ß-Cyclodextrin Derivative Grafted on Silica Gel Represents a New Polymeric Sorbent for Extracting Nitisinone from Model Physiological Fluids.

Nitisinone (NTBC) is used in the treatment of disorders affecting the tyrosine pathway, including hereditary tyrosinemia type I, alkaptonuria, and neuroblastoma. An inappropriate dosage of this therapeutic drug causes side effects; therefore, it is necessary to develop a rapid and sensitive method to monitor the content of NTBC in patients' blood. This study aimed to develop anew polymeric sorbent containing ß-cyclodextrin (ß-CD) derivatives grafted on silica gel to effectively extract NTBC from model physiological fluids. The inclusion complex formed between ß-CD and NTBC was examined by proton nuclear magnetic resonance spectroscopy. The novel sorbents with derivatives of ß-CD were prepared on modified silica gel using styrene as a comonomer, ethylene glycol dimethacrylate as a crosslinking agent, and 2,2'-azo-bis-isobutyronitrile as a polymerization initiator. The obtained products were characterized via Fourier transform infrared spectroscopy and then used as sorbents as part of a solid phase extraction technique. High NTBC recovery (70%indicated that the developed polymeric sorbent may be suitable for extracting this compound from patients' blood samples.

Tannins and Tannin-Related Derivatives Enhance the (Pseudo-)Halogenating Activity of Lactoperoxidase.

Several hydrolyzable tannins, proanthocyanidins, tannin derivatives, and a tannin-rich plant extract of tormentil rhizome were tested for their potential to regenerate the (pseudo-)halogenating activity, i.e., the oxidation of SCN- to hypothiocyanite -OSCN, of lactoperoxidase (LPO) after hydrogen peroxide-mediated enzyme inactivation. Measurements were performed using 5-thio-2-nitrobenzoic acid in the presence of tannins and related substances in order to determine kinetic parameters and to trace the LPO-mediated -OSCN formation. The results were combined with docking studies and molecular orbital analysis. The -OSCN-regenerating effect of tannin derivatives relates well with their binding properties toward LPO as well as their occupied molecular orbitals. Especially simple compounds like ellagic acid or methyl gallate and the complex plant extract were found as potent enzyme-regenerating compounds. As the (pseudo-)halogenating activity of LPO contributes to the maintenance of oral bacterial homeostasis, the results provide new insights into the antibacterial mode of action of tannins and related compounds. Furthermore, chemical properties of the tested compounds that are important for efficient enzyme-substrate interaction and regeneration of the -OSCN formation by LPO were identified.

Nutrition effects on the biofilm immobilization and 3,5-DNBA degradation of Comamonas testosteroni A3 during bioaugmentation treatment.

The survival of inoculated microbes is critical for successful bioaugmentation in wastewater treatment. The influence of readily available nutrients (RANs) on the colonization of two functional bacteria, Pseudomonas putida M9, a strong biofilm-forming strain, and Comamonas testosteroni A3, a 3,5-dinitrobenzoic acid (3,5-DNBA)-degrading strain, in biofilms was studied with 3,5-dinitrobenoic acid synthetic wastewater (DCMM) complemented with various ratios of Luria-Bertani broth (LB). With the increase in LB rate, the biofilm biomass was increased, the percentage of gfp-labeled M9 measured in the mixed culture enhanced, and also M9 became dominant. In laboratory-scale sequencing batch biofilm reactors, with the increase in 3,5-DNBA concentration and extension of the running time, the 3,5-DNBA removal in DCMM wastewater complemented with RANs tended to be more efficient and its removal rates increased gradually over the experimental period. Our study demonstrated that supplementing RANs could be a useful strategy for enhancing colonization of degrading bacteria in wastewater treatment systems.

Isolation of a nitrobenzoate from the leaves of Cinnamomum tenuifolium.

Investigation of leaf extract of Cinnamomum tenuifolium (Lauraceae) led to the isolation of one new nitrobenzoate, ethyl 3,5-dihydroxy-4-nitrobenzoate (1), along with four known compounds. This is the first report of Cinnamomum metabolites with a nitro benzenoid derivative. Their structures were determined through in-depth spectroscopic and mass spectrometric analyses.

Monitoring tyrosinaemia type I: Blood spot test for nitisinone (NTBC).

BACKGROUND: Quantification of nitisinone, 2-(nitro-4-trifluoromethylbenzoyl)1,3-cyclohexanedione (NTBC) has been repeatedly described. Nevertheless monitoring of NTBC has not yet become part of routine therapy surveillance in tyrosinaemia type I (OMIM 276700). We developed a blood spot test to facilitate collection and transport of samples. Furthermore, the test material can be used for determination of other parameters like tyrosine and succinylacetone. METHOD: For quantification of NTBC in blood spots filter paper discs of 3.2mm diameter were extracted with 150 µL methanol containing mesotrione as internal standard (IS). Analysis was done by UPLC-MS/MS on a Xevo mass spectrometer (ESI+), (MRM). Parent ions were 330.05 for NTBC and 340.05 for IS, daughter ions were m/z 217.95 and m/z 125.95 for NTBC, and m/z 227.95 and m/z 103.95 for IS. RESULTS: The calibration curve for NTBC in blood spots was linear from 0.1 µmol/L to 100 µmol/L. Recovery exceeded 73.1%, CV intraday and interday were below 9.6%. Instrumental run time was 2.5 min. Sensitivity of the method was 0.1 µmol/L. NTBC concentrations in plasma were higher than in blood spots by a factor of 1.56 ± 0.13. CONCLUSION: As demonstrated in patients with tyrosinaemia type I quantification of NTBC by UPLC-MS/MS in blood spots is feasible and gives valuable information for monitoring NTBC treatment.

HPLC with polysaccharide chiral stationary phase in polar-organic phase mode: application to the asymmetric epoxidation of allylic alcohols.

A simple and rapid HPLC method using a polysaccharide-based chiral stationary phase (Chiralpak AD-H) in polar-organic phase mode has been developed for direct resolution of glycidyl nitrobenzoate (GNB) and 2-methyl glycidyl nitrobenzoate (MGNB) enantiomers. ACN and methanol were used as mobile phase and the effects of the addition of ethanol and 2-propanol as organic modifier in the mobile phase, flow rate and the column temperature were tested. The optimized conditions were: methanol/ethanol (80:20) at a flow rate of 0.9 mL/min and 40 degrees C. Analysis time was < or = 13 min and the chiral resolution was > or = 2. The method was validated and resulted to be selective, precise and accurate. The method was found to be linear in 2-300 microg/mL range (R(2) >0.999) with an LOD nearly 0.5 microg/mL for four enantiomers. GNB and MGNB enantiomers were obtained by asymmetric epoxidation of allyl alcohol and 2-methyl allyl alcohol, respectively, using chiral titanium-tartrate complexes as catalyst and dichloromethane as solvent after in situ derivatization of the intermediate glycidols derivatives. The quite simple and rapid validated method was applied successfully for direct determination of the enantiomeric excess (> or = 90%) and yield obtained in real samples of asymmetric epoxidation of allylic alcohols without further purification, workup or solvent removal. The method provides a useful and value-added tool for controlling the enantiomeric purity of the synthesized epoxides.

Isolation and preliminary characterization of a o-nitrobenzaldehyde-degrading Alcaligenes sp. ND1 / Isolamento e caracterização preliminar de lcaligenes SP ND1 degradador de o-nitrobenzaldeido

This paper reports the isolation and characterization of a new o-nitrobenzaldehyde (ONBA)-degrading bacterium, Alcaligenes sp. ND1. ND1 degraded almost all ONBA (100 mg L-1) in M9 medium within 36 hours. The key enzyme(s) involved in the initial biodegradation was a constitutively intracellular enzyme(s). This bacterium has great potential utility for bioremediation.

Esse trabalho relata o isolamento e a caracterização de uma nova bactéria degradadora de o-nitrobenzaldeido (ONBA), Alcaligenes sp ND1. A bactéria ND1 decompôs todo o ONBA (100 mg.L-1) do meio M9 em 36 horas. A enzima-chave envolvida na biodegradação inicial foi uma enzima constitutiva intracelular. Esta bactéria apresenta um potencial de aplicação para bioremediação.

Biomass characteristics in three sequencing batch reactors treating a wastewater containing synthetic organic chemicals.

The physical and biochemical characteristics of the biomass in three lab-scale sequencing batch reactors (SBR) treating a synthetic wastewater at a 20-day target solids retention time (SRT) were investigated. The synthetic wastewater feed contained biogenic compounds and 22 organic priming compounds, chosen to represent a wide variety of chemical structures with different N, P and S functional groups. At a two-day hydraulic retention time (HRT), the oxidation-reduction potential (ORP) cycled between -100 (anoxic) and 100 mV (aerobic) in the anoxic/aerobic SBR, while it remained in a range of 126+/-18 and 249+/-18 mV in the aerobic sequencing batch biofilm reactor (SBBR) and the aerobic SBR reactor, respectively. A granular activated sludge with excellent settleability (SVI=98+/-31 L mg(-1)) developed only in the anoxic/aerobic SBR, compared to a bulky sludge with poor settling characteristics in the aerobic SBR and SBBR. While all reactors had very good COD removal (>90%) and displayed nitrification, substantial nitrogen removal (74%) was only achieved in the anoxic/aerobic SBR. During the entire operational period, benzoate, theophylline and 4-chlorophenol were completely removed in all reactors. In contrast, effluent 3-nitrobenzoate was recorded when its influent concentration was increased to 5 mg L(-1) and dropped only to below 1 mg L(-1) after 300 days of operation. The competent (active) biomass fractions for these compounds were between 0.04% and 5.52% of the total biomass inferred from substrate-specific microbial enumerations. The measured competent biomass fractions for 4-chlorophenol and 3-nitrobenzoate degradation were significantly lower than the influent COD fractions of these compounds. Correspondent to the highest competent biomass fraction for benzoate degradation among the test SOCs, benzoate oxidation could be quantified with an extant respirometric technique, with the highest specific oxygen uptake rate (SOUR(benzoate), 0.026 g O2 h(-1) g(-1) XCOD) in the anoxic/aerobic SBR. These combined results suggest that operating SBRs with alternative anoxic/aerobic cycles might facilitate the formation of granular sludge with good settleability, and retain comparable removal of nitrogen and synthetic organic compounds. Hence, the practice of anoxic/aerobic cycling should be considered in wastewater treatment systems whenever possible.

Supported liquid membrane stability in chiral resolution by chemically and physically modified membranes.

In the present work some stability studies on Supported Liquid Membranes (SLMs) to be used for chiral separations were realized. In particular, primary aim was to determine how a modification of the support surface influences the SLM stability. First, the procedure for support modification was optimised, making a screening of various compounds (sulphuric acid, nitric acid, chromic acid, sodium dodecyl sulphate (SDS), glycerol, oleic alcohol, propylene glycol (PPG), bovine serum albumin (BSA)) and testing their performance by means of contact angle measurements. Next, a second screening was realized by permeation tests in a stirred cell. Finally, to compare the stability of modified with unmodified support in a process of interest for chemical and/or biochemical industries, some permeation tests for resolution of DNB-DL-Leucine were realized in a re-circulation system. Results showed a better surface hydrophilization of chemically modified support and better stability of the sulphonated support. However, in operating conditions a little high stability of the unmodified support was obtained.

Enantiomeric separations by capillary electrochromatography using a macrocyclic antibiotic chiral stationary phase.

Racemic mixtures of tryptophan and dinitrobenzoyl leucine have been successfully resolved by capillary electrochromatography (CEC) using the macrocyclic antibiotic teicoplanin, covalently bonded to a 5 microns silica support. Modification of a previously published packing procedure was required to pack reliable capillaries, capable of performing enantiomeric separations. Good levels of enantioselectivity were obtained in all cases, with optimised separations being performed in less than 6 min. Retention times, resolution and reproducibility are discussed.

Resolution of gram quantities of racemates by high-speed counter-current chromatography.

Gram quantities of (+/-)-dinitrobenzoyl amino acids were separated by high-speed counter-current chromatography (CCC) using N-dodecanoyl-L-proline-3,5-dimethylanilide as a chiral selector (CS). Standard and pH-zone-refining CCC techniques were compared. By using the standard technique, 10 mg to a maximum of 1 g of samples was resolved in 2-9 h simply by increasing the concentration of the CS in the stationary phase. By using pH-zone-refining CCC, even more sample (2 g) was efficiently separated in less time (3 h). In both techniques, leakage of CS from the column was negligible. The method requires no solid support and the same column can be used repeatedly to separate a variety of enantiomers by dissolving appropriate chiral selectors in the stationary phase.

Capillary zone electrophoresis study of naphthylethylcarbamoylated beta-cyclodextrins.

Regiospecifically monosubstituted 1-(1-naphthyl)ethylcarbamoylated beta-cyclodextrins (NEC-beta-CDs) were successfully employed as chiral additives in capillary zone electrophoresis (CZE) to achieve chiral separation of N-(3,5-dinitrobenzoyl)phenylglycine (3,5-DNB-PG), phenylalanine (3,5-DNB-PA), and homophenylalanine (3,5-DNB-HPA). The enantioselectivity of the various site-substituted NEC-beta-CDs in CZE was compared with that of native beta-CD. Complexation constants of the three 3,5-DNB amino acids with beta-CD were determined from the CZE results: 3,5-DNB-L-HPA, 473 +/- 9 M-1; 3,5-DNB-D-HPA, 460 +/- 10 M-1; 3,5-DNB-L-PA, 260 +/- 4 M-1; 3,5-DNB-D-PA, 161 +/- 3 M-1; and 3,5-DNB-D, L-PG, 43 +/- 4 M-1.

Chromatographic separation of the diastereomers of a dihydropyridine-type calcium channel antagonist as the bis-3,5-dinitrobenzoates.

The diastereomeric components of the calcium channel antagonist RS-93522-004 are separated as the bis-3,5-dinitrobenzoate ester by reversed-phase high-performance liquid chromatography. The diastereomer ratio determination is shown to be precise, accurate and sensitive, and is not affected by the reaction yield of the derivatization with 3,5-dinitrobenzoyl chloride. The four individual stereoisomers of RS-93522-004 were independently shown to have equal reactivity toward 3,5-dinitrobenzoyl chloride.

[Contribution to the standardization of results in paper and thin-layer chromatography (author's transl)]. / Beitrag zur Standardisierung der Ergebnisse in der Papier-und Dünnschichtchromatographie.

Chromatographic indexes that facilitate a better reproducibility of paper chromatographic results have been proposed, using the homologous series of coloured derivatives of secondary amines as standard substances. When using dimethylformamide as the stationary phase the value of the indexes proved to be independent of the concentration of the impregnating solution of the stationary phase, of the temperature changes during the chromatographic process, of the ways of impregnation and of the skills of workers. It depends only on the type of the stationary phase and on the composition of the mobile phase. The obtained indexes can contribute to a better reproducibility of RF values and to an easier and more objective identification of substances.