Rubber stoppers used with sulphidic media: a note of caution

Common black rubber stoppers, made from natural rubber and styrene–butadiene, may cause a loss of hydrogen sulphide from aqueous media and impede the growth of sulphate-reducing bacteria under thermophilic conditions. This revised version was published online in July 2006 with corrections to the Cover Date.     

Combined Screw-Cap and Rubber-Stopper Closure for Hungate Tubes (Pre-reduced Anaerobically Sterilized Roll Tubes and Liquid Media)

A double-closured method introduced for Hungate tubes eliminates clamps or special racks to hold the rubber stoppers in place while the tubes are being processed. The system prevents stoppers from being dislodged by gas-producing anaerobes and keeps the stoppers sterile so that the closed system of transfer can be conveniently utilized. Open and closed systems for handling a gas-producing anaerobe were compared. Ten tubes containing Clostridium perfringens were opened; the organism was detected in the air by use of an impingement collector. The gloved hand of the operator also revealed the test organism. A similar trial with culture material removed by the closed system (syringe transfer) resulted in no detectable contamination of the environment.     

Problems in the measurement of zinc using heparin as an anticoagulant

Although several reports imply that anticoagulants and preservatives contain zinc, the quantity of zinc in heparin, if any, has not been documented. Zinc concentration was determined by flame atomic absorption spectroscopy in varying dilutions of multiple commercially obtained samples of purified sodium heparin N = 15 (microgram Zn/1000 Units heparin). Rubber stoppers of sterile heparin vials and of blood evacuation tubes were incubated in pre-analyzed water or saline on a mechanical shaker with fluid aliquots obtained up to 27 hours and analyzed for zinc content (microgram Zn/0.1 ml). Heparin, with contact or without contact with rubber stoppers, recorded identical zinc concentrations. Zinc concentrations varied from 0.222 +/- 0.01 (mean +/- SE) to 3.49 +/- 0.005 microgram Zn/1000 Units heparin. Leaching of zinc from rubber stoppers of vacutainer tubes (N = 9) was noted only with those containing known chelators of zinc. These results indicate that zinc is present in certain lots of sodium heparin and that caution must be exercised when reporting zinc concentrations of blood samples that contain sodium heparin as the anticoagulant.     

Minerals leached into drinking water from rubber stoppers.

Drinking water and its delivery system are potential sources of variation in animal research. Concern arose that rubber stoppers used to cork water bottles might be a source of some nutritionally required minerals which could leach into drinking water. Six types of stoppers, each having different compositions, were cleaned with stainless-steel sipper tubes inserted into them and attached to polypropylene bottles filled with either deionized water (pH 4.5) or acidified-deionized water (pH 2.5). After six days of contact, water levels of copper, magnesium, iron, manganese, zinc, chromium, and selenium were determined by atomic absorption spectroscopy. Additionally, three of the stopper types were analyzed for mineral content. Minerals were present in both stoppers and drinking water. Acidified-deionized water generally leached minerals from the stoppers than did deionized water. The black stopper which is commonly used in animal facilities contained and leached measurable levels of some minerals, but it still can be recommended for typical animal husbandry uses, although other types of stoppers would be more suitable for specific nutritional and toxicologic studies.     

冻干麻疹减毒活疫苗残余水分影响因素的探讨

影响冻干制品残余水分的因素很多,但西林瓶胶塞的含水量是其关键因素。使用经干烤工艺处理和未干烤处理的药用溴丁基胶塞进行麻疹减毒活疫苗冻干试验,结果显示,用未经干烤工艺处理及处理的药用溴丁基胶塞冻干的麻疹减毒活疫苗,存储18个月后,水分上升的平均值分别为2.03%和0.08%,经单一因素的方差分析,两者有统计学意义(P0.01)。证明经处理的药用溴丁基胶塞,对保持麻疹减毒活疫苗在存储期内的残余水分稳定作用效果明显。     

Factors Affecting the Viability of Mycoplasma mycoides in Bottled Contagious Bovine Pleuropneumonia Vaccine

S ummary . Some of the factors contributing to the loss of viability of V5 contagious bovine pleuropneumonia vaccine in BVFOS medium during storage were investigated. Results showed that: 1, some types of rubber stoppers were highly toxic to Mycoplasma mycoides ; 2, loss of viability was greater when the vaccine was stored in smaller bottles and was related to the proportionately greater air space; 3, the addition of glycerol to BVFOS medium resulted in slightly better survival during storage at 37° but decreased survival at 4°; 4, small bottles can be filled to higher levels, and the air space reduced, if the mouth of the bottle is protected during incubation by a metal cover instead of a cotton wool plug. It is recommended that the amount of air space should not exceed 40% of the volume of the bottle, the vaccine should be stoppered with non-toxic stoppers after incubation and the BVFOS medium should not be supplemented with glycerol. Vaccine prepared in this way should maintain a viable count of 10⁷ or more organisms/ml during storage at 4° for at least 12 weeks.     

Thermistor probe for testing an antipyretic suppository in rabbits

The thermistor probe for estimating the effects of an antipyretic suppository after its administration into the rectum of the rabbit was studied. A thermistor probe with three rubber disk stoppers was confirmed to be able to prevent the leakage of drugs from the rectum of a rabbit restrained in a neck stock. By using this newly devised thermistor probe or the usual thermistor probe without a stopper, the febrile response was determined in rabbits injected with bacterial pyrogen. There was no difference in the ability to detect rectal temperature between the two thermistor probes. From these results, it could be concluded that this newly devised thermistor prove was useful in studying the effects of antipyretic suppositories in rabbits.     

Mycobiota in Portuguese 'normal' and 'green' cork throughout the manufacturing process of stoppers

The compounds responsible for the so-called 'cork taint' include, among others, some microbial metabolites which can be produced by the microbial population colonizing the unprocessed cork and stoppers. This study was intended to obtain information on the mycobiota associated with Portuguese cork throughout the manufacturing process of stoppers. Samples of barks and stoppers of both 'normal' and 'green' cork were examined. Moulds were isolated from 'normal' and 'green' cork throughout the entire cork stopper manufacturing process. Yeasts were rarely detected in the corks. Fungal contamination was not detected in finished stoppers from the company under study.     

Influence of the glass packing on the contamination of pharmaceutical products by aluminium. Part III: Interaction container-chemicals during the heating for sterilisation

The interaction of chemicals with the container materials during heating for sterilisation was investigated, storing the components of parenteral nutrition solutions individually in sealed glass ampoules and in contact with a rubber stopper, and heating the system at 121 degrees C for 30 min. Subsequently, the aluminium content of the solutions was measured by atomic absorption spectrometry (AAS). The assay was also carried out with acids, alkalis and some complexing agents for Al. The containers were decomposed and also assayed for aluminium. 30 different commercial solutions for parenteral nutrition, stored either in glass or in plastic containers, were assayed measuring the aluminium present in the solutions and in the container materials. The results of all investigated container materials revealed an aluminium content of 1.57% Al in glass, 0.05% in plastic and 4.54% in rubber. The sterilisation procedure showed that even pure water was able to extract Al from glass and rubber, 22.5 +/- 13.3 microg/L and 79.4 +/- 22.7 microg/L respectively, while from plastic the aluminium leached was insignificant. The Al released from glass ampoules laid between 20 microg/L for leucine, ornithine and lysine solutions and 1500 microg/L for solutions of basic phosphates and bicarbonate; from rubber stoppers it reached levels over 500 microg/L for cysteine, aspartic acid, glutamic acid and cystine solutions. Ion-exchange properties and influence of pH can explain the interaction of glass with some chemicals (salts, acids and alkalis), but only an affinity for aluminium could explain the action of some amino acids and other chemicals, as albumin and heparin, on glass and rubber, considering the aluminium release. Experiments with complexing agents for Al allowed to conclude that the higher the stability constant of the complex, the higher the Al release from the container material.     

An automatic feeder for infant rats

An automatic feeder for infant rats was developed which consisted of a rectangular container in which tubes were passed through the front and back walls and sloped slightly from the back to the front. Nursing bottles were inserted into the tubes from the back until the nipples projected 5 mm from holes in the rubber stoppers which closed the tubes at the front. At the surface of the front wall around the nipples, there was material similar in feel to that of the skin and fur of mother rats. Milk was fed from a tank to the nursing bottles by a peristaltic pump. After training 3-day-old infants which were nursed by mother rats during the first three days, they found the nipples and suckled milk by themselves. At the time of weaning, the automatically-fed infants gained as much weight as the mother nursed-infants.     

A closed system for the filtration of Mycobacterium bovis liquid cultures using disposable capsule filters

A filtration system was designed to sterilize large volumes of Mycobacterium bovis BCG Tokyo culture safely, needed to purify protein antigens for immunodiagnosis of bovine tuberculosis. A closed system consists of culture bottles connected to three disposable filter capsules of decreasing pore size in series : a depth prefilter over a 1·2 μm filter ; a 0·8 μm prefilter over a 0·45 μm filter ; and a 0·2 μm sterile filter. Low air pressure (3 psi) forces liquid from below the bacillary pellicle. The system features a stainless steel clamp to hold rubber stoppers on the culture bottles, pleated filters to exclude bacillary clumps, a quick disconnector to minimize aerosols, and a closed system with plastic disposable filters that can be autoclaved as a unit without dismantling.     

Exogenous bacterial contamination of donor blood.

The Canadian Red Cross blood transfusion service has followed a set protocol for phlebotomy and collection of a unit of blood. Recent requirements for automated testing have necessitated that a second tube of blood be obtained from the blood line following collection of the unit. Evaluation of the techniques used, however, has indicated the possibility of bacterial contamination from the skin of donors, from insertion of the needle through an unsterile rubber stopper, and through backflow from a nonsterile vacuum tube. To test these possibilities swabs were taken from skin and stoppers of vacuum tubes. Further, vacuum tubes were deliberately contaminated with Escherichia coli. The normal sampling procedure, which involves stripping the donor line to refill and mix the blood, was then followed. This resulted in contamination of the segments and even the blood bag. These findings led to modification of the standard bleeding technique, whereby stripping was eliminated and sterile vacuum tubes were to be used at all times.     

Growth of Bacteria in Inorganic Medium at Different Levels of Airborne Organic Substances

Invasion rates of airborne organic substances into sterile mineral medium were compared by using flasks closed with cotton stoppers, silicone stoppers, and screw caps with Teflon gaskets. The resulting increases of dissolved organic carbon were 0.5, 0.2, and 0 mg/liter per week, respectively. The compounds supported the growth of lake water bacteria and a strain of Pseudomonas fluorescens. Growth rates were correlated to the permeability of the stoppers used. The measured input of organic carbon in the sterile mineral medium is considered to be a minimum value for the actual contribution of organic compounds by the air. Multiplication rates of the bacteria suggest that the organisms prevent the escape of volatile organic substances from the medium by rapid utilization. The steady nutrient supply through the air should be considered in growth experiments with bacteria at low concentrations of nutrients.     

Loss of preservative from a tuberculin solution in rubber stoppered vials fastened with different seals

A tuberculin purified protein derivative (PPD) solution containing 0.3% phenol as a preservative was dispensed in glass vials closed with rubber stoppers fastened in three different ways, namely with Tear-off seals, Flip-off seals and partial seals. After various times of storage at 5 degrees C and 37 degrees C, the phenol content in the tuberculin solution was determined. It was found that the Flip-off seals allowed the phenol to escape at a faster rate than the Tear-off seals and that vials closed with partial seals showed the highest loss of phenol. Although these losses were much more pronounced at 37 degrees C than at 5 degrees C, the phenol content at the latter temperature was, over a period of three years, within the limits of acceptability for tuberculin products capped with Tear-off or Flip-off seals. A loss of phenol also occurred from tuberculin solution stored at -28 degrees C in vials capped with either Tear-off or partial seals. In addition to the Tear-off and Flip-off seals other seals such as the "controlled score' Flip-off seal and the Alcoa Steri-Twist cap were evaluated for their imperviousness to air. Except for the Alcoa Steri-Twist cap none of the seals we have investigated were air tight and hence entirely satisfactory to prevent losses of phenol by evaporation from tuberculin products.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sizing the protein translocation pathway of colicin Ia channels

The bacterial toxin colicin Ia forms voltage-gated channels in planar lipid bilayers. The toxin consists of three domains, with the carboxy-terminal domain (C-domain) responsible for channel formation. The C-domain contributes four membrane-spanning segments and a 68-residue translocated segment to the open channel, whereas the upstream domains and the amino-terminal end of the C-domain stay on the cis side of the membrane. The isolated C-domain, lacking the two upstream domains, also forms channels; however, the amino terminus and one of the normally membrane-spanning segments can move across the membrane. (This can be observed as a drop in single-channel conductance.) In longer carboxy-terminal fragments of colicin Ia that include /=90 mV, even a 26-A stopper is translocated. Upon reduction of their disulfide bonds, all of the stoppers are easily translocated, indicating that it is the folded structure, rather than some aspect of the primary sequence, that slows translocation of the stoppers. Thus, the pathway for translocation is >/=26 A in diameter, or can stretch to this value. This is large enough for an alpha-helical hairpin to fit through.     

Ethylene enhances shoot formation in cultures of the peach rootstock GF-677 (Prunus persica x P. amygdalus)

The influence of ethylene on shoot formation from GF-677 (Prunus persica × P. amygdalus) shoot tip explants was studied in vitro. Cultures in test tubes were placed inside 5 1 glass jars and supplemented with various ethylene concentrations (0–10 ppm). Ethylene at 0.1 ppm, applied during the first 2 weeks of culture, increased the number and the length of shoots produced in vitro. Test tubes with cultures sealed with various types of closure accumulated in their atmosphere different levels of ethylene ranging from 0.1 to 1.2 ppm, depending on the type of closure. Test tubes with cotton-wool bungs had the least while those with serum stoppers had the highest amount of ethylene. The maximum number of shoots was recorded in test tubes covered with serum stoppers. The ethylene concentration was related linearly (R=0.974) to the shoot number and exponentially (R=0.975) to the shoot length.Abbreviations BA benzyladenine - IAA indoleacetic acid - HSD honestly significant difference     

Ethylene generation and reversal of ethylene effects during development in vitro of rapid-cycling Brassica campestris L

A rapid-cycling line of Brassica compestris L was grown from seed to maturity in vitro. Ethylene accumulated in culture containers throughout the growth period when these were tightly sealed (i.e. closed with aluminum foil, tight plastic caps, parafilm or rubber stoppers), whereas no C₂H₄ was detected in vented containers (i.e. closed with foam or cotton plugs, or loosely fitted plastic caps). Under both closure conditions, mean levels of CO₂, O₂ and N₂ were similar to atmospheric levels. The accumulation of C₂H₄ in sealed containers was associated with inhibition of plant development. Plants grown in a sealed condition had swollen hypocotyls, fewer and smaller leaves and either abortive flowers or no floral buds whereas plants in vented containers developed normally to maturity. Significant reversion to normal development was observed when plants growing in sealed containers were treated with 2,5-norbornadie e (NBD), an anti-C₂H₄ compound. Treatment with NBD offers a promising way of regulating C₂H₄ effects in vitro.     

Trapping noctuid moths with synthetic floral volatile lures

Male and female noctuid moths were collected from plastic bucket traps that were baited with different synthetic floral chemicals and placed in peanut fields. Traps baited with phenylacetaldehyde, benzyl acetate, and a blend of phenylacetaldehyde, benzyl acetate, and benzaldehyde collected more soybean looper moths, Pseudoplusia includens (Walker), than benzaldehyde-baited or unbaited traps. Females comprised over 67% of the moths captured and most were mated. At peak capture, over 90 male and female moths per night were collected. In another experiment, phenylacetaldehyde delivered in plastic stoppers attracted more P. includens moths than traps baited using other substrates, but this chemical delivered in wax attracted more velvetbean caterpillar moths (Anticarsia gemmatalis Hübner). Other noctuid male and female moths collected included Agrotis subterranea (F.), Argyrogramma verruca (F.), Helicoverpa zea (Boddie), and several Spodoptera species. Aculeate Hymenoptera were collected in large numbers, especially in traps baited with phenylacetaldehyde delivered from stoppers.     

Metabolism of ethylene in olive leaves

Olive tree tissues are able to metabolize ethylene. This metabolism is inhibited by heat killing and carbon disulfide. Rubber stoppers usually employed to close the incubation vials release carbon disulfide, thereby modifying the values obtained for ethylene. The ethylene consumption rate has been found to be 9 nl/h.g, an intermediate value among the different plant tissues so far examined.     

Cork stoppers industry: defining appropriate mould colonization

AIMS: The main aims of this work were the study of cork slabs moulds colonization and the evaluation of the moulds diversity during cork processing steps, in different cork stoppers factories. Simultaneously, it was envisaged to perform an evaluation of the air quality. METHODS AND RESULTS: Moulds were isolated and identified from cork slabs and cork samples in four cork stoppers factories. The identification was based on morphological characters and microscopic observation of the reproductive structures. Airborne spore dispersion was assessed using a two stage Andersen sampler. It was observed that Chrysonilia sitophila was always present on cork slabs during the maturing period, but mould diversity appeared to be associated to the different factory configurations and processing steps. CONCLUSIONS: Spatial separation of the different steps of the process, including physical separation of the maturation step, is essential to guarantee high air quality and appropriate cork slabs colonization, i.e. C. sitophila dominance. The sorting and cutting of the edges of cork slabs after boiling and before the maturing step is also recommended. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is very important for the cork stopper industry as it gives clear indications on how to keep high quality manufacturing standards and how to avoid occupational health problems.