Sharifudin, Shaiful Adzni
(2024)
Evaluation of antioxidant, antimicrobial and potential anticancer activities of papaya-based kombucha beverages on colon cancer.
PhD thesis, University of Nottingham.
Abstract
Traditional kombucha is prepared by fermenting sugar and tea extract with probiotic microorganisms collectively named as Symbiotic Culture of Bacteria and Yeast (SCOBY). Recently, fruits or leaves juices have become a new, favourable alternative medium for kombucha making. The employment of various starting materials and starter cultures during fermentations might result in the creation of several variable metabolites that may have various bioactivities. Thus, this study was aimed to determine the secondary metabolite content, antioxidant activities, antimicrobial effect against selected food borne pathogens and anticancer activities of the kombucha juices of papaya pulp and leaves fermented using locally isolated kombucha cultures, namely: yeast (Dekkera bruxellensis) and acetic acid producing bacteria (Komagataeibacter rhaeticus).
Tolerance of these selected cultures against low pH and different bile salt concentrations were evaluated. D. bruxellensis MFS1 showed tolerance to pH 2 and higher and significant tolerance to bile salt at 0.3 %, 0.5%, and 1%. In contrast, K. rhaeticus MFS1 was only tolerant to pH 3 and higher and exhibited poorer tolerance to bile salt. Then, the kombucha juices supernatant were subjected to the profiling of organic acids and phenolic compounds using UPLC-PDA; quantification of total phenolic content; antioxidant activity measurement using FRAP, DPPH, and antimicrobial effect by well diffusion assay. The concentrations of acetic acid, L-malic acid, kojic acid and quinic acid increased in both fermented kombucha samples, with acetic acid being the most abundant form of organic acid being produced. Besides, chlorogenic acid, ellagic acid and 2,5-dihydroxybenzoic acid were also significantly increased after fermentation. In terms of antioxidant activities, papaya leaf kombucha (PLK) revealed higher activities of antioxidant compounds in the range of 89.56% compared to papaya pulp kombucha (PPK) (54.95%) DPPH inhibition. In addition, supernatant of fermented papaya leaves kombucha were able to inhibit the growth of pathogenic strains with higher zone of inhibition on most of the pathogens when compared to non-fermented and acetate samples.
Other than that, anticancer properties of papaya-based kombucha were evaluated using in vitro and in vivo approaches. Using the MTT assay, higher concentrations of both kombucha pulp and leaves were shown to induce cellular growth suppression in two colon cancer cell lines, HT29 and SW 480. Both treated HT29 and SW480 shared a comparable cell population percentage pattern change from viable cells to early apoptotic, subsequently to the late apoptotic cell, as indicated by the Annexin V-FITC/PI assay. To determine how papaya kombucha administration affected the advancement of the cell cycles in HT29 and SW480, cell cycle analysis utilising flow cytometry was performed. Conclusively, kombucha treatment from PPK and PLK successfully inhibited the transition of the HT29 and SW480 cancer cells in vitro.
Next, an in vivo study model named the AOM/DSS-induced ICR mice model was employed to evaluate the effect of papaya kombucha on the progression of colon cancer. The colon adenocarcinoma tumourigenesis process was delayed when the mice were fed using a low dose (0.7mL/kg) of papaya-based kombucha twice a day. On the contrary, feeding the mice with a high dose (1.8mL/kg) of similar kombucha treatment two times a day hastened colon carcinogenesis. Overall, significant dysbiosis in gut microbiota was observed in the control and AOM/DSS-induced groups. As the most predominant gut microorganisms, Bacteroidetes and Firmicutes demonstrated fluctuating abundances in healthy and ill mice. For instance, Bacteroidetes proliferation was significantly repressed by the growing of colon adenocarcinoma. Combining treatment with kombucha juices of papaya pulp and leaves and increasing Firmicutes growth was shown to help restore the bacterial load of several genera to near control levels.
Using both in vitro and in vivo study designs, papaya kombucha beverages were shown to possess promising anticancer effects. This is because it contains potentially advantageous probiotic microbes, phenolic compounds, and substantial organic acid. Moreover, the acidic nature of kombucha enables it to regulate the pH balance within the intestine, helping to establish a healthy and balanced gut environment. The aforementioned positive results demonstrate the prospective benefits of incorporating papaya kombucha into one's diet as a functional food.
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