Apolipoprotein B100 (apoB100) is an essential component of very low density lipoprotein (VLDL) and low-density lipoprotein (LDL), both independent markers of cardiovascular risk. Nicotinic acid (NA) is an efficacious drug for decreasing VLDL and LDL, but the underlying mechanisms are unclear. For this purpose, six obese insulin-resistant dogs were given 350 mg/day of NA for 1 week and then 500 mg/day for 3 weeks. Turnover of apoB100-containing lipoproteins was investigated using stable isotope-labeled tracers. Multicompartmental modeling was used to derive kinetic parameters before and at the end of NA treatment. Hepatic diacylglycerol acyltransferase 2 (DGAT2), microsomal triglyceride transfer protein (MTP), hepatic lipase (HL), and adipose lipoprotein lipase (LPL) mRNA expression was also determined. NA treatment decreased plasma triglyceride (TG) (p < 0.001), VLDL-TG (p < 0.05), total cholesterol (p < 0.0001), and LDL cholesterol (p < 0.05), whereas plasma nonesterified fatty acids were unchanged. The decrease in VLDL-apoB100 concentration (p < 0.001) was the result of a lower absolute production rate (APR) (p < 0.001), despite a moderate decrease (p < 0.05) in fractional catabolic rate (FCR). LDL-apoB100 concentration was reduced (p < 0.05), an effect related to a decrease in LDL APR (p < 0.05) and no change in FCR. NA treatment reduced DGAT2 expression (p < 0.05), whereas MTP, HL, and LPL expression was unchanged. Our results suggest that NA treatment reduced VLDL and LDL concentration as a consequence of a decrease in VLDL production.