Thesis (Ph. D.)--University of Texas at Austin, 1998. / Vita. Includes bibliographical references. Available also in a digital version from Dissertation Abstracts.

The oxidation of lipids and antioxidants has been extensively studied in human plasma but little attention has been given to how plasma proteins are oxidised. Proteins make up the majority of biomolecules in cells and plasma and therefore are the most likely reactants with oxidants and free radicals. Previous studies in the laboratory had shown that peroxyl radicals generated by the thermolytic decay of 2-azobis (2-amdinopropane) dihydrochloride (AAPH) generated significant amounts of protein hydroperoxides, but only after a six hour lag period. In this study the existence of the six hour lag period was confirmed and shown by dialysis of the plasma to be due to the presence of low molecular weight antioxidants. The addition of both uric acid and ascorbic acid to the dialysed plasma restored the lag phase suggesting that in vivo these antioxidants act to prevent protein hydroperoxide formation. Lipid oxidation was also observed in the plasma but only after a two hour lag phase. This was the first time lipid oxidation has been observed in the absence of protein oxidation. The lipid lag phase was also abolished by dialysis of the plasma and restored by the addition of ascorbic acid and uric acid. The kinetics of tocopherol loss suggests that the tocopherol radicals act to inhibit lipid oxidation by transferring the electrons to the water-soluble ascorbate. The loss of ascorbate appears to cause the formation of a tocopherol radical mediate the lipid peroxidation process. Overall the data shows ascorbic acid scavenging the peroxyl radicals while uric acid acts to reduce the overall AAPH generated radical flux. In a separate investigation, the production of protein-bound DOPA (PB-DOPA) on albumin during X-ray radiolysis and copper mediate Fenton oxidation was investigated using a fluorescence based derivatisation method (ED-DOPA), which was compared with the more specific acid hydrolysis and HPLC analysis method. The ED-DOPA method consistently gave a much higher reading that the HPLC based methods, suggesting that the ED-DOPA method was measuring DOPA plus DOPA oxidation products. This was confirmed by oxidising X-ray radiolysis generated PB-DOPA with Cu++ to cause DOPA oxidation. The Cu++ treatment drastically increased the level of signal given by the ED-DOPA assay while HPLC analysis showed all the DOPA had been oxidised.

Os recém-nascidos possuem o sistema antioxidante imaturo, por haver baixa tensão de oxigênio no ambiente intrauterino durante a vida fetal. Logo após o nascimento, as alterações súbitas das condições fisiológicas e ambientais causam significativo aumento no consumo de oxigênio, desencadeando, assim, a produção de radicais livres. Tais condições promovem vulnerabilidade dos neonatos ao efeito negativo do estresse oxidativo, o que potencialmente podem prejudicar a vitalidade neonatal. O presente estudo teve como objetivo comparar o perfil antioxidante e estresse oxidativo de neonatos e fêmeas caninas no periparto eutócico vaginal ou cesariana eletiva, e avaliar a influência da condição obstétrica para o estado oxidativo. Foram selecionadas 21 cadelas gestantes, as quais, constituíram dois grupos amostrais, de acordo com a condição obstétrica: Eutocia Vaginal (n = 10) e Cesariana Eletiva (n = 11); e seus respectivos neonatos foram alocados em subgrupos de acordo com a condição obstétrica e momento do nascimento: Eutocia Vaginal Inicial (n=10), Eutocia Vaginal Final (n = 9), Cesariana Eletiva Inicial (n = 11) e Cesariana Eletiva final (n= 10). As cadelas foram avaliadas no período pródromo do parto, intraparto; uma hora e três dias pós-parto, quando amostras de sangue foram colhidas para análise do perfil antioxidante [dosagem das enzimas superóxido dismutase (SOD), glutationa peroxidase (GPx), dosagem da concentração de tióis totais e determinação do status antioxidante total(TAC)] e do estresse oxidativo [dosagem da peroxidação lipídica (TBARS) e da oxidação de proteínas]. Os neonatos foram avaliados quanto ao escore Apgar aos 0 e 60 minutos do nascimento; avaliação clínica (frequências cardíaca e respiratória; escore de tônus muscular, irritabilidade reflexa e coloração de mucosa, aferição da temperatura corporal), lactatemia sanguínea, oximetria de pulso, determinação do perfil antioxidante e do estresse oxidativo e aferição do peso corporal aos 0, 60 minutos, às 12, 24 horas e ao 3º dia pós nascimento. As cadelas do Grupo Eutocia Vaginal apresentaram maior peroxidação lipídica, oxidação de proteínas e atividade de SOD e menor atividade de GPx e concentração de tióis totais em comparação ao Grupo Cesariana Eletiva. A capacidade antioxidante total elevou-se após 1h do parto em comparação aos outros momentos de avaliação no Grupo Cesariana Eletiva. Embora os neonatos do Grupo Eutocia Vaginal tenham apresentado melhores parâmetros de vitalidade neonatal, em comparação ao Grupo Cesariana Eletiva, todos os neonatos apresentaram adequada evolução do escore Apgar, coloração de mucosa, irritabilidade reflexa, tônus muscular e oxigenação periférica após 1h do nascimento. A lactatemia sanguínea foi maior no Grupo Eutocia Vaginal, bem como nos neonatos nascidos ao final do parto. A peroxidação lipídica foi superior nos neonatos nascidos por eutocia vaginal em comparação aos nascidos por cesariana eletiva, enquanto a oxidação de proteínas mostrou-se maior nos primeiros neonatos nascidos por eutocia vaginal em comparação aos nascidos ao final do parto. Porém, resultado contrário foi verificado para o Grupo Cesariana Eletiva, pois os neonatos nascidos ao final da cirurgia apresentaram maior valor de oxidação de proteínas. Ademais, para os neonatos nascidos ao final do parto, o Grupo Cesariana Eletiva apresentou maior oxidação proteica em comparação ao Grupo Eutocia Vaginal. A atividade da GPx foi superior nos neonatos nascidos por cesariana eletiva. Em conclusão, a condição obstétrica impõe diferenças no perfil oxidativo e antioxidante em cadelas e neonatos, os quais apresentam estado oxidativo semelhante, denotando influência materna sobre o equilíbrio oxidativo dos recém-nascidos. / Newborns have an immature antioxidant system, due to low oxygen exposure in intrauterine environment during fetal life. Immediately after birth, sudden changes of physiological and environmental conditions cause a significant increase in oxygen consumption, resulting in the production of free radicals. These conditions turn the newborn vulnerable to the negative effects of oxidative stress, which potentially can impair neonatal vitality. This study aimed to compare the antioxidant profile and oxidative stress of neonates and canine females during vaginal labour or elective cesarean section, and to evaluate whether the obstetric condition influences their oxidative status. For this purpose, 21 pregnant bitches were subjected to two experimental groups, according to the obstetric condition: Vaginal Eutocia (n = 10) and Elective Cesarian Section (n = 11) and their respective newborns were allocated into subgroups according to the obstetric condition and moment of birth: Inicial Vaginal Eutocia (n=10), Final Vaginal Eutocia (n = 9), Inicial Elective Cesarian Section (n = 11) and Final Elective Cesarian Section (n= 10). Bitches were evaluated during the preparatory phase of whelping, intrapartum; one and 72 hours postpartum, when blood samples were collected for analysis of the antioxidant profile [Superoxide Dismutase (SOD) and Glutathione Peroxidase (GPx) activity enzymes assays, Total Antioxidant Capacity (TAC) assay and Total Thiols Concentration assay] and oxidative stress [lipid peroxidation (TBARS) and protein oxidation assays]. Neonates were evaluated for the Apgar score at 0 and 60 minutes of birth; clinical evaluation (heart and respiratory rates; muscle tone, irritability reflex and mucous color score; and body temperature), blood lactate, pulse oximetry, determination of antioxidant profile and oxidative stress, and body weight measurement at 0, 60 minutes, 12, 24 and 72 hours after birth. The Vaginal Eutocia bitches had higher lipid peroxidation, protein oxidation and SOD activity and lower GPx activity and total thiols concentration in comparison to the Elective Cesarian Section Group. Total antioxidant capacity was higher 1 hour postpartum compared to the others evaluation moments in the Elective Cesarian Section Group. Although neonates from the Vaginal Eutocia Group presented better neonatal vitality than those from the Elective Cesarian Section, all neonates presented adequate evolution of the Apgar score, mucous color, irritability reflex, muscle tone and pulse oximetry 1 hour postpartum. Blood lactatemia was higher in the Vaginal Eutocia Group, as well as for the last neonates. Lipid peroxidation was higher in neonates born by vaginal eutocia compared to those born by elective cesarean section, whereas protein oxidation was higher in the first neonates born by vaginal eutocia compared to those born at the end of delivery. Conversely, Elective Cesarian Section neonates born at the end of surgery had higher protein oxidation. In addition, for those neonates born at the end of delivery, the Elective Cesarian Section group presented higher protein oxidation compared to the Vaginal Eutocia group. Furthermore, GPx activity was higher in neonates born by elective caesarean section. In conclusion, the obstetric condition imposes differences in the oxidative and antioxidant profile in bitches and neonates with similar oxidative status, denoting maternal influence on the oxidative balance of the newborns.

Os recém-nascidos possuem o sistema antioxidante imaturo, por haver baixa tensão de oxigênio no ambiente intrauterino durante a vida fetal. Logo após o nascimento, as alterações súbitas das condições fisiológicas e ambientais causam significativo aumento no consumo de oxigênio, desencadeando, assim, a produção de radicais livres. Tais condições promovem vulnerabilidade dos neonatos ao efeito negativo do estresse oxidativo, o que potencialmente podem prejudicar a vitalidade neonatal. O presente estudo teve como objetivo comparar o perfil antioxidante e estresse oxidativo de neonatos e fêmeas caninas no periparto eutócico vaginal ou cesariana eletiva, e avaliar a influência da condição obstétrica para o estado oxidativo. Foram selecionadas 21 cadelas gestantes, as quais, constituíram dois grupos amostrais, de acordo com a condição obstétrica: Eutocia Vaginal (n = 10) e Cesariana Eletiva (n = 11); e seus respectivos neonatos foram alocados em subgrupos de acordo com a condição obstétrica e momento do nascimento: Eutocia Vaginal Inicial (n=10), Eutocia Vaginal Final (n = 9), Cesariana Eletiva Inicial (n = 11) e Cesariana Eletiva final (n= 10). As cadelas foram avaliadas no período pródromo do parto, intraparto; uma hora e três dias pós-parto, quando amostras de sangue foram colhidas para análise do perfil antioxidante [dosagem das enzimas superóxido dismutase (SOD), glutationa peroxidase (GPx), dosagem da concentração de tióis totais e determinação do status antioxidante total(TAC)] e do estresse oxidativo [dosagem da peroxidação lipídica (TBARS) e da oxidação de proteínas]. Os neonatos foram avaliados quanto ao escore Apgar aos 0 e 60 minutos do nascimento; avaliação clínica (frequências cardíaca e respiratória; escore de tônus muscular, irritabilidade reflexa e coloração de mucosa, aferição da temperatura corporal), lactatemia sanguínea, oximetria de pulso, determinação do perfil antioxidante e do estresse oxidativo e aferição do peso corporal aos 0, 60 minutos, às 12, 24 horas e ao 3º dia pós nascimento. As cadelas do Grupo Eutocia Vaginal apresentaram maior peroxidação lipídica, oxidação de proteínas e atividade de SOD e menor atividade de GPx e concentração de tióis totais em comparação ao Grupo Cesariana Eletiva. A capacidade antioxidante total elevou-se após 1h do parto em comparação aos outros momentos de avaliação no Grupo Cesariana Eletiva. Embora os neonatos do Grupo Eutocia Vaginal tenham apresentado melhores parâmetros de vitalidade neonatal, em comparação ao Grupo Cesariana Eletiva, todos os neonatos apresentaram adequada evolução do escore Apgar, coloração de mucosa, irritabilidade reflexa, tônus muscular e oxigenação periférica após 1h do nascimento. A lactatemia sanguínea foi maior no Grupo Eutocia Vaginal, bem como nos neonatos nascidos ao final do parto. A peroxidação lipídica foi superior nos neonatos nascidos por eutocia vaginal em comparação aos nascidos por cesariana eletiva, enquanto a oxidação de proteínas mostrou-se maior nos primeiros neonatos nascidos por eutocia vaginal em comparação aos nascidos ao final do parto. Porém, resultado contrário foi verificado para o Grupo Cesariana Eletiva, pois os neonatos nascidos ao final da cirurgia apresentaram maior valor de oxidação de proteínas. Ademais, para os neonatos nascidos ao final do parto, o Grupo Cesariana Eletiva apresentou maior oxidação proteica em comparação ao Grupo Eutocia Vaginal. A atividade da GPx foi superior nos neonatos nascidos por cesariana eletiva. Em conclusão, a condição obstétrica impõe diferenças no perfil oxidativo e antioxidante em cadelas e neonatos, os quais apresentam estado oxidativo semelhante, denotando influência materna sobre o equilíbrio oxidativo dos recém-nascidos. / Newborns have an immature antioxidant system, due to low oxygen exposure in intrauterine environment during fetal life. Immediately after birth, sudden changes of physiological and environmental conditions cause a significant increase in oxygen consumption, resulting in the production of free radicals. These conditions turn the newborn vulnerable to the negative effects of oxidative stress, which potentially can impair neonatal vitality. This study aimed to compare the antioxidant profile and oxidative stress of neonates and canine females during vaginal labour or elective cesarean section, and to evaluate whether the obstetric condition influences their oxidative status. For this purpose, 21 pregnant bitches were subjected to two experimental groups, according to the obstetric condition: Vaginal Eutocia (n = 10) and Elective Cesarian Section (n = 11) and their respective newborns were allocated into subgroups according to the obstetric condition and moment of birth: Inicial Vaginal Eutocia (n=10), Final Vaginal Eutocia (n = 9), Inicial Elective Cesarian Section (n = 11) and Final Elective Cesarian Section (n= 10). Bitches were evaluated during the preparatory phase of whelping, intrapartum; one and 72 hours postpartum, when blood samples were collected for analysis of the antioxidant profile [Superoxide Dismutase (SOD) and Glutathione Peroxidase (GPx) activity enzymes assays, Total Antioxidant Capacity (TAC) assay and Total Thiols Concentration assay] and oxidative stress [lipid peroxidation (TBARS) and protein oxidation assays]. Neonates were evaluated for the Apgar score at 0 and 60 minutes of birth; clinical evaluation (heart and respiratory rates; muscle tone, irritability reflex and mucous color score; and body temperature), blood lactate, pulse oximetry, determination of antioxidant profile and oxidative stress, and body weight measurement at 0, 60 minutes, 12, 24 and 72 hours after birth. The Vaginal Eutocia bitches had higher lipid peroxidation, protein oxidation and SOD activity and lower GPx activity and total thiols concentration in comparison to the Elective Cesarian Section Group. Total antioxidant capacity was higher 1 hour postpartum compared to the others evaluation moments in the Elective Cesarian Section Group. Although neonates from the Vaginal Eutocia Group presented better neonatal vitality than those from the Elective Cesarian Section, all neonates presented adequate evolution of the Apgar score, mucous color, irritability reflex, muscle tone and pulse oximetry 1 hour postpartum. Blood lactatemia was higher in the Vaginal Eutocia Group, as well as for the last neonates. Lipid peroxidation was higher in neonates born by vaginal eutocia compared to those born by elective cesarean section, whereas protein oxidation was higher in the first neonates born by vaginal eutocia compared to those born at the end of delivery. Conversely, Elective Cesarian Section neonates born at the end of surgery had higher protein oxidation. In addition, for those neonates born at the end of delivery, the Elective Cesarian Section group presented higher protein oxidation compared to the Vaginal Eutocia group. Furthermore, GPx activity was higher in neonates born by elective caesarean section. In conclusion, the obstetric condition imposes differences in the oxidative and antioxidant profile in bitches and neonates with similar oxidative status, denoting maternal influence on the oxidative balance of the newborns.

The oxidation of lipids and antioxidants has been extensively studied in human plasma but little attention has been given to how plasma proteins are oxidised. Proteins make up the majority of biomolecules in cells and plasma and therefore are the most likely reactants with oxidants and free radicals. Previous studies in the laboratory had shown that peroxyl radicals generated by the thermolytic decay of 2-azobis (2-amdinopropane) dihydrochloride (AAPH) generated significant amounts of protein hydroperoxides, but only after a six hour lag period. In this study the existence of the six hour lag period was confirmed and shown by dialysis of the plasma to be due to the presence of low molecular weight antioxidants. The addition of both uric acid and ascorbic acid to the dialysed plasma restored the lag phase suggesting that in vivo these antioxidants act to prevent protein hydroperoxide formation. Lipid oxidation was also observed in the plasma but only after a two hour lag phase. This was the first time lipid oxidation has been observed in the absence of protein oxidation. The lipid lag phase was also abolished by dialysis of the plasma and restored by the addition of ascorbic acid and uric acid. The kinetics of tocopherol loss suggests that the tocopherol radicals act to inhibit lipid oxidation by transferring the electrons to the water-soluble ascorbate. The loss of ascorbate appears to cause the formation of a tocopherol radical mediate the lipid peroxidation process. Overall the data shows ascorbic acid scavenging the peroxyl radicals while uric acid acts to reduce the overall AAPH generated radical flux. In a separate investigation, the production of protein-bound DOPA (PB-DOPA) on albumin during X-ray radiolysis and copper mediate Fenton oxidation was investigated using a fluorescence based derivatisation method (ED-DOPA), which was compared with the more specific acid hydrolysis and HPLC analysis method. The ED-DOPA method consistently gave a much higher reading that the HPLC based methods, suggesting that the ED-DOPA method was measuring DOPA plus DOPA oxidation products. This was confirmed by oxidising X-ray radiolysis generated PB-DOPA with Cu++ to cause DOPA oxidation. The Cu++ treatment drastically increased the level of signal given by the ED-DOPA assay while HPLC analysis showed all the DOPA had been oxidised.

Orientador: Walkiria Hanada Viotto / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-23T15:48:33Z (GMT). No. of bitstreams: 1 Urzedo_AnaCarolinaBorgesde_D.pdf: 4425198 bytes, checksum: ca9f39f7b13a21e55c2d558014da6722 (MD5) Previous issue date: 2013 / Resumo: A foto-oxidação do leite é o fator determinante na vida de prateleira do leite microfiltrado pasteurizado. A exposição do leite à luz, nas gôndolas dos supermercados, desencadeia a foto-oxidação da riboflavina e das proteínas do leite, provocando mudanças na cor e formação de off-flavors. O objetivo do trabalho foi verificar, em condições reais, a influência do tipo de embalagem e da intensidade da luz na degradação da riboflavina, formação de lumicromo, oxidação de proteína, cor e vida de prateleira do leite desnatado microfiltrado pasteurizado. Leite desnatado microfiltrado pasteurizado foi acondicionado em garrafas de vidro e de polietileno de alta densidade, e armazenados no escuro (controle) e sob incidência de luz (500 e 1200 lux), durante 14 dias de estocagem refrigerada. Análises de espectroscopia de fluorescência da riboflavina, lumicromo, triptofano e ditirosina e análise de cor instrumental foram realizadas para acompanhar a foto-oxidação dos componentes do leite. Para estimar a vida de prateleira do leite microfiltrado pasteurizado, sob diferentes condições de luz e embalagem, contagens de micro-organismos mesófilos aeróbios e análise sensorial com assessores treinados, teste de aceitação e intenção de compra foram realizados durante o tempo de armazenamento refrigerado. Assessores foram treinados para avaliação da cor, aroma e sabor característicos de leite desnatado e de leite desnatado oxidado. Aos 14 dias de estocagem refrigerada, 76,1% da riboflavina foi degradada no leite exposto à radiação de 500 lux. Já a 1200 lux, esse valor foi de 86,4%. A degradação da riboflavina, a formação de lumicromo e a fotodegradação do triptofano foram maiores e mais rápidas quando a intensidade de luz foi mais intensa (1200 lux). Durante o armazenamento refrigerado, a oxidação das proteínas resultou em desnovelamento da estrutura terciária, e consequentemente, em exposição e posterior degradação do triptofano. No período estudado, houve somente formação de ditirosina para os leites submetidos à intensidade de luz mis intensa (1200 lux). A vida de prateleira dos leites armazenados no escuro (controle) foi de 10 a 14 dias. O aparecimento do sabor oxidado, proveniente da foto-oxidação dos componentes do leite, foi o parâmetro determinante para o fim da vida de prateleira dos leites armazenados sob luz. O tipo de embalagem somente influenciou a vida de prateleira do leite, quando a intensidade de exposição à luz foi mais baixa (500 lux). Nessa intensidade de radiação luminosa, a vida de prateleira do leite pasteurizado aumentou de 4-6 dias para 10-13 dias, quando a embalagem de vidro foi substituída pela de polietileno / Abstract: Photo-oxidation of milk is probably the main cause for the end of shelf life of a microfiltered pasteurized milk. Milk is inevitably exposed to light on the supermarket shelves, which triggers the photo-oxidation of riboflavin and milk proteins, affecting the sensory quality with changes in color and formation of off-flavors. The objective was to verify, in real conditions, the influence of the type of packaging and the light intensity on the riboflavin degradation, protein oxidation, color, shelf life of microfiltered pasteurized skim milk. After processing, milk was packaged in glass and high density polyethylene bottles and stored in the dark (control) and under influence of light (500 and 1200 lux), during 14 days of refrigerated storage. Analyses of fluorescence spectroscopy of riboflavin, lumicrome, tryptophan and dityrosine and instrumental color were performed to monitor the photo-oxidation of milk components. The shelf life of pasteurized microfiltered skim milk, under different light conditions and packaging was estimated by standard plate count of aerobic mesophilic and sensory analysis with trained assessors, acceptance testing, and purchase intent, during refrigerated storage time. Assessors were trained to evaluate sensorially the color, aroma and flavor of skim milk and oxidized skim milk. At 14 days of refrigerated storage, 76.1% of riboflavin was degraded in milk exposed to radiation of 500 lux. However, at 1200 lux, degradation of riboflavin reached 86.4% of its initial content in milk. Riboflavin degradation, lumicrome and tryptophan formation were higher and faster when light intensity was more intense (1200 lux). During storage time, the oxidation of proteins resulted in the tertiary structure unfolding, and exposure and subsequent degradation of tryptophan. During this period of time, there was formation of dityrosine only for the milks exposed to more intense light radiation (1200 lux). The shelf life of milk stored in the dark (control) was 10-14 days. The development of oxidized flavor, derived from the photo-oxidation of milk components, was the main parameter for determining the ending of the shelf life of milk stored under light. Packaging material influenced the milk shelf life when the intensity of light was lower (500 lux). In this condition, the shelf life of pasteurized milk increased from 10-13 days to 4-6 days when the glass container was replaced by polyethylene bottle / Doutorado / Tecnologia de Alimentos / Doutora em Tecnologia de Alimentos

This study investigated the influence of dietary antioxidants and quality of oil on the oxidative and enzymatic properties of chicken broiler meat stored in an oxygen-enriched package (HiOx: 80% O2/20% CO2) in comparison with air-permeable polyvinylchloride (PVC) or skin (SK) packaging systems during retail display 2–4 °C for up to 14, 7, and 21 d, respectively. Broilers were fed a diet either with a low-oxidized oil (peroxide vale POV 23 meq O2/kg) or with a high-oxidized oil (POV 121 meq O2/kg), supplemented with an antioxidant pack (200 ppm EconomasE and organic minerals Se, Zn, Cu, Mn, and Fe as in Bioplex) in substitution for vitamin E and inorganic minerals for 42 d. In all packaging systems, lipid oxidation and protein oxidation were inhibited by up to 65% with an antioxidant-supplemented diet when compared to diets without antioxidant supplements. Antioxidant enzyme activities were significantly higher (P < 0.05) in the antioxidant-supplemented diets compared with control diets, regardless of oil quality. Meat samples from the antioxidant-supplemented group, irrespective of oil quality, has less purge and cooking loss compared to control diets. In all packaging systems, meat shear force was higher (P < 0.05) for broilers fed high-oxidized diets than the low-oxidized groups. Comparison between muscle types (breast as white vs. thigh as red) showed a similar trend in muscle susceptibility to oxidized oil in the diet but greater protection of antioxidant supplements for thigh meat in both physiochemical and textural properties. Dietary regimen influenced protein expression in broiler breast meat. Three protein spots from 2-dimensional gel electrophoresis, identified by mass spectrometry as glyceraldehyde 3-phosphate dehydrogenase, creatine kinase, and heat shock protein beta-1 were over-abundant in muscle from low-oxidized diets. The differential proteomes that suggested down regulation of the genes encoding antioxidative proteins upon feeding oxidized oil may be implicated in the broiler meat quality deterioration during storage. In summary, feeding diets with poor oil quality increased the vulnerability of lipids and proteins to oxidation in broiler breast and thigh meat during refrigerated and / or frozen storage in various packaging conditions, yet these effects were alleviated upon dietary supplementation with antioxidants.

Oxidative stress, caused by increased levels of reactive oxygen species (ROS)and reactive nitrogen species (RNS) and/or a decrease in antioxidant capacity, can result in the oxidation of various bio-molecules, such as proteins, lipids and deoxyribonucleic acid (DNA). These oxidized bio-molecules may contribute to pathologies such as cardiovascular diseases, neurodegenerative disorders and cancer. The Sympathetic Activity and Ambulatory Blood Pressure in Africans (SABPA) study was initiated in 2008 to investigate the coping styles and catecholamine metabolic markers of Africans, contributing to their higher sympathetic output and poorer psychosocial wellbeing. This study forms part of the SABPA study, but with a specific aim to investigated lipid and protein oxidation markers in hypertensive Africans versus their normotensive counterparts. Analytical methods for the quantification of specific lipid and protein oxidation markers were optimized and validated. Urine samples from 172 urbanized black South Africans were collected and 3-nitrotyrosine (3NT) and thiobarbituric acid reactive substances (TBARS) were quantified in these samples, using the optimized spectrophotometric and LC-MS/MS methods. Statistical analyses showed that in both males and females, TBARS and 3NTcorrelated with each other. In males, 3NT also correlated with physical activity level (PAL) and C-reactive protein (CRP), while TBARS also correlated with body mass index (BMI). In females 3NT correlated with BMI, while TBARS correlates with PAL. These correlations meant that they could influence the calculations of the true effect of 3NT and TBARS levels between normotensive and hypertensive subjects. After analyses of covariance (ANCOVA) analyses it was determined that the hypertensive male subjects had higher TBARS values than the normotensive male subjects did (p-value = 0.03) and the normotensive female subjects had higher 3NT levels compared to the hypertensive female subjects (p-value = 0.04). These results partially supported the hypothesis that that elevated concentrations of specific urinary lipid and protein oxidation markers will be observed in the hypertensive test subjects compared to their normotensive counterparts. The results also indicated that there were indeed a difference in lipid and protein oxidation between hypertensive and normotensive subject. / Thesis (MSc (Biochemistry))--North-West University, Potchefstroom Campus, 2013

Oxidative stress, caused by increased levels of reactive oxygen species (ROS)and reactive nitrogen species (RNS) and/or a decrease in antioxidant capacity, can result in the oxidation of various bio-molecules, such as proteins, lipids and deoxyribonucleic acid (DNA). These oxidized bio-molecules may contribute to pathologies such as cardiovascular diseases, neurodegenerative disorders and cancer. The Sympathetic Activity and Ambulatory Blood Pressure in Africans (SABPA) study was initiated in 2008 to investigate the coping styles and catecholamine metabolic markers of Africans, contributing to their higher sympathetic output and poorer psychosocial wellbeing. This study forms part of the SABPA study, but with a specific aim to investigated lipid and protein oxidation markers in hypertensive Africans versus their normotensive counterparts. Analytical methods for the quantification of specific lipid and protein oxidation markers were optimized and validated. Urine samples from 172 urbanized black South Africans were collected and 3-nitrotyrosine (3NT) and thiobarbituric acid reactive substances (TBARS) were quantified in these samples, using the optimized spectrophotometric and LC-MS/MS methods. Statistical analyses showed that in both males and females, TBARS and 3NTcorrelated with each other. In males, 3NT also correlated with physical activity level (PAL) and C-reactive protein (CRP), while TBARS also correlated with body mass index (BMI). In females 3NT correlated with BMI, while TBARS correlates with PAL. These correlations meant that they could influence the calculations of the true effect of 3NT and TBARS levels between normotensive and hypertensive subjects. After analyses of covariance (ANCOVA) analyses it was determined that the hypertensive male subjects had higher TBARS values than the normotensive male subjects did (p-value = 0.03) and the normotensive female subjects had higher 3NT levels compared to the hypertensive female subjects (p-value = 0.04). These results partially supported the hypothesis that that elevated concentrations of specific urinary lipid and protein oxidation markers will be observed in the hypertensive test subjects compared to their normotensive counterparts. The results also indicated that there were indeed a difference in lipid and protein oxidation between hypertensive and normotensive subject. / Thesis (MSc (Biochemistry))--North-West University, Potchefstroom Campus, 2013

Doctor of Philosophy(PhD) / Atherosclerosis manifests a state of increased oxidative stress characterized by comparable lipid and protein oxidation in the affected arterial wall. While oxidative modification of low density lipoprotein (LDL) has been extensively studied, increasing attention has been focused recently on oxidation of high-density lipoproteins (HDL) and its functional consequences in relation to atherosclerosis. Oxidative modification is thought to generate “dysfunctional” HDL that has lost anti-atherosclerotic activities, including the ability to remove cholesterol from lipid-laden cells. Therefore, there has been much interest in the detection of oxidized HDL. Unfortunately, available methods to detect oxidized HDL are limited at present, in part because oxidative modification of HDL is a complex process and ‘oxidized HDL’ is not a chemically defined entity. What is known however is that conversion of methionine (Met) residues of apolipoprotein (apo) A-I to methionine sulfoxide (MetO) is a process that occurs commonly as HDL undergoes oxidative modification. For example, human apoA-I+16 (containing MetO86 or MetO112) and apoA-I+32 (MetO86 plus MetO112) are generated when apoA-I reacts with lipid hydroperoxides formed as a consequence of the lipoprotein being exposed to 1e−oxidants. The formation of MetO in apoA−I induced by 2e−oxidants (i.e., hydrogen peroxide, hypochlorous acid or myeloperoxidase/hydrogen peroxide/chloride system) is associated with an impaired ability of the apolipoprotein to facilitate reactions relevant to reverse cholesterol transport. In addition, a previous study has suggested the plasma content of apoA-I+32 to be increased in certain subjects that have an increased risk to develop cardiovascular disease (CVD). Moreover, the MetO content in circulating, HDL−associated apoA−I is elevated in type 1 diabetes, a disorder commonly associated with increased oxidative stress and a risk factor for atherosclerosis. Therefore, in the present study, an existing HPLC method was applied to HDL samples from the Fletcher−Challenge study, a nested case control study, to test the potential usefulness of MetO-containing apoA-I as a marker of oxidative stress and/or CVD in a general population. Plasma samples whose HDL contained detectable apoA-I+16 and/or apoA-I+32 had significantly elevated levels of F2-isoprostanes, a marker of in vivo lipid oxidation, consistent with MetO-containing apoA-I being a useful marker of in vivo protein oxidation. Despite this however, there was no significant difference between controls and cases in their concentrations of HDL apoA-I+16 and apoA-I+32 or F2-isoprostanes, suggesting that markers of protein and lipid oxidation are not associated with the risk of coronary heart disease (CHD) in this general population. A limitation of the Fletcher−Challenge study was that only 22% of the 534 HDL samples analyzed contained apoA-I+16 and/or apoA-I+32. In addition, the HPLC−based method used is expensive and time−consuming and may lack the sensitivity needed for apolipoproteins to clinical studies. Thus, a mouse monoclonal anti-human apoA-I+32 antibody (MOA−1) was raised using HPLC−purified apoA-I+32 as immunogen. A sensitive ELISA was then developed using a commercial anti-human apoA-I monoclonal antibody as capture and biotinylated MOA−1 as detection antibody, respectively. The assay detected lipid−free HPLC−purified human apoA-I+32 in a concentration-dependent manner and with a significantly lower limit of detection (i.e., 3 ng/mL) than the HPLC method (1 μg/mL). The ELISA also detected lipid-free apoA-I modified by 2e-oxidants (hydrogen peroxide, hypochlorous acid, peroxynitrite), and HDL oxidized by 1e- or 2e-oxidants and present in buffer or human plasma. Moreover, the extent of recognition of MetO by MOA−1 increased with increasing numbers of MetO in apoA−I, as assessed by the experiments with H2O2−oxidized forms of apoA−I mutants, in which one, two or three Met residues were replaced with Leu. Their detection was concentration-dependent, reproducible, and exhibited a linear response over a physiologically plausible range of concentrations of oxidized HDL. In contrast, MOA-I failed to recognize native apoA-I, native apoA-II, apoA-I modified by hydroxyl radicals or metal ions, or LDL modified by 2e-oxidants. Furthermore, MOA−1 did not detect other Met−containing proteins oxidized by either hypochlorous acid or hydrogen peroxide. Taken together, the results showed that recognition of oxidized proteins by MOA−1 is limited to MetO contained in apoA−I. Finally, in a pilot study, plasma samples obtained from subjects with coronary artery disease (CAD) proven by angiography, and samples from CAD patients undergoing percutaneous coronary intervention (PCI) were analyzed by the ELISA. The preliminary data obtained showed elevated levels of MetO-containing apoA-I in plasma samples of CAD patients compared to those of corresponding control subjects. Unexpectedly, levels of MetOcontaining apoA-I decreased PCI compared to before PCI. A possible explanation for these results is that HDL−associated apoA−I become displaced by acute phase proteins, such as serum amyloid A, in response to PCI. In summary, the ELISA developed here specifically detects apoA-I containing MetO in HDL and human plasma. As such it may provide a useful tool for investigating the relationship between oxidized HDL and CAD.