Kombucha

Kombucha is a fermented beverage made from tea and sugar, where the addition of inoculum and subsequent fermentation is thought to produce unique health bioactives.^[1] This inoculum is commonly referred to as 'tea fungus' and contains a mixture of bacteria and fungi which act during the fermentation process. A pellicle which forms during this process is referred to as the 'fungus', and the fungus produces alcohol which helps the bacteria produce the aforementioned bioactives.^[2]

The tea used is from the Camellia sinensis plant and usually refers to the black tea, although green tea is sometimes used. When black tea is used (as black tea itself requires fermentation to be produced) the final kombucha product is sometimes referred to as being doubly fermented.

Kombucha is produced by making a tea from the plant Camellia sinensis (basic extraction being called 'green tea'), fermenting it initially to make black tea, and then fermenting it again with a mixture of bacteria and fungi and sugars to produce kombucha; in this sense, it is doubly fermented.

The essential organisms that make up the tea "fungus" are an acetic-acid producing strain of bacteria (usually the Acetobacter genus, both and being identified) and yeast.^[1] Strains of lactic acid-producting (Lactobacillus) and gluconic acid-producing (Gluconobacter oxydans being identified) bacteria can also be present.^[1]

There are various yeast fungi that are seen, including Brettanomyces/Dekkera, Candida, Kloeckera, Pichia, Saccharomyces, Saccharomycoides, Shizosaccharomyces, Torulospora and Zygosaccharomyces.^[1][3] Although most have been unidentified, up to 163 strains of yeast have been detected,^[1] with four major yeasts being Zygosaccharomyces bailii, T.delbrueckii, C.stellata, and S.pombe.^[1]

Although kombucha contains a variety of bacteria and fungi, it appears to be safe for human consumption when properly processed and consumed in moderation (details in the safety and toxicology section).

The strains of bacteria used in kombucha fermentation are those that are acid-resistant and produce acids upon metabolizing ethanol and sugars, and while there are no standard yeast fungi used during this process, the ones that are most-often found also tend to also be acid-resistant and acid-producing.

Components of kombucha that are preexisting in the tea (Camellia sinensis) prior to fermentation include:

• Green tea catechins, which experience variable degradation rates (18-48%) with less degradation seen with green tea relative to black tea and less with EGCG relative to other catechins;^[2] epigallocatechin (EGC) and epicatechin (EC) were noted to be markedly (30-50%) elevated after 12 days fermentation, thought to be due to the degragation or their gallated forms (EGCG and ECG, respectively).^[2]
• Theaflavins found in black teas, where 5% is lost during 18 days fermentation^[2]
• Thearubigins found in black tea, where 11% is lost during 18 days fermentation^[2]

The standard tea polyphenols found in the Camellia sinensis plant and those produced during the intitial fermentation to get black tea (theaflavins and thearubigins) persist in kombucha, with the losses during the second fermentation being fairly minimal.

Components of kombucha tea produced during the fermentation process include:

• Alcohol (produced from added sugars via yeast) reaching 0.6g/100mL after 10 days^[4]
• Acetic acid (produced from the alcohol via bacteria)^[5] reaching 1.6g/100mL after 10 days;^[4] this may be a high estimate, as other studies have noticed a plateau at 0.95g/100mL after 15 days followed by a decline^[2]
• D-Saccharic acid 1,4-lactone (saccharolactone)^[6]
• Succinic acid reaching 0.65g/100mL after 10 days^[4]
• Lactic acid^[5] peaking after three days of fermentation (when other acids required 15 days)^[2] resulting in around 0.01g/100mL after 12 days^[2]
• Gluconic acid^[5] reaching 0.20g/100mL after 10 days^[4]
• Glucuronic acid produced from glucose in the medium^[7] reaching 0.38g/100mL after 10 days^[4] although elsewhere a plateau of 0.23g/100mL has been noted around 7-12 days^[2][7]
• Usnic acid^[8][5]
• Citric acid has been noted to transiently occur after three days fermentation (less than 0.01g/100mL) although it's undetectable after 12 days^[2]
• Carbon dioxide (produced from the acetic acid via bacteria), produced which separates the pellicle from the broth and creates an anaerobic/serum-starved environment^[2]

Studies comparing the fermentation of green tea and black tea by the same fungal and bacterial colonies have failed to find any significant differences in the production of acids, aside from possibly more acetic acid with green tea relative to black tea.^[2]

The fungal fermentation that produces kombucha creates a large variety of small acidic compounds, with the most notable one (thought to mediate the 'detoxification' effects) being D-Saccharic acid 1,4-lactone (Saccharolactone)

Kombucha is known to have a somewhat specific processing method, and similar to most fermentation products (requiring heat) there is a possibility for contamination during the cooling phase.^[9]

The formation of alcohol during the fermentation process is necessary for the production of acetic acid, and while the alcohol content of kombucha is usually under 1% after fermentation,^[9] excessive fermentation for one month has been noted to raise it up to 3%;^[10] commercial products tend to have less than 0.5% alcohol content (to avoid being mandated as an alcohol-containing product).^[9]

Overfermentation of kombucha during the standard 7-10 day period is possible if not refrigerated soon after, which may cause an elevation of acetic acid levels above what is desirable.^[9] Acetic acid has the potential to leech metals from the container in which kombucha is fermented, so care should be taken to ferment kombucha in nonmetallic containers.^[9]

Improper processing of kombucha, either contamination or by needlessly prolonging the fermentation period, is known to cause bacterial and fungal overgrowth and has been noted to covert kombucha into a toxic food product.

The brewing of kombucha from teas of Camellia sinensis (green or black) results in a pH around 5, which can be reduced to around 2.5 (variable between 2.3-2.8) after one week.^[9][7] The increase in acidity (occurring within one day of fermentation^[1]) is due to production of organic acids during bacterial fermentation^[9] (although a perfect correlation does not exist between pH and organic acid content, thought to be due to some buffering agents in the medium^[7]). This is necessary for proper fermenting since it, as well as antimicrobial metabolites produced from the tea, are thought to prevent competing bacterial and fungal strains from contaminating the final product.^[1]

The pH of the final product is increased (acidity reduced) after 12 days, perhaps explaining why traditional fermentation is halted at around this time.^[2] It is also around this time that sucrose, continually produced into increasing quantities of fructose and glucose, reaches peak levels where afterwards they decline.^[7]

The standard processing of kombucha starts with boiling of water and adding both the tea and sugar to steep for 10 minutes, although unlike other teas which would be drunk at this stage, manufacturing kombucha requires that the tea leaves be removed and an inoculum (bacteria and fungi which would conduct the fermentation) be added. This is then left to ferment at room temperature over the course of 7-10 days and then refrigerated.^[9]

The yeast tends to proliferate after two to four days fermentation when the pH drops, with levels of yeast in the pellicle (removed from final product) spiking after four days and remaining steady until the end of standard fermentation (10 days), where a slight decline is seen.^[1]

If not consumed locally, kombucha is then packaged with additional measures taken to prevent excessive microbial growth (pasteurization or addition of sodium benzoate and potassium sorbate, for example).^[9]

It has been hypothesized that kombucha can increase glucuronidation in the body after ingestion, either directly by providing dietary glucuronic acid^[7] or secondary to inhibiting the β-Glucuronidase enzyme (which hydrolyzes the bond between glucuronide and its conjugation target).^[6] D-saccharic acid 1,4-lactone (saccharolactone) is a competitive inhibitor of β-Glucuronidase with an IC₅₀ of 3.6µM^[11] and exerts complete inhibition at 1mM.^[12] Fecal β-Glucuronidase is inhibited in both healthy controls and in subjects with colon cancer (who have elevated β-Glucuronidase concentrations^[13]) at a concentration of 30-150µg/mL.^[14]

The inhibition of β-Glucuronidase and supposed augmentation of glucuronic acid's conjugation ability seen with saccharolactone is thought to also underly the 'detoxifying' anticancer properties of kombucha by facilitating the elimination of toxic substances from the body,^[15] similar to the mechanism of calcium-D-glucarate.

The 'detoxifying' properties of kombucha refer to the ability of some acids produced during the fermentation process to increase glucuronidation in the human body, which is involved in eliminating some drugs and xenobiotics from the body by conjugating them.

When tested in vitro in lymphocytes exposed to γ-radiation, kombucha at 250-1000µL in whole blood samples prior to irradiation appeared to dose-dependently preserve lymphocyte chromosomal structure, reaching approximately 50% preservation relative to control.^[16] Kombucha at 1000µL by itself did not alter lymphocyte structure without any irradiation relative to control.^[16]

The antioxidative properties of kombucha appear to preserve white blood cell integrity in vitro when exposed to radiation, which is an expected effect from antioxidant compounds; practical significance of this information is not known.

A study of male rats examined the protective effects of black tea (from Camellia sinensis) or kombucha prepared from the aforementioned batch of black tea to CCl₄-induced hepatotoxicity; it found that doses of both black tea and kombucha of 2.5mL/kg for 30 days prior to (preventative) or alongside (curative) induced hepatotoxicity were protective, as assessed by liver enzymes and malondialdehyde in the liver, but the reductions noted with black tea (50-74% in preventative and 61-65% in curative) were less than kombucha (75-83% and 70-76%, respectively).^[4]

Protective effects from kombucha have been noted elsewhere in rats against acetominophen-induced liver toxicity^[17] and isolated liver cells subject to oxidative death via TBHP,^[18] thought to be due to the D-Saccharic acid 1,4-lactone content of the tea which may function through antioxidation or by increasing the glucuronidation and elimination of toxic substances through inhibiting β-glucuronidase.^[17] This substance alone is also known to exert hepatoprotective effects.^[19]

Kombucha, at least when given to rodents, appears to be beneficial in reducing toxicity of known stressors on the liver. This is likely due to the saccharolactone content, and is thought to be due to either antioxidative mechanisms or increased glucuronidation of toxins (perhaps a combination of both).

While the protective effects are thought to be due to a combination of antioxidative mechanisms plus the potential increase in glucuronidation of the toxins via saccharolactone, the toxic effects of kombucha (thought to be due to improper preparation) can manifest themselves as hepatotoxicity^[20] or gastrointestinal toxicity.^[21]

When kombucha is improperly processed, the potential benefits are lost and ingestion will result in hepatotoxicity instead of hepatoprotection.

Kombucha in general appears to be associated with numerous case studies where the subject was harmed. These instances include an increase in oral intake from 4oz to 14oz in a person making kombucha at home (this person may have had a predisposition to acidosis), resulting in death.^[9] Death has been reported in other instances,^[22] and there exist numerous cases of nonlethal hepatoxicity,^[23][20] instances of gastrointestinal toxicity with and without jaundice,^[21] cutaneous anthrax,^[23] unspecified acute illness (resulting in hospitalization),^[22] and acute renal failure.^[24]

Based on these case studies, it has been suggested to limit daily kombucha ingestion to 4oz (125mL)^[9] or to omit it fully due to the risk of contamination from nonsterile production.^[25]

Kombucha tea can be produced safely, but even then the recommended intake of safely-produced tea is still quite low (half of a metric cup); such a low dose may preclude the health benefits seen in rat studies and expected to occur with the D-Saccharic acid 1,4-lactone content. Kombucha also carries the possibility to exert a wide variety of negative effects due to mishandling of the fungal and bacterial strains used in its production.