Lipid quality and oxidative stability in food products from animal origin, as affected by breeding factor

Nowadays it is requested more investigations on alternative rearing systems that are able to improve poultry welfare and to warrant high-quality and safe meat products. This thesis work was focused on the evaluation of the oxidative stability of poultry meats, obtained with different rearing systems, diets (supplemented with bioactive compounds), and packaging conditions. The thesis work was divided into the following parts: - Evaluation of the effects of different rearing systems on the quality, fatty acid composition and oxidative stability of poultry thigh and breast meat belonging to different product categories (“rotisserie” and “cut-up” carcasses); - Evaluation of the effects of different rearing systems and packaging conditions on the shelf-life of poultry thigh meat stored at 4°C for 14 days, and the effects of feed supplementation with thymol (control diet and diet with 2 different concentration of thymol) and packaging conditions on lipid oxidation of poultry thigh meat shelf-life (stored at 4°C for 14 days). The oxidative stability of poultry meat was studied by means of the spectrophotometric determinations of peroxide value and thiobarbituric acid reactive substances. - Evaluation of anti-inflammatory effects of different flavonoids (thymol, luteolin, tangeretin, sulforaphane, polymethoxyflavones, curcumin derivates) to detect their biological activity in LPS-stimulated RAW 264.7 macrophage cells in vitro, in order to study more in depth their action mechanisms. It was evaluated the cell vitality (MTT assay), nitrite concentration and protein profile. The study was focused on the identification of potential dietary bioactive compounds in order to investigate their biological activity and possible synergic effects, and to develop new suitable strategies for long-term promotion of human health, in particular against cancer.

Farming and processing stategies for improving poultry meat quality

The purpose of the PhD research was the identification of new strategies of farming and processing, with the aim to improve the nutritional and technological characteristics of poultry meat. Part of the PhD research was focused on evaluation of alternative farming systems, with the aim to increase animal welfare and to improve the meat quality and sensorial characteristics in broiler chickens. It was also assessed the use of innovative ingredients for marination of poultry meat (sodium bicarbonate and natural antioxidants) The research was developed by studying the following aspects: - Meat quality characteristics, oxidative stability and sensorial traits of chicken meat obtained from two different farming systems: free range vs conventional; - Meat quality traits of frozen chicken breast pre-salted using increasing concentrations of sodium chloride; - Use of sodium bicarbonate in comparison with sodium trypolyphosphate for marination of broiler breast meat and phase; - Marination with thyme and orange essential oils mixture to improve chicken meat quality traits, susceptibility to lipid oxidation and sensory traits. The following meat quality traits analyseswere performed: Colour, pH, water holding capacity by conventional (gravimetric methods, pressure application, centrifugation and cooking) and innovative methods (low-field NMR and DSC analysis) ability to absorb marinade soloutions, texture (shear force using different probes and texture profile analysis), proximate analysis (moisture, proteins, lipids, ash content, collagen, fatty acid), susceptibility to lipid oxidation (determinations of reactive substances with thiobarbituric acid and peroxide value), sensorial analysis (triangle test and consumer test).

Fine tuning of on-growing diets for farmed marine fish species as a key tool to sustain the aquaculture development

The aim of this thesis was to investigate some important key factors able to promote the prospected growth of the aquaculture sector. The limited availability of fishmeal and fish oil led the attention of the aquafeed industry to reduce the dependency on marine raw materials in favor of vegetable ingredients. In Chapter 2, we reported the effects of fishmeal replacement by a mixture of plant proteins in turbot (Psetta maxima L.) juveniles. At the end of the trial, it was found that over the 15% plant protein inclusion can cause stress and exert negative effects on growth performance and welfare. Climate change aroused the attention of the aquafeed industry toward the production of specific diets capable to counteract high temperatures. In Chapter 3, we investigated the most suitable dietary lipid level for gilthead seabream (Sparus aurata L.) reared at Mediterranean summer temperature. In this trial, it was highlighted that 18% dietary lipid allows a protein sparing effect, thus making the farming of this species economically and environmentally more sustainable. The introduction of new farmed fish species makes necessary the development of new species-specific diets. In Chapter 4, we assessed growth response and feed utilization of common sole (Solea solea L.) juveniles fed graded dietary lipid levels. At the end of the trial, it was found that increasing dietary lipids over 8% led to a substantial decline in growth performance and feed utilization indices. In Chapter 5, we investigated the suitability of mussel meal as alternative ingredient in diets for common sole juveniles. Mussel meal proved to be a very effective alternative ingredient for enhancing growth performance, feed palatability and feed utilization in sole irrespectively to the tested inclusion levels. This thesis highlighted the importance of formulating more specific diets in order to support the aquaculture growth in a sustainable way.

Farming and processing factors for improving quality properties of poultry and rabbit meat

Recently, global meat market is facing several dramatic changes due to shifting in diet and life style, consumer demands, and economical considerations. Firstly, there was a tremendous increase in the poultry meat demand. Furthermore, current forecast and projection studies pointed out that the expansion of the poultry market will continue in future. In response to this demand, there was a great success to increase growth rate of meat-type chickens in the last few decades in order to optimize the production of poultry meat. Accordingly, the increase of growth rate induced the appearance of several muscle abnormalities such as pale-soft-exudative (PSE) syndrome and deep-pectoral-myopathy (DPM) and more recently white striping and wooden breast. Currently, there is growing interest in meat industry to understand how much the magnitude of the effect of these abnormalities on different quality traits for raw and processed meat. Therefore, the major part of the research activities during the PhD project was dedicated to evaluate the different implications of recent muscle abnormalities such as white striping and wooden breast on meat quality traits and their incidence under commercial conditions. Generally, our results showed that the incidence of these muscle abnormalities was very high under commercial conditions and had great adverse impact on meat quality traits. Secondly, there is growing market share of convenient, healthy, and functional processed meat products. Accordingly, the remaining part of research activities of the PhD project was dedicated to evaluate the possibility to formulate processed meat products with higher perceived healthy profile such as phosphate free-marinated chicken meat and low sodium-marinated rabbit meat products. Overall all findings showed that sodium bicarbonate can be considered as promising component to replace phosphates in meat products, while potassium chloride under certain conditions was successfully used to produce low marinated rabbit meat products.

From fossil to sustainable diets: an assessment of farming energy footprint

The dependence of industrial agricolture on fossil fuels has been assessed in two comparative case studies between Italy (Emilia-Romagna and Piemonte)and Missouri. The first is related to dairy farming; 15 different farms were surveyed, divided into three different groups: grain based, pasture based and organic. The second is devoted to rice cropping; 12 holdings were examined divided into two groups: conventional and organic. Energy footprint was determined for structures, machinery, fertilizers, pesticides, fuel, electricity, feed and seeds. Possible scenarios of transition to a more sustainable agricolture based on renewable energy sources were analized in detail for all the farms analized.

Tackling of unhealthy diets, physical inactivity, and obesity: health effects and cost-effectiveness

The obesity epidemic is spreading to low-income and middle-income countries as a result of new dietary habits and sedentary ways of life, fuelling chronic diseases and premature mortality. In this report we present an assessment of public health strategies designed to tackle behavioural risk factors for chronic diseases that are closely linked with obesity, including aspects of diet and physical inactivity, in Brazil, China, India, Mexico, Russia, and South Africa. England was included for comparative purposes. Several population-based prevention policies can be expected to generate substantial health gains while entirely or largely paying for themselves through future reductions of health-care expenditures. These strategies include health information and communication strategies that improve population awareness about the benefits of healthy eating and physical activity; fiscal measures that increase the price of unhealthy food content or reduce the cost of healthy foods rich in fibre; and regulatory measures that improve nutritional information or restrict the marketing of unhealthy foods to children. A package of measures for the prevention of chronic diseases would deliver substantial health gains, with a very favourable cost-effectiveness profile.

Polyunsaturated fatty acid-enriched diets used for the treatment of canine chronic enteropathies decrease the abundance of selected genes of cholesterol homeostasis

Lipids are important for cell function and survival, but abnormal concentrations may lead to various diseases. Cholesterol homeostasis is greatly dependent on the active transport by membrane proteins, whose activities coordinate lipid status with cellular function. Intestinal Niemann-Pick C1-Like 1 protein (NPC1L1) and scavenger receptor B1 (SR-B1) participate in the uptake of extracellular cholesterol, whereas ATP binding cassette A1 (ABCA1) mediates the efflux of excessive intracellular cholesterol. Caveolin-1 binds cholesterol and fatty acids (FA) and participates in cholesterol trafficking. Sterol response element binding protein-2 (SREBP-2) is a sensor that regulates intracellular cholesterol synthesis. Given that cholesterol is a constituent of chylomicrons, whose synthesis is enhanced with an increased FA supply, we tested the hypothesis that feeding polyunsaturated FA (PUFA)-enriched diets in treatment of canine chronic enteropathies alters the mRNA expression of genes involved in cholesterol homeostasis. Using quantitative reverse transcriptase polymerase chain reaction (RT-PCR), we compared the mRNA abundance of NPC1L1, SR-B1, ABCA1, caveolin-1, and SREBP-2 in duodenal mucosal biopsies of dogs with food-responsive diarrhea (FRD; n=14) and inflammatory bowel disease (IBD; n=7) before and after treatment with cholesterol-free PUFA-enriched diets and in healthy controls (n=14). The abundance of caveolin-1, ABCA1, and SREBP-2 were altered by PUFA-enriched diets (P<0.05), whereas that of NPC1L1 and SR-B1 mRNA remained unchanged. The gene expression of caveolin-1, ABCA1, and SREBP-2 was down-regulated (P<0.05) by PUFA-enriched diets in IBD dogs only. Our results suggest that feeding PUFA-enriched diets may alter cholesterol homeostasis in duodenal mucosal cells of dogs suffering from IBD.

Plasma PCSK9 concentrations during an oral fat load and after short term high-fat, high-fat high-protein and high-fructose diets

Background PCSK9 (Proprotein Convertase Subtilisin Kexin type 9) is a circulating protein that promotes hypercholesterolemia by decreasing hepatic LDL receptor protein. Under non interventional conditions, its expression is driven by sterol response element binding protein 2 (SREBP2) and follows a diurnal rhythm synchronous with cholesterol synthesis. Plasma PCSK9 is associated to LDL-C and to a lesser extent plasma triglycerides and insulin resistance. We aimed to verify the effect on plasma PCSK9 concentrations of dietary interventions that affect these parameters. Methods We performed nutritional interventions in young healthy male volunteers and offspring of type 2 diabetic (OffT2D) patients that are more prone to develop insulin resistance, including: i) acute post-prandial hyperlipidemic challenge (n=10), ii) 4 days of high-fat (HF) or high-fat/high-protein (HFHP) (n=10), iii) 7 (HFruc1, n=16) or 6 (HFruc2, n=9) days of hypercaloric high-fructose diets. An acute oral fat load was also performed in two patients bearing the R104C-V114A loss-of-function (LOF) PCSK9 mutation. Plasma PCSK9 concentrations were measured by ELISA. For the HFruc1 study, intrahepatocellular (IHCL) and intramyocellular lipids were measured by 1H magnetic resonance spectroscopy. Hepatic and whole-body insulin sensitivity was assessed with a two-step hyperinsulinemic-euglycemic clamp (0.3 and 1.0 mU.kg-1.min-1). Findings HF and HFHP short-term diets, as well as an acute hyperlipidemic oral load, did not significantly change PCSK9 concentrations. In addition, post-prandial plasma triglyceride excursion was not altered in two carriers of PCSK9 LOF mutation compared with non carriers. In contrast, hypercaloric 7-day HFruc1 diet increased plasma PCSK9 concentrations by 28% (p=0.05) in healthy volunteers and by 34% (p=0.001) in OffT2D patients. In another independent study, 6-day HFruc2 diet increased plasma PCSK9 levels by 93% (p<0.0001) in young healthy male volunteers. Spearman’s correlations revealed that plasma PCSK9 concentrations upon 7-day HFruc1 diet were positively associated with plasma triglycerides (r=0.54, p=0.01) and IHCL (r=0.56, p=0.001), and inversely correlated with hepatic (r=0.54, p=0.014) and whole-body (r=−0.59, p=0.0065) insulin sensitivity. Conclusions Plasma PCSK9 concentrations vary minimally in response to a short term high-fat diet and they are not accompanied with changes in cholesterolemia upon high-fructose diet. Short-term high-fructose intake increased plasma PCSK9 levels, independent on cholesterol synthesis, suggesting a regulation independent of SREBP-2. Upon this diet, PCSK9 is associated with insulin resistance, hepatic steatosis and plasma triglycerides.

Maternal and Postweaning High-Fat Diets Disturb Hippocampal Gene Expression, Learning, and Memory Function

We tested the hypothesis that excess saturated fat consumption during pregnancy, lactation, and/or postweaning alters the expression of genes mediating hippocampal synaptic efficacy and impairs spatial learning and memory in adulthood. Dams were fed control chow or a diet high in saturated fat before mating, during pregnancy, and into lactation. Offspring were weaned to either standard chow or a diet high in saturated fat. The Morris Water Maze was used to evaluate spatial learning and memory. Open field testing was used to evaluate motor activity. Hippocampal gene expression in adult males was measured using RT-PCR and ELISA. Offspring from high fat-fed dams took longer, swam farther, and faster to try and find the hidden platform during the 5-day learning period. Control offspring consuming standard chow spent the most time in memory quadrant during the probe test. Offspring from high fat-fed dams consuming excess saturated fat spent the least. The levels of mRNA and protein for brain-derived neurotrophic factor and activity-regulated cytoskeletal-associated protein were significantly decreased by maternal diet effects. Nerve growth factor mRNA and protein levels were significantly reduced in response to both maternal and postweaning high-fat diets. Expression levels for the N-methyl-D-aspartate receptor (NMDA) receptor subunit NR2B as well as synaptophysin were significantly decreased in response to both maternal and postweaning diets. Synaptotagmin was significantly increased in offspring from high fat-fed dams. These data support the hypothesis that exposure to excess saturated fat during hippocampal development is associated with complex patterns of gene expression and deficits in learning and memory.

Speciating Campylobacter jejuni and Campylobacter coli isolates from poultry and humans using six PCR-based assays

Six previously published polymerase chain reaction (PCR) assays each targeting different genes were used to speciate 116 isolates previously identified as Campylobacter jejuni using routine microbiological techniques. Of the 116 isolates, 84 were of poultry origin and 32 of human origin. The six PCR assays confirmed the species identities of 31 of 32 (97%) human isolates and 56 of 84 (67%) poultry isolates as C. jejuni. Twenty eight of 84 (33%) poultry isolates were identified as Campylobacter coli and the remaining human isolate was tentatively identified as Campylobacter upsaliensis based on the degree of similarity of 16S rRNA gene sequences. Four of six published PCR assays showed 100% concordance in their ability to speciate 113 of the 116 (97.4%) isolates; two assays failed to generate a PCR product with four to 10 isolates. A C. coli-specific PCR identified all 28 hippuricase gene (hipO)-negative poultry isolates as C. coli although three isolates confirmed to be C. jejuni by the remaining five assays were also positive in this assay. A PCR-restriction fragment length polymorphism assay based on the 16S rRNA gene was developed, which contrary to the results of the six PCR-based assays, identified 28 of 29 hipO-negative isolates as C. jejuni. DNA sequence analysis of 16S rRNA genes from four hipO-negative poultry isolates showed they were almost identical to the C. jejuni type strain 16S rRNA sequences ATCC43431 and ATCC33560 indicating that assays reliant on 16S rRNA sequence may not be suitable for the differentiation of these two species.

Substituting Wet Distillers Grains or Condensed Distillers Solubles for Corn Grain in Finishing Diets for Yearling Heifers

A feeding trial was conducted with 790-lb yearling heifers fed an average of 121 days to evaluate replacing cracked corn and supplemental urea with wet distillers grains or condensed distillers solubles. Wet distillers grains were evaluated at 16%, 28% and 40% of diet dry matter. Condensed distillers solubles were added at 6.5% of diet dry matter. Control diets were supplemented with urea or a combination of urea and soybean meal. Feeding 16% wet distillers grains or condensed distillers solubles increased gain of heifers compared with those fed the control urea diet. Increasing the amount of wet distillers grains tended to decrease feed intake and reduce gain. The calculated apparent net energy based on gain of the heifers was greatest for the heifers fed 16% wet distillers grains. The apparent energy of the wet distillers grains declined as the quantity fed was increased. The calculated net energy values were 1.09 and 1.35 Mcal/lb of dry matter for the average of the three concentrations of wet distillers grains and condensed distillers solubles. These results confirm the high energy values of wet distillers grains relative to cracked corn as observed in a previous steer feeding trial.

Evaluation of Wet Distillers Grains in Finishing Diets for Yearling Steers

A feeding trial was conducted with 940-lb yearling steers fed 113 days to determine the feeding value of distillers grains relative to corn grain. Replacing corn and urea with wet distillers grains for 20% of the diet dry matter tended to increase gain with no increase in feed consumption, resulting in improved feed conversion. Replacing 40% of diet dry matter with wet distillers grains decreased feed intake without affecting gains, and improved feed efficiency. The overall average estimated net energy value of wet distillers grains was 1.20 Mcal NEg per pound dry matter. This experiment confirmed the observations in previous cattle feeding experiments, that for finishing cattle wet distillers grains have a high energy value compared with cracked corn grain. Another objective of the study was to determine if cattle being fed wet distillers grains could be suddenly changed to a different diet if the supply of wet feed was suddenly disrupted. It was found that if intake is managed during the change, that distillers grains portion of the diet can be suddenly changed from wet to dry and then changed back to wet after a week, without sacrificing performance of the cattle.

Evaluation of Condensed Porcine Solubles as a Source of Supplemental Nitrogen in Corn-Based Finishing Diets for Yearling Steers

A feeding trial was conducted with 860-lb yearling steers fed 121 days to evaluate Condensed Porcine Solubles (Porcine Solubles) as a source of supplemental nitrogen for finishing cattle. Diets containing 5% soybean meal, 1.46% urea, and 2% or 4% Porcine Solubles were compared. When first offered, cattle did not want to consume feed containing the Porcine Solubles. Following adaptation, feed containing up to 4% Porcine Solubles was readily consumed. During the first 56 days, steers fed soybean meal gained faster and were more efficient than steers fed urea or Porcine Solubles. At the end of the trial there were no differences among the nitrogen supplements in feed intake, gain, or feed conversion. There were no significant differences in carcass weight or measures of carcass quality.

Evaluation of a Mixture of Corn Steep Liquor and Distillers Solubles as a Replacement for Corn and Supplement in Cattle Finishing Diets

Finishing yearling steers fed a corn-based diet containing steep liquor had statistically similar live performance as steers fed the control diet. Numerically steers fed the steep containing diet were 6% more efficient. Steers fed steep liquor tended to contain less carcass fat (as measured by intramuscular marbling) less kidney, heart and pelvic fat, and less backfat thickness. When priced at $50/ton adding steep liquor at 10% of diet dry matter reduced feed cost for gain 9%.