# Potential Prebiotic and Anti-Obesity Effects of _Codium fragile_ Extract
## 4 Discussion  
The cultivated green algae, _C. fragile_, contains large amounts of structural sulfated polysaccharides. Sulfated galactan from _C. fragile_ consists of a large amount of galactose residues with trace arabinose and the presence of pyruvate and sulfate as substituents [41; 42]. In addition, the polysaccharide composed of \(\beta\)-D-Gal\(p\)-(1 \(\rightarrow\), \(\rightarrow\) 3)-\(\beta\)-d-Gal\(p\)-(1\(\rightarrow\) and \(\rightarrow\) 3,6)-\(\beta\)-d-Gal\(p\)-(1 \(\rightarrow\) with a small amount of \(\rightarrow\) 6)-Gal\(p\)-(1 \(\rightarrow\) and \(\rightarrow\) 3,4)-Gal\(p\)-(1 \(\rightarrow\) residues and sulfates are mainly linked at O-4 of \(\rightarrow\) 3)-Gal\(p\)-(1 \(\rightarrow\) and \(\rightarrow\) 3,6)-Gal\(p\)-(1 \(\rightarrow\) residues [15]. Partial degradation products of sulfated structural polysaccharides, produced during fermentation by the human gut microbiota, have been shown to have bioactivity [6; 16; 18; 19]. Furthermore, the gut microbiome is well known for its role in regulating obesity and other metabolic disease. CFEs extracted with water comprise many galactose residues with traces of arabinose, and the presence of sulfate as substituents. In accordance with these results, [PERSON] et al. [15] reported that sulfated polysaccharides isolated from _C. fragile_ were mainly composed of galactose, arabinose, and glucose. Therefore, our results indicate that CFEs belongs to the galactan/arabinan group (or arabinocalatan). Moreover, [PERSON] et al. [43] reported that the elemental composition and bioaccessibility of marine algae were different depending on pretreatment such as seaweed rehydration. Therefore, further research on the processing of seaweed for use as a functional material is required. To evaluate the growth effect of CFE on probiotics, CFE was used as the single carbon source. CFE was able to promote growth of all the probiotic bacterial strains used in this study, while there was no increase in growth of pathogenic bacteria. CFE enhanced the growth of _Ly_ and _Lh_ to a greater extent than GOS. These results are consistent with a previ  
Figure 3: CFE inhibits lipid accumulation in 3T3-L1 cells. (**a**) Confirmation of cell viability on adipocyte differentiation using the MTT assay. (**b**) Confirmation of the inhibitory effect on adipocyte differentiation using oil red O staining. (**c**) Confirmation of CFE-induced changes in the expression of protein related to adipocyte differentiation using Western blot analysis. Data are representative of more than three separated experiments and are presented as mean \(\pm\) SD. Significant differences are noted as * \(p<0.05\), ** \(p<0.01\)** as compared to differentiated 3T3-L1 cells without CFE treatment. MDI: methylisobutylxanthine, dexamethasone, and insulin, NAC: N-Acetyl-cysteine, **# \(p<0.05\)** as compared to normal cells. ous report that sulfated galactan can promote the proliferation of lactic acid bacteria [44]. Therefore, fermentation involving of \(Lp\) and \(Lh\) with CFE as a substrate was further studied. The fermentation of CFE with \(Lp\) and \(Lh\) resulted in an increase in acidity of the culture medium. Application of lactic acid bacteria in the fermentation of CFE showed increased acidity, which correlated with the drop in pH. These results suggest that CFE can be employed as carbon source for lactic acid bacteria and affected medium acidity and secondary metabolite production, such as SCFA. The main SCFAs produced during lactic fermentation are acetate, propionate and butyrate [45; 46]. SCFAs can also inhibit the invasion and colonization of pathogens by lowering the intestinal pH [47; 48; 49]. [PERSON] et al. [50], reported that accumulation of acids during fermentation can decrease growth rate. CFE fermented with probiotics showed the highest viable cell count with 24 h of fermentation, following which the viable cell count decreased.