Selective cellular and molecular effects of beryllium on lymphocytes†

An involvement of the immune system has been implicated in many of the whole animal effects of beryllium (Be) and its compounds and thus lymphocytes are a relevant cell model to study some of the cellular and molecular actions of the metal. Recent results indicate micromolar concentrations of Be salts are weakly mitogenic to lymphocytes in vitro probably as a result of direct interactions with the cell membranes. Furthermore evidence has been obtained that high affinity lymphocyte Be‐binding may be predictive of the potential to express delayed Be hypersensitivity in vivo. Although sub mitogenic concentrations of Be salts may actually activate accessory (macrophage type) cells, concentrations normally mitogenic to lymphocytes were found to inhibit their proliferative response to accessory cell dependent mitogens as a result of a Be²⁺ concentration dependent destruction of the adherent cell population. Higher concentrations of Be salts proved cytostatic to lymphocytes possibly by the established property of intracellularly accumulated Be²⁺ to inhibit cell division. Lymphocyte proliferation can be related to increases in protein phosphoryl‐ation capacity which in vitro can be inhibited by Be²⁺ and preliminary evidence indicates that tyrosine phosphorylation may be particularly sensitive.     

Speciation studies on simulated gastro‐intestinal digests of foodstuffs†

The factors affecting the solubilisation of metals during digestion of food have been examined in a two‐stage model system comprising simulated gastric and intestinal juices. The solubilisation of iron, copper and zinc was examined in digests of white and wholemeal bread. Zinc solubility was principally governed by pH and the simulated digestion had little additional effect. In contrast both iron and copper solubility were markedly affected by digestion of the bread. In the case of copper the enzymolysis increased solubility appreciably at neutral pH. All of the cadmium in a simulated gastric digest of canned crab meat was associated with soluble low molecular weight species of less than 1000 daltons. Cadmium solubility resulted from the acidic conditions of the digest rather than enzymatic solubilisation of binding proteins. Ninety percent of the cadmium in the digest became rebound to the insoluble fraction on adjustment to pH7.0. Subsequent simulated intestinal digestion increased the level of soluble cadmium to 20%; 1.0–1.5% was associated with soluble species in the range 26,000–37,000 daltons and 15.0–20.0% with soluble species of less than 3,000 daltons.     

Genotoxicity of chromium in vitro on yeast: Interaction with DNA†

The genotoxicity of chromium chloride was investigated in cells of D₇ strain of Saccharomyces cerevisiae harvested either from logarithmic or stationary growth phase.

A weak induction of mifotic gene conversion and point reverse mutation was obtained when the incubations were performed using phosphate buffer. No genetic effect was observed when the incubations were performed using Tris‐HCl buffer.

The experiments with ⁵¹Cr radiotracer demonstrated that Cr³⁺ ion enters the yeast cells and binds to DNA even if the incubation mixture was performed with Tris HCl buffer. This behaviour could be due to the highest concentration of CrCl₃ that cause some damages to cytoplasmatic membrane.     

Comments on the Carcinogenicity and Mutagenicity of metals and their compounds†

The carcinogenicity of metals has received extensive study, both epidemiologically and in the laboratory. These have included case reports of occasional human occurrence or clusters of cancer cases as well as extensive epidemiologic studies; in addition, there has been significant laboratory research on the whole animals and in vitro systems. This body of information will be examined selectively.

I will not in this paper attempt a comprehensive review of the mutagenicity and carcinogenicity of metals and their compounds. Rather, I will attempt to set forth some historical perspectives, and to comment on some current gaps and needs.

Other papers in this workshop have presented thorough and very current reviews of most of the topics briefly noted in this presentation and do not require repetition here.

The cancer issue has been studied and reported on far more extensively than that relating to heritable mutations. There has been in recent years increasing interest in the use of short term tests for mutagenicity and cell transformation. These, however, are primarily with respect to their relationship to cancer production rather than to germ cell injury. Interest in cancer from metal compounds goes back a long time; in fact, one of the earliest reports was on the carcinogenicity of arsenic not many decades after the pioneering report of Sir Perceval Pott on cancer in chimney sweeps. Since then cancer has been definitely associated in humans with chromium compounds, nickel, and with less assurance but probably definitely with beryllium and cadmium. The confirmation of these findings in laboratory animals has been uneven. In the case of arsenic, for example, there has been only limited success in the production of cancer in laboratory animals with arsenic.

Many other metals have been found in laboratory studies to produce cancer, although with most of these, evidence of production of cancer in humans is either absent or extremely uncertain.

The extensive body of recent information relating to the testing of metals with a variety of short term tests will be briefly reviewed.     

Photoreduction of parathion ethyl †

Photodegradation of parathion ethyl in presence of unsaturated biomolecules takes place by sufficient reduction of the phenyl nitro group. Main products are N,0‐dialkenyl hydroxylamino, cis/trans azo, and azoxy derivatives of parathion.     

Influence of interspecific competitors on behavioral thermoregulation: developmental or acute plasticity?

Behavioral Ecology and Sociobiology - Deep-diving sperm whales have a complex social structure and the largest brain of any animal, but very little is known about the ontogeny of their diving,...     

Mutagenic and carcinogenic effects of cadmium†

There is limited evidence to suggest that certain compounds of cadmium may be mildly mutagenic. Some compounds of cadmium have given rise to fibrosarcoma and rhabdomyosarcoma following subcutaneous or intramuscular injection in the rat and also to interstitial cell tumours of the testis as a sequel to cellular damage and testicular atrophy. Cadmium chloride aerosol inhalation has induced a high incidence of dose‐related lung cancer in the rat. Epidemiological observations suggest that mortality from prostatic and possibly from lung cancer has been increased in cadmium workers who had experienced the very high levels of cadmium, mainly as oxide dust or fume, which existed in the past. The more recent epidemiological studies have not added further evidence in support of the carcinogenic role of cadmium.     

Analysis of particulates by Raman microprobe†

We describe a new technique of local analysis of heterogeneous samples which is analogous to the electron or ion microprobe but is based on molecular spectra of materials to characterize the different components of a microscopic sample and (or) to obtain images giving their surface distribution.

The sample is illuminated by a laser beam causing characteristic Raman lines of the different components to be emitted. Using these Raman lines, the components can then be detected, identified and located. For this purpose, the microscopic area of the preparation which is illuminated is analysed by a conventional optical microscope coupled with a tunable band pass filter and a multichannel or monochannel detection system.

One main advantage of this technique is the ability to study samples not under vacuum but in air, under a controlled atmosphere or even inside a transparent matrix. Normal Raman spectra can be obtained from particles of 1 μm size and larger, and are the basis for the qualitative identification of molecular constituents. The technique has been applied to molecular microanalysis of matter in the particulate form (i.e., Airborne particles—stack particulates, flyash, asbestos minerals, defects and inclusions in industrial materials,...). It is especially interesting for organic materials which are not detected by other methods. Different examples illustrate this technique.     

Biogeochemistry of aromatic hydrocarbons in the benthos of microcosms†

Investigations into the fate of petroleum compounds in the marine environment were carried out using experimental microcosms of two sizes and designs. Aromatic hydrocarbons or No. 2 fuel oil were spiked to the water of a 13 m³ continuous flow system and to a 2281 recycled flow system. The transport and alteration of this oil was traced in the sediment and benthic organisms (Glycera americana, Crepidula sp., and Nephtys incisa) of these microcosms. Measurable contamination was found in both sample types. The aromatic hydrocarbon distribution, including relative isomeric distribution (e.g., C₂‐phenanthrenes) was found to be different in sediment and in organisms from that which was originally introduced to the experimental microcosm. Differences in isomer distribution between Glycera and Crepidula were also detected. Based on the experimental data: molecular weight and specific isomeric form, biochemical processes, solubility, and particle adsorption/desorption influence the fate of petroleum compounds in benthic ecosystems.     

RP-HPLC determination of phytoplankton pigments—comparison of calibration results for two columns

In the experiment reported the results obtained with two different columns for the RP-HPLC analysis of phytoplankton pigment components were compared. LiChroCART Hypersil ODS and LiChroCART LiChrospher 100 RP18e columns were used exchangeably. Two groups of high-purity pigment standards of reference phytoplankton strains (23 chlorophylls and carotenoids in the first case and 25 in the second case) were subjected to chromatographic analysis. All calibration parameters (retention times, calibration curves, spectrum shapes, detection thresholds) for qualitative and quantitative identification of pigments were determined. The results indicate that both columns used in the analysis ensure sufficient resolution and can be used interchangeably for the analysis of pigments present in naturally existing phytoplankton, even when present in only trace amounts.Communicated by O. Kinne, Oldendorf/Luhe     

The polymorphism of the human serum paraoxonase †

The activity of paraoxonase, the enzyme which hydrolyses paraoxon, 0,0‐diethyl‐0–4‐nitrophenylphosphate, in human serum shows a genetically determined polymorphism with strong interethnic differences. The serum paraoxonase genotype has a significant influence on the paraoxon clearance. Individuals with high serum paraoxonase activity may be better protected against the toxic effects of parathion (0,0‐diethyl‐0–4‐nitrophenylthiophosphate).

In Caucasians the polymorphism is governed by two alleles. The first allele has a gene frequency p _low of 0.67 to 0.78, and is manifested in both the form of a first homozygotic group with low activities and a second heterozygotic group with medium activities. About 50% of all Europeans belong to the low activity group. The second allele with a gene frequency q _high of 0.22 to 0.33 is manifested in the second heterozygotic and a third homozygotic group with medium resp. high activities. The Hardy‐Weinberg rule for a two allele model is valid for the distribution.

The percentage of the low activity group decreases as one moves from Europe to Africa and Asia. In most of the Mongoloids and Negroids only 5 to 20% of the population can be included in the low activity group, which is not even demonstrable in Aborigines, Maoris, Tonga and some African and Indian (Central America) tribes. The validity of the Hardy‐Weinberg rule for a two‐ or three‐allele model must be rejected in non‐Caucasians.     

Bioavailability and metabolism of hexavalent chromium compounds †

Intratracheal instillation of ⁵¹CrCl₃ in anaesthetized rabbits resulted in partial absorption. In blood, the absorbed material was entirely confined to the plasma compartment. Only trace amounts were deposited in liver and kidney. By contrast, after similar application of Na, ⁵¹CrO₄ the bulk of blood radioactivity was present in red blood cells (RBC). Substantial deposition occurred in liver and kidneys. It is concluded that Cr(VI) may enter the body unreduced via the lung and is partially deposited in cells over a prolonged period of time.

Since chromium was accumulated in liver after administration of Cr(VI) we investigated the intracellular disposition of Cr(VI) in the isolated perfused liver. No significant sex differences in chromium distribution were observed. At the end of the experiments (1 h), 60% of the applied dose (312μg Cr/liver) was located in the cytosol, whilst 14% was in the mitochondria, 9% in the microsomal pellet and 2% was associated with the nuclei. Gel chromatography of the cytosolic compartment showed that the overwhelming part of chromium was eluted in fractions with an apparent molecular weight of 6,000 dalton. These fractions exhibited absorption maxima at 410nm and 548nm. It is concluded, that cytosolic reduction might be the main intracellular redox pathway for chromates. This view was confirmed by monitoring the reaction of Cr(VI) with GSH in vitro. GSH reduced Cr(VI) without further cofactors under formation of GSH‐chromium complexes, which possibly represent major intermediates in the metabolism of Cr(VI).     

The use of the embryo‐culture techniques in the research on the Teratogenic and Mutagenic properties of metals†

Actually, embryos can be cultured from the one‐cell stage up to the blastocyst stage, and their development can be easily monitored at any time: severe effects caused by toxic compounds are traduced by rapid embryonic death, less pronounced effects can be expressed by a lowered cleavage activity or by an arrest of the development from a particular stage. The system can be improved by transferring the embryos at the blastocyst stage in another more complete medium where they can “implant”; and form an inner cell mass with differentiated ectoderm and endoderm. Since last years, it has also become possible to culture postimplantation rodent embryos for short periods involving a number of particularly critical stages of organogenesis, such as the formation and closure of the anterior neuropore. Embryo‐culture also represents a useful system to study cytogenetic effects of chemicals which are often linked to lethal or teratogenic effects. These different possibilities are illustrated by examples of studies already performed with metals, and dealing with their teratogenic and/or cytogenetic effects on pre‐ and postimplantation rodents.     

Alkylation of metals and the activity of metal‐alkyls†

The biochemical basis for resistance to metal ion toxicity is emerging though it is complicated by the different resistance mechanisms. Several strategies for resistance to toxic metal ions have been identified:

1. The development of energy driven efflux pumps which keep toxic element levels low in the interior of the cell. Such mechanisms have been described for Cd(II) and As(V).

2. Oxidation (e.g. AsO^2‐ to AsO₄ ^3‐) or reduction (e.g. Hg²⁺ to Hg⁰) can enzymatically and intracellularly convert a more toxic form of an element to a less toxic form.

3. The biosynthesis of intracellular polymers which serve as traps for the removal of metal ions from solution such as traps have been described for cadmium, calcium, nickel and copper.

4. The binding of metal ions to cell surfaces.

5. The precipitation of insoluble metal complexes (e.g. metal sulfides and metal oxides) at cell surfaces.

6. Biomethylation and transport through cell‐membranes by diffusion controlled processes.

In this short review I shall discuss the implications of biomethylation as a detoxification mechanism for microorganisms as well as for certain higher organisms.     

Source and Distribution of Heavy Metals in Food†

Diet is the major route of exposure to contaminants, therefore food monitoring programs are essential to estimate the degree of human risk. We present a study undertaken in Austria dealing with the total uptake of mercury, cadmium, arsenic and lead. The dates were calculated from diet as well as from commercial food. Higher contaminated foodstuffs are registered and a comparison was made between body burden of babies and adults on the basis of bodyweight. It was pointed out, that cooked or washed food ready to eat has a lower burden of lead and cadmium than non prepared food due to household processing, which may decontaminate food to some extent’ Arsenic content was not reduced, because it is bound chemically and not primarily deposited on the surface of food such as in the case of lead. It was mentioned that the type of compound plays an important role in the assessment of action of those elements on humans.

Their biological activity is not necessarily related to their concentration in food. As in the case of selenium and chromium one has to take into consideration that there is a narrow range between benefit and harm, toxicity, and minor supply.

Therefore we conclude, that due to interactions of elements, their different biological activity and due to the difference of compounds present in food, we have to relativate the terms toxicity and essentiality. Then we can get a more profound insight into the problems of trace elements in food.     

Multielement techniques for determination of Inorganics in Watert†

Several analytical techniques offer the capability of simultaneously determining 5–40 elements. Of those techniques available, two are receiving extensive attention for analysis of water samples. Optical emission spectrometry utilizing plasma excitation has now evolved to a status in which 20–40 elements can be simultaneously determined on a routine basis. Instrumentation which relies on ion exchange chromatography separations is available which allows the routine determination of several cationic and/or anionic species.

The present discussion will briefly describe each of the above multielement analysis systems, summarize their respective capabilities and limitations with respect to water analysis, and project areas of future development.     

Optical–digital system applied to the identification of five phytoplankton species

An optical–digital system was used to identify five phytoplankton species of the genus Ceratium from Todos Santos Bay off the California Peninsula, Mexico. The system recognized species with at least 90% accuracy, regardless of their size, rotation, or location within the field of view. An optical filter was constructed for each species for correlation with each of six test images. The test images were prepared from phytoplankton samples containing one or more of the five species. The correlations demostrate that each optical filter recognized its corresponding species. To evaluate the effectiveness of the correlation technique, we used images of 100 different samples. The species of the Ceratium genus analyzed are important indicators, since some are responsible for red-tide blooms in areas surrounding Todos Santos Bay, Baja California, Mexico. In the near future, based on the method proposed, we plan to implement an automated plankton-identification system to determine marine eco-biological properties. Received: 9 June 1997 / Accepted: 3 June 1998     

Influence of grazing practices on cow milk quality: a case study on the Comarnic–Poieni bauxite quarry, Romania

The current study represents a preliminary investigation made into the influence of cattle grazing in the area of a bauxite quarry (Comarnic–Poieni, Romania) on the cow milk chemistry. Weathering and surface runoff in the bauxite quarry contaminate the local chemistry of the soil, vegetation and water. During cattle transhumance, cyclic feeding patterns occur, with grazing alternating between clean pastures and the area of the quarry. Soil and water samples were collected from the contaminated area of the quarry. Raw milk samples were collected during two stages, corresponding to the periods of grazing on clean pasture and the quarry area, respectively. Based on the obtained data, the relationship between cattle grazing and the composition of milk was interpreted. Preliminary results indicated a direct correlation of increased concentration of Al in the milk, following grazing in the bauxite quarry.     

Biological effects of selenium and relationships with carcinogenesis†

Selenium is an element that is both essential, at low levels of dietary intake, and toxic, at high levels, to man and a wide range of animals. The purpose of the communication is to review the evidence for the involvement of hyper‐ and hypo‐selenosis in the aetiology of cancer. High levels of selenium intake have been blamed for an increase in cancer, dental caries and an array of other conditions. Critical evaluation of the evidence, however, leads to the conclusion that such claims have little substance. On the other hand, low levels of selenium intake may be associated with an increased incidence of certain forms of cancer and there appears to be an inverse relationship between blood selenium levels and cancer death rate in the United States. Keshan Disease, in the People's Republic of China has only been shown to occur in populations severely depleted of selenium. In well nourished populations there is no evidence to indicate that selenium supplements have any prophylactic or therapeutic benefit against human diseases such as cancer, cardiovascular disease or cystic fibrosis.