Microalgae in Terms of Biomedical Technology: Probiotics, Prebiotics, and Metabiotics

Abstract

Green, red, brown, and diatomic algae, as well as cyanobacteria, have been in the focus of attention of scientists and technologists for over 5 decades. This is due to their importance as efficient and economical producers of food additives, cosmetics, pharmaceuticals, biofertilizers, biofuels, and wastewater bioremediation agents. Recently, the role of microalgae has increasingly been considered in terms of their probiotic function, i.e., of their ability to normalize the functioning of the microbiota of humans and agricultural animals and to produce biologically active substances, including hormones, neurotransmitters, and immunostimulators. A separate brief subsection of the review deals with the potential functions of microalgae with respect to the brain and psyche, i.e., as psychobiotics. Moreover, algal polysaccharides and some other compounds can be broken down to short fragments that will stimulate the development of useful intestinal microorganisms, i.e., function as efficient prebiotics. Finally, many components of microalgal cells and chemical agents produced by them can exert important health-promoting effects per se, which indicates that they are as potentially valuable metabiotics (the term preferred by late Prof. B.A. Shenderov), which are alternatively denoted as postbiotics in the literature.

The present work deals with an interdisciplinary topic that represents an intersection point between ecology, biotechnology, and medicine [1–6]. It is concerned with the potential functions of microalgae in the human body, as probiotics, psychobiotics, prebiotics, and metabiotics. All these functions can be fulfilled by microalgae within the framework of the human (animal) body–microbiota consortium. This consortium is of significant importance in physiological and medical terms and it resembles other multispecies associations that include microalgae as essential components involved in trophic and regulatory interactions within the framework of natural and human-made ecosystems in water bodies and in soil.

Using microalgae in biotechnology. In accord with a somewhat arbitrary classification, algae are subdivided into two main groups: the macro- and the microalgae. While macroalgae are macroscopic multicellular organisms that are up to 65 m in size, microalgae that are considered in this work are microscopic unicellular, colonial, or filamentous organisms; their size is ~1 to ~900 μm. Presumably, about 800 000 species of microalgae exist; at least 50 000 species have been described [7]. The large number of microalgal species is a prerequisite for a wide spectrum of their possible applications. In terms of taxonomy, many microalgae are eukaryotes, including representatives of the kingdom Viridiplantae such as green (division Chlorophyta) and red (division Rhodophyta) algae and of the kingdom Stramenophila (yellow–green algae including Ochrophyta and Prymnesiophyta). Other microalgae are prokaryotes; they belong to cyanobacteria (divisions Nostocales, Oscillatoriales, etc.). Microalgae are of paramount importance as sources of pharmaceuticals, nutraceuticals (e.g., food additives), cosmetics, biofuel, biofertilizers, animal feed, and wastewater treatment agents [5]. All the aforementioned uses of microalgae are related to biotechnology as “industrial application of biological processes and agents on the basis of highly efficient forms of microorganisms and the cultures of plant/animal cells and tissues with desired properties” [8], or, according to an alternative definition of biotechnology, of any technological application that uses biological systems, living organisms, or derivatives thereof, to make or modify products or processes for specific use (https://www.cbd.int/convention/text, see also [9]).

The “work horses” of modern biotechnology (Table 1) include a large number of species of green algae of the genera Chlorella, Dunaliella, Scenedesmus, Haematococcus, Chlamydomonas, Botyococcus, and Chlorococcum that are actually used or potentially applicable in pharmaceutical, dietary, and cosmetic terms. Biotechnology, including its medical subfields, also involves microscopic red algae (division Rhodophyta), e.g., the species of the genus Porphyridium. As for cyanobacteria, widely used producers of valuable compounds ranging from cosmetics and food additives to drugs are of note. They include the species of the genera Arthrospira (e.g., A. platensis, the outdated name is Spirulina platensis) that were used as food long ago by Aztecs in Central America and tribes that inhabited the region around Lake Chad in Africa), Nostoc (N. commune finds application in bioremediation of industrially polluted soils [10]), Anabaena, and Aphanizomenon.

Table 1.

Examples of microalgae (including cyanobacteria) used in biomedical technologies

Genus and representative species	Important applications [4, 5, 14–16, 19]
Cyanobacteria
Arthrospira A. platensis A. maxima	Functional food additive (FFA), cancer and allergy treatment, cosmetics
Nostoc N. commune	FFA, cosmetics
Aphanizomenon А. flos-aquae	FFA (necessity of toxicity tests is emphasized in the literature)
Green algae (Chlorophyta)
Chlorella С. vulgaris C. pyrenoidosa	FFA, immunostimulant, cosmetics
Scenedesmus S. quadricauda S. obliquus	FFA, cosmetics
Dunaliella D. salina D. maritima	FFA, source of pharmaceutics based on β-carotene and glycerol, cosmetics, cardioprotector
Haematococcus H. pluvialis	FFA, source of astaxanthin as an antioxidant, cosmetics, obesity treatment
Chlorococcum C. infusionum	Source of astaxanthin
Red algae (Rhodophyta)
Porphyridium P. cruentum	Cosmetics, cardioprotector, anti-inflammatory agent
Ochrophyta
Phaeodactylum P. tricornutum	FFA, obesity treatment
Nitzschia N. frigida	Obesity treatment
Chaetoceros C. affinis	Antimicrobial agent
Nannochloropsis N. oculate	FFA (enriched in ω-3PUFAs), cosmetics, obesity treatment
Haptophyta
Isochrysis I. galbana	Cosmetics, cardioprotector, anti-inflammatory agent

FFA is a functional food additive; this term implies a microbiota-normalizing and physical and mental health-promoting effect along with a more specific influence of the microalgae in question on the organism (exemplified by rejuvenating, tranquilizing, or anti-allergenic activities).

It has recently been estimated that the global market of microalgal products in 2022 will be worth 3.3 billion USD and the most important contribution will be made by pharmaceutical and food industry, taking the fact into account that people are concerned about their health and interested in environment-friendly alternatives to chemically synthesized products especially as chronic diseases are increasingly widely spread around the globe [5]. The leading companies in the field of biotechnology include Algae Tec (Australia), Pond Biofuels Incorporated (Canada), Cyanotech (United States), and Algae Systems (United States).

The prospects of modern biotechnology and especially its biomedical aspects are largely associated with synthetic biology as the engineering approach to the genome that can be modified and rearranged in order to change gene functions [11]. As an example, a microalga can be developed that will flocculate (form a sediment easily separable by filtration for harvesting the biomass) whenever it is necessary for a biotechnologist; it will also synthesize the target product in response to supplementing the medium with bacterial quorum-sensing signals provided that the necessary genes are inserted into the microalgal genome from bacterial DNA [11].

Further prospects for using microalgae in biotechnology are also associated with widely used phenomics that is based on extensive databases containing the important features (phenotypes) of microalgae. Phenomics enables target-oriented screening of natural and genetically modified (GM) organisms for obtaining optimum biotechnological producers that combine a high growth rate with a high specific yield of the target product, such as the antioxidant asthaxanthine that is expected to produce health-promoting effects [11].

In addition, the Internet of Things (IoT) is of significant practical importance; it involves automatons, sensors, and learning robots for developing self-adaptable biotechnological processes that respond to and the adjust to environmental changes; a biotechnological process is supplemented with its sensors-provided information-dependent digital “twin,” i.e., a virtual process that enables predicting the development of the actual process and making necessary technological changes for increasing the yield of biomass and target products and reducing the waste product amounts [11].

Medical applications. Microalgae and their products hold much promise for revolutionizing the pharmaceutical industry because they represent an environmentally friendly alternative to chemically synthesized preparations for treating or preventing diverse diseases including various types of diabetes, metabolic syndrome and obesity, cardio-vascular problems, malignant tumors, inflammatory processes, Alzheimer’s disease, depression, and other psychiatric disorders, as well as various bacterial, fungal, and viral infections. The medically useful effects of microalgae are caused by their components that exhibit antioxidant, light-protective, gel-forming, moistening, antimicrobial, and other activities. They are due to the presence of polysaccharides (especially their sulfated derivatives), carotenoids, phycobiliproteins (in cyanobacteria), lipid components (especially polyunsaturated fatty acids (PUFAs)), vitamins, and a large number of other chemical compounds [4–6, 12–17].

The capacity of algal components to quench free radicals is of paramount clinical importance. In particular, microalgal carotenoids can use at least three different reactions to quench free radicals, especially reactive oxygen species; this is one of the main factors that are responsible for their light-protective, anti-inflammatory, rejuvenating, and anticarcinogenic effects [18]:

Car + R* → Car⁺ + R^– (electron transfer),

Car + R* → [Car…R] (formation of the radical adduct),

Car + R* → Car(-H)* + RH (proton transfer),

where Car is a carotenoid and R* is a free radical. The above processes are particularly characteristic of astaxanthin, an active radical quencher that contains polar groupings interacting with radicals [18].

Both separate microalgal components and whole biomass preparations find application as pharmaceuticals and preventives. Chlorella sp. biomass promotes the healing of gastric ulcers and wounds, accelerates muscular tissue synthesis, and increases the secretion of anabolic hormones. Per diem addition of 6 g of chlorella biomass to food substantially reduces the risk of pregnancy-associated complications (anemia, proteinuria, and edema) [5, 18].

In their capacity as food products or food additives, the microalgae of the genera Chlorella, Dunaliella, and Haematococcus and the cyanobacteria of the genera Arthrospira, Aphanizomenon, and Nostoc conform with the Generally Regarded As Safe (GRAS) designation but a prerequisite for their practical use is a test for toxicity [4, 19]. Microalgae are important protein sources (the global demand for algal protein exceeded 700 million USD in 2019, which has continued increasing [5]). It has been predicted that, by the mid-21st century, up to 18% of the protein in the global market will be provided by algae [20]. Microalgae are valuable sources of essential amino acids, carbohydrates, e.g., glucose and starch, vitamins В₁, В₂, В₅, В₆, В₉, В₁₂, A, C, E, and biotin, as well as carotenoids and other nutritionally and pharmacologically important pigments, including the phycobiliproteins of cyanobacteria [12, 13, 20].

Microalgae are used either as self-contained food items, in the form of liquid cultures, capsules, pills, powders, or as additives to diverse food products (sauces, desserts, yoghurts, cheeses including molten and cottage cheese, pasta, bread, steaks, sausages, etc. [4, 5, 19]). Gel-forming and stabilizing properties of the biomass components of many algae and their application as food thickeners provide the development of a wide range of biomass-derived products such as alginate and carrageenan [19].

The main component of chlorella biomass, β-1,3-glucan, is an immunostimulant and an antioxidant; it also decreases lipid concentrations in the blood [4, 5]. In the capacity of nutraceuticals and biologically active additives (BAAs), C. vulgaris and other chlorella species are produced by such companies as Chlorella Manufacturing and Co. (Taiwan), Klötze (Germany), and Ocean Nutrition (Canada) [5]. In Russia, a popular trademark is Orgtium® (tableted chlorella; https://fitomarket.ru/catalog/zdorovoe_pitanie/superfudy/khlorella; the same trademark applies to Arthrospira biomass denoted Spirulina). There is a Chinese analog, Natural Chlorella Tablet, that is manufactured by the Qingdao Vital Nutraceutical Ingredients Bioscience Co. company (https://www.made-in-china.com/products-search/hot-china-products) (see also [21]).

The use of polysaccharides, especially those with sulfate groups, that are obtained from microalgae including cyanobateria and possess antioxidant, anti-inflammatory, immunomodulating, and antiviral properties, is of considerable interest; these are also used for treating joint problems. Popular microalgal species include Tetraselmis sp., Isochrysis sp., Porphyridium cruentum, and Porphyridium purpureum, in addition to various chlorella species [22].

The advantages of chlorella biomass include high concentrations of ascorbic acid and K⁺, Na⁺, Mn²⁺, and Ca²⁺ ions. Chlorella vulgaris competes with Arthrospira platensis as a valuable vegetarian source of vitamin В₁₂ contained in chlorella biomass in the biologically active form of methylcobalamine that is suitable for the human body [23]. Cultivation at a low light intensity facilitates the accumulation of valuable PUFAs, in particular, α-linolenic acid, in chlorella biomass [5].

The global market features such microalgal pharmaceutically important substances as astaxanthin from Haematococcus pluvialis under the trademark Spirulina (Earth Spirulina Group, ES Co., South Korea) that is recommended for decreasing lipid content [24, 25], eicosapentaenoic acid (EPA) from Nannochloropsis sp. (Almega®PL and Qualitas Health, United States) that reduces the cholesterol level in the blood [5, 26], and docosahexaenoic acid (DHA) from Schizochytrium limacinum (Maris DHA Oil, IOI, Germany), a drug for rheumatoid arthritis [27].

Apart from asthaxanthin, biotechnologically important microalgal pigments include other carotenoids, especially β-carotene, lycopene, zeaxanthin, canthaxanthin, etc. The intensely colored orange β-carotene is the precursor of visually and immunologically important vitamin A, and it also possesses antioxidant properties and is used as a valuable food dye (colorant) [4, 5]. The green microalga Dunaliella salina contains up to 12–14% of β-carotene (by dry weight). β-Carotene from D. salina is produced by the companies Earthrise Nutritionals (United States) and Nature Beta Technologies Cognis (Australia). Due to a lack of a rigid cell wall and its high protein content, the biomass of the algae of the genus Dunaliella is easily ingested; it is used in baking industry and also as feed for fish and cattle [5, 13, 28–30].

Microalgae are employed for preparing extracts (used, e.g., in beauty salons), alginic acids (as masks), and essential oils. The useful effects of microalgal preparations are due to their efficiency as skin moisturizers (or, conversely, excessive liquid removers), thickeners, pigments, sunscreens, and rejuvenating and skin-bleaching agents [4, 5, 22].

The products of genetically modified (GM) microalgae find increasingly wide application [12]. As an example, antibodies [31], vaccines, erythropoietin, viral protein 28 (VP 28) [32], and immunoconjugated cytotoxins for target-oriented elimination of cancer cells [11] were obtained from Chlamydomonas reinhardtii using genetic engineering techniques. Antibodies in the form that enables immediate introduction into the body and elimination of pathogens therein are released into the medium by GM cells of Phaeodactylum tricornutum [11].

Microalgae as probiotics. The actual and prospective clinical applications of microalgae raise the possibility that they can be used as probiotics. This term was originally coined by the German nutritionist Werner Collath in 1950; he contrasted probiotics with risky antibiotics [33]. According to the World Health Organization [34], probiotics are live organisms that, “when administered in adequate amounts, confer a health benefit on the host.”

The term probiotic derives from the Greek words pro and bios that mean “for life”; it predominantly refers to microorganisms that promote the health of their consumers, provided that they are ingested in adequate doses and form a part of a well-balanced diet [13]. Probiotics enter the market in the form of biologically active food additives and wholefood ingredients.

Probiotics and their advantages were considered in recent works by Prof. Boris A. Shenderov [35–39].

A debatable issue is whether microalgae are to be regarded as probiotics. Serious objections have been raised in the literature. Even though addition of live microalgae to the feed improves the health and viability of marine animals (fish and invertebrates), for most of them there is no evidence that these microalgae are efficient probiotics. The fate of microalgae administered with feed and their impact on the intestinal microbiota have not yet been elucidated [13]. No convincing data have been obtained on the survival of microalgae and the retention of a sufficiently dense microalgal population in the gut, in contrast to classical probiotics, such as lactobacilli, lactococci, bifidobacterial, and others. Nonetheless, the authors believe that special hopes should be pinned on the heterotrophic lifestyle that is characteristic of a sufficiently large number of microalgae. For comparison, it should be noted that probiotic lactobacilli are expected to persist in the gut as a robust population with a minimum density of 10⁶ colony-forming units per 1 g of intestinal content [40].

In the global literature available to the authors no evidence has been presented that microalgae conform to all requirements met by probiotics, such as tolerance to the conditions of the gastro-intestinal tract (GIT) including specific physical and chemical stress caused by low pH values, a high redox potential, and a high osmotic pressure [30].

There is no direct evidence that microalgae can attach to the host gut mucosa, which is considered a prerequisite for the probiotic role in a large number of recent publications [35, 41–43]. Nonetheless, such attachment seems to be possible in view of the proclivity of many microalgae for biofilm formation with adherence to substrates. The authors of the present review article are planning to conduct target-oriented research at their laboratory to test the capacity of prospective probiotic microalgae to attach to the gut mucosa.

Recently, much attention has been given to mixed cultures of microalgae and probiotic bacteria, both in natural water bodies (in aquaculture) and under laboratory conditions. The components of such a mixed culture produce a synergistic effect that results in accelerating the growth of probiotic bacteria and algae and stimulating the synthesis of products that are important for the health of the host organism. Data have been presented on the acceleration of the growth of the microalga Isochrysis galbana in mixed cultures with various probiotic microorganisms [44]. In aquaculture, addition of microalgae along with bacterial probiotics enables improving the functioning of the gut and increasing the production yield in experiments with fish, mussels, and shrimp [40, 45, 46]. Introducing a Chlorella sorokiniana culture together with the probiotics Lactobacillus plantarum and Bifidobacterium longum results in prolonging the survival period of these probiotics in a cooled flan (at 4°С). In addition, C. sorokiniana-produced metabolites enhance the antiviral effect of both probiotics vis-à-vis rotaviruses [47].

In the following, the applicability of some other important probiotic criteria to microalgae will be discussed, drawing on a number of recent publications [29, 30, 39].

1. Probiotics facilitate the optimization of the qualitative and quantitative composition of the GIT microbiota; they increase its stability and robustness and suppress noxious microorganisms (competitive exclusion) by competing for ecological niches, nutrients and growth factors in the host organism and by producing antimicrobial compounds (short-chain fatty acids, bacteriocines and their analogs, hydrogen peroxide, nitric oxide, etc.) [48–50]. Although of paramount importance, this probiotic function has not been sufficiently tested in studies with microalgae. However, it was established that, e.g., Arthrospira platensis stimulates the development of such useful symbiotic bacteria of the GIT as Lactobacillus casei, L. acidophilus, Streptococcus thermophilus, and Bifidobacteria. The microalga inhibits the growth of the opportunistic pathogens Proteus vulgaris, Bacillus subtilis, and B. pumulis [13, 51–55]. Due to their polysaccharides, Chlorella pyrenoidosa and C. ellipsoidea suppress the proliferation of the cells of the pathogenic bacterium Listeria monocytogenes and the yeast Candida albicans [13, 51]. A significant positive influence is exerted on the intestinal microbiota by microalgae-produced omega-3-unsaturated fatty acids [56].

There is evidence for a probiotic effect of microalgae on animal microbiota. Administering Nannochloropsis oculata to the seahorse Hippocampus reidi or feeding the microalgae Chaetoceros sp., Pavlova sp., and Isochrysis sp. (separately or in combination) to oysters promoted the survival of these animals and decreased the quantity of viable pathogenic bacteria in their organisms [13, 57]. Importantly, probiotics do not disrupt the operation of the symbiotic microbiota of the GIT, in contrast to antibiotics [43]. Taken together, these facts raise the possibility of using microalgae as new therapeutics for maintaining a healthy microbial consortium of the GIT [13].

2. Probiotics eliminate toxins and metabolites that are harmful for the host organism. This probiotic function was emphasized by Boris Shenderov [30, 37–39, 48–50]; it is characteristic of classical bacterial probiotics. Nevertheless, it is to be expected that a mixed algae–bacteria culture will prove to be still more efficient in eliminating nocuous substances, since a synergistic stimulatory effect is typical of such a mixed culture and involves both its components [40]. In addition, microalgae per se actively bind various harmful substances. They effectively purge the environment of compounds containing sulfur, selenium, and, still more important, heavy metals including zinc, copper, lead, mercury, chrome, cadmium, nickel, iron, manganese, and vanadium [5, 58, 59]. All these elements accumulate in water and soil and affect humans and agricultural animals; this is the reason that this potential biotechnological niche of microalgae as candidate probiotics is so important. Of special interest in this context are yoghurts, juices, and other microalgal biomass-supplemented beverages that enable reducing the transfer of metals and other noxious agents into the human organism, especially under urban conditions. Thanks to their active enzymes, many microalgae can detoxify harmful organic compounds, including those accumulating in drinking water and food and deriving from pharmaceutics and cosmetics. As an example, Scenedesmus obliquus and Chlorella pyrenoidosa break down the hormone-based contraceptives progesterone and norgestrel [11, 60].

3. Probiotics form low-molecular-weight nutrients, antioxidants, protective compounds, and other biologically active substances (BASs) that influence the water-salt, lipid, amino acid, and energy metabolism, the redox balance at the local (intestinal) and systemic (organismic) levels, and the development and operation of the peripheral and central nervous system; they regulate host gene expression, impact the functioning of the innate and adaptive immune system, and eliminate toxic and carcinogenic compounds [30, 37, 48–50]. These capacities of probiotics remain to be further tested with respect to microalgae. Nonetheless, microalgal components, especially carotenoids, undoubtedly possess antioxidant properties, which enables their cardioprotective and anti-atherosclerotic effects. Dunaliella salina, which contains up to 10–13% of β-carotene, protects both mice and humans against the development of atherosclerosis. The D. salina-specific mixture of trans- (~40%) and cis-isomeres of β-carotene reduces the levels of total lipids, cholesterol, and triglycerides in the organism to a greater extent than synthetic β-carotene that contains the trans-isomer only [61, 62]. The cis-isomer of β-carotene from Dunaliella bardawil suppressed the development of atherosclerosis in aged mice that were on a fat-enriched diet [63, 64]. Cardioprotective properties are characteristic of PUFAs, especially of omega-3 acids produced by a number of microalgal species, e.g., Porphyridium purpureum and Isochrysis galbana that lower the blood cholesterol level and facilitate the normalization of the blood pressure. Docosahexaenoic acid (DHA) preparations are commercially available [5]. A Nannochloropsis sp. strain exposed to bright sunlight in an open pond produced significant amounts of eicosapentaenoic acid (EPA) that formed the basis of the commercial preparation A2EPA Pure^TM (United States) [65].

Under the rubric of BASs, of note are numerous signal molecules including hormones and neurotransmitters. As an example, neuroactive biogenic amines also serve as a chemical language for communication between the host organism and the microbiota, including putative probiotic microalgae. Biogenic amines (norepinephrine, dopamine, serotonin, histamine, etc.) bring about specific responses such as growth stimulation and accelerated development of microalgal cultures exemplified by Chlorella vulgaris, Scenedesmus quadricauda, and a number of other species [66–71].

4. Probiotics exert an anticarcinogenic effect. This is characteristic of a large number of microalgae and their components, such as astaxanthin, β-carotene, luteine, violaxanthin, fucoxanthin, and other microalgal carotenoids, as well as of the phycobiliproteins of cyanobacteria including Arthronema africanum and Arthrospira platensis [5, 12, 13, 72]. Ñ-phycocyanin exerts an inhibitory influence on the liver cancer (HepG2) [73], leukemia (Ê562) [74], and lung cancer (À549 and NSCLC) [75, 76] cell lines. It was established that phycocyanin from Limnothrix sp. potentiates the effect of the antitumor drug topotecan on the prostate cancer cell line [77]. Monoacylglycerides forming a part of Skeletonema marinoi lipids can activate caspase 3/7 and, therefore, induce the apoptosis (programmed death) of colon cancer (HCT-116) and hematological cancer (U-937) cells but not normal cells [56]. PUFA-containing microalgal lipids exhibit antitumor activity with respect to cervical and breast cancer. EPA and DHA suppress blood vessel growth in the tumor tissue and enhance peroxide-dependent stress in the endoplasmic reticulum, which results in tumor cell destruction [56]. The sulfated polysaccharide fucoidan from Fucus vesiculosis, Sargassum henslowianum, Cladosiphon fucoidan, and Coccophora longdorfii inhibits blood vessel formation and metastasis development by inducing, via caspase 3/7 activation, the apoptosis of the lymphoma, melanoma, lung carcinoma, promyeloid leukemia, colon and breast cancer cell lines [78]. Anticarcinogenic activity was found to be exhibited by violaxanthin that is synthesized, e.g., by Dunaliella tertiolecta [22].

5. Probiotics are characterized by anti-inflammatory, anti-allergic, and antidiabetic activities. The anti-inflammatory effect that is largely associated with the impact on the immune system (see a special passage below) is typical of the sulfated polysaccharides of Chlorella spp., Tetraselmis sp., Isochrysis sp., Porphyridium spp. [22], and other algae as well as of their pigments, especially β-carotene and astaxanthin that are in the focus of attention of influential international networks such as the International Carotenoid Society, Eurocaroten, IBERCAROT, and CaRed [13]. In experiments with rats as models, the dried powder of Dunaliella bardawil biomass mitigated acetic acid-induced small intestine inflammation [79]. As for patients with nonalcoholic fatty liver disease (NAFLD), treating them with tableted C. vulgaris biomass resulted in a verifiable decrease in proinflammatory cytokine TNF-á content [79]. Arthrospira phycocyanins inhibit the NADPH oxidase enzyme that is implicated in inflammatory processes [12, 80]. Astaxanthin, a red pigment (a food and cosmetic colorant) and an antioxidant that is particularly characteristic of Haematococcus pluvialis, mitigates the “cytokine storm” (excessive immune system activator production) during the COVID-19 infection [12, 80]. The PUFAs of algae facilitate the treatment of inflammatory diseases including arthritis [13, 81].

A bacterial probiotics-specific antidiabetic effect was revealed, e.g., in the cyanobacteria of the genus Arthrospira, which is attributable to their high vitamin and γ-linolienic acid content [12]. Antidiabetic activity is also exhibited by the aforementioned astaxanthin [13] and phycocyanin [82] pigments. A debatable issue is whether microalgae produce an anti-allergic effect. In the authors’ opinion, they should be first tested for an opposite effect, i.e., they might cause allergic complications during systematic administration to patients.

6. Probiotics facilitate metabolism normalization, body weight reduction, and obesity (metabolic syndrome) treatment. In addition, probiotics can be used to treat diametrically opposite health problems, i.e., anorexia and emaciation. It was experimentally demonstrated that probiotics contribute to normalizing the health state of a rodent after food deprivation [83]. Therapeutical effects were revealed in in vitro and in vivo studies, including clinical tests, for the microalgae Euglena gracilis, Phaeodactylum tricornutum, Arthrospira maxima, A. platensis, and Nitzschia laevis. Microalgae suppress the differentiation of preadypocytes (adipose tissue cell precursors) and reduce total lipogenesis (lipid synthesis) and, more specifically, triacylglyceride accumulation [82]. The resulting increased lipolysis and fatty acid oxidation are accompanied by enhanced energy loss via thermogenesis activation in the brown adipose tissue and the browning of the white adipose tissue. Along with reduced fat accumulation in the organism, microalgal treatment mitigates other symptoms in obese people, including an increased plasma lipid level, insulin resistance (posing the diabetes threat), and mild chronic systemic inflammation [82]. Addition of Arthrospira platensis or Chlorella sp. biomass powder to bread or cookies helps decrease lipid and cholesterol levels and replace the feeling of hunger with that of satiety [13].

7. Our hopes for staving off progressive senescence symptoms are pinned on probiotics. This statement, made in the work by Shenderov [48–51], draws on the ideas suggested by Elia Metchnikoff in Etudes sur la nature humaine; essai de philosophie optimiste [84] and is in line with data on the aging-decelerating and rejuvenating influence of microalgae, which, in turn, is linked to their antioxidant, protective, anti-inflammatory, and metabolism-normalizing effects.

Skin aging is associated with a decrease in the synthesis of the structural components of the skin matrix (collagen, elastin, and hyaluronic acid) and concomitant activation of proteases that degrade these components. Accordingly, aging deceleration and rejuvenation caused by microalgal preparations are partly due to their restrictive effect on matrix component proteolysis. Of significant importance is also the fact that microalgae contain antioxidant effects-producing substances that quench free radicals, especially reactive oxygen species (ROS). Microalgal carotenoids including β-carotene (Dunaliella salina), lutein (D. salina, Scenedesmus spp., Chlorella spp., and Mougeotia sp.), and lycopene protect the skin from the ultraviolet light effect, quench ROSs and slow down skin aging. The rejuvenating influence of microalgae is also attributed to the regulatory effect of their phytohormones (auxins, cytokinins, abscisic acid, gibberellins, etc.) on the human body [5, 22].

8. Probiotics promote blood vessel growth (angiogenesis) in the intestinal tissue by producing vascular endothelial growth factor (VEGF) [43]. In traditional Chinese medicine, spirulina (denoted as Arthrospira platensis currently) and a number of other microalgae (https://www.ginsen-london.com/blog/benefits-of-spirulina, see also [85]) are used for treating duodenal ulcer because they speed up vascularization, i.e., blood vessel elongation, in the gut wall. Target-oriented research aimed at enabling the biosynthesis and release into the medium of VEGF or its functional analogs represents a growth point in the area of research dealing with the probiotic functions of microalgae. Of interest in the context of angiogenesis stimulation in the intestinal tissue is the important therapeutical influence of microalgae including those of the genus Chlorella on the leaky gut syndrome, with chlorella contributing to gut wall tissue regeneration [86].

9. Some probiotics exert a prominent pain-relieving effect. This effect is characteristic of the aqueous extracts of Chlorella stigmatophora and Phaeodactylum tricornutum, which is attributed to their polysaccharide components [22, 87, 88]; these extracts also exhibit anti-inflammatory activity and can quench free radicals.

10. Probiotics mitigate stress; this is not only typical of classical bacterial probiotics such as the bifidobacteria and lactobacilli that form a part of fermented dairy products [89]. Such dairy items can be enriched with microalgal preparations that produce anti-stress effects. As an example, evidence was presented that Chlorella vulgaris possesses not only anti-infection and anticarcinogenic, but also anti-stress properties. In experiments with Wistar rats that were exposed to stress (placing in wet cages or tanks with cold or hot water, disrupting the day–night rhythm, etc.), a C. vulgaris culture mitigated : (1) the behavioral consequences of stress such as anhedonia, i.e., a lack of interest in drinking sucrose solution that rats normally find tasty (after administering chlorella, the rats return to normal behavior: they prefer sweetened solution to plain water); (2) the biochemical effects of stress, such as an increase in blood cholesterol level; chlorella verifiably decreased this level [90].

11. Probiotics interact with gut epithelial cells and, therefore, regulate the activity of the immune system and, in direct fashion, of its intestinal part, the gut-associated lymphoid tissue (GALT); they modulate immune responses, normalize the balance between pro- and anti-inflammatory cytokines, and decrease the antigen pressure exerted on the GALT. Probiotics reduce gut wall permeability, increase immunoglobulin IgA secretion, activate anti-inflammatory Treg cells [91], and facilitate the production of anti-inflammatory interleukin IL-10.

For microalgae as “candidate probiotics,” an active immunostimulatory influence was detected with β-1,3-glucan and other polysaccharides (containing the residues of mannose, glucose, rhamnose, arabinose, etc.) of the representatives of the genus Chlorella [13, 22, 79]. One of the mechanisms of immune system regulation involves stimulation of monocyte, macrophage, and neutrophil proliferation and an increase in phagocyte activity and secretion of immune mediators such as cytokines. Polysaccharide-containing Chlorella stigmatophora, Skeletonema costatum, and S. dohrni extracts activated macrophage-dependent phagocytosis in the abdominal cavity of mice [79]. Omega-3 PUFAs contained in microalgal biomass also stimulate macrophage activity [56]. Administering Chlorella vulgaris biomass as dried powder to human subjects increased natural killer activity in the monocyte fraction of peripheral blood and the interferon γ and interleukin IL-1β and IL-12 content in the blood serum [79]. In similar fashion, feeding dried Dunaliella salina biomass to mice resulted in activating their macrophages and natural killers (NK cells) and also increased the viability of mice with leukemia [79].

The literature available to the authors contains no data on the direct impact of microalgae and preparations obtained therefrom on the activity of the intestinal part of the immune system (GALT) that is inherent in many representatives of classical bacterial probiotics [29, 30]. This area of research remains a sufficiently important growth point for subsequent studies. A similar issue (also related to immune system activity) to be raised is whether microalgae, their components, and preparations obtained therefrom regulate the activity of natural barriers such as the gut–blood barrier and the blood–brain barrier (BBB) via enhancing the expression of proteins involved in intercellular tight contacts. The capacity for strengthening the BBB and other important barriers was detected with respect to bacterial probiotics [92]. Under stress, probiotics improve the protective function of the intestinal barrier, decrease the concentrations of circulating corticosteroids and proinflammatory cytokines and concomitantly increase those of anti-inflammatory cytokines. They are implicated in restoring the integrity of the BBB and the gut–blood barrier and mitigate systemic inflammation [93].

12. The useful effects of probiotics on the nervous system, the operation of the brain, and psychological features including cognitive capacities, memory and social behavior give grounds for classifying some probiotics into the subgroup of psychobiotics. These are live microorganisms that when administered in adequate amounts, confer a health benefit on patients with psychiatric problems [94, 95]. An increasing body of evidence indicates that probiotics can influence the brain, behavior, and, most important, mood and cognitive capacities, both in the experimental and the clinical setting [96]. As an example, it was demonstrated that the psychobiotic strain Lactobacillus rhamnosus JB-1 acted on the GABA-dependent system in the brain and suppressed the anxious behavior of mice in a complex maze and in an illuminated open field as well as their depressive behavior in a forced swimming test [97, 98]. It was hypothesized that humans would not be able to achieve the modern-day level of cognitive capacities without the microbiota [99].

As far as microalgae are concerned, important data have been obtained recently on the neuroprotective effects of various microalgae and their components such as polysaccharides, lipids (especially those with PUFAs), carotenoids, phycobilins, and other compounds. The nervous system is protected by them against oxidative stress, aging, and neurodegenerative disorders (Alzheimer’s and Parkinson’s disease, dementia, etc.). It suffices to point out that representatives of the genus Arthrospira possess neuroprotective properties and facilitate the performance of normal brain functions [40]. Their extracts improve brain fatigue symptoms, prevent or mitigate cerebral circulation problems, and promote cognitive, locomotive, and verbal capacities. This was established in studies in which undernourished children were treated with Arthrospira (“Spirulina”) preparations [40].

Microalgal PUFAs, especially DHA and EPA, are mandatory for the normal development of the nervous system and serve as an important supplement for baby formula products, vitamin-enriched foods and drinks, and dietary food additives. As an example, the baby formula that is produced by the Dutch State Mines Company (Netherlands) contains preparations from the biomass of the dinoflagellate Crypthecodinium cohnii; DHA accounts for up to 60% of its total acid fraction [11].

Probiotic bacteria are known to produce substances involved in the operation of the nervous system (neurotransmitters) or their precursors that can reach the brain via the BBB. Such precursors include 2,3-dihydrophenylalanine (DOPA), the catecholamine precursor, and 5-hydroxytryptophan (5-HTP), the serotonin precursor [29, 30, 39]. Of much interest in this context is the fact that many algal species (exemplified by representatives of Chlorophyta, Charophyta, Ochrophyta, Rhodophyta) synthesize significant amounts of dopamine, serotonin, histamine, tyramine, acetylcholine, and other neurotransmitters [66]; such algae are to be envisaged as potential probiotics, and they could be used to improve the functioning of the brain, promote mental health, and treat brain disorders, e.g., Parkinson’s disease associated with dopamine deficiency in the substantia nigra of the brain.

Importantly, each of the useful effects of probiotics is due not only to individual microbial substances but also to intricate complexes of low-molecular-weight compounds that are produced by probiotic microorganisms either in their functional form or as precursors [37, 38]. These complexes of microbial substances affect the host and its microbiota against the background of the impact of other biologically active substances that either enter with food or are produced by the resident microbiota. In connection with the probiotic role of microalgae, it should be noted that various phenols, fatty acids, indole, terpenes, acetogenins, and some volatile halogenated hydrocarbons obtained from microalgae exhibit antimicrobial activity. Supercrticical (СО₂-extracted) extracts of the microalga Chaetoceros muelleri produce antimicrobial effects because of their lipid composition [100]. Another growth point of present-day research involves studies on the antibacterial, antiprotozoal, antifungal, and antiviral activities of many microalgae and their components [12].

Microalgae as prebiotics. Apart from the aforementioned data in support of the probiotic function of microalgae, the question is raised in the literature of whether microalgal polysaccharide components and other organic constituents perform a prebiotic function. Prebiotics are indigestible food components that bring about specific changes in the composition and/or activities of the GIT microbiota and, therefore, exert a positive influence on health. In accord with the official WHO/FAO definition, prebiotics are to be construed as nonliving edible products that improve health by altering the microbiota [101].

Typical representatives of probiotics are indigestible oligosaccharides degradable by beneficial microorganisms in the gut, which produce short-chain fatty acids and other organic compounds holding much value for the host [42]. Optimizing the diet by enriching it in such prebiotics as fructans should contribute to the proliferation of useful bacteria such as Bifidobacterium [49, 102]. Prebiotics can exert anti-inflammatory effects that are attributable to polysaccharides’ capacity for direct interaction with the intestinal epithelium, regardless of gut bacteria, which significantly reduces the production of proinflammatory cytokines [93].

Importantly, the polysaccharide components of microalgae can be broken down to short fragments (oligosaccharides) that possess prebiotic properties. They are exemplified by inulin, galactooligosaccharides, xylooligosaccharides, and the oligosaccharides that are obtained from agarose, alginate, and carrageenan, as well as by arabinoxylans, galactans, and β-glucans [13, 14]. Microalgal oligosaccharides are either not fermented or only partially degraded by the usual gut microbiota of humans or animals. However, these compounds selectively stimulate the growth and activity of specific beneficial bacteria exemplified by lactobacilli and bifidobacterial, provided that the oligosaccharides translocate to the colon in order to promote the host’s health, i.e., to function as prebiotics [13].

Presumably, the prebiotic components of microalgae contribute to the aforementioned (see item 1 in the list of the potential probiotic features of microalgae) positive influence on the host microbiota, as exemplified by the stimulatory effect of Arthrospira platensis on the viability of bacteria that form a part of the intestinal microbiota, including Lactobacillus casei, Streptococcus thermophilus, Lactobacillus acidophilus, and Bifidobacteria as well as its negative impact on the pathogens Proteus vulgaris, Bacillus subtilis, and Bacillus pumulis that were suppressed by A. platensis [13] in in vitro studies.

The prebiotic and the probiotic roles of microalgae are mutually complementary: algae can be used as live cultures and produce the probiotic effects listed above; in addition, their carbohydrate components can provide the raw material for preparing efficient prebiotics in order to stimulate the beneficial microbiota and, therefore, to promote human health.

Microalgae as sources of metabiotics. Metabiotics were defined in the literature as biologically active substances that are produced as a result of the metabolic activities of symbiotic (probiotic) microorganisms and exert a positive influence on various kinds of physiological processes [35]. The meaning of the term metabiotics actually is closely related to that of the relatively popular term postbiotics. It denotes bacterial products that, in the absence of viable bacterial cells, can exert an influence, in analogy to those cells, on the signaling pathways and the barrier functions of the organism. Metabiotics contain bacteriocins, organic acids, ethanol, and diacetyl [29, 30, 35, 43]. Metabiotics are exemplified by heated cells of probiotic bacteria; polysaccharide A formed by Bacteroides fragilis that activates the immune system and protects mice (in an experiment) from Helicobacter hepaticus-induced colitis; and the preparation skeleton P-CWS (Propionibacterium acne cell wall fragments) that increases the cytotoxic activity of macrophages and, therefore, produces an anticarcinogenic effect [30].

In the authors’ opinion, the aforementioned interpretation of metabiotics is generally applicable to microalgae-produced valuable products that improve human physical and mental health, including carotenoids (β-carotene, astaxanthin, lycopene, luteine, zeaxanthin, violaxanthin, canthaxanthin, fucoxanthin, etc.); chlorophylls; phycobiliproteins (especially phycocyanin, allophycocyanin, and phycoerythrin); carbohydrates (exemplified by β-glucan, fucoidan, and other sulfated polysaccharides); lipids (especially PUFA-containing triacylglycerides); vitamins; plant hormones (auxins, cytokinins, etc.), and other regulatory molecules; K⁺, Na⁺, Mn²⁺, and Ca²⁺ ions; and other important components [5, 12, 13, 16–18, 24]. Interestingly, PUFAs facilitate the treatment of cardiovascular diseases, hypertension [13], thrombosis of heart coronary vessels, malignant tumors, asthma, bowel inflammatory diseases, and psychiatric problems such as schizophrenia and anxiety disorders [15].

To sum up, the present work is focused on the present-day knowledge concerning the practical applications of microalgae, including cyanobacteria, as agents used for treating and preventing various diseases, promoting health, and decelerating aging. Despite a lack of sufficient data on the long-term survival of useful microalgal cultures in the human/animal gastro-intestinal tract, the statement can be made that a large number of algal cultures fully conform to many important probiotic criteria. Some of their components, e.g., polysaccharides, seem to provide the raw material for producing short fragments to be used as prebiotics, or, alternatively, hold much pharmaceutical and dietary value as metabiotics, as emphasized by Boris Shenderov [35, 36].

COMPLIANCE WITH ETHICAL STANDARDS

The authors declare that they have no conflicts of interest. This article does not contain any studies involving animals or human participants performed by any of the authors.