Protein Extraction from Chlorella pyrenoidosa Using Bacillus spp. Isolated from Jeotgal: Strain isolation, Characterization, and Fermentation

Abstract

Chlorella pyrenoidosa, a versatile microalga with a rich nutritional profile and functional components, has various applications. However, rigid cell walls pose challenges for the effective extraction of proteins. Microbial fermentation is a promising solution for large-scale production and industrial applications. This study aimed to isolate Bacillus spp. with high enzymatic activity from Jeotgal, a Korean traditional fermented food, and enhance protein extraction from C. pyrenoidosa using microbial fermentation with the isolated Bacillus spp. Twenty-two strains of Bacillus spp. were isolated, and eight Bacillus species were selected based on their ability to produce cellulases, proteases, and lipases. Microbial safety was further assessed by testing for biogenic amine production and hemolytic activity. All eight strains exhibited γ-hemolysis, with four strains not producing biogenic amines. Notably, fermentation using Bacillus amyloliquefaciens F2 showed the highest protein extraction yield at 35.45 ± 1.21% (v/v). In conclusion, this study demonstrates the potential of microbial fermentation for protein extraction from C. pyrenoidosa, offering a novel approach for its utilization in the food industry.

Keywords: Bacillus spp. fermentation; Chlorella pyrenoidosa; Jeotgal; protein extraction.

Fig. 1. Phylogenetic tree of Bacillus spp. isolated from Jeotgal products.

Evolutionary analyses were conducted using the MEGA11 software, based on 16S rRNA sequences using the maximum likelihood method. Bootstrap values (1,000 re-sampling) are shown at the branch point. The final dataset comprised 1,076 positions.

Fig. 2. Analysis of three enzymatic activities of isolated Bacillus species.

(A) Cellulase activity, (B) protease activity, (C) lipase activity. Enzyme activity was calculated as the clearance zone size (mm) divided by the colony size (mm). Values with different letters vary significantly (p < 0.05).