The significant increase in LDH activity at maximumlevels of MPs concentrations can be explained by the occurrence of cellulardamage in organs and tissues due to anoxic situations during stressfulconditions (Abhijith et al., 2016), as previously identified in E. andrei with histopathological damage(Rodríguez-Seijo et al., 2017). These resultswere also in agreement with other studies that assessed the depletion of energyreserves and LDH levels among marine worms (Wright et al., 2013), mussels(Paul-Pont et al., 2017) and terrestrial organism, such as mice (Deng et al.,2017) when exposed to different polymers.
Whenexposed to several contaminants, earthworms are highly susceptible to oxidativestress stemming from the peroxidation of lipid tails in the membrane structure,due to their high content of polyunsaturated fatty acids (Markad et al., 2012;Mwaanga et al., 2014).
Organisms exposed tohigher MPs concentrations (500 and 1000 mg kg -1) had the highestmean values of TBARS per mg protein, with statistically significant differenceswhen compared with control earthworms and those exposed to other concentrations(ANOVA: F = 70.73; d.f. 5, 48; p = 0.000) (Figure 2).
At the same time,at the lowest level of MPs concentrations, the lowest TBARS content was observed.These results indicate that thelowest MPs levels activated theearthworms’ defensive systems to prevent excess ROS production, asindicated by Paul-Pont et al. (2016) for marine mussels exposed to polystyrenemicroplastics. On the other hand, the increaseof TBARS levels in E. fetida may haveresulted from the incapacity of the defence system to counteract oxidativestress in earthworms exposed to the highest concentrations of MPs (r = 0.668; p < 0.01) (Table 1) (Lin et al., 2012;Zhang et al.
, 2013; Paul-Pont et al., 2016; Babic et al., 2016; Chao et al.
,2016). Oliveira et al. (2013) also detected no damage to lipidperoxidation when P. microps were exposed to polyethylenemicrospheres, although at lower MPs concentrations than those herein tested (intheir case, 200 ug L-1 and 1-5 µm). In thissense, the biomarker data collected in the present study are complementary tothe histopathological and molecular analyses conducted in the previous study,as the incrementing concentrations of lipids, proteins and polysaccharides canoccur after lipid peroxidation (García et al.
, 2005) or through coelomocytesinvolved in eliminating foreign material (Kauschke et al., 2007).According to Deng et al. (2017), MPs exposuremay cause inflammatory responses, which were also observed in our previous research,and thus lead to lipid disturbance, as reported for mussels (von Moos et al., 2012),or fishes (Oliveira et al.
, 2013). The absence of mortality orsignificant changes in weight suggested that antioxidant enzymes may play a key role in the health status of the organismand can prevent damages at several levels. However, the antioxidant responsewas insufficient to hold the oxidative stress from exposure to MPs (Markad etal.
, 2015), as was previously observed through histopathological and molecularchanges in E. andrei (Rodríguez-Seijoet al., 2017).