Anything we dont need for S is not stored N waste products Labeled AA has 2

# Anything we dont need for s is not stored n waste

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certain proportion is taken out for protein S. Anything we don’t need for S is not stored N waste products - Labeled AA has 2 fates once it enters this pool 1. Synthesized into proteins or 2. Oxidized and made into a N-containing waste products which would be found in the urine (measure amount of 15 N in collected urine) If found in very low amounts in the urine means labeled AA was not oxidized and had to be used to synthesize proteins o § There’s an indirect correlation: if you don’t find the 15 N in the urine then you can assume that it had to be used for protein S 9. DETERMINING RATE OF S USING SPM VIA TRACER ENRICHMENT - Rate of protein S can be determined by measuring 15 N enrichment in urine over time o When you administer the tracer AA: You know the rate of administration (mg labeled AA/ hr) so you know how much is going into this pool (I) You know the rate at which is being degraded b/c you’re collecting urine (E) You can calculate synthesis rate from Q = S + E - Enrichment measured by a graph ( 15 N/ 14 N) o Insert graph here - Eventually all AA body pools will be equally enriched/ labeled with tracer-AA (original studies administered the label AA at a very slow rate over time intravenously) o Enrichment = 15 N/ 14 N (if measuring urea it would be 15 N urea/ 14 N urea) As you begin slowly infusing the 15 N-Gly, it goes into circulation and enriches/goes into all the different cells and tissues of the body This takes time and eventually changes the 15 N/ 14 N Gly in every body tissue until all the pools get labeled simultaneously This means the 15 N Gly is treated the same way as unlabeled Gly ( 14 N- Gly) & rate of entry of 15 N- Gly is = rate of exit from the pool (SA) o Either they’ll go into protein S together or they ’l l be excreted as urea of 15 N-NH 3 together
SFLORES_2/24/2014 7 Labeling of all pools takes ~24 hrs of constant infusion 10. SPECIFIC ACTIIVTY PLATEAU - Specific activity of 15 N (SA): Rate of entry of 15 N-Gly = Rate of exit from the pool o Q=amount of N entering the pool = amount exiting the pool o Turnover (Q) = ???? ?? ?????? 𝑖????𝑖?? (?? ?????/ℎ?/?? ?𝑊) 𝑆? ??????? SA = 15 N/ 14 N If looking at 15 N vs 14 N urea this is saying that maybe 1 hr into infusion, 0.01% of all the N atoms in urea is due to 15 N (very low amount) That’s the enrichment: out of all the N atoms from urea in the urine only 0.01% is enriched with the 15 N label. o As you continue to infuse the label, it labels all of the body tissues together and that begins to the amount of enrichment until you reach steady state (can continue for long periods of time and it ll remain the same). Steady state is where the plateau was reached. This is where you know the rate of entry of the isotope = rate of exit of the isotope, and you can take measurement. 11. USING DIFFERENT LABELS BESIDES AAs - Can also measure other things: 1. Label urea and measuring NH 3 . Measuring 15 N-NH 3 / 14 N-NH 3 (any N waste products) This is going to be a lot less b/c we excrete a lot less NH 3 than urea but you can do that 2. Use C label in carboxyl group ( 14 C-radioactive or 13 C-stable) and measure CO 2 . When you decarboxylate an AA, you breathe out the -COOH as CO 2 and collect that

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• Spring '14
• Knutson,MitchellD
• NRG

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