Kombucha: Challenges for Health and Mental Health

Background: Increasing research into probiotics is showing potential benefits for health in general and mental health in particular. Kombucha is a recent beverage and can be considered a probiotic drink, but little is known about its effects on physical and mental health. This product is experiencing growth in the market; however, there are no scientific results to support its potential for physical and mental health. Aim: This review article aims to draw attention to this issue and to highlight the lack of studies in this area. Key findings and conclusions: The lack of legislation for the correct marketing of this product may also constrain clinical studies. However, clinical studies are of utmost importance for an in-depth understanding of the effects of this product on the human body. More research is needed, not only to better understand the impact of Kombucha on the human body, but also to ensure the application of regulatory guidelines for its production and marketing and enable its safe and effective consumption.


Introduction
Nowadays people are increasingly concerned about their own health.People want to become healthier, and the adoption of healthier lifestyles is necessary to improve quality of life and to reduce the numerous pathologies and comorbidities associated with nutrient inadequacy or poor diet choices [1].People are concerned about their body, their mind and their health, and so their search for healthy foods or specific foods that improve health, such as functional foods, has been increasing [2][3][4].Functional foods have been linked to improved health and greater quality of life.These products are defined as dietary products (processed or natural foods) that when consumed regularly in a balanced and diversified diet, improve both the mental and physical state of health and/or reduce the risk of disease [5,6].These functional foods can be classified from the product point of view as follows: fortified (with additional nutrients), enriched (new nutrients or components), altered (substance(s) replacement by other(s) with beneficial effects), and/or enhanced products (new feed composition, genetic manipulation) [5].Thus, in addition to their nutritional value as conventional foods, these foods present potential to enhance health and can reduce the risk of developing non-communicable diseases, such as cancer, diabetes, cardiovascular diseases, and mental illnesses [7].Although there is not yet a worldwide acceptable definition for functional foods, a definition that may become integrative is that of the Functional Food Centre (FFC).The FFC defines functional food as 'natural or processed foods that contain known or unknown biologically-active compounds which, in defined, effective, and non-toxic amounts, provide a clinically proven and documented health benefit for the prevention, management or treatment of a chronic disease' [8].
According to the literature, some of the main components of functional foods are polyunsaturated fatty acids, probiotics/prebiotics/synbiotics, and antioxidants [6].The current research highlights the role of probiotics in functional foods and their impacts on mental health.The development of functional foods and beverages with probiotics has shown a marked increase over the last years, and the consumer and market demand for these products is increasing all over the world [9].According to Fortune Business Insights the global market was valued at USD 48 billion in 2019 and it is projected to reach USD 94 billion by 2027 [10].Probiotics have been in the spotlight lately mainly due to their impact on the modulation of the gut microbiota and to other irrefutable health benefits, such as neurological and immunological effects [11,12].Currently, it is known that physical and mental health are controlled by the brain and one's environment, but recent discoveries have placed a special role on the gut microbiota [13,14].Recent findings show that there is a relation between what we consume and our mental health, more precisely, between the brain and the gut [13], which means that both emotional and behavioral processing are affected not only by the brain and the environment, but also by the gut microbiota [13,15].
The consumption of probiotics is increasing, and there is a great diversity of probiotic products on the market [11,12].These products are displayed in several forms such as supplements (e.g., powder, capsules), food (e.g., yoghurts, cheese, butter, soy products, fermented vegetables, processed food, among others), and beverages [16].

The Kombucha Beverage
One popular beverage that has been gaining visibility is Kombucha.Kombucha is a traditional Chinese beverage made of sweetened tea (mainly black or green; however, other variants of tea and also other ingredients, such as cereals, milk, and even mushrooms, may be used) fermented by a specific combination of probiotic microorganisms.Kombucha mainly comprises tea polyphenols, amino acids, vitamins, minerals, organic acids, sugars, and probiotic microorganisms.This is a functional beverage that has a high amount of plant extracts and metabolites resulting from the metabolism of the symbiotic culture of bacteria (mainly Lactobacillus and Bifidobacterium species) and yeasts (mainly Saccharomyces species), known as SCOBY, [17,18].According to the literature, Kombucha has been shown to have physiological benefits, such as anti-inflammatory, antibacterial, and antioxidant activities, alongside other properties.Consequently, it qualifies as a functional beverage [19][20][21] (see Figure 1).
According to the literature, some of the main components of functional foods are polyunsaturated fatty acids, probiotics/prebiotics/synbiotics, and antioxidants [6].The current research highlights the role of probiotics in functional foods and their impacts on mental health.The development of functional foods and beverages with probiotics has shown a marked increase over the last years, and the consumer and market demand for these products is increasing all over the world [9].According to Fortune Business Insights the global market was valued at USD 48 billion in 2019 and it is projected to reach USD 94 billion by 2027 [10].Probiotics have been in the spotlight lately mainly due to their impact on the modulation of the gut microbiota and to other irrefutable health benefits, such as neurological and immunological effects [11,12].Currently, it is known that physical and mental health are controlled by the brain and one's environment, but recent discoveries have placed a special role on the gut microbiota [13,14].Recent findings show that there is a relation between what we consume and our mental health, more precisely, between the brain and the gut [13], which means that both emotional and behavioral processing are affected not only by the brain and the environment, but also by the gut microbiota [13,15].
The consumption of probiotics is increasing, and there is a great diversity of probiotic products on the market [11,12].These products are displayed in several forms such as supplements (e.g., powder, capsules), food (e.g., yoghurts, cheese, butter, soy products, fermented vegetables, processed food, among others), and beverages [16].

The Kombucha Beverage
One popular beverage that has been gaining visibility is Kombucha.Kombucha is a traditional Chinese beverage made of sweetened tea (mainly black or green; however, other variants of tea and also other ingredients, such as cereals, milk, and even mushrooms, may be used) fermented by a specific combination of probiotic microorganisms.Kombucha mainly comprises tea polyphenols, amino acids, vitamins, minerals, organic acids, sugars, and probiotic microorganisms.This is a functional beverage that has a high amount of plant extracts and metabolites resulting from the metabolism of the symbiotic culture of bacteria (mainly Lactobacillus and Bifidobacterium species) and yeasts (mainly Saccharomyces species), known as SCOBY, [17,18].According to the literature, Kombucha has been shown to have physiological benefits, such as antiinflammatory, antibacterial, and antioxidant activities, alongside other properties.Consequently, it qualifies as a functional beverage [19][20][21] (see Figure 1).Kombucha is very rich in various bioactive compounds, including acetic and gluconic acid, complex B vitamins, minerals, amino acids, and polyphenols, that have been recognized as having health benefits, such as anti-inflammatory, antioxidant, antidiabetic, and antimicrobial activity [22].Different studies have shown that the type and amount of these bioactive compounds are associated not only with the properties of the black or green tea used (other materials can also be used, such as cereals, milk, and even mushrooms) but also with the fermentation conditions, namely, the type of microorganisms and their interactions, time and temperature [22,23].
The literature reports the impact of the biological properties of Kombucha on health and several disorders, such as inflammatory diseases, arthritis, allergies, cardiovascular diseases, and cancer [24].Some studies highlight that Kombucha's composition impacts hypocholesterolemic activity, as well as anti-hypertensive and antidiabetic activities, by inhibiting the enzymes angiotensin-converting enzyme, α-amylase and β-glucosidase, respectively [24].
On the other hand, some studies have correlated the consumption of probiotics with reductions in anxiety and depression [25][26][27], and a positive impact on cognitive and emotional functions [28].However, it is not well established how its consumption can affect the human "psyche".It is known that probiotics modulate the intestinal microbiota which subsequently influence brain responses, such as neurotransmitter biosynthesis and the emotional state, through the gut-brain axis [29].
Thus, the present review focuses on the role of probiotic Kombucha in mental health and associated diseases and intends to draw attention to this thematic, summarizing the latest publications in this field and aiming to understand its potential impact on health.

Mental Health via Gut-Brain Axis
The literature contains increasing evidence that corroborates the mutual influence between the brain and the gut microbiota [25,30].The gut microbiota-brain axis is a bidirectional structure that connects these two organic systems and has effects on the modulation of human organic and psychological functions [30].For example, substances released by the gut microbiota can reach the brain via the circulatory pathway.On the other hand, the brain may also be influencing the gut microbiota by neuronal and endocrine pathways.The literature reports that this communication occurs through three main pathways: the autonomic nervous system (including the enteric nervous system and vagus nerve), immune system and neuroendocrine system [14,31,32].The autonomic nervous system (ANS) is a neural network that comprises the sympathetic and parasympathetic systems.This network in connection with hypothalamicpituitary-adrenal axis establishes the communication between the brain and the gut [14].The ANS is responsible for controlling physiological homeostasis without conscious effort, namely the gastrointestinal function.The ANS in connection with neuronal (e.g., vagus nerve) and neuroendocrine signaling can induce changes in the gastrointestinal autonomic activation, which can be triggered by interoceptive afferent feedback or by cognitive and emotional efferent modulation [14,32,33].On the other hand, the ANS controls several gastrointestinal functions, such as gut permeability, fluids production, and the mucosal immune response, and it can be modulated by gut microbiota metabolites like short-chain fatty acids (SCFAs) and lipopolysaccharides [14,32].Therefore, the ANS provides the gut with the most direct neurological response available; however, the intestinal microbiota can interact with gut ANS synapses, by microbiota-derived neuromodulatory metabolites (e.g., serotonin (SER), gamma-aminobutyric acid (GABA), catecholamines).This process explains the microbiota-gut-brain axis bidirectional communication.Thus, changes in the gut microbiota can influence brain functions (e.g., cognitive, memory, emotion, decision-making) and behaviors (e.g., mood) [14,32,34].
The role of the immune system's pathway role in gut-brain signaling is growing [35].It is known that neuroinflammation increases the odds of developing psychiatric disorders, such as autism spectrum disorders, epilepsy, Alzheimer's disease, Parkinson's disease and cerebrovascular diseases [14,35,36].The literature reports that various gut metabolites, including SCFAs (e.g., butyrate, propionate, acetate), bacteriocins and neuromodulators (e.g., glutamate), seem to activate the immune system and so affect and regulate cytokine secretion and microglial activation [14].Thereby, cytokines, neurotransmitters, neuropeptides, SCFAs and other microbiota metabolites can pass through the blood and lymphatic systems and be in constant communication with both the brain and the gut [14,32,37].
Finally, the other most studied pathway in this axis communication is the neuroendocrine pathway.As with the immune system, gut metabolites can enter into the systemic circulation, activate the hypothalamic-pituitary-adrenal axis, and modify the secretion of gut hormones and neurotransmitters, and consequently regulate body metabolism functions [14,33].The literature shows that homeostatic deregulation can be caused by mental disorders, such as stress, anxiety, or depression [35].For example, in stress disorders, there is a release of the adrenocorticotrophic hormone by the pituitary gland and cortisol by the adrenal cortex, which seems to be associated with the neuroimmune response and cytokine release, leading to an increase in gastrointestinal permeability [14,38,39].
Such a complex mechanism that involves not only internal but also external factors, such as stress, diet, exercise, and the environment, has interested researchers and, in consequence, much research has been conducted to understand these interactions [38].

Probiotics/Kombucha Impact on Mental Health
As mentioned above, probiotics are functional components.According to the World Health Organization (2001), probiotics are live microorganisms that when incorporated into the daily diet in adequate amounts, confer physical and mental health benefits on the host [40,41].Their therapeutic potential has been explored in several pathologies, such as obesity, cancer, inflammatory bowel disease, diabetes, arthritis, and mental disorders.Therefore, the impacts of probiotic products on the gut-brain relation and how altered gut microbiota affects mental illness (e.g., depression, anxiety, bipolar disorder, schizophrenia) and its prevention and/or treatment have been studied [28].
The literature suggests that the consumption of these probiotic foods and supplements may enhance cognitive function.With that, the reduction in negative neurologic or neuropsychiatric conditions, such as depression, anxiety, stress related symptoms, emotional deregulation, some neurodegenerative diseases (Alzheimer's, and Parkinson's disease) can even help with antisocial and aberrant behaviors associated with autism disorder [29,[45][46][47].
Although the literature reports the advantages of probiotics and probiotic food in mental health, less is known about the role of these foods in the gut-brain relation, their importance in CNS function, and in neurodevelopment.Some animal and human studies have reported that probiotics contribute to neuronal modulations through the release of their metabolites, such as GABA, Serotonin, SCFAs, Glutamate, and others.For example, Hur and collaborators (2022) reported that in their animal study, treatment with probiotics relieved hyperactivation of the hypothalamic-pituitary-adrenal axis, which decreased cortisol levels and reduced anxiety and depressive symptoms [34].The probiotics' antiinflammatory effect modulates the proinflammatory cytokines and can potentially decrease depressive symptoms.Probiotics have also been shown to alleviate depressive mood in humans; however, studies in humans are still scarce [34].Although other animal studies have been conducted [48], further studies, both in animals and humans, are needed to validate the beneficial effects of probiotic supplementation on mental health conditions (depression, anxiety and other brain functions).
One probiotic food that has gained particular attention is Kombucha, and the present review aims to investigate the impacts of this specific product on mental health.Regarding the impact of Kombucha on mental health, several studies have reported the health benefits of this functional beverage, and some of its beneficial properties include antioxidant, antiinflammatory [19,20], and antimicrobial activities [49], as well as improving immunologic and liver functions, particularly the hepaprotective effect [20,50,51].Although the literature refers to the possible effects of Kombucha on mental health, there are few studies to support these claims, and hence the need for this review.

Methodology
This review intended to analyze the relationship between "Kombucha and mental health".However, due to the lack of studies in the area, the search was refined and a review on "Kombucha and health" was included to identify information on in vitro, in vivo and human assays.To this end, a systematic search was performed using Pubmed, Science Direct and Web of Science databases from 2 June to 29 July 2023.Several descriptors were used: "Kombucha" AND "health" AND ("in vitro" OR "in vivo" OR "human assays") (Figure 1).The screening of the articles was conducted by applying the following eligibility criteria: (a) Inclusion: research articles, clinical assays, publications written in English.(b) Exclusion: outside the scope of the subject, other types of publications (reviews, comments, editorials, discussions, correspondence, letters, short communications), publications in languages other than English, and studies whose full texts were not available.
This review was conducted using the PRISMA criteria for preferred reporting items within systematic reviews and meta-analyses (PRISMA) [52,53].The collected information was compiled and analyzed regarding the year of publication, authors, sample, country, methodology/type of study, results, conclusions, and research aims.The bibliographic references were compiled through the computer program EndNote bibliographic referencing.

Results
This systematic review identified 476 scientific articles published in international journals indexed to the digital databases used in this search.After screening, duplicate publications and research that fitted the exclusion criteria were removed.Thus, after the eligible criteria were applied, fifty-three publications met the defined inclusion criteria, as shown in the PRISMA flow diagram presented in Figure 2. A summary of the most important characteristics of these articles is presented in Table 1.
It should be noted that no specific studies evaluated the impact of Kombucha on mental health (brain impact, cognitive functions, or brain disorders).Most studies reported the characteristics of the Kombucha beverage, highlighting the physicochemical and biological properties of the product.In vitro and in vivo studies with animal models were included, as well as studies of new products (incorporation of new ingredients) and new processing methods (e.g., fermentation times).
Considering the abovementioned collected data, the lack of clinical assays related to Kombucha is evident (Table 1).Human assays are scarce, and in vivo assays, although they exist, are not yet fully conclusive.Most of the studies that aimed to identify and evaluate the properties of this functional beverage (some studies reported on commercialized Kombucha already in the market and others on Kombucha still in the development process) focused on the physiochemical and biological characterization, using in vitro assays.Based on the analysis of the included studies, it is also worth highlighting the lack of information about the SCOBY used, which is why its inclusion in Table 1 was not justified.
Regarding the studies presented, most of the time, it is difficult to understand the pathology with which they are associated.However, some studies highlight using Kombucha treatments mainly for oncological diseases, diabetes, and cardiac diseases.There is a need for more practical studies about the impact of the consumption of Kombucha on the brain and brain-related diseases.Table 1.Summary information of the studies included from the literature search using the descriptors "Kombucha" and "health", and "in vitro", "in vivo" and "human assays".Olive leaves (polyphenol-rich substrate) may represent a viable strategy to valorize agro-industrial waste, reduce production costs, preserve the antioxidant properties of Kombucha and enrich its sensory profile. [57] To produce Kombucha using coffee infusion, and to evaluate the physicochemical profile, antioxidant, microbial, and possible toxic effects in vivo.
Kombucha producion is feasible since the changes in the physicochemical characteristics, such as reduction in the contents of total soluble solids and reducing sugars and increase in acidity expressed as acetic acid proved the metabolic activity of SCOBY.
According to the pH value, coffee infusion can be considered safe from a microbiological point of view.In vivo assay -n = 50 (5 groups with 10 Galleria mellonella larvae); -Toxicity assays.[58] To evaluate the hangover relieving effect of ginseng berry Kombucha (GBK) fermented with Saccharomyces cerevisiae and Gluconobacter oxydans in in vitro and in vivo models.

Black tea infusion. Ginseng berries 30 • C 18 days
In vitro assay -Radical scavenging activity; -Content of ginsenosides; -Cell viability assay (HepG2 cells); -mRNA expression; -Analysis of genes related to OS and alcohol metabolism; -Malondialdehyde assay.
The radical scavenging activity of GBK was increased by fermentation.In HepG2 cells in which oxidative stress was induced, GBK significantly increased the expression of antioxidant enzymes by upregulating the Nrf2/Keap1 pathway.GBK significantly reduced blood ethanol and acetaldehyde concentrations in ethanol-treated mice.GBK significantly increased the levels of alcohol-metabolizing enzymes, including alcohol dehydrogenase and acetaldehyde dehydrogenase.
The behavioral assays revealed that high-dose GBK significantly ameliorated ethanol-induced behavioral changes.GBK exerted a protective effect against ethanol-induced liver damage by regulating the Nrf2/Keap1 pathway.
In vivo assay -n = 30 ICR mice (5 groups); -Blood ethanol and acetaldehyde concentrations; -ADH and ALDH activities in the liver tissue; -Serum alanine aminotransferase, aspartate aminotransferase, glucose, and lactate dehydrogenase levels; -Behavioral analysis following ethanol administration. [59] To determine the glycemic index and insulin index responses when a standard high carbohydrate, high GI meal is consumed with a complex living Kombucha, compared with either soda water or diet soft drink.
Organic Kombucha (The Good Brew Company Pty Ltd., Victoria, Australia) Not reported Human assay -Randomised, single-blinded, placebo-controlled crossover design with 11 healthy adults with normal glucose tolerance and BMI, aged between 18 and 45 years.
There wase no statistically significant difference in GI or II between the standard meal consumed with soda water or diet soft drink.In contrast, when Kombucha was consumed, there was a clinically significant reduction in GI and II compared with the meal consumed with soda water. [60] To carry out the physicochemical, antioxidant, and enzymatic characterization of green tea and Kombucha.
Green tea leaves Commercial Kombucha, Viva Mais ® brand,
-AOA; -thiobarbituric acid reactive substances; -EIC.Kombucha had lower pH, higher acidity, and solids content compared with green tea.TPC showed no significant difference between the beverages.Green tea and Kombucha presented significant antioxidant capacity, and significant inhibitory activity of the α-glucosidase enzyme; however, green tea presented superior inhibitory potential.Kombucha exhibited pro-oxidant activity.Changes in the Kombucha phenolic profile take place during fermentation, which may lead to the higher bioctive potential and contribute to a better understanding of the Kombucha fermentation process.
[65] To develop and evaluate the protective effect of blueberry fermented and unfermented Kombucha beverages after induction of gastric ulcer in mice.
The use of blueberry for fermentation of Kombucha was favorable in relation to the physicochemical aspects: increased TPC, tannins and anthocyanins contents.However, the fermentation product had lower antioxidant activity when compared with the unfermented beverage.The gastroprotective effect of the fermented Kombucha beverage: the biochemical parameters remained at normal levels; however, some animals showed gastric lesions in an expressive area.
In vivo assays -n = 24 (4 groups (n = 6)) Swiss mice; -Biochemical analysis; -Lipid peroxidation and advance oxidation protein products determination. [66] To incorporate butterfly pea into an innovative drink through a SCOBY fermentation and to evaluate the biological activity (in vitro and in vivo).
A total of 79 secondary metabolite compounds were successfully identified in KBPF.
In vitro studies showed the potential activity of KBPF in inhibiting not only ABTS, but also lipid (lipase) and carbohydrate (α-amylase, α-glucosidase) hydrolyzing enzymes.
In the in vivo study, the administration of KBPF (130 mg/kg BW) significantly alleviated metabolic disorders caused caused by a high-fat diet.Lipid profile (HDL, LDL, TC, TG), blood glucose, markers of oxidative stress, metabolic enzymes (lipase, amylase), and markers of inflammation (PGC-1α, TNF-α, and IL-10) were, in most cases, restored to normal values.The gut microbiota community analysis showed that KBPF has a positive effect on both the Bacteroidetes phylum and the Firmicutes phylum.
-Study design of treatments; -Biomedical analysis of collected blood samples; -Gut microbiota community. [67] To explore the effects of both temperature and time on the dynamic changes induced during fermentation on the properties of laver Kombucha.
Dried laver (Porphyra dentata) sheets 25 or 30  The water-holding capacity of the Kombucha cellulose hydrolysates was higher than for the intact Kombucha cellulose, while the oil-holding capacity was lower.
The hydrolysates of Kombucha cellulose and the intact Kombucha cellulose were used to make deep-fried donuts.
In vitro digestion results suggested that there would be no adverse health effects from substituting Kombucha cellulose hydrolysates into the deep-fried donut formula.
2021 [70] To evaluate the impact of the addition of pitanga and umbu-cajá fruits on the physicochemical parameters, volatiles, phenolics and antioxidant capacity of Kombucha.

• C 48 h
Characterization and in vitro assay -pH and TA; -TSS and TPC; -Profile of organic acids and sugars; -Profile of volatile compounds; -AOA; -Digestion.
After a simulated gastrointestinal digestion, the phenolic contents in all Kombuchas decreased, resulting in a significant drop in the antioxidant capacity.
The findings demonstrated that pitanga and umbu-cajá contribute to diversifying and improving the chemical and bioactive characteristics of the Kombucha, resulting in a sweeter beverage, with a tendency to fruity aromas. [71] To investigate the effects of Kombucha tea based on blood glucose levels, total cholesterol, and PGC-1α in Swiss albino mice that were given CFED.
These preliminary results showed the potential of using these isolated strains as starter cultures for the production of novel functional fermented beverages from papaya pulp and leaves. [73] To produce unfermented and Kombucha beverages, and assess their physicochemical characteristics, in vivo toxicities, antioxidant activities and antimicrobial properties.
It was possible to elaborate a fermented beverage using a non-conventional edible plant.Beverages produced with Malvaviscus showed antioxidant activity but not antimicrobial activity.
Beverages produced with green tea showed high antioxidant and antimicrobial activity.
First time the use of this in vivo model of toxicity.In vivo assay -n = 50 Galleria mellonella larvae model (5 groups of 10 larvae); -Toxicity assay.[74] To evaluate the effects of supplementation with Kombucha and green banana flour on Wistar rats fed with a cafeteria diet.
The intake of caffeine altered the lipid and liver profile of the animals and the consumption of Kombucha and green banana flour did not prevent these changes.
The high polyphenols level in Kombucha did not exert a hepatoprotective effect as an antioxidant.There is great potential for using medicinal mushrooms as a substrate in Kombucha fermentation.Hot water extraction of produced mushroom fruiting bodies enabled the generation of substrate, which stimulated total acid production and consequently provided shorter fermentation time.
The Kombucha products possess significant amounts of bioactive components.
The analysis of the biological properties of Kombucha polysaccharides suggests they have highly desirable immunomodulatory properties in human cell cultures. [77] To evaluate the natural potential of combination therapy of this natural product with doxorubicin as a chemotherapeutic agent.
Kombucha beverage inhibited antiapoptotic function Kombucha beverages prepared from green tea possess interesting antiproliferative properties associated with significant antiapoptotic activity in cellular and molecular scales. [78] To study the effects of tea leaves on the antioxidant capacity, phenolic content, and bioaccessibility of Kombucha teas during fermentation, investigated by the simulated in vitro gastrintestinal digestion method.Kombucha fermentation with tea leaves led to an increase in antioxidant capacity, TPC and bioaccessibility.Among the kombucha tea types, the green tea Kombucha had the highest antioxidant capacity levels.The bioaccessibility levels did not significantly change during the fermentation.The TPC content in the extractable fractions was higher compared with hydrolysable fractions due to complex interactions among the constituents. [79] To evaluate the effects of long-term storage at 4 • C on the pH, total phenolic and flavonoid contents, and free radical scavenging properties of Kombucha during 9 months with a sampling interval of 30 days, aiming to determine the period during which these parameters are stable.

Not reported 30 days Characterization
-Monitoring of pH; -TPC and TFC; -AOA.
After 4 months, the phenolic content decreased significantly from the initial value, as well its antioxidant capacity.
The pH value increased from 2.82 to 3.16.
The novel findings of this pilot study revealed that Kombucha from sugared black tea can be stored at refrigerator temperature for four months.After this period the antioxidant properties of Kombucha are no longer retained. [80] To investigate the possibility of the fortification of traditional Kombucha beverage with different medicinal plant infusions.
After fermentation, the antioxidant capacity of the Kombucha increased.On days 0 and 9 of storage, the bioaccessibility of the total phenolics and antioxidant capacity in all of the samples showed a significant increase after gastric and intestinal digestion when compared with pregastric digestion.The antioxidant capacity after in vitro digestion at the beginning and end of storage in all of the beverages also increased after gastric digestion compared with pregastric digestion; however, it decreased after intestinal digestion. [81] To characterize fermented foods produced at small scale, archaeal and bacterial microbiota composition were determined and a preliminary in vitro gut microbiota experiment was conducted to assess the potential prebiotic or probiotic properties of the kraut product and Kombucha.

Black tea Room temperature 7 days
In vitro assay -Fermentation study; -Microbial DNA extraction; -Dual-index bacterial amplicon library preparation and sequencing.
The Kombucha fermentation was dominated by Gluconacetobacter, but also characterized by a high abundance of Bacteroides.
The microbiota composition and dynamics were very different between the two Kombucha batches tested, suggesting redundancy in microorganisms' fermentative roles.
A preliminary in vitro fermentation study was indicative of a potential bifidogenic effect of microbial metabolites from Kombucha.Metagenomic DNA from the solid and liquid phases of 3 Kombucha consortia revealed differences across Kombuchas and between the two phases.Fermentation kinetics showed an association between the microbiota, sugar consumption, and secondary metabolite production.
The most abundant species of bacteria were the same in all the samples, differing only in relative abundance.
The yeast populations differed most considerably.Microbiota differences were observed in the biological profiles of the obtained teas. [84] To evaluate the antimicrobial activity of green tea Kombucha at two fermentation time points against Alicyclobacillus spp.
Green tea (local grower in Maringá, Paraná) Room temperature 7 and 14 days Characterization -AOA; -AMA; -Compounds characterization; Kombuchas fermented for different times showed effective inhibitory and bactericidal activities at low concentrations against all evaluated Alicyclobacillus spp.The length of fermentation affects the antimicrobial activity of Kombucha against spoilage bacteria: the longer the fermentation time, the greater the antimicrobial activity.Fermented green tea showed a higher number of metabolites than green tea not fermented by symbiotic bacteria. [85] To isolate and identify a yeast strain from Kombucha and evaluate in vitro its potential as a novel starter in beverage fermentation.Cytotoxicity tests showed that the biodegraded products were less toxic than pure AFB1. [87] To investigated the anti-virulence activity of a polyphenolic fraction previously isolated from Kombucha.

Black tea 28 • C 14 days Characterization
-ABA; -Effect on bacterial motility and proteolytic activity; The overall results imply that Kombucha might be considered as a potential alternative source of anti-virulence polyphenols against V. cholerae.This is the first report on the anti-virulence activity of Kombucha, mostly attributed to its polyphenolic content. [88] To determine the antioxidant capacities, and antibacterial and antiproliferative activities.
A greater diversity and abundance of phenolic compounds were detected in black tea Kombucha, which resulted in a higher antioxidant capacity.The green tea Kombucha was the only one that presented antibacterial activity against all the bacteria tested and an increased antiproliferative activity against the cancer cell lines.
The type of tea used in the Kombucha production influences its bioactive composition and properties.Results for the ferments were compared with the green coffee extract that was not fermented.
The fermentation time has a positive effect on the content of bioactive compounds and antioxidant properties.The highest values were recorded for the tested samples after 28 days of fermentation.After 14 days of the fermentation process, it was observed that the analyzed ferments were characterized by low cytotoxicity to keratinocytes and fibroblasts.On the other hand, the short fermentation time of 7 days had a negative effect on the properties of the analyzed ferments.
2019 [91] To compare the antidiabetic activity of snake fruit Kombucha, black tea Kombucha and metformin in streptozotocin-induced diabetic rats.

Black tea
Room temperature 14 days Characterization -pH and total acidity; -RS and AOA; -TPC and TSS; -Tannins.
The 3 treatments (snake fruit Kombucha, black tea Kombucha and metformin) were effective as diabetes therapy agents in the rat model by lowering FPG, improving oxidation stress status and lipid profiles.Improvements in the pancreas by these 3 treatments.The snake fruit Kombucha was as effective as the metformin in managing the induced diabetes, and more effective than the black tea Kombucha.In vivo assay -n = 25 Wistar rats (5 groups) -FPG, SOD-malondialdehyde levels; -Lipid profiles; -Pancreas immunohistochemistry staining.[92] To investigate the feasibility of transforming soy whey into a novel functional beverage using Kombucha consortium."Commercial Kombucha" 28 • C 7 days Characterization -pH and TA; -RS; -TFC; -Glucuronic acid, organic acids and isoflavones contents; -AOA and AMA; -Sensory analysis.
The antioxidant capacity of kombucha-fermented soy whey was significantly enhanced and showed antibacterial activity against Staphylococcus aureus, Bacillus subtilis and Escherichia coli.
The fermentation produced new aroma-active volatiles (esters and higher aldehydes) which imparted the fruity flavor of soy whey and improved its sensory quality. [93] To provide a deep insight into traditionally prepared Kombuchas and gain new knowledge on the use of rooibos herbal tea, which has never been considered as a substrate, in spite of its well-known bioactivity and potential health benefits.
All of the Kombuchas showed similarity in bacterial composition, with the dominance of Komagataeibacter spp.The yeast community was significantly different among all tea substrates, between 7 and 14 days of fermentation and between the biofilm and Kombucha, indicating the influence of the substrate on the fermenting microbiota.Kombucha from rooibos has a low ethanol concentration, and a glucuronic acid level comparable to black tea.Although antioxidant activity was higher in black and green kombucha compared with rooibos, the latter showed an important effect on the recovery of oxidative damage on fibroblast cell lines against oxidative stress. [94] To evaluate if black goji berry and red goji berry fruits are suitable for Kombucha beverage production.

• C 48 h
Characterization and in vitro assay -pH and total acidity; -Determination of water soluble solids.
Total phenolic content of all Kombucha samples fluctuated during fermentation and storage.All Kombucha samples had higher antioxidant activity than their infusions.Total phenolic content and antioxidant capacity in in vitro predigestion and postdigestion of samples were ranked as follows: black tea Kombucha > black goji berry Kombucha > red goji berry Kombucha.Kombucha beverages were successfully produced on yarrow infusions and subcritical water extracts.The most suitable substrate was obtained with the following fermentation process parameters: subcritical water extract at 115 • C, and 2.26 g of yarrow flowers in 500 mL water.Organic acids (oxalic, formic, acetic, succinic and malic) content was higher in beverages produced from subcritical water extracts.Vitamin C values were higher in beverages produced with infusions.Total phenols and flavonoids contents depended directly on the amount of herb.
Yarrow kombucha produced by fermentation on subcritical water extracts showed higher antioxidant activity but lower antimicrobial and antiproliferative activity in comparison with products obtained by infusions.Kombucha beverages produced from subcritical water extracts of yarrow had the highest sensory score. [ To investigate fermentation of snake fruit with the Kombucha consortium.
To evaluate the physicochemical and sensory properties of the fermented products.
To assess the antioxidant and antibacterial activities of the most promising cultivars and their bioactive compounds.
Sugared snake fruit juices from five Indonesian snake fruit cultivars (Salak Doyong, Salak Madu, Salak Pondoh, Salak Segaran, and Salak Suwaru) can potentially be used to produce fermented beverages with the Kombucha consortium.
The fermentation affected the physicochemical and sensory properties of the juices, enhanced the antioxidant and antibacterial activities of the resulting products, and increased the levels of the bioactive compounds, indicating the desirable overall properties of the fermented beverage. [96] To enhance the health-related benefits of pollen by fermentation with a Kombucha/SCOBY consortium.
The pollen addition increased the proportion of Lactic acid bacteria in the total population of SCOBY microbial strains.Scanning electron microscopy images highlighted the adhesion of the SCOBY bacteria to pollen.The content of bioactive compounds (polyphenols, soluble silicon species and short chain fatty acids) was higher in the fermented pollen and the product shows a moderate antitumoral effect on Caco-2 cells.The health benefits of pollen are enhanced by fermentation with a Kombucha consortium.

[97]
To investigate the antioxidant activity and anti-inflammatory effects of Kombucha analogues from oak by examining their modulation ability on macrophage-derived TNF-alpha and IL-6.
Herbal infusions from oak and black tea 28 • C 7 days Characterization and in vitro assay -Chemical characterization; -Sugar, gluconic and glucuronic acid content; -TPC; -AOA and AIA; -Cell viability (THP-1) -Suppression of TNF-alpha and IL-6 release; -Nitric oxide and OS determination.
Kombucha analogues from oak showed good antioxidant properties, attributed particularly to their phenolic composition.The major effect detected was their ability to suppress lipopolysaccharide-induced production of nitric oxide, TNF-alpha and IL-6, showing an important anti-inflammatory activity.
2016 [98] To investigate the antifungal activity of Kombucha tea ethyl acetate fraction (KEAF) against Malassezia species obtained from the patients with seborrheic dermatitis.

• C 14 days
In vitro assay -Culture and identification of Malassezia species; -DNA sequencing; -Antifungal properties.
The results of the DNA sequence analysis indicated that M. furfur was the predominant species, followed by M. globosa, M. sloofie, M. sympodialis, and M. restricta, respectively.The findings of the study highlight the antifungal properties of KEAF.KEAF showed inhibitory activity against Malassezia species.KEAF had the lowest and highest MIC value against M. sloofie and M. restricta, respectively.The Kombucha fermented milk product showed similar trend of changes in pH, degree of proteolysis, and sensory properties as products obtained by probiotic and yoghurt starters.Significant ACE inhibitory was determined in all fermented products, which increased during storage, and the Kombucha product had the highest ACE activity at the end of storage compared with probiotic and yoghurt products.In all products, higher radical scavenging activity was determined, while both activities slightly decreased during storage.
2013 [101] To investigat the antidiabetic and antioxidant effects of Kombucha in comparison with unfermented black tea, in alloxan monohydrate (ALX)-induced diabetic rats.

Black tea
Room temperature 14 days Characterization -TPC and TFC; -Organic acids composition; -AOA.
ALX exposure lowered the body weight and plasma insulin by about 28.12% and 61.34%, respectively, and elevated blood glucose level and glycated Hb.
The oxidative stress-related parameters, including lipid peroxidation end products (increased), protein carbonyl content (increased) and glutathione content (decreased), and antioxidant enzyme activities, were also altered in the pancreatic, hepatic, renal and cardiac tissues of diabetic animals.
The results showed the significant antidiabetic potential of the fermented beverage as it effectively restored ALX-induced pathophysiological changes.It could ameliorate DNA fragmentation and caspase-3 activation in the pancreatic tissue of diabetic rats.
Although unfermented black tea is effective in the above pathophysiology, Kombucha tea was found to be more efficient.This might be due to the formation of some antioxidant molecules during the fermentation period.
-Antioxidant enzymes; -Glutathione level DNA fragmentation assay. [102] To examine the anti-angiogenic effect of Kombucha on angiogenesis stimulators/regulators in a human androgen-independent prostate cancer cell line.
Kombucha baby mat with its mother liquor "commercial" Compared with black tea, Kombucha tea was a better inhibitor of α-amylase and lipase activities in the plasma and pancreas and a better suppressor of increased blood glucose levels.Kombucha was noted to induce a marked delay in the absorption of LDL-cholesterol and triglycerides and a significant increase in HDL-cholesterol.Histological analyses showed an ameliorative action on the pancreas and effective protection of liver-kidney function in diabetic rats, evidenced by significant decreases in aspartate transaminase, alanine transaminase, and gamma-glytamyl transpeptidase activities in the plasma, as well as in the creatinine and urea contents.

[104]
To elucidate the relationship between the fermentation time and antioxidant activities of Kombucha.
To determine changes in free-radical scavenging abilities along with pH and total phenolic compounds of Kombucha tea during fermentation.
Green tea, and black tea, and tea waste material.
The prepared Kombucha teas have excellent antioxidant activities.Kombucha exhibited increased free-radical scavenging activities during fermentation.The extent of these activities depends upon the fermentation time, type of tea material and the normal microbiota of the Kombucha culture, which in turn determined the forms of their metabolites.
2006 [105] To study the effects of the origins of Kombucha on antioxidant ability of Kombucha collected from households throughout Taiwan during fermentation.
-Inhibition of linoleic acid peroxidation measure.
The Kombucha exhibited increased antioxidant activities during fermentation.
The extent of activity depended on the culture period and starter origins, which in turn determined the forms of their metabolites.

Discussion
This systematic review was designed to provide insights into the impact of Kombucha consumption on health in general, and its effects on mental health or brain disorders, in particular.Due to the appeal of Kombucha among consumers, and its potential higher consumption, together with the lack of evidence about its impact on health, specifically in mental health, careful in-depth study of these products is of utmost importance.
According to our results, it is clear there is an absence of scientific evidence/studies that can confirm the veracity of the commonly misused expression 'the beneficial impact of Kombucha in mental health'.In consequence, readers should be cautious, not only when accessing online information, but also when it comes to Kombucha consumption.
Despite the ambiguous discussions about Kombucha found in the literature, in the obtained data, and after using several descriptors, we were only able to identify two articles with experimental designs related to this thematic and these articles were without a clear focus.Permatasari and collaborators reported the potential of Kombucha in antiageing [71].However, the authors also emphasizes that evidence is needed for determining the functional potential of Kombucha consumption in humans.On the other hand, in the research works of Barbosa and collaborators [65] and other research teams (see Table 1), study emphasis was placed on the antioxidant potential of Kombucha, highlighting also the need for further studies.
Most of the studies we found in these three credible scientific databases focused on identifying and characterizing the physicochemical and biological properties of this functional beverage.Some in vitro and in vivo studies are presented, although in vivo assays are also few, and they are not yet fully conclusive.Human assays are scarce, and there is an urgent need to better understand the impact of this specific probiotic beverage on health.However, the lack of legislation and regulatory guidelines for the production process may be one reason for the lack of research.The literature has highlighted the beneficial properties of Kombucha, but the clinical evidence is limited, especially in relation to brain disorders.
The literature has reported the role of Kombucha as a possible neuroprotective agent for neurodegenerative diseases and brain damage lesions, mainly due to its antioxidant properties that are responsible for decreases in neural cell death [24,71,106].In the studies presented in Table 1, there is clear evidence of the antioxidant potential of this functional beverage; however, it is important to transpose these findings to clinical trials.This biological property is of great importance, since the antioxidant activity is at the basis of preventing the development of neurodegenerative diseases and other non-neurological diseases.There are strong arguments and evidence for the possible correlation between this Kombucha activity and reductions in the development of neurodegenerative diseases; nevertheless, this correlation must be tested in order to draw solid conclusions [71].
Furthermore, studies have shown that glutamate, one of the most abundant metabolites in Kombucha, is the most important excitatory neurotransmitters in the brain associated with learning, memory, and neural development [83,107].Furthermore, glutamate with GABA, effectively reduces the symptomology of anxiety, depression, and bipolar disorders [108,109].This information helps us to understand not only the therapeutic potential of Kombucha, but also its possible preventive effect against psychiatric and neurological disorders.However, it is quite ambitious and, in fact, misleading to say that there is a cause-effect relation given the lack of studies addressing this question.Therefore, it is of paramount importance to investigate the glutamate role in Kombucha, how it reacts with other Kombucha components and if it really has a significant impact on mental health.
In addition, the anti-inflammatory properties associated with Kombucha must also be better analyzed as there is limited scientific evidence reported by only few studies [66,83,97].This property is essential to understand neurodegenerative diseases because in these diseases, inflammation is a complex multifactorial process involving the central nervous system.These beneficial properties of the product are due not only to the substrates used in its composition and other elements (e.g., phenolic compounds, proteins, vitamins, etc.), but also to the composition of the applied SCOBY.The literature vaguely mentions these microorganisms, providing little information on their type and quantity.Most of the articles included in this review did not report this information, and so details about SCOBY were not included in Table 1.
In the literature, one can find several studies about probiotic strains and their impact on mental health.However, the validation of their effects should be extended to specific products.Studies in the literature report how the consumption of probiotic supplements (composed mainly of Bifidobacterium, Lactobacillus and Actinobacteria) as part of a therapy can significantly reduce depression symptomatology, improve cognitive functions (e.g., memory and neuroplasticity), and regulate emotional behavior.Nevertheless, it is crucial to assess specific types of probiotics [25,47,[110][111][112], as well as particular food matrices, which are sources of these probiotics.Some studies indicate that treatment with probiotic supplements, containing mainly Lactobacillus and Bifidobacterium, with quantities around 2 × 10 9 CFU/g, primarily for 8-12 weeks, can significantly reduce Alzheimer's and other neurodegenerative disease symptoms as well as elevate the production of serotonin, dopamine, norepinephrine and GABA, thereby reducing depression symptomology [113].A specific study showed that a probiotic treatment consisting of supplementation of 8 × 10 9 CFU/day over 12 weeks for schizophrenia could reduce the condition's general symptomology [114,115].It is obvious that probiotic really do impact our health and reduce the development of certain diseases, specifically neurological ones.Still, given that Kombucha consumption is increasing as a probiotic source, more studies must be performed in order to understand how this beverage can particularly influence the improvement in neurological and psychological conditions and to determine suitable doses to treat different diseases.
Thus, information about SCOBY deserves special attention, especially concerning the creation of regulatory guidelines for the development/production and commercialization of this product to ensure its quality and safety.
Another critical factor that influences Kombucha properties concerns the manufacturing process, namely the fermentation process.This review highlighted some studies that report that the fermentation conditions and time affect the antimicrobial and antioxidant activities and phenolic, flavonoid, and bioactive compounds of Kombucha [54,84,90].For example, Aung and collaborators showed that using higher fermentation temperatures boosts the microorganism growth, maintains important total flavonoid compounds and enhances the α-amylase inhibitory activity [67].
These characterization studies are extremely important, but there is an urgent need for further studies in humans.These studies are clearly scarce but are urgent because the consumption of this product is increasing.Clinical human assays should seek to understand the mechanism of action of probiotic foods and drinks on brain function, particularly on how this interaction occurs and what results from it in terms of cerebral activity.As Kombucha is one of the probiotic beverages with potential for expansion in the market, the implications of its consumption on the human body must be more deeply investigated [116], particularly its main effects on general health, as well as brain activity.On the other hand, the lack of legislation may be an obstacle to the research into impactful novel properties of this beverage [117].Thus, more human clinical trials need to be conducted in order to guarantee the efficacy and safety of this product's production and consumption.Only with clinical studies will it be possible to improve knowledge about this product, clearly understand its benefits, and explore the associated risks, particularly its potential toxicity.
The data reported in this review are intended to draw attention to this issue and its importance.It is extremely important to understand the real impact of Kombucha consumption on health and mental health through clinical trials and to define national and international regulatory guidelines for the production and commercialization of this beverage in order to standardize its consumption and ensure consumer health and safety [117].
The lack of such regulations and artisanal production can compromise the quality and safety of the beverage [116].This drink involves microbiological complexity, which must be preserved according to specific guidelines.Some limitations in the present review should be highlighted.Important limitations were the difficulty in selecting the specific descriptors and the lack of studies in the area, especially in mental health, which hampered the review process.

Conclusions and Future Perspectives
The literature has shown a link between the consumption of probiotics and the benefits associated with mental health, although the scientific evidence about the mental health benefits of the Kombucha beverage is not clear.
Even though probiotic supplements may have positive effects on the prevention and treatment of mental disorders, more studies are needed to explore the different probioticrich functional products in the market.
This review intended to draw attention to the lack of up-to-date scientific evidence about the health effects of Kombucha, particularly mental health-related benefits.In particular, the neuroprotective effect so often associated with Kombucha needs to be carefully studied and its potential biological activity validated.In addition, to ensure safe consumption, it is of utmost importance to apply regulatory guidelines to its production and marketing.Also, the lack of existing legislation regarding this product may also constrain the execution of clinical trials.

Figure 1 .
Figure 1.Composition and properties of Kombucha beverage.Figure 1. Composition and properties of Kombucha beverage.

Figure 1 .
Figure 1.Composition and properties of Kombucha beverage.Figure 1. Composition and properties of Kombucha beverage.
Kombucha treatment also induced weight loss and increased levels of liver SOD.Kombucha improved lipid profiles: reducing total cholesterol, TG, LDL, and increasing HDL levels compared with CFED and normal groups.This Kombucha has good potential as a functional beverage with anti-obese and lipid improving activity.

Table 1 .
Cont.Black and green tea beverages showed the highest antibacterial activity.The bacteria E. coli and Salmonella sp. were the most sensitive to the effects of Kombucha.A total of 17 bioactive compounds were found in Kombucha.Kombucha contained many elements such as aluminium, calcium, iron, potassium, magnesium, sodium, phosphorus, and sulphur.Tea mushroom microflora contained the following microorganisms: Gluconacetobacter xylinus, Acetobacter xylinum, Bacterium gluconicum, Gluconobacter oxydans, Leuconostoc mesenteroides, Propionibacterium spp., Acetobacter nitrogenifigens, Gluconacetobacter kombucha, Saccharomyces cerevisiae, Candida vini, Schizosaccharomyces pombe, Pichia membranefaciens, Kloeckera apiculate, Kluyveromyces marxianus, and Pichia kluyveri.The aroma most highly rated by the evaluators was that of Kombucha No. 4 from green tea and coconut sugar, while Kombucha No. 1 from black tea and cane sugar had a less desirable aroma.