3.6.2. Antioxidant Activity

The antioxidant activity of the EOs and NEs, obtained from aromatic plants, is conferred by their phenolic compounds. The DPPH free radical method is an antioxidant assay based on an electron-transfer that produces a violet solution in methanol. This free radical, stable at room temperature, is reduced in the presence of an antioxidant molecule (active compound), giving rise to a colorless solution [77]. The use of the DPPH assay provides an easy and rapid way to evaluate antioxidants using spectrophotometry. In the conservation of foods, substances with character antioxidants are of grea<sup>t</sup> interest because the main cause of food deterioration results from enzymatic reactions that trigger the oxidation of lipids and carbohydrates [78,79].

The percent inhibition and the equivalent concentration in micrograms of trolox per gram of sample of the neat OEO, RE, GTE, and of the electrospun fibers and films of the PHBV containing the active substances are shown in Table 7. One can observe that all the EOs and NEs showed DPPH radical scavenging activity. OEO presented the highest percentage of inhibition (91.96%) attributed to its main active compounds (e.g., carvacrol, thymol, p-cymene, *γ*-terpinene) [64]. Similarly, Chun et al. [80] reported an inhibition percentage of DPPH of 82% for oregano extracts. RE presented a percentage of inhibition of 75.24%, which was in agreemen<sup>t</sup> with, for instance, Bajalan et al. [81] who reported an inhibition percentage of DPPH of 73.69%. GTE showed an inhibition of 71.77%, conferred by the relative amount of catechins and GA [82]. Afroz Bakht et al. [83] studied the antioxidant activity from DPPH of five commercial teas, finding percentages of inhibition in the 24–71% range. In addition, Lu and Chen [84] reported percentages of inhibition between 33% and 62%. In this sense, it is important to highlight that antioxidant activity is dependent on the quantity of secondary metabolites that the plant manages to synthesize in its development stage, which is influenced by the variety of the plant, the environmental conditions [85–87], and the extraction method used [83].

One can observe that the antioxidant activity decreased in the electrospun PHBV fibers containing OEO, RE, and GTE. After the electrospinning process, the fibers with OEO showed a percentage of inhibition of 43.14%, whereas the fibers containing RE and GTE showed values of 25.82% and 22.12%, respectively. In all cases, there was a decrease in the antioxidant activity of between 20% and 30% compared to the pure OEO, RE, and GTE. The lower antioxidant inhibition of the active compounds inside the fibers could be related to polarity differences between the solvent and the polymer, the stirring process applied to the active solution prior to electrospinning, and the loss of volatiles compounds during the electrospinning process [44].

**Table 7.** Inhibition percentage (%) of 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) and concentration (eq. trolox/g sample) of DPPH for oregano essential oil (OEO), rosemary extract (RE), and green tea tree extract (GTE) and the electrospun fibers of poly(3-hydroxybutyrate-*co*-3-hydroxyvalerate) (PHBV) containing OEO, RE, and GTE.


Table 8 shows the percentages of inhibition and the equivalent concentration in micrograms of trolox per gram of sample of the electrospun PHBV films containing OEO, RE, and GTE, evaluated in the so-called open and closed systems on days 1, 8, and 15. Films containing OEO exhibited the highest inhibition of DPPH at day 1 (24.54%), followed by films with the RE (15.59%) and films with the GTE (11.14%). Over time, all the films decreased their antioxidant activity, obtaining no significant differences in the results between each storage system. Thus, after 15 days, an inhibition percentage in the range of 8.83–10.55% was obtained for the films containing OEO. At this time, the inhibition percentage ranges for the films containing RE and GTE were 6.91–7.31% and 5.45–6.68%, respectively. As can be observed, the antioxidant activity decreased when the EOs and NEs were included in the PHBV fibers and, more intensively, when the films were formed. Likewise, during the days of storage, a continuous release of the characteristic volatile compounds of each EOs and NEs was produced, which was evidenced by the low percentage of DPPH inhibition at day 15 for all the samples. Previous reports have indicated that the degree of antioxidant power of biodegradable films is generally proportional to the amount of added antioxidant additives, while the thermal process for obtaining the films also affects the bioactivity, since most of the bioactive compounds are sensitive to temperatures above 80 ◦C [47,88]. Regardless of this, all the films presented antioxidant performance and they can, therefore, be applied in antioxidant active packaging systems to extend the shelf life of packaged food products, thus minimizing the development of off-flavors, color and flavor changes, and nutritional losses [89,90].


**Table 8.** Inhibition percentage (%) of 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) and concentration (eq. trolox/g sample) of DPPH for the electrospun poly(3-hydroxybutyrate-*co*-3-hydroxyvalerate) (PHBV) films containing oregano essential oil (OEO), rosemary extract (RE), and green tea tree extract (GTE).
