Antioxidant Activity in Various Oxidative Environments
In this section, the antiradical capacities of alkyl gallates and gallic
acid were examined using different chemical methods: β-carotene
bleaching, DPPH● scavenging, and Rancimat assays.
These assays were carried to evaluate the impact of -COOH group and the
ester chain length on the antioxidative activity of gallic acid
derivatives in various oxidative environments.
The ability of antioxidative compounds for scavenging DPPH free radicals
is presented in Table 2. The antiradical activities of gallic acid and
alkyl gallates enhanced as the antioxidant concentrations increased from
10 to 50 μM. The percent values of for gallic acid at all concentrations
were significantly higher than the alkyl gallates. Considering the
IC50, the gallic acid concentration required to scavenge
50% of DPPH● was considerably lower than its alkyl
ester derivatives. These results were in accordance with
Asnaashari et al. (2019) and
Mansouri et al. (2020) that reported the
anti-DPPH• activity of gallic acid was significantly more potent
compared to methyl gallates.
The O-H bond disassociation enthalpy (O-H BDE), intramolecular hydrogen
bonds (IHB), and ionization potential are the three critical parameters
that affect the antiradical activity of antioxidant agents. Given that
the O-H BDE, IHB, and ionization potential of alkyl gallates are
similar, thus the steric impact can be a plausible explanation for the
difference in antiradical activity of alkyl gallates. The ester
structure affects the polarity and solubility of antioxidant compounds
as well as their molecular mobility and availability to oxidative
components (Lu et al., 2006). Our results
showed that the radical scavenging potency of gallic acid as affected by
an extra carboxyl group than alkyl gallates. In fact, the dissociation
possibility of the carboxyl group in polar media is the main reason for
increased the electron-donating and H-atom transfer activities of gallic
acid than the alkyl gallates. In contrast to the electron-donating
impact of carboxylate anion, -COOH is considered an electron-withdrawing
group that increases the O-H BDE of the phenolic ring
(Correia et al., 2004). As can be seen in
Table 2, the polarity (log P) of gallic acid was significantly higher
than the alkyl gallates, which is leading to more hydrophilic
interactions between the antioxidant and the polar molecules of methanol
as a reaction solvent.
In the β-carotene bleaching assay, gallic acid and alkyl gallates
prevented the extent of β-carotene bleaching by neutralizing the
linoleate-free radical and other free radicals formed in the emulsion.
As shown in Table 2, a concentration-dependent antioxidant potential was
observed for all antioxidant components studied. A comparison of
antioxidant capacity in the emulsion system showed that gallic acid was
the lowest reactive antioxidant than alkyl gallates. So that, the
IC50 value of antioxidant agents was as follows: dodecyl
gallate < octyl gallate < propyl gallate
~ methyl gallate < gallic acid <
stearyl gallate. This means that the alkyl gallates concentration
required for scavenging %50 of linoleate-free radicals was
significantly lower than the gallic acid. In general, due to the
appropriate length of the alkyl chain, dodecyl gallate had a more
effective interfacial performance at the water-oil interfaces. The
nonlinear behavior or cut-off effect was observed for inhibitory
activities of homologous series of antioxidative compounds in the W/O
emulsion. The interfacial activities of gallic acid derivatives enhanced
as the lengths of the alkyl chain were reached until a critical chain
length (12 carbon atoms); after that, further chain length extension
causes a decrease in antioxidant activities.
Medina et al. (2009),
Laguerre et al. (2010) and
Meireles et al. (2019) reported similar
findings showing that the lipophilic esters with short medium-chain
compared to long-chain esters improved the activity of antioxidative
compounds in the multiphase system.
Laguerre et al. (2009) also reported in
the emulsified medium the antiradical potency enhances as the alkyl
chain is lengthened, with a threshold for the dodecyl chain, after which
further chain extension causes a drastic decrease in antiradical
activity. The increased antiradical activities of the alkyl gallates in
β-carotene-linoleic acid emulsion compared to gallic acid can be
attributed to increased hydrophobicity (log P), surface-active
characteristic, and the precise placement of antioxidants in the
water-oil interface.
In the Rancimat test, the oxidative stability index (OSI) of stripped
soybean oil (0.32 h) was significantly promoted with the antioxidants
added. The OSI values of bulk soybean oil treatments with antioxidant
agents were as follow: propyl gallate > octyl gallate
~ dodecyl gallate > stearyl gallate
~ methyl gallate > gallic acid. A higher
induction period was obtained for soybean oil containing propyl gallate
compared to other antioxidants means that propyl gallate was the most
effective antioxidant to increase oil oxidative stability at high
temperature (120 °C). This result was in accordance with
Phonsatta et al. (2017) that reported the
OSI of bulk oil containing alkyl gallates, especially propyl gallate,
was significantly more potent compared to gallic acid. Concerning the
carboxyl group’s electron-withdrawing character, which is not markedly
dissociated in the hydrophobic media, gallic acid had the lowest
inhibitory activity than alkyl gallates in the lipid system. Studies
have shown that the esterification of gallic acid with fatty alcohols
can be increasing its hydrophobicity as well as surface-active
properties (Chaiyasit et al., 2005).
According to log P results (Table 2) the hydrophobicity of alkyl
gallates was higher than the gallic acid. Therefore, the increased
antioxidant activities of the alkyl gallates, especially propyl gallate,
in bulk phase oil compared to gallic acid can be attributed to improved
amphiphilic properties by increasing the alkyl chain length and the
precise placement of antioxidants in the actual site of oxidation. Our
findings agreed with those reported by
Kikuzaki et al. (2002). The results
clearly showed the nonlinear antioxidative activities for alkyl gallates
due to enhance in the length of the alkyl chain. The size of
antioxidants affects their activity by changing their mobility in the
bulk phase oils, leading to a cut-off effect. Lipophilic antioxidative
components by long alkyl chains have lower mobility than those with a
smaller size because of steric hindrance, so decreased diffusibility
toward the reactive centers such as free radicals and oxidizable
substrates. Furthermore, enhance in the length of the alkyl chain
increase the possibility of hydrophobic interaction among antioxidative
components and the environment such as the membrane or emulsifiers,
which cause a decrease in mobility and diffusion of the antioxidative
components to the oxidation site of bulk oil
(Laguerre et al., 2015).