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Assessment of bilirubin clearance capacity of a newly developed ion-exchange adsorption column and its possible use as a supportive therapy in hepatorenal syndrome.

We assessed the bilirubin reduction capacity of three different types of devices in vitro: a high-permeable membrane column for double-filtration plasmapheresis (DFP) (Evaflux 2A, Kuraray, Japan), and non-coated charcoal column for hemoperfusion (HP) (N-180, Asahi Medical, Japan), and ion-exchange columns for plasma adsorption (PA) (BR-350, Asahi Medical, Japan, and B-001, Kuraray, Japan). A column for DFP reduced the concentration of low-molecular proteins effectively such as plasma bilirubin and bile acids in an albumin-dependent manner. A charcoal column adsorbed low-molecular substances preferentially. But in these two columns, the loss of fibrinogen is a limiting factor for determining the processing plasma volume. Ion-exchange columns for PA adsorbed bile acids, disconjugated bilirubin, and monoconjugated bilirubin more efficiently compared with delta-bilirubin and unconjugated bilirubin. Pretreatment of the column with heparin reduced the loss of fibrinogen to less than 10%. We applied the BR-350 ion-exchange column in vivo for treatment of three patients with hyperbilirubinemia. After treatment, an alcoholic hepatitis patient with the hepatorenal syndrome (HRS) recovered from acute renal failure. However, in a patient with primary biliary cirrhosis and in a patient with fulminant hepatitis, the decrease of serum bilirubin was transient and no obvious beneficial responses were noted. The capacity and ability of the BR-350 column to adsorb plasma bilirubin was shown sufficient to treat deeply jaundiced patients, because 4 liters of the plasma of a patient with 108 mg/dl of initial total bilirubin concentration was able to be processed continuously without an obvious decrease in bilirubin adsorption capacity.(ABSTRACT TRUNCATED AT 250 WORDS)

Adsorption↗

Purification of recombinant adeno-associated virus type 8 vectors by ion exchange chromatography generates clinical grade vector stock.

Recombinant vectors based on the recently isolated AAV serotype 8 (rAAV-8) shows great promise for gene therapy, particularly for disorders affecting the liver. Transition of this vector system to the clinic, however, is limited by the lack of an efficient scaleable purification method. In this report, we describe a simple method for purification of rAAV-8 vector particles based on ion exchange chromatography that generates vector stocks with greater than 90% purity. The average yield of purified rAAV-8 from five different vector preparation was 41%. Electron microscopy of these purified stocks revealed typical icosohedral virions with less than 10% empty particles. Liver targeted delivery of ion-exchange purified rAAV-8 vector encoding the human factor IX (hFIX) gene, resulted in plasma hFIX levels approaching 30% of normal in immunocompetent mice, which is 20-fold higher than observed with an equivalent number of rAAV-5 ion exchange purified vector particles. The method takes less then 5 h to process and purify rAAV-8 vector from producer cells and represents a significant advance on the CsCl density centrifugation technique in current use for purification of rAAV-8 vector systems and will likely facilitate the transition of the rAAV-8 vector system to the clinic.

Animals↗

Elevated HbA1 due to foetal haemoglobinaemia in children. A common artifact of certain ion-exchange chromatography methods.

Foetal haemoglobin (HbF) is known to elute with glycosylated haemoglobins (HbA1) in mini-column ion-exchange chromatography, thus producing falsely elevated values for HbA1 in subjects with foetal haemoglobinaemia. We studied the occurrence of this methodological artifact in 266 patients with suspected or known insulin-dependent diabetes mellitus (IDDM) and in 205 healthy children of various age groups. HbA1 was measured using ion-exchange chromatography with prefabricated mini-columns and HbA1c and HbF by high-performance liquid chromatography (HPLC). In healthy children of over 5 years of age, the HbA1 values were usually within the reference limits for adults. However, 69% of the children younger than 1 year of age and 7% of those 1-5 years of age had HbA1 levels above 10%. Their HbA1c values were normal but the HbF values were high. Six of the 223 patients with IDDM (2.7%) and three of the 43 patients with stress glucosuria (7%) had much higher HbA1 than HbA1c values. The discrepancy was again related to the presence of raised concentrations of HbF. We conclude that foetal haemoglobinaemia is more prevalent than was previously known, especially among children younger than 5 years of age. Presence of HbF will bias the use of HbA1 as a diagnostic aid or as an index of metabolic control in children with IDDM. Methods specific for HbA1c, such as HPLC, should be used in paediatric patients.

Adolescent↗

Fractionation of low-molecular-mass heparin by centrifugal partition chromatography in the ion-exchange displacement mode.

Centrifugal partition chromatography in the ion-exchange displacement mode allowed a preparative and efficient fractionation of low-molecular-mass heparins from enoxaparin sodium. Amberlite LA2--a lipophilic liquid secondary amine--was chosen as a weak anion exchanger. The biphasic system methyl isobutyl ketone-water was selected. Protonated LA2 (10%, v/v) was added to the organic stationary phase. Hydroxide (Na+, OH-) was chosen as a displacer in the aqueous mobile phase. The observed pH and concentration profiles are typical of displacement chromatography, as supported by numerical simulation. The Dubois test for the analysis of sugar content and an analysis of sulfur content (and consequently sulfatation rate) were carried out to monitor the effectiveness of the procedure. Moreover, the fractions were analyzed by high-performance size-exclusion chromatography and the 1H NMR spectra confirmed the fractionation of the sample of enoxaparin sodium.

Chromatography, Gel↗

Preparation of an ion-exchange chromatographic support by a "grafting from" strategy based on atom transfer radical polymerization.

A new "grafting from" strategy based on surface-initiated atom transfer radical polymerization (ATRP) was first used for the preparation of a polymer-based ion-exchange support for HPLC. The most important property of the proposed method is to be applicable for the synthesis of any type of ion exchanger in both the strong and the weak forms. Monodisperse, porous poly(glycidyl methacrylate-co-ethylene dimethacrylate), poly(GMA-co-EDM) particles 5.8 mum in size were synthesized by "modified seeded polymerization". Poly(dihydroxypropyl methacrylate-co-ethylene dimethacrylate), poly(DHPM-co-EDM) particles were then obtained by the acidic hydrolysis of poly(GMA-co-EDM) particles. The ATRP initiator, 3-(2-bromoisobutyramido)propyl(triethoxy)silane was covalently attached onto poly(DHPM-co-EDM) particles via the reaction between triethoxysilane and diol groups. In the next stage, the selected monomer carrying strong cation exchanger groups, 3-sulfopropyl methacrylate (SPM), was polymerized on the initiator-immobilized particles via surface-initiated ATRP. The degree of polymerization of SPM (i.e., length of polyionic ligand) on the particles was precisely controlled by adjusting ATRP conditions. Poly(SPM)-grafted poly(DHPM-co-EDM) particles obtained with different ATRP formulations were tried as chromatographic packing in the separation of proteins by ion-exchange chromatography. The proteins were successfully separated with higher column yields with respect to the previously proposed materials. The plate heights between 100 and 150 mum were achieved with the column packed with the particles carrying the shortest poly(SPM) chains. The plate height showed no significant increase with increasing flow rate in the range of 0.5-16 cm/min.

Journal Article↗

Ion exchange extraction of heavy metals from wastewater sludges.

Heavy metals are common contaminants of some industrial wastewater. They find their way to municipal wastewaters due to industrial discharges into the sewerage system or through household chemicals. The most common heavy metals found in wastewaters are lead, copper, nickel, cadmium, zinc, mercury, arsenic, and chromium. Such metals are toxic and pose serious threats to the environment and public health. In recent years, the ion exchange process has been increasingly used for the removal of heavy metals or the recovery of precious metals. It is a versatile separation process with the potential for broad applications in the water and wastewater treatment field. This article summarizes the results obtained from a laboratory study on the removal of heavy metals from municipal wastewater sludges obtained from Ardhiya plant in Kuwait. Data on heavy metal content of the wastewater and sludge samples collected from the plant are presented. The results obtained from laboratory experiments using a commercially available ion exchange resin to remove heavy metals from sludge were discussed. A technique was developed to solubilize such heavy metals from the sludge for subsequent treatment by the ion exchange process. The results showed high efficiency of extraction, almost 99.9%, of heavy metals in the concentration range bound in wastewater effluents and sludges. Selective removal of heavy metals from a contaminated wastewater/sludge combines the benefits of being economically prudent and providing the possibility of reuse/recycle of the treated wastewater effluents and sludges.

Industrial Waste↗

Factorial experimental design for recovering heavy metals from sludge with ion-exchange resin.

Wastewaters containing heavy metals are usually treated by chemical precipitation method in Taiwan. This method can remove heavy metals form wastewaters efficiently, but the resultant heavy metal sludge is classified as hazardous solid waste and becomes another environmental problem. If we can remove heavy metals from sludge, it becomes non-hazardous waste and the treatment cost can be greatly reduced. This study aims at using ion-exchange resin to remove heavy metals such as copper, zinc, cadmium, and chromium from sludge generated by a PCB manufacturing plant. Factorial experimental design methodology was used to study the heavy metal removal efficiency. The total metal concentrations in the sludge, resin, and solution phases were measured respectively after 30 min reaction with varying leaching agents (citric acid and nitric acid); ion-exchange resins (Amberlite IRC-718 and IR-120), and temperatures (50 and 70 degrees C). The experimental results and statistical analysis show that a stronger leaching acid and a higher temperature both favor lower heavy metal residues in the sludge. Two-factors and even three-factor interaction effects on the heavy metal sorption in the resin phase are not negligible. The ion-exchange resin plays an important role in the sludge extraction or metal recovery. Empirical regression models were also obtained and used to predict the heavy metal profiles with satisfactory results.

Analysis of Variance↗

[Sorption of acid ribonuclease from Penicillium brevi-compactum by various ion-exchange resins].

Sorption of acid ribonuclease from Penicillium brevi-compactum F-399 was studied. The effect of nature and structure of various ion-exchange resins on the sorption capacity and reversibility was determined. High rate and reversibility of the enzyme sorption on the strong base anion-exchange fiber TsM-A2 were demonstrated. It is suggested that sorption of acid ribonuclease of the TsM-A2 fiber proceeds as ion exchange and on carboxyl cation exchange resins as molecular sorption.

Chromatography, Ion Exchange↗

High-performance ion-exchange chromatography and adsorption of plasma proteins.

Resolution and recovery are primary concerns in protein chromatography. Separations are often by size, ion exchange, or hydrophobic-hydrophilic properties of the support, eluent and protein. Adsorption of a protein to a synthetic surface plays an essential role in this complex process. In this study, we examined the adsorption properties of three representative plasma proteins (albumin, fibrinogen, and immunoglobulin G) on nonporous column materials containing either quaternary amine or sulfopropyl functional groups. The adsorption properties were studied at 37 degrees C and pH 7.4. Salt gradients were used to examine the adsorption/desorption properties of each of the proteins on each type of surface. The salt concentrations at desorption were measured and compared to the protein isoelectric points. In addition, we examined protein recoveries as a function of desorption time. Our results suggest that protein recoveries depend not only on the protein, eluent and surface, but also the residence time and overall charge concentration during the initial adsorption process. Finally, we correlated the number of charge sites on a molecule with the width of a chromatographic band at half height. The data produced as a result of this study may be used to determine the actual unfolding time of a protein, given a certain set of conditions. The data may also help in understanding the chemistry and dynamics of the protein adsorption processes in ion-exchange chromatography as well as provide key structural information about the proteins.

Adsorption↗

Selective extraction of benzoic acid from landfill leachate by solid-phase extraction and ion-exchange chromatography.

In this work a simple method was described for selective extraction of benzoic acid from landfill leachate samples. The samples were submitted to solid-phase extraction (SPE) with XAD-4 resin as the stationary phase and ion-exchange chromatography (IEC) using the ion-exchange resin Amberlyst A-27. The instrumental analysis was performed by gas chromatography with mass spectrometric detection (GC-MSD). Benzoic acid was isolated, identified and quantified. The extraction process is rapid, simple and of low cost. It was also environmental friendly, that is, it was used a minimum amounts of hazardous organic solvents and produced also minimum quantities of residues.

Benzoic Acid↗

[Ion exchange interactions on silica gel layers. III. Salts of organic bases with organic acids. Organic acids and sodium salts].

By thin-layer chromatographic and spectroscopic (UV, IR) methods it has been proved that bass and acid are formed from salts of organic bases with organic acids during primary interaction because of the ion exchange behaviour of the silica gel layer. The organic acids formed have interacted with metals on the layer during secondary ion exchange and they migrate further as salts again. It has also been established that the extent of secondary ion exchange between metal silanate groups and acids depends on pKS values of acids. On the basis of investigations it can be ascertained with great probability that organic acids of pKS greater than 7 do not interact with metal silanate groups of silica gel layer. Sodium salts of organic acids of pKS less than 3 values similarly do not interact on the other hand sodium salts of organic acids of pKS greater than 3 values are able to interact with silanol groups. It has been established by flame atomic emission method that salicylic acid interacts with sodium ion and 86% of it migrates further as sodium salicylate.

Carboxylic Acids↗

A new strategy for exploiting ion exchange in sequential injection analysis: in-line phytic acid separation/determination in foods as an example.

A novel strategy for exploiting ion exchange in sequential injection systems is proposed. The procedure is based on the selection of a defined volume of a resin suspension, which is introduced and packed in the analytical path, establishing a resin mini-column in the system. The passage of a selected sample volume through the resin mini-column leads to the retention of the analyte, while the sample matrix is discarded. The analyte is eluted during the passage of the eluant/reagent by the packed beads, being the analytical signal monitored (absorbance) in the liquid phase. The beads are then aspirated back to the holding coil and directed to a recovery flask, linked at the selection valve; then the system is ready to begin a new cycle. With the proposed strategy, the main characteristics of the sequential injection system are kept as any new artifact is added to the manifold and system reconfiguration is not required. The feasibility of the approach is demonstrated by the phytic acid determination in food samples. For this specific application, AG1-X8 was selected as ion exchanger, and a solution containing Cl- and Fe(III)-salicylate complex was used as eluant and spectrophotometric reagent.

Anion Exchange Resins↗

Rat small-intestinal beta-galactosidases. Studies on the fractionation of "acid" beta-galactosidase with isoelectric focusing, gel filtration and ion-exchange chromatography.

1. Different forms of the rat small-intestinal ;acid' beta-galactosidase were separated by using the isoelectric-focusing technique. The isoelectric points of the different forms were at pH4.2, 4.6, 5.4, 6.1 and approx. 8. 2. The two forms of ;acid' beta-galactosidase isoelectric at pH4.2 and 4.6 were completely excluded from the Sephadex G-200 gel, whereas the form isoelectric at pH8 had K(av.) 0.4. The concentration and pH of the elution buffer influenced the distribution of enzyme activity between different forms. Thus, under certain conditions of ionic strength and pH, the enzyme seems to form high-molecular-weight aggregates with low isoelectric points. These may be homopolymeric aggregates or the result of binding of enzyme to, for example, membrane fragments. The forms isoelectric at pH5.4 and 6.1 are probably aggregates of intermediate size. 3. During ion-exchange chromatography at pH6.0 one fraction of ;acid' beta-galactosidase was not retained on the column and was isoelectric at pH8 and another fraction was eluted when the buffer concentration in the eluate had increased to about 50mm. The main part of enzyme eluted in this second fraction was also isoelectric at pH8, indicating that the elution of this fraction is not a simple ion-exchange procedure but probably also involves a splitting of high-molecular-weight aggregates, originally retained because of their low isoelectric points. The enzyme subunits have a higher isoelectric point, and are therefore no longer bound to the ion-exchange resin.

Animals↗

Influence of Anodic Surface Treatment of Activated Carbon on Adsorption and Ion Exchange Properties.

The effect of anodic surface treatment of activated carbon on adsorption and ion exchange characteristics was investigated in the condition of 35 wt% NaOH electrolyte for 60 s. The acid and base values were determined by a titration technique, and surface and pore structures were studied in terms of BET volumetric measurement with N(2) adsorption. The ion exchange capacity of the anodized activated carbons was characterized by a dry weight capacity technique. It was observed that an increase in current intensity leads to an increase in the surface functional groups of activated carbons, resulting in increasing pH, acid-base values, and anion-cation exchange capacities, without significant change of surface and pore structures (i.e., specific surface area, total pore volume, micropore volume, and average pore diameter). Also, anodically treated activated carbons are more effectively evaluated on the base value or cation exchange capacity than on the opposite properties in this electrolytic system. Copyright 1999 Academic Press.

Journal Article↗

Liquid ion exchangers in paper chromatography of steroidal glucosiduronic acids, glucosiduronic esters and free steroids. Influence of concentration of exchanger and counterion.

The chromatographic mobility of steroidal glucosiduronic acids on paper in chloroform-formamide increases as the concentration of ion exchanger in the chloroform phase increases; mobility decreases as the concentration of counterion in formamide increases. Mobility of glucosiduronic esters and of hydroxylated free steroids increases with an increase in concentration of exchanger; small changes in concentration of counterion in the stationary phase do not alter the migration of these nonionizable compounds. Data are presented which suggest that partition of the glucosiduronic acids between the two phases occurs predominantly by an ion-exchange process and that hydrogen bonding plays a secondary role. Partition of the glucosiduronic esters and hydroxylated free steroids appears to occur primarily by a hydrogen-bonding process.

Cholesterol↗