Abstract:Os sideróforos provenientes de Xylella fastidiosa de videiras foram investigados. Tais metabólitos seqüestram ferro, um elemento essencial, do hospedeiro, o que os torna um potencial fator de patogenicidade. Em um meio de cultura em placa com limitação de ferro, tais sideróforos foram detectados pela reação com o complexo cromoazurol S (CAS). Diferentes métodos de análise instrumentais foram utilizados para caracterização dos sideróforos, como: cromatografia de afinidade por metal imobilizado (IMAC), cromatogr… Show more
“…As shown in Fig. 5, the purified catechol siderophore DHB-Gly-Thr (M = 312), which is the single unit of bacillibactin, could be obtained by (Hart et al 2002;Barnouin et al 2005), and hydroxamate-containing siderophores [Ni(II)-IMAC, Fe(III)-IMAC] (Braich and Codd 2008;Simionato et al 2010;Ejje et al 2013). As bacillibactin is a secondary metabolite with low yield, and only the single unit of bacillibactin DHB-Gly-Thr was separated in this study.…”
Section: Purification Results From Fe(iii)-based Imacmentioning
confidence: 86%
“…When sample was loaded, the incoming active substances were bound to the immobilized metal and separated from the rest of sample. Thus, Fe 3+ -immobilized IMAC columns may be successfully used for the extraction of siderophore from complex samples (Simionato et al 2010). Najwa Braich and Rachel Codd had already successfully captured hydroxamate-containing siderophores and other Fe(III)-binding metabolites directly from bacterial culture supernatants with nickel(II)-based IMAC (Braich and Codd 2008), but there are no records of Fe(III)-based IMAC for the separation of catechol siderophore.…”
Catechol siderophore plays an important role in microbial ecology, agriculture, and medicine, but its research is often limited by the difficulty in acquisition of it in large quantities. Based on evidence from the coordination chemistry and chemical biology, catechol siderophore could chelate Fe with high affinity. Therefore, Fe(III)-based immobilized metal-affinity chromatography (IMAC) was applied to capture siderophore from the culture filtrate of CD36. The ethanol-precipitated sample and the separated sample from Fe(III)-based IMAC were analyzed by liquid chromatography-mass spectrometry. According to the result, the pure siderophore DHB-Gly-Thr could be extracted from the ethanol-precipitated sample. Compared with other purifications, Fe(III)-based IMAC was convenient and had fewer steps. In addition, it also reduced the use of toxic chemical solvents in some traditional extraction process, such as extraction and ion exchange chromatography. Fe(III)-based IMAC was successfully used in separation of the catechol siderophore from CD36. The results revealed that Fe(III)-based IMAC was an efficient and environmentally friendly method for the separation and purification of catechol siderophore.
“…As shown in Fig. 5, the purified catechol siderophore DHB-Gly-Thr (M = 312), which is the single unit of bacillibactin, could be obtained by (Hart et al 2002;Barnouin et al 2005), and hydroxamate-containing siderophores [Ni(II)-IMAC, Fe(III)-IMAC] (Braich and Codd 2008;Simionato et al 2010;Ejje et al 2013). As bacillibactin is a secondary metabolite with low yield, and only the single unit of bacillibactin DHB-Gly-Thr was separated in this study.…”
Section: Purification Results From Fe(iii)-based Imacmentioning
confidence: 86%
“…When sample was loaded, the incoming active substances were bound to the immobilized metal and separated from the rest of sample. Thus, Fe 3+ -immobilized IMAC columns may be successfully used for the extraction of siderophore from complex samples (Simionato et al 2010). Najwa Braich and Rachel Codd had already successfully captured hydroxamate-containing siderophores and other Fe(III)-binding metabolites directly from bacterial culture supernatants with nickel(II)-based IMAC (Braich and Codd 2008), but there are no records of Fe(III)-based IMAC for the separation of catechol siderophore.…”
Catechol siderophore plays an important role in microbial ecology, agriculture, and medicine, but its research is often limited by the difficulty in acquisition of it in large quantities. Based on evidence from the coordination chemistry and chemical biology, catechol siderophore could chelate Fe with high affinity. Therefore, Fe(III)-based immobilized metal-affinity chromatography (IMAC) was applied to capture siderophore from the culture filtrate of CD36. The ethanol-precipitated sample and the separated sample from Fe(III)-based IMAC were analyzed by liquid chromatography-mass spectrometry. According to the result, the pure siderophore DHB-Gly-Thr could be extracted from the ethanol-precipitated sample. Compared with other purifications, Fe(III)-based IMAC was convenient and had fewer steps. In addition, it also reduced the use of toxic chemical solvents in some traditional extraction process, such as extraction and ion exchange chromatography. Fe(III)-based IMAC was successfully used in separation of the catechol siderophore from CD36. The results revealed that Fe(III)-based IMAC was an efficient and environmentally friendly method for the separation and purification of catechol siderophore.
“…Between the matrix and the chelating agent, a short alkyl chain (the socalled spacer arm) is grafted to ensure accessibility. The IMAC column can be purchased already filled with the matrix grafted with the complexing agent [59,[83][84][85] or the matrix can be bought separately [86][87][88][89][90]. Inversely, an initially empty column can be filled with resins [15,57,91,92].…”
Section: The Matrixmentioning
confidence: 99%
“…Metal ions are frequently loaded onto resins in their hydrochloride form [106][107][108] or as sulfates salts [15,90,91]. The metal ions loaded in excess are removed using either sodium chloride solutions, a weak complex agent such as acetate [59,109], a low concentration of a strong complexing agent (phosphate, <50 mM) [110,111], or simply with deionized water.…”
Section: Metal Ion Loading and Equilibration Of The Column With Buffe...mentioning
confidence: 99%
“…Unbound components are removed using the loading buffer or deionized water [15,88,[90][91][92]106,113]. When water is used for washing, it is followed by a second wash with a weak complexing agent such as formate or acetate [45,59,94,94,109,115].…”
Section: Removing the Unbound Componentsmentioning
Metal-Chelating Peptides (MCPs), obtained from protein hydrolysates, present various applications in the field of nutrition, pharmacy, cosmetic etc. The separation of MCPs from hydrolysates mixture is challenging, yet, techniques based on peptide-metal ion interactions such as Immobilized Metal Ion Affinity Chromatography (IMAC) seem to be efficient. However, separation processes are time consuming and expensive, therefore separation prediction using chromatography modelling and simulation should be necessary. Meanwhile, the obtention of sorption isotherm for chromatography modelling is a crucial step. Thus, Surface Plasmon Resonance (SPR), a biosensor method efficient to screen MCPs in hydrolysates and with similarities to IMAC might be a good option to acquire sorption isotherm. This review highlights IMAC experimental methodology to separate MCPs and how, IMAC chromatography can be modelled using transport dispersive model and input data obtained from SPR for peptides separation simulation.
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