Calibration and validation of a carbon oxidation system and determination of the bicarbonate retention factor and the dietary phenylalanine requirement, in the presence of excess tyrosine, of adult, female, mixed-breed dogs1,2,3

Abstract

Carbon oxidation methods have been used as rapid and sensitive methods to determine whole-body AA requirements in multiple species. The objectives of the current studies were to validate complete CO recovery, determine the bicarbonate retention factor, and estimate the Phe requirement, in the presence of excess Tyr, in adult dogs using the direct oxidation technique. In this series of studies, 2 oxidation chambers were constructed and calibrated to ensure accurate collection of breath CO. First, 104.6 ± 7.1% CO was recovered from chambers and suggests that the chambers were appropriately designed for complete and efficient CO recovery. Second, we determined bicarbonate retention in 5 dogs using repeated oral dosing of a bicarbonate tracer (NaHCO) with small meals. At isotopic and physiological steady state, 102.5 ± 2.6% of the delivered NaHCO was recovered in breath. Third, the Phe requirement, when Tyr was supplied in excess, was determined by the rate of appearance of CO in the breath (CO). Dogs ( = 5) were fed test diets with different concentrations of Phe ranging from deficient to excessive for 2 d prior to conducting the tracer studies. The mean Phe requirement (when Tyr was supplied in excess) was 0.535% of diet (upper 95% confidence interval = 0.645% diet) on an as-fed basis or 0.575% of diet (upper 95% confidence interval = 0.694% of diet) on a DM basis and was based on a calculated (modified Atwater calculation) dietary ME density of 3.73 Mcal/kg DM. These data support the use of carbon oxidation methods and oral dosing of isotope to measure whole-body requirements of indispensable AA in adult dogs and suggest the current recommendations may be low.

Dietary phenylalanine requirements are similar in small, medium, and large breed adult dogs using the direct amino acid oxidation technique1

“…The effect of Phe content in the test diet on F 13 CO 2 was analyzed using PROC MIXED of SAS (v. 9.4; SAS Institute Inc., Cary, NC) with diet as a fixed effect and dog as a random effect. The estimate of the mean Phe requirement and the upper 95% confidence intervals (CI) for individual dog breeds and pooling data from all breeds were derived by breakpoint analysis of the F 13 CO 2 /kg BW using a two-phase linear regression model as previously reported (Shoveller et al, 2017). Differences in AA concentration in blood were determined using orthogonal contrast comparing each dietary Phe content against the lowest Phe diet (0.24%).…”

“…During the study, dogs were pair-housed in kennels (2.4 × 2.4 m) in a temperature-controlled building (22 °C) and with a lighting schedule of 12 h:12 h light:dark. Dogs received similar amounts of daily socialization, exercise, and regular veterinary care as previously reported (Shoveller et al, 2017).…”

“…The constants 44.6 (mmol/mL) and 60 (min/h) convert the FCO 2 to micromoles per hour; Arg = arginine; Cys = cysteine; His = histidine; Ile = isoleucine; Leu = leucine; Lys = lysine; Met = methionine; Phe = phenylalanine; Thr = threonine; Trp = tryptophan; Tyr = tyrosine; Val = valine. the factor 100 changes APE to a fraction; and, the 1.0 is the retention factor of CO 2 in the body due to bicarbonate fixation as reported previously (Shoveller et al, 2017). Resting and fed energy expenditure (EE) was calculated based on VO 2 and VCO 2 based on modified Weir equation (Weir, 1949).…”

“…NRC (2006) states that: "No individual dose-response peer-reviewed reports could be found for the minimal requirements of any of these amino acids -including Phe-for maintenance on dogs." Shoveller et al, (2017) determined phenylalanine (Phe) requirements for adult dogs. However, there are vast phenotypic differences among dog breeds that may contribute to differences in macronutrient metabolism, potentially modifying AA requirements that have not been accounted for.…”

“…Studies in humans, pigs, and recently in dogs have validated the use of carbon oxidation techniques to determine AA requirements at a whole body-tissue level (Bross, et al 1998;Moehn, et al, 2004;Pencharz and Ball, 2003;Shoveller et al, 2017). Comparison of AA recommendations based on nitrogen balances or carbon oxidation techniques suggests that nitrogen balance underestimates AA requirements by ~40% (Zello et al, 1995;Elango et al, 2008).…”