Skip to main content
SpringerLink
Log in

Amino acid flooding doses for measuring rates of protein synthesis

  • Review Article
  • Published:
Amino Acids Aims and scope Submit manuscript

Summary

The development, advantages and disadvantages of using the amino acid flooding dose technique to determinein vivo rates of protein synthesis are examined in this review. A discussion of the use of this procedure in animals greater than 5 kg is included. The flooding dose procedure reduces the disparity between isotope enrichment in different amino acid precursor pools, which should theoretically improve the precision and accuracy of protein synthesis measurements. However, the possibility must be considered that the large doses of amino acids injected or infused in conjunction with this technique may influence protein turnover due to attendant metabolic effects. Therefore, a judicious choice of an amino acid and an evaluation of the experimental parameters involved in this procedure are required to optimize the accuracy of results obtained.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Airhart J, Vidrich A, Khairallah EA (1974) Compartmentation of free amino acids for protein synthesis in rat liver. Biochem J 140: 539–548

    PubMed  Google Scholar 

  • Attaix D, Arnal M (1987) Protein synthesis and growth in the gastrointestinal tract of the young preruminant lamb. Br J Nutr 58: 159–169

    PubMed  Google Scholar 

  • Attaix D, Manghebati A, Grizard J, Arnal M (1986) Assessment ofin vivo protein synthesis in lamb tissues with [3H]valine flooding doses. Biochim Biophys Acta 882: 389–397

    PubMed  Google Scholar 

  • Attaix D, Papet I, Grizard J, Arnal M (1987) Absence d'effets de l'injection de surcharges de L-valine et L-leucine sur les teneurs plasmatiques en insuline et en glucagon de l'agneau préruminant. Reprod Nutr Develop 27: 297–298

    Google Scholar 

  • Attaix D, Aurousseau E, Bayle G, Rosolowska-Huszcz D, Arnal M (1988a) Respective influences of age and weaning on skeletal and visceral muscle protein synthesis in the lamb. Biochem J 256: 791–795

    PubMed  Google Scholar 

  • Attaix D, Aurousseau E, Manghebati A, Arnal M (1988b) Contribution of liver, skin and skeletal muscle to whole-body protein synthesis in the young lamb. Br J Nutr 60: 77–84

    PubMed  Google Scholar 

  • Attaix D, Aurousseau E, Bayle G, Rosolowska-Huszcz D, Manghebati A, Arnal M (1989) Influences of age and weaning onin vivo pancreatic protein synthesis in the lamb. J Nutr 119: 463–470

    PubMed  Google Scholar 

  • Baracos VE, Brun-Bellut J, Marie M (1991) Tissue protein synthesis in lactating and dry goats. Br J Nutr 66: 451–465

    PubMed  Google Scholar 

  • Davis SR, Barry TN, Hughson GA (1981) Protein synthesis in tissues of growing lambs. Br J Nutr 46: 409–419

    PubMed  Google Scholar 

  • Dorup I, Clausen T (1989) Effects of potassium deficiency on growth and protein synthesis in skeletal muscle and the heart of rats. Br J Nutr 62: 269–284

    PubMed  Google Scholar 

  • Early RJ, Thompson JR, Christopherson RJ (1989) Net blood exchange of branched-chain amino andα-keto acid across the portal-drained viscera and hindlimb of cattle during infusions of leucine and insulin. Can J Anim Sci 69: 131–140

    Google Scholar 

  • Early RJ, McBride BW, Ball RO (1990) Growth and metabolism in somatotropin-treated steers: III. Protein synthesis and tissue energy expenditures. J Anim Sci 68: 4153–4166

    PubMed  Google Scholar 

  • Everett AW, Prior G, Zak R (1981) Equilibration of leucine between the plasma compartment and leucyl-tRNA in the heart, and turnover of cardiac myosin heavy chain. Biochem J 194: 365–368

    PubMed  Google Scholar 

  • Ferrara M, Arnal M, Fauconneau G (1977) Mesure de la synthèse protéiquein vivo à l'aide du14C lysyl-tRNAlys dans le foie d'agneau en croissance. Ann Biol anim Bioch Biophys 17: 1061–1068

    Google Scholar 

  • Ford GC, Cheng KN, Halliday D (1985) Analysis of (1-13C)leucine and (13C)KIC in plasma by capillary gas chromatography/mass spectrometry in protein turnover studies. Biomed Mass Spec 12: 432–436

    Google Scholar 

  • Funabiki R, Yagasaki K, Hara H, Nyumura N, Takeda A (1990) Measurement of the rate of whole body protein synthesis by intraperitoneal injection of a large dose of alanyltyrosine with [14C]tyrosine. Agr Biol Chem Tokyo 54: 113–119

    Google Scholar 

  • Garlick PJ, Grant I (1988) Amino acid infusion increases the sensitivity of muscle protein synthesisin vivo to insulin: effect of branched chain amino acids. Biochem J 254: 579–584

    PubMed  Google Scholar 

  • Garlick PJ, Millward DJ, James WPT (1973) The diurnal response of muscle and liver protein synthesisin vivo in meal fed rats. Biochem J 136: 935–945

    PubMed  Google Scholar 

  • Garlick PJ, McNurlan MA, Preedy VR (1980) A rapid and convenient technique for measuring the rate of protein synthesis in tissues by injection of [3H]phenylalanine. Biochem J 192: 719–723

    PubMed  Google Scholar 

  • Garlick PJ, Burns HJG, Palmer R (1988) In: Burns HJG (ed) Baillière's clinical gastroenterology, vol. 2, no. 4. Baillière-Tindall, London, pp 915–940

    Google Scholar 

  • Garlick PJ, Wernerman J, McNurlan MA, Essen P, Lobley GE, Milne E, Calder GA, Vinnars E (1989) Measurement of the rate of protein synthesis in muscle of postabsorptive young men by injection of a “flooding dose” of [1-13C]leucine. Clin Sci 77: 329–336

    PubMed  Google Scholar 

  • Gibson JNA, Morrison JL, Rennie MJ (1985) Elevation of blood branched chain amino acid concentration has no effect on protein synthesis or breakdown in human leg tissues. Eur J Clin Invest 15: A9

  • Hammarqvist F, Wernerman J, von der Decken A, Vinnars E (1988) The effects of branched chain amino acids upon post-operative muscle protein synthesis and nitrogen balance. Clin Nutr 7: 171–175

    Google Scholar 

  • Hammarqvist F, Wernerman J, Rutsom A, von der Decken A, Vinnars E (1989) Addition of glutamine to total parenteral nutrition after elective abdominal surgery spares free glutamine in muscle, counteracts the fall in muscle protein synthesis and improves nitrogen balance. Ann Surg 209: 455–461

    PubMed  Google Scholar 

  • Harper AE, Miller RH, Block KP (1984) Branched-chain amino acid metabolism. Ann Rev Nutr 4: 409–454

    Google Scholar 

  • Hasselgren PO, LaFrance R, Pedersen P, James JH, Fischer JE (1988) Infusion of a branchedchain amino acid-enriched solution andα-ketoisocaproic acid in septic rats: effects on nitrogen balance and skeletal muscle protein turnover. J Parent Enter Nutr 12: 244–249

    Google Scholar 

  • Heys SD, Park KGM, McNurlan MA, Calder AG, Buchan V, Blessing K, Eremin O, Garlick PJ (1991) Measurement of tumour protein synthesisin vivo in human colorectal and breast cancer and its variability in separate biopsies from the same tumour. Clin Sci 80: 587–593

    PubMed  Google Scholar 

  • Holt TL, Ward LC, Thomas BJ, Davey JF, Shepherd RW (1990) The effect of an anabolic steroid, methenolone enanthate, on growth, body composition and skelelal muscle protein synthesis in the growing rat. Nutr Res 10: 535–545

    Google Scholar 

  • Irvine JW, Ottaway JH, Saunderson CL (1990) Turnover of sarcoplasmic proteins in the breast muscle of rapidly growing chicks. Int J Biochem 22: 187–192

    PubMed  Google Scholar 

  • Khairallah EA, Mortimore GE (1976) Assessment of protein turnover in perfused rat liver: Evidence for amino acid compartmentation from differential labeling of free and tRNA-bound valine. J Biol Chem 251: 1375–1384

    PubMed  Google Scholar 

  • Khairallah EA, Airhart J, Bruno MK, Puchalsky D, Khairallah L (1977) Implications of amino acid compartmentation for the determination of rates of protein catabolism in livers in meal fed rats. Acta Biol Med Germ 36: 1735–1745

    PubMed  Google Scholar 

  • Krishnamurti CR, Schaefer AL (1987) Measurement of plasma leucine flux and protein synthesis in pregnant ewes using gas chromatography and mass spectrometry. Nutr Rep Intl 35: 683–692

    Google Scholar 

  • Lin FD, Smith TK, Bayley HS (1988) A role for tryptophan in regulation of protein synthesis in porcine muscle. J Nutr 118: 445–449

    PubMed  Google Scholar 

  • Lobley GE, Milne V, Lovie JM, Reeds PJ, Pennie K (1980) Whole body and tissue protein synthesis in cattle. Br J Nutr 43: 491–502

    PubMed  Google Scholar 

  • Lobley GE, Connell A, Buchan V, Milne E, Calder AG, Skene PA (1990a) Use of a large dose, stable isotope procedure to measure muscle protein synthesis in sheep. Proc Nutr Soc 49: 134A

  • Lobley GE, Connell A, Milne E, Buchan V, Calder AG, Anderson SE, Vint H (1990b) Muscle protein synthesis in response to testosterone administration in wether lambs. Br J Nutr 64: 691–704

    PubMed  Google Scholar 

  • Lundholm K, Ternell M, Zachrisson H, Moldawer L, Lindstrom L (1991) Measurement of hepatic protein synthesis in unrestrained mice-evaluation of the “flooding technique”. Acta Physiol Scand 141: 207–219

    PubMed  Google Scholar 

  • MacLennan PA, Smith K, Weryk B, Watt PW, Rennie MJ (1988) Inhibition of protein breakdown by glutamine in perfused rat skeletal muscle. FEBS Lett 237: 133–136

    PubMed  Google Scholar 

  • MacLennan PA, Edwards RHT (1990) Protein turnover is elevated in muscle of mdx micein vivo. Biochem J 268: 795–797

    PubMed  Google Scholar 

  • Martin AF, Rabinowitz M, Blough R, Prior G, Zak R (1977) Measurements of half-life of rat cardiac myosin heavy chain with leucyl-tRNA used as precursor pool. J Biol Chem 252: 3422–3429

    PubMed  Google Scholar 

  • McKee EE, Cheung JY, Rannels DE, Morgan HE (1978) Measurement of the rate of protein synthesis and compartmentation of heart phenylalanine. J Biol Chem 253: 1030–1040

    PubMed  Google Scholar 

  • McNurlan MA, Tomkins AM, Garlick PJ (1979) The effect of starvation on the rate of protein synthesis in rat liver and small intestine. Biochem J 178: 373–379

    PubMed  Google Scholar 

  • McNurlan MA, Fern EB, Garlick PJ (1982) Failure of leucine to stimulate protein synthesisin vivo. Biochem J 204: 831–838

    PubMed  Google Scholar 

  • McNurlan MA, Essen P, Heys SD, Buchan V, Garlick PJ, Wernerman J (1991) Measurement of protein synthesis in human skeletal muscle: further investigation of the flooding technique. Clin Sci 81: 557–564

    PubMed  Google Scholar 

  • Millward DJ (1990) Muscle glutamine and the regulation of muscle protein balance in endotoxemia. Clin Nutr 9: 34

    Google Scholar 

  • Millward DJ, Davies CTM, Halliday D, Wolman SL, Matthews DE, Rennie MJ (1982) Effect of exercise on protein metabolism in humans as explored with stable isotopes. Fed Proc 41: 2686–2691

    PubMed  Google Scholar 

  • Mitch WE, Clark AS (1984) Specificity of the effects of leucine and its metabolites on protein degradation in skeletal muscle. Biochem J 222: 579–586

    PubMed  Google Scholar 

  • Morgan HE, Chua BHL (1984) In: Adibi SA, Fekl W, Langenbeck U, Schauder P (eds) Branched chain amino and keto acids in health and disease. Karger, Basel, pp 15–21

    Google Scholar 

  • Morgan JB, Jones SJ, Calkins CR (1989) Muscle protein turnover and tenderness in broiler chickens fed cimaterol. J Anim Sci 67: 2646–2654

    PubMed  Google Scholar 

  • Morrison WL, Gibson JNA, Rennie MJ (1988) Skeletal muscle and whole-body protein turnover in cardiac cachexia: influence of branched chain amino acid administration. Eur J Clin Invest 18: 648–654

    PubMed  Google Scholar 

  • Mortimore GE, Woodside KH, Henry JE (1972) Compartmentation of free valine and its relation to protein turnover in perfused rat liver. J Biol Chem 247: 2776–2784

    PubMed  Google Scholar 

  • Muramatsu T, Takai D, Kita K, Okumura JI (1990) Studies on the measurement of whole-body protein degradationin vivo in the chicken. Int J Biochem 22: 153–158

    PubMed  Google Scholar 

  • Obled C, Barre F, Millward DJ, Arnal M (1989) Whole body protein synthesis: studies with different amino acids in the rat. Am J Physiol 257: E639-E646

    PubMed  Google Scholar 

  • Okita M, Watanabe A, Tsuji T (1988) Effect of branched chain amino acid on15N-incorporation into liver and skeletal muscle proteins following15N-ammonium chloride administration to carbon tetrachloride-intoxicated rats. J Nutr Sci Vitaminol 34: 85–96

    PubMed  Google Scholar 

  • Papet I, Breuille D, Glomot F, Arnal M (1988a) Nutritional and metabolic effects of dietary leucine excess in preruminant lamb. J Nutr 118: 450–455

    PubMed  Google Scholar 

  • Papet I, Gardy-Godillot M, Dalle M, Grizard J, Arnal M (1988b) Absence d'effets d'un exces alimentaire en leucine sur les cinetiques postprandiales du glucagon et du cortisol plasmatiques chez l'agneau préruminant. Reprod Nutr Develop 28: 161–162

    Google Scholar 

  • Papet I, Grizard J, Gardy-Godillot M, Dalle M, Arnal M (1989) Lack of an effect of a high leucine test meal on plasma insulin in preruminant lambs. Nutr Res 9: 941–945

    Google Scholar 

  • Parmacek MS, Magid NM, Lesch M, Decker RS, Samarel AM (1986) Cardiac protein synthesis and degradation during thyroxine-induced left ventricular hypertrophy. Am J Physiol 251: C727-C736

    PubMed  Google Scholar 

  • Preedy VR, Garlick PJ (1984) Reduced rates of muscle protein synthesis during tail-vein infusion of rats. Biochem Soc Trans 12: 700–701

    Google Scholar 

  • Preedy VR, Garlick PJ (1986) The response of muscle protein synthesis to nutrient intake in post-absorptive rats: The role of insulin and amino acids. Biosci Rep 6: 177–183

    PubMed  Google Scholar 

  • Preedy VR, Peters TJ (1989) Protein synthesis of muscle fractions from the small intestine in alcohol-fed rats. Gut 31: 305–310

    Google Scholar 

  • Preedy VR, Peters TJ (1990) The acute and chronic effects of ethanol on cardiac muscle protein synthesis in the ratin vivo. Alcohol 7: 97–102

    PubMed  Google Scholar 

  • Preedy VR, Hammond B, Bottiglieri TG, Marway JS, Peters TJ (1990a) Measurement of protein synthesis by the phenylalanine flooding dose technique: Effect of phenylalanine and anaesthesia on plasma electrolyte, enzyme and metabolite levels. J Pharm Pharmacol 42: 851–856

    PubMed  Google Scholar 

  • Preedy VR, Marway JS, Salisbury JR, Peters TJ (1990b) Protein synthesis in bone and skin of rat are inhibited by ethanol: implications for whole body metabolism. Alcoholism 14: 165–168

    PubMed  Google Scholar 

  • Rannels DE, Wartell SA, Watkins CA (1982) The measurement of protein synthesis in biological systems. Life Sci 30: 1679–1690

    PubMed  Google Scholar 

  • Reeds PJ, Fuller MF (1983) Nutrient intake and protein turnover. Proc Nutr Soc 42: 463–471

    PubMed  Google Scholar 

  • Rennie MJ, Edwards RHT, Halliday D, Matthews DE, Wolman SL, Millward DJ (1982) Muscle protein synthesis measured by stable isotope techniques in man: the effects of feeding and fasting. Clin Sci 63: 519–523

    PubMed  Google Scholar 

  • Robinson ME, Samarel AM (1990) Regional differences inin vivo myocardial protein synthesis in the neonatal rabbit heart. J Mol Cell Cardiol 22: 607–618

    PubMed  Google Scholar 

  • Rodwell VW (1985) In: Martin DW, Mayes PA, Rodwell VW, Granner DK (eds) Harper's review of biochemistry, 20th edn. Lange Medical Publications, Los Altos CA, pp 293–318

    Google Scholar 

  • Samarel AM (1991)In vivo measurements of protein turnover during muscle growth and atrophy. FASEB J 5: 2020–2028

    PubMed  Google Scholar 

  • Sayegh JF, Lajtha A (1989)In vivo rates of protein synthesis in brain, muscle, and liver of five vertebrate species. Neurochem Res 14: 1165–1168

    PubMed  Google Scholar 

  • Schaefer AL, Davis SR, Hughson GA (1986) Estimation of tissue protein synthesis in sheep during sustained elevation of plasma leucine concentration by intravenous infusion. Br J Nutr 56: 281–288

    PubMed  Google Scholar 

  • Schaefer AL, Scott SL, Lepage P (1991) The effect of a valine flooding dose on plasma and intracellular fluid aminograms in sheep. Can J Anim Sci 71: 1279 (Abstr.)

    Google Scholar 

  • Scott SL (1991) Effect of a cold environment on protein synthesis and endocrine kinetics in growing cattle. Ph.D. Thesis, University of Alberta, Canada

    Google Scholar 

  • Scott SL, Christopherson RJ, Thompson JR, Baracos VE (1992) The effect of a cold environment on protein and energy metabolism in calves. Br J Nutr (in press)

  • Smith K, Barua JM, Watt PW, Scrimgeour CM, Rennie MJ (1992) Flooding with L-[1-13C]leucine stimulates human muscle protein incorporation of continuously infused L-[1-13C]valine. Am J Physiol 262: E372-E376

    PubMed  Google Scholar 

  • Smith TK (1985) Effect of leucine-rich dietary protein onin vitro protein synthesis in porcine muscle. Proc Soc Exp Biol Med 180: 538–543

    PubMed  Google Scholar 

  • Southorn BG, Kelly JM, McBride BW (1991) Protein synthesis measurements in sheep using a “flooding” dose technique. Can J Anim Sci 71: 1279 (Abstr.)

    Google Scholar 

  • Thibault L, Roberge AG (1987) Dietary protein-induced changes in serum dopamineβ-hydroxylase activity, glucose, insulin, cortisol and pyridoxyl-5′-phospate levels in normal and immobilized cats. J Clin Biochem Nutr 3: 55–64

    Google Scholar 

  • Thibault L, Roberge AG (1988) Dietary protein and carbohydrate effects on blood parameters related to stress in cats. Physiol Behav 42: 1–5

    PubMed  Google Scholar 

  • Trenkle A (1986) In: Dutt RH (ed) Proceedings of the symposium on food animal research, Lexington, Kentucky 1986. Intersociety Research Committee, Lexington, pp 80–91

    Google Scholar 

  • Vinnars E, Hammarqvist F, von der Decken A, Wernerman J (1990) Role of glutamine and its analogs in posttraumatic muscle protein and amino acid metabolism. J Parent Enter Nutr 14 [Suppl]: 125S-129S

    Google Scholar 

  • Waterlow JC, Garlick PJ, Millward DJ (1978) Protein turnover in mammalian tissues and in the whole body. North-Holland Publishing Company, Amsterdam

    Google Scholar 

  • Watkins CA, Rannels DE (1980) Measurement of protein synthesis in rat lungs perfusedin situ. Biochem J 188: 269–278

    PubMed  Google Scholar 

  • Watt PW, Lindsay Y, Scrimgeour CM, Chien PAF, Gibson JNA, Taylor DJ, Rennie MJ (1991) Isolation of aminoacyl-tRNA and its labeling with stable-isotope tracers: use in studies of human tissue protein synthesis. Proc Natl Acad Sci 88: 5892–5896

    PubMed  Google Scholar 

  • Wu G, Thompson JR (1990) The effect of glutamine on protein turnover in chick skeletal musclein vitro. Biochem J 265: 593–598

    PubMed  Google Scholar 

  • Yang RD, Matthews DE, Bier DM, Lo C, Young VR (1984) Alanine kinetics in humans: influence of different isotopic tracers. Am J Physiol 247: E634-E638

    PubMed  Google Scholar 

  • Young LH, Gelfand RA, King JE, Revkin JH, Barrett EJ (1989) In: Kinney JM, Borum PR (eds) Perspectives in clinical nutrition. Urban and Schwarzenberg, Baltimore, pp 261–273

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Lacombe Research Station, Agriculture Canada, Bag Service 5000, T0C 1S0, Lacombe, Alberta, Canada

    A. L. Schaefer & S. L. Scott

Authors
  1. A. L. Schaefer
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. S. L. Scott
    View author publications

    You can also search for this author in PubMed Google Scholar

Additional information

Scientific Paper No. 710. Agriculture Canada, Lacombe Research Station, Bag Service 5000, Lacombe, Alberta T0C 1S0.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Schaefer, A.L., Scott, S.L. Amino acid flooding doses for measuring rates of protein synthesis. Amino Acids 4, 5–19 (1993). https://doi.org/10.1007/BF00805797

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00805797

Keywords

Search

Navigation