The Control of Reactive Oxygen Species Influences Porcine Oocyte In Vitro Maturation

The aim of this study was to examine the effect of varying intracellular reactive oxygen species (ROS) levels during oocyte in vitro maturation with enzymatic ROS production systems (xanthine + xanthine oxidase or xanthine + xanthine oxidase + catalase), scavenger systems (catalase or superoxide dismutase + catalase) or cysteine on porcine oocyte maturation. Oocyte ROS levels showed an increase when H₂O₂ or O₂?^‐ production systems were added to the culture medium (p < 0.05). On the other hand, the presence of ROS scavengers in the maturation medium did not modify oocyte ROS levels compared with the control after 48 h of maturation, but the addition of cysteine induced a decrease in oocyte ROS levels (p < 0.05). The ROS production systems used in this work did not modified the percentage of oocyte nuclear maturation, but increased the decondensation of sperm head (p < 0.05) and decreased the pronuclear formation (p < 0.05). In turn, the addition of O₂?^‐ and H₂O₂ scavenging systems during in vitro maturation did not modify the percentage of oocytes reaching metaphase II nor the oocytes with decondensed sperm head or pronuclei after fertilization. However, both parameters increased in the presence of cysteine (p < 0.05). The exogenous generation of O₂?^‐ and H₂O₂ during oocyte in vitro maturation would not affect nuclear maturation or later sperm penetration, but most of the spermatozoa cannot progress to form the pronuclei after fusion with the oocyte. The decrease in endogenous ROS levels by the addition of cysteine would improve pronuclear formation after sperm penetration.     

Determination of reference intervals for metabolic profile of Hanwoo cows at early,middle and late gestation periods

Background

Metabolic profile was initially designed as a presymptomatic diagnostic aid based on statistical analyses of blood metabolites to provide an early warning of certain types of metabolic disorder. However, there is little metabolic profile data available about Korean Hanwoo cows. Therefore, this study aimed to determine the reference intervals of metabolic profile for Korean Hanwoo cows.

Methods

Healthy animals (2,205) were selected and divided into early (day 1 to 95), middle (day 96 to 190) and late (day 191 to 285) period according to their gestating period. Metabolic profile including total protein (TP), albumin (Alb), urea (UREA), glucose (Glu), total cholesterol (T-Cho), long-chain fatty acid (LCFA), aspartate aminotransferase (AST), gamma-glutamyl transpeptidase (GGT), creatinine (Crea), calcium (Ca), inorganic phosphorous (iP) and magnesium (Mg) were analyzed using a TBA-40FR automatic biochemical analyzer. The data of Korean Hanwoo cows were then compared to those of the Japanese Wagyu cows.

Results

Most of the data of the Korean Hanwoo cows were relatively higher than those of Japanese Wagyu cows, with the exception of Glu and GGT. This may indicate that the nutritional level of feed for the Korean Hanwoo cows was higher than that of the Japanese Wagyu cows because of the different feeding system. In particular, relatively higher levels of UREA and LCFA were observed in the Korean Hanwoo cows, and this may also contribute to the low reproduction efficiency.

Conclusions

These findings may provide some theoretical basis for understanding the reproductive and feeding situation of Korean Hanwoo cows.     

Baicalein induction of hydroxyl radical formation via 12-lipoxygenase in human platelets: an ESR study

The pro-oxidant activities of baicalein, morin, myricetin, quercetin, and rutin were examined in various cell-containing systems including human platelets, rat vascular smooth muscle cells, human umbilical vein endothelial cells (HUVECs), human THP-1 cells, and fibroblast cells. Electron spin resonance (ESR) results showed that only baicalein generated hydroxyl radicals in a resting human platelet suspension, whereas the other flavonoids showed no effects on any of the resting cell systems. A low concentration of arachidonic acid (AA) increased the intensity of hydroxyl radicals, but a high concentration inhibited it. Collagen and thrombin, platelet aggregatory agents that can cause the release of AA by platelets, enhanced baicalein-induced hydroxyl radical formation, whereas ADP and U44619 showed no significant effects. Quinacrine and 5,8,11,14-eicosatetraenoic trifluoromethyl ketone, both PLA2 inhibitors, significantly attenuated baicalein-induced hydroxyl radical formation. These results suggest that baicalein-induced hydroxyl radical formation is associated with AA metabolite enzymes in human platelets. The formation of hydroxyl radicals was significantly inhibited by lipoxygenase inhibitors including nordihydroguaiaretic acid, (-)-epicatechin, (-)-epicatechin gallate, and hinokitiol, but was not affected by desferroxamine or the heme protein inhibitors KCN and NaN3. On the other hand, semiquinone free radicals were generated when baicalein was incubated with horseradish peroxidase/H2O2 or platelets/AA. The semiquinone radicals formed in the platelets/AA system could be extensively inhibited by desferroxamine, diethylenetriaminepentaacetic acid, KCN, and NaN3, indicating that prostaglandin H synthase (PGHS)-peroxidase may be involved. The results of this study led to the proposal that baicalein induces hydroxyl radical formation via 12-lipoxygenase and induces semiquinone radical formation via PGHS-peroxidase in human platelets.     

Improved HPLC-MS/MS method for determination of isoxaflutole (balance) and its metabolites in soils and forage plants

A robust multi-residue procedure is needed for the analysis of the pro-herbicide isoxaflutole and its degradates in soil and plant materials at environmentally relevant (<1 microg kg-1) levels. An analytical method using turbo-spray and heat-nebulizer high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was developed for the analysis of isoxaflutole (IXF) and its two metabolites, diketonitrile (DKN) and the benzoic acid metabolite (BA), at sub-microgram per kilogram levels in soil and plant samples. The average recoveries of the three compounds in spiked soil and plant samples ranged from 84 to 110% and 94 to 105%, respectively. The limits of quantification were validated at 0.06 microg kg-1 for soil and 0.3 microg kg-1 for plant samples. The limits of detection (LOD) for soil analysis were 0.01, 0.002, and 0.01 microg kg-1 for IXF, DKN, and BA, respectively. Corresponding LOD for the plant analysis method were 0.05, 0.01, and 0.05 microg kg-1. The developed method was validated using forage grass and soil samples collected from a field lysimeter study in which IXF was applied to each of four forage treatments. Forage plants and soils were sampled for analyses 25 days after IXF application to the soil. In soils, IXF was not detected in any treatment, and DKN was the predominant metabolite found. In forage plants, the concentrations of DKN and BA were 10-100-fold higher than that in soil samples, but IXF was not detected in any forage plants. The much higher proportion of BA to DKN in plant tissues (23-53%), as compared to soils (0-5%), suggested that these forages were capable of detoxifying DKN. The developed methods provided LODs at sub-microgram per kilogram levels to determine the fate of IXF and its metabolites in soils and forage plants, and they also represent considerable improvements in extraction recovery rates and detection sensitivity as compared to previous analytical methods for these compounds.     

Effect of dips in a 1-methylcyclopropene-generating solution on 'Harrow Sun' plums stored under different temperature regimes

The effect of postharvest dips in a 1-methylcyclopropene-generating solution of the formulation AFxRD-038 (Rohm & Haas) on plum fruit (Prunus salicina Lindell cv. 'Harrow Sun') quality and ripening during storage was determined. Fruit weight loss, tissue firmness, soluble solids content (SSC), titratable acidity (TA), ethylene production, respiration, and the activities of the cell wall modifying enzymes polygalacturonase (PG), 1,4-beta-D-glucanase/glucosidase (EGase), beta-galactosidase (beta-gal), and pectin methylesterase (PME) were measured. Fruit reddening, anthocyanin content, and phenylalanine ammonia-lyase (PAL) activity were also analyzed. The 1-MCP-treated fruit showed reduced ethylene production and respiration rate and delayed softening, which was associated with the reduction in the activity of PG, EGase, and beta-gal. The immersion in 1-MCP-generating solutions also decreased weight and acidity loss without modifying the fruit SSC. The immersion treatment was particularly effective in the fruit stored at 5 degrees C, keeping higher overall quality, maintaining lower levels of anthocyanins and PAL activity, and preventing flesh reddening. The present data show that beneficial effects in delaying plum fruit ripening and controlling chilling injury can be obtained by dipping the fruits in a solution of this novel 1-MCP-generating formulation.     

Determination of anabolic steroids in muscle tissue by liquid chromatography-tandem mass spectrometry

A specific and sensitive method based on liquid chromatography-tandem mass spectrometry using atmospheric pressure chemical ionization (LC-APCI-MS/MS) has been developed for the determination of four anabolic steroids [trenbolone, methylboldenone, methyltestosterone, and norethandrolone] in bovine muscle. Methyltestosterone- d 3 was used as internal standard. The procedure involved enzymatic hydrolysis, extraction with tert-butyl methyl ether, defattening, and final cleanup with solid-phase extraction with Oasis HLB cartridges. The analytes were analyzed by reversed-phase LC-MS/MS, acquiring two diagnostic product ions from the chosen precursor [M + H] (+) for the unambiguous confirmation of hormones. The method was validated according to the European Commission Decision 2002/657/EC for the detection and confirmation of residues in products of animal origin. The limits of detection (LOD) and limits of quantitation (LOQ) were found to be 0.3 ng/g and 1.0 ng/g, respectively. The accuracy and precision have been determined, with recoveries ranging from 83% to 104% and the CV factor not exceeding the value of 7%. The decision limits CCalpha were calculated and ranged from 0.05 to 0.15 ng/g while the detection capabilities CCbeta ranged from 0.09 to 0.25 ng/g. The method proved to be sensitive and reliable and thus renders an appropriate means for residue analysis studies.     

Autoimmune hepatitis-specific antibodies against soluble liver antigen and liver cytosol type 1 in patients with chronic viral hepatitis

Background

Non-organ specific autoantibodies are highly prevalent in patients with chronic hepatitis C (HCV). Among them, anti-liver kidney microsomal type 1 (LKM1) antibody – the serological marker of type 2 autoimmune hepatitis (AIH-2)- is detected in up to 11% of the HCV-infected subjects. On the other hand, anti-liver cytosol type 1 antibodies (anti-LC1) – either in association with anti-LKM1, or in isolation- and anti-soluble liver antigen antibodies (anti-SLA) have been considered as useful and specific diagnostic markers for AIH. However, their specificity for AIH has been questioned by some recent studies, which have shown the detection of anti-LC1 and anti-SLA by immunoprecipitation assays in HCV patients irrespective of their anti-LKM1 status. The aim of the present study was to test the anti-LC1 and anti-SLA presence by specific enzyme linked immunosorbent assays (ELISAs), in a large group of Greek HCV-infected patients with or without anti-LKM1 reactivity as firstly, immunoprecipitation assays are limited to few specialized laboratories worldwide and cannot be used routinely and secondly, to assess whether application of such tests has any relevance in the context of patients with viral hepatitis since antibody detection based on such ELISAs has not been described in detail in large groups of HCV patients.

Methods

One hundred and thirty eight consecutive HCV patients (120 anti-LKM1 negative and 18 anti-LKM1 positive) were investigated for the presence of anti-LC1 and anti-SLA by commercial ELISAs. A similar number (120) of chronic hepatitis B virus (HBV) infected patients seronegative for anti-LKM1 was also tested as pathological controls.

Results

Six out of 18 (33%) anti-LKM^pos/HCV^pos patients tested positive for anti-LC1 compared to 1/120 (0.83%) anti-LKM^neg/HCV^pos patients and 0/120 (0%) of the anti-LKM1^neg/HBV^pos patients (p < 0.001 for both comparisons). Anti-SLA antibodies were not present in any of the HCV (with or without anti-LKM1) or HBV-infected patients.

Conclusion

We showed that anti-LC1 and anti-SLA autoantibodies are not detected by conventional assays in a large group of anti-LKM1 negative patients with chronic hepatitis B and C infections. Based on these results we cannot find any justification for the application of anti-LC1 and anti-SLA tests in the routine laboratory testing of viral hepatitis-related autoantibody serology with the only potential exception being the anti-LC1 screening in anti-LKM1^pos/HCV^pos patients.

     

油用亚麻主要品质和农艺性状的变异分析

为深入了解油用亚麻主要品质和农艺性状的遗传变异,以253份油用亚麻种质为研究对象,在3个环境下（内蒙古呼和浩特市、集宁市和锡林郭勒盟太仆寺旗）对其全生育天数、株高、工艺长度、单株果数、每果粒数、单株粒重、千粒重和分枝数8个农艺性状及棕榈酸、硬脂酸、油酸、亚油酸、亚麻酸、粗脂肪含量6个品质性状进行鉴定。结果表明,8个农艺性状的变异系数为5.66%~42.65%;6个品质性状的变异系数为4.10%~30.14%。全生育天数在太仆寺旗表现最长,为112.51 d;千粒重和单株粒重在集宁地区表现最大,分别为5.94和0.55 g;果粒数和单株果数在呼和浩特地区表现最多,分别为5.65和16.90;粗脂肪和亚麻酸含量在太仆寺旗地区表现最大,分别为39.53%和53.45%;亚油酸和棕榈酸含量在集宁地区最大,分别为16.41%和5.09%;油酸和硬脂酸含量在呼和浩特地区表现最大,分别为24.03%和8.31%。聚类分析表明,253份油用亚麻种质被划分为4个类群,相同地理来源的油用亚麻种质被聚到1个类群,为油用亚麻种质资源的收集保存与繁殖提供了科学依据。     

Effect of protein source and level on growth and performance of the captive freshwater prawn,Macrobrachium rosenbergii

Growth, feed conversion and survival were determined for juvenile Macrobrachium rosenbergii held in tanks under semi-controlled environmental conditions. Feeding trials incorporated water-stable diets at three levels of protein (15, 25 and 35%). The principal protein source combinations consisted either of soybean and tuna meal, or of soybean, tuna and shrimp meal. In a 244-day comparison of these diets, higher protein content produced larger prawns (P < 0.01), but differences between sources were not significant. No significant differences existed between feed conversion ratios (range 1.36–1.72) or percentage survival (range 90.3–93.6%). Trials of several other diets were also conducted, including soybean and Tilapia meal, and copra and Tilapia meal (25% protein level) as principal protein source combinations. After 167 days on these diets, growth was inferior to that obtained with soybean and tuna meal or soybean, tuna and shrimp meal combinations. No significant differences existed between feed conversion ratios or percentages of survival.For the 244 days, a control group of prawns received no formulated diet. Growth and survival in this group during the first 110 days suggested that naturally occurring algae contributed substantially to the prawns' nutrition.Mean prawn length after 244 days on the best diet (35% protein from soybean and tuna meal) was 73 mm, and growth rate was equivalent to that achieved under pilot pond conditions.     

The Chinese grass carp, Ctenopharyngodon idella, its biology,introduction, control of aquatic macrophytes and breeding in the Sudan

The paper describes the distribution and biology of the grass carp following its introduction into Sudan. It provides aspects on the culture of the grass carp and its breeding under Sudanese conditions with emphasis on future research.