Mitochondrial fraction

Creatine supplementation has been shown to increase the rate of PCr resynthesis in the recovery period after ischemic exercise (Greenhaff et al., 1993a). This could be attributed to an acceleration of oxidative phosphorylation by increased free creatine content available to the mitochondrial fraction of the creatine kinase enzyme, as previously suggested (Bessman and Fonyo, 1966). 

Polarographic studies of a mitochondrial fraction from Hymenolepis diminuta showed that of four substrates tested, DL-glycerol-3-phosphate was the most rapidly oxidized, but the highest respiratory control ratio (1.7) was obtained with dl-isocitric acid. With isocitrate as substrate oxyclozanide at 1.61 nM stimulated O uptake and relieved oligomycin inhibition of adinosine diphosphate-stimulated respiration, but at concentrations above 2 pM progressively inhibited O uptake. Rafoxanide, niclosamide, 3,4,5-tribromo-salicylanilide, nitroxynil, resorantel, di-chlorophen, and 2,4-dinitrophenol exhibited effects similar to those of oxyclozanide on the respiration in cestode mitochondria. The relative potencies were compared and the possible mode of action discussed [38]. 

Numerous studies were dedicated to the effects of flavonoids on microsomal and mitochondrial lipid peroxidation. Kaempferol, quercetin, 7,8-dihydroxyflavone and D-catechin inhibited lipid peroxidation of light mitochondrial fraction from the rat liver initiated by the xanthine oxidase system [126]. Catechin, rutin, and naringin inhibited microsomal lipid peroxidation, xanthine oxidase activity, and DNA cleavage [127]. Myricetin inhibited ferric nitrilotriacetate-induced DNA oxidation and lipid peroxidation in primary rat hepatocyte cultures and activated DNA repair process [128]. 

Rhodanese is widely distributed in the body, but activity levels in mammals are highest in the mitochondrial fraction of liver. Rhodanese activity levels in catalyzing the transformation of thiosulfate to thiocyanate are limited by the availability of sulfur. Minor detoxification pathways for cyanide include exhalation in breath as HCN, and as C02 from oxidative metabolism of formic... 

Mercuric chloride is a potent nephrotoxicant in the adult rat, but has little effect on the newborn [222], There are significant maturational changes in organ, cellular and subcellular distribution of mercury during the first 4 weeks after birth. With increasing age, mercury is redistributed from the renal cytosolic fraction to the nuclear/mitochondrial fraction, where it may be more damaging. 

Careful examination of the yellowish sediment obtained after spinning down the crude mitochondrial fraction showed it was frequently overlaid with loosely packed, fluffy material —the fluffy layer. Experiments from de Duve s and, later, Novikoff s laboratories in the 1950s demonstrated that the lighter, lysosomal fraction was enriched in a number of hydrolases including acid phosphatase, aryl sulphatase, B glucuronidase, RNAase, and a peptidase, cathepsin. All the enzymes had optimal pHs in the acid range (pH 5-pH 6). Density... 

There was considerable epoxidase activity in the mitochondrial fraction but the highest activity was in the washed microsome fraction. Microsomes washed in KC1 were more active than unwashed microsomes. No epoxidase activity was detected in the post-microsomal supernatant, or when PBO was added to microsomes (Table VIII). 

Found in mitochondrial fraction of liver mRNA not detected in rat brain (Mellon, 1993). 

C atropine to the mitochondrial fraction of the rat brain. Edgewood Arsenal Technical Report 4042, 1966. 

KMBA Oxidative stress parameter OCPs, nitroaromatics, PAHs Mitochondrial fraction l.t. pooled... 

Effect of standardized P. ginseng extract G115 after exhaustion exercise glutathion peroxidase (GPX) enzymatic activity (red gastrocnemius) "mitochondrial fraction."... 

Compart mentation in fi Oxidation Free palmitate is activated to its coenzyme A derivative (palmitoyl-CoA) in the cytosol before it can be oxidized in the mitochondrion. If palmitate and [ CJcoenzyme A are added to a liver homogenate, palmitoyl-CoA isolated from the cytosolic fraction is radioactive, but that isolated from the mitochondrial fraction is not. Explain. 

Many of the biochemical processes that generate chemical energy for the cell take place in the mitochondria. These organelles contain the biochemical equipment necessary for fatty acid oxidation, di- and tricarboxylic acid oxidation, amino acid oxidation, electron transport, and ATP generation. In this experiment, a mitochondrial fraction will be isolated from beef heart muscle. The mitochondria will be analyzed for protein content and fractionated into submitochondrial particles. Each fraction will be analyzed for malate dehydrogenase and monoamine oxidase activities. 

Rather, the isolated subcellular fraction is a mitochondrial fraction, which indicates that its major components are mitochondria. Other cellular components that may be present are lysosomes, cell fragments, nuclear fragments, and microbodies (peroxisomes). The purity of the fraction depends on the source of the extract and the method chosen for isolation. 

The general procedure for preparation of a mitochondrial fraction consists of the following steps ... 

Select and procure a suitable biological material. Mitochondrial fractions have been prepared from many animal organs including liver, heart, muscle, and brain, many plant tissues including spinach and avocado, and yeast. 

The heavy mitochondrial fraction is suspended in buffer and homogenized for a brief period in a glass-Teflon homogenizer to complete cell lysis. 

This experiment describes the preparation of a mitochondrial fraction from beef heart muscle. Heart muscle is an excellent choice of tissue because isolated mitochondria are stable and most enzyme activities remain high for a long period of time. Since heart muscle is more fibrous than other tissues, some problems are encountered in homogenizing the tissue. The preparation described here is suitable for the study of characteristic enzymatic activity, electron transport, and ADP phosphorylation. 

Mitochondrial fractions may be characterized by testing for the presence of known enzyme activities as previously discussed. The relative purity of each fraction can be estimated by measuring the specific activity of marker enzymes. Table El0.1 identifies marker enzymes for the matrix and membranes. Malate dehydrogenase, the tricarboxylic acid cycle enzyme that catalyzes the interconversion of malate and oxaloacetate (Equation E10.1), serves as a marker for the matrix enzymes. 

Malate dehydrogenase activity would be expected in intact mitochondria, but not in SMPs. The activity of this enzyme in mitochondrial fractions may be estimated by a spectrophotometric assay. Oxaloacetate and NADH are incubated, and the disappearance of NADH is monitored at 340 nm. NAD+ does not have strong absorption at this wavelength. Note that the reverse reaction is studied because the reaction as shown above is very unfavorable in thermodynamic terms ACT = +30 kj/mol). 

A. Isolation of the mitochondrial fraction—2 hours. It is recommended that the isolation procedure up to step 13 be done as a class project with each student group completing two or three sequential steps. The centrifugation step (step 13) may be done in several 50-mL tubes and each student group receives a centrifuge tube to continue through step 20 on their own. 

Mitochondrial fraction from part A Sucrose-Tris isolation solution Sonicator Ice-salt bath... 

SMP fraction from part C Mitochondrial fraction from part A - Phosphate buffer, 0.2 M, pH 7.4... 

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