Difference between total analysis, fractionation and speciation

1. Difference between total analysis, fractionation and speciation KJM MEF 4010 Module 19

2. Module plan, Week 1 • Thursday February 15th; • Lecture, Auditorium 3, hr. 08:15 – 10:00 • Difference between total analysis, fractionation and species • The significance of species activities rather than total concentration in terms of mobility and toxicity • Chemical analytical speciation and fractionation (Al) methods • Water sampling from different compartments of the environment • Sampling strategies for environmental samples • Field work, Vansjø, hr. 10:00 ~ 17:00 • Visit to Morsa – Vansjø watershed • Sampling of water samples from forest streams, tributeries to Vansjø and Vansjø • Friday, February 16th; • Lecture, Auditorium 3, hr. 08:15 – 09:00 • Important species in natural water samples • Central equilibriums in natural water samples • Concentrations and activities • Labwork, hr. 09:15 ~ 17:00Analysis of: pH, Alkalinity, TOC, Major Anions and Cations on IC, Al-fractions and P-fractions

3. Module plan, Week 2 • Thursday, February 22th; • Lecture, Auditorium 3, hr. 09:15 – 11:00 • Challenges with simultaneous equilibrium • Speciation programs (MINEQL) • PC lab, hr. 11:15 – 16:00 • Practice in using MINEQL • Friday, February 23rd • Individual report writing

4. Total analysis • Most standard chemical analytical methods determine the total amount (component) of an element in the sample • AAS, ICP and IC • The sample is typically digested where all analyte is transferred to its aqueous form • Me(H2O)2,4 or 6n+ prior to analysis

5. What is speciation and fractionation? • Speciation* “ Specific form of an element defined as to isotopic composition, electronic or oxidation state, and/or complex or molecular structure. ” * D.M. Templeton, F. Ariese, R. Cornelis, L- G. Danielsson, H. Muntau, H.P. Van Leuwen , and R. Lobinski, Pure Appl. .Chem.,2000, 72, 1453

6. Fractionation* “Process of classification of an analyte or a group of analytes from a certain sample according to physical (e.g., size, solubility) or chemical (e.g., bonding, reactivity) properties.” • “Chemical Fractionation = Classification according to chemical properties.” * D.M. Templeton, F. Ariese, R. Cornelis, L- G. Danielsson, H. Muntau, H.P. Van Leuwen , and R. Lobinski, Pure Appl. .Chem.,2000, 72, 1453.

7. Why is chemical speciation important? • Mobility and solubility of a compound depend on in which form it can exist in solution • The bioavailability of metals and their physiological and toxicological effects depend on the actual species present – not on the total concentration • Examples: • Methylmercury (CH3Hg+) is kinetically inert, and readily passes through cell walls. It is far more toxic than inorganic forms • Organometallic Hg, Pb, and Sn are more toxic than inorganic • Organometallic As are less toxic than inorganic • Copper toxicity correlates with free Cu-ion conc.- has reduced toxicity in the presence of organic matter • Cr (VI) more toxic than Cr (III) • Inorganic Al are more toxic to aquatic organism than Al bound to organic ligand

8. Aluminum • Toxicity of Al, and its environmental and biological effect are associated with the forms present in aquatic system • In aquatic systems, Al exists mainly as: • free Al 3+ , AlOH 2+ , Al(OH)+, Al(OH)3 and Al(OH) 4 – • AlF 2+ , AlF +2, AlF3, • monomeric SO42– complexes, Al-Org • Al speciation depend on soln pH & conc. of ligand • Toxicity: • Al 3+, AlOH 2+, Al(OH)+(more toxic) • Al-F and Al-Org (less toxic)

9. Mobility of metals in soil and soil solution G.W Brummer In the : Bernhard M, Brickman FE, Sadler PJ (eds) The importance of chemical “speciation” in environmental process. Springer, Berlin Heidelberg New York, 1986, p 170

10. The significance of species activities in terms of mobility • The distribution of an element among different compounds profoundly affects its transport by determining such properties as: charge, solubility, and diffusion coefficient • Electronic and oxidation states • Profoundly affect mobility • E.g. The Fe(II) ion is soluble, whereas Fe(III) is more prone to hydrolysis and subsequent precipitation • Inorganic compounds • E.g. NiCl2 and NiSO4 are water soluble while NiO and Ni3S2 are highly insoluble in water • Inorganic complexes • Formation of hydroxides is often a key determinant of element solubility • Organic complexes • Organometallic compounds • E.g. Biomethylationof Hg by S reducing bacteria in soil is believed to be a main cause for increased conc. of Hg in surface waters in Scandinavia. • Macromolecular compounds and complexes • Dissolved natural organic matter (DNOM) complex heavy metals and sorb organic micro pollutants enhancing thereby solubility and mobility

11. The significance of species activities in terms of toxicity • The distribution of an element among different compounds profoundly affects its bioavailability by determining such properties as: Charge, solubility, and diffusion coefficient • Isotopic composition • Not very essential for toxicity • Electronic and oxidation states • Profoundly affect toxicity • E.g. Cr(III) may be an essential element, but Cr(VI) is genotoxic and carcinogenic • Toxic effect of As and its compound decreases in sequence As (III)>As (V) • Inorganic compounds • E.g. Occupational exposure to Ni and its compounds. Ni3S2 is a potent carcinogen • Inorganic complexes • E.g. Transient polymeric aluminum-hydroxo complexes with high toxicity • Organic complexes • Organometallic compounds • Hydrophobicity and volatility are important • Bioaccumulation in fatty tissues and penetration of membrane barriers • E.g. MeHg • Macromolecular compounds and complexes • Heavy metals and organic micro pollutants bound to DNOM are generally concidered less toxic.

12. Some metals of “environmental” concern

13. Effect of a pollutant • Determined by its concentration and physical, chemical and biological characteristics of the • Pollutant • Solubility in water and organic solvent • Bio-accumulability (Kow) • Degradability, persistence (t½) • Organic complexability (Kex) • Density • Recipient • Stagnant conditions (thermocline) • Hardness (Ca+Mg) • pH (speciation) • Dissolved Organic Carbon (DOC)

14. Chemical analytical speciation methods • Isotopic composition • Mass spectrometry (MS) (e.g. 14C/12C) • Electronic and oxidation states • A. Selective organic complexation with spectrophotometric detection • Separation with HPLC, detection with e.g. ICP • Inorganic compounds and complexes • B. Potentiometric determination of the activity of Free aqueous species (e.g. ion selective electrodes for Free F, Ca) • Organic complexes • C. Anodic stripping voltametry on electroactive species • Organometallic compounds • D. Separation with GC or HPLC, detection with e.g. ICP • Macromolecular compounds and complexes • Size exclusion, ion-exchange, affinity and reversed phase chromatography (e.g. P and Al fractionation) Speciation

15. Species/element specific techniques A. Spectroscopy • Less application because of low sensitivity • Oxy-anions of Mn, Cr absorb UV/vis • Many species can be determined by addition of selective color forming reagents • Molecular Absorption spectrophotometry (MAS) • For metal species using chromogenic reagents • Al-8HQ Speciation

16. Species/element specific techniques B & C. Electro analytical techniques • Ion selective Electrode (ISE) • Detection of free-aqua ions (F-, Cu2+, Cd2+, Pb2+) • Low sensitivity (µmol/L –mmol/L) • Less applicable for the speciation of metals • E.g. Fluoride Selective electrode - Al speciation • Polarographic and Voltametric methods Differential Pulse Polarography (DPP) • Gives separated signals for free-metal ion and metal complex • DC- polarography • Can be used to distinguish between red-ox states Speciation

17. Species/element specific techniques D. Separation and detection techniques Separation • Chromatography • Liquid chromatography/HPLC • Size, affinity to mobile/stationary phase • Gas chromatography (GC) • Volatility Detection techniques • Element specific techniques • HPLC: Photometer, Refractometer, Diode-array, Electrochemical and Conductivity • GC:TCD, FID • Element specific: FAAS , GFAAS, ICP-AES, ICP-MS Speciation

18. Liquid chromatography-HPLC • Samples introduce to chromatographic column of stationary phase (SP) • Mobile phase (MP) pumped through column • Separation based on the interaction of the analyte with the SP and MP • Separated speciation: • inorganic species, such as cations, anions (IC) • metal complexes • organometallic compounds Speciation

19. Speciation of mercury in soil and sediment by selective solvent and acid extraction * MP: Methanol+2-mercaptoethanol+NH3AC SP: C-18 column *Y. Han , H. M. Kingston , H. M. Boylan, G. M. M. Rahman, S. Shah, R. C. Richter, D. D. Link, S. Bhandari, Anal. Bioanal. Chem.,2003, 375,428.

20. Gas chromatography • Separation • Volatile and thermal stability • Species to be analyzed organometallic • Volatile species :-Me2Hg, Me2 Se, Me4Sn, Me3Sb Tetraalkylated lead • Non-volatile species after derivatization: • Different methods.. Speciation

21. GC-ICP-TOF-MS for the speciation analysis of organo-lead compounds in environmental water samples* • Toxicity increases with increase the number of alkyl group i.e speciation of organolead cpds necessary • In aqueous solution ionic …Derivitization NaEt4 • GC-oven program 600C to 2000C at 30 0C min-1 * M. Heisterkamp, F. C. Adams, Fresenius J. Anal. Chem.,2001, 370,597

22. Problem • Often, chemical species present in a given sample are not stable enough to be determined as such • During the measurement process the partitioning of the element among its species may be changed • This behavior can be caused by, for example, a change in pH necessitated by the analytical procedure, or by intrinsic properties of measurement methods that affect the equilibrium between species • Detection limit problems

23. Solution: Chemical analytical fractionation • Isolate various classes of species of an element and determine the sum of its concentrations in each class • In some instances, fractionation may be refined by supplementary speciation analysis. • With further analyses and calculations the inorganic fraction can be subdivided into individual species. Fractionation

24. Extraction • Soil sample • Leaching method • Sonication, stirring, shaking or soxhlet with organic solvent • Sequential Extraction (Tessier) • Supercritical Fluid Extraction (SFE) • Water sample • Solvent extraction • Ion exchange resins • Solid Phase Extraction (SPE) Fractionation

25. Ion exchange resins for fractionation of metals in water • Retains: • Free metal ions • Labile metal organic complex • Labile inorganic complexes • Eluted: • Non-labile metal complexes (Strong complexes) Fractionation

26. Example;Al fractionation • Fractionation of monomeric aluminium from polymeric forms is accomplished by 20 sec. complexation with 8-hydroxyquinoline at pH 8.3 with subsequent extraction into MIBK organic phase • Organic bound monomeric aluminium is separated from inorganic aluminium (mainly labile) by trapping the latter fraction on an Amberlight IR-120 ion exchange column • The Al concentrations in the organic extracts are determined photometrically

27. Chemical analytical fractionation; Shortcomings • The discrimination inherent in the method can be more or less selective but it is not absolute • Small variations in the methodical cutoff may cause significant variations in the output • Operationally defined Small variations in the cutoff will often give large variations in the results

28. Summary Sample prep. Sampling Chemical SpeciationTechniques Sample prep. Fractionation Techniques • Separation • HPLC • GC • Liquid phase • Extraction • Ion exchange resins • Solid phase (soil) • Sequential extraction • Leaching Element specific detection Techniques • GFAAS • ICP-AES • ICP-MS

29. Water sampling from different compartments of the environment

30. Sampling and sample preparation • Soil • Sample genetic horizons • Drying • Storage • Sieving (2 mm) • Grinding/Homogenizing • Soil water • Samples at different depth (5-40cm) • Conservation • Filtration (through 0.45µm filter)

31. Problem associated with sampling , storage and sample preparation for speciation/fractionation • The procedure should not disturb the chemical equilibrium between the different forms of the element that exist in a given matrix • Species transformations stimulated by change in: • Temperature, light, pH (in case of liquid samples)

32. Water sampling equipment • Deposition • Bulk precipitation • Wet only • Canopy throughfall • Ground vegetation throughfall • Soil water • Percolation lysimeter • Suction lysimeter • Runnoff • V-notch weir

33. The Vansjø Problem • In 1964 the level of nutrients in the water course was at average level with an evenly distributed algae growth • Ten years later the water course was starting to become eutrophic with increased algae growth and an almost doubling of the levels of nutrients as well as blossoming of bluegreen alga. • Bluegreen alga produce toxins • A number of remedial actions have not given the required effect.

34. Watershed • 698 km2 • Forest and agricultural land • 8 communes • Below the marine limit • Drinking water source for 60.000 people

35. Sampling strategy • Spatial and temporal variation • Worst case or representative • Regional variation: Regional or Hotspots • Temporal variation: Climate (e.g. runoff intensity)

36. Temporal variation • Runoff concentrations of both solutes and suspended material show large variations both over time and as a function of discharge. • Water sampling routines should be utilised which best can handle these variations. Discharge and nitrate concentration measured continuously in runoff water from a small agricultural catchment in Norway Deelstra et al., 1998, Sampling technique and strategy. In: Measuring runoff and nutrient losses from agricultural land in Nordic countries. TemaNord, Nordic Council of Ministers, 1998:575

37. Point sampling strategies • Different water sampling strategies depending on the objectives of the measurements. • Study processes • e.g. event studies • Study of total loss • Study of chemical and/or biological conditions

38. Point sampling strategies • Point sampling with variable time interval • Rainfall - and snow melt events, which often lead to high soil and nutrient losses, influence to a high degree the sampling frequency. • Calculations of soil and nutrient losses based on this method are biased. • Point sampling with fixed time intervals • The accuracy of the result is much influenced by the sampling frequency • Volume proportional point sampling • Point sampling is triggered each time a certain volume of water has passed the monitoring station. In general, load estimates based on this system leads to an improvement • Flow proportional composite water sampling • An alternative to point sampling systems is volume proportional mixed water samples. In this case a small water sample is taken, each time a preset volume of water has passed the monitoring station • Combined sampling • Sampling systems might be combined so as best to suit its purpose. • It is assumed that the chemical concentration of runoff water during low flow periods can be considered constant

39. Integrated monitoring