A novel bioassay for screening and quantification of taxanes

We report a novel method for screening and quantifying tubulin-binding antimitotic agents that promote microtubule formation. The method is based on the shift in the peak of the fluorescence emission difference spectrum of tubulin complexed to metal free tetrakis(4-carboxyphenyl)porphyrin (TCPP) in the presence of antimitotic agents. Taxol (paclitaxel), an anti-tumor drug approved for the treatment of a variety of cancers, caused the appearance of a new fluorescence peak at 645 nm at concentrations as low as 125 nM, the intensity of which was a function of the paclitaxel concentration. Cephalomannine caused the induction of a new fluorescence peak at 651 nm only above 1 muM. Baccatin did not induce the appearance of any new peak within detectable operating measurement conditions. These observations are in accordance with the biological activities/cytotoxicities of these compounds. Accordingly, it is proposed that the new method can be used for high throughput screening of antimitotic compounds.     

Über triterpene—XXIX : Zur struktur des avenacins

From the roots of oat (Avena sativa) a second antibiotic active glycoside, avenacine B, was isolated in addition to the main glycoside avenacine A. In the aglycone of A the double bond could not be established contrary to the statement in the literature but an aldehyde group in addition to the keto group was identified and therefore the proposed structure has to be revised. In the aglycone of B the —CH₂OH-group of A has been substituted by CH₃. The structures of the sugar chains in both glycosides have been determined, in avenacine A as [β-d-gluco-pyranosyl(1 → 4)-[β-d-glucopyranosyl (1 → 2)]-α-l-arabinosyl (1)- and in B as [β-d-glucopyranosyl (1 → 4)]-β-d-glucopyranosyl (1)-.     

Photometric determination of trivalent gallium,indium and thallium with Xylenol orange

Summary Xylenol orange reacts very sensitively with gallium(III), indium (III) and thallium(III) to form reddish violet colored chelates having _max 560 nm in case of Ga and In and _max 590 nm in case of Ti at P_H 4.0. The molar ratio for all the chelates is 1 1 (metal reagent). Optimum conditions including the range for adherence to Beer's law, effect of P_H on the color intensity, effect of excess reagent, and sensitivity are reported for the photometric determination of these metal ions using Xylenol orange.

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Zusammenfassung Xylenolorange reagiert sehr empfindlich mit Gallium(III), Indium(III) und Thallium(III) unter Bildung rötlich-violetter Chelate mit einem Absorptionsmaximum bei 560 nm im Falle von Ga und In bzw. 590 nm für Tl bei p_H 4,0. Das Molverhältnis ist in jedem Fall 11. Die besten Arbeitsbedingungen, der Gültigkeitsbereich des Beerschen Gesetzes, der Einfluß des P_H auf die Farbintensität und des Reagensüberschusses sowie die Empfindlichkeit werden für die photometrische Bestimmung der genannten Ionen angegeben.

     

Role of palladium in the redox electrochemistry of ferrocene monocarboxylic acid encapsulated within ORMOSIL networks

We report herein the effect of palladium on the redox electrochemistry of ferrocene monocarboxylic acid encapsulated within an organically modified sol-gel glass network (ORMOSIL). It has been found that amount of palladium and its geometrical distribution significantly alter the redox electrochemistry of FcMCA. The geometrical distribution of palladium has been controlled by two methods: (i) palladium is allowed to link within nanostructured network of the ORMOSIL which was subsequently availed from the reactivity of palladium chloride and trimethoxysilane; (ii) palladium powder is encapsulated together FcMCA thus allowing the presence of palladium within the nanoporous domain. The content of palladium is varied by controlling the reaction dynamics of palladium chloride and trimethoxysilane interaction. For this we initially allowed to trigger hydrolysis, condensation and poly-condensation of trimethoxysilane and dimethyldiethoxysilane in acidic medium and subsequently partially dried ORMOSIL film was allowed to interact with palladium chloride. Even with partially dried ORMOSIL derived from trimethoxysilane and dimethyldiethoxysilane undergoes rapid interaction with palladium chloride and the transparent color of ORMOSIL changed to a black colour due to the formation of palladium silicon linkage. The palladium-silicon linkage has been identified by NMR, UV-VIS and transmission electron spectroscopy. The electrochemistry of FcMCA encapsulated within such an ORMOSIL matrix has been studied. Excellent redox electrochemistry of ferrocene monocarboxylic acid having peak potential separation tending to 0 for a multilayered electrode was investigated. The palladium content has been found to affect the redox electrochemistry of ferrocene as well as electrocatalytic efficiency of new ORMOSIL material. The electroanalysis of NADH is reported. The modified electrode is very sensitive to NADH with lowest detection limit of < 1 microM.     

Hydrophobic interaction chromatography coupled with atomic fluorescence spectrometric detection Effect of the denaturation on the determination of thiolic proteins

Hydrophobic interaction chromatography coupled online with chemical vapour atomic fluorescence spectrometry (HIC-CVGAFS) has been optimized for the analysis of thiolic proteins in denaturing conditions. Proteins are pre-column simultaneously denatured and derivatized in phosphate buffer solution containing 8.0 mol dm⁻³ urea and p-hydroxymercurybenzoate (PHMB) and the derivatized denatured proteins are separated on a silica HIC Eichrom Propyl column in the presence of 8.0 M urea in the mobile phase. Post-column online reaction of derivatized denatured proteins with bromine, generated in situ by KBr/KBrO₃ in HCl medium, allowed the fast conversion of the uncomplexed PHMB and of the PHMB bound to proteins to inorganic mercury also in presence of urea. Hg²⁺, present in solution as Hg²⁺-urea complex, is selectively detected by AFS in a Ar/H₂ miniaturized flame after sodium borohydride reduction to Hg. Under optimized conditions, online bromine treatment gives a 100±2% recovery of both free and protein-complexed PHMB. Denatured glyceraldehyde-3-phosphate dehydrogenase, aldolase, lactate dehydrogenase, trioso phosphate isomerase and β-lactoglobulin have been examined. As the sensitivity and limit of detection of proteins in the HIC-CVGAFS apparatus depends on number of SH groups reacting with PHMB, the denaturation process, which increases the number of PHMB-reactive thiolic groups in proteins, improves the analytical performances of the described system in protein analysis. The detection limit for the denatured proteins examined was found in the range of 10⁻¹⁰-10⁻¹² mol dm⁻³, depending on the considered protein, with linear calibration curves spanning over four decades of concentration.     

Ternary complexes of copper(II), nickel(II) and zinc(II) with nitrilotriacetic acid as a primary ligand and some phenolic acids as secondary ligands

Summary The formation of ternary complexes of the MAL^3– type [where M = Cu^II, Ni^II and Zn^II ; A = nitrilotriacetic acid (NTA); L = 1-hydroxy-2-naphthoic acid (1,2 HNA) and 2-hydroxy-1-naphthoic acid (2,1 HNA)] have been studied potentiometrically in 50% v/v aqueous — ethanol (25° and µ = 0.1). Under identical conditions the binary complexes of the 1,2- and 2,1-HNA ligands have also been examined. The values of mixed ligand formation constants K_MAL have been found to be lower than K_ML (first step formation constant of binary complexes) and even less than (second step formation constant of binary complexes).     

Pharmacokinetics of ICG and HPPH-car for the detection of normal and tumor tissue using fluorescence, near-infrared reflectance imaging: a case study

We present in vivo fluorescent, near-infrared (NIR), reflectance images of indocyanine green (ICG) and carotene-conjugated 2-devinyl-2-(1-hexyloxyethyl) pyropheophorbide (HPPH-car) to discriminate spontaneous canine adenocarcinoma from normal mammary tissue. Following intravenous administration of 1.0 mg kg-1 ICG or 0.3 mg kg-1 HPPH-car into the canine, a 25 mW, 778 nm or 70 mW, 660 nm laser diode beam, expanded by a diverging lens to approximately 4 cm in diameter, illuminated the surface of the mammary tissue. Successfully propagating to the tissue surface, ICG or HPPH-car fluorescence generated from within the tissue was collected by an image-intensified, charge-coupled device camera fitted with an 830 or 710 nm bandpass interference filter. Upon collecting time-dependent fluorescence images at the tissue surface overlying both normal and diseased tissue volumes, and fitting these images to a pharmacokinetic model describing the uptake (wash-in) and release (wash-out) of fluorescent dye, the pharmacokinetics of fluorescent dye was spatially determined. Mapping the fluorescence intensity owing to ICG indicates that the dye acts as a blood pool or blood persistent agent, for the model parameters show no difference in the ICG uptake rates between normal and diseased tissue regions. The wash-out of ICG was delayed for up to 72 h after intravenous injection in tissue volumes associated with disease, because ICG fluorescence was still detected in the diseased tissue 72 h after injection. In contrast, HPPH-car pharmacokinetics illustrated active uptake into diseased tissues, perhaps owing to the overexpression of LDL receptors associated with the malignant cells. HPPH-car fluorescence was not discernable after 24 h. This work illustrates the ability to monitor the pharmacokinetic delivery of NIR fluorescent dyes within tissue volumes as great as 0.5-1 cm from the tissue surface in order to differentiate normal from diseased tissue volumes on the basis of parameters obtained from the pharmacokinetic models.     

Kinetics and mechanism of oxidation of methanol by acid bromate

The kinetics of oxidation of methanol by bromate ion in hydrochloric acid medium has been investigated. A mechanism consistent with the experimental observations is suggested.

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Analysis of fragrances in cosmetics by gas chromatography–mass spectrometry

A gas chromatographic (GC)-mass spectrometric (GC-MS) method has been developed for the routine analysis of 11 fragrance substances in cosmetics: cinnamic alcohol, cinnamic aldehyde, eugenol, hydroxy citronellal, α-amyl cinnamic aldehyde, geraniol, isoeugenol, coumarin, dihydrocoumarin, citronellal and citral. Methods for sample preparation of various types of cosmetic products, prior to GC analysis, have also been developed and proved to be rugged. Detection limits of all of target fragrance substances were approximately 1 ppm. Calibration curves of the target fragrance substances analyzed by GC were found to be linear in the investigated concentration range, 0.005% – 0.50%. The recoveries of the target fragrances from various types of cosmetic products were 80% – 116% and the relative standard deviations of the quantitative analysis of the target fragrance substances were within 5%.     

Development of a bionematicide with Paecilomyces lilacinus to control Meloidogyne incognita

Root-knot disease caused by Meloidogyne incognita is a matter of grave concern because it affects several economically important crop plants. The use of solid-state fermentation (SSF) may help to elaborate efficient formulations with fungi to be employed in the biologic control of nematodes. Attempts were made to select low-cost substrates for spore production of a strain of Paecilomyces lilacinus with known nematicide capacity. Coffee husks, cassava bagasse, and defatted soybean cake were utilized as substrates, and sugarcane bagasse was used as support. Fermentations were carried out in flasks covered with filter paper at 28°C for 10 d. The products obtained by SSF were evaluated for their nematicide activity in pot experiments containing one seedling of the plant Coleus inoculated with the nematode M. incognita. The plants were evaluated 2 mo after inoculation. Fermented products showed a reduction in the number of nematodes. The best results were obtained with defatted soybean cake, which showed almost 100% reduction in the number of nematodes; the reduction with coffee husk was 80% and with cassava bagasse was about 60%.