Chemical Composition of Kombucha

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This entry is adapted from the peer-reviewed paper 10.3390/beverages8030045

Kombucha is a fermented sweetened tea with a mixed fermenting culture of yeast and acetic acid bacteria. The proponents of kombucha consumption tout the varied health benefits it can provide. The final kombucha flavor and composition is a function of both the initial tea used and the fermentation process.

kombucha fermentation tea

1. Introduction

Kombucha is sweetened tea that has been fermented by an inoculation of symbiotic culture of bacteria and yeast (SCOBY) ^[1]. The origins of kombucha are not well understood; however, evidence suggests that it originated in the Manchuria region, which is located in Northern China ^[2]. There is also evidence to suggest that kombucha has a long history of consumption in China, Russia, and Germany ^[3]. In addition, kombucha is brewed in several Asian countries, such as Japan, India, Korea, Java, and the Philippines. While kombucha is an old beverage it has become increasingly popular in Western society recently. Now it is possible to find kombucha in a large portion of the world, including Eastern Europe, the United Kingdom, the United States, Canada, and Brazil ^[4].

The fermented tea and kombucha market is globally worth USD 1.84 and the leading consumer of kombucha is the North American market, with the United States consuming about half of all the kombucha produced ^[5]. Another leading consumer of kombucha is the European market, with the United Kingdom and Russia leading the consumption within that market share. Probiotic drink consumption has increased in Asian countries, most prominently in China. Additionally, markets in the Middle East and North Africa have increased the consumption of kombucha because of the beverage’s health benefits and non-alcoholic nature. The global kombucha market is forecast to grow by 500% over the next 7 years ^[5].

The popularity and availability of Kombucha in the United States has been, in part, driven by social media, highlighting the positive health benefits that can be achieved from regular consumption. It has been suggested that kombucha can increase vitality, combat acne, eliminate wrinkles, purify the gall bladder, improve constipation, increase weight-loss, relieve arthritis, increase immune response, reduce blood pressure and cholesterol, reduce kidney calcification, and even inhibit cancer proliferation and cure aids ^[2]^[3]^[6]^[7]^[8]. These ascribed associated health effects have certainly caused an increased interest in this product, yet there is limited published scientific research attributing these effects to the consumption of kombucha. However, it should be noted that there are some indications that the consumption of kombucha may indeed provide some health prophylaxis and recovery through detoxification, antioxidants, energizing, and immune-stimulating effects ^[9].

2. Chemical Composition

Kombucha is composed of a number of organic acids, sugars, vitamins, amino acids, biogenic amines, purines, pigments, lipids, proteins, some hydrolytic enzymes, ethanol, caffeine, carbon dioxide, polyphenols, anions, minerals, D-saccharic acid-1, 4-lactone (DSL), bacterial metabolites ^[2]. Several examples are shown in Figure 1. The chemical composition of the tea leaves used to produce kombucha has been well studied and will impact the concentration of the compounds within the kombucha ^[4]. The presence and quantitates of certain chemical compounds are dependent upon the microorganisms found with the SCOBY, fermentation parameters (time and temperature), sucrose concentration, type of tea used, and the analytical method used for quantification ^[10]. If sucrose is used as the primary carbon source for the fermentation, acetic acid will be the predominate metabolite produced. Other organic acids like gluconic and glucuronic are also produced during the fermentation process. If the fermentation process is allowed to go on for too long the pH will drop to low and it becomes undrinkable ^[11]^[12].

Figure 1. Several chemical structures for example compounds found in kombucha including: (A) vitamin C, (B) β-catechin, (C) ethanol, (D) lactic acid, (E) glucuronic acid, and (F) caffeine.

2.1. Compositions

2.1.1. Vitamins

Although the clinical evidence regarding the benefits of kombucha on health is lacking, scientists have confirmed through research that kombucha is comprised of a wide array of chemical components that come from both green and black tea. Some of these components are known to positively affect the human immune system and metabolic processes in the body. Kombucha fermented with green or black tea contains high levels of Vitamin C or ascorbic acid, and trace amounts of some B vitamins. Vitamins are necessary components for numerous biochemical and physiological processes that take place in the body. Vitamins cannot be synthesized within the body; therefore, they must be supplemented in the diet to obtain healthy levels ^[13]. The water-soluble Vitamin C and Vitamin B (thiamine, riboflavin, niacin, pantothenic acid, B6, biotin, B9, and cobalamin) have been reported in kombucha ^[14].

Water-soluble vitamins are less likely to be stored within the body like fat-soluble vitamins (Vitamins A, D, E or K) because they are quickly transported through the blood stream following consumption ^[14]. Vitamin C supports human health by using defensive antioxidants, the formation of collagen to aid in connective tissue, and during an immune response. At the height of an infection, levels of Vitamin C are quickly depleted ^[14]. The mechanism behind this action can be explained by the fact that Vitamin C does not require a coenzyme, although it acts as a cofactor for an enzyme called “prolyl hydrolase”, which aids in the formation of collagen ^[15].

2.1.2. Minerals

Kombucha is a complex beverage composed of a number of compounds, some of which are minerals (F, K Mn) that come from the tea itself ^[16]. Vitamins and minerals are used by the body for a number of metabolic pathways along with physiological functions ^[17]. Minerals are inorganic substances in which play an important role in the human body. Small amounts are required by the body for normal function, growth, and maintenance ^[18]. Essential minerals, such as potassium (K⁺), cobolt (Co²⁺), manganese (Mn⁴⁺), copper (Cu²⁺), iron (Fe²⁺), magnesium (Mg²⁺), and fluoride ions (F⁻) ^[18]^[19], can be found in kombucha made from green and black tea. Bauer-Petrovska and Petrushevska-Tozi (2000) quantified the content of manganese, iron, nickel, copper, zinc, lead, cobalt, chromium, and cadmium in kombucha ^[18]. Mineral concentration can range from 0.004 μg/mL for cobalt to 0.462 μg/mL for magnesium ^[18]. The authors also analyzed known toxic metals. Lead (0.005 μg/mL) and chromium (0.001 μg/mL) were detected in kombucha, while cadmium was not. Studies have shown that certain essential minerals (Cu, Fe, Mn, Ni, and Zn) increase due to the fermentation process, while others, such as cobalt, did not.

Tea, and ultimately kombucha, contain various minerals including fluoride (F). Fluorine plays an important role in the hard tissue mineralization process. However, there is a fine line between enough and too much fluorine. Approximately 85% of fluoride comes from food and beverages (tea, herbal infusions alcoholic beverages, and coffee), while the remaining 15% comes from toothpaste and drinking water ^[16]. It is well known that certain compounds will decrease sugar concentration), while others will increase it (ethanol and acetic acid). Little is known about what happens to certain ions, such as fluoride, during the fermentation process. Jakubcyk et al. (2021) examined the fluoride content during the fermentation process. Four different tea (white, red, green, black) infusions were analyzed, and the fluoride content ranged from 0.42–0.93 mg/L ^[16]. The authors saw that the white tea kombucha had the lowest levels, while green had the highest. It should be noted that the authors used distilled water during their experiment; however, if a kombucha brewery uses a municipal water source that will also contribute to the overall concentration of fluoride in the finished product ^[16]. The recommended dietary allowance (RDA) for adults is 3 mg/L (women)–4 mg/L (men) ^[20]. A single glass of kombucha has the ability to contribute significantly to an individual’s RDA for fluoride ^[16].

2.1.3. Polyphenols

Polyphenols are bioactive substances that contain more than one phenol structure per molecule. Polyphenols represent the largest group of phytochemicals and are the most abundant antioxidants in the diet. People are estimated to ingest upwards of 1 g/day of polyphenols ^[21]. Green, black, and many other teas are rich in water soluble polyphenols; these components make up the aroma and taste of the tea. These compounds may account for up to 30% of the dry weight of the tea leaves, according to the literature. The primary polyphenols found in fresh tea leaves are flavonoids flavanols, flavanol gallate, and flavanol glycosides ^[22]. Flavonoids are a group of bioactive compounds produced during plant metabolism. Flavonoids can be found in fruits and vegetables, such as raspberries, blueberries, and spinach, along with beverages, such as wine and tea ^[23]. Flavonoids are composed of two six figure rings linked together by a three chalcone structure.

Catechins, often referred to as flavanols, are a type of bioactive compound that is a subclass of flavonoids. These compounds are also the primary secondary metabolites found in tea ^[22]^[23]. The major catechins are: α-epigallocatechin-3-gallate (EGCG), α-epigallocatechin (EGC), α-epicatechin-3-gallate (ECG), α-epicatechin (EC), α-epicatechin-3-gallate (ECG), α-gallocatechin, and β-catechin ^[22]. It should be noted that the concentration of catechins can vary based on the tea type and style. Catechin levels in green tea are relatively stable, unlike black tea, because in contrast to black tea, green tea does not undergo any sort of oxidative process during manufacturing. Due to the higher concentration of catechins, this is the main reason that green tea’s characteristic flavor profile is often described as being bitter and astringent. The catechins in black tea are oxidized to form theaflavins and thearubigins, which causes the catechins levels to drop by 85% when compared to green tea. This is the reason black tea is darker and less bitter ^[23]. The antioxidant properties of polyphenols are responsible for the health benefits associated with tea and kombucha, such as the prevention of cancer, increased immunity, reduced inflammation, and arthritis ^[24]^[25].

The total polyphenols and individual concentrations of specific polyphenols in kombucha can vary based on the type of tea leaves used. Cardoso et al. (2020) analyzed kombucha brewed with green and black tea leaves and found that the total polyphenol concentrations varied from 0.70 mg GAE/mL (green)–1.09 mg GAE/mL (black) ^[26]. Yang et al. (2022) analyzed nine (black, green, and mixture black/green) kombucha samples for total polyphenols and four tea catechins (catechin (C), (-)-epicatechin (EC), (-)-epicatechin gallate (ECG), and (-)-epigallocatechin gallate (EGCG)). The total polyphenols measured ranged from 120 μg/L GAE–380 μg/L GAE. Of the samples analyzed, the black tea had the highest concentration of total polyphenols, while a mixed kombucha (black/green) had the lowest ^[27]. Ozyurt (2020) found similar results to both Cardoso and Yang when analyzing the total polyphenols for green (312 μgGAE/mL) and black (422 μgGAE/mL) ^[28]. These results are consistent with previously reported results from other research groups ^[6]^[25].

2.1.4. Ethanol

Ethanol, a byproduct of yeast fermentation, can also be found in kombucha. The concentration of ethanol in kombucha will continue to increase as the fermentation progresses. Chen and Liu (2000) found in their study that the ethanol concentration reached its maximum value of 5.5 g/L on day 20 of the fermentation, followed by a slow decrease ^[29]. The entire chemical profile (sugars, carbon dioxide, organic acids, etc.) of kombucha plays a role in the final flavor and aroma profile of the finished beverage, which is why it is important to control ones fermentation parameters to obtain the desired characteristics of the final product ^[29]. Traditional kombucha does contain ethanol. The Food and Drug Administration (FDA) has investigated the range of kombucha to be between 0.7–1.3% alcohol by volume (ABV). However, craft breweries are starting to make what is known as “hard kombucha”. Kombucha with higher amounts of alcohol. Hard kombucha is known to have an alcohol content of around 3.5–5.5% ABV or higher ^[30].

2.1.5. Organic Acids

Kombucha is made up of a number of organic acids, such as acetic, gluconic, glucuronic, citric, L-lactic, malic, tartaric, malonic, oxalic, succinic, pyruvic, and usnic ^[2]. The composition and metabolite concentration within kombucha can vary greatly due to the starter culture used ^[31], sugar and tea concentration ^[32], fermentation time ^[29], and the fermentation temperature ^[25].

Yeast and bacteria hydrolyze sucrose into glucose and fructose using the enzyme invertase. Yeast within the matrix then produces ethanol via glycolysis, using fructose as the primary substrate. Acetic acid bacteria uses the glucose to make gluconic acid and also utilizes the ethanol produced by the yeast and turns it into acetic acid ^[33]. Acetic acid is the organic compound responsible for the vinegary flavor and aroma commonly associated with kombucha. The concentration of this acid can vary; however, it tends to reach its peak at 11 g/L on day 30 of the fermentation process and will drop to 8 g/L by day 60. The reason for the reduction in acetic acid is due to the microorganisms within the kombucha that utilize acetic acid as a carbon source after they have depleted all the sugar and ethanol within the fermentation matrix ^[2]. The ethanol and the acetic acid found in kombucha have been reported to provide antiseptic properties, inhibiting the growth of pathogenic microbes ^[33].

Lactic acid is primarily found in kombucha made from green tea, rather than other teas, for example, black. Microorganisms produce glucuronic acid from the oxidation of glucose during the fermentation process. Glucuronic acid is the most significant detoxifier in the body because of its ability to bind with toxic compounds in the liver. Once those toxic compounds are bond the body is able to excrete them via the kidneys ^[31].

Glucuronic acid (GlcUA) plays a significant role in detoxifying the liver because of its ability to combine toxic molecules, which are eliminated by the organisms. This acid is very involved with endobiotic elimination. Bilirubin is a well-known endobiotic, which is eliminated by GlcUA (glucuronidation) to prevent toxic pigments from harming the organism. Most bilirubin is excreted through bile, while a smaller portion is excreted in the urine, which is why high concentrations of bilirubin are found in the urine are an indication of damage somewhere in the process ^[34]. GlcUA also plays an important role in increasing the bioavailability of polyphenols. Phenols will conjugate with GlcUA, thus improving its ability to transport, along with it its bioavailability ^[9]. Glucuronic acid is also a precursor for the biosynthesis of vitamin C ^[31].

2.1.6. Caffeine

Caffeine is a naturally occurring xanthine alkaloid found in a number of plants, such as coffee, tea, and cocoa ^[4]. Caffeine, and to a lesser extent theobromine and theophylline, are well known components of tea ^[35]. The caffeine within the plant is used as a pesticide to protect the plant from insects. Humans tend to consume products with caffeine in them for the stimulating effect on the nervous system that they provide, along with an increase in energy. However, some people are sensitive to caffeine and watch their daily consumption of caffeine ^[4]. Caffeine makes up approximately 3% to 6% of the tea leaves. The concentration of caffeine within the tea leaves varies based upon cultivation conditions and the further processing of the tea leaves ^[22]. When it comes to kombucha, caffeine plays an important role during the fermentation process, by providing the yeast and bacteria with the nitrogen necessary for metabolic processes and building new cells, as well providing energy for the yeast and bacteria so they can undergo the fermentation process ^[4].

2.1.7. Amino Acids and Biogenic Amine (BAs)

Amino acids found in foods and beverages are essential for the human body. There are a number of amino acids that have been identified in tea, such as aspartic acid, threonine, glutamic acid, glycine, a-alanine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine, glutamine, asparagine, tryptophan, and theanine ^[35]^[36]. Theanine is the most abundant amino acid found in tea, accounting for 50% of the total amino acids and 1% of the dry weight of the tea ^[19].

While amino acids play an important role in the human body, they can transform into biogenic amines (BAs). BAs are the result of microbial decarboxylation during the fermentation process ^[37]. Some BAs are essential for cellular metabolism, while others can be harmful if consumed in high concentrations, such as histamine tyramine, putrescine, cadaverine, β-phenylethylamine, agmatine, tryptamine, serotonin (SRT), spermidine, and spermine ^[38]. Typically, consuming low amounts of BAs tends to not have an impact on the human body; however, toxic elevated levels of BAs can be found in fermented foods ^[39]. Several factors can influence the formation of BAs, such as starting materials, starter culture, processing, and storage conditions ^[37]^[39].

BAs are an important group of compounds to control and monitor because of the potential toxicological and health implications on the human body but are also used as quality, safety, and product freshness indicators for various types of foods (meat and fish) and beverages (wine) ^[37]^[38]. The number of articles and reviews that discuss the concentration of BAs in kombucha are limited. Researchers have focused on analyzing the individual amino acids and the BAs in the SCOBY (tea fungus) itself; however, no researcher has discussed the concentration of the kombucha itself (broth) that is typically consumed. Researchers have found that the SCOBY contained higher concentrations of lysine, isoleucine, and leucine and lower levels of phenylalanine, valine, methionine, threonine, tryptophan, glutamic acid, alanine, aspartic acid, and proline ^[10]. Of the amino acids identified in the SCOBY only three (lysine, phenylalanine, and tryptophan) are precursors for potentially harmful BAs ^[39]. Ethylamine, choline, and adenine are the only identifiable BAs found in kombucha that have been discussed in the literature, none of which are harmful ^[40]. Bromley (2021) examined the BAs concentration in kombucha broth and found there were no biogenic amines in the sample ^[40].

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