Immobilized betulinic acid column and its interactions with phospholipase A2 and snake venom proteins

Betulinic acid (BA) is a plant-derived pentacyclic triterpenoid. Although BA has been found to have diverse pharmacological effects, including anti-tumor and anti-inflammatory actions and potential as inhibitor of phospholipase A2 (PLA2), its cellular targets remain unclear. In this study, BA was immobilized onto an acrylamide matrix. The immobilized-BA column could retain the purified PLA2 of bovine pancreas or the PLA2 of snake venom from Naja nigricollis. The bound PLA2 were not eluted by high salt concentrations but were eluted by either acid or calcium free buffer. Besides the PLA2, a group of basic proteins of snake venom with molecular weights of about 7 kDa were also strongly bound by immobilized BA. One of these proteins was identified as gamma-cardiotoxin. The usefulness of immobilized BA for exploring the cellular targets of BA is discussed.     

4D-QSAR analysis of a series of antifungal p450 inhibitors and 3D-pharmacophore comparisons as a function of alignment

A training set of 55 antifungal p450 analogue inhibitors was used to construct receptor-independent four-dimensional quantitative structure-activity relationship (RI 4D-QSAR) models. Ten different alignments were used to build the models, and one alignment yields a significantly better model than the other alignments. Two different methodologies were used to measure the similarity of the best 4D-QSAR models of each alignment. One method compares the residual of fit between pairs of models using the cross-correlation coefficient of their residuals of fit as a similarity measure. The other method compares the spatial distributions of the IPE types (3D-pharmacophores) of pairs of 4D-QSAR models from different alignments. Optimum models from several different alignments have nearly the same correlation coefficients, r(2), and cross-validation correlation coefficients, xv-r(2), yet the 3D-pharmacophores of these models are very different from one another. The highest 3D-pharmacophore similarity correlation coefficient between any pair of 4D-QSAR models from the 10 alignments considered is only 0.216. However, the best 4D-QSAR models of each alignment do contain some proximate common pharmacorphore sites. A test set of 10 compounds was used to validate the predictivity of the best 4D-QSAR models of each alignment. The "best" model from the 10 alignments has the highest predictivity. The inferred active sites mapped out by the 4D-QSAR models suggest that hydrogen bond interactions are not prevalent when this class of P450 analogue inhibitors binds to the receptor active site. This feature of the 4D-QSAR models is in agreement with the crystal structure results that indicate no ligand-receptor hydrogen bonds are formed.     

Analytical expressions for the electrical potential near planar, cylindrical, and spherical surfaces for symmetric electrolytes

The conjecture of Tuinier (J. Colloid Interface Sci. 258 (2003) 45) for the electrical potentials near a cylindrical surface and near a spherical surface under the conditions of symmetric electrolyte and large scaled radius are derived by solving the corresponding Poisson-Boltzmann equation. The surface charge density-surface potential relations for these surfaces are also derived under the conditions of constant surface potential. We show that the level of surface charge density for planar, cylindrical, and spherical surfaces follows the order spherical surface > cylindrical surface > planar surface.     

Automated,continuous, and dynamic speciation of urinary arsenic in the bladder of living organisms using microdialysis sampling coupled on-line with high performance liquid chromatography and hydride generation atomic absorption spectrometry

An on-line and fully automated method was developed for the continuous and dynamic in vivo monitoring of four arsenic species [arsenite (AsIII), arsenate (AsV), monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA)] in urine of living organisms. In this method a microdialysis sampling technique was employed to couple on-line with high performance liquid chromatography (HPLC) and hydride generation atomic absorption spectrometry (HGAAS). Dialysates perfused through implanted microdialysis probes were collected with a sample loop of an on-line injector for direct and automated injection into HPLC system hyphenated with HGAAS. The saline (0.9% NaCl) solution was perfused at the rate of 1 microl min(-1) through the microdialysis probe and the dialysate was loaded into 50 microl of sample loop. The separation conditions were optimally selected to be in phosphate buffer solution at a pH 5.2 with a flow rate of 1.2 ml min(-1). The effluent from the HPLC was first mixed on-line at the exit of the column with HCl (1 M) solution and then mixed with a NaBH4 (0.2% m/v) solution. Based on the optimal conditions obtained, linear ranges of 2.5-50 ng ml(-1) for AsIII and 6.75-100 ng ml(-1) for the other three arsenic species were obtained. Detection limits of 1.00, 2.18, 1.03 and 2.17 ng ml(-1) were obtained for AsIII, DMA, MMA and AsV, respectively. Typical precision values of 3.4% (AsIII), 5.4% (DMA), 3.6% (MMA) and 7.5% (AsV) were obtained, respectively, at a 25 ng ml(-1) level. Recoveries close to 100%, relative to an aqueous standard, were observed for each species. The average in vivo recoveries of AsIII, DMA, MMA and AsV in rat bladder urine were 56+/-5%, 60+/-9%, 49+/-3% and 55+/-7%, respectively. The use of an on-line microdialysis-HPLC-HGAAS system permitted the determination of four urinary arsenic species in the bladder of an anesthetized rat with a temporal resolution of 50 min sampling.     

Dynamic chirality: keen selection in the face of stereochemical diversity in mechanically bonded compounds

The template-directed syntheses, employing bisparaphenylene-[34]crown-10 (BPP34C10), 1,5-dinaphthoparaphenylene-[36]crown-10 (1/5NPPP36C10), and 1,5-dinaphtho-[38]crown-10 (1/5DNP38C10) as templates, of three [2]catenanes, whereby one of the two bipyridinium units in cyclobis(paraquat-p-phenylene) is replaced by a bipicolinium unit, are described. The crude reaction mixtures comprising the [2]catenanes all contain slightly more of the homologous [3]catenanes, wherein a "dimeric" octacationic cyclophane has the crown ether macrocycles encircling the alternating bipyridinium units with the bipicolinium units completely unfettered. X-ray crystallography, performed on all three [2]catenanes and two of the three [3]catenanes reveals co-conformational and stereochemical preferences that are stark and pronounced. Both the [3]catenanes crystallize as mixtures of diastereoisomers on account of the axial chirality associated with the picolinium units in the solid state. Dynamic (1)H NMR spectroscopy is employed to probe in solution the relative energy barriers for rotations by the phenylene and pyridinium rings in the tetracationic cyclophane component of the [2]catenanes. Where there are co-conformational changes that are stereochemically "allowed", crown ether circumrotation and rocking processes are also investigated for the relative rates of their occurrence. The outcome is one whereby the three [2]catenanes containing BPP34C10, 1/5NPPP36C10, and 1/5DNP38C10 exist as one major enantiomeric pair of diastereoisomers amongst two, four, and eight diastereoisomeric pairs of enantiomers, respectively. The diastereoisomerism is a consequence of the presence of axial chirality together with helical and/or planar chirality in the same interlocked molecule. These [2]catenanes constitute a rich reserve of new stereochemical types that might be tapped for their switching and mechanical properties.     

The Synthesis of Alanine Oligopeptides

The oligopeptides of alanine (penta-, deca-, and pentadeca-) were synthesized by solid phase method and new peptide-resin cleaving reagent TFA-BBr₃ was used to cleave the peptides from resin-support. Penta-alanine was also synthesized by liquid phase for the comparision and FAB-Mass method was used to determine the molecular weight of penta-alanine. HPLC separation of these new oligopeptides were described.     

Structures and properties of self-assembled monolayers of bistable [2]rotaxanes on Au (111) surfaces from molecular dynamics simulations validated with experiment

Bistable [2]rotaxanes display controllable switching properties in solution, on surfaces, and in devices. These phenomena are based on the electrochemically and electrically driven mechanical shuttling motion of the ring-shaped component, cyclobis(paraquat-p-phenylene) (CBPQT(4+)) (denoted as the ring), between a tetrathiafulvalene (TTF) unit and a 1,5-dioxynaphthalene (DNP) ring system located along a dumbbell component. When the ring is encircling the TTF unit, this co-conformation of the rotaxane is the most stable and thus designated the ground-state co-conformer (GSCC), whereas the other co-conformation with the ring surrounding the DNP ring system is less favored and so designated the metastable-state co-conformer (MSCC). We report here the structure and properties of self-assembled monolayers (SAMs) of a bistable [2]rotaxane on Au (111) surfaces as a function of surface coverage based on atomistic molecular dynamics (MD) studies with a force field optimized from DFT calculations and we report several experiments that validate the predictions. On the basis of both the total energy per rotaxane and the calculated stress that is parallel to the surface, we find that the optimal packing density of the SAM corresponds to a surface coverage of 115 A(2)/molecule (one molecule per 4 x 4 grid of surface Au atoms) for both the GSCC and MSCC, and that the former is more stable than the latter by 14 kcal/mol at the optimum packing density. We find that the SAM retains hexagonal packing, except for the case at twice the optimum packing density (65 A(2)/molecule, the 3 x 3 grid). For the GSCC and MSCC, investigated at the optimum coverage, the tilt of the ring with respect to the normal is theta = 39 degrees and 61 degrees, respectively, while the tilt angle of the entire rotaxane is psi = 41 degrees and 46 degrees , respectively. Although the tilt angle of the ring decreases with decreasing surface coverage, the tilt angle of the rotaxane has a maximum at 144 A(2)/molecule (the 4 x 5 grid/molecule) of 50 degrees and 51 degrees for the GSCC and MSCC, respectively. The hexafluorophosphate counterions (PF(6)(-)) stay localized around the ring during the 2 ns MD simulation. On the basis of the calculated density profile, we find that the thickness of the SAM is 40.5 A at the optimum coverage for the GSCC and 40.0 A for MSCC, and that the thicknesses become less with decreasing surface coverage. The calculated surface tension at the optimal packing density is 45 and 65 dyn/cm for the GSCC and MSCC, respectively. This difference suggests that the water contact angle for the GSCC is larger than for the MSCC, a prediction that is verified by experiments on Langmuir-Blodgett monolayers of amphiphilic [2]rotaxanes.     

On the convergence of the affine-scaling algorithm

The affine-scaling algorithm, first proposed by Dikin, is presently enjoying great popularity as a potentially effective means of solving linear programs. An outstanding question about this algorithm concerns its convergence in the presence of degeneracy. In this paper, we give new convergence results for this algorithm that do not require any non-degeneracy assumption on the problem. In particular, we show that if the stepsize choice of either Dikin or Barnes or Vanderbei, et al. is used, then the algorithm generates iterates that converge at least linearly with a convergence ratio of , wheren is the number of variables and (0,1] is a certain stepsize ratio. For one particular stepsize choice which is an extension of that of Barnes, we show that the cost of the limit point is within O(/(1–)) of the optimal cost and, for sufficiently small (roughly, proportional to how close the cost of the nonoptimal vertices are to the optimal cost), is exactly optimal. We prove the latter result by using an unusual proof technique, that of analyzing the ergodic convergence of the corresponding dual vectors. For the special case of network flow problems with integer data, we show that it suffices to take = 1/(6mC), wherem is the number of constraints andC is the sum of the cost coefficients, to attain exact optimality.This research is partially supported by the U.S. Army Research Office, contract DAAL03-86-K-0171 (Center for Intelligent Control Systems), by the National Science Foundation, grant NSF-ECS-8519058, and by the Science and Engineering Research Board of McMaster University.     

Triisopropyltriazacyclononane copper(II): an efficient phosphodiester hydrolysis catalyst and DNA cleavage agent

A 6000-fold rate enhancement has been observed for the hydrolysis of bis(p-nitrophenyl)phosphate (BNPP) in the presence of 0.2 mM Cu(i-Pr(3)[9]aneN(3))(2+) at pH 9.2 and 50 degrees C. In a direct comparison, the rate of hydrolysis of BNPP is accelerated at least 60-fold over the previously reported catalyst Cu([9]aneN(3))(2+). As observed for Cu([9]aneN(3))(2+), hydrolysis is selective for diesters over monoesters. Hydrolysis of BNPP by Cu(i-Pr(3)[9]aneN(3))(2+) is catalytic, exhibiting both rate enhancement and turnover. The reaction is inhibited by both p-nitrophenyl phosphate and inorganic phosphate. The reaction is first-order in substrate and half-order in metal complex, with a k(1.5) of 0.060 +/- 0.004 M(-1/2) s(-1) at 50 degrees C. The temperature dependence of the rate constant results in a calculated activation enthalpy (Delta H(++) of 51 +/- 2 kJ mol(-1) and activation entropy (Delta S(++)) of -110 +/- 6 J mol(-1) K(-1). The kinetic pK(a) of 7.8 +/- 0.2 is close to the thermodynamic pK(a) of 7.9 +/- 0.2, consistent with deprotonation of a coordinated water molecule in the active form of the catalyst. The active catalyst [Cu(i-Pr(3)[9]aneN(3))(OH)(OH(2))](+) is in equilibrium with an inactive dimer, and the formation constant for this dimer is between 216 and 1394 M(-1) at pH 9.2 and 50 degrees C. Temperature dependence of the dimer formation constant K(f) indicates an endothermic enthalpy of formation for the dimer of 27 +/- 3 kJ mol(-1). The time course of anaerobic DNA cleavage by Cu(i-Pr(3)[9]aneN(3))(2+) is presented over a wide range of concentrations at pH 7.8 at 50 degrees C. The concentration dependence of DNA cleavage by Cu([9]aneN(3))(2+) and Cu(i-Pr(3)[9]aneN(3))(2+) reveals a maximum cleavage efficiency at sub-micromolar concentrations of cleavage agent. DNA cleavage by Cu(i-Pr(3)[9]aneN(3))(2+) is twice as efficient at pH 7.8 as at pH 7.2.     

Hepatoprotective Activity of Nelumbo nucifera Gaertn. Seedpod Extract Attenuated Acetaminophen-Induced Hepatotoxicity

The aim is to investigate the effect of lotus (Nelumbo nucifera Gaertn.) seedpod extract (LSE) on acetaminophen (APAP)-induced hepatotoxicity. LSE is rich in polyphenols and has potent antioxidant capacity. APAP is a commonly used analgesic, while APAP overdose is the main reason for drug toxicity in the liver. Until now, there has been no in vitro test of LSE in drug-induced hepatotoxicity responses. LSEs were used to evaluate the effect on APAP-induced cytotoxicity, ROS level, apoptotic rate, and molecule mechanisms. The co-treatment of APAP and LSEs elevated the survival rate and decreased intracellular ROS levels on HepG2 cells. LSEs treatment could significantly reduce APAP-induced HepG2 apoptosis assessed by DAPI and Annexin V/PI. The further molecule mechanisms indicated that LSEs decreased Fas/FasL binding and reduced Bax and tBid to restore mitochondrial structure and subsequently suppress downstream apoptosis cascade activation. These declines in COX-2, NF-κB, and iNOS levels were observed in co-treatment APAP and LSEs, which indicated that LSEs could ameliorate APAP-induced inflammation. LSE protected APAP-induced apoptosis by preventing extrinsic, intrinsic, and JNK-mediated pathways. In addition, the restoration of mitochondria and inflammatory suppression in LSEs treatments indicated that LSEs could decrease oxidative stress induced by toxic APAP. Therefore, LSE could be a novel therapeutic option for an antidote against overdose of APAP.