## 3 Discussion
The important role of gut microbiota in health has been gradually gaining recognition in science; although studies regarding the physiological functions of BA have mainly involved pigs as a research model for humans [22], benzoic acid administration has been found to improve gut functions including digestion and absorption, and also could improve gut barrier function, including nonspecific barrier mechanisms and specific immunological responses [22]. The lower concentrations of hippuric acid and benzoic acid found in the bladder cancer group might be associated with gut microbial pathways, decreased activity of the hippurate hydrolase, or increased activity of the glycine \(N\)-benzoyltransferase [14]. Another down-regulated metabolite was \(N\)-Acetyl-phenylalanine, which also participates in the phenylalanine metabolism pathway. Acetylphenylalanine is a product of enzyme phenylalanine \(N\)-acetyltransferase [20; 23], and lower concentrations of this compound in saliva have been previously reported in oral squamous cell carcinoma patients [23]. Histidine and carnosine also exhibited decreased levels in BC patients. The latter plays a role in histidine, beta-alanine, and nitrogen metabolism as well as in the aminoacyl-tRNA biosynthesis pathway. Histidine is an \(\alpha\)-amino acid that is used in the biosynthesis of proteins. One of the proteins significantly related to tumor progression is fragile histidine triad protein (FHIT), a protein that has been proven to have suppressor properties. The lack of FHIT protein or its reduced level is observed in many types of cancer as well as in various cancer cell lines [24; 25; 26]. Reduced concentration of histidine in bladder cancer patients may correlate with a lower activity of FHIT protein [24]. FHIT protein deficiency has been also reported in bladder cancer, mainly in high-grade tumors. This phenomenon correlates with poorer prognosis in BC patients [26; 27; 28]. Histidine is also a precursor for carnosine biosynthesis. This dipeptide links histidine and the beta-alanine metabolism pathway. While carnosine has not been reported in the literature as a metabolite of significant importance in bladder cancer, numerous studies have shown that carnosine exhibits some antioxidant effects [29; 30; 31]. Carnosine's ability to scavenge reactive oxygen species (ROS) may be important in the prevention of cancers. In this study, carnosine levels were found to be higher in urine of the control group, which would be in line with these assumptions. The 2-Acetyl-1-alkyl-sn-glycero-3-phosphocholine levels in urine exhibited relevant differences in comparisons of metabolomic profiles of bladder cancer patients and healthy volunteers. This compound is also known as a platelet-activating factor (PAF), playing a role in platelet aggregation, inflammation, and allergic reactions. PAF and its receptor have been implicated in malignant processes such as tumor development, growth, and metastatic angiogenesis [32]. [PERSON] et al. observed significant increases in PAF accumulation in bladder cancer cells following cigarette smoke extract exposure [12]. The increased levels of PAF in BC patients are in accordance with the current study; however, the number of smokers in both studied groups was similar, which might suggest that smoking is not the only factor influencing PAF accumulation in the bladder. A higher level of 4-hydroxycinnamic acid was found in the urine of healthy volunteers compared to patients with bladder cancer. This organic compound, also known as \(p\)-Cumaric acid, is a naturally occurring phenolic acid present in most plants, including commonly consumed vegetables and fruits [33]. There are numerous reports describing its antioxidant and anticancer properties [34; 35; 36]. [PERSON] et al. have shown that \(p\)-coumaric acid inhibits tumor growth through reduction of angiogenesis within the tumor [36].