were significantly lower than those in the Fc300mol/L medicated and non-medicated groups.Conclusions: Patients undergoing HD have low carnitine levels. l-Carnitine can effectively increase Fc concentration and improve clinical symptoms; however, only the proper Fc range can reduce complications caused by dialysis. Thus, this range needs to be determined.