The effect of hyperlipidemia (HL) on the pharmacokinetics of nelfinavir (NFV), a human immunodeficiency virus protease inhibitor, was investigated, focusing on the change of NFV distribution in plasma using poloxamer 407-induced HL model rats (HL rats). The plasma unbound fraction (f_u) in HL rats (0.20—0.39%) was significantly lower than the control (0.92—1.47%). Lipoprotein level in HL rats was about 5 times higher and low- and very low-density liproteins ratio were 1.7—4.5 times higher than the control. NFV recovery in the triglyceride-rich lipoprotein such as chylomicron, low- and very low-density liproteins fractions of HL rats were significantly higher. The area under the plasma concentration–time curve (AUC) of NFV after intravenous (i.v.: 5 mg/kg) and intraduodenal (i.d.: 30 mg/kg) administration to HL rats (i.v.: 6.12±0.48, i.d.: 24.4±2.2 μg·h/ml) were higher than the control (i.v.: 1.62±0.21, i.d.: 5.00±0.36 μg·h/ml). The steady state volume of distribution (Vd_ss) in HL rats (0.60±0.07 l/kg) was lower than the control (6.25±0.55 l/kg). Systemic availability (F) in HL rats (66.6%) was higher than the control (51.4 %). Directly absorbed NFV from the gastrointestinal tract to the lymphatic system in HL rats was about 2 times higher than the control. From the results of this study, it was concluded that the increase of AUC was caused by decreasing f_u and Vd_ss due to the increase of the triglyceride-rich lipoprotein level. In addition, it was suggested that the increase of absorbed NFV through the lymphatic system, which did not receive the first-pass effect, was one reason for the increase of F in HL rats.