There are existing guidelines for sampling and storage with the aim of total element determinations (Versieck & Cornelis, 1989;
Cornelis et al., 1996). Additional information about species sam- pling procedures is given in Brereton et al. (2003), Emons (2003), and De Cremer (2003). The main issue is to keep the elemental species unaffected by the procedure both in composition and in concentration. This remark can be simply put as: avoid contamina- tion and keep the species unchanged. Of the two threats, contam- ination hazards may be the easier to master. Suppose we are interested in methylmercury in serum, urine, or hair. In this case, any fortuitous contamination with inorganic mercury goes unnoticed, as only methylmercury will be specifically isolated from the matrix, and there is no danger that methylmercury will be formed during the procedure. The same reasoning is valid when analysing organolead compounds. Although PbII contaminations are ubiquitous, those by organolead species are not that widespread. There still remains the
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possibility of fortuitous binding of trace element impurities with ligands in the sample. In principle, good analytical practice under well controlled, clean conditions can avoid such problems.
As mentioned previously, the integrity of the elemental species throughout the analysis is highly dependent upon the nature of the species. A first, general guideline is to store biological specimens at below 7 °C when it is only for a few days, but to deep-freeze them for longer periods. This may be insufficient. It is important to be aware that elemental species in powders may also suffer from lack of stability due to residual humidity. This was documented in the case of arsenic species in rice powder, kept at +20 °C, + 4 °C, and í
the rice contained AsV, AsIII, MMA, and DMA. It was observed that MMA demethylated completely to AsIII and also that all of the AsV was gradually reduced to AsIII at the end of a 2-month period. The freezing temperature did little to preserve MMA and AsV in their original form. The arsenic content in rice originates from the water in which it is grown. It is thought that grinding the rice and storing it with about 18% humidity may have led to this unwelcome conversion of species, possibly induced by anaerobic activity. The arsenic species in the rice powder standard reference material (SRM) with very low residual humidity, issued by the National Institute of Standards and Technology (NIST SRM 1568a), was found to be stable.
Long-term freezing of samples is generally acceptable, although some exceptions have been reported. Arsenobetaine in sample extracts stored at 4 °C for 9 months was found to decompose to trimethylarsine oxide and two other species (Le et al., 1994). Deep- freezing samples will generally minimize any bacterial or enzyme degradation or loss from volatility. Poorly sealed sample containers let in oxygen, so that the species are oxidized, or loss of species occurs if the compounds are volatile. Bacterial degradation of the sample should be avoided. Bacteria can convert inorganic arsenic to methylated forms, and steps should be taken to preserve the original samples. Sample cleanup from a biological or complicated matrix can present problems. Ultimately, a stability study using samples spiked with known arsenic species is necessary to validate a sample storage and treatment procedure (B’Hymer & Caruso, 2004).
20 °C. The results are shown in Table 4. At the start of the study,
Table 4. Stability study of rice powder sample with 18% residual humidity during 2 months of storage at room temperature, 4 ºC, and í20 ºCa
X SD (μg/kg) Arsenic
species T (ºC) t = 0 Month 1 Month 2
AsIII 20
4
í20 46.99 0.75
88.3 4.9 81.9 7.4 82.2 7.4
94.9 2.2 92.9 2.3 93.2 1.9 DMA 20
4
í20 28.33 1.11
29.7 3.6 27.0 1.6 26.0 2.3
27.5 2.5 24.9 0.8 23.9 0.6 MMA 20
4
í20 18.10 1.7
nd nd nd
nd nd nd
AsV 20
4
í20 24.45 1.09
10.7 3.9 10.5 4.7 11.0 5.7
nd nd nd nd = not detected; SD = standard deviation
a Adapted from Brereton et al. (2003).
It appears that for biological samples, long-term preservation of species can be guaranteed only when they are kept in the dark and at very low temperatures. To prevent microbiological activity over many years, the specimens should be kept at below í130 °C. The other approach is to remove all the residual water, when temper- atures of only í20 °C may be acceptable. Anyway, it is absolutely necessary to do a stability study for each individual elemental species in its particular matrix (Emons, 2003).
An interesting study was published on the stability during stor- age of arsenic, selenium, antimony, and tellurium species in, among others, urine and fish. The species studied were AsIII, AsV, arsenobetaine, MMA, DMA, phenylarsonic acid, SeIV, SeVI, selenomethionine, SbIII, SbV, and TeVI (Lindemann et al., 2000).
Best storage conditions for aqueous mixtures of these species were achieved at 3 °C; at í20 °C, species transformation, especially of selenomethionine and SbV, took place, and a new selenium species appeared within a period of 30 days.
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Special attention is needed for sampling techniques and storage of airborne metal species in the workplace (Dabek-Zlotorzynska &
Keppel-Jones, 2003). The filter media must be chosen carefully.
General criteria that must be considered when selecting them are 1) representative sampling for particulates of 0.3 μm and greater in size, 2) low hygroscopicity, since hygroscopicity exceeding 1 mg per piece leads to serious errors in mass concentration measure- ments, and hence to the improper estimate of the environmental con- centration, and 3) absence of impurities that might interfere with the analysis. As an example of the latter, glass fibre or Teflon filters were found to be unsuitable for the sampling of airborne dust with low platinum content (Alt et al., 1993). Only polycarbonate and cellulose gave blank values as low as 5 pg of platinum per total filter.
Moreover, filters should be mechanically and thermally stable and should not interact with the deposit, even when subject to a strong extraction solvent (Dabek-Zlotorzynska & Keppel-Jones, 2003). This is particularly relevant in the case of the analysis of CrIII/CrVI in air particulate matter. Spini et al. (1994) reported the reduction of CrVI to CrIII when cellulose filters were extracted with an alkaline solution containing a known amount of CrVI. The same results were obtained by an acid (sulfuric acid) dissolution of the filters, which can be explained by cellulose’s well documented reducing properties. Therefore, cellulose filters cannot be used for chromium speciation in airborne particulates. Polycarbonate mem- brane filters (Scancar & Milaüiü, 2002) and borosilicate microfibre glass discs (Christensen et al., 1999) are suitable for this type of analysis.
Sample integrity during storage of particulate matter is another important issue. Some changes can be anticipated — for example, reduction of CrVI due to interaction not only with the collection substrate during sample storage but also with the air. Erroneous results may occur due to redox reactions. The enrichment of parti- cles on the filter gives rise to enhanced contact of the chromium species with gaseous species (e.g. sulfur dioxide, nitrogen oxides, oxygen, ozone) and/or with material collected on the filter (e.g.FeII [iron oxide, or magnetite] and AsIII-containing components) (Dabek- Zlotorzynska & Keppel-Jones, 2003). Such changes may be mini- mized by storing the samples in closed polypropylene vessels under
an inert atmosphere (nitrogen or argon) (Dyg et al., 1994;
Christensen et al., 1999). The shorter the time between collection and analysis, the better. Anyway, the reverse — namely, the oxidation of CrIII to CrVI — is most unlikely under usual conditions of storage and sample treatment.
The rule of thumb is that when no data are available from reliable studies by other research groups, the effect of sampling and storage conditions on the stability of the species in the matrix should be studied. Many species are thermodynamically unstable. The simple act of sampling and storing the species may alter them. The information is then irreversibly lost.