Mass isotopomer distribution analysis at eight years: theoretical, analytic, and experimental considerations

被引:273
作者
Hellerstein, MK [1 ]
Neese, RA
机构
[1] Univ Calif Berkeley, Dept Nutr Sci, Berkeley, CA 94720 USA
[2] Univ Calif San Francisco, Dept Med, Div Endocrinol & Metab, San Francisco, CA 94110 USA
来源
AMERICAN JOURNAL OF PHYSIOLOGY-ENDOCRINOLOGY AND METABOLISM | 1999年 / 276卷 / 06期
关键词
D O I
10.1152/ajpendo.1999.276.6.E1146
中图分类号
R5 [内科学];
学科分类号
1002 ; 100201 ;
摘要
Mass isotopomer distribution analysis (MIDA) is a technique for measuring the synthesis of biological polymers. First developed approximately eight years ago, MIDA has been used for measuring the synthesis of lipids, carbohydrates, and proteins. The technique involves quantifying by mass spectrometry the relative abundances of molecular species of a polymer differing only in mass (mass isotopomers), after introduction of a stable isotope-labeled precursor. The mass isotopomer pattern, or distribution, is analyzed according to a combinatorial probability model by comparing measured abundances to theoretical distributions predicted from the binomial or multinomial expansion. For combinatorial probabilities to be applicable, a labeled precursor must therefore combine with itself in the form of two or more repeating subunits. MIDA allows dilution in the monomeric (precursor) and polymeric (product) pools to be determined. Kinetic parameters can then be calculated (e.g., replacement rate of the polymer, fractional contribution from the endogenous biosynthetic pathway, absolute rate of biosynthesis). Several issues remain unresolved, however. We consider here the impact of various deviations from the simple combinatorial probability model of biosynthesis and describe the analytic requirements for successful use of MIDA. A formal mathematical algorithm is presented for generating tables and equations (APPENDIX), on the basis of which effects of various confounding factors are simulated. These include variations in natural isotope abundances, isotopic disequilibrium in the precursor pool, more than one biosynthetic precursor pool, incorrect values for number of subunits present, and concurrent measurement of turnover from exogenously labeled polymers. We describe a strategy for testing whether isotopic inhomogeneity (e.g., an isotopic gradient or separate biosynthetic sites) is present in the precursor pool by comparing higher-mass (multiply labeled) to lower-mass (single- and double-labeled) isotopomer patterns. Also, an algebraic correction is presented for calculating fractional synthesis when an incomplete ion spectrum is monitored, and an approach for assessing the sensitivity of biosynthetic parameters to measurement error is described. The different calculation algorithms published for MIDA are compared; all share a common model, use overlapping solutions to computational problems, and generate identical results. Finally, we discuss the major practical issue for using MIDA at present: quantitative inaccuracy of instruments. The nature and causes of analytic inaccuracy, strategies for evaluating instrument performance, and guidelines for optimizing accuracy and reducing impact on biosynthetic parameters are suggested. Adherence to certain analytic guidelines, particularly attention to concentration effects on mass isotopomer ratios and maximizing enrichments in the isotopomers of interest, reduces error. Improving instrument accuracy for quantification of isotopomer ratios is perhaps the highest priority for this field. In conclusion, MIDA remains the "equation for biosynthesis," but attention to potentially confounding factors and analytic performance is required for optimal application.
引用
收藏
页码:E1146 / E1170
页数:25
相关论文
共 41 条
  • [1] Contributions of de novo synthesis of fatty acids to total VLDL-triglyceride secretion during prolonged hyperglycemia hyperinsulinemia in normal man
    Aarsland, A
    Chinkes, D
    Wolfe, RR
    [J]. JOURNAL OF CLINICAL INVESTIGATION, 1996, 98 (09) : 2008 - 2017
  • [2] BENYON JH, 1960, MASS SPECTROMETRY IT
  • [3] TESTING GAS-CHROMATOGRAPHIC MASS-SPECTROMETRIC SYSTEMS FOR LINEARITY OF RESPONSE
    BERGNER, EA
    LEE, WNP
    [J]. JOURNAL OF MASS SPECTROMETRY, 1995, 30 (05): : 778 - 780
  • [4] BYERLEY LO, 1993, FASEB J, V7, pA288
  • [5] Comparison of mass isotopomer dilution methods used to compute VLDL production in vivo
    Chinkes, DL
    Aarsland, A
    Rosenblatt, J
    Wolfe, RR
    [J]. AMERICAN JOURNAL OF PHYSIOLOGY-ENDOCRINOLOGY AND METABOLISM, 1996, 271 (02): : E373 - E383
  • [6] DEINES P, 1980, HDB ENV ISOTOPE GEOC, V1, pCH9
  • [7] Glucose homeostasis in children with falciparum malaria: Precursor supply limits gluconeogenesis and glucose production
    Dekker, E
    Hellerstein, MK
    Romijn, JA
    Neese, RA
    Peshu, N
    Endert, E
    Marsh, K
    Sauerwein, HP
    [J]. JOURNAL OF CLINICAL ENDOCRINOLOGY & METABOLISM, 1997, 82 (08) : 2514 - 2521
  • [8] DIETSCHY JM, 1974, J LIPID RES, V15, P508
  • [9] Fagerquist CK, 1998, J MASS SPECTROM, V33, P144, DOI 10.1002/(SICI)1096-9888(199802)33:2<144::AID-JMS618>3.0.CO
  • [10] 2-F