Activity of anchored human matrix metalloproteinase-1 catalytic domain on Au (111) surfaces monitored by ESI-MS

Matrix metalloproteinases (MMPs) are a family of Zn-dependent endo-peptidases known for their ability to cleave several components of the extracellular matrix, but which can also cleave many non-matrix proteins. There are many evidences that MMPs are involved in physiological and pathological processes, and a huge effort has been put in the development of possible inhibitors that could reduce the activity of MMPs, as it is clear that the ability to monitor and control such activity plays a pivotal role in the search for potential drugs aimed at finding a cure for several diseases such as pulmonary emphysema, rheumatoid arthritis, fibrotic disorders and cancer.A powerful method currently available to study enzyme-inhibitor interactions is based on the use of the surface plasmon resonance (SPR) technique. When MMP interactions are studied, a procedure by which inhibitors are normally anchored on sensor chips and SPR technique is used in order to study their interaction with MMPs molecules is usually followed. This is because it is currently believed that MMPs cannot be anchored on the sensor-chip surface without losing their activity. However, this approach gives rise to problems, as the anchoring of low-molecular-weight inhibitors on gold surfaces easily affects their ability to interact with MMPs. For this reason, the anchoring of MMPs is highly desirable.A new experimental protocol that couples the Fourier transform-SPR (FT-SPR) technique with electrospray ionization-mass spectroscopy (ESI-MS) is described here for the evaluation of the activity of MMP-1 catalytic domain (cdMMP-1) anchored on gold surfaces. The cdMMP-1 surface coverage is calculated by using FT-SPR and the enzyme activity is estimated by ESI-MS. The proposed method is label-free.     

New insights into the acid-promoted reaction of caffeic acid and its esters with nitrite: decarboxylation drives chain nitrosation pathways toward novel oxime derivatives and oxidation/fragmentation products thereof

In 0.05 M acetate buffer, pH 4, containing 1% methanol, caffeic acid (1a) (2 x 10(-3) M) reacted smoothly with nitrite (NO(2)(-)) (4 x 10(-3) M) to afford as main products the novel 2-hydroxy- and 2-methoxyaldoximes 7a,b, the 2-oxoaldoxime 9a, 3,4-dihydroxybenzoic acid, 3,4-dihydroxybenzaldehyde, and the known furoxan 3c and benzoxazinone 4b in smaller amounts. At lower 1a concentration (e.g., 1 x 10(-4) M), 7a was the main product, whereas with 0.1 M 1a and 0.5 M NO(2)(-) 3c and 9a were prevailing. At pH 2, 7a was still the most abundant product, together with 3,4-dihydroxybenzaldehyde and some 9a, whereas at pH 1 9a and 3,4-dihydroxybenzaldehyde were formed in higher yields. No evidence for ring nitration products, including the previously reported 4,5-dihydroxy-2-nitrobenzaldehyde, was obtained. At 2 x 10(-3) M concentration and at pH 4, caffeic acid methyl ester (1b) reacted with NO(2)(-) chiefly via ring nitration and/or dimerization to give 5a, the novel nitrated neolignan derivative 10, and the parent 6. Chlorogenic acid (1c) afforded only the ring nitrated derivative 5b. A unifying mechanism for the reaction of 1a and its esters with NO(2)(-) is proposed involving reversible formation of nitroso intermediates via chain nitrosation at the 2-position of the (E)-3-(3,4-dihydroxyphenyl)propenoic system. In the case of 1a, decarboxylation would drive the nitroso intermediates toward the formation of oximes 7a,b and 3c, reflecting nucleophilic addition of water, methanol, and NO(2)(-), and their oxidation or breakdown products, viz. 9a, 3,4-dihydroxybenzaldehyde, 3,4-dihydroxybenzoic acid, and the benzoxazinone 4b. In the case of esters 1b,c, to which decarboxylation is precluded, ring nitration or dimerization become the favored routes, triggered by preliminary oxidation at the catechol moiety.     

Feasibility of employing permanent chemical modifiers for the determination of cadmium in coal using slurry sampling electrothermal atomic absorption spectrometry

Iridium and ruthenium, alone and in combination with tungsten, thermally deposited on the platform of a transversely heated graphite tube, were investigated for their suitability as permanent chemical modifiers for the determination of cadmium in coal slurries by electrothermal atomic absorption spectrometry (ET AAS). The conventional mixed palladium and magnesium nitrates (Pd–Mg) modifiers, added in solution, were also investigated for comparison. The latter one showed the best performance for aqueous solutions, and the mixed W–Ir and W–Ru permanent modifiers had the lowest stabilizing power. All of the investigated modifiers lost some of their stabilizing power when coal slurries were investigated. The Pd–Mg modifier, pure Ir and Ru, and a mixture of 300 μg W + 200 μg Ir could stabilize Cd at least to a pyrolysis temperature of 600 °C, whereas all the other combinations already failed at temperatures above 500–550 °C. Additional investigations of the supernatant liquid of the slurries supported the assumption that the high acid concentration of the slurries and/or a concomitant leaching out of the coal might be responsible for the reduced stabilizing power of the modifiers. The maximum applicable pyrolysis temperature of 600 °C was not sufficient to reduce the background absorption to a manageable level in the majority of the coal samples. High-resolution continuum source ET AAS revealed that the continuous background absorption was exceeding values of A = 2, and was overlapping with the analyte signal. Although the latter technique could correct for this background absorption, some analyte was apparently lost with the rapidly vaporizing matrix so that the method could not be considered to be rugged. A characteristic mass of 1.0 pg and a detection limit of 0.6 ng g^− 1 could be obtained under these conditions.     

Efficient synthesis of an enantiopure beta-lactam as an advanced precursor of thrombin and tryptase inhibitors

A new and efficient synthesis of a beta-lactam that is an advanced precursor of inhibitors of thrombin and tryptase is reported. The reaction sequence is based on the use of an inexpensive enantiomerically pure starting material and is designed to allow access to both enantiomers of the target molecules by epimerization of a side-product obtained along the synthesis. An improved procedure for the epimerization step that takes advantage of the use of a polymer-supported and recyclable phase-transfer catalyst is described.     

Development of an integrated capillary electrophoresis/sensor for L-ascorbic acid detection

A CE/biosensor for measuring ascorbic acid was developed by coupling a polyaniline optical sensor and capillary electrophoresis (CE). The capillary column was partially modified with a thin film of polyaniline redox sensitive material. Ascorbic acid was detected by monitoring the changes in optical absorbance occurring to the polyaniline film upon the reduction reaction. The sensor response (change in optical absorbance at 650 nm) is proportional to the concentration of ascorbic acid over a range of 2.5-250 mg/L and the response range has shown a clear dependence on the characteristics of the polymerized film. High specificity and sensitivity of the present method, low sample consumption, short times of response (ca. 2 min) and the reproducibility of the results demonstrate that the CE/polyaniline-sensor could be further employed in the study of the relation between the content of L-ascorbic acid in body fluids and clinical parameters, e.g., cell ageing.     

QSAR Models for Human Carcinogenicity: An Assessment Based on Oral and Inhalation Slope Factors

Carcinogenicity is a crucial endpoint for the safety assessment of chemicals and products. During the last few decades, the development of quantitative structure–activity relationship ((Q)SAR) models has gained importance for regulatory use, in combination with in vitro testing or expert-based reasoning. Several classification models can now predict both human and rat carcinogenicity, but there are few models to quantitatively assess carcinogenicity in humans. To our knowledge, slope factor (SF), a parameter describing carcinogenicity potential used especially for human risk assessment of contaminated sites, has never been modeled for both inhalation and oral exposures. In this study, we developed classification and regression models for inhalation and oral SFs using data from the Risk Assessment Information System (RAIS) and different machine learning approaches. The models performed well in classification, with accuracies for the external set of 0.76 and 0.74 for oral and inhalation exposure, respectively, and r² values of 0.57 and 0.65 in the regression models for oral and inhalation SFs in external validation. These models might therefore support regulators in (de)prioritizing substances for regulatory action and in weighing evidence in the context of chemical safety assessments. Moreover, these models are implemented on the VEGA platform and are now freely downloadable online.     

Total synthesis of five thapsigargins: guaianolide natural products exhibiting sub-nanomolar SERCA inhibition

Herein we describe the total synthesis of five guaianolide natural products: thapsigargin, thapsivillosin C, thapsivillosin F, trilobolide and nortrilobolide. Prodrug derivatives of thapsigargin have shown selective in vivo cytotoxicity against prostate tumours and the need for further investigation of this phenomenon highlights the importance of these total syntheses. The first absolute stereochemical assignment of thapsivillosin C is also delineated.     

Interaction between the DNA model base 9-ethylguanine and a group of ruthenium polypyridyl complexes: kinetics and conformational temperature dependence

The binding capability of three ruthenium polypyridyl compounds of structural formula [Ru(apy)(tpy)Ln-](ClO4)(2-n) [1a-c; apy = 2,2'-azobis(pyridine), tpy = 2,2':6',2'-terpyridine, L = Cl, H2O, CH3CN] to a fragment of DNA was studied. The interaction between each of these complexes and the DNA model base 9-ethylguanine (9-EtGua) was followed by means of 1H NMR studies. Density functional theory calculations were carried out to explore the preferential ways of coordination between the ruthenium complexes and guanine. The ruthenium-9-EtGua adduct formed was isolated and fully characterized using different techniques. A variable-temperature 1H NMR experiment was carried out that showed that while the 9-EtGua fragment was rotating fast at high temperature, a loss of symmetry was suffered by the model base adduct as the temperature was lowered, indicating restricted rotation of the guanine residue.     

Tuning the selectivity/specificity of fluorescent metal ion sensors based on N2S2 pyridine-containing macrocyclic ligands by changing the fluorogenic subunit: spectrofluorimetric and metal ion binding studies

Two new fluorescent chemosensors for metal ions have been synthesized and characterized, and their photophysical properties have been explored; they are the macrocycles 5-(2-quinolinylmethyl)-2,8-dithia-5-aza-2,6-pyridinophane (L5) and 5-(5-chloro-8-hydroxyquinolinylmethyl)-2,8-dithia-5-aza-2,6-pyridinophane (L6). Both systems have a pyridyl-thioether-containing 12-membered macrocycle as a binding site. The coordination properties of these two ligands toward CuII, ZnII, CdII, HgII, and PbII have been studied in MeCN/H2O (1:1 v/v) and MeCN solutions and in the solid state. The stoichiometry of the species formed at 25 degrees C have been determined from absorption, fluorescence, and potentiometric titrations. The complexes [CuL5](ClO4)(2).1/2MeCN, [ZnL5(H2O)](ClO4)2, [HgL5(MeCN)](ClO4)2, [PbL5(ClO4)2], [Cu3(5-Cl-8-HDQH-1)(L6H-1)2](ClO4)(3).7.5H2O (HDQ=hydroxyquinoline), and [Cu(L6)2](BF4)(2).2MeNO2 have also been characterized by X-ray crystallography. A specific CHEF-type response of L5 and L6 to the presence of ZnII and CdII, respectively, has been observed at about pH 7.0 in MeCN/H2O (1:1 v/v) solutions.