School of Chemistry UNIVERSITY OF KWAZULU-NATAL, WESTVILLE NOVEMBER 2008 EXAMINATIONS CHEM 781: HONOURS ELECTIVES CHEM781: Speciation, Toxicity and Bioavailability DURATION: 1½ Hours TOTAL MARKS: 100 Internal Examiner: Prof. S.B. Jonnalagadda External examiner: Prof: O.J. Okonkwo, TUT, Pretoria NOTE: This paper consists of 3 pages. Please check that you have them all. Answer ALL questions Students are requested, in their own interests, to write legibly.
Page 2 Special Topics: Speciation, Toxicity and Bioavailability 1. Discuss the speciation of lead and copper and it s bioavailability with the help of the following Pourbaix diagrams. (15)
Page 3 2. Describe the mechanisms of metal toxicity on human body and the ways in which the body responds to it. (12) 3. Write the structures of common methylated species and volatile hydrides of arsenic. (5) 4. Roxarsone (3-nitro-4-hydroxyphenylarsonic acid) and other similar pesticides are used as poultry feed additives to control coccidial intestinal parasites. Discuss the possible pathways for biotransformation of Roxarsone. (13) 5. List four major variables that influence the metal mobility in waters. (4) 6. Sketch a diagram to illustrate the cycling of mercury in the aquatic system and atmosphere. (10) 7. Discuss why speciation is paramount in bioavailability and toxicity. Justify your answer taking any metal as an example, which is not used in other answers. (10) 8. Discuss the role and speciation of selenium with respect to human health. (6) 9. Describe the heavy metal adsorption mechanisms in soils, commenting on the heavy metal interactions, metal-organic matter interaction and effects of redox processes. (15) 10. Discuss the implications of interactions between the metals and how they impact on plants and organisms. (10) End of Examination Paper.
Page 4 B.Sc. Honours Examinations: 2008: Special Topics: Speciation, Toxicity and Bioavailability M. Scheme/Solutions Duration: 90 minutes. Answer ALL the questions. 1. Important factor to consider is the chemical stability of copper species. A stability diagram showing regions of thermodynamic stability for different species under particular conditions is called a Pourbaix, phase, ph-potential diagram or Eh-pH-diagrams. These diagrams are compiled at fixed temperatures and under specific chemical conditions with axes of ph and Eh. The ph is the negative logarithm of acidity and alkalinity; the H + concentration. The Eh is a measure of the reduction or oxidation (electrochemical) potential of the system. Low Eh values indicating reducing conditions where electrons are supplied by the system to the species and high Eh values represent oxidizing conditions (Cu(II) + H 2 O 2 = CuO + H 2 O) where electrons are removed from the species. The simplest Eh-pH-diagrams are of a single element in water, such as the Cu-H 2 O system, in which several species can be depicted including the dissolved or precipitated ions (oxides, hydroxides, etc.). The Eh redox potential axis is scaled on the Standard Hydrogen Electrode. The ph described the supply and removal of protons from the system, being acidic and basic conditions, respectively. Only predominant species in a system are represented in these diagrams, where the lines represent equilibrium between two species under the same Eh-pH conditions. Horizontal lines involve reactions with electrons and no proton transfer, diagonal lines (positive or negative slopes) involve reactions with electron and proton movement, and vertical lines involve reactions where only protons are transferred. Figure 4 shows the Eh-pH stability diagram for copper and chloride (seawater concentration) together with an assumed carbonate equilibrium with air. The total carbonate content includes CO 3 2-, HCO 3 -, H 2 CO 3. The stability regions for the red Cu 2 O cuprite, the blue-green malachite and paratacamite (Cu 2 (OH) 3 Cl) are represented. These Eh-pH-diagrams can be of importance to researchers as they can be used as a tool to predict the form of the metal of concern under different conditions. This can be used to determine what percentage of the total metal content of a system is a threat to becoming bioavailable. Copper becomes bioavailable to an organism when it is in its desorbed form, thus if ph and redox conditions allow for the adsorption of copper to mineral surfaces or complexation of copper to organic matter the bioavailability can be reduced, and thus limit the degree of bioaccumulation in a biological system. Similar explanation for lead. 2. Mechanisms of toxicity Metals generally produce their toxicity by forming complexes (called "ligands") with organic compounds. The modified molecules loose their ability to function properly, which leads to the malfunction or death of the affected cells. Metals commonly bind to biological compounds containing oxygen, sulphur and nitrogen, which may inactivate certain enzyme systems or affect protein structure 1, 5. In addition to this, light toxic metals may compete with or replace similar metals, for example lithium competes with the similar metal sodium. In acute poisoning, large excesses of metal ions can cause disruption of membrane and mitochondrial function and the generation of free radicals. Due to this, generalized clinical effects, including weakness and malaise feature in most cases 1. The diagram below shows how many metals specifically exert their toxicity: How the body reacts to an intake of metal:
Page 5 Despite the considerable potential for metals to disrupt metabolic process, the body actually has several defenses against them. These are described in the paragraphs below: 1. Sequestration with metallotheonein Metallothionein is a low molecular weight intracellular protein, which is able to bind to at least 18 different metals, rendering them harmless. High concentrations of the protein are found in the liver, kidney, intestine and pancreas, and it is thought to be involved in the regulation of copper and zinc metabolism, and in the detoxification of heavy metals, particularly cadmium. Its synthesis can be induced in response to many substances, including zinc, cadmium, copper, mercury, gold, bismuth, and some non-metallic compounds (although often at non-physiological concentrations). It is also part of the acute phase response to inflammation, and it is thought that this may be a method by which the body can increase zinc availability for protein synthesis. Metallothionein is a potential preventative treatment for metal poisoning: studies have shown that induction of metallothionein synthesis by pre-treatment with zinc reduces the toxicity of many elements in mammals, and cadmium in humans 1. 2. Formation of nuclear inclusion bodies Lead, bismuth and mercury all cause the formation of these eosinophilic nuclear inclusion bodies in renal tubular cells. These are insoluble protein complexes, which contain the metal and stop it from further damaging the cells. The protein haemosiderin can sequester excess iron in the body in a similar mechanism 3. Biotransformation Several toxic elements are metabolised within the body, some to less harmful compounds, but some to more harmful ones. Some examples of this are given below: Mercury, arsenic and selenium can be methylated and demethylated by several pathways Selenium: excess selenium is excreted as trimethylselonium unless exposure is great, in which case volitile dimethylselonium is formed. This compound gives the breath the characteristic garlic odour. The methylation of mercury, lead and tin produces metabolites which are more harmful to the body. 3. Info attached 4. Info attached. 5. ph, redox conditions, inorganic ligands, organic ligands and competition from other ions 6. Sketch attached 7. Explanation of speciation term and justification taking any example not used any other answer. 8. Selenium in the +4 state occurs naturally as selenite. In alkaline solution, it tends to oxidize slowly to the +6 state, if oxygen is present, but not in an acid medium. It is readily reduced to elemental selenium by a number of reducing reagents, ascorbic acid or sulfur dioxide being commonly used for this purpose. It readily reacts with certain o-diamines, and this is used as the basis for some analytical methods. Selenium dioxide, the anhydride of selenious acid, sublimes at 317 C. This is important with regard to air pollution through the combustion of materials containing the element, and also to air sampling procedures. Dietary selenium can protect against the toxicity of several heavy metals, such as mercury or cadmium, and certain xenobiotics, such as paraquat, but the mechanism of these protective effects is not known.
Page 6 Selenium has been suspected of being a carcinogen in the past, but more recent research suggests that it may be able to protect against certain types of cancers in experimental animals. Effects on man: General population exposure Tthe main environmental pathway of selenium exposure in the general population is through food. Nutritional surveys have shown that extreme dietary intakes range from 11-5000 µg/day, but on most diets intakes between 20 and 300 µg/day can be considered as more typical. The extremes in intake are reflected in extreme levels of selenium in blood, mean reported values ranging from 0.021 to 3.2 mg/litre. The highest blood-selenium levels ever observed in the general population were found in an area of the People's Republic of China in which an episode of intoxication reported as selenosis had occurred some years earlier. In this respect, as well as in at least two other studies in over-exposed populations, hair loss and nail pathology were the most marked and readily documented toxic signs. The Task Group, being aware of the hepatotoxicity of selenium compounds observed in animal studies, noted that no clinical signs of hepatotoxicity were observed in the studies of people exposed to high levels of dietary selenium, but concluded that there is a need for more thorough evaluation of hepatic function in persons with high selenium exposure. Tooth decay was also observed in several studies on over-exposed populations, but in evaluating its significance the Task Group was unable to exclude interference by other environmental factors. The Task Group recognized the difficulty in establishing an exact dose-response with respect to selenium in the above studies. The range of blood values noted was 0.44-3.2 mg/litre; in these studies no adverse effects were reported at the lower level, whereas clear effects on the hair and nails were observed at and above a level of 0.813 mg/litre. The lowest blood-selenium levels ever reported in a general population were seen in regions of the People's Republic of China where Keshan disease and Kashin-Beck disease are known to be endemic. I intensive research is being carried out concerning the involvement of selenium in the multifactorial etiology of these diseases and the use of selenium compounds in their prevention. With current understanding it is difficult to conclude the possible relationship between low levels of selenium intake and a high incidence of cancer. 9. Discuss the heavy metal adsorption mechanisms in soils, commenting on the heavy metal interactions, metal-organic matter interaction and effects of redox processes. Heavy metal Adsorption mechanisms Soils have ability to adsorb metal ions from aqueous solution by various mechanisms which include physical and chemical adsorption, precipitation, and solid state diffusion. Metal ion transfer occurs at the solid solution interface consisting of inorganic colloids (e.g., clay), metal oxides and hydroxides, metal carbonates and phosphates, organic matter, and living microorganisms (algae and bacteria). Another influencing parameter is the ligands in the solution responsible for the distribution of metal ions, such as humic and fulvic acids. Anthropogenic ligands like Nitrilotriacetic acid (NTA), Ethylene-diamine Tetraacetic Acid (EDTA) and polyphosphates are also may be present in natural waters and soil. However, the most significant role in heavy metal retention, mobility and bioavailability is played by oxides of Fe, Al, and Mn as well as soil organic matter. The factors controlling exchange between heavy metal in the solution and the soil particles are: soil type, metal speciation, metal concentration, soil ph, solid:solution mass ratio, multiple ions in the solution (interacting ions) and contact time. Among them soil ph has the greatest effect of any single factor on the solubility or retention of metals, with a greater retention and lower solubility of metal cations which occurs at high soil ph
Page 7 Heavy metal interaction Heavy metals in soil or lechate can be present in the form of free and exchangeable ions and organic and inorganic complexes and can be precipitated by oxides of Fe, Al and Mn. They can also be bound within the crystalline lattice structure of primary minerals. Among the different forms, the water-soluble metals are the most mobile. These are free ions and soluble organic and inorganic complexes. The most stable heavy-metal forms are those incorporated into a crystal lattice structure by isomorphic substitution. The most unstable heavy metals are those exchangeable between the soil solution and the zone affected by the charged colloidal surfaces or the double diffuse layer. Soil ph plays an important role as it establishes H + concentration. H + ions are strongly attracted by the soil surface negative charges and H + are capable of replacing other cations. Factors affecting the replacing power of any ion in the cation exchange complex are valence of the ion, its hydrated diameter and other ions in the solution. In most cases, higher valency provides greater degree of adsorption. The strength with which cations of identical charge are held to the soil particle surfaces is inversely proportional to their hydrated radius. In the presence of multiple ions, competition between the metallic ions for adsorption sites occurs, affects sorption in soil and consequently, gives rise to difficulties in assessing the process. The preferred soil adsorption of one heavy metal over another species is called selective adsorption and it is dominantly influenced by the ionic size of heavy metals. On the basis of unhydrated radii, the expected order of selectivity is: Pb 2+ (0.12nm) > Cd 2+ (0.097nm) > Zn 2+ (0.074nm) > Cu 2+ (0.072nm). Metal-Organic matter interaction A number of organic compounds, such as humic and fulvic acids, can form complexes with metal ions. Depending on the stability of the complexes, they can be soluble or insoluble in the soil solution. When containing organic matter with a low C/N ratio, a high ph, and a high Ca 2+ content in the exchange complex, soils are considered to have a high adsorptive capacity. Such organic matter with a low C/N ratio is highly humified and is slightly soluble. The binding mechanism of heavy metals with organic matter includes complexation, adsorption, and chelation. The common inorganic ligands forming complexes with the metallic ion include OH -, Cl, SO 4 2-, CO 3 2-, PO 3 3-, and CN. The complexes produced may be +ve, -ve, or neutral. Complexation by co-ordination with multidentate ligands is termed as chelation. To form complexes, metal ions need to be chelated by two or more functional groups of the organic fraction, such as carbonyl, carboxyl, alcohol, phenol, and methoxyl. Complex formation in the soil solution results in competition between the ligands and the soil solids for the adsorption of heavy metals. The complication in predicting heavy metal mobility arises when complexes, such as soluble organic ligands (fulvic acid) are unable to adsorb onto the soil-particle surfaces. Metallic-ion complexes with Cl-ions, sulphates, and organics interfere with their adsorption by the soil particles. The water solubility of organic complexes depends on their stability. The soil ph plays a vital role in the stability of heavy metal complexes since with increasing ph, the ionization of the functional groups (carboxyl, phenolic, alcoholic and carbonyl groups) is increased accordingly. Effects of REDOX processes Heavy metals such as Cr, Fe, Hg, As, Mn, and Se are referred to as redox elements or redox couples, since they have more than one possible oxidation state. However, Ag, Cu, Cd, and Zn,
Page 8 with only one preferred valence state, can also be influenced by redox processes. Under very low redox conditions, Pb and Cd, with one oxidation state, form insoluble sulphide minerals. However, at a ph of 7 to 8, where redox conditions are not so low, they form insoluble carbonate minerals. Arsenic, with two oxidation states, is insoluble with sulphide under very low redox conditions but does not form a carbonate under any Eh-pH conditions. The changes in redox potential affect the soil ph. Reducing conditions enhance ph value while oxidation brings ph down. Oxidation of pyrite (FeS 2 ) in soil can significantly decrease ph. 10. Interactions between chemical elements may be both antagonistic and synergistic, and their imbalanced reactions may cause a real chemical stress in plants. Antagonism occurs when the combined physiological effect of two or more elements is less than the sum of their independent effects, and synergism occurs when the combined effects of these elements is greater. These interactions may also refer to the ability of one element to inhibit or stimulate the absorption of other elements in plants (fig.). All these reactions are quite variable and may occur inside the cells, within the membrane surfaces, and also surrounding plant roots. Antagonistic effects occur most often in two ways -the macronutrient may inhibit trace element absorption and, in turn, the trace element may inhibit absorption of a macronutrient. These reactions have been observed especially for phosphate, but also have been reported for other macronutrients whose uptake and metabolic activity may be inhibited by several trace elements. Most important for practical application are the antagonistic effects of Ca and P on heavy metals such as Be, Cd, Pb, and Ni that often constitute a health hazard. It is noteworthy that although the antagonistic effects of P and Ca on many trace cations and anions are frequently reviewed in the literature, the antagonistic impact of Mg on trace metals is only occasionally reported. Interactions observed within plants between trace elements have also indicated that these processes are quite complex, being at times both antago-nistic and synergistic in nature, and occasionally are involved in the metabolism of more than two elements (Fig). The greatest number of antagonistic reactions have been observed for Fe, Mn, Cu, and Zn which are, obviously, the key elements in plant physiology (Table). These trace metals are linked to processes of absorption by plants and to the enzymatic pathway. The other trace elements often involved in antagonistic processes with these four trace metals are Cr, Mo and Se. Synergistic interactions between trace elements are not commonly observed. Those reported for Cd and other trace metals such as Pb, Fe, and Ni may be artifacts resulting from the destruction of physiological barriers under the stress of excessive concentrations of heavy metals. Moreover, several reactions that occur in the external root media and affect root uptake should not be directly related to metabolic interactions, but the two reactions are not easily separated. One water quality parameter affecting cadmium uptake is the Ca 2+ and Mg 2+ concentration (hardness) of the water. Increasing Ca 2+ concentration reduces cadmium uptake through fish gills, cadmium accumulation, and cadmium toxicity for fish. Two mechanisms can be distinguished for the Ca 2+ -mediated reduction in cadmium uptake. The first is an inhibitory effect on uptake into gill tissue, while the second is related to the adaptive response of the fish to increased Ca 2+ concentrations. Mg 2+ also reduces cadmium uptake through fish gills but at 5 times higher concentrations than Ca 2+.
Page 9 Zinc also has been shown to reduce cadmium uptake through the gills (Wicklund, 1990). Like cadmium, zinc is assumed to enter the epithelial cell by facilitated diffusion (Spry & Wood, 1989) and, furthermore, Ca 2+ acts antagonistically on zinc uptake.