Antibody production in Chinese hamster ovary (CHO) cell culture was enhanced by supplementing the culture medium with barley shochu distillation by-product supernatant (BX2). To predict antibody production following BX2 addition, fed-batch culture experiments were conducted under varying BX2 conditions using a response surface methodology. BX2 supplementation was predicted to improve antibody production by 138 %, 146 %, 120 %, and 240 % in IgG-producing CHO-MK1, CHO-MK2, CHO-DG44, and Fc-fusion protein-producing CHO-DG44 cells, respectively, compared to controls without BX2. BX2 consisted primarily of 27 % (w/w) bound amino acids, 19 % (w/w) free amino acids, 15 % (w/w) organic acids, 14 % (w/w) glycerol, 14 % (w/w) bound sugars, 6 % (w/w) ash, and 5 % (w/w) free sugars within the soluble solids. BX2 was roughly fractionated into free components (e.g., sugars, amino acids, organic acids, and glycerol fraction) and bound components (e.g., polysaccharides, peptides, peptide-linked sugars, and glycosylated amino acids fraction) using reverse-phase high-performance liquid chromatography. In cultures of IgG-producing CHO-MK2 cells, the addition of BX2 or its bound components enhanced IgG production by 170 % and 117 %, respectively. In contrast, the free components fraction reduced IgG production to 73 % of that observed without BX2. These results suggest that BX2 enhances antibody production through the combined action of multiple components rather than a single component. This study demonstrates that BX2 is an effective additive for enhancing antibody production in CHO cells cultured in high-performance medium typically used in industrial applications.
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