Purification of low‐concentration phenazine‐1‐carboxylic acid from fermentation broth of <i>Pseudomonas</i> sp. M18 via free flow electrophoresis with gratis gravity

Shao, Jing; Fan, Liu-Yin; Zhang, Wei; Guo, Chaowan; Li, Si; Xu, Yuquan; Cao, Cheng-Xi

doi:10.1002/elps.201000213

Cited by 16 publications

(27 citation statements)

References 26 publications

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“…The use of hydrostatic herein is distinctly dissimilar to that in the previous works on the LS-FFE [40][41][42]. In the QC-FFE, the hydrostatic-induced pressure difference is formed between the liquid level of the background buffer reservoir and that of outlets positioned at the 16-channel pump (Fig.…”

Section: Hydrostatic-induced Uniform Flow In Qc-ffementioning

confidence: 80%

“…Evidently, the size of developed QC-FFE device was slightly higher than the size of C-FFE (20-40 mm Â 20-56 mm Â 20-60 mm, the corresponding volume 20-134 mL) [39], but greatly less than that of LS-FFE (e.g. 400 mm Â 182 mm Â (0.4-1.2) mm 5 56.2-87.4 mL [40][41][42]). …”

Section: Adjustable Volume Of Qc-ffe Chambermentioning

confidence: 99%

“…Figure 1 shows the simple disassembly of QC-FFE chamber. It was the modification of the previous LS-FFE chamber [40][41][42], The original LS-FFE device has been described in detail in [40][41][42]. The main modifications were that the size of separation chamber was 45 mm Â 30 mm Â (80-500) mm.…”

Section: Fractionation Of Reassemblable and Rewashable Qc-ffementioning

confidence: 99%

“…In the QC-FFE, the hydrostatic-induced pressure difference is formed between the liquid level of the background buffer reservoir and that of outlets positioned at the 16-channel pump (Fig. 3A), whereas in the LS-FFE with hydrostatic-induced uniform background flow, the pressure difference is induced between the liquid level in gas cushion injector and that in sample collecting tubes [40][41][42].…”

Section: Hydrostatic-induced Uniform Flow In Qc-ffementioning

confidence: 99%

“…It has 16 outlets for fractionation, hydrostatic-induced uniform background buffer and sample flows, and 108-654 mL chamber higher than that of C-FFE [39] but less than the one of LS-FFE [40][41][42]. The device could be well used for the micropurification of model enzyme of trypsin in crude sample.…”

Section: Introductionmentioning

confidence: 99%

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Reassemblable quasi‐chip free‐flow electrophoresis with simple heating dispersion for rapid micropreparation of trypsin in crude porcine pancreatin

et al. 2011

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An increasing number of small biosamples (e.g. proteins and enzymes) need micropreparation in lab. However, neither large-scale free-flow electrophoresis (LS-FFE) nor chip FFE (C-FFE) could fit the growing demands. Herein, a simple quasi-chip FFE (QC-FFE) was constructed. In contrast to C-FFE, the features of QC-FFE are as follows: (i) its separation chamber is reassemblable and rewashable avoiding discard of C-FFE due to blockage of solute precipitation in chamber; (ii) its chamber size is 45 mm × 30 mm × (80-500) μm (108-654 μL volume) having function of micropreparation; (iii) there are up to 16 outlets in QC-FFE bestowing fine fraction for micropurification. The QC-FFE was used for the micropurification of model enzyme of self-digestible trypsin in crude pancreatin. Under the given conditions, the purification factor of enzyme was 11.7, the specific activity reached 6236 U/mg, the run time for 19 μL sample purification was 45 s and the throughput of trypsin was 3.34 mg/h, and the yield of pure trypsin was 55.2%. All of the results show the feasibility of enzyme micropreparation via QC-FFE. The developed device and procedure have potential use to other micropurification of protein or peptide sample.

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Section: Hydrostatic-induced Uniform Flow In Qc-ffementioning

confidence: 80%

Section: Adjustable Volume Of Qc-ffe Chambermentioning

confidence: 99%

Section: Fractionation Of Reassemblable and Rewashable Qc-ffementioning

confidence: 99%

Section: Hydrostatic-induced Uniform Flow In Qc-ffementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Reassemblable quasi‐chip free‐flow electrophoresis with simple heating dispersion for rapid micropreparation of trypsin in crude porcine pancreatin

et al. 2011

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Enhancing separation of histidine from amino acids via free‐flow affinity electrophoresis with gravity‐induced uniform hydrodynamic flow

et al. 2012

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In this paper, a novel mode of free-flow affinity electrophoresis (FFAE) was developed to indirectly enhance the separation of free-flow electrophoresis (FFE). In the mode of FFAE, a Ni(II) with high electric charge density and histidine (His) is chosen as a model ligand and target solute, respectively. Through the controlling of experimental conditions (10 mM pH 6.0 Na(2)HPO(4)-NaH(2)PO(4) with 2.0 mM NiCl(2)·6H(2)O background buffer), Ni(II) can combine with His and the combination leads to the high electric charge density of affinity complex of His-Ni(II) in contrast to the low density of free His molecule. But the ligand has weak interaction with uninterested amino acids. Thus, the mobility of His existing as His-Ni(II) is greatly increased from 14.5×10(-8) m(2) V(-1) s(-1) to 30.2 × 10(-8) m(2) V(-1) s(-1), while those mobilities of uninterested amino acids are almost constant. By virtue of the mode, we developed the FFAE procedure and conducted the relevant experiments. The experiments demonstrated the following merits of the FFAE technique: (i) clear enhancement of separation between the target solute of His and uninterested amino acids; (ii) simplicity, and (iii) low cost. Furthermore, the technique was used for the continuous separation of His from its complex sample, and the purity of His was near to 100%. All of the results demonstrate the feasibility of affinity separation in FFE. The developed FFAE may be used in the separation and pretreatment of some biological molecules (e.g. peptides).

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Simply enhancing throughput of free‐flow electrophoresis via organic‐aqueous environment for purification of weak polarity solute of phenazine‐1‐carboxylic acid in fermentation of Pseudomonas sp. M18

et al. 2012

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Herein, a simple novel free-flow electrophoresis (FFE) method was developed via introduction of organic solvent into the electrolyte system, increasing the solute solubility and throughput of the sample. As a proof of concept, phenazine-1-carboxylic acid (PCA) from Pseudomonas sp. M18 was selected as a model solute for the demonstration on feasibility of novel FFE method on account of its faint solubility in aqueous circumstance. In the developed method, the organic solvent was added into not only the sample buffer to improve the solubility of the solute, but also the background buffer to construct a uniform aqueous-organic circumstance. These factors of organic solvent percentage and types as well as pH value of background buffer were investigated for the purification of PCA in the FFE device via CE. The experiments revealed that the percentage and the types of organic solvent exerted major influence on the purification of PCA. Under the optimized conditions (30 mM phosphate buffer in 60:40 (v/v) water-methanol at an apparent pH 7.0, 3.26 mL/min background flux, 10-min residence time of injected sample, and 400 V), PCA could be continuously purified from its impurities. The flux of sample injection was 10.05 μL/min, and the recovery was up to 93.7%. An 11.9-fold improvement of throughput was found with a carrier buffer containing 40% (v/v) methanol, compared with the pure aqueous phase. The developed procedure is of evident significance for the purification of weak polarity solute via FFE.

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Purification of low‐concentration phenazine‐1‐carboxylic acid from fermentation broth of Pseudomonas sp. M18 via free flow electrophoresis with gratis gravity

Cited by 16 publications

References 26 publications

Reassemblable quasi‐chip free‐flow electrophoresis with simple heating dispersion for rapid micropreparation of trypsin in crude porcine pancreatin

Reassemblable quasi‐chip free‐flow electrophoresis with simple heating dispersion for rapid micropreparation of trypsin in crude porcine pancreatin

Enhancing separation of histidine from amino acids via free‐flow affinity electrophoresis with gravity‐induced uniform hydrodynamic flow

Simply enhancing throughput of free‐flow electrophoresis via organic‐aqueous environment for purification of weak polarity solute of phenazine‐1‐carboxylic acid in fermentation of Pseudomonas sp. M18

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