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PubMed1984

[Analysis of residues of the fungicide cymoxanil in grapes using multicolumn HPLC].

Residues of the contact fungicide Cymoxanil in grapes were determined by a multidimensional multicolumn high pressure liquid chromatography technique (MC-HPLC). The sample pretreatment is a simple water extraction. Further clean-up and trace enrichment of an aliquot of the aqueous acidic sample solution is performed on-line via an automatized microprocessor controlled MC-HPLC system. In the range of 0.05-2.0 mg/kg grapes good linearity is achieved. Recoveries of Cymoxanil added to untreated grapes range between 70% and 80% at 0.05 and 2.0 mg/kg values, respectively, with a reproductibility of s +/- 3% at 0.2 mg/kg (n = 5). The total time for analysis is approximately 30 min.

Acetamides↗

PubMed2006

Recent developments in methods and technology for analysis of biological samples by MALDI-TOF-MS.

Matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF-MS) is widely used in a variety of fields because it has the characteristics of speed, ease of use, high sensitivity, and wide detectable mass range for obtaining molecular weights and for structural characterization of macromolecules. In this article we summarize recent developments in matrix additives, new matrices, and sample-pretreatment methods using off-probe or on-probe techniques or nanomaterials for MALDI-TOF-MS analysis of biological samples.

Analytic Sample Preparation Methods↗

PubMed1994

Immunoturbidimetry of serum apolipoproteins A-I and B on the Cobas Bio centrifugal analyzer: method validation and reference values.

We describe an immunoturbidimetric assay for estimating the concentration of apolipoproteins (apo) A-I and B in serum with a Cobas Bio centrifugal analyzer. Several analytical variables were studied: pH, temperature, wavelength, reaction time, sample pretreatment, antisera dilution, concentration of NaCl, phosphate buffer, and PEG 6000; a good optical response was obtained with small volumes of sample and antisera. Imprecision was low, with intrarun coefficient of variance (CV) from 1.1 to 2.8% and interrun CV from 1.8 to 3.2%. We demonstrated a good correlation between our method and radial immunodiffusion for apo A-I (r = 0.975) and apo B (r = 0.980). Reference values were calculated from a sample of 494 healthy subjects, and separate intervals for the ratio of apo A-I to apo B in gender and age subgroups were established.

Adult↗

PubMed2004

Behavior and quantification studies of amiloride drug using cyclic and square-wave adsorptive stripping voltammetry at a mercury electrode.

The cyclic voltammograms of amiloride at the hanging mercury drop electrode showed a single well-defined four-electron irreversible cathodic peak in Britton-Robinson (B-R) buffer of pH 2. At higher pH values (pH > or =3), two irreversible cathodic peaks corresponding to the transfer of four (first peak) and two (second peak) electrons, were obtained The peak potentials were shifted to more negative values on the increase of pH of the medium, implying the involvement of protons in the electrode reaction and that the proton-transfer reaction precedes the proper electrode process. The 4-electron single peak (pH 2) or the first peak (pH > or = 3) may be attributed to the cleavage of the -CH=NH double bond of the N-imidino amide group with the release of NH(3) molecule. While the second peak may be due to the saturation of the C?O double bond of the carboxamide moiety. Based on the interfacial adsorptive character of the drug onto the mercury electrode surface, a simple, sensitive and low cost square-wave adsorptive cathodic stripping (SWAdCS) voltammetric procedure was optimized for analysis of the drug. The optimal operational conditions of the proposed procedure were: accumulation potential E(acc)= -0.7 V, accumulation time t(acc)= 60-65s, scan increment= 10 mV, pulse-amplitude = 50-60 mV, frequency =120 Hz using a B-R buffer of pH 8 as a supporting electrolyte. The linear concentration range was found to be 2 x 10(-9) to 2 x 10(-7) M amiloride with limits of detection (LOD) and quantitation (LOQ) of 1.9 x 10(-10) and 6.3 x 10(-10) M, respectively. The procedure was successfully applied for determination of amiloride in pharmaceutical formulation and spiked in human serum. The LOD and LOQ of amiloride spiked in human serum were 5.7 x 10(-10) and 1.9 x 10(-9) M amiloride, respectively. The procedure did not require sample pretreatment or any time-consuming extraction or evaporation steps, other than deproteinization and then centrifugal separation of protein from serum sample prior to analysis of the drug.

Amiloride↗

PubMed1997

Determination of irinotecan (CPT-11) and its active metabolite SN-38 in human plasma by reversed-phase high-performance liquid chromatography with fluorescence detection.

Sensitive high-performance liquid chromatographic assays have been developed to determine the levels of the lactone and lactone plus carboxylate (total) forms of the antitumor agent irinotecan (CPT-11) and its active metabolite SN-38, in human plasma. The related compound camptothecin was used as the internal standard. The selective sample pretreatment for the lactone forms involved a single solvent extraction with acetonitrile-n-butyl chloride (1:4, v/v), whereas the sample clean-up for the total forms was a simple protein precipitation with aqueous perchloric acid-methanol (1:1, v/v), which results in the conversion of the carboxylate to the lactone forms. Chromatography was carried out on a Hypersil ODS column, with detection performed fluorimetrically. The methods have been validated, and stability tests under various conditions have been performed. The lower limits of quantitation are 0.5 and 2.0 ng/ml for the lactone and total forms, respectively. The assays have been used in a single pharmacokinetic experiment in a patient to investigate the applicability of the method in vivo.

Antineoplastic Agents, Phytogenic↗

PubMed2001

A factorial analysis of silanization conditions for the immobilization of oligonucleotides on glass surfaces.

The modification of glass surfaces with (3-mercaptopropyl)trimethoxysilane and the application of this to DNA chip technology are described. A range of factors influencing the silanization method, and hence the number of surface-bound, chemically active thiol groups, were investigated using a design of experiment approach based on analysis of variance. The number of thiol groups introduced on glass substrates were measured directly using a specific radiolabel, [14C]cysteamine hydrochloride. For liquid-phase silanization, the number of surface-bound thiol groups was found to be dependent on both postsilanization thermal curing and silanization time and relatively independent of silane concentration, reaction temperature, and sample pretreatment. Depending on the conditions used in liquid-phase silanization, (1.3-9.0) x 10(12) thiol groups/cm2 on the glass samples were bound. The reliability and repeatability of liquid- and vacuum-phase silanization were also investigated. Eighteen-base oligonucleotide probes were covalently attached to the modified surfaces via a 3'-amino modification on the DNA and subsequent reaction with the cross-linking reagent N-(gamma-maleimidobutyryloxy) succinimide ester (GMBS). The resulting probe levels were determined and found to be stoichiometric with that of the introduced thiol groups. These results demonstrate that silanization of glass surfaces under specific conditions, prior to probe attachment, is of great importance in the development of DNA chips that use the simple concept of the covalent attachment of presynthesized oligonucleotides to silicon oxide surfaces.

Factor Analysis, Statistical↗

PubMed2004

Direct determination of lead isotopes (206Pb, 207Pb, 208Pb) in arctic ice samples at picogram per gram levels using inductively coupled plasma-sector field MS coupled with a high-efficiency sample introduction system.

Adopting strict cleanroom procedures, ice samples from the Canadian High Arctic have been analyzed for Pb concentrations and Pb isotopes (206Pb, 207Pb, 208Pb) using ICP-SMS. The detection limit for Pb (0.06 pg g(-1)) was approximately 2 orders of magnitude lower than the lowest concentration of Pb in the ice samples (range, 4.3-1660 pg g(-1); median, 45 pg g(-1)). Acidification of ice samples with high-purity HNO3 for stabilization purposes contributed only 0.004 pg of Pb g(-1), which is an insignificant source of Pb. Using a new sample introduction system consisting of a heated (140 degrees C) minicyclonic spray chamber and a Peltier cooled condenser (2 degrees C) and by replacing the conventional sample cone with a high-performance cone, signal intensities for Pb were increased by approximately 1 order of magnitude. Thus, it was possible not only to measure Pb isotope ratios directly using ICP-SMS but also to achieve reasonable precision (approximately 0.2%) at low picogram per gram concentrations of total Pb. This precision is comparable to that achievable by thermal ionization mass spectrometry at such low Pb concentrations, but the ICP-SMS requires much less sample volume (approximately 2 mL), needs no sample pretreatment, and therefore is considerably faster and less expensive than the conventional approach. Even though absolute Pb concentrations in two ice samples dating from 1974 and 1852 were very similar (9 and 6 pg g(-1)) their fundamentally different isotopic signature (206Pb/207Pb: 1.169 +/- 0.002 vs 1.147 +/- 0.003) clearly indicates different sources of Pb. The analytical procedures described here, therefore, offer great promise for fingerprinting the predominant sources of atmospheric Pb in polar snow and ice.

Journal Article↗

PubMed1997

Miniaturized supported liquid membrane device for selective on-line enrichment of basic drugs in plasma combined with capillary zone electrophoresis.

A hollow fiber miniaturized supported liquid membrane (SLM) device for sample preparation is connected on-line with capillary electrophoresis and used for determination of a basic drug, bambuterol, in human plasma. The analyte is extracted from the outside of the hollow fiber (donor) through the liquid membrane (pores of the fiber impregnated with organic solvent) into the acceptor solution in the fiber lumen. The process is driven by differences in pH between the donor and acceptor solution. The whole volume of the acceptor solution can then be injected into the CZE capillary by using the double-stacking procedure for large volume-injection. Very clean extracts of low ionic strength are obtained from the SLM treatment, making this sample pretreatment method compatible with the CZE double-stacking procedure, which in turn makes it possible to inject large volumes of sample onto the separation capillary. Good performance of the whole procedure is demonstrated, and detection limits in the low nanomolar range were obtained in spite of the relatively weak UV absorbance of bambuterol. Extractions through the miniaturized SLM unit can be performed for 5-6 h without regenerating the fiber. The regeneration procedure was tested, and no relevant changes in the performance of the extraction could be found after seven regenerations, allowing the same fiber to be used for a week.

Electrophoresis, Capillary↗

PubMed2003

Influence of different mineral and Organic pesticide treatments on Cd(II), Cu(II), Pb(II), and Zn(II) contents determined by derivative potentiometric stripping analysis in Italian white and red wines.

This paper deals with the use of derivative potentiometric stripping analysis (dPSA) as a rapid and precise method to determine Cd(II), Cu(II), Pb(II), and Zn(II) levels in red and white wine samples from Sicily, Campania, and Tuscany and to investigate the possible connection between the content of these metals and the pesticide treatments used in vine-growing to control plant diseases and pests. dPSA allowed direct quantitation of heavy metals in acidified wines without any sample pretreatment. Mean recoveries of Cd(II), Cu(II), Pb(II), and Zn(II) ranged from 95.5 to 99.2% for white wine samples and from 96.1 to 100.0% for red wine samples. The obtained results showed that Cd(II) was not found in any sample and that Cu(II), Pb(II), and Zn(II) levels were always lower than the toxicity limits in both fungicide- and water-treated wines. Nevertheless, the contents of metals were increased in samples from organic and inorganic pesticides treatment with respect to the water-treated samples. In particular, quinoxyfen, dinocap-penconazole, and dinocap applications considerably increased Cu(II) and Zn(II) contents in white and red wines. The levels of lead were significantly raised by azoxystrobin and sulfur treatments.

Cadmium↗

PubMed2003

Neamin as an immunogen for the development of a generic ELISA detecting gentamicin, kanamycin, and neomycin in milk.

A broad-specific ELISA using one antibody preparation for the detection of gentamicin, kanamycin, and neomycin in milk is reported for the first time. For the immunization of rabbits, neamin was used as the generic hapten on the basis of the facts that it is a two-ring fragment of neomycin and, in shape and charge, it resembles parts of kanamycin and gentamicin. Neamin was linked to the preactivated carrier protein keyhole limpet hemocyanin by EDC coupling. The specificity and sensitivity of the polyclonal antibodies for the aminoglycoside antibiotics were tested in a competitive assay using homologous and heterologous conjugates coupled by various conjugation procedures as the ELISA solid phase. In contrast to the homologous assay recognizing only neomycin, the heterologous assay could be used for the detection of the whole subclass of deoxystreptamin antibiotics in buffer and raw milk. Gentamicin, kanamycin, and neomcyin were detected in artificially contaminated undiluted raw milk (without sample pretreatment) with 50% inhibition levels at 9, 21, and 113 ng mL(-)(1), respectively. Neomycin levels were also measured in milk samples obtained from a cow suffering from mastitis and treated with an antibiotic cocktail including neomycin. Levels below the EU maximum residue levels for deoxystreptamin antibiotics could readily be measured in this generic ELISA.

Animals↗

PubMed1993

Clarithromycin vs. amoxicillin suspensions in the treatment of pediatric patients with acute otitis media.

Clarithromycin is a new macrolide antibiotic that is active in vitro against a variety of organisms that are responsible for acute otitis media in children. The parent compound is metabolized to microbiologically active 14-hydroxy clarithromycin, which is especially active against Haemophilus influenzae. The safety and efficacy of clarithromycin and amoxicillin suspensions were compared in the treatment of acute otitis media in children 1 to 12 years of age inclusive. This was a Phase III, single blind (investigator-blind), randomized, multicenter clinical trial. Clarithromycin oral suspension was given in a dose of 7.5 mg/kg (maximum, 500 mg) twice daily, and amoxicillin suspension in a dose of 20 mg/kg (maximum, 750 mg) was given twice daily for 7 to 10 days in a 1:1 ratio. Clinical evaluations were performed pretreatment, within 48 hours posttreatment and 10 to 14 days posttreatment. Myringotomy was performed in every child to obtain a microbiologic sample pretreatment and at subsequent visits as clinically indicated. A total of 79 children were enrolled, 39 in the clarithromycin and 40 in the amoxicillin treatment group. Thirty-two children were excluded from the efficacy analysis for various reasons. Clinical success (cure and improvement) rates at 0 to 4 days posttreatment were 93% for clarithromycin and 90% for amoxicillin (P > 0.999). Altogether 17 children (10 receiving clarithromycin, 7 receiving amoxicillin) experienced some adverse event, with gastrointestinal disorders being the most common complaint. No clinically significant differences in laboratory tests were found between the groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute Disease↗

PubMed1992

Cathepsin B/L-, elastase-, tryptase-, trypsin- and dipeptidyl peptidase IV-like activities in gingival crevicular fluid. A comparison of levels before and after basic periodontal treatment of chronic periodontitis patients.

20 chronic periodontitis patients were given a full periodontal examination, including measurements of probing depth, clinical attachment loss, gingival index, bleeding index and plaque index. At a second visit, gingival crevicular fluid (GCF) was collected from the deepest accessible probing site of each tooth. The patients then received scaling, root planing and other appropriate nonsurgical treatment. GCF was collected from the same sites as sampled pretreatment and clinical parameters were measured again. Cathepsin B/L-, elastase-, tryptase-, trypsin-, and dipeptidyl peptidase IV-like activities in GCF samples were determined by fluorimetric assay with peptidyl derivatives of 7-amino-4-trifluoromethyl coumarin. Following treatment, there were reductions in all clinical parameters and all protease activities. Most were statistically significant both on a patient level using average patient values and on a site level using either individual patient or pooled patient data. As in previous pre-treatment comparisons, post-treatment protease levels correlated positively and significantly with the corresponding clinical parameters at patient and site levels. The reductions and correlations were more marked for total enzyme activities than concentrations. GCF protease levels appear to reflect the clinical status of periodontal lesions and may thus be of value in monitoring disease activity.

Adult↗

PubMed2005

Utility of specific IgG4 response in saliva and serum samples for the diagnosis and follow up of human neurocysticercosis.

This study was aimed to analyze the Ig subtypes and IgG1 and IgG4 subclass responses to crude soluble extract (CSE) antigen and Ag B of Cysticercus cellulosae in pre and post treatment (PT) saliva and serum samples for the diagnosis and follow-up of neurocysticercosis (NCC) patients. Saliva and serum samples collected from 55 patients (15 highly suggestive of NCC clinically and radiologically, 10 hydatidosis, 10 other helminthic infections, 10 tubercular meningitis, 10 neurological disorders other than NCC), 15 normal healthy subjects and 10 NCC patients at 1, 3 and 6 months following albendazole therapy were analyzed for specific IgG, IgG1, IgG4, IgM and IgA antibody responses by ELISA. With the use of CSE Ag, the rank orders in saliva for sensitivity was IgG (71.4%) > IgG1 (68.2%) > IgG4 (65.2%) > IgM (57.7%) > IgA (55.5%) and specificity IgG1 = IgA (93.2%) > IgG = IgG4 = IgM (91.6%) while in serum, sensitivity was IgG (78.9%) > IgG1 (71.4%) > IgG4 (68.2%) > IgA (65.2%) > IgM (62.5%) and specificity IgG1 = IgG4 (90.2%) > IgA (85.9%) > IgG (83.3%) > IgM (82.1%). With the use of Ag B in saliva, the sensitivity was IgG (65.2%) > IgG1 = IgG4 (62.5%) > IgM = IgA (55.5%) and specificity with the use of saliva was IgG1 = IgG4 = IgM (94.8%) > IgG (93.2%) > IgA (91.6%) while with serum the sensitivity was IgG = IgG1 (68.2%) > IgG4 (65.2%) > IgA (62.5%) > IgM (57.7%) and specificity was IgG1 (93.2%) > IgG4 = IgM (91.6%) > IgA (90.2%) > IgG (87.3%). Comparative analysis of antibody responses in patients with single Vs multiple CT scan lesions indicated higher sensitivity in multiple lesion patients. Antibody responses in PT samples indicated that the undetectable IgG4, IgM and IgA responses in saliva samples correlated well with the CT scan reports while in serum samples, responses persisted longer. In conclusion, this study indicated that due to the lower sensitivity of IgM and IgA responses in pretreatment samples, detection of IgG4 subclass in saliva to either CSE Ag or AgB may serve better marker in the NCC follow-up.

Antigens, Helminth↗

PubMed1993

Solid phase extraction of morphine from whole blood by means of Bond Elut Certify columns.

The use of Bond Elut Certify columns for the isolation of morphine from whole blood was evaluated. In order to monitor possible losses and the elution profile of morphine, a small amount of the tritiated analogue was added to the samples. Four sample pretreatment methods, three protein precipitation methods and one sonication/dilution method, were tested. The latter one gave the best results. The blood sample was applied onto the column at pH 3.3 after sonication and dilution with 0.1 M phosphate buffer (pH 3.3). The retention of morphine was affected by the pH of the samples, and the loss of morphine during sample application was minimized at low pH (3.3). The interferences were removed by washing the column with the phosphate buffer, 0.01 M acetic acid (pH 3.3), and methanol, sequentially. Ammoniated methanol, 2 mL at 2%, was selected to elute morphine. As a result, more than 80% of 3H-morphine was recovered for concentrations of morphine ranging from 5 to 4000 ng/mL.

Forensic Medicine↗

PubMed1993

Collagen I carboxyterminal propeptide in human gingival crevicular fluid before and after periodontal treatment.

The amount of procollagen I carboxyterminal propeptide (PICP) in crevicular fluid (CF) was measured in three periodontitis patients. Samples were collected from 29 sites before treatment (scaling, root planing, and curettage) and 2, 5, 10, 20, and 40 days after treatment, by placing two paper strips in periodontal pockets for 5 s. The amount of fluid in strips was measured by the Periotron device. Control samples were collected from subjects with minimal gingival inflammation. PICP was extracted into saline solution and determined by a radioimmunologic method. Plaque index, papilla bleeding index, and pocket depth were recorded before and 40 days after treatment. The CF PICP mean concentration was 4.2 mg/l in the pretreatment samples. Five days after treatment a statistically significant increase in PICP concentration was seen in all subjects. The peak appeared on days 5 or 10 in 27 sites. The mean peak PICP concentrations of the subjects were 5-10 times higher than the pretreatment values. Twenty days after treatment, mean PICP concentration decreased to pretreatment level. PICP concentrations did not correlate with the clinical parameters. In control samples PICP amounts were below the detection limit. CF PICP is a new marker of type I collagen metabolism in periodontal tissues. It was concluded that elevated PICP concentrations in CF after periodontal treatment reflected increased type I collagen synthesis in periodontal tissues and that the peak in type I collagen synthesis takes place 5-10 days after treatment.

Adult↗

PubMed2005

Amperometric biosensor for determining human salivary phosphate.

An amperometric biosensor was constructed for analysis of human salivary phosphate without sample pretreatment. The biosensor was constructed by immobilizing pyruvate oxidase (PyOD) on a screen-printed electrode. The presence of phosphate in the sample causes the enzymatic generation of hydrogen peroxide (H(2)O(2)), which was monitored by a potentiostat and was in proportion to the concentration of human salivary phosphate. The sensor shows response within 2s after the addition of standard solution or sample and has a short recovery time (2 min). The time required for one measurement using this phosphate biosensor was 4 min, which was faster than the time required using a commercial phosphate testing kit (10 min). The sensor has a linear range from 7.5 to 625 microM phosphate with a detection limit of 3.6 microM. A total of 50 salivary samples were collected for the determination of phosphate. A good level of agreement (R(2)=0.9646) was found between a commercial phosphate testing kit and the phosphate sensor. This sensor maintained a high working stability (>85%) after 12h operation and required only a simple operation procedure. The amperometric biosensor using PyOD is a simple and accurate tool for rapid determinations of human salivary phosphate, and it explores the application of biosensors in oral and dental research and diagnosis.

Biosensing Techniques↗

PubMed1989

Solid-phase extraction and high-performance liquid chromatographic determination of flumequine and oxolinic acid in salmon plasma.

Two methods for determination of oxolinic acid and flumequine in salmon plasma are described. The first method applies sample pretreatment on C2 disposable solid-phase extraction columns. The second method is based on direct plasma injection and on-line sample clean-up on a polystyrene-divinylbenzene precolumn. After column-switching, the analytes are separated on a polystyrene-divinylbenzene analytical column and detected with a fluorescence detector. Validation of the methods showed good sensitivity, precision and reproducibility. Both methods are well suited for determination of plasma levels of the drugs in pharmacokinetic studies in Atlantic salmon.

Animals↗

PubMed2004

Sulfated nanozirconia: an investigation on acid-base properties and n-butane isomerization activity.

Hydrated zirconia was synthesized by an organo-inorganic route employing surfactant and was sulfated using aqueous ammonium persulfate, followed by drying at 110 degrees C. The sample thus obtained was calcined at 600 degrees C to obtain sulfated zirconia and was characterized by several physicochemical methods. Crystallite sizes of sulfated zirconia were calculated from X-ray line broadening using the Debye-Scherer equation and were found to be in the range of 25 nm. When pretreated in air, the catalyst was found to exhibit butane isomerization activity at a temperature as low as 35 degrees C under atmospheric pressure. It showed conversion as high as 37% at 100 degrees C under normal pressure when pretreated in air, whereas nitrogen-pretreated catalyst showed zero activity under similar conditions. NH(3) and CO(2) temperature-programmed desorption studies on air- and helium-pretreated samples indicated that the catalyst surface changes appreciably during air pretreatment. Results on butane isomerization in conjunction with TPD studies suggest that zirconium-oxy sites play an important role in butane activation during the reaction.

Butanes↗