Extraction of equine chorionic gonadotrophin from pregnant mare plasma by direct adsorption on chromatographic media

González, Gabriel Alcántar; Castro, Beatriz; Massaldi, Hugo

doi:10.1002/(sici)1097-0290(19980105)57:1<22::aid-bit3>3.0.co;2-x

Cited by 4 publications

(4 citation statements)

References 11 publications

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“…From the data it can be seen, that the precipitation with meta‐phosphoric acid and EtOH resulted in a partial loss of the available activity, which cannot be explained by the physical loss of the target alone. eCG is stable in a pH range of 4 to 8, but a pH of less than 3, which occurs during the precipitation step using meta‐phosphoric acid, the noncovalent linkage between the subunits is gradually decreased and a loss of sialic acid residues can occur . In case of sialic proteins, the sialic acids in the molecule maintain the hormone in circulation.…”

Section: Resultsmentioning

confidence: 99%

“…Analogous to the packed bed chromatography procedure for eCG purification from horse serum described in literature, selected anion exchange (AEX) materials in 1 mL packed bed plastic columns connected to an ÄKTApurifier 100 workstation (GE Healthcare, Uppsala, Sweden), were tested. For the purification, weak (DEAE Sepharose Fast Flow) and strong (Q Sepharose Fast Flow (Q FF), and Q Sepharose XL (Q XL)) AEX materials were chosen.…”

Section: Methodsmentioning

confidence: 99%

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Purification of equine chorionic gonadotropin (eCG) using magnetic ion exchange adsorbents in combination with high‐gradient magnetic separation

Müller

Heidenreich

Franzreb

et al. 2014

Biotechnology Progress

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Current purification of the glycoprotein equine chorionic gonadotropin (eCG) from horse serum includes consecutive precipitation steps beginning with metaphosphoric acid pH fractionation, two ethanol precipitation steps, and dialysis followed by a numerous of fixed-bed chromatography steps up to the specific activity required. A promising procedure for a more economic purification procedure represents a simplified precipitation process requiring only onethird of the solvent, followed by the usage of magnetic ion exchange adsorbents employed together with a newly designed 'rotor-stator' type High Gradient Magnetic Fishing (HGMF) system for large-scale application, currently up to 100 g of magnetic adsorbents. Initially, the separation process design was optimized for binding and elution conditions for the target protein in mL scale. Subsequently, the magnetic filter for particle separation was characterized. Based on these results, a purification process for eCG was designed consisting of (i) pretreatment of the horse serum; (ii) binding of the target protein to magnetic ion exchange adsorbents in a batch reactor; (iii) recovery of loaded functionalized adsorbents from the pretreated solution using HGMF; (iv) washing of loaded adsorbents to remove unbound proteins; (v) elution of the target protein. Finally, the complete HGMF process was automated and conducted with either multiple single-cycles or multicycle operation of four sequential cycles, using batches of pretreated serum of up to 20 L. eCG purification with yields of approximately 53% from single HGMF cycles and up to 80% from multicycle experiments were reached, with purification and concentration factors of around 2,500 and 6.7, respectively.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Purification of equine chorionic gonadotropin (eCG) using magnetic ion exchange adsorbents in combination with high‐gradient magnetic separation

Müller

Heidenreich

Franzreb

et al. 2014

Biotechnology Progress

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“…Fig. 1 eCG is a highly glycosylated protein with a total carbohydrate content of 45% (González et al 1998). Glycoproteins containing more than 10% carbohydrate bind less SDS per g than unglycosylated proteins (Moore et al 1980).…”

Section: Resultsmentioning

confidence: 99%

Simplified purification of equine chorionic gonadotropin (eCG)––an example of the use of magnetic microsorbents for the isolation of glycoproteins from serum

et al. 2011

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Simplified purification of equine chorionic gonadotropin (eCG)-an example of the use of magnetic microsorbents for the isolation of glycoproteins from serum. Biotechnology Letters, Springer Verlag, 2011, 33 (5) AbstractClassical purification of the glycoprotein equine chorionic gonadotropin (eCG) from serum includes pH fractionation with metaphosphoric acid, two ethanol precipitation steps as well as dialysis followed by fixed-bed chromatography. A simplified process requiring only 1/3 of the solvent and improving the yield from 53 to 65% has been developed. The process comprises an ultra-/diafiltration step after the first ethanol precipitation, directly followed by an 2 adsorption/desorption procedure based on magnetic microadsorbents with N,N-diethyl-ammonium functionalization. The process reaches an overall purification factor of eCG of more than 1800 and an average product activity of 1300 IU ELISA /mg. After adapting the parameters of the fractionation and the type of magnetic microadsorbents, the new concept is likely to be transferable to other serum proteins.

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“…In the finite batch adsorption experiment, bovine serum albumin (BSA) was incubated with the ion exchange and a diluted suspension of yeast cell under investigation of pH and ion conductivity as to establish a proper ratio of BSA adsorption to ion conductivity in cell suspension [11]. Prior to the experiment, the ion exchanges were equilibrated with 50mL Tris-HCl (pH 7.6).…”

Section: Adsorption Experimentsmentioning

confidence: 99%

A bioreactor with an internal contactor for primary recovery of bovine serum albumin from yeast suspension

Yoon¹,

Sim²,

Kim

et al. 2009

Korean J. Chem. Eng.

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In this paper the internal contactor is a newly developed device for the primary recovery of protein from crude feedstock. Ion exchanges (DEAE-Streamline) are confined inside the internal contactor in a stirred tank. Interactions between the ion exchange in the internal contactor and protein (BSA) in yeast suspension have been studied. For better performance, two strategies are considered: to determine the ion conductivity of a simulated yeast suspension, and to select the optimum process time for adsorption. In this system, advantages of both batch adsorption and expanded bed adsorption were obtained. Furthermore, in denser cell concentration (50 g/L) where EBA cannot be operated, the primary recovery was carried out in 1-2 hr. The efficiency of yield is higher than 80% in this condition.

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Extraction of equine chorionic gonadotrophin from pregnant mare plasma by direct adsorption on chromatographic media

Cited by 4 publications

References 11 publications

Purification of equine chorionic gonadotropin (eCG) using magnetic ion exchange adsorbents in combination with high‐gradient magnetic separation

Purification of equine chorionic gonadotropin (eCG) using magnetic ion exchange adsorbents in combination with high‐gradient magnetic separation

Simplified purification of equine chorionic gonadotropin (eCG)––an example of the use of magnetic microsorbents for the isolation of glycoproteins from serum

A bioreactor with an internal contactor for primary recovery of bovine serum albumin from yeast suspension

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