Revisiting the Chrome Azurol S Assay for Various Metal Ions

Abstract

Siderophores play an important role in the solubilisation and mobilization of iron (III) and various metal ions. To have a useful method to test siderophores in culture supernatants for their metal binding affinity, we redesigned and optimized the liquid CAS-assay for selected metal ions. CAS-assay solutions were calibrated with desferrioxamine B in different concentrations to calculate DFOB-equivalents to get a semi-quantitative evaluation. With these assay solutions, we were able to test siderophores in culture supernatants for their ability to chelate with Fe, Al, Ga, Cu, V and As.

Bacterial Metabolites Produced Under Iron Limitation Kill Pinewood Nematode and Attract Caenorhabditis elegans

“…This is the first time that siderophore production was described in more detail by a strain of the genus Rouxiella and the respective siderophore was identified as DFOE. The HPLC-enriched fraction of DFOE and related molecules (Figure 2) showed affinity to various metals and metalloids as described previously for other desferrioxamines obtained from soil bacteria (Mehnert et al, 2017; Wiche et al, 2017; Schwabe et al, 2018).…”

“…As described above both strains, A41C3 and Arv20#4.1, produced different siderophores which have different properties. As various metals and metalloids may play a role in the rhizosphere ecosystem it is of interest to verify the capability of the siderophores to bind various types of ions (Mehnert et al, 2017; Wiche et al, 2017; Schwabe et al, 2018). Therefore, we decided to choose a selection of ions which can form a CAS-complex and thus allow to show and to discriminate affinity of siderophores for Fe, Ga, Cu, V, and Al (Figure 2).…”

“…Supernatants obtained from cultures of strains Arv20#4.1 (bacterium carried by PWN from United States) and A41C3 (endophytic bacterium of P. pinaster from Portugal) were assessed for the siderophore activities by a modified CAS assay and a modified Csáky test (Esuola et al, 2016; Mehnert et al, 2017). EDTA and desferrioxamine B (DFOB) were used as positive controls.…”

“…EDTA and desferrioxamine B (DFOB) were used as positive controls. The modified CAS assay was used to determine Fe, Ga, Cu, V, and Al chelating compounds in comparison to the same control reagents as for the Csáky test (Mehnert et al, 2017). Fe-, Ga-, Al-CAS solutions were prepared as follows (100 ml): 1.5 ml of 1 mM FeCl 3 × 6H 2 O (in 10 mM HCl) or 1.5 ml of 1 mM Ga 2 (SO 4 ) 3 or 1.5 ml of 1 mM AlCl 3 × 6 H 2 O was added to 7.5 ml of 2 mM aqueous CAS solution.…”

“…Metallophores have distinct biochemical characteristics and are divided into various families, i.e., hydroxamates, catecholates, carboxylates, and mixed types (Hider and Kong, 2010; Saha et al, 2013). Different types of siderophores have different metal binding capabilities and are often not restricted to iron but can scavenge different metals like copper, zinc, vanadium, gallium, and cadmium from the environment (Mehnert et al, 2017; Wiche et al, 2017; Schwabe et al, 2018). There are several biotechnological applications of siderophores, for example, enhancing of plant growth, acting as biological control agents (BCA), to perform bioremediation of environments contaminated with metals, to recover rare earth elements, and to act as antibiotic carriers or inhibitors of metalloenzymes (Ahmed and Holmström, 2014; Kurth et al, 2016).…”