[Study of acetone metabolism in children. 2. Effect of fatty amino acids and metabolic inhibiting substances on synthesis of liver acetone bodies].
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Amino Acids↗
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Involvement of an ATP-dependent carboxylase in a CO2-dependent pathway of acetone metabolism by Xanthobacter strain Py2.
The metabolism of acetone by the aerobic bacterium Xanthobacter strain Py2 was investigated. Cell suspensions of Xanthobacter strain Py2 grown with propylene or glucose as carbon sources were unable to metabolize acetone. The addition of acetone to cultures grown with propylene or glucose resulted in a time-dependent increase in acetone-degrading activity. The degradation of acetone by these cultures was prevented by the addition of rifampin and chloramphenicol, demonstrating that new protein synthesis was required for the induction of acetone-degrading activity. In vivo and in vitro studies of acetone-grown Xanthobacter strain Py2 revealed a CO2-dependent pathway of acetone metabolism for this bacterium. The depletion of CO2 from cultures grown with acetone, but not glucose or n-propanol, prevented bacterial growth. The degradation of acetone by whole-cell suspensions of acetone-grown cells was stimulated by the addition of CO2 and was prevented by the depletion of CO2. The degradation of acetone by acetone-grown cell suspensions supported the fixation of 14CO2 into acid-stable products, while the degradation of glucose or beta-hydroxybutyrate did not. Cultures grown with acetone in a nitrogen-deficient medium supplemented with NaH13CO3 specifically incorporated 13C-label into the C-1 (major labeled position) and C-3 (minor labeled position) carbon atoms of the endogenous storage compound poly-beta-hydroxybutyrate. Cell extracts prepared from acetone-grown cells catalyzed the CO2- and ATP-dependent carboxylation of acetone to form acetoacetate as a stoichiometric product. ADP or AMP were incapable of supporting acetone carboxylation in cell extracts. The sustained carboxylation of acetone in cell extracts required the addition of an ATP-regenerating system consisting of phosphocreatine and creatine kinase, suggesting that the carboxylation of acetone is coupled to ATP hydrolysis. Together, these studies provide the first demonstration of a CO2-dependent pathway of acetone metabolism for a strictly aerobic bacterium and provide direct evidence for the involvement of an ATP-dependent carboxylase in bacterial acetone metabolism.
Acetoacetates↗
Acetone metabolism during diabetic ketoacidosis.
The presence and the importance of acetone and its metabolism in diabetic ketoacidosis has largely been ignored. Therefore, we studied acetone metabolism in nine diabetic patients in moderate to severe ketoacidosis. The concentration of acetone in plasma, urine, and breath, and the rates of acetone production and elimination in breath and urine were determined and the rates of vivo metabolism were calculated. Plasma acetone concentrations (1.55-8.91 mM) were directly related and were generally greater than acetoacetate concentrations (1.16-6.08 mM). The rates of acetone production ranged from 68 to 581 mumol/min/1.73 m2, indicating the heterogeneous nature of the patients studied. The average acetone production rate was 265 mumol/min/1.73 m2 and accounted for about 52% of the estimated acetoacetate production rate. Urinary excretion of acetone remained constant and accounted for about 7% of the acetone production rate in all patients. There was a positive linear relationship between the percentage of the acetone production rate accounted for by excretion in breath and the plasma acetone concentration. At low plasma acetone concentrations, approximately 20%, and at high plasma acetone concentrations, approximately 80% of the production rate was accounted for by breath acetone. In contrast, there was a negative linear relationship between the percentage of acetone production rate undergoing in vivo metabolism and plasma acetone concentration. At low plasma acetone concentrations, approximately 75%, and at high concentrations, approximately 20% of acetone production rate was accounted for by in vivo metabolism. Radioactivity from 2-[14C]-acetone was variably present in plasma acetone, glucose, lipids and proteins. No radioactivity was found in plasma acetoacetate, beta-hydroxy butyrate or free fatty acids or other anionic compounds. Exchange rates of acetone into other metabolites could not be estimated because of non-steady-state precursor product relationships in these patients.
Acetone↗
The metabolism of acetone in rat.
Intraperitoneal injection of 5 mumol of acetone/g, body weight, into 3 rats previously fed 1% acetone (v/v) in their drinking water resulted in the appearance in blood serum of 16 +/- 2 nmol of 1,2-propanediol/ml and 8 +/- 1 nmol of 2,3-butanediol/ml. No detectable 1,2-propanediol or 2,3-butanediol was found in the serum of animals after acetone or saline injection without prior addition of acetone to drinking water or in the serum of animals injected with saline after having been maintained on drinking water containing 1% acetone. These data suggest that acetone both acts to induce a critical enzyme or enzymes and serves as a precursor for the production of 1,2-propanediol. It is also clear from these data that chronic acetone feeding plays a role in 2,3-butanediol production in the rat. Microsomes isolated from the liver of animals maintained on drinking water supplemented with 1% acetone contained two previously unreported enzymatic activities, acetone monooxygenase which converts acetone to acetol and acetol monooxygenase which converts acetol to methylglyoxal. Both activities require O2 and NADPH. Prior treatment with acetone increased serum D-lactate from 9 nmol/ml +/- 9 nmol/ml in control animals to 77 +/- 36 nmol/ml in acetone-fed animals after injection with 5 mumol of acetone/g, body weight. This is consistent with methylglyoxal being a by-product of acetone metabolism. Two pathways for the conversion of acetone to glucose are proposed, the methylglyoxal and the propanediol pathways. The methylglyoxal pathway is responsible for the conversion of acetone to acetol, acetol to methylglyoxal, and the subsequent conversion of methylglyoxal to glucose. The propanediol pathway involves the conversion of acetol to L-1,2-propanediol by an as yet unknown process. L-1,2-Propanediol is converted to L-lactaldehyde by alcohol dehydrogenase, and L-lactaldehyde is converted to L-lactic acid by aldehyde dehydrogenase. Expression of these metabolic pathways in rat appears to be dependent on the induction of acetone monooxygenase and acetol monooxygenase by acetone.
Acetone↗
Net glucose production from acetone in isolated murine hepatocytes. The effect of different pretreatments of mice.
1. To evaluate the condition under which net glucose production from acetone, added as sole substrate, occurs different pretreatments of mice, in combination with starvation, were used; (i) acetone pretreatment (acetone is a known inducer of cytochrome P-450 isozymes involved in this pathway), (ii) fructose pretreatment (to induce NADPH+H+ generating enzymes) or (iii) their combination. 2. There was net glucose formation from acetone only in that case, when the cells were prepared from 48 hr fasted animals pretreated with both acetone and fructose. However, using 2-14C-acetone, incorporation of 14C-carbon into glucose could be detected in all the cases and, at the same time, acetone was without any effect on protein synthesis. 3. The addition of acetone increased gluconeogenesis from alanine in almost all the cases. The only exception from this general rule was that the case, when hepatocytes were prepared from acetone pretreated 48 hr starved mice where, instead of the elevation of glucose formation, a decrease of that was caused by acetone. 4. Acetone decreased 14C-carbon incorporation into glucose from 14C-(U)-alanine added at saturating concentration in hepatocytes prepared from starved mice. 5. Similarly to acetone there was no net glucose formation from acetone either when added alone, however, it enhanced gluconeogenesis from alanine at non-saturating concentrations of the amino acid. 6. Methylglyoxal proved gluconeogenic in all the cases. 7. It is concluded that net glucose formation from acetone as sole substrate occurs only under those conditions which are far from a physiological situation, however, when gluconeogenesis from another substrate takes place, acetone can contribute to net glucose formation in hepatocytes prepared from fasted mice.
Acetone↗
Evidence for an inducible nucleotide-dependent acetone carboxylase in Rhodococcus rhodochrous B276.
The metabolism of acetone was investigated in the actinomycete Rhodococcus rhodochrous (formerly Nocardia corallina) B276. Suspensions of acetone- and isopropanol-grown R. rhodochrous readily metabolized acetone. In contrast, R. rhodochrous cells cultured with glucose as the carbon source lacked the ability to metabolize acetone at the onset of the assay but gained the ability to do so in a time-dependent fashion. Chloramphenicol and rifampin prevented the time-dependent increase in this activity. Acetone metabolism by R. rhodochrous was CO2 dependent, and 14CO2 fixation occurred concomitant with this process. A nucleotide-dependent acetone carboxylase was partially purified from cell extracts of acetone-grown R. rhodochrous by DEAE-Sepharose chromatography. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggested that the acetone carboxylase was composed of three subunits with apparent molecular masses of 85, 74, and 16 kDa. Acetone metabolism by the partially purified enzyme was dependent on the presence of a divalent metal and a nucleoside triphosphate. GTP and ITP supported the highest rates of acetone carboxylation, while CTP, UTP, and XTP supported carboxylation at 10 to 50% of these rates. ATP did not support acetone carboxylation. Acetoacetate was determined to be the stoichiometric product of acetone carboxylation. The longer-chain ketones butanone, 2-pentanone, 3-pentanone, and 2-hexanone were substrates. This work has identified an acetone carboxylase with a novel nucleotide usage and broader substrate specificity compared to other such enzymes studied to date. These results strengthen the proposal that carboxylation is a common strategy used for acetone catabolism in aerobic acetone-oxidizing bacteria.
Acetoacetates↗
Measuring airway exchange of endogenous acetone using a single-exhalation breathing maneuver.
Exhaled acetone is measured to estimate exposure or monitor diabetes and congestive heart failure. Interpreting this measurement depends critically on where acetone exchanges in the lung. Health professionals assume exhaled acetone originates from alveolar gas exchange, but experimental data and theoretical predictions suggest that acetone comes predominantly from airway gas exchange. We measured endogenous acetone in the exhaled breath to evaluate acetone exchange in the lung. The acetone concentration in the exhalate of healthy human subjects was measured dynamically with a quadrupole mass spectrometer and was plotted against exhaled volume. Each subject performed a series of breathing maneuvers in which the steady exhaled flow rate was the only variable. Acetone phase III had a positive slope (0.054+/-0.016 liter-1) that was statistically independent of flow rate. Exhaled acetone concentration was normalized by acetone concentration in the alveolar air, as estimated by isothermal rebreathing. Acetone concentration in the rebreathed breath ranged from 0.8 to 2.0 parts per million. Normalized end-exhaled acetone concentration was dependent on flow and was 0.79+/-0.04 and 0.85+/-0.04 for the slow and fast exhalation rates, respectively. A mathematical model of airway and alveolar gas exchange was used to evaluate acetone transport in the lung. By doubling the connective tissue (epithelium+mucosal tissue) thickness, this model predicted accurately (R2=0.94+/-0.05) the experimentally measured expirograms and demonstrated that most acetone exchange occurred in the airways of the lung. Therefore, assays using exhaled acetone measurements need to be reevaluated because they may underestimate blood levels.
Acetone↗
Identification of ethanol-inducible P-450 isozyme 3a as the acetone and acetol monooxygenase of rabbit microsomes.
Treatment of rabbits with 1% (v/v) acetone for 1 week resulted in the appearance in blood serum of 88 +/- 14 14 nmol/ml 1-hydroxyacetone (acetol) and 70 +/- 9 nmol/ml 1,2-propanediol. Untreated rabbits had no detectable 1,2-propanediol or acetol. Hepatic microsomes from control, ethanol-, and acetone-treated rabbits catalyzed the hydroxylation of acetone at rates of 0.32 +/- 0.01, 2.01 +/- 0.43, and 3.64 +/- 0.23 nmol/min/mg of protein, respectively. The same microsomal preparations catalyzed the hydroxylation of acetol at rates of 0.33 +/- 0.04, 0.94 +/- 0.20, and 1.08 +/- 0.12 nmol/min/ mg of microsomal protein, respectively. Isozyme 3a purified from acetone- or ethanol-treated rabbits was identical as judged by comparison of the high performance liquid chromatographic profiles of tryptic digests of the two proteins. Antibody to isozyme 3a inhibited greater than 90% of the acetone monooxygenase activity from untreated, acetone-, or ethanol-treated rabbits. In contrast, the antibody only inhibited 30% of the acetol monooxygenase activity of microsomes from untreated rabbits. The inhibition was increased to about 70% after acetone or ethanol treatment. Although the activities were inhibited to different extents, a comparison of the rates attributable to isozyme 3a from antibody inhibition experiments indicated that both activities were induced to a similar extent by ethanol. Similarly, acetone also increased both activities to the same extent but was more effective than ethanol. In a reconstituted system, isozyme 3a was the only isozyme of six forms from rabbit liver to exhibit acetone monooxygenase activity. Isozyme 3a was the most active enzyme in the hydroxylation of acetol, but isozymes 2, 3b, and 4 also were able to catalyze the reaction. Antibody to isozyme 3a also inhibited greater than 90% of the acetone hydroxylase activity and 70% of the acetol hydroxylase activity of microsomes from acetone-treated rats. Two proteins were immunochemically stained on Western blots of microsomes from untreated and acetone-treated rats, one of which was increased by acetone treatment. These results suggest that isozyme 3a in rabbit and an immunochemically homologous enzyme in rat are responsible for acetone and acetol hydroxylation, the initial steps in the proposed gluconeogenic pathways for acetone.
Acetone↗
Correlation between acetone-potentiated CCl4-induced liver injury and blood concentrations after inhalation or oral administration.
In studies of acetone-potentiated liver injury induced by haloalkanes, acetone is usually given by gavage, whereas industrial exposure to acetone normally occurs by inhalation. It was of interest to verify if the route of administration influences the potentiation. Male Sprague-Dawley rats were exposed for 4 hr to acetone vapors or treated orally with acetone; the minimal effective dose (MED) levels for potentiating CCl4-induced liver injury were estimated to be 2500 ppm and 0.25 ml/kg, respectively. Groups were treated with acetone using 0.4, 1, 2, 4, or 6 times the MED. Half of each group was killed at various time intervals after treatment for blood acetone measurements by gas chromatography; the other half was challenged with CCl4 (0.1 ml/kg, ip) 18 hr after acetone, and killed 24 hr later. Plasma alanine aminotransferase (ALT) activity and bilirubin concentrations were measured. Inhalation and oral administration of acetone both potentiated CCl4 toxicity. Rats exposed repetitively to acetone vapors (10 daily exposures) and subsequently challenged with CCl4 exhibited liver toxicity that was not significantly different from that of rats subjected to a single exposure. Correlations between ALT activities and maximal blood acetone concentrations were found to be linear (positive) and significant for both routes. For a given blood acetone concentration, however, toxicity was least severe following acetone exposure by inhalation. When the concept of threshold concentrations was applied to the data, the severity of the toxic response was dependent on the blood acetone concentration above the threshold, irrespective of the route of administration.
Acetone↗
Acetone potentiation of acute acetonitrile toxicity in rats.
The purpose of these studies was to investigate the nature and mechanism of a toxicologic interaction between acetonitrile and acetone. Results of oral dose-response studies utilizing a 1:1 (w/w) mixture of acetonitrile and acetone, or varying doses of acetonitrile administered together with a constant dose of acetone, indicated that acetone potentiated acute acetonitrile toxicity three- to fourfold in rats. The onset of severe toxicity (manifested by tremors and convulsions) was delayed in the groups dosed with both solvents compared to the groups that received acetonitrile or acetone alone. Blood cyanide (a metabolite of acetonitrile) and serum acetonitrile and acetone concentrations were measured after oral administration of 25% aqueous solutions of acetonitrile, acetone, or acetonitrile plus acetone. Concentrations of cyanide in the blood of rats given acetonitrile plus acetone remained near baseline, in contrast to the high concentrations found in rats dosed with acetonitrile alone. At 34-36 h, high blood cyanide concentrations were found in rats dosed with both of the solvents. This delayed onset of elevation of blood cyanide coincided with the occurrence of clinical signs and with the disappearance of serum acetone. In further pharmacokinetic studies, blood cyanide concentrations were measured after similar dosage regimens of acetone and acetonitrile. Peak cyanide concentrations were found to be significantly greater in rats dosed with both solvents than in rats given only acetonitrile. Administration of either sodium thiosulfate or a second dose of acetone prevented the toxicity associated with exposure to both solvents. These results suggest that the effects of acetone on acetonitrile toxicity are due to a biphasic effect on the metabolism of acetonitrile to cyanide, that is, an initial inhibition followed by a stimulation of this metabolism upon acetone elimination.
Acetone↗
Infrared spectroscopy of acetone-methanol liquid mixtures: hydrogen bond network.
Acetone and methanol mixtures covering the whole solubility range are studied by Fourier transform infrared attenuated total reflectance spectroscopy. The strong bathochromic shifts observed on methanol OH and acetone CO stretch IR bands are related to hydrogen bonds between these groups. Factor analysis separates the spectra into four acetone and four methanol principal factors. A random molecular model developed for the acetone-water system [Max and Chapados, J. Chem. Phys. 119, 5632 (2003); 120, 6625 (2004)] was modified for the acetone-methanol system. This model, which takes into account H bonds accepted by methanol and acetone, is made up of 12 methanol and 11 acetone species. The 23 species abundances are regrouped according to evolving patterns or spectral similarities to compare them to the eight experimental factors. Methanol acetone mixtures are almost but not exactly random: the methanol oxygen atoms have stronger capacities than acetone to accept H bonds from methanol in the proportion 1.5 to 1. Since oxygen atoms are in excess, all labile hydrogen atoms will form H bonds. As acetone is added to methanol, its OH stretch band blueshifts as the number of accepted H bonds decreases. When methanol gives one H bond and accepts one, an H-bonding network is formed that was coined "chained organization." However, the acetone molecules do not sequester any methanol molecules by breaking or increasing the H-bond methanol network. Similarly, the methanol molecules do not sequester any acetone molecules. Consequently no acetone-methanol complex is formed in the mixtures. Gaussian simulation of the four principal factors in the methanol OH stretch region gave three distinct absorption regimes consisting of the OH stretch bands and their satellites that are identified as MeOH(1), MeOH(2), and MeOH(3) (subscript indicates the number of H, covalent and H bond, which surround the oxygen). These regimes are related to those identified in the water-acetone system as OH(2), OH(3), and OH(4).
Journal Article↗
Purification and characterization of acetone carboxylase from Xanthobacter strain Py2.
Acetone metabolism in the aerobic bacterium Xanthobacter strain Py2 proceeds by a carboxylation reaction forming acetoacetate as the first detectable product. In this study, acetone carboxylase, the enzyme catalyzing this reaction, has been purified to homogeneity and characterized. Acetone carboxylase was comprised of three polypeptides with molecular weights of 85,300, 78,300, and 19,600 arranged in an alpha2beta2gamma2 quaternary structure. The carboxylation of acetone was coupled to the hydrolysis of ATP and formation of 1 mol AMP and 2 mol inorganic phosphate per mol acetoacetate formed. ADP was also formed during the course of acetone consumption, but only accumulated at low, substoichiometric levels ( approximately 10% yield) relative to acetoacetate. Inorganic pyrophosphate could not be detected as an intermediate or product of acetone carboxylation. In the absence of CO2, acetone carboxylase catalyzed the acetone-dependent hydrolysis of ATP to form both ADP and AMP, with ADP accumulating to higher levels than AMP during the course of the assays. Acetone carboxylase did not have inorganic pyrophosphatase activity. Acetone carboxylase exhibited a Vmax for acetone carboxylation of 0.225 micromol acetoacetate formed min-1.mg-1 at 30 degrees C and pH 7.6 and apparent Km values of 7.80 microM (acetone), 122 microM (ATP), and 4. 17 mM (CO2 plus bicarbonate). These studies reveal molecular properties of the first bacterial acetone-metabolizing enzyme to be isolated and suggest a novel mechanism of acetone carboxylation coupled to ATP hydrolysis and AMP and inorganic phosphate formation.
Adenosine Triphosphate↗
Urodynamics in acetone-induced cystitis of anesthetized rats.
We examined the bladder function of cystitis models induced by intravesical acetone instillation in urethane-anesthetized rats. Acetone (0.35 ml) at 10, 30, or 50% concentration or deionized water (sham-treatment) was instilled into the bladder via the cannula which was inserted into the lumen. Acetone was withdrawn 90 sec after instillation and the bladder lumen was washed with saline after 15 min. One hour later, the cystometrogram induced by transvesical infusion of saline (3.3 ml/hr) was measured. During cystometrography of normal (non-treated) or sham-treated group, the time required to cause micturition, reflecting bladder capacity, was 9.6 +/- 0.9 (n = 7) or 10.0 +/- 0.8 min (n = 6), respectively. In the 10% acetone-treated group, the bladder capacity was similar to that in the normal or sham group. In the 30% acetone group, the time to micturition was 4.4 +/- 0.4 min (n = 7), indicating decreased bladder capacity, although the micturition pressure and the threshold pressure were not significantly different from those in the normal or sham group. However, in the 50% acetone group, the micturition reflex disappeared. In isolated rat bladder strips, contractile responses to carbachol or electrical field stimulation in the sham and 30% acetone group were similar. While, both responses in isolated strips from the 50% acetone group were reduced. The degree of damage from degeneration and desquamation of epithelium and hemorrhage in the bladder tissue from the 30% acetone group was less prominent than in the 50% acetone group. Additionally, some tissue from the 50% acetone group showed degeneration of muscle layer. The effects of three drugs were investigated in the 30% acetone group which showed increased urinary frequency. Baclofen (100 microg/kg, i.v.) and morphine (100 microg/kg, i.v.) increased significantly the bladder capacity and the threshold pressure. Atropine (10 microg/kg, i.v.) decreased the micturition pressure. These results suggest that cystitis models induced by intravesical instillation of 30% acetone may be valuable for evaluating drugs for the treatment of urinary frequency.
Acetone↗
Relationship between cytochrome P450 2E1 and acetone catabolism in rats as studied with diallyl sulfide as an inhibitor.
Previous studies have demonstrated that cytochrome P450 2E1 (P450 2E1) catalyzes the oxidation of acetone in vitro. The present study was designed to determine the importance of P450 2E1 in the catabolism of acetone in rats using diallyl sulfide (DAS) as an inhibitor of this enzyme. After a single intragastric dose of DAS, blood samples were collected from rats at different time points, and blood acetone concentrations were measured by gas chromatography. In a low DAS dose (50 mg/kg body weight) group, the maximum acetone level of 6-fold higher than the normal level was reached at 6 hr; the acetone level returned to normal at 48 hr. In a high dose (200 mg/kg) group, the maximum acetone level of 9-fold higher than the normal level was reached at 12 hr; the acetone level returned to normal at 60 hr. The turnover time and fractional turnover rate of elevated acetone were 15.8 +/- 0.5 hr and 0.054 +/- 0.001 hr-1, respectively, for the low dose, and 19.2 +/- 0.6 hr and 0.046 +/- 0.005 hr-1, respectively, for the high dose. In a chronic experiment, DAS (50 and 200 mg/kg, i.g.) was given to rats daily for 29 days, and elevated blood acetone levels were observed during the entire experimental period: 2.0 to 2.8 micrograms/mL for the low dose and 3.4 to 3.9 micrograms/mL for the high dose at 24 hr after the 1st, 7th, 14th and 28th doses versus 0.8 to 0.9 micrograms/mL for the control. The increase of blood acetone level was closely related to the decreases of N-nitrosodimethylamine (NDMA) demethylase activity and P450 2E1 content in liver microsomes. Consistent with the lack of cumulative effect from the multiple doses of DAS on acetone level, rather stable levels of the DAS metabolites, diallyl sulfoxide (45.0 micrograms/mL, range: 33.8 to 58.6 micrograms/mL) and diallyl sulfone (11.7 micrograms/mL, range: 6.9 to 15.6 micrograms/mL), were observed at 24 hr after the 1st, 7th, 21st and 28th doses with DAS (200 mg/kg) in the chronic experiment. It is likely that the inactivation and inhibition of P450 2E1 by DAS and its metabolites block the oxidation of acetone and cause its elevation in blood. The results strongly suggest an important role of P450 2E1 in acetone catabolism under physiological conditions.
Acetone↗
Correlation between positive chronotropic effect and norepinephrine release induced by acetone in the rat right atrium.
The effects of acetone on contraction rate and norepinephrine (NE) release of rat right atrium were investigated. Acetone, in the concentration range 10-210 mM increased the atrial contraction rate (ACR) in a dose-dependent manner, but in concentrations exceeding 210 mM, caused a gradual reduction in the ACR from the peak. The positive chronotropic effect of acetone on ACR can be reduced by adding 0.002 mM of propranolol (a non-selective beta 1- and beta 2-adrenergic receptor blocker), or by pretreating the rat with reserpine (an NE depleter) (5 mg/kg body weight, i.p., 24 h prior to experiment). These findings lead to the hypothesis that the increase in ACR induced by acetone may be partly due to an increase in NE release from sympathetic nerve endings in the atrium by acetone. To test this speculative hypothesis, the effect of acetone on [14C]NE + cold NE release from the right atrium, preinoculated with 0.25 microM of [14C]NE + cold NE for 20 min at 35 degrees C, was carried out using a simple technique developed in our laboratory. The acetone (10-1000 mM)-stimulated peak of NE release (pmol/g atrium/min) and summed NE release (pmol/g atrium/5 min) in excess of the basal spontaneous NE release were analyzed. The dose-response curves of the effect of 10-210 mM of acetone on the ACR and the peak of NE release were parallel. However, acetone concentrations above 210 mM caused a gradual drop of the curve of the ACR from its maximum while still enhancing the curve of the NE release. This continued gradually up to an acetone concentration of 500 mM. The atrial NE release reached a maximum at acetone concentrations between 500 and 1000 mM. This indicates that the increase in ACR caused by acetone in the range 10-210 mM may be partly due to an increase in NE release from the sympathetic nerve terminals in the atrium. However, an acetone concentration above 210 mM may be too toxic to the muscle fibers and/or the pacemaker and/or the conducting system, and so even though the NE release is still increasing, it can not enhance the ACR any further. It is generally speculated that the release of NE from the sympathetic nerve endings in the heart induced by the great number of organic solvents and general anesthetics contributes to the cause of tachycardia, arrhythmia and fibrillation of the heart.(ABSTRACT TRUNCATED AT 400 WORDS)
Acetone↗
On the mammalian acetone metabolism: from chemistry to clinical implications.
Despite the description of the ways of acetone metabolism, its real role(s) is (are) still unknown in metabolic network. In this article, a trial is made to ascertain a comprehensive overview of acetone research extending discussion from chemistry to clinical implications. Mammals are quite similar regarding their acetone metabolism, even if species differences can also be observed. By reviewing experimental data, it seems that plasma concentration of acetone in different species is in the order of 10 microm range and the concentration-dependent acetone metabolism is common to all mammals. At low concentrations of plasma acetone, the C3 pathways are operative, while at higher concentrations, the metabolism through acetate becomes dominant. Glucose formation from acetone may also contribute to the maintenance of a constant blood glucose level, but it seems to be only a minor source for that. From energetical point of view, an interorgan cooperation is suggested because transportable C3 fragments produced in the liver can serve as alternative sources of energy for the peripheral tissues in the short of circulating glucose. The degradation of acetoacetate to acetone contributes to the maintenance of pH buffering capacity, as well. Special attention is paid to the discussion of acetone production in diseases amongst which endogenous and exogenous acetonemiae have been defined. Acetonemiae of endogenous origin are due to the increased rate of acetone production followed by an increase of degrading capacity as cytochrome p450IIE1 (CYPIIE1) isozymes become induced. Exogenous acetonemiae usually resulted from intoxications caused by either acetone itself or other exogenous compounds (ethanol, isopropyl alcohol). It is highlighted that, on the one hand, isopropanol is also a normal constituent of metabolism and, on the other hand, the flat opinion that the elevation of its plasma level is a sign of alcoholism cannot further be held. The possible future directions of research upon acetone are depicted by emphasizing the need for the clear-cut identification of mammalian acetoacetate decarboxylase, and the investigation of race differences and genetic background of acetone metabolism.
2-Propanol↗
Bacterial acetone carboxylase is a manganese-dependent metalloenzyme.
Bacterial acetone carboxylase catalyzes the ATP-dependent carboxylation of acetone to acetoacetate with the concomitant production of AMP and two inorganic phosphates. The importance of manganese in Rhodobacter capsulatus acetone carboxylase has been established through a combination of physiological, biochemical, and spectroscopic studies. Depletion of manganese from the R. capsulatus growth medium resulted in inhibition of acetone-dependent but not malate-dependent cell growth. Under normal growth conditions (0.5 microm Mn2+ in medium), growth with acetone as the carbon source resulted in a 4-fold increase in intracellular protein-bound manganese over malate-grown cells and the appearance of a Mn2+ EPR signal centered at g = 2 that was absent in malate-grown cells. Acetone carboxylase purified from cells grown with 50 microm Mn2+ had a 1.6-fold higher specific activity and 1.9-fold higher manganese content than cells grown with 0.5 microm Mn2+, consistently yielding a stoichiometry of 1.9 manganese/alpha2beta2gamma2 multimer, or 0.95 manganese/alphabetagamma protomer. Manganese in acetone carboxylase was tightly bound and not removed upon dialysis against various metal ion chelators. The addition of acetone to malate-grown cells grown in medium depleted of manganese resulted in the high level synthesis of acetone carboxylase (15-20% soluble protein), which, upon purification, exhibited 7% of the activity and 6% of the manganese content of the enzyme purified from acetone-grown cells. EPR analysis of purified acetone carboxylase indicates the presence of a mononuclear Mn2+ center, with possible spin coupling of two mononuclear sites. The addition of Mg.ATP or Mg.AMP resulted in EPR spectral changes, whereas the addition of acetone, CO2, inorganic phosphate, and acetoacetate did not perturb the EPR. These studies demonstrate that manganese is essential for acetone carboxylation and suggest a role for manganese in nucleotide binding and activation.
Acetone↗
Application of a physiologically based pharmacokinetic model for reference dose and reference concentration estimation for acetone.
Recent health risk assessments to propose a Reference Dose (RfD) for acetone (Forsyth, 2001; U.S. EPA, 2001) have been based on the results of an oral subchronic study conducted in rats and mice (Dietz et al., 1991; NTP, 1991). These assessments have utilized the traditional concept of establishing the RfD by determining the lowest experimentally determined No-Observed-Adverse-Effect Level (NOAEL) and applying various Uncertainty Factors (UFs) (U.S. EPA, 1988). This article describes a risk assessment for acetone based on the systemic toxicity observed in subchronic and developmental toxicity studies to estimate an RfD and an inhalation reference concentration (RfC) for acetone. Specifically, this approach examined the subchronic study by Dietz et al. (1991), as well as an inhalation developmental toxicity study on acetone (Mast et al., 1988) and several toxicology studies of isopropanol (IPA). This was accomplished by applying a physiologically based pharmacokinetic (PBPK) model developed previously for IPA and its metabolite acetone (Clewell et al., 2001). The incorporation of the PBPK model into the derivation of an RfD and RfC for acetone allowed for a tissue-based approach rather than an external exposure-based approach, making it possible to derive an oral RfD from an inhalation study. In addition, the use of the PBPK model to analyze data from chronic and reproductive/developmental studies conducted with IPA enabled an assessment of the potential for acetone to produce any of the effects observed in the IPA studies. This analysis provided sufficient information to reduce the need for UFs in the adjustment of the NOAEL from the oral subchronic study for the determination of an RfD. Using the PBPK model in the acetone risk assessment supports a composite UF of 60 for the subchronic study, compared to composite factors of 300 to 3000 in the other recent risk assessments. This difference resulted in an RfD of 16 mg/kg/d, compared to the values of 0.3 to 3 that have previously been estimated (Forsyth, 2001; U.S. EPA, 2001). Considering the results from the inhalation developmental study (Mast et al., 1988) resulted in an RfD of 8.7 mg/kg/d. Using this study also fills a data gap for acetone that exists if only the oral database for acetone is considered for RfD derivation. An RfC of 29 ppm was also estimated for acetone using the Mast et al. (1988) study results in combination with the PBPK model. The potential impact of endogenous acetone on a risk assessment for acetone is also discussed.
Acetone↗