Andirobeira is an Amazonian tree, the seeds of which produce a commercially valuable oil that is used in folk medicine and in the cosmetic industry. Andiroba oil contains components with anti-inflammatory, cicatrizing and insect-repellant actions. However, virtually nothing is known of the safety of this oil for humans. The aim of this work was therefore to investigate the hematotoxicity, genotoxicity and mutagenicity of andiroba oil using the comet and micronucleus assays, and to assess its antioxidant properties and lipidome as a means of addressing safety issues. For the experiments, andiroba oil was administered by gavage for 14 consecutive days in nulliparous female Swiss mice randomly distributed in four groups: negative control and three doses of oil (500, 1000 and 2000 mg/kg/day). These doses were chosen based on recommendations of the OECD guideline no. 474 (1997). GC/MS was used to investigate the free fatty acid, cholesterol and triterpene content of andiroba oil in a lipidomic analysis. No clinical or behavioral alterations were observed throughout the period of treatment, and exposure to andiroba oil at the doses and conditions used here did not result in hematotoxic, genotoxic or mutagenic effects. Tests in vitro showed that oil sample 3 from southwestern of Brazilian Amazon had a high antioxidant capacity that may protect biological systems from oxidative stress, although this activity remains to be demonstrated in vivo.

BackgroundHolocentric chromosomes occur in approximately 750 species of eukaryotes. Among them, the genus Tityus (Scorpiones, Buthidae) has a labile karyotype that shows complex multivalent associations during male meiosis. Thus, taking advantage of the excellent model provided by the Buthidae scorpions, here we analyzed the chromosomal distribution of several repetitive DNA classes on the holocentric chromosomes of different populations of the species Tityus obscurus Gervais, 1843, highlighting their involvement in the karyotypic differences found among them.ResultsThis species shows inter- and intrapopulational karyotype variation, with seven distinct cytotypes: A (2n = 16), B (2n = 14), C (2n = 13), D (2n = 13), E (2n = 12), F (2n = 12) and G (2n = 11). Furthermore, exhibits achiasmatic male meiosis and lacks heteromorphic sex chromosomes. Trivalent and quadrivalent meiotic associations were found in some cytotypes. In them, 45S rDNAs were found in the terminal portions of two pairs, while TTAGG repeats were found only at the end of the chromosomes. In the cytotype A (2n = 16), the U2 snRNA gene mapped to pair 1, while the H3 histone cluster and C 0 t-1 DNA fraction was terminally distributed on all pairs. Mariner transposons were found throughout the chromosomes, with the exception of one individual of cytotype A (2n = 16), in which it was concentrated in heterochromatic regions.ConclusionsChromosomal variability found in T. obscurus are due to rearrangements of the type fusion/fission and reciprocal translocations in heterozygous. These karyotype differences follow a geographical pattern and may be contributing to reproductive isolation between populations analyzed. Our results also demonstrate high mobility of histone H3 genes. In contrast, other multigene families (45S rDNA and U2 snRNA) have conserved distribution among individuals. The accumulation of repetitive sequences in distal regions of T. obscurus chromosomes, suggests that end of chromosome are not covered by the kinetochore.

Andiroba oil (AO) is obtained from an Amazonian plant and is used in traditional medicine. We carried out a comparative study to test the cytotoxicity, genotoxicity, and hematotoxicity of the oil and its nanoemulsion (AN) in vitro (fibroblasts, lineage NIH/3T3) and in vivo (Swiss mice). The AN was characterized by DLS/Zeta, and its stability was investigated for 120 days. The biological activity of AN was assessed in vitro by MTT test and cell morphology analyses and in vivo by micronucleus, comet, and hematotoxicity tests. The AN presented a hydrodynamic diameter (Hd) of 142.5 ± 3.0 and PDI of 0.272 ± 0.007 and good stability at room temperature. The MTT test evidenced the cytotoxicity of AO and of AN only at their highest concentrations, but AN showed lower cytotoxicity than AO. A lower cytotoxicity of AN, when compared to AO, is in fact an interesting data suggesting that during therapeutic application there will be a lower impact in the cell viability of healthy cells. Cytotoxicity, genotoxicity, and hematotoxicity were not observed in vivo. These tests on the biological and toxicological effects of andiroba oil and nanostructured oil are still initial ones but will give a direction to future application in cosmetics and/or the development of new phytotherapics.

Various species and lineages that until recently were identified as Microsternarchus bilineatus (Hypopomidae, Gymnotiformes) have a widespread distribution in the Amazon and Orinoco River basins and across the Guiana shield. Recent molecular studies show five distinct lineages for Microsternarchus from different localities. These results suggest that this previously monotypic genus actually consists of more than one species. Here, we describe the karyotype of M. aff. bilineatus from the Cururutuia River (Bragança, Pará, Brazil). The diploid number of 48 chromosomes (14 meta-submetacentric/34 subtelo-acrocentric) is found for males and females, with an XX/XY sex chromosome system. The nucleolar organizer region is found in the short arm of pair 9. Constitutive heterochromatin occurs in the pericentromeric region of all chromosomes, in the distal region of 3p, 5p, 7p, 8q, 9q, 16q, and Xq, in the interstitial region in 2p, 10q, 11q, and 12q and all along 4p, and in a large block of the Y chromosome. These results indicate extensive karyotype divergence between this population and samples from Igarapé Tarumã Grande (Negro River, Amazonas, Brazil) studied by other researchers. Moreover, despite the diversity of sex chromosome systems found in Gymnotiformes, the XX/XY sex chromosome system of M. aff. bilineatus is the first case of Y chromosome degeneration in this order. The present data are valuable to help understand karyotype evolution in Hypopomidae.

Gymnotus coatesi is a small and rare species of banded knife fish that was originally described by LaMonte in 1935, found along the main stretch of the Amazon River. There is no described cytogenetic data on this species. We analyzed the karyotype of five specimens of G. coatesi collected from Cururutuia Stream in Bragança, Pará, Brazil. The obtained diploid number is 50 and the karyotypic formula is 24 m/sm +26 st/a. The constitutive heterochromatin is DAPI positive and distributed mainly in the centromeric and pericentromeric regions of the chromosomes. Ag-nucleolus organizer regions staining showed nine active sites. The 5S rDNA probe hybridized chromosome pair 17 in the interstitial part of the long arm. Fluorescence in situ hybridization (FISH) with telomeric probes revealed signals only at terminal regions of the chromosomes. The 18S rDNA probe hybridized to 21 sites, and these signals colocalized with the telomeric sequences. This relatively high number of 18S rDNA sites may reflect gene duplication mediated by transposable elements. These results indicate that although the diploid number of G. coatesi is within the range previously observed for other members of the genus, various karyotypic characteristics distinguish G. coatesi from the other species of the genus and members of the Gymnotiform order.

Crenicichla (Cichliformes, Cichlidae) present a highly conserved diploid number 2n=48 with fundamental numbers varying between 52 and 62. We analyzed four species in order to investigate the role of repetitive DNA in chromosome evolution in the genus. Crenicichla johanna , Crenicichla cf. saxatilis and Crenicichla cf. regani have 2n=48 (8 m/sm and 40st/a) and FN=56, while Crenicichla sp. ‘Xingu I’ has 2n=48 (48 st/a) and FN=48. Different patterns of constitutive heterochromatin distribution were observed including pericentric, interstitial and whole arm C bands. A single chromosome bears 18S rDNA clusters in most species, except C. johanna , where population variation exists in terms of the quantity and distribution of clusters and their association with interstitial telomeric sequences. All species showed hybridization of 5S rDNA sequences in an interstitial region on an acrocentric chromosome pair. The karyotypic differences and maintenance of the diploid number supports chromosome evolution mediated by inversions in Crenicichla . The telomeric and 18S rDNA sequence association in various chromosomes of C. johanna are proposed to represent hotspots for breakage, favoring intra-chromosomal rearrangements. The results suggest that repetitive sequences can contribute to microstructural cytogenetic diversity in Crenicichla .

The Carapa guianensis (andiroba) oil is commonly used by the Amazon population for medicinal purposes. The objective of this study was to determine the genotoxic and antigenotoxic potential of the andiroba oil (AO) and nanoemulsion (AN) using Swiss mice. Therefore, we used the comet assay and micronucleus test. The AO predominant compounds were oleic (39.13%), palmitic (33.22%), and linoleic (16.86%) acids. AN composition obeyed the surfactant/oil ratio of 0.69, and the Tween 80/Span 80 ratio was held at 0.9. Our results showed no cytotoxicity or genotoxicity in the mice treated with AO and AN alone. However, there was a significant reduction in the polychromatic erythrocytes (PCEs) numbers in all groups treated with doxorubicin (DOX), including those pretreated with AO and AN. Thus, the samples tested did not protect against DOX. On the other hand, our results showed a large increase in micronucleus (MN) formation when the mice were treated with DOX alone; these numbers were reduced when the animals were pretreated with AO and AN. The results indicate a protective effect of andiroba on MN formation and show no evidence of genotoxicity in mice.

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