Reactions of a sulfonamide antimicrobial with model humic constituents: assessing pathways and stability of covalent bonding

The mechanism of covalent bond formation of the model sulfonamide sulfathiazole (STZ) and the stronger nucleophile para-ethoxyaniline was studied in reactions with model humic acid constituents (quinones and other carbonyl compounds) in the absence and presence of laccase. As revealed by high resolution mass spectrometry, the initial bonding of STZ occurred by 1,2- and 1,4-nucleophilic additions of the aromatic amino group to quinones resulting in imine and anilinoquinone formation, respectively. Experiments using the radical scavenger tert-butyl-alcohol provided the same products and similar formation rates as those without scavenger indicating that probably not radical coupling reactions were responsible for the initial covalent bond formation. No addition with nonquinone carbonyl compounds occurred within 76 days except for a slow 1,4-addition to the β-unsaturated carbonyl 1-penten-3-one. The stability of covalent bonds against desorption and pressurized liquid extraction (PLE) was assessed. The recovery rates showed no systematic differences in STZ extractability between the two product types. This suggests that the strength of bonding is not controlled by the initial type of bond, but by the extent of subsequent incorporation of the reaction product into the formed polymer. This incorporation was monitored for (15)N aniline by (1)H-(15)N HMBC NMR spectroscopy. The initial 1,2- and 1,4-addition bonds were replaced by stronger heterocyclic forms with increasing incubation time. These processes could also hold true for soils, and a slow nonextractable residue formation with time could be related to a slow increase of the amount of covalently bound sulfonamide and the strength of bonding.     

Adsorption of sulfonamide antimicrobial agents to clay minerals

Adsorption of three sulfonamide antimicrobials to clay minerals was investigated as a function of pH, ionic strength, and type of exchangeable cation. Sulfonamide antimicrobial adsorption exhibited pronounced pH dependence consistent with sorbate speciation and clay properties. Sulfonamide antimicrobials did not intercalate into montmorillonite, and surface charge density influenced sorption by determining adsorption domain size. Adsorption edge data were best fit to a model including terms for the cationic and uncharged species. Adsorption of uncharged sulfamethazine to montmorillonite was relatively insensitive to pH, ionic strength, and type of exchangeable cation, while that to kaolinite was highly sensitive to ionic strength. Adsorption of cationic sulfamethazine to montmorillonite exceeded that of the neutral species by 1-2 orders of magnitude, but was unimportant for kaolinite atthe pH values examined. Cation exchange appeared to contribute to sorption of cationic sulfonamide species to montmorillonite. Anionic sulfamethazine adsorption was negligible. The nature of the sulfonamide R group influenced the degree of adsorption of cationic and neutral species. Our results highlight the importance of considering sulfonamide speciation and clay surface charge density in predicting the transport of these antimicrobials.     

NMR investigation of enzymatic coupling of sulfonamide antimicrobials with humic substances

Phenoloxidases mediate the oxidative transformation of soil phenolic constituents, contributing to the formation of humic substances and the chemical incorporation of some xenobiotic organic compounds into natural organic matter. We previously demonstrated phenoloxidase-mediated covalent coupling of sulfonamide antimicrobials with model humic constituents. Here, we investigate fungal peroxidase-mediated covalent coupling of 13C-sulfamethazine and 15N-sulfapyridine to humic substances. 1H-13C heteronuclear single quantum correlation (HSQC) nuclear magnetic resonance spectroscopy provided an initial indication of peroxidase-mediated covalent binding of 13C-sulfamethazine to humic acid. To confirm the role of the sulfonamide anilinic nitrogen in coupling to humic acid and to determine the nature of the covalent linkage, we incubated 15N-sulfapyridine with humic acid and peroxidase and examined reaction products in 1H-15N heteronuclear multiple bond (HMBC) experiments. The HMBC spectra revealed the presence of Michael adducts (i.e., anilinohydroquinones, anilinoquinones) and possibly other covalent linkages. No evidence for Schiff base formation was observed. Analogous experiments with the model humic constituent catechol provided corroborating evidence for these assignments. Michael adducts are expected to exhibit greater environmental stability than imine linkages that can form between sulfonamides and 2,6-dimethoxyphenols. Because the free anilinic nitrogen is required for the bioactivity of sulfonamide antimicrobials, nucleophilic addition occurring through this moiety could result in the biochemical inactivation of these compounds.     

Essential oils and their principal constituents as antimicrobial agents for synthetic packaging films

Spices and herbal plant species have been recognized to possess a broad spectrum of active constituents that exhibit antimicrobial (AM) activity. These active compounds are produced as secondary metabolites associated with the volatile essential oil (EO) fraction of these plants. A wide range of AM agents derived from EOs have the potential to be used in AM packaging systems which is one of the promising forms of active packaging systems aimed at protecting food products from microbial contamination. Many studies have evaluated the AM activity of synthetic AM and/or natural AM agents incorporated into packaging materials and have demonstrated effective AM activity by controlling the growth of microorganisms. This review examines the more common synthetic and natural AM agents incorporated into or coated onto synthetic packaging films for AM packaging applications. The focus is on the widely studied herb varieties including basil, oregano, and thyme and their EOs.     

Interaction of the macrolide antimicrobial clarithromycin with dissolved humic acid

The association of the cationic macrolide antimicrobial clarithromycin (CLA) with dissolved Elliot soil humic acid (ESHA) was investigated as a function of solution chemistry. CLA-ESHA association was strongly pH-dependent, reaching a maximum near pH 6.5, and was modeled successfully using FITEQL as a 1:1 complexation reaction between CLA+ and discreet deprotonated acidic functional groups with pKa values of 4 and 6. Approximate order of magnitude increases in ionic strength produced approximately 10-fold decreases in CLA+ -ESHA association. Coefficients for CLA+ -ESHA association were significantly smaller in the presence of K+ vs Na+. Sorption data were well-fit by the Freundlich model; the Freundlich exponent was <1, suggesting CLA+ interacted with sites having a range of binding energies. Sorption appeared largely reversible; little sorption-desorption hysteresis was observed. The affinities of erythromycin and CLA+ for ESHA association sites were indistinguishable, suggesting interaction with specific sorption sites. Comparison of experimentally determined CLA-ESHA association coefficients with those predicted from single-parameter linear free energy relationships based on log Kow suggested limited contribution of hydrophobic interactions to CLA-ESHA association at environmentally relevant pH values. CLA-ESHA association constants were similar in magnitude (10(3.9)-10(4.6) to those of many nonpolar organic contaminants, and macroscopic binding data were consistent with cation exchange dominating CLA+ -ESHA association.     

植物源抑菌剂的研究进展

植物源抑菌剂是通过物理或化学分离提取方法从天然植物中获得的具有抑菌作用的生物活性物质,具有安全、高效、毒副作用小等优点,在抑制病原菌及有害微生物生长繁殖、保障食品安全方面有较高的开发潜力和研究价值。植物源抑菌剂种类丰富,主要包括多酚、多糖、黄酮、挥发油、生物碱、萜类化合物等。本文分析总结了常见植物源抑菌剂的提取及纯化方法,讨论了提取方法选择过程中存在的问题,探究了不同种类植物源抑菌剂的作用机理及在食品保藏、加工中的应用现状。植物源抑菌剂在抑菌防腐领域具有较广阔的应用前景,未来可在提取工艺优化、有效成分分析及抑菌基因表达等方面进行深入研究,以期为植物源抑菌剂的综合利用提供一定的理论依据。     

Biopolymer-based films as carriers of antimicrobial agents

Films were prepared by incorporating different levels of antibacterial agents such as oregano oil, sodium lactate (NaL) and ɛ-polylysine (ɛ-PL) into sorbitol-plasticized whey protein isolate (WPI) films. The moisture uptake behaviour and the water vapour permeability (WVP) of the films were only affected by NaL, as the water sorption and permeability increased with addition of NaL into the protein matrix. An increase of the glass transition temperature of the sorbitol regions, as assessed by Dynamic Mechanical Thermal Analysis (DMTA), was caused by the addition of ɛ-PL, while incorporation of the oregano oil caused plasticization of the film that was depicted by a decrease in the transition temperature of the polymer-rich regions. On the other hand, incorporation of NaL into the films did not significantly alter their thermo-mechanical properties. However, the addition of NaL or ɛ-PL in the film forming solution resulted in a decline of maximum tensile strength (σ_max). Wrapping of beef cuts with the antimicrobial films resulted in a significant reduction of the bacterial population levels. The maximum specific growth rate (μ_max) of total flora (Total Viable Count, TVC) and pseudomonads was significantly reduced (P<0.05), with the use of antimicrobial films containing relatively high levels of oregano oil (1.5% w/w in the film forming solution) or ɛ-PL (0.75% w/w in the film forming solution), while the growth of Lactic Acid Bacteria (LAB) was completely inhibited.     

Efficacy of polyethylene-based antimicrobial films containing principal constituents of basil

The feasibility of low-density polyethylene (LDPE)-based films containing linalool or methylchavicol as antimicrobial (AM) packages to retard microbial growth on food surfaces was investigated. The AM LDPE-based films were tested for inhibition against selected microorganisms. Both compounds retained their AM activity, after an extrusion film-blowing process, against Escherichia coli in solid medium. Cheddar cheese was wrapped with the AM films and the packaged cheese samples were stored at 4 °C. The changes in the mesophilic aerobic bacteria and coliform, as well as yeast and mould counts were monitored. In addition, cheese samples inoculated with E. coli or Listeria innocua were wrapped with the AM films, stored at refrigerated (4 °C) or at abuse (12 °C) temperatures and the count of these microorganisms was monitored as a function of time. The results showed an inhibitory effect of these AM films against microbial growth in naturally contaminated cheese and in inoculated samples. The effect on suppression of E. coli and L. innocua growth was more pronounced at the abuse temperature. Methylchavicol-LDPE-based film exhibited a higher efficacy of inhibition than that of linalool-LDPE-based film. In addition, a sensory evaluation was performed with regards to possible taint in the flavour of the cheese. Taint in flavour as affected by linalool or methylchavicol was not significantly detectable by the panelists at the end of the storage period of 6 weeks. This study shows the potential use of polymeric films containing the principal constituents of basil as the AM components for enhancing quality and safety of cheeses.     

Susceptibility of Lactobacillus spp. to antimicrobial agents

Bacteria used as probiotics or in starter cultures may serve as hosts of antibiotic resistance genes, which can be transferred to pathogenic bacteria. Before launching a starter culture or a probiotic product into the market, it is therefore important to verify that the single bacterial isolates (strains) do not contain transferable resistance genes. A study has been undertaken to establish the levels of susceptibility of Lactobacillus spp. to various antimicrobial agents. This is a prerequisite for differentiating putative transferable resistance from natural resistance. A selection of 62 strains has been screened with the use of the Etest (ABBiodisk, Stockholm, Sweden) for their susceptibility to 25 antimicrobial agents. The strains belonged to the following species: Lactobacillus plantarum/pentosus, L. rhamnosus, L. paracasei, L. sakei, L. curvatus and species of the L. acidophilus group: L. johnsonii, L. crispatus, L. gasseri, and L. acidophilus.The results from the Etests have shown that the level of susceptibility to the antimicrobial agents is species-dependent. For the following antimicrobial agents, susceptibility varied several folds between species: vancomycin, teicoplanin, tetracycline, norfloxacin, ciprofloxacin, fusidic acid, and clindamycin. The differences between the species were more subtle for the rest of the tested antimicrobial agents. On the basis of the result, it was possible to suggest minimal inhibition concentrations (MICs) for the individual Lactobacillus species to be used as a microbiological breakpoint when screening strains for transferable resistance genes.     

香草精油作为抗菌剂的应用

细菌对多种抗生素具有耐药性是一个健康问题。精油(EOs)具有抗菌作用,并已筛选出潜在的新型抗菌化合物。萜类化合物是从精油(EOs)中分离出来的。这些精油(EOs)具有抑制金黄色葡萄球菌活性的作用。香芹酚对金黄色葡萄球菌和表皮葡萄球菌具有特定的作用。紫苏油具有抑制α-毒素、葡萄球菌肠毒素A和B以及中毒性休克综合征毒素表达的作用。香叶醇对一些革兰氏阴性菌在调节耐药性方面具有良好的活性。精油(EOs)作为生物保鲜剂,能减少或消除致病菌并提高动植物食品的整体质量。虽然临床研究很少,但精油(EOs)局部给药并作为防腐剂的渗透增强剂具有很好前景。也有少部分用于口服。     

Controlled fermentation of kimchi using naturally occurring antimicrobial agents

Kimchi is a traditional Korean fermented food. Since it ferments continuously during distribution and storage, the extension of shelf life by preventing over-acidification is a major concern in the kimchi industry. One of the most frequently attempted ways to delay fermentation is to add naturally occurring antimicrobial agents. Many researchers have investigated ways to delay over-acidification by adding minor ingredients, fruits or fruit seed extracts, extracts of medicinal herbs, culinary herbs and spices, and other miscellaneous substances to kimchi. The addition of naturally occurring antimicrobial agents may enhance the acceptability of kimchi to consumers over a longer period of time but may also have a disadvantage in that it may cause changes in sensory quality, especially if added in large amounts. To avoid undesirable sensory changes, application of hurdle technologies (i.e., multifactor preservative systems) which involve using combinations of low amounts of various naturally occurring antimicrobial agents as ingredients should be explored with the goal of controlling fermentation. If synergistic or additive antimicrobial effects can be achieved using small amounts of a combination of natural agents, changes in sensory qualities will be minimized, thereby prolonging shelf life. Research findings summarized in this review provide a basis for developing effective hurdle technologies using naturally occurring antimicrobial agents to extend shelf life of kimchi and perhaps other types of traditional fermented foods.     

抗菌剂抗菌机理简述

该文对现有抗菌剂进行了简单分类,主要包括无机系、有机系和天然生物抗菌剂三大类,并且简单阐述了无机系、有机系和天然生物抗菌剂的种类、特点以及抗菌机理,并指出不同类别抗菌剂存在的问题,对抗菌剂未来发展的方向进行了展望。     

Decontamination of cattle hides prior to slaughter using washes with and without antimicrobial agents

Two trials were conducted to determine the efficacy of cattle wash treatments in reducing pathogens on hides of cattle before slaughter. In trial I, live cattle (n = 120) were washed in an automated, commercial cattle wash system with one of four treatments (single water wash, double water wash, water wash with 0.5% L-lactic acid, or water wash with 50 ppm chlorine). Samples were collected at three locations (brisket, belly, and inside round) pre- and posttreatment to evaluate the effectiveness of treatments on the reduction of aerobic plate counts, coliforms, Escherichia coli and the incidence of Salmonella. For all three locations, bacterial numbers increased from 0.1 to 0.8 log CFU/cm2 posttreatment. In trial II, hide samples were inoculated in the laboratory with 6.0 log CFU/cm2 of rifampicin-resistant Salmonella serotype Typhimurium. Hide wash treatments included higher concentrations of chlorine (100, 200, and 400 ppm) and L-lactic acid (2, 4, and 6%), as well as other antimicrobial agents such as ethanol (70, 80, and 90%), acetic acid (2, 4, and 6%), and Oxy-Sept 333 (0.5, 2, and 4%). Spray wash treatments with ethanol and 4 to 6% concentrations of lactic acid had greater (P < 0.05) mean log reductions than 2% solutions of acetic or lactic acid, as well as 100, 200, and 400 ppm chlorine and the control water wash treatment. Spray wash treatments with Oxy-Sept 333 and 100, 200, or 400 ppm chlorine were not effective (P > 0.05) in reducing Salmonella Typhimurium compared to the (control) distilled water spray wash treatment. Several effective cattle hide interventions were identified in a controlled laboratory setting, but the high concentrations required for effectiveness would likely present problems from an animal welfare standpoint.     

Bark extractives from spruce as constituents of plywood bonding agents

The principles of the use of tannins and resins extracted from wood bark have been known since the 1950's. But only in recent years research on tannins has become lively. The literature review at the beginning of this paper concentrates mainly on the latest investigations in this field. The experimental part investigates the amount and quality of spruce bark extracts and their suitability as extenders in plywood adhesives. The use of chemically treated bark (bark dispersion) in plywood gluing is studied in the last part. If spruce bark is extracted with distilled water, the extract yield is 11.2% at the most, but when 10% of NaOH is added the yield increases to 40.0%. The extract content of drum-debarked pulpwood bark was clearly lower than that of unfloated logs, debarked in the sawmill. The results obtained reinforce the earlier belief that spruce bark extract can substitute Quebracho as an extender for plywood adhesives. Bark dispersion can also be used for this purpose. Exterior-grade birch plywood can be manufactured, using glues which contain spruce bark extract and dispersion. The extract and dispersion can also substitute part of the phenolic resin in the glue mixture, up to 20% at best, without any deterioration in the quality of gluing.     

Chemical investigation and antimicrobial properties of mastic water and its major constituents

Mastic water is a commercial flavouring obtained during the steam distillation of mastic resin (the resin of Pistacia lentiscus var. chia) for the production of mastic oil. The mastic water extracts were analysed by GC−MS. The major compounds identified were verbenone, α-terpineol, linalool and trans-pinocarveol. Overall the composition was found to be very different from that of mastic oil. Additional GC−MS revealed the enantiomeric ratio of the chiral constituents of mastic water. The antimicrobial activity of mastic water extract, as well as that of its major constituents, was examined against Escherichia coli, Staphylococcus aureus and Candida spp. including ATCC wild clinical and food-borne strains. Linalool and α-terpineol were found to be the most potent antimicrobial constituents. Finally the stability of mastic water at different temperatures was studied, showing no change in the GC−MS profile of the organic extract for a period of 4 months at storage temperatures up to 4 °C.     

Improved agar diffusion method for detecting residual antimicrobial agents

The improved agar diffusion method for determination of residual antimicrobial agents was investigated, and the sensitivities of various combinations of test organisms and assay media were determined using 7 organisms, 5 media, and 31 antimicrobial agents. Bacillus stearothermophilus and synthetic assay medium (SAM) showed the greatest sensitivity for screening penicillins (penicillin G and ampicillin). The combination of Bacillus subtilis and minimum medium (MM) was the most sensitive for tetracyclines (oxytetracycline and chlortetracycline), B. stearothermophilus and SAM or Micrococcus luteus and Mueller-Hinton agar (MHA) for detecting tylosin and erythromycin, B. subtilis and MHA for aminoglycosides (streptomycin, kanamycin, gentamicin, and dihydrostreptomycin), B. stearothermophilus and SAM for polyethers (salinomycin and lasalocid), and B. subtilis and MM or Clostridium perfringens and GAM for polypeptides (thiopeptin, enramycin, virginiamycin, and bacitracin). However, gram-negative bacterium Escherichia coli ATCC 27166 and MM were better for screening for colistin and polymixin-B. For detecting the synthetic drugs tested, the best combination was B. subtilis and MM for sulfonamides, E. coli 27166 and MM for quinolones (oxolinic acid and nalidixic acid), B. subtilis and MM for furans (furazolidone), and the bioluminescent bacterium Photobacterium phosphoreum and luminescence assay medium for chloramphenicol and oxolinic acid. The results showed that the use of four assay plates, B. stearothermophilus and SAM, B. subtilis and MM, M. luteus and MHA, and E. coli 27166 and MM, was superior to the currently available techniques for screening for residual antimicrobial agents in edible animal tissues.     

海水小球藻中脂类物质的成分分析及其抗菌活性研究

采用GC-MS方法对小球藻中脂类成分进行分析,从中鉴定出9种成分,其中十六烷酸最高,占24.88%。EPA含量为18.45%。抗菌活性实验显示,脂肪酸甲酯成分对金黄色葡萄球菌和枯草杆菌的抑菌效果要好于大肠杆菌和绿脓杆菌,而且EPA甲酯的抑菌效果要好于粗脂甲酯和PUFA甲酯。      

Mitigating the antimicrobial activities of selected organic acids and commercial sanitizers with various neutralizing agents

This study was conducted to evaluate the abilities of five neutralizing agents, Dey-Engley (DE) neutralizing broth (single or double strength), morpholinepropanesulfonic acid (MOPS) buffer, phosphate-buffered saline (PBS), and sodium thiosulfate buffer, in mitigating the activities of acetic or lactic acid (2%) and an alkaline or acidic sanitizer (a manufacturer-recommended concentration) againt the cells of Shiga toxin-producing Escherichia coli (STEC; n = 9). To evaluate the possible toxicity of the neutralizing agents to the STEC cells, each STEC strain was exposed to each of the neutralizing agents at room temperature for 10 min. Neutralizing efficacy was evaluated by placing each STEC strain in a mixture of sanitizer and neutralizer under the same conditions. The neutralizing agents had no detectable toxic effect on the STEC strains. PBS was least effective for neutralizing the activity of selected organic acids and sanitizers. Single-strength DE and sodium thiosulfate neutralized the activity of both acetic and lactic acids. MOPS buffer neutralized the activity of acetic acid and lactic acid against six and five STEC strains, respectively. All neutralizing agents, except double-strength DE broth, had a limited neutralizing effect on the activity of the commercial sanitizers used in the study. The double-strength DE broth effectively neutralized the activity of the two commercial sanitizers with no detectable toxic effects on STEC cells.     

海水小球藻中脂类物质的成分分析及其抗菌活性研究

采用GC-MS方法对小球藻中脂类成分进行分析,从中鉴定出9种成分,其中十六烷酸最高,占24.88％.EPA含量为18.45％.抗菌活性实验显示,脂肪酸甲酯成分对金黄色葡萄球菌和枯草杆菌的抑菌效果要好于大肠杆菌和绿脓杆菌,而且EPA甲酯的抑菌效果要好于粗脂甲酯和PUFA甲酯.