Combination of On-Line Solid-Phase Extraction with LC-MS for the Determination of Potentially Hazardous Glycoalkaloids in Potato Products

A simple and rapid method for the determination of naturally occurring, potentially hazardous glycoalkaloids (GAs) in potatoes and their products has been developed. The procedure is based on the on-line solid-phase extraction of the acetic acid extracts from potato products and combined with liquid chromatography (LC)-mass spectrometry (MS) in a fully automated system (Symbiosis™, Spark Holland Instruments, Emmen, The Netherlands). As sorbent material HySphere™ 18HD was used for alkaloid enrichment. GAs were eluted with the LC gradient and directly analysed by MS. Detection of the analytes was achieved in the sensitive multiple reaction monitoring mode using two characteristic ions (m/z 98 as a qualifier for GAs and m/z 868.3 as a quantifier for α-solanine or m/z 852.4 for α-chaconine). Typical validation data for method precision (v _k α-solanine = 5.3–6.5, v _k α-chaconine = 3.4–15.4), accuracy (average recovery of α-solanine = 84%, average recovery of α-chaconine = 87%) and linearity over the range from 1 to 1,000 ng ml⁻¹ (R ² = 0.9915 for α-solanine, R ² = 0.9939 for α-chaconine) with detection limits of 0.3 ng ml⁻¹ for α-chaconine and 0.5 ng ml⁻¹ for α-solanine were obtained. GA contents of commercial potato products were determined by the new on-line method and afterwards compared with those obtained with an established high-performance LC routine procedure. Better performance of the on-line procedure was obvious from the standard deviations of both methods. Other advantages included a strong reduction of overall analysis time, human intervention and solvent consumption as well as waste production. The time required for the on-line analysis was 5 min, which would allow processing of almost 100 samples in 8 h.     

Inheritance and detoxification enzyme levels in <Emphasis Type="Italic">Tetranychus urticae</Emphasis> Koch (Acari: Tetranychidae) strain selected with chlorpyrifos

Changes in insecticide susceptibilities and detoxifying enzyme activities were measured in a strain of Tetranychus urticae Koch following repeated exposure to the organophosphate insecticide, chlorpyrifos. Twelve consecutive selection at the LC₆₀ of the parental strain increased resistance from 8.58 to 91.45 fold. The interaction of some synergists [piperonyl butoxide, triphenyl phosphate and S-benzyl-O,O-diisopropyl phosphorothioate (IBP)] with chlorpyrifos was analyzed in the selected strain. Solely IBP showed a low synergistic effect with chlorpyrifos. The selected strain also demonstrated resistance against abamectin, propargite, clofentezine and fenpyroximate. The mode of resistance inheritance to chlorpyrifos was found to be incompletely dominant, and not sex-linked. Non-specific esterase enzyme activity was raised from 19.35 to 33.59 mOD/min/mg proteins during the selection period and it was observed that esterase band intensities visualized by polyacrylamide gel electrophoresis increased. This study has investigated the selection of resistance to chlorpyrifos and documented resistance to abamectin, propargite, clofentezine and fenpyroximate in Turkish T. urticae. Esterase enzymes may be playing a role in chlorpyrifos resistance while glutathione S-transferase (GST) and P450 enzymes do not appear to have any significant involvement.     

Identification of oyster-derived hypotensive peptide acting as angiotensin-I-converting enzyme inhibitor

Angiotensin-I-converting enzyme (ACE) plays a crucial role in the crisis of hypertension. Some peptides that originate from protease hydrolysates are known to suppress ACE activity in vitro and in vivo. Here, we investigated whether trypsin hydrolysate of oyster Crassostrea gigas showed hypotensive activity and ACE inhibition. The hydrolysate significantly suppressed systolic blood pressure and ACE activity in spontaneously hypertensive rats following a one-shot oral administration and a long-term feeding experiment lasting 9 weeks. Each hydrolysate from oyster tissue showed ACE inhibitory activity, indicating the hypotensive effect was due to synergism. One potent ACE inhibitory peptide, Asp-Leu-Thr-Asp-Tyr, was identified from the hydrolysate of the striate muscle, and the peptide exhibited hypotensive activity in vivo. Protease digestion analysis suggested that Asp-Tyr could be the real effector of this penta-peptide in vivo.     

Thin layer chromatographic identification and quantitation of niacin and niacinamide in pharmaceutical preparations

A safe and simple thin layer chromatographic method has been developed for the estimation of niacin and niacinamide in pharmaceutical preparations containing other vitamins, enzymes, herbs, antiamoebic drugs, etc. The method involves removal of excipients from the preparation by ethanol precipitation and isolation of niacin or niacinamide from other ingredients by TLC on silica gel with water as developing solvent, followed by extraction in 0.1N HCl, and spectrophotometric estimation of the vitamin at 262 nm. The percent recoveries for niacin and niacinamide were 100.1 +/- 1.9 and 100.2 +/- 1.5, respectively.     

Historical processes and landscape context influence genetic structure in peripheral populations of the collared lizard (<Emphasis Type="Italic">Crotaphytus collaris</Emphasis>)

Populations at the periphery of a species’ range often show reduced genetic variability within populations and increased genetic divergence among populations compared to those at the core, but the mechanisms that give rise to this core-periphery pattern in genetic structure can be multifaceted. Peripheral population characteristics may be a product of historical processes, such as founder effects or population expansion, or due to the contemporary influence of landscape context on gene flow. We sampled collared lizards (Crotaphytus collaris) at four locations within the northern Flint Hills of Kansas, which is at the northern periphery of their range, to determine the genetic variability and extent of genetic divergence among populations for ten microsatellite loci (n = 229). We found low genetic variability (average allelic richness = 3.37 ± 0.23 SE; average heterozygosity = 0.54 ± 0.05 SE) and moderate population divergence (average F_ST = 0.08 ± 0.01 SE) among our sample sites relative to estimates reported in the literature at the core of the species’ range in Texas. We also identified differences in dispersal rates among sampling locations. Gene flow within the Flint Hills was thus greater than for other peripheral populations of collared lizards, such as the Missouri glade system where most of the mesic grasslands have been converted to forest since the last glacial retreat, which appears to have greatly impeded gene flow among populations. Our findings signify the importance of considering landscape context when evaluating core-peripheral trends in genetic diversity and population structure.     

Habitat modification by invasive crayfish can facilitate its growth through enhanced food accessibility

Background

Invasive ecosystem engineers can facilitate their invasions by modifying the physical environment to improve their own performance, but this positive feedback process has rarely been tested empirically except in sessile organisms. The invasive crayfish Procambarus clarkii is an ecosystem engineer that destroys aquatic macrophytes, which provide a physical refuge for animal prey, and this destruction is likely to enhance vulnerability to predators. Using two series of mesocosm experiments, we tested the hypothesis that the invasive crayfish increases its feeding efficiency on animal prey by reducing submerged macrophytes, thus increasing its individual growth rate in a positive density-dependent manner.

Results

In the first experiment, increasing crayfish density reduced both macrophytes and animal prey (dragonfly and chironomid larvae) and, importantly, increased the growth rate of individual crayfish, in accordance with our expectation. In the second experiment, we used artificial macrophytes to clarify whether the physical architecture of macrophytes itself protects animal prey and limits crayfish growth rate. Increasing the artificial macrophyte quantity not only increased the survival of animal prey, but also retarded the crayfish growth rate.

Conclusions

We conclude that macrophytes strengthen bottom-up control of crayfish, but this effect can be relaxed by increasing the density of crayfish via reduction in macrophytes. This positive feedback process may explain the crayfish outbreaks and regime shifts occasionally observed in invaded freshwater ecosystems.

     

Immunogene expression in head kidney and spleen of common carp (<Emphasis Type="Italic">Cyprinus carpio</Emphasis> L.) following thermal stress and challenge with Gram-negative bacterium, <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

To evaluate the effect of thermal and microbial stress on the immune response of common carp (Cyprinus carpio L.), relative mRNA expression level of pro-inflammatory cytokines [tumor necrosis factor alpha (TNF-α) and interleukin (IL)-1β] and other genes related to immune or stress response [inducible nitric oxide synthase (iNOS), heat shock protein 70 (Hsp70), superoxide dismutase one (SOD1), and glucocorticoid receptor (GR)] was measured by quantitative PCR (qPCR). In addition, total protein and total immunoglobulin level in blood plasma of experimental common carp was also assayed. All the above parameters were estimated 24 h post-challenge with Gram-negative bacterium, Aeromonas hydrophila. Common carp (54.89?±?6.90 g) were initially exposed to 20 °C (control group) and 30 °C (thermal stress group) water temperature for 30 days, followed by experimental challenge with 2.29?×?10⁸ colony forming unit/mL (CFU/mL; LD₅₀ dose) of A. hydrophila. Exposure of fish to thermal stress and subsequently challenge with A. hydrophila significantly (P?<?0.05) increases the IL-1β mRNA expression in head kidney and spleen of common carp by ~?39.94 and ~?4.11-fold, respectively. However, TNF-α mRNA expression in spleen decreased ~?5.63-fold in control fish challenged with A. hydrophila. Thermal stress and challenge with bacterium suppresses the iNOS and GR mRNA expression in spleen of common carp. Moreover, significant (P?<?0.05) increase in total protein content of blood plasma (~?43 mg/g) was evident in fish exposed to thermal stress and challenged with A. hydrophila. In conclusion, our study highlights the importance of elevated temperature stress and microbial infection in differential regulation of expression of several immunogenes in common carp.     

Ovicidal activity of seven <Emphasis Type="Italic">Pochonia chlamydosporia</Emphasis> fungal isolates on <Emphasis Type="Italic">Ascaris suum</Emphasis> eggs

The ovicidal effect of the nematophagous fungus Pochonia chlamydosporia on eggs of Ascaris suum was tested under laboratory conditions. A. suum eggs were plated on 2% water–agar with seven fungal isolates (Isol. 5, Isol. 31, Isol. 1, VC1, Isol. 12, Isol. 22 and VC4) and control without fungus. After 5, 7, 10, 14, 15 and 21 days of incubation, approximately 100 eggs were removed from the plates and classified according to the following parameters: type 1, biochemical and physiological effect without morphological damage to the eggshell, type 2, lytic effect with morphological alteration of the eggshell and embryo and type 3, lytic effect with morphological alteration of eggshell and embryo showing hyphal penetration and internal egg colonization. The isolates effectively destroyed A. suum eggs and all types of effects were observed during the experiment. There was no variation in ovicidal capacity (type 3 effect) among the isolates (p > 0.05) throughout the experiment. After 21 days, isolate 5 showed the highest percentages of type 3 effect (58.33%). The results indicated that P. chlamydosporia (Isol. 5, Isol. 31, Isol. 1, VC1, Isol. 12, Isol. 22 and VC4) can destroy A. suum eggs and is, therefore, a potential biological control agent of nematodes.     

Comparison of biological parameters between the invasive B biotype and a new defined Cv biotype of <Emphasis Type="Italic">Bemisia tabaci</Emphasis> (Hemiptera: Aleyradidae) in China

The sweetpotato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a widely distributed and destructive agricultural pest on various host plants. The biology of two biotypes of B. tabaci: the invasive B and a new defined Cv biotype, on a range of host plants (hibiscus, laurel, poinsettia, collard, cucumber and tomato) were studied in the laboratory. Results revealed that the developmental periods of the B biotype immatures were not significantly different on the tested host plants except those between laurel and collard. The Cv biotype immatures developed significantly slower on cucumber and tomato than on the other plants. B. tabaci B biotype had the highest survivorship on collard (68.55%), and the lowest on laurel (33.24%), while the Cv biotype had the highest and lowest survivorships on laurel (61.63%) and tomato (36.74%). Host plants did not significantly affect the pre-ovipostion period regardless of biotype. The longest averaged longevity and highest fecundity of B biotype were both recorded on collard: 25.15 days and 143.0 eggs. The highest fecundity of Cv biotype was 196.49 eggs on laurel and its longest longevity was on hibiscus (19.62 days). The intrinsic rate of natural increase (r _m) of B biotype on the three vegetables were all higher than those on the three ornamentals whereas the r _m of Cv biotype on the three ornamentals were all higher that those on the three vegetables. Our research indicates that B. tabaci B and Cv biotypes have different host plant suitabilities. The three tested vegetables were more suitable for B biotype while the three tested ornamental plants were more suitable for Cv biotype. The potential mechanism for the different suitability of B and Cv biotypes on various host plants is also discussed.