Biomedical subjects

C Baggiani

Publications and source records attributed to C Baggiani.

14 recordsLinked to original sources

PubMed2006

Molecularly imprinted solid-phase extraction method for the high-performance liquid chromatographic analysis of fungicide pyrimethanil in wine.

A method for molecularly imprinted solid-phase extraction (MISPE) of the fungicide pyrimethanil from wine samples has been investigated. The molecular imprinted polymer was obtained by iniferter-mediated grafting on porous chloromethylated polystyrene beads, using methacrylic acid as the functional monomer and ethylene glycol dimethacrylate as the cross-linker. The imprinted beads were evaluated for use as a solid-phase extraction sorbent, in order to develop the extraction protocol in aqueous standards and red wine samples. The optimised extraction protocol resulted in a reliable MISPE method suitable for HPLC analysis (stationary phase: Cromolith Performance C18 column, 100 mm x 4.6 mm; mobile phase: acetonitrile-water (3:2, v/v), flow-rate: 1.00 ml/min; detection 270 nm). It was selective for pyrimethanil and the related pyrimidinic fungicides cyprodinil and mepanipyrim, while the non-pyrimidinic fungicides benalaxyl, chlozolinate, furalaxyl, iprodione, metalaxyl, nuarimol, procymidone and vinclozolin were not extracted. Recoveries performed on a wine matrix spiked with pyrimethanil at three different concentration levels were reproducible and were in good agreement with the recoveries performed on buffer, coming out between 80 and 90% (85+/-7.0% at 0.50 microg/ml, 79+/-1.6% at 2.0 microg/ml and 87+/-5.6% at 20 microg/ml). Preconcentration and quantitative extraction of pyrimethanil from wine samples was shown to be feasible down to 0.1 microg/ml.

Chromatography, High Pressure Liquid↗

PubMed2004

Multivariate analysis of the selectivity for a pentachlorophenol-imprinted polymer.

A pentachlorophenol (PCP)-imprinted polymer (MIP) was obtained by thermal polymerization of a mixture of template, 4-vinylpyridine and ethylene glycol dimethacrylate with molar ratio 1 +3 + 27, using as porogenic solvent methanol-water ( 3 + 1(v/v)). The polymer was packed in an HPLC column and selectivity towards 52 PCP-related phenols (22-chloro-, 21-alkyl-, 4-aryl-, 3-methoxy- and 6-polyphenols) was measured using acetonitrile-acetic acid (99 + 1(v/v)) as mobile phase. The same was made for a reference polymer obtained without pentachlorophenol (NIP). The molecular recognition properties of the imprinted polymer were expressed in terms of selectivity index (SI), calculated for each phenol as k(NIP)/k(MIP). Sixteen molecular descriptors were calculated for each molecule: qO, the partial charge of the phenolic oxygen atom; qH, the partial charge of the phenolic hydrogen atom; Deltaq, the absolute value of the difference qO - qH; HOMO, the highest occupied molecular orbital; LUMO, the lowest unoccupied molecular orbital; Deltaorb, absolute value of the difference HOMO - LUMO; micro(2), the square of total dipole moment; MW, the molecular weight; SAS, the solvent-accessible molecular surface area; hSAS, the hydrophobic solvent-accessible molecular surface area; Svdw, the van der Waals molecular surface area; hSvdw, the hydrophobic part of Svdw; MOv, the molecular ovality; RG, the radius of gyration; logP, the logarithm of n-octanol-water partition coefficient; pK, the phenolic dissociation constant. Correlations between selectivity index and these descriptors were searched utilizing multivariate principal component analysis (PCA). The multivariate model obtained by regression on the principal components correlate collectively several of the calculated descriptors with the polymer selectivity. The magnitude of the model's parameters shows that selectivity is strongly influenced by molecular descriptors having structural character, such as MW, hSvdw and logP, while the effect of molecular descriptors having electronic character, such as qO and pK, is much less marked.

Multivariate Analysis↗

PubMed2001

Molecularly imprinted solid-phase extraction sorbent for the clean-up of chlorinated phenoxyacids from aqueous samples.

A molecularly imprinted polymer (MIP) was synthesized using the herbicide 2,4,5-trichlorophenoxyacetic acid as a template, 4-vinylpyridine as an interacting monomer, ethylendimethacrylate as a cross-linker and a methanol-water mixture as a porogen. The binding properties and the selectivity of the polymer towards the template were investigated by frontal and zonal liquid chromatography. The polymer was used as a solid-phase extraction material for the clean-up of the template molecule and some related herbicides (2,4-dichlorophenoxyacetic acid, fenoprop, dichlorprop) from river water samples at a concentration level of ng/ml with quantitative recoveries comparable with those obtained with a traditional C18 reversed-phase column when analyzed by capillary electrophoresis. The results obtained show that the MIP-based approach to the solid-phase extraction is comparable with the more traditional solid-phase extraction with C18 reversed-phase columns in terms of recovery, but it is superior in terms of sample clean-up.

2,4,5-Trichlorophenoxyacetic Acid↗

PubMed2001

A molecular imprinted polymer with recognition properties towards the carcinogenic mycotoxin ochratoxin A.

A molecularly imprinted polymer which recognises the mycotoxin ochratoxin A was prepared using the mimic N-(4-chloro-1-hydroxy-2-naphthoylamido)-(L)-phenylalanine as a template. The polymer was obtained by dissolving the template, methacrylic acid and ethylendimethacrylate in chloroform and polymerising the mixture by thermal treatment at 60 degrees C. The monolith obtained was crushed, sieved to 30-90 microm and extensively washed till the template could no longer be found in the washing solution. The binding properties towards the template, ochratoxin A and several related molecules were measured by eluting with acetonitrile and chloroform a HPLC column packed with the imprinted polymer. The experimental results show that the polymer recognises not only the template well, but also the ochratoxin A. The specific molecular recognition effect is due to hydrogen bond interactions but in order to assure the full recognition effect adjunctive steric factors are necessary. The magnitude of these interactions can be controlled by the use of limited amounts of acetic acid in the mobile phase. From the measurement of the relative selectivity it was found that only the simultaneous presence of the carboxyl, the phenolic hydroxyl and certain peculiar substructures such as the chlorine atom assures the whole recognition of the template.

Carcinogens↗

PubMed2000

Chromatographic characterization of molecularly imprinted polymers binding the herbicide 2,4,5-trichlorophenoxyacetic acid.

Two polymers binding the herbicide 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) were prepared by utilising the technique of the non-covalent molecular imprinting polymerisation in an aqueous medium. The polymers obtained were packed in HPLC columns and the effects of the mobile phase composition on the retention of the imprinting molecule and the selectivity of the stationary phases towards several analogous structures were studied by liquid chromatography. The columns showed a good level of selectivity towards the template and strictly related molecules. It was found that the molecular recognition mechanism acting on the columns was dependent on a combination of ion pair and hydrophobic interactions.

2,4,5-Trichlorophenoxyacetic Acid↗

PubMed2000

Application of an ELISA to the determination of benalaxyl in red wines.

The applicability of an ELISA for detection and quantification of benalaxyl in red wine samples is described. The study of the influence of this matrix on the reliability of the assay indicates that red wine samples require a rapid and simple cleanup step before ELISA assay. Recovery and precision of the method were evaluated by spiking red wine samples with benalaxyl in the 0.5-24 ng/mL range. Benalaxyl can be determined with good accuracy and precision up to 0. 5 ng/mL in starting red wine samples (detection limit of 0.13 ng/mL). No false negative or positive results were obtained. Authentic red wine samples were analyzed by ELISA and by RP-HPLC. The amounts of benalaxyl found by ELISA were in good agreement with RP-HPLC analysis.

Alanine↗

PubMed1999

A general method to perform a noncompetitive immunoassay for small molecules.

A new general method to perform a noncompetitive immunoassay for low-molecular-mass analytes (less than 6000 Da) is described and checked using cortisol as a model system. The method is based on the use of a "polydentate ligand" (cortisol-poly(L-lysine) conjugate) able to block the antibody sites unoccupied by the analyte, followed by the replacement of an antibody-bound analyte by an enzyme-labeled analyte (cortisol-horseradish peroxidase), and permits the direct measurement of the analyte bound sites. The observed signal shows a near-linear correlation with the analyte concentration. The characteristics of interactions between the analyte and polydentate ligand with the specific antibody were studied to perform a preliminary evaluation of the noncompetitive immunoassay for cortisol. The noncompetitive assay was compared with a competitive immunoassay obtained under the same conditions and using the same reagents. The results of the experiments showed a lower detection limit for the noncompetitive model (0.15 ng mL-1 rather than 0.72 ng mL-1), emphasizing that the model is successful. Moreover, as the polydentate ligand is prepared from the same hapten used for the immunogen synthesis, this type of noncompetitive immunoassay appears generally applicable to all small molecules for which antibodies have been obtained.

Animals↗

PubMed1996

Strategy for fractionating high-affinity antibodies to steroid hormones by affinity chromatography.

A general strategy for fractionating high-affinity antibodies to steroid hormones has been developed and applied to the fractionation of an antiserum to testosterone 3-(O-carboxymethyl)oxime-bovine serum albumin. If the antibodies interacting with a stationary phase containing a low concentration of immobilized steroid are considered as monovalent binders, a simple equation can be applied to show that the affinity of the antibody-stationary phase interaction must be higher than about 2 x 10(6) l mol-1 in order to avoid the loss of antibodies during the loading and washing of the column. Conversely, to elute the retained antibodies, the affinity must be decreased to a value lower than about 2 x 10(5) l mol-1 and the dissociation rate constants of the antibody-steroid complexes must be >> 1 s-1. In order to prepare an affinity column that satisfies these conditions, the ligand to be immobilized was selected on the basis of the cross-reactions of the antiserum with several testosterone derivatives. Moreover, the dissociation rate constants of several antibodies of known affinity were measured, together with the effect of acidic buffers and various organic solvents on the antiserum-testosterone interaction. Then, an affinity column, prepared by coupling testosterone 17 beta-acetate to AH-Sepharose 4B, was used to load the antiserum without loss of antibodies during the washing step. The retained antibodies were successfully eluted by a mixture of 30% dioxane in phosphate-citrate buffer (pH 3.4). The affinity of the eluted antibodies was in the range 7 x 10(9)-2 x 10(11) l mol-1 and was linearly related to the retention volume. These results confirm that high-affinity antibodies can be fractionated to steroid hormones by a proper choice of the ligand on the stationary phase and the eluent composition.

Antibodies↗

PubMed1995

Fractionation of an antiserum to progesterone by affinity chromatography: effect of pH, solvents and biospecific adsorbents.

Several progesterone-AH Sepharose 4B matrices were prepared as biospecific adsorbents suitable for affinity chromatography to fractionate antibodies of different affinity and specificity from a polyclonal antiserum to progesterone-11 alpha-hemisuccinate-BSA. From an affinity column of progesterone-11 alpha-hemisuccinate-AH Sepharose 4B no antibodies can be eluted, even with glycine buffer (pH 2.6) and 30% of 2-methoxyethanol. The use of biospecific adsorbents, prepared by coupling with AH Sepharose 4B progesterone derivatives [5-pregnene-3,20-dione di(ethyleneacetal)-11 alpha-ol-11 alpha-hemisuccinate; 4-pregnene-11,20 beta-diol-3-one-11 alpha-hemisuccinate 20 beta-benzoate; progesterone-3-carboxymethyloxime] having a low cross-reactivity with the antiserum, makes the elution of various antibody fractions of variable affinity and specificity possible. 2-Methoxyethanol or N,N-dimethylformamide gradients, in acetate or TRIS buffer, were equally efficient for fractionating the antiprogesterone serum, while a decreasing pH gradient was less effective and eluted antibody fractions that were further separated into various binding components by a solvent gradient. Antibodies eluted from the affinity columns by an eluent containing a high solvent concentration have affinities higher than antibodies eluted at lower solvent concentration.

Absorption↗

PubMed1994

Immunochemical methods for environmental monitoring.

Immunochemical methods for environmental analysis must be taken into consideration more for their ability to expand the potential of analytical measures rather than for substituting current methodologies. Moreover, the full potential of these methods has yet to be realized. Indeed, the terms and concepts of immunology are new to most analytical chemists, even if environmental science has always been an interdisciplinary field. On the other hand, the clinical development of immunoassays means that much experience has been gained in the analysis of blood, urine, and tissue samples. The immunochemical analysis of samples from soils, ground water, waste chemicals, poses new challenges in sample preparation that have yet to be extensively studied, and in the future there may be immunoassays better suited for the particular problems associated with environmental monitoring.

Environmental Monitoring↗

PubMed1993

Solvent effect on testosterone-antitestosterone interaction.

The inhibition of the binding between testosterone and antitestosterone antiserum caused by organic solvents was studied at pH 7.4, 298 K. Inhibition curves were obtained at variable ranges of molar fractions for the following solvents: methanol (range 0-0.4), ethanol (0-0.317), 1-propanol (0-0.082), 2-propan-ol (0-0.260), t-butanol (0-0.223), ethylenglycol (0-0.189), 2-methoxyethanol (0.036), 2-butoxyethanol (0-0.063), 1,4-dioxan (0-0.124), tetrahydrofuran (0-0.238) and acetonitrile (0-0.392). Steroid-antibody binding decreases with increasing molar fraction of solvent in the reaction mixture for all but tetrahydrofuran and acetonitrile, which enhance binding at low molar fraction then cause a sharp inhibition. Molar fraction of solvent that causes a 50% binding inhibition is uncorrelated to some solvent properties (i.e. dielectric constant, polarity index, dipole moment) but is inversely correlated to the molecular mass of the solvent. The correlation becomes better by taking into account the length of the solvent molecule, or the Randic molecular connectivity index, suggesting that binding inhibition could be related to the length of the solvent molecules that displace water around the steroid molecule. However, the increase of binding observed at low molar fraction with tetrahydrofuran and acetonitrile, together with very different shapes of inhibition curves suggest that a molecular mechanism based on the differential solvation of the steroid by solvent and water molecules must be taken into account to explain adequately the solvent effect on testosterone-antitestosterone interaction.

Antibodies↗

PubMedDate not supplied

Functionalized biopolymers as soluble macromolecular chelating agents.

Two different conjugates of bovine serum albumin (BSA) with lysine and a derivative of imidazole have been synthesized to obtain watersoluble macromolecules with binding properties against bivalent transition metal ions. Syntheses have been carried out using the 60 aminogroups or the 99 carboxylic groups on BSA for the coupling reactions, with such molar ratios able to produce highly substituted BSA. The skill of each conjugate to bind metal ions in aqueous medium was studied through the use of titration curves with some metal ions, characterized by a good affinity for the free ligand. Both the conjugates allow us to recover a high number of metal ions per protein molecule, close to the number of ligand molecules on the BSA surface in the case of the lysine conjugate, whereas in the case of the imidazole conjugate M3L complexes are performed.

Biopolymers↗

PubMedDate not supplied

The complexation of mercury (II) and organomercurial compounds by 8-hydroxyquinoline-bovine serum albumin conjugates.

The complexing properties of conjugates between 8-hydroxyquinoline and bovine serum albumin (Ox-BSA) towards inorganic and organic mercury were studied. Two Ox-BSA conjugates (different substitution ratio) were prepared and their complexing properties were studied. Through the use of titration curves with mercury (II), methylmercury and ethylmercury an evaluation of the complex stoichiometry and stability was obtained, showing that Ox-BSA has good affinity for all investigated mercuric compounds and that the stability increases in the order: Hg (II) < CH3Hg+ < C2H5Hg+, whatever conjugate is considered. Complexes show a stoichiometry of 1:1 between mercury and 8-hydroxyquinoline residues, except with the high substituted conjugate and Hg2+ ion. The skill of the high substituted conjugate to bind inorganic and organic mercury in the presence of NaCl was also studied. Organic mercuric complexes do not show significant modification due to NaCl. Nevertheless, considering inorganic mercury, the number of retained metal ions per protein molecule increases if the NaCl concentration becomes higher than 0.1 M, probably because at high NaCl concentrations 1:1 complexes between mercury and 8-hydroxyquinoline are preferred to 1:2 complexes.

Animals↗

PubMedDate not supplied

Separation and characterization of a yeast alcohol dehydrogenase conjugate with theophylline.

The purpose of this work is to characterize the structural difference of conjugates in order to condition the immunoreactivity of the enzymatic tracer in the homogeneous immunoassays. Conjugates between yeast alcohol dehydrogenase (ADH) and 7-theophyllincarboxyalkyl acids were obtained by the mixed-anhydride method, and characterized with kinetic, electrophoretic and chromatographic techniques (IEF, MCC, CF). The enzyme activities were found to be inversely proportional to the substitution grade but not correlated to the number of carbon atoms of the carboxyalkyl spacer arms. Only the oxidative form (ADH2) of the enzyme was found capable of reacting with theophylline derivatives in the experimental conditions. Enzyme conjugates were resolved into three components, due to the different distribution of theophylline moieties on the surface of the protein, as confirmed by examination of the tryptic patterns of the single components.

Alcohol Dehydrogenase↗