Sephadex Gel Column Chromatography Research Articles

Determination of high polymer in flucloxacillin sodium capsule by sephadex gel column chromatography

Determination of high polymer in flucloxacillin sodium capsule by sephadex gel column chromatography

Separation of aromatic monomers from oxidatively depolymerized products of lignin by combining Sephadex and silica gel column chromatography

Abstract The conversion of lignin into value-added aromatic monomers has attracted much attention as these monomers can be potentially used to prepare liquid fuel rather than heavy oil by further hydrodeoxygenation. A new process combining Sephadex and silica gel column chromatography was employed to obtain aromatic monomers with higher purity. It was found that pH and polarity of mobile phase were the most important factors affecting the separation effect for Sephadex G-10 and silica gel column chromatography, respectively. When the pH of mobile phase reached 10.5, the ionization of hydroxyl together with carboxyl groups resulted in electrostatic repulsion between the aromatic compounds and Sephadex G-10 gel, which counteracted the adsorption caused by hydrogen bond between aromatics and gel, causing the increase of the separation effect. Under the condition of pH above 10.5, Sephadex gel column chromatography gave 78.1% of purity of aromatic monomers (PAM) and 99.5% of recovery of aromatic monomers (RAM) due to the elimination of non-size exclusion effect. Silica gel column chromatography was further conducted to obtain 99.8% of PAM and 73.9% of RAM. The polar mobile phase was favorable for eluting aromatic monomers, while the non-polar mobile phase was beneficial to remove oligomers and increasing the PAM.

Isolation and characterization of an antifungal compound 5-hydroxy-7,4'-dimethoxyflavone from Combretum zeyheri.

BackgroundCombretum zeyheri, belongs to the family Combretaceae and is one of the most popular herbal plants in tropical and subtropical countries. The leaves of Combretum zeyheri have been used as herbal medicine and have been reported to have pharmacological activity which includes anti-bacterial, anti-fungal, anticancer and antioxidant properties. The goal of this study was to isolate, identify and characterize compounds from C. zeyheri leaves which are responsible for its antifungal activity.MethodsThe preliminary isolation of C. zeyheri active compounds was carried out using chromatographic techniques which include sephadex gel column chromatography, silica gel column chromatography and thin-layer chromatography (TLC). The isolated compounds were then investigated for their antifungal activity using broth dilution assay. The combined effect of the most potent compound and an antifungal drug miconazole was investigated using the checkerboard assay. Time-kill assays were conducted for the combinations using the colony counting method. The mechanism of action of 5-hydroxy-7,4’-dimethoxyflavone as a potent antifungal agent was investigated by determining its inhibitory activity on Candida albicans drug efflux pumps using the ciprofloxacin assay. The ability of 5-hydroxy-7,4’-dimethoxyflavone to inhibit antioxidant enzymes as well as the biosynthesis of ergosterol were also investigated.ResultsA total of four pure compounds (A-D) were isolated from C. zeyheri leaf extract. Compound B (5-hydroxy-7,4’-dimethoxyflavone) was found to be active against Candida albicans using broth dilution method. This compound was also found to have synergistic activity on growth of C. albicans when combined with miconazole, completely inhibiting growth after only 4 hrs of incubation. Analysis of ergosterol content from Candida albicans showed a time-dependent decrease to 91 % and 63 % at 16 and 24 hrs respectively, in cells treated with ½ MIC of 5-hydroxy-7,4’-dimethoxyflavone. The compound 5-hydroxy-7,4’-dimethoxyflavone also showed inhibition of both the drug efflux pumps (with IC50 = 51.64 μg/ml) and the antioxidant enzymes (at 5 μM).ConclusionThe compound 5-hydroxy-7,4’-dimethoxyflavone may be partly responsible for the reported antifungal activity of C. zeyheri, and may serve as a potential source of lead compounds that can be developed as antifungal phytomedicines.

Antitumor Constituents of the Wetland Plant Nymphoides peltata: A Case Study for the Potential Utilization of Constructed Wetland Plant Resources

The efficient utilization of plant resources is a necessary and important measure for sustainable management of constructed wetlands. Screening bioactive metabolites from wetland plants could reveal potential solutions for the utilization of constructed wetland plant resources. In this study, the constructed wetland macrophyte Nymphoides peltata was screened for constituents with antitumor activity. The secondary metabolites of N. peltata were extracted and separated by MCI gel, silica gel, and Sephadex gel column chromatography. Antitumor tests were then carried out with MTT assay against the human prostate cancer cell PC3 and the human osteosarcoma cell U2OS. The secondary metabolite group with the most significant antitumor activity was further examined, and four constituents were obtained and identified. This study provides a scientific basis for the potential efficient utilization of N. peltata and other constructed wetland plant resources.

Sea Buckthorn (Hippophae rhamnoides) Proanthocyanidins Inhibit In Vitro Enzymatic Hydrolysis of Protein

Interactions of phenolics with other food constituents and digestive enzymes are likely to have interference with the digestion and bioavailability of food and phenolics. In this study the effect of sea buckthorn proanthocyanidins on in vitro digestion of protein was evaluated. Optimization of the extraction conditions showed that maximum recovery of sea buckthorn proanthocyanidins was obtained with acidified acetone; water mixture (60% to 70%, v/v). Crude proanthocyanidin extracts thus prepared were purified using sephadex gel column chromatography and their average degree of polymerization and the effects on enzymatic hydrolysis of bovine serum albumin as influenced by their protein precipitation capacities were studied. Average degree of polymerization of proanthocyanidins in berry pulp, kernel, seed coat, and leaves was 7.4, 5.6, 8.2, and 10.6, respectively. The EC50 values for the protein precipitation by the PA of berry pulp, kernel seed coat, and leaves were 44.2, 44.1, 65.8, and 39.8 μg, respectively. Relative enzymatic hydrolysis of the protein-proanthocyanidin complexes was 44.1% to 60.3% for pepsin and 57.5% to 67.7% for trypsin. Interactions of sea buckthorn proanthocyanidins with food proteins and digestive enzymes might alter the protein digestibility and phenolic bioavailabilty.

Persistent Splenic Visualization on Technetium-99m DISIDA Hepatobiliary Studies

Tc-99m DISIDA is widely used for studying a variety of biliary conditions. It is generally recognized that the spleen is normally visualized on DISIDA imaging only during the blood pool phase. Upon review of our experience with DISIDA, visualization of the spleen was found more common than generally acknowledged. All DISIDA examinations performed at our two institutions were retrospectively reviewed; 95 studies were performed at Institution #1 and 150 studies at Institution #2. There were 138 men and 107 women. Quality control to confirm radiochemical purity included sephadex gel column chromatography. Of 245 patient studies, persistent splenic visualization was identified in one patient (1.0%) at Institution #1 and in 11 patients (7.3%) at Institution #2. There was no correlation between age, sex or final diagnosis and persistent visualization of the spleen. It is suggested that persistent splenic visualization may be caused by an elevated concentration of reduced hydrolized technetium (colloid) in DISIDA.

Characterization of an intracellular insulin-degrading enzyme in human erythrocytes

Using conventional techniques of ammonium sulfate fractionation and Sephadex gel column chromatography, insulin-degrading enzyme was partially purified from lysate of human erythrocytes. The enzymatic activity was measured by the trichloroacetic acid precipitation method. Compared to trypsin, the enzyme was highly specific for insulin. The apparent molecular weight of the enzyme was 160,000 Da, the optimum pH was the 7.4 to 7.8 range, and the K m value for insulin for the partially purified enzyme was 162 n m. Bacitracin and N-ethylmaleimide were potent inhibitors, while chloroquine, ethylenediaminetetraacetate, antipain, and soybean trypsin inhibitor failed to inhibit the activity of the enzyme. Like most nucleated cells, human erythrocytes not only have the membranal insulin receptors, but also possess the cytosolic specific insulin-degrading enzyme. Insulin internalization and degradation are shown to be due to the receptor and the enzyme acting in concert as in many nucleated cells. Anucleated erythrocytes have both these entities for possible internalization and degradation of insulin.

The plasma protein binding of HIDA

By using Sephadex gel column chromatography to separate substances into their various components according to molecular weight, we have investigated the effect of incubating several "brands" of HIDA in plasma, in vitro. The results show that such incubation has no effect on either dimethyl HIDA, or diethyl HIDA, but that in the case of para-butyl HIDA, incubation in plasma increases the Rf value to that of HSA (human serum albumin). This indicates that para-butyl HIDA becomes bound to plasma proteins, in contrast to both dimethyl HIDA and diethyl HIDA.

Immunohistochemical localization of aldose reductase. I. Enzyme purification and antibody preparation--localization in peripheral nerve, artery, and testis.

Aldose reductase (AR) was purified from rat and bovine seminal vesicles using DEAE-cellulose, hydroxylapatite, and Sephadex-gel column chromatography. The purification resulted in the obtention of an AR pool and a contaminating pool. Antibodies were raised in rabbits against both enzymes by subcutaneous injection of the AR pool. The antisera was judged to be specific for AR by immunoprecipitation of AR activity and by Ouchterlony double immunodiffusion and immunoelectrophoretic methods. Antibodies against rat AR were used in the unlabeled antibody-enzyme (PAP) technique to demonstrate the cellular location of the enzyme in a number of tissues known to be sites of diabetic lesions. Antibodies against bovine AR were not cross reactive with the rat enzyme, as determined by Ouchterlony and competitive protein-binding studies. AR was localized in rat tissues to the Schwann cell sheath of peripheral nerve, arterial endothelium, and the sustentacular (Sertoli) cells and mature spermatids of the testis.

Étude Qualitative et Quantitative des Inhibiteurs de Croissance Présents dans les Exsudats de Racines de Chénopode Blanc (Chenopodium album L.) au Début de sa Floraison en Culture Hydroponique et sous Conditions Contrôlées

The exudation of Chenopodium album L. in distilled water was studied under the controlled conditions of a growth room. The exudates of plants beginning to flower show a growth inhibitory effect, measured by the elongation of the main root of maize developing seeds. The inhibitory effect increases with the time of exudation (from 2 to 6 days) and with the exudate concentration of Chenopodium album L. (from 2,5 to 20 g fresh matter in 10 ml water extract). Differences in the degree of inhibition are observed according to the methods used to obtain exudates: at the flowering stage, the exudates of common lambsquarters collected in fields or grown in pots are more inhibitory than those of common lambs quarters grown on hydroponic culture. These differences may be explained by the results obtained after the purification of exudates: Sephadex gel column chromatography enabled us to separate two inhibitory fractions in the exudate of common lambsquarters grown on soil and only one in the exudate of common lambs quarters grown on nutrient solution. This last inhibitory fraction is common to both exudates, whatever the culture medium tested may be; it contains oxalic acid identified by 13C-NMR spectrometry. Oxalic acid is found in exudates of common lambsquarters at concentrations high enough (2,5 to 5 10-3 M/l) to be responsible for the growth inhibitory effect observed.

Séparation et Identification des Bétalaines Synthétisées par les Tissus de Tige de Myrtillocactus geometrizans Cultivés «in vitro»

Summary Sephadex gel column chromatography enabled us to separate betalains present in stem tissue cultures as well as in fruits of Myrtillocactus geometrizans. The identification of the isolated fractions was attempted by means of paper chromatography, electrophoresis, spectrum analysis and Sephadex gel chromatography. Three pigments — one betaxanthin and two betacyanins -, have been identified in both materials. Indicaxanthin and betanin were characterized with relative certainty. Another betaxanhin was tentatively identified as phyllocactin.

Séparation des pigments bétalaïques par une méthode fondée sur la chromatographie sur gel de Séphadex appliquée à des plantules d' Amarantus caudatus L. var. pendula

A simple method based on Sephadex gel column chromatography and requiring only small quantities of plant material, enabled not only the separation of betacyanins from betaxanthins but also the isolation of several different species of each of these types of pigment. Amarantus caudatus L. var. pendula seedlings, cultivated on 3,4-dihydroxyphenylalanine solutions in light conditions, were selected as the experimental material. The identification of the components of the isolated fractions was attempted by means of paper chromatography and electrophoresis. Two betacyanins, amaranthin and betanin, could be characterized with relative certainty. Moreover, two betaxanthins, present in small quantities in the extracts, were tentatively identified as being miraxanthin and vulgaxanthin.