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Mar. Drugs 2015, 13, 6226-6246; doi:10.3390/md13106226
marine drugs
ISSN 1660-3397
www.mdpi.com/journal/marinedrugs
Review
Marine Carotenoids against Oxidative Stress: Effects on
Human Health
Maria Alessandra Gammone 1,*, Graziano Riccioni 1,2, and Nicolantonio D’Orazio 1
1 Human and Clinical Nutrition Unit, Department of Medical Oral and Biotechnological Sciences,
Via Dei Vestini, “G. D’Annunzio” University, Chieti 66013, Italy;
E-Mails: griccioni@hotmail.com (G.R.); ndorazio@unich.it (N.D.)
2 Cardiology Unit, San Camillo De Lellis Hospital, Manfredonia 71043, Foggia, Italy
* Author to whom correspondence should be addressed; E-Mail: m.alessandra.gammone@gmail.com;
Tel.: +39-0871355731.
Academic Editor: Paul Long
Received: 29 May 2015 / Accepted: 22 September 2015 / Published: 30 September 2015
Abstract: Carotenoids are lipid-soluble pigments that are produced in some plants, algae,
fungi, and bacterial species, which accounts for their orange and yellow hues. Carotenoids
are powerful antioxidants thanks to their ability to quench singlet oxygen, to be oxidized, to
be isomerized, and to scavenge free radicals, which plays a crucial role in the etiology of
several diseases. Unusual marine environments are associated with a great chemical
diversity, resulting in novel bioactive molecules. Thus, marine organisms may represent an
important source of novel biologically active substances for the development of therapeutics.
In this respect, various novel marine carotenoids have recently been isolated from marine
organisms and displayed several utilizations as nutraceuticals and pharmaceuticals. Marine
carotenoids (astaxanthin, fucoxanthin, β-carotene, lutein but also the rare siphonaxanthin,
sioxanthin, and myxol) have recently shown antioxidant properties in reducing oxidative
stress markers. This review aims to describe the role of marine carotenoids against oxidative
stress and their potential applications in preventing and treating inflammatory diseases.
Keywords: marine carotenoids; oxidative stress; antioxidants; inflammatory diseases
OPEN ACCESS
Mar. Drugs 2015, 13 6227
1. Introduction
Oceans cover most of the earth’s surface, constituting a wide resource for the discovery of potential
therapeutics. Over the last decades, numerous substances with interesting pharmaceutical activities have
been identified in marine organisms. The diversity of marine environments provides an important source
of bioactive compounds. This may lead to potential new drug candidates of natural origin with efficacy
and low toxicity in the therapeutic strategy against many diseases characterized by cellular redox
state alterations.
In recent years it has become evident that the oxidation of lipids is a fundamental step in the
pathogenesis of several diseases, in both adult and infant patients. Lipid peroxidation is a process naturally
generated in small amounts in the body, mainly by the effect of some reactive oxygen species (ROS),
such as hydroxyl radical and hydrogen peroxide. Both enzymatic and non-enzymatic natural antioxidant
defenses exist; however, these protective mechanisms may be overcome. Then a self-propagating
chain-reaction starts and oxidative stress can result in significant tissue damage.
Oxidative stress and chronic inflammation are the main pathophysiological factors contributing to the
development of chronic inflammatory diseases, such as diabetes, atherosclerosis, and hypertension.
Appropriate and effective interventions, including nutrition, pharmacology, and physical exercise, are
necessary in order to activate the expression of cellular antioxidant systems, thus preventing
inflammatory and degenerative processes. In fact, inflammatory diseases derive from a continuum of
patho-physiological processes: for example, cardiovascular disorders advance from a local redox
disequilibrium to endothelial dysfunction, inflammation, and excessive vascular remodeling, which
slowly leads to atherosclerosis and subsequent cardiovascular accidents such as coronary artery disease,
myocardial infarction, and stroke [1]. A nutritional approach through natural antioxidant substances
represents an important new frontier in both the prevention and treatment of cardiovascular diseases.
Scientific evidence supports the beneficial roles of phytochemicals against some inflammatory and
chronic diseases. Several naturally-occurring antioxidant bioactives have been associated with
their prevention.
For example, many carotenoids with great antioxidant properties displayed a risk reduction both in
epidemiological studies and supplementation human trials, indicating the presence of a strong link
between oxidative stress, a pro-inflammatory systemic environment, and a wide number of chronic
diseases [2]. Consequently, consistent dietary improvement may shift human health toward decreased
morbidity and mortality as well as a better quality of life.
2. Oxidative Stress: The Role of Antioxidants
ROS are molecules containing oxygen, with unpaired valence electrons. They are generated as a
natural product of normal cellular functioning and oxygen metabolism and have important roles in both
cell signaling and intercellular homeostasis. ROS effects on cells include not only roles in apoptosis
(programmed cell death) but also positive effects, such as the induction of host defense genes [3], the
stimulation of the adaptive immune system via the recruitment of leukocytes, and mobilization of ionic
transport systems in the so-called redox or oxidative signaling. However, ROS levels can augment
dramatically due to environmental stress, such as ionizing radiation, UV, or heat exposure [4]; this
Mar. Drugs 2015, 13 6228
increase can result in significant cellular damage, which cumulatively constitutes oxidative stress. ROS
can damage DNA, RNA, and proteins; they can determine oxidations of both amino acids in proteins
and polyunsaturated fatty acids in lipids and can oxidatively inactivate specific enzymes by oxidation of
co-factors, thus contributing to the physiology of aging. Oxidative stress caused by the imbalance
between ROS and biological antioxidant systems and consequent oxidative stress can lead to
modification of these macromolecules (Figure 1); subsequently, in the case of excessive amounts, ROS
can determine deleterious effects [5]. Free radicals play a crucial role in the progression of many
pathologies, such as atherosclerotic processes [6], myocardial and cerebral ischemia [7], renal failure [8],
rheumatoid arthritis [9], inflammatory bowel disease, retinopathy of prematurity, asthma, Parkinson’s
disease, kidney damage, preeclampsia [10], and more general inflammation, as well as all the chronic
degenerative diseases. In fact, ROS can attack the polyunsaturated fatty acids in the cell membrane,
initiating a self-propagating chain reaction: this peroxidative rupture of cellular membranes and the
end-products of such lipoperoxidation reactions are dangerous for cells and tissues, in a self-propagating
chain-reaction whereby the initial oxidation of only a few molecules can result in significant tissue
damage. Oxidative stress is a powerful contributor to aging, although the accumulation of oxidative
damage and its implications for senescence depends on the particular tissue type where it occurs.
Normally, cells defend themselves against ROS damage through intracellular and extracellular defenses,
in particular through enzymes such as superoxide dismutases (SOD), catalases (CAT), lactoperoxidases,
and glutathione peroxidases. Exogenous antioxidants such as ascorbic acid (vitamin C), tocopherol
(vitamin E), and polyphenols also play important roles in preventing ROS damage by scavenging
free radicals.
Exogenous Antioxidant systems Endogenous
factors dysfunction factors
OXIDATIVE STRESS
DNA oxidation
*
Lipid
peroxidation Protein oxidation Glycoxidation
8-OHdG MDA Protein carbonyl AGP
8-nitroguanine Isoprostane Nitrotyrosine
POLYUNSATURATED
FATT Y AC ID
LIPID FREE RADICAL
LIPID PEROXYL RADICAL
LIPID HYDROPEROXIDE
MALONDIALDEHYDE
*
R –CH =CH –CH
2
–R
R –CH =CH –C’H –R
R –CH =CH –CH –R
|
O
|
O
R –CH =CH –CH –R
|
O
|
OH
R –CH =CH
2
–CH –R
|| ||
O O
(a) (b)
Figure 1. Sources, effects, and markers of oxidative stress. In detail, (a) schematic steps of
MDA formation from polyunsaturated fatty acids (MDA: malondialdehyde; AGP: advanced
glycation end-products; 8-OHdG: 8-hydroxy-2′-deoxyguanosine); (b) Lipid peroxidation
and MDA production.
In a similar manner, carotenoids’ pigments, which have been shown to possess strong antioxidant
properties, have been attracting increasing attention due to their beneficial effects on human health, in
particular because of their potential against cancer and cardiovascular diseases [11].
Mar. Drugs 2015, 13 6229
3. Bioactivity and the Protective Effects of Natural Carotenoids: New Perspectives from the Sea
Carotenoids are lipid-soluble pigments produced by some plants, algae, fungi, and bacterial species.
They are responsible for some food’s orange-yellow hues. Carotenoids, which play a crucial role in the
complex network of antioxidant phytochemicals, should certainly be constituents of a healthy diet. They
are excellent light filters and efficient quenchers of both singlet oxygen and excited triplet state
molecules. Their lipophilicity determines their peculiar sub-cellular distribution: they are more
represented in membranes and lipophilic cell compartments. This makes them suitable photo-protectants,
not only for plants but also for humans. In fact, carotenoids absorb light, thus providing photo-protection
and defense from photo-oxidative damage not only to photo-synthetic organisms, but also to the eye and
the skin. Skin protection involves the carotenes β-carotene and lycopene, while protection of the macula
involves the xanthophylls zeaxanthin and lutein.
Carotenoids are antioxidants thanks to their ability to quench singlet oxygen, to be oxidized, and to
be isomerized. The protection mechanisms involve singlet oxygen quenching and free radicals scavenging.
However, they scavenge reactive free radicals and become carotenyl radicals after reaction through
hydrogen abstraction: this process can lead to a switch from a beneficial antioxidant process to a
damaging pro-oxidative one [12]. This potential antioxidant role has been suggested to be the main
mechanism for their preventive effects against cancer and inflammatory diseases. About 700 carotenoids
with different structures have been isolated from natural source; the evaluation of their pharmaceutical
potential may be a promising field of medical research.
However, the carotenoid species so far studied for this purpose are restricted to a small number,
including the dicyclic β-carotene, β-cryptoxanthin, canthaxanthin, α-carotene and lutein, and the acyclic
carotenoid lycopene. Typical carotenoids, as well as marine ones, displayed a wide range of beneficial
effects on human health.
In this respect, novel marine carotenoids, such as fucoxanthin, astaxanthin, zeaxanthin, and, more
recently, rare marine carotenoids such as sioxanthin, saproxanthin, myoxol, and siphoxanthin, are
gaining attention and need to be evaluated for their important potential as development materials for
pharmaceuticals or functional foods, in order to prevent such disorders as cancer and cardiovascular
diseases. Marine carotenoids are important bioactive compounds principally derived from algae, with
antioxidant activities deriving from their chemical structure and interaction with biological membranes.
These bioactive substances recently showed unique and remarkable properties that explain their
potentially beneficial effects on human health.
The potential benefits of marine carotenoids have been studied particularly in astaxanthin and
fucoxanthin, which are the main marine carotenoids [13]. Both carotenoids show strong antioxidant
activity, which is attributed to quenching singlet oxygen and scavenging free radicals.
3.1. Astaxanthin
Astaxanthin (Figure 2), a red carotenoid pigment belonging to the xanthophylls class, was shown to
prevent lipid peroxidation in biological membranes and to support human health even more effectively
than other antioxidants. It has been approved as a nutraceutical by the United States Food and Drug
Administration since 1999 [14].
Mar. Drugs 2015, 13 6230
Figure 2. The molecular structure of astaxanthin.
Astaxanthin, used in nutritional supplements, is usually a mixture of configurational isomers
produced by the microalga Haematococcus pluvialis. As an antioxidant, it scavenges free radicals and
other oxidants and protects the lipid bilayer from peroxidation with its polar ionic rings and
non-polar conjugated carbon–carbon bonds, with an antioxidant property 10-fold greater than other
carotenoids, including lutein, canthaxanthin, and β-carotene. In fact, astaxanthin and some derivatives
can scavenge superoxide anion radicals (O2−) [15]; astaxanthin and its derivatives inhibit H2O2-mediated
activation of the transcription factor NF-κB (nuclear factor kappa-light-chain-enhancer of activated B
cells), which controls the expression of inducible genes such as heme oxygenase 1 (HO-1), which
represents a marker of oxidative stress) and iNOS, implicated in the inflammatory response and engaged
in cellular defenses against oxidative stress. Subsequently, astaxanthin also blocks its downstream
cytokine production by modulating protein tyrosine phosphatase-1 expression [16]. Astaxanthin can
interact with different radicals because the pattern of conjugated double bonds in its polyene backbone
explains its ability to quench singlet oxygen [17] through the transfer of excitation energy to the
carotenoid. In this process of quenching, astaxanthin remains intact so that it can undergo further cycles
of singlet oxygen quenching [18]. As a consequence, astaxanthin supplementation not only lowers ROS
levels but also leads to an important functional recovery of the antioxidant network including both the
enzyme SOD, which alternately catalyzes the dismutation of O2− into molecular oxygen (O2) and H2O2,
and CAT, which protects the cell from oxidative damage by catalyzing the decomposition of H2O2− to water
and O2 [19]. Astaxanthin displayed antioxidant properties even stronger than vitamin E and β-carotene [20]:
it provides protection against UV light damage and age-related diseases; it promotes the immune
response in the liver and kidneys, but also in the eyes and joints; it protects phospholipids from
peroxidation [21]; and it was responsible for important shifts in phlogistic response. For example, the
treatment of Helicobacter pylori-infected mice with astaxanthin was demonstrated to reduce gastric
inflammation and bacterial load and even to modulate cytokine release by splenocytes [22]. In addition,
recent clinical studies showed a significant reduction in the cardiovascular risk markers of oxidative
stress and inflammation [23], as well as an important improvement in blood status [24]. Astaxanthin has
considerable potential for both the prevention and treatment of various chronic inflammatory disorders,
such as cancer, asthma, rheumatoid arthritis, metabolic syndrome, diabetes, and diabetic nephropathy,
as well as gastrointestinal, hepatic, and neurodegenerative diseases [2]. In addition, the effects of
astaxanthin and n-3 polyunsaturated fatty acids in combination are synergistic: astaxanthin offered both
antioxidant and anti-apoptotic activities to neutrophils, through improving their glutathione-based
redox equilibrium.
Mar. Drugs 2015, 13 6231
Thus, habitual consumption of marine products such as fish and microalgae, which are natural sources
of both astaxanthin and n-3 polyunsaturated fatty acids (PUFAs), may be associated with a significant
improvement in immune response and may lower risks for both vascular and infective diseases [25].
Astaxanthin also displayed a neuro-protective property due to its antioxidant activities: in this respect,
its age-dependent and region-specific antioxidant action in the mouse brain was recently investigated.
Treated animals were given 2 mg/kg/day astaxanthin for four weeks. The level of non-enzymatic
oxidative marker malondialdehyde (MDA), nitric oxide (NO), advanced protein oxidation product, and
glutathione (GSH), and the activity of enzymatic antioxidants SOD and CAT were determined from the
isolated brain regions. Astaxanthin supplementation markedly decreased the level of MDA, NO, and
advanced protein oxidation product in the cortex, striatum, hypothalamus, hippocampus, and cerebellum.
Treatment with astaxanthin increased the activity of CAT and SOD enzymes as well as the level of GSH
in the brain. This noticeable improvement in oxidative markers was more important in the young group
compared to the aged one, thus resulting in an age-dependent antioxidant effect of astaxanthin [26]. A
similar result was achieved in animal models of autism (which had been obtained through prenatal
exposure to valproic acid), where astaxanthin displayed neuro-protective effects due to its antioxidant
mechanism [27]. In fact, oxidative stress leads to rapid changes in the antioxidant system, such as the
dropping of the cellular endogenous antioxidant GSH, and may result in cell damage and even cell death,
which may be responsible for autistic disorders [28]. Behavioral tests were conducted wherein oxidative
stress markers (such as lipid peroxidation, advanced protein oxidation product, NO, and GSH) and the
activity of SOD and CAT were estimated to confirm the mouse model of autism and assess the effect of
astaxanthin. An increased level of oxidative stress was found by determining these different oxidative
stress markers and astaxanthin significantly reduced the oxidative stress in the brain and liver, thus
improving the behavioral disorder [27]. Hence, prenatal exposure to valproic in pregnant mice leads to
the development of autism-like features, while astaxanthin improves the impaired behavior presumably
by its antioxidant activity.
In addition, a very recent study demonstrates that astaxanthin also protects steroidogenesis from
oxidative stress in Leydig cells. In fact, H2O2 induces oxidative stress and influences protein kinase A
(PKA), a family of enzymes whose activity is dependent on cellular levels of cyclic adenosine
monophosphate (c-AMP), which has several cell functions including regulation of glycogen, sugar, and
lipid metabolism. Oxidative stress attenuates the post-PKA pathway, thus resulting in suppressed
expression of the mature form of steroidogenic acute regulatory protein (StAR), a transport protein that
regulates cholesterol transfer. Astaxanthin prevents the downregulation of the mature form of the StAR
protein and restores steroidogenesis in the Leydig cell through a reduction of ROS formation caused by
H2O2 [29].
Some evidence also revealed a potential therapeutic value of astaxanthin in pulmonary fibrosis
treatment through promotion of myofibroblast apoptosis. A very recent study investigated the anti-fibrotic
effect of astaxanthin on the promotion of myofibroblast apoptosis based on dynamin-related protein-1
(Drp1)-mediated mitochondrial fission in vivo and in vitro [30]. Results showed that astaxanthin can
inhibit lung parenchymal distortion and collagen deposition, as well as promote myofibroblast apoptosis.
Astaxanthin demonstrated pro-apoptotic function in myofibroblasts by contributing to mitochondrial
fission, thereby leading to apoptosis by increasing Drp1 expression and enhancing Drp1 translocation
into the mitochondria. Drp1-associated genes, such as Bcl-2-associated X protein, cytochrome c, tumor
Mar. Drugs 2015, 13 6232
suppressor gene p53, and a p53-upregulated modulator of apoptosis, were highly upregulated in the
astaxanthin group [30]. Hence, astaxanthin provides a potential preventive and therapeutic strategy in
pulmonary fibrosis by promoting myofibroblast apoptosis through a Drp1-dependent molecular pathway.
Therefore, daily consumption of such marine products is a beneficial strategy in human health
management. It can also help with fighting oxidative stress in healthy subjects whose free radical
production is accentuated because of physical exercise, such as athletes [31].
These health-promoting effects of astaxanthin make it a high-value carotenoid and a novel potential
treatment for oxidative stress and inflammation, not only against cardiovascular pathology [32] but also
against other important inflammatory diseases.
3.2. Fucoxanthin
Fucoxanthin (Figure 3), another carotenoid, can be found in brown seaweeds such as Undaria
pinnatifida, Hijikia fusiformis, Laminaria japonica, and Sargassum fulvellum. It belongs to the class of
xanthophylls and non-provitamin A carotenoids. Its structure possesses an unusual allenic bond, an
epoxide group, and a conjugated carbonyl group in a polyene backbone, which confers antioxidant
properties [33]. Dietary ingested fucoxanthin is converted to amarouciaxanthin A, which is stored in
abdominal white adipose tissue (WAT) via fucoxanthinol in mice. It is hydrolyzed to fucoxanthinol in
the gastrointestinal tract by digestive enzymes such as lipase and cholesterol esterase, and then converted
to amarouciaxanthin A in the liver [34]. These metabolites are considered to be the active forms exerting
physiological and biological functions in the body. Fucoxanthin was demonstrated to be effective in the
reduction of major cardiovascular risk factors such as obesity, diabetes, hypertension, chronic
inflammation, plasma, and hepatic triglyceride levels, as well as cholesterol concentrations [35]. In
particular, fucoxanthin was found to induce in white adipose tissue (WAT) both protein and mRNA
expression of UCP1, a protein situated in the mitochondrial inner membrane dissipating the pH gradient
of oxidative phosphorylation, thus releasing chemical energy as heat, and which is normally expressed
only in brown adipose tissue (BAT). UCP1 induction by fucoxanthin metabolites accumulated in WAT
increases the amount of energy released as heat in fat tissue and leads to oxidation of fatty acids and heat
production in WAT [36]. Physiologic bodily metabolism determines heat production; this process is
named thermogenesis, and UCP-1 dissipates the pH-gradient generated by oxidative phosphorylation,
through releasing chemical energy as heat. UCP1 gene expression, which is stimulated by many factors,
such as cold, β3-agonists, adrenergic stimulation, and thyroid hormones, represents a significant part of
body energy expenditure, its dysfunction being an important cause of weight gain and a significant
cofactor for the development of obesity. Fucoxanthin augments the amount of energy that is released as
heat in fat tissue, thus stimulating thermogenesis [36]. UCP-1 and mRNA could be detected in WAT
when experimental animals received Undaria lipids containing fucoxanthin: 0.2% fucoxanthin in their
diet significantly attenuated weight gain in mice by increasing UCP-1 expression. This UCP1 induction
in white adipose tissue (WAT) by fucoxanthin and its derivatives leads to fatty acid oxidation and heat
production in WAT [35]. This adaptive thermogenesis plays a crucial role in energy expenditure as heat,
in order to limit weight gain and favor weight loss.
Mar. Drugs 2015, 13 6233
Figure 3. The molecular structure of fucoxanthin.
Fucoxanthin was found to promote not only UCP1 protein and mRNA expression in WAT of obese
animals but also β3-adrenergic receptor (Adrb3), which is responsible for lipolysis and thermogenesis [36].
This increased sensitivity to sympathetic nerve stimulation may lead to a further upregulation of fat
oxidation in WAT. A clinical study on humans [37] tested the effects of 16-week supplementation with
fucoxanthin in obese patients with non-alcoholic fatty liver disease (NAFLD), providing a significant
reduction of body weight, fat, and systolic/diastolic blood pressure, decreased levels of TG, C-reactive
protein (CRP), and some enzymes such as glutamic pyruvic transaminase (GPT), glutamic oxaloacetic
transaminase (GOT), and gamma-glutamyl transpeptidase (gamma-GT), and a significant increase in
resting energy expenditure (REE) measured by indirect calorimetry. Supplementation with 4.0 mg/day
fucoxanthin led to an important increase in REE and an even greater increase was observed in the group
taking fucoxanthin at a dose of 8 mg. A significant reduction in body weight and fat in obese subjects
results in the downregulation of inflammatory markers and helps prevent metabolic syndrome. The
potential anti-diabetic action of fucoxanthin is attributable to the ability of this marine bioactive to induce
weight loss and WAT reduction, so that it helps decrease insulin resistance. In fact, the chronic
low-grade inflammation elicited by pro-inflammatory mediators in the WAT leads to decreased insulin
sensitivity [38]. A recent study showed that the metabolite fucoxanthinol also prevents inflammation
and insulin resistance by inhibiting NO and prostaglandin E2 (PGE2) production through the
downregulation of both iNOS and cyclooxygenase-2 (COX-2) mRNA expression, which are related to
the pathogenesis of inflammation, as well as adipokine secretion in WAT [39].
In addition, fucoxanthin might alter the plasma leptin level in order to achieve its anti-obesity action.
Many previous studies reported that leptin secretions are elevated due to the accumulation of fat in
adipocytes; leptin could control body weight and adipose fat pads through the regulation of energy
expenditure. In particular, Park et al. [39] performed a study evaluating the beneficial effect of
Undaria pinnatifida ethanol extract on C57BL/6J mice and found that fucoxanthin could significantly
decrease the plasma leptin level and that it was associated with a significant decrease in the epididymal
adipose tissue weight. In this study, fucoxanthin supplementation reduced the adipocyte size remarkably
compared to the control group. Fasting blood glucose, plasma leptin, and insulin levels were significantly
higher in the control group by 1.5- to 2.3-fold. Fucoxanthin significantly lowered blood glucose levels
by 19.8% and blood insulin levels by about 33%, compared to the control. The plasma leptin concentration
showed a positive correlation with body weight and was lower after fucoxanthin supplementation.
Another relevant study displayed that fucoxanthin downregulates stearoyl-coenzyme A desaturase-1
(SCD1), with subsequent improvement of insulin and leptin sensitivity, thus contributing to the
prevention of obesity [36].
Mar. Drugs 2015, 13 6234
Another metabolic benefit of fucoxanthin was the promotion of docosahexaenoic acid (DHA) synthesis
in the rodent liver, thus resulting in improved lipid profiles. Supplementation of fucoxanthin or its
derivatives consistently attenuated body and visceral fat weight gain as well as lipid accumulation in the
liver, decreases insulin resistance, and improves the plasma lipid profile in rodents fed a high-fat diet.
However, it should be noted that in diabetic/obese KK-Ay mice with genetically compromised insulin
signaling, fucoxanthin might increase the plasma levels of cholesterol and low-density lipoproteins. These
beneficial metabolic effects of fucoxanthin are apparently mediated by the hormones leptin and
adiponectin through their common target, adenosine monophosphate-activated protein kinase (AMK),
resulting in a downregulation of lipogenic enzymes and an upregulation of lipolytic enzymes [40]. In
addition, experiments on stroke-prone spontaneously hypertensive rats show the possible protective role
of fucoxanthin against cerebrovascular accidents, even if the metabolic boost from taking fucoxanthin
did not stimulate the central nervous system. Thus fucoxanthin might have a potential role in the
modulation and prevention of human diseases, particularly in reducing the incidence of cardiovascular
diseases [41]. Fucoxanthin was proved to be safe and without side effects, and provided numerous health
benefits. Therefore, a fucoxanthin-rich diet could reduce body fat accumulation and modulate blood
glucose and insulin levels through the regulation of cytokine secretions from WAT.
Apart from the metabolic benefits, fucoxanthin and its metabolite fucoxanthinol have recently been
evaluated against proliferation of estrogen-sensitive MCF-7 and estrogen-resistant MDA-MB-231 breast
cancer cell lines [42]. These cell lines were stimulated with 10–20 μM of fucoxanthin or fucoxanthinol,
followed by cell viability assays and immunofluorescence to evaluate apoptosis, as well as mRNA and
protein extractions for changes in NF-κB expressions and nuclear translocations. Fucoxanthin and
fucoxanthinol reduced the viability of MCF-7 and MDA-MB-231 cells in a time-dependent manner as a
result of increased apoptosis. In both cell lines, the modulatory action of fucoxanthinol on members of
the NF-κB pathway was more pronounced than fucoxanthin in reducing nuclear levels of NF-κB
members p65 and p52 and the transcriptional factor RelB [42]. Hence, fucoxanthinol and fucoxanthin
could be effective for the treatment and/or prevention of breast cancer, thus opening new frontiers in
anticancer research.
3.3. Zeaxanthin
Zeaxanthin (Figure 4) is another oxygenated non-provitamin A carotenoid that consists of a
40-carbon hydroxylated compound identical to lutein [43]. Dietary sources of this xanthophyll include
corn, eggs, orange, honeydew melon, and green leafy vegetables, but it can be also found in marine
sources. Gramella oceani sp., a zeaxanthin-producing bacterium of the family Flavobacteriaceae, was
recently isolated from marine sediment off coastal Taiwan [44]. Xanthophyll has also been identified in
Gramella planctonica sp. nov., Aquibacter zeaxanthinifaciens sp. nov., and Kordia aquimaris, and in
algae such as Rhodophyta spp. and Spirulina spp. [45].
Mar. Drugs 2015, 13 6235
Figure 4. The molecular structure of zeaxanthin.
The human retinal area involved in central vision is named macula lutea due to its yellow coloration
from lutein, but it also contains zeaxanthin, whose localization is more centralized than lutein. Both
lutein and zeaxanthin intake are positively correlated with augmented macular pigment density, which
lowers the risk for age-related macular degeneration (AMD). Numerous population studies indicate
lower rates of AMD among subjects with higher blood levels of zeaxanthin, mostly because of its
antioxidant protection of retinal tissue but also because of its ability to filter out the damaging blue light [46].
Zeaxanthin not only directly scavenges ROS but also prevents protein, lipid, or DNA oxidative damage
by regulating other antioxidant mechanisms such as intracellular GSH. This protective effect of
zeaxanthin is comparable to α-tocopherol: supplementation with either zeaxanthin or α-tocopherol
reduces oxidized glutathione (GSSG) and augments intracellular reduced glutathione levels and the
GSH:GSSG ratio in response to oxidative stress [47]. In this sense, zeaxanthin acts as a direct antioxidant
but also an indirect one, by regulating GSH synthesis and levels, so that the intracellular redox status
upon oxidative stress is ameliorated while susceptibility to (H2O2)-induced cell death declines [48].
An important clinical trial demonstrating the long-term benefit of supplemental carotenoids was the
Age-Related Eye Disease Study (AREDS), which meant to learn more about the natural history and risk
factors of AMD and cataracts. It evaluated the effect of high doses of vitamin C, vitamin E, β-carotene,
and zinc, showing that high levels of these antioxidants significantly decreased AMD progression and
vision loss risk [49]. Subsequently, ARDS2 introduced some modifications to the initial formulation by
substituting lutein and zeaxanthin for β-carotene, since prior studies had shown an increased risk of lung
cancer in smokers. Lutein and zeaxanthin together appeared to be a safe and effective alternative
to β-carotene.
Recent studies have shown that, in addition to traditional mechanisms, zeaxanthin can influence cell
viability and function through various signal pathways or transcription factors.
It has been reported that zeaxanthin decreased the upregulation of vascular endothelial growth factor
(VEGF) in the retina of diabetic rats and in apolipoprotein-deficient mice [50].
Zeaxanthin was very recently shown to block hypoxia-induced VEGF secretion in cultured human
retinal pigment epithelial cells; additionally, it may have a broader effect on the control of angiogenesis
caused by factors other than VEGF through inhibition of hypoxia-induced accumulation of
hypoxia-inducible factors-1α (HIF-1α). Taken orally, zeaxanthin could be used as an adjunct to
intravitreal anti-VEGF therapy, thus enabling a decreased frequency of injections with a consequently
reduced risk of local side effects [51]. Therefore, it could be a promising agent to be explored for the
prevention and treatment of a variety of retinal diseases associated with revascularization.
In addition, similarly to β-carotene, zeaxanthin was inversely correlated with common carotid artery
stiffness, as well as elastic modulus and pulse wave velocity. The Beijing Atherosclerosis Study and the
Los Angeles Atherosclerosis Study had already shown the inverse association between plasma lutein
Mar. Drugs 2015, 13 6236
and early atherosclerosis; their follow-up trials and further studies confirmed that higher plasmatic levels
of zeaxanthin could be protective against early atherosclerosis, too [52]. These results indicated that
zeaxanthin may be beneficial not only to eyes but also to cardiovascular health.
Even if evidence from AREDS2 and other studies suggests that lutein and zeaxanthin could be more
appropriate than β-carotene in supplements [53], more prolonged follow-up will provide further
information on the biological mechanisms, duration of trial effects, and potential late effects of
intervention with these antioxidants.
3.4. β-Cryptoxanthin
β-Cryptoxanthin (Figure 5) is a xanthophyll with pro-vitamin A activity; its best dietary sources are
orange fruits such as oranges, peaches, tangerines, and tropical fruits, especially papaya, but also marine
sources, such as Nanochlorum eucaryotum, a novel marine alga with unusual biological characteristics [54].
Figure 5. The molecular structure of β-cryptoxanthin.
Epidemiological studies showed that β-cryptoxanthin improves respiratory function and lowers lung
cancer rates; prospective studies recognized its dietary intake as protective [55] since its antioxidant
potential protects against the oxidative damage that can result in inflammation. In tissue culture,
β-cryptoxanthin demonstrated a direct stimulatory effect on bone formation and, at the same time, an
inhibitory effect on bone re-absorption [56]. Large prospective population-based studies, such as the
European Prospective Investigation of Cancer Incidence (EPIC)-Norfolk study [57] as well as the Iowa
Women’s Health Study [58], showed that increased β-cryptoxanthin intake was associated with a
reduced risk of developing inflammatory diseases such as inflammatory poly-arthritis, synovitis, and
rheumatoid arthritis. These studies did not show the same beneficial effect for β-carotene, lutein, and
zeaxanthin; the influence of β-cryptoxanthin on some inflammatory markers is probably stronger than
other carotenoids. Further epidemiologic studies showed that CRP and oxidized low-density lipoprotein
(LDL)-cholesterol plasmatic levels are inversely linked to circulating antioxidants, including β-cryptoxanthin
concentrations [59]. In addition, a recent report showed an inverse correlation between β-cryptoxanthin
serum concentration and obesity, which was directly related to CRP in the general population instead [60].
Therefore, β-cryptoxanthin may also be associated with a decreased cardiovascular risk and
consequently with a reduced morbidity and mortality.
In addition, the cancer preventive effect of β-cryptoxanthin has been widely described in population
studies. The relation of head and neck cancer (HNC) risk with the intake of carotenoids was recently
explored: the analysis included over 6000 subjects with oral, laryngeal, and pharyngeal cancer,
categorized by quintiles of carotenoid intake from natural sources. Higher intake of β-cryptoxanthin was
associated with a reduction of at least 18% in the rate of oral and pharyngeal cancer and a 17% reduction
Mar. Drugs 2015, 13 6237
in the rate of laryngeal cancer. The overall protective effect of β-cryptoxanthin on HNC was stronger for
subjects reporting greater tobacco or alcohol consumption [61]. Hence, a diet rich in carotenoids may
protect against HNC, especially in persons with high risk.
A very recent human intervention study [62] focused on the therapeutic potential of β-cryptoxanthin
individually and in combination with oxaliplatin in colon cancer: β-cryptoxanthin decreased the
proliferation of cancer cells and cooperated with oxaliplatin to induce apoptosis through the negative
regulation of NH2-terminally truncated p73 (ΔNP73). The administration of anti-tumoral drugs such as
oxaliplatin can decrease in the presence of β-cryptoxanthin to achieve same percentage of growth
inhibition. Thus, a putative novel therapeutic strategy for the treatment of colon cancer could be based
on the combination of β-cryptoxanthin and oxaliplatin. The combined regimen produced greater benefits
than either individual modality, without increasing side effects. Additionally, the concentration-limiting
toxicity of oxaliplatin is reduced in presence of this antioxidant carotenoid [62].
In addition, the anti-metastatic effect of β-cryptoxanthin (0.2 μM) was assessed in Taiwanese and
American populations using human hepatocarcinoma SK-Hep-1 cells [63]. Results revealed an additive
inhibition on invasion, migration, and adhesion at 48 h of incubation. The anti-metastatic action of
β-cryptoxanthin and multicarotenoids involved additive reduction on the activities of matrix
metalloproteinase (MMP-2 and MMP-9) and the protein expression of Rho and Rac 1, but additive
promotion on the protein expression of MMP tissue inhibitors (TIMP-1 and TIMP-2) [63]. However,
more in vivo studies are needed to confirm these findings.
3.5. Rare Marine Carotenoids: Siphonaxanthin, Saproxanthin, and Myxol
Recent years have seen a rising trend in exploring microalgae as the demand for lutein and other
carotenoids in global markets increased dramatically. New marine resources are now under examination
and novel entities are emerging. Among these, siphonaxanthin (Figure 6) is a specific keto-carotenoid,
present in edible green algae such as Codium fragile, Caulerpa lentillifera, and Umbraulva japonica,
constituting approximately 0.1% of their dry weight, whose bio-functional properties are going to be
identified [64]. Differently from fucoxanthin, siphonaxanthin does not possess either epoxide or an
allenic bond in its structure, but contains an additional hydroxyl group on the 19th carbon that could
contribute to its strong apoptosis-inducing effect. Siphonaxanthin seems to facilitate an efficient energy
transfer of carotenoids to chlorophylls [65] and to have a light-harvesting function in underwater
habitats [66].
Figure 6. The molecular structure of siphonaxanthin.
Siphonaxanthin also proved to be a powerful inhibitor of human leukemia HL-60 cells’ viability
through induction of their apoptosis (even more than fucoxanthin) due to its double cellular uptake. This
strong pro-apoptotic effect was accompanied by reduced Bcl-2expression and subsequent activation of
Mar. Drugs 2015, 13 6238
caspase-3 and upregulation of death receptor 5 (DR5) expression [67]. Tumor Necrosis Factor (TNF)-related
apoptosis-inducing ligand (TRAIL) determines selective apoptosis in cancer cells by binding to the
trans-membrane receptors TRAIL-R1/DR4 and TRAIL-R2/DR5 with no effects on normal cells [68].
This pathway constitutes an attractive strategy in anti-cancer research and in this sense siphonaxanthin
may be a more potent growth inhibitor in cancer cells, compared to fucoxanthin, and could be a potential
chemo-preventive or chemotherapeutic agent. It is already known that dietary carotenoids exert an
anti-inflammatory effect by suppressing mast cell degranulation in vivo: astaxanthin, fucoxanthin,
β-carotene, and zeaxanthin significantly block antigen-induced degranulation of rat basophilic leukemia
cells (RBL-2H3) and bone marrow-derived mast cells through inhibition of antigen-induced
translocation of the high-affinity IgE receptor FcεRI to lipid rafts [69]. In a similar way, siphonaxanthin
was reported to exert inhibitory effects on the antigen-induced degranulation of mast cells, because it
modifies the functions of lipid rafts by localizing in the cell membrane and inhibiting the translocation
of FcεRI to lipid rafts [70].
In studies on human umbilical vein endothelial cells and the rat aortic ring, siphonaxanthin also
showed a significant anti-angiogenic activity, due to signal transduction downregulation by fibroblast
growth factor receptor-1 (FGFR-1) in vascular endothelial cells; in particular, siphonaxanthin suppresses
the mRNA expression of fibroblast growth factor 2 (FGF-2), its receptor FGFR-1, and their
trans-activation factor (EGR-1) [71]. This potential prevention of angiogenesis under pathological
conditions, such as cancer, atherosclerosis, diabetic retinopathy, and rheumatoid arthritis [72], results in
a promising approach in the prevention of cancer and other inflammatory, pro-angiogenic diseases.
Recently, three novel marine bacteria belonging to the family Flavobacteriaceae have been
isolated [73]. Two rare carotenoids, saproxanthin and myxol (Figure 7), were identified from these
strains and reported to possess powerful antioxidant action. 3R-saproxanthin has been previously
extracted from Saprospira grandis of the family Saprospiraceae; 3R,2′S-myxol has been found in marine
bacterial strain P99-3, belonging to the family Flavobacteriaceae, and in cyanobacterium Anabaena
variabilis [74]. If 2′-hydroxylase works on saproxanthin, this carotenoid is capable of being converted
into myxol. The antioxidant potential of saproxanthin and myxol has been explained by their inhibitory
activity against lipid peroxidation induced by free radicals in a rat brain homogenate and by their
neuro-protective effect against l-glutamate toxicity on the neuronal hybridoma cell line [75].
(a)
(b)
Figure 7. The molecular structures of saproxanthin (a) and myxol (b).
Saproxanthin or myxol supplementation may determine reinforcement and stabilization of biological
membranes, which decreases their permeability to oxygen and enhances protection against radical-induced
Mar. Drugs 2015, 13 6239
peroxidation. The antioxidant activities of saproxanthin and myxol were even superior to those of
zeaxanthin and β-carotene [73]. These rare novel and monocyclic marine carotenoids containing
γ-carotene skeleton need to be carefully evaluated for their potential as development materials for
pharmaceuticals or functional foods, in order to prevent oxidative stress-related diseases such as cancer
and cardiovascular pathologies.
4. Antioxidant and Pro-Oxidant Activities of Carotenoids
The ability of dietary carotenoids to act as antioxidants in biological systems is dependent upon a
number of factors. Even if their structure, in particular the conjugated double bond system, gives rise to
many fundamental properties of these molecules, it also affects how these molecules are incorporated
into biological membranes. This, in turn, alters the way these molecules interact with ROS, so that their
behavior may be different in vivo, compared to in solution. The effectiveness of carotenoids as
antioxidants is also dependent upon their interaction with other co-antioxidants, such as vitamins E and
C [76]. In addition, rather than acting as proper scavenging agents, some carotenoids and their
sub-products/metabolites activate the Nrf2-system, which triggers antioxidant gene expression in
particular cells and tissues.The transcription factor Nrf2 (nuclear factor erythroid-2-related factor 2)
activates the transcription of over 500 genes in the human genome, most of which have cytoprotective
functions. Nrf2 acts via transcription of these genes, in order to raise antioxidant responses,
mitochondrial biogenesis, energy metabolism, detoxification of carbon-containing xenobiotics or toxic
metals, and autophagy of toxic protein aggregates and dysfunctional organelles, but it also greatly lowers
many inflammatory responses [77]. Therefore, a great number of chronic diseases characterized by
oxidative stress, inflammation and impaired mitochondrial function can be treated and/or prevented by
raising Nrf2, at least in animal models. Nrf2 has been proposed to produce both lifespan and health span
extension, given the many diseases of aging characterized by oxidative stress, inflammation, and
mitochondrial dysfunction.
Health-promoting nutrients such as carotenoids, terpenoids and phenolic compounds [78] act at least
in part by raising Nrf2. Other health-promoting Nrf2-raising factors include low level oxidative stress
(hormesis), exercise, and caloric restriction. Raising Nrf2 has been found to prevent and treat a large
number of chronic inflammatory diseases, including various cardiovascular, kidney or lung diseases,
toxic liver damage, metabolic syndrome, sepsis, autoimmune disorders, inflammatory bowel disease,
human immunodeficiency virus (HIV) infection, and epilepsy. So, the induction of Nrf2-antioxidant
response elements (ARE)-mediated antioxidant enzymes by carotenoids provides a cellular defense
against oxidative stress.
In addition, some carotenoids, in particular astaxanthin, in combination with low concentrations of
docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), which are important nutritional
ingredients found in fish oil, showed a synergistic antioxidant effect in a HepG2-C8-ARE-luciferase cell
line system. The three compounds alone and in combination elevated cellular GSH levels, increased the
total antioxidant activity, and induced mRNA expression of Nrf2 and Nrf2 downstream target genes
NQO1 (which encodes NAD(P)H dehydrogenase-quinone 1), HO-1 (which encodes for heme oxygenase
1, an enzyme that is induced in response to stress), and GSTM2 (which encodes for the enzyme
glutathione S-transferase Mu 2), with synergistic antioxidant effects at low concentrations. The
Mar. Drugs 2015, 13 6240
Nrf2/ARE pathway plays an important role in the antioxidative effects induced by astaxanthin, DHA,
and EPA [79].
On the other hand, carotenoids may lose their effectiveness as antioxidants at high concentrations or
at high partial pressures of oxygen [77]. Their benefits not only seem to disappear but they can also be
converted into negative effects. In particular, a 9-cis-β-carotene-rich diet reduced mRNA levels of
CYP7a, the rate-limiting enzyme of bile acid synthesis [80], and consequently decreased cholesterol
absorption in the intestine. β-Carotene also decreased the expression of interleukin (IL)-1a, vascular cell
adhesion molecule-1 (VCAM-1), E-selectin, and genes involved in cholesterol metabolism and
excretion, such as ABCG1, ABCG5, and ABCG8; this suggests its potential to inhibit atherosclerosis
progression and, more generally, the inflammatory process in humans [32]. β-Carotene was also involved
in body fat store control [81]: in mature adipocytes, it is metabolized to arachidonic acid (RA), which
decreases the expression of peroxisome proliferator-activated receptor (PPAR)-alpha and
CCAAT/enhancer-binding protein, which are key lipogenic transcription factors, and reduces the lipid
content of mature adipocytes. A diet rich in β-carotene and fat tends toward energy expenditure;
otherwise, adipocytes store energy as fat. In fact, circulating β-carotene levels are inversely correlated
with risk of obesity and type 2 diabetes [82]. However, when β-carotene is administered as a
pharmacological supplement, it has harmful effects in some sub-populations, thus acting as a pro-oxidant
under specific conditions. Synthetic all-trans β-carotene seems to increase the incidence of lung cancer
and cardiovascular disease in smokers [83]: the Alpha-Tocopherol Beta-Carotene Cancer Prevention
(ATBC) Study, a randomized, double-blind, placebo-controlled primary prevention trial, reported that
male smokers who took beta-carotene had an 18% increased incidence of lung cancer and an 8%
increased overall mortality. The adverse effects of β-carotene were even stronger in the presence of
alcohol consumption [84]. These results, in conjunction with those from the Beta-Carotene and Retinol
Efficacy Trial (CARET) Study [85], continue to support the recommendation that β-carotene
supplementation should be avoided by smokers.
5. Conclusions
In conclusion, natural bioactives may be used for therapeutic purposes, in order to protect cells against
oxidative conditions. In particular, rare carotenoids need to be evaluated for their potential as
development materials for pharmaceuticals and/or functional foods. It is hoped that this review will
promote exploration of marine carotenoids best utilized.
Author Contributions
Maria Alessandra Gammone researched data and wrote the manuscript. Nicolantonio D’Orazio
designed the project. Graziano Riccioni contributed to constructive discussions.
Conflicts of Interest
The authors declare no conflict of interest.
Mar. Drugs 2015, 13 6241
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