J. Bernhard Wehr

Alkali hydroxide-induced gelation of pectin

The mechanism of pectin gelation depends on the degree of methoxylation. High methoxyl pectin gels due to hydrophobic interactions and hydrogen bonding between pectin molecules. Low methoxyl pectin forms gels in the presence of di- and polyvalent cations which cross link and neutralise the negative charges of the pectin molecule. Monovalent cations normally do not lead to gel formation with high methoxyl pectin solutions free of divalent cations, especially Ca. The present study found that alkali (NaOH or KOH) added to high methoxyl pectin leads to gel formation in a concentration-depended manner. It was also found that monovalent cations (Na and K) induce gelation of low methoxyl pectin and the time required for gel formation (setting time) depends on the cation concentration. The results indicate that a combined char-e neutralisation and ionic strength effect is responsible for the monovalent cation-induced gelation of pectin

The rhizotoxicity of metal cations is related to their strength of binding to hard ligands

Mechanisms whereby metal cations are toxic to plant roots remain largely unknown. Aluminum, for example, has been recognized as rhizotoxic for approximately 100 yr, but there is no consensus on its mode of action. The authors contend that the primary mechanism of rhizotoxicity of many metal cations is nonspecific and that the magnitude of toxic effects is positively related to the strength with which they bind to hard ligands, especially carboxylate ligands of the cell-wall pectic matrix. Specifically, the authors propose that metal cations have a common toxic mechanism through inhibiting the controlled relaxation of the cell wall as required for elongation. Metal cations such as Al(3+) and Hg(2+), which bind strongly to hard ligands, are toxic at relatively low concentrations because they bind strongly to the walls of cells in the rhizodermis and outer cortex of the root elongation zone with little movement into the inner tissues. In contrast, metal cations such as Ca(2+), Na(+), Mn(2+), and Zn(2+) , which bind weakly to hard ligands, bind only weakly to the cell wall and move farther into the root cylinder. Only at high concentrations is their weak binding sufficient to inhibit the relaxation of the cell wall. Finally, different mechanisms would explain why certain metal cations (for example, Tl(+), Ag(+), Cs(+), and Cu(2+)) are sometimes more toxic than expected through binding to hard ligands. The data presented in the present study demonstrate the importance of strength of binding to hard ligands in influencing a range of important physiological processes within roots through nonspecific mechanisms.

Effects of Ca, Cu, Al and La on pectin gel strength: implications for plant cell walls

Rheology of Ca-pectate gels is widely studied, but the behaviour of pectate gels formed by Cu, Al and La is largely unknown. It is well known that gel strength increases with increasing Ca concentration, and it is hypothesised that this would also be the case for other cations. Pectins are a critical component of plant cell walls, imparting various physicochemical properties. Furthermore, the mechanism of metal toxicity in plants is hypothesised to be, in the short term, related to metal interactions with cell wall pectin. This study investigated the influence of Ca, Cu, Al and La ion concentrations at pH 4 on the storage modulus as a function of frequency for metal-pectin gels prepared from pectin (1%) with a degree of esterification of 30%. Gels were formed in situ over 6d in metal chloride solution adjusted daily to pH 4. Cation concentration was varied to develop a relationship between gel strength and cation concentration. At similar levels of cation saturation, gel strength increased in the order of La

Metal ion effects on hydraulic conductivity of bacterial cellulose-pectin composites used as plant cell wall analogs

Low concentrations of some trace metals markedly reduce root elongation rate and cause ruptures to root rhizodermal and outer cortical cells in the elongation zone. The interactions between the trace metals and plant components responsible for these effects are not well understood but may be linked to changes in water uptake, cell turgor and cell wall extensibility. An experiment was conducted to investigate the effects of Al, La, Cu, Gd, Sc and Ru on the saturated hydraulic conductivity of bacterial cellulose (BC)-pectin composites, used as plant cell wall analogs. Hydraulic conductivity was reduced to approximately 30% of the initial flow rate by 39 microM Al and 0.6 microM Cu, approximately 40% by 4.6 microM La, 3 microM Sc and 4.4 microM Ru and approximately 55% by 3.4 microM Gd. Scanning electron microscopy (SEM) revealed changes in the ultrastructure of the composites. The results suggest that trace metal binding decreases the hydraulic conductivity through changes in pectin porosity. The experiment illustrates the importance of metal interactions with pectin, and the implications of such an interaction in plant metal toxicity and in normal cell wall processes.

Hydrolysis and speciation of Al bound to pectin and plant cell wall material and its reaction with the dye chrome azurol S

Hydrolysis of aluminum (Al) in solution increases at pH >or= 4 and with an Al concentration. Pectin, an important anionic polysaccharide of plant cell walls, adsorbs Al, but this phenomenon is poorly understood. This study showed that Al(3+) hydrolysis results in binding of Al to pectin in excess of the stoichiometric equivalent, leading to oversaturation of the pectin with Al. However, the degree of pectin methyl-esterification did not affect the extent of Al hydrolysis. Binding of Al to purified cell wall material also resulted in Al hydrolysis in a pH- and soluble Al concentration-dependent manner, but the source of cell wall material had no effect at fixed pH. Staining of Al-treated pectin and cell wall material from wheat ( Triticum aestivum L.) and sunflower ( Helianthus annuus L.) with the Al-specific dye, chrome azurol S (CAS), resulted in the formation of a purple color, with the intensity related to the extent of Al hydrolysis.

Rhizotoxic effects of silver in cowpea seedlings

Silver (Ag) is highly toxic to aquatic organisms, including algae, invertebrate animals, and fish, but little information exists on Ag rhizotoxicity in higher plants. In two solution culture experiments with approximately 1,000 microM Ca(NO3)2 and 5 microM H3BO3 (pH 5.4), 20 to 80% of added Ag (< or =2 microM) was lost from solution within approximately 30 min, with a further decrease after 48 h root growth. Using measured Ag concentrations at the start of the experiments, the median effective concentration (EC50) for root elongation rate of cowpea (Vigna unguiculata [L.] Walp. cv. Caloona) was 0.010 microM Ag in the first 4 h of exposure (0.021 microM in the first 8 h). This demonstrates that Ag (as Ag+) is rapidly rhizotoxic to cowpea seedlings at concentrations similar to those that are toxic to freshwater biota. Rupturing of rhizodermal and outer cortical layers was evident after 48 h with 0.13 to 0.57 microM Ag initially in solution, being most severe at 0.13 or 0.25 microM Ag. An additional experiment showed that ruptures were first evident after 20 h exposure to 0.17 microM Ag, with increased severity of rupturing over time. The rhizotoxic effects of Ag are similar to those of some other trace metals (e.g., Cu, Al, La) that bind strongly to hard ligands and weakly to soft ligands. The similarity of rupturing effects, despite the difference in strong binding to soft ligands by Ag and to hard ligands by the other metals, suggests a distinctive metabolic effect of Ag that binds only weakly to hard ligands.

Comparison between methods using copper, lanthanum, and colorimetry for the determination of the cation exchange capacity of plant cell walls

The determination of the cation exchange capacity (CEC) of plant cell walls is important for many physiological studies. We describe the determination of cell wall CEC by cation binding, using either copper (Cu) or lanthanum (La) ions, and by colorimetry. Both cations are strongly bound by cell walls, permitting fast and reproducible determinations of the CEC of small samples. However, the dye binding methods using two cationic dyes, Methylene Blue and Toluidine Blue, overestimated the CEC several-fold. Column and centrifugation methods are proposed for CEC determination by Cu or La binding; both provide similar results. The column method involves packing plant material (2-10 mg dry mass) in a chromatography column (10 mL) and percolating with 20 bed volumes of 1 mM La or Cu solution, followed by washing with deionized water. The centrifugation method uses a suspension of plant material (1-2 mL) that is centrifuged, and the pellet is mixed three times with 10 pellet volumes of 1 mM La or Cu solution followed by centrifugation and final washing with deionized water. In both methods the amount of La or Cu bound to the material was determined by spectroscopic methods.

Comparative hydrolysis and sorption of Al and La onto plant cell wall material and pectic materials

Pectin, which is an important component of plant cell walls, strongly binds Al and this may play a role in expression of Al toxicity. Sorption of aluminium (Al) and lanthanum (La) from aqueous solutions onto pectic acid, Ca-pectate and plant cell wall material was pH dependent. For Al at pH 3, sorption was less than the available sorption sites (i.e., the cation exchange capacity) on all three sorbents, whereas at pH 4, sorption of Al was in excess of available sorption capacity. By contrast, sorption of the trivalent Al analogue La corresponded to the available sorption capacity on all three sorbents at pH 5. This indicates, therefore, that Al hydrolyses at ≥ pH 4, and hydrolysis increases with Al concentration in solution. Further, it is proposed that the sorption of Al to pectin leads to deprotonation of the galacturonic acid (GalA) residues. Sorption of Al to pectin limits hydrolysis of Al, thereby shifting the pH of hydrolysis to a higher value. Hydrolysis of Al results in its sorption in excess of the stoichiometric equivalent (assuming the free Al3+ ion), leading to oversaturation of the pectin with Al. Staining of the metal-pectate complexes with the metachromatic dye eosin showed that with increasing Al saturation (but not La saturation), the complex developed a positive net charge, due to formation of some positively charged Al-complexes. The development of a positive charge on the Al-pectate complex may have major effects on cellular transmembrane potential and nutrient acquisition by plant roots. This is the first report of Al binding in excess of binding sites and development of a net positive charge on Al-pectate.

Ferric minerals and organic matter change arsenic speciation in copper mine tailings

Arsenic (As) is commonly associated with Cu ore minerals, with the resultant risk that As can be released offsite from mine tailings. We used synchrotron-based fluorescence X-ray absorption near-edge spectroscopy (XANES) imaging to provide in situ, laterally-resolved speciation of As within tailings which differed in magnetite content (5-12%) and organic matter content (0-5%). Although the total As content was lower in tailings with low magnetite (LM), the soluble (pore water) As was actually 7-times higher in LM tailings than in high magnetite (HM) tailings. Additionally, amendment with 5% sugarcane mulch residues (SMR) (for revegetation) further increased soluble As due to the dissolution and oxidation of arsenopyrite or orpiment. Indeed, in HM tailings, arsenopyrite and orpiment initially accounted for 88% of the total As, which decreased to 48% upon the addition of SMR - this being associated with an increase in AsV-ferrihydrite from 12% to 52%. In LM tailings, the pattern of As distribution and speciation was similar, with As as AsV-ferrihydrite increasing from 57% to 75% upon the addition of SMR. These findings indicate that changes in ore processing technology, such as the recovery of magnetite could have significant environmental consequences regarding the As mobilisation and transformation in mine tailings.

Aluminium effects on mechanical properties of cell wall analogues.

Aluminium (Al) toxicity adversely impacts plant productivity in acid soils by restricting root growth and although several mechanisms are involved the physiological basis of decreased root elongation remains unclear. Understanding the primary mechanisms of Al rhizotoxicity is hindered due to the rapid effects of soluble Al on root growth and the close proximity of many cellular components within the cell wall, plasma membrane, cytosol and nucleus with which Al may react. To overcome some of these difficulties, we report on a novel method for investigating Al interactions with Komagataeibacter xylinus bacterial cellulose (BC)-pectin composites as cell wall analogues. The growth of K. xylinus in the presence of various plant cell wall polysaccharides, such as pectin, has provided a unique in vitro model system with which to investigate the interactions of Al with plant cell wall polysaccharides. The BC-pectin composites reacted in a similar way with Al as do plant cell walls, providing insights into the effects of Al on the mechanical properties of the BC-pectin composites as cell wall analogues. Our findings indicated that there were no significant effects of Al (4-160 μM) on the tensile stress, tensile strain or Youngs modulus of the composites. This finding was consistent with cellulose, not pectin, being the major load bearing component in BC-pectin composites, as is also the case in plant cell walls.