Separation of Heparin into Subtractions by DEAE-Cellulose Chromatography

1979 ◽  
Vol 42 (05) ◽  
pp. 1452-1459 ◽  
Author(s):  
Robert H Yue ◽  
Toby Starr ◽  
Menard M Gertler
Keyword(s):  
High Activity ◽  
Uronic Acid ◽  
Deae Cellulose ◽  
Exchange Chromatography ◽  
Net Charge ◽  
Uronic Acid Content ◽  
Successful Separation ◽  
Salt Gradient ◽  
Structure And Activity ◽  
Low Activity

SummaryCommercial porcine heparin can be separated into three distinct subtractions by using DEAE-cellulose chromatography and a stepped salt gradient. Gram quantities of heparin can be fractionated by this technique. All three heparin subtractions can accelerate the inhibition of thrombin by antithrombin III with different efficiency. The specific activities of the high activity heparin, intermediate activity heparin and low activity heparin are 228 units/mg, 142 units/mg and 95 units/mg, respectively. Both the uronic acid content and the quantity of N-SO4 for all three heparin subfractions have been evaluated. The high activity heparin has the lowest uronic acid and N-SO4 content. The successful separation of commercial heparin into three distinct subfractions by means of ion-exchange chromatography suggests that the net charge on these three heparin components will serve as a model system in the elucidation of the structure and activity relationship to the biological function of heparin.

scholarly journals The purification and properties of carbonic anhydrases from guinea-pig erythrocytes and the mucosae of the gastrointestinal tract

1970 ◽  
Vol 120 (4) ◽  
pp. 797-808 ◽  
Author(s):  
M. J. Carter ◽  
D. S. Parsons
Keyword(s):  
Gastrointestinal Tract ◽  
Carbonic Anhydrase ◽  
Guinea Pig ◽  
High Activity ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Carbonic Anhydrases ◽  
Low Activity

Procedures for isolating carbonic anhydrase (EC 4.2.1.1) enzymes from the erythrocytes and the mucosae of the gastrointestinal tract of guinea pigs are described. From a haemolysate, haemoglobin was removed by the addition of ammonium sulphate, and also by two other methods, namely by gel filtration or by adsorption on DEAE-Sephadex. The crude enzyme thus obtained was resolved into the different isoenzymes by chromatography with DEAE-cellulose. From particle-free supernatants of homogenates of some gastrointestinal tissues, carbonic anhydrases were purified by ammonium sulphate fractionation, gel filtration, and ion-exchange chromatography with DEAE-cellulose. The major isoenzymes from blood, stomach, proximal colonic mucosa and caecal mucosa were homogeneous during ion-exchange chromatography, acrylamide-gel electrophoresis, and centrifugal examination. From these tissues, carbonic anhydrase was isolated as two major isoenzymes. They resemble the pairs of isoenzymes discovered in the bloods of other species. The carbon dioxide hydratase activity of one isoenzyme (`high activity' carbonic anhydrase) was 40 times that of the other isoenzyme (`low activity' carbonic anhydrase), as measured at a single substrate concentration. Two other minor components of the enzyme are also found in guinea-pig erythrocytes. All of the enzymes isolated had molecular weights of nearly 30000 (sedimentation equilibrium). `High activity' carbonic anhydrases from blood and gastrointestinal tissues were indistinguishable according to some chemical, physical and kinetic measurements; similarly `low activity' carbonic anhydrases from those tissues were indistinguishable. `High activity' carbonic anhydrase was markedly different from the `low activity' carbonic anhydrase with respect to its amino acid composition, chromatographic behaviour and isoelectric pH value. Marked differences were also found in the tissue concentrations of the major isoenzymes. It is suggested that the characteristic and selective distribution of the different forms of carbonic anhydrase in the guinea-pig tissues is related to the specific and different physiological functions of the enzymes.

scholarly journals Carbonic anhydrase isoenzymes in the erthrocytes and dorsolateral prostate of the rat

1969 ◽  
Vol 114 (3) ◽  
pp. 463-476 ◽  
Author(s):  
J. E. A. McIntosh
Keyword(s):  
Carbonic Anhydrase ◽  
High Activity ◽  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Exchange Chromatography ◽  
Kinetic Properties ◽  
Efficient Catalyst ◽  
Molecular Weights ◽  
Hydrolysis Of ◽  
Low Activity

1. Three forms of the zinc-containing enzyme carbonic anhydrase (EC 4.2.1.1) were isolated from the erythrocytes of the rat and two forms from the dorsolateral prostate of the rat. Several additional minor components were observed but not isolated. Separation of the isoenzymes was achieved by ion-exchange chromatography, polyacrylamide-gel electrophoresis and isoelectric focusing. 2. The general properties of the isolated isoenzymes, their molecular weights and their contents of zinc were closely similar. As catalysts of the hydration of carbon dioxide, however, they were distinctly different. The two most abundant isoenzymes of the erythrocytes, which were found in equal proportions, differed 70-fold in specific activity, whereas the isoenzymes of the dorsolateral prostate were similar to one another and resembled the high-activity component of the erythrocytes. The inhibition of the latter by acetazolamide (5-acetamido-1-thia-3,4-diazole-2-sulphonamide) was mainly competitive, whereas in identical conditions the low-activity erythrocyte component and the dorsolateral prostate isoenzymes were non-competitively inhibited. 3. The use of chloroform–ethanol to remove haemoglobin from the rat haemolysate was found (a) to bring about changes in the kinetic properties of the soluble isoenzymes and (b) to cause the appearance of an additional isoenzyme. 4. The actions were compared of the inhibitors acetazolamide, 1,1-dimethylaminonaphthalene-5-sulphonamide and ethoxzolamide (6-ethoxybenzothiazole-2-sulphonamide) on the hydrolysis of p-nitrophenyl acetate catalysed by the isoenzymes. 5. The low-activity erythrocyte isoenzyme was an efficient catalyst of the hydrolysis of β-naphthyl acetate whereas the high-activity forms were much less active towards this ester. Neither of the isoenzymes present in the dorsolateral prostate catalysed this reaction. 6. Carbonic anhydrase in the rat dorsolateral prostate accounts for no more than 5% of the unusually high content of zinc in this organ.

scholarly journals Chemical composition diversity of fucoidans isolated from three tropical brown seaweeds (Phaeophyceae) species

2020 ◽  
Vol 21 (7) ◽  
Author(s):  
FADILAH NOR LAILI LUTFIA ◽  
Alim Isnansetyo ◽  
Ratna Asmah Susidarti ◽  
Muhammad Nursid
Keyword(s):  
Chemical Composition ◽  
Uronic Acid ◽  
Dry Matter ◽  
Deae Cellulose ◽  
Brown Seaweed ◽  
Exchange Chromatography ◽  
Chemical Components ◽  
Brown Seaweeds ◽  
Total Carbohydrates ◽  
High Sulfate

Abstract. Lutfia FN, Isnansetyo A, Susudarti RA, Nursid M. 2020. Chemical composition diversity of fucoidans isolated from three tropical brown seaweeds (Phaeophyceae) species. Biodiversitas 21: 3170-3177. Fucoidan is a polysaccharide with high sulfate content, found in the cell walls matrix of brown seaweed. Its bioactivities vary depending on the algae species and the chemical structure. The aims of this research were to extract, purify, and characterize fucoidan from three Indonesian brown seaweed, Sargassum sp., Turbinaria sp., and Padina sp. The extraction was carried out using acid method followed by precipitation with ethanol and CaCl2, while its purification using DEAE-cellulose ion-exchange chromatography. Characterization was performed by FTIR and 1H-NMR spectroscopic analyses. Chemical components of fucoidan determined were total carbohydrates, sulfate residue, uronic acid residue, and monosaccharide components. The results revealed that the Turbinaria sp. produced highest yield of fucoidan (4.8% dry matter), followed by Sargassum sp. (2.7% dry matter) and Padina sp. (2.6% dry matter). The carbohydrate contents of Sargassum sp., Turbinaria sp. and Padina sp. fucoidan were 64.55±0.12%, 67.42 ± 0.03% and 62.90 ± 0.04% with sulfate residues of 0.013% ± 4.71; 19.47±0.002% and 0.016%±8.81; and uronic acid residues of 25.19±0.03%; 12.69±0.03% and 12.91±0.01%, respectively. Sargassum sp., Turbinaria sp., and Padina sp. fucoidans consist of fucose and some other minor monosaccharides.

scholarly journals Studies on the structure and activity of rabbit C1q (a subcomponent of the first component of complement)

1974 ◽  
Vol 143 (2) ◽  
pp. 265-272 ◽  
Author(s):  
Diane M. Lowe ◽  
Kenneth B. M. Reid
Keyword(s):  
Amino Acids ◽  
Deae Cellulose ◽  
Exchange Chromatography ◽  
Haemolytic Activity ◽  
Subunit Structure ◽  
Carboxypeptidase A ◽  
Rapid Loss ◽  
Collagenase Digestion ◽  
Polypeptide Chains ◽  
Structure And Activity

1. The subunit structure of rabbit subcomponent C1q was examined in a previous publication (Reid et al., 1972). The present paper describes some aspects of the structure of the polypeptide chains derived from the molecule. 2. The three polypeptide chains, produced by performic oxidation, of rabbit subcomponent C1q were isolated by ion-exchange chromatography in 8m-urea on DEAE-cellulose. 3. Each chain was found to contain 15–18% glycine and significant amounts of the amino acids hydroxyproline and hydroxylysine. 4. By means of collagenase digestion it was shown that all three chains of rabbit subcomponent C1q contain collagen-like sequences of amino acids which constitute about 40% of each chain. 5. By use of carboxypeptidase A it was established, indirectly, that the collagen-like sequences, in one of the chains, are probably located near, or at, the N-terminal end of the chain. 6. Collagenase digestion and heating at 52°C (but not at 49°C) caused rapid loss of native rabbit subcomponent C1q haemolytic activity.

scholarly journals Biochemical analyses of EDTA extracts and collagenase digests from bone and skin of Wistar rats

1982 ◽  
Vol 16 (3) ◽  
pp. 256-264 ◽  
Author(s):  
J. M. Mbuyi ◽  
J. Dequeker
Keyword(s):  
Sialic Acid ◽  
Plasma Proteins ◽  
Uronic Acid ◽  
Deae Cellulose ◽  
Organic Matrix ◽  
Rat Skin ◽  
Acid Glycoprotein ◽  
Uronic Acid Content ◽  
The Matrix ◽  
Biochemical Analyses

Using EDTA extraction and collagenase digestion, rat bone and rat skin were compared in terms of their content of hydroxyproline, hexoses, uronic acid, sialic acid and plasma proteins. The collagen content of the organic matrix from both tissues was similar. Greater differences were observed in the sialic acid and uronic acid content of the matrix, bone containing higher amounts; smaller differences were found in the levels of hexoses, albumin, IgG and α1 acid-glycoprotein, which are higher in EDTA extracts from bone. The DEAE-cellulose chromatography of the EDTA extracts and soluble collagenase digests indicated the presence of a variety of glycoproteins and a proteoglycan fraction. An acidic glycoprotein, corresponding to sialoprotein, was present in bone but not in skin extracts.

scholarly journals Phosphatase activity of Poa pratensis seeds. I. Preliminary studies on acid phosphatase II

2015 ◽  
Vol 42 (3) ◽  
pp. 369-377
Author(s):  
I. Lorenc-Kubis ◽  
B. Morawiecka
Keyword(s):  
Acid Phosphatase ◽  
High Activity ◽  
Phosphatase Activity ◽  
Sodium Acetate ◽  
Poa Pratensis ◽  
Deae Cellulose ◽  
Acetate Buffer ◽  
Sodium Acetate Buffer ◽  
Nitrophenyl Phosphate ◽  
Low Activity

Acid phosphatase (EC 3.1.3.2) was extracted with 0.1 M sodium acetate buffer pH 5.1 from <i>Poa pratensis</i> seeds, and separated into three fractions by chromatography on DEAE cellulose. The highest activity was found in fraction Il-b (acid phosphatase II). The activity of the enzyme was optimal at pH 4.9. It hydrolyzed p-nitrophenyl phosphate most readily among the various phosphomonoesters examined. Acid phosphatase II showed also a high activity toward β-naphtyl phosphate and phenyl phosphate, very low activity towards β-glycero phosphate, 5'-GMP and no activity with glucose-1 phosphate. The enzyme was inhibited by Ca<sup>2+</sup> and fluoride, but activated by Mg<sup>2+</sup>. EDTA had no influence on the activity of the enzyme.

Extracellular polysaccharides from suspension cultures of Nicotiana plumbaginifolia

2020 ◽  
Author(s):  
Ian Sims ◽  
A Bacic
Keyword(s):  
Uronic Acid ◽  
Cell Suspension Cultures ◽  
Anion Exchange Chromatography ◽  
Nicotiana Plumbaginifolia ◽  
Suspension Cultures ◽  
Exchange Chromatography ◽  
Arabinogalactan Protein ◽  
Extracellular Polysaccharides ◽  
Selective Precipitation ◽  
The Individual

The soluble polymers secreted by cell-suspension cultures of Nicotiana plumbaginifolia contained 78% carbohydrate, 6% protein and 4% inorganic material. The extracellular polysaccharides were separated into three fractions by anion-exchange chromatography using a gradient of imidazole-HCl at pH 7 and the individual polysaccharides in each fraction were then isolated by selective precipitation and enzymic treatment. Monosaccharide and linkage compositions were determined for each polysaccharide after reduction of uronic acid residues and the degree of esterification of the various uronic acid residues in each polysaccharide was determined concurrently with the linkage types. Six components were identified: an arabinoxyloglucan (comprising 34% of the total polysaccharide) and a galactoglucomannan (15%) in the unbound neutral fraction, a type II arabinogalactan (an arabinogalactan-protein, 11%) and an acidic xylan (3%) in the first bound fraction, and an arabinoglucuronomannan (11%) and a galacturonan (26%) in the second bound fraction. © 1995.

Binding of lead and copper(II) cations to hemicelluloses of rye bran

1986 ◽  
Vol 51 (10) ◽  
pp. 2250-2258 ◽  
Author(s):  
Rudolf Kohn ◽  
Zdena Hromádková ◽  
Anna Ebringerová
Keyword(s):  
Uronic Acid ◽  
Equilibrium Solution ◽  
Glucuronic Acid ◽  
The Other ◽  
Carboxyl Groups ◽  
Stoichiometric Ratio ◽  
Molar Ratios ◽  
Free Ions ◽  
Uronic Acid Content ◽  
The Relationship

Several fractions of acid hemicelluloses isolated from rye bran were characterized by molar ratios of saccharides (D-Xyl, L-Ara, D-Glc, D-Gal) and 4-O-methyl-D-glucuronic acid and protein content. Binding of Pb2+ and Cu2+ ions to these acid polysaccharides was considered according to function (M)b = f([M2+]f), expressing the relationship between the amount of metal (M)b bound to 1 g of the substance and the concentration of free ions [M2+]f in the equilibrium solution and according to the association degree β of these cations with carboxyl groups of uronic acid at a stoichiometric ratio of both components in the system under investigation. Acid hemicelluloses contained only a very small portion of uronic acid ((COOH) 0.05-0.18 mmol g-1); the model polysaccharide, 4-O-methyl-D-glucurono-D-xylan of beech, was substantially richer in uronic acid content ((COOH) 0.73 mmol g-1). Consequently, the amount of lead and copper bound to acid hemicelluloses is very small ((M)b 0.017-0.025 mmol g-1) at [M2+]f = 0.10 mmol l-1. On the other hand, much greater amount of cations ((M)f 0.09-0.10 mmol g-1) was bound to the glucuronoxylan. The association degree β was like with the majority of samples (β = 0.31-0.38). The amount of lead and copper(II) bound to acid hemicelluloses from rye bran is several times lower than that bound to dietary fiber isolated from vegetables (cabbage, carrot), rich in pectic substances.

scholarly journals Physical Activity Trajectories and Associated Changes in Physical Performance in Older Men: The MrOS Study

2020 ◽  
Vol 75 (10) ◽  
pp. 1967-1973
Author(s):  
Deepika R Laddu ◽  
Neeta Parimi ◽  
Katie L Stone ◽  
Jodi Lapidus ◽  
Andrew R Hoffman ◽  
...  
Keyword(s):  
Physical Activity ◽  
Grip Strength ◽  
High Activity ◽  
Physical Performance ◽  
Gait Speed ◽  
Functional Independence ◽  
Performance Measure ◽  
Least Square ◽  
Activity Group ◽  
Low Activity

Abstract Background Physical activity (PA) is important to maintaining functional independence. It is not clear how patterns of change in late-life PA are associated with contemporaneous changes in physical performance measures. Methods Self-reported PA, gait speed, grip strength, timed chair stand, and leg power were assessed in 3,865 men aged ≥ 65 years at baseline (2000–2002) and Year 7 (2007–2009). Group-based trajectory modeling, using up to four PA measures over this period, identified PA trajectories. Multivariate linear regression models (adjusted least square mean [95% confidence interval {CI}]) described associations between-PA trajectories and concurrent changes in performance. Results Three discrete PA patterns were identified, all with declining PA. Linear declines in each performance measure (baseline to Year 7) were observed across all three PA groups, but there was some variability in the rate of decline. Multivariate models assessing the graded response by PA trajectory showed a trend where the high-activity group had the smallest declines in performance while the low-activity group had the largest (p-for trend &lt; .03). Changes in the high-activity group were the following: gait speed (−0.10 m/s [−0.12, −0.08]), grip strength (−3.79 kg [−4.35, −3.23]), and chair stands (−0.38 [−0.50, −0.25]), whereas changes in the low-activity group were the following: gait speed (−0.16 [−0.17, −0.14]), grip strength (−4.83 kg [−5.10, −4.55]), and chair stands (−0.53 [−0.59, −0.46]). Between-group differences in leg power trajectories across PA patterns were not significant. Conclusions Declines in functional performance were higher among those with lower PA trajectories, providing further evidence for the interrelationship between changes in PA and performance during old age.

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