connection between lifespan and stress-resistance for a variety of
stresses, including oxidative stress [23]. Thus, an enhanced stress
resistance may also result in an extended lifespan.
Several studies have used C. elegans for testing antioxidant
potential of compounds such as selenite [24], cocoa powder [11],
and tyrosol [25]. The protective effect of specific carotenoids or
food matrices enriched in carotenoids against oxidative stress has
been also assessed using C. elegans as experimental animal model.
In a representative experiment, the nematodes are fed with NGM
or NGM supplemented with a certain concentration of the test
compound (e.g., carotenoid or carotenoid extract) and subsequently subjected to oxidative stress induced by addition to the
nematode culture medium (solid or liquid) of an oxidant stressor
[11, 16–18]. In all of the assays there is an increase of ROS levels in
the nematode to a point where survival is decreased. After a period
of incubation with the stressor, the survival rate of the nematodes
fed with the carotenoid prior to receiving oxidative treatment
(acute or chronic) is compared with the survival rate of control
nematode population. To study the protection capacity of a specific
compound (e.g., pure carotenoid or food matrix carotenoidenriched) it is advisable, if possible, to evaluate a wide range of
concentrations of the test-compound in order to perform a doseresponse experiment. Carotenoid concentrations below the threshold level to accumulate and reach the specific tissue or cellular
targets may not enhance resistance to ROS but elevated doses
may also fail due to a prooxidant effect. It should be keep in mind
that the results obtained are highly influenced by the type of ROS,
rate of oxidative stress exposure, and the target subcellular compartment affected by the specific stressor [26, 27]. Moreover, it is
also possible that nematodes sensitive to oxidative stress at one age
are resistant to the same agent at a different age. Therefore, when
comparing results obtained in different trials and laboratories, all
these factors to design the oxidative stress experiment have to be
considered and clearly report all conditions and parameters.
By using different oxidant stressors and procedures, the
increased resistance to oxidative stress has been reported for several
carotenoids such as astaxanthin [28, 29], lutein [30], β-carotene,
lycopene, and β-cryptoxanthin [31]. This type of in vivo antioxidant assays is not only restricted to evaluate pure carotenoids or
mixtures but can also be used for testing the protection of complex
food matrices as orange juice. A successful example was reported for
sweet oranges with increased levels of β-carotene obtained through
metabolic engineering which showed 20% higher antioxidant effect
compared to control juice [32].
In this chapter we report in the animal model C. elegans a
simple and quick assay to evaluate the protection of pure carotenoids or carotenoid extracts against an acute oxidative stress by
using H 2 O 2 as oxidant agent. The protocol also includes the
390
Patricia Martorell et al.
stresses, including oxidative stress [23]. Thus, an enhanced stress
resistance may also result in an extended lifespan.
Several studies have used C. elegans for testing antioxidant
potential of compounds such as selenite [24], cocoa powder [11],
and tyrosol [25]. The protective effect of specific carotenoids or
food matrices enriched in carotenoids against oxidative stress has
been also assessed using C. elegans as experimental animal model.
In a representative experiment, the nematodes are fed with NGM
or NGM supplemented with a certain concentration of the test
compound (e.g., carotenoid or carotenoid extract) and subsequently subjected to oxidative stress induced by addition to the
nematode culture medium (solid or liquid) of an oxidant stressor
[11, 16–18]. In all of the assays there is an increase of ROS levels in
the nematode to a point where survival is decreased. After a period
of incubation with the stressor, the survival rate of the nematodes
fed with the carotenoid prior to receiving oxidative treatment
(acute or chronic) is compared with the survival rate of control
nematode population. To study the protection capacity of a specific
compound (e.g., pure carotenoid or food matrix carotenoidenriched) it is advisable, if possible, to evaluate a wide range of
concentrations of the test-compound in order to perform a doseresponse experiment. Carotenoid concentrations below the threshold level to accumulate and reach the specific tissue or cellular
targets may not enhance resistance to ROS but elevated doses
may also fail due to a prooxidant effect. It should be keep in mind
that the results obtained are highly influenced by the type of ROS,
rate of oxidative stress exposure, and the target subcellular compartment affected by the specific stressor [26, 27]. Moreover, it is
also possible that nematodes sensitive to oxidative stress at one age
are resistant to the same agent at a different age. Therefore, when
comparing results obtained in different trials and laboratories, all
these factors to design the oxidative stress experiment have to be
considered and clearly report all conditions and parameters.
By using different oxidant stressors and procedures, the
increased resistance to oxidative stress has been reported for several
carotenoids such as astaxanthin [28, 29], lutein [30], β-carotene,
lycopene, and β-cryptoxanthin [31]. This type of in vivo antioxidant assays is not only restricted to evaluate pure carotenoids or
mixtures but can also be used for testing the protection of complex
food matrices as orange juice. A successful example was reported for
sweet oranges with increased levels of β-carotene obtained through
metabolic engineering which showed 20% higher antioxidant effect
compared to control juice [32].
In this chapter we report in the animal model C. elegans a
simple and quick assay to evaluate the protection of pure carotenoids or carotenoid extracts against an acute oxidative stress by
using H 2 O 2 as oxidant agent. The protocol also includes the
390
Patricia Martorell et al.