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The stability of four major arsenic (As) species during application of the BARGE (Bioaccessibility Research Group of Europe) unified bioaccessibility method (UBM) has been assessed. The concentrations of As species in the UBM gastric and gastro-intestinal (gastric + intestinal) phases were determined using HPLC-ICP-MS whilst the total As content in the samples was determined using ICP-MS alone. The arsenic species studied were arsenite As(III), arsenate As(V), dimethylarsinic acid (DMA) and monomethylarsonic acid (MMA). These species were separated in 10 min using an anion exchange column (Hamilton PRP-X100) with a mobile phase containing 20 mmol L−1 NH4H2PO4/1% methanol (pH 6.0). The recoveries of arsenic species spiked into the gastric and gastro-intestinal fluids were in the range 90–108%. No interconversion between As species was observed as a result of applying the BARGE UBM, which is a particularly important finding for the reliability of As(III) measurements. The accuracy of the BARGE UBM for in vitro extractable As(V) was verified using British Geological Survey (BGS) guidance material 102 (an ironstone soil). For a commercial rice sample, the bioaccessibility sequence of As was DMA > As(III) > As(V) for the gastric phase and As(III) > DMA > As(V) for the gastro-intestinal phase.

Objective Management and diagnosis of multiple human cancers remains a challenge and search for a common biomarker is still debatable. In this manuscript we have evaluated the use of monoclonal antibody UNIVmAb, to detect the protein (H11) as a common biomarker for all cancers irrespective of the grade and origin. We have shown by both ELISA and Western Blot that the H11 protein, is a unique hyaluronan binding protein that has not been detected earlier. H11 protein was fractionated in an anion exchange column followed by cibacron blue gel exclusion chromatography. Hyaluronan binding H11 protein reacted with Monoclonal antibody UNIVmAb and b-HA inspite of b-Hyaluronan (biotinylated Hyaluronan) interaction and HA-Oligo (Hyaluronan oligosaccharides) competition from various grades of Human cancers sera. Results ELISA, Western blot and b-Hyaluronan interactions clearly showed an over-expression of UNIVmAb reacted H11 protein in all fifty cancer’s sera when compared with seventy normal sera. UNIVmAb reactive H11 protein can be used as a common biomarker. We believe, UNIVmAb detected H11 protein, is a unique hyaluronan binding protein, that can be used as a common biomarker for all cancers.

A bacterial strain was isolated from Pasinler hot spring, Erzurum, Turkey. The purified thermophilic isolate was identified as Geobacillus pallidus P26 and used to produce extracellular pectin lyase (EC 4.2.2.10). Pectin lyase enzyme was purified 34 fold by using DEAE-cellulose anion exchange column chromatography and characterized. Molecular weight of the enzyme was determined as 56 kDa by using Sephadex G-100 gel filtration chromatography. Purification of enzyme was verified by SDS-PAGE. The pH- and temperature optima of enzyme were determined (pH 9.0 and 60°C, respectively). Pectin lyase was mostly stable at 50 oC for 24 hours. Its’ activity decreased to 50 % for 24 h at 60°C. KM and Vmax were calculated as 24.8 mg/mL and 2.28 μmol/L min, respectively. Purified pectin lyase was inhibited by Fe3+, Zn2+, Cu2+, Ca2+, Co2+ and Hg2+ but not by Mg2+. The purified pectin lyase enzyme was used for getting fruits juices. It was found that yields of fruits juices increased when they were compared with control.

Samples of the vitreous were analysed in order to identify changes of soluble proteins in vitreo-retinal disease. The soluble proteins of the vitreous were separated on an anion exchange column (Mono-Q). The degree of neutral proteolytic activity in vitreous body was also measured. The vitreous from cataract cases without vitreoretinal disease was characterized by its low content of soluble proteins equivalent to about 1% of that of serum. Albumin and transferrin were the major identified components and their concentrations were approximately 0.85 and 0.03 g/l, respectively. In cases with vitreoretinal disease the vitreous showed changes of total soluble protein and the appearance of additional protein peaks. In patients with PVR the albumin concentration in the vitreous was found to be three times higher as compared to the control group consisting of patients with cataract. Neutral proteolytic activity in the vitreous was relatively low in both normal and pathological vitreous.

Background: The small amount of allantoin present in human serum results from free radical (FR) action on urate and may provide a stable marker of free radical activity in vivo. We describe a gas chromatography–mass spectrometry (GC–MS) assay for serum allantoin and report a reference range in healthy individuals. Methods: Fasting blood samples were obtained from 134 healthy middle-aged volunteers (56 men, mean age 55, range 45–72; 78 women, mean age 55, range 50–72) Allantoin was assayed using 15N2 allantoin as an internal standard. After isolation from aqueous standards or serum by extraction onto an anion exchange column (AG-MP1), allantoin was derivatised with N-methyl-N-(tert-butyldimethylsilyl) trifluoroacetamide (MTBSTFA). Derivatives were injected onto an HP-1 column and analysed using a Mass Selective Detector with Single Ion Monitoring at 398 and 400 m/z. Results: The distribution of serum allantoin concentrations in men and women was non-Gaussian and log transformation was used for the analysis of data. Women (10.8±1.7 μmol/l (mean±S.D.)) had significantly lower serum allantoin levels than men (13.4±1.6 μmol/l, p=0.015). Reference ranges (95% CI) for middle-aged healthy subjects were 7.4–46.8 μmol/l (men) and 3.7–31.2 μmol/l (women). Conclusion: Gas chromatography–mass spectrometry provides a reliable and accurate method for the determination of serum allantoin.

Using a biochemical approach we identified families of ubiquitin-activating and ubiquitin-conjugating enzymes in Chiamydomonas reinhardtii. The family of ubiquitin-activating enzymes, characterized by their ability to form thioesters with ubiquitin and eluting off a ubiquitin affinity column by ATP-depletion probably consists of at least four members. Whereas one of these enzymes is active under a broad range of pH values, thioesterof the other UBAs with ubiquitin is restricted to pH 7.5. Two ubiquitin-activating enzymes are metabolically phosphory-lated which is assumed to be an activity control mechanism. Most of the 7 ubiquitin-conjugating enzymes detected in this study were found to bind rather tightly to an anion exchange column, and eluted off the column at specific salt concentra tions. Two of the ubiquitin-conjugating enzymes described here did, however, not bind to this column. These enzymes can, as all other C. reinhardtii ubiquitin-conjugating enzymes, perform thioester formation with ubiquitin regardless of the source (plant/animal) of the ubiquitin-activating enzyme.

Plant material contains carbohydrates and phenolic compounds that interfere with derivatization and HPLC analysis of polyamines. The use of strong anion-exchange resins was investigated for purification of polyamines in plant samples. Results obtained indicate that anion-exchange resins produce equally good results as cation-exchange resins. Polyamines are eluted from the anion-exchange resin with NaOH, and derivatization can be performed directly on the eluate, resulting in enormous time savings. Optimum results were achieved with low cross-linkage strong anion-exchange resins and the conditions for maximum recoveries of polyamines are reported.

The selectivity of a rapid (90)Sr bioassay technique over (241)Am, (238/239)Pu, (210)Po, (137)Cs and (60)Co has been investigated. Similar to (90)Sr, these radionuclides are likely to be used in radiological dispersive devices. The purpose of this study was to demonstrate the degree to which the (90)Sr bioassay technique is free from interference by these radionuclides if present in a urine matrix. The interfering radionuclides were removed (from (90)Sr) by their retention on an anion exchange column. While, recovery of the target radionuclide ((90)Sr) was found to be >or= 90 %, contributions from (241)Am, (242)Pu and (208)Po were found to be

The development of a moderate-capacity anion-exchange resin has resulted in new ion chromatography (IC) methods for the determination of trace anions is concentrated acids. Suppressed microbore IC offers a higher suppression capability, which allows higher-capacity analytical columns to be used. As a result, higher-capacity columns permit higher concentrations of acids to be injected into the column without overloading, thus improving trace anion detection limits. In addition, using a selective matrix elimination method, high concentrations (% levels) of weak acids ( e.g. hydrofluoric acid and acetic acids) may be eliminated prior to analysis by IC.

A method for the preparation of insulin derivatives which have protected sulfhydryl group(s) at definite site(s) on the molecule is described. Porcine insulin reacts with S-acetylmercaptosuccinic anhydride to afford four species of insulin derivatives that have 2 (or 3)-acetylmercapto-3-carboxypropanoyl group(s) at i) Gly(A1), ii) Gly(A1) and Phe(B1), iii) Gly(A1) and Lys(B29), and iv) Gly(A1), Phe(B1) and Lys(B29) positions. The derivatives are efficiently separated in a preparative scale by anion-exchange high-performance liquid chromatography on a TSKgel DEAE-2SW column. The four derivatives are all readily deacetylated with hydroxylamine to give the corresponding sulfhydryl insulin derivatives.

The analysis of blood Esterase D (ESD) phenotypes was performed by using high performance liquid chromatography (HPLC). The hemolysate was first treated with a cation exchanger to remove hemoglobin, injected into HPLC on a weak anion exchange column, and developed to become fluorescent with reaction reagent. The chromatogram of each ESD phenotype was distinct from those of the other phenotypes. ESD protein was confirmed by isoelectric focusing in the HPLC fraction that was referred for ESD typing. This HPLC method can be used to determine the common ESD phenotypes in blood.

A rapid and simple high performance liquid chromatographic analytical method is described for the simultaneous determination of norfloxacin and furprofen in rat plasma. Following dichloromethane extraction, the solution was chromatographed in a Vydac anion exchange column using a mobile phase of 0.05 M phosphate buffer (pH = 7.0): acetonitrile (80:20, v/v) at a flow-rate of 1.8 mL/min. The drugs were detected by UV absorption at 278 nm. The total chromatographic analysis time was 10 min. The response was linear, 0.1-5.0 micrograms/mL for norfloxacin and 0.1-3.0 micrograms/mL for furprofen, respectively. This method is useful for pharmacokinetic studies of these compounds and will facilitate detailed investigations into the interactions between quinolones and furprofen.

The separation of dilute protein mixtures was achieved using simple monovalent buffering species to form retained, internally produced pH gradients on a strong-basic anion-exchange column. Highly focused proteins bands localized on stepwise pH transitions were produced experimentally under trace and volume overloaded feed conditions. Numerical simulations were performed that accurately predict the pH profile and protein band shapes in the column effluent. Experimental results were combined with numerical investigations to explore strategies for designing efficient preparative-scale chromatofocusing systems using simple, inexpensive buffers and adsorbents.

A rapid, reproducible and sensitive high performance liquid chromatography (HPLC) method for the determination and purification of metallothionein-I (MT-I) and metallothionein-II (MT-II) in mouse and rabbit livers has been developed. Methallothioneins (MTs) were separated by an HPLC anion exchange column, eluted through a linear gradient of Tris buffer and the peak containing MTs was determined by atomic absorption spectrophotometry. Furthermore, the content of MT-I or MT-II was calculated by protein peak area in a short time (about 20 min). The sample to be tested was homogenized, centrifuged and saturated by cadmium. MT-I and MT-II were eluted at 15.9 and 19.3 min, respectively. The following mouse liver cytosols were tested: controls, Cd-injected samples and 60Co-irradiated samples. A detection limit of 5 micrograms/g liver was established for this method. We have analysed more than 100 biological samples and obtained satisfactory results.

A method of purifying Sarcocystis sporozoites following in vitro excystation is described. After passage of sporozoites through a modified DE 52 anion-exchange cellulose column, the mean recovery was 88% for S. capracanis, 71% for S. hircicanis, 77% for S. ovicanis and 74% for S. arieticanis. Most of the sporozoites were motile, and the parasite suspension was free of excystation debris.

The concentrations of D-glucose and D-galactose in plasma of galactosemic rats were quantitatively measured by a liquid chromatographic method, based on retention of the weakly ionized monosaccharides by an anion exchange column under alkaline conditions, elution with 9 mmol/L NaOH, and electrochemical detection. This method is both simple and sensitive, since very dilute plasma samples can be directly analysed.

A rapid analytical method for the determination of ochratoxin A (OCTA), without using chloroform or an immunoaffinity column, in coffee beans and cereals such as rice, wheat, barley and corn was developed. OCTA was extracted from spiked food samples with acetonitrile-1% H3PO4 (99:1) and cleaned up using cartridge anion exchange column (Bond Elut DEA) chromatography. The test solution was chromatographed on a Capcell Pak C8 column with acetonitrile-water-acetic acid (40:58:2). OCTA was detected using a fluorescence detector (excitation: 335nm; emission: 465nm). Recoveries from samples spiked with 10ng/g OCTA and 50ng/g in coffee beans, unpolished rice, wheat, barley and corn averaged 93.9±3.9% and 96.1±3.5%, respectively. The detection limits corresponded to 0.05ng/g in green coffee beans, roasted coffee beans, unpolished rice, wheat, barley and corn.

The separation of large, single-stranded oligodeoxyribonucleotides, prepared by chemical synthesis, was carried out on a DEAE-5PW anion-exchange column with eluents containing only volatile salts. It was found that gradient elution is essential and that a high resolution can be achieved at elevated temperatures ( ca. 50°C). Also, neutral pH of the eluents was found to be desirable for the separation. Under optimum conditions, a linear relationship between the elution volumes and the number of bases in the substrates was observed.

The miniature anion-exchange/centrifugation (AEC) technique, developed originally for the detection of low parasitaemias in laboratory rodents, was adapted to field use for the diagnosis of trypanosomiasis in man in Africa and was tested in The Gambia. During this field study it was found that microfilariae of Dipetalonema perstans could also pass through the anion exchange column and appear in the centrifugate as 'medusa heads'. One locality-group (Mansafa Bolon) showed a generally higher prevalence and prevalences in women over 40 years old were higher than in the corresponding male groups in every locality. The potential usefulness of this technique in epidemiological studies of filariasis and ways of improving the accuracy of numerical estimates are briefly discussed.

A method is reported for the determination of 4-hydroxy-3-methoxymandelic acid (HMMA) by HPLC with electrochemical detection. The preparation of sample prior to HPLC has been studied and an efficient and selective extraction procedure described. Bonded-phase silica extraction columns and a vacuum manifold were used for the simple and rapid processing of batches of urine samples. Combining a reverse-phase C18 and an anion exchange column ensures selective isolation of HMMA. This simplified greatly the subsequent chromatography. The method was combined into a simple scheme for the routine analysis of urine HMMA, catecholamines and 5-hydroxyindole-3-acetic acid. The HPLC was standardised such that a single mobile phase was used with minor modification for each of the assays.

This paper describes a method of estimating the nucleotides in red blood cells. The technique used is based on that of Cohn and Carter (1950), who have shown that the nucleotides, adenosinemonophosphate (A.M.P.), adenosinediphosphate (A.D.P.), and adenosinetriphosphate (A.T.P.) can be separated by an anion exchange column. Bartlett, Savage, Hughes, and Marlow (1953) have successfully used this method for estimations on 100 ml. of whole blood; however, this volume is inconveniently large, especially if serial estimates are to be carried out. The present method uses only 1 ml. of red cells. Quantitative measurements have been made using the characteristic ultra-violet light absorption of adenine. In order to reduce the quantity of other substances also absorbing ultra-violet light, preliminary precipitation of the nucleotides as barium salts is carried out before placing them on the column. After elution of the nucleotides in separate fractions, the ultra-violet light absorption at three wavelengths is measured and correction for impurities made by the method of Morton and Stubbs (1946). This paper contains a detailed account of the method and findings of the nucleotide content of fresh red cells from 12 normal subjects. In order to demonstrate the use of the method, the results obtained before and after incubating stored red cells with adenosine are described.

A simple and rapid method is described for the determination of 6-thiouric acid in the urine of patients receiving azathioprine (Imuran, Burroughs Wellcome). The technique makes use of anion exchange column chromatography to isolate the 6-thiouric acid which is then measured by its absorption at 356 nm.