With the global population increasing, seeking alternative food sources has become an urgent issue to face the spread of hunger, which doesn’t decrease despite technological progress.
On the other hand, the demand for functional food has gained popularity on the market. This term defines food that is characterized by another function, that can be disease-preventing and/or health-promoting, in addition to its nutritive value.
Because of the rising concerns about the risks of food additives on human health, customers are looking for more natural and sustainable alternatives.
In this scenario, phycocyanins market value has significantly increased in the last years because of their several beneficial effects and the wide range of applicability. [1]
Phycocyanins are peptide-pigment compounds belonging to the phycobiliproteins (PBP) family that can be found in phycobilisomes (PBS). These systems are found in red algae, cryptomonads, cyanelles, and, most of all cyanobacteria.
In these organisms, PBS can represent up to 50 % of the total cellular protein. [2]
What is Phycocyanin and Where Can You Find It?
PBS are light-harvesting complexes located in the outer part of the lipidic thylakoid membrane, which explains the high water solubility of PBPs, that differentiates them from other proteins. [3]
PBSs’ function is to capture light and transfer it with an electron chain mechanism to photosystems I and II (PSI and PSII), where the proper photosynthetic reaction occurs and that is located inside the thylakoid membrane.
Because of their key role in capturing light, it’s no surprise that all phycobiliproteins have bright and intense colors.
Furthermore, their role is to increase the absorption range in organisms that are often exposed to low-light conditions, by covering the regions of the spectrum where usual pigments – such as chlorophylls and carotenoids- don’t show any absorption peak. [2]
PBPs are divided into three groups, according to their absorption spectrum:
- Phycoerythrins (λ max = 540 to 570 nm);
- Phycocyanins (λ max = 610 to 620 nm);
- Allophycocyanins (λ max = 650 to 655 nm). [1]
The first component of PBPs is peptidic and is divided into two subunits, α and β, that can assume different conformations.
In the case of phycocyanin, which is the most common phycobiliprotein in cyanobacteria, a ring-shaped trimeric form (αβ)3 and a hexameric form [(αβ)3]2 exist. [3]
Each subunit is linked to a chromophore belonging to the phycobilin class. These molecules are noncyclic tetrapyrroles linked by α,β, and γ single-carbon bridges. [2]
Until now, we have used the plural “phycocyanins” because three forms of phycocyanin exist, C- and R-II- and R-III- phycocyanin. Chromophores are precisely what differentiate these compounds from one another.
R-phycocyanins are only found in a few cyanobacteria species, whilst C-phycocyanin (C-PC) is present almost in all the species and its chromophore is phycocyanobilin, linked through a thioether bond to the peptide structure and responsible for the blue color of the protein.[1]
Because of its abundance compared to the other phycocyanins, articles always use the general term phycocyanin (PC) to speak of C-PC and from now on, this terminology will be adopted here too.
All the Benefits for Human Health
Phycocyanin has been proven to be safe and non-toxic for human health. Besides, the majority (35%) of articles on phycocyanin report an antioxidant activity, hence preventing the production and/or scavenging of reactive oxygen species (ROS). [1]
The imbalance between antioxidant systems and ROS is related to several diseases such as cancer, neurodegenerative and inflammatory diseases, and aging processes.
It has been proven that both the peptidic structure and the chromophores are responsible for antioxidant activity, resulting in the scavenging of the majority of radicals. [3]
PCB scavenging methods are easier to determine and usually involve the oxidation of tetrapyrrole double bonds.
On the other hand, the apoprotein (α and β subunits) has been proven to scavenge OH radicals – increasing the pH from 7 to 11 resulted in better scavenging due to the change of the protein charge- and HOCl radical by reacting with cysteine and methionine residues.
Other amino acids such as tyrosine, tryptophan, and histidine were able to scavenge ROO. [3]
Many assays can be used to measure the antioxidant activity of phycocyanin, but the main one is with 1-diphenyl-2-picrylhydrazyl (DPPH). [1]
Furthermore, once the antioxidant activity was demonstrated, its biological efficacy was also tested in vivo and in vitro towards pathological disorders of the heart, brain, liver, lungs, eyes, kidneys, and pancreas. [3]
Interestingly, the antioxidant activity has also been tested in real food, through the in vitro digestion of milk-based ice-creams. Antioxidant activity of PC-containing icecreams was 2- or 13-fold higher than control, depending on the method used. [1]
Extraction and Purification
The majority of articles on phycocyanin deal with its extraction and purification methods. Depending on its purity index (PI), indeed, PC is considered to be food, cosmetic, reactive, and analytical grade. Many chemical and physical parameters influence PC extraction.
- Temperature: it has been reported that extraction yield increases by switching the temperature from 30 to 50 °C. However, above 50 °C protein denaturation occurs, which leads to the unfolding of the hexameric structure. Since the peptidic structure has a protective effect on the chromophore, with the reduction of the H bonds network due to denaturation, the tetrapyrrole structure tends to close with a consequent loss of color and antioxidant power;
- pH and solvent: phycocyanin isoelectric point (IP) can vary between 4.1 and 6.3 depending on the cyanobacteria. Extractions at pH > 7 have reported higher yields due to the increased protein’s global charge and protein-solvent interactions;
- Biomass/solvent ratio: a higher biomass/solvent ratio is generally characterized by a higher extraction yield, along with a loss of extract’s purity. [4]
Several methods have been used to obtain PC, using both conventional (solvent, maceration, percolation…) and novel extraction techniques (ultrasound or microwave-assisted extraction, high-pressure processing, high electric fields).
A key step in PC extraction and purification is cell disruption which can be improved by coupling two techniques.
Freezing and thawing is one of the privileged techniques in the laboratory since it allows to reach highly pure extracts. It has been reported that the best yield value has been obtained with 4 cycles of freezing and thawing. However, it is a long procedure which makes it difficult to scale up. New technologies have been set up to respond to industrial applications.
Some of the most suitable methods for such an application are:
- Ultrasound-assisted extraction (UAE);
- Pulsed electric field (PEF);
- Aqueous two-phase system (ATPS) extraction.
Furthermore, to purify phycocyanin chromatographic methods (ion exchange, membrane, gel filtration, and hydrophobic interaction chromatography) are often coupled to extraction methods.
However, PC stability remains a challenge and several efforts are being made to find stabilizing agents such as small sugars that could act as co-solute, preserving protein structure and chromophore stability. [4], [5]
Other Possible Uses
Phycocyanin properties can be used for other important applications. First of all its blue color makes this compound very attractive on the market as a possible alternative natural blue colorant.
Indeed, the United States have approved only two artificial blue colorants as food additives: triarylmethane dye Brilliant Blue FCF and Indigo Carmine, while the European Union has authorized another triarylmethane dye Patent Blue V in addition to the previous ones.
Compared to other pigments, blue and purple colors are relatively less common in nature and can be found in fruits and vegetables with anthocyanins or in cyanobacteria. [1] This explains why phycocyanin is so attractive to the global market.
More on Fluorescence
Another important property of phycocyanin is its fluorescence. Indeed, during extraction procedures, phycobilisomes are disrupted. The energy that is usually harvested by phycobiliproteins that can’t be transferred to an electron acceptor, is then released through an intense fluorescent emission.
PC’s fluorescence is characterized by a higher quantum yield and molar extinction coefficient than other natural fluorochromes, making these molecules interesting as fluorescent markers for several applications.
Final Word
In the end, phycocyanin is being studied in cancer therapies, where the compound is incorporated into several kinds of nanoparticles. [1]
Phycocyanin is a promising molecule in many different fields that has raised a particular interest among researchers in the last decade.
However, its low stability represents a challenge both for extraction and purification methods and for applications.
Many efforts are being made to find solutions since the amount and the properties of this compound make it very attractive from a sustainable development perspective.
References:
- Ashaolu, T. J.; Samborska, K.; Lee, C. C.; Tomas, M.; Capanoglu, E.; Tarhan, Ö.; Taze, B.; Jafari, S. M. Phycocyanin, a Super Functional Ingredient from Algae; Properties, Purification Characterization, and Applications. International Journal of Biological Macromolecules. Elsevier B.V. December 15, 2021, pp 2320–2331. https://doi.org/10.1016/j.ijbiomac.2021.11.064;
- Pagels, F.; Guedes, A. C.; Amaro, H. M.; Kijjoa, A.; Vasconcelos, V. Phycobiliproteins from Cyanobacteria: Chemistry and Biotechnological Applications. Biotechnology Advances. Elsevier Inc. May 1, 2019, pp 422–443. https://doi.org/10.1016/j.biotechadv.2019.02.010;
- Fernández-Rojas, B.; Hernández-Juárez, J.; Pedraza-Chaverri, J. Nutraceutical Properties of Phycocyanin. Journal of Functional Foods. Elsevier Ltd 2014, pp 375–392. https://doi.org/10.1016/j.jff.2014.10.011;
- Pez Jaeschke, D.; Rocha Teixeira, I.; Damasceno Ferreira Marczak, L.; Domeneghini Mercali, G. Phycocyanin from Spirulina: A Review of Extraction Methods and Stability. Food Research International. Elsevier Ltd May 1, 2021. https://doi.org/10.1016/j.foodres.2021.110314;
- Fratelli, C.; Burck, M.; Amarante, M. C. A.; Braga, A. R. C. Antioxidant Potential of Nature’s “Something Blue”: Something New in the Marriage of Biological Activity and Extraction Methods Applied to C-Phycocyanin. Trends in Food Science and Technology. Elsevier Ltd January 1, 2021, pp 309–323. https://doi.org/10.1016/j.tifs.2020.10.043.
