Sand intake by laying hens and its effect on egg production parameters.

Soil intake may be the most prominent source of environmental contaminants for free range and organic hens, but there are no quantitative data concerning soil intake by domestic hens. Consumption of soil of 14-32 g a day can be estimated from literature, but such a dilution of nutrient intake seems incompatible with high productivity. In this study laying hens were fed pelleted diets with 0%, 10%, 20%, 25% and 30% of sand addition to determine its effect on productivity. Feed intake, feed and nutrient (feed minus sand) conversion ratio, egg production, egg weight and body weight gain were measured over a 4-week period. Acid insoluble ash concentration in the faeces was measured to determine the accuracy of estimating the soil ingestion by the soil-ingestion equation for wildlife as a way to determine soil ingestion of free range and organic hens under practical circumstances. The hens were able to compensate the dilution of the diet with 20%, 25% and 30% of sand by increasing their feed intake. Feed intake increased significantly and feed to egg conversion ratio decreased significantly with increasing sand levels in the diet. The nutrient to egg conversion ratio of the diet without sand tended to be worse than for the diets with sand, presumably due to the total absence of coarse material in the diet. There were no differences in egg production and egg weight between hens fed the different diets but body weight gain was significantly lower for the hens fed the diets with 20%, 25% and 30% of sand. Estimation of sand ingestion was done by the soil-ingestion equation for wildlife. Provided that the actual dry matter digestibility coefficient of the nutrient part of the diet is taken into account, estimating the soil ingestion according to the soil-ingestion equation for wildlife seems an appropriate way to determine soil ingestion for free range and organic hens under practical circumstances.

Residues of dioxins (PCDD/Fs) and PCBs in eggs, fat and livers of laying hens following consumption of contaminated feed.

Laying hens were fed with feed from the Belgian dioxin incident diluted ten-fold with non-contaminated feed, resulting in concentrations of 61 ngTEQkg(-1) PCDD/Fs, 23 ngTEQkg(-1) non-ortho PCBs, 116 ngTEQkg(-1) mono-ortho PCBs and 3.2 mgkg(-1) of the seven indicator PCBs. Following exposure for seven days, feed was replaced by non-contaminated feed for a period up to six weeks. Concentrations of PCDD/Fs in eggs showed a maximum of 214 pgTEQg(-1) fat after nine days and decreased to 44 pgTEQg(-1) after seven weeks. Dioxin concentrations in abdominal fat of chickens killed just after the last treatment, or after 1, 3 or 6 weeks on clean feed were 69, 84, 54 and 41 pgTEQg(-1) fat, respectively. Concentrations in livers decreased more rapidly, being 35, 7, 4 and 3 pgTEQg(-1) tissue, respectively. In both eggs and tissues, total TEQ concentrations were 3-4 times higher. Concentrations of the seven indicator PCBs in egg fat showed a stronger decrease with concentrations of 16.5 microgg(-1) at the peak (day 9) and 2.2 microgg(-1) after seven weeks. Corresponding concentrations in abdominal fat were, respectively, 4.6 and 2.6 microgg(-1) fat, and in livers 0.77 and 0.14 microgg(-1) tissue. The ratio of indicator PCBs to PCDD/Fs in feed was 52200. In eggs this ratio was initially higher (85000), but decreased towards 50000 after six weeks on clean feed. In abdominal fat the ratio varied between 49000 and 67000. In livers, the ratio was initially low (22000) but increased to 45000 towards the end of the study. It is concluded that the behaviour of PCDD/Fs and PCBs in laying hens is comparable and that the use of indicator PCBs appears to be a good alternative for PCDD/Fs but only in the case of co-exposure to both PCBs and PCDD/Fs, such as in incidents with PCB oil.

A toxicokinetic model for the carry-over of dioxins and PCBs from feed and soil to eggs.

A mathematical model for the kinetics of carry-over of dioxins and dioxin-like PCBs from feed mixed with contaminated oil to eggs has been developed. This model incorporates uptake of the compounds over the gut wall and their subsequent transport by blood, distribution over the body, hepatic metabolism and excretion through egg yolk fat. The model is analysed with respect to the possibility of identifying as yet unknown model parameters by fitting these to the experimental data. The model was fitted to the experimental data on the carry-over from feed to eggs. The calibrated model was applied to calculate the steady-state concentrations in eggs which were compared to European Maximum Residue Levels for dioxins in feed and eggs, showing that these limits do not match. The feed limit of 0.75 ng TEQ/kg should be reduced to about 0.17 ng TEQ/kg in order to guarantee egg levels below the residue limit of 3 pg TEQ/g fat. Experimental results of carry-over from contaminated soil were used to estimate the absorption of dioxin-like compounds from soils as compared to the absorption from feed, resulting in a value around 40 to 60% absorption from soil as compared to around 90% absorption from feed.

Carry-over of dioxins and PCBs from feed and soil to eggs at low contamination levels-- influence of mycotoxin binders on the carry-over from feed to eggs.

Laying hens were fed with compound feed containing six different levels of dioxins, dioxin-like PCBs and indicator PCBs for a period of 56 days. This was followed by a period of 56 days on clean feed. Dioxin levels in feed varied from background levels to three times the current EU tolerance limit of 0.75 ng TEQ/kg. At all dose levels a rapid increase was observed in the dioxin levels in eggs. There was a clear linear dose-response relationship between the dioxin levels in eggs and feed. The feed containing 0.4 ng TEQ dioxins per kg resulted in egg levels just above the EU limit of 3 pg TEQ/g fat. Dioxin-like and indicator PCB residues followed a pattern very similar to that of dioxins. Exposure to the highest indicator PCB level of 32 microg/kg resulted in egg levels around 300 ng/g fat. Exposure to dioxins through contaminated soil, mixed at 10% into the feed, resulted in a similar carry-over as from feed. Mycotoxin binders, mixed at 0.5% into the feed, had little effect on the carry-over of dioxins from the feed to the egg. It can be concluded that consumption of feed or soil with even moderate levels of dioxins and dioxin-like PCBs rapidly results in increased levels in eggs. The current EU dioxin limit for feed cannot guarantee egg dioxin levels below the EU-limit.

Evaluation of broiler performance when fed insect-protected, control, or commercial varieties of dehulled soybean meal.

We evaluated the nutritional value of broiler diets containing approximately 35% soybean meal from insect-protected soybean containing CrylAc protein, or from a similar nontransgenic control, or from 7 reference commercial soybean varieties. The feeding trial lasted 41 d, and each treatment consisted of 10 replicates of 1-d-old Ross 508 broilers (5 pens males and 5 pens females). Each pen contained 12 birds, and at d 13, birds were randomly removed until 9 birds remained. Body weight and feed intake were measured on pen basis at 41 d. At d 42, 4 broilers per pen were slaughtered. The carcasses were dissected, and cut-up yields were determined. Dry matter, protein, and fat contents of breast meat as well as shear force of breast meat were determined. The data were analyzed by ANOVA. The BW and feed conversion ratio at d 41 averaged 2,435 g and 1.52, respectively. There were no significant treatment x sex interactions. Data for final BW, feed conversion, carcass yield, and breast meat data were not different (P < 0.05) between broilers fed insect-protected and those fed commercial soybean meal varieties. Thus, insect-protected soybean meal was nutritionally equivalent to nongenetically modified soybean varieties when fed to broilers.

Evaluation of broiler performance when fed Roundup-Ready wheat (event MON 71800), control, and commercial wheat varieties.

We evaluated the nutritional value of broiler diets containing approximately 40% wheat grain from Roundup Ready wheat (MON 71800), its similar nontransgenic control (MON 71900), or reference commercial wheat varieties. The feeding trial lasted 40 d, and each treatment consisted of 10 replicates of 1-d-old Ross 308 broilers (5 pens of males and 5 pens of females). Each pen contained 12 birds, and at d 13 birds were randomly removed until 9 birds remained. Body weight and feed intake were measured on pen basis at 40 d. At d 41, four broilers per pen were slaughtered. The carcasses were dissected, and cut-up yields were determined. Dry matter, protein, and fat contents of breast meat were determined. The data were analyzed by an ANOVA procedure. The BW and feed conversion at d 40 averaged 2,450 g and 1.52, respectively. There were no significant treatment x sex interactions, except for evisceration yield with significant differences (P < 0.05) in yield between birds fed 2 commercial wheat varieties. Data for final BW, feed conversion, carcass yield, and breast meat were not statistically different (P < 0.05) between broilers fed MON 71800 or MON 71900 or the population of birds fed commercial wheat varieties, except a lower carcass yield at d 41 for birds fed the nontransgenic control wheat. Thus MON 71800 was nutritionally equivalent to nongenetically modified wheat varieties when fed to broilers.

Residues of dioxins and PCBs in fat of growing pigs and broilers fed contaminated feed.

To investigate the kinetics of PCBs and dioxins, 3 week old broilers and 3 month old pigs were fed with a 10-fold diluted feed from the Belgium crisis for one week, followed by a period on clean feed. In the case of broilers this resulted in levels for dioxins, non-ortho and mono-ortho PCBs in fat of 102, 84 and 216 ng TEQ/kg, summarized to 402 ng TEQ/kg. Total levels decreased to 217 and 109 ng TEQ/kg after 1 and 3 weeks on clean feed. Indicator PCB levels decreased from an initial 6.2 mg/kg fat to respectively 3.2 and 1.5 mg/kg. The ratio of indicator PCBs to dioxins TEQs was stable over this period. Levels in back fat of pigs at the end of the exposure period were 26, 15, 82 and 123 ng TEQ/kg for respectively dioxins, non-ortho PCBs, mono-ortho PCBs and the sum. Total TEQ levels decreased to respectively 95, 70, 40, 22 and 12 ng TEQ/kg after 1, 2, 4, 8 or 12 weeks on clean feed. After 12 weeks dioxin levels were around 1 ng TEQ/kg. Indicator PCB levels decreased from 3.48 mg/kg to 2.65, 2.01, 1.25, 0.76 and 0.45 mg/kg fat, again after 1, 2, 4, 8 or 12 weeks on clean feed. Levels of dioxins decreased more rapidly than those of indicator PCBs, also reflected by the ratio of indicator PCBs to dioxins, being 133,000 at the end of the exposure period and 357,000 after week 12. It is concluded that the use of indicator PCBs for dioxins, in the case of a combined exposure, is a safe alternative for screening and in the case of pigs results in an overestimation rather than underestimation of the dioxin levels.

Residues of veterinary drugs in eggs and their distribution between yolk and white.

Veterinary drugs and feed additives (especially some coccidiostats) can be absorbed by the digestive tract of laying hens and transferred to the egg. Physicochemical characteristics of these compounds determine their pharmacokinetic behavior and distribution to and within the egg. Traditionally the quite lipid soluble drugs and additives are expected to yield residues only in the fat-rich yolk. However, the quite lipid soluble drug doxycycline--as well as many other drugs--showed during long-term administration higher residues in white than in yolk. In a model study with 11 sulfonamides differing in pK(a) value and lipid solubility, their distribution in vivo between yolk and white was determined. Neither differences in pK(a) values nor those in lipid solubility could explain the distributions found. Binding to egg white macromolecules in vivo as an explanatory factor was tested with five sulfonamides, and no correlation between binding and the distribution of sulfonamides between white and yolk was found. Literature data on the distribution of drugs between egg white and yolk showed a reasonable consistency within drugs and a large variability among drugs (as could be expected). This larger database also did not provide a clue as to what factor determines the distribution of a drug between egg white and yolk when given to laying hens.

Flubendazole residues in eggs after oral administration to laying hens: determination with reversed phase liquid chromatography.

Flubendazole residues in eggs were experimentally induced by providing groups of 8 laying hens feed with approximately 3, 10 and 30 mg kg-1 flubendazole for 21 days. Eggs were sampled during this period and one week after the administration. Samples of both whole egg and egg white/yolk were analysed separately. Flubendazole analysis was performed by reversed phase HPLC and UV detection at 250 nm (eggs) or 320 nm (feed). The limit of detection (LOD) for flubendazole in feed was 0.3 mg kg-1 and in whole egg 0.012 mg kg-1. Both the hydrolysed and reduced metabolites of flubendazole were also determined quantitatively. The eggs of control hens housed in the same room during the study period did not contain any detectable flubendazole or metabolite residue. The eggs from the lowest dosed group (3 mg kg-1 feed) did contain residues, but most of them were only slightly higher than the LOD. Residues in eggs collected from the laying hens that obtained feed with 10 and 30 mg kg-1 flubendazole reached a plateau level after some 10 days and there was a good dose response relation between levels in feed and those in eggs. The residues of parent compound and metabolites almost exclusively occurred in yolk, the metabolites accounting for some 60-65% of the total residue. The residues of the parent compound and its metabolites declined below 100 micrograms kg-1 5 days after the administration of dosed feed had ended.

Induction and characterization of multianalyte antibodies against penicillins in egg yolk.

Antibodies against penicillins were induced in eggs of laying hens after immunization with 6-aminopenicillanic acid (6-APA) coupled to key-hole limpet hemocyanin (KLH). Development of the antibody titer was monitored by an indirect enzyme-linked immunosorbent assay (ELISA), with 6-APA coupled to ovalbumin as antigen for coating microtiter plates. Different characteristics (time course, affinity) of the immune reaction were observed by testing eggs of individual hens. Titer values varied between 150 and 2000. Antibodies were isolated by polyethylene glycol precipitation and affinity chromatography, using a hapten sorbent with 6-APA as ligand. Glycine buffer, pH 3.0, was used to elute the immunoglobulins. Antibody specificity was determined in a competitive ELISA with 7 penicillins and the cephalosporin cephalexin as competitors. Cross reactivities for the penicillins were between 100 and 290% (6-APA = 100%). Cephalexin cross reacted only marginally (3%).

Liquid chromatographic multicomponent method for determination of residues of ipronidazole, ronidazole, and dimetridazole and some relevant metabolites in eggs, plasma, and feces and its use in depletion studies in laying hens.

A simple, rapid liquid chromatographic (LC) method that uses UV/VIS detection has been developed for the determination in eggs of residues of the histomonostats dimetridazole (DMZ), ronidazole (RON), ipronidazole (IPR), and side-chain hydroxylated metabolites of DMZ and RON. Sample pretreatment includes an aqueous extraction, purification with an Extrelut cartridge, and acid partitioning with isooctane. An aliquot of the final aqueous extract is injected into a reverse-phase LC system; detection is performed at 313 nm. The limits of determination are in the 5-10 microgram/kg range. A UV/VIS spectrum can be obtained at the 10 microgram/kg level by using diode-array UV/VIS detection. Recoveries are between 80 and 98% with a coefficient of variation of about 5%. Some 20 samples can be analyzed per day. A side-chain hydroxylated metabolite of IPR can also be detected with this method, as demonstrated with samples from animal experiments. After a single oral dose of the drugs to laying hens, residues of the parent compound and/or the hydroxylated metabolites could be detected in eggs 5-8 days after dosing. Plasma distribution and excretion in feces were established both with and without deconjugation. DMZ and IPR were extensively metabolized to hydroxylated nitroimidazole metabolites; RON was excreted mainly as the parent compound.

Cadmium and lead in muscle tissue and organs of broilers, turkeys and spent hens and in mechanically deboned poultry meat.

Cadmium and lead levels have been determined in meat, livers and kidneys of turkeys, in livers and kidneys of spent hens, in kidneys of broilers and in mechanically deboned poultry meat (MDPM), by dry ashing of the sample with Mg(NO3)2 followed by differential pulse anodic stripping voltammetric analysis. The possible age-dependent cadmium and lead accumulation in livers and kidneys of turkeys was also studied. The cadmium concentrations in meat and organs of poultry are in good agreement with data reported for other countries, but lead concentrations were low compared with most data reported in the literature. The cadmium levels in kidneys and livers of turkeys increased with age, whilst the lead concentrations were unrelated to age. The cadmium and lead concentrations in industrially prepared MDPM from broiler carcasses were very low (less than 0.001-0.004 mg/kg and less than 0.01-0.06 mg/kg, respectively). In MDPM prepared from backs (including the kidneys) and necks, elevated cadmium concentrations were found. The increase was related to the cadmium concentrations in the kidneys. In MDPM prepared from broilers, turkeys and spent hens the cadmium concentrations varied between 0.001 and 0.002 mg/kg, 0.005 and 0.006 mg/kg and 0.007 and 0.028 mg/kg, respectively. For lead the concentrations in meat and MDPM were similar. It can be concluded from the results that in MDPM prepared from spent hens and possibly turkeys, when incorporating the kidneys, cadmium concentrations greater than 0.05 mg/kg, (the legal limit for cadmium in poultry meat in The Netherlands) may occur.

An automated HPLC determination of meticlorpindol in eggs with UV absorbance detection, using on-line dialysis and pre-concentration as sample clean-up; occurrence in and carry over to eggs.

A fully automated HPLC determination of the coccidiostat meticlorpindol in whole egg, egg white and yolk is described. The sample homogenate is dialysed on-line against water. The dialysate is concentrated on-line on a short reversed-phase (RP) column. The contents of this column are transferred to the reversed-phase analytical column by means of the mobile phase. Meticlorpindol is detected using an absorbance detector at 270 nm. Linear calibration graphs are obtained in the range 40-900 ng/g in whole egg and egg white (detection limit 10 ng/g) and 80-1800 ng/g in yolk (detection limit 20 ng/g). Out of 111 commercially obtained egg samples 12 contained meticlorpindol with levels varying from 10 to 433 ng/g. A group of laying hens, kept in cages, received 10 mg/kg of Lerbek (meticlorpindol and methylbenzoquate; Dow Chemical) in the feed for 10 days. Meticlorpindol residues in the eggs rose to a level of 622 ng/g. Meticlorpindol was found in the eggs until 6 days after withdrawal of the medicated feed. Another group received 110 mg/kg in the feed. Meticlorpindol residues rose to levels of 4480 ng/g in the eggs, 5880 ng/g in the egg white and 2660 ng/g in the yolk. Meticlorpindol was found in the eggs and the egg white until 14 days and in the yolk until 8 days after withdrawal of the medicated feed.

[Amprolium residues in eggs following administration of amprolium/ethopabate in laying hens and breeding hens]. / Residuen van amprolium in eieren na toediening van amprolium/ethopabaat aan leghennen en opfokleghennen.

Amprolium may be used as a coccidiostat in rearing hens and is a therapeutical agent used in laying hens. As a result of cross contamination, low amprolium levels may occur in feed. Feed containing a concentration of amprolium ranging from 5 to 250 mg/kg was therefore supplied to groups of laying hens. The amprolium residues in the yolks during and after treatment were subsequently determined. These levels varied from 1.75 mg/kg in the group fed 250 mg/kg to 0.2 mg/kg in the group fed 5 mg/kg. Amprolium levels in the whites of eggs were much lower than those in the yolks. The residues in yolks decreased below detectable levels (less than 0.005 mg/kg) within approximately ten days after treatment. Rearing hens in a tiered wire floor system were given amprolium in their feed until the first egg was laid. Amprolium residues in yolks were detected for well over a fortnight after the onset of laying. The amprolium residues determined in yolk did not exceed US tolerance levels of 8 mg/kg.

Plasma disposition and renal clearance of sulphadimidine and its metabolites in laying hens.

Plasma disposition of sulphadimidine (SDM) and its metabolites was studied in laying hens after 100 mg SDM kg-1 doses were administered as a single intravenous dose, a single oral dose and multiple oral doses once daily for five consecutive days. SDM was extensively metabolised by acetylation and hydroxylation. In plasma, the metabolite observed with the highest concentration was N4-acetylsulphadimidine (N4-SDM) followed by hydroxymethylsulphadimidine (CH2OH) and 5-hydroxysulphadimidine. Following intravenous administration a biphasic elimination (as seen for a capacity limited reaction) pattern for SDM and its metabolites was observed. Multiple (5x) SDM dosing revealed plasma SDM concentrations ranging between 7 and 108 micrograms ml-1; within 96 hours of termination of the multiple SDM dosing, the plasma SDM concentration was below 0.01 micrograms ml-1. The renal clearances of N4-SDM and the hydroxy metabolites were approximately 10 times greater than that of SDM. The SDM mass balance (faecal/urinary recovery) showed a loss of 56 per cent after intravenous dosage and of 67 per cent after a single oral dosage; the hydroxy metabolites accounted for the highest percentage in faeces/urine. Thus additional metabolic pathways must exist in laying hens.

Residues of sulphadimidine and its metabolites in eggs following oral sulphadimidine medication of hens.

The depletion of sulphadimidine (SDM) and its N4-acetyl and hydroxy metabolites was studied in eggs laid by hens after administration of either a single or multiple oral dosages of 100 mg SDM/kg. During medication and until 1 day after the last dose, the SDM and its metabolite concentrations in the egg white exceeded those in the egg yolk and reflected the plasma levels. In the period starting 2 days after the (last) dosage, the SDM concentration in the yolk became higher than in the egg white, and the drug depletion curves ran parallel. The mean maximum amount of SDM found in the whole egg was 1500 micrograms after a single and 1280 micrograms after multiple dosage. In eggs, traces of the N4-acetyl and 6-methylhydroxy metabolites could be detected (mainly in the egg white), and their concentrations were approximately 40 times lower than those of the parent drug. A highly significant correlation (P less than 0.005) was found between the development stage of the oocyte at the time of (last) medication and the amount of SDM found in the egg that developed from it. A period of 7 or 8 days after the (last) dosage of 100 mg SDM/kg/day is required to obtain SDM levels below 0.1 micrograms/g egg.

Biological activity of technical Aroclor 1254 compared to Aroclor 1254 residues: swine fat residues fed to broiler cockerels.

Fat from Aroclor 1254-treated swine was rendered and incorporated into the diets of broiler chicks for 3-4 weeks. The technical Aroclor 1254 which was fed to the swine was also mixed into control lard for comparison at dietary concentrations of 0.07-9.0 mg/kg. The swine-residue PCB seemed to have a higher proportion of strong microsomal inducers, but the technical PCB was slightly more effective in inducing ethoxy resorufin and p-nitro-anisole (pNA) O-dealkylases than the swine-residue PCB. No overt signs of toxicosis were apparent and none of the diets resulted in changes in growth, relative organ weights, microsomal protein or high affinity pNA O-dealkylase. Increases in cytochrome(s) P-450 were significant only at the higher dietary concentrations (approx. 9 mg/kg) while ethoxyresorufin O-dealkylase was induced at dietary concentrations below 1 mg/kg.