Naturally Drug-Loaded Chitin: Isolation and Applications.

Naturally occurring three-dimensional (3D) biopolymer-based matrices that can be used in different biomedical applications are sustainable alternatives to various artificial 3D materials. For this purpose, chitin-based structures from marine sponges are very promising substitutes. Marine sponges from the order Verongiida (class Demospongiae) are typical examples of demosponges with well-developed chitinous skeletons. In particular, species belonging to the family Ianthellidae possess chitinous, flat, fan-like fibrous skeletons with a unique, microporous 3D architecture that makes them particularly interesting for applications. In this work, we focus our attention on the demosponge Ianthella flabelliformis (Linnaeus, 1759) for simultaneous extraction of both naturally occurring ("ready-to-use") chitin scaffolds, and biologically active bromotyrosines which are recognized as potential antibiotic, antitumor, and marine antifouling substances. We show that selected bromotyrosines are located within pigmental cells which, however, are localized within chitinous skeletal fibers of I. flabelliformis. A two-step reaction provides two products: treatment with methanol extracts the bromotyrosine compounds bastadin 25 and araplysillin-I N20 sulfamate, and a subsequent treatment with acetic acid and sodium hydroxide exposes the 3D chitinous scaffold. This scaffold is a mesh-like structure, which retains its capillary network, and its use as a potential drug delivery biomaterial was examined for the first time. The results demonstrate that sponge-derived chitin scaffolds, impregnated with decamethoxine, effectively inhibit growth of the human pathogen Staphylococcus aureus in an agar diffusion assay.

The research of antimicrobial efficacy of antiseptics decamethoxin, miramistin and their effect on nuclear DNA fragmentation and epithelial cell cycle.

OBJECTIVE: Introduction: Nowadays, the study of biological safety of modern cationic surface-active antiseptics with a wide antimicrobial spectrum has acquired particular importance. The aim was to study antimicrobial effectiveness of antiseptics decamethoxin, miramistin and their influence on nuclear DNA fragmentation and cellular cycle. PATIENTS AND METHODS: Materials and methods: A comparative microbiological study of antimicrobial efficacy and a cytometric study of the effect of decamethoxin 0,02% and miramistin 0,01% on the cellular cycle were carried out. Antimicrobial activity of decamethoxin and miramistin was estimated by their minimal inhibitory and minimal microbicidal concentrations against opportunistic microorganisms using serial double dilution technique. Decamethoxin and miramistin cytotoxicity on anterior corneal epithelial cells, after their two-week daily instillation into the eyes of a Vistar line male rats was studied using flow cytometry. The parameters of epithelial cellular cycle, nuclear DNA fragmentation and apoptosis under the influence of antiseptics were registered. RESULTS: Results: High antimicrobial effect of decamethoxin and miramistin against Gram-positive, Gram-negative bacteria with the significant advantages of decamethoxin were found (р<0,001). Decamethoxin caused minimal influence on anterior corneal epithelial cells, the insignificant decrease of their proliferation index, low increase of apoptosis (0.68%), no difference of mitotic activity (p>0.05). But the use of miramistin resulted in the significant increase of nuclear DNA fragmentation, decrease of proliferative activity (р<0.05). CONCLUSION: Conclusions: Higher antimicrobial effect against a wide range of opportunistic pathogens is proved in decamethoxin 0,02% comparably to miramistin 0,01% (р<0,001). In prolonged antiseptic use of the first one there were found no cytotoxic and no pro-apoptotic effects on the epithelium (р<0,05).

[АNTISEPTICS: MODERN STRATEGY OF STRUGGLE WITH CAUSING AGENTS OF THE ІNFECTION COMPLICATIONS].

Etiology of infective complications was investigated in 71 injured persons, suffering severe burns. There was established, that the main causing agents in patients, suffering burn disease, are S. aureus(in 35.9% of observations), A. baumannii (in 25%), P. aeruginosa (in 12.82%), P. mirabilis (in 5.12%). Resistance of conditionally pathogenic microorganisms towards cephalosporins, аminoglycosides, іmipenem, meropenem, doxycycline was determined. Effective bactericidal activity of antiseptic solutions of decasan, miramistinum, chlorhexidine was proved. High antimicrobial properties of dressing materials, which contain decametoxine, chlorhexidine, furagin, silver ions against Staphylococcus were noted. Clinical efficacy of application of materials, impregnated by antimicrobial composition decametoxine with carboxymethylstarch, oxyethylcellulose and polyvynilacetate, for prophylaxis and treatment of infective purulent­inflammatory complications in patients, suffering burns, was proved.

[Kinetics of decamethoxine, an antimicrobial agent].

The kinetics of decamethoxine liberation from medical antimicrobial textiles was studied. The elution of decamethoxine was shown to be a complicated diffusive-kinetic process dependent on the exposure and concentration of decamethoxine.

[Use of antiseptic dekasan in complex treatment of peritonitis].

Experience in treatment of 91 patients with peritonitis on various genesis using antiseptic Dekasan are presented. A marked clinical efficacy compared with that of other antiseptics in the complex treatment of peritonitis was noted.

[Sanation of biliary system using antiseptic decasan in complex treatment of cholangitis].

Experience of treatment of 17 patients, suffering cholangitis of various genesis, using antiseptic Decasan, is presented. Clinical efficacy of the preparation in complex treatment of cholangitis, confirmed by results of the bile bacteriological investigation, was noted.

[Antimicrobial properties of antiseptic composite with prolonged action].

Antimicrobial properties of a composite based on decamethoxine and modified polysaccharides (carboxymethylamylum, oxyethyl-cellulose) were studied. The composite was shown to have high antimicrobial activity against grampositive and gramnegative bacteria under different conditions of the experiment.

[Local impact of antiseptic medical textile on tissues of organism].

Morphological investigation for studying of a local impact on the tissues, localized in the antiseptic textile implantation zone, was conducted. The textile was impregnated by composition of decametoxine with modified polysaccharides. Basing on the investigation result there was established the absence of a toxic impact of antiseptic medical textile on the macroorganism tissues, the regenerative processes course, the wounds epithelization, antioedematous and anti-inflammatory effects.

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.

[Application of decamethoxin solution in the treatment of surgical peritonitis].

Basing on analysis of results of clinical and experimental investigations, there was established, that application of the cationic antiseptics solution (including 0.02% solution of decametoxin) for the abdominal cavity sanation permits to reduce the microbal soiling while sanation performance and as well so on--the postoperative complications rate and mortality in surgical peritonitis.

CE methods for analysis of isoforms of prostate-specific antigen compatible with online derivatization for LIF detection.

Prostate-specific antigen (PSA) is the usual biomarker for prostate cancer (PCa). However, its lack of selectivity has lead to the search for new biomarkers. PSA glycosylation seems to depend on the pathophysiological conditions of the individual. Thus, methods to separate PSA isoforms (peaks) to study their role as PCa markers are needed. In this work, CE methods for PSA isoforms separation, based on the use of different dynamic coatings, are developed using UV detection. Three complementary CE methods allowing the separation of 8 or 9 PSA isoforms are selected. The longest method takes only 17 min, while the shortest one separates 9 isoforms in < 8 min. Depending on the isoforms of interest for their use as PCa biomarker, the CE method to be used can be chosen or various of them can be combined. A remarkable aspect of these methods is that the BGEs employed are devoid of compounds with primary amino groups, making the CE methods compatible with fluorescent on-column derivatization through amino residues. As a proof-of-concept, a preliminary result shows that LIF detection of labeled PSA analyzed by one of the three developed methods permits detection of glycoprotein isoforms.

[Application of antiseptic decasan in purulent infections of pararectal region].

The results of studying of antiseptic preparation Decasan for the treatment of patients, suffering purulent infections of pararectal region, are adduced. Efficacy of Decasan was analyzed in 102 patients. The results of investigation obtained are trusting hig antiseptic efficacy of preparation, witnessing possibility of its application as a preparation of choice for the treatment of patients, suffering purulent infections of pararectal region.

Activation and inhibition of mouse muscle and neuronal nicotinic acetylcholine receptors expressed in Xenopus oocytes.

Transgenic mouse models with nicotinic acetylcholine receptor (nAChR) knockouts and knockins have provided important insights into the molecular substrates of addiction and disease. However, most studies of heterologously expressed neuronal nAChR have used clones obtained from other species, usually human or rat. In this work, we use mouse clones expressed in Xenopus oocytes to provide a relatively comprehensive characterization of the three primary classes of nAChR: muscle-type receptors, heteromeric neuronal receptors, and homomeric alpha7-type receptors. We evaluated the activation of these receptor subtypes with acetylcholine and cytisine-related compounds, including varenicline. We also characterized the activity of classic nAChR antagonists, confirming the utility of mecamylamine and dihydro-beta-erythroidine as selective antagonists in mouse models of alpha3beta4 and alpha4beta2 receptors, respectively. We also conducted an in-depth analysis of decamethonium and hexamethonium on muscle and neuronal receptor subtypes. Our data indicate that, as with receptors cloned from other species, pairwise expression of neuronal alpha and beta subunits in oocytes generates heterogeneous populations of receptors, most likely caused by variations in subunit stoichiometry. Coexpression of the mouse alpha5 subunit had varying effects, depending on the other subunits expressed. The properties of cytisine-related compounds are similar for mouse, rat, and human nAChR, except that varenicline produced greater residual inhibition of mouse alpha4beta2 receptors than with human receptors. We confirm that decamethonium is a partial agonist, selective for muscle-type receptors, but also note that it is a nondepolarizing antagonist for neuronal-type receptors. Hexamethonium was a relatively nonselective antagonist with mixed competitive and noncompetitive activity.

[Dekasan application in treatment of infected pancreatic necrosis and its complications].

The experience of the use of domestic antiseptic dekasan for topical use in patients over the infected pancreatic necrosis and its complications. There was significant improvement in the results of patients treatment.

Drug-templated mesoporous silica nanocontainers with extra high payload and controlled release rate.

The possibility of one-step creating of pH-sensitive mesostructured silica-based nanocontainers with exceptionally high payload using associates of two antiseptics (including hydrolyzable one) as templates is demonstrated. The effects of the template nature and the conditions of the sol-gel process on the porous structure of silica nanocontainers are studied and discussed. The kinetics of the templating drug release from such containers is studied and some features of this process are analyzed. It is shown that the drug release rate can be tuned by varying the medium pH. The bactericidal activity of two encapsulated antiseptics against the Staphylococcus aureus is evaluated in vitro by agar diffusion method with replacement of agar with agarose. The diameters of the inhibition zones for silica-based containers loaded with antiseptics increased with the pre-diffusion time at 4 °C. At the same time, empty containers (after elimination of antiseptics by etching) did not reveal any bactericidal properties.

Stabilization of Torpedo californica acetylcholinesterase by reversible inhibitors.

The dimeric form of Torpedo californica acetylcholinesterase provides a valuable experimental system for studying transitions between native, partially unfolded, and unfolded states since long-lived partially unfolded states can be generated by chemical modification of a nonconserved buried cysteine residue, Cys 231, by denaturing agents, by oxidative stress, and by thermal inactivation. Elucidation of the 3D structures of complexes of Torpedo californica acetylcholinesterase with a repertoire of reversible inhibitors permits their classification into three categories: (a) active-site directed inhibitors, which interact with the catalytic anionic subsite, at the bottom of the active-site gorge, such as edrophonium and tacrine; (b) peripheral anionic site inhibitors, which interact with a site at the entrance to the gorge, such as propidium and d-tubocurarine; and (c) elongated gorge-spanning inhibitors, which bridge the two sites, such as BW284c51 and decamethonium. The effects of these three categories of reversible inhibitors on the stability of Torpedo californica acetylcholinesterase were investigated using spectroscopic techniques and differential scanning calorimetry. Thermodynamic parameters obtained calorimetrically permitted quantitative comparison of the effects of the inhibitors on the enzyme's thermal stability. Peripheral site inhibitors had a relatively small effect, while gorge-spanning ligands and those binding at the catalytic anionic site, had a much larger stabilizing effect. The strongest effect was, however, observed with the polypeptide toxin, fasciculin II (FasII), even though, in terms of its binding site, it belongs to the category of peripheral site ligands. The stabilizing effect of the ligands binding at the anionic subsite of the active site, like that of the gorge-spanning ligands, may be ascribed to their capacity to stabilize the interaction between the two subdomains of the enzyme. The effect of fasciculin II may be ascribed to the large surface area of interaction (>2000 A(2)) between the two proteins. Stabilization of Torpedo californica acetylcholinesterase by both divalent cations and chemical chaperones was earlier shown to be due to a shift in equilibrium between the native state and a partially unfolded state ( Millard et al. ( 2003 ) Protein Sci. 12 , 2337 - 2347 ). The low molecular weight inhibitors used in the present study may act similarly and can thus be considered as pharmacological chaperones for stabilizing the fully folded native form of the enzyme.

Acetylcholine binding to Torpedo electroplax: relationship to acetylcholine receptors.

Binding of [(3)H]acetylcholine to a particulate fraction of Torpedo electroplax was measured by equilibrium dialysis. Two high-affinity sites present on phospholipoproteins bound acetylcholine reversibly, and binding was blocked by nicotinic drugs. Characteristics of this binding suggest that these phospholipoproteins may be acetylcholine receptors.

Electrical phenomena associated with the activity of the membrane-bound acetylcholinesterase.

Treatment of isolated electroplax with physiological solutions supplemented with either 1 molar sodium chloride, 2 molar urea, or 2 molar sucrose renders the cell insensitive to carbamylcholine, phenyltrimethylammonium, or decamethonium even at high concentrations. The treated cells have a residual resting potential of -20 +/- 10 millivolts (negative inside) and are depolarized by acetylcholine at concentrations larger than 10(-3) mole per liter. This response is not affected by d-tubocurarine but is blocked by physostigmine, diisopropylphosphorofluoridate, or strong buffers and thus depends on the catalytic activity of the membrane-bound acetylcholinesterase.

[Application of antiseptic dekasan in urgent abdominal surgery].

Experience in local application of antiseptic Dekasan in the complex treatment of abdominal infections of various origins are presented. The clinical and economic efficacy of the drug application is notice.