9 Analytical Protocols in Antioxidant Capacity Measurement
205
competitive trials puts them in advantage as compared to other methods as it confers
close representation to biological mechanisms (Apak et al. 2016).
According to Rodrigues et al. (2012), to ensure an unequivocal measurement,
it is essential to guarantee the solubilization of the antioxidant compound in the
medium since, among the existing protocols, there is variability in terms of polarity.
Consequently, the structural characteristics of the compound to be analyzed are taken
into account since some are lipophilic, such as carotenoids and chlorophylls, and
others are water-soluble, such as phycobiliproteins and phenolic compounds.
All of these details culminate in several analytical factors before choosing the
protocol to be carried out on microalgae extracts. In this sense, this chapter provides
a comprehensive description of the main protocols used to investigate the in vitro
antioxidant capacity of microalgae compounds, including the characteristics bioactive compounds to be analyzed, reaction medium, principles, mechanisms involved,
limitations and advantages of the protocols used, as well as a database with existing
measurements.
9.2 Microalgae Bioactive Compounds: Structure
and Antioxidant Capacity
Over the years, microalgae have become the focus of scientific exploration as
they consist of different metabolites with significant bioactive potentials, such as
carotenoids, chlorophylls, phycobiliproteins, and phenolic compounds (Raposo et al.
2013; Fernandes et al. 2017; Jacob-Lopes et al. 2019; Nascimento et al. 2019).
In terms of carotenoids, several compounds have been structurally defined and
reported in the literature. Britton et al. (2004) report over 750 structures, whereas
in the last compilation by Yabuzaki (2017), approximately 1181 were reported.
In microalgae, a range of these pigments has been isolated, the most common
being β-carotene, α-carotene, lutein, zeaxanthin, astaxanthin, violaxanthin, neoxanthin, crocoxanthin, fucoxanthin, canthaxanthin, echinenone, and myxoxanthophyll
(Rodrigues et al. 2014; Jacob-Lopes et al. 2019), and their structures, as well as their
preferred antioxidant mechanisms, are illustrated in Fig. 9.1.
Carotenoids are fat-soluble natural pigments that may impart yellow, orange, or
red color due to their basic chemical structures consisting of a linear and symmetrical
skeleton with a series of conjugated double bonds (CDBs) (Rodriguez-Amaya 2001).
This series of CDBs generates a resonance system of π electrons moving along the
entire polyenic chain; so these compounds are very reactive and absorb light in the
visible region of the spectrum (around 450 nm) (Mercadante et al. 2008).
Structurally, they are grouped into carotenes and xanthophylls: carotenes are
formed only by carbon and hydrogen (e.g., β-carotene and α-carotene), while xanthophylls contain oxygen-containing substituents (e.g., lutein, zeaxanthin, astaxanthin,
violaxanthin, neoxanthin) (Fernandes et al. 2018).
205
competitive trials puts them in advantage as compared to other methods as it confers
close representation to biological mechanisms (Apak et al. 2016).
According to Rodrigues et al. (2012), to ensure an unequivocal measurement,
it is essential to guarantee the solubilization of the antioxidant compound in the
medium since, among the existing protocols, there is variability in terms of polarity.
Consequently, the structural characteristics of the compound to be analyzed are taken
into account since some are lipophilic, such as carotenoids and chlorophylls, and
others are water-soluble, such as phycobiliproteins and phenolic compounds.
All of these details culminate in several analytical factors before choosing the
protocol to be carried out on microalgae extracts. In this sense, this chapter provides
a comprehensive description of the main protocols used to investigate the in vitro
antioxidant capacity of microalgae compounds, including the characteristics bioactive compounds to be analyzed, reaction medium, principles, mechanisms involved,
limitations and advantages of the protocols used, as well as a database with existing
measurements.
9.2 Microalgae Bioactive Compounds: Structure
and Antioxidant Capacity
Over the years, microalgae have become the focus of scientific exploration as
they consist of different metabolites with significant bioactive potentials, such as
carotenoids, chlorophylls, phycobiliproteins, and phenolic compounds (Raposo et al.
2013; Fernandes et al. 2017; Jacob-Lopes et al. 2019; Nascimento et al. 2019).
In terms of carotenoids, several compounds have been structurally defined and
reported in the literature. Britton et al. (2004) report over 750 structures, whereas
in the last compilation by Yabuzaki (2017), approximately 1181 were reported.
In microalgae, a range of these pigments has been isolated, the most common
being β-carotene, α-carotene, lutein, zeaxanthin, astaxanthin, violaxanthin, neoxanthin, crocoxanthin, fucoxanthin, canthaxanthin, echinenone, and myxoxanthophyll
(Rodrigues et al. 2014; Jacob-Lopes et al. 2019), and their structures, as well as their
preferred antioxidant mechanisms, are illustrated in Fig. 9.1.
Carotenoids are fat-soluble natural pigments that may impart yellow, orange, or
red color due to their basic chemical structures consisting of a linear and symmetrical
skeleton with a series of conjugated double bonds (CDBs) (Rodriguez-Amaya 2001).
This series of CDBs generates a resonance system of π electrons moving along the
entire polyenic chain; so these compounds are very reactive and absorb light in the
visible region of the spectrum (around 450 nm) (Mercadante et al. 2008).
Structurally, they are grouped into carotenes and xanthophylls: carotenes are
formed only by carbon and hydrogen (e.g., β-carotene and α-carotene), while xanthophylls contain oxygen-containing substituents (e.g., lutein, zeaxanthin, astaxanthin,
violaxanthin, neoxanthin) (Fernandes et al. 2018).