Amino acid analysis for three experimental
diets was carried out by ion-exchange
chromatography under the experimental
conditions. For protein hydrolysis, samples
containing 5.0 mg of protein were hydrolyzed
with 1.0 mL of 6 N HCl in vacuum-sealed
hydrolysis vials at 110°C for 22 h. Norleucine
was added to HCl as an internal standard. After
hydrolysis, the hydrolysis vials were opened
and stored for five to six days in a desiccator
containing NaOH pellets under vacuum until
dry. Then the residue was dissolved in a
suitable volume of a sample dilution of Na-S
buffer and filtered through a Millipore
membrane (0.22 µm pore size). The amino
acids were analyzed by ion-exchange
chromatography in a Beckman instrument
(model 7300, USA) equipped with an automatic integrator.