Impact of functional satellite groups on the antimicrobial activity and hemocompatibility of telechelic poly(2-methyloxazoline)s

Polyoxazolines with a biocidal quarternary ammonium end-group are potent biocides. Interestingly, the antimicrobial activity of the whole macromolecule is controlled by the nature of the group at the distal end. These nonreactive groups are usually introduced via the initiator. Here we present a study with a series of polymethyloxazolines with varying satellite groups introduced upon termination of the polymerization reaction. This allowed us to introduce a series of functional satellites, including hydroxy, primary amino, and double-bond-containing groups. The resulting telechelic polyoxazolines were explored regarding their antimicrobial activity and toxicity. It was found that the functional satellite groups greatly controlled the minimal inhibitory concentrations against the bacteria Staphylococcus aureus and Escherichia coli in a range of 10 to 2500 ppm. Surprisingly, the satellite groups also controlled the hemotoxicity but in a different way than the antimicrobial efficiency.     

A facile synthesis of azido-terminated heterobifunctional poly(ethylene glycol)s for "click" conjugation

New azido-terminated heterobifunctional poly(ethylene glycol) (PEG) derivatives having primary amine and carboxyl end groups, (Azide-PEG-NH 2 and Azide-PEG-COOH, respectively) were synthesized with high efficiency. An alpha-allyl-omega-hydroxyl PEG was prepared as the first step to Azide-PEG-X (X = NH 2 and COOH) through the ring-opening polymerization of ethylene oxide (EO) with allyl alcohol as an initiator, followed by two-step modification of the hydroxyl end to an azido group. To introduce primary amino or carboxyl functional groups, amination and carboxylation reactions of the allyl terminal ends was then conducted by a radical addition of thiol compounds. Molecular functionalities of both ends of the PEG derivatives thus prepared were characterized by (1)H, (13)C NMR, and MALDI-TOF MS spectra, validating that the reaction proceeded quantitatively. The terminal azido functionality is available to conjugate various ligands with an alkyne group through the 1,3-dipolar cycloaddition reaction condition ("click chemistry").     

Synthesis, characterization and in vitro biological evaluation of some novel diarylsulfonylureas as potential cytotoxic and antimicrobial agents

A series of novel diarylsulfonylureas (1-28) have been synthesized and characterized by FTIR, (1)H NMR, (13)C NMR and LC mass spectral analysis. All the synthesized compounds were evaluated for their in vitro cytotoxicity and antimicrobial activities. Among the tested compounds for cytotoxicity using Brine Shrimp Lethality assay, compounds 18 and 22 exhibited significant cytotoxicity at ED(50) values 3.96±0.21 and 4.02±0.19μg/mL, respectively. This level of activity was found comparable to that of the reference drug podophyllotoxin with ED(50) value 3.61±0.17μg/mL and it could be a remarkable starting point to develop new lead molecules with major cytotoxicity. Antimicrobial activity was screened using agar well diffusion assay method against selected Gram-positive, Gram-negative and fungal strains. Most of the compounds showed promising antibacterial and antifungal activity and the activity expressed as the minimum inhibitory concentration (MIC) in μg/mL.     

防御素构效关系研究进展

防御素是第一大类内源性抗微生物肤,具有广泛的抗菌谱作用,是生物体先天防御系统的重要组成成分.综述了近年国际上对防御素的一级结构、二级结构、三级结构和四级结构及其构效关系研究的最新进展.     

Enzymatic preparation of novel thermoplastic di-block copolyesters containing poly[(R)-3-hydroxybutyrate] and poly(epsilon-caprolactone) blocks via ring-opening polymerization

Enzymatic modification of a microbial polyester was achieved by the ring-opening polymerization of epsilon-caprolactone (CL) with low-molecular weight telechelic hydroxylated poly[( R)-3-hydroxybutyrate] (PHB-diol) as initiator and Novozym 435 (immobilized Candida antarctica Lipase B) as catalyst in anhydrous 1,4-dioxane or toluene. The ring-opening polymerization was investigated at different conditions with two different types of PHB-diols: PHB-diol(P), containing a primary OH and a secondary OH end groups, and PHB-diol(M), consisting of 91% PHB-diol(P) and 9% PHB-diol containing two secondary OH end groups. The reactions were followed by GPC analyses of the resulting polymers at different time points, and the optimal conditions were established to be 70 degrees C at a weight ratio of CL/enzyme/solvent of 8:1:24. The ring-opening polymerization of CL with PHB-diol(M) (Mn of 2380, NMR) at the molar ratio of 50:1 under the optimal conditions in 1,4-dioxane gave the corresponding poly[HB(56 wt %)-co-CL(44 wt %)] with Mn (NMR) of 3900 in 66% yield. Polymerization of CL and PHB-diol(P) ( Mn of 2010, NMR) at the same condition in toluene gave the corresponding poly[HB(28 wt %)-co-CL(72 wt %)] with Mn (NMR) of 7100 in 86% yield. Both polymers were characterized by (1)H and (13)C NMR and IR analyses as di-block copolyesters containing a PHB block with a secondary OH end group and a poly(epsilon-caprolactone) (PCL) block with a primary OH end group. NMR analyses and control experiments suggested no formation of random copolymers and no change of the PHB block during the reaction. The enzymatic ring-opening polymerization was selectively initiated by the primary OH group of PHB-diol, whereas the secondary OH group remained as an end group in the final polymers. The thermal properties of the di-block poly(HB-co-CL)s were analyzed by DSC, with excellent T g values for the elastomer domain: poly[HB(56 wt %)- co-CL(44 wt %)] with M n (NMR) of 3900 demonstrated a T g of -57 degrees C, Tm of 145, 123, and 53 degrees C; and poly[HB(28wt%)-co-CL(72wt%)] with Mn (NMR) of 7100 gave a Tg of -60 degrees C, Tm of 147 and 50 degrees C. Thus, the selective enzymatic ring-opening polymerization with PHB-diol as macro-initiator provides a new method for the preparation of PHB-based block copolymers as biomaterials with good thermoplastic properties and novel structures containing functional end groups.     

Synthesis,antimicrobial and anti-biofilm activities of novel Schiff base analogues derived from methyl-12-aminooctadec-9-enoate

A novel library of Schiff base analogues (5a–q) were synthesized by the condensation of methyl-12-aminooctadec-9-enoate and different substituted aromatic aldehydes. The synthesized compounds were thoroughly characterized by spectroscopic techniques (FT-IR, ¹H NMR, ¹³C NMR, ESI-MS and HRMS). The Schiff base analogues with different substitutions were screened for in vitro antibacterial activity against 7 different bacterial strains. Among these, the compounds with electron withdrawing substituent, namely chlorine (5a) and electron donating substituents, namely hydroxy (5n) and methoxy (5o), were found to exhibit excellent to good antimicrobial activities (MIC value 9–18 μM) against Staphylococcus aureus MTCC 96, Staphylococcus aureus MLS-16 MTCC 2940 and Bacillus subtilis MTCC 121. The products were also screened for anti-biofilm and MBC (Minimum Bactericidal Concentration) activities which exhibited promising activities.     

Block copolymerization of α-amino acid N-carboxyanhydride initiated by vinyl polymers having a terminal amino group and the characterization of the block copolymers

Poly(methyl methacrylate) and polystyrene having terminal amino groups were synthesized by the radical polymerization of those monomers in the presence of 2-mercaptoethylammonium chloride as a chain-transfer agent. By the terminal group analysis and the molecular weight determination of the polymers, 0.5–1.3 amino groups were found in a chain of poly(methyl methacrylate) and 0.5–2.5 amino groups in a chain of polystyrene. Using these polymers having a terminal amino group as an initiator, the block polymerization of α-amino acid N-carboxyanhydride (NCA) was carried out. In the polymerizations of Glu(OBzl) NCA and Lys(Z) NCA by the poly(methyl methacrylate) initiator, the terminal amino group underwent a nucleophilic addition reaction to NCA and initiated the polymerization, yielding A-B-type block copolymers in a high yield. The same was observed in the polymerizations of Gly(OBzl) NCA and Lys(Z) NCA by the polystyrene initiator. By eliminating the protecting groups of the side chains of the polypeptide segment, the block copolymers poly(methyl methacrylate)-poly(Glu), poly(methyl methacrylate)-poly(Lys), polystyrene-poly(Glu) and polystyrene-poly(Lys) were synthesized with little side reactions. The side chain amino groups of poly(Lys) segment in the poly(methyl methacrylate)-poly(Lys) block copolymers were sulphonated or stearoylated successfully.     

Quaternization of N-aryl chitosan derivatives: synthesis, characterization, and antibacterial activity

Chemical modification of chitosan by introducing quaternary ammonium moieties into the polymer backbone renders excellent antimicrobial activity to the adducts. In the present study, we have synthesized 17 derivatives of chitosan consisting of a variety of N-aryl substituents bearing either electron-donating or electron-withdrawing groups. Selective N-arylation of chitosan was performed via Schiff bases formed by the reaction between the 2-amino groups of the glucosamine residue of chitosan with aromatic aldehydes under acidic conditions, followed by reduction of the Schiff base intermediates with sodium cyanoborohydride. Each of the derivatives was further quaternized using N-(3-chloro-2-hydroxypropyl)trimethylammonium chloride (Quat-188) as the quaternizing agent that reacted with either the primary amino or hydroxyl groups of the glucosamine residue of chitosan. The resulting quaternized materials were water soluble at neutral pH. Minimum inhibitory concentration (MIC) antimicrobial studies of these materials were carried out on Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive) bacteria in order to explore the impact of the extent of N-substitution (ES) on their biological activities. At ES less than 10%, the presence of the hydrophobic substituent, such as benzyl and thiophenylmethyl, yielded derivatives with lower MIC values than chitosan Quat-188. Derivatives with higher ES exhibited reduced antibacterial activity due to low quaternary ammonium moiety content. At the same degree of quaternization, all quaternized N-aryl chitosan derivatives bearing either electron-donating or electron-withdrawing substituents did not contribute antibacterial activity relative to chitosan Quat-188. Neither the functional group nor its orientation impacted the MIC values significantly.     

Insights in the antibacterial action of poly(methyloxazoline)s with a biocidal end group and varying satellite groups

The antimicrobial activity of poly(2-methyl-1,3-oxazoline)s (PMOX) with the antimicrobial N,N-dimethyldodecylammonium (DDA) end group is greatly dependent on the nature of the group at the distal end of the polymer, the satellite group. Three comparable PMOX with a DDA end group and different satellite groups (methyl, decyl, hexadecyl) were investigated with respect to the reasons for the huge differences in their biocidal behavior. Static light scattering (SLS) and pulsed field gradient diffusion NMR measurements revealed that the samples show comparable aggregation conduct, thus, not being responsible for the varying biological activity. Experiments using different liposomal systems as models for bacterial cell membranes have been performed. It was found that differential interactions between the respective polymers and the phospholipid membranes constitute the reason for the varying effectiveness observed in antimicrobial susceptibility determinations.     

Hyperbranched poly(amino ester)s with different terminal amine groups for DNA delivery

Hyperbranched poly(amino ester)s containing tertiary amines in the core and primary, secondary, and tertiary amines in the periphery, respectively, were evaluated for DNA delivery in vitro. The same core structure facilitated the investigation on the effects of the terminal amine type on the properties of hyperbranched poly(amino ester)s for DNA delivery. The hydrolysis of the poly(amino ester)s was monitored using (1)H NMR. The results reflected that the terminal amine type had negligible effects on the hydrolysis rate but was much slower than that of linear poly(amino ester)s, probably due to the compact hyperbranched spatial structure preventing the accessibility of water. In comparison with PEI 25 K, the hyperbranched poly(amino ester)s showed much lower cytotoxicity in Cos7, HEK293, and HepG2 cells. Gel electrophoresis indicated that poly(amino ester)s could condense DNA efficiently, and the zeta potentials and sizes of the complexes formed with different weight ratios of hyperbranched poly(amino ester)s and DNA were measured. Remarkably, all the hyperbranched poly(amino ester)s showed DNA transfection efficiency comparable to PEI 25 K in Cos7, HEK293, and HepG2 cells regardless of the terminal amine type. Therefore, the terminal amine type had insignificant effects on the hydrolysis rate, cytotoxicity, DNA condensation capability, and in vitro DNA transfection efficiency of the hyperbranched poly(amino ester)s.     

L-Phe end-capped poly(L-lactide) as macroinitiator for the synthesis of poly(L-lactide)-B-poly(L-lysine) block copolymer

A poly(L-lactide)-b-poly(Nepsilon-(Z)-L-lysine) (PLLA-b-PZLys) block copolymer was synthesized through the ring-opening polymerization of Nepsilon-(Z)-lysine-N-carboxyanhydride using L-Phe-terminated PLLA as a macroinitiator. The L-Phe-terminated PLLA was prepared through a novel three-step process. First, the hydroxyl-terminated PLLA was synthesized through the ring-opening polymerization of L-lactide initiated by n-butanol under the existence of tin(II) ethylhexanoate. Subsequently, the complete capping of the hydroxyl end group of PLLA with BOC-L-Phe was achieved by using a mixed anhydride of BOC-L-Phe under the catalysis of 4-(1-pyrrolidinyl) pyridine. Finally, the free amino end group was obtained by removal of the t-butoxycarbonyl group through trifluoroacetic acid treatment under anhydrous condition. All these treatments were conducted under mild conditions, thus avoiding the breakdown of the PLLA backbone. Poly(L-lactide)-b-poly(L-lysine) block copolymer was produced after deprotection treatment of PLLA-b-PZLys. The structure of the block copolymer was confirmed by 1H NMR, IR, and GPC. Adjustment of the ratio of the NCA monomer to the macroinitiator could control the chain length of the PLys block.     

Novel urea and thiourea derivatives of thiazole-glutamic acid conjugate as potential inhibitors of microbes and fungi

Since discovery and development of effective as well as safe drugs has brought a progressive era in human healthcare that is accompanied by the appearance of drug resistant bacterial strains, there is constant need of new antibacterial agent having novel mechanisms of action to act against the harmful pathogens. In the present study, several N-terminal substituted urea/thiourea derivatives were synthesized by the reaction of glutamic acid and 3-(1-piperazinyl)-1,2-benzisothiazole with various substituted phenyl isocyanates/isothiocyanates. Elemental analysis, IR, ¹H NMR, ¹³C NMR and mass spectral data confirmed the structure of the newly synthesized compounds. The derivatives were investigated for their antibacterial and antifungal activities against various pathogens of human origin by agar well diffusion method and microdilution method. The preliminary antimicrobial bioassay reveals that the compounds containing fluoro and bromo as substituents showed promising antimicrobial activity.     

Synthesis, characterization and antimicrobial activity of new aliphatic sulfonamide

A series of novel aliphatic sulfonamide derivatives (1-7) were synthesized and characterized by elemental analyses, FT-IR, (1)H NMR, (13)C NMR and LC-MS techniques. All the synthesized compounds were evaluated in vitro as antimicrobial agents against representative strains of Gram-positive (Staphylococcus aureus ATCC 25953, Bacillus cereus ATCC 6633 and Listeria monocytogenes ATCC Li6 (isolate), Gram-negative bacteria (Escherichia coli ATCC 11230) and antifungal agent against Candida albicans (clinical isolate) by both disc diffusion and minimal inhibition concentration (MIC) methods. All these bacteria and fungus studied were screened against some antibiotics to compare with our chemicals' zone diameters. Our aliphatic sulfonamides have highest powerful antibacterial activity for Gram-negative bacteria than Gram-positive bacteria and antibacterial activity decreases as the length of the carbon chain increases.     

Purification of a novel antibacterial short peptide in earthworm Eisenia foetida

Earthworms live in an environment with abundantpathogens. These pathogens are, firstly, bacteria living inwater or soil that are ingested during feeding or introducedinto the body following injury. Parasites, particularlylarval forms, which represent the dissemination phase,are another important group of potentially pathogenicagents. During the course of evolution, earthworms havedeveloped defense strategies against these living patho-gens [1,2]. Earthworms lack true antibodies and hence anada…     

Exploring symbiotic nitrogen fixation and assimilation in pea root nodules by in vivo 15N nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry

Nitrogen (N) fixation and assimilation in pea (Pisum sativum) root nodules were studied by in vivo (15)N nuclear magnetic resonance (NMR) by exposing detached nodules to (15)N(2) via a perfusion medium, while recording a time course of spectra. In vivo (31)P NMR spectroscopy was used to monitor the physiological state of the metabolically active nodules. The nodules were extracted after the NMR studies and analyzed for total soluble amino acid pools and (15)N labeling of individual amino acids by liquid chromatography-mass spectrometry. A substantial pool of free ammonium was observed by (15)N NMR to be present in metabolically active, intact nodules. The ammonium ions were located in an intracellular environment that caused a remarkable change in the in vivo (15)N chemical shift. Alkalinity of the ammonium-containing compartment may explain the unusual chemical shift; thus, the observations could indicate that ammonium is located in the bacteroids. The observed (15)N-labeled amino acids, glutamine/glutamate and asparagine (Asn), apparently reside in a different compartment, presumably the plant cytoplasm, because no changes in the expected in vivo (15)N chemical shifts were observed. Extensive (15)N labeling of Asn was observed by liquid chromatography-mass spectrometry, which is consistent with the generally accepted role of Asn as the end product of primary N assimilation in pea nodules. However, the Asn (15)N amino signal was absent in in vivo (15)N NMR spectra, which could be because of an unfavorable nuclear Overhauser effect. gamma-Aminobutyric acid accumulated in the nodules during incubation, but newly synthesized (15)N gamma-aminobutyric acid seemed to be immobilized in metabolically active pea nodules, which made it NMR invisible.     

Synthesis, characterization, and antibacterial activity of N,O-quaternary ammonium chitosan

N,N,N-Trimethyl O-(2-hydroxy-3-trimethylammonium propyl) chitosans (TMHTMAPC) with different degrees of O-substitution were synthesized by reacting O-methyl-free N,N,N-trimethyl chitosan (TMC) with 3-chloro-2-hydroxy-propyl trimethyl ammonium chloride (CHPTMAC). The products were characterized by ¹H NMR, FTIR and TGA, and investigated for antibacterial activity against Staphylococcus aureus and Escherichia coli under weakly acidic (pH 5.5) and weakly basic (pH 7.2) conditions. TMHTMAPC exhibited enhanced antibacterial activity compared with TMC, and the activity of TMHTMAPC increased with an increase in the degree of substitution. Divalent cations (Ba²⁺ and Ca²⁺) strongly reduced the antibacterial activity of chitosan, O-carboxymethyl chitosan and N,N,N-trimethyl-O-carboxymethyl chitosan, but the repression on the antibacterial activity of TMC and TMHTMAPC was weaker. This indicates that the free amino group on chitosan backbone is the main functional group interacting with divalent cations. The existence of 100 mM Na⁺ slightly reduced the antibacterial activity of both chitosan and its derivatives.     

Geminal dialkyl derivatives of N-substituted pantothenamides: synthesis and antibacterial activity

As a key precursor of coenzyme A (CoA) biosynthesis, pantothenic acid has proven to be a useful backbone to elaborate probes of this biosynthetic pathway, study CoA-utilizing systems, and design molecules with antimicrobial activity. The increasing prevalence of bacterial strains resistant to one or more antibiotics has prompted a renewed interest for molecules with a novel mode of antibacterial action such as N-substituted pantothenamides. Although numerous derivatives have been reported, most are varied at the terminal N-substituent, and fewer at the β-alanine moiety. Modifications at the pantoyl portion are limited to the addition of an ω-methyl group. We report a synthetic route to N-substituted pantothenamides with various alkyl substituents replacing the geminal dimethyl groups. Our methodology is also applicable to the synthesis of pantothenic acid, pantetheine and CoA derivatives. Here a small library of new N-substituted pantothenamides was synthesized. Most of these compounds display antibacterial activity against sensitive and resistant Staphylococcus aureus. Interestingly, replacement of the ProR methyl with an allyl group yielded a new N-substituted pantothenamide which is amongst the most potent reported so far.     

Novel synthesis of functionalized poly(3-hydroxybutanoic acid) and its copolymers.

Novel feasibility of fuctionalized poly(3-hydroxybutanoic acid), PHB, and its copolymers synthesis via ring-opening of beta-butyrolactone (ROP) mediated by activated anionic initiators or enzymes in vitro is presented. Using these new synthetic approaches, PHB with defined chemical structure of the end groups as well as block, graft and random copolymers have been obtained and characterized by IR, NMR, ESI-MS and GPC techniques. The relationship between the structure and properties of the novel polymeric materials prepared is discussed.     

Structure‐activity relationship studies of shortened analogues of the antimicrobial peptide EeCentrocin 1 from the sea urchin Echinus esculentus

EeCentrocin 1 is a potent antimicrobial peptide isolated from the marine sea urchin Echinus esculentus. The peptide has a hetero‐dimeric structure with the antimicrobial activity confined in its largest monomer, the heavy chain (HC), encompassing 30 amino acid residues. The aim of the present study was to develop a shorter drug lead peptide using the heavy chain of EeCentrocin 1 as a starting scaffold and to perform a structure‐activity relationship study with sequence modifications to optimize antimicrobial activity. The experiments consisted of 1) truncation of the heavy chain, 2) replacement of amino acids unfavourable for in vitro antimicrobial activity, and 3) an alanine scan experiment on the truncated and modified heavy chain sequence to identify essential residues for antimicrobial activity. The heavy chain of EeCentrocin 1 was truncated to less than half its initial size, retaining most of its original antimicrobial activity. The truncated and optimized lead peptide ( P6 ) consisted of the 12 N‐terminal amino acid residues from the original EeCentrocin 1 HC sequence and was modified by two amino acid replacements and a C‐terminal amidation. Results from the alanine scan indicated that the generated lead peptide ( P6 ) contained the optimal sequence for antibacterial activity, in which none of the alanine scan peptides could surpass its antimicrobial activity. The lead peptide ( P6 ) was also superior in antifungal activity compared to the other peptides prepared and showed minimal inhibitory concentrations (MICs) in the low micromolar range. In addition, the lead peptide ( P6 ) displayed minor haemolytic and no cytotoxic activity, making it a promising lead for further antimicrobial drug development.     

Synthesis and in vitro antibacterial activity of 3-[N-methyl-N-(3-methyl-1,3-thiazolium-2-yl)aminolmethyl cephalosporin derivatives

The new cephalosporins having N-linked quarternary ammonium salt at C-3 position were prepared from the reaction of 2-methylimino-3-methyl-1,3-thiazoline derivatives with cephalosporins. Also antimicrobial activities of those compounds were determined.