Organic acid profiling

A volume equivalent to 0.25 mg of creatinine from 500 mL of urine of a healthy adult donor is analyzed to exclude unusual findings or presence of drugs and/or other interfering compounds in the organic acid profile. If the specimen is deemed free of interference and suitable to be used as a QC normal control, the collection is diluted with RO water so that 3 ml equals 0.25 mg creatinine equivalent, and aliquoted into screw-cap vials during continued mixing then stored frozen. 

Figure 3.1.3a shows a normal urine organic acid profile. Panel represents an expansion of the same profile (5% of original abundance) to emphasize the underlying complexity of the urine organic acid fraction and the need to appreciate how informative findings could be hidden within a background of comparable intensity (see section 2.1.8.3). For this reason, exclusive reliance on computer-driven identification is not a safe option because visual evaluation is required, for example in the case of the critical overlaps discussed later in this chapter. 

On the other hand, the SIM analysis of a predefined set of analytes (Fig. 3.1.4a) require visual inspection, but only to exclude unexpected interferences in the signal of a target compound. For the purpose of detecting unexpected interferences, it is necessary, and is also a CAP accreditation requirement, to monitor at least two ion species per compound (Fig. 3.1.4b and c), so as to recognize artificial variations in the intensity ratios between two ion chromatograms. Figure 3.1.5 shows examples of characteristic organic acid profiles in patients with selected conditions [32]. 

Fig. 3.1.6 GC-MS TIC organic acid profile of a patient with methylmalonic acidemia, a sample volume corresponding to 0.25 mg creatinine (standard procedure), b 10 x dilution (0.025 mg creatinine). Peak legend 1 methylmalonic acid (note the characteristic broad shape and spike, 2 urea, 3 citric acid, 4 2-methylcitric acid (two peaks). There was a fourfold difference in the measurement of methylmalonic acid excretion between the two analyses, with a substantial underestimation in the undiluted specimen...

Fig. 3.1.7 Detection of HG by the GC-MS TIC method in the urine of patients with MCAD deficiency collected at different clinical statuses. A, left panel Organic acid profile of an acutely ill patient. The arrow indicates the portion of the chromatogram shown in the middle panel. Peak labeling 1 HG, 2 4-hydroxyphenylacetic acid. Right panel extracted ion chromatograms of the [M-15]+ ion of HG (m/z 230 red) and 4-hydroxyphenylacetic acid (m/z 281). Patient recovering from an acute episode. C Asymptomatic patient. The latter profile represents a situation where there is a high probability that HG may not be detected by a GC-MS TIC method...

Fig. 3.1.11a GC-MS TIC organic acid profile of a patient with 4hydroxyphenylpyruvate dioxygenase deficiency (hawkinsinuria) b,c Electron impact mass spectra of two peaks of 2-cystein-S-yl-l,4-dihydroxycyclohex-5-en-l-yl) acetic acid...

Acetazolamide can produce severe lactic acidosis, with an increased lactaterpyruvate ratio, ketosis with a low beta-hydroxybutyrateracetoacetate ratio, and a urinary organic acid profile consistent with pyruvate carboxylase deficiency. The acquired enzymatic injury that results from inhibition of mitochondrial carbonic anhydrase V, which provides bicarbonate to pyruvate carboxylase, can damage the tricarboxylic acid cycle. 

All enteral feeds were stopped. The baby was given intravenous glucose, L-arginine, sodium benzoate, and sodium phenylacetate. Hemodialysis was initiated. At this time, there were no spontaneous respirations, there was no response to painful stimuli, and brainstem reflexes were absent. The plasma amino acid results revealed a glutamine level of 1500 pmol/L (normal 254-823), and citrulline was undetectable (normal 10-34 pmol/L). Quantitative carnitine, plasma acylcarnitine, and urine organic acid profiles were normal. The urine orotic acid concen-... 

Figure 55-8 Partial urine organic acid profiles 15-23 minute portion of a 33 minute run) of two patients with tyrosinemia type i. A, Acutely III patient with markedly elevated excretion of succiny[acetone, pre-NTBC treatment.The insert shows the selected ion chromatogram of the [M-15] ion of succinylacetone O-TMS-oxime TMS ester, m/z 212 B, Fifteen month old patient, succinylacetone was not detected by either total ion current (orrow) or selected ion chromatogram in three different urine specimens.This patient was later shown to be compound heterozygote for the French Canadian common splice mutation (IVS12+5G>A) and another previously unreported mutation. Peak legend I, Succinylacetone (oxime, peak I) 2, succinylacetone (oxime, peak II) 3, 4-hydroxy phenyllactic acid 4, 4-hydroxy phenylpyruvic add (oxime).The symbol marks the internal standard (pentadecanoic acid), signal abundance is normalized to the intensity of the internal standard peak.

Table 55-3 shows a summary of known disorders of organic acid metabolism, including information about their incidence, major clinical features and biochemical patterns, availability of prenatal diagnosis and newborn screening, and association with sudden unexpected death. The incidence of individual inborn errors of organic add metabolism varies from 1 10,000 to >1 1,000,000 live births. All possible disease entities included, the incidence of conditions in which informative organic acid profiles could be detected in urine is likely to approach 1 1000 live births. 

Figure 55-12 Partial urine organic acid profiles (7-28 minute portion of a 33 minute run). A, Ethylmalonic encephalopathy.

Soyer, Y, Koca, N., and Karadeniz, F., Organic acid profile of Turkish white grapes and grape juices. J. Food Comp. Anal., 16, 629-636, 2003. 

Fig. 8.4 An abnormal urine organic acid profile indicat- acidemia (and organic acid disorder of leucine metabo-...

Liquid chromatography Organic acids profile, minor acids, citric/ isocitric acid ratio Addition of cheaper juice, e.g., grapefruit added to orange juice can be detected by tartaric acid... 

Fruit juices each have very distinct organic acid profiles that can be used as fingerprint for establishing authenticity. HPLC methods that compare ratios of the major acid components are limited in most cases because the adulterators can add synthetic acids so that the ratios are consistent with those found in pure juice. Some minor acids analysis is a more powerful technique, since it is not economically feasible to adjust the levels of all acids, many of which are expensive. D-isocitric acid in lower... 

In the food industry LC is frequently used to determine the quality of the finished product. For example, it has been shown that the flavor quality of grape juice is directly linked to the amount and spectrum of organic acids present. Likewise, in the production of sour cream by fermentation, the quality of the finished product quality is related directly to the organic acid profile. Last, but not least and as pointed out above, LC techniques are also crucial in determining the quality of recombinant protein products. 

Deficiency of fumarase causes marked elevations of fumaric acid and often other Krebs cycle intermediates in the urinary organic acids profile. The diagnosis is confirmed by measurement of fumarase activity in cultured skin fibroblasts, leukocytes or affected organs. For defining the carrier status of family members, fumarase activity measurement in blood mononuclear cells appears to be a good, easy screening tool [7-9]. Mutation analysis can confirm the results of enzyme studies [11]. 

The organic acid profile might also show elevations of other Krebs cycle intermediates succinate, fumarate citrate and malate. 

Sometimes, the organic acid profile also shows elevations of 2-ketoglutarate. ... 

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