Inhibition by oxalates of spinach chloroplast phenolase in unfrozen and frozen states

Spinach chloroplast phenolase was inhibited by oxalic acid and its salts. Complete inhibitions were induced instantly in the acidic region (e.g. by 1 and 5 mM oxalate at pH 5 and 5.5, respectively), and in the neutral region pre-incubation of the enzyme with oxalates could also lead to complete loss of activity. The inhibition mode was non-competitive for phenol substrate with K_i of 0.9 mM pH 6.8. Reduction of enzyme activity in a crude extract of chloroplasts induced by freezing at neutral pH was due to the presence of ammonium oxalate. With 0.5 mM oxalate, the inhibition attained 75% under frozen conditions, whilst no inhibition could be detected in the enzyme which had not been frozen. Free oxalic acid and K⁺ and Na⁺ salts also caused freezing inhibition. Glyoxylic and oxamic acids acted as inhibitors with less efficiency. With a pure mushroom tyrosinase (phenolase), essentially the identical results were obtained using the same conditions.     

Biochemical mechanisms of stone alteration carried out by filamentous fungi living in monuments

Biochemical weathering mechanisms carried out by Penicillium frequentans and Cladosporium cladosporoides on unaltered sandstone, granite and limestone were studied using FTIR, X-ray diffraction, atomic absorption and flame photometry. Strains belonging to both fungal species, isolated from the façades of two Spanish Cathedrals, were used.Large amounts of oxalic, citric and gluconic acids were produced by P. frequentans in broth cultures. These metabolites caused extensive deterioration of clay silicates, micas and feldspars from both sandstone and granite and also of calcite and dolomite from limestone, as a result of high cation release and organic salts formation such as calcium, magnesium and ferric oxalates and calcium citrates. Comparatively, the biodegradative effect brought about by C. cladosporoides was much less than that caused by P. frequentans. Neither organic acids nor organic salts were formed by C. cladosporiodes samples.It is concluded that filamentous fungi are able to cause an extensive weathering of stone, due principally to organic acid excretion, although other metabolites participate to a lesser extent in these deteriorative processes. Ecological adaptative mechanisms, such micronutrients uptake and trivalent cations chelation (Fe³⁺ and Al³⁺) are derived from fungal growth on stone monuments.     

Species-specific Cd-stress Response in the White Rot Basidiomycetes Abortiporus biennis and Cerrena unicolor

The effect of cadmium (Cd) on fungal growth, Cd bioaccumulation and biosorption, and on the formation of potential heavy metal response indicators such as thiols, oxalate, and laccase was investigated in the white rot fungi Cerrena unicolor andAbortiporus biennis. Only the highest Cd concentration employed (200 μM) inhibited growth of C. unicolor, whereas already lower Cd concentrations caused decreasing mycelia dry weights in A. biennis. Cd biosorption onto the mycelial surface was the predominant Cd sequestration mechanism in C. unicolor. Surface-bound and bioaccumulated Cd concentrations were essentially in the same range in A. biennis, leading to considerably higher intracellular Cd concentrations in A. biennis than in C. unicolor. Oxalate and laccase were produced by both of the fungal strains and their extracellular levels were elevated upon Cd exposure. Oxalate concentrations and laccase titres were considerably higher in C. unicolor than in A. biennis. Both fungi responded to increasing Cd concentrations by increasing intracellular amounts of thiol compounds (cysteine, γ-glutamylcysteine, glutathione in both its reduced and oxidized form) but Cd application increased the amounts of thiols to a higher extend in A. biennis. Taken together, these species-specific responses towards Cd suggest that C. unicolor possesses a more efficient system than A. biennis to keep intracellular Cd concentrations low.     

SORPTION OF FERROUS AND FERRIC IRON BY EXTRACELLULAR POLYMERIC SUBSTANCES (EPS) FROM ACIDOPHILIC BACTERIA

The sorption of Fe(II) and Fe(III) by extracellular polymeric substances (EPS) of acidophilic bacteria Acidiphilium 3.2Sup(5) and Acidithiobacillus ferrooxidans, harvested from the ecosystem of the Tinto River (Huelva, Spain), was investigated. EPS from mixed cultures of both bacteria (EPS_mixed) and pure cultures of A. 3.2Sup(5) (EPS_pure) were extracted with ethylenediamine tetraacetic acid (EDTA) and were characterized by Fourier-transform infrared (FTIR), electron photoemission (XPS), x-ray diffraction (DRX), and energy dispersive x-ray (EDX) spectroscopy and scanning electron microscopy (SEM). EPS _pure were loaded, in sorption tests, with Fe(II) and Fe(III). The results obtained indicate that the biochemical composition and structure of EPS_mixed was very similar to that of EPS_pure. Besides, results indicate that EPS_mixed adsorbed Fe(II) and Fe(III) by preferential interaction with the carboxyl group, which favored the formation of Fe(II)/Fe(III) oxalates. These species were also formed in EPS_pure loaded with Fe(II)/Fe(III). All this behavior suggested that the sorption of iron by EPS_mixed was similar to sorption of EPS_pure, which fitted the Freundlich model. Thus, the iron uptake of EPS_mixed reached 516.7 ± 23.4 mg Fe/g-EPS at an initial concentration of 2.0 g/L of Fe_total and Fe(II)/Fe(III) ratio of 1.0.     

Biocorrosion and biodeterioration of antique and medieval glass

Over the past few years we have examined various antique and medieval glasses with regard to general biogenic damage, biopitting (crater erosion), bio‐crusts, and opalescent and white biogenic films. Experiments were carried out on pieces from Roman glass bottles excavated near Abu Tor, Sinai, some pieces of green and blue glass from Cologne Cathedral, some pieces from a little church in Evreux, glass samples from the fortress of the former Dukedom of Delmenhorst near Oldenburg, and some neolithic flint tools from the Negev Desert, Israel. Modern glass from a pigsty (19th century) additionally has been used for laboratory experiments on the attack of glass surfaces by fungi and bacteria. Some of the bacteria used in these experiments were isolated from the ancient pieces of glass. Biopitting with structures very similar to the biopitting of marble and limestone was found on almost all specimens. Lichens were not identified directly, but fungi and algae were observed in the pits as well as under the thin layers exfoliating from the Roman glass bottles. Initial steps of colonization and the potential for heavy‐metal accumulation by the isolated bacteria have been shown in laboratory experiments. A fractal dimension of diffusion‐limited disaggregation (DLD) is suggested as one possible explanation for the characteristic form and structure of the microbially induced and shaped biopitting patterns. A biopitting classification is suggested.     

Oxalate patinas on ancient monuments: the biological hypothesis

Summary Whewellite and weddellite, calcium oxalate monohydrate and dihydrate respectively, have been found in the form of thin surface layers on limestone and marble monuments and artifacts of various historical periods at different sites. Experimental results indicate that the formation of both minerals must be attributed essentially to the action of oxalic acid secreted by microorganisms (lichens) which live and proliferate on the stone. Oxalic acid attacks the calcium carbonate of the stone surface giving rise to the precipitation of calcium oxalate.     

Ecological approach to the genesis of calcium oxalate patinas on stone monuments

Summary The genesis of calcium oxalate patinas on stone monuments gives rise to controversial opinions. One of the proposed hypotheses links this phenomenon to the past presence of lichens on the exposed surfaces of monuments. However, the growth of a biological species cannot occur if environmental conditions are not compatible with its autoecology. Analysis of variations of the environmental factors that can act as «limiting factors» shows that in most monuments, the various exposures are not always compatible with biological growth. The environmental factor that seems to be the most limiting is the amount of surface water that is frequently below the range of tolerance of even the most xerophylous species. In the case of Trajan's column in Rome, the distribution of oxalate layers shows an opposite trend with respect to what we would expect for lichen colonization. Presently other kinds of biological colonization cannot be excluded.     

Impact of oxalic acid on rumen function and bacterial community in sheep

Oxalic acid (OA) is a secondary compound occurring in a wide range of plants consumed by ruminants, especially in saline lands or in arid and semi-arid regions. However, its impact on the rumen microbial community and its changes over time, as well as the potential consequences on ruminal function, remain unknown. To examine this impact, five ewes fitted with a ruminal cannula and fed low-quality grass hay were dosed daily with 0.6 mmol of OA/kg body weight through the cannula for 14 days. On days 0 (before the start), 4, 7 and 14 of the administration period, samples of ruminal digesta were collected throughout the day (0, 3, 6 and 9 h after the morning feeding) for analysis of the bacterial community and fermentation parameters (pH, ammonia and volatile fatty acid (VFA) concentrations). In addition, two feedstuffs were incubated in situ using the nylon bag technique to estimate ruminal degradation. Terminal restriction fragment length polymorphism was employed to monitor the dynamics of total bacteria, and quantitative real-time PCR was used to investigate the abundance of the oxalate-degrading Oxalobacter formigenes. Neither pH nor total VFA concentrations were affected. Nevertheless, OA dosing altered molar proportions of most individual VFA and ammonia concentrations (P < 0.001). The dry matter disappearance of alfalfa hay was reduced on days 7 and 14 and that of barley straw only on day 7 (P < 0.01). These slight changes were related to others observed in the relative frequency of a number of terminal restriction fragments. Variations in the ruminal microbiota occurred rapidly with OA administration, which did not modify the bacterial diversity significantly but altered the structure of the community. However, many of these changes were reversed by the end of the experiment, with no significant differences between days 0 and 14 of dosing. These results suggest a rapid adaptation of the rumen bacterial community linked to the estimated increase in the abundance of O. formigenes (from 0.002% to 0.007% of oxc gene in relation to the total bacteria 16S rDNA; P < 0.01), which is assumed to be responsible for oxalate breakdown.