Yeast Fermentation Broth Research Articles

Recovery of 2,3-Butanediol by Vacuum Membrane Distillation∗

Abstract A vacuum membrane distillation process using a microporous polytetrafluoroethylene (PTFE) membrane was developed to concentrate 2,3-butanediol from model 2,3-butanediol solutions and fermentation broths. A 0.22-μm PTFE membrane passed water vapor while retaining 2,3-butanediol. Water flux through the membrane was 4 kg/m2·h at 35°C and 12–14 kg/m2·h at 55°C. Medium components, especially yeast extract and fermentation broth, reduced flux as compared to model solutions. Using this process, butanediol was concentrated from 40 g/L to about 650 g/L in the model solution, and when integrated to a fed-batch reactor, 2,3-butanediol was concentrated to over 430 g/L.

Covalent immobilization of glucose oxidase on silanized platinum microelectrode for the monitoring of glucose

The use of glucose oxidase (GOD) in electrochemical sensors for glucose detection is reported. The aim of this study was to show the feasibility and the performances of enzymatic platinum electrodes prepared by direct covalent coupling of GOD. The glucose oxidase enzyme electrode is used in a batch and flow injection analysis (FIA) system. It has been demonstrated that oxidized Pt can be silanized with surface densities comparable to those of silica; the grafting densities do not depend on the preparation method of Pt oxide, but on the nature of the monofunctional silane used. The best sensitivities for glucose detection are obtained using the coupling of 1,2,4,5-benzene tetracarboxylic acid dianhydride (BTCAD) and succinic anhydride with the aminosilanized electrodes. This coupling procedure leads to a dynamic range of response up to 30 mM in batch conditions because of no limitation by diffusion. In comparison, the BSA crosslinking technique results in a much higher sensitivity (about 200-fold) but also in the dynamic range limited to 2 mM. The sensitivity after a two-month storage remains 50% of its initial level for BTCAD coupling. The sensitivity can be amplified by performing an additional coupling process using the CDI reagent on a BTCAD-coupled enzyme electrode, whereas the opposite effect is obtained with GA. Nevertheless, the succinic anhydride-coupled electrode exhibits high non-specific signals in yeast fermentation broth samples because of the presence of electroactive interferents.

Purification and characterization of the trefoil peptide human spasmolytic polypeptide (hSP) produced in yeast

Recombinant human spasmolytic polypeptide (r-hSP) has been produced in relatively large amounts in Saccharomyces cerevisiae. The two intronless trefoil domains of the hSP-DNA were cloned separately by PCR from human genomic DNA, and the remaining parts of the gene synthezised. Recombinant plasmids were constructed to encode a fusion protein consisting of a hybrid leader sequence and the hSP sequence. The leader sequence serves to direct the fusion protein into the secretory pathway of the cell and to expose it to the Kex 2 processing enzyme system. The secreted r-hSP was found in a glycosylated and an non-glycosylated form. The two forms of r-hSP were purified from the yeast fermentation broth by a combination of ion-exchange chromatography and preparative HPLC. The overall yield from 8 litres of fermentation broth was 160 mg r-hSP and 219 mg glycosylated r-hSP corresponding to 50% and 34%, respectively. The structure of the r-hSP and the glycosylated r-hSP was determined by amino acid analysis and carbohydrate composition analysis as well as by peptide mapping, amino acid sequencing and mass spectrometric analysis.

HPLC quantification of rDNA polypeptides like insulin precursors produced in yeast

A reversed-phase HPLC system for the quantification of recombinant DNA insulin precursors in yeast fermentation broth is described. HPLC monitoring of the polypeptide can be achieved without interference from substrate components or proteins stemming from the host cell, thus serving to determine the optimum harvesting time. Furthermore, it is demonstrated how different amino acid links of the insulin A- and B-chains affect the chromatographic behaviour of the insulin precursor making it possible to identify each precursor by its retention on the reversed phase support.