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SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS | Dry Ashing
S SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS Contents
Dry Ashing Oxygen Flask Combustion Wet Digestion Microwave Digestion
Dry Ashing M Hoenig, Veterinary and Agrochemical Research Centre, Tervuren, Belgium & 2005, Elsevier Ltd. All Rights Reserved.
Introduction Nowadays, trace element determinations are performed using very sensitive analytical techniques. However, with a lowering of detection limits, the risk of errors suddenly appearing due to sample handling is increased. Prior to commercial introduction of ultrasensitive instrumentation as inductively coupled plasma-mass spectrometry (ICP-MS), these ‘new’ errors were practically imperceptible to the determination of relatively high analyte concentrations that were measured with less responsive techniques. The danger of contamination is now increasingly present: the choice of sample preparation procedure, the quality of its application, and the need for an adequate laboratory environment have therefore become most critical points defining successful trace element determinations. In most cases, preparation of solid samples involves several stages: drying, homogenization, and/or grinding, followed by mineralization and dissolution of a subsample. The solution so obtained is ultimately diluted to volume. Ideally, the organic fraction of the sample has been decomposed and completely eliminated during these preparation steps and only dissolved inorganic compounds constitute the dissolved residue to be analyzed.
Before the analysis, samples of organic or of a mixed nature are subject to two distinct steps, which often take place simultaneously: mineralization and dissolution. Samples of purely inorganic composition are simply dissolved. The composition of biological and environmental samples varies from purely inorganic to purely organic, but, generally, they are an intermediate combination of these extremes. This implies that the total dissolution of samples usually cannot be achieved in a single step using a single reagent. In practice, the necessary number of steps and reagents is dictated by the matrix composition. Purely organic or mixed samples are usually brought into solution by some type of oxidation process combined with an acid dissolution of the resulting residue, as well as of the initial inorganic part of the matrix. Already in 1844, Fresenius and Von Babo had developed a method for the destruction of animal tissues prior to elemental analysis. In the intervening years, many procedures were described for this purpose. However, despite numerous possible variations, almost all of the methods fall into one of two main classes, i.e., dry ashing and wet digestion. Dry ashing methods are especially appropriate for samples having high organic matter content. The first step of the method ensures the decomposition of organic matter by heating the sample to a relatively high temperature, with atmospheric oxygen serving as the oxidation agent. Chemical compounds (the socalled ashing aids) may sometimes be added to help this process. The second step of a dry ashing method is the subsequent solubilization of the resultant ash using an appropriate acid or a mixture of acids. Depending on the sample type, the dissolution procedure generally involves several steps. Here, the terminology is precise: the term ‘mineralization’
132 SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing
relates to samples having a totally or partly organic matrix only (animal and plant tissues, food samples, soils, etc.). Prior to the analysis, any organic compounds present must be decomposed and/or completely eliminated by the mineralization procedure. Using various reagents, the organic matter is decomposed into carbon dioxide, nitrogen oxides, and water, thus liberating into solution all elements initially associated with it. After the mineralization procedure, the resulting sample residue should be essentially inorganic: it will be subject to a final dissolution step similar to that used for a sample having an initially total inorganic composition (rocks, metals, etc.). For more complex samples (organic plus inorganic composition: soils, sludge, plant samples, etc.), chemical reagents and physical means are most often used to ensure these two roles (mineralization and dissolution) are simultaneously achieved. The objective of the sample preparation stage is usually to bring all available means into play in order to determine as readily as possible the elements of interest. First, these means have to ensure the transformation and simplification of the matrix (mineralization: wet digestion, dry ashing). Second, they should convert the sample to a form compatible with the measurement technique utilized (generally a dissolution). Dry ashing methods compete with the following techniques for sample dissolution of organic and biological species for elemental analysis: * * * *
wet ashing in open or closed vessels, oxygen flask techniques, combustion tube techniques, and microwave digestion techniques in open or sealed vessels.
General Principles After the appearance, at the end of the 1970s, of commercial advertisings praising the universality and absolute necessity of wet digestion microwave heating devices for trace element analysis, several scientific papers have radically condemned dry ashing procedures, despite their long record of usefulness. In contrast, many respected institutions as well as numerous laboratories carry on the use of classical dry ashing in practical analyses of a number of materials of biological origin. Our own extensive experience in the field of sample preparation has shown that, better than the other known mineralization procedures, dry ashing methods ensure the quantitative decomposition and elimination of organic matter. Usually, these procedures
are performed by calcination at atmospheric pressure in programmable muffle furnaces. The commonly utilized temperature for this step is B4501C. In addition to conventionally heated muffle furnaces generally employed for dry ashing purposes, the market now also provides microwave furnaces especially adapted to attain elevated temperatures. The unique advantage of the latter is the capacity to ensure application of very fast heating ramps. However, this property is not directly interesting for usual dry ashing procedures, where precisely slow heating ramps are needed. Additionally, a low-temperature ashing (LTA) procedure in electronically excited oxygen plasma exists, very desirable for sample preparation when volatile elements are to be determined. The instrumentation is, unfortunately, very expensive and not readily available at present. In addition, LTA is a particularly time-consuming procedure. In the usual high-temperature ashing, fresh or dried (generally 103–1051C) samples are weighed into suitable ashing vessels (vitreous silica, porcelain, platinum) and placed in the furnace. The temperature is then progressively elevated, following a convenient heating program, to attain 4501C, and then maintained for several hours. The resulting inorganic residue (ash) is dissolved using an appropriate acid. The solution is diluted to a known volume and analyzed. Depending on the initial sample condition, results are expressed based on a freshor dry-weight basis. The application of dry ashing methods is simple and large series of samples may be treated at the same time. This is not their unique advantage – compared with wet digestions, dry ashing procedures present several other interesting characteristics: 1. Possibility of treating large sample amounts and dissolving the resulting ash in a small volume of acid. This permits preconcentration of trace elements in the final solution, which is useful when very low analyte concentrations are to be determined. Such an advantage is not realizable with wet digestion methods. Additionally, heterogeneity is a typical property of many biological materials. The possibility of processing larger masses of sample, which, upon mineralization, provides a homogeneous solution, helps to minimize subsampling errors. 2. The resulting ash is completely free of organic matter. This is a prerequisite for ensuring accuracy with some analytical techniques (e.g., ICP-MS or electrochemical methods) wherein analyte response may be influenced by the presence of residual carbon or some undigested organic molecules. The resulting solutions are of very acceptable aspect
SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing 133
(clear, colorless, and odorless), rarely the case when wet digestion methods are used and residual carbon content often attains elevated values. This is systematically observed not only for both conventionally heated and open microwave digestions, but also for closed microwave-assisted digestions. With highpressure bombs, the residual carbon content may be lower but it is never quantitatively eliminated. 3. Reagent volumes and their handling are reduced compared to wet digestion methods. 4. The acidity of final solutions can efficiently be controlled: the acid is added directly to the ash and only a small fraction is consummated during its dissolution. With wet digestion procedures, added acids must also ensure the destruction of the organic matter and their effective amounts utilized during these chemical reactions vary quite significantly. This results in unknown acid concentrations in the final solutions to be analyzed. This fact is at variance with well-known requirements for all methods based on atomic spectroscopy concerning the need for similarity of acidities between standards and samples. In some situations, this similarity is absolutely obligatory, e.g., for the determination of nickel by ICP-MS. The commonly used nickel cones in the ICP-MS interface usually produce relatively high nickel backgrounds due to their finite dissolution by the aerosol being introduced. With variable acid concentrations, the background can vary significantly from one sample to another, resulting in erratic nickel results. Such unfavorable conditions are avoided using dry ashing methods that ensure a practically constant acid concentration from sample to sample, allowing more consistent ICP-MS determinations of nickel to be made. Despite these several advantages, one must also accept several drawbacks to dry ashing procedures: 1. The chemistry of charring process is very complex and the actual temperature in the sample remains unknown; in some cases, it may be several hundred degrees above that of the furnace. This may result in volatilization losses and then to poor recoveries of some elements. 2. Under particular experimental conditions and for some types of samples, retention phenomena can occur, resulting also in incomplete recoveries. 3. Dangers of contaminations by the ambient air are greater than in wet digestion procedures performed in closed systems.
Chemistry of the Method
Results of studies concerning the dynamics and chemistry of organic matter degradation revealed
the strong exothermic character of decomposition during charring but also a possible variability, depending on the specific type of biological material. The former findings call for sufficient moderation of the charring step (ramp heating) in order to prevent local overheating of the sample and subsequent risk of loss of a fraction of the analyte due to its mechanical removal in the form of solid particles of aerosol (smoke). Only under such controlled conditions can classical dry ashing have the potential to yield accurate results. The term ‘dry oxidation’ usually characterizes procedures wherein organic matter is oxidized by reaction with gaseous oxygen, generally with the supply of energy in some form. Included in this general term are methods in which the sample is heated to a relatively high temperature in an open vessel (conventional dry ashing), or in a stream of oxygen or air. In addition, related low temperature techniques employing excited oxygen, bomb methods using oxygen under pressure, and the classical oxygen flask technique in which the sample is ignited in a closed system must also be included. All these methods involve two main processes: they provoke evaporation of the moisture and of volatile materials and ensure the progressive oxidation of the nonvolatile residue until all organic matter is destroyed. Although these processes occur in all dry oxidations, it is not always possible to distinguish them as separate events. They are probably most easily separated in the conventional ashing procedure in which the organic material is heated in an open vessel with free access to air or oxygen. In usual analytical practice, the first steps of such a procedure are usually conducted at a temperature much lower than that used to complete the oxidation. This is largely to prevent the ignition of the volatile and inflammable material produced by the process of destructive distillation and partial oxidation, as this would lead to an uncontrolled rise in the temperature resulting in an increased danger of analyte losses.
Volatility Losses The long experience has repeatedly shown that the most severe element losses are systematically observed during the heating ramp, not as a consequence of too high final ashing temperature. An inadequate heating ramp may provoke the autoignition of the sample and the resulting rapid temperature increase results in volatilization losses. The most often utilized means of avoiding ignition problems is direct insertion of the sample into the cold muffle furnace, followed by heating with an appropriately slow ramp.
134 SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing
Before the last stage of the process – progressive oxidation of the nonvolatile residue – the material remaining after the preliminary treatment is a more or less porous mass of charred organic matter containing variable amounts of inorganic material distributed throughout it. In reality, this picture is highly variable and it will depend not only on the type and the composition of samples analyzed, but also on the action of possible reagents added that can change the initial chemistry of the process. Consequently, the kinetics of oxidation of such material will be dependant on the nature of the material itself, the inorganic substances it contains, and its particle size and porosity. Tentative findings derived from such reactions with pure carbon or graphite can only be applied with caution to the complex chars existing in dry ashing of real samples. The temperatures generally recommended for dry ashing are apparently low compared with those reported for the oxidation of graphite, but the chars produced are probably far more reactive due to unknown catalytic effects of the inorganic constituents present. In dry ashing procedures used for the analysis of environmental or biological samples (animal and plant tissues, food samples, blood, milk, etc.), the final temperature is maintained for several hours. If the oxidation is achieved under optimal conditions, it leads to white or light gray colored ashes, easily soluble in acids. Sometimes, depending on the sample type, the oxidation of organic matter is not completely achieved; in this case, the ash exhibits darker spots (dark gray to black) attributable to insufficiently oxidized carbon. Because this phenomenon is always responsible for a difficult subsequent dissolution (often resulting in incomplete recoveries for several elements), such a residue must be re-treated using a few drops of nitric acid and briefly recalcinated at the usual ashing temperature. After this treatment, ashes generally become clear and easily soluble. During the oxidation process, the analyte(s) will behave in one or more of a number of ways. Ideally, they will quantitatively remain in the residue (ash) arising from the oxidation, and in a form in which they can be readily recovered, generally by a simple dissolution of the ash in an appropriate acid. Fortunately, for the usefulness of the method, this is the case for most analytes and samples. In some cases, a part (or the total) of the analyte may be converted to a volatile form that may escape from the vessel (i.e., volatilization losses) or may be combined with the vessel surface or with some components of the inorganic residue remaining after oxidation (i.e., retention losses). In practical trace element analysis, the most often reported volatilization losses pertain mainly to mercury, arsenic, and selenium.
Retention Losses Retention losses result in poor recoveries of one or more analytes using the normal procedure for solubilization of the ash. They are generally observed for a particular quality of ashing vessel or in the presence of silicates or other insoluble compounds in the sample matrix. During intercomparison studies involving analyses of plant matrices, significant discrepancies amongst results are often observed between laboratories using simple mineralization procedures and those that apply procedures that include a hydrofluoric acid attack followed by evaporation to dryness. In the former, the values obtained are systematically lower because complete digestion is not achieved. In the often utilized wet digestion procedures, mixtures of various acids with hydrogen peroxide may also lead to poor recoveries due to the presence of silicate compounds in the sample or to (co)precipitation phenomena. As a consequence, Al, Fe, Cu, and Mn, in particular, are not completely recovered, depending upon the specific plant matrix, probably related to the binding of analytes with the insoluble residue. Several authors have noted similar problems with agricultural matrices such as composts, animal meats, or brewers yeast. These statements, among others, are supported by studies concerning retention losses occurring during dry ashing of plant samples. For several plants, analyses of insoluble residues have revealed that a significant fraction of elements (major, minor, trace) is retained, depending on the type of sample and the element studied. The most affected element is Al (sometimes up to 95% retained). Consequently, it is useful to use the recovery of aluminum as a marker for the procedure quality; if the Al recovery is incomplete, it may be concluded that the dissolution step was not performed under good conditions and that many other elements may be affected in a similar fashion. For an efficient dissolution of the ash of samples with silicate compounds in the matrix, such problems highlight the absolute necessity of utilizing an HF step followed by evaporation to dryness if the objective is the determination of total element content. However, this problem, typically associated with plant samples, is similar when applying a wet digestion procedure: if an insoluble residue remains, an additional HF step, followed by evaporation to dryness, must also be performed. Another example highlighting this problem is associated to ICP-MS interferences from residual silicon resulted in up to 30% positive bias in intensities from 63Cu, 65Cu, and 55Mn derived from soil and sewage sludge digests, due to spectral interferences
SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing 135
from 28Si27Al þ , 28Si35Cl þ , and 28Si37Cl þ . In such a case, the interfering Si will be easily removed by the above-mentioned HF step. Such a procedure is not always easy to achieve with most commercially available microwave heating (closed) devices and is, in any case, much more difficult to apply than with a dry ashing procedure. Finally, plants are often considered as purely organic samples with some trace elements present. It is clear that the aforementioned problem is comparable to that encountered in soil, sediment, sludge, and rock samples where silicon is typically the primary matrix element. In this case, however, all analysts are aware of the absolute necessity of dissolving the entire sample if the total analyte content has to be determined. An additional retention problem encountered with dry ashing procedures is that posed by the sequestering action of some materials produced during ignition. The binding of iron by condensed phosphates produced by the action of heat on simple phosphates, or retention of several elements on silicate compounds present in the sample are the best known examples. Nevertheless, many studies in this field often present contradictory observations, illustrating clearly the complexity of the retention problems and the need for an adequate dissolution step. On the other hand, with the exception of arsenic and selenium, these considerations indicate that several losses reported as being due to volatilization were, in reality, due to retention problems.
Methodology Dry ashing methods can be applied to mineralization of organic materials, biological tissues, and liquids, plant, and foodstuffs, sludge, etc. Well mastered, they ensure total destruction of the organic matter; the associated elements are generally transformed to carbonate or oxide forms. At present, they are generally performed using fully programmable (ramp and holding times/temperatures) muffle furnaces characterized by an efficient temperature control and reproducible thermal programs. Required intermediate evaporations to dryness are usually achieved on sand baths or on hot plates. It is mandatory to select an ashing temperature that ensures the quantitative decomposition of organic matter without partial or total loss of analytes by volatilization or by their incorporation into a residue, which is insoluble in usual reagents. The latter may result from formation of refractory oxides, from combinations with other sample constituents present, as well as from reactions with the walls of the crucible. As noted earlier, one
of the causes of losses during dry ashing procedures is the reaction of the analyte with some of the solid matter present in the system. In order for a reaction of this nature to constitute a problem, it is first necessary that it occurs to a significant extent and, second, that the product of the reaction be insoluble in the reagents generally used for dissolving the resultant ash. The solid matter available for such a reaction is generally the material of the ashing vessel and the residue from the sample itself. It is obvious then that their nature will have a considerable effect on the extent of the losses. The most common used ashing vessels are made of silica, porcelain, or platinum. Vitreous silica is a glass consisting almost entirely of SiO2, with some Na, Al, Fe, Mg, and Ti oxide impurities, whereas the glaze on porcelain ware is a more complex material containing Al, K, Ca, and Na oxides (up to 30%) in addition to silica. For both vitreous silica and porcelain, the obvious reaction is between the oxide of the analyte and the ashing vessel to produce a complex silicate, resulting in a loss. Studies with radiotracers have shown that retention of metals by reaction vessels made of vitreous silica may be very significant during dry ashing. This type of reaction clearly occurs but is dependent on many factors. Some oxides react much more readily than others and, even if silicates are formed, some will be stable to subsequent acid attack while others will readily be decomposed and so cannot be considered to cause losses. These reactions will, of course, be exacerbated if the ashing vessel is made of silica or porcelain, exhibits a marked weakening of the silicate structure, or a surface worn by extensive use. Because the extent of such reactions remains unknown, the alternative practice of using essentially inert platinum crucibles is much more reliable. This metal is virtually unaffected by any of the usual acids, including hydrofluoric, but cannot be used for aqua regia digestion procedures. Of course, the initial cost for platinum is significantly higher than for other types of ashing vessel, but its lifetime is practically unlimited.
Ashing Aids In its initial form, a dry ashing procedure cannot be considered appropriate for preparation of samples to be used for the determination of As and Se. However, many of the reported ashing methods describe the addition of some extra inorganic compounds to the sample to improve the efficiency of the procedure. These added materials are generally called ashing aids, and they serve one or both of two purposes, i.e.,
136 SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing
to facilitate the decomposition of the organic matter, or to improve the recovery of the element to be determined. The most common aid, used to purely hasten the oxidation of organic material, is nitric acid. It is generally added toward the end of the ashing process to decompose small amounts of remaining carbonaceous material. Since the ash from most biological materials contains up to several tens of percent carbonates, nitrates are formed in situ after the addition of nitric acid. Additional ashing is then, in fact, melting with nitrates and should help to remove the most resistant degradation products present in the organic matrix. This step, leading to the production of a clean ash, has to be performed with care because when appreciable amounts of organic material are still present, it can cause the ignition of the residue when it is returned to the furnace, resulting in a possible loss of material. Some substances serve as auxiliary oxidants as well as serving other purposes. These are commonly the nitrates of light metals such as magnesium, calcium, or aluminum, which decompose on heating to yield oxides of nitrogen. These auxiliary oxidants also fulfill the important function of being inactive diluents in the process. As the organic matter in a sample is progressively decomposed, the analytes are brought into closer contact with the material of the vessel and other constituents of the residue. If a reaction with them is feasible, then the increased proximity will increase the chance of its occurrence. Under these circumstances, dilution of the ash with an inert material, such as magnesium oxide, should greatly reduce the possibility of undesirable solid-state reactions, resulting in improvement of recoveries. The well-known utilization of relatively unstable magnesium nitrate as an ashing aid likely offers both the advantage of more rapid oxidation and of decreased retention losses. These oxidative-dilution agents improve recoveries without entering into any reaction with the sample itself. Another group of ashing aids achieves the same end by altering the chemical nature of some of the constituents. The best example of this is the use of sulfuric acid to convert volatile chlorides to nonvolatile sulfates; this may prevent losses of Cd, Pb, or Cu up to B7501C. Arsenic and selenium determinations can, in some cases and under particular conditions, also benefit from the advantages offered by a dry ashing procedure. The addition of ashing aids – generally MgO and/or MgNO3 – can give rise to less volatile As or Se compounds during the ashing procedure. The successful use of ashing aids is, of
course, strongly dependent on the initial form of the analyte. In any case, utilization of ashing aids is a particularly delicate step because some successful examples cannot lead to generalizations: for routine use, the procedure necessitates a serious and time-consuming validation for each type of sample analyzed. In addition, the utilization of ashing aids significantly increases the total dissolved solids content of solutions and enhances the dangers of contamination, limiting strongly the use of this approach for trace element analysis. See also: Sample Dissolution for Elemental Analysis: Oxygen Flask Combustion; Wet Digestion; Microwave Digestion.
Further Reading Anderson R (1987) Sample Pretreatment and Separation, ACOL series. Chichester: Wiley. Bock R (1979) A Handbook of Decomposition Methods in Analytical Chemistry. Glasgow: International Textbook Company. Dunneman L, Begerow J, and Bucholski A (1994) Sample preparation for trace analysis. In: Ullmann’s Encyclopedia of Industrial Chemistry. Weinheim: Verlag Chemie. Gorsuch TT (1970) Destruction of Organic Matter. Oxford: Pergamon. Hoenig M, Baeten H, Vanhentenrijk S, Vassileva E, and Quevauviller Ph (1998) Critical discussion on the need for an efficient mineralization procedure for the analysis of plant material by atomic spectrometric methods. Analytica Chimica Acta 358: 85–94. Hoenig M (2001) Preparation steps in environmental trace element analysis – facts and traps. Talanta 54: 1021– 1038. Hoenig M (2003) Dry ashing. In: Mester Z and Sturgeon R (eds.) Sample Preparation for Trace Element Analysis. Comprehensive Analytical Chemistry 41. Amsterdam: Elsevier. Kinoshita K (1988) Carbon – Electrochemical and Physicochemical Properties. New York: Wiley. Mader P, Sza´kova´ J, and Curdova´ E (1998) Combination of classical dry ashing with stripping voltammetry in trace element analysis of biological materials: Review of literature published after 1978. Talanta 43: 521–534. Sulcek Z and Povondra P (1989) Methods of Decomposition in Inorganic Analysis. Boca Raton, FL: CRC Press. Ure AM, Butler LRP, Scott RO, and Jenkins R (1997) Preparation of materials for analytical atomic spectroscopy and other related techniques In: Nomenclature, Symbol, Units and Their Usage in Spectrochemical Analysis, Part X. Spectrochimica Acta B 52: 409–420.
Dry Ashing Oxygen Flask Combustion Wet Digestion Microwave Digestion
Dry Ashing M Hoenig, Veterinary and Agrochemical Research Centre, Tervuren, Belgium & 2005, Elsevier Ltd. All Rights Reserved.
Introduction Nowadays, trace element determinations are performed using very sensitive analytical techniques. However, with a lowering of detection limits, the risk of errors suddenly appearing due to sample handling is increased. Prior to commercial introduction of ultrasensitive instrumentation as inductively coupled plasma-mass spectrometry (ICP-MS), these ‘new’ errors were practically imperceptible to the determination of relatively high analyte concentrations that were measured with less responsive techniques. The danger of contamination is now increasingly present: the choice of sample preparation procedure, the quality of its application, and the need for an adequate laboratory environment have therefore become most critical points defining successful trace element determinations. In most cases, preparation of solid samples involves several stages: drying, homogenization, and/or grinding, followed by mineralization and dissolution of a subsample. The solution so obtained is ultimately diluted to volume. Ideally, the organic fraction of the sample has been decomposed and completely eliminated during these preparation steps and only dissolved inorganic compounds constitute the dissolved residue to be analyzed.
Before the analysis, samples of organic or of a mixed nature are subject to two distinct steps, which often take place simultaneously: mineralization and dissolution. Samples of purely inorganic composition are simply dissolved. The composition of biological and environmental samples varies from purely inorganic to purely organic, but, generally, they are an intermediate combination of these extremes. This implies that the total dissolution of samples usually cannot be achieved in a single step using a single reagent. In practice, the necessary number of steps and reagents is dictated by the matrix composition. Purely organic or mixed samples are usually brought into solution by some type of oxidation process combined with an acid dissolution of the resulting residue, as well as of the initial inorganic part of the matrix. Already in 1844, Fresenius and Von Babo had developed a method for the destruction of animal tissues prior to elemental analysis. In the intervening years, many procedures were described for this purpose. However, despite numerous possible variations, almost all of the methods fall into one of two main classes, i.e., dry ashing and wet digestion. Dry ashing methods are especially appropriate for samples having high organic matter content. The first step of the method ensures the decomposition of organic matter by heating the sample to a relatively high temperature, with atmospheric oxygen serving as the oxidation agent. Chemical compounds (the socalled ashing aids) may sometimes be added to help this process. The second step of a dry ashing method is the subsequent solubilization of the resultant ash using an appropriate acid or a mixture of acids. Depending on the sample type, the dissolution procedure generally involves several steps. Here, the terminology is precise: the term ‘mineralization’
132 SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing
relates to samples having a totally or partly organic matrix only (animal and plant tissues, food samples, soils, etc.). Prior to the analysis, any organic compounds present must be decomposed and/or completely eliminated by the mineralization procedure. Using various reagents, the organic matter is decomposed into carbon dioxide, nitrogen oxides, and water, thus liberating into solution all elements initially associated with it. After the mineralization procedure, the resulting sample residue should be essentially inorganic: it will be subject to a final dissolution step similar to that used for a sample having an initially total inorganic composition (rocks, metals, etc.). For more complex samples (organic plus inorganic composition: soils, sludge, plant samples, etc.), chemical reagents and physical means are most often used to ensure these two roles (mineralization and dissolution) are simultaneously achieved. The objective of the sample preparation stage is usually to bring all available means into play in order to determine as readily as possible the elements of interest. First, these means have to ensure the transformation and simplification of the matrix (mineralization: wet digestion, dry ashing). Second, they should convert the sample to a form compatible with the measurement technique utilized (generally a dissolution). Dry ashing methods compete with the following techniques for sample dissolution of organic and biological species for elemental analysis: * * * *
wet ashing in open or closed vessels, oxygen flask techniques, combustion tube techniques, and microwave digestion techniques in open or sealed vessels.
General Principles After the appearance, at the end of the 1970s, of commercial advertisings praising the universality and absolute necessity of wet digestion microwave heating devices for trace element analysis, several scientific papers have radically condemned dry ashing procedures, despite their long record of usefulness. In contrast, many respected institutions as well as numerous laboratories carry on the use of classical dry ashing in practical analyses of a number of materials of biological origin. Our own extensive experience in the field of sample preparation has shown that, better than the other known mineralization procedures, dry ashing methods ensure the quantitative decomposition and elimination of organic matter. Usually, these procedures
are performed by calcination at atmospheric pressure in programmable muffle furnaces. The commonly utilized temperature for this step is B4501C. In addition to conventionally heated muffle furnaces generally employed for dry ashing purposes, the market now also provides microwave furnaces especially adapted to attain elevated temperatures. The unique advantage of the latter is the capacity to ensure application of very fast heating ramps. However, this property is not directly interesting for usual dry ashing procedures, where precisely slow heating ramps are needed. Additionally, a low-temperature ashing (LTA) procedure in electronically excited oxygen plasma exists, very desirable for sample preparation when volatile elements are to be determined. The instrumentation is, unfortunately, very expensive and not readily available at present. In addition, LTA is a particularly time-consuming procedure. In the usual high-temperature ashing, fresh or dried (generally 103–1051C) samples are weighed into suitable ashing vessels (vitreous silica, porcelain, platinum) and placed in the furnace. The temperature is then progressively elevated, following a convenient heating program, to attain 4501C, and then maintained for several hours. The resulting inorganic residue (ash) is dissolved using an appropriate acid. The solution is diluted to a known volume and analyzed. Depending on the initial sample condition, results are expressed based on a freshor dry-weight basis. The application of dry ashing methods is simple and large series of samples may be treated at the same time. This is not their unique advantage – compared with wet digestions, dry ashing procedures present several other interesting characteristics: 1. Possibility of treating large sample amounts and dissolving the resulting ash in a small volume of acid. This permits preconcentration of trace elements in the final solution, which is useful when very low analyte concentrations are to be determined. Such an advantage is not realizable with wet digestion methods. Additionally, heterogeneity is a typical property of many biological materials. The possibility of processing larger masses of sample, which, upon mineralization, provides a homogeneous solution, helps to minimize subsampling errors. 2. The resulting ash is completely free of organic matter. This is a prerequisite for ensuring accuracy with some analytical techniques (e.g., ICP-MS or electrochemical methods) wherein analyte response may be influenced by the presence of residual carbon or some undigested organic molecules. The resulting solutions are of very acceptable aspect
SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing 133
(clear, colorless, and odorless), rarely the case when wet digestion methods are used and residual carbon content often attains elevated values. This is systematically observed not only for both conventionally heated and open microwave digestions, but also for closed microwave-assisted digestions. With highpressure bombs, the residual carbon content may be lower but it is never quantitatively eliminated. 3. Reagent volumes and their handling are reduced compared to wet digestion methods. 4. The acidity of final solutions can efficiently be controlled: the acid is added directly to the ash and only a small fraction is consummated during its dissolution. With wet digestion procedures, added acids must also ensure the destruction of the organic matter and their effective amounts utilized during these chemical reactions vary quite significantly. This results in unknown acid concentrations in the final solutions to be analyzed. This fact is at variance with well-known requirements for all methods based on atomic spectroscopy concerning the need for similarity of acidities between standards and samples. In some situations, this similarity is absolutely obligatory, e.g., for the determination of nickel by ICP-MS. The commonly used nickel cones in the ICP-MS interface usually produce relatively high nickel backgrounds due to their finite dissolution by the aerosol being introduced. With variable acid concentrations, the background can vary significantly from one sample to another, resulting in erratic nickel results. Such unfavorable conditions are avoided using dry ashing methods that ensure a practically constant acid concentration from sample to sample, allowing more consistent ICP-MS determinations of nickel to be made. Despite these several advantages, one must also accept several drawbacks to dry ashing procedures: 1. The chemistry of charring process is very complex and the actual temperature in the sample remains unknown; in some cases, it may be several hundred degrees above that of the furnace. This may result in volatilization losses and then to poor recoveries of some elements. 2. Under particular experimental conditions and for some types of samples, retention phenomena can occur, resulting also in incomplete recoveries. 3. Dangers of contaminations by the ambient air are greater than in wet digestion procedures performed in closed systems.
Chemistry of the Method
Results of studies concerning the dynamics and chemistry of organic matter degradation revealed
the strong exothermic character of decomposition during charring but also a possible variability, depending on the specific type of biological material. The former findings call for sufficient moderation of the charring step (ramp heating) in order to prevent local overheating of the sample and subsequent risk of loss of a fraction of the analyte due to its mechanical removal in the form of solid particles of aerosol (smoke). Only under such controlled conditions can classical dry ashing have the potential to yield accurate results. The term ‘dry oxidation’ usually characterizes procedures wherein organic matter is oxidized by reaction with gaseous oxygen, generally with the supply of energy in some form. Included in this general term are methods in which the sample is heated to a relatively high temperature in an open vessel (conventional dry ashing), or in a stream of oxygen or air. In addition, related low temperature techniques employing excited oxygen, bomb methods using oxygen under pressure, and the classical oxygen flask technique in which the sample is ignited in a closed system must also be included. All these methods involve two main processes: they provoke evaporation of the moisture and of volatile materials and ensure the progressive oxidation of the nonvolatile residue until all organic matter is destroyed. Although these processes occur in all dry oxidations, it is not always possible to distinguish them as separate events. They are probably most easily separated in the conventional ashing procedure in which the organic material is heated in an open vessel with free access to air or oxygen. In usual analytical practice, the first steps of such a procedure are usually conducted at a temperature much lower than that used to complete the oxidation. This is largely to prevent the ignition of the volatile and inflammable material produced by the process of destructive distillation and partial oxidation, as this would lead to an uncontrolled rise in the temperature resulting in an increased danger of analyte losses.
Volatility Losses The long experience has repeatedly shown that the most severe element losses are systematically observed during the heating ramp, not as a consequence of too high final ashing temperature. An inadequate heating ramp may provoke the autoignition of the sample and the resulting rapid temperature increase results in volatilization losses. The most often utilized means of avoiding ignition problems is direct insertion of the sample into the cold muffle furnace, followed by heating with an appropriately slow ramp.
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Before the last stage of the process – progressive oxidation of the nonvolatile residue – the material remaining after the preliminary treatment is a more or less porous mass of charred organic matter containing variable amounts of inorganic material distributed throughout it. In reality, this picture is highly variable and it will depend not only on the type and the composition of samples analyzed, but also on the action of possible reagents added that can change the initial chemistry of the process. Consequently, the kinetics of oxidation of such material will be dependant on the nature of the material itself, the inorganic substances it contains, and its particle size and porosity. Tentative findings derived from such reactions with pure carbon or graphite can only be applied with caution to the complex chars existing in dry ashing of real samples. The temperatures generally recommended for dry ashing are apparently low compared with those reported for the oxidation of graphite, but the chars produced are probably far more reactive due to unknown catalytic effects of the inorganic constituents present. In dry ashing procedures used for the analysis of environmental or biological samples (animal and plant tissues, food samples, blood, milk, etc.), the final temperature is maintained for several hours. If the oxidation is achieved under optimal conditions, it leads to white or light gray colored ashes, easily soluble in acids. Sometimes, depending on the sample type, the oxidation of organic matter is not completely achieved; in this case, the ash exhibits darker spots (dark gray to black) attributable to insufficiently oxidized carbon. Because this phenomenon is always responsible for a difficult subsequent dissolution (often resulting in incomplete recoveries for several elements), such a residue must be re-treated using a few drops of nitric acid and briefly recalcinated at the usual ashing temperature. After this treatment, ashes generally become clear and easily soluble. During the oxidation process, the analyte(s) will behave in one or more of a number of ways. Ideally, they will quantitatively remain in the residue (ash) arising from the oxidation, and in a form in which they can be readily recovered, generally by a simple dissolution of the ash in an appropriate acid. Fortunately, for the usefulness of the method, this is the case for most analytes and samples. In some cases, a part (or the total) of the analyte may be converted to a volatile form that may escape from the vessel (i.e., volatilization losses) or may be combined with the vessel surface or with some components of the inorganic residue remaining after oxidation (i.e., retention losses). In practical trace element analysis, the most often reported volatilization losses pertain mainly to mercury, arsenic, and selenium.
Retention Losses Retention losses result in poor recoveries of one or more analytes using the normal procedure for solubilization of the ash. They are generally observed for a particular quality of ashing vessel or in the presence of silicates or other insoluble compounds in the sample matrix. During intercomparison studies involving analyses of plant matrices, significant discrepancies amongst results are often observed between laboratories using simple mineralization procedures and those that apply procedures that include a hydrofluoric acid attack followed by evaporation to dryness. In the former, the values obtained are systematically lower because complete digestion is not achieved. In the often utilized wet digestion procedures, mixtures of various acids with hydrogen peroxide may also lead to poor recoveries due to the presence of silicate compounds in the sample or to (co)precipitation phenomena. As a consequence, Al, Fe, Cu, and Mn, in particular, are not completely recovered, depending upon the specific plant matrix, probably related to the binding of analytes with the insoluble residue. Several authors have noted similar problems with agricultural matrices such as composts, animal meats, or brewers yeast. These statements, among others, are supported by studies concerning retention losses occurring during dry ashing of plant samples. For several plants, analyses of insoluble residues have revealed that a significant fraction of elements (major, minor, trace) is retained, depending on the type of sample and the element studied. The most affected element is Al (sometimes up to 95% retained). Consequently, it is useful to use the recovery of aluminum as a marker for the procedure quality; if the Al recovery is incomplete, it may be concluded that the dissolution step was not performed under good conditions and that many other elements may be affected in a similar fashion. For an efficient dissolution of the ash of samples with silicate compounds in the matrix, such problems highlight the absolute necessity of utilizing an HF step followed by evaporation to dryness if the objective is the determination of total element content. However, this problem, typically associated with plant samples, is similar when applying a wet digestion procedure: if an insoluble residue remains, an additional HF step, followed by evaporation to dryness, must also be performed. Another example highlighting this problem is associated to ICP-MS interferences from residual silicon resulted in up to 30% positive bias in intensities from 63Cu, 65Cu, and 55Mn derived from soil and sewage sludge digests, due to spectral interferences
SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing 135
from 28Si27Al þ , 28Si35Cl þ , and 28Si37Cl þ . In such a case, the interfering Si will be easily removed by the above-mentioned HF step. Such a procedure is not always easy to achieve with most commercially available microwave heating (closed) devices and is, in any case, much more difficult to apply than with a dry ashing procedure. Finally, plants are often considered as purely organic samples with some trace elements present. It is clear that the aforementioned problem is comparable to that encountered in soil, sediment, sludge, and rock samples where silicon is typically the primary matrix element. In this case, however, all analysts are aware of the absolute necessity of dissolving the entire sample if the total analyte content has to be determined. An additional retention problem encountered with dry ashing procedures is that posed by the sequestering action of some materials produced during ignition. The binding of iron by condensed phosphates produced by the action of heat on simple phosphates, or retention of several elements on silicate compounds present in the sample are the best known examples. Nevertheless, many studies in this field often present contradictory observations, illustrating clearly the complexity of the retention problems and the need for an adequate dissolution step. On the other hand, with the exception of arsenic and selenium, these considerations indicate that several losses reported as being due to volatilization were, in reality, due to retention problems.
Methodology Dry ashing methods can be applied to mineralization of organic materials, biological tissues, and liquids, plant, and foodstuffs, sludge, etc. Well mastered, they ensure total destruction of the organic matter; the associated elements are generally transformed to carbonate or oxide forms. At present, they are generally performed using fully programmable (ramp and holding times/temperatures) muffle furnaces characterized by an efficient temperature control and reproducible thermal programs. Required intermediate evaporations to dryness are usually achieved on sand baths or on hot plates. It is mandatory to select an ashing temperature that ensures the quantitative decomposition of organic matter without partial or total loss of analytes by volatilization or by their incorporation into a residue, which is insoluble in usual reagents. The latter may result from formation of refractory oxides, from combinations with other sample constituents present, as well as from reactions with the walls of the crucible. As noted earlier, one
of the causes of losses during dry ashing procedures is the reaction of the analyte with some of the solid matter present in the system. In order for a reaction of this nature to constitute a problem, it is first necessary that it occurs to a significant extent and, second, that the product of the reaction be insoluble in the reagents generally used for dissolving the resultant ash. The solid matter available for such a reaction is generally the material of the ashing vessel and the residue from the sample itself. It is obvious then that their nature will have a considerable effect on the extent of the losses. The most common used ashing vessels are made of silica, porcelain, or platinum. Vitreous silica is a glass consisting almost entirely of SiO2, with some Na, Al, Fe, Mg, and Ti oxide impurities, whereas the glaze on porcelain ware is a more complex material containing Al, K, Ca, and Na oxides (up to 30%) in addition to silica. For both vitreous silica and porcelain, the obvious reaction is between the oxide of the analyte and the ashing vessel to produce a complex silicate, resulting in a loss. Studies with radiotracers have shown that retention of metals by reaction vessels made of vitreous silica may be very significant during dry ashing. This type of reaction clearly occurs but is dependent on many factors. Some oxides react much more readily than others and, even if silicates are formed, some will be stable to subsequent acid attack while others will readily be decomposed and so cannot be considered to cause losses. These reactions will, of course, be exacerbated if the ashing vessel is made of silica or porcelain, exhibits a marked weakening of the silicate structure, or a surface worn by extensive use. Because the extent of such reactions remains unknown, the alternative practice of using essentially inert platinum crucibles is much more reliable. This metal is virtually unaffected by any of the usual acids, including hydrofluoric, but cannot be used for aqua regia digestion procedures. Of course, the initial cost for platinum is significantly higher than for other types of ashing vessel, but its lifetime is practically unlimited.
Ashing Aids In its initial form, a dry ashing procedure cannot be considered appropriate for preparation of samples to be used for the determination of As and Se. However, many of the reported ashing methods describe the addition of some extra inorganic compounds to the sample to improve the efficiency of the procedure. These added materials are generally called ashing aids, and they serve one or both of two purposes, i.e.,
136 SAMPLE DISSOLUTION FOR ELEMENTAL ANALYSIS / Dry Ashing
to facilitate the decomposition of the organic matter, or to improve the recovery of the element to be determined. The most common aid, used to purely hasten the oxidation of organic material, is nitric acid. It is generally added toward the end of the ashing process to decompose small amounts of remaining carbonaceous material. Since the ash from most biological materials contains up to several tens of percent carbonates, nitrates are formed in situ after the addition of nitric acid. Additional ashing is then, in fact, melting with nitrates and should help to remove the most resistant degradation products present in the organic matrix. This step, leading to the production of a clean ash, has to be performed with care because when appreciable amounts of organic material are still present, it can cause the ignition of the residue when it is returned to the furnace, resulting in a possible loss of material. Some substances serve as auxiliary oxidants as well as serving other purposes. These are commonly the nitrates of light metals such as magnesium, calcium, or aluminum, which decompose on heating to yield oxides of nitrogen. These auxiliary oxidants also fulfill the important function of being inactive diluents in the process. As the organic matter in a sample is progressively decomposed, the analytes are brought into closer contact with the material of the vessel and other constituents of the residue. If a reaction with them is feasible, then the increased proximity will increase the chance of its occurrence. Under these circumstances, dilution of the ash with an inert material, such as magnesium oxide, should greatly reduce the possibility of undesirable solid-state reactions, resulting in improvement of recoveries. The well-known utilization of relatively unstable magnesium nitrate as an ashing aid likely offers both the advantage of more rapid oxidation and of decreased retention losses. These oxidative-dilution agents improve recoveries without entering into any reaction with the sample itself. Another group of ashing aids achieves the same end by altering the chemical nature of some of the constituents. The best example of this is the use of sulfuric acid to convert volatile chlorides to nonvolatile sulfates; this may prevent losses of Cd, Pb, or Cu up to B7501C. Arsenic and selenium determinations can, in some cases and under particular conditions, also benefit from the advantages offered by a dry ashing procedure. The addition of ashing aids – generally MgO and/or MgNO3 – can give rise to less volatile As or Se compounds during the ashing procedure. The successful use of ashing aids is, of
course, strongly dependent on the initial form of the analyte. In any case, utilization of ashing aids is a particularly delicate step because some successful examples cannot lead to generalizations: for routine use, the procedure necessitates a serious and time-consuming validation for each type of sample analyzed. In addition, the utilization of ashing aids significantly increases the total dissolved solids content of solutions and enhances the dangers of contamination, limiting strongly the use of this approach for trace element analysis. See also: Sample Dissolution for Elemental Analysis: Oxygen Flask Combustion; Wet Digestion; Microwave Digestion.
Further Reading Anderson R (1987) Sample Pretreatment and Separation, ACOL series. Chichester: Wiley. Bock R (1979) A Handbook of Decomposition Methods in Analytical Chemistry. Glasgow: International Textbook Company. Dunneman L, Begerow J, and Bucholski A (1994) Sample preparation for trace analysis. In: Ullmann’s Encyclopedia of Industrial Chemistry. Weinheim: Verlag Chemie. Gorsuch TT (1970) Destruction of Organic Matter. Oxford: Pergamon. Hoenig M, Baeten H, Vanhentenrijk S, Vassileva E, and Quevauviller Ph (1998) Critical discussion on the need for an efficient mineralization procedure for the analysis of plant material by atomic spectrometric methods. Analytica Chimica Acta 358: 85–94. Hoenig M (2001) Preparation steps in environmental trace element analysis – facts and traps. Talanta 54: 1021– 1038. Hoenig M (2003) Dry ashing. In: Mester Z and Sturgeon R (eds.) Sample Preparation for Trace Element Analysis. Comprehensive Analytical Chemistry 41. Amsterdam: Elsevier. Kinoshita K (1988) Carbon – Electrochemical and Physicochemical Properties. New York: Wiley. Mader P, Sza´kova´ J, and Curdova´ E (1998) Combination of classical dry ashing with stripping voltammetry in trace element analysis of biological materials: Review of literature published after 1978. Talanta 43: 521–534. Sulcek Z and Povondra P (1989) Methods of Decomposition in Inorganic Analysis. Boca Raton, FL: CRC Press. Ure AM, Butler LRP, Scott RO, and Jenkins R (1997) Preparation of materials for analytical atomic spectroscopy and other related techniques In: Nomenclature, Symbol, Units and Their Usage in Spectrochemical Analysis, Part X. Spectrochimica Acta B 52: 409–420.