Semi-preparative scale purification of enterococcal bacteriocin enterocin EJ97, and evaluation of substrates for its production |
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Authors: | Rosario Lucas López Ma Teresa García Hikmate Abriouel Nabil Ben Omar Ma José Grande Magdalena Martínez-Cañamero Antonio Gálvez |
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Institution: | (1) área de Microbiología, Facultad de Ciencias Experimentales, Universidad de Jaén, 23071 Jaén, Spain |
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Abstract: | The influence of substrate composition on the production of enterocin EJ97 and the conditions for semi-preparative bacteriocin
recovery have been studied. Final bacteriocin concentrations of 12.5 or 15.6 mg/l were obtained in the commercial media brain
heart infusion broth (BHI) and tryptic soya broth, respectively. The bacteriocin was also produced in the complex medium CM
(8.75 mg/l), in which the vitamin supplement was essential for production. Some combinations of meat peptone and yeast extract
plus either soy peptone or BHI also supported bacteriocin production, at concentrations of 6.25–7.5 mg/l. In cow milk (whole,
half-skimmed, and skimmed), the final bacteriocin concentrations obtained ranged from 7.5 to 11.25 mg/l. Highest bacteriocin
activity was obtained by using pasteurised milk whey as growth substrate (up to 25 mg/l), suggesting that this bacteriocin
can be obtained on a large scale by using this cheap food-grade industrial by-product. Highest bacteriocin titres were always
obtained after 8 h of incubation at 37 °C. Semi-preparative concentration and purification of enterocin EJ97 produced in a
complex medium was achieved by bulk cation exchange chromatography without previous cell separation, followed by reversed-phase
chromatography. This two-step procedure allowed preparation of milligram quantities of purified bacteriocin, which is an improvement
compared to purification procedures established for most other bacteriocins (35). The availability of purified enterocin EJ97
will facilitate other studies such as the elucidation of its molecular structure and its interaction with target bacteria. |
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Keywords: | Bacteriocins Biopreservation Fermentation |
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