Aluminon: its limited application as a reagent for the detection of aluminum species

The triammonium salt of aurin tricarboxylic acid, commonly referred to as aluminon, forms a dye that has been used for the colorimetric determination of Al(III) species. We have reviewed the pertinent literature on the reaction of aluminon with respect to the metallic species that form colored aluminon complexes. The effects of experimental variables, such as time, temperature, and pH, upon the color development of the aluminon complex are also presented. Organic and inorganic species, particularly Be(II) and Fe(III), which can affect color formation, are described. The use of aluminon as a histochemical staining agent for the detection of aluminum requires verification by atomic absorption spectrophotometric analysis or other quantitative techniques.     

Use of a Technovit 7200 VLC to Facilitate Integrated Determination of Aluminum by Light and Electron Microscopy

Technovit 7200 VLC is an excellent embedding medium for both inorganic histochemistry by light microscopy and X-ray microanalysis by scanning and transmission electron microscopy. Liver samples from rats after intraperitoneal treatment with aluminum chloride were fixed in glutaraldehyde and embedded in the resin. Thick sections were easily cut on an ultramicrotome and stained with aluminon for aluminum (Al). An intense positive reaction with aluminon was observed in the Kupffer cells by light microscopy. The surface structures of the same resin block cut for light microscopy were observed under a scanning electron microscope fitted with an energy dispersive X-ray spectrometer. The Kupffer cells appeared white in the backscattered mode. Localization of Al in the Kupffer cells was confirmed by an X-ray distribution map in the scanning electron microscope. Subcellular localization of Al in the Kupffer cells was performed on the same semithin sections using a transmission electron microscope equipped with an energy dispersive X-ray spectrometer. Most Al was found in lysosomes of the Kupffer cells. The resin was stable in the electron beam and chlorine-free.     

Characterization and Separation of Some of the Coordination Compounds of Chromium and Aluminon

Chromic nitrate and aluminon (ATA) react to form highly colored lakes. Examination of these lakes by the Vosburgh and Cooper method has led to the identification of a pigment [(ATA)Cr(H₂O)_x] which is insoluble in water and absolute alcohol, a deep red anionic component [(ATA)_BCr(H₂O)_x]^-y, and a purplish-red cationic component [(ATA)Cr₄(H₂O)_x]^+y. These components of the lake have been isolated and separated in the dry form. The anionic compound is apparently a simple coordination compound and can be used as a cytoplasmic stain while the cationic component is a chelate and can be used as a rather selective nuclear stain. In addition to these components, a number of other components were also found. Not only the ratio between chromium and aluminon but also the concentration of the reactants influences the formation of these different components. The amount of pigment formed is maximal with a molar ratio of 1 and a concentration of 10^-2 M or 10^-1 M. The anionic component is maximal with a molar ratio of 1 Cr to 6 aluminon at the 10^-1 M concentration, the cationic component is maximal with a molar ratio of 4 chromium to 1 aluminon at the 10^-1 M concentration. None of these are formed at the 10^-4 M concentration but only a deep redpurple soluble compound at the 1 : 1 ratio, which was not further investigated.     

Localization of aluminium in the leaves of some aluminium-accumulating species

Summary Transverse sections of leaves of some aluminium-accumulating and nonaccumulating species of the cerrado vegetation of central Brazil were coloured using aluminon to identify the tissues where aluminium occurs or is deposited. None of the tissues of the nonaccumulating species showed evidence of high concentrations of Al. All of the aluminium accumulating species showed high concentrations of Al in all of the elements of the phloem of the midrib and the secondary veins and total absence of it in the vessel members, xylem fibres and the palisade parenchyma. Walls and contents of the collenchyma of the midrib, epidermal cells, guard cells of the stomata and spongy parenchyma showed evidence of high concentrations of Al in the accumulating species.     

Laser Microprobe Mass Analysis (LAMMA) to Verify the Aluminon Staining of Bone

Triammonium aurin tricarboxylate (aluminon) has been used to localize aluminum in 2 μm sections of undecalcified, methyl methacrylate embedded bone obtained from patients with terminal chronic renal failure. Aluminum appeared in four cases as bright red lines at the mineralized-bone boundary. In two cases, however, purplish lines were found and one patient showed red as well as purplish lines. Laser microprobe mass analysis (LAMMA) identified aluminum at the location of the red lines and both aluminum and iron at the purplish lines. Furthermore, both iron and aluminum were found in histiocytic bone marrow cells, which showed brownish aluminon staining. It appears that when aluminum and iron occur together, aluminon staining may yield aberrant results. This study shows that LAMMA can be used for the identification of elements sought by histochemical methods and thus permits the evaluation of their staining effects.     

普通荞麦资源的耐铝性研究

利用小容器溶液培养法对耐铝性鉴定条件和52份普通荞麦栽培品种资源的耐铝性进行了研究。结果发现普通荞麦耐铝性鉴定的适宜条件为发芽种子于500μmol/LAlCl3溶液(pH4.5)处理3d,以发芽种子在这三天内的根伸长量衡量耐铝性程度。在该处理条件下,普通荞麦不同品种间的耐铝性有显著差异。其中,陕西大红花甜荞品种、日本大粒荞、织金红花甜荞的耐铝毒胁迫能力最强,值得在荞麦耐铝性育种和耐铝机制研究中利用。     

Aluminium reactions with polygalacturonate and related organic ligands

Aluminium (Al), in inorganic monomeric forms, has been recognised as a limiting factor for root growth in many acid soils. Plant tolerance to Al may be achieved by the detoxification (complexation) of Al by organic ligands present in the rhizosphere. The Al-complexing ability of seven organic ligands, citric, oxalic, gluconic, glucuronic, mucic, galacturonic and polygalacturonic (pectin) acids, was investigated. The proportion of organically-complexed Al was determined using colorimetric methods based on differences in reaction rate with pyrocatechol violet or aluminon. The colorimetric methods confirmed that citric acid forms a strong complex with Al at pH 4.2. In contrast, pectin and related organic ligands weakly complexed Al in acidic conditions. In an additional study, the Al-binding ability of pectin and Ca-pectate was compared at a biologically significant concentration of 32 µM Al. Only 29% of free Al remained in solution in the presence of Ca-pectate, while 54% remained when pectin was present. This suggests that Ca-pectate, rather than pectin, is responsible for binding Al in root cell walls and consequently plays an important role in Al toxicity to plants. Root growth of mungbean (Vigna radiata (L.) Wilczek) confirmed differences in the ability of citrate, oxalate and galacturonate to complex Al.     

Aluminium hydroxide uptake in the stomach and in the intestine of the rat: a histochemical study.

Using the histochemical stains aluminon, solochrome azurine and solochrome cyanine, intracellular binding of aluminium was examined in the mucosa of the stomach, duodenum, jejunum and ileum of adult rats. A first group of rats (n = 42) was sacrificed 1, 3, 6, 12, 24, 48 and 96 h after a single (300 mg x kg-1) oral administration of aluminium hydroxide. A second group of animals (n = 30) received daily the same dose of Al(OH)3 and was euthanatized after 3, 4, 5, 6 and 7 days of treatment. Aluminium deposits occurred only in the antral glands of the stomach and in rats treated for at least 3 days. The reactive deposits are located in the cytoplasm of the upper glandular cells and in the lumen of the antral glands. These results suggest that aluminium is absorbed through the antral mucosa and may be re-excreted through the glandular mucus flow into the digestive lumen where it will be absorbed again. We hypothesize that the metal could act as a delayed-effect drug.     

Silicon amelioration of aluminium toxicity in teosinte (Zea mays L. ssp. mexicana)

The influence of different Al concentrations, (0, 60 and 120 M Al) on growth and internal concentrations of Al, Si and selected organic acids was analysed in plants of teosinte (Zea mays L. ssp. mexicana), a wild form of maize from acid soils from Mexico. The plants were grown in nutrient solutions (pH 4.0) with or without 4 M silicon. Analysis with the GEOCHEM speciation program did not reveal differences between free activities of Al³⁺ in solutions with and without 4 M Si, but solutions with Si yielded lower concentrations of monomeric Al species, [Al]_mono, when analysed by a modified aluminon method. Plants grown on solutions with similar [Al]_mono, but differing in silicon, showed highly significant differences in growth and tissue concentrations of Al and organic acids. Silicon prevented growth inhibition at [Al]_mono concentrations as high as 35 M, while plants grown without Si suffered severe growth reductions with 33 M [Al]_mono. In solutions with similar [Al]_mono concentrations plants with Si had lower tissue Al concentrations and higher concentrations of malic acid than plants without Si. In view of both the significant influence of Si on the response of plants to Al toxicity and the fact that some soluble Si is always present in soil solutions, the addition of low Si concentrations to nutrient solutions used for Al-tolerance screening is recommended.     

Responses of meristematic callus cells of two Cynodon dactylon genotypes to aluminium

Responses to Al3+ of embryogenic callus cells of an Al-sensitive (Al-S) and Al-resistant (Al-R) Cynodon dactylon genotype were evaluated with regard to Al3+ toxicity and resistance. A chemical equilibrium speciation model (MINTEQA2) was used to ensure the availability of the Al3+ ion in culture media, which was supplied as 0.08-2.3 mM Al3+ for 2-8 weeks. Increasing Al3+ concentration and exposure time had a greater negative impact on the Al-S than on the Al-R genotype, in terms of callus growth rate and frequency of non-embryogenic cells. Exposure to 0.8 mM Al3+ for 2 weeks resulted in an 88% reduction in the Al-S meristematic cell number, whereas that of the Al-R genotype remained unaffected. In addition, the Al-S cells accumulated three times more Al in the nucleus than did the Al-R cells, suggesting that Al interfered with mitosis. The Al-R cells appeared to exclude Al3+ from its cells through an increase in extracellular pH (4.34 in Al-R and 4.08 in Al-S) and by the immobilisation of Al in the cell wall (33% more in Al-R). The results showed that by studying the cellular responses to Al3+ it is possible to discriminate between the Al-S and Al-R C. dactylon genotypes.     

Laser microprobe mass analysis (LAMMA) to verify the aluminon staining of bone

Triammonium aurin tricarboxylate (aluminon) has been used to localize aluminum in 2 micron sections of undecalcified, methyl methacrylate embedded bone obtained from patients with terminal chronic renal failure. Aluminum appeared in four cases as bright red lines at the mineralized-bone boundary. In two cases, however, purplish lines were found and one patient showed red as well as purplish lines. Laser microprobe mass analysis (LAMMA) identified aluminum at the location of the red lines and both aluminum and iron at the purplish lines. Furthermore, both iron and aluminum were found in histiocytic bone marrow cells, which showed brownish aluminon staining. It appears that when aluminum and iron occur together, aluminon staining may yield aberrant results. This study shows that LAMMA can be used for the identification of elements sought by histochemical methods and thus permits the evaluation of their staining effects.     

Comparison of water-soluble and exchangeable forms of Al in acid forest soils

Soil acidification promotes Al release from minerals and parent bedrocks; it also affects Al mobilization and speciation. Speciation of KCl extractable and water-extractable Al in forest soils was done by means of HPLC/IC method. Species Al3+ were the most abundant Al forms in the KCl extracts (around 93%). Prevailing Al forms (more than 70%) in aqueous extracts were Al(X)1+, [i.e., Al(OH)2+, Al(SO4)+, AlF2+, Al(oxalate)+, Al(H-citrate)+, etc.] species. It is assumed that most of KCl and water-extractable Al is bound in soil sorption complex (i.e., highly dispersed colloidal fraction of the soil solid phase creating negative charge) where majority of Al exists in the form of Al3+ species. The ECEC values, total carbon content and parameters related to soil organic matter composition (N and S content) have apparent effect on Al speciation. The most toxic Al3+ species are more concentrated in the B horizons compared to the A and E horizons. Aqueous extracts simulate Al release to soil solution under normal conditions; it can thus exhibit the actual Al toxicity. On the other hand, KCl extraction describes a potential threat for case of strong disturbance of natural soil conditions.     

Adsorption of Al3+ to phosphatidylcholine vesicles

Aluminum toxicity to soil and aquatic organisms is widespread, but the mechanisms of toxicity are unknown. To understand these mechanisms, it is important to know how aluminum reacts with cell surfaces. In this report, we studied adsorption of Al3+ to liposomes composed of phosphatidylcholine, the most abundant phospholipid in plasma membranes of eukaryotic cells. Our equilibrium dialysis and electrophoresis experiments both showed that Al3+ has a 560-fold higher affinity for the phosphatidylcholine surface than Ca2+. Unlike previous reports for adsorption of divalent metals, adsorption of Al3+ to phosphatidylcholine was predicted only approximately by the Stern model. Adsorption of AlF2+ and AlF2+ to the surface was not detectable at the activities we used. From our data, we calculate that Al3+ at 5 x 10(-6) activity could neutralize the surface charge on plant cell plasma membranes and cause a surface potential shift from -30 to +11 mV. This is consistent with non-specific Al3+ inhibition of cation uptake by root cells. Al3+ adsorption to phosphatidylcholine may also play a role in aluminum uptake into cytoplasm by endocytosis.     

Sorption of Aluminum to Plasma Membrane Vesicles Isolated from Roots of Scout 66 and Atlas 66 Cultivars of Wheat

To further elucidate the mechanisms of differential genotypic tolerance to Al, plasma membrane (PM) vesicles were isolated from whole roots, root tips, and tipless roots of Al3+-sensitive and Al3+-tolerant cultivars (cv) of wheat (Triticum aestivum L. cv Scout 66 and cv Atlas 66, respectively). Vesicles from cv Scout root tips sorbed more Al than vesicles prepared from any other source. The intrinsic surface-charge density of vesicles isolated from cv Scout was 26% more negative than vesicles from cv Atlas (-37.2 versus -29.5 millicoulombs m-2). Growth experiments indicated that cv Scout is slightly more sensitive to La3+ than is cv Atlas, that the cultivars are equally sensitive to H+, and that cv Atlas is slightly more sensitive to SeO42-. The difference in sensitivity to Al3+ was very large; for a 50% inhibition, a 16-fold greater activity of Al3+ was required for cv Atlas. Using a newly developed Gouy-Chapman-Stern model for ion sorption to the PM together with growth-response curves, we estimate that the difference in surface-charge density can account for the slightly greater sensitivity of cv Scout to cationic toxicants and the slightly greater sensitivity of cv Atlas to anionic toxicants. According to our estimates the differences in PM surface negativity and Al sorptive capacity probably account for some of the difference in sensitivity to Al3+, but the greater part of the difference probably arises from other tolerance mechanisms expressed in cv Atlas root tips that reduce the amount of Al3+ that can reach the PM.     

铝、镉对小麦幼苗生长的影响及其DNA损伤效应研究

本研究旨在探索Al^3＋、Cd^2＋对小麦幼苗生长的影响及其DNA损伤效应。结果表明：（1）所试浓度Al^3＋、Cd^2＋不仅不同程度抑制2d龄和12d龄幼苗根系和地上部分的生长,而且表现明显的DNA损伤效应;（2）除Al^3＋对幼苗地上部分鲜重、干重的影响及Cd^2＋对幼苗根系生长速率的影响外,其余生长指标显示2d龄幼苗比12d龄幼苗对Al^3＋、Cd^2＋更敏感,这与导致2d龄幼苗DNA损伤的Al^3＋、Cd^2＋浓度明显低于12d龄幼苗的结果一致。研究结果表明,DNA损伤可能是Al^3＋、Cd^2＋抑制2d龄和12d龄小麦幼苗生长的重要原因之一。     

Cd2+、Al3+对蚕豆(Vicia faba)DNA合成及修复的影响

利用＾３Ｈ－ＴｄＲ掺入方法,研究了不同浓度单金属离子Ｃｄ＾２＋、Ａｌ＾３＋对蚕豆ＤＮＡ合成、ＤＮＡ修复（以ＵＤＳ为指标）的影响。结果表明：在低浓度Ｃｄ＾２＾＋、Ａｌ＾３＋（Ｃｄ＾２＋浓度〈２００ｍｇ／ｌ,Ａｌ＾３＋浓度１００ｍｇ／ｌ）处理后,蚕豆ＤＮＡ合成加快,并且不同程度地诱导了ＵＤＳ的发生;但在高于此浓度的Ｃｄ＾２＋、Ａｌ＾３＋作用下,蚕豆ＤＮＡ合成受抑制,浓度越高,抑制作用超强;并且几乎不表     

Complexes of DNA with trivalent metal ions in presence of manganese ions

The interaction of DNA with Fe3+, Al3+, Co(NH3)6(3+) in a solution containing MnCl2 was studied. It was shown that there exists a competition for the binding sites between Mn2+ and Al3+, while the binding of Mn2+ to DNA does not depend on the presence of Fe3+ and Co(NH3)6(3+) in solution. We proposed that Fe3+ and Co(NH3)6(3+) ions prefer to bind to phosphates, and Al3+ ions are capable to bind to the nitrogen bases of DNA.     

Effect of Al on the phytosiderophore-mediated solubilization of Fe and uptake of Fe-phytosiderophore complex in wheat (Triticum aestivum)

The effect of Aluminum (Al) on phytosiderophore-mediated solubilization of insoluble Fe and the uptake of phytosiderophore-Fe³⁺ complex was examined in wheat ( Triticum aestivum L. cv. Atlas 66). Al addition did not affect the Fe solubilization by 2'-deoxymugineic acid (DMA), although Cu addition significantly inhibited the solubilization capacity. Addition of ten times more Al than Fe to the solution of DMA-Fe³⁺ complex did not decrease the absorption of the DMA-Fe³⁺ complex at 375 nm. Furthermore, NMR study indicated that Al did not shift the proton chemical shifts of DMA. All these results suggest that Al could not form a complex with the phytosiderophore, and is thereby unlikely to affect the process of phytosiderophore-mediated solubilization of Fe. Exposure of root to Al up to 100 μ M for 3 h did not inhibit the DMA-Fe³⁺ uptake by the roots, but longer pretreatment (>6 h) inhibited the uptake of the DMA-Fe³⁺ by more than 50%. Neither the uptake of DMA-Fe³⁺ nor root elongation was inhibited by 24 h pretreatment with 10 μ M Al, but both uptake and root elongation were inhibited by higher Al (>20 μ M ) pretreatment. These results suggest that Al did not directly block the transport of the phytosiderophore-Fe³⁺ complex, and that the decreased uptake of the phytosiderophore-Fe³⁺ complex resulted from the roots being damaged by Al.     

Colorimetric determination of aluminum in soil extracts using haematoxylin

Summary A method for the determination of small amounts of aluminum using haematoxylin has been developed for use with soil extracts. The effects of initial and final pH, time, variations in the amounts of haematoxylin and acetate buffer and interference of ions on colour development were studied. As compared with the standard aluminon method, this method is six times more sensitive. Moreover, aluminum in the soil extracts of complex forming reagents such as fluoride, oxalate, citrate or EDTA can be determined. The recovery of added aluminum in the fluoride extract of the soil was complete.     

植物铝毒害及遗传育种研究进展

本文简单概述了目前植物铝毒害及遗传育种方面的研究进展。Al3+可以通过与细胞骨架的作用,影响根的正常生理功能和形态建成。 植物可以通过根尖分泌有机酸或磷酸等将离子态的铝变成螯合态的铝,通过吸收H+提高根尖周围的pH,将Al3+变成难溶性的 Al(OH)3或磷酸铝从而解除铝毒害, 也可以通过在细胞内与Al3+形成无毒害的复合结构从而解除铝毒害。国外通过基因工程和突变体筛选已经获得了一批耐铝的植物材料,国内一些研究者通过突变体筛选也获得了一些耐铝的植物材料。 对植物耐铝性的遗传研究表明, 植物的耐铝性既可以是受单基因控制的,也可以是受多基因控制的。