A composition of dietary fat considerably affects the metabolism of nutrients, especially high-fat diet is associated with various metabolic disorders, diabetes and cardiovascular diseases in humans. Studies on rodents are an established model to investigate the pathophysiology of excessive fat in diets, hence the aim of the study was to evaluate the effect of dietary fats different in composition on plasma lipids, glucose and insulin levels in non-genetically modified rats instead of genetic models of obesity or diabetes. Animals were fed purified low- or high-fat (12 and 40% energy from fat, respectively) diets containing the same fat sources – lard, sunflower oil or menhaden oil for six weeks. Feeding menhaden oil, both in low- and high-fat diets, resulted in decreased plasma triglyceride, total cholesterol and high-density lipoprotein cholesterol levels in comparison to diets composed of lard or sunflower oil. However, high-fat diet with menhaden oil caused an increase in plasma glucose and insulin levels, which resulted in the increased HOMA-IR (Homeostatic Model Assessment – Insulin Resistance) index in relation to low-fat diet with the same oil. The obtained results demonstrate that high menhaden oil intake has the hypolipidemic effect but can impair glucose homeostasis in non-genetically modified rats.