Determination of Threonine Requirement in Adult Males With Crohn's Disease Using the Indicator Amino Acid Oxidation (IAAO) Methodology

The goal of the current study is to measure the requirement for threonine in patients with CD using the IAAO method and compare the requirement to previously determined threonine requirement estimated in young adults using the IAAO technique. It is hypothesize that the requirement for threonine in patients with CD will be higher than the threonine requirement previously determined in young adults using the IAAO method. Up to 10 clinically stable patients with CD will be recruited from the IBD Clinic at Mt. Sinai Hospital, Toronto, and subsequently followed up at the Clinical Research Center (CRC), The Hospital for Sick Children (SickKids), Toronto, Canada. Before the study begins, the participants will be required to visit the CRC (Room 5500 Hill Wing, The Hospital for Sick Children) for a pre-study assessment of their height, weight, fat mass, fat free mass, resting metabolic rate and medical history. These assessments will take about 3 hours to complete. They will need to have been fasted for 10 hours prior to the pre-study assessment. The pre-study assessment is needed to calculate their dietary requirements for the study, and to assess health status. After signing the consent form, the subjects will complete the screening procedures (height, weight, fasting blood sample and medical history questionnaire, BIA, Skinfold and calorimetry).

Each study will consist of a 2-day adaptation period to a prescribed diet in accordance with the energy requirement of the subject and 1-study day. The diet will provide an adequate amount of protein, of 1 g protein/kg/d. The 2-day adaptation period is to allow the body to adapt to an adequate amount of protein as it has been shown that protein kinetics is altered without it.

Dietary intakes during this time will be provided in the form of lactose-free milk shakes (Scandishake) with added carbohydrate (SolCarb) and protein (beneprotein) to meet the subjects' requirement.

Following the 2 days of adaptation is the study day where threonine intake will be randomly assigned and phenylalanine (Phe) kinetics will be measured with the use of isotopically labelled Phe. On this day, VCO2 will be measured by calorimetry immediately after the 5th meal for a period of 20 minutes.

On the study day (3rd day of each 3-day period), the diet will be provided as 8 hourly isocaloric, isonitrogenous meals made up of a flavored liquid formula and protein free cookies developed for use in amino acid kinetic studies. Each meal will represent 1/12th of the subject's total daily requirements. The nitrogen (protein) content of the diet will be provided in the form of a crystalline amino acid mixture based on the amino acid composition of egg protein.

• A daily multivitamin supplement will be provided during the study period.
• No other food or beverages will be consumed on the adaptation days except water, 1 cup clear tea, or 1 cup clear coffee.
• During the 8-hr study day, no other food or drink will be consumed except water.
• Urine and breath samples will be collected at baseline and at isotopic steady state.
• Breath samples will be collected simultaneously with urine samples.
• Five baseline breath samples will be collected 60, 45, 30, 15 min, and just before the tracer protocol begins.
• Three baseline urine samples will be collected 60, 30 min, and just before the tracer protocol begins.
• Four plateau breath samples will be collected every 15 minutes 2.5 h after the tracer protocol begins.
• Three plateau breath samples will be collected every 30 minutes 2.5 h after the tracer protocol begin
• Breath samples will be collected with subjects breathing into an Exetainers plastic tube and samples will be stored in pre-evacuated glass tubes at room temperature until analysis.
• Urine samples will be collected in Eppendorf tubes and stored at - 20 º C until analyzed for 1-13C phenylalanine enrichment.
• The rate of CO2 production (VCO2) will be measured on each testing day using a ventilated hood indirect calorimeter at meal 5 to quantify 13CO2 excretion in breath.

Subjects can choose to withdraw from the study at any time and for any reason, based on his/her individual judgment. In particular, if a subject is unable to tolerate the diet, whether it is regards to taste, loose stools or constipated stools, he/she has the right to withdraw at any time during the study.