Probiotics need to be alive – their efficacy is based on live microorganisms conferring health benefits. The issues for their inclusion in any food are well recognised because of the uncertainty around their viability and shelf-life. Postbiotics do not seem or need this reliance because they are based on non-living material (inanimate cells and/or components) derived from probiotics. Better understanding into the mechanisms by which microorganisms alter one’s microbiome has led to increased interest from product developers. They are eager to exploit the opportunities that a ‘ new news’ ingredient will bring.
Postbiotic health benefits have been demonstrated in the research canon. The reports on their properties have steadily increased in the scientific literature over the past five years, albeit without a consensus definition of the term. Although this has been addressed there is still much debate and it is not yet recognised as such by national bodies. The healthiness of postbiotics include but are not limited to modulation of immune responses and systemic metabolism, improved epithelial barrier function, enhanced cognitive function via nervous system signaling, and amelioration of symptoms derived from irritable bowel and other gut dysfunction syndromes.
What are Postbiotics?
It helps to do a quick recap on the other terms we know. Probiotics are those microorganisms that we ingest and do us good by fermenting our food and turning it into useful componentry. Prebiotics are what probiotics and our gut bacteria live on and serve as the raw material for all those beneficial microorganisms. It includes materials such as dietary fibre.
Postbiotics are actually all the good healthy components that our gut microorganisms produce. It includes metabolites such as short-chain fatty acids (SCFAs), broken down protein and peptides etc. To all intents and purposes it is waste material from a microbial gut fermentation but the term postbiotics is reserved for all the really beneficial material.
The term ‘postbiotics’ is new to many of us. Whilst this term seems to be very recent the components such as ingredients and products that make it up have been researched and discussed for many years. Back in 1998 the health benefits of cultured milk products using living and non-living bacteria were being discussed (Ouwehand & Salminen, 1998). That article was identifying the very components in fermented milk which would start to be described as postbiotics.
Why should we be interested in these so-called postbiotics? It all stems from the EU novel food regulations to some extent where this body would prefer any novel microbes with notable health benefits to be in an inactive form for the purposes of an effective safety assessment.
It is well established that postbiotics are getting better known and amongst the health food fraternity, its popularity is rising. The Japanese have for many years enjoyed products containing non-viable lactic acid bacteria which are now recognised for their benefits in other markets.
Unfortunately, given the nature of unscrupulous operatives, there is probably and deliberate misuse of the term besides genuine misinterpretation of what these ingredients are. It has created confusion when there is no need for it.
In 2019, the International Scientific Association for Probiotics and Prebiotics (ISAPP) formed a multidisciplinary consensus group and attempted to produce a concise and modern definition for postbiotics. They had earlier developed definitions for probiotics, prebiotics and synbiotics so they were naturally placed to develop this next definition. Accordingly, such a definition had to meet both a regulatory, safety and science-based framework so that removed confusion in the nomenclature. It should also help the general public in their understanding.
ISAPP is also providing guidance to various stakeholders as well as customers and are suggesting they use the Nature Journal’s definition of a biotic on a formal basis. They have also published various findings in Nature Reviews in Gastroenterology and Hepatology (Salminen et al., 2021) and an open access article in Foods (2022).
A response paper to the ISAPP definition refuted their findings. This was published in Nature Reviews in Gastroenterology and Hepatology. It was titled ” when simplification fails to clarify” and they refuted (Aguilar-Toala et al., 2021) the ISAPP definition and reaffirmed their viewpoint that the previous definition from a 2013 paper was still the most appropriate. Aguilar-Toala et al., (2021) say “Any factor resulting from the metabolic activity of a probiotic or any released molecule capable of conferring beneficial effects to the host in a direct or indirect way“.
Both publications give an up to date perspective of where the scientific community currently stands with the use of that word.
Alternative Names For Postbiotics
One of the confusing issues is the number of different names for postbiotics. We have parabiotics, paraprobiotics, modified probiotics, non-viable probiotics, inactivated probiotics and even ghostbiotics! Other terms include peribiotics which are just intact and activated cells. Psychobiotics – confer specifically mental health benefits through the gut-brain access. Another term with wide spread use is synbiotic which covers the synergistic effects of probiotics and prebiotics in a combination. New names are constantly being derived from the probiotic or biotic space and each of which could be considered a postbiotic.
You can see in the research literature how many different names there are: Lew & Liong, 2013; Kolling et al., 2015; Bajpai et al., 2018; Deshpande et al., 2018; Piqué et al., 2019; Wegh et al., 2019). They all refer to the same type of product but you can see how it would cause a muddle.
Currently the best approach is to use the term postbiotics with this consensus definition:-
“Preparation of inanimate microorganisms and/or their components that confers a health benefit on the target host“.
It certainly helps the consumer and research scientist because there has been a 1,300% increase in searches on postbiotics and the targets of postbiotics. However, whilst it is a consensus there is still considerable antipathy to this definition and some as we’ve already mentioned do not accept the consensus definition.
What are we referring to as postbiotic products?
So these are essentially products derived from killed microbes, from probiotics and from food fermentations.
The current list is now:-
- non-viable microorganisms
- SCFAs
- Microbial cell fractions
- functional proteins
- extracellular polysaccharides
- cell lysates
- peptidoglycan-derived muropeptides
- cell wall & membranes
- pili-type structures
- nucleic acids
- bacteriocins and other anti-microbials
What Type Of Foods Contain Postbiotics?
If you think of the foods that contain probiotics then they will most certainly contain postbiotics. If they have been heat treated then they should contain non-viable microbes along with their cellular components and metabolites.
Get postbiotics by eating the following:
- Yogurt containing live and active cultures
- Sauerkraut
- kefir
- kombucha
- kimchi
- tempeh
- miso
- sourdough
These types of food are familiar to many so selling the idea will not prove too much of a stretch. Japan is a good place to start for brands and products supporting postbiotics. You can find ice cream, popcorn (M-1), natto (S-903), instant miso soup, pancake powder (M-1), chewing tablets, beverages with inactivated Lactobacillus gasseri CP2305 and heat-killed L. paracasei K71. That’s quite a broad range of foods which suggests to product developers that using postbiotics should not be an issue.
Quality and Efficacy of Postbiotics
Redefining Efficacy
Unlike probiotics, postbiotics are not tied to the colony forming unit (CFU) which is the gold standard way to characterise microorganism potency. Limitations on how CFU is measured include intrinsic variability of the methods and the degree of specificity. Evaluating postbiotics for efficacy present a unique opportunity to redefine this aspect as well as measure quantity, using more sophisticated techniques.
Traditional probiotic potency methods are not applicable. Characterisation of efficacy is based on the microorganism used as the starting material, how that microorganism is inactivated since different processes can lead to different postbiotic compositions, and a description and quantification of the final postbiotic composition itself.
Inactivation processes can include but are not limited to high heat, high pressure, and exposure time to oxygen for strict anaerobes. Since the composition of the postbiotic can change depending on the inactivation process, its is imperative manufacturers keep this process consistent from batch to batch and the same as that used for any clinical trials demonstrating a health benefit. That’s especially so of the analyte being measured for efficacy that is performed pre-processing.
The question then becomes how to measure or quantify the inactivated materials and its components. Methods used to evaluate postbiotic content need to be scientifically valid, reliable, repeatable and accessible. These characteristics are imperative to proper quality control, and to facilitate replication in research.
Efficacy depends then on processing and the type of molecule under investigation.
How to measure efficacy?
No of inactivated cells: You can use flow cytometry, digital PCR, direct cell counting – these offer benefits as long as the cells remain roughly intact.
Benefits of flow cytometry include real time quantification, distinguishing live, injured and dead cells, based on cellular characteristics such as membrane integrity as well as better repeatability and precision compared to traditional plate counts. Limitations include the fact that the representative quantity is an active or non-active fluorescing unit, and not a CFU, and that there isn’t a singular correlation between the two values. It also poses issues with accuracy if the cells are not completely broken down into cell fragments which may skew results to be higher than they should be due to a single cell being counted as multiple AFUs. Perhaps the CFU levels of the microorganisms before processing can be used to determine efficacy of the final product.
Quantitative or digital droplet PCR could prove useful for evaluating postbiotics – the advantages include specificity of target DNA, but the need for optimization could prove this technology to be too limiting for wide range use. Quantification of postbiotics could also rely on direct microscopic observation assuming complete inactivation or with the use of the enzymatic dye such as Trypan Blue to confirm cell death.
Other molecular dependent analyte measurements that are more chemical in nature include enzyme-linked immunoassay approaches. These could be more suited to certain postbiotic materials depending on what cellular component is deemed to confer the health benefit. It could be as simple as measuring the efficacious dose by weight in milligrams.
A specific example has been to use a metabolomics approach based on untargeted LC-QTOF-MS/MS (liquid chromatography time-of-flight mass spectrometry) to reveal the bioactive components in probiotic fermented coffee brews (Chan et al., 2021). These were coffee brews fermented with L. rhamnosus GG, and/or S. boulardii CNCM-I745. The untargeted approach was coupled with multivariate analyses which showed 37 different metabolites. The postbiotics produced by bacteria were aromatic amino acid catabolites such as indole-3-lactate, p-hydroxyphenyllactate, 3-phenyllactate whilst yeast were producing isopropylmalic acid and fatty acids. The precise approach relied on good fermentation design for both bacteria and yeast in a coffee brew versus appropriate controls. The products were analysed using LC-QTOF-MS/MS followed by data acquisition, pathway analysis to understand where the postbiotics were generated in the metabolic cycle, statistical analysis and then links to the therapeutic benefits.
Health Benefits of Postbiotics
For many of us, postbiotics are compounds which could have a wide variety of potential benefits. If you think how good other-biotics happen to be than you can identify them with reducing inflammation, modifying and stimulating the immune system, reducing cholesterol as in their hypocholesterolemic effect and having antioxidant benefits.
ISAPP described 5 different mechanisms of action (International Probiotics):
- Modulation of resident microbiota
- Enhancement of epithelial barrier functions in the intestine
- Modulation of local and systemic immune responses
- Modulation of systemic metabolic responses
- Systemic signaling via the nervous system
The main focus is the production of short-chain fatty acids (SCFAs) and these have been known for many years to be associated with good gut health. The SCFAs are acetate, propionate and especially butyrate.
Potential Reduction in Gut Inflammation
One of the main health issues of our time is chronic inflammation as opposed to acute inflammation. It’s linked to other conditions too like type 2 diabetes and metabolic syndrome, cancer and obesity.
The generation of SCFAs especially propionate is linked to lowering inflammation as well as possibly reducing the risk of developing cancer in the gut (Zolkiewicz et al., 2020). One effect these agents as well as other materials are potentially having is raising the acidity level of the stomach which not only offers buffering support but also causes the stomach lining in particular to produce more mucus. Such mucus protects the stomach from ulcers and digestive issues.
Recent research shows that postbiotics can reduce inflammation and potentially in the long-term too (Jastrzab et al., 2021). The SCFAs are probably not the only agents helping to reduce inflammation.
In a more generic sense, it is already known that many preparations of live bacteria and yeast which were used as probiotics for their gut health benefits also contain a mix of postbiotic materials too. These however have been overlooked in the past until the term ‘postbiotic’ came to prominence.
Take the yeast Saccharomyces cerevisiae var. boulardii, the only yeast with a probiotic status. It has long been used in a live form to ameliorate diarrhoeal conditions (e.g. antibiotic associated, travellers’ diarrhoea, acute), irritable bowel syndrome, and Helicobacter pylori infection.
Obesity
It’s now recognised that obesity can occur when the intestines lose their microbial balance. Muramyl dipeptide is a postbiotic component which relieves glucose intolerance by increasing insulin sensitivity. It implies that postbiotics could reduce the risk of obesity.
Healthy Bowel Movements
Postbiotics can reduce the growth of damaging bacteria in the gut, especially in the colon. They are linked to the production of a toxic gas called hydrogen sulphide which is naturally produced by bacteria. It seems farfetched but it is thought that this gas at particular levels kills various cells including cancer cells as well as reducing inflammation.
Inflammatory Bowel Conditions
A host of inflammatory bowel conditions could be managed using postbiotics. There is still plenty of research ongoing but because one of the SCFAs butyrate is linked to better health in the gut, it possibly helps in reducing really damaging conditions like ulcerative colitis as well as inflammatory bowel syndrome (IBS).
Treating Diarrhoea
Postbiotics could prevent or treat diarrhoea. A study in children especially showed that these supplements shorten the length of diarrhoea.
Cancer
The effects of different components such as culture supernatants, cytoplasmic extracts, cell-wall extracts and live cells of Lactobacillus gasseri and Lactobacillus crispatus on proliferative and apoptotic responses of normal and tumour cervical cells have been reported by Motevaseli et al. (2013). The exopolysaccharide from L. plantarum 70,180 has been reported to exert antitumor activity against colon carcinoma cells (Wang et al., 2014).
Postbiotics and Immunity
The gut is highly influential in supporting the immune response. A number of physicians are of the view that a healthy gut is directly linked to gut immunity and our ability to ward off diseases. Again, SCFAs are linked to this phenomenon because of the role they play is reducing inflammation as well as reducing the growth of pathogens (Yoo et al., 2020).
The Safety of Postbiotics
Postbiotics should be regarded as a food or dietary ingredient much like any other functional ingredient that you would find in fortified foods and dietary supplements. The same regulations on food ingredient safety such as toxicology etc. apply as they do with similar ingredients depending on the format. This will differ from country to country.
For many businesses, cell-free supernatants are preferred rather than formulations using dead cells (Moradi et al., 2020). This is often from the perspective of them servicing as antimicrobials with food applications in food biopreservation, for food packaging, to cause biodegradation of contaminants and to control biofilms.
It can be inferred that postbiotics would have a better safety profile than probiotics since there is no longer a need to consider antibiotic resistance genes and virulence factors. Considering only postbiotics for a food rather than postbiotics means eliminating exposure to live microorganisms even if those risks are rare because they are now removed from the product. It means there could be treatment for susceptible populations such as neonates, the very ill and the elderly. Some examples of this benefit compared to probiotics include maintaining colonic health (Konstantinov et al., 2013).
We are of course already familiar with particular postbiotics such as nisin. This antimicrobial and antibiotic has been explored in its interactions with specific ingredients and food matrices (Younes et al., 2017). As a converse argument, there are postbiotics such as EPS (exopolysaccharides) that can protect pathogens from environmental stress such as changes in pH and temperature and encourage pathogenic (microbial) proliferation. It’s feasible that a food ingredients have a relationship with active postbiotic metabolites and in turn would damage the performance of metabolites that are meant to be inhibitory to pathogenic growth (Hartmann et al., 2011).
Some inferences about postbiotic performance can be gleaned from research on feeding studies with animals. One typical example was an investigation of a yeast cell wall blend (PYCW) fed to newly-weaned pigs to reduce the impact of mycotoxins. These mycotoxins included aflatoxin B1 and deoxynivalenol. It appears that feeding yeast cell wall in this instance did limit the damage such mycotoxins do to animals (Holanda et al., 2020).
The traditional measurement of safety for these ingredients, both chemical and microbiological, is applicable to this category of ingredient as with any other functional ingredient. We must however not assume safety based on the inanimate nature of the ingredient. Safety assessments will always be required for the intended use of the product prior to first use in the market.
The Mechanistic Action of Postbiotics
A number of suppliers are beginning to offer postbiotics by removing the live bacteria from the mix and leaving all those healthy compounds. Check out Cargill and ADM on their approaches to supply and marketing.
One of the issues is overhyping the concept. At the moment the research is truly promising but there are no claims for these ingredients just as there are no claims for probiotics. Much of the research is conducted in ‘in vitro‘ systems with no explicit human clinical studies to speak of.
The ingredients business, ADM offer probiotic strains including BPL-1 which is linked to reducing visceral fat. If you heat treat the microorganisms then it inactivates it but offers roughly the same benefits as the live microorganism. One benefit is that it is probably easier to sell the concept as a non-living ingredient. It will remain stable and not be altered further because it is no longer living and this relatively easier to work with. There are shelf-life issues when it comes to proving the presence of live bacteria at the end of shelf-life but this aspect is taken out of the equation.
In the USA, heat-treated BPL-1 has GRAS status. The material is associated with an important fatty acid known as lipoteichoic acid (LTA) which has specific cell signaling properties. The biochemistry is only just being worked out.
Cargill have a yeast fermentate called Epichor® which also has a general wellness function. The material is produced from a specialized anaerobic fermentation process using Saccharomyces cerevisiae. The material is also known as Saccharomyces cerevisiae fermentation product (SCFP). It is comprised of a unique collection of metabolites including proteins, peptides, antioxidants, polyphenols, organic acids, nucleotides, polysaccharides (beta-1,3/1,6-glucans) and mannans. The ingredient has immunity, gut and microbiome health benefits. It is stated not to cause bloating.
The same type of material has been used as an additive to lower Salmonella enterica in broiler chickens (Chaney et al., 2022). This is called Original XPC™ (Diamond V, Cedar Rapids, IA, USA). In vitro gut fermentation models have associated SCFP with the modulation of the microflora composition in a manner to synergistically reduce Salmonella Typhimurium and Campylobacter jejuni concentrations. An SCFP called Olimond BB has been shown to improve the efficacy of the influenza vaccine in racehorses (Lucassen et al., 2021).
LAC-Shield is heat-killed Lacticaseibacillus paracasei MCC1849 from Morinaga Milk Industry Co., Ltd. which has clinically proven activity in enhancing immune activity. This product induces the production of interleukin-12 (IL-12) which is a potent activator of both innate and adaptive immunity. According to Morinaga’s white paper this material boosts the natural defence system of IgA production. When orally administered it is said to be recognised by antigen-presenting cells including the dendritic cells found in Peyer’s patches where it induces IL-12 production.
In terms of a high lipolytic activity the bacteria Lactobacillus paracasei is a good organism for countering metabolic syndrome. The postbiotic benefit comes from components in the mix influencing lipid metabolism based on the lipolytic action. This is by limiting the formation of complex lipid forms or indirectly through influencing the activity of peroxisome proliferator-activated receptor α (PPARα), which plays a crucial role in controlling lipid metabolism (Marra & Svengliati-Baroni, 2018).
Urolithin A is produced by bacteria in the gut. This material comes from conversion of ellagic acid and ellagitannins found in particular fruits such as pomegranates and berries. It is the first postbiotic that affects mitophagy. This is the process whereby damaged mitochondria are removed and recycled into new cell organelles and components. It is known that a damaged mitophagy process is part of many age-related processes involved in cardiovascular disease, cancer and neurodegenerative diseases. If mitophagy is supported there may be a positive role in aging and the amelioration of chronic illnesses.
Postbiotic Applications and Advantages
Unlike probiotics, which relies on live microorganisms that are susceptible to heat and oxygen, postbiotics are extremely stable at room temperature, making storage and shipping less challenging, and less expensive.
Probiotic manufacturers will formulate with significant overages so as to have an efficacious dose at the end of shelf-life assuming degradation over time. Postbiotic applications limit this requirement. Consideration also needs to be taken over the fact that even those microorganisms that have naturally degraded over time may actually possess a health benefit, which may redefine probiotic efficacy in and of itself, because it is currently only based on viable cells.
Structure-function claims and probiotic clinical trials can be attributed to specific cellular components that may be present in the material from the dead cells available. So do the cells need to be viable at the end of shelf-life to pose a health benefit or can it be a total count? Both the live and the dead cells will define efficacy. We understand a different approach to quantification will be required if the industry shifts to this change in understanding.
Lacking live MOS postbiotics may make it easier for product developers to maintain ownership under intellectual property laws, since these materials are not tied to a specific bacteria strain. The strain with which the postbiotic materials is derived cannot be isolated from the commercial product. This being said, stating the microorganisms that was used to produce the postbiotic will be essential for scientific reproducibility, in order to progress the developing field. Functional foods and beverages currently fortified with traditional lactic acid bacterial probiotic need to be in a condition such that the microorganisms remain viable. This severely limits the formats with which probiotics can be applied. Postbiotics on the other hand, are similar to probiotic bacterial spore formers can be added to functional foods and beverages that cannot support traditional probiotic efficacy. These include foods that require heat or other types of processing not conducive to bacterial survivability, low pH materials, products with a high moisture content, an unfavourable probiotic conditions, and products with ambient storage conditions.
The use of postbiotics as an alternative therapeutic agent to probiotics can also expand the geographical scope of where the fortified products can be consumed, such as regions with poor and unreliable cold chains, or whose ambient temperature conditions causes problems for storage of traditional probiotic microorganisms. To date the USA FDA is yet to address postbiotics. When it does address the use of postbiotics, they will likely base regulations on the specific category chosen for a product under development and the use, efficacy and safety of the product will need to meet the standard for the applicable regulatory category.
If the postbiotic is to be used as food ingredient it will need to be determined whether the ingredient is GRAS and/or undergo pre-market approval as a food additive. Health claims will be treated similarly to other functional ingredients where health claims will need to be identified as the ability to reduce the risk of disease or as a non-approved structure-function claim where the food can influence the normal structure of function of the human body.
Other categories that postbiotics will inevitably fall under drugs and medical devices, or subcategories of food, such as dietary supplements, infant formula foods for special dietary use and medical foods. Postbiotics can be used in alternative microorganism applications in susceptible populations.
In summary, it goes without saying that postbiotics are a new and exciting ingredient in the food and supplement industry with a number of advantages:
- the health benefits are similar to probiotics, but without the need to keep them alive
- it’s easier to maintain ownership under current intellectual property laws since they are not tied to any one specific bacterial strain
- an ability to add to a food or beverage considered inappropriate for use with probiotics
- expands functional food marketplace in scope and geography
- meets regulatory considerations in the USA
- alternative “-biotic” for susceptible populations.
Revision: This article has been revised with more extensive examination of the definition of postbiotics and additional material on the conditions to be treated from 14th December 2020.
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