María Juliana Restrepo Vega
1, José Hipólito Isaza Martínez
1*, Ana Julia Colmenares Dulcey
11 Research Group on Natural Products and Food (GIPNA), Department of Chemistry, Faculty of Natural and Exact Sciences, Ed. 320, Of 2096., Ciudad Universitaria-Meléndez, Universidad del Valle, Cali, Colombia
Abstract
A novel and selective
method to quantify lipid peroxidation in Pisum
Sativum by measuring one of its secondary products, malondialdehyde, by its
reaction with N-methyl-2-phenylindole was validated in this study. It has been reported that indirect effect of
heavy metals in plant macromolecules via reactive oxygen species (ROS)
production is more toxic and fast than its direct effect. The OH• is produced
by metal-catalized Fenton reaction, as a consequence, metal presence in soils
may potentiate lipid peroxidation. Enzymes such as cyclooxygenase and
lipoxygenase may initiate lipid peroxidation too, therefore, either ROS scavenging
or LOX or COX inhibition is convenient routes to diminish lipid peroxidation
occurrence. Two way anova was performed
to determine if interaction between time and treatment result in a significant
difference between MDA levels in twelve plants. Bonferoni test revealed that
treatment was not consistent with time. The day that resulted in significant
difference of MDA was the eighth day. MDA to perform the reaction was extracted
from groups of twelve 8-day plants by sonication in hydrochlorhydric acid. A
calibration curve using 1,1,3,3, tetramethoxypropane as MDA standard, HCl 1 N
in methanol, acetonitrile-methanol (3:1) as solvent was performed. Its linear
range was 0,0078 μM to 1 μM with a 99,8% correlation. Sensibility of the method
was enhanced by variation of time and temperature. Repeatability reported as
variation coefficient yielded 7,8% for the standard curve and 19,6% for the
samples, being these latter representative of combined biological and chemical
variability. A natural flavonoid rich fraction extracted from Siparuna
gigantotepala and a synthetic pyrimidine-type compound were evaluated as
potential lipid peroxidation inhibitors. Flavonoid rich fraction inhibits the
reaction by scavenging radicals while pyrimidine-type compound inhibits LOX and
COX enzyme-catalized lipid peroxidation. Both potential inhibitors showed lipid
peroxidation inhibition as evidenced by returning MDA concentration in stressed
plants to that in control plants.
Keywords: Lipid peroxidation, Pisum sativum, N-methyl-2-phenylindol method, Malondialdehyde, Flavonoids, Oxidative Stress