## 2 Materials and Methods
#### 3.2.3 Plasma Oxidative Status
On the other hand, the total antioxidant capacity (measured by the FRAP assay) differed between GP and basal diet fed animals [14]. In laying hens, the dietary supplementation with Roselle (_Hibiscus sabbariffia_ L) calyx (crude extracts powder at the 2.4% concentration administered at the dose of 1% or 2% of the diet) did not change the TBARS values in plasma in the first 4 weeks of an 8 week feeding trial among the different inclusion levels in the diet or in comparison with the VitE supplementation (250 mg/kg). Nevertheless, an overall tendency in decreasing this parameter was determined by Roselle calyx in yolk [31]. In broilers fed a linseed oil rich diet (high PUFA), the addition of SCW (0.3% of the diet) reduced the lymphocyte DNA damage (by 42.0%) [57]. Lower relative transcripts levels (by 594.3% on average) of HSP70 gene (expression of which usually increases in heat stress) in the heart tissue of GSE fed broilers were found when animals were reared under heat stress. A reduction (by 199.9%) in the expression of HSP70 gene in the liver of GSE fed broilers exposed to heat stress was also found [40]. The effects of PFA on selected immunological parameters in comparison with synthetic vitamins were also investigated in poultry. More specifically, different grape polyphenols (GPP) to VitE ratios (C, 0:100; LGPP, 25:75; MGPP, 50:50; HGPP, 75:25) in broiler diets were tested. Broilers supplemented with the highest GPP:VitE ratio (HGPP) compared with those receiving only VitE (100 mg/kg) improved their antibody titers (by a mean value of 59.5%) against Newcastle disease and infectious bursal disease (IBD) [59]. However, antibody titers against IBD were also higher in HGPP compared with LGPP and MGPP groups (by an average proportion of 389.0%) [59]. In contrast to these results, dietary supplementation with roscamy powder (RPO; 0%, 0.5%, and 1% of the diet) or VitE for 42 days did not affect broilers' antibody titers against viruses and sheep red blood cells (SRBC) [60]. In most of the cases, the dietary addition of PFS was able to reduce the MDA content of plasma and tissues. Based on the type of PFA, this decrease was accompanied by changes in antioxidant enzymes activities and total antioxidant capacity. Grape pomace and grape polyphenols positively affected poultry immunity due to an increase in antibody titers. Effects on Macro- and Microscopic Anatomical Features and on Intestinal Microbiome Profile (Table S3 in Supplementary Materials)  
Several studies suggested an immunostimulant activity exerted by polyphenols on different signal transduction pathways, which are responsible for a wide range of cell functions related, for instance, to immune organs growth and proliferation [61]. The broiler spleen and bursa of Fabricius were not affected when their diet was supplemented with different levels of GP (5%, 7.5%, and 10% of the diet) or with VitE (200 mg/kg) [34]. Similarly, lower GP supplementation levels (1.5% to 6% of the diet) did not affect relative spleen weight in broilers; only those that were supplemented with low GP levels (1.5% of the diet) compared with VitE showed improvement in this morphological parameter (by 20.0%) [14]. Similar findings concerning weights of thymus, spleen, and bursa of Fabricius in RPO (0.5% and 1% of the diet) and VitE (100 and 200 mg/kg) fed broilers were found [60]. Furthermore, some studies also tested PFA effects on small intestine and gut gross morphometry, which have relevance for digestion capability. Broilers supplemented with GP (1.5% of the diet) compared with VitE improved relative length of duodenum and ceca (by about 9.0%) [14]. On the other hand, RPO addition (0.5-1% of the diet) in broilers' diet negatively affected jejunum length and weight, colon length and weight, and right eccum weight, but no significant differences were reported concerning these anatomical traits compared with those consuming two different levels of VitE (100 and 200 mg/kg) [28].