Aspetti di sostenibilità in agricoltura a diverse scale spaziali e temporali. Valutazioni ambientali, economiche ed energetiche.

La Tesi analizza le relazioni tra i processi di sviluppo agricolo e l’uso delle risorse naturali, in particolare di quelle energetiche, a livello internazionale (paesi in via di sviluppo e sviluppati), nazionale (Italia), regionale (Emilia Romagna) e aziendale, con lo scopo di valutare l’eco-efficienza dei processi di sviluppo agricolo, la sua evoluzione nel tempo e le principali dinamiche in relazione anche ai problemi di dipendenza dalle risorse fossili, della sicurezza alimentare, della sostituzione tra superfici agricole dedicate all’alimentazione umana ed animale. Per i due casi studio a livello macroeconomico è stata adottata la metodologia denominata “SUMMA” SUstainability Multi-method, multi-scale Assessment (Ulgiati et al., 2006), che integra una serie di categorie d’impatto dell’analisi del ciclo di vita, LCA, valutazioni costi-benefici e la prospettiva di analisi globale della contabilità emergetica. L’analisi su larga scala è stata ulteriormente arricchita da un caso studio sulla scala locale, di una fattoria produttrice di latte e di energia elettrica rinnovabile (fotovoltaico e biogas). Lo studio condotto mediante LCA e valutazione contingente ha valutato gli effetti ambientali, economici e sociali di scenari di riduzione della dipendenza dalle fonti fossili. I casi studio a livello macroeconomico dimostrano che, nonostante le politiche di supporto all’aumento di efficienza e a forme di produzione “verdi”, l’agricoltura a livello globale continua ad evolvere con un aumento della sua dipendenza dalle fonti energetiche fossili. I primi effetti delle politiche agricole comunitarie verso una maggiore sostenibilità sembrano tuttavia intravedersi per i Paesi Europei. Nel complesso la energy footprint si mantiene alta poiché la meccanizzazione continua dei processi agricoli deve necessariamente attingere da fonti energetiche sostitutive al lavoro umano. Le terre agricole diminuiscono nei paesi europei analizzati e in Italia aumentando i rischi d’insicurezza alimentare giacché la popolazione nazionale sta invece aumentando.

Quality and safety of oils and fats obtained as co-products or by-products of the food chain and destined to animal production

The aim of the first part of this thesis was to evaluate the effect of trans fatty acid- (TFA), contaminant, polycyclic aromatic hydrocarbon (PAH)- and oxidation productenriched diets on the content of TFA and conjugated linoleic acid (CLA) isomers in meat and liver of both poultry and rabbit. The enriched feedings were prepared with preselected fatty co-and by-products that contained low and high levels of TFA (low, palm fatty acid distillate; high, hydrogenated palm fatty acid distillate), environmental contaminants (dioxins and PCBs) (two different fish oils), PAH (olive oil acid oils and pomace olive oil from chemical refining, for low and high levels) and oxidation products (sunflower-olive oil blend before and after frying), so as to obtain single feedings with three enrichment degrees (high, medium and low) of the compound of interest. This experimental set-up is a part of a large, collaborative European project (http://www.ub.edu/feedfat/), where other chemical and health parameters are assessed. Lipids were extracted, methylated with diazomethane, then transmethylated with 2N KOH/methanol and analyzed by GC and silver-ion TLC-GC. TFA and CLA were determined in the fats, the feedings, meat and liver of both poultry and rabbit. In general, the level of TFA and CLA in meat and liver mainly varied according to those originally found in the feeding fats. It must be pointed out, though, that TFA and CLA accumulation was different for the two animal species, as well as for the two types of tissues. The TFA composition of meat and liver changes according to the composition of the oils added to the feeds with some differences between species. Chicken meat with skin shows higher TFA content (2.6–5.4 fold) than rabbit meat, except for the “PAH” trial. Chicken liver shows higher TFA content (1.2–2.1 fold) than rabbit liver, except for the “TRANS” and “PAH” trials. In both chicken and rabbit meats, the TFA content was higher for the “TRANS” trial, followed by the “DIOXIN” trial. Slight differences were found on the “OXIDATION” and “PAH” trends in both types of meats. In both chicken and rabbit livers, the TFA content was higher for the “TRANS” trial, followed by those of the “PAH”, “DIOXIN” and “OXIDATION” trials. This trend, however, was not identical to that of feeds, where the TFA content varied as follows: “TRANS” > “DIOXIN” >“PAH” > “OXIDATION”. In chicken and rabbit meat samples, C18:1 TFA were the most abundant, followed by C18:2 TFA and C18:3 TFA, except for the “DIOXIN” trial where C18:3 TFA > C18:2 TFA. In chicken and rabbit liver samples of the “TRANS” and “OXIDATION” trials, C18:1 TFA were the most abundant, followed by C18:2 TFA and C18:3 TFA, whereas C18:3 TFA > C18:2 in the “DIOXIN” trial. Slight differences were found on the “PAH” trend in livers from both species. The second part of the thesis dealt with the study of lipid oxidation in washed turkey muscle added with different antioxidants. The evaluation on the oxidative stability of muscle foods found that oxidation could be measured by headspace solid phase microestraction (SPME) of hexanal and propanal. To make this method effective, an antioxidant system was added to stored muscle to stop the oxidative processes. An increase in ionic strength of the sample was also implemented to increase the concentration of aldehydes in the headspace. This method was found to be more sensitive than the commonly used thiobarbituric acid reactive substances (TBARs) method. However, after antioxidants were added and oxidation was stopped, the concentration of aldehydes decreased. It was found that the decrease in aldehyde concentration was due to the binding of the aldehydes to muscle proteins, thus decreasing the volatility and making them less detectable.

Food lipids: their effects on quality and oxidative stability of animal tissues and emulsions

Lipolysis and oxidation of lipids in foods are the major biochemical and chemical processes that cause food quality deterioration, leading to the characteristic, unpalatable odour and flavour called rancidity. In addition to unpalatability, rancidity may give rise to toxic levels of certain compounds like aldehydes, hydroperoxides, epoxides and cholesterol oxidation products. In this PhD study chromatographic and spectroscopic techniques were employed to determine the degree of lipid oxidation in different animal products and its relationship with technological parameters like feeding fat sources, packaging, processing and storage conditions. To achieve this goal capillary gas chromatography (CGC) was employed not only to determine the fatty acids profile but also, after solid phase extraction, the amount of sterols (cholesterol and phytosterols) and cholesterol oxidation products (COPs). To determine hydroperoxides, primary products of oxidation and quantify secondary products UV/VIS absorbance spectroscopy was applied. Beef and pork meat in this study were analysed. In actual fact, lipid oxidation is a major deterioration reaction in meat, meat products and results in adverse changes in the colour, flavour, texture of meat and develops different compounds which should be a risk to human health as oxysterols. On beef and pork meat, a study of lipid fraction during storage was carried out to evaluate its shelf-life and some nutritional features life saturated/unsaturated fatty acids ratio and sterols content, in according to the interest that has been growing around functional food in the last years. The last part of this research was focused on the study of lipid oxidation in emulsions. In oil-in-water emulsions antioxidant activity of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) was evaluated. The rates of lipid oxidation of 1.0% stripped soybean oil-in-water emulsions with DOPC were followed by monitoring lipid hydroperoxide and hexanal as indicators of primary and secondary oxidation products and the droplet surface charge or zeta potential (ζ) of the emulsions with varying concentrations of DOPC were tested. This manuscript reports the main results obtained in the three activities briefly summarized as follows: 1. study on effects of feeding composition on the photoxidative stability of lipids from beef meat, evaluated during storage under commercial retail conditions; 2. evaluation of effects of diets and storage conditions on the oxidative stability of pork meat lipids; 3. study on oxidative behavior of DOPC in stripped soybean oil-in-water emulsions stabilized by nonionic surfactant.

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.

Application of chromatographic and spectroscopic techniques in the evaluation of the lipid fraction of animal products

Lipolysis and oxidation of lipids in foods are the major biochemical and chemical processes that cause food quality deterioration, leading to the characteristic, unpalatable odour and flavour called rancidity. In addition to unpalatability, rancidity may give rise to toxic levels of certain compounds like aldehydes, hydroperoxides, epoxides and cholesterol oxidation products. In this PhD study chromatographic and spectroscopic techniques were employed to determine the degree of rancidity in different animal products and its relationship with technological parameters like feeding fat sources, packaging, processing and storage conditions. To achieve this goal capillary gas chromatography (CGC) was employed not only to determine the fatty acids profile but also, after solid phase extraction, the amount of free fatty acids (FFA), diglycerides (DG), sterols (cholesterol and phytosterols) and cholesterol oxidation products (COPs). To determine hydroperoxides, primary products of oxidation and quantify secondary products UV/VIS absorbance spectroscopy was applied. Most of the foods analysed in this study were meat products. In actual fact, lipid oxidation is a major deterioration reaction in meat and meat products and results in adverse changes in the colour, flavour and texture of meat. The development of rancidity has long recognized as a serious problem during meat handling, storage and processing. On a dairy product, a vegetal cream, a study of lipid fraction and development of rancidity during storage was carried out to evaluate its shelf-life and some nutritional features life saturated/unsaturated fatty acids ratio and phytosterols content. Then, according to the interest that has been growing around functional food in the last years, a new electrophoretic method was optimized and compared with HPLC to check the quality of a beehive product like royal jelly. This manuscript reports the main results obtained in the five activities briefly summarized as follows: 1) comparison between HPLC and a new electrophoretic method in the evaluation of authenticity of royal jelly; 2) study of the lipid fraction of a vegetal cream under different storage conditions; 3) study of lipid oxidation in minced beef during storage under a modified atmosphere packaging, before and after cooking; 4) evaluation of the influence of dietary fat and processing on the lipid fraction of chicken patties; 5) study of the lipid fraction of typical Italian and Spanish pork dry sausages and cured hams.

Application of different instrumental techniques to the selective evaluation of bio-sensible compounds in foodstuff of animal and vegetable origin

This PhD thesis describes the application of some instrumental analytical techniques suitable to the study of fundamental food products for the human diet, such as: extra virgin olive oil and dairy products. These products, widely spread in the market and with high nutritional values, are increasingly recognized healthy properties although their lipid fraction might contain some unfavorable components to the human health. The research activity has been structured in the following investigations: “Comparison of different techniques for trans fatty acids analysis” “Fatty acids analysis of outcrop milk cream samples, with particular emphasis on the content of Conjugated Linoleic Acid (CLA) and trans Fatty Acids (TFA), by using 100m high-polarity capillary column” “Evaluation of the oxidited fatty acids (OFA) content during the Parmigiano-Reggiano cheese seasoning” “Direct analysis of 4-desmethyl sterols and two dihydroxy triterpenes in saponified vegetal oils (olive oil and others) using liquid chromatography-mass spectrometry” “Quantitation of long chain poly-unsatured fatty acids (LC-PUFA) in base infant formulas by Gas Chromatography, and evaluation of the blending phases accuracy during their preparation” “Fatty acids composition of Parmigiano Reggiano cheese samples, with emphasis on trans isomers (TFA)”

Food safety and zoonotic enteric pathogens: sources, risk factors and transmission routes of human salmonellosis and campylobacteriosis

Salmonella and Campylobacter are common causes of human gastroenteritis. Their epidemiology is complex and a multi-tiered approach to control is needed, taking into account the different reservoirs, pathways and risk factors. In this thesis, trends in human gastroenteritis and food-borne outbreak notifications in Italy were explored. Moreover, the improved sensitivity of two recently-implemented regional surveillance systems in Lombardy and Piedmont was evidenced, providing a basis for improving notification at the national level. Trends in human Salmonella serovars were explored: serovars Enteritidis and Infantis decreased, Typhimurium remained stable and 4,[5],12:i:-, Derby and Napoli increased, suggesting that sources of infection have changed over time. Attribution analysis identified pigs as the main source of human salmonellosis in Italy, accounting for 43–60% of infections, followed by Gallus gallus (18–34%). Attributions to pigs and Gallus gallus showed increasing and decreasing trends, respectively. Potential bias and sampling issues related to the use of non-local/non-recent multilocus sequence typing (MLST) data in Campylobacter jejuni/coli source attribution using the Asymmetric Island (AI) model were investigated. As MLST data become increasingly dissimilar with increasing geographical/temporal distance, attributions to sources not sampled close to human cases can be underestimated. A combined case-control and source attribution analysis was developed to investigate risk factors for human Campylobacter jejuni/coli infection of chicken, ruminant, environmental, pet and exotic origin in The Netherlands. Most infections (~87%) were attributed to chicken and cattle. Individuals infected from different reservoirs had different associated risk factors: chicken consumption increased the risk for chicken-attributed infections; animal contact, barbecuing, tripe consumption, and never/seldom chicken consumption increased that for ruminant-attributed infections; game consumption and attending swimming pools increased that for environment-attributed infections; and dog ownership increased that for environment- and pet-attributed infections. Person-to-person contacts around holiday periods were risk factors for infections with exotic strains, putatively introduced by returning travellers.