Influence of the glass packing on the contamination of pharmaceutical products by aluminum. Part I: Salts, glucose, heparin and albumin

The presence of aluminium (Al) in pharmaceutical products used parenterally as sodium and potassium chlorides, glucose, heparin and albumin were investigated with respect to their storage in glass containers. As glasses can have aluminium in their composition, the aluminium may be released from the glass into the solution. The action of the substances above mentioned were investigated storing their solutions in glass and plastic containers, and measuring the aluminium in solution at determined time intervals. The aluminium present in the commercial pharmaceutical products, stored in both plastic and glass containers were also measured. All glass containers were analysed to determine their aluminium content. The aluminium determinations were done by atomic absorption spectrometry. The results showed that aluminium is present in all analysed glasses in a percentage of 0.6 to 3%. Although all substances already have a residual aluminium contamination, the major contribution comes from the glass containers in which their solutions were stored. The contamination arising from glass depends too much on the nature of the substance. While the salts extracted about 400 μg Al/l in 60 days, glucose extracted 150 μg Al/l, and albumin and heparin about 500 μg Al/l in the same time interval. Commercial solutions of glucose contain about 10 μg Al/l when stored in polyethylene and from 350 to 1000 μg Al/l when in glass ampules. Considering all commercial products, solutions stored in plastic containers contained no more than 20 μg Al/l whereas in glass the aluminium contamination reached 1000 μg/l, and in all of them the aluminium increases with the age of the product.     

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.     

Influence of the glass packing on the contamination of pharmaceutical products by aluminium. Part II: Amino acids for parenteral nutrition

The presence of aluminium in amino acids parenteral nutrition solutions can be related to the affinity of the amino acids for aluminium present in glass containers used for storage. For this study solutions of 19 amino acids used in parenteral nutrition were stored individually in glass flasks and the aluminium measured at determined time intervals. Solutions of complexing agents for aluminium, as ethylene-diaminetetraacetic acid, nitrilotriacetic acid, citrate, oxalate and fluoride ions were also stored in the same flasks and the aluminium measured during the same time interval. The measurements were made by electrothermal atomic absorption spectrometry. The aluminium content of the glass containers was also measured. The results showed that the glasses have from 0.6% to 0.8% Al. Only solutions of cysteine, cystine, aspartic acid and glutamic acid became contaminated by aluminium. As the same occurred with the complexing agents, aluminum can be released from glass due to an affinity of the substances for aluminium. Comparing the action of complexing agents and amino acids for which the stability constants of aluminium complex are known, it is possible to relate the magnitude of the stability constant with the aluminium leached from glass, the higher the stability constant, the higher the aluminium released. The analysis of commercial formulations with and without cysteine, cystine, glutamic acid or aspartic acid stored in glass containers confirms that the presence of these amino acids combined with the age of the soLution are, at least partially, responsible for the aluminium contamination. The resuLts demonstrated that the contamination is an ongoing process due to the presence of aluminium in glass combined with the affinity of some amino acids for this element.     

Heparin modulates the growth and adherence and augments the growth-inhibitory action of TNF-alpha on cultured human keratinocytes

Previous works suggest the involvement of mast cells in the epithelialization of chronic wounds. Since heparin is a major mediator stored in the secretory granules of mast cells, the purpose of this work was to elucidate the function of heparin in epithelialization using in vitro culture models. For this, low- and high-calcium media in monolayer and epithelium cultures of keratinocytes were used. Also, an assay based on keratinocyte adherence onto plastic surface was used as well. Heparin (0.02-200 microg/ml) inhibited keratinocyte growth in a non-cytotoxic and dose-dependent manner in low- and high-calcium media, Keratinocyte-SFM and DMEM, in the absence of growth factors and serum. Also, heparin inhibited the growth of keratinocyte epithelium in the presence of 10% fetal calf serum and DMEM. Instead, in the presence of Keratinocyte-SFM and growth factors, heparin at 2 microg/ml inhibited the growth by 18% but at higher heparin concentrations the inhibition was reversed to baseline. TNF-alpha is another preformed mediator in mast cell granules and it inhibited keratinocyte growth in monolayer and epithelium cultures. Interestingly, heparin at 2-20 microg/ml augmented or even potentiated this growth-inhibitory effect of TNF-alpha. The association of TNF-alpha with heparin was shown by demonstrating that TNF-alpha bound tightly to heparin-Sepharose chromatographic material. However, heparin could not augment TNF-alpha-induced cell cycle arrest at G0/G1 phase or intercellular adhesion molecule-1 expression in keratinocytes. In the cell adherence assay, heparin at 2 microg/ml inhibited significantly by 12-13% or 33% the adherence of keratinocytes onto the plastic surface coated with fibronectin or collagen, respectively, but this inhibition was reversed back to baseline at 20 or 200 microg/ml heparin. Also, heparin affected the cell membrane rather than the protein coat on the plastic surface. In conclusion, heparin not only inhibits or modulates keratinocyte growth and adherence but it also binds and potentiates the growth-inhibitory function of TNF-alpha.     

Capillary electrophoresis-single strand conformation polymorphism for the detection of multiple mutations leading to tuberculosis drug resistance

For rate determinations of anaerobic metabolism it is essential to maintain strictly anoxic conditions throughout the experiment. However, even if oxygen contamination can be avoided while preparing the incubation containers, it is still possible that the incubation containers themselves contaminate the samples by oxygen diffusing from or through their plastic or rubber components. In this study, we investigated the sources and extent of oxygen contamination during anoxic incubations, and present solutions to minimize oxygen contamination. In particular, we investigated oxygen contamination in Labco® Exetainers, glass vials with a butyl rubber septum in the screw cap, which are frequently used in microbiological experiments. Our results show that significant oxygen contamination occurred at different stages during the incubation. Contamination occurred when Exetainers were either filled or incubated for more than 16 h under oxic atmosphere, but also under an oxygen-free atmosphere due to diffusion of oxygen out of the butyl rubber septum. Therefore, to avoid oxygen contamination during incubations, we suggest (1) filling and incubating the incubation containers under anoxic atmosphere (glove bag) and (2) deoxygenating all elastomers in sample processing and incubation equipment. If initial oxygen contamination cannot be avoided, introduction of an anoxic headspace might help extract oxygen from the incubated sample and present a buffer against oxygen diffusing out of the septum. We modeled the amount of oxygen diffusing out of butyl rubber septa under different conditions, and results fitted well with the observed oxygen contamination. Thus, the model can be used to predict oxygen contamination under varying conditions and for differently sized septa.     

Effect of sub-lethal concentrations of aluminium on the filtration activity of the freshwater mussel Anodonta cygnea L. at neutral pH

Significant amounts of aluminium (Al) are commonly present in rivers and lakes, largely in particulate form in neutral waters. Freshwater bivalves, as filter feeders are therefore exposed to both particulate and dissolved metal and are potentially vulnerable to Al. The effect of Al on filtering behaviour of the freshwater mussel Anodonta cygnea L. was investigated during short (1 hour) and long-term (15 days) exposure to environmentally relevant concentrations (250 and 500 microg l(-1)) at neutral pH. Water flow through the outflow siphon was monitored as an indicator of pumping capacity. Short-term (1 hour) exposure to 500 microg l(-1) added Al produced an irreversible decrease in the duration of filtering periods, presumably as an avoidance response to the toxicant. One-hour exposure 250 microg l(-1) Al had no detectable effect. When mussels were exposed to 250 or 500 microg l(-1) added Al for 15 days, siphon activity measured in days 11-15 of exposure was inhibited by 50% and 65%, respectively, compared to pre-exposure levels. Recovery occurred following transfer of mussels to uncontaminated water. Interaction between Al and freshwater bivalves at neutral pH may affect both the performance of the mussels and the chemical speciation of the metal in the natural environment.     

Migration of phthalates from plastic products to model solutions

The aim of this investigation was to determine the level and rate of migration of phthalates, compounds used as plastic softeners, from various plastic products into model solutions and to assess the possible adverse effects of the phthalate amounts released on human health, thus to contribute to harmonization of the opinions on the maximal allowed human exposure to these compounds through environmental factors. Nine specimens of plastic toys, 16 specimens of plastic food containers and 10 specimens of other plastic consumer goods were analyzed. The specimens of plastic products were submitted to 10-day action of model solutions. Three model solutions were used: distilled water, 10% ethyl alcohol, and 3% acetic acid. Identification and quantification of the phthalates released were performed by the method of gas chromatography on days 1, 5 and 10 of exposure, at a detection limit of 0.005 microgram/kg. On day 10, the highest level of released phthalates (54.5 mg/kg) was measured in distilled water, followed by 44.4 mg/kg in 3% acetic acid and 32.3 mg/kg in 10% ethyl alcohol. According to plastic product categories, the highest pooled level of phthalates released to all three solutions was recorded for plastic toys (66.2 mg/kg), followed by food containers (37.6 mg/kg) and other consumer goods (27.4 mg/kg). According to plastic product categories, toys showed the most rapid phthalate release, with 65.4% (43.3 of 66.2 mg/kg) of the pooled level of phthalates released to all three solutions recorded on day 1. As indicated by the study results, the levels of phthalates released would not present a hazard for human health, not even over a prolonged period of time. However, data on the highest and fastest pooled phthalate release from plastic toys, and this especially to distilled water simulating salivary action, point to the need of continuous evaluation and amendments of the legislation on phthalates in consumer goods.     

An In Vitro Study of the Effect of Aluminum and the Combined Effect of Strontium, Aluminum, and Fluoride Elements on Early Enamel Carious Lesions

The purpose of this in vitro study was to evaluate the effect of aluminum and of combined strontium, aluminum, and fluoride treatments on enamel demineralization and remineralization. During a 6-day pH-cycling protocol, pre-softened bovine enamel slabs were immersed twice daily for 1 min in the following experimental solutions: (a) distilled water [W] (negative control); (b) 1,000 ppm F [F] (positive control); (c) 1,000 ppm Al [Al]; (d) 1,000 ppm Al,1,000 ppm F applied interchangeably [Al-F]; (e) 1,000 ppm Al, 1,000 ppm F, applied in sequential order [Al+F]; (f) combined 1,000 ppm Al and 150 ppm Sr [Al+Sr]; and (g) combined 150 ppm Sr and 1,000 ppm F [Sr+F]. Subsequently, the specimens were subjected to a 5-day acid resistance test. Lesions were evaluated quantitatively by performing surface microhardness and qualitatively by using polarized light microscopy. According to the results, solutions [Sr+F] and [Al-F] enhanced remineralization and inhibited demineralization as effectively as the [F] solution and significantly superiorly compared to [Al+Sr] and [Al] solutions. All tested solution groups, except for the [Al+Sr] group, presented significantly increased resistance to acidic attack, compared to [W]. PLM examination revealed that all solution groups, except for the [W] group, developed an acid-resistant zone at lesion surfaces. In conclusion, under the present experimental conditions, the combined strontium-fluoride and aluminum-fluoride treatments presented similar anti-caries efficacy compared to fluoride treatment alone, but they did not show evidence of synergistic activity on pre-softened enamel.     

Survival of Azotobacter spp. in Dry Soils

Dry soils stored in glass containers in the laboratory and protected from contamination for periods of 22 to 24 years yielded numerous colonies of Azotobacter chroococcum and other members of the family Azotobacteraceae. These results were compared with those reported in 1974, and the findings are uniformly consistent in terms of surviving populations. The data prove that these bacteria remain viable after prolonged periods of dormancy in much the same way as do the endospores of gram-positive bacteria.     

Effect of climate and type of storage container on aflatoxin production in corn and its associated risks to wildlife species

The effects of grain storage containers on aflatoxin production, and the relationship between the level of aflatoxin and the number and weight of fluorescing kernels were determined in corn (Zea maize) stored in controlled climate regimes. Two hundred and forty 100-g samples were held up to 3 mos using four types of storage containers placed in four climates. Storage containers included corn placed in metal cans, paper bags, plastic bags, and paper bags placed in plastic bags. Climates were constant during the duration of the project and included a combination of temperatures and humidities. Temperatures were 29-32 C and 14-18 C; relative humidities were 85-88% and 35-40%. In addition, corn was exposed to environmental conditions conductive for aflatoxin production and 100 g samples were randomly collected, examined under ultraviolet light for fluorescence, and then quantified for aflatoxin levels. Corn samples tested negative for aflatoxin at the beginning of the project. Main (i.e., container, climate, and month) and interactive effects were not observed. Mean levels of aflatoxin ranged from 0 to 151 microg/kg. Aflatoxin was produced regardless of type of storage container, time of storage, and climatic conditions; however, only 8% of the samples produced aflatoxin levels that exceeded 50 microg/kg. Fluorescing corn ranged from 0 to 19 kernels per sample, while aflatoxin levels ranged from 0 to 1,375 microg/kg for the same samples. No relationships were found between the number and weight of fluorescing kernels of corn and aflatoxin levels. The black light test yielded a false negative rate of 23% when in fact the aflatoxin concentrations exceeded 50 microg/kg. Therefore, quantifying fluorescing grain under UV light should not be considered a feasible alternative for aflatoxin testing of grain intended for wildlife.     

Bacteriological stability and growth kinetics of Pseudomonas aeruginosa in bottled water

Bacteriological stability of water bottled in plastic containers and the growth kinetics of Pseudomonas aeruginosa were determined. Samples of water from the source, water to be bottled, finished product and sterile water bottled in non-returnable and returnable containers were analysed for aerobic colony count, coliforms, Escherichia coli and Ps. aeruginosa. The samples were examined for up to 30 d storage. Aerobic colony count increased 6 d after bottling to between 10³ and 10⁵ cfu ml⁻¹. Coliforms and E. coli were not found in any sample. Pseudomonas aeruginosa was isolated from commercial products bottled in returnable plastic containers due to the contamination from the containers and the subsequent multiplication utilizing trace nutrients. The predominant Ps. aeruginosa strains showed high doubling time (26 h) due to competition from the accompanying flora. In the absence of competing flora Ps. aeruginosa reached higher density than the maximum reached by aerobic flora, with a doubling time of only 3·6 h. After 30 d storage, this micro-organism was predominant.     

Inhibition of Babesia spp. by plastics.

Five commercially available plastic containers were compared with glass for toxicity toward Babesia rodhaini and Babesia bigemina. Comparisons were made by using infectivity tests in mice (B. rodhaini) and cattle (B. bigemina). Low-density polypropylene and polystyrene containers were not toxic, but two of the three polyvinyl chloride containers tested significantly reduced the viability of both species of Babesia. Plasticizer present in various amounts on the surface of the toxic containers was most likely the inhibitory material.     

四种基质表面上常规杀虫剂对四纹豆象的药效评价（英文）

在害虫治理中, 在消费或贮藏粮食加工产品的建筑设施或场所进行害虫防治需要将杀虫剂施用在各种基质表面上。为了测定不同基质表面上杀虫剂的药效, 将四纹豆象Callosobruchus maculatus (F.)成虫接触田间推荐剂量的阿维菌素、 溴氰菊酯和毒死蜱。结果表明： 施用在玻璃、 瓷砖、 塑料和纸盘表面上, 阿维菌素对四纹豆象成虫的致死率分别为63.33%, 22.41%, 12.9%和11.9%, 而溴氰菊酯在这4种基质表面上对四纹豆象成虫的致死率分别为 55%, 44.2%, 41.3%和 37.4%。在所有基质表面上接触毒死蜱, 四纹豆象成虫的死亡率均为100%。对数据进行的Probit 分析表明, 毒死蜱制剂在玻璃、 瓷砖、 塑料和纸盘上对四纹豆象成虫 的LC₅₀ 值分别为 8.66, 13.6, 29.16和 56.5 μg/mL, 阿维菌素制剂的相应数值分别为119.4, 446.2, 774.2 和 836.4 μg/mL, 溴氰菊酯制剂的相应数值分别为 1 008, 1 131, 1 210和 1 336 μg/mL。据此推断, 毒死蜱对四纹豆象的毒性最强, 且在玻璃、 瓷砖、 塑料和纸盘表面上的毒性依次降低。     

Effect of electrostatic charge on the contamination of plastic food containers by airborne bacterial spores

Electrostatic charge of approximately -10 kv was produced by friction on polystyrene food container samples. This charge quickly decayed to a lower, more stable, level. Exposure of samples to positively charged red and negatively charged green fluorescent particles resulted in a particle-distribution pattern on the plastic surface. The dynamic attraction of fluorescent particles was illustrated by time-lapse photography. Similar distribution patterns of airborne bacterial spores were shown to develop. In controlled bacterial aerosol exposure tests, an increase in surface contamination of the plastic samples was found to be quantitatively related to an increase in negative electrostatic charge on the plastic. Static charge was found to accumulate on plastic food containers during their manufacture, and to remain indefinitely on many of the finished products. This charge was of the intensity and polarity to attract positively charged bacterial cells if such particles were present in the air.     

Matrix isolation of free radicals from carbohydrates. II. Reactions of H. with glucose and derivatives in acidic glasses

Photolytically produced H.-atoms in 6 mol dm-3 H2SO4/H2O glasses trapped at 77 K react upon annealing to 130 K with dissolved carbohydrates to form carbon-located free radicals by abstraction of carbon-bound protons. Analysis of electron spin resonance (e.s.r.) spectra at various annealing stages from alpha- and beta-D-glucose together with 6,6-d2-D-glucose, 6-deoxy-D-glucose, 2-deoxy-D-glucose, glucose-1-phosphate, D-xylose, D-allose and D-mannose indicates radical formation at all possible carbon sites with a strong preference for C1 and a somewhat enhanced contribution of C4 over the statistical expectation. The corresponding component spectra are analysed either by spectra isolation or simulation and their parameters are given. Intramolecular radical transformation at temperatures of 140-160 K is explained by acid-catalysed H2O-elimination. The findings are discussed in relation to the radiation-chemistry of aqueous glucose solutions. We thus show that the system of photolyzed Fe2+ in acidic glasses at low temperatures containing 10 mmol dm-3 carbohydrate is suitable for studying H(D.)-reactions by means of e.s.r. spectroscopy. Unlike previously used glasses containing carbohydrates, contributions of oxidation and reduction by direct effects or mixtures of direct and indirect effects and phase-effects due to incomplete glass formation are avoided.     

Long-term storage of Pennisetum glaucum (L.) R. Br. pollen

Summary Pearl millet [Pennisetum glaucum (L.) R. Br.] pollen has been successfully stored for 2,615 and 2,911 days at -18° and -73 °C, respectively, and continues to be viable. Viability of pollen stored at -73 °C appears to be little affected either by pollen storage moisture contents below 7.2% or by storage in glass vial or zip-lock plastic bag containers. Pollen moisture content appears to be more critical for maintaining viability at -18°C than at -73°C. Glass vials appear to be more desirable for longer term (>3 years) storage at -18°C.     

On the stability of salivary progesterone under various conditions of storage

The concentrations of progesterone in saliva of women exhibited significant decreases when the fluid was stored in plastic vials for 3 days at room temperature or 37 C. The addition of antibiotics or a variety of metabolic poisons to the saliva prior to storage did not prevent the progesterone decrement. However, the addition of albumin (2 g/dl) was protective, suggesting that the protein impeded adsorption of salivary progesterone by the plastic container. Saliva could be maintained at 37 C for 3 days in glass vials or at -20 C in plastic containers for indefinite periods without loss of progesterone titers. These data indicate that a patient under luteal function assessment may collect saliva samples in glass vials at regular intervals during the latter half of her cycle and store them in the freezer compartment of the refrigerator until shipment by mail to the laboratory for progesterone assay. With special care, plastic vials charged with albumin may also be used.     

Type of closure prevents microbial contamination of cosmetics during consumer use.

The dispensing closure used for containers plays an important role in protecting cosmetics from in-use microbial contamination. This hypothesis was tested by aseptically packing unpreserved shampoo and skin lotion into containers with three different closure types which provided various degrees of protection against consumer and environmental microbial insults. Shampoo was packed in containers with slit-cap (n = 25), flip-cap (n = 25), or screw-cap (n = 28) closures. Skin lotion was packed in containers with pump-top (n = 21), flip-cap (n = 18), or screw-cap (n = 21) closures. The products were then used by volunteers under actual in-use conditions for 3 (shampoo) or 2 (skin lotion) weeks. After use, the products were evaluated for microbial contamination by using standard methods for enumeration and identification. The standard screw-cap closure provided only minimal protection against microbial contamination of both the shampoo (29% contamination incidence) and the skin lotion (71%). The slit-cap closure on the shampoo container and the flip-cap closure on the skin lotion container provided slightly enhanced degrees of protection (21 and 39% contamination incidence, respectively). The greatest amount of protection (i.e., lowest contamination incidence) was provided by the flip-cap closure for the shampoo container (0%) and the pump-top closure for the skin lotion container (10%). As a result, closure type plays an important role in protecting poorly preserved products from in-use microbial contamination.     

The oxidative mechanism of heparin interferes with radical production by glucose and reduces the degree of glycooxidative modifications on human serum albumin.

Among substances which may prove useful in preventing or reducing the progression of glycooxidative modifications of proteins, heparin plays a unique role. To elucidate the mechanism whereby heparin may favourably influence the protein structure during glycation, human serum albumin (HSA) was glycated with both 25 and 50 mM glucose in the absence and presence of 12 microg.mL(-1) low-molecular-mass heparin. Glycation caused: (a) modifications of fluorescence emission and excitation spectra consistent with the covalent attachment of glucose to protein; (b) a significant increase in the esterase activity of HSA on p-nitrophenyl acetate; (c) a reduced susceptibility to tryptic digestion and (d) enhanced formation of high-molecular mass aggregates of HSA. These alterations were accompanied by oxidative reactions, as the EPR spectra showed a clear-cut radical signal, dependent on glucose concentration, further confirmed by measurement of the carbonyl content of HSA, as an indirect proof of oxidative damage. In the presence of heparin all the above alterations, especially at 25 mM glucose, turned out to be antagonized. The effects of heparin were dependent on its specific binding to HSA, which triggered an oxidative mechanism strikingly different from that caused by glucose. In the presence of heparin, only the radical species catalyzed by heparin was detected across all samples of glycated HSA, irrespective of glucose concentration. In addition, at 25 mM glucose, enhancement of the oxidative capacity of heparin was also observed. The results demonstrate that the oxidative mechanism sustained by heparin mediates biological effects that may be beneficial in reducing the extent of glycooxidative damage on HSA.     

Microscopy with Plastic Substitutes for Cover Glasses

To determine whether plastic substitutes for cover glasses on microscope slides affect the performance of the microscope, their optical constants were determined. The plastic covers and glass cover glasses were mounted also on silvered slides to form Star Test Plates which were studied by competent observers. The thicker plastic cover glasses, now on the market, are satisfactory when mounted to give a plane surface. Optically inhomogeneous materials, of irregular thickness, those that curl or do not have plane surfaces, adversely affect the performance of the microscope and should not be used. Since the substitutes are softer than glass they must be protected from abrasion. It is recommended that thicknesses of 0.18 mm., and none outside of a range of 0.12 to 0.20 mm. be used. For critical observation with unimmersed objectives of high aperture, best results are obtained with the correction collar set at the position corresponding to the actual thickness of the cover slip, just as would be done with glass cover glasses.