25 Microalgae of the Chlorophyceae Class : Potential Nutraceuticals Reducing Oxidative Stress Intensity and Cellular Damage

Nutraceutical is a term combining the words nutrition and pharmaceutical. It is a food or food product that provides health and medical benefits, including the prevention and treatment of disease. A nutraceutical has beneficial effects because it possesses many compounds with antioxidant and intracellular signalling-pathway modulator effects. In recent years, it has been demonstrated that microalgae of the Chlorophyceae class could be excellent nutraceuticals because they contain polyphenols, chlorophyll, -carotene, ascorbic acid, lycopene, -tocopherol, xanthophylls, and PUFAs. For this reason, some research groups, including ours, have studied the nutraceutical properties of the genera Dunalliela, Haematococcus, and Chlorella. However, our research group has put special emphasis on the genera Chlorella and Chlamydomonas. For these genera, we present new results that reveal antioxidant effects in different models of oxidative stress and cell damage


Introduction
Nutraceutical is a term combining the words nutrition and pharmaceutical.It is a food or food product that provides health and medical benefits, including the prevention and treatment of disease.A nutraceutical has beneficial effects because it possesses many compounds with antioxidant and intracellular signalling-pathway modulator effects.In recent years, it has been demonstrated that microalgae of the Chlorophyceae class could be excellent nutraceuticals because they contain polyphenols, chlorophyll, -carotene, ascorbic acid, lycopene, -tocopherol, xanthophylls, and PUFAs.For this reason, some research groups, including ours, have studied the nutraceutical properties of the genera Dunalliela, Haematococcus, and Chlorella.However, our research group has put special emphasis on the genera Chlorella and Chlamydomonas.For these genera, we present new results that reveal antioxidant effects in different models of oxidative stress and cell damage

Nutraceuticals
For a long time, natural products obtained from plants have been used as prominent sources of prophylactic agents for the prevention and treatment of disease in humans, animals, and in plants.started "let food be your medicine and medicine be your food".Now, the relationship between food and drugs is getting closer.
As we enter the third millennium, with increased life expectancy and greater media coverage of the health care issue, consumers are understandably more interested in the potential benefits of nutritional support for disease control or prevention.A recent survey in Europe concluded that diet is rated more highly by consumers than exercise or the hereditary factor for achieving good health (Hardy, 2000).For that reason, many entrepreneurs seek to introduce different products into the health and nutritional market.Marketing strategies have exploited the words "functional food" and "nutraceuticals" in their advertisements.Nutraceuticals and functional foods are the fastest growing segment of today´s food industry, although nutraceuticals should be treated as pharmaceutical products as we will detail.Nutraceuticals and functional foods are a market estimated at between $6 billion US and $60 billion US and it is growing at 5% per annum.Unfortunately, entrepreneurs in an effort to make money attract, as irresponsible market entrants, products that do not comply with biosafety tests.This is because there are few laws that regulate the production and sale of such products.Because the products are not submitted for standardized toxicology testing, sometimes they may be toxic for human consumption.There are no specific regulation in any country to control nutraceuticals, and they need to be established and should be considered under the same laws that regulate pharmaceuticals and food (Bernal et al., 2011).For our purposes, we will first define "nutraceuticals" and "functional foods" and how the microalgae could be excellent nutraceuticals.
The term nutraceutical was first mentioned in 1989 to describe the union between nutrition and pharmaceuticals, both key contributors to human wellness.Stephen DeFelice MD is the founder and chairman of the Foundation for Innovation in Medicine (FIM) and he defined a nutraceutical as a food (or part of the food) that provides medicinal health benefits, including the prevention or treatment of a disease.It was proposed that a nutraceutical is not a drug, which is a pharmacologically active substance that potentiates, antagonizes, or otherwise modifies any physiological function.A nutraceutical may be a single natural nutrient in powder, tablet, capsule, or liquid form.It is not necessarily a complete food but equally not a drug (Hardy, 2000).Also, it was proposed that a nutraceutical is a product that delivers a concentrated form of a presumed bioactive agent from a food, presented in a nonfood matrix, and it is used with the purpose of enhancing health in a dosage that exceeds those that could be obtained from normal food (Zeisel, 1999).
Functional food and nutraceutical are terms used incorrectly and indiscriminately for nutrients or nutrient-enriched food that can prevent or treat disease.Functional food is a product that resembles traditional food but it possesses demonstrated physiological benefits (Shahidi, 2009).For example a functional food could be a lutein-rich food as chicken, spinach, tomatoes, or oranges, or the omega-3 fatty acids found in fish oil.All functional foods are processed and consumed as food.A nutraceutical is not a nutritional supplement because the latter are nutrients that are added to the diet to correct or prevent deficiencies of vitamins, minerals, and proteins, and often used in the recovery of a patient suffering an illness or has undergone surgery, and also taken to improve overall health (Mandel et al., 2005).The beneficial effects of nutraceuticals and functional foods have been attributed to their components, such as polyphenols, polyunsatured fatty acids (PUFAs), terpenes, chlorophyll, and accessory pigments of the photosynthetic apparatus in cyanobacteria such as Spirullina.In general these compounds are antioxidants that reduce intensity of oxidative stress or modulate intracellular communication

Nutraceutical effects of polyphenols, particularly flavonoids
The polyphenols are compounds characterized by a benzene ring bearing one or more hydroxyl groups attached to the ring.They are ubiquitous in the plants, vegetables, fruit, vines, tea, coffee and microalgae.The polyphenols in food originate from one of the main classes of secondary metabolites in plants.They are involved in the growth and reproduction and are produced as a response to defend injured plants against pathogens, and to participate in the defense mechanism against ultraviolet radiation (Biesalski, 2007).Polyphenols have different nutraceutical properties, such as an antioxidant, antiinflammatory (Biesalski, 2007), anticancer (Oz & Ebersole, 2010), antibacterial (Du et al., www.intechopen.comMicroalgae of the Chlorophyceae Class: Potential Nutraceuticals Reducing Oxidative Stress Intensity and Cellular Damage 583 2011), antiatherogenic, and antiangiogenic (Rimbach et al., 2009).There are now polyphenols with therapeutic properties for which the mechanism of action at the molecular level has been discovered and they are used in clinical trials, e.g.flavonoids.
Flavonoids comprise the most common group of polyphenols and provide much of the flavor and color to fruit and vegetables.More than 6000 different flavonoids have been described and it is estimated that humans consume about 1 g/day.
The structure of flavonoids is C6-C3-C6 and they consist of two aromatic rings linked through three carbons usually forming an oxygenated heterocycle nucleus, named the flavan nucleus, and shown in figure 1.In general, the flavonoids are classified into six groups (Grassi et al., 2009).
1. Flavones: These kinds of flavonoids are used by angiosperms to color their flowers.
Some authors have proposed that aurones are another flavonoid group, however we consider that aurones are derived from chalcones (Fowler & Koffas, 2009).
The flavonoid synthesis is shown in figure 1 The beneficial effects can be divided into 1.Antioxidants: Flavonoids suppress the formation of reactive oxygen species (ROS) either by inhibiting enzymes or chelating trace elements involved in free radical production.Thus flavonoids help maintain an ROS steady state in the case of physical and chemical injury of the cell (Corradini et al., 2011).Not all flavonoids are ROS scavengers because some flavonoids, as nucleophiles, trap electrons from the ROS and become a free radical themselves, which then propagate a chain reaction causing a deleterious effect in the cell (Grassi et al., 2009).
The mechanism of bioavailability and metabolism of particular flavonoids has been demonstrated in mammals.In general it has been shown that flavonoid absorption and metabolism occurs in a common pathway and it begins in the stomach and intestinal tract.In the small intestine flavonoids pass into the bloodstream in the form of glycosides, though esters or polymers cannot be absorbed.Some intestine cell enzymes or microorganisms of microflora hydrolyze them to be absorbed.In the bloodstream there are different thermodynamic pathways.They could interact with cells to modify intracellular communication.The polyphenols can be conjugated in the intestine or liver to form methylated, glucuronidated, or sulphated metabolites that reach the body via urinary and biliary excretion.The microflora also metabolizes some metabolized flavonoids that are secreted in the bile into the small intestine.Thus, there is a recycling of polyphenols that allow them more time in the plasma (Erdman et al., 2007;Manach et al., 2004).In general, the microalgae produce low quantities of polyphenols.For this reason, in the following parts of this chapter we give special attention to pigments and PUFAs.

Nutraceutical effects of terpenes
The terpenes are other secondary metabolites that have nutraceutical properties.The terpenes are not only the largest group of plant natural products, comprising at least 30,000 compounds, but also contain the widest assortment of structural types.Hundreds of different monoterpene (C10), sesquiterpene (C15), diterpene (C20), and triterpene (C30) carbon skeletons are known.The wealth of terpene carbon skeletons can be attributed to an enzyme class known as the terpene synthases (EC 4.2.3.20).These catalysts convert the acyclic prenyl diphosphates and squalene into a multitude of cyclic and acyclic forms.The chief causes of terpene diversity are the large number of different terpene synthases and that some terpene synthases produce multiple products.An excellent review of terpene synthase and the diversity of products were published by Degenhard and coworkers (Degenhardt et al., 2009).Microalgae produce terpenes in the form of carotenoids.These compounds offer therapeutic effects.Carotenoids are tetraterpenoid organic pigments that are naturally occurring in the chloroplasts and chromoplasts of photosynthetic organisms.The use of carotenoids by animals is because they cannot synthetize them.Animals obtain carotenoids in their diets, and they may employ them in various ways in their metabolism.
There are over 600 known carotenoids and they are divided into two classes, xanthophylls (that contain oxygen) and carotenes (that are purely hydrocarbons and contain no oxygen).Carotenoids in general absorb blue light.They serve two key roles in plants and algae; they absorb light energy for use in photosynthesis and they protect chlorophyll from photodamage (Armstrong & Hearst, 1996).
The biosynthesis of carotenes is explained in figure 2. The carotenogenesis differ somewhat among organisms and the current knowledge on the biosynthesis of carotenoids has been gained mainly from studies of bacteria and vascular plants (Armstrong & Hearst, 1996).In Figure 2, we proposed the model of Lohr for the carotenogenesis in Chlamydomonas.This is probably related to other microalgae of Chlorophyceae class (Lohr et al., 2005;Lohr, 2008).There are other major divisions in different organisms, such as diatoms (Bertrand, 2010) or plants (Cazzonelli & Pogson, 2010;Zhu et al., 2010), which references the readers can check to deepen their knowledge in this area.
There has been much interest in carotenoids, especially their effect on human health, because they have a market value of several hundred million Euros.Their chemical synthesis is still a demanding challenge for chemists.The major dietary source of vitamin A for mammals, including humans, is derived from carotenoids.Vitamin A is an essential micronutrient for cell growth, embryonic development, vision, and the function of the immune system (Jackson et al., 2008).
In general carotenoids exert their mechanism on health via an antioxidant pathway or by modulating intracellular communication.
1. Antioxidant properties: This property of carotenoids was characterized by the ability to quench singlet oxygen, the inhibition of peroxide formation, and the correlation of antioxidant dependency with oxygen partial pressures.The ketocarotenoids, such as astaxanthin and canthaxanthin, were the best radical scavengers that did not contain conjugated terminal carbonyl functions (see figure 2).These findings suggest that the keto function in conjugation with the polyene backbone is able to stabilize carboncentered radicals more effectively than the polyene backbone alone (Jackson et al., 2008).2. Modulation of intracellular communication: Carotenes modulate the intracellular communication because they or their metabolites interact with nuclear receptors like the pregnant-X-receptor (PXR) or retinoic acid receptor (RAR).For PXR it has been postulated that -carotene activated the PXR more than its metabolites.Following this pathway, the -carotene-PXR enhanced the metabolism of xenobiotics, bile acids, and retinoids (Ruhl, 2005).The carotenoids can be converted into two molecules of 9-cisretinal, which is oxidized to 9-cis-retinoic acid.The RXR binds the 9-cis-retinoic acid with high affinity to modulate cell functions (Heyman et al., 1992).Carotenoids like lycopene modulate mevalonate and Ras pathways to modify cell growth inhibition of cancerous cells (Palozza et al., 2010), and it changes Wnt and hedgehog proteins in those cells (Sarkar et al., 2010).The PI3K-Akt and MAPK pathways are stimulated in kidney by lycopene (Chan et al., 2009).
Carotenoids are lipid soluble and in general they follow the same absorption pathway as lipids, however other mechanisms of absorption have been proposed.To learn more, read the review of Kotake-Nara and Nagao (Kotake-Nara & Nagao, 2011).Once in the bloodstream, carotenes are fundamentally ligated to low density lipoprotein (LDL) whereas the xanthophylls are more evenly distributed between high density lipoproteins (HDL) and low density lipoproteins (LDL).Nonpolar carotenoids (lycopene, -carotene, -carotene) are located in the hydrophobic core and the polar (xanthophylls) would be, at least in part, on the surface of lipoproteins (Furr & Clark, 1997).For the microalgae, carotenoids are synthesized in high concentrations under several different environmental conditions, and humans exploited these as nutraceuticals in food.

Nutraceutical effects of chlorophylls, PUFA and other vitamins
There are other components in microalgae that could modulate redox environment to prevent oxidative stress and can affect intracellular communication.These components are chlorophyll, PUFAs, and vitamins such as vitamin A, B, C, and E.
Microalgae, like all chloroplast-containing photosynthetic eukaryotes, synthesize chlorophyll pigments.In Chlorophyceae chorophylls a and b are the most predominant.The chlorophylls have a porphyrin ring structure similar to heme, but with a central nonreactive magnesium ion instead of iron.To review chlorophyll biosynthesis in microalgae, read the chapter of Beale (Beale, 2008).The information about the biological activities of chlorophyll as nutraceuticals is scarce.They do have antipoliferative (Wu et al., 2010) and antioxidant (Serpeloni et al., 2011) activities.The chlorophyllin-cooper complex, a water-soluble commercial version of chlorophyll, possesses antimutagenic (Chernomorsky et al., 1997) and anticancer activities (Chernomorsky et al., 1997).The other components of microalgae; PUFAs, and vitamins A, B, C, and E, could be a nutraceutical because there is much evidence of how they modulate intracellular signals and act as antioxidants.

Chlorella genus as nutraceutic
Chlorella species are encountered in all water habitats having cosmopolitan occurrences.The species of this genus have a simple form, a unicellular green alga belonging to the Chlorophyceae family.The Chlorella sp. is morphologically classified into four types; a) spherical cells (ratio of the two axes equals one), b) ellipsoidal cells (ratio of the longest axis to the shorter axis 1.45 to 1.60), spherical or ellipsoidal cells, and globular to subspherical cells.Their reproduction is asexual.Each mature cell divides usually producing four or eight (and more rarely 16) autospores, which are freed by rupture or dissolution of the parental walls.
Our research group has used Chlorella vulgaris as nutraceutical, particularly against mercury-caused oxidative stress and renal damage.For that we used male mice that were assigned into six groups; 1) a control group that received 100 mM phosphate buffer (PB) ig and 0.9% saline ip, 2) PB + HgCl 2 5 mg/kg ip, 3) PB + 1000 mg/kg Chlorella vulgaris ig, and three groups receiving HgCl 2 + 250, 500, or 1000 mg/kg Chlorella vulgaris ig.The administration of the microalgae or PB was made 30 min before saline or HgCl 2 for 5 days.
Our results demonstrated that HgCl 2 caused oxidative stress and cellular damage, whereas Chlorella vulgaris administration prevents oxidative stress (figure 3) and cellular damage (figure 4) in the kidney (Blas-Valdivia et al., 2011).We proposed that Chlorella vulgaris's carotenes play an important role in preventing HgCl 2 -caused lipid peroxidation.Carotenes have a wide pharmacological spectrum of effects.The inhibition of lipid peroxidation may be caused by the free radical scavenging property of these compounds (Miranda et al., 2001).Carotenes can scavenge singlet oxygen and they terminate peroxides by their redox potential because of the hydroxyl group in its structure.Thus, the ROS-steady state is maintained in the kidney damage lower than in animals with mercury intoxication.The biochemical behavior of this microalgae against mercury-caused oxidative stress is similar to the purified component of cyanobacteria such as Pseudoanabaena tenuis (Cano-Europa et al., 2010) or Spirulina maxima (Sharma et al., 2007).
.  Here are some experiments that demonstrated the nutraceutical use of Chlorella (Table 3).

Study Evidences
The administration of Chlorella sp.reduces endotoxemia, intestinal oxidative stress and bacterial traslocation in experimental biliary obstruction (Bedirli et al., 2009) Chlorella administration inhibits bacterial culture and it avoids oxidative stress.
Hot water extract of Chlorella vulgaris induced DNA damage and apoptosis (Yusof et al., 2010) The extract of Chlorella vulgaris inhibited DNA synthesis, causing apoptosis and it increases p53, caspase-3, and Bax expression in hepatoma cells (HEpG2) Attenuating effect of Chlorella supplementation on oxidative stress and NFκB.Activation in peritoneal macrophages and liver of C57BL/6 mice fed on atherogenic diet (Lee et al., 2003) Chlorella supplementation decreases the NFκB activation and superoxide anion production and because it increases SOD and catalase activity Chlorella accelerates dioxin excretion in rats (Morita et al., 1999) Chlorella enhanced dioxin metabolism and excretion by feces Effect of Chlorella and its fractions on blood pressure, cerebral stroke lesions, and life-span in stroke-prone spontaneously hypertensive rats (Sansawa et al., 2006) A Chlorella supplemented diet decreases blood pressure and the incidence rate of cerebral stroke in SHRSP.

Hypocholesterolemic mechanism of
Chlorella: Chlorella and its indigestible fraction enhance hepatic cholesterol 7hydroxylase in rats (Shibata et al., 2007) Chlorella powder increases the expression of CYP7A1, a limiting enzyme of the main pathway of the cholesterol catabolism, lowering the concentration of LDL in plasma Chlorella vulgaris triggers apoptosis in hepatocarcinogenesis-induced rats (Mohd Azamai et al., 2009) Chlorella vulgaris inhibits the anti-apoptotic protein Bcl-2 Effect of Chlorella vulgaris on lipid metabolism in Wistar rats fed high fat diet (Lee et al., 2008) Chlorella vulgaris decreases HDL cholesterol concentration by a reduction in the intestinal absortion Antioxidant effect of the marine algae Chlorella vulgaris against naphthaleneinduced oxidative stress in the albino rats (Vijayavel et al., 2007) Chlorella vulgaris inhibits production of free radicals, decreasing lipoperoxidation, and increasing the activity of antioxidant enzymes as SOD, catalase, GPX and reduced glutathione, preventing from the toxicity of naftalene Six-week supplementation with Chlorella has favorable impact on antioxidant status in Korean male smokers (Lee et al., 2010) Chlorella supplement exhibits antioxidant activity decreasing ROS and increasing the activity of SOD and catalase Chlorella pyrenoidosa supplementation reduces the risk of anemia, proteinuria and edema in pregnant women (Nakano et al., 2010) Chlorella pyrenoidosa exhibits an antiinflammatory activity regulated by cytokine.It increased the production of IL-10

Study
Evidences Effect of Chlorella intake on cadmium metabolism in rats (Shim et al., 2009) Chlorella inhibits cadmium absorption and it promotes the excretion through the feces.Also, it stimulates the production of metallothionein in the small intestine.Isolation of phophorylated polysaccharides from algae: the inmmunostimulatory principle of Chlorella pyrenoidosa (Suarez et al., 2010) The Chlorella polysaccharides increases the production of NO in macrophages enhancing the innate immune response, mediated by Toll-like receptors (TLR-4) Influence of Chlorella powder intake during swimming stress in mice (Mizoguchi et al., 2011) Chlorella vulgaris exhibits an antioxidant activity, reducing the lipoperoxidation, avoiding the DNA damage.However it does not show hypoglycemic activity Table 3. Nutraceutical evidences of Chlorella.

Chlamydomonas genus as nutraceutic
Chlamydomonas spp.are unicellular algae with cell walls and with either two or four flagella.The genus Chlamydomonas is of worldwide distribution and is found in a diversity of habitats including temperate, tropical, and polar regions.Chlamydomonas species have been isolated from freshwater ponds and lakes, sewage ponds, marine and brackish waters, snow, garden and agricultural soil, forests, deserts, peat bogs, damp walls, sap on a wounded elm tree, an artificial pond on a volcanic island, mattress dust in the Netherlands, roof tiles in India, and a Nicaraguan hog wallow.These algae belong to the family Chlamydomonadaceae that consists of approximately 30 genera.DNA sequence analysis clearly demonstrates, however, that this family is composed of multiple phylogenetic lineages that do not correspond to the morphologically defined genera.Although the identities of the species are uncertain, it is noteworthy that the traits in which they differed included body shape, thickness of the cell wall, presence or absence of the apical papilla, lateral vs. basal position of the chloroplast, chloroplast position, and shape of the eyespot, all of which were later used as criteria to separate species.Although cell-body shape and size vary among Chlamydomonas species (as defined by morphological criteria), the overall polar structure, with paired apical flagella and basal chloroplast surrounding one or more pyrenoids, is constant.Cells are usually free-swimming in liquid media but on solid substrata may be nonflagellated, and are often seen in gelatinous masses similar to those of the algae Palmella or Gloeocystis in the order Tetrasporales.This condition has been referred to as a palmelloid state.Some species may also form asexual resting spores, or akinetes, in which the original vegetative cell wall becomes much thicker, and carotenoids, starch, and lipids may accumulate (Harris et al., 2008).
Our group has studied the nutraceutical properties of Chlamydomonas gloeopara, a microalgae collected from a eutrophic reservoir (La Piedad Lake) in Cuautitlan Izcalli, Mexico.That reservoir is located at 19°39´N (latitude) and 99°14´W (longitude).Our research group has used Chlamydomonas gloeopara as a nutraceutical, particularly against mercury-caused oxidative stress and renal damage.For that we used male mice that were assigned into six groups; 1) a control group that received 100 mM phosphate buffer (PB) ig and 0.9% saline ip, 2) PB + HgCl 2 5 mg/kg ip, 3) PB + 1000 mg/kg Chlamydomonas gloeopara ig, and three groups receiving HgCl 2 + 250, 500, or 1000 mg/kg Chlamydomonas gloeopara ig.The administration of the microalgae or PB was made 30 min before saline or HgCl 2 for 5 days.Our results demonstrated that Chlamydomonas gloeopara as well as Chlorella prevents renal damage (figure 5, panel A-F) by reducing the oxidative stress of lipid peroxidation (figure 5, panel G).

Haematococcus genus as nutraceutic
Haematococcus are green microalgae; single-celled aquatic organisms.It is known that Haematococcus is the primarily source of astaxanthin, a ketocarotenoid that is a natural nutritional component.In the marine environment, astaxanthin is biosynthesized in the food chain, within the microalgae or phytoplankton, at the primary production level.When these algae are exposed to harsh environmental conditions and ultraviolet light, they accumulate the highest level of astaxanthin and in this process, the algae become blood red.Astaxanthin accumulates 2% to 3% of dry weight and constitutes 85% to 88% of the total carotenoids.Chemically it is a ketocarotenoid (3,3´-dihydroxy-, -carotene-4,4´dione) and is the principal pigment of salmonoids and shrimp.Astaxanthin has a higher antioxidant activity than lutein, lycopene, or -carotene, and -tocopherol.Astaxanthin has 100 times and 10 times greater antioxidant activity than vitamin E and -carotene (Guerin, 2003).
Morphological studies have shown that the algae have a life cycle.The, green vegetative cells with two flagellae grow autotrophycally in the light and heterotrophically in the dark.In culture, H. pluvialis has the typical characteristics of a motile stage, with biflagellate spherical cells.The growth in a bioreactor, with mechanical stirring, favors the occurrence of more or less mature aplanospores.This stage becomes dominant together with the evolution of growth.The aplanospore color turns gradually red, because of the accumulation of carotenoids in the chloroplast, and especially outside of them in lipid globules (astaxanthin).The red aplanospores are known as haematocysts.This stage may appear under stress conditions caused by light, high temperature, increased salinity, nutritional limitation, or change of carbon source.During the growth stage, the cells with a diameter of 30 µm were spherical to ellipsoid and enclosed by a cell wall.The cells had 2 flagellae of equal length emerging form an anterior papilla.As they age, the cells ceased to be mobile, yet the cellular structure remained the same without the flagellae.Under stress conditions, the volume of the cells increased to a diameter of > 40 µm and the cell wall became resistant.The maturation of the cyst cells was accompanied by enhanced carotenoid biosynthesis and a gradual change in cell color to red.When the cystic cells were transferred to optimal growth conditions, daughter cells were released from the cystic cells, and then vegetative cells regenerated from daughter cells (Cysewski & Todd Lorenz, 2004).
Haematococcus has the potential as a nutraceutical because there is various evidence of this.In table 4, we show some articles that employed Haematococcus or its astaxanthin.

Study
Evidences Haematococcus astaxanthin: applications for human health and nutrition (Guerin, 2003) This is a review about the uses of astaxantin from Haematococcus in health Optimization of microwave-assisted extraction of astaxanthin from Haematococcus pluvialis by response surface methodology and antioxidant activities of the extracts (Zhao et al., 2009) The extracts have a high antioxidant capacity, inhibit peroxidation of linoleic acid, and neutralize free radicals Cardioprotection and myocardial salvage by a disodium disuccinate astaxanthin derivative (Cardax™) (Gross & Lockwood, 2004) The astaxanthin is an antioxidant, antiinflammatory, and cardioprotective.reducer of levels of nitric oxide, tumor necrosis factor alpha, and prostaglandin E2 Ulcer preventive and antioxidative properties of astaxanthin from Haematococcus pluvialis (Kamath et al., 2008) The astaxanthin exerts its gastroprotection of gastric ulceration by activation of antioxidant enzyme such as catalase, superoxide dismutase, and glutathione peroxidase.It inhibits the activity pump Na-K ATPase Safety assessment of astaxanthin-rich microalgae biomass: acute and subchronic toxicity studies in  (Stewart et al., 2008) Astaxanthin, a carotenoid with potential in human health and nutrition (Hussein et al., 2006).

The antihypertensive and neuroprotective potentials of the compound
Protective effects of Haematococcus astaxanthin on oxidative stress in healthy smokers (Kim et al., 2011).
The results suggest that ASX supplementation might prevent oxidative damage in smokers by suppressing lipid peroxidation and stimulating the activity of the antioxidant system in smokers Astaxanthin-rich extract from the green alga Haematococcus pluvialis lowers plasma lipid concentrations and enhances antioxidant defense in apolipoprotein E knockout mice (Yang et al., 2011) It results suggest that supplementation of astaxanthin-rich Haematococcus extract improves cholesterol and lipid metabolism as well as antioxidant defense mechanisms, all of which could help mitigate the progression of atherosclerosis.

Dunaliella genus as nutreutic
Dunaliella salina is a unicelular green alga belonging to the Chlorophyceae family.Dunaliella cells are ovoid, spherical, pyriform, fusiform, or ellipsoid with sizes varying from 5 to 25 µm in length and from 3 to 13 µm in width.The cells also contain a single chloroplast, which mostly has a central pyrenoid surrounded by starch granules.Dunaliella multiplies by lengthwise division, but sexual reproduction does occur rarely by isogametes with a conjugation process.It proliferates in extremely varied salinities from 0.5 to 5.0 M NaCl.The alga cells do not contain a rigid cell wall; instead a thin elastic membrane surrounds them.It is known to accumulate carotenoids under various stress conditions.It possesses a remarkable degree of environmental adaptation by producing an excess of -carotene and glycerol to maintain its osmotic balance.-carotene occurs naturally as its isomers, namely, all-trans, 9-cis, 13-cis, and 15-cis forms and functions as an accessory light harvesting pigment, thereby protecting the photosynthetic apparatus against photo damage in all green plants including algae.-carotene, a component of the photosynthetic reaction center is accumulated as lipid globules in the interthylakoid spaces of the chloroplasts of Dunaliella.They protect the algae from damage obtained during excessive irradiance by preventing the formation of reactive oxygen species, by quenching the triplet-state chlorophyll, or by reacting with singlet oxygen ( 1 O 2 ) and also act as a light filter (Ben-Amotz, 2004).Dunaliella nutraceutical properties are shown in table

Study
Conclusion In vivo antioxidant activity of carotenoids from Dunaliella salina a green microalga (Chidambara-Murthy et al., 2005) Carotenoids provide protection against CCl 4 -caused hepatic damage by restoring the activity of hepatic enzymes like peroxidase, super oxide dismutase, and catalase, which reduce ROS and lipid peroxidation.

Study
Conclusion 9-cis -carotene-rich powder of the alga Dunaliella bardawil increases plasma HDLcholesterol in fibrate-treated patients (Shaish et al., 2006) Dunaliella treatment increases plasma HDL-cholesterol and lower plasma triglyceride levels Ethanol extract of Dunaliella salina induces cell cycle arrest and apoptosis in A545 human non-small cell lung cancer cells (Sheu et al., 2008) Ethanol extract of Dunaliella salina inhibits cell proliferation and causes apoptosis possibly via p53 and p21 promoting the protein expression of Fas and FasL Protective effects of Dunaliella salina against experimental induced fibrosarcoma on Wistar rats (Raja et al., 2007).
The chlorophyta has a protective effect against experimentally caused fibrosarcoma Bioavailability of the isomer mixture of phytoene and phytofluene-rich alga Dunaliella bardawil in rat plasma and tissues (Werman et al., 2002).9-cis phytoene has a stronger antioxidative effect than the all trans isomer Hypercholesterolemia induced oxidative stress is reduced in rats with diets enriched with supplement from Dunaliella salina algae (Bansal & Sapna, 2011).
Dunaliella salina components inhibit lipid peroxidation and also increases Type1 5´iodothyronine deiodinase (5´-DI) expression, which leads to a T 3 level increase Evaluation of carotenoid extract from Dunaliella salina against cadmium-induced cytotoxicity and transforming growth factor 1 induced expression of smooth muscle -actin with rat liver cell lines (Jau-Tien et al., 2011).
Carotenoid extract of Dunaliella salina contains abundant cis and trans carotenes.These antioxidants decrease the lipid peroxidation and also inhibit activation of hepatic stellate cells (HSCs).
Protective effects of Dunaliella salina-a carotenoids-rich alga, against carbon tetrachloride-induced hepatotoxicity in mice (Hsu et al., 2008).Carotenoids of D. salina inhibit the lipid peroxidation and increases the antioxidant enzyme activity Table 5. Nutraceutical evidences of Dunaliella.

Final remarks
The functional food and nutraceutical market is growing.However, to promote health the active compounds must be ingested in high concentration.This is a great problem because sometimes the components such as carotenoids, polyphenols, and chlorophylls are extracted from vegetables or plants.In their production, we are modifying the environment, thus the use of biotechnology of microalgae or other microorganisms like bacteria or fungus could be an alternative because they may be environmentally friendly.The sun can be used as energy source and the medium could be fresh or sea water, with the carbon source as CO 2 and other inorganic or organic sources.In this chapter we show the evidence of some genera, particularly of Chlorophyceae class as Chlorella, Chlamydomonas, Haematococcus, and Dunaliella.It is evident that their components modulate intracellular communication and they act as antioxidants.

Fig. 3 .
Fig. 3. Quantification of relative kidney weight (A) and the score of kidney damage (B) of mice treated with HgCl 2 and Chlorella vulgaris.In A each bar represents the mean ± S.E.M.In B each box represents the median ± intercuartilic space.*P < 0.05 vs. control.Author right permission.Springer ©.

Fig. 4 .
Fig. 4. Quantification of lipid peroxidation (A) and reactive oxygen species in the kidneys of mice treated with HgCl 2 and Chlorella vulgaris.Bar represents the mean ± S.E.M.* P < 0.05 vs. control.Author right permission.Springer ©.

Fig. 5 .
Fig. 5. Effect on Chlamydomonas gloeopara administation on HgCl 2 -caused renal damage (panel A-F) and oxidative stress (panel G and H).Photomicrographs of renal cortex .Panel A shows control group.Panel B shows group treated with HgCl 2 .Panel C shows group treated with Chlamydomonas gloeopara 1000 mg/kg .Panels D, E and F show groups treated with Chlamydomonas gloeopara 250, 500 and 1000 mg/kg plus HgCl 2 .The tissue was stained by hematoxylin-eosin.Treatment with HgCl 2 causes cell atrophy, hyperchromatic nuclei, and edema.Histological alterations were partially ameliorated in groups treated with Chlamydomonas gloeopara.Chlamydomonas gloeopara administration reduced lipid peroxidation (G) and reactive oxygen species (H) in the kidneys of mice treated with HgCl 2 and Chlorella vulgaris.Bar is the mean ± SE * P < 0.05 vs. control.