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The miniaturization and optimization of a white rot fungal bioremediation experiment is described in this paper. The optimized procedure allows determination of the degradation kinetics of anthracene. The miniaturized procedure requires only 2.5 ml of culture medium. The experiment is more precise, robust, and better controlled comparing it to classical tests in flasks. Using this technique, different parts, i.e., the culture medium, the fungi, and the cotton seal, can be analyzed. A simple sample preparation speeds up the analytical process. Experiments performed show degradation of anthracene up to approximately 60 % by Irpex lacteus and up to approximately 40 % by Pleurotus ostreatus in 25 days. Bioremediation of anthracene by the consortium of I. lacteus and P. ostreatus shows the biodegradation of anthracene up to approximately 56 % in 23 days. At the end of the experiment, the surface tension of culture medium decreased comparing it to the blank, indicating generation of surfactant compounds.

Background For over two decades, a racemic mixture of oxamniquine (OXA) was administered to patients infected by Schistosoma mansoni, but whether one or both enantiomers exert antischistosomal activity was unknown. Recently, a ~30 kDa S. m ansoni sulfotransferase (SmSULT) was identified as the target of OXA action. Methodology/Principal Findings Here, we separate the OXA enantiomers using chromatographic methods and assign their optical activities as dextrorotary [(+)-OXA] or levorotary [(-)-OXA]. Crystal structures of the parasite enzyme in complex with optically pure (+)-OXA and (-)-OXA) reveal their absolute configurations as S- and R-, respectively. When tested in vitro, S-OXA demonstrated the bulk of schistosomicidal activity, while R-OXA had antischistosomal effects when present at relatively high concentrations. Crystal structures R-OXA•SmSULT and S-OXA•SmSULT complexes reveal similarities in the modes of OXA binding, but only the S-OXA enantiomer is observed in the structure of the enzyme exposed to racemic OXA. Conclusions/Significance Together the data suggest the higher schistosomicidal activity of S-OXA is correlated with its ability to outcompete R-OXA binding the sulfotransferase active site. These findings have important implications for the design, syntheses, and dosing of new OXA-based antischistosomal compounds., Author Summary Schistosomes, parasites that cause the disease schistosomiasis in humans, are blood flukes that infect an estimated 200 million people in 76 countries. Control of schistosomiasis is currently based on repeated doses of the drug praziquantel (PZQ). Parasites showing reduced susceptibility to PZQ have been recovered from patients that failed PZQ treatment and have been obtained by experimental selection. New anti-schistosomal drugs are therefore needed that can be used with PZQ to minimize the probability of resistance. The older anti-schistosomal drug oxamniquine (OXA) has an excellent efficacy and safety record but is only active against one of the three species infecting humans. Recently, a combination of genetic and structural analyses resulted in the determination of the structure of OXA in complex with its target enzyme in the parasite, providing opportunity for structure-guided modifications of OXA to make it more effective against all three schistosome species. Synthesis of OXA results in a racemic mixture. Here, we isolate OXA enantiomers and find that one is more effective than the other at killing schistosomes. Crystal structures of both OXA enantiomers bound to the target enzyme suggest a molecular basis for this observation that should be considered in ongoing and future OXA-based drug design efforts.

The lipophilicity of some cardiovascular drugs was determined by capillary electrophoresis (CE). Mexiletine, amlodipine and indapamide, the drugs considered, were in contact with liposomial vescicles for 2, 4 or 6 h. After the contact time the drugs, penetrated into liposomial vesicles, were determined by CE using phosphate buffer (pH 6.3 or 7.4) or borate buffer (pH 9). The lipophilicity of three drugs was determined considering the drug percentage penetrated into liposomial vesicles. The found lipohilicity order was amlodipine > mexiletine > indapamide.

The lipophilicity of pipemidic, nalidixic and oxolinic acids was determined by forming phospholipidic micelles directly in an electrophoretic capillary. Phosphatidylcholine derivatives, namely l -α-dilauroyl phosphatidylcholine (DLPC) or l -α-dimiristoyl phosphatidylcholine (DMPC), were added in the run buffer (50 mM phosphate buffer at pH 7.4). To obtain a mixed micelle, phospholipidic derivatives and sodium cholate were together added in the run buffer. Considering the increasing of migration time when phosphatidylcholine derivative is added in the run buffer, Ks can be determined and then quinolones lipophilicity.

Midodrine hydrochloride is a peripheral α1-adrenoreceptor agonist that induces venous and arterial vasoconstriction. Midodrine, after oral or intravenous administration, undergoes enzymatic hydrolysis and releases deglymidodrine, a pharmacologically active metabolite. Midodrine and deglymidodrine have a chiral carbon in the 2-position. To investigate the bioactivity of racemates and enantiomers of the drug and metabolite, three chromatographic chiral stationary phases, Chiralcel OD-H, Chiralcel OD-R, and α1-AGP, were evaluated for enantiomeric resolution. Good enantioseparation of midodrine racemate was obtained using the Chiralcel OD-H column. This stationary phase was then used to collect separately the midodrine enantiomers. By alkaline hydrolysis of rac-midodrine and each separated enantiomer, rac-deglymidodrine and its enantiomers were prepared. The control of the enantiomeric purity was carried out by α1-AGP stationary phase, while the hydrolysis of rac-midodrine and its enantiomers was controlled by capillary electrophoresis using trimethyl-β-cyclodextrin as chiral selector. The pharmacological activity of the two racemates and the two enantiomeric pairs was tested in vitro on a strip of rabbit descending thoracic aorta. The tests continued that the activity of the drug and metabolite is due only to the (−)-enantiomer because neither of the (+)-enantiomers is active. Chirality 16:356–362, 2004. ©2004 Wiley-Liss, Inc.

The organochlorine pesticide γ-hexachlorocyclohexane (γ-HCH, lindane) and its non-insecticidal isomers α-, β-, and δ- continue to pose serious environmental and health concerns, although their use has been restricted or completely banned for decades. The present study reports the first results on the ability of two Arthrobacter strains, not directly isolated from a HCH-polluted site, to grow in a mineral salt medium containing α-, β-, or γ-HCH (100 mgl(-1)) as sole source of carbon. Growth of cultures and HCHs degradation by Arthrobacter fluorescens and Arthrobacter giacomelloi were investigated after 1, 2, 3, 4, and 7 days of incubation by enumerating colony forming units and GC with ECD detection, respectively. Both bacteria are able to metabolize the HCHs: A. giacomelloi is the most effective one, as after 72 h of incubation it produces 88 % degradation of α-, 60 % of β-, and 56 % of γ-HCH. The formation of possible persistent compounds was studied by GC/MS and by HPLC analysis. Pentachlorocyclohexenes and tetrachlorocyclohexenes have been detected as metabolites, which are almost completely eliminated after 72 h of incubation, while no phenolic compounds were found.

Humic acids are able to interact with a wide range of pollulants and can influence their solubility, transport and bioavailability. In order to study the interaction between polar aromatic hydrocarbons and these macromolecules an affinity capillary electrophoretic method, the Hummel-Dreyer in its modified version, has been used. Two acidic metabolites of Phenanthrene:1-hydroxy-2-naphthoic acid (1-HNA) and 3,4-dihydroxybenzoic acid (3,4-DBA) were studied. The analysis for the binding studies were carried out by injecting a solution of HA in 25 mM ammonium hydrogen carbonate buffer (pH 9) into an uncoated fused silica capillary, filled with buffer solutions of 1-HNA or 3,4-DBA in varying and increasing amounts. The results obtained indicate that both compounds bind to HA, as had been confirmed by dialysis experiments and literature data. Capillary Electrophoresis proved to be a useful technique for investigating the link between xenobiotics and environmental macromolecules.

Diltiazem (DTZ) is an optically active calcium channel blocker having a benzodiazepine structure. The drug used in therapy is (+)-cis-diltiazem with configuration (2S,3S). To describe the analytical profile of DTZ different stationary phases (RP-18, RP-8, monolithic support) were tested. The best separation of DTZ from A, B, E and F was obtained using as stationary phase a RP-8 or a monolithic RP-18. The characterization of impurities was carried out using two analytical systems, HPLC and HPLC/MS.

Pethidine, predominantly a u-receptor agonist, is a phenyl-piperidinic synthetic drug. It is used in the management of moderate to several pain. A possible hydrolytic degradation of an ester group can generate a very toxic compound, the N-methyl-4-1,2,3,6 tetrahydropyridine (MPTP) which contaminates the drug. Because of the toxicity of MPTP a suitable method for its determination must be selective and sensitive. Afterwards we propose simple methods to determine pethidine and MPTP by capillary electrophoresis (CE), MECK and RP-high performance liquid chromatography (HPLC) looking at the limit of detection obtained using these three techniques. CE was carried out using as running buffer ammonium acetate (pH 8.3). MECK was performed with a borate buffer (pH 8.3) containing sodium dodecylsulphate and trimethyl-beta-cyclodextrins. RP-HPLC was carried out on a RP18 stationary phase, using as mobile phase a mixture of phosphate buffer (ph7) containing acetonitrile and 1% of diethylamine. 2003 Elsevier Science B.V. All rights reserved.

Capillary electrophoresis (CE) and capillary electrochromatography (CEC) have been used to determine losartan and hydrochlorothiazide. The CE separation was carried out in an uncoated capillary filled with a 100 mM sodium borate pH 9 solution containing trimethyl-beta-cyclodextrin. CEC was performed using a capillary packed with a RP-18 stationary phase. The mobile phase was a mixture of 50 mM ammonium acetate pH 7, water and acetonitrile (1/1.5/7.5). By CE and CEC suitable methods to determine simultaneously losartan and hydrochlorothiazide in working standard mixture or pharmaceutical form were obtained. The proposed methods are very simple and both gave accurate and precise results.

The quality control of drugs is generally made by HPLC. This control could be made also by capillary electrochromatography (CEC). In this paper we report the analysis by CEC of ibuprofen, a well-known anti-inflammatory non steroidic drug, and some of its impurities. The analyses were performed in a 100-microm inner diameter (I.D.) fused silica capillary, packed with RP-18 stationary phase. The mobile phase was a mixture of 100 mM formic acid solution (pH 2.5), water and acetonitrile (ACN). The ACN percentage in the mobile phase and the applied voltage were carefully studied to well resolve the drug from each impurity. The results, obtained determining ibuprofen and related compounds by CEC, showed the selectivity and efficiency of the method.

Fenticonazole is a chiral antifungal agent, used in therapy as the racemic mixture. The investigation on the chirality of fenticonazole is reported in this study. rac-Fenticonazole was resolved by HPLC and by capillary electrophoresis (CE). The chiral stationary phase (CSP), used in HPLC, was Daicel OD-H, a commercial phase, which allowed the separate collection of the two enantiomers. The chiral selectors used for CE were some cyclodextrin derivatives. The analysis time required from CE was about the half the HPLC enantioseparation time. The biological activity of the rac-mixture and each individual enantiomer was tested against Cryptococcus neoformans and two Aspergillus nidulans strains. The minimum inhibitory concentration (MIC) evaluation showed that the eutomer was the enantiomer chromatographically more retained and had a longer migration time in the electrophoretic enantioseparation. The CD spectrum of the eutomer showed a positive Cotton effect. © 2002 Wiley-Liss, Inc.

Fenticonazole, a topical antifungal agent containing a stereogenic centre, is used in therapy as a racemic mixture. Five related compounds can be found as impurities in the drug. We propose a HPLC method using as stationary phase a RP-8 column eluted with different gradients of acetonitrile/phosphate buffer (pH 6) for simultaneous determination of the drug and these impurities. We also studied a high performance capillary electrophoresis (HPCE) method for quality control of fenticonazole. The separation of fenticonazole from all its impurities by HPCE was obtained in a relative short capillary (40 cm, effective length 34 cm, 50 μm ID) with a running buffer of 30 mM phosphate (pH 3) containing 8 mM trimethyl-β-cyclodextrin. Under these experimental conditions very good separation of fenticonazole from each individual impurity is obtained in less than 20 minutes. The choice of cyclodextrin added to the running buffer was dictated by the chiral nature of fenticonazole; trimethyl-β-cyclodextrin is mainly used as chiral selector. The optimised HPLC and HPCE methods are compared.

Silybine (SBN), isosilybine (ISBN), silycristine (SCN), silydianine (SDN), and taxifoline (TXF) are the main active flavanoids, generally found in the dried fruits of silybum marianum. The concentrations of these compounds, excepted TXF, are all together usually expressed as silymarine content. In this paper the determination of the silymarine titre was made by high performance liquid chromatography (HPLC), and high performance capillary electrophoresis (HPCE). Two reversed stationary phases, RP-18 and RP-8, were observed comparing the resolutions of all considered flavanoids with each stationary phases. The HPCE was carried out considering the possible improvement in the resolution of SBN, CN, SDN and TXF using, 8-cyclodextrines or organic modifier. The qualitative and quantitative data obtained by HPLC and HPCE were compared.