Homogenization medium

2010 ◽  
Vol 2010 (11) ◽  
pp. pdb.rec12341-pdb.rec12341
Keyword(s):  
Homogenization Medium

The Para-O-Methylation of Apigenin to Acacetin by Cell-Free Extracts of Robinia pseudoacacia L.

1981 ◽  
Vol 36 (11-12) ◽  
pp. 916-920 ◽  
Author(s):  
Gary Kuroki ◽  
Jonathan E. Poulton
Keyword(s):  
Reaction Rate ◽  
Callus Tissue ◽  
Robinia Pseudoacacia ◽  
Methylation Pattern ◽  
Maximum Activity ◽  
Significant Activity ◽  
Coumaric Acid ◽  
Methyltransferase Activity ◽  
Robinia Pseudoacacia L ◽  
Homogenization Medium

Abstract Crude extracts from young Robinia pseudoacacia seedlings, shoots, and callus tissue catalyze the para-O-methylation of apigenin to acacetin using S-adenosyl-ʟ-methionine as methyl donor. Optimum activity was exhibited at pH 9.0, and Mg2+ was not required for maximum activity. EDTA (10 mᴍ) did not affect the reaction rate, but 47% inhibition was observed with SAH (100 μᴍ). β-Mercaptoethanol (5 mᴍ) was required in the homogenization medium for optimum O-methyltransferase activity. Apigenin (Km, 50 μᴍ) was the best substrate, but significant activity was shown towards caffeic acid, 5-hydroxyferulic acid, naringenin, and quercetin. Paracoumaric, ferulic, and sinapic acids were not methylated. The Km for S-adenosyl-ʟ-methionine was 31 μᴍ. Our demonstration of a para-O-methyltransferase activity methylating apigenin, but not para-coumaric acid, strongly supports the conclusion that the B-ring methylation pattern of acacetin is determined at the C15-level in Robinia pseudoacacia.

scholarly journals Activation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in homogenates of developing rat brain

1979 ◽  
Vol 182 (2) ◽  
pp. 367-370 ◽  
Author(s):  
W A Maltese ◽  
J J Volpe
Keyword(s):  
Rat Brain ◽  
Cholesterol Synthesis ◽  
Specific Activity ◽  
Postnatal Period ◽  
Developing Brain ◽  
Brain Maturation ◽  
Assay Mixture ◽  
Coa Reductase ◽  
Developing Rat ◽  
Homogenization Medium

The specific activity of 3-hydroxy-3-methylglutaryl-CoA reductase increases when homogenates of developing rat brain are incubated at 37 degrees C or kept on ice. This increase is completely blocked by the addition of F- to the homogenization medium and the assay mixture. The capacity for activation of the reductase is greatest during the early postnatal period and declines as brain maturation proceeds. The data suggest that catalytic modification of the reductase may play a role in the regulation of cholesterol synthesis in the developing brain.

Characteristics of rat pancreatic zymogen granules prepared by different methods

1986 ◽  
Vol 251 (3) ◽  
pp. G421-G429
Author(s):  
C. Niederau ◽  
J. H. Grendell ◽  
S. S. Rothman
Keyword(s):  
Digestive Enzyme ◽  
Percoll Gradient ◽  
Zymogen Granules ◽  
Experimental Conditions ◽  
Amylase Release ◽  
Ionic Solutions ◽  
Ph Values ◽  
Sucrose Solutions ◽  
Tissue Homogenates ◽  
Homogenization Medium

Zymogen granules isolated from tissue homogenates by differential centrifugation in isotonic sucrose solutions show substantial release of digestive enzyme when suspended in isotonic NaCl and in sucrose solutions at pH values above neutrality. A recent study reported a new method for isolating granules, involving the use of a complex homogenization medium and a Percoll gradient that was claimed to produce "stable" granules, i.e., granules that do not release their content in salt solutions and at pH values at or above neutrality. In the present study, we compare granules prepared in both ways, particularly in terms of their tendency to release amylase in isotonic ionic solutions and as a function of pH. The relative absence of amylase release from granules isolated by the new technique was found to be attributable to simple differences in the details of the experimental procedures that were used and not to actual differences in the characteristics of the two granule preparations. For example, previous studies with granules prepared in sucrose solutions reported substantial salt-induced release at 37 degrees C, whereas the recent study reporting the absence of salt-induced release from granules obtained from a Percoll gradient was done at 24 degrees C. Under the identical experimental conditions as used in the present study, little amylase release was seen at 24 degrees C for granules isolated by either technique, but substantial release was seen for both at 37 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals KCl-Dependent Release of Mitochondrial Membrane-Bound Arginase Appears to Be a Novel Variant of Arginase-II

Scientifica ◽  
2016 ◽  
Vol 2016 ◽  
pp. 1-10
Author(s):  
Mishra Suman ◽  
Mishra Rajnikant
Keyword(s):  
Outer Membrane ◽  
Mitochondrial Membrane ◽  
Functional Anatomy ◽  
Mitochondrial Enzymes ◽  
Novel Variant ◽  
Immunological Surveillance ◽  
Membrane Bound ◽  
Arginase Ii ◽  
Arginase I ◽  
Homogenization Medium

Arginase regulates arginine metabolism, ornithine-urea cycle, and immunological surveillance. Arginase-I is predominant in cytosol, and arginase-II is localised in the mitochondria. A mitochondrial membrane-bound arginase has also been proposed to be adsorbed with outer membrane of mitochondria which gets released by 150 mM potassium chloride (KCl). It is presumed that inclusion of 150 mM KCl in the homogenization medium would not only facilitate release of arginase bound with outer membrane of mitochondria but also affect functional anatomy of mitochondria, mitochondrial enzymes, and proteins. Therefore, it has been intended to characterize KCl-dependent release of mitochondrial membrane-bound arginase from liver of mice. Results provide advancement in the area of arginase biology and suggest that fraction of mitochondrial membrane-bound arginase contains mitochondrial arginase-II and a variant of arginase-II.

scholarly journals Tissue and subcellular distribution of the lectin from Datura stramonium (thorn apple)

1979 ◽  
Vol 184 (2) ◽  
pp. 215-219 ◽  
Author(s):  
D C Kilpatrick ◽  
M M Yeoman ◽  
A R Gould
Keyword(s):  
Cell Walls ◽  
Specific Activity ◽  
Soluble Fraction ◽  
Protein Body ◽  
Datura Stramonium ◽  
Immunofluorescent Staining ◽  
Lectin Activity ◽  
Thorn Apple ◽  
Mature Seeds ◽  
Homogenization Medium

Plants of Datura stramonium (thorn-apple) were dissected into their component tissues and examined for the presence of the Datura lectin. This lectin was easily detected in seeds and in various parts of the flowers of adult plants. Traces were also found in green (emerged) cotyledons and roots of seedlings. The specific lectin activity in seeds contained within the fruits increased as the seeds matured. Mature seeds were homogenized in sucrose and separated by differential centrifugation into four fractions, three of which were clearly of distinct composition. Most of the lectin activity sedimented with the low-speed (cell-wall/protein-body) pellet, but a similar specific activity was recovered from the other fractions. However, if EDTA was included in the homogenization medium, three or four times more lectin activity was recovered in the soluble fraction. Immunofluorescent staining of formaldehyde-fixed sections showed that the lectin was localized in the cytoplasm, with little associated with cell walls. The possible relevance of these results to the function of the lectin in plant cells is discussed.

Adaptation of the Phosphotungstate Method for the Determination of Vitamin C Contents in Animal and Human Tissues

2002 ◽  
Vol 57 (11-12) ◽  
pp. 1062-1065 ◽  
Author(s):  
Maciej Rutkowski ◽  
Krzysztof Grzegorczyk ◽  
Janusz Greger
Keyword(s):  
Vitamin C ◽  
Variation Coefficient ◽  
Human Tissues ◽  
Analytical Curve ◽  
Tissue Homogenates ◽  
The Right ◽  
Homogenization Medium

The usefulness of phosphotungstate reagent for vitamin C determination in tissue homogenates has been confirmed. An optimal homogenization medium was selected: 1.8 м solution of HPO3 in 1.3 м CH3COOH. With this medium the analytical curve (at 700 nm) demonstrated the right linearity, correlation and recovery coefficients were appropriately high (0.999 and 99.8%) and the values of intraserial and interserial variation coefficient were low (< 5% and < 10%, respectively). It makes this method sensitive, easily repeatable, and useful for vitamin C determination in animal and human tissues, including neoplastic ones.

scholarly journals Diurnal variation in the fraction of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in the active form in the mammary gland of the lactating rat

1986 ◽  
Vol 239 (2) ◽  
pp. 285-293 ◽  
Author(s):  
R A Smith ◽  
B Middleton ◽  
D W West
Keyword(s):  
Reductase Activity ◽  
Active Form ◽  
Total Activity ◽  
Maximum Activity ◽  
Covalent Modification ◽  
Light Phase ◽  
Hmg Coa Reductase ◽  
Enzyme Molecules ◽  
Coa Reductase ◽  
Homogenization Medium

‘Expressed’ and ‘total’ activities of 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) were measured in freeze-clamped samples of mammary glands from lactating rats at intervals throughout the 24 h light/dark cycle. ‘Expressed’ activities were measured in microsomal fractions isolated and assayed in the presence of 100 mM-KF. ‘Total’ activities were determined in microsomal preparations from the same homogenates but washed free of KF and incubated with exogenously added sheep liver phosphoprotein phosphatase before assay. Both ‘expressed’ and ‘total’ activities of HMG-CoA reductase underwent a diurnal cycle, which had a major peak 6 h into the light phase and a nadir 15 h later, i.e. 9 h into the dark period. Both activities showed a secondary peak of activity (around 68% of the maximum activity) at the time of changeover from dark to light, with a trough in the value of the ‘expressed’ activity that was close to the nadir value. ‘Expressed’ activity was lower than ‘total’ at all time points, indicating the presence of enzyme molecules inactivated by covalent phosphorylation. Nevertheless the ‘expressed’/‘total’ activity ratio was comparatively constant and varied only between 43% and 75%. Immunotitration of enzyme activity, with antiserum raised in sheep against purified rat liver HMG-CoA reductase, confirmed the presence of both active and inactive forms of the enzyme and indicated that at the peak and nadir the variation in ‘expressed’ HMG-CoA reductase activity resulted from changes in the total number of enzyme molecules rather than from covalent modification. The sample obtained after 3 h of the light phase exhibited an anomalously low ‘total’ HMG-CoA reductase activity, which could be increased when Cl- replaced F- in the homogenization medium. The result suggests that at that time the activity of the enzyme could be regulated by mechanisms other than covalent phosphorylation or degradation.

Prunasin Biosynthesis by Cell-Free Extracts from Black Cherry (Prunus serotina Ehrh.) Fruits and Leaves

1983 ◽  
Vol 38 (5-6) ◽  
pp. 369-374 ◽  
Author(s):  
Jonathan E. Poulton ◽  
Sun-In Shin
Keyword(s):  
Benzoic Acid ◽  
Phosphate Buffer ◽  
Mandelic Acid ◽  
Optimum Concentration ◽  
Prunus Serotina ◽  
Black Cherry ◽  
Significant Activity ◽  
Uridine Diphosphate ◽  
Diphosphate Glucose ◽  
Homogenization Medium

Immature fruits and leaves of black cherry (Prunus serotina Ehrh.) accumulate the cyanogenic glucoside prunasin (the β-glucoside of (R)-mandelonitrile). Cell-free extracts from these tissues catalysed the stereospecific glucosylation of (R,S)-mandelonitrile to (β)-prunasin at rates of 0.2-2.0 μmol/h/mg protein. Uridine diphosphate glucose (Km = 0.32 mᴍ) acted as glucose donor. The optimum concentration of (R,S)-mandelonitrile was 20 mᴍ. Highest activity was exhibited at pH 7.0-8.0 in Tris-phosphate buffer, and no additional cofactors were required. β- Mercaptoethanol (14.5 mᴍ), provided in the homogenization medium to prevent browning of homogenates, did not stimulate the rate of prunasin production. In addition to (R)-mandelonitrile (100%), significant activity was shown towards mandelamide (21%), benzyl alcohol (15%), mandelic acid (8%) and benzoic acid (153%), but not towards prunasin. Mandelonitrile glucosyltransferase activity was most stable at - 20 °C in the presence of 10% glycerol.

scholarly journals THE ISOLATION OF CELL NUCLEI FROM RAT BRAIN

1962 ◽  
Vol 15 (1) ◽  
pp. 109-120 ◽  
Author(s):  
Michael B. Sporn ◽  
Theodor Wanko ◽  
Wesley Dingman
Keyword(s):  
Rat Brain ◽  
Sucrose Solution ◽  
Cell Nuclei ◽  
Adult Rat ◽  
Adult Rat Brain ◽  
Pure Cell ◽  
Liver Nuclei ◽  
Homogenization Medium ◽  
Isopycnic Centrifugation

A method for preparing highly pure cell nuclei from adult rat brain, using both differential and isopycnic centrifugation in sucrose media, is described. The morphology of these preparations was examined by both phase contrast and electron microscopy. The isolated nuclei retained many aspects of their in situ morphology; in particular, the nuclear envelope was double layered and interrupted by pore-like discontinuities, and the nucleoli consisted of irregular masses of densely packed granules. Analyses of these nuclear preparations for cytochrome oxidase and cholinesterase activity, as well as RNA/DNA ratio, indicated minimal contamination with mitochondria and microsomes. Problems involving the homogenization technique, choice of ionic conditions in the homogenization medium, and choice of optimal density of the sucrose solution used for the final purification of nuclei are discussed. Results of application of the technique to isolation of adult rat liver nuclei are also reported.

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