Determination of a steady-state isotope dilution protocol for carbon oxidation studies in the domestic cat

The present study aimed to develop an isotope protocol to achieve equilibrium of ¹³CO₂ in breath of cats during carbon oxidation studies using L-[1-¹³C]-Phenylalanine (L-[1-¹³C]-Phe), provided orally in repeated meals. One adult male cat was used in two experiments. In each experiment, three isotope protocols were tested in triplicate using the same cat. During carbon oxidation study days, the cat was offered thirteen small meals to achieve and maintain a physiological fed state. In experiment 1, the isotope protocols tested (A, B and C) had a similar priming dose of NaH¹³CO₃ (0⋅176 mg/kg; offered in meal 6), but different priming [4⋅8 mg/kg (A) or 9⋅4 mg/kg (B and C); provided in meal 6] and constant [1⋅04 mg/kg (A and B) or 2⋅4 mg/kg (C); offered in meals 6-13] doses of L-[1-¹³C]-Phe. In experiment 2, the isotope protocols tested (D, E and F) had similar priming (4⋅8 mg/kg; provided in meal 5) and constant (1⋅04 mg/kg; provided in meals 5-13) doses of L-[1-¹³C]-Phe, but increasing priming doses of NaH¹³CO₃ (D: 0⋅264, E: 0⋅352, F: 0⋅44 mg/kg; provided in meal 4). Breath samples were collected using respiration chambers (25-min intervals) and CO₂ trapping to determine ¹³CO₂:¹²CO₂. Isotopic steady state was defined as the enrichment of ¹³CO₂, above background samples, remaining constant in at least the last three samples. Treatment F resulted in the earliest achievement of ¹³CO₂ steady state in the cat's breath. This feeding and isotope protocol can be used in future studies aiming to study amino acid metabolism in cats.