Amalia Pérez-Jiménez

Digestive enzymatic profile of Dentex dentex and response to different dietary formulations

Digestive physiology of on-growing common dentex (Dentex dentex), including protease, amylase and lipase activity in stomach, pyloric caeca, anterior and posterior intestine, was evaluated. The influence of dietary macronutrient balance on these digestive processes was also assessed. Four experimental diets with different protein:lipid:carbohydrate ratios (43/16/28; 43/24/4; 38/19/28 and 38/24/13) were formulated. The highest activity for acid proteases was located in the stomach at pH 1.5. Alkaline proteolytic activities showed the highest values in the pyloric caeca and posterior intestine at pH 8.5-9.0. Dentex showed substantial amylase activity in the pyloric caeca and posterior intestine. Lipase activity was higher in the pyloric caeca, anterior and posterior intestine and was not detected in the stomach. Feed composition influenced alkaline protease activity in the anterior and posterior intestine and was higher for the diet with less protein and more carbohydrates. Enhanced amylase activity was observed in the pyloric caeca and posterior intestine in those groups fed on higher carbohydrate and lower lipid level diets. High dietary carbohydrate levels produced the highest lipase activity but this only occurred in the anterior intestine. We can conclude that the digestive tract of dentex adapts well to protein digestion and possesses a high potential for digesting the other dietary macronutrients, too. Dietary carbohydrate content seems to induce changes in protease, amylase and lipase activity.

Effect of fishmeal replacement by soy protein concentrate with taurine supplementation on hepatic intermediary metabolism and antioxidant status of totoaba juveniles (Totoaba macdonaldi).

The effect of dietary incorporation of soy protein concentrate (SPC) and the concomitant supplementation with taurine on hepatic intermediary metabolism and antioxidant status of totoaba (Totoaba macdonaldi) juveniles was assessed. Four isoproteic and isolipidic diets were formulated containing either 30 or 60% of SPC (diets SP30 and SP60), supplemented or not with 1% of taurine (diets SP30T and SP60T). A fish meal (FM) based diet, without SPC and taurine supplementation, was used as a control. Triplicate groups of 32 totoaba juveniles (average body mass=7.5g) were fed these diets over 45days. Results revealed that dietary FM replacement by SPC depressed the overall intermediary metabolism. Activity of key enzymes of amino acid catabolism and gluconeogenesis was significantly reduced and a trend to reduce glycolysis and glucose-6-phosphate dehydrogenase activity was observed. The incorporation of the highest level of SPC also significantly increased hepatic lipid peroxidation and the activity of superoxide dismutase. Concomitant taurine supplementation restored the activity of amino acid catabolic and gluconeogenic enzymes and hexokinase to levels similar of those of the control diet. Taurine supplementation also led to a significant increase of glucose-6-phosphate dehydrogenase and catalase activity, as well as to a significant reduction of liver lipid peroxidation. These results suggest that taurine may play an important metabolic modulation action on totoaba fed SPC based diets, contributing to the enhancement of the overall metabolism and to the reduction of liver oxidative damage.

The effect of hypoxia on intermediary metabolism and oxidative status in gilthead sea bream (Sparus aurata) fed on diets supplemented with methionine and white tea.

The present study evaluates the influence of previous nutritional status, fish fed on diets supplemented with tea and methionine, on acute hypoxia tolerance and subsequent recovery of Sparus aurata juveniles. Four isonitrogenous (45% of protein) and isolipidic (18% lipid) diets were formulated to contain 0.3% methionine, 2.9% white tea dry leaves or 2.9% of white tea dry leaves+0.3% methionine. An unsupplemented diet was used as control. Hepatic key enzymes of intermediary metabolism and antioxidant status, superoxide dismutase isoenzyme profile, glutathione (total, reduced and oxidized) and oxidative damage markers were determined under normoxia, hypoxia challenge and during normoxia recovery. Dietary white tea inclusion decreased plasma glucose levels under normoxia and seemed to induce an increase in anaerobic pathways as showed by enhanced liver lactate dehydrogenase activity. Hypoxia challenge reversed some of the responses induced by diet tea supplementation. Hypoxia decreased plasma glucose levels, increased glucose 6-P-dehydrogeanse activity, decreased superoxide dismutase activity (especially Mn-SOD and CuZn-SOD isoforms) and increased glutathione peroxidase activity in all dietary treatments. Catalase activity during hypoxia varied with dietary treatments and glutathione reductase was not modified. Antioxidant defenses were insufficient to avoid an oxidative stress condition under hypoxia, independently of dietary treatment. In general, pre-challenge values were recovered for almost all parameters within 6 h recovery time.

Effects of temperature and dietary protein level on hepatic oxidative status of Senegalese sole juveniles (Solea senegalensis)

Effects of 55 and 45% dietary protein levels (55P and 45P diets, respectively) and temperature (12 and 18 °C) on hepatic activity of superoxide dismutase (SOD), catalase, glutathione peroxidase, glutathione reductase (GR), glucose-6-phosphate dehydrogenase and lipid peroxidation (LPO) levels of Solea senegalensis juveniles were studied. Further, effects of acute thermal shocks provoked by a drop (18 °C to 12 °C) or a rise (12 °C to 18 °C) of water temperature on sole oxidative state was also evaluated. Dietary protein reduction increased LPO levels though no major alterations were found on antioxidant enzyme activities between dietary treatments. At 12 °C GR activity was higher and SOD activity was lower than 18 °C but LPO levels were not affected. In both thermal shock cases, LPO levels increased in 55P group, probably due to insufficient antioxidant enzyme activation. In contrast, fish of 45P group under acute exposition to warmer and colder temperature exhibited no substantial changes and a significant decrease on LPO levels, respectively, along with no major changes in antioxidant enzymes. Overall, results suggest that independently of rearing temperatures 45P group was more susceptible to oxidative stress than 55P group. Thermal shock either due to rise or drop of temperature seemed to induce oxidative stress in 55P group.

Effect of temperature and short chain fructooligosaccharides supplementation on the hepatic oxidative status and immune response of turbot (Scophthalmus maximus).

In this study the effect of diet supplementation with different levels of short-chain fructooligosaccharides (scFOS) on the hepatic oxidative status, hematology and innate immune parameters was evaluated in turbot reared at 15 °C and 20 °C. Four practical diets containing half of the protein provided by plant ingredients and the other half by fish meal were supplemented with scFOS at 0%, 0.5%, 1.0% and 2.0% and fed to turbot juveniles for 9 weeks. Independently of the rearing temperature, diet with 1% scFOS increased the haematocrit (Ht) while 2% scFOS augmented the mean corpuscular haemoglobin concentration (MCHC). Mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), white blood cells (%) and lysozyme were higher in fish reared at 15 °C, whereas red blood cells and neutrophil numbers increased in fish reared at 20 °C. Catalase (CAT) and glutathione peroxidase (GPX) activities were affected by rearing temperature being lower in fish reared at 20 °C. Compared to the control diet, at 15 °C, turbot fed 0.5 or 1% scFOS presented lower activities of CAT and glutathione reductase (GR). At 20 °C turbot fed the 2% scFOS diet presented lower activities of CAT and GPX. Lipid peroxidation (LPO) and glucose 6-phosphate dehydrogenase (G6PDH) activity were not affected by temperature nor dietary prebiotic incorporation. Results of this study suggest scFOS has no effect on innate immunology or hematology. High temperature (20 °C) does not induce turbot oxidative stress, but the recommended dietary scFOS incorporation level for counteracting oxidative stress may differ with other rearing temperature.

Dietary carbohydrate and lipid sources affect differently the oxidative status of European sea bass (Dicentrarchus labrax) juveniles.

This study aimed to evaluate the effects of dietary lipid source and carbohydrate content on the oxidative status of European sea bass (Dicentrarchus labrax) juveniles. For that purpose, four diets were formulated with fish oil (FO) and vegetable oils (VO) as the lipid source and with 20 or 0 % gelatinised starch as the carbohydrate source, in a 2×2 factorial design. Liver and intestine antioxidant enzyme activities (catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPX), glutathione reductase (GR), glucose-6-phosphate dehydrogenase (G6PD)), hepatic and intestinal lipid peroxidation (LPO), as well as hepatic oxidative stress index (OSI), were measured in fish fed the experimental diets for 73 d (n 9 fish/diet). Carbohydrate-rich diets promoted a decrease in hepatic LPO and OSI, whereas the lipid source induced no changes. Inversely, dietary lipid source, but not dietary carbohydrate concentration, affected LPO in the intestine. Lower intestinal LPO was observed in VO groups. Enzymes responsive to dietary treatments were GR, G6PD and CAT in the liver and GR and GPX in the intestine. Dietary carbohydrate induced GR and G6PD activities and depressed CAT activity in the liver. GPX and GR activities were increased in the intestine of fish fed VO diets. Overall, effects of diet composition on oxidative status were tissue-related: the liver and intestine were strongly responsive to dietary carbohydrates and lipid sources, respectively. Furthermore, different metabolic routes were more active to deal with the oxidative stress in the two organs studied.

Nutritional and metabolic responses in common dentex (Dentex dentex) fed on different types and levels of carbohydrates.

The present study was aimed to evaluate the capacity of common dentex (Dentex dentex) to efficiently use dietary carbohydrates. So, the effects of different type and levels of carbohydrates on growth performance, feed utilization, fish composition, plasma metabolites and key metabolic pathways in liver and white muscle of common dentex are presented. Nine isonitrogenous (43%) and isoenergetic (22 MJ kg(-1)) diets were formulated combining three types, pregelatinized starch (PS), dextrin (Dx) and maltodextrin (Mx), and three levels (12, 18 and 24%) of carbohydrates. Growth performance was not significantly influenced by treatments. The best feed utilization was observed in 18% Mx group. Higher hepatic lipid content was found in fish fed lower dietary carbohydrate levels. PS induced higher liver and white muscle hexokinase and pyruvate kinase activities compared to the lower values observed for Mx. Malic enzyme and glucose 6-phosphate dehydrogenase in liver and white muscle were lower in Mx group. The influence of dietary carbohydrates source was more noticeable than those induced by the carbohydrate level, when glycolysis and lipogenesis pathways were considered. Common dentex is able to use properly dietary carbohydrates, although optimal dietary inclusion levels are below 24%. The greater protein-sparing effect was promoted by the less complex carbohydrate (maltodextrin) and the best feed utilization indices were obtained at intermediate levels of inclusion (18%).

Maslinic Acid, a Triterpene from Olive, Affects the Antioxidant and Mitochondrial Status of B16F10 Melanoma Cells Grown under Stressful Conditions

Maslinic acid (MA) is a natural compound whose structure corresponds to a pentacyclic triterpene. It is abundant in the cuticular lipid layer of olives. MA has many biological and therapeutic properties related to health, including antitumor, anti-inflammatory, antimicrobial, antiparasitic, antihypertensive, and antioxidant activities. However, no studies have been performed to understand the molecular mechanism induced by this compound in melanoma cancer. The objective of this study was to examine the effect of MA in melanoma (B16F10) cells grown in the presence or absence of fetal bovine serum (FBS). We performed cell proliferation measurements, and the reactive oxygen species (ROS) measurements using dihydrorhodamine 123 (DHR 123) and activities of catalase, glucose 6-phosphate dehydrogenase, glutathione S-transferase, and superoxide dismutase. These changes were corroborated by expression assays. FBS absence reduced cell viability decreasing IC50 values of MA. The DHR 123 data showed an increase in the ROS level in the absence of FBS. Furthermore, MA had an antioxidant effect at lower assayed levels measured as DHR and antioxidant defense. However, at higher dosages MA induced cellular damage by apoptosis as seen in the results obtained.

The oleanolic acid derivative, 3-O-succinyl-28-O-benzyl oleanolate, induces apoptosis in B16–F10 melanoma cells via the mitochondrial apoptotic pathway

Oleanolic acid (1) is a pentacyclic triterpene present in olive pomace, which is known to induce apoptosis and to have anti-tumor properties; however, high concentrations of this product are necessary to produce cytotoxic effects. The 3-O-succinyl-28-O-benzyl oleanolate derivative (4) presents greater cytotoxicity and apoptosis effects than its natural precursor, oleanolic acid, or its benzyl derivative (2). This study examines the response of B16–F10 melanoma cells to treatment with compound 4, in comparison to 1 and 3. Our studies show that treatment with 4 results in a significant inhibition of cell proliferation in a dose-dependent manner and causes apoptotic cell death. At concentrations inhibiting cell growth by 50% and 80%, compound 4 induces strong G0/G1 cell-cycle arrest, around 72–95% apoptosis, and mitochondrial disturbances confirmed by FACS analysis, which probably involve the activation of the intrinsic apoptotic route. Morphological changes including cell shrinkage, chromatin condensation, and loss of nuclear architecture were also observed. In this report, we demonstrated for the first time that in melanoma cancer cells, compound 4 exerts a significant anti-proliferation effect by inducing the apoptotic process with mitochondrial depolarization. These findings support the role of compound 4 as a new, potential therapeutic tool against aberrant cell proliferation in melanoma.

Dietary carbohydrates improve oxidative status of common dentex (Dentex dentex) juveniles, a carnivorous fish species

Common dentex (Dentex dentex) is an appreciated carnivorous fish with high growth rate and life cycle adaptable to existing farming techniques. Since the use of carbohydrates is an economic and sustainable alternative for a protein-sparing effect, the study of how this macronutrient affects the welfare of carnivorous species must be studied. The aim of the present study was to evaluate the effects of different types and levels of carbohydrates on common dentex oxidative status. Nine isonitrogenous (43%) and isoenergetic (22MJkg-1) diets were formulated combining three types (pregelatinized starch-PS, dextrin-Dx and maltodextrin-Mx) and three levels (12, 18 and 24%) of carbohydrates. The activities of catalase-CAT, superoxide dismutase-SOD, glutathione peroxidase-GPX, glutathione reductase-GR and glucose 6-phosphate dehydrogenase-G6PDH, SOD isoenzymatic profile, lipid peroxidation-LPO and protein oxidation-PO were determined in liver and white muscle. SOD and CAT were not affected. GPX in liver and white muscle and GR in liver increased at higher inclusion carbohydrates levels. The lowest levels of GR and G6PDH in both tissues and LPO in liver were observed in maltodextrin groups. No significant effects by carbohydrate source were observed in liver PO and white muscle LPO. Regarding carbohydrate level effect, 18% and 24% dietary inclusion level decreased LPO in white muscle and PO in liver. LPO in liver was also decreased at 24% inclusion level. Altogether, results indicate the use of carbohydrates as an alternative energy source does not produce negative effects on oxidative status of common dentex, on the contrary, even contribute to their oxidative protection.