1. Nonessential Amino Acid Metabolism in Healthy Adult Males Leah Cooper Department of Pediatrics, University of British Columbia & Child and Family Research Institute, BC Children's Hospital

2. Outline  Background  Protein  Amino Acids  Indicator Amino Acid Oxidation Technique  Objective & Hypothesis  Methods  3 Steps in Study  Results  Conclusion  Future Directions

3. Background - Protein  Structural component of the body  Enzymes, Hormones, Neurotransmitters, Transport Molecules  Made of 20 Amino Acids  Need all 20 in correct proportions for protein synthesis to occur in the body www.onlymyhealth.com

4. Background – Amino Acids www.thehealthsite.comwww.rpi.edu

5. Background - Evolution http://www.nature.com/scitable/topicpage/an-evolutionary-perspective-on-amino-acids-14568445

6. Background - Nitrogen  Play many metabolic roles in the body  The requirement for total nitrogen is the primary determinant of total protein requirement  If total amino nitrogen is inadequate, synthesis of nonessential amino acids and thus protein is less than maximal

7.  Some nonessential amino acids (NEAA) may supply nitrogen better than others  Rose et al. (1948) showed increased growth by 70- 75% in rats fed essential amino acids (EAA) + NEAA  Rechcigl et al. (1957) showed different growth rates in rats  Glutamate gave the largest increase in growth  Followed by Alanine, Aspartate, Asparagine, Proline, and Glutamine  Smallest increases in growth were Glycine and Serine Background – Nitrogen Supply

8. Background - Indicator Amino Acid Oxidation Technique  Previous Methods (N balance) were long in duration, very invasive and impractical  The stable isotope based IAAO is rapid, minimally invasive and accurate  IAAO measures the oxidation of the tracer 13 C-PHE to 13 CO 2 in breath  Oxidation is inversely proportional to protein synthesis  High Oxidation = Low Protein Synthesis

9. Objective: To determine the metabolic demand of 9 of the NEAA using the indicator amino acid oxidation (IAAO) method in healthy adult males Hypothesis: There will be a hierarchy among the 9 NEAA, more specifically Alanine, Arginine, Asparagine, Aspartate, Glutamine, Glutamate, Glycine, Proline, and Serine, to supply Nitrogen to the body for the de novo synthesis of the other NEAA and for protein

10. Methods Preliminary Study AdaptationStudy Day

11. Preliminary Study  Target Population:  Healthy adult males  Between the ages 20-40  classified as normal body weight and BMI (18.5-25 kg/m 2 )  Background Information:  Medical & Dietary histories  Anthropometric measurements  Body composition  Resting energy expenditure (REE)  Informed consent

12. Methods Preliminary Study AdaptationStudy Day

13.  2 days prior to each study day  Adaptation diets contain a protein intake of 1.0 g/kg and a caloric intake of 1.7 × REE  Provided as 6 milkshakes. Adaptation products.nestle.ca

14. Methods Preliminary Study AdaptationStudy Day

15.  10 Study Days  Randomized  Diet:  The Base Diet (0.34 g/kg/day)  Only EAA  The Base Diet + 1 NEAA (0.73 g/kg/day)  50:50 ratio of EAA: NEAA  5 th Meal starts the isotope tracer protocol with an oral dose of L-1- 13 C- Phenylalanine Study Day Diet Base + ALA BASE (EAA) Base + SER Week 1 Week 2 Base + GLN BASE + PRO Base + ARG Week 4 Week 5 Etc…for 10 weeks Study Day 1Study Day 2Study Day 3 Study Day 4Study Day 5Study Day 6

16. Study Day Sampling  Sample Collection  Breath and Urine  Baseline  Isotopic Steady State  Carbon dioxide production (VCO2)

17.  13 CO 2 oxidation values analyzed using repeated measures ANOVA (Graphpad PRISM) Sample Analysis & Statistics Multi-flow Continuous Isotope Ratio Mass Spectrometer

18. Results – 50 Study Days

19. The first study to look at NEAA in humans We suggest all NEAA can satisfy the metabolic demand for total nitrogen. Conclusion

20.  Determine whether a difference in NEAA metabolic demand exists in more vulnerable populations Future Directions www.vancitychiro.com

21. Acknowledgments  Dr. Rajavel Elango  Lab members  Ajinomoto Inc. Co.  Child and Family Research Institute  University of British Columbia  All my participants!

22.  Rose, W.C., Oesterling, M.J., and Womack, M. (1948). Comparative growth on diets containing ten and nineteen amino acids with further observations upon the role of glutamic and aspartic acids. J Biol Chem 176: 753-762.  Rechcigl Jr., M., Loosli, J. K., Williams, H. H. (1957). The net utilization of non-specific nitrogen sources for the synthesis of non-essential amino acids I. Growth and nitrogen utilization. J Nutr 63: 177-192.  Millward DJ. Methodological considerations. Proc Nutr Soc 2001;60: 3–5.  Food and Agriculture Organization of the United Nations. Energy and Protein Requirements (1991). References

23. QUESTIONS ?? Thank you for listening !!

24. Study Diet Study Day Base Diet