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. 2021 Sep 10;9(9):1927. doi: 10.3390/microorganisms9091927

Figure 3.

Figure 3

Purification of the anti-Vibrio bacteriocin produced by B. methylotrophicus NTBD1. (A) Anti-Vibrio activity (left) and anti-Listeria activity (right) of the bacterial cultures obtained by different cultivation times. Wells (3)–(48) reflect cultures collected after 3–48 h, respectively. (B) Purification of bacteriocin crude extract by hydrophobic-interaction chromatography. Anti-Vibrio activity of crude extract (S), crude extract after loading through column (GT), the column-wash with 2% methanol (0.1% TFA) (W), column-elutes with 30% (30), 50% (50), 70% (70), 70% isopropanol (0.1% TFA) (I), and 100% of methanol (0.1% TFA) (100), respectively. Each step was conducted with three column volumes (code (1–3) follows each solvent concentration), with each solvent used as a negative control (–). The results indicated that the column eluted with 70% isopropanol (I) showed anti-Vibrio activity. (C) Purification of this active fraction (I) by reverse-phase HPLC resulted in a bacteriocin peak at a retention time of 25.6 min, monitored only with a UV wavelength of 220 nm.