Developmental endpoints of chronic exposure to suspected endocrine-disrupting chemicals on benthic and hyporheic freshwater copepods.

The aims of this study were: (i) to assess if carbamate pesticides and ammonium, widely detected in European freshwater bodies, can be considered ecologically relevant endocrine-disrupting chemicals (EDCs) for benthic and interstitial freshwater copepods; and (ii) to evaluate the potential of copepods as sentinels for monitoring ecosystem health. In order to achieve these objectives, four species belonging to the harpacticoid copepod genus Bryocamptus, namely B. (E.) echinatus, B. (R.) zschokkei, B. (R.) pygmaeus and B. (B.) minutus, were subjected to chronic exposures to Aldicarb and ammonium. A significant deviation from the developmental time of unexposed control cultures was observed for all the species in test cultures. Aldicarb caused an increase in generation time over 80% in both B. minutus and B. zschokkei, but less than 35% in B. pygmaeus and B. echinatus. Ammonium increased generation time over 33% in B. minutus, and 14, 12 and 3.5% for B. pygmaeus, B. zschokkei and B. echinatus, respectively. On the basis of these results it can be concluded that chronic exposure to carbamate pesticides and ammonium alters the post-naupliar development of the test-species and propose their potential role as EDCs, leaving open the basis to search what are the mechanism underlying. A prolonged developmental time would probably produce a detrimental effect on population attributes, such as age structure and population size. These deviations from a pristine population condition may be considered suitable biological indicators of ecosystem stress, particularly useful to compare polluted to unpolluted reference sites. Due to their dominance in both benthic and interstitial habitats, and their sensitivity as test organisms, freshwater benthic and hyporheic copepods can fully be used as sentinel species for assessing health condition of aquatic ecosystems as required by world-wide water legislation.

Relative sensitivity of hyporheic copepods to chemicals.

The sensitivity of harpacticoid copepods was tested against selected pollutants. Acute toxicity tests were carried out for five hyporheic species exposed to pesticides, ammonia, and metals. The stygoxene Bryocamptus zschokkei, B. minutus, B. pygmaeus and Attheyella crassa; and the stygophilous B. echinatus were sampled and cultured during 8 months in controlled conditions. A first test protocol is presented. The acute endpoints among species fell within one order of magnitude. The sensitivity among various species evaluated in this study is consistent and the choice of species for further sediment/groundwater assessment is not specific to a chemical class. These potential test organisms would be more suitable to protect meiofaunal communities.

Analysis of pulegone and its enanthiomeric distribution in mint-flavoured food products.

A procedure for the extraction and determination of pulegone enanthiomers in mint essential oils and mint products (syrups, dried leaves, toothpaste, lozenges, candy and chewing-gum) was developed. The compounds were recovered from the food matrices by employing a simultaneous distillation-extraction (SDE) technique with a Likens-Nickerson apparatus using dichloromethane as an extraction solvent. The analyses were performed by capillary gas chromatography mass spectrometry (GC/MS). Experiments on food products spiked at different pulegone concentrations showed recoveries ranging from 95 to 106%. The detection limit was about 5?mg?l(-1) for both pulegone enanthiomers and good linearity was found in the concentration range 0.5-25?mg?l(-1). In a number of repeated analyses, the pulegone peak height repeatability (RSD) was 0.2%. The pulegone enanthiomers were separated and quantified by enanthioselective multidimensional gas chromatography. The results of analyses conducted on essential mint oils and mint-flavoured food products are reported.

Analysis of free and esterified ergosterol in tomato products.

A new extraction and chromatographic procedure to quantify free and esterified ergosterol in tomato products was devised. The extraction solution was composed of a dichloromethane/methanol mixture in a 2:1 (v/v) ratio. This extraction solvent allowed for higher ergosterol recovery from tomato products (an average of 25% more) compared to hexane, which is frequently employed for ergosterol extraction. Both free and esterified ergosterol were determined by HPLC reverse-phase chromatography employing a Nova-Pak C-18 column (300 x 3.9 mm), filled with 4 mm average particle size and a guard column of the same material. The elution was performed at a flow rate of 1 mL. min(-1) with a linear gradient of solvent A (methanol/water, 80:20, v/v) and solvent B (dichloromethane). The gradient, starting at sample injection, was from 0 to 50% B for 20 min for the free ergosterol analysis and additional 15 min at 50% B to analyze the ergosterol esters. This technique has proven to be more sensitive for ergosterol determination than other reported chromatographic procedures. Moreover, ergosterol esters, extracted from various fungal sources, separated well and were easily quantified.

Determination of residual pectin methylesterase activity in food products.

A method for the determination of a low level of pectin methylesterase activity from vegetable products is described. The method is based on an affinity chromatography technique that employs a resin-bound pectin methylesterase inhibitor, purified from kiwi fruit, which selectively binds the pectin methylesterase. The resin has the capacity to concentrate the enzyme, allowing measurement of enzyme activities too low to be determined by commonly employed techniques and commensurate with those found in pasteurized food products. The enzyme is eluted from the resin at alkaline pH (9.5) and assayed by a pH-stat method. Depulped orange juices containing different amounts of pectin methylesterase were prepared and used to determine enzyme recovery. The results show a recovery of 90% with a standard deviation of 6.8%.

A glycoprotein inhibitor of pectin methylesterase in kiwi fruit. Purification by affinity chromatography and evidence of a ripening-related precursor.

The pectin methylesterase inhibitor from kiwi fruit (Actinidia chinensis) was purified by a single-step procedure based on affinity chromatography. Partially purified tomato pectin methylesterase was covalently bound to Sepharose. The affinity resin strongly and selectively binds the inhibitor, which could be eluted in high yield as a single, homogeneous and sharp peak by high salt concentration at pH 9.5 without loss of inhibitory activity. The purified protein possesses a molecular mass of 18 kDa, as estimated by SDS/PAGE, whereas by gel filtration under native conditions, its molecular mass appears to be 25 kDa. The inhibitor interacts with pectin methylesterase, forming a 1:1 complex, as demonstrated by gel-filtration experiments. The inhibitor was glycosylated. Its glycidic portion can be removed by digestion with N-glycosidase F after protein denaturation and, to a minor extent, by digestion with N-glycosidase H. No glycidic residue could be removed by digesting the native protein with those N-glycosidases. Antibodies against pectin methylesterase inhibitor were raised in rabbits and used to evidence protein expression during fruit ripening. The results showed that the inhibitor is present in the unripe fruit as an inactive precursor with a higher molecular mass (30 kDa) and is transformed into the active protein, most likely by proteinase action, during the course of the ripening process.

Effects of exposing broiler breeders to nicarbazin contaminated feed.

Ten-mo-old broiler feeds were fed nicarbazin (NCZ) at 0, 25, 50 and 100 ppm of their diet for 2, 4, or 6 days to simulate accidental contamination of their feed with the medicant. Reduced egg production was observed in all treatments except 25 and 50 ppm NCZ for 2 days. A consistent reduction in egg weight occurred only at the maximum treatment level of 100 ppm for 6 days. Reduction in hatchability was generally evident by Days 5 and 6 of the experiment except for the lowest treatment of 25 ppm NCZ for 2 days. Due partially to the low number of eggs set, no statistically significant reduction in hatchability was seen for the group receiving 50 ppm NCZ for 4 days, but hatchability had dropped over 17 percentage points (from 93.3 to 75.5%) by Days 5 and 6 of the experiment, and continued to drop to a low of 31% on Days 11 and 12 of the experiment. Shell pigmentation was the most sensitive characteristic measured, with significant depigmentation occurring after only 2 days of feeding 25 ppm NCZ. Generally, the severity and duration of effects were in proportion to medicant concentration and length of treatment time. Fertility was not influenced by the medicant.

A glycoprotein inhibitor of pectin methylesterase in kiwi fruit (Actinidia chinensis).

The finding of a powerful inhibitor of pectin methylesterase in ripe kiwi fruit is reported. The inhibitor was revealed to be a glycoprotein. It was purified to homogeneity and found to have a molecular mass of about 28 kDa, as estimated by gel filtration chromatography, SDS/PAGE and analytical ultracentrifugation. The sugar portion is composed of galactose, arabinose and rhamnose, the latter being much less represented. The amino acid composition showed a very high content of acidic residues compared to basic ones, which is the reason for the very low isoelectric point of the protein (less than 3.5). The kind of inhibition on kiwi pectin methylesterase was found to be competitive with an apparent Ki of 0.22 microM, using citrus pectin as a substrate. Moreover, the inhibitor is effective in inhibiting pectin methylesterase in the pH range 3.5-7.5. Kiwi inhibitor appears to be specific for pectin methylesterase, inasmuch as it was found to be ineffective against other polysaccharide-degrading enzymes, such as polygalacturonase and amylase. Conversely, it appears to be completely aspecific as far as the pectin methylesterase source is concerned. In fact, it was found to inhibit this enzyme effectively from all the sources we assayed, i.e. orange, tomato, apple, banana, potato.

Growth and carcass composition of female turkeys implanted with anabolic agents and fed high-protein and low-protein diets.

This laboratory investigated the anabolic effect of the synthetic steroid trienbolone acetate (TA) and found it effective in male and female meat turkeys without any apparent gross abnormalities (Poultry Sci., 61: 1386, 1982). The present study was undertaken to characterize the response of female turkeys, fed equicaloric diets varying in dietary protein density to anabolic agents implanted at 13 wks. TA and zeranol (Z) were tested singly and in combination (TAZ). Body weight gain and feed conversion at 16 wks of age were improved (P less than 0.01) with TA and TAZ. The response to TA was enhanced as dietary protein density increased. Carcass fat, protein, ash, energy, potassium, and calcium were not altered by implant treatment. A trend existed toward increased carcass fat with zeranol implantation. Carcass moisture (P less than 0.01) and sodium (P less than 0.05) were increased in the TA treatment. Blood plasma electrolytes were not affected by implant treatments but plasma calcium was decreased (P less than 0.05) by TAZ. No synergism was noted between TA and Z with respect to growth, feed conversion, carcass composition, plasma electrolytes with the exception of plasma calcium. No interaction was observed between dietary protein density and implant treatment. Performance variables increased and carcass fat decreased with increasing dietary protein density. These results confirm our earlier finding with respect to TA and provide additional evidence that dietary protein density influences the response of meat turkeys to TA. The data show that zeranol lacks an anabolic effect in turkeys.

Shell gland adenosine triphosphatase in hens producing strong and weak egg shells.

1. Two groups of 4 hens at the end of their first production cycle were classified as producers of strong egg shells (greater than 70 mg/cm2) and weak egg shells (less than 60 mg/cm2) on the basis of shell surface density. 2. Shell gland mucosa was homogenised and fractionated into nuclear, mitochondrial, microsomal and supernatant fractions, and ATPase activity determined. 3. ATPase activity in the total homogenate was significantly greater for hens producing strong shells (SES) than for hens producing weak shells (WES). 4. ATPase activities detected in the nuclear fraction, mitochondria and microsomes were significantly greater for SES than for WES birds.

Production and egg-quality responses of White Leghorn layers to anticoccidial agents.

Two studies were conducted to determine the effects of anticoccidial agents on the production and reproduction of White Leghorns. In Experiment 1, nicarbazin (NCZ) was fed at 0, 20, 50, and 100 ppm. Hen-day egg production, egg weight, the egg-yolk DNC (4-4'-dinitrocarbanilide) level, and egg-yolk mottling were affected by the treatments. When response was evidenced, the relationship between those variables and the level of NCZ was basically linear. Decreased egg production occurred from Days 5 and 6 of the treatment through Days 1 and 2 of withdrawal. On Days 9 and 10 of treatment, the control hens peaked at 92% hen-day production, while hens fed 20, 50, and 100 ppm of NCZ peaked late--at 90, 82, and 80%, respectively. Compared to the controls, egg weight was reduced linearly as the level of dietary NCZ increased. The egg-yolk DNC level increased from Days 3 and 4 of treatment through Days 9 and 10 of withdrawal. Egg yolk mottling generally increased along with the level and duration of feeding NCZ. If the NCZ was mistakenly fed to White Leghorn layers, ill effects would be alleviated within 10 days after drug withdrawal. In Experiment 2, halofuginone (3 ppm), maduramicin (5 ppm), monensin (100 ppm), narasin (70 ppm), nicarbazin (125 ppm), robenidine (33 ppm), and salinomycin (60 ppm) were fed to White Leghorn hens at the levels specified in parentheses. Nicarbazin reduced egg production, depressed egg weight, reduced shell thickness, and caused egg-yolk mottling; but internal egg quality, as measured by Haugh Units, was unaffected. Halofuginone, maduramicin, monensin, narasin, robenidine, and salinomycin did not have a meaningful effect on the variables measured when fed to White Leghorn layers.

Reproduction responses of broiler-breeders to anticoccidial agents.

Two experiments were conducted to determine the effects of anticoccidial agents on production and reproduction of broiler breeders. In Experiment 1, nicarbazin (NCZ) was fed at 20, 50, and 100 ppm. There was no depression in egg production, egg weight, or fertility from feeding these levels. As level of NCZ increased, there was a linear decrease in hatchability. The amount of 4,4'-dinitrocarbanilide (DNC) in the egg yolks increased linearly as the levels of NCZ went up; the degree of egg-shell depigmentation was directly related to the level of NCZ fed starting at 50 ppm. Experiment 2 utilized a different strain of broiler breeders. Halofuginone (3 ppm), maduramicin (5 ppm), monensin (100 ppm), narasin (70 ppm), NCZ (125 ppm), robenidine (33 ppm), and salinomycin (60 ppm) were fed to broiler breeders at the levels listed. Only NCZ reduced egg production. Narasin induced a reduction in egg weight. Both narasin and salinomycin caused a significant drop in hatchability. Feeding NCZ also induced a rapid and more severe decrease in hatchability. Monensin was the only anticoccidial agent that reduced fertility. Halofuginone, maduramicin, and robenidine had no biologically significant effect on henday production, egg weight, hatch of fertile eggs, or shell depigmentation. Feeding NCZ at 125 ppm caused a complete bleaching of brown-shell eggs by the 3rd consecutive day of treatment; but 7 days after NCZ was withdrawn from the feed, pigmentation returned to the pretreatment level.

Plasma calcitriol in chickens producing strong and weak egg shells and the response of hens to exogenous calcitriol.

Calcitriol (CAL) regulates intestinal calcium transport by inducing an increase in membrane phosphatidylcholine (PC) and calbindin. A positive correlation has been reported between shell gland PC and egg shell strength. Three experiments were conducted to test the hypothesis that the improvement in egg shell strength associated with increased PC is mediated via changes in the concentration of CAL. Sera from aged laying hens, identified as either strong (SES) or weak (WES) egg shell producers, were obtained from blood samples collected at 16 h after oviposition and assayed for CAL. Laying hens were injected (i/m) with 1.0 or 0.4 micrograms CAL/d for 28 d. Serum CAL in the WES hens was 70% of that in the SES hens (P less than .05). Hens producing extremely weak shells did not respond to 0.4 micrograms CAL/d for 10 d. Administration of CAL did not improve egg shell strength nor was serum CAL increased when WES hens were molted and shell gland PC increased. The results show that serum CAL is lower in aged hens producing weak egg shells but shell strength is not responsive to exogenous CAL and increased shell gland PC in WES hens after molting is not associated with higher serum CAL.

Characterization of shell gland lipids from chickens (Gallus domesticus) producing strong or weak egg shells.

1. The reduction in egg shell strength in hens at the end of the first reproductive cycle was not associated with abnormal uterine or body fat accumulation. 2. The phosphatidylcholine concentration was greater in the shell glands of hens producing strong egg shells compared to that of producing weak egg shells. 3. Differences were not detected in the total lipid or cholesterol concentrations or the fatty acid profiles of phosphatidylcholine or phosphatidylethanolamine fractions. 4. Following an induced molt, the fatty acid profile of the total lipid fraction of the shell gland was altered with a decrease in stearic acid and an increase in oleic acid. 5. The results demonstrate that alterations in egg shell strength are mainly associated with changes in shell gland phospholipid polar head group composition and not with changes in phospholipid fatty acid profile or cholesterol concentration.

The American Dental Network. Dentistry's information source.

A brief history of the development of telematics and electronic information retrieval was presented. Electronic information services of special interest to the dental professional were outlined. The American Dental Network, which provides the most comprehensive online services specifically for dentists, was described in detail.

Supplementary inorganic sulfate and methionine for young turkeys.

Two replicated experiments utilizing a total of 864 Large White Turkeys were conducted to determine the effects and possible interactions of supplementary inorganic sulfate and DL-methionine on body weight gain and feed consumption. Experimental diets, in which a corn-soybean meal basal diet was supplemented with inorganic sulfate either in the form of sodium sulfate (0, .0952, and .1904%) or potassium sulfate (0, .1180, and .2360%), and with methionine (0, .05, .10 and .15%) in the place of glucose monohydrate, were fed in a 3 X 2 X 4 experimental design. Each diet was fed to 9 males and 9 females from 1 day to 7 weeks of age. For 0, .05, .10, and .15% added methionine, 7-week body weights were 2020, 2162, 2238, and 2243 g, and feed consumptions were 3346, 3478, 3587, and 3571 g, respectively, producing significant (P less than .001) increases. For 0, .0215, and .0430% added sulfur from sulfates, body weights were 2163, 2167, and 2167 g, and feed consumptions were 3483, 3510, and 3494 g, respectively, but none were significantly different. Interactions were not significant; thus, sulfates did not replace or alter the requirement for added methionine in diets of young turkeys in this study.