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PubMed2001

Sensitive gas chromatographic determination of the cyclophosphamide metabolite 2-dechloroethylcyclophosphamide in human plasma.

Cyclophosphamide (CP) is one of the most frequently used anticancer agents. It is a prodrug requiring activation before exerting cytotoxicity. CP is deactivated to 2-dechloroethylcyclophosphamide (2-DCECP) with formation of an equimolar amount of chloroacetaldehyde. The aim of this study was to develop and validate a sensitive and simple assay for 2-DCECP in plasma of patients treated with CP. Sample pre-treatment consisted of solid-phase extraction of 500 microl of plasma over OASIS HLB (1 ml) cartridges with trofosfamide as internal standard. Separation and detection of underivatized 2-DCECP was performed with capillary gas chromatography with nitrogen/phosphorous selective detection. Extraction recovery of 2-DCECP exceeded 87%. No interference from endogenous compounds, other metabolites of CP and frequently coadministered drugs was detected. The assay was linear in the range of 5-5000 ng/ml in plasma. Accuracy, within-day and between-day precision were less than 11% for the complete concentration range. In plasma, 2-DCECP was stable for at least 1 month when kept at -70 degrees C. Analysis of samples from patients treated with CP demonstrated the applicability of the assay. In conclusion, a sensitive and simple assay for 2-DCECP in plasma, which meets the current requirements for bioanalytical assays, was developed.

Antineoplastic Agents, Alkylating↗
PubMed2004

Efficiency of different extraction solvent mixtures used in analyses of aflatoxins from a certified peanut meal reference material.

Four analytical methods with different extraction solvent mixtures were used to compare the efficiency of the extraction step in the analysis of aflatoxins from a reference material with a certified value of 206 +/- 13 microg aflatoxin B(1) kg(-1). The extraction mixtures used were chloroform-water (100 + 10), acetonitrile-water (60 + 40), acetone-water (85 + 15) and methanol-water (80 + 20). The analytical values determined from the extraction mixtures after correction for recovery were 211, 204, 163 and 120 microg kg(-1), respectively. The small amounts of aflatoxin B(2) in the reference material followed the same pattern. The selection of extraction solvent mixture is very important for achieving the true value and that correction for recovery does not always fully compensate for getting the correct analytical value.

Acetone↗
PubMed2003

Chemical forms and extractability of iron in sediments of three contrasting lakes of China and UK.

Iron is a limiting factor for the eutrophication of lakes, especially those lakes that are enriched with phosphorus. Extractability of iron in sediments of West Lake and Taihu Lake in China and Lower Lough Erne in Northern Ireland of UK was comparatively investigated on the basis of analysing chemical forms of iron using different extractants. It was shown that extractable iron in sediments of the lakes was greatly different using various extractants. Reactive iron or easily released iron such as "active" iron oxides, total free iron oxide and water-soluble iron was not high, only accounting for 0.01%-0.15% of total iron. The efficiency of the extractants for exchangeable iron was decreased in the sequence 0.1 mol/L HCl >> DTPA + TEA mixed solution > 1 mol/L NH4OAc > 0.5 mol/L MgCl2 = 0.5 mol/L CaCl2. It seems that the complexion of iron by organic matter was not strong because the concentration of organically bound iron was significantly lower than the concentration it was forecasted. Extractable iron is not entirely consistent with or dependent on total iron in lake sediments. To a certain extent, phosphate can inhibit the release of iron in sediments of the lakes. The selection of extractants is thus the first key step to evaluate bioavailability of iron in lake sediments.

Biological Availability↗
PubMed1996

Comparison of supercritical CHF3 and CO2 for extraction of sulfonamides from various food matrices.

Supercritical CHF3 and methanol-modified CHF3 were compared with supercritical CO2 and methanol-modified CO2 for the extraction of sulfonamides from a spiked sand sample. In addition, a fortified nonfat milk powder, fortified egg yolk, and fortified beef liver were studied. The results showed CHF3 has higher solvating power and selectivity for extraction of two of the three sulfonamides than CO2. Extraction efficiencies of sulfamethazine and sulfadimethoxine using pure CHF3 were more than 45% higher from spiked sand samples and over 200% higher from fortified beef liver samples than pure CO2.

Animals↗
PubMed2002

Subcritical water extraction to evaluate desorption behavior of organic pesticides in soil.

We evaluated the feasibility of extracting organic pesticides in soil using a hot-water percolation apparatus at 105 degrees C and 120 kPa pressure. Efficiency of the method was assessed by extracting six selected pesticides (acetochlor, atrazine, diazinon, carbendazim, imidacloprid, and isoproturon) from previously equilibrated soil at 13.6-65.8 mg/kg concentration range. Studies were performed on brown forest soil with clay alluviation (Luvisol). The method developed was compared to the traditional batch equilibrium method in terms of desorbed amount of pesticides from soil and extraction time. Pesticides in the liquid phase from the batch sorption experiment and in the effluent from the hot-water percolation were quantified by high-performance liquid chromatography with UV detection. The results of the percolation experiment are in close correlation with those of the conventional soil testing method. Desorbed quantities by hot-water percolation were 85% acetochlor, 62% atrazine, 65% carbendazim, 44% diazinon, 95% imidacloprid, and 84% isoproturon, whereas using batch equilibrium method 101, 66, 64, 37, 81, and 90% were desorbed, expressed as the percentage of the adsorbed amount of pesticide on soil following equilibration. The average time for hot-water extraction was 3.45 min, in contrast to the 16 h time consumption of the traditional batch method. The effect of temperature on stability of selected compounds was also evaluated using pesticide-spiked sand without soil. Recoveries of analytes ranged between 84.6 and 91.1% with reproducibility of 7.9-10.2%, except for diazinon, for which recovery was 59.4% with 14.4% relative standard deviation since decomposition occurred at elevated temperature. The percolation process has been described by a first-order kinetic equation. The parameters calculated from the equation provide an opportunity to estimate the amount of compound available for desorption, the rate of desorption processes in the studied soil-pesticide-water system, and modeling the leaching process to obtain additional information on the environmental behavior of the examined pesticide.

Atrazine↗
PubMed1993

Cell-specific expression of helix-loop-helix transcription factors encoded by the E2A gene.

The E2A gene encodes transcription factors of the helix-loop-helix family that are implicated in cell-specific gene expression as part of dimeric complexes that interact with E box enhancer elements. It has previously been shown that transcripts of the E2A gene can be detected in a wide range of cell types. We have now examined expression of the mouse E2A gene at the protein level using polyclonal antisera directed against distinct portions of the E2A protein to probe blots of cellular extracts. A 73 kDa protein was identified by this analysis: this protein is highly enriched in cell lines of B lymphoid origin as compared to pancreatic beta-cells and fibroblast cells. The detection of this protein selectively in extracts of lymphoid cells correlates with the presence of the E box-binding activity LEF1/BCF1 in these cells; this binding activity was previously shown to be efficiently recognized by antiserum directed against E2A gene products. Transfection of cells with full length E2A cDNA leads to appearance of protein co-migrating with the 73 kDa protein on SDS gel electrophoresis and co-migrating with LEF1/BCF1 on mobility shift analysis. Our results are consistent with the view that the DNA-binding activity LEF1/BCF1 is a homodimer of E2A proteins; the selective appearance of this putative cell-specific transcription factor in B lymphoid cells seems to be attributable, at least in part, to the elevated E2A protein concentrations in these cells.

Animals↗
PubMed2006

Mangifera indica L. extract (Vimang) inhibits 2-deoxyribose damage induced by Fe (III) plus ascorbate.

Vimang is an aqueous extract of selected species of Mangifera indica L, used in Cuba as a nutritional antioxidant supplement. Many in vitro and in vivo models of oxidative stress have been used to elucidate the antioxidant mechanisms of this extract. To further characterize the mechanism of Vimang action, its effect on the degradation of 2-deoxyribose induced by Fe (III)-EDTA plus ascorbate or plus hypoxanthine/xanthine oxidase was studied. Vimang was shown to be a potent inhibitor of 2-deoxyribose degradation mediated by Fe (III)-EDTA plus ascorbate or superoxide (O2-). The results revealed that Vimang, at concentrations higher than 50 microM mangiferin equivalent, was equally effective in preventing degradation of both 15 mM and 1.5 mM 2-deoxyribose. At a fixed Fe (III) concentration, increasing the concentration of ligands (either EDTA or citrate) caused a significant reduction in the protective effects of Vimang. When ascorbate was replaced by O2- (formed by hypoxanthine and xanthine oxidase) the protective efficiency of Vimang was also inversely related to EDTA concentration. The results strongly indicate that Vimang does not block 2-deoxyribose degradation by simply trapping *OH radicals. Rather, Vimang seems to act as an antioxidant by complexing iron ions, rendering them inactive or poorly active in the Fenton reaction.

Ascorbic Acid↗
PubMed1996

Ca2+ entry induced by calcium influx factor and its regulation by protein kinase C in rabbit neutrophils.

Extracellular application of acid extract from platelet-activating factor- or thapsigargin-treated rabbit neutrophils induced a rise of cytosolic free calcium concentration ([Ca2+]i) in neutrophils and adrenal chromaffin cells suspended in Ca(2+)-containing, but not in Ca(2+)-deficient, medium. The ability of the extract to selectively induce Ca2+ entry was also confirmed by the increase in 45Ca2+ uptake and failure to stimulate Ca2+ release in digitonin-permeabilized neutrophils. 12-O-tetradecanoylphorbol-13-acetate (TPA) inhibited the extract-induced [Ca2+]i rise in a staurosporine (ST)-sensitive fashion, neither of which had any effect on its production. SK&F 96365 and econazole also reduced extract-induced Ca2+ entry. These results suggest that a Ca2+ entry-inducible substrate (calcium influx factor) is extracted from Ca2+ store-depleted neutrophils, and that its action may be regulated by protein kinase C and certain pharmacological agents.

Adrenal Glands↗
PubMedDate not supplied

Staphylococcosis of turkeys. 4. Characterization of a bacteriocin produced by an interfering Staphylococcus.

A Staphylococcus epidermidis isolate designated strain 115, which is used as an interfering agent against staphylococcosis of turkeys, produces a bacteriocin that was partially purified and characterized in this study. This bacteriocin diffused through agar media, but it was not found in appreciable quantities in the supernatant fluid of broth cultures. Extraction of the bacterial cells with 7 M urea, 1% sodium dodecyl sulfate, or 1% Triton X-100 caused considerable amounts of the bacteriocin to go into solution. This substance was partially purified by selective chemical extraction and by gel filtration chromatography using a Sephacryl S-300 column. This bacteriocin had two active forms: an aggregate, and a small-molecular-weight form estimated by gel filtration chromatography to be less than 6500. Activity was not affected by heat, repeated freeze-thaw cycles, pH 2 and pH 10, or a variety of proteolytic enzymes, nucleases, a lipase, and lysozyme.

Animals↗
PubMed1978

Rapid radioimmunoassay of progesterone in unextracted bovine plasma.

A rapid radioimmunoassay for determining progesterone levels in unextracted bovine plasma has been developed. Two antisera have been tested, anti-deoxycorticosterone-21-hemisuccinate human serum albumin and antiprogesterone-11-hemisuccinate bovine serum albumin. The usefulness of an antiserum for rapid assays has been based on its agreement with assays of organic extracts or chromatographically purified extracts of selected bovine samples. To avoid large blank effects and non-specific interference, a plasma volume of 20 microliter that provided adequate sensitivity was not exceeded. All correlations between rapid assays, assays using organic extraction or assays after chromatographic purification were greater than 0.9 and showed regression slopes approximating unity. It was concluded that individual lots of antisera should be evaluated thoroughly before use in a rapid assay.

Animals↗
PubMed2003

Plasticizer-free polymer containing a covalently immobilized Ca2+-selective ionophore for potentiometric and optical sensors.

A derivative of a known Ca2+-selective ionophore, ETH 129, was synthesized to contain a polymerizable acrylic moiety (AU-1) and covalently grafted into a methyl methacrylate-co-decyl methacrylate polymer matrix. The polymer containing AU-1 was prepared via a simple one-step homogeneous polymerization method. It exhibited mechanical properties suitable for the fabrication of plasticizer-free ion-selective membrane electrodes and bulk optode films by solvent-casting and spin-coating techniques, respectively. The segmented sandwich membrane technique was utilized to assess the binding constant of free and covalently bound ionophores to calcium and to study their diffusion coefficients in the membrane phase. Diffusion was greatly diminished for the bound ionophore. This was confirmed in ion-selective electrode membranes containing no calcium ions in the inner solution, which should normally show apparent super-Nernstian response slopes in dilute calcium solutions. The response slope was Nernstian down to submicromolar concentration levels, indicating slow mass transport of calcium in the membrane. Optical-sensing films with the new copolymer matrix, unblended and blended with PVC-DOS, also confirmed that covalently bound ionophores are fully functional for maintaining selective ion extraction and binding properties of the sensing membrane.

Biosensing Techniques↗
PubMed1989

Geranyl pyrophosphate synthase: characterization of the enzyme and evidence that this chain-length specific prenyltransferase is associated with monoterpene biosynthesis in sage (Salvia officinalis).

Cell-free homogenates from sage (Salvia officinalis) leaves convert dimethylallyl pyrophosphate and isopentenyl pyrophosphate to a mixture of geranyl pyrophosphate, farnesyl pyrophosphate, and geranylgeranyl pyrophosphate, with farnesyl pyrophosphate predominating. These prenyltransferase activities were localized primarily in the soluble enzyme fraction, and separation of this preparation on Sephadex G-150 revealed the presence of a partially resolved, labile geranyl pyrophosphate synthase activity. The product of the condensation reaction between [1-14C]dimethylallyl pyrophosphate and [1-3H]isopentenyl pyrophosphate was verified as [14C,1-3H]geranyl pyrophosphate by TLC isolation, enzymatic hydrolysis to geraniol, degradative studies, and the preparation of the crystalline diphenylurethane. The cis-isomer, neryl pyrophosphate, was not a product of the enzymatic reaction. By employing a selective tissue extraction procedure, the geranyl pyrophosphate synthase activity was localized in the leaf epidermal glands, the site of monoterpene biosynthesis, suggesting that the role of this enzyme is to supply the C10 precursor for the production of monoterpenes. Glandular extracts enriched in geranyl pyrophosphate synthase were partially purified by a combination of hydrophobic interaction chromatography on phenyl-Sepharose and gel permeation chromatography on Sephadex G-150. Substrate and product specificity studies confirmed the selective synthesis of geranyl pyrophosphate by this enzyme, which was also characterized with respect to molecular weight, pH optimum, cation requirement, inhibitors, and kinetic parameters, and shown to resemble other prenyltransferases.

Cell-Free System↗
PubMed2005

Braces and orthoses for treating osteoarthritis of the knee.

BACKGROUND: Patients with osteoarthritis of the knee can be treated with a brace or orthosis (shoe insole). The main purpose of these aids is to reduce pain, improve physical function and, possibly, to slow disease progression. OBJECTIVES: To assess the effectiveness of a brace or orthosis in the treatment of osteoarthritis of the knee. SEARCH STRATEGY: We searched Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE and EMBASE (Current contents, Health STAR) up to October 2002. The reference lists of the publications in the identified trials were also screened. SELECTION CRITERIA: Extracted studies were included in the final analysis if they met the pre-defined inclusion criteria: 1) a randomised controlled clinical trial or a controlled clinical trial, 2) all patients had osteoarthritis of the knee, 3) the intervention in one of the studied groups was a brace or an orthosis. DATA COLLECTION AND ANALYSIS: Two reviewers independently selected the trials and assessed the methodological quality using the Delphi-list and one additional question about care programs. Three reviewers independently extracted the data on the intervention, type of outcome measures, follow-up, loss to follow-up, and results, using a pre-tested standardized form. Study authors were contacted for additional information. MAIN RESULTS: Four trials involving a total of 444 people were included in this review. One study investigated a knee brace and three studies examined different types of orthoses for medial compartment osteoarthritis of the knee. Two studies were of high methodological quality while the other two studies were low. Notably, the randomisation and the blinding procedures were either insufficient or not described. The follow-up period (six weeks to six months) was too short to demonstrate long-term results. Pooling was difficult primarily due to the heterogeneity of the data and the way the information was presented. The pain, stiffness and physical function (WOMAC and MACTAR) scores of a brace group showed greater improvement at six months compared with a neoprene sleeve group, which showed greater improvement compared with a control group. The numbers of days of non-steroidal anti-inflammatory drug (NSAID) intake decreased significantly (relative percentage difference 23.9%) compared with baseline in a group with laterally wedged insoles,and remained unchanged in the neutrally wedged group. Patient compliance with the laterally wedged insole was significantly better compared with the neutrally wedged insole. In one study, the Visual Analogue Pain (VAS) pain score was significantly decreased from baseline in a strapped insole group (RPD - 24%), but not in the traditional lateral wedge group, but this strapped insole showed more adverse effects (popliteal pain, low back pain, and foot sole pain) compared with the traditional lateral wedge insole. Pain during bed rest, after getting up, after getting up from seated position and walking distance was significantly improved in a subtalar strapped group compared with baseline, and no improvement was found in a sock type group. No studies were found that assessed the effectiveness of a brace or orthosis to treat lateral compartment osteoarthritis or general osteoarthritis of the knee, or that compared a knee brace with a wedge insole, or that compared a brace or orthosis with operative treatment. AUTHORS' CONCLUSIONS: Based on one brace study we conclude there is limited evidence that: a brace has additional beneficial effect (WOMAC, MACTAR, function tests) for knee osteoarthritis compared with medical treatment alone.(Silver) a sleeve has additional beneficial effect (WOMAC, function tests) for knee osteoarthritis compared with medical treatment alone.(Silver) a brace is more effective (WOMAC, function tests) than a neoprene sleeve.(Silver) Based on 3 orthoses studies, of which 2 were high quality, (n=2) we conclude there is limited evidence that: a laterally wedged insole decreases NSAID intake compared with a neutral insole. (Silver) patient compliance is better in the laterally wedged insole compared with a neutral insole. (Silver) a strapped insole has more adverse effects than a lateral wedge insole. (Silver).

Braces↗
PubMed2006

Carotenoids extraction from Japanese persimmon (Hachiya-kaki) peels by supercritical CO(2) with ethanol.

The extraction of carotenoids from Japanese persimmon peels by supercritical fluid extraction (SFE), of which the solvent was CO(2), was performed. In order to enhance the yield and selectivity of the extraction, some portion of ethanol (5 - 20 mol%) was added as an entrainer. The extraction temperature ranged from 313 to 353 K and the pressure was 30 MPa. The effect of temperature on the extraction yield of carotenoids was investigated at 10 mol% of the ethanol concentration in the extraction solvent, and a suitable temperature was found to be 333 K among the temperatures studied with respect to the carotenoid yield. With increasing the entrainer amount from 0 to 10 mol% at a constant temperature (333 K), the carotenoid yield in the extraction was improved, whereas the selectivity of the extracted carotenoids was drastically depressed. We also conducted qualitative and quantitative analyses for the carotenoid components in the extract by HPLC, and analyzed the extraction behavior of each individual carotenoid (alpha-carotene, beta-carotene, beta-cryptoxanthin, lycopene, lutein, and zeaxanthin). The selectivity of each carotenoid changed with the elapsed time and its time evolution was dependent on the carotenoid component, indicating that the location profile and the content can be important factors to understand the SFE behavior of each carotenoid in persimmon peels.

Carbon Dioxide↗
PubMed1998

Human colostrum contains IgA antibodies reactive to enteropathogenic Escherichia coli virulence-associated proteins: intimin, BfpA, EspA, and EspB.

BACKGROUND: In Brazil, enteropathogenic Escherichia coli diarrhoea is endemic among infants born into low economic levels, and it is one of the main causes of morbidity and mortality in this group. Binding of enteropathogenic E. coli to the brush border mucosa triggers a cascade of transmembrane and intracellular signals, causing cytoskeletal reorganization and formation of a specific lesion, termed the attaching and effacing lesion. Several enteropathogenic E. coli gene products have been implicated in formation of attaching and effacing lesions. Evaluation of pathogen-specific protective factors shows that breast feeding is effective against enteropathogenic E. coli infection. To investigate the nature of the protection, defatted colostrum and secretory immunoglobulin A obtained from mothers living in Sao Paulo were investigated for the ability to recognise selected enteropathogenic E. coli-associated virulence factors. METHODS: Western blot analysis was used to investigate the IgA repertoire in pooled colostrum that is reactive with specific enteropathogenic E. coli proteins. Whole enteropathogenic E. coli bacterial cell extracts, nonpathogenic E. coli strains overexpressing specific virulence factors, and purified polypeptides were used as antigen sources in this study. RESULTS: Reaction of the colostrum samples in Western blots of whole bacterial cell extracts and selected purified enteropathogenic E. coli proteins showed that they contained a secretory immunoglobulin A reactive with all the virulence-associated proteins studied. CONCLUSION: These results suggest that maternal antibodies may protect infants from enteropathogenic E. coli infection by interfering with adherence processes (anti-intimin and anti-bundle-forming pili antibodies) and cell signaling (anti-enteropathogenic Escherichia coli-secreted protein A and B antibodies.

Adhesins, Bacterial↗
PubMedDate not supplied

Chemical extraction methods to assess bioavailable arsenic in soil and solid media.

Soil ingestion by children is an important pathway in assessing public health risks associated with exposure to arsenic-contaminated soils. Soil chemical methods are available to extract various pools of soil arsenic, but their ability to measure bioavailable arsenic from soil ingestion is unknown. Arsenic extracted by five commonly used soil extractants was compared with bioavailable arsenic measured in vivo by immature swine (Sus scrofa) dosing trials. Fifteen contaminated soils that contained 233 to 17 500 mg kg(-1) arsenic were studied. Soil extractants were selected to dissolve surficially adsorbed and/or readily soluble arsenic (water, 1 M sodium acetate, 0.1 M Na2HPO4/0.1 M NaH2PO4) and arsenic in Fe and Mn oxide minerals (hydroxylamine hydrochloride, ammonium oxalate). The mean percent of total arsenic extracted was: ammonium oxalate (53.6%) > or = hydroxylamine hydrochloride (51.7%) > phosphate (10.5%), acetate (7.16%) > water (0.15%). The strongest relationship between arsenic determined by soil chemical extraction and in vivo bioavailable arsenic was found for hydroxylamine hydrochloride extractant (r = 0.88, significant at the 0.01 probability level). Comparison of the amount of arsenic extracted by soil methods with bioavailable arsenic showed the following trend: ammonium oxalate, hydroxylamine hydrochloride > in vivo > phosphate, acetate > water. The amount of arsenic dissolved in the stomach (potentially bioavailable) is between surficially adsorbed (extracted by phosphate or acetate) and surficially adsorbed + nonsurficial forms in Fe and Mn oxides (extracted by hydroxylamine hydrochloride or ammonium oxalate). Soil extraction methods that dissolve some of the amorphous Fe, such as hydroxylamine hydrochloride, can be designed to provide closer estimates of bioavailable arsenic.

Animals↗
PubMed2004

Free radical scavenging activity of extracts prepared from fresh leaves of selected Chinese medicinal plants.

The free radical scavenging activity of the 80% methanolic extracts from fresh leaves of 300 Chinese medical woody plants was assessed with the aid of the stable DPPH radical. Among the plants screened, 56 species had strong free radical scavenging capacities, with IC50 values lower than 0.5 mg leaves per milliliter. Analysis of the medical uses of these plants showed that most of them are employed for their effects on hemostasis, as anti-inflammatory, antimicrobial or for treatment of dysentery. These uses may be directly linked to the content in tannins and flavonoids and consequently to their free radical scavenging activities.

Biphenyl Compounds↗
PubMedDate not supplied

Anti-acne effects of Oriental herb extracts: a novel screening method to select anti-acne agents.

The acne-therapeutic effects of Oriental herb extracts were investigated in terms of antichemotactic effect on polymorphonuclear leucocytes, antilipogenic actions, antibacterial activity against Propionibacterium acnes and resistance induction potency in the bacteria. The ethanol extract (0.01%) of Angelica dahurica markedly suppressed neutrophil chemotaxis, comparable to the effect of erythromycin (0.01%), whereas a strong antilipogenic effect was obtained with rhizoma coptidis (Coptis chinensis) extract (0.01%), leading to a higher efficacy than that of retinoic acid (0.01%). Interestingly, only Glycyrrhiza glabra showed a remarkable antibacterial activity against P. acnes, resulting in negligible induction of resistance, in comparison with a marked development of resistance in the bacteria treated with erythromycin. We suggest that an appropriate formulation containing A. dahurica, rhizoma coptidis and G. glabra could be helpful for the prevention and treatment of acne lesions.

Acne Vulgaris↗