Bray · Nutrients 2020 · randomized controlled parallel-arm trial · n=23

Effect of Overeating Dietary Protein at Different Levels on Circulating Lipids and Liver Lipid: The PROOF Study.

Cited 2 times in the scientific literature.

Level 2 - randomized trial

Individual randomized controlled trial

PubMed 33322340 · doi:10.3390/nu12123801 · record verified 2026-08-27

What was done

Twenty-three healthy adults (14 men, 9 women) were enrolled in an 8-week parallel-arm randomized trial and overfed by approximately 40% above baseline energy requirements. Participants were assigned to diets containing 5% (low protein, LPD), 15% (normal protein, NPD), or 25% (high protein, HPD) protein, with fixed carbohydrate intake (causing dietary protein and fat to vary inversely). Body composition was measured using dual-energy X-ray absorptiometry (DXA), intrahepatic lipid/liver density was evaluated via computed tomography (CT), and circulating lipids and metabolites were assessed.

What was found

Overfeeding with the low-protein diet was associated with significant increases in HDL-cholesterol (p < 0.001) and free fatty acids (p = 0.034), along with significant decreases in fat-free mass (p < 0.0001) and liver density (indicating increased intrahepatic lipid, p = 0.038). In statistical modeling, dietary protein intake independently predicted changes in fat-free mass (p = 0.0040), but not blood or liver lipids. Instead, dietary fat intake was the primary predictor of changes in HDL-cholesterol (p = 0.014), free fatty acids (p = 0.0016), and liver fat (p = 0.0007).

Why it matters

These findings differentiate the metabolic effects of macronutrients during weight gain, demonstrating that while protein level dictates lean tissue accrual or loss, the accompanying fat content of the hypercaloric diet drives hepatic steatosis and circulating lipid alterations.

Limits

The total sample size was very small (n = 23 across three parallel arms). Because carbohydrate content was kept constant, protein and fat intake were co-dependent, making it impossible to evaluate protein effects completely independent of dietary fat variations. Long-term metabolic outcomes beyond the 8-week intervention were not evaluated.

Cited by