Species differences in the interaction of serum albumin with antibiotics

Interaction of oxacillin, cloxacillin, dicloxacillin, phenoxymethylpenicillin, methicillin, nafcillin and benzylpenicillin with human serum albumin (HSA) was studied with flow microcalorimetry and differential scanning calorimetry. The measured thermodynamic parameters of complex formation between the penicillins and HSA were compared with similar characteristics of their binding to bovine serum albumin. It was shown that there were species differences between these two globular proteins in their interaction with the above antibiotics in relation to both the number of the biopolymer active sites and the nature of the molecular forces in the complex formation. The effect of the first bound molecule of oxacillin, cloxacillin, dicloxacillin, nafcillin, phenoxymethylpenicillin and benzylpenicillin on HSA conformation was observed. It was demonstrated that there was thermostabilization of HSA on its interaction with the above drugs with preserving cooperative nature of thermal denaturation of the complexes in relation to HSA melting.     

Simultaneous determination of six penicillins in cows'' raw milk by a multiresidue high-performance liquid chromatographic method

A high-performance liquid chromatographic method based on C₁₈ solid-phase extraction and ultraviolet detection at 323 nm of analytes derivatized with benzoic anhydride and 1,2,4-triazole mercuric chloride solution was developed for the simultaneous determination of amoxicillin, penicillin G (benzylpenicillin), ampicillin, oxacillin, cloxacillin and dicloxacillin residues in raw milk. The detection limit of the method was 1.3 μg/l for penicillin G; 1.4 μg/l for amoxicillin, oxacillin and cloxacillin, 1.5 μg/l for ampicillin and 2.7 μg/l for dicloxacillin. The mean recovery was 95–102% for amoxicillin, penicillin G and ampicillin, 92–98% for oxacillin and cloxacillin and 87–94% for dicloxacillin, measured by using an internal standard. The relative repeatability standard deviation was 4–9% on level 4–15 μg/l, respectively, 2–7% on level 30–40 μg/l.     

Simultaneous determination of seven penicillins in muscle, liver and kidney tissues from cattle and pigs by a multiresidue high-performance liquid chromatographic method

A high-performance liquid chromatographic (HPLC) method based on solid-phase extraction (SPE) was developed for determination of amoxicillin, penicillin G (benzylpenicillin), ampicillin, oxacillin, cloxacillin, nafcillin and dicloxacillin in muscle, liver and kidney tissues of pigs and cattle. The compounds were extracted in aqueous solution by precipitation of organic materials with a mixture of sulphuric acid and sodium tungstate. The extract was cleaned up by SPE on a divinylbenzene-co-N-vinylpyrrolidone polymeric sorbent. Further clean-up was performed by liquid–liquid partition with diethyl ether. The extract was derivatised with benzoic anhydride and 1,2,4-triazole mercury (II) reagent. Chromatography was performed by reversed-phase gradient HPLC on a C₁₈ column with ultraviolet detection at 323 nm. The limits of detection estimated by a conservative model were in the range 8.9–11.1 μg/kg for amoxicillin, penicillin G, ampicillin, oxacillin, cloxacillin and nafcillin and 18.3–20.9 μg/kg for dicloxacillin. The mean recovery range was 66–77% for amoxicillin, 73–75% for penicillin G, 81–82% for ampicillin, 73–76% for oxacillin, 74–75% for cloxacillin, 66–72% for nafcillin and 58–65% for dicloxacillin.     

Detection and side-chain specificity of IgE antibodies to flucloxicillin in allergic subjects

Unlike studies on the antigenicity of penicillins in laboratory animals, limited information is available on the allergenicity of penicillins in man, especially with regard to fine structural allergenic differences between the many different penicillins. Inconsistent with the earlier conclusions of others, our studies suggest that side-chain structures to flucloxacillin-reactive IgE antibodies. Quantitative hapten inhibition studies revealed potent inhibition by flucloxacillin and three structurally related penicillins: oxacillin, cloxacillin and dicloxacillin. Analysis of the results showed that the side-chain group of flucloxacillin, 3-2(-chloro-6-fluorophenyl)-5-methyl-4-isoxazolyl, is recognized by some antibodies and that the 5-methyl-3-phenyl-4-isoxazolyl group, with or without halogen substituents, accounts for the reactivity of other antibodies and for the cross-reactions seen with some other penicillins. Since it is the side-chain group that distinguishes the many different types of penicillin, and since IgE antibodies in many of the allergic reactions recognize the side-chain groups, it is becoming clear that specific assays are required for the detection of IgE antibodies to each of the different penicillins.     

Penicillin combinations against multi-resistant urinary pathogens as an alternative to gentamycin treatment.

A total of 42 multi-resistant urinary pathogens, sensitive to gentamycin only, were investigated with regard to the effects of dicloxacillin and ampicillin or cloxacillin/flucloxacillin and amoxycillin in combination. A synergistic antibiotic effect with resulting low MIC levels was demonstrated for 81% of the strains. It was also found that a further increase in sensitivity might be achieved by adjustment of the pH level.     

Dicloxacillin: its chemotherapeutic action and experimental kinetic characteristics]

Activity against 50 clinical staphylococcal strains, chemotherapeutic efficiency, absorption and distribution of dicloxacillin in animals were studied in comparison with oxacillin. It was found that dicloxacillin was superior to oxacillin with respect to a number of their properties. The antibiotic was characterized by a higher antistaphylococcal activity, especially with respect to methicillin resistant strains. It was more effective in treatment of staphylococcal pneumonia and septicopiemia of albino mice. The drug was better absorpted when used orally and was detected in the blood in therapeutic concentrations for long periods of time. The antibacterial titer of the blood serum after dicloxacillin administration was 4 times higher than that of oxacillin.     

Cefoxitin,a New Semi-synthetic Cephamycin: An In-vitro and In-vivo Comparison with Cephalothin

The activity of cefoxitin was compared with that of cephalothin against 229 bacterial strains. Cefoxitin was more active against most Gram-negative strains, notably against indole-producing Proteus spp., which are usually resistant to the cephalosporins. Cefoxitin was not susceptible to any significant extent to degradation by β-lactamases produced by Gram-negative organisms. Against Gram-positive organisms, however, cefoxitin was considerably less active than cephalothin, but minimum inhibitory concentrations for Staphylococcus aureus were well within therapeutically attainable blood levels.Pharmacokinetic studies in 18 volunteers showed a higher and longer sustained antibiotic activity in serum and urine after injections of cefoxitin than after equal doses of cephalothin. Urinary recovery of cefoxitin activity was also much higher than that of cephalothin. No evidence of toxicity due to cefoxitin was found. Cefoxitin was slightly less painful after intramuscular injection than cephalothin.     

Comparative Inhibition of Methicillin-resistant Strains of Staphylococcus aureus by Lysostaphin and Otner Antibiotics

Sixteen methicillin-resistant strains of Staphylococcus aureus obtained from Europe were found to be sensitive to the lytic activity of lysotaphin. With only minor exceptions, the strains were found to be sensitive to novobiocin, erythromycin, fusidic acid, and lincomycin, and slightly less sensitive to vancomycin and chloramphenicol. All strains were resistant to tetracycline, penicillinase-sensitive penicillins (benzylpenicillin, ampicillin, and propicillin), penicillinase-resistant penicillins (methicillin, nafcillin, ancillin, oxacillin, cloxacillin, and dicloxacillin), and two cephalosporin antibiotics (cephalothin and cephaloridine).     

Selection and characterization of a carrot cell line tolerant to glyphosate

Cultured carrot (Daucus carota L.) cells were adapted to growing in 25 millimolar glyphosate by transfer into progressively higher concentrations of the herbicide. Tolerance was increased 52-fold, and the adaptation was stable in the absence of glyphosate. The uptake of glyphosate was similar for adapted and nonadapted cells. Activity of the enzyme 5-enolpyruvylshikimic acid-3-phosphate synthase was 12-fold higher in the adapted line compared to nonadapted cells, while activities of shikimate dehydrogenase and anthranilate synthase were similar in the two cell types. The adapted cells had higher levels of free amino acids—especially threonine, methionine, tyrosine, phenylalanine, tryptophan, histidine, and arginine—than did nonadapted cells. Glyphosate treatment caused decreases of 50 to 65% in the levels of serine, glycine, methionine, tyrosine, phenylalanine, and tryptophan in nonadapted cells, but caused little change in free amino acid levels in adapted cells.

The adaptation reported here supports the growing body of evidence linking tolerance to glyphosate with increased levels of the enzyme 5-enolpyruvylshikimic acid-3-phosphate synthase. The elevated levels of aromatic amino acids, which may confer resistance in adapted cells, suggest that control of the shikimate pathway may be altered in these cells.

     

ISOLATION OF NERVE ENDINGS FROM THE POSTERIOR PITUITARY GLAND : Electron Microscopy of Fractions Obtained by Centrifugation

Bovine posterior pituitary glands were homogenized in 10 per cent sucrose and fractionated by differential centrifugation. The following centrifugation procedure resulted in the most satisfactory separation: 1000 g for 15 minutes—nuclei, connective tissue, basement membranes with associated endothelium, giant nerve endings, and whole pituicytes; 4200 g for 15 minutes—free nerve endings, including Herring bodies; 17,000 g for 15 minutes—mitochondria; 68,000 g for 15 minutes—neurosecretory granules. Electron microscopic examination was carried out on whole tissue and on the isolated fractions. Isolated nerve endings were examined also by negative staining techniques. Isolated nerve endings retain an apparently normal complement of mitochondria, neurosecretory granules, and microvesicles ("synaptic" vesicles). The free nerve endings closely resemble those observed in sections of intact posterior pituitary tissue. Free microvesicles were not observed in any of the fractions isolated and apparently sediment at centrifugal forces higher than those employed in this study.     

Naturally produced opsonizing antibodies restrict the survival of Mycobacterium tuberculosis in human macrophages by augmenting phagosome maturation

This study investigated the hypothesis that serum antibodies against Mycobacterium tuberculosis present in naturally infected healthy subjects of a tuberculosis (TB) endemic area could create and/or sustain the latent form of infection. All five apparently healthy Indian donors showed high titres of serum antibodies against M. tuberculosis cell membrane antigens, including lipoarabinomannan and alpha crystallin. Uptake and killing of bacilli by the donor macrophages was significantly enhanced following their opsonization with antibody-rich, heat-inactivated autologous sera. However, the capability to opsonize was apparent for antibodies against some and not other antigens. High-content cell imaging of infected macrophages revealed significantly enhanced colocalization of the phagosome maturation marker LAMP-1, though not of calmodulin, with antibody-opsonized compared with unopsonized M. tuberculosis. Key enablers of macrophage microbicidal action—proinflammatory cytokines (IFN-γ and IL-6), phagosome acidification, inducible NO synthase and nitric oxide—were also significantly enhanced following antibody opsonization. Interestingly, heat-killed M. tuberculosis also elevated these mediators to the levels comparable to, if not higher than, opsonized M. tuberculosis. Results of the study support the emerging view that an efficacious vaccine against TB should, apart from targeting cell-mediated immunity, also generate ‘protective’ antibodies.     

Effects of exposure to pile-driving sounds on the lake sturgeon,Nile tilapia and hogchoker

Pile-driving and other impulsive sound sources have the potential to injure or kill fishes. One mechanism that produces injuries is the rapid motion of the walls of the swim bladder as it repeatedly contacts nearby tissues. To further understand the involvement of the swim bladder in tissue damage, a specially designed wave tube was used to expose three species to pile-driving sounds. Species included lake sturgeon (Acipenser fulvescens)—with an open (physostomous) swim bladder, Nile tilapia (Oreochromis niloticus)—with a closed (physoclistous) swim bladder and the hogchoker (Trinectes maculatus)—a flatfish without a swim bladder. There were no visible injuries in any of the exposed hogchokers, whereas a variety of injuries were observed in the lake sturgeon and Nile tilapia. At the loudest cumulative and single-strike sound exposure levels (SEL_cum and SEL_ss respectively), the Nile tilapia had the highest total injuries and the most severe injuries per fish. As exposure levels decreased, the number and severity of injuries were more similar between the two species. These results suggest that the presence and type of swim bladder correlated with injury at higher sound levels, while the extent of injury at lower sound levels was similar for both kinds of swim bladders.     

Treatment of subclinical mastitis in Damascus goats during lactation

This study was carried out to determine the efficacy of three treatment protocols for subclinical mastitis during lactation in Damascus goats. For this purpose intramammarian ampicillin dicloxacillin, intramuscular amoxycillin clavulonic acid and the combination of intramammarian ampicillin dicloxacillin and intramuscular amoxycillin clavulonic acid were used in goats with subclinical mastitis during lactation.The microbiological treatment rates in the intramammarian ampicillin dicloxacillin, intramuscular amoxycillin clavulonic acid and the combination of intramammarian ampicillin dicloxacillin and intramuscular amoxycillin clavulonic acid groups in the 7th and 21st days after the treatment were 62.5% and 92.5%, 62.5% and 70%, 87.5% and 87.5%, respectively. The intramammarian and intramuscular combination group was found to have a statistically higher treatment rate than other two groups on the 7th day. On the 21st day intramammarian and combination groups were found to have statistically better treatment rates than that of intramuscular group.It was concluded that the goat subclinical mastitis could be successfully treated during lactation. While the intramammarian ampicillin dicloxacillin treatment had the best treatment rates, the combination of intramuscular amoxycillin clavulonic acid was also successful. Intramuscular amoxycillin clavulonic acid as the sole treatment was not as effective as intramammarian therapy.     

Microbiological Activities of Lysostaphin and Penicillins Against Bacteriophage 80/81 Strains of Staphylococcus aureus

Using 20 clinical isolates of S. aureus (all bacteriophage 80/81 type), we found that lysostaphin inhibits the growth of all cultures at concentrations significantly lower than those observed with any of eight penicillins, a penicillin-like compound (cephalothin), or fusidic acid (a steroid antibiotic). All test cultures were shown to be resistant to penicillin G, ampicillin, and propicillin. Of the remaining penicillins (all penicillinase-insensitive), oxacillin, nafcillin, cloxacillin, and cephalothin were approximately equal in antimicrobial activity. Ancillin was slightly less active, and methicillin was even lower in potency. Cultures varied more widely in susceptibility to fusidic acid. None of the clinical isolates tested was found to be resistant to lysostaphin.     

Localization and Quantitative Determination of Ferredoxin-NADP Oxidoreductase, a Thylakoid-Bound Enzyme in the Cyanobacterium Anabaena sp. Strain 7119

Thylakoid membrane preparations obtained from mechanically disrupted (sonicated) cells of the cyanobacterium Anabaena sp. strain 7119 show a membrane-bound ferredoxin-NADP⁺ oxidoreductase (EC 1.18.1.2) as determined either by specific antibodies or by using the ferredoxin-dependent NADPH-cytochrome c reductase activity, which is a specific test for this enzyme. However, in contrast with higher plant thylakoids, a low yield of the cyanobacterial reductase—only about 20% of the total amount of this protein estimated in whole cell homogenates—was obtained as a membrane-bound form when Mg²⁺ was present during the disruption treatment. It is noteworthy that the addition of water-soluble nonionic polymers, namely polyethylene glycol and polyyinylpyrrolidone, dramatically increased the yield of the thylakoid-bound reductase, reaching values up to 80 to 85% of the total enzyme. Using these thylakoid membrane preparations, a quantitative determination of the reductase has been performed for the first time for cyanobacterial thylakoids. The value determined by immunoelectrophoresis—from 8 to 10 nanomoles per micromole of chlorophyll—is clearly higher than those reported for chloroplast thylakoids.     

Novel N-benzylpyridinium moiety linked to arylisoxazole derivatives as selective butyrylcholinesterase inhibitors: Synthesis,biological evaluation,and docking study

A novel series of N-benzylpyridinium moiety linked to arylisoxazole ring were designed, synthesized, and evaluated for their in vitro acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities. Synthesized compounds were classified into two series of 5a-i and 5j-q considering the position of positively charged nitrogen of pyridinium moiety (3- or 4- position, respectively) connected to isoxazole carboxamide group. Among the synthesized compounds, compound 5n from the second series of compounds possessing 2,4-dichloroaryl group connected to isoxazole ring was found to be the most potent AChE inhibitor (IC₅₀ = 5.96 µM) and compound 5j also from the same series of compounds containing phenyl group connected to isoxazole ring demonstrated the most promising inhibitory activity against BChE (IC₅₀ = 0.32 µM). Also, kinetic study demonstrated competitive inhibition mode for both AChE and BChE inhibitory activity. Docking study was also performed for those compounds and desired interactions with those active site amino acid residues were confirmed through hydrogen bonding as well as π-π and π-anion interactions. In addition, the most potent compounds were tested against BACE1 and their neuroprotectivity on Aβ-treated neurotoxicity in PC12 cells which depicted negligible activity. It should be noted that most of the synthesized compounds from both categories 5a-i and 5j-q showed a significant selectivity toward BChE. However, series 5j-q were more active toward AChE than series 5a-i.     

A Meta-Analysis of Oxidative Stress Markers in Depression

ObjectStudies have suggested that depression was accompanied by oxidative stress dysregulation, including abnormal total antioxidant capacity (TAC), antioxidants, free radicals, oxidative damage and autoimmune response products. This meta-analysis aims to analyse the clinical data quantitatively by comparing the oxidative stress markers between depressed patients and healthy controls.MethodsA search was conducted to collect the studies that measured the oxidative stress markers in depressed patients. Studies were searched in Embase, Medline, PsychINFO, Science direct, CBMDisc, CNKI and VIP from 1990 to May 2015. Data were subjected to meta-analysis by using a random effects model for examining the effect sizes of the results. Bias assessments, heterogeneity assessments and sensitivity analyses were also conducted.Results115 articles met the inclusion criteria. Lower TAC was noted in acute episodes (AEs) of depressed patients (p<0.05). Antioxidants, including serum paraoxonase, uric acid, albumin, high-density lipoprotein cholesterol and zinc levels were lower than controls (p<0.05); the serum uric acid, albumin and vitamin C levels were increased after antidepressant therapy (p<0.05). Oxidative damage products, including red blood cell (RBC) malondialdehyde (MDA), serum MDA and 8-F₂-isoprostanes levels were higher than controls (p<0.05). After antidepressant medication, RBC and serum MDA levels were decreased (p<0.05). Moreover, serum peroxide in free radicals levels were higher than controls (p<0.05). There were no differences between the depressed patients and controls for other oxidative stress markers.ConclusionThis meta-analysis supports the facts that the serum TAC, paraoxonase and antioxidant levels are lower, and the serum free radical and oxidative damage product levels are higher than controls in depressed patients. Meanwhile, the antioxidant levels are increased and the oxidative damage product levels are decreased after antidepressant medication. The pathophysiological relationships between oxidative stress and depression, and the potential benefits of antioxidant supplementation deserve further research.     

Regulation of Lipid Droplet Size in Mammary Epithelial Cells by Remodeling of Membrane Lipid Composition—A Potential Mechanism

Milk fat globule size is determined by the size of its precursors—intracellular lipid droplets—and is tightly associated with its composition. We examined the relationship between phospholipid composition of mammary epithelial cells and the size of both intracellular and secreted milk fat globules. Primary culture of mammary epithelial cells was cultured in medium without free fatty acids (control) or with 0.1 mM free capric, palmitic or oleic acid for 24 h. The amount and composition of the cellular lipids and the size of the lipid droplets were determined in the cells and medium. Mitochondrial quantity and expression levels of genes associated with mitochondrial biogenesis and polar lipid composition were determined. Cells cultured with oleic and palmitic acids contained similar quantities of triglycerides, 3.1- and 3.8-fold higher than in controls, respectively (P < 0.0001). When cultured with oleic acid, 22% of the cells contained large lipid droplets (>3 μm) and phosphatidylethanolamine concentration was higher by 23 and 63% compared with that in the control and palmitic acid treatments, respectively (P < 0.0001). In the presence of palmitic acid, only 4% of the cells contained large lipid droplets and the membrane phosphatidylcholine concentration was 22% and 16% higher than that in the control and oleic acid treatments, respectively (P < 0.0001). In the oleic acid treatment, approximately 40% of the lipid droplets were larger than 5 μm whereas in that of the palmitic acid treatment, only 16% of the droplets were in this size range. Triglyceride secretion in the oleic acid treatment was 2- and 12-fold higher compared with that in the palmitic acid and control treatments, respectively. Results imply that membrane composition of bovine mammary epithelial cells plays a role in controlling intracellular and secreted lipid droplets size, and that this process is not associated with cellular triglyceride content.     

The Staphylococcus aureus Protein Sbi Acts as a Complement Inhibitor and Forms a Tripartite Complex with Host Complement Factor H and C3b

The Gram-positive bacterium Staphylococcus aureus, similar to other pathogens, binds human complement regulators Factor H and Factor H related protein 1 (FHR-1) from human serum. Here we identify the secreted protein Sbi (Staphylococcus aureus binder of IgG) as a ligand that interacts with Factor H by a—to our knowledge—new type of interaction. Factor H binds to Sbi in combination with C3b or C3d, and forms tripartite Sbi∶C3∶Factor H complexes. Apparently, the type of C3 influences the stability of the complex; surface plasmon resonance studies revealed a higher stability of C3d complexed to Sbi, as compared to C3b or C3. As part of this tripartite complex, Factor H is functionally active and displays complement regulatory activity. Sbi, by recruiting Factor H and C3b, acts as a potent complement inhibitor, and inhibits alternative pathway-mediated lyses of rabbit erythrocytes by human serum and sera of other species. Thus, Sbi is a multifunctional bacterial protein, which binds host complement components Factor H and C3 as well as IgG and β₂-glycoprotein I and interferes with innate immune recognition.     

(—) S-adenosyl-L-methionine-magnesium Protoporphyrin Methyltransferase, an Enzyme in the Biosynthetic Pathway of Chlorophyll in Zea mays

The enzyme (—) S-adenosyl-L-methionine-magnesium protoporphyrin methyltransferase, which catalyzes the transfer of the methyl group from (—) S-adenosyl-L-methionine to magnesium protoporphyrin to form magnesium protoporphyrin monomethyl ester, has been detected in chloroplasts isolated from Zea mays.

Zinc protoporphyrin and free protoporphyrin also act as substrates in the system, although neither one is as active as magnesium protoporphyrin.

The following scheme of chlorophyll synthesis in higher plants is proposed: δ-aminolevulinic acid → → → protoporphyrin → magnesium protoporphyrin → magnesium protoporphyrin monomethyl ester → → → chlorophyll a.