Development of Sediment Reference Sample for Toxicity Tests

Development of depository Reference Sample for Toxicity TestsDevelopment of deposition credit rating sample for oto toxicity rendering victimisation Microtox steadfast physique canvas and Metal Fractionation using iodine originsAbstract chemic characterisation of pollutants using fractional process techniques and bioassays argon personaful monitor assholes for fix t unrivaled of voice judicial decision. However, a common criticism of down payment bioassays is the lack of an withdraw beginning positation sample which sample repository toxicity potbelly be comparatively assessed. In this contract an approach of obtaining a reference sediment sample by cleaning the sediment samples with metals was tested. Metal fractional process was carried out by applying single declivity techniques modified from a back-to-back extraction lineation proposed by Tessier et al (1979). The full metal concentrations were characterised using nitrous acid digestion. The sediment samples before and after the extractions were analysed using the Microtox Solid Phase Test (SPT). Comparison of total metal concentration with several(a) sediment tincture guidelines suggests that the sediments atomic number 18 polluted due to risqueer concentrations of Cu , Ni , Pb , Cd and Zn. The fractionation studies reveal that metals atomic number 18 contained in the main at bottom Fe-Mn Oxide frame.The comparison of the firmness of purposes of the SPT with various sediment classification methods suggests that the sediments argon slightly toxic to non toxic. However, the results of changes in the toxicity of sediment residues obtained after apiece extraction comp atomic number 18d to rough sediment toxicity results atomic number 18 non statistically signifi jackpott. still the comparison of toxicity results of sediment residues obtained after HNO3 and NaOAc digestion with the toxicity value of replicate1 of crude(a) sediment suggests a marginal decr comfort in the toxicity of sediments fleck the comparison of toxicity value of MgCl2 , NH2OH.HCl, HNO3+H2O2 indicates an increase in the toxicity of sediment residued. The comparison of toxicity values of all sediment residues with that of replicate2 of unprocessed sediment indicates an increase in the toxicity of the sediments after extractions.Get wisdom, keep netherstanding. recognition is supreme therefore get wisdom. Though it represents all you piddle, get understanding (Proverbs 4 5,7). Then you forget know the truth, and the truth will set you free (John 8 32). CHAPTER 1INTRODUCTIONBackgroundWith the suppuration interest in the rules that govern the fate of pollutants in urban purlieus, the sediments of urban rivers pose a particularly challenging scientific problem as m whatsoever persistent taints (e.g. metals, persistent essential pollutants (POPs)) tend to concentrate in river bed sediments. Therefore, the perspicacity of sediment quality is recognised as a critical ste p in understanding the risks associated with man made contaminant in the riverine system (De Miguel et al , 2005). Depending upon the conditions in the river, pollutants springiness to sediment whitethorn shape bioavailable and impose toxicity on aquatic organisms. Chemical synopsis alone is non adequate to explain effects of chemics present in the sediment (Beg and Ali, 2008) as they do not prove that adverse effects ar occurring (Luoma et al , 1995) , olibanum for optimal characterization and assessment of taint , issues concerning both concentration and toxicity should be addressed (Mowat et al , 2001).Therefore, be go of the request to establish a cause -effect relationship amidst the concentration of pollutants and attendant environmental damage and to measure the possible synergistic effect of Byzantine mixture of chemicals(Girotti et al , 2008), Microbial toxicity tests ground on bacterium have been astray use in environmental toxicity wake due to the simila rity of conf utilize biochemical function in bacterium and higher organisms (Mowat et al , 2001) .Among the bioassays unassailable chassis tests are useable and replete(p)ly apply as test organisms are exposed to whole sediments which include peeing soluble and non polar substances and thus offer a high relative realism for toxicity assessment of sediments. However, sediment toxicity tests dominate reference sediment exclusive of contaminant with similar physico chemical characteristics as the test sediments (Guzzella , 1998).The microtox test based on bacterial bioluminescence which uses V. Fischeri bacteria as test organism represent one of the most suitable test for sediment toxicity assessment as it corporation be used on extracts as well as directly to the sediment (solid phase test) ( Calace et al , 2005).As it is now widely recognised that the total concentrations of sour Metals indicate the extent of contamination, but they provide little information round the forms in which Heavy Metals are present, or about their potential for mobility and bioavailability in the environment (Lake et al , 1987) , knowledge on metal speciation in the sedimentary environment may be of more importance for hazard assessment than the total metal concentrations( Farkas et al , 2007). For this reason, in series(p) extraction mathematical processs are comm still applied because they provide information about the fractionation of metals in the diametric lattices of the sediments and other solid samples (Margui et al , 2004).It is against this background that an investigation into establishing a reference sediment sample for solid phase bioassays was undertaken in relation to Microtox solid phase test utilising single extractions of metal fractions using -same conditions and summonss set forth in the sequential extraction procedure mentioned in Tessier et al 1979 .Aims and ObjectivesThe main sire of the study is to assess whether the approach of cleaning t he sediment with metals using single extraction steps of sequential extraction is an appropriate alternative to acquire a sediment reference sample or not.In rules of company to obtain a reference sample exclusive of metals, the following procedure was adoptedEach extraction step described in the Tessier scheme was applied to separate aliquots of sediment samples using the same extraction conditions and chemicals described in the scheme (see section 3.8 for details). After the extraction step water-washed and dried residue sediment samples were analysed for toxicity using the Microtox solid phase test. A reduction in the toxicity could be expected as the metals were removed using chemicals. Microtox solid phase test was also conducted on unprocessed sediment so that a relative comparison amidst toxicity measurements could be made.The objectives of the investigation are summarised as follows To characterise the sediments for total bio available metal concentration for eight heavy metals (Cd, Cr, Cu,Fe,Mn,Zn,Pb,Ni) using nitric acid digestion method.To characterise various fractions of metals as described in the Tessier dodging using single extraction procedures.To determine the level of toxicity associated with unprocessed and processed sediment sample using the Microtox solid phase test.CHAPTER 2LITERATURE REVIEW2.1 urban River Sediments and PollutionUrban rivers have been joined with water quality issues since the nineteenth century when it was usual practice to solve untreated domestic and industrial waste into water courses. Since then the position has been improved due to e.g. the management curtailment of pollution at cloaca treatment plants. However, because of high population densities in urban areas due to motley of sources of pollution the degradation of urban rivers is still important today (Goodwin et al , 2003).When released into the river environment many anthropogenic chemicals bind or adsorb on to particulate involvement matter and dep ending upon river morphology and hydrological conditions such particulate matter along with associated contaminants can settle out along the water course and become part of the bottom sediments(Vigano et al , 2003). outcomely , sediments are considered as repositories for somatogenic and biological debris and for many pollutants (Calmano et al , 1996).Further more , under various physical , biological and chemical conditions (e.g. aqueous solubility ,pH, oxidoreduction , affinity for sediment natural carbon , grain size of sediments , sediment mineral constituents and quantity of acid volatile sulfides) these contaminants may become bioavailable and result in a toxic impact on aquatic biological science(Ingersoll et al , 1995).Nowdays, escalating evidence of environmental degradation have been confirmed where water quality guidelines for contaminants are not surpassed but, still organisms in or near the sediments are adversely affected (Ingersoll et al , 1995).Thus, with a vis ion to cheering aquatic biota, improving water quality and managing problems of re dangling and the vote down deposition of dredged materials, sediment quality assessment has been a crucial scientific and legislative issue in recent years. ( Calmano et al 1996 Nipper et al 1998).2.2 Water Framework directive (WFD) The atomic number 63an Unions(EU) Water Framework Directive (WFD) which came in effect on 22 December 2000, is one of the most important pieces of environmental code and is likely to transform the way water quality is being monitored within all member states ( Allan et al , 2006).The main objective of the Directive is to improve, protect and prevent further deterioration of water quality across Europe and it aims to achieve and ensure good quality status of all water bodies through and throughout Europe by 2015. Thus the necessity of addressing water quality issues associated with urban rivers has been increased within Member States(Goodwin et a, 2003). Under the WFD , tercet modes of monitoring strategies are specified and at each strategy level chemical monitoring , biological/ecological assessment , physico-chemical and hydro morphological tools have been cover to assess the water quality status of the body(Allan et al,2006).In the WFD, EU focal point places emphasis on establishing quality standards related to the concentrations of priority substances and substances which may cause harm in water , sediment or biota . (Crane , 2003).2.3 Sediment and Pollutants Sources in Urban Rivers Urban river system is much more complex in its sediments and pollutant sources. Sediments may be released into urban rivers due to erosion of land surface through categorization of physical and chemical processes, the rapid scat off from impervious surfaces, routing through waste pipe network, retention tanks and winter gritting roads (Goodwin et al, 2003). These sediments may contain or associated with pollutants such as hydrocarbons , garden and beast w astes , fertilisers , pesticides , oils , detergents , deicing chemicals , street litter (Hall, 1984 Chapman, 1996) and trace and heavy metals (Collins et al, 2007). Moreover, Combined Sewer flowage (CSO) events also augment the pollutant and sediment load due to its own contaminant load and the erosion and wash out of in-sewer sediments (Fierros et al , 2002). Due to the wide variety of sources and river dynamics there exist a wide spacial and temporal variation in the properties of sediments.2.4 River Sediment Composition and dynamics River sediments are mainly composed of mineral specks originated from the parent rocks due to erosion process, particulate radical matter adsorbed on mineral particles or particle surface organic matter which originates from plant detritus and savage debris, adsorbed nutrients and toxic inorganic and organic pollutants (Chapman , 1996). However , with respect to their behaviour in nature , sediments can be classified in two distinctively diff erent groups a) fair sediments with particles smaller than 50m (i.e silt and clay) and b) coarse sediments with size exceeding 50m ( i.e. sands and gravels) (Salomons et al , 1984).The erosion, transportation and deposition of sediment is a function of river flow velocity, particle size, water content of the material (Chapman , 1996) , channel structure and degree of turbulence(Goodwin et al , 2003). Under certain hydraulic conditions sediments can be transported in suspension or by traction along the bottom which is often called pick out buck. The suspension mechanism initiates the movement of fine particles maculation the Bed Load causes the movement of coarse particles (Chapman , 1996). More over, within urban catchments rapid outpouring and CSO events trigger river flow events with short efflorescence times and high peak flows which step up transport of sediments and associated pollutants (Goodwin et al , 2003).2.5 Sediment Quality sound judgementHistorically, the assessm ent of sediment quality has often been limited to chemical characterisation. It helps to kick downstairs what are the contaminants and what is their concentrations(McCauley et al , 2000) and it provides information about the condition of sediments and processes within them(Wolska et al , 2007). However, quantifying contaminant concentration alone can not provide large information to evaluate adequately potential adverse effects, possible fundamental interaction among chemicals or the time dependent availability of these materials to aquatic organisms ( Ingersoll et al , 1995) because it is meshuggeneh to analyze all the compounds and their synergistic/antagonistic effects contributing to toxicity(Plaza et al , 2005). As the bioavailability of pollutants to aquatic biota and their effects on the biota is the find out concern in sediment risk assessment , ecotoxicological testing (bioassays) of sediments which study the toxic effects of sediment contaminants on living organisms ( e.g. fish , plants , bacteria , algae) has been extensively used ( Rand et al , 1995).Thus, to understand the fate of pollutants in sediments and their impacts on aquatic biota , a tiered biological and chemical assessment methods have been implemented (Calmano et al , 1996) . The sediment quality triad methodology, one of the most widely used tiered approach based on weight of evidence combines 1) Identification and quantification of contaminants (i.e. chemical analyses ) , 2) Measurement and quantification of Toxicity based on bioassays (toxicity tests) and 3) Evaluation of in situ biological effects(e.g. Benthic community structure) (Calmano et al , 1996 McCauley et al , 2000 ).Principal advantages are that it can be used for any sediment type (Calmano et al ,1996) and as both biological and chemical components are used , environmental significance of contaminated sediments is addressed (McCauley et al , 2000). However the cause -effect relations are not always set due to the synergistic/antagonistic effects of chemicals causing toxicity in sediments (Calmano et al , 1996 McCauley et al , 2000) . Furthermore, the assessment is very site specific and does not allow empirical calculations of chemical specific guidelines ( Mc Cauley , 2000).2.6 Metals in Urban Sediments and Sources Metals are natural components of biosphere (Luoma , 1983) and they are introduced in to the aquatic environment through many lithogenic and anthropogenic sources(Zhou et al , 2008). Chemical leaching of bedrocks , water drainage basins and run off from banks are considered as the major lithogenic sources of metals (Zhou et al , 2008) while emissions from industrial processes ( e.g. mining , smelting , finishing , plating , paint an dye manufaturing) (Rand et al , 1995) and through urban sewage, house hold effluents, drainage water, business effluents , atmospheric deposition and traffic related emissions transported with storm water (Karvelas et al , 2003) are the major anthr opogenic sources of metals in the aquatic environment. Upon discharge to the aquatic environment metals are partitioned between solid and liquid phase (Luoma , 1983) and eventually as a result of settling metals associated with solid phase accumulate in bottom sediments(Farkas et al , 2007).Thus , sediments are main sink of metals in aquatic environment(Morillo et al , 2002).A comparison of typical concentration of metals in urban river sediments is presented in the Table 2.1.Table 2.1 Concentration of metals in urban river sediments(g/g) (reproduced from De Miguel et al , 2005)CrCuFe(%)MnNiPbZnRiver Henares, Spain(97-180)(7-270)(0.8-3.16)(150-445)(11-128)(17-1280)River seine , France842.91162429River Sowe , UK47.9164411786Semarang , Indonesia(12.3-448)(5.2-2666)(53.7-1257)Danube River, Austria43.553.9187Tiber river , Italy(18.2-54.2)(13.3-45.5)(3.6-33.5)(12.4-43.1)(53.4-417.6)River Po, Italy(118-223)(45.2-179.9)(4.5-5.2)(355-1159)(99-237)(39.3-71.8)(127-519)River Sherbourne38712.9 48119118196River Manzanares(18-1260(11-347)(1.9-9.1)(305-1276)(5-47)(42-371)(70-591)In brackets minimum- maximum values in italic arithmatic mean values2.7 Toxic metals and their forms in sediments Although rough metals are essential micronutrients (e.g. Mn, Fe, Cu,Zn) , almost all metals are toxic to aquatic organisms and human health if exposure levels are sufficiently high (Luoma , 1983). Among the toxic metals cadmium, chromium, copper, lead, nickel, zinc, mercury and arsenic are of prime importance due to their railroad tie with anthropogenic inputs. Under different physical, biological or chemical conditions the toxicity of metals in sediments is a matter of bio availability (Jennett et al ,1980).Thus in order to estimate the bio availability of metals and their potential toxicity it is desirable not only to determine the total concentration but also the different chemical forms or ways of binding between metals and sediments(Albores et al , 2000).In sediments depending up on various physical, chemical and biological conditions , metals partitioned into different chemical forms associated with a variety of organic and inorganic phases (Farkas et al , 2007).In river sediments metal can be leaping to various compartments e.g. adsorbed onto clay surfaces or iron and manganese oxy hydroxides, present in lattice of secondary minerals such as carbonates, sulphates or oxides, occluded within amorphous material such as iron and manganese oxyhydroxides, complexed with organic matter or lattice of primary minerals such as silicates (Gismera et al , 2004). Due to natural and anthropogenic environmental changes these associations can be alter and metals can become more or less bio available or mobilized within different phases. These influential factors include pH, temperature , redox potential , organic matter decomposition , leaching and ion exchange processes and microbial activity(Filgueiras et al ,2002). Thus in relation to their mobility and bioavailabili ty, in order of decreasing interest the major metal fractions are 1) Exchangeable ,2) Bound to carbonates , 3) Bound to Fe-Mn Oxides , 4) Bound to organic matter and 5) Residual .2.7.1 Exchangeable Metals In this fraction , weakly adsorbed metals retained on the solid surface by relatively weak unchanging forces that can be released by ion exchange processes in the sediment are included(Filgueiras et al , 2002). These metals are considered the most available form of metals present in the sediments (Morrison , 1985).2.7.2 Metals Bound to Carbonates Metals in this fractions are co-precipitated with carbonates which exist as cement and cover (Morrison , 1985) and this phase can be an important adsorbent for metals in the absence seizure of organic matter and Fe-Mn oxides (Filgueiras et al , 2002).2.7.3 Metals bound to Fe-Mn Oxides Metals in this fraction are associated with smoothing iron and Manganese oxides which exist as nodules , concretion and cement between particles or simpl y as a coating on particles. Iron and Manganese oxides are considered as excellent scavengers of metals and are thermodynamically impermanent under anoxic conditions (Tessier et al , 1979).2.7.4 Metals bound to organic matter In this fractions metals associated with various forms of organic materials such as living organisms, plant and animal detritus or coatings on mineral particles are included. This fraction is considered to be less mobile due to their associations with humic substances of higher molecular weights(Filgueiras et al , 2002).2.8 Sequential Extractions A sequential extraction procedure (SEP) also cognize as sequential extraction scheme (SES) can be used to determine above mentioned binding fractions of metals in the sediment. In this process, given sediment sample is subjected to a series of increasingly strong , phase specific reagents under controlled condition which extract our metals from the particular physic-chemical phase of interest(Bird et al , 2005).Depen ding upon can fractions, a wide variety of chemical extractants can be used (see fig.2.1) and thus in the literature various sequential extraction schemes are available which differ in the use of extractant, intention phase and the order of attack to separate particular form of metals. The majority of the schemes are variants of a scheme proposed by Tessier et al (1979). Many researchers have reported difficulties in comparing the results of SES due to their wide variation in the use of chemicals and target phase. Thus, in an effort to harmonize the different methodologies and to make the comparison of results easier , union Bureau of Reference (BCR) proposed a three step extraction procedure along with a reference sediment material to certify the protocol (Mossop and Davidson , 2003).2.9 Advantages and problems of sequential extractions The application of sequential extraction techniques , though time down provide valuable information about the origin , mode of occurrence, bio logical and physic-chemical availability , mobilisation and transport of metals within the sedimentary matrices(Tokalioglu et al , 2000).However, since their initial study, sequential extraction schemes have been criticized for the lack of selectivity of reagents, issues of re adsorption and redistribution of metals solubilised during extraction and changes in speciation due to sample pre- treatment and its general methodology ( Gleyzes et al , 2002).In the sequential extraction scheme, the reagents are expected to attack only the target phase without solubilising the other phases. However, it has been tack that the reagents are not discriminating and may affect other phases also. Thus the sequential extractions are termed as operationally defined fractionation techniques. This lack of selectivity may cause re-adsoprtion and re distribution of metals among the target phases. Moreover, incomplete dissolution of some phases and changes in pH may also lead toward re adsorption and r edistribution problems (Gleyzes et al, 2002). Various researchers have reported the problem of re adsorption and redistribution for many sequential extractions for each phase.Despite these limitations sequential extractions are widely accepted for metal fractionation in sediment samples to assess the mobility and bioavailability of metals .2.10 Single Extractions To reduce lengthy procedures and thus making sequential extractions a part of routine analysis, various alternatives(e.g. nuke heating and ultrasonic shaking) to conventional extraction procedures have been employed (Albores et al , 2000). One of the alternatives to reduce the lengthy and laborious sequential process is to use single extractions. In single extractions the same reagents and operating conditions as the sequential extractions are applied to different sub- sample (Albores et al ,2000) and, except for start step , the metal concentrations in each individual step can be obtained by subtracting the results obtai ned in two successive steps(Filgueiras et al , 2002). ab initio this technique was suggested by Tack et al (1996) in which first three steps mentioned in Tessiers Scheme were extracted simultaneously while, for organic matter bound metals, it was suggested that the sample should be extracted first for reducing metals and should then be re treated with hydrogen peroxide step to remove organic matter and thus release metals bound to this phase.2.11 Bioassays A useful monitoring toolBioassays measure changes in physiology and behaviour of living organisms resulting from stress induced by biological or chemical toxic compounds which can cause rupture of e.g. metabolism. Thus, bioassays help to establish cause / effect relationship between the concentrations of pollutants and consequent environmental damage (Girrotti et al , 2008).Historically fish and macro instruction invertabrates bioassays are the first in the series of toxicity bioassays involving animals. As these bioassays wer e found useful in assessing the acute toxicity of chemicals and effluents and often predicted their effects of aquatic biota and habitat, they have been extensively used in the screening of chemicals and regulatory deference monitoring (Blaise et al , 1998). However , these conventional bioassays require longer test duration along with additional time(e.g. acclimatisation) for preparations of the test (Ribo and Kaiser , 1987). Moreover toxicity was found a trophic level property and thus it was realized that security department of aquatic resources could not be ensured by conducting bioassays solely at macro organism level (Rand et al , 1995).Therefore an urgent requirement of cost effective , multi trophic and faster bioassays was strongly felt which led to development of micro scale testing procedures involving bacteria , protozoa , micro algae and micro invertabrate (Blaise et al , 1998). Distinct advantages of microbial testing procedures include 1) ease of handling ,2 ) short testing time , 2) reproducibility of results (Mowat et al , 2001) and 4) cost effectiveness (Wadhia and Thompson , 2007).2.12 Sediment Toxicity Tests As Van Beelen (2003) stated, toxicity is not a substance property only , but it is the c