What happens inside Kombucha Vinegar?

How to Hack Your Brew – adjusting sugar, fructose, alcohol, and acidity by changing the brew time.

A detailed look at what happens with acidity and sugar levels in kombucha when brewed a long time. kombucha research. kombucha science. fructose.

Have you ever wondered what goes on inside your batch of kombucha when you leave it too long? What about when you’re deliberately making kombucha vinegar? What are the changes that happen?

Or maybe you’re trying to hack your kombucha brew to get a particular effect. If you want to minimize the sugar, how long should you leave it? What happens to the sugar? How much alcohol gets produced and when? How acidic does it get and when? How much fructose gets produced?

Well, you are not alone in asking those questions!

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This month’s research article was published in 2000 by C. Chen and B.Y. Liu from the Department of Food Science, National Chung-Hsing University, Taiwan.

Changes in major components of tea fungus metabolites during prolonged fermentation. Journal of Applied Microbiology 2000, 89, 834-839

When these two scientists were working on this project, nobody had studied kombucha during a long fermentation. All the science that had been done up until this date had been done on the standard instructions of fermenting it for 5 to 10 days.

So, they wondered. What does happen if we leave it for longer? What about 60 days?

(If you’re not that interested in reading all about the experiment and just want to know how to maximize your alcohol or minimize your sugar, for instance, scroll down to the So What? section, below 😉 )

How to hack your brew. Adjust sugar, acidity and alcohol levels in your kombucha. Kombucha research. Kombucha science.What they wanted to find out

How does microbial activity change through an extended fermentation?

In other words, how do the yeasts and bacteria grow and change when the kombucha is left for 60 days?

How do the metabolites change through a long fermentation?

In other words, what happens with the chemicals and substances that the bacteria and yeast ferment?

In this experiment, they measured sugar, alcohol, overall acidity, and the amounts of acetic and gluconic acids.

Read this post for a discussion about the best sugar to use for kombucha.

What they already knew

  • Acetic acid, alcohol and gluconic acid are present in measurable quantities in kombucha
  • Standard fermentation had been studied by others, but nobody had looked at longer fermentation times
  • Different kombucha brews can differ greatly from each other, depending on the source culture and the fermentation conditions

What they did

They sourced scobys from 9 different households scattered throughout Taiwan

Using a fairly standard recipe they made 8 batches of kombucha from each of the 9 parent scobys. This way they could take 8 different samples at different times without fear of contaminating the remaining cultures.

They tested the samples at days 0, 3, 6, 9, 14, 20, 30, and 60.

They grew cultures of the bacteria and yeast from each sample, in order to count and identify the microbes from both the scoby and from the broth.

At the same time they measured:

  • Ethanol levels
  • Sugar concentration (glucose, sucrose, and fructose)
  • Acidity
  • Gluconic acid amounts
  • Acetic acid amounts

Now, from our 2017 perspective, the glaring omission here is that they didn’t test for lactic acid bacteria or lactic acid amounts.

Presumably, it just didn’t occur to them to test for it, since the research into the benefits of lactic acid bacteria for humans didn’t really get underway for a few more years. This paper that I found, from 2006, says “no specific health or nutritional claims can yet be made for the lactic acid bacteria.”

So. I’m pretty sure that were this research being conducted today, lactic acid and lactic acid bacteria would be on the list.

What they found

Numbers of living cells hit maximum levels at 6 to 14 days (differed depending on the culture).The numbers of living cells remained high after this point, but there began to be a slight die off near the 60-day mark.

Figure from the research paper. Sugar and ethanol concentrations. Open circles are sucrose, triangles are glucose, squares are fructose, and closed circles are ethanol.

Sugar concentration decreased linearly for 30 days, then continued to decrease, but slowly.

Fructose accumulated in the broth. Glucose was further metabolized into gluconic acid and cellulose, so did not accumulate to the same levels as fructose.

Ethanol concentration steadily increased until day 14, where it held steady for another week to 10 days, decreasing after that and returning to nearly zero by 60 days. The maximum average concentration of ethanol was 0.55% but ranged from 0.4% and 0.7% in different samples. The maximum ethanol concentration never reached even 1%.

Acidity increased steadily throughout fermentation.

Figure from the research paper. Acidity and acid concentrations. Open circles are titratable acidity (tartness), squares are gluconic acid, and triangles are acetic acid.

Now, don’t get confused, but when they measured the acidity they didn’t measure the pH. What they measured was the ‘titratable acidity’. There are two very different ways of thinking about and measuring how acidic something is. A good explanation can be found here at E.C. Kraus’ excellent website on winemaking. But basically, what they tested was the ‘tartness’ of the kombucha over time, not necessarily how ‘strong’ the acid was at any point.

They did mention that the average pH was 2.5 after 30 days’ fermentation, but didn’t show this measurement in their results.

Acetic acid levels increased steadily for the first 30 days then slowly decreased over the next 30. Gluconic acid was not detectable until day 6. Then it increased steadily for the rest of the fermentation. Once gluconic acid began to be produced, there was more gluconic acid than acetic acid in every sample taken.

So what?

Okay, so these researchers fermented kombucha for 2 months, and measured some stuff that went on in the brew through that time. What does that actually mean for us as home kombucha brewers?

I’m so glad you asked that 😉


In the first place, if we’re looking to make vinegar, it lets us know that brewing kombucha for 2 months gives us a pretty solid acid. It might not taste as sharp as, say, distilled white vinegar from the store, but the pH is likely to be just as strong. (Remember, they said it got to pH 2.5 after 30 days)

The reason for this is that the dominant acid being produced at the 2-month mark is gluconic acid – not acetic acid, which is the dominant acid in distilled white vinegar – and gluconic acid tastes milder than acetic acid.

I was really interested in this point because a couple of months ago I set up an Apple Cider Kombucha Vinegar brew, just to see how it would turn out. It was mild and delicious and a great addition to salad dressings, but I had questions about how acidic it was. It didn’t taste very sharp or tart at all. My mouth identified it as ‘vinegar’, but it didn’t pucker up like I’d expect from sipping a teaspoon of white vinegar or lemon juice, for example.

Your tongue can only tell you how tart something tastes – it is not a reliable way to test pH

I don’t have any pH strips at home, so I made a mental note to just use the Apple Cider Kombucha Vinegar for salad dressings and other things that don’t rely on the vinegar being strong. I figured I’d pick up some pH strips next time I was at the gardening store, and check out my next batch of vinegar to see what was going on.

I’ll still do that, but I’m feeling much more reassured after reading this research paper. I also followed a bunny-trail of information about how food scientists test for and adjust ‘tartness’ and ‘acidity tastes’ (here, here, and especially here, if you want to read them yourself).

It turns out that there are at least three different things that contribute to a food being identified as an acid by our tongues when we taste it – and the least important of these is its pH!

Which means that if you’re wanting a low-pH acid to use for pickling, for example, the only way to know for sure that your vinegar is strong enough is by testing it with a pH strip, or a pH meter. Your tongue is not a reliable source of information.


Now, how about if you’re wanting to minimize the amount of sugar in your kombucha?

Maybe you’ve got diabetes, or you’ve Quit Sugar. Or you’re following a paleo autoimmune protocol or trying to keep your metabolism in ketosis, perhaps.

This paper demonstrates that by leaving your brew for 30 days, most of the sucrose has been used up. Sure, there’s a little bit left (between 2% and 3%), but leaving it for another 30 days only reduces this to about 1%.That second 30 days also increases the acidity quite a lot, so you’d have to be Very Keen to want to drink it as kombucha at that point.

The tea when you set up the kombucha brew is 9% to 10% sugar. After the standard 6 day fermentation, there is about 8% to 8.5% sugar. 20 days fermentation leaves you with 4% to 5% sugar.

So, pick your comfort level. There’s always a trade-off when you’re looking at sugar levels – too high and it’s not good for you, too low and it’s unpleasant to drink. Only you can decide what your goal is and when you’re happy to drink it.


Now, I’ll just make a quick mention of fructose here. Some people find that fructose is something their gut has real problems with – quite a few people diagnosed with Irritable Bowel Syndrome (IBS) are finding that they really have Fructose Malabsorption Syndrome. Or maybe you’re following a low-FODMAP diet, and want to reduce your fructose intake for that reason.

If this is you, then you should know that fructose gradually builds up in the broth during the fermentation.

Sucrose gets broken into glucose and fructose by the yeast cells. The microbes in the kombucha brew use that glucose to make the new baby scoby, and to make gluconic acid. But they don’t use much, if any, of the fructose. So the fructose levels build up as more and more sucrose is broken down.

According to the Wikipedia entry, it’s the amount of fructose in combination with the amount of glucose that is important, as well as the overall amount of fructose.

“Foods and beverages containing greater than 0.5 g fructose in excess of glucose per 100 g … should be avoided. Foods with >3 g of fructose per serving are termed a ‘high fructose load’ and possibly present a risk of inducing symptoms”

So, let’s have a look at this.

After the standard fermentation of 6 days, fructose is only about 1%, but it reaches 2% to 3% after 30 days and is sitting around 6% after 60 days.

At the same time points, glucose is at less than 1% at 6 days, reaches 1% at 30 days, and stays there for the next 30 days.

Given the guidelines quoted above, then, if you have Fructose Malabsorption, you can probably drink kombucha safely at 6 days fermentation, because there are roughly equal amounts of fructose and glucose then. But by 30 days it’s definitely into ‘high fructose load’ territory, and the levels of glucose are far below what might be able to compensate for it.

More information about Fructose Malabsorption at this great blog, here.


Like all yeast-based fermentations, kombucha produces alcohol.

The amount of alcohol increased for the first 15 days, reaching a high of about 0.4% to 0.7%, depending on which scoby they measured – remember that they used scobys from 9 different households, and each one behaves slightly differently. Alcohol percentages stayed at their high point for about another week before dropping down again. By day 60 the alcohol levels had returned almost to zero.

Now, I have seen other reports of kombucha reaching alcohol percentages of 1.5%, so don’t rely just on this study for information about what is going on in your own brew at home. But the general trend will probably hold true – an increase for two weeks, holding roughly steady for one week, before dropping slowly off and becoming negligible at the two month mark.

Gluconic acid

I particularly want to chat about gluconic acid while we’re here. Mostly to point out that gluconic acid and glucuronic acid are not the same thing.

You might have read somewhere about the detoxifying effects of glucuronic acid, and how useful it is in the body, and about how kombucha has got it. All of which is true. Glucuronic acid is produced by your liver to help it detox things, and it is also found in kombucha.

The jury is out about whether or not it’s a useful thing to actually drink it, but it’s generally seen as one of the candidate molecules for kombucha’s detoxifying effects (as we saw in this blog post of mine, and this other one)

But that’s not the chemical this study is measuring. This study is measuring a cousin molecule, gluconic acid.

Gluconic acid doesn’t appear to have any specific role in the body, and they were measuring it mostly because they knew it was there and would be contributing to the acidity and flavor of the kombucha. It’s a common food additive, and is also found naturally in foods like honey, fruit, and wine. (Read more about gluconic acid’s use in the food industry here)

So. I mention this mostly so that if you’re looking for information about glucuronic acid in kombucha, you don’t get mixed up and look at this study for that. This study says nothing at all about glucuronic acid.

What can we take away from this research?

This research paper gives us a window into what is happening in a kombucha brew when you leave it to brew for longer than the standard 6 to 10 days.

We can now use this information to adjust our brew to suit our own particular dietary desires – whether that involves sugar concentrations, alcohol levels, or fructose amounts.

I also learned something new about vinegars and acids! Now I know that the acidic taste of a vinegar is not really related to how strong the acid is, and I’ll measure the acid with a pH stick if that’s what I want, rather than relying on my taste-buds.


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Remember, Stacey is not a registered health professional of any type, and this does not constitute medical advice in any way 🙂

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Author: stacey

Stacey lives and works in the South Island of New Zealand with her husband, their 4 children, 4 hens, and a rabbit that they secretly think must have watched Monty Python's Holy Grail movie.