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Drosophila melanogaster alcohol dehydrogenase. Biochemical properties of the NAD+-plus-acetone-induced isoenzyme conversion.

The NAD+ + acetone-induced isoenzyme conversion of the Drosophila melanogaster AdhS alleloenzyme was studied. Absorption and fluorescence spectra as well as electrophoretic and kinetic methods show that the conversion process proceeds through three steps. Initially a binary enzyme-NAD+ complex is formed, followed by a ternary enzyme-NAD+-acetone complex with a KEO,Ac of 1.7 M. The last step is a rate-limiting irreversible process in which NAD+ and acetone are covalently linked to the enzyme. A Vm of 2.4 min-1 was obtained at pH 8.6.

Acetone↗

Purification and characterization of an acetone-inducible cytochrome P-450 from hamster liver microsomes.

A form of cytochrome P-450 has been purified to electrophoretic homogeneity from the hepatic microsomes of Syrian golden hamsters treated with acetone. This P-450 form, designated ha P-450j, had an M(r) of approximately 55,000, bound dimethyl sulphoxide and exhibited a CO-reduced absorbance maximum at 451 nm. The absolute spectra of its oxidized form indicated that ha P-450j was predominantly in the low-spin state. In a reconstituted system, ha P-450j showed relatively low catalytic activities towards 7-ethoxycoumarin, 7-ethoxyresorufin, aminopyrine, ethylmorphine and benzphetamine, whereas it catalysed the oxidation of aniline, acetone and thiobenzamide with a high catalytic-centre activity. In addition, ha P-450j catalysed at a high rate the high-affinity component of dimethylnitrosamine N-demethylase; in contrast, only the low-affinity component of diethylnitrosamine N-de-ethylase was efficiently catalysed. The addition of cytochrome b5 to the reconstitution system decreased the Km value for dimethylnitrosamine N-demethylase by a factor of 5 and increased the Vmax. value, and slightly enhanced the other activities. Thiobenzamide and diethyldithiocarbamate were found to be the most effective inhibitors of the ha-P-450j-dependent aniline hydroxylation. Polyclonal antibodies against rat P-450j recognized ha P-450j in immunoblots of control and treated hamster liver microsomes. Treatment of hamsters with acetone increased the apparent abundance of ha P-450j in microsomes, whereas phenobarbital and beta-naphthoflavone did not induce it. Analysis of N-terminal amino acid sequences demonstrated that ha P-450j has a high degree of sequence identity with rat P-450j. All the evidence presented in this study indicates that ha P-450j could represent the hamster orthologue of the previously described CYP2E1(s) of other species.

Acetone↗

Human barrier recovery after acute acetone perturbation: an irritant dermatitis model.

The efficacy of a topical agent in barrier recovery was evaluated after acetone-induced acute water loss barrier disruption in vivo in humans. The upper back of several volunteers was rubbed with acetone-soaked cotton balls until elevated rates of transepidermal water loss (TEWL) occurred (> 20 g/m2h, or greater). The topical agent was then applied to the acetone-treated skin sites once daily for 5 days. Resolution evaluation used TEWL measurements and the data were expressed as the percentage recovery in water barrier function. In comparison with placebo control the topical agent significantly enhanced barrier recovery, especially within the first 72 h (P < 0.05). This model offers a simple method of examining chemicals accelerating (or inhibiting) repair of this form of acute skin damage in man.

Acetone↗

Isolation of three crystalline proteins from beef liver after partial purification by acetone fractionation.

The isolation of three proteins in crystalline form from ground beef liver is described. These proteins are FTBL protein (Arch. Biochem. Biophys. 188, 251-265 (1978), crotonase, and catalase. Crotonase is isolated by crystallization from a 32% acetone extract of the ground liver. FTBL protein and catalase can subsequently be isolated from the same extract. For optimal yield and ease of isolation, FTBL protein is isolated from a 46.5% acetone extract from which catalase can subsequently be crystallized by dialysis. The isolation of FTBL protein as well as the isolation of catalase involves a preliminary fractional precipitation and solution before crystallization can be achieved. Isopropanol can be substituted for acetone in the isolation of the above three proteins and in the case of catalase, results in an exceptionally high yield. Methods for the recrystallization of the proteins are presented and the role of organic solvents in recrystallization is discussed.

Acetone↗

Influence of gender and acetone pretreatment on benzene metabolism in mice exposed by nose-only inhalation.

Benzene (BZ) requires oxidative metabolism catalyzed by cytochrome P-450 2E1 (CYP 2E1) to exert its hematotoxic and genotoxic effects. We previously reported that male mice have a two-fold higher maximum rate of BZ oxidation compared with female mice; this correlates with the greater sensitivity of males to the genotoxic effects of BZ as measured by micronuclei induction and sister chromatid exchanges. The aim of this study was to quantitate levels of BZ metabolites in urine and tissues, and to determine whether the higher maximum rate of BZ oxidation in male mice would be reflected in higher levels of hydroxylated BZ metabolites in tissues and water-soluble metabolites in urine. Male and female B6C3F, mice were exposed to 100 or 600 ppm 14C-BZ by nose-only inhalation for 6 h. An additional group of male mice was pretreated with 1% acetone in drinking water for 8 d prior to exposure to 600 ppm BZ; this group was used to evaluate the effect of induction of CYP 2E1 on urine and tissue levels of BZ and its hydroxylated metabolites. BZ, phenol (PHE), and hydroquinone (HQ) were quantified in blood, liver, and bone marrow during exposure and postexposure, and water-soluble metabolites were analyzed in urine in the 48 h after exposure. Male mice exhibited a higher flux of BZ metabolism through the HQ pathway compared with females after exposure to either 100 ppm BZ (32.0 2.03 vs. 19.8 2.7%) or 600 ppm BZ (14.7 1.42 vs. 7.94 + 0.76%). Acetone pretreatment to induce CYP 2E1 resulted in a significant increase in both the percent and mass of urinary HQ glucuronide and muconic acid in male mice exposed to 600 ppm BZ. This increase was paralleled by three- to fourfold higher steady-state concentrations of PHE and HQ in blood and bone marrow of acetone-pretreated mice compared with untreated mice. These results indicate that the higher maximum rate of BZ metabolism in male mice is paralleled by a greater proportion of the total flux of BZ through the pathway for HQ formation, suggesting that the metabolites formed along this pathway may be responsible for the genotoxicity observed following BZ exposure.

Acetone↗

Effect of acetone on the toxicity of atrazine towards photosynthesis in Anabaena.

The effect of acetone on the toxicity of atrazine towards photosynthesis in the blue-green algae Anabaena inaequalis, A. variabilis and A. cylindrica was investigated. The order of sensitivity to atrazine was A. inaequalis greater than A. variabilis greater than A. cylindrica. Acetone and atrazine interacted additively, antagonistically, and synergistically, depending upon the concentrations of acetone and atrazine used. The EC50 of atrazine towards photosynthesis was dependent upon the type of solvent-pesticide interaction.

Acetone↗

A longitudinal study of ammonia, acetone and propanol in the exhaled breath of 30 subjects using selected ion flow tube mass spectrometry, SIFT-MS.

Selected ion flow tube mass spectrometry, SIFT-MS, has been used to monitor the volatile compounds in the exhaled breath of 30 volunteers (19 males, 11 females) over a 6 month period. Volunteers provided breath samples each week between 8:45 am and 1 pm (before lunch), and the concentrations of several trace compounds were obtained. In this paper the focus is on ammonia, acetone and propanol. It was found that the concentration distributions of these compounds in breath were close to log-normal. The median ammonia level estimated as a geometric mean for all samples was 833 parts per billion (ppb) with a multiplicative standard deviation of 1.62, the values ranging from 248 to 2935 ppb. Breath ammonia clearly increased with increasing age in this volunteer cohort. The geometric mean acetone level for all samples was 477 parts per billion (ppb) with a multiplicative standard deviation of 1.58, the values ranging from 148 to 2744 ppb. The median propanol level for all samples was 18 ppb, the values ranging from 0 to 135 ppb. A weak but significant correlation between breath propanol and acetone levels is apparent in the data. The findings indicate the potential value of SIFT-MS as a non-invasive breath analysis technique for investigating volatile compounds in human health and in the diseased state.

1-Propanol↗

Comparison of formalin- and acetone-fixation for immunohistochemical detection of carcinoembryonic antigen (CEA) and keratin.

The purpose of the present study was to compare the relative merits of cold acetone and buffered formalin as fixatives for the detection of carcinoembryonic antigen and keratin in permanently embedded tissues using a peroxidase-antiperoxidase (PAP) immunohistochemical procedure. The effect of treatment with the proteolytic enzyme pronase also was examined in the formalin-fixed tissues. Cold acetone was found to be superior to formalin for the preservation of CEA and keratin antigenic activities in a variety of benign and malignant tissues. Pronase treatment markedly increased the staining intensities of both antigens in formalin-fixed tissues. For many tissues, however, superior results were obtained using the cold acetone method, and this technic is recommended for the optimum retention of antigenic activity in permanently embedded tissues.

Acetone↗

Quantitation of formaldehyde, acetaldehyde, and acetone in sidestream cigarette smoke by high-performance liquid chromatography.

A method is described for the collection and quantitation of formaldehyde, acetaldehyde, and acetone in sidestream cigarette smoke. Using the "fishtail" chimney procedure (1), sidestream smoke is drawn by vacuum through the chimney, a Cambridge filter pad (Performance Systematics Inc.; Caledonia, MI), and into an impinger containing a solution of 2,4-dinitrophenylhydrazine (DNPH), whose solubility in acetonitrile was increased by using diglyme as a constituent of the collection derivatization solution. The chimney is rinsed with DNPH solution to remove "formaldehyde", and the rinse is added to the impinger solution. This combined wash and impinger solution is used to extract the Cambridge pad. An aliquot of the extract is analyzed by high-performance liquid chromatography using a reversed-phase column, ultraviolet detection, and a mobile phase gradient with tetrahydrofuran as a modifier. Acrolein is detected but gives poor precision and recovery because of dimer formation. For formaldehyde, acetaldehyde, and acetone, both the precision (7.30, 2.49, 2.88% relative standard deviation, respectively) and recovery (90.0, 95.2, 102.8%, respectively) are acceptable. The response for formaldehyde, acetaldehyde, and acetone is linear and a function of the number of cigarettes smoked. The minimum detectable quantity for the analytes ranges from 5 to 16 micrograms per cigarette.

Acetaldehyde↗

Vapors from collodion and acetone in an EEG laboratory.

In the many EEG laboratories, the collodion-acetone technique has lost favor because of offensive vapors. We measured vapor concentrations of diethyl ether and acetone, the two principal vapors from this technique, to determine whether they reached toxic levels. We found that diethyl ether vapors usually reached the olfactory threshold, but acetone concentration did not. Neither reached concentrations that were systemically toxic. We then developed an inexpensive, effective method of reducing concentrated vapors during electrode application and removal and documented a significant reduction in vapor concentrations. With this information and with an inexpensive, "in house" vapor extraction system, technologist and patient satisfaction with the collodion method should greatly improve.

Acetone↗

Attempt to induce cataracts in rabbits by cutaneous application of acetone.

Acetone is used as a solvent for chemicals in skin sensitization studies. Its low cost, high solvent powers, and comparative lack of toxicity make it one of the most widely used industrial solvents. Absorption of acetone through the skin is not considered hazardous; however, in an earlier study we found that acetone caused cataracts in a large group of guinea pigs when small multiple doses were administered either topically or intracutaneously on the backs of these animals. No cataracts were found when two rabbits were tested. The purpose of the present study is to repeat the earlier study using more rabbits. Here, none of the 10 experimental rabbits or the 10 control rabbits developed any lens abnormalities. Those drugs and toxic substances that have produced cataracts in rabbits are described.

Acetone↗

Prekallikrein activator and kallikrein in acetone- and kaolin-activated rat plasma.

Activation of plasminogen-free rat citrated plasma (RCPL-P) with acetone/kaolin yielded BAEe-esterase activities of 0.6--0.8 U/ml. Gel filtration demonstrated one single peak of BAEe-esterase activity (mol. wt. approximately 135000) with a kininogenase-esterase ratio (3.3) close to that known for human plasma kallikrein (2.7). Similarly activated rat citrated plasma (RCPL) revealed on gel filtration an additional esterase peak (mol. wt, approximately 47,000) with a low kininogenase-esterase ratio (0.3), and should accordingly not be used for a BAEe-esterase assay of rat plasma kallikrein. Acetone activation of RCPL-P and of RCPL yielded prekallikrein activator (PKA) activities which were about doubled by treatment with kaolin to 1.9--2.1 and 3.5--4.2 PKA-U/ml respectively. Gel filtration of acetone-activated RCPL-P or RCPL revealed two peaks of PKA activity, mol. wt. approximately 110,000 corresponding to activated factor XII (XIIa), and mol. wt. approximately 33,000 corresponding to XII fragments (XIIf). Kaolin-treatment of acetone-activated RCPL-P, but not of RCPL, caused an extensive fragmentation of XIIa to the 4--6 times more active XIIf. The lower yield of PKA-activity in acetone/kaolin-activated RCPL-P, as compared with activated RCPL, seems to be due to the absence of a factor of significance for the activation of factor XII, which is not plasmin, plasma kallikrein, or high molecular weight kininogen.

Acetone↗

Activation of factor XII in acetone-treated human plasma: significance of the functional state of plasma kallikrein for the extent of activation.

The kaolin-induced activation of factor XII (XII) to XIIa was studied in plasminogen-free human citrated plasma treated with acetone in the presence of benzamidine 7.5 mM. XIIa was assayed as prekallikrein (PK) activator. The significance of the concentrations of XII, PK and high molecular weight kininogen (HMrK) was examined using mixtures of normal plasma and plasma genetically deficient in these factors. At the high plasma dilution used (1 + 23 v/v in the kaolin incubate) a joint estimation of the factors was obtained. A reduction in amount of XII, PK or HMrK resulted in a correspondingly reduced yield of XIIa. Plasma kallikrein present was assayed as S-2302 amidase. The concentration of PK in XII-deficient plasma was normal, in HMrK-deficient plasma about 30% of normal. The activation of XII was studied in fresh plasma as well as in plasma stored for 3-6 months at -70 degrees, and the activation with acetone was carried out in the presence and in the absence of benzamidine, EDTA or purified HMrK. In previous work benzamidine was found to protect the cofactor function of purified HMrK in the assay system used, and EDTA was found to inhibit purified human plasma kallikrein assayed as plasminogen activator. The present results support the previous observations, and indicate that acetone treatment of fresh human plasma (benzamidine present) results in the activation of plasma kallikrein in a functional state that requires kinin-free, but otherwise native HMrK as a cofactor for the activation of XII.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetone↗

Effect of acetone fixation on infectivity and antigenicity of respiratory syncytial virus and adenovirus in the fluorescent antibody test.

An investigation was made on the effect of acetone fixation on infectivity and immunofluorescent antigenicity of respiratory syncytial virus and adenovirus type 5, lipid- and nonlipid-containing viruses, respectively. Viruses were allowed to replicate in HEp-2 cells, and the cells were then fixed in acetone at 5 C for periods ranging from 30 s to 7 days. Treatment for 10 min was sufficient to inactivate respiratory syncytial virus, whereas infectious adenovirus type 5 could be isolated from cells immersed in acetone for 7 days. There was a gradual reduction, to 50% of that observed at 30 s, in the intensity of fluorescent antibody staining of both viruses with increasing fixation time, but no significant decreases in fluorescent antibody end-point titers of antisera to either virus were observed.

Acetone↗

Patch test reactions to the acetone-soluble fraction of human dander in atopic dermatitis.

An acetone-soluble fraction of human dander provoked positive patch test reactions in 45 (38%) of 120 patients with atopic dermatitis. The positive patch tests were rare in normal controls and clinical controls. The positive patch test reactions histologically showed spongiosis and mononuclear cell migration in the epidermis, edema and mononuclear cell infiltrate in the upper dermis. Scratch tests with the acetone-soluble human dander fraction were positive in only 2 (2%) of the 120 patients examined. It was suggested that in a considerable number of patients with atopic dermatitis, an acetone-soluble fraction of human dander provokes a delayed-type skin reaction without inducing an immediate-type skin reaction.

Acetone↗

Evaluation of formalin-acetone sedimentation in the concentration of stool for intestinal parasites.

Formalin-acetone sedimentation was compared with the formalin-ether method for the concentration of stool for intestinal parasites. Of 80 stool specimens, 45 (56.25%) were positive for parasites by the formalin-acetone method. The figures for the two methods were formalin-ether 35 (43.75%) and for the direct lacto-phenol cotton blue wet mount method 17 (21.25%). There was no statistically significant difference in the parasite recovery rate between the two methods. Acetone is more stable, safer, and a cheaper fat solvent and promises to be a useful alternative to ether.

Acetone↗

Exhaled isoprene and acetone in newborn infants and in children with diabetes mellitus.

A new analytical method gas chromatography combined with UV spectrophotometry was used to measure isoprene and acetone in expired breath collected from four different groups of children: 1) healthy newborn babies, 2) healthy preschool children, 3) healthy school children, and 4) diabetic children in different metabolic states. Both isoprene and acetone could readily be determined in one single analysis of a 250-mL air sample. Newborn babies during the first postnatal week had undetectable or very low levels of isoprene in their expired air irrespective of catabolic or anabolic state. Breath isoprene increased with age, and healthy school children had higher levels than did healthy preschool children. No significant differences in breath isoprene were found between healthy and diabetic children. Breath acetone was found to correlate with metabolic state both in newborn babies and in diabetic children. These findings illustrate the potential use of a new technique for breath analysis in children with metabolic disturbances.

Acetone↗

Effects of cytochrome b5 on aniline hydroxylation catalyzed by a reconstituted system containing acetone or 2,2'-bipyridine.

Cytochrome b5 did not exert any effect on NADPH-dependent aniline hydroxylation in the absence of acetone or 2,2'-bipyridine, whereas cytochrome b5 exhibited a stimulatory effect on the reaction in the presence of acetone or 2,2'-bipyridine. In addition, cytochrome b5 did not have any significant effect on the cumene hydroperoxide-dependent reaction in the presence of acetone or 2,2'-bipyridine.

2,2'-Dipyridyl↗