Striking antitumor activity of a methinium system with incorporated quinoxaline unit obtained by spontaneous cyclization.

A novel pentamethinium salt was synthesized with an unforeseen expanded conjugated quinoxaline unit directly incorporated into a pentamethinium chain. The compound exhibited high fluorescence intensity, selective mitochondrial localization, high cytotoxicity, and selectivity toward malignant cell lines, and resulted in remarkable in vivo suppression of tumor growth in mice.

Cellular membrane phospholipids act as a depository for quaternary amine containing drugs thus competing with the acetylcholine/nicotinic receptor.

We previously demonstrated that ammonium- or guanidinium-phosphate interactions are key to forming noncovalent complexes (NCXs) through salt bridge formation with G-protein coupled receptors (GPCR), which are immersed in the cell membrane's lipids. The present work highlights MALDI ion mobility coupled to orthogonal time-of-flight mass spectrometry (MALDI IM oTOF MS) as a method to determine qualitative and relative quantitative affinity of drugs to form NCXs with targeted GPCRs' epitopes in a model system using, bis-quaternary amine based drugs, α- and ß- subunit epitopes of the nicotinic acetylcholine receptor' (nAChR) and phospholipids. Bis-quaternary amines proved to have a strong affinity for all nAChR epitopes and negatively charged phospholipids, even in the presence of the physiological neurotransmitter acetylcholine. Ion mobility baseline separated isobaric phosphatidyl ethanolamine and a matrix cluster, providing an accurate estimate for phospholipid counts. Overall this technique is a powerful method for screening drugs' interactions with targeted lipids and protein respectively containing quaternary amines and guanidinium moieties.

Capillary ion electrophoresis of inorganic anions and uric acid in human saliva using a polyvinyl alcohol coated capillary column and hexamethonium chloride as additive of background electrolyte.

A combination of polyvinyl alcohol chemically coated capillary (PVA capillary) and background electrolyte (BGE) with ion-pair reagent (hexamethonium dichloride, HMC) was used on capillary ion electrophoresis-UV detection (CIE-UV) for analysis of Br⁻, I⁻, NO2⁻, NO3⁻, SCN⁻ and uric acid in human saliva. The PVA capillary prepared in our laboratory minimized electro-osmotic flow (EOF) at the BGE in pH 3-10, and did not affect the UV detection at 210 nm by the PVA-layer on capillary wall. Therefore, use of the PVA capillary was suitable for sensitive UV detection for analyte anions, as well as suppression of protein adsorption. In this study, we optimized the BGE of 10 mM phosphate plus 10 mM HMC with applying a voltage of -15 kV. HMC as an additive to BGE could manipulate the electrophoretic mobility of anions, without electrostatic adsorption to the PVA capillary. The CIE-UV could separate and determine analyte anions in human saliva containing proteins by the direct injection without pretreatments such as dilution or deproteinization within 13 min. The relative standard deviations (n=10) were ranged of 0.5-1.6% in migration times, 2.2-6.8% in peak heights and 2.8-8.4% in peak areas. The limits of detection (S/N=3) were ranged of 3.42-6.87 µM. The peak height of anions in this system was gradually decreased through the successive injections of saliva samples, but the problem was successfully solved by periodically conditioning the PVA capillary. The quantifiability of anions in human saliva samples by the CIE-UV was evaluated through the recoveries by standard addition methods and comparison of other representative analytical methods, as well as identification by ion chromatography (IC). From the anion analyses in 12 different saliva samples, the CIE-UV demonstrated that can obtain obvious differences in concentrations of SCN⁻ between of smoker and non-smoker and those of uric acid between male and female with satisfactory results.

Properties of acetylcholine-induced relaxation of smooth muscle isolated from the proximal colon of the guinea-pig.

The properties of mechanical responses elicited by stimulation with acetylcholine (ACh) were investigated in circular smooth muscle preparations isolated from the proximal colon of guinea-pig. Application of ACh (10(-8)-10(-6) M) for 3-5 min produced a biphasic response, with an initial contraction followed by a relaxation. Atropine inhibited the initial contraction, while N(ω)-nitro-L-arginine (L-NA) inhibited the relaxation, suggesting that the former was produced by activation of muscarinic receptors while the latter was produced by an elevated production of nitric oxide (NO). In the presence of atropine, the ACh-relaxation was attenuated by removal of the mucosa and abolished by removal of both submucosal and mucosal layers. The ACh-induced relaxation was also attenuated by either tetrodotoxin (TTX, 3 × 10(-7) M) or hexamethonium (10(-6) M). In the presence of atropine, transmural nerve stimulation (TNS) elicited a biphasic response, with an initial phasic contraction followed by a relaxation. The amplitude of TNS-induced relaxation was significantly reduced by hexamethonium or L-NA and was abolished by TTX. Both ACh and TNS produced relaxation in preparations isolated from the proximal colon, but not in those from the middle part of colon. Immunohistochemistry for neuronal nitric oxide synthase revealed no difference in the distribution of nitrergic nerves between the proximal and middle part of the colon, with nitrergic nerves in both the mucosal and submucosal layers as well as in the smooth muscle and myenteric layers. These results suggest that ACh induces NO production by excitation of postganglionic nerves distributed mainly in the mucosal and submucosal layers. In circular smooth muscle preparations isolated from the middle part of colon, ACh or TNS produced contractile responses alone, with no associated relaxation, suggesting that the ACh-activated postganglionic nitrergic nerves are distributed in the mucosal and submucosal layers of the proximal colon but not in the middle part of the colon.

Electrophoretic behaviors of human hepatoma HepG2 cells.

The electrophoretic mobility of HepG2 cells was measured and a charge-regulated model was proposed to simulate the results obtained. Here, a cell was simulated by a rigid core and an ion-penetrable membrane layer containing both acidic and basic functional groups. The influences of the key parameters, including the pH, the ionic strength, the thickness of the membrane layer of a cell, the density and the dissociation constant of the dissociable functional groups in the membrane layer, and the binding constant of divalent cations on the electrophoretic mobility of a cell were investigated. In particular, the role of the buffer used in the experiment was discussed; this effect was neglected in almost all the relevant theoretical analyses in the literature. We showed that the binding ability of divalent cations to the dissociated functional groups in the membrane layer of a cell ranks as Ca(2+)>Mg(2+)>hexamethonium.

Amazing stability of phosphate-quaternary amine interactions.

We have previously used MALDI mass spectrometry to highlight ammonium- or guanidinium-aromatic interactions via cation-pi bonding and ammonium- or guanidinium-phosphate interactions through salt bridge formation. In the present work, the gas-phase stability and dissociation pathways of the interaction between phosphorylated peptides and compounds containing quaternary amines are demonstrated using electrospray ionization mass spectrometry. The presence of one quaternary amine in a compound is enough to form a noncovalent complex with a phosphorylated residue. However, if two quaternary amines are present in one molecule, the electrostatic interactions of the quaternary amines with the phosphate results in a "covalent-like" stability, and these bonds can withstand fragmentation by collision-induced dissociation at energies similar to those that fragment covalent bonds. Such interactions are important in accounting for physiological, pathophysiological, and pharmacological effects of many therapeutic compounds and small molecules containing quaternary amines or phosphates.

Detection of hexamethonium-perchlorate association complexes using NACE-MS.

Perchlorate (ClO(4) (+)) and other chlorine oxide anions were observed to complex weakly with hexamethonium (1,6-bis-(trimethylammonium)-hexane) in both aqueous and polar nonaqueous solvents. The resultant positively charged complexes were resolved by NACE using 2-propanol/acetone electrolytes prior to mass spectrometric detection using an Agilent(3D)CE system coupled to a Bruker Esquire 3000+ quadrupole IT mass detector. Using electrokinetic injection, the method detection limit for perchlorate in nonaqueous media was 10 microg/L. The isotope patterns due to the presence of (35)Cl and (37)Cl in complex mass spectra allowed for unambiguous identification of perchlorate, chlorate (ClO(3) (+)), chlorite (ClO(2) (+)), and chloride (Cl(+)) in photoreaction samples.

Muscarinic allosteric enhancers of ligand binding: pivotal pharmacophoric elements in hexamethonio-type agents.

Bisphthalimidopropyl-substituted hexamethonio compounds have been established as allosteric modulators of ligand binding to muscarinic acetylcholine receptors. Enhancers of ligand binding are of special interest. This study aimed to unravel the structural elements inducing positive cooperativity with the binding of an antagonist. [(3)H]-N-methylscopolamine binding to muscarinic M(2) receptors was measured in porcine heart homogenates. Dimethylation, but not monomethylation, of the lateral propyl chain in combination with an affinity increasing aromatic imide moiety, such as a 5-methylphthalimide and naphthalimide, on the same side of the molecule shifts the cooperativity toward positive values, resulting in enhancers of antagonist binding. Thus, lateral side chain dimethylation is a pivotal pharmacophoric element for positive cooperativity in hexamethonio-type muscarinic allosteric agents.