The objective of this study was to assess the protein quality of the myofibrillar and connective tissue proteins of chicken gizzard. Protein fractions were isolated from White Leghorn chicken gizzards and quantified by detailed amino acid analysis. This quantification involved repeated extractions of ground gizzards first with Triton X-100, then with low ionic strength imidazole-buffered saline (pH 7.1), followed by either 2% SDS or by 5 M guanidine hydrochloride. The total soluble intracellular protein fraction averaged 86.3% of the total protein and the insoluble extracellular connective tissue proteins comprised the remaining 13.7%. These fractions differed significantly in their essential amino acid (EAA) profiles, with the soluble intracellular fraction having the highest percentage EAA9 (48.6 to 49.0%) and the insoluble connective tissue fraction varying from 20.8 to 23%, compared to the FAO/WHO reference pattern value of 33.9% for a 2- to 5-yr-old child. Calculated protein efficiency ratios (PER) for intracellular proteins averaged 3.02 compared with a value of 1.65 for the extracellular matrix proteins. These results provide an accurate assessment of the protein quality of smooth muscle proteins of chicken gizzard and may prove valuable for industrial control of the amount of connective tissue added to formulations of meats and poultry products.