ABSTRACT
Research attention is growing for biochar as amendment for anaerobic digestion (AD), as it may improve both the AD process and digestate properties. In this study, two biochars (from insect frass or the woody fraction of green waste, both pyrolyzed at 450 °C) were added (5 % w/w) during semi-continuous AD of organic kitchen waste and chicken manure. Biochar was mixed either during the AD process or to the digestate post-treatment. The impact of biochar addition on the AD process, NH3 and greenhouse gas emissions, as well as C and N mineralization following soil application, was examined. Biochar did not affect the biogas yield, but did lead to a reduction in NH3 levels in the biogas and NH4+-N sorption onto the biochars, reducing the risk of AD inhibition. N sorbed as NH4+-N onto the biochar was more available for mineralization than N sorbed as NH3. N sorption on biochar-amended digestates did not result in trade-offs when these digestates were applied to soil: the sorbed N was not released as NH3 or N2O after soil application and was not released as mineral N in either the short or medium term. The C-rich biochars increased the OC content of the digestate, which enhanced its value as soil improver. Additionally, mixing biochar with digestate decreased the soil CO2 emissions up to 33 % when it was added after AD; no priming effects were observed. These findings highlight the potential benefits of incorporating biochar into AD systems and the subsequent application of biochar-amended digestate in agricultural practices.
Subject(s)
Biofuels , Charcoal , Anaerobiosis , Soil , Fertilizers/analysis , Nitrous Oxide/analysisABSTRACT
UNLABELLED: The rostral ventrolateral medulla (RVLM) controls the vascular system and may contribute to postoperative hypertension. It comprises adrenergic cardiovascular neurons, a site for action of alpha2-adrenergic agonists. Because alpha2 agonists minimize perioperative circulatory activation, we asked the following question: do alpha2 agonists, such as clonidine and mivazerol, blunt the catecholamine activation observed in the RVLM on emergence from anesthesia? Halothane-anesthetized, paralyzed rats had their ventilatory, circulatory, and acid-base stability controlled. All pressure points and incisions were infiltrated with local anesthetics. With in vivo electrochemistry, a catechol signal was recorded in the RVLM during 150 min of stable halothane anesthesia (saline-halothane group); for 120 min after halothane discontinuation (saline-emergence group); after emergence and administration of the reference alpha2 agonist, clonidine 7 microg/kg or 21 microg/kg I.V. (50% or 90% effective dose [ED50 or ED90], respectively); and after emergence and administration of a new alpha2 agonist, mivazerol 20 microg/kg or 150 microg/kg I.V. (ED50 or ED90). Under halothane, dose-response curves for the RVLM catecholamine signal were constructed for mivazerol and an alpha2 antagonist, idazoxan (ED50 2.3 mg/kg I.V.). Stable halothane anesthesia (n = 5) led to no change in mean arterial pressure (MAP), heart rate (HR), or catechol signal (CAOC). During emergence from anesthesia, the MAP, HR, and CAOC increased (n = 5). Clonidine led to a near total suppression of the RVLM catecholamine activation noticed on emergence from anesthesia (n = 5). Hypertension was partially blunted with clonidine 7 microg/kg (n = 5). Tachycardia was partially blunted with mivazerol 20 microg/kg (n = 5). Pretreatment with idazoxan suppressed all the effects of mivazerol (n = 5). IMPLICATIONS: On emergence from anesthesia, alpha2 agonists modify the activity of adrenergic cardiovascular neurons located within the vasomotor center, as assessed by in vivo electrochemistry. We provide a rationale for the use of alpha2 agonists on emergence from anesthesia in coronary/hypertensive patients.
Subject(s)
Adrenergic alpha-Agonists/pharmacology , Anesthesia , Catecholamines/metabolism , Clonidine/pharmacology , Imidazoles/pharmacology , Medulla Oblongata/drug effects , Vasomotor System/drug effects , Adrenergic alpha-2 Receptor Agonists , Animals , Idazoxan/pharmacology , Male , Medulla Oblongata/metabolism , Rats , Rats, Sprague-DawleyABSTRACT
Ischaemic injury in a number of animal models is reduced by mivazerol (2-hydroxy-3-[(1-H-imidazol-4-yl) methyl]-benzamide, CAS 125472-02-8). This effect was accompanied by a reduction in heart rate. The effect of mivazerol on myocardial blood flow and lactate production in the ischaemic myocardium was examined at constant heart rate by right atrial pacing in an anaesthetised open-chest dog model. Three periods of ischaemic were induced by coronary occlusion for 5 min. The first (sham) and the second in the absence of the drug and the third 15 min after 10 nmol/kg i.v. Arteriovenous differences in plasma lactate using a local vein and coronary sinus draining the ischaemic and non-ischaemic myocardium, respectively, were measured before and after 4 min after coronary occlusion. Blood flow (microspheres) was determined at 3 min of ischaemia. Mivazerol reduced lactate production by the ischaemic area from 2.6 +/- 1.2 to 1.5 +/- 0.9 mmol/l (paired t-test, p < 0.01), but blood flow to the ischaemic sub-endocardium was not changed: 0.19 +/- 0.1 vs 0.21 +/- 0.12 ml.g-1.min-2. Mean ST segment elevation tended to be reduced 1.6 +/- 1.0 vs 3.8 +/- 3.0 mV (one-sided paired t-test, p = 0.05). Mivazerol exerts its anti-ischaemic effect at least in part by a reduction in ischaemic lactate production but not by increasing ischaemic blood flow.
Subject(s)
Adrenergic alpha-Agonists/pharmacology , Coronary Circulation/drug effects , Coronary Vessels/physiology , Electrocardiography/drug effects , Imidazoles/pharmacology , Lactic Acid/metabolism , Myocardial Ischemia/drug therapy , Myocardium/metabolism , Anesthesia, General , Animals , Blood Pressure/drug effects , Dogs , Heart Rate/drug effects , Myocardial Ischemia/physiopathologyABSTRACT
Differential and isopycnic centrifugation of rat liver homogenates showed that, besides its established localization in peroxisomes and endoplasmic reticulum, dihydroxyacetone-phosphate acyltransferase is also present in mitochondria. The three activities differed in a number of properties (pH optimum, palmitoyl-CoA and dihydroxyacetone-phosphate dependence, and sensitivity toward N-ethylmaleimide) and are therefore likely associated with three distinct proteins. Glycerol 3-phosphate (5 mM) did not inhibit peroxisomal dihydroxyacetone-phosphate acyltransferase but inhibited the extraperoxisomal activities virtually completely. Peroxisomal dihydroxyacetone-phosphate acyltransferase was located at the inner aspect of the peroxisomal membrane, but the enzyme was not latent. Purified microsomes, from which intact peroxisomes had been removed, were still contaminated with peroxisomal membranes as deduced from the presence of two dihydroxyacetone-phosphate acyltransferase activities: a glycerol 3-phosphate-resistant activity with properties similar to those of peroxisomal dihydroxyacetone-phosphate acyltransferase and a glycerol 3-phosphate-sensitive "true" microsomal dihydroxyacetone-phosphate acyltransferase. We propose that, assayed in the presence of 5mM glycerol 3-phosphate, dihydroxyacetone-phosphate acyltransferase can be used as a marker enzyme for peroxisomal membranes. Such a marker enzyme has not hitherto been available. The differential effect of 5 mM glycerol 3-phosphate on peroxisomal and extraperoxisomal dihydroxyacetone-phosphate acyltransferases enabled us to determine the relative contribution of these activities to overall dihydroxyacetone-phosphate acylation in whole liver homogenates. At near-physiological pH and at near-physiological concentrations of unbound palmitoyl-CoA and of dihydroxyacetone-phosphate plus glycerol 3-phosphate, peroxisomes contributed 50-75%. The remaining percentage was mostly accounted for by the microsomal enzyme. At near-physiological concentrations of glycerol 3-phosphate plus dihydroxyacetone-phosphate, glycerolphosphate acyltransferase contributed 93% and dihydroxyacetone-phosphate acyltransferase 7% to overall glycerolipid synthesis in homogenates. This suggests that the dihydroxyacetone-phosphate pathway is of minor quantitative importance in overall hepatic glycerolipid synthesis but that its main function lies in the synthesis of ether lipids, which have acyldihydroxyacetone-phosphate as obligatory precursor.(ABSTRACT TRUNCATED AT 400 WORDS)
Subject(s)
Acyltransferases/metabolism , Glycerides/biosynthesis , Liver/enzymology , Microbodies/enzymology , Acyltransferases/isolation & purification , Animals , Cell Fractionation , Endoplasmic Reticulum/enzymology , Hydrogen-Ion Concentration , Isoenzymes/isolation & purification , Isoenzymes/metabolism , Kinetics , Male , Microsomes, Liver/enzymology , Mitochondria, Liver/enzymology , Rats , Rats, Inbred StrainsABSTRACT
Our results demonstrate that benfluorex at doses that are strongly hypotriglyceridemic does not increase hepatic peroxisomal enzyme activities, whereas fenofibrate at doses that are only slightly hypolipidemic induces a dramatic increase in the activity of these enzymes. Thus, the biochemical approach used in this study reveals that the hypolipidemic drug benfluorex does not belong to the class of hypolipidemic compounds known to induce hepatomegaly, hepatic peroxisome proliferation and hepatocarcinoma in rodents. Morphological studies should confirm the absence of peroxisomal induction.
Subject(s)
Fenfluramine/analogs & derivatives , Fenofibrate/toxicity , Hypolipidemic Agents/toxicity , Liver/drug effects , Microbodies/drug effects , Mitochondria, Liver/drug effects , Propionates/toxicity , Animals , Catalase/analysis , Fenfluramine/toxicity , Fenofibrate/analogs & derivatives , Glutamate Dehydrogenase/analysis , Male , Microbodies/enzymology , Mitochondria, Liver/enzymology , Rats , Rats, Inbred StrainsABSTRACT
In this brief review a survey is presented of our present knowledge of mitochondrial and peroxisomal fatty acid oxidation. For mitochondrial fatty acid oxidation, emphasis is placed on the key regulatory role of the carnitine acyltransferase system and its modulation by malonyl-Co-A. Also, the concept of opposite regulation of fatty acid synthesis and oxidation in response to nutritional or hormonal stimuli is presented. For peroxisomal fatty acid oxidation, the key features of the system are summarized together with various approaches used to measure the contribution of the peroxisomal pathway to overall fatty acid oxidation. The conclusion is reached that this contribution is minor, at least for the most abundant fatty acids palmitate and oleate.
Subject(s)
Fatty Acids/metabolism , Liver/metabolism , Microbodies/metabolism , Mitochondria, Liver/metabolism , Acyl Coenzyme A/metabolism , Animals , Diabetic Ketoacidosis/metabolism , In Vitro Techniques , Liver/ultrastructure , Mitochondria, Liver/enzymology , Oxidation-Reduction , RatsABSTRACT
Peroxisomes were purified from liver homogenates from rats, treated with the peroxisome proliferator clofibrate, by a combination of differential centrifugation and isopycnic centrifugation in iso-osmotic self-generating Percoll gradients. Structural integrity of the peroxisomes appeared to be preserved as evidenced by a high degree of catalase latency, the absence of catalase release during purification and the exclusion of inulin (mol.wt. +/- 5000). Spaces for water and solutes were measured after incubation of the peroxisomes in iso-osmotic sucrose with radioactive water or solutes and separation of the organelles from their media by centrifugation through an organic layer. Extraperoxisomal water was corrected for by the use of radioactive dextran or inulin. The sucrose, glucose, urea, methanol and acetate-accessible spaces were identical, suggesting that these spaces represent the volume in which molecules that can cross the membrane distribute. This volume equalled 50-65% of the water space. Urate and NAD+, a cofactor of peroxisomal beta-oxidation of fatty acids, also distributed in this volume, but were also partly bound. Urate and NAD+ binding was not abolished by sonication, which released the bulk of matrix catalase activity, but NAD+ binding was seriously diminished. The peroxisomal water and sucrose spaces were estimated to be 107 microliters and 55 microliters per g of liver tissue from a clofibrate-treated rat. From quantitative morphometric data [Anthony, Schmucker, Mooney & Jones (1978) J. Lipid Res. 19, 154-165] and our marker enzyme analyses, as well as from our experimentally determined water spaces of mitochondrial and microsomal fractions, it could be calculated that the volume contamination by lysosomes, mitochondria and microsomes did not exceed 1, 8 and 6% respectively. Our data indicate that apparently intact peroxisomes are permeable to a number of small molecules, including NAD+. Whether the NAD+-binding sites in sonicated peroxisomes mirror the likely existence of a membrane carrier requires further investigation.
Subject(s)
Microbodies/metabolism , NAD/metabolism , Organoids/metabolism , Animals , Biological Transport , Body Water/metabolism , Cell Fractionation , Cell Membrane Permeability , In Vitro Techniques , Liver/metabolism , Male , Proteins/metabolism , Rats , Rats, Inbred Strains , Uric Acid/metabolismABSTRACT
1. Subfractionation by isopycnic density-gradient centrifugation in self-generating Percoll gradients of peroxisome-rich fractions prepared by differential centrifugation confirmed the presence of acyl-CoA synthetase in peroxisomes. Peroxisomes did not contain nicotinamide or adenine nucleotides other than CoA. 2. The gradient fractions most enriched in peroxisomes were pooled and the peroxisomes sedimented by centrifugation, resulting in a 50-fold-purified peroxisomal preparation as revealed by marker enzyme analysis. 3. Palmitate oxidation by intact purified peroxisomes was CoA-dependent, whereas palmitoyl-CoA oxidation was not, demonstrating that the peroxisomal CoA was available for the thiolase reaction, located in the peroxisomal matrix, but not for acyl-CoA synthetase. This suggests that the latter enzyme is located at the cytoplasmic side of the peroxisomal membrane. 4. Additional evidence for this location of peroxisomal acyl-CoA synthetase was as follows. Mechanical disruption of purified peroxisomes resulted in the release of catalase from the broken organelles, but not of acyl-CoA synthetase, indicating that the enzyme was membrane-bound. Acyl-CoA synthetase was not latent, despite the fact that at least one of its substrates appears to have a limited membrane permeability, as evidenced by the presence of CoA in purified peroxisomes. Finally, Pronase, a proteinase that does not penetrate the peroxisomal membrane, almost completely inactivated the acyl-CoA synthetase of intact peroxisomes.
Subject(s)
Coenzyme A Ligases/metabolism , Liver/enzymology , Microbodies/enzymology , Organoids/enzymology , Repressor Proteins , Saccharomyces cerevisiae Proteins , Adenine Nucleotides/metabolism , Animals , Catalase/metabolism , Cell Fractionation , Centrifugation, Density Gradient , Coenzyme A Ligases/antagonists & inhibitors , Cytoplasm/enzymology , In Vitro Techniques , Intracellular Membranes/enzymology , Male , NAD/metabolism , Pronase/metabolism , Rats , Rats, Inbred StrainsABSTRACT
1. Glycerol 3-phosphate content of isolated hepatocytes from starved rats and of glycogen-depleted hepatocytes from fed rats was low and severely limited triacylglycerol synthesis. 2. Raising the glycerol 3-phosphate content by addition of precursors to the cells resulted in a hyperbolic-like relationship between triacylglycerol synthesis and cellular glycerol 3-phosphate content. Statistical analysis of the curves showed no significant differences between the nutritional states either at saturating or at subsaturating glycerol 3-phosphate content. 3. V(max.) of glycerophosphate acyltransferase measured in homogenized hepatocytes was decreased by 30-40% in starvation. There was no change in apparent K(m) for glycerol 3-phosphate. Since at saturating glycerol 3-phosphate content esterification rates in hepatocytes of both nutritional states were identical, the enzyme is not limiting esterification under this condition. 4. At subsaturating glycerol 3-phosphate content the flux through glycerophosphate acyltransferase necessarily limits esterification. Therefore one would expect a decrease in esterification in starvation under this condition. This was the case when triacylglycerol synthesis was plotted against intracellular glycerol 3-phosphate concentration, calculated from the cellular glycerol 3-phosphate content and the intracellular water space, which was smaller in hepatocytes from starved rats. 5. The data obtained in hepatocytes were extrapolated to the intact liver by using the number of parenchymal cells per g of liver as determined from marker-enzyme analysis and the liver weight per 100g body weight. The extrapolation suggested that glycerol 3-phosphate is limiting esterification in vivo for contents below 0.3-0.4 and 0.5-0.65mumol/g for livers from fed and starved animals respectively. Also for a given fatty acid load and a glycerol 3-phosphate content below 0.3mumol/g the liver may esterify less in the starved state. However, at the glycerol 3-phosphate contents measured in freeze-clamped livers (0.30 and 0.44mumol/g for the fed and starved state respectively), livers in both nutritional states seemed capable of esterifying similar amounts of fatty acids.
Subject(s)
Acyltransferases/metabolism , Glycerol-3-Phosphate O-Acyltransferase/metabolism , Glycerophosphates/metabolism , Liver/metabolism , Triglycerides/biosynthesis , Animals , Esterification , In Vitro Techniques , Intracellular Fluid/metabolism , Kinetics , Liver/cytology , Liver/enzymology , Male , Rats , Rats, Inbred Strains , Starvation/enzymology , Starvation/metabolismSubject(s)
Lipolysis , Liver/metabolism , Triglycerides/metabolism , Animals , Cell Membrane/enzymology , Cyclic AMP/metabolism , Endothelium/metabolism , Fatty Acids/metabolism , Glucagon/pharmacology , Heparin/pharmacology , Hydrogen-Ion Concentration , Insulin/pharmacology , Lipase/metabolism , Liver/drug effects , Lysosomes/enzymology , Models, BiologicalABSTRACT
Triacylglycerol synthesis by glycogen-depleted hepatocytes from fed rats that have low glycerol 3-phosphate contents was stimulated by the addition of glycerol 3-phosphate precursors. Glucagon decreased triacylglycerol synthesis only when it also lowered glycerol 3-phosphate content. The hyperbolic-like relationship between glycerol 3-phosphate content and rates of triacylglycerol synthesis was identical in the absence or presence of glucagon, indicating that the glucagon effect on triacylglycerol synthesis was not mediated through changes in enzyme activities of the esterification pathway but through changes in cellular glycerol 3-phosphate content.
Subject(s)
Glucagon/pharmacology , Glycerophosphates/metabolism , Liver/metabolism , Triglycerides/biosynthesis , Animals , Depression, Chemical , Dihydroxyacetone Phosphate/metabolism , In Vitro Techniques , Liver/cytology , Liver/drug effects , Male , Palmitic Acid , Palmitic Acids/metabolism , Rats , Rats, Inbred StrainsSubject(s)
Fatty Acids/metabolism , Liver/metabolism , Microbodies/metabolism , Mitochondria, Liver/metabolism , Organoids/metabolism , Adenosine Diphosphate/pharmacology , Adenosine Triphosphate/pharmacology , Animals , Clofibrate/pharmacology , Coenzyme A/metabolism , Kinetics , Liver/drug effects , NAD/metabolism , Oleic Acid , Oleic Acids/metabolism , Oxidation-Reduction , Palmitic Acid , Palmitic Acids/metabolism , RatsABSTRACT
1. The first dehydrogenation step of peroxisomal beta-oxidation involves the reduction of O2 to H2O2. Production rates of H2O2 and acetyl units by purified rat liver peroxisomes oxidizing palmitoyl-CoA were equal, indicating that H2O2 production is a reliable index for the release of acetyl units during peroxisomal fatty-acid oxidation. 2. Measurements of H2O2 and acid-soluble oxidation products during [1-14C]palmitoyl-CoA oxidation by purified peroxisomes revealed that the number of acetyl units released per molecule of palmitoyl-CoA oxidized rapidly decreased with increasing unbound palmitoyl-CoA concentrations. Structural damage to the peroxisomes caused by detergents or other treatments also decreased the number of acetyl units released. Under conditions where oxidation proceeded linearly with time the theoretical maximum of 5 acetyl units released per molecule of palmitoyl-CoA oxidized [Lazarow (1978) J. Biol. Chem. 253, 1522--1528] was never reached. 3. Expressed in terms of acetyl units produced and measured at low unbound-palmitoyl-CoA concentrations, mitochondrial oxidation was 10--20-fold higher than peroxisomal oxidation. 4. ATP stimulated peroxisomal palmitoyl-CoA oxidation approx. 2-fold. The ATP effect required the presence of Mg2+ and was lost when peroxisomal membranes were disrupted by Triton X-100 or high concentrations of unbound palmitoyl-CoA. 5. Disruption of peroxisomes by detergents, freeze--thawing, osmotic or mechanical treatment did not stimulate palmitoyl-CoA oxidation in the presence of ATP, indicating that peroxisomal fatty-acid-CoA oxidation was not latent. In the absence of ATP, Triton X-100 stimulated peroxisomal palmitoyl-CoA oxidation approx. 2-fold.
Subject(s)
Acyl Coenzyme A/metabolism , Liver/metabolism , Microbodies/metabolism , Organoids/metabolism , Palmitoyl Coenzyme A/metabolism , Adenosine Triphosphate/pharmacology , Animals , Hydrogen Peroxide/metabolism , In Vitro Techniques , Liver/drug effects , Male , Microbodies/drug effects , Mitochondria, Liver/metabolism , Oxidation-Reduction , Rats , Serum Albumin/metabolismSubject(s)
Lipase/metabolism , Lipid Mobilization , Liver/enzymology , Animals , Hydrogen-Ion Concentration , Kinetics , Rats , Subcellular Fractions/enzymologyABSTRACT
Mitochondrial and peroxisomal fatty acid oxidation were compared in whole liver homogenates. Oxidation of 0.2 mM palmitoyl-CoA or oleate by mitochondria increased rapidly with increasing molar substrate:albumin ratios and became saturated at ratios below 3, while peroxisomal oxidation increased more slowly and continued to rise to reach maximal activity in the absence of albumin. Under the latter condition mitochondrial oxidation was severely depressed. In homogenates from normal liver peroxisomal oxidation was lower than mitochondrial oxidation at all ratios tested except when albumin was absent. In contrast with mitochondrial oxidation, peroxisomal oxidation did not produce ketones, was cyanide-insensitive, was not dependent on carnitine, and was not inhibited by (+)-octanoylcarnitine, malonyl-CoA and 4-pentenoate. Mitochondrial oxidation was inhibited by CoASH concentrations that were optimal for peroxisomal oxidation. In the presence of albumin, peroxisomal oxidation was stimulated by Triton X-100 but unaffected by freeze-thawing; both treatments suppressed mitochondrial oxidation. Clofibrate treatment increased mitochondrial and peroxisomal oxidation 2- and 6- to 8-fold, respectively. Peroxisomal oxidation remained unchanged in starvation and diabetes. Fatty acid oxidation was severely depressed by cyanide and (+)-octanoylcarnitine in hepatocytes from normal rats. Hepatocytes from clofibrate-treated rats, which displayed a 3- to 4-fold increase in fatty acid oxidation, were less inhibited by (+)-octanoylcarnitine. Hydrogen peroxide production was severalfold higher in hepatocytes from treated animals oxidizing fatty acids than in control hepatocytes. Assuming that all H2O2 produced during fatty acid oxidation was due to peroxisomal oxidation, it was calculated that the contribution of the peroxisomes to fatty acid oxidation was less than 10% both in cells from control and clofibrate-treated animals.