An in vitro method for the quantitative determination of the antimicrobial efficacy of silver-containing wound dressings

Treatment with silver-containing wound dressings is becoming an increasingly popular strategy to eliminate growth of opportunistic wound pathogens during the healing process. However, there are concerns over the possible side-effects of silver to the patient; coupled to the cost of silver as an ingredient there is a desire to ensure that wound dressings contain the least quantity of active ingredient to ensure the minimum bactericidal concentration (MBC) of silver is maintained in the wound environment. This requires the ability to determine the efficacy of silver directly within the wound environment; an extremely complicated task that is difficult using classical (plate counting) microbiological assays because these cannot be conducted in situ. Here, we report a quantitative method for determining the efficacy of silver in wound dressings using an isothermal calorimetric method. The growth curves of P. aeruginosa (NCIMB 8628) were recorded in growth medium and in growth medium containing AQUACEL Ag Hydrofiber dressing. It was found that 10 mg of dressing was sufficient to ensure no detectable growth of organism in 2.5 mL of medium inoculated to 10(6) cfu/mL. This corresponded to a silver load of 1.1x10(-6) moles (equivalent to 4.4x10(-4) M, in the volume of medium used in the experiment). Experiments conducted with silver nitrate rather than dressing indicated the MBC of silver against P. aeruginosa was 1x10(-4) M. The results suggested that not all of the silver in the dressing was bioavailable, at least over the lifetime of the experiment. One advantage of this effect would be the lack of excess availability of the silver, which allays fears of potential toxicity to the patient and may provide an extended period of time over which the dressing is bactericidal.     

New translational assays for preclinical modelling of cognition in schizophrenia: the touchscreen testing method for mice and rats

We describe a touchscreen method that satisfies a proposed 'wish-list' of desirables for a cognitive testing method for assessing rodent models of schizophrenia. A number of tests relevant to schizophrenia research are described which are currently being developed and validated using this method. These tests can be used to study reward learning, memory, perceptual discrimination, object-place associative learning, attention, impulsivity, compulsivity, extinction, simple Pavlovian conditioning, and other constructs. The tests can be deployed using a 'flexible battery' approach to establish a cognitive profile for a particular mouse or rat model. We have found these tests to be capable of detecting not just impairments in function, but enhancements as well, which is essential for testing putative cognitive therapies. New tests are being continuously developed, many of which may prove particularly valuable for schizophrenia research.     

New cytokine dressings. I. Kinetics of the in vitro rhG-CSF, rhGM-CSF, and rhEGF release from the dressings.

Wound repair-stimulatory activities of various cytokines and growth factors depend on successful delivery of these factors to the injured sites. Here were present the design and preparation of the new collagen- and polyurethane-based dressings containing the recombinant human cytokines-rhG-CSF, rhGM-CSF or rhEGF. To test the efficacy of the retrieval of the incorporated cytokines, their controlled release from the dressings was carried out over three consecutive days using polyurethane sponge as a collector of the extracts. The maximum quantities of the released rhG-CSF, rhGM-CSF and rhEGF reached approximately 25, 50, and 10%, respectively, of the total cytokine contents of the dressings, as assessed by the specific ELISA tests. These data indicate that collagen- and polyurethane dressings containing rhGM-CSF and rhG/CSF may serve as effective tools for the topical delivery of cytokines to wounded tissues. Copyright     

New real-time PCR-based method for in vitro susceptibility testing of Anaplasma phagocytophilum against antimicrobial agents

Up to now, only a few isolates of Anaplasma phagocytophilum have been tested for their susceptibility against a small number of antimicrobial agents. In addition, as with other fastidious or intracellular bacteria, the test methods are laborious and neither minimal inhibitory concentration (MIC) definitions, nor the test conditions and the inocula are standardised to date. A new 16S-rDNA-based real-time PCR assay has been developed and used under standardised conditions to analyse the activity of seven antimicrobial agents against two A. phagocytophilum isolates. After 72 h incubation, MICs were determined by software-assisted calculation of bacterial growth in samples and controls from semi-quantitative PCR results. In our study, the rank order of potency on a mg/l basis for the antimicrobial agents with enhanced in vitro activity against A. phagocytophilum was moxifloxacin (MIC: ≤0.03 mg/l) > doxycycline (MIC: ≤0.125 mg/l) > ciprofloxacin (MIC: 0.125 mg/l). Gentamicin, ampicillin, azithromycin and cethromycin showed no activity against the isolates tested in this investigation. Our new 16S-rDNA-PCR-based microdilution test system was shown to be sensitive, reproducible and reliable. The assay is capable of testing larger numbers of isolates and antimicrobial agents under standardised and very precise test conditions and may therefore offer a competent technical solution of the difficulties known to be associated with in vitro testing of other bacterial pathogens that grow intracellularly, such as chlamydia or rickettsia.     

葛根素注射液的细菌内毒素检查法

目的 建立葛根素注射液的细菌内毒素检测法.方法 根据中国药典2005年版二部收载的细菌内毒素检查的要求进行实验.结果 葛根素注射液稀释到2.083 g·L-1浓度1→24后可消除对鲎试剂的干扰,用灵敏度为0.25 EU·ml - 1的鲎试剂检测内毒素是有效的,并与家兔法的结果一致.结论 对葛根素注射液可用内毒素检查法替代家兔法的热原检查.     

Optimization of HPLC method for stability testing of bacitracin

A stability indicating HPLC assay for bacitracin has been developed and validated. The assay is based on a gradient elution, reversed phase column and UV diode array detection. On the basis of our previous analytical work several additional systematic HPLC tests for optimization of analytical method were performed. In order to achieve the highest selectivity of HPLC method, tests were conducted with extremely complex samples – zinc bacitracin feed grade as food additive for animals. The influence of pH of mobile phase and type of columns on chromatographic separation of active (A, B₁ and B₂) and inactive (F) polypeptide components of bacitracin were investigated in detail. It was found also that the peak B₁ comprises three and the peak F two subunits – probably isomers. The obtained analytical procedure proved to be very selective and effective for the simultaneous determination of active polypeptide A, B₁ and B₂, impurities, known and unknown degradation products and ballast material.     

注射用盐酸艾司洛尔细菌内毒素检查法研究

目的：建立注射用盐酸艾司洛尔的细菌内毒素检查方法。方法：按《中国药典》2015年版四部通则进行实验和结果判断。结果：将注射用盐酸艾司洛尔样品溶解液加适量稀释剂Ⅰ稀释,用BET水稀释至1.25mg.mL-1时对细菌内毒素检查无干扰,细菌内毒素限值为0.10EU.mg-1。对3批注射用盐酸艾司洛尔进行常规检查,结果均符合规定。结论：所建立的细菌内毒素检查法可用于注射用盐酸艾司洛尔的细菌内毒素检查。     

A multiple toe-pinch method for testing analgesic drugs

In guinea-pigs and rats, an immediate squeak was one of the most consistent and readily observed responses to application of a light artery clip to the base of a toe. Morphine and related drugs suppressed this response. Squeak-responses from each toe of an experimental animal formed the basis of a technique for measuring activity of analgesic drugs. A statistical method was developed to analyse the correlated quantal observations obtained. It provided an estimate of the increase of information from several toes compared with one. Testing all toes of each animal yielded a substantial increase of information, because the correlation between responses of different toes was low. Among drugs having an analgesic action in man, 1-(β-diethyl-aminoethyl)-2-(p-ethoxybenzyl)-5-nitrobenzimidazole, methadone, morphine, pethidine and codeine (in descending order of potency) were active in this test in guinea-pigs. Acetylsalicylic acid, amidopyrine, amphetamine, chlorpromazine, 4-hydroxyisophthalic acid, lysergic acid diethylamide, mephenesin, nalorphine, pentetrazole, phenobarbitone, phencyclidine, phenytoin, salicylamide, strychnine and troxidone showed little or no activity. The time-courses of active drugs were estimated, and morphine had the longest action.     

An in vivo method for testing GABAergic compounds

This report describes an intracerebroventricular technique of drug injection which enables compounds with GABAergic properties to be rapidly identified in vivo. In addition, this method allows for the testing of compounds that poorly penetrate the blood brain barrier for GABAergic activities. Subcutaneous administration of apomorphine (1.5 mg/kg, SC) elicits climbing behavior in mice. The apomorphine-induced climbing behavior is inhibited by the intracerebroventricular or intraperitoneal administration of known GABA (gamma aminobutyric acid) agonists including muscimol (10–50 ng) and GABA (2–10 μg). This inhibitory effect of GABA or muscimol on apomorphine-induced climbing behavior can be reversed by picrotoxin (2 mg/kg, IP), a known GABA-receptor antagonist.     

注射用兰索拉唑细菌内毒素检查法的可行性研究

目的对注射用兰索拉唑进行干扰试验,建立注射用兰索拉唑细菌内毒素检查的试验方法。方法采用《中国药典》2010年版二部附录细菌内毒素检查法进行试验。结果将注射用兰索拉唑稀释至浓度为50μg/ml时,对细菌内毒素检查法无干扰作用。结论应用鲎试剂进行注射用兰索拉唑细菌内毒素检查是可行的。