scholarly journals Intracellular localization of aldehyde dehydrogenase in rat liver

1972 ◽  
Vol 127 (4) ◽  
pp. 633-639 ◽  
Author(s):  
Leo Marjanen
Keyword(s):  
Rat Liver ◽  
Aldehyde Dehydrogenase ◽  
Gradient Centrifugation ◽  
Intracellular Localization ◽  
Total Activity ◽  
Mitochondrial Fraction ◽  
Point Of View ◽  
Ethanol Metabolism ◽  
Acetaldehyde Oxidation

1. Distribution of aldehyde dehydrogenase activity in rat liver was studied by measuring the rate of disappearance of acetaldehyde in the presence of each of the subcellular fractions. These were obtained by rough separation of particulate fractions from the soluble portion of the cell, by differential centrifugation, and by isopycnic gradient centrifugation. 2. The maximal rate of acetaldehyde oxidation was 3.7 μmol/min per g, with an apparent Km value below 10-5m. The highest rate of activity was observed in phosphate buffers of high Pi concentration (above 60mm). 3. The activity measured was completely dependent on NAD+. 4. The microsomal fraction and the nuclei were inactive in the assay. Of the total activity 80% was found in the mitochondrial fraction and the remaining 20% in the cytoplasm. 5. The distribution pattern is important from the point of view of acetaldehyde oxidation during ethanol metabolism. The apparent discrepancy of the results obtained by different workers and the localization of acetaldehyde oxidation in vivo is discussed.

scholarly journals Is the mechanism of nitroglycerin tolerance associated with aldehyde dehydrogenase activity? A contribution to the ongoing discussion

2019 ◽  
Author(s):  
Anna Bilska-Wilkosz ◽  
Magdalena Kotańska ◽  
Magdalena Górny ◽  
Barbara Filipek ◽  
Małgorzata Iciek
Keyword(s):  
Rat Liver ◽  
Aldehyde Dehydrogenase ◽  
Lipoic Acid ◽  
Total Activity ◽  
Nitrate Tolerance ◽  
Aldh Activity ◽  
Liver Homogenates ◽  
Development Of Tolerance ◽  
Aldehyde Dehydrogenase Activity

The aim of the study presented here was an attempt to answer the question posed in the title: Is the mechanism of nitroglycerin tolerance associated with aldehyde dehydrogenase (ALDH) activity? Here, we investigated the effect of administration (separately or jointly) of lipoic acid (LA), nitroglycerin (GTN), and disulfiram (DSF; an irreversible in vivo inhibitor of all ALDH isozymes (including ALDH2)), on the development of tolerance to GTN. We also assessed the total activity of ALDH in the rat liver homogenates. Our data revealed that not only DSF and GTN inhibited the total ALDH activity in the rat liver, but LA also proved to be an inhibitor of this enzyme. At the same time, the obtained results demonstrated that the GTN tolerance did not develop in GTN, DSF and LA jointly treated rats, but did develop in GTN and DSF jointly treated rats. This means that the ability of LA to prevent GTN tolerance is not associated with the total ALDH activity in the rat liver. In this context, the fact that animals jointly receiving GTN and DSF developed tolerance to GTN, and in animals that in addition to GTN and DSF also received LA such tolerance did not develop, is – in our opinion – a sufficient premise to conclude that the nitrate tolerance certainly is not caused by a decrease in the activity of any of the ALDH isoenzymes present in the rat liver, including ALDH2. However, many questions still await an answer, including the basic one: What is the mechanism of tolerance to nitroglycerin?

scholarly journals Role of cytosolic rat liver aldehyde dehydrogenase in the oxidation of acetaldehyde during ethanol metabolism in vivo

1975 ◽  
Vol 152 (3) ◽  
pp. 709-712 ◽  
Author(s):  
C J Eriksson ◽  
M Marselos ◽  
T Koivula
Keyword(s):  
Aldehyde Dehydrogenase ◽  
Oxidation Rate ◽  
Ethanol Metabolism ◽  
Minor Role ◽  
Liver Cytosol ◽  
A Minor ◽  
Liver Aldehyde Dehydrogenase ◽  
Phenobarbital Induction

The activity of a high-Km aldehyde dehydrogenase in the liver cytosol was increased by phenobarbital induction. No corresponding increase in the oxidation rate of acetaldehyde in vivo was found, and it is concluded that cytosolic aldehyde dehydrogenase plays only a minor role in the oxidation of acetaldehyde during ethanol metabolism.

scholarly journals Hydrogen transfer between ethanol molecules during oxidoreduction in vivo

1985 ◽  
Vol 229 (2) ◽  
pp. 315-322 ◽  
Author(s):  
T Cronholm
Keyword(s):  
Alcohol Dehydrogenase ◽  
Isotope Effect ◽  
Hydrogen Transfer ◽  
Ethanol Oxidation ◽  
Internal Standard ◽  
Ethanol Metabolism ◽  
Acetaldehyde Oxidation ◽  
Extensive Formation ◽  
Rate Limiting

Rates of exchange catalysed by alcohol dehydrogenase were determined in vivo in order to find rate-limiting steps in ethanol metabolism. Mixtures of [1,1-2H2]- and [2,2,2-2H3]ethanol were injected in rats with bile fistulas. The concentrations in bile of ethanols having different numbers of 2H atoms were determined by g.l.c.-m.s. after the addition of [2H6]ethanol as internal standard and formation of the 3,5-dinitrobenzoates. Extensive formation of [2H4]ethanol indicated that acetaldehyde formed from [2,2,2-2H3]ethanol was reduced to ethanol and that NADH used in this reduction was partly derived from oxidation of [1,1-2H2]ethanol. The rate of acetaldehyde reduction, the degree of labelling of bound NADH and the isotope effect on ethanol oxidation were calculated by fitting models to the found concentrations of ethanols labelled with 1-42H atoms. Control experiments with only [2,2,2-2H3]ethanol showed that there was no loss of the C-2 hydrogens by exchange. The isotope effect on ethanol oxidation appeared to be about 3. Experiments with (1S)-[1-2H]- and [2,2,2-2H3]ethanol indicated that the isotope effect on acetaldehyde oxidation was much smaller. The results indicated that both the rate of reduction of acetaldehyde and the rate of association of NADH with alcohol dehydrogenase were nearly as high as or higher than the net ethanol oxidation. Thus, the rate of ethanol oxidation in vivo is determined by the rates of acetaldehyde oxidation, the rate of dissociation of NADH from alcohol dehydrogenase, and by the rate of reoxidation of cytosolic NADH. In cyanamide-treated rats, the elimination of ethanol was slow but the rates in the oxidoreduction were high, indicating more complete rate-limitation by the oxidation of acetaldehyde.

scholarly journals Heterogeneity of rat liver mitochondrial fractions and the effect of tri-iodothyronine on their protein turnover

1970 ◽  
Vol 118 (1) ◽  
pp. 111-121 ◽  
Author(s):  
S. S. Katyare ◽  
P. Fatterpaker ◽  
A. Sreenivasan
Keyword(s):  
Rat Liver ◽  
Mitochondrial Fraction ◽  
Liver Mitochondria ◽  
Heavy Fraction ◽  
Rat Liver Mitochondria ◽  
Turnover Rates ◽  
Loosely Coupled ◽  
Mitochondrial Fractions ◽  
Isoelectric Ph

1. Rat liver mitochondria were separated into heavy, light and fluffy fractions by differential centrifugation under standard conditions. 2. All mitochondrial fractions possessed soluble as well as membrane-bound enzymes typical of mitochondria. 3. The heavy fraction represented the stable mitochondrial structures and the fluffy particles appear to be loosely coupled. 4. The light mitochondrial fraction lacked the ability of coupled phosphorylation. 5. A study of mobility and isoelectric pH indicated a similarity in the basic membrane structure of all the mitochondrial fractions. 6. The turnover rates of proteins in the heavy and fluffy particles were almost identical; however, this rate was rapid for the light mitochondrial fraction. 7. On treatment with 3,3′,5-tri-iodo-l-thyronine, succinoxidase activity was maximally stimulated much earlier in the light mitochondrial fraction than in the heavy fraction. The activity of the fluffy particles, however, remained almost unaffected. 8. Malate dehydrogenase activity in all the mitochondrial fractions was stimulated only at 40h after tri-iodothyronine treatment. 9. The pattern of incorporation of dl-[1-14C]leucine in vivo in the tri-iodothyronine-treated animals indicated a rapid initial incorporation and high synthetic ability of the light mitochondrial fraction. 10. The turnover pattern of proteins of the mitochondrial fractions from animals receiving repeated doses of tri-iodothyronine was remarkably different from the normal pattern and suggested that preformed soluble protein units may be incorporated in the light mitochondrial fraction during maturation to form the stable heavy mitochondria. 11. The amount of light-mitochondrial proteins decreased by 40% on thyroidectomy and increased by 160% on treatment with tri-iodothyronine. 12. The possible significance of these results is discussed in relation to mitochondrial genesis.

scholarly journals Subcellular localization and properties of rat liver adenosine diphosphatase

1980 ◽  
Vol 192 (2) ◽  
pp. 527-535 ◽  
Author(s):  
G P Smith ◽  
G D Smith ◽  
T J Peters
Keyword(s):  
Rat Liver ◽  
Gradient Centrifugation ◽  
Intracellular Localization ◽  
Sucrose Density Gradient ◽  
Subcellular Fractionation ◽  
Single Step ◽  
Sucrose Density Gradient Centrifugation ◽  
Ph Optimum ◽  
Outer Membranes ◽  
Selective Membrane

ADPase (adenosine diphosphatase) was assayed in rat liver homogenates with [beta-32P]ADP as substrate. The activity had a pH optimum of 8.0 and was strongly activated by Mg2+. The intracellular localization was determined by analytical subcellular fractionation with single-step sucrose-density-gradient centrifugation. Selective membrane perturbants were used to enhance the resolution of the various organelles. ADPase was localized to the mitochondria. Mitochondria were isolated by differential centrifugation and subfractionated by selective disruption of the inner and outer membranes. The intramitochondrial localization of ADPase was compared with various marker enzymes and was shown to be concentrated in the outer-membrane fractions. The effects of various inhibitors on the ADPase activity were determined and the possibility that the activity could be due to known enzyme systems was considered. It is concluded that ADP degradation is due to a hitherto unrecognized mitochondrial enzyme.

scholarly journals In vivo measurement of ethanol metabolism in the rat liver using magnetic resonance spectroscopy of hyperpolarized [1-13C]pyruvate

2009 ◽  
Vol 62 (2) ◽  
pp. 307-313 ◽  
Author(s):  
Daniel M. Spielman ◽  
Dirk Mayer ◽  
Yi-Fen Yen ◽  
James Tropp ◽  
Ralph E. Hurd ◽  
...  
Keyword(s):  
Magnetic Resonance ◽  
Magnetic Resonance Spectroscopy ◽  
Rat Liver ◽  
Ethanol Metabolism ◽  
Resonance Spectroscopy ◽  
In Vivo Measurement

Effect of Pyrazole on Ethanol Metabolism in Ethanol-Tolerant Rats

1975 ◽  
Vol 53 (3) ◽  
pp. 416-422 ◽  
Author(s):  
H. Kalant ◽  
J. M. Khanna ◽  
L. Endrenyi
Keyword(s):  
Maximum Level ◽  
Total Activity ◽  
Alcohol Concentration ◽  
Male Rats ◽  
Ethanol Metabolism ◽  
Control Group ◽  
Blood Alcohol ◽  
Chronic Alcohol ◽  
Alcohol Group

Adult male rats were pair-fed liquid diets, providing 37% of calories as ethanol or sucrose, for 1 month. Alcohol dehydrogenase (ADH) activity in the cytosol fractions of liver homogenates from the two groups did not differ with respect to total activity per 100 g body weight, Km for ethanol, or Ki for pyrazole. Other rats, fed in the same way, were fasted for 18–24 h, then given an intraperitoneal injection of pyrazole followed 1 h later by an injection of ethanol, 3 g/kg. Blood alcohol curves showed an unexplained slower rise to maximum level in the chronic alcohol group. Both groups showed a period of several hours in which the blood alcohol stayed at the respective maximum concentrations, which were higher in the control group. After 7–8 h the alcohol concentration began to fall in both groups, significantly more rapidly in the chronic alcohol-fed animals. A kinetic analysis shows that the results are adequately explained by the known effects of pyrazole on the ADH–mitochondrial system. The results are interpreted as evidence against the function of any microsomal ethanol oxidizing system in vivo.

scholarly journals Subcellular distribution of cytochrome c in rat liver. Methods for its extraction and purification

1967 ◽  
Vol 105 (2) ◽  
pp. 427-442 ◽  
Author(s):  
N. F. González-Cadavid ◽  
P. N. Campbell
Keyword(s):  
Cytochrome C ◽  
Rat Liver ◽  
Subcellular Distribution ◽  
Microsomal Fraction ◽  
Gradient Centrifugation ◽  
Ammonium Phosphate ◽  
Mitochondrial Fraction ◽  
Subcellular Fractions ◽  
Nuclear Fraction ◽  
Extraction And Purification

1. A method for the extraction and purification of cytochrome c from rat liver is described. The method depends on multiple chromatography on Amberlite IRC-50 with elution with ammonium phosphate buffers of differing ionic composition and pH, interspersed with gel filtration with Sephadex G-25. Conditions leading to denaturation are avoided and the product is chromatographically pure. 2. The method may be used for the quantitative analysis of cytochrome c either in unfractionated liver or in subcellular fractions. 3. Two pools of cytochrome c were detected, one extractable at pH4·0 with distilled water and the other extracted from the residues of the first extraction with 0·15m-sodium chloride. 4. For subcellular distribution studies the liver was homogenized in 0·3m-sucrose and a nuclear fraction (washed thoroughly to remove trapped mitochondria), a mitochondrial fraction, a heavy microsomal fraction, a standard microsomal fraction and the cell sap were isolated. The mitochondrial fraction was subfractionated further by density-gradient centrifugation. Each fraction was analysed for protein, RNA, DNA, succinate–neotetrazolium oxidoreductase and glucose 6-phosphatase. 5. A total of 123μg. of cytochrome c was obtained/g. wet wt. of rat liver. 6. Values for the percentage subcellular distribution of cytochrome c are: nuclear fraction, 24·4; mitochondrial fraction, 57·2; heavy microsomal fraction, 5·2; standard microsomal fraction, 10·6; cell sap, 2·7. 7. Three out of the eight mitochondrial subfractions separated by gradient centrifugation contained 76% of the cytochrome c and 85% of the succinate–neotetrazolium oxidoreductase present in the mitochondrial fraction. 8. In unfractionated liver 94% of the cytochrome c was extracted at pH4·0 with water whereas in most of the subcellular fractions the corresponding value was approx. 75–80%.

scholarly journals A procedure for the rapid isolation from rat liver of plasma membrane vesicles exhibiting Ca2+-transport and Ca2+-ATPase activities

1984 ◽  
Vol 223 (3) ◽  
pp. 733-745 ◽  
Author(s):  
R J Epping ◽  
F L Bygrave
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Rat Liver ◽  
Gradient Centrifugation ◽  
Membrane Surface ◽  
Membrane Vesicles ◽  
Mitochondrial Fraction ◽  
Plasma Membrane Vesicles ◽  
Percoll Density Gradient Centrifugation ◽  
Percoll Density Gradient

A technique is described for the isolation of a plasma membrane-enriched preparation from a rat liver post-mitochondrial fraction by using discontinuous Percoll density-gradient centrifugation. The procedure is simple, of high reproducibility and yield and requires a total isolation time of only 90 min. The preparation consists almost exclusively of membrane vesicles and is enriched approx. 26-fold in plasma membrane-localized enzymes with minor contamination (less than 10%) with membranes derived mainly from the endoplasmic reticulum and Golgi apparatus. Approx. 20% of the fraction comprises tightly-sealed vesicles in the inverted orientation which are capable of accumulating calcium ions and exhibiting vanadate-insensitive Ca2+-ATPase activity. The properties of these activities, including insensitivity to vanadate, oxalate, and to p-chloromercuribenzoate as well as a lack of requirement for added Mg2+, contrast markedly with the reported properties of Ca2+ transport by the endoplasmic reticulum isolated from rat liver. The technique may have wide application in the study of plasma membrane-associated activities in rat liver, particularly in relation to sinusoidal membrane surface-related events.

Sign in / Sign up

Export Citation Format

Share Document