The effect of concomitant use of systemic antibiotics in patients with Clostridium difficile infection receiving metronidazole therapy.

Management of Clostridium difficile infection (CDI) involves discontinuation of the offending antibiotic agent as soon as possible. However, the ongoing infection does not allow discontinuation of the offending antibiotic. We aimed to retrospectively investigate the predictors of treatment failure and impact of the concomitant use of systemic antibiotics in patients receiving metronidazole therapy. This study was conducted among patients hospitalised at a second care academic hospital from January 2013 to December 2014. Eligible patients were identified by reviewing stool toxin enzyme immunoassay results for C. difficile. Diarrhoea was defined as the passage of at least three loose or watery stools within 24 h. Among 314 patients with CDI receiving metronidazole therapy, 62 (19.7%) showed treatment failure and 105 (33.4%) received concomitant antibiotics. Underlying dialysis, fever >38.3 °C, low median serum albumin levels and concomitant use of antibiotics were independent predictors of treatment failure in patients with CDI receiving metronidazole therapy. The concomitant use of antibiotics increased the rates of treatment failure and 30-day mortality in patients receiving metronidazole therapy. These results suggest that metronidazole should be used in mild cases of CDI only after discontinuation of the offending antibiotics.

[Diagnosis of a case with Williams-Beuren syndrome with nephrocalcinosis using chromosome microarray analysis].

Objective: To explore the clinical phenotypes and the genetic cause for a boy with unexplained growth retardation, nephrocalcinosis, auditory anomalies and multi-organ/system developmental disorders. Method: Routine G-banding and chromosome microarray analysis were applied to a child with unexplained growth retardation, nephrocalcinosis, auditory anomalies and multi-organ/system developmental disorders treated in the Department of Pediatrics of Tongji Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology in September 2015 and his parents to conduct the chromosomal karyotype analysis and the whole genome scanning. Deleted genes were searched in the Decipher and NCBI databases, and their relationships with the clinical phenotypes were analyzed. Result: A six-month-old boy was refered to us because of unexplained growth retardation and feeding intolerance.The affected child presented with abnormal manifestation such as special face, umbilical hernia, growth retardation, hypothyroidism, congenital heart disease, right ear sensorineural deafness, hypercalcemia and nephrocalcinosis. The child's karyotype was 46, XY, 16qh+ , and his parents' karyotypes were normal. Chromosome microarray analysis revealed a 1 436 kb deletion on the 7q11.23(72701098_74136633) region of the child. This region included 23 protein-coding genes, which were reported to be corresponding to Williams-Beuren syndrome and its certain clinical phenotypes. His parents' results of chromosome microarray analysis were normal. Conclusion: A boy with characteristic manifestation of Williams-Beuren syndrome and rare nephrocalcinosis was diagnosed using chromosome microarray analysis. The deletion on the 7q11.23 might be related to the clinical phenotypes of Williams-Beuren syndrome, yet further studies are needed.

MicroRNAs function primarily in the pathogenesis of human anencephaly via the mitogen-activated protein kinase signaling pathway.

Anencephaly is one of the most serious forms of neural tube defects (NTDs), a group of congenital central nervous system (CNS) malformations. MicroRNAs (miRNAs) are involved in diverse biological processes via the post-transcriptional regulation of target mRNAs. Although miRNAs play important roles in the development of mammalian CNS, their function in human NTDs remains unknown. Using a miRNA microarray, we identified a unique expression profile in fetal anencephalic brain tissues, characterized by 70 upregulated miRNAs (ratio ≥ 2) and 7 downregulated miRNAs (ratio ≤ 0.5) compared with healthy human samples. Ten miRNAs with altered expression were selected from the microarray findings for validation with real-time quantitative reverse transcription-polymerase chain reaction. We found that in anencephalic tissues, miR-22, miR-23a, miR-34a, miR-103, miR-125a, miR-132, miR-134, miR-138, and miR-185 were significantly upregulated, while miR-149 was significantly downregulated. Furthermore, 459 potential target genes within the validated miRNAs were revealed using combined four target prediction algorithms in the human genome, and subsequently analyzed with the Molecule Annotation System 3.0. A total of 119 target genes were ultimately identified, including those involved in 22 singular annotations (i.e., transcription, signal transduction, and cell cycle) and 55 functional pathways [i.e., mitogen-activated protein kinase (MAPK) signaling pathway, and actin cytoskeleton regulation]. Six target genes (HNRPU, JAG1, FMR1, EGR3, RUNX1T1, and NDEL1) were chosen as candidate genes and associated with congenital birth abnormalities of the brain structure. Our results, therefore, suggest that miRNA maladjustment mainly contributes to the etiopathogenesis of anencephaly via the MAPK signaling pathway.

The population pharmacokinetics of fentanyl in patients undergoing living-donor liver transplantation.

Fentanyl, an opioid analgesic with a high hepatic extraction ratio, is frequently used to supplement general anesthesia during liver transplantation and is also continuously infused to provide postoperative analgesia. However, because fentanyl is metabolized mainly in the liver, the pharmacokinetics of fentanyl may vary widely during the different phases of the surgery, potentially leading to adverse events. Using nonlinear mixed-effects modeling, we characterized the pharmacokinetics of fentanyl in 15 patients (American Society of Anesthesiologists Physical Status Classification 2 or 3) undergoing living-donor liver transplantation (LDLT). Fentanyl was continuously infused at the rate of 200-400 µg/h throughout the operation. The time course of the fentanyl plasma concentration levels was best described in terms of a two-compartment model. Estimates were made of the pharmacokinetic parameters during the preanhepatic, anhepatic, and neohepatic phases: central volume of distribution (V(1)) (l): 59.0 + hourly volume infused by rapid infusion system (RIS) × 42.5, 113.0, and 189.0, respectively, × (body weight/69)(1.3); peripheral volume of distribution (V(2)) (l): 94.3, 412.0, and 427.0, respectively; intercompartmental clearance (Q) (l/h): 96.4 × (cardiac output (CO)/6.7)(2.5), 22.6, and 28.2, respectively; metabolic clearance (Cl) (l/h): 21.7 during the preanhepatic and neohepatic phases, and 0 during the anhepatic phase. The preanhepatic central volume of distribution was found to be markedly influenced by the massive infusion of fluids and blood products. The more hyperdynamic the circulation was during the preanhepatic phase, the higher the distributional clearance.

Acute myocardial infarction due to sinus of Valsalva aneurysm in a patient with Behçet's disease.

Cardiovascular manifestations have been reported in 7-38% of patients with Behçet's disease (BD), and mortality occurs in up to 20% of those with marked vascular involvement. Sporadic cases of endocarditis, myocarditis, pericarditis, acute myocardial infarction, aortic aneurysm, ventricular thrombosis, congestive cardiomyopathy, and valvular dysfunction have been reported. Here we report a case of acute myocardial infarction that resulted from the compression of coronary arteries by a sinus of Valsalva aneurysm in a patient with BD.

R-enantioselective hydrolysis of 2,2-dimethylcyclopropanecarboxamide by amidase from a newly isolated strain Brevibacterium epidermidis ZJB-07021.

AIMS: To isolate new micro-organisms with R-stereospecific amidase activity and to examine their potential as biocatalysts in enantioselective hydrolysis of 2,2-dimethylcyclopropanecarboxamide (1). METHODS AND RESULTS: A novel R-stereospecific amidase-producing strain ZJB-07021 was isolated through a sophisticated colorimetric screening method. Based on morphology, physiological tests, Biolog system (GP2) and 16S rRNA sequence, the new isolate was identified as Brevibacterium epidermidis. After 70 min of bioconversion at 35 degrees C, kinetic resolution of (R,S)-1 by the amidase afforded (S)-1 in 41.1% yield (>99% ee) and (R)-2 in 49.9% yield (69.7% ee) with an average E-value of 23. The enantioselectivity was found to be temperature dependent and enhanced from 12.6 at 45 degrees C to 65.9 at 14 degrees C. CONCLUSIONS: A novel bacterial strain of B. epidermidis ZJB-07021 producing R-stereospecific amidase was isolated and characterized. The isolate exhibited high E values for kinetic resolution of racemic-1 to (S)-1. SIGNIFICANCE AND IMPACT OF THE STUDY: To our knowledge, this was the first report on the species B. epidermidis that harboured R-stereospecific amidase. Strain ZJB-07021 could be further improved as a suitable biocatalyst for the stereoselective bioconversion of racemic-1 after optimization of culture and biotransformation process.

Molecular organization of the ribosomal RNA transcription unit and the phylogenetic study of Zymomonas mobilis ZM4.

Previously we reported that Zymomonas mobilis ZM4 contains three ribosomal transcription units (rrnA to C operons) which are clustered around the 50 min region, PacI fragments 13 and 6, on the physical map of Z. mobilis ZM4 [Kang, H. L. and Kang, H. S. (1998) Gene 206, 223-228]. The physical map reveals that the rrnA gene set is located on the 76 kb PacI fragment 13. The complete nucleotide sequence of the rrnA gene set has been determined. The total number of nucleotides of the rrnA gene set is about 6,250 bp. The structural genes of the 16S, 23S and 5S rRNA code for the 1,478 nt, 2,786 nt and 121 nt RNA chains, respectively. The length of the spacer regions between the 16S and 23S rRNA genes, and between the 23S and 5S rRNA genes, are 606 bp and 101 bp, respectively. Two tRNA genes, Ile-tRNA and Ala-tRNA, are found between the 16S and 23S rRNA genes and a fMet-tRNA gene is identified downstream of the 5S rRNA gene. Thus, the molecular organization of this rrnA gene set is the order of 5'-16S rRNA-tRNAIle-tRNAAla-23S rRNA-5S rRNA-tRNAfMet-3'. The secondary structure models of 16S, 23S and 5S rRNA are proposed. The phylogenetic tree, based on the 16S rRNA sequence, was constructed by neighbor-joining and maximum-parsimony methods. Zymomonas belongs to a group in which Aqorbacterium, Rhodobacter and Sphingomonas are included.

Transcriptional activation of Agrobacterium tumefaciens virulence gene promoters in Escherichia coli requires the A. tumefaciens RpoA gene, encoding the alpha subunit of RNA polymerase.

The two-component regulatory system, composed of virA and virG, is indispensable for transcription of virulence genes within Agrobacterium tumefaciens. However, virA and virG are insufficient to activate transcription from virulence gene promoters within Escherichia coli cells, indicating a requirement for additional A. tumefaciens genes. In a search for these additional genes, we have identified the rpoA gene, encoding the alpha subunit of RNA polymerase (RNAP), which confers significant expression of a virB promoter (virBp)::lacZ fusion in E. coli in the presence of an active transcriptional regulator virG gene. We conducted in vitro transcription assays using either reconstituted E. coli RNAP or hybrid RNAP in which the alpha subunit was derived from A. tumefaciens. The two forms of RNAP were equally efficient in transcription from a sigma(70)-dependent E. coli galP1 promoter; however, only the hybrid RNAP was able to transcribe virBp in a virG-dependent manner. In addition, we provide evidence that the alpha subunit from A. tumefaciens, but not from E. coli, is able to interact with the VirG protein. These data suggest that transcription of virulence genes requires specific interaction between VirG and the alpha subunit of A. tumefaciens and that the alpha subunit from E. coli is unable to effectively interact with the VirG protein. This work provides the basis for future studies designed to examine vir gene expression as well as the T-DNA transfer process in E. coli.

Reduction pathway of cis-5 unsaturated fatty acids in intact rat-liver and rat-heart mitochondria: assessment with stable-isotype-labelled substrates.

Besides the conventional isomerase-mediated pathway, unsaturated fatty acids with old-numbered double bonds are also metabolized by reduction pathways with NADPH as cofactor. The relative contributions of these pathways were measured in intact rat-liver and rat-heart mitochondria with a novel stable isotope tracer technique. A mixture of equal amounts of unlabelled cis-5-enoyl-CoA and 13C4-labelled acyl-CoA of equal chain lengths was incubated with mitochondria. The isotope distribution of 3-hydroxy fatty acids produced from the first cycle of beta-oxidation was analysed with selected ion monitoring by gas chromatograph-mass spectrometer. 3-Hydroxy fatty acids produced from the reduction pathway of unsaturated fatty acids were unlabelled (m + 0) whereas those produced from saturated fatty acids were labelled (m + 4). The m + 0 content serves to indicate the extent of reduction pathway. Rotenone treatment was used to switch the pathway completely to reduction. The extent of m + 0 enrichment in untreated mitochondria normalized to the m + 0 enrichment of rotenone-treated mitochondria was the percentage of reduction pathway. With this technique, cis-4-decenoate was found to be metabolized completely by the reduction pathway in both liver and heart mitochondria. cis-5-Dodecenoate was metabolized essentially by the reduction pathway in liver mitochondria, but only to 75% in heart mitochondria. When the chain length was extended to cis-5-tetradecenoate, the reduction pathway in liver mitochondria decreased to 86% and that in heart mitochondria to 65%. The effects of carnitine, clofibrate and other conditions on the reduction pathway were also studied. Enrichments of the label on saturated fatty acids and 3-hydroxy fatty acids indicated that the major pathway of reduction was not by the direct reduction of the cis-5 double bond. Instead, it is most probably by a pathway that does not involve forming a reduced saturated fatty acid first.

Oxidation of cis-5-unsaturated fatty acids in intact rat liver mitochondria: the operation of reduction pathways.

The metabolism of cis-5 unsaturated fatty acids was studied in intact rat liver mitochondria to assess the operation of a reduction pathway. By using direct quantification of metabolites with a capillary-column gas chromatography, 3-hydroxydodecanoate was identified among other metabolites when cis-5-dodecenoate was metabolized in intact rat liver mitochondria. The formation of 3-hydroxydodecanoate supports the existence of a reduction pathway in the metabolism of cis-5-unsaturated fatty acids. This metabolite cannot be produced from the conventional isomerase-mediated pathway. However, the data also indicated the possible operation of the conventional isomerase-mediated pathway in intact rat liver mitochondria. The reduction pathway appears to account for at least 61% of the pathway for cis-5-dodecenoate. This reduction pathway was likely to proceed from the dehydrogenation to trans-2,cis-5-dodecadienoyl-CoA, which was isomerized to delta 3, delta 5-dodecadienoyl-CoA, then to trans-2,trans-4-dodecadienoate. The reduction was mediated by 2,4-dienoyl-CoA reductase by the conversion of trans-2,trans-4-dodecadienoyl-CoA into trans-3-dodecenoyl-CoA. However, direct reduction of the cis-5 double bond was also shown to be operating, although to a lesser extent.

Comparison of metabolic fluxes of cis-5-enoyl-CoA and saturated acyl-CoA through the beta-oxidation pathway.

The metabolic fluxes of cis-5-enoyl-CoAs through the beta-oxidation cycle were studied in solubilized rat liver mitochondrial samples and compared with saturated acyl-CoAs of equal chain length. These studies were accomplished using either spectrophotometric assay of enzyme activities and/or the analysis of metabolites and precursors using a gas chromatographic method after conversion of CoA esters into their free acids. Cis-5-enoyl-CoAs were dehydrogenated by acyl-CoA oxidase or acyl-CoA dehydrogenases at significantly lower rates (10-44%) than saturated acyl-CoAs. However, enoyl-CoA hydratase hydrated trans-2-cis-5-enoyl-CoA at a faster rate (at least 1.5-fold) than trans-2-enoyl-CoA. The combined activities of 3-hydroxyacyl-CoA dehydrogenase and 3-ketoacyl-CoA thiolase for 3-hydroxy-cis-5-enoyl-CoAs derived from cis-5-enoyl-CoAs were less than 40% of the activity for the corresponding 3-hydroxyacyl-CoAs prepared from saturated acyl-CoAs. Rat liver mitochondrial beta-oxidation enzymes were capable of metabolizing cis-5-enoyl-CoA via one cycle of beta-oxidation to cis-3-enoyl-CoA with two less carbons. However, the overall rates of one cycle of beta-oxidation, as determined with stable-isotope-labelled tracer, was only 15-25%, for cis-5-enoyl-CoA, of that for saturated acyl-CoA. In the presence of NADPH, the metabolism of cis-5-enoyl-CoAs was switched to the reduction pathway.(ABSTRACT TRUNCATED AT 250 WORDS)

Isomerization of trans-2,delta 5-dienoyl-CoA's to delta 3,delta 5-dienoyl-CoA's in the beta-oxidation of delta 5-unsaturated fatty acids.

The NADPH-dependent reduction pathway for the metabolism of delta 5-unsaturated fatty acids involves the isomerization of trans-2,delta 5-dienoyl-CoA, initially formed from the dehydrogenation of delta 5-enoyl-CoA, to isomeric delta 3,delta 5-dienoyl-CoA. The latter intermediates were then isomerized to trans-2,trans-4-dienoyl-CoA, which then follows the NADPH-dependent pathway mediated by 2,4-dienoyl-CoA reductase. The isomerization from trans-2,delta 5-dienoyl-CoA to delta 3,delta 5-dienoyl-CoA is catalyzed by delta 3,delta 2-enoyl-CoA isomerase. In this investigation, we identified the stereoisomers of delta 3,delta 5-dienoates that were formed in the reaction. Starting from trans-2,cis-5-decadienoyl-CoA, the isomerization produced cis-3,cis-5- and trans-3,cis-5-decadienoates. On the other hand, trans-2,trans-5-decadienoyl-CoA yielded cis-3,trans-5- and trans-3,trans-5-decadienoates. In addition to purified rat liver delta 3,delta 2-enoyl-CoA isomerase, acyl-CoA oxidase from Arthrobacter also catalyzed the isomerization from trans-2,cis-5-dienoyl-CoA. However, this acyl-CoA oxidase could not catalyze the similar isomerization of trans-2,trans-5-dienoyl-CoA. delta 3,delta 5-t-2,t-4-Dienoyl-CoA isomerase used cis-3,cis-5-, trans-3,cis-5-, and cis-3,trans-5-dienoyl-CoA's as substrates and converted them to trans-2,trans-4-dienoyl-CoA. In contrast, trans-3,trans-5-dienoyl-CoA was not a substrate for this isomerization. Extensive purification of acyl-CoA oxidase through column chromatography could not remove or diminish the isomerization activity associated with acyl-CoA oxidase. Acyl-CoA oxidases derived from Candida and rat liver also possess isomerization activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Purification and mechanism of delta 3,delta 5-t-2,t-4-dienoyl-CoA isomerase from rat liver.

A new enzyme, i.e., delta 3,delta 5-t-2,t-4-dienoyl-CoA isomerase, required in the NADPH-dependent metabolic pathway of odd-numbered double bond, unsaturated fatty acids, was isolated and purified to apparent homogeneity from rat liver. In the oxidation of odd-numbered double bond, unsaturated fatty acids, stepwise beta-oxidation leads to cis-5-enoyl-CoA, which is then dehydrogenated and isomerized to delta 3,delta 5-dienoyl-CoA. delta 3,delta 5-t-2,t-4-Dienoyl-CoA isomerase converts delta 3,delta 5-dienoyl-CoA to trans-2,trans-4-dienoyl-CoA, which is a substrate for NADPH-dependent 2,4-dienoyl-CoA reductase. This enzyme was purified through Matrex gel red A, blue Sepharose, DEAE-cellulose, CM-cellulose, hydroxylapatite, and Sepharose CL6B column chromatography of an ammonium sulfate precipitated fraction (30-80%) of rat liver homogenate. A native molecular weight of 200,000 with four subunits of 55,000 each was determined. The isoelectric point was 6.5. This enzyme was located in mitochondria and was inducible by clofibrate treatment. Using delta 3,delta 5-decadienoyl-CoA, delta 3,delta 5-dodecadienoyl-CoA, and delta 3,delta 5-tetradecadienoyl-CoA as substrates, the Vmax ratio was 1:0.5:0.4 and the Km's were 10.9, 5.9, and 1.4 microM, respectively. The specific activity of purified enzyme was 7 units/mg using delta 3,delta 5-decadienoyl-CoA as substrate. The mechanism of isomerization was studied by deuterium labeling. Consistent with the deuterium labeling pattern of the products, the isomerization from trans-2,cis-5-dienoyl-CoA to trans-2,trans-4-dienoyl-CoA was a two-step process through an intermediate delta 3,delta 5-dienoyl-CoA.(ABSTRACT TRUNCATED AT 250 WORDS)

[A histogenetic study on the carcinogenesis of colonic mucous membrane glands induced by 1,2-dimethylhydrazine in rats].

The relationship between atypical proliferation and carcinogenesis of colonic mucous membrane glands induced by different doses of 1,2-dimethylhydrazine (DMH) was studied by use of histopathological methods and sequential endoscopic observation. The results showed a significant increase of atypical glands in the flat mucosa with the increase in number of DMH injections, the degree of atypical proliferation becoming more serious with the increase of glandular ducts. The following changes were observed under sequential endoscopy: small elevations were first seen on the flat mucosa, then in sequence-flat elevations, hemisphere elevations, substem elevations and finally, stemmed elevations. At the same time, changes such as rubifaction, erosion, central depression and hemorrhage on the tumor surface were frequently encountered. It is concluded that atypical glandular proliferation in the flat mucosa of the colon is a precancerous lesion of colonic carcinoma.

Incomplete fatty acid oxidation. The production and epimerization of 3-hydroxy fatty acids.

3-Hydroxydicarboxylic acids are major urinary metabolites derived from fatty acid metabolism. These compounds are produced from the omega-oxidation of 3-hydroxy fatty acids. The production of the precursor 3-hydroxy fatty acids from incomplete beta-oxidation of fatty acids in rat liver mitochondria was investigated. Independent of the chain length or the concentration of fatty acid substrates, the accumulation of 3-hydroxyacyl intermediates was relatively constant at the concentration of 3-5 nmol/mg of mitochondrial protein. The extent of the incomplete oxidation was the same in Percoll gradient-purified mitochondria. Rotenone treatment increased the production of 3-hydroxy fatty acids. 3-Hydroxy fatty acids did not exist as pure L-enantiomer as expected from beta-oxidation. Instead, these metabolites were epimerized to a near racemic mixture of D- and L-isomers with a slightly dominant D-isomer (58 +/- 3%). By using deuterium-isotope labeling, the mechanism of epimerizartion was shown to be a rapid dehydration-rehydration through trans-2-enoyl-CoA. In addition, cis-3 and trans-3 fatty acids were produced; these metabolites were derived from the isomerization of trans-2-enoyl-CoA. Epimerase and isomerase were thought to be enzymes involved in the oxidation of unsaturated fatty acids. Current data have shown that the metabolism of these acids is actually through NADPH-dependent reduction pathways. The activities of epimerase and isomerase detected in rat liver mitochondria possibly function mainly in the metabolism of saturated fatty acids in a reverse role to the conventional concept.

Biogenesis of dicarboxylic acids in rat liver homogenate studied by 13C labeling.

The aim of this investigation is to assess whether long-chain fatty acids can be a substrate for omega-oxidation and the subsequent beta-oxidation to produce medium-chain dicarboxylic acids normally found in urine. Isolated rat liver 10,000 g supernatant and pellet fractions were used as the source of enzymes. The metabolism of palmitate was studied using [1,2,3,4-13C4]hexadecanoic acid as tracer. Selected ion monitoring mass spectrometry was utilized for the determination of isotope enrichments in precursor and products. Palmitate was found to be a good substrate for omega-oxidation; the rate was only slightly slower than decanoate. The beta-oxidation of [1,2,3,4-13C4]hexadecanedioic acid yielded labeled adipic, suberic, and sebacic acids. Isotope distribution in these dicarboxylic acids consisted mostly of unlabeled molecules (M + 0) and molecules labeled with four 13C (M + 4), in agreement with a beta-oxidation initiated equally from both carboxyl ends of the precursor. Significant enrichments (1-8%) with only two 13C labels (M + 2) indicate a partial bidirectional beta-oxidation. The direct metabolic conversion of hexadecanedioate to succinate was documented by the significant enrichment (1.40-1.90%) in M + 4 of succinate. These data indicate that long-chain fatty acids can be a substrate for the production of medium-chain dicarboxylates and the eventual direct conversion to succinate.

Spiropentaneacetic acid as a specific inhibitor of medium-chain acyl-CoA dehydrogenase.

To study the structure-activity relationship between pentanoic acid analogues and the inhibition of fatty acid oxidation, a number of 4-pentenoic and methylenecyclopropaneacetic acid derivatives were prepared. All compounds inhibited palmitoylcarnitine oxidation in rat liver mitochondria, with 50% inhibition occurring at a concentration between 6 and 100 microM. However, only methylenecyclopropaneacetic acid (MCPA) and spiropentaneacetic acid (SPA) showed in vivo inhibitory activity in rats as indicated by the occurrence of dicarboxylic aciduria. Rats treated with SPA excreted metabolites derived only from fatty acid oxidation whereas MCPA-treated rats also excreted metabolites derived from branch-chained amino acid and lysine metabolism. SPA is a specific inhibitor of fatty acid oxidation without affecting amino acid metabolism. The site of inhibition is medium-chain acyl-CoA dehydrogenase (MCAD). In contrast, MCPA inhibited both MCAD and short-chain acyl-CoA dehydrogenase with a stronger inhibition toward the latter. The inhibition of fatty acid oxidation by both inhibitors was partially reversible by glycine or l-carnitine. Since SPA does not form a ring-opened nucleophile such as that proposed for MCPA in the inhibition of FAD prosthetic group in acyl-CoA dehydrogenases, we propose that the irreversible inhibition by SPA occurs by a tight complex without forming a covalent bond to the isoalloxazine ring in FAD.