Automated multistep column chromatography on ÄKTA pure system using in-line sample dilution

2020 ◽  
Vol 240 ◽  
pp. 116556 ◽  
Author(s):  
Leonildo Delgado ◽  
Bastian Franke ◽  
Tuomo Frigård ◽  
Shin Isogai
Keyword(s):  
Column Chromatography ◽  
Pure System ◽  
Sample Dilution ◽  
Line Sample

Heparin Excretion in Intact and Hepatectomized Rats

1981 ◽  
Vol 45 (02) ◽  
pp. 146-149
Author(s):  
Ray Losito ◽  
Harry Gattiker ◽  
Ginette Bilodeau
Keyword(s):  
Column Chromatography ◽  
Normal Values ◽  
Anticoagulant Activity ◽  
Biliary Fistula ◽  
Kinetics Of ◽  
Intact Rats

SummaryMetabolism and kinetics of 3H-heparin were compared in intact and hepatectomized rats. Rats were divided into three groups: 1) intact rats with biliary fistulas and cystostomies 2) intact rats with only cystostomies and 3) hepatectomized rats with cystostomies. Radioactivity in blood, bile and urine besides anticoagulant activity in blood and urine were examined. In addition, column chromatography of urine was used to isolate possible metabolites. Seventy percent and 80% of the radioactive dose was found in the urine of intact rats at 24 hr and 48 hr. Close to 5% of the radioactivity was found in bile or rats with a biliary fistula after 48 hr. The APTT declined to near normal values at 1 hr whether rats had a biliary fistula or not. In contrast, only 25 % of the radioactivity could be excreted into the urine of hepatectomized rats in 24 hr; the APTT did not decline as fast and at 5 hr, it was still 100 seconds. Only one radioactive component could be isolated on chromatography from all urines of these animals and appears to be similar to the original heparin. Thus, the liver has an important role to play in regulating the anticoagulant effects and excretion of heparin.

Autoantibodies to Thrombomodulin: Development of an Enzyme Immunoassay and a Survey of their Frequency in Patients with the Lupus Anticoagulant

1992 ◽  
Vol 67 (05) ◽  
pp. 507-509 ◽  
Author(s):  
John Gibson ◽  
Margaret Nelson ◽  
Ross Brown ◽  
Hatem Salem ◽  
Harry Kronenberg
Keyword(s):  
Enzyme Immunoassay ◽  
Lupus Anticoagulant ◽  
Specific Binding ◽  
Technical Problem ◽  
Serum Samples ◽  
Citrate Buffer ◽  
The Mean ◽  
Sample Dilution ◽  
Negative Population ◽  
Serum Components

SummaryIn order to investigate the possibility that autoantibodies to thrombomodulin (TM) may exist in patients with the lupus anticoagulant (LA) and perhaps be implicated in the pathogenesis of recurrent thrombosis seen in such patients, we developed an enzyme-immunoassay to screen serum samples for anti-human TM activity. The major technical problem encountered in developing this assay was to reduce the non-specific binding of serum components from both the LA positive and the negative population. Considerable reduction of non-specific binding was achieved by use of a phosphate/citrate buffer at pH 8.0 and the use of an optimal sample dilution of 1/40. In addition, samples were always tested in parallel in blank wells and results are expressed as an OD ratio. Samples from 113 patients with the LA were assayed and compared to 78 patients referred for LA testing but found to be negative. The mean OD values for the LA positive patients (± SD) was 1.36 (0.44) with a range of 0.78-2.57. This was virtually identical to the values for the LA negative population (1.38 ± 0.40, range 0.76-2.77). The results of this study indicate that there is no evidence for the presence of a significant autoantibody activity to TM in patients with the LA when compared to LA negative patients. If such autoantibodies do exist their frequency must be quite low.

GAGs-Potentiated Inhibition of Thrombin, Factor Xa and Plasmin in Plasma and in a Purified System Containing Antithrombin III – Correlation with Total Charge Density

1981 ◽  
Vol 46 (04) ◽  
pp. 749-751 ◽  
Author(s):  
E Cofrancesco ◽  
A Vigo ◽  
E M Pogliani
Keyword(s):  
Charge Density ◽  
Chemical Structure ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Total Charge ◽  
Pure System ◽  
At Iii ◽  
Total Charge Density ◽  
The Difference ◽  
Inhibition Of Thrombin

SummaryThe ability of heparin and related glycosaminoglycans (GAGs) to accelerate the inhibition of thrombin, factor Xa and plasmin in plasma and in a purified system containing antithrombin III (At III) was studied using chromogenic peptide substrate assaysThere was a good correlation between the charge density of the mucopolysaccharides and the activities investigated. While the difference between potentiation of the antithrombin activity by GAGs in plasma and in the purified system was slight, the inhibition of factor Xa in plasma was more pronounced than in the presence of purified At III, indicating the mechanisms for GAGs-potentiated inhibition of thrombin and factor Xa are not identical.For the antiplasmin activity, there was a good correlation between the chemical structure and biological activity only in the pure system, confirming that the antithrombin-GAG complex plays a very limited role in the inactivation of plasmin in plasma.

VANILLINSÄURE ALS ENDPRODUKT DES STOFFWECHSELS VON ADRENALIN UND NORADRENALIN. II.

1963 ◽  
Vol 44 (1) ◽  
pp. 101-106 ◽  
Author(s):  
Wilhelm Dirscherl ◽  
Helmut Thomas
Keyword(s):  
Rat Liver ◽  
Paper Chromatography ◽  
Column Chromatography ◽  
Vanillic Acid ◽  
Hippuric Acid ◽  
Single Spot ◽  
Solvent Systems ◽  
Perfusion Experiment ◽  
Kinetics Of

ABSTRACT Perfusion of rat liver with vanillic acid yielded only one metabolite. In paper chromatography with three different solvent systems, the substance showed the same RF-values as vanillyolglycine (3-methoxy-4-hydroxyhippuric acid) and in mixed chromatograms there was only one single spot. After separation by column chromatography, the UV- and IRspectra of the reaction product were identical with those of 3-methoxy4-hydroxy-hippuric acid. During the perfusion experiment, the kinetics of the conjugation were investigated.

DIE ZUSAMMENSETZUNG DER JODHALTIGEN VERBINDUNGEN UND IHRE BEZIEHUNG ZUM JODUMSATZ IN EUTHYREOTEN STRUMEN

1962 ◽  
Vol 41 (4) ◽  
pp. 584-594 ◽  
Author(s):  
Dankwart Reinwein ◽  
Erich Klein
Keyword(s):  
Column Chromatography ◽  
Iodine Content ◽  
High Plasma ◽  
Secretion Rate ◽  
Total Iodine ◽  
Iodine Compound ◽  
Hormonal Secretion ◽  
Carrier Free ◽  
Iodine Metabolism ◽  
Diffuse Goiter

ABSTRACT The iodine metabolism was investigated in 29 patients with euthyroid non-endemic diffuse goiter. 1 to 14 days before thyroidectomy the patients received carrier-free 131I. The chemical iodine fractions (PBI, BEI and iodide) of the thyroid and the labelled iodine compound were analyzed by means of paper- and column chromatography. In one gland the total iodine content varied only by ± 19.6% of the average, the relative shares of PBI, BEI and iodide as well as that of the iodoamino acids being equal. Monoiodotyrosine, diiodotyrosine and thyroxine were found in the thyroid homogenate without hydrolysis. The homogenate after hydrolysis contained more iodotyrosines at the expense of iodothyronines than do normal glands. 17 goiters with normal 131I-uptake showed a high total iodine content (14.2 ± 5.0 mg) whilst 9 goiters with an increased 131I-uptake had low values (3.58 ± 0.6 mg). The opposite was found for the relative shares of BEI with the chromatographically isolated iodothyronines thyroxine, triiodothyronine and an unidentified iodine compound. Goiters with »high plasma PB131I« were characterized by a faster transfer of 131I into the more heavily iodinated compounds than is found in glands with a normal hormonal secretion rate. The highest values for the iodothyronines were found in goiters with increased 131I-uptake together with a high hormonal secretion rate. From this study it appears that the changes in the iodine-poor glands are due to a defective exo- or endogenous iodine supply. The observed alterations in iodine-rich glands are probably induced by a faulty iodine utilization characterized by an incomplete iodotyrosyl-coupling defect.

Improvement of monitoring of tertiary filtration with particle counting

2006 ◽  
Vol 6 (1) ◽  
pp. 1-9
Author(s):  
V. Miska ◽  
J.H.J.M. van der Graaf ◽  
J. de Koning
Keyword(s):  
Particle Surface ◽  
Minor Effect ◽  
Particle Size Distributions ◽  
Particle Volume ◽  
Mass Distributions ◽  
Particle Counting ◽  
Total Particle ◽  
Particle Counts ◽  
A Minor ◽  
Sample Dilution

Nowadays filtration processes are still monitored with conventional analyses like turbidity measurements and, in case of flocculation–filtration, with phosphorus analyses. Turbidity measurements have the disadvantage that breakthrough of small flocs cannot be displayed, because of the blindness regarding changes in the mass distributions. Additional particle volume distributions calculated from particle size distributions (PSDs) would provide a better assessment of filtration performance. Lab-scale experiments have been executed on a flocculation–filtration column fed with effluent from WWTP Beverwijk in The Netherlands. Besides particle counting at various sampling points, the effect of sample dilution on the accuracy of PSD measurements has been reflected. It was found that the dilution has a minor effect on PSD of low turbidity samples such as process filtrate. The correlation between total particle counts, total particle volume (TPV) and total particle surface is not high but is at least better for diluted measurements of particles in the range 2–10 μm. Furthermore, possible relations between floc-bound phosphorus and TPV removal had been investigated. A good correlation coefficient is found for TPV removal versus floc-bound phosphorus removal for the experiments with polyaluminiumchloride and the experiments with single denitrifying and blank filtration.

Protein Solvent Interaction: Transition of Protein-solvent Interaction Concept from Basic Research into Solvent Manipulation of Chromatography

2018 ◽  
Vol 20 (1) ◽  
pp. 34-39 ◽  
Author(s):  
Tsutomu Arakawa ◽  
Yoshiko Kita
Keyword(s):  
Column Chromatography ◽  
Rapid Development ◽  
Academic Research ◽  
Basic Research ◽  
Protein Stabilization ◽  
Special Issue ◽  
Solvent Interaction ◽  
Protein Formulations ◽  
Solvent Additive ◽  
Downstream Processes

Previously, we have reviewed in this journal (Arakawa, T., Kita, Y., Curr. Protein Pept. Sci., 15, 608-620, 2014) the interaction of arginine with proteins and various applications of this solvent additive in the area of protein formulations and downstream processes. In this special issue, we expand the concept of protein-solvent interaction into the analysis of the effects of solvent additives on various column chromatography, including mixed-mode chromatography. Earlier in our research, we have studied the interactions of such a variety of solvent additives as sugars, salts, amino acids, polymers and organic solvents with a variety of proteins, which resulted in mechanistic understanding on their protein stabilization and precipitation effects, the latter known as Hofmeister series. While such a study was then a pure academic research, rapid development of genetic engineering technologies and resultant biotechnologies made it a valuable knowledge in fully utilizing solvent additives in manipulation of protein solution, including column chromatography.

A Cytotoxic Natural Product from Patrinia villosa Juss

2019 ◽  
Vol 19 (11) ◽  
pp. 1399-1404 ◽  
Author(s):  
Yangcheng Liu ◽  
Wei Liu ◽  
Changlan Chen ◽  
Zheng Xiang ◽  
Hongwei Liu
Keyword(s):  
Cell Cycle ◽  
Cell Lines ◽  
Silica Gel ◽  
Column Chromatography ◽  
Bioactive Compound ◽  
Antitumor Agent ◽  
2D Nmr ◽  
Preparative Hplc ◽  
Dependent Manner ◽  
2D Nmr Spectra

Background and Purpose:: Patrinia villosa Juss is an important Chinese herbal medicine widely used for thousands of years, but few reports on the ingredients of the herb have been presented. In this study, we aim to isolate the bioactive compound from the plant. Material and Methods:: The air-dried leaves of P. villosa (15kg) were extracted three times with 70% EtOH under reflux. The condensed extract was suspended in H2O and partitioned with light petroleum, dichloromethane and n-BuOH. The dichloromethane portion was then subjected to normal-phase silica gel column chromatography, ODS silica gel column chromatography and semi-preparative HPLC to yield compound 1. Cytotoxicities of 1 were assayed on HepG2, A549 and A2780 cell lines. The mechanism of apoptosis and cell cycle on A549 was confirmed subsequently. Results: A new impecylone (Impecylone A) was isolated from the leaves of Patrinia villosa Juss, and its structures were established using 1D, 2D-NMR spectra and HR-ESI-MS. Impecylone A could selectivity inhibit HepG2 and A549 cell lines. The compound could induce apoptosis of A549 and arrest the cell cycle at G2/M phase in a dose-dependent manner. Conclusion: Impecylone A is a novel compound from Patrinia villosa Juss and could be a potential antitumor agent especially in the cell lines of A549.

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