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Kombucha is a fermented beverage made from green or black tea, sugar, and a polymicrobial community consisting of both yeast and bacteria. This community has one of the best acronyms for a microbial community and is known as a SCOBY (symbiotic culture of bacteria and yeast). The composition of this SCOBY varies depending on many environmental factors, which in turn can influence the fermentation process as well as the flavor profile of a finished product. That is why we take our partnership with our kombucha clients very seriously.

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Health Benefits

For probiotics to provide consumers with improved gut health, they must be alive and in sufficient numbers, to deliver its probiotic effects. However, putting a specific volume of probiotics in your product does not guarantee the number of live probiotics in the finished product. That is because some of these organisms will not survive.

While it is essential to know the probiotic concentrations as your product leaves the facility, this is only a snapshot in time. Over the course of your products shelf-life, the microorganism(s) populations may change due to storage conditions. Twin Arbor labs has your solution to this dilemma by utilizing our probiotic shelf-life study testing services. Please see our microbial services for more information.

Image by Anshu A
Image by Anshu A
How is it made?

The overall fermentation process is similar to vinegar fermentation in that the organic acids produced contribute to the vinegary flavor profile; however, kombucha is unique in that it is produced through a double fermentation known as primary and secondary fermentation.


Generally, the process includes inoculating a sweetened tea with the SCOBY from a previous batch. It is then covered with a clean porous cloth and incubated at room temperature for about 7–10 days. Primary fermentation begins wherein the yeast begin to consume the added sugar and produce ethanol and CO2 as byproducts. The ethanol is converted into organic acids by bacteria while the CO2 escapes to the environment through the porous cloth cover. 

Once the primary fermentation is complete the resulting product flavor profile is often found to be plain and flat. The SCOBY is removed and the primary fermentation product is transferred to a final air-tight container. The addition of a new sugar source (often in the form fruit juice) allows the yeast to begin a secondary fermentation over the course of 1-10 days. Because CO2 is not allowed to escape, this secondary fermentation provides a natural carbonation in the final product.

Traditional kombucha typically is a lightly carbonated, slightly sour, and refreshing drink, which is composed of several acids, 14 amino acids, vitamins, and some hydrolytic enzymes. With consumer demand increasing kombucha manufactures have begun to expand the consumer options by creating bold new flavor profiles with the additions of botanical extracts or herbs and spices.

Here is one of our favorite Kombucha review papers.

Safety Concerns

Exceeding ethanol (ABV) limits in non-alcoholic kombucha:

As the tea fermentation industries continue to grow, quality is becoming the distinguishing characteristic of a successful kombucha brewer. Preventive process controls must be established to ensure food safety of the finished product. For a kombucha to be considered non-alcoholic, manufactures must ensure their product is below 0.5% ABV. You could estimate the ethanol with theoretical equations OR you can send your product to Twin Arbor Labs to ensure accurate ABV readings. We utilize a headspace gas chromatography method which is recommended by Kombucha Brewers International (KBI).


The consumption of large amounts of kombucha has been linked to a number of serious illnesses, including metabolic acidosis and liver damage. Additionally, there have been instances of food poisoning or other infections from contaminated home brews. The pasteurization of commercial brews limits this risk significantly, though the process renders any probiotic properties of the drink inert. As such, kombucha is not recommended for people with compromised immune systems.

Physical (bottling issues):

Bottling an actively fermenting kombucha beverage is considered a physical hazard that needs to be carefully controlled. CO2 can build up inside the container causing pressure to exceed the ability of the container to hold it. Leakage or breakage is the results with bottles and/or caps, forming projectile hazards. There are two main mitigation strategies for this hazard.


  1. The final product can be filtered or sterilized to reduce the amount of active yeast. Unfortunately this also reduces the natural beneficial probiotic bacteria. In these instances, starter probiotic cultures, such as Bacillus coagulans, can be reintroduced to the final product as a post-production ingredient. Utilizing Twin Arbor Labs qPCR services, you can verify that your starter culture is not contaminated, accurately quantify additions of Bacillus coagulans to your final product, as well as test its stability over the shelf-life.

  2. Alternatively, to slow secondary fermentation, the product can be stored under refrigerated conditions during the entirety of its shelf-life. However, there are some strains of yeast (Saccharomyces bayanus, Saccharomyces  pastorianus, and Saccharomyces uvarum) that are known as "cold fermenters" and can continue secondary fermentation under refrigeration until explosive pressures are reached. If you utilize this mitigation technique, we recommend our qPCR assays specifically targeting these troublesome cold fermenting yeast.

Suggested Analytical Services

  • Alcohol by volume (ABV)

  • pH

    • Generally, the kombucha operational target for pH is 2.5, and the pH will continue to lower as the Kombucha is allowed to continue fermenting.

    • pH must be measured until the time of sale, serving or bottling. pH below 2.5 is unsafe for consumption.  Kombucha with a pH below 2.5 or that tastes especially acidic should not be sold to consumers.

    • A fresh brewed tea or distilled water can be used to dilute the high acidity until pH is greater than 2.5, but should never be higher than pH 4.2.

  • Total acidity

  • Total sugar (glucose, fructose, sucrose)

  • Caffeine

  • Fingerprint of volatile organic compounds

Suggested Microbial Services

  • qPCR analyses:

    • Track the microbial communities’ composition during pre and post fermentation, or the final product.

    • In addition to screening the liquid portion of the kombucha, we can also quantify microbes present in the SCOBY, tea, and starter cultures.

    • This system can also be used to verify that new processes, such as filtration or sterilization, are effective by screening the product before and after.

  • Probiotic shelf-life studies:

    • This reveals the stability of the probiotics in your product. In other words, it tells you how well the organisms are surviving over the life of your product.

    • The data produced provides the manufacture with the length of time until the product no longer has a sufficient concentration of live organisms to meet product specifications. This information is important for supporting label claims as well as determining expiration and best by dates.

    • In addition, other analytical tests can be added to the stability study that provide insight into whether decomposition of flavor compounds is occurring, or whether heavy metals are present due to leaching or corrosion of the packaging material.

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