**2.4 Therapeutic applications of microalgal nutraceutical substances**

Microalgae are known for being a fantastic source of protein, which enables it to meet the ever-growing demands of a growing population. Microalgae-derived protein products exhibit a high concentration of protein with complete essential amino acid profiles. Other than proteins, microalgae are known to contain crucial vitamins such as A, B12, C, D, E and minerals such as iron, calcium, potassium, and more. Foods derived from common microalgae species *Chlorella sp.* and *Spirulina platensis* are known for their high protein content and nutritional value and are available in form of capsules, powders, and liquids. These have great abilities to combat various types of diseases due to their antimicrobial, anti-inflammatory, anticancer, and immunosuppressive properties [24].

Generation of reactive oxygen species induced by prooxidants leads to oxidative stress and is a cause of swelling and lysis of mitochondria and mutagenic actions. They accelerate the aging process and contribute to various threatening chronic and degenerative diseases such as diabetes mellitus, rheumatoid arthritis, and cancer. Enzymatic and nonenzymatic antioxidants from microalgae have the capability to quench free radicals and have high radical scavenging activities [25]. The antimicrobial properties of microalgae-based food products are mainly due to its lipid composition. Fatty acids affect the membranes of microbes resulting in the damage of internal constituents. This has a direct impact on the cells' metabolism [26].

Monounsaturated and polyunsaturated fatty acids (PUFAs) in microalgae derived food products also contribute to their antibacterial properties. Microalgal fatty acids intrusively interfere with bacterial mechanisms and can result in fatal effects such as cell leakage, reduced uptake of nutrients and inhibition of cellular respiration. Antiviral and antifungal properties of microalgae food products are attributed to polysaccharides and lipid fractions respectively [27]. High concentrations of polyphenols, polysaccharides, and phycobiliproteins in microalgae-derived nutraceuticals are the reason for their exhibition of anticancer properties due to their abilities to induce apoptosis in tumor cells [28]. The biological response due to diverse factors such as pathogens, allergens, irritants is termed as inflammation, and this is mainly observed in damaged or infected cells and tissues [29]. Chronic inflammation can directly contribute to chronic illnesses such as diabetes, cardiovascular and neurodegenerative disorders, and several inflammatory responses are a direct consequence of oxidative stress. Anti-inflammatory compounds such as carotenoids produced by microalgae reinvigorate the body's immunity and accelerate healing [27].

Despite the obvious benefits, due to lack of awareness, microalgae are underexploited as a potential food source. There are a plethora of microalgal species that have the ability to produce therapeutic bioactive compounds with their respective functional properties, which are impacted by the varying biotic and abiotic stresses [27].

Carotenoids produced by microalgae have excellent therapeutic effects on humans due to their antioxidant properties, thus yielding protection against oxidative and free radical stresses [24].

They play a significant role in the inhibition of oxidative injury to cells and tissues. Two of the industrially important carotenoids include β-carotene and astaxanthin. The 9-cis-isomer of β-carotene has been observed to possibly prevent the advancement of artherosclerosis in humans. Studies have also evaluated the positive effects of β-carotene in cardiovascular health. Other than its addition in multivitamin tablets, β-carotene can also be used in food products such as cheese and butter. Astaxanthin is another important carotenoid that is known for its exceptional antioxidant properties. It has been proved to reduce oxidative stress and inflammation and aids in improving the immune system to combat cardiovascular diseases [27].

Another valuable bioactive compound extracted from the microalgal biomass is chlorophyll, which possesses powerful antioxidant and antimutagenic properties. Studies have demonstrated its wound healing properties and stimulation of tissue growth. Due to its facilitation of rapid oxygen and carbon dioxide interchange, it is used in the treatment of oral sepsis and ulcers. By trapping mutagens in the gastrointestinal tract, chlorophyll contributes to prevention of cancer [30]. Chlorophyll also consists of a component called chlorophyllin, which has the ability to target multiple carcinogen pathways and invade their cell cycle [24]. Despite the various benefits, utilization of chlorophyll for dietary needs has disadvantages of its own. Apart from being an expensive natural food dye, it is also unstable to changes in pH conditions, and to ensure its stability, it has to undergo chemical modification [30].

Carbohydrates from microalgae are in the form of reducing sugars such as sucrose, lactose, fructose, and polysaccharides. Despite limited food applications, microalgal polysaccharides have garnered attention due to their nontoxicity and biocompatibility [31]. They boost the immune system functionality and have the ability to block tumorigenesis. Polysaccharides such as immulina and immurella demonstrate great anticancer properties while β-1,3 glucan from *Chlorella* species alleviates gastric ulcers and atherosclerosis [27].

Most microalgae consist of polar lipids such as phospholipids and galactolipids in the exponential growth phase and then tend to accumulate triacylglycerols under stressful conditions [31]. In general, microalgae are rich in poly monounsaturated fatty acids, which play crucial roles in cellular metabolism, balancing of membrane fluidity and electron-oxygen transport. External administration of lipids is important for humans due to the lack of ability of synthesizing lipids for the maintenance of homoeostasis. Poly monounsaturated fatty acids have also been proven to decrease the prevalence of various chronic diseases and have demonstrated health benefits with respect to the nervous system [27]. Microalgal fatty acids have also shown therapeutic effects against inflammation and numerous cardiovascular diseases such as myocardial infarction and cardiac arrhythmia. Docosahexaenoic acid (DHA) shows excellent cardiovascular and nervous system benefits and is therefore used as a nutritional supplement in infant formula [32]. Species of microalgae such as *Cryptothecodininum cohnii* are cultivated in fermenters for the mass production of DHA [31].

Microalgal protein extraction has a lot of benefits regarding enhanced nutritional value, productivity, and efficiency [33]. Phycobiliproteins are hydrophilic complexes used in popsicles, gum, soft drinks, and dairy products. These pigments are found

#### **Figure 2.**

*This figure summarizes some of the various diseases that can be targeted by bioactive compounds derived from microalgae based food products [27, 35].*

majorly in Cyanobacteria and have innate wound healing, antiviral, antioxidative properties and can act as neuroprotective agents. It also has the ability to function as an immunotoxin against B-cell lymphoma and also has an enhanced resistance to UV-induced stress [24, 27].

Microalgal species contain a plethora of vitamins (e.g., Pro-Vitamin A, Vitamin B12, Vitamin C, Vitamin D, Vitamin E), which are used as food supplements and are proven to help detoxify, revitalize cells and are involved in the activation of the immune system. The precursor of Vitamin A produced by microalgae has been seen to hinder the development of tumors in various types of cancers such as bladder, lung, and skin cancers. Cobalamin (Vitamin B12) has been demonstrated to aid in DNA repair, whereas ascorbic acid acts as an immunomodulatory agent for the prevention of severe diseases. Through the blockage of cell cycle progression, Vitamin D has demonstrated anticancer properties. Tocopherols and tocotrienols (Vitamin E) meanwhile have shown beneficial effects against atherosclerosis and pancreatic cancer [27, 34].

Microalgae consist of high levels of sterols and are components of the cellular membrane. The sterol composition is directly impacted by algal strain and the external conditions they are subjected to. Microalgae-derived sterols have been reported to demonstrate good anticancer, antioxidant, and anti-inflammatory properties. These have also been utilized as supplements for lowering cholesterol levels leading to the decrease in cardiovascular disease risks [27]. **Figure 2** summarizes some of the various diseases that can be targeted by bioactive compounds derived from microalgae based food products.

#### **2.5 Challenges and future prospects**

Despite the discovery of several compounds of high biological value and health benefits, microalgae continue to remain one of the most sparsely explored groups of organisms with an overwhelming majority of microalgal compounds yet to be isolated and identified. Although several thousand species of microalgae are believed to exist, only a small handful of these are currently cultivated and used for industrial applications.

This dearth of information calls for intensive research in the field of microalgal bioprospecting. These developments, however, give rise to a new set of challenges.

While the nutritional content of a few microalgal species has been thoroughly researched upon, their palatability, digestibility, and nutritional value ultimately depend upon the genetic makeup of the individual strains used, as well as the processes involved in large-scale biomass production. A major constraint in the use of large amounts of microalgae for human nutrition is the presence of excessive quantities of nucleic acids, which when metabolized in the human body to form urea may prove deleterious to human health. Microalgal cultivation strategies in the past include the use of natural lagoons or artificial ponds. While these open air systems are relatively economical, they suffer from a number of challenges including frequently varying climatic conditions, the prevalence of non-axenic cultures, etc. To combat these issues, the recent years have seen a rise in the use of closed systems. However, major constraints encountered with the use of closed culturing systems are the lack of inexpensive equipment and the inefficient use of light, both of which make scale-up problematic. Despite these challenges, closed reactors are promising candidates for the production of high-value nutraceuticals by microalgae including their production by recombinant technologies. Food safety is a highly pertinent matter in microalgal technology, particularly when open air systems are involved, and these products are thoroughly scrutinized by regulatory bodies in developed countries to determine their safety and efficacy prior to human consumption.

Despite being rich sources of proteins, microalgal foods and nutraceuticals have failed to carve a market niche for themselves due to the unpleasant odors and tastes associated with them, and better marketing strategies need to be developed to attract a larger consumer base There are also several considerations involving the specific doses to be prescribed to consumers. Proper labeling and marketing must be ensured to specify the quantities of all the ingredients, including details on allergenics and side effects.

Another key area of future research is the whole-genome sequencing of microalgal species producing high quantities of nutraceuticals to better understand the genes, enzymes, and metabolic pathways involved in nutraceutical production as well as the

**Figure 3.** *Depicts challenges faced during microalgal nutraceuticals production.*

mechanisms involved in the upregulation/downregulation of these genes and pathways. The development of engineered microalgae to boost protein expression, metabolism, photosynthesis, etc., is integral to large-scale production in the future. **Figure 3** depicts challenges faced during microalgal nutraceuticals production.
