The etiology of the hypertriglyceridemia in alloxan-diabetic rabbits was studied by two independent methods. Production and removal rates of VLDL triacylglycerol were measured in diabetic rabbits by injection of [3H]palmitate-labelled VLDL. Similarly, triacylglycerol total removal rates were determined in non-diabetic rabbits which were infused with Intralipid to mimic the plasma triacylglycerol concentrations of diabetic rabbits. Compared to nondiabetic rabbits, triacylglycerol removal rats were decreased in diabetic rabbits, particularly at higher levels of plasma triacylglycerol. During cholesterol and triacylglycerol supplementation of the diet, post-heparin plasma lipoprotein lipase activity of diabetic rabbits with severe hypertriglyceridemia averaged 36% of that of nondiabetics, suggesting an impaired triacylglycerol removal capacity. Furthermore, plasma triacylglycerol was inversely related to post-heparin plasma lipoprotein lipase activity among diabetic rabbits. VLDL triacylglycerol production increased with increasing plasma triacylglycerol concentration among diabetic cholesterol-fed rabbits with moderately severe hypertriglyceridemia, but reached an apparent plateau among rabbits with plasma triacylglycerol concentrations from approx. 2000-9000 mg/dl. Thus, severe hypertriglyceridemia in this model of insulin deficiency can be attributed only partially to VLDL hypersecretion, whereas a removal defect, resulting in saturation of the triacylglycerol removal mechanism, appears to be largely responsible. The impaired removal of plasma triacylglycerol is also related to the presence of cholesterol predominantly in lipoproteins of increased size. The data support the hypothesis that protection against atherosclerosis in cholesterol-fed diabetic rabbits results from exclusion of very large cholesterol-containing lipoproteins from the arterial wall.