To study the effects of an extract of plant flavonoids [Bioflavex (FL)] in cattle fed highconcentrate diets, 2 experiments were designed. In the first experiment, the effects of Bioflavex on the development of rumen acidosis was evaluated in 8 Holstein-Friesian crossbreed heifers (451 kg; SEM 14.3 kg of BW) using a crossover design. Each experimental period lasted 22 d; from d 1 to 20, the animals were fed rye grass, on d 21 the animals were fasted, and on d 22, rumen acidosis was induced by applying 5 kg of wheat without [Control: (CTR) heifers who did not receive Bioflavex] or with flavonoids [heifers who received FL; 300 mg/kg DM] through a rumen cannula. Rumen pH was recorded continuously (from d 19 to d 22). On d 22, average rumen pH was significantly (P < 0.01) higher in the FL animals (6.29; SEM = 0.031) than it was in the CTR heifers (5.98; SEM = 0.029). After the wheat application, the rumen VFA concentration increased (P < 0.01), the proportion of acetic acid decreased (P < 0.01), and lactate concentration (mmol/L) increased, but the increase was not as great (P = 0.09) in the FL as it was in the CTR heifers (0.41 to 1.35 mmol/L; SEM = 0.24). On d 22, Streptococcus bovis and Selenomonas ruminantium titers increased after the wheat application, but Megasphaera elsdenii titers increased (P < 0.05) only in the FL heifers. In the second experiment, the effect of Bioflavex on the performance and rumen fermentation in finishing heifers was evaluated. Forty-eight Fleckvieh heifers (initial BW = 317 kg; SEM = 5.34) were used in a completely randomized design. Heifers were assigned to 1 of 4 blocks based on their BW and, within each block, assigned to 1 of 2 pens (6 heifers/pen). In addition, 16 heifers (2/pen) were rumen cannulated. Individual BW and group consumption of concentrate and straw were recorded weekly until the animals reached the target slaughter weight. Supplementation with FL did not affect ADG, feed consumption, or feed conversion ratio. Rumen pH and molar proportions of propionate were greater (P < 0.01) and acetate proportion was less in the FL (P < 0.01) than they were in the CTR heifers. Flavonoid supplementation might be effective in improving rumen fermentation and reducing the incidence of rumen acidosis. This effect of flavonoids may be partially explained by increasing the numbers of lactateconsuming microorganisms (e.g., M. elsdenii) in the rumen.

Està subjecte a una llicència de Reconeixement-NoComercial-SenseObraDerivada 4.0 de Creative Commons The effect of Bioflavex ® and its pure flavonoid components on in vitro fermentation parameters and methane production in rumen fluid from steers given high concentrate diets

One hundred fourty-four bulls (164.8 ± 5.91 kg BW and 135 ± 7.2 d of age) were randomly allocated to one of 8 pens and assigned to control (C) or citrus flavonoid (BF) treatments (Citrus aurantium, 0.4 kg per ton of concentrate of Bioflavex CA, > 20% naringin; BF). Each pen had one drinker, one separate five-space straw feeder, and one separate three-space feeder where mash concentrate containing mostly corn, barley, DDG and wheat was offered. Concentrate intake was recorded daily, whilst BW and animal behavior were recorded fortnightly. Animals were slaughtered after 168 d of study (12 periods of 14 d), and HCW and carcass quality were recorded, and rumen papillae samples were collected. Final BW (437.9 ± 1.85 kg), HCW (238.7 ± 2.02 kg), and concentrate intake (7.1 ± 0.13 kg/d) were not affected by treatment. Concentrate feed conversion ratio (kg of concentrate/ kg of BW) tended (P < 0.10) to be lesser in BF than in C bulls (5.11 vs. 5.36 ± 0.108 kg/kg). Percentage of animals eating concentrate during visual scan was greater (P < 0.01) in BF compared with C bulls (10.02 % vs. 7.97 % ± 0.512). Oral non-nutritive behaviors, agonistic interactions (fighting, butting, and chasing) and sexual behaviors (flehmen, attempted and complete mounts) were greater (P < 0.01) in C than in BF bulls. In the rumen epithelium, gene expression of bitter taste receptor 7, bitter taste receptor 16, bitter taste receptor 38 and bitter taste receptor 39 was greater (P < 0.05) in C compared with BF bulls, as well as was gene expression of free fatty acid receptor 2, pancreatic polypeptide receptor 1, cholecystokinin receptor 4, cytokine IL-25, Toll-like receptor-4 and β-defensin1. In conclusion, supplementation with flavonoids extracted from Citrus aurantium in bulls fed high-concentrate diets tends to improve efficiency, and reduces oral non-nutritive behaviors, agonistic interactions and sexual behavior. Moreover, flavonoid supplementation modifies the expression of genes in the rumen epithelium that could be related with eating and animal behavior regulation.

Feed Science and Technology © Elsevier after peer review. To access the final edited and published work see https://doi.org/10.1016/j.anifeedsci.2018.08.010 FLAVONOIDS AND RUMEN HEALTH AND PERFORMANCE 1 2 Effects of flavonoids extracted from Citrus aurantium on performance, eating and 3 animal behavior, rumen health, and carcass quality in Holstein bulls fed high-4 concentrate diets. 5 6 MABSTRACT 22The effects of flavonoids extracted from Citrus aurantium (Bioflavex® CA) on eating 23 pattern, performance, carcass quality, and rumen wall health of Holstein bulls fed on a 24 single feeder were studied. One hundred ninety-eight bulls (195.3 ± 19.6 kg of body 25 weight and 149 ± 6.8 d of age) were used in a complete block randomized design. 26 Groups of animals with the same mean and coefficient of variation of body weight 27(replicates) were randomly allocated in 1 of 6 pens (20 animals per pen), and each 28 pen was assigned to one of 6 pens and assigned to a Control (C) diet or to a diet 29 supplemented with flavonoids (Bioflavex® CA, Interquim S.L., Spain) (BF, 0.4 kg 30 per ton of concentrate of Bioflavex® CA) in two consecutive fattening cycles. 31Concentrate intake was recorded daily, and BW fortnightly. Animal behavior was 32 monitored by visual scan procedure every fourteen days. Animals were slaughtered after 33 168 d of study, hot carcass weight and carcass quality were recorded, and internal 34 rumen wall was examined. Concentrate intake was higher (P < 0.05) in C than in BF 35 bulls; however, ADG and concentrate efficiency were not affected by treatments. The 36 final BW tended (P = 0.06) to be higher in C than in BF bulls, but this difference 37 disappeared for carcass weight. In the finishing phase, the proportion of meal size 38 values above 750 g was higher (P < 0.05) in C compared with BF bulls. Throughout the 39 study exhibited more displacements and fighting than C bulls, whilst C bulls performed 40 more (P < 0.05) oral behaviors. During the finishing phase, sexual behaviors such as 41 flehmen and complete mounts were higher (P < 0.01 and P < 0.05, respectively) in C 42 bulls as well, and C bulls tended (P = 0.10) to perform more attempted mounts 43 compared with BF bulls. In the slaughterhouse, color of rumen wall tended (P = 0.06) 44 to be lighter for BF compared with C bulls, and presence of baldness areas in the rumen 45 was lesser (P = 0.01) in BF animals. In conclusion, when bulls were supplemented with 46 3 Bioflavex® CA, feed intake was reduced. Flavonoids supplementation increased time 47 eating straw, reduced agonistic behaviors throughout the study and sexual interactions 48 during the finishing phase, potentially improving animal welfare. Rumen wall 49 parameters analyzed were indicative of a better rumen health in BF than in C bulls, 50 which maybe due to the reduction of large meal sizes. 51

Two experiments were performed to study the effects of the citrus flavonoid extract Bioflavex (BF; Interquim SA, FerrerHealthTech, Sant Cugat, Barcelona, Spain) or its components on the rumen fermentation of a high-concentrate diet. In an in vivo experiment, eight Friesian steers (398 ± 12.2 kg bodyweight) fitted with a rumen cannula were given a basal concentrate (CTR) or a CTR supplemented with BF (450 mg/kg dry matter, DM) in a 2 × 4 crossover design. No differences were observed in performance parameters of BF and CTR steers. Diet BF increased pH values and the molar proportion of propionate and reduced lactate concentration as a result of an increase in the relative abundance of lactate-consuming microorganism Selenomomas ruminantium (P < 0.01) and Megaesphaera elsdenii (P = 0.06). In an in vitro experiment, the effect of BF and its pure flavonoid components added to the incubation medium was studied separately. Bioflavex and its main components naringine, neohesperidine (NH) and poncirine (PC) were added to the incubation medium at 500 µg/g DM, with the unsupplemented substrate also included as a control (CTR). After 12 h of incubation, flavonoid mixture and NH and PC reduced (P < 0.01) the volume of gas produced and the molar proportion of acetate (P < 0.01), and increased that of propionate (P < 0.01). PC reduced the relative quantification of Streptococcus bovis, whereas NH and BF increased the relative quantification of M. elsdenii in relation to CTR (P < 0.01). Bioflavex supplementation in steers in feedlot was effective in preventing a collapse in pH and it enhanced rumen fermentation efficiency through modifying the activity of lactate-consuming bacteria and a greater molar proportion of propionate and a reduction of that of acetate, suggesting its positive role in modulating the activity of rumen microbiota.