Measurement of metabolites

Measurements of metabolite concentrations in muscle fibers before and after fatiguing stimulation have shown that ATP decreases from 6 to 4.6 mM and PCr decreases from 35 to 2.4 mM with a calculated increase in Pj from 3 to 38 mM (Dawson et al., 1978 Nassar-Gentina et al., 1978). The free ADP concentration was calculated to increase from 30 to 200 pM. At the same time pH decreased from 7.0 to 6.5 (Dawson et al., 1978 Juel, 1988 Westerblad and Lannergren, 1988). The effect of these metabolic changes has been studied in skinned muscle fibers, i.e., fibers in which the cell membrane has been removed. The skinning of the fibers... 

Lin, E.L.C., S.M. Cornier, and R.N. Racine. 1994. Synchronous fluorometric measurement of metabolites of polycyclic aromatic hydrocarbons in the bile of brown bullhead. Environ. Toxicol. Chem. 13 707-715. 

In the Utell et al. (1997) study, urine samples were collected prior to exposure, at the end of exposure (0-4 h), and at 4-12 h and 12-24 h for measurement of metabolites (Tong et al. 1998). The major metabolite was 2,2-dichloro-2-fluoroethyl glucuronide. Excretion of the metabolite was dose-dependent with the highest amounts present in the 4-12 h urine samples. A small amount of dichlorofluoroacetic acid was present. HCFC-141b was not detected in blood 24 h after exposure. 

Measurements of metabolites of endrin can also be useful in monitoring exposure to endrin. The glucuronide of anti-12-hydroxyendrin and 12-ketoendrin have been detected in feces and urine (Baldwin and Hutson 1980). The anti-12-hydroxyendrin glucuronide marker is the most sensitive and specific... 

The scaled elasticities of a reversible Michaelis Menten equation with respect to its substrate and product thus consist of two additive contributions The first addend depends only on the kinetic propertiesand is confined to an absolute value smaller than unity. The second addend depends on the displacement from equilibrium only and may take an arbitrary value larger than zero. Consequently, for reactions close to thermodynamic equilibrium F Keq, the scaled elasticities become almost independent of the kinetic propertiesof the enzyme [96], In this case, predictions about network behavior can be entirely based on thermodynamic properties, which are not organism specific and often available, in conjunction with measurements of metabolite concentrations (see Section IV) to determine the displacement from equilibrium. Detailed knowledge of Michaelis Menten constants is not necessary. Along these lines, a more stringent framework to utilize constraints on the scaled elasticities (and variants thereof) as a determinant of network behavior is discussed in Section VIII.E. 

As already discussed in Section VII.B.2, reactions close to equilibrium are dominated by thermodynamics and the kinetic properties have no, or only little, influence on the elements of the Jacobian matrix. Furthermore, thermodynamic properties are, at least in principle, accessible on a large-scale level [329,330]. In some cases, thermodynamic properties, in conjunction with the measurements of metabolite concentrations described in Section IV, are thus already sufficient to specify some elements of the Jacobian in a quantitative way. 

Aging is of clinical interest in the treatment of poisoning because cholinesterase reactivators such as pralidoxime (2-PAM, Protopam) chloride are ineffective after aging has occurred. Measurement of metabolites of methyl parathion, p ra-nitrophenol, and dimethylphosphate in the urine has been used to monitor exposure to workers. ... 

Breath analysis for TCE has provided a more accurate index of exposure than the measurement of metabolites (trichloroethanol and trichloroacetic acid) in the urine. ... 

Measurement of Metabolites. The elimination of drug usually happens through metabolites that are more polar than the drug itself to enhance their excretion via urine or bile. Some special situations exist in bioavailability/bioequivalence that require the determination of metabolites ... 

Stable-isotope-labeled compounds are the most important standard reagents used in most MS-based assays that require accurate quantification. Measurement of metabolites using stable isotopes is commonly referred to as isotope... 

These studies demonstrated that DNA-binding can be a reliable probe of metabolic activation. In contrast to studies of metabolites per se, which usually involve large numbers of metabolite intermediates, DNA-binding monitors only chemically reactive metabolites. Also, if there is no selective repair of specific adducts, DNA-binding monitors the cumulative production of metabolites over time, while direct measurement of metabolites can show the metabolite spectrum only at the time observed. This can be particularly critical for studies of activation of complex chemicals such as polycyclic aromatic hydrocarbons whose primary metabolites are subject to secondary and tertiary metabolism (8). 

Measurement of metabolite formation is preferred over substrate-depletion approaches, and linearity with incubation time and protein must be ensured. 

We will show that the probative advantages of hair testing depend critically on the methods used for analysis wash procedures, kinetic analysis of the wash data, the digestion of the hair for the complete release of anal)des, the measurement of metabolites, and adoption of appropriate cutoff levels. These analytical procedures will be described in the present paper along with certain unique chemical and physical properties of hair which contribute greatly to the reliability of the analytical result. The advantages of a recently published proprietary method will also be discussed. 

For the accurate interpretation of a positive hair analysis result, it is necessary to subject hair to specially developed wash and extraction procedures. During the development of these procedures, we discovered several important properties of hair, properties which were essential for defining three unique wash kinetic criteria and their cutoff levels which, in conjunction with washing, enhance the acciuacy of the interpretation of a positive hair analysis result. Additional certainty can be achieved by the measurement of metabolites or metabolite/drug ratios, by the development of ultrasensitive gas chromatography/tandem mass spectrometry (GC/MS/MS) or ion... 

Cellular Responses In Trained Muscle. Recent evidence, obtained from appropriate measurements of metabolites within the cell during contractions, suggests that skeletal muscle of trained individuals Is better able to adjust, as compared to skeletal muscle of untrained Individuals, to the energy demands of a submaxlmal contraction effort. This Is apparent since metabolic conditions altered by contractions within trained muscle change less than in untrained muscle from that found at rest. For example, the PCr content of... 

Despite these findings, other data suggest that mitochondrial membranes may be a target for glucagon emd phenylephrine action. Measurements of metabolite gradients in situ have shown that endogenous adenine nucleotide levels are increased by... 

Assay of drug in blood, plasma or serum Measurement of drug excretion in urine Measurement of metabolite(s) in blood and/or urine... 

The advantage of such methods over measurement of metabolite levels is that the measured parameter varies in response to changes in culture viability and not cell metabolism. 

Metabolites, biologically relevant compounds with mass less than 1000 Da, are the downstream products of cellular functions and represent virtually every metabolic function in living systems. Altered metabolic levels often represent defects in the functions of specific genes, transcripts, or enzymes. Hence, measurement of metabolite concentrations using biofluids or tissue specimens offers many avenues, for example, for diagnosing and managing diseases assessing... 

Quantitative metabolomics, on the other hand, can be described as a targeted approach focused on the analysis of specific metabolite species. In this method, multivariate statistical analysis follows metabolite identification and quantitation. Because of the reliable peak identification and measurement of metabolite integrals, quantitative metabolomics promises greater insights into the dynamics and fluxes of metabolites, as well as robust statistical models for distinguishing classes with better classification accuracy. A major requirement for quantitative metabolomics is good-quality spectral analysis to provide reliable peak assignments and metabolite identification. 

---