## 3 Discussion  
WG was developed to solve the problems of cultivation and the depletion of natural resources. To extend the utilization of the newly developed _Glycyrrhiza_ variety, WG, several biological activities have been elucidated. In this study, the anti-inflammatory effect of WG on DSS-induced ulcerative colitis was studied. UC is a chronic and complex autoimmune inflammatory disorder, also called chronic IBD, and is characterized by an uncontrolled inflammatory condition of the intestinal mucosa [27, 28, 29, 30]. Symptoms of UC can include abdominal pain, blood diarrhea, and weight loss [31]. In particular, the continuous progression of UC increases the risk of colorectal cancer [32]. Using 5-ASA, also known as mesalazine, yields immunosuppressive effects such as prostaglandin restriction and pro-inflammatory cytokine inhibition. The development of complementary treatments is needed for the effective mitigation of UC symptoms for safe long-term use due to the side effects of sulfasalazine [33, 34]. Traditionally, GR is used for the treatment of allergies, bowel disease, and coughs [35]. GR possesses various bioactive constituents, including flavonoids, coumarins, and saponins [36]. Thus, we hypothesized that WG could also have a beneficial effect on UC. As shown in Figure 2, the DSS control group showed significantly increased body weight loss and decreased colon length in comparison with the control group. Previous studies have shown that the length change of the colon and the severity of ulcerative colitis are correlated [37]. In this study, we found that WG and GR effectively inhibited DSS-induced colon length, DAI, and weight loss--symptoms and pathological changes of colitis in our DSS model (Figure 3 a, b). Macrophages and neutrophils penetrate the colon during UC and secrete large amounts of inflammatory cytokines, such as IL-6, IL-1\(\beta\), and TNF-\(\alpha\)[38, 39]. In IBD, immune cells such as macrophages and T lymphocytes secrete inflammatory cytokines [21, 40]. These activated cells regulate the balance between increased TNF-\(\alpha\) and IL-6 pro-inflammatory cytokine expression in the rectal mucosa of UC patients [41]. In this study, WG and GR suppressed the DSS-induced increase in TNF-\(\alpha\) and IL-6 levels in mouse serum (Figure 4). However, even with significant inhibition, the rates were variable by cytokine. In a previous study, GR extract inhibited IL-6, IL-1\(\beta\), and TNF-\(\alpha\) in a different manner. IL-6 level was strongly suppressed by a treatment of 100 \(\mu\)g/mL in RAW 264.7 cells [42]. However, the TNF-\(\alpha\)  
Figure 7: Stool sample microbiota from mice with DSS-induced ulcerative colitis, supplemented with or without WG and GR. (**a**) The relative abundance of bacteria in the stool was determined using terminal restriction fragment length polymorphism (T-RFLP) analysis. Data are shown as a percentage of the total identified sequences per group. (**b**) Effect of WG and GR on the ratio of operational taxonomic units (OTUs) before and after intervention with DSS, WG, GR, and 5-ASA. level showed less inhibition at same dose. This variability could mean that WG and GR selectively affect the production of cytokines based on dosing. It is likely that the anti-inflammatory activities of WG and GR may be caused by glycyrrhetic acid, a metabolite of glycyrrhizin [43], as glycyrrhetic acid showed anti-inflammatory activity via the inhibition of nuclear factor \(\kappa\)B(NF-\(\kappa\)B) activation. As this mechanism has previously been reported, NF-\(\kappa\)B was not analyzed in this study. However, NF-\(\kappa\)B's involvement in the anti-inflammatory activities of WG and GR is presumed, as glycyrrhizin was detected in relatively high quantities in both WG and GR extracts. In the inflammatory response, cyclooxygenase(COX)-1 protein activity rarely changes; however, COX-2 production increases rapidly, leading to increased production of prostaglandin(PG)s [44], with especially high levels of PGE\({}_{2}\) in the bowel of IBD patients [22].