The seeds were obtained from avocados sold in a super market in the city of Maracay, Aragua state, Venezuela.

To obtain the alcoholic extract, 10 g of plant material were weighed and previously crushed, placing them in a 400 mL beaker with 300 mL of 70% v/v methanol/water. It was macerated and left to rest for 24 h, and then it was subjected to microwave radiation for 15 s with a power of 125 MHz. The extracts were filtered, using Whatman filter paper number 4. The extract was left to macerate for 24 h, filtering with Whatman filter paper number 4[9].

For the determination of total phenolics, 50 μL of the alcoholic extract were mixed with 250 μL of the Folin-Ciocalteu 1 N reagent (Analytical grade, Merck). It was left to stand for 8 minutes and then 750 µL of 20% Na2CO3 and 950 µL of distilled water were added. Was incubated for 30 min at room temperature and the absorbance was read on a Genesis 20 UV/VIS spectrophotometer (Thermo Scientific, Waltham, Massachusetts, USA). A calibration curve for Gallic Acid (Sigma-Aldrich, Germany) was prepared with concentrations of 50, 100, 200, 300, 400, 500 and 1000 ppm. The results were expressed in mg of Gallic Acid Equivalents (GAE) / g of PM [10].

Ferric-Reducing Antioxidant Power (FRAP) Assay

The FRAP test was used to determine the reducing capacity of infusions [11]: 100 μL of infusion were mixed with 3 mL of FRAP reagent consisting of a mixture of 300 mM sodium acetate and acetic acid buffer, 10 mM TPTZ solution (2,4,6-tri(2-pyridyl)-s-triazine), and 20 mM FeCl3 solution, in a volume ratio of 10:1:1.

Figure 1

Avocado seeds

The reaction was carried out at room temperature for 4 min, and absorbance was recorded at 593 nm using a Genesis 20 UV/VIS spectrophotometer (Thermo Scientific, Waltham, Massachusetts, USA). FeSO₄ was used as standard, and the results were expressed as µmol Fe ²⁺/g PM.

Prior to the evaluation of the antioxidant activity of the extracts, the concentration of total Phenolics was determined in triplicate, obtaining a concentration of 26.32±2.45 mg GAE/g PM in the aqueous extract and 57, 32 ±2.45 mg GAE/g PM for the methanolic extract, observing a statistical difference (p=0.035). The difference observed is similar to that reported in several studies carried out with Persea americana Mill, which indicate that the extraction of phenolic compounds is influenced by the type of solvent used [14,15]

The antioxidant capacity was determinate as recommended by several studies, which indicate that at least two methods should be used to evaluate the antioxidant activity of an extract [16-18]. The antioxidant effect by the DPPH and FRAP method is shown in Table 1. The antioxidant capacity observed by the DPPH and FRAP methods shows that the extracts possess an important source of antioxidant compounds, a conclusion specially valid when compared to other studies worldwide, which indicate that avocado seeds can be used as a functional food ingredient or as an antioxidant additive [19,20].

Table 1

Antioxidant activity of extracts

For each method, means ± SD. Significantly different (p < 0.05).

Fig 2

Evaluation of antioxidant activity by ABTS method

Among the methods used to determine the capacity of an antioxidant to capture free radicals, the ABTS radical is one of the most applied, as it is considered highly sensitive, practical, fast and very stable [21]. The ABTS assay showed higher inhibition in the methanolic extract with statistical difference from the aqueous extract (Figure 2). These results coincide with what was reported in Bogota Colombia, whose study consisted in determining the phenolic profile of avocado byproducts, among them the seed, indicating that the extracts showed an antioxidant capacity (ABTS method) related to their content of bioactive compounds, considering these byproducts for pharmaceutical and industrial use [15].