Dietary omega-3 polyunsaturated fatty acids induce plasminogen activator activity and DNA damage in rabbit spermatozoa.

The aim of this study was to determine the effect(s) of dietary omega-3 polyunsaturated fatty acids (ω-3 PUFA) on rabbit semen. Adult rabbit bucks were assigned to two groups that were given two diets, a standard diet (control) and a diet supplemented with ω-3 PUFA. Sperm samples were collected from all bucks with the use of an artificial vagina in 20-day intervals, for a total period of 120 days. The enrichment of membranes in ω-3 PUFA was manifested by the elevation of the 22:5 ω-3 (docosapentaenoic acid [DPA]) levels within 40 days. This increase in DPA content did not affect semen characteristics (i.e., concentration, motility and viability). However, it was associated with the induction of lipid peroxidation in spermatozoa, as determined on the basis of the malondialdehyde content. Lipid peroxidation was associated with DNA fragmentation in ω-3 PUFA-enriched spermatozoa and a concomitant increase in plasminogen activator (PA) activity. The effects of ω-3 PUFA on sperm cells were evident within 40 days of ω-3 PUFA dietary intake and exhibited peack values on day 120. Our findings suggest that an ω-3 PUFA-rich diet may not affect semen characteristics; however, it may have a negative impact on the oxidative status and DNA integrity of the spermatozoa, which was associated with an induction of PAs activity.

Effect of supplementation of the laying hen diet with olive leaves (Olea europea L.) on lipid oxidation and fatty acid profile of α-linolenic acid enriched eggs during storage.

1. The aim of this study was to evaluate the effect of supplementation of the layer diet with olive leaves (Olea europea L.) on lipid oxidation and fatty acid profile of α-linolenic acid enriched eggs during refrigerated storage, and to compare this effect with α-tocopheryl acetate supplementation. 2. A total of 72 brown Lohmann laying hens, equally allocated to 3 groups, were fed on diets supplemented with 40 g/kg linseed oil, or linseed oil and olive leaves at 10 g/kg or linseed oil and α-tocopheryl acetate at 200 mg/kg. Collected eggs were analysed for fatty acid profile and lipid oxidation either fresh or following 60 d storage at 4°C. 3. Results showed that olive leaves or α-tocopheryl acetate supplementation reduced lipid hydroperoxide concentration in fresh eggs but had no effect on their fatty acid profile and malondialdehyde (MDA) content compared to controls. 4. Refrigerated storage for 60 d decreased the proportions of PUFAs but increased those of MUFAs in eggs from the control diet, whilst it had no effect on the fatty acid composition of eggs from the diets supplemented with olive leaves or α-tocopheryl acetate, which in turn showed decreased concentrations of lipid hydroperoxides and MDA.

Lipid oxidation of stored eggs enriched with very long chain n-3 fatty acids, as affected by dietary olive leaves (Olea europea L.) or α-tocopheryl acetate supplementation.

The antioxidant potential of dietary olive leaves or α-tocopheryl acetate supplementation on lipid oxidation of refrigerated stored hen eggs enriched with very long-chain n-3 fatty acids, was investigated. Ninety-six brown Lohmann laying hens, were equally assigned into three groups. Hens within the control group were given a typical diet containing 3% fish oil, whereas other groups were given the same diet further supplemented with 10 g ground olive leaves/kg feed or 200mg α-tocopheryl acetate/kg feed. Results showed that α-tocopheryl acetate or olive leaves supplementation had no significant effect on the fatty acid composition and malondialdehyde (MDA) levels of fresh eggs but reduced their lipid hydroperoxide levels compared to controls. Storage for 60 d decreased the proportions of polyunsaturated fatty acids (PUFAs) but increased those of monounsaturated fatty acids (MUFAs) in eggs from the control group, while had no effect on the fatty acid composition of the eggs from the other two groups, which showed decreased levels of lipid hydroperoxides and MDA. Therefore, the very long chain n-3 PUFAs in eggs were protected from undergoing deterioration partly by olive leaves supplementation and totally by α-tocopheryl acetate supplementation. In addition, incorporating tocopherols into eggs might also provide a source of tocopherols for the human diet.

Oxidative stability and microbial growth of turkey breast fillets during refrigerated storage as influenced by feed supplementation with olive leaves, oregano and/or α -tocopheryl acetate.

1. The aim of this study was to evaluate the inhibitory potential of feed supplementation with olive leaves, oregano and/or α-tocopheryl acetate on microbial growth and lipid oxidation of turkey breast fillets during refrigerated storage. 2. A total of 40 turkeys, allocated to 5 groups of 8 birds each, were fed on diets supplemented with olive leaves at 10 g/kg, oregano at 10 g/kg or α-tocopheryl acetate at 150 or 300 mg/kg. Following slaughter, fillets from breast were stored at 4°C in the dark for 12 d, and lipid oxidation and microbial growth were assessed. 3. Results showed that dietary olive leaves were more effective than oregano at inhibiting lipid oxidation, but were inferior to dietary supplementation of 300 mg α-tocopheryl acetate/kg. In turn, α-tocopheryl acetate supplementation at 150 mg/kg was effective at inhibiting lipid oxidation compared to the control but inferior to oregano supplementation. 4. Total viable counts, lactic acid bacteria, Enterobacteriaceae and psychrotrophic bacteria counts were all increased in breast fillets of all groups throughout the refrigerated storage. Diet supplementation with α-tocopheryl acetate had no effect on the bacterial counts recorded in the control group, but diet supplementation with olive leaves or oregano resulted in a decrease of all bacterial counts at d 2 of storage and thereafter; during this period, oregano was more effective at inhibiting bacterial growth compared with olive leaves. 5. Therefore, if shown clinically to be safe and having beneficial effects in vivo, olive leaves and oregano might be utilised in novel applications as nutritional supplements or functional food components.

Potential of long-term dietary administration of rosemary in improving the antioxidant status of rat tissues following carbon tetrachloride intoxication.

In this study, 24 Wistar rats were allocated to 4 groups of 6 animals each. Groups 1 and 2 were fed a basal diet, while groups 3 and 4 were fed the basal diet supplemented further with ground rosemary at 1% level. Following 6-weeks feeding, groups 2 and 4 were injected 1 ml CCl(4)/kg bw and after six hours all animals were sacrificed. Results showed that feeding rosemary before CCl(4) treatment resulted in decline (P<0.05) of the increased aspartate transaminase, alanine transaminase and alkaline phosphatase activities and increase (P<0.05) of the reduced cholesterol and triacylglycerols in serum. It also decreased (P<0.05) lipid peroxidation and increased (P<0.05) the reduced hydroxyl anion radical and hydrogen peroxide scavenging activities in serum, liver, kidney and heart tissues. In addition, it increased (P<0.05) the reduced ABTS radical cation and the superoxide anion scavenging activities in all tissues except in heart and in kidney and heart tissues, respectively. These results suggest that dietary rosemary has the potential to become a promising functional food component.

Effect of long-term dietary administration of oregano on the alleviation of carbon tetrachloride-induced oxidative stress in rats.

This study aimed at evaluating the protective effect of long-term dietary oregano on the alleviation of carbon tetrachloride-induced oxidative stress in rats. Twenty-four female Wistar rats were allocated to four groups of six animals each. Groups 1 (control) and 2 (CCl 4) were fed a basal diet, while groups 3 (oregano) and 4 (oregano + CCl 4) were fed the basal diet supplemented further with ground oregano at 1% level. Following six-week feeding, the rats of groups 2 and 4 were given a single intraperitoneal injection of CCl 4 at a dose of 1 mL/kg bw. Six hours after the CCl 4 injection, all animals were sacrificed, and serum, liver, kidney, and heart tissue samples were collected. Analysis results showed that the addition of oregano significantly increased the total phenolic content and the Trolox equivalent antioxidant capacity of the basal diet but had no effect on its lipid peroxidation index. Treatment with CCl 4 of rats from the CCl 4 group caused a significant increase in aspartate transaminase (AST), alanine transaminase (ALT), and alkaline phosphatase (ALP) in serum, whereas it decreased cholesterol and triglyceride content as compared to the control. It also increased the lipid peroxidation index and decreased the scavenging activities of the 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid diammonium salt (ABTS) radical cation, the hydroxyl anion radical, the superoxide anion radical, and the hydrogen peroxide in all tested tissues, as compared to that of the control. Without CCl 4 treatment, diet supplementation with oregano had no effect on these biochemical parameters, excluding the hydroxyl radical scavenging activity, which was increased in all tested tissues as compared to that of the control. Feeding oregano before CCl 4 treatment resulted in a significant decline of the increase in AST, ALT, and ALP activities ( P < 0.05 vs CCl 4 group), but the recorded values could not attain those of the control group ( P < 0.05 vs control group). It significantly increased the reduced cholesterol and triglycerides ( P < 0.05 vs CCl 4 group) to values not differing from those of the control. It also resulted in a significant reduction of the increased malondialdehyde ( P < 0.05 vs CCl 4 group) to values that could not attain the levels of the control but had no significant effect ( P > 0.05) on the reduced ABTS radical cation scavenging activity. It increased significantly the reduced hydroxyl anion radical scavenging activity ( P < 0.05 vs CCl 4 group) to values that could not attain those of the control in all tested tissues except kidney. Additionally, it resulted in a significant elevation of the decreased superoxide anion radical scavenging activity in serum and liver but had no effect in kidney and heart, whereas it also resulted in a significant elevation of the decreased hydrogen peroxide scavenging activity in liver, kidney, and heart but had no effect in serum. These results suggest that dietary oregano may effectively improve the impaired antioxidant status in CCl 4-induced toxicity in rats.

The antimicrobial effect of thyme essential oil, nisin and their combination against Escherichia coli O157:H7 in minced beef during refrigerated storage.

The antimicrobial effect of thyme essential oil (EO) at supplementation levels of 0.3%, 0.6% or 0.9%, nisin at 500 or 1000IU/g, and their combination, on Escherichia coli O157:H7 was examined in both tryptic soy broth (TSB) and minced beef meat. EO at 0.3% possessed a weak antibacterial activity against the pathogen in TSB, whereas at 0.9% showed unacceptable organoleptic properties in minced meat. Thus, only the level of 0.6% of EO was further examined against the pathogens in minced meat. Treatment of minced beef meat with EO at 0.6% showed an inhibitory activity against E. coli O157:H7 during storage at 10°C, but not at 4°C. Treatment of minced beef meat or TSB with nisin at 500 or 1000IU/g did not show any antibacterial activity against E. coli O157:H7. The combination of EO at 0.6% and nisin at 500 or 1000IU/g showed an additive effect against the pathogen, which was higher during storage at 10°C than at 4°C.

The antimicrobial effect of thyme essential oil, nisin, and their combination against Listeria monocytogenes in minced beef during refrigerated storage.

The antimicrobial effect of thyme essential oil (EO) at 0.3%, 0.6%, or 0.9%, nisin at 500 or 1000IU/g, and their combination against Listeria monocytogenes was examined in both tryptic soy broth (TSB) and minced beef meat. Thyme EO at 0.3% possessed a weak antibacterial activity against the pathogen in TSB, whereas at 0.9% showed unacceptable organoleptic properties in minced meat. Thus, only the level of 0.6% of EO was further examined against the pathogen in minced meat. Treatment of minced beef meat with nisin at 500 or 1000IU/g showed antibacterial activity against L. monocytogenes, which was dependent on the concentration level of nisin and the strains used. Treatment of minced beef meat with EO at 0.6% showed stronger inhibitory activity against L. monocytogenes than treatment with nisin at 500 or 1000IU/g. All treatments showed stronger inhibitory activity against the pathogens at 10 degrees C than at 4 degrees C. The combined addition of EO at 0.6% and nisin at 500 or 1000IU/g showed a synergistic activity against the pathogen. Most efficient among treatments was the combination of EO at 0.6% with nisin at 1000IU/g, which decreased the population of L. monocytogenes below the official limit of the European Union recently set at 2logcfu/g, during storage at 4 degrees C.

The incorporation of dehydrated rosemary leaves in the rations of turkeys and their impact on the oxidative stability of the produced raw and cooked meat.

Thirty-six 12-week-old turkeys were distributed into six groups and were raised for 4 weeks on rations containing 0%, 0.5% or 1.0% dehydrated rosemary leaves as antioxidant in the presence of alpha-tocopheryl acetate from 10 to 300 mg/kg. Following slaughtering, breast and thigh meat samples, raw or cooked, from all six groups were collected to be refrigerated at 4 degrees C for 9 days. All stored samples were submitted to analysis for their concentration in malondialdehyde (MDA), a lipid oxidation marker, and alpha-tocopherol. The results showed that the rations containing 300 mg/kg alpha-tocopheryl acetate increased the mean alpha-tocopherol content of the breast and thigh significantly (P <0.05) compared with the respective control values. No significant (P>0.05) changes could be observed in the alpha-tocopherol content of breast and thigh of turkeys consuming rations containing up to 1% dehydrated rosemary leaves. The refrigeration of the meats led to spontaneous increase in the MDA content of the breast and thigh meat samples. Samples from turkeys fed rations containing 300 mg/kg alpha-tocopheryl acetate showed the lowest mean levels of MDA after the 9-day refrigerated period. The incorporation of rosemary in the rations led to a modest decrease in the formation of MDA in the meats compared with the respective mean control values. The combination of alpha-tocopheryl acetate and rosemary was not associated with an additional decrease in MDA formation.

Effect of dietary saffron (Crocus sativus L.) on the oxidative stability of egg yolk.

1. The effects of dietary inclusion of red stigmas of Greek saffron (Crocus sativus L.) on the oxidative stability of shell eggs and liquid yolks were investigated and compared with those of dietary a-tocopherol. 2. Ninety-six Lohmann laying hens, 38 weeks old, distributed into 4 groups with 4 replicates each, were given either a control diet, diets enriched with 10 (SAF10) or 20 (SAF20) mg/kg saffron, or a diet enriched with 200 mg/kg a-tocopheryl acetate (VE200). 3. Following 6 weeks of feeding, eggs were collected and the rate of lipid oxidation was determined in refrigerated stored shell eggs, as well as in yolks adjusted to a pH of 6.2 or 4.2 and stored in the presence of light. 4. The results showed that the extent of lipid oxidation in shell eggs, as measured by malondialdehyde (MDA) formation, differed between dietary treatments, but did not change with storage time. In stored shell eggs, MDA levels differed between dietary treatments at all time points. 5. Yolks from the control group adjusted to pH 6.2 gave MDA values higher than those of the SAF10 group, which in turn were higher than those of the SAF20 group, a finding suggesting that saffron exerted a dose-dependent antioxidative activity. The VE200 group gave lower MDA values than the other groups at all time points. The oxidation profile of yolks at pH 4.2 showed a similar pattern but the rate of oxidation was greater.

Performance of rabbits and oxidative stability of muscle tissues as affected by dietary supplementation with oregano essential oil.

The effect of dietary supplementation with oregano essential oil on the performance of rabbits, and the susceptibility of the produced raw and thermally treated muscle tissue to lipid oxidation during refrigerated storage, were investigated. A total of 96 weaned rabbits were separated into four equal groups with three subgroups each. One group was given the basal diet and served as control, two groups were administered diets supplemented with oregano essential oil at levels of 100 and 200 mg/kg diet, whereas the remaining group was given a diet supplemented with alpha-tocopheryl acetate at 200 mg/kg. During the 42-day experimental period, body weight and feed intake were recorded weekly and the feed conversion ratio was calculated. Feeding the experimental diets to rabbits, performance parameters were not affected. Therefore, dietary oregano essential oil exerted no growth-promoting effect on rabbits. With increased supplementation of oregano essential oil, malondialdehyde values decreased in both raw and thermally treated muscles during refrigerated storage. This finding suggests that dietary oregano essential oil exerted a significant antioxidant effect. Dietary supplementation of oregano essential oil at the level of 200 mg/kg was more effective in delaying lipid oxidation compared with the level of 100 mg/kg, but inferior to dietary supplementation of 200 mg alpha-tocopheryl acetate per kg. This study indirectly provides evidence that antioxidant compounds occurring in oregano essential oil were absorbed by the rabbit and increased the antioxidative capacity of tissues.

Dietary versus post-mortem use of oregano oil and/or alpha-tocopherol in turkeys to inhibit development of lipid oxidation in meat during refrigerated storage.

The dietary and post-mortem uses of oregano oil in turkeys to inhibit development of lipid oxidation in breast and thigh meat during refrigerated storage were investigated. Using minced meat, patties were prepared from turkey meat post-mortem added with either 200 mg oregano oil or alpha-tocopherol/kg, meat from turkeys dietary supplemented with either 200 mg oregano oil or alpha-tocopheryl acetate/kg feed, and control meat. All patties were cooked, placed in a refrigerated cabinet at 4 degrees C, and lipid oxidation was assessed by monitoring malondialdehyde formation after 3, 6 and 9 days of storage. Treatments significantly (P<0.05) retarded lipid oxidation in both breast and thigh meat patties at all storage times compared with controls. The dietary supplementation of either oregano oil or alpha-tocopheryl acetate exhibited the highest antioxidative activity compared with the other treatments. Post-mortem addition of either oregano oil or alpha-tocopherol to the minced meat also retarded lipid oxidation in the prepared patties compared with controls; however, this effect was inferior to that of the dietary supplementation even though the post-mortem alpha-tocopherol supplemented meat contained 90-fold more alpha-tocopherol than patties from the dietary supplemented meat. Thigh meat was more susceptible to oxidation than breast meat, although the former contained alpha-tocopherol at markedly higher levels. Supplementing the diet with 200 mg oregano oil/kg, alpha-tocopherol levels in the breast and thigh meat significantly (P<0.05) increased compared with control. This increase could not be attributed to the alpha-tocopherol already present in the oregano oil since post-mortem addition of oregano oil to control breast and thigh meat at the same dose could not actually increase the alpha-tocopherol concentrations.

Blockade of Kupffer cells by gadolinium chloride reduces lipid peroxidation and protects liver from ischemia/reperfusion injury.

BACKGROUND/AIMS: The implication of lipid peroxidation in the inhibitory effect of GdCl3 (gadolinium chloride) on Kupffer cells activation has not been extensively investigated. The aim of this study was to examine the effect of GdCl3 inhibition of Kupffer cells activation on lipid peroxidation after severe total hepatic ischemia/reperfusion. METHODOLOGY: Male Wistar rats (n = 40) were randomly divided into a sham-operation group, a control ischemia/reperfusion group, and two ischemia/reperfusion groups pretreated with GdCl3 (10 mg and 20 mg/kg bw intravenously, 48 and 24 h prior to operation). Following 60 min of total hepatic ischemia and 120 min of reperfusion, the rats were sacrificed, and liver samples were taken for determination of malondialdehyde and light microscopy examination. Blood samples were also taken for assay of aspartate and alanine transaminase. Additional animals (n = 60) were followed up for a 7-day survival rate determination. RESULTS: Ischemia/reperfusion decreased the survival rate to 13.3%, increased (p < 0.001) the levels of aspartate and alanine transaminase in serum to 2387 +/- 75 and 2157 +/- 87 IU/L, respectively, and increased (p < 0.001) malondialdehyde levels in liver to 1.609 +/- 0.096 nmoles/g compared with 1.164 +/- 0.060 in the sham operation group. Pretreatment with GdCl3 increased the survival rate to 60%, and decreased (p < 0.001) the levels of aspartate transaminase in serum to 1549 +/- 66 and 1496 +/- 55 IU/L, the levels of alanine transaminase in serum to 1302 +/- 48 and 1305 +/- 63 IU/L, and the levels of malondialdehyde in liver to 1.132 +/- 0.034 and 1.149 +/- 0.57 nmoles/g for the lower and the higher doses of GdCl3, respectively. Histological examination showed protection of liver parenchyma in the animals treated with GdCl3. CONCLUSIONS: Experimental data suggest that GdCl3 inhibition of Kupffer cells activation protects liver from ischemia/reperfusion injury by a mechanism that reduces lipid peroxidation.

Effect of dietary oregano oil and alpha-tocopheryl acetate supplementation on iron-induced lipid oxidation of turkey breast, thigh, liver and heart tissues.

Twenty-five 12-week-old turkeys randomly divided into five groups were given a basal diet, or a basal diet supplemented with 200 mg alpha-tocopheryl acetate/kg, or 100 mg oregano oil/kg or 200 mg oregano oil/kg, or 100 mg oregano oil plus 100 mg alpha-tocopheryl acetate/kg diet, for 4 weeks prior to slaughter. Breast, thigh, liver and heart tissues were subjected to iron-induced lipid oxidation, the extent of which was determined by third-order derivative spectrophotometry. Results showed that dietary oregano oil at the inclusion level of 200 mg oregano oil/kg diet was more effective in delaying lipid oxidation compared with the inclusion level of 100 mg/kg, but equivalent to the inclusion of 200 mg alpha-tocopheryl acetate/kg diet, which in turn was inferior to the combined inclusion of 100 mg oregano oil plus 100 mg alpha-tocopheryl acetate/kg, which was superior to all dietary treatments. Thigh tissue was more susceptible to oxidation than breast tissue, although it contained alpha-tocopherol at higher concentrations. Also, lipid oxidation in heart was relatively high, although it contained the highest alpha-tocopherol levels. This indicates that tissue alpha-tocopherol is one important factor influencing the level of lipid oxidation, but the distribution of lipids, iron and oregano oil in tissues must also be taken into consideration. Tissue alpha-tocopherol levels responded to dietary intake of 30-200 mg alpha-tocopheryl acetate/kg in the order heart > liver > thigh > breast. Breast, thigh and heart tissues from the oregano groups presented significantly (p < 0.05) higher levels of alpha-tocopherol compared with the control, the increase being positively correlated with the supplementation level. The increased levels of alpha-tocopherol in these tissues indicated that the dietary oregano oil exerted a protective action on alpha-tocopherol.

Effect of dietary supplementation with oregano essential oil on performance of broilers after experimental infection with Eimeria tenella.

A study was carried out to examine the effect of dietary supplementation of oregano essential oil on performance of broiler chickens experimentally infected with Eimeria tenella at 14 days of age. A total of 120 day-old Cobb-500 chicks separated into 4 equal groups with three replicates each, were used in this study. Two groups, one infected with 5 x 10(4) sporulated oocysts of E. tenella and the other not, were given a basal diet and served as controls. The other two groups also infected with E. tenella were administered diets supplemented with oregano essential oil at a level of 300 mg/kg, or with the anticoccidial lasalocid at 75 mg/kg. Following this infection, survival rate, bloody diarrhoea and oocysts excretion as well as lesion score were determined. Throughout the experimental period of 42 days, body weight gain and feed intake were recorded weekly, and feed conversion ratios were calculated. Two weeks after the infection with E. tenella supplementation with dietary oregano oil resulted in body weight gains and feed conversion ratios not differing from the non-infected group, but higher than those of the infected control group and lower than those of the lasalocid group. These parameters correspond with the extent of bloody diarrhoea, survival rate, lesion score and oocyst numbers and indicated that oregano essential oil exerted an anticoccidial effect against E. tenella, which was, however, lower than that exhibited by lasalocid.

The effects of dietary oregano essential oil and α-tocopheryl acetate on lipid oxidation in raw and cooked turkey during refrigerated storage.

The effects of dietary oregano essential oil and α-tocopheryl acetate supplementation on the susceptibility of raw and cooked turkey breast and thigh meat to lipid oxidation during refrigerated storage for 9 days were examined. Thirty 12-week-old turkeys were divided into five groups and fed a basal diet containing 30 mg α-tocopheryl acetate kg(-1) feed as control, or basal diet plus 200 mg α-tocopheryl acetate kg(-1), or basal diet plus 100 mg oregano oil kg(-1), or basal diet plus 200 mg oregano oil kg(-1), or basal diet plus 100 mg oregano oil and 100 mg α-tocopheryl acetate kg(-1), for 4 weeks prior to slaughter. Lipid oxidation was assessed by monitoring malondialdehyde formation in raw and cooked meat at 0, 3, 6 and 9 days of refrigerated storage, through use of a third-order derivative spectrophotometric method. Results showed that all dietary treatments significantly (P<0.05) increased the stability of both raw and cooked turkey meat to lipid oxidation compared with the control. Oregano oil at 200 mg kg(-1) was significantly (P<0.05) more effective in delaying lipid oxidation compared to the level of 100 mg kg(-1), equivalent to α-tocopheryl acetate at 200 mg kg(-1), but inferior (P<0.05) to oregano oil plus α-tocopheryl acetate at 100 mg kg(-1) each, which in turn was superior (P<0.05) to all dietary treatments, indicating a synergistic effect. Thigh muscle was more susceptible to oxidation compared with breast muscle in all treatments, although it contained α-tocopherol at significantly (P<0.05) higher levels.

Distribution and stability of aflatoxin M1 during production and storage of yoghurt.

Yoghurt from cow's milk artificially contaminated with aflatoxin M1 (AFM1) at levels of 0.050 and 0.100 g l(-1) was fermented to reach pHs 4.0 and 4.6. Yoghurt fermented to pH 4.6 was also used for preparing strained yoghurt. Yoghurts were stored at 4 degrees C for up to 4 weeks. Analysis of AFM1 in milk, yoghurt, strained yoghurt and yoghurt whey was carried out using immunoaffinity column extraction and liquid chromatography coupled with fluorometric detection. AFM1 levels in yoghurt samples showed a significant decrease (p < 0.01) compared with those initially added to milk. Growth of culture lactic acid bacteria was not affected in the AFM1 contaminated yoghurts, with the exception of Streptococcus thermophilus that showed a significantly (p < 0.01) lower increase in the yoghurt containing the toxin at high concentration. Following fermentation, AFM1 was significantly lower (p < 0.01) in yoghurts with pH 4.0 than in yoghurts with pH 4.6 at both contamination levels. During refrigerated storage, AFM1 was rather more stable in yoghurts with pH 4.6 than with pH 4.0. The percentage loss of the initial amount of AFM1 in milk was estimated at about 13 and 22% by the end of the fermentation, and 16 and 34% by the end of storage for yoghurts with pHs 4.6 and 4.0, respectively. The percentage distribution ratio of AFM1 in strained yoghurt/yoghurt whey of the initial toxin present in the yoghurt was about 90/10 and 87/13 for the lower and the higher contamination levels, respectively.

Occurrence of aflatoxin M(1) in raw and market milk commercialized in Greece.

From December 1999 to May 2000, 114 samples of pasteurized, ultrahigh temperature-treated (UHT) and concentrated milk were collected in supermarkets, whereas 52 raw milk samples from cow, sheep and goat were obtained from different milk producers all over Greece. Sample collection was repeated from December 2000 to May 2001 and concerned 54 samples of pasteurized milk, 23 samples of bulk-tank raw milk and 55 raw milk samples from cow, sheep and goat. The total number of samples analysed for aflatoxin M(1) (AFM(1)) contamination by immunoaffinity column extraction and liquid chromatography was 297. In the first sampling, the incidence rates of AFM(1) contamination in pasteurized, UHT, concentrated and cow, sheep and goat raw milk were 85.4, 82.3, 93.3, 73.3, 66.7 and 40%, respectively, with only one cow raw milk and two concentrated milk samples exceeding the EU limit of 50 ng l(-1). In the second sampling, the incidence rates of AFM(1) contamination in pasteurized, bulk-tank and cow, sheep and goat raw milk were 79.6, 78.3, 64.3, 73.3 and 66.7%, respectively, with only one cow and one sheep raw milk samples exceeding the limit of 50 ng l(-1). The results suggest that the current regulatory status in Greece is effective.

Effect of dietary oregano essential oil on performance of chickens and on iron-induced lipid oxidation of breast, thigh and abdominal fat tissues.

1. We studied the effect of dietary oregano essential oil (50 and 100 mg/kg of feed) on the performance of broilers, and determined the susceptibility of the resulting broiler meat to iron-induced lipid oxidation. 2. Performance of the birds was unaffected by the experimental diets. Therefore, dietary oregano oil exerted no growth-promoting effect on broilers. 3. Iron-induced lipid oxidation showed that as oregano oil increased in the diet, malondialdehyde values decreased in tissue samples, suggesting that the oil, particularly at 100 mg/kg of feed, exerted an antioxidant effect on chicken tissues. 4. Dietary alpha-tocopheryl acetate supplementation at 200 mg/kg of feed displayed greater antioxidant activity than oregano oil at either supplementation rate. 5. Thigh muscle was more susceptible to oxidation than breast muscle, although the former contained alpha-tocopherol at higher concentration. Muscle alpha-tocopherol is an important factor influencing lipid oxidation, but the influence of polyunsaturated fatty acids and content of pro-oxidants must be taken into consideration too.

New simple liquid chromatographic method for the determination of trimethoprim, sulfadiazine and N4-acetylsulfadiazine in plasma of broilers.

A new method for simultaneous quantification of trimethoprim, sulfadiazine and N4-acetylsulfadiazine in plasma of broilers at levels down to 13-16 ng/ml has been developed. Samples were deproteinized with acetonitrile, defatted with hexane, and extracted with dichloromethane. Chromatographic analysis was carried out on a C18 column in the presence of tetrabutylammonium hydrogen sulfate, a competing base, while detection was performed at 240 nm for trimethoprim, and 270 nm for both sulfadiazine and N4-acetylsulfadiazine. Accuracy and precision data showed recoveries and relative standard deviation values better than 87.3% and 3.1%, respectively, for all three analytes. The good analytical characteristics of the method could allow limits of detection in the low ng/ml range to be realised. The method was successfully applied to determine drug concentrations in plasma samples from broilers administered a combination of sulfadiazine and trimethoprim.