Investigation of the Performance of the Disintegration Test for Dietary Supplements

The aim of this study was to investigate how beaker size, basket assembly, use of disk, and immersion medium impact the disintegration time of dietary supplements. The disintegration times were determined for five tablet and two capsule products. A two-station disintegration tester was used with Apparatus A or Apparatus B as described in the United States Pharmacopeia (USP) chapters, < 701 > and < 2040 >. Two beakers complying with the harmonized specifications were used, one with a volume of 1,000 mL and one with a 1,500-mL volume. The disintegration data were analyzed using ANOVA for the following factors: beaker size, equipment (App A and B) and condition (with/without disk). Two tablet products were not sensitive to any changes in the test conditions or equipment configurations. One product was only partially sensitive to the test conditions. The other products showed impact on the disintegration time for all test conditions. The results revealed that these tablet products might pass or fail current USP disintegration requirements depending on the equipment configuration. Similar results were obtained for the two investigated capsule formulations. One product might fail current USP disintegration requirements if the large beaker was used, but might pass the disintegration requirements when the small beaker was used. Hydroxy propyl methyl cellulose capsules were mostly influenced if sodium instead of a potassium buffer was used as the immersion medium. The results demonstrate that the current harmonized ICH specifications for the disintegration test are insufficient to make the disintegration test into reliable test for dietary supplements.

Offspring size effects in the marine environment: A field test for a colonial invertebrate

A central tenet of life-history theory is the presence of a trade-off between the size and number of offspring that a female can produce for a given clutch. A crucial assumption of this trade-off is that larger offspring perform better than smaller offspring. Despite the importance of this assumption empirical, field-based tests are rare, especially for marine organisms. We tested this assumption for the marine invertebrate, Diplosoma listerianum, a colonial ascidian that commonly occurs in temperate marine communities. Colonies that came from larger larvae had larger feeding structures than colonies that came from smaller larvae. Colonies that came from larger larvae also had higher survival and growth after 2 weeks in the field than colonies that came from smaller larvae. However, after 3 weeks in the field the colonies began to fragment and we could not detect an effect of larval size. We suggest that offspring size can have strong effects on the initial recruitment of D. listerianum but because of the tendency of this species to fragment, offspring size effects are less persistent in this species than in others.

Effect of Test Environment and Microstructure on the Flexural Strength of Dental Porcelains

Purpose: The objective of this study was to verify the influence of test environment on the flexural strength of dental porcelains with distinct microstructures. Material and Methods: Disk-shaped specimens from three dental porcelains with distinct leucite content (VM: zero; CE: 12; NS: 22 vol%) were manufactured and tested for biaxial flexural strength in air and immersed in artificial saliva. The results were analyzed by means of two-way ANOVA and Tukey`s test (alpha = 0.05). Results: The flexural strength (MPa) obtained for ambient air and artificial saliva environments, respectively, were: 110.0 +/- 16.0 and 81.5 +/- 10.8 for VM; 51.9 +/- 4.0 and 42.0 +/- 4.7 for CE; 72.0 +/- 11.5 and 63.6 +/- 5.8 for NS. A numerical decrease in the mean flexural strength was observed for all groups when specimens were tested under artificial saliva; however, the difference was only statistically significant for VM. Conclusions: The results indicate that the effect of water immersion on the flexural strength of dental porcelains varies according to their leucite content, as only the material without leucite in its microstructure (VM) showed significant strength degradation when tested under water.

The development of a positive non-infectious control for the detection of Perkinsus using the Ray test

To establish a noncontagious control for the Ray thioglycollate test for the detection of Perkinsus in mollusks we evaluated nonviable stages of P. olseni for enlargement of hypnospores and blue/black iodine stain. Trophozoites made nonviable with formalin, irradiation or colchicine failed to swell in thioglycollate. They remained small and did not differentially stain in iodine. Trophozoites that had already developed into hypnospores in thioglycollate were rendered inactive by freezing, ethanol or formalin immersion. They retained their iodinophilic properties and thus could provide a partial control for the Ray Test.

Methods for field detection of resistance to temephos in simuliids. Larval esterase level and topical application of the insecticide to adults

Two practical field methods for indirect detection of simuliid populations resistant to temephos are proposed. The first is based on high esterase activity in resistant larvae and involves adaptations of a filter paper test in which faintly stained spots indicate susceptible populations and strongly stained ones reveal populations resistant to temephos. The second is based on the resistance to the larvicide when adults are topically exposed, and involves the use of diagnostic doses obtained by the comparison between the LD50 for susceptible and resistant populations. The relevance of such methods is discussed in order to help resistance detection in Simulium pertinax Kollar control programmes.

Larval dispersal and predation in experimental populations of Chrysomya albiceps and Cochliomyia macellaria (Diptera: Calliphoridae)

In this study we investigated the larval dispersal associated with larval predation in experimental populations of Chrysomya albiceps and Cochliomyia macellaria. Frequency distribution of sampling units (G test) in the substrate was used to evaluate variation in larval dispersal. An experimental acrylic channel (1 x 0.1 x 0.2 m) covered with wood shavings was used to observe larval dispersal prior to pupation. The acrylic channel was graduated at 0.05 m intervals, each representing a sampling unit; hence, 20 sampling units were set up. A Petri dish containing third instar larvae of single and double species was deposited at one edge of the acrylic channel allowing larvae to disperse. The number of buried pupae (0, 1, 2, …n) present in each sampling unit was recorded. For double species, the number of recovered larvae of C. albiceps was similar to the number initially released on the dish Petri. On the other hand, the number of recovered larvae of C. macellaria was significantly smaller than the initially released number. The results show that C. albiceps attacks C. macellaria larvae during the larval dispersal process. The larval distribution of C. albiceps did not differ significantly from C. macellaria in double species, but it differed significantly in single species. The larval aggregation level of C. macellaria decreased when C. albiceps was present and the larval aggregation level of C. albiceps increased when C. macellaria was present. The implications of such findings for the population dynamics of these species are discussed.

Larval biometry of Simulium rubrithorax (Diptera: Simuliidae) and size comparison between populations in the states of Minas Gerais and Roraima, Brazil

The number of larval instars of Simulium (Hemicnetha) rubrithorax Lutz (Diptera: Nematocera) was determined using the lateral length of the head capsule. In this study 1,035 larvae, of different sizes, were measured (639 from the state of Roraima and 396 from the state of Minas Gerais). A frequency distribution analysis was carried out on the measurements of the lateral length of the head capsule to determine the number of larval instars. The limits of each instar were defined by the lower frequency of the measurements falling in a range of values, by the presence of the "egg burster" that characterizes the first larval instar, and by the developmental stage of the gill histoblast. The determination of the instar number was tested using a Student's t-test (p < 0.05), the Dyar rule and the Crosby growth rule. The results indicate the existence of 7 larval instars for this species, although this result was not in accordance to the Crosby rule. Last-instar larvae from two widely separated geographical populations (Roraima and Minas Gerais), collected in habitats with different water temperature were compared and no differences (p > 0.05) were observed between them.

Western blotting using Strongyloides ratti antigen for the detection of IgG antibodies as confirmatory test in human strongyloidiasis

The present study was conducted to evaluate the frequency of antigenic components recognized by serum IgG antibodies in Western blotting (WB) using a Strongyloides ratti larval extract for the diagnosis of human strongyloidiasis. In addition, the WB results were compared to the enzyme-linked immunosorbent assay (ELISA) and the indirect immunofluorescence antibody test (IFAT) results. Serum samples of 180 individuals were analyzed (80 with strongyloidiasis, 60 with other intestinal parasitoses, and 40 healthy individuals). S. ratti was obtained from fecal culture of experimentally infected Rattus rattus. For IFAT, S. ratti larvae were used as antigen and S. ratti larval antigenic extracts were employed in WB and ELISA. Eleven S. ratti antigenic components were predominantly recognized by IgG antibodies in sera of patients with strongyloidiasis. There was a positive concordance for the three tests in 87.5% of the cases of strongyloidiasis. The negative concordance in the three tests was 94% and 97.5%, in patients with other intestinal parasitoses and healthy individuals, respectively. In cases of positive ELISA and negative IFAT results, diagnosis could be confirmed by WB. ELISA, IFAT, and WB using S. ratti antigens showed a high rate of sensitivity and specificity. In conclusion, WB using S. ratti larval extract was able to recognize 11 immunodominant antigenic components, showing to be a useful tool to define the diagnosis in cases of equivocal serology.

An evaluation of the dot-ELISA procedure as a diagnostic test in an area with a high prevalence of human Toxocara canis infection

The aim of this work was to evaluate a dot-enzyme-linked immunosorbent assay (dot-ELISA) using excretory-secretory antigens from the larval stages of Toxocara canis for the diagnosis of toxocariasis. A secondary aim was to establish the optimal conditions for its use in an area with a high prevalence of human T. canis infection. The dot-ELISA test was standardised using different concentrations of the antigen fixed on nitrocellulose paper strips and increasing dilutions of the serum and conjugate. Both the dot-ELISA and standard ELISA methods were tested in parallel with the same batch of sera from controls and from individuals living in the problem area. The best results were obtained with 1.33 µg/mL of antigen, dilutions of 1/80 for the samples and controls and a dilution of 1/5,000 for the anti-human IgG-peroxidase conjugate. All steps of the procedure were performed at room temperature. The coincidence between ELISA and dot-ELISA was 85% and the kappa index was 0.72. The dot-ELISA test described here is rapid, easy to perform and does not require expensive equipment. Thus, this test is suitable for the serological diagnosis of human T. canis infection in field surveys and in the primary health care centres of endemic regions.

Micronucleus test on Triturus carnifex as a tool for environmental biomonitoring.

The amphibian micronucleus test has been widely used during the last 30 years to test the genotoxic properties of several chemicals and as a tool for ecogenotoxic monitoring. The vast majority of these studies were performed on peripheral blood of urodelan larvae and anuran tadpoles and to a lesser extent adults were also used. In this study, we developed protocols for measuring micronuclei in adult shed skin cells and larval gill cells of the Italian crested newt (Triturus carnifex). Amphibians were collected from ponds in two protected areas in Italy that differed in their radon content. Twenty-three adult newts and 31 larvae were captured from the radon-rich pond, while 20 adults and 27 larvae were taken from the radon-free site. The animals were brought to the laboratory and the micronucleus test was performed on peripheral blood and shed skins taken from the adults and on larval gills. Samples from the radon-rich site showed micronucleus frequencies higher than those from the radon-free site and the difference was statistically significant in gill cells (P < 0.00001). Moreover, the larval gills seem to be more sensitive than the adult tissues. This method represents an easy (and noninvasive in the case of the shed skin) application of the micronucleus assay that can be useful for environmental studies in situ. Environ. Mol. Mutagen. 56:412-417, 2015. © 2014 Wiley Periodicals, Inc.

Soluble sugars and germination of Annona emarginata (Schltdl.) H. Rainer seeds submitted to immersion in GA3 up to different water contents

The objective of this study was to evaluate the effect of different water contents achieved by Annona emarginata (Schltdl.) H. Rainer seeds during immersion in GA3 solutions, in variation of soluble sugars levels and germination. Seeds with 10% of initial water content were submitted to imbibition in GA3 solutions with concentrations of 0; 250; 500; 750 and 1000 mg L-1 and when they reached the water content of 15%, 20%, 25%, 30% and 35%, the quantification of soluble sugars levels and germination test were performed. Seeds immersed up to they reach 15% of water with GA3 and immersed up to the water acquisition of 20% without GA3, presented higher soluble sugars levels and germination percentage, which were decreased when the seeds reached 30% and 35% of water, independently of the presence of the plant growth regulator. It was conclude that different water contents reached by the seeds in immersion treatments with GA3 affect the soluble sugars levels and germination percentage of Annona emarginata seeds. Thus, in treatments with Annona emarginata, the seeds must remain immersed in water without GA3 up to they reach 20% of water, as higher water contents (35%) reduce the soluble sugars levels and the seed germination percentage.

Viability of barley seeds by the tetrazolium test

The tetrazolium test is used to control seed quality of various plant species since it allows a rapid evaluation of viability. Freshly harvested barley seeds show dormancy that can make the germination test ineffective for an immediate evaluation. Therefore, the development of more efficient methods, such as the tetrazolium test, is necessary. The objective of this research work was to study various procedures for performing the tetrazolium test on barley seeds. Five lots of cv. BRS 195 barley seeds were used and subjected to the following treatments: two different methods of seed preconditioning (direct immersion in H2O and between sheets of moistened paper towels); two types of preparation for staining (longitudinal cross-section of the seed through the embryo with immersion of one half in a 2,3,5 triphenyl tetrazolium chloride solution or placing both halves on top of filter paper moistened with the tetrazolium salt solution); two methods of staining (on top of filter paper and direct immersion in the tetrazolium salt solution). Three concentrations of the tetrazolium salt solution (0.1%, 0.5%, and 1.0%) were used. It was concluded that the tetrazolium test on barley seeds may be accomplished with preconditioning by direct immersion in H2O and staining by immersing in a 0.1% or 0.5% concentration of tetrazolium salt solution or staining on top of filter paper moistened with such solution at a 1.0% concentration.

The joint effects of larval density and C-14-cypermethrin on the life history and population growth rate of the midge Chironomus riparius

1. Chemical effects on organisms are typically assessed using individual-level endpoints or sometimes population growth rate (PGR), but such measurements are generally made at low population densities. In contrast most natural populations are subject to density dependence and fluctuate around the environmental carrying capacity as a result of individual competition for resources. As ecotoxicology aims to make reliable population projections of chemical impacts in the field, an understanding of how high-density or resource-limited populations respond to environmental chemicals is essential. 2. Our objective was to determine the joint effects of population density and chemical stress on the life history and PGR of an important ecotoxicological indicator species, Chironomus riparius, under controlled laboratory conditions. Populations were fed the same ration but initiated at different densities and exposed to a solvent control and three concentrations of C-14-cypermethrin in a sediment-water test system for 67 days at 20 +/- 1 degreesC. 3. Density had a negative effect on all the measured life-history traits, and PGR declined with increasing density in the controls. Exposure to C-14-cypermethrin had a direct negative effect on juvenile survival, presumably within the first 24 h because the chemical rapidly dissipated from the water column. Reductions in the initial larval densities resulted in an increase in the available resources for the survivors. Subsequently, exposed populations emerged sooner and started producing offspring earlier than the controls. C-14-cypermethrin had no effect on estimated fecundity and adult body weight but interacted with density to reduce the time to first emergence and first reproduction. As a result, PGR increased with cypermethrin concentration when populations were initiated at high densities. 4. Synthesis and applications. The results showed that the effects of C-14-cypermethrin were buffered at high density, so that the joint effects of density and chemical stress on PGR were less than additive. Low levels of chemical stressors may increase carrying capacity by reducing juvenile competition for resources. More and perhaps fitter adults may be produced, similar to the effects of predators and culling; however, toxicant exposure may result in survivors that are less tolerant to changing conditions. If less than additive effects are typical in the field, standard regulatory tests carried out at low density may overestimate the effects of environmental chemicals. Further studies over a wide range of chemical stressors and organisms with contrasting life histories are needed to make general recommendations.

Comparison between cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) in short-term skeletal muscle recovery after high-intensity exercise in athletes-preliminary results

In the last years, phototherapy has becoming a promising tool to improve skeletal muscle recovery after exercise, however, it was not compared with other modalities commonly used with this aim. In the present study we compared the short-term effects of cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) with placebo LEDT on biochemical markers related to skeletal muscle recovery after high-intensity exercise. A randomized double-blind placebo-controlled crossover trial was performed with six male young futsal athletes. They were treated with CWIT (5A degrees C of temperature [SD +/- 1A degrees]), active LEDT (69 LEDs with wavelengths 660/850 nm, 10/30 mW of output power, 30 s of irradiation time per point, and 41.7 J of total energy irradiated per point, total of ten points irradiated) or an identical placebo LEDT 5 min after each of three Wingate cycle tests. Pre-exercise, post-exercise, and post-treatment measurements were taken of blood lactate levels, creatine kinase (CK) activity, and C-reactive protein (CRP) levels. There were no significant differences in the work performed during the three Wingate tests (p > 0.05). All biochemical parameters increased from baseline values (p < 0.05) after the three exercise tests, but only active LEDT decreased blood lactate levels (p = 0.0065) and CK activity (p = 0.0044) significantly after treatment. There were no significant differences in CRP values after treatments. We concluded that treating the leg muscles with LEDT 5 min after the Wingate cycle test seemed to inhibit the expected post-exercise increase in blood lactate levels and CK activity. This suggests that LEDT has better potential than 5 min of CWIT for improving short-term post-exercise recovery.

Ultramorphology and histochemistry of fat body cells from last Instar larval of the Pachycondyla (=Neoponera) villosa (Fabricius) (Formicidae: Ponerinae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)