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Cleidocranial dysplasia (CCD) is an autosomal dominant skeletal dysplasia characterized by persistent open skull sutures with bulging calvaria, hypoplasia, or aplasia of clavicles permitting abnormal opposition of the shoulders; wide public symphysis; short middle phalanx of the fifth fingers; and vertebral, craniofacial, and dental anomalies. It is a rare disease, with a prevalence of 1-9/1,000,000, high penetrance, and variable expression. The gene responsible for CCD is the Runt-related transcription factor 2 (RUNX2) gene. We characterize the clinical, genetic, and bioinformatic results of four CCD cases: two cases within Mexican families with six affected members, nine asymptomatic individuals, and two sporadic cases with CCD, with one hundred healthy controls. Genomic DNA analyses of the RUNX2 gene were performed for Sanger sequencing. Bioinformatics tools were used to predict the function, stability, and structural changes of the mutated RUNX2 proteins. Three novel heterozygous mutations (c.651_652delTA; c.538_539delinsCA; c.662T>A) and a previously reported mutation (c.674G>A) were detected. In silico analysis showed that all mutations had functional, stability-related, and structural alterations in the RUNX2 protein. Our results show novel mutations that enrich the pool of RUNX2 gene mutations with CCD. Moreover, the proband 1 presented clinical data not previously reported that could represent an expanded phenotype of severe expression.

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Accurate and reliable measurements of optical properties are crucial for a wide range of industrial and commercial applications. However, external illumination fluctuations can often make these measurements challenging to obtain. This work proposes a new technique based on digital lock-in processing that enables the use of CCD spectrometers in optical spectroscopy applications, even in uncontrolled lighting conditions. This approach leverages digital lock-in processing, performed on each pixel of the spectrometer's CCD simultaneously, to mitigate the impact of external optical interferences. The effectiveness of this method is demonstrated by testing and recovering the spectrum of a yellow LED subjected to other light sources in outdoor conditions, corresponding to a Signal-to-Noise Ratio of -70.45 dB. Additionally, it was possible to demonstrate the method's applicability for the spectroscopic analysis of gold nanoparticles in outdoor conditions. These results suggest that the proposed technique can be helpful for a wide range of optical measurement techniques, even in challenging lighting conditions.

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Canine Cognitive Dysfunction is a neurological condition, that causes dogs to experience a wide variety of clinical signs. On rare occasions the symptoms may be unusual and severe, therefore they reminiscent of another disease. In this case report a 16 year and 8-month-old intact female poodle presented with circling, head pressing, and generalized ataxia. Prior clinical and neurologic examinations indicated the neurolocalisation to be forebrain. Morphometric brain parameters in MRI indicated otherwise. Quantitative MRI parameters such as the ventricle-brain index, interthalamic adhesion thickness, area, and the ratio of the interthalamic adhesion thickness to brain height may aid in the diagnosis of CCD.

A Disfunção Cognitiva Canina é uma condição neurológica, que faz com que os cães apresentem uma grande variedade de sinais clínicos. Em raras ocasiões, os sintomas podem ser incomuns e graves, portanto, lembram outras doenças. Neste relato de caso, uma cadela, da raça poodle, inteira de 16 anos e 8 meses de idade apresentou-se com movimentos circulares, pressão de cabeça e ataxia generalizada. Exames clínicos e neurológicos prévios indicaram que a neurolocalização era prosencéfalo. Os parâmetros morfométricos do cérebro na ressonância magnética indicaram o contrário. Parâmetros quantitativos de RM, como índice ventrículo-cérebro, espessura de adesão intertalâmica, área e a relação entre a espessura de adesão intertalâmica e a altura do cérebro podem auxiliar no diagnóstico de DCC.

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Coloring is one of the most important characteristics in commercial flowers and fruits, generally due to the accumulation of carotenoid pigments. Enzymes of the CCD4 family in citrus intervene in the generation of ß-citraurin, an apocarotenoid responsible for the reddish-orange color of mandarins. Citrus CCD4s enzymes could be capable of interacting with the thylakoid membrane inside chloroplasts. However, to date, this interaction has not been studied in detail. In this work, we present three new complete models of the CCD4 family members (CCD4a, CCD4b, and CCD4c), modeled with a lipid membrane. To identify the preference for substrates, typical carotenoids were inserted in the active site of the receptors and the protein-ligand interaction energy was evaluated. The results show a clear preference of CCD4s for xanthophylls over aliphatic carotenes. Our findings indicate the ability to penetrate the membrane and maintain a stable interaction through the N-terminal α-helical domain, spanning a contact surface of 2250 to 3250 Å2. The orientation and depth of penetration at the membrane surface suggest that CCD4s have the ability to extract carotenoids directly from the membrane through a tunnel consisting mainly of hydrophobic residues that extends up to the catalytic center of the enzyme.

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Dendritic cells (DCs) are the most potent antigen-presenting cells, unique to initiate and coordinate the adaptive immune response. In pigs, conventional DCs (cDCs), plasmacytoid DCs (pDCs), and monocyte-derived DCs (moDCs) have been described in blood and tissues. Different pathogens, such as viruses, could infect these cells, and in some cases, compromise their response. The understanding of the interaction between DCs and viruses is critical to comprehend viral immunopathological responses. Porcine reproductive and respiratory syndrome virus (PRRSV) is the most important respiratory pathogen in the global pig population. Different reports support the notion that PRRSV modulates pig immune response in addition to their genetic and antigenic variability. The interaction of PRRSV with DCs is a mostly unexplored area with conflicting results and lots of uncertainties. Among the scarce certainties, cDCs and pDCs are refractory to PRRSV infection in contrast to moDCs. Additionally, response of DCs to PRRSV can be different depending on the type of DCs and maybe is related to the virulence of the viral isolate. The precise impact of this virus-DC interaction upon the development of the specific immune response is not fully elucidated. The present review briefly summarizes and discusses the previous studies on the interaction of in vitro derived bone marrow (bm)- and moDCs, and in vivo isolated cDCs, pDCs, and moDCs with PRRSV1 and 2.

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Navicula incerta is a marine microalga distributed in Baja California, México, commonly used in aquaculture nutrition, and has been extended to human food, biomedical, and pharmaceutical industries due to its high biological activity. Therefore, the study aimed to optimize culture conditions to produce antioxidant pigments. A central composite experimental design and response surface methodology (RSM) was employed to analyze the best culture conditions. The medium (nitrogen-deficient concentrations), salinity (PSU = Practical Salinity Unity [g/kg]), age of culture (days), and solvent extraction (ethanol, methanol, and acetone) were the factors used for the experiment. Chlorophyll a (Chl a) and total carotenoids (T-Car), determined spectroscopically, were used as the response variables. The antioxidant capacity was evaluated by DPPH• and ABTS•+ radical inhibition, FRAP, and anti-hemolytic activity. According to the overlay plots, the optimum growth conditions for Chl a and T-Car production were the following conditions: medium = 0.44 mol·L-1 of NaNO3, salinity = 40 PSU, age of culture: 3.5 days, and solvent = methanol. The pigment extracts obtained in these optimized conditions had high antioxidant activity in ABTS•+ (86.2-92.1% of inhibition) and anti-hemolytic activity (81.8-96.7% of hemolysis inhibition). Low inhibition (33-35%) was observed in DPPH•. The highest value of FRAP (766.03 ± 16.62 µmol TE/g) was observed in the acetonic extract. The results demonstrated that RSM could obtain an extract with high antioxidant capacity with potential applications in the biomedical and pharmaceutical industry, which encourages the use of natural resources for chemoprevention of chronic-degenerative pathologies.

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The treatment by Advanced Oxidation Processes (AOPs) of wastewater polluted with dyes is of particular interest in the field of environmental engineering, especially for the removal azo-dyes, representing over 50% of the global annual production of dyes. Unfortunately, most azo-dyes are non-biodegradable and can be toxic to aquatic organisms. This is the first data article that applies the methodology of response surface for the optimization of decolorization of an azo-compound using cobalt in a homogeneous medium as the catalyst of a bicarbonate activated hydrogen peroxide (BAP) system which, in turn, is an emerging technology for wastewater treatment. The Response Surface Methodology (RSM) based on a Central Composite Design (CCD) was used to evaluate and optimize the influence of three experimental variables (stoichiometric dosage of H2O2, molar ratio H2O2/NaHCO3 and cobalt concentration) on the decolorization of Ponceau 4R. Reactions were performed at 25 °C, pH 8.3 with a reaction time of 2 h. Analysis of variance (ANOVA) showed values of R2 and adjusted-R2 of 0.9815 and 0.9648, and experimental data were fit to a second-order regression model. The optimal conditions to achieve a maximum decolorization (96.31%) of a Ponceau 4R aqueous solution of 20 mg/l were: 4.73 times stoichiometric dosage of H2O2, molar ratio H2O2/NaHCO3 of 1.70 and cobalt concentration of 11.16 µM. Under the optimal reaction conditions, the influence of temperature (20, 25, 30 and 35 °C) on decolorization was evaluated and data were adjusted to second order kinetics. To verify the efficiency of the BAP system on the decolorization of Ponceau 4R, under the optimal conditions of reaction, UV-Vis spectra, at different reaction times, were measured. Additionally, blank experiments in order to evaluate the effect of individual factors in the Ponceau 4R decolorization, using BAP system, were carried out. Data showed that the Co(II)-NaHCO3-H2O2 system is a suitable technology for the decolorization of azo-dyes aqueous solutions.

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The coloration of Citrus fruits is related with the concentration of carotenoids, isoprenoid pigments of 40 carbon atoms (C40). Rodrigo et al. and Ma et al. reported a CCD4-type citrus dioxygenase responsible for the generation of C30 apocarotenoids providing a reddish-orange pigmentation to the peel of many mandarins and oranges. Among them, CCD4b was the first case described of a dioxygenase that cleaves carotenoids C40 in the double bond 7', 8' or 7, 8, generating ß-citraurin or 8-ß-apocarotenal. Here we report the three-dimensional structures of CCD4a and CCD4b, modeled by sequence homology (2BIW) and validated by molecular dynamics (MD). Docking calculations were performed in CCD4a and CCD4b structures with thousands of rotated initial carotenoid conformations and all the possible poses in the active site were found. The interaction energy was measured by means of ASE scoring, Amber99 refinement and London ΔG rescoring. For the case of CCD4a model, the results showed London ΔG score of -19, -17 and -15 kcal/mol for zeaxanthin, ß-cryptoxanthin and ß-carotene, respectively. The same sequence in the estimated interaction strength for the three ligands was obtained using MD. The interaction energy of CCD4b indicated that, in agreement with experimental data, zeaxanthin and ß-cryptoxanthin could be cleaved by the enzyme, ß- and α-carotene have chances to be oxidized and lycopene has not good interaction energy to be predicted as substrate. These findings will be discussed considering the potential in vivo substrates and products, and the physiological role in Citrus fruits. Communicated by Ramaswamy H. Sarma.

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According to Quality by Design (QbD) concept, quality should be built into product/method during pharmaceutical/analytical development. Usually, there are many input factors that may affect quality of product and methods. Recently, Design of Experiments (DoE) have been widely used to understand the effects of multidimensional and interactions of input factors on the output responses of pharmaceutical products and analytical methods. This paper provides theoretical and practical considerations for implementation of Design of Experiments (DoE) in pharmaceutical and/or analytical Quality by Design (QbD). This review illustrates the principles and applications of the most common screening designs, such as two-level full factorial, fractionate factorial, and Plackett-Burman designs; and optimization designs, such as three-level full factorial, central composite designs (CCD), and Box-Behnken designs. In addition, the main aspects related to multiple regression model adjustment were discussed, including the analysis of variance (ANOVA), regression significance, residuals analysis, determination coefficients (R2, R2-adj, and R2-pred), and lack-of-fit of regression model. Therefore, DoE was presented in detail since it is the main component of pharmaceutical and analytical QbD.

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Com o aumento na expectativa de vida dos animais de companhia há um maior aparecimento de doenças neurodegenerativas, como a Disfunção Cognitiva Canina (DCC). A DCC apresenta diversas semelhanças à Doença de Alzheimer (DA) presente em seres humanos. Entre estas semelhanças é possível citar o nível reduzido de acetilcolina, o que justifica a busca por tratamentos que tenham como foco, a inibição da acetilcolinesterase (AChE), enzima responsável pela degradação deste neurotransmissor. Neste trabalho foi avaliado o potencial inibitório dos chás de maçã, de hortelã e de chá verde sobre a atividade de AChE presente em cérebro de cães. O chá verde foi o chá mais eficaz na inibição desta enzima, apresentando uma IC50 de aproximadamente 1.7 mg/mL. A quercetina, flavonol que segundo a literatura está presente no chá verde e ausente nos chás de maçã e hortelã, pode ser um dos constituintes responsáveis por esta inibição. Desta forma, também foi avaliado o potencial anticolinesterásico da quercetina em cérebro canino, obtendo-se uma IC50 de 0,41 mM. Uma vez que a quercetina já tem sido utilizada na medicina veterinária para outros fins, o potencial anticolinesterásico deste flavonol em cérebro canino mostra-se de grande relevância para os estudos da DCC.

Canine Cognitive Dysfunction (CCD) is a neuodegenerative disease with very similar characteristics to humans Alzhemer’s disease. Currently, the increase of pets life expectancy leads to the more frequent appearance of neurodegenerative diseases in these species. Among the main hypotheses for the appearance of CCD has the reduction of acetylcholine levels in the synaptic cleft. This hypothesis justifies many researches that search CCD treatment based on acetylcholinesterase (AChE) inhibition, an acetylcholine degrading enzyme. In this work, inhibitory potential of some commecial teas (apple tea, peppermint tea and green tea) toward brain AChE activity of dogs was evaluated. Green tea is tea more effective to inhibit this enzyme, getting a IC 50 of approximately 1.7 mg / mL. Quercetin, flavonol which according to the literature is present in green tea and absent in apple and peppermint teas, may be one of the molecules that contribute to AChE inhibition. In this way, quercetn anticholinesterase potential in canine brain was also evaluated, obtaining an IC 50 of 0.41 mM. Since quercetin has already been used in veterinary medicine for other fins, the anticholinesterase potential of this flavonol toward canine brain is of great relevance for CCD studies.

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RESUMO

Com o aumento na expectativa de vida dos animais de companhia há um maior aparecimento de doenças neurodegenerativas, como a Disfunção Cognitiva Canina (DCC). A DCC apresenta diversas semelhanças à Doença de Alzheimer (DA) presente em seres humanos. Entre estas semelhanças é possível citar o nível reduzido de acetilcolina, o que justifica a busca por tratamentos que tenham como foco, a inibição da acetilcolinesterase (AChE), enzima responsável pela degradação deste neurotransmissor. Neste trabalho foi avaliado o potencial inibitório dos chás de maçã, de hortelã e de chá verde sobre a atividade de AChE presente em cérebro de cães. O chá verde foi o chá mais eficaz na inibição desta enzima, apresentando uma IC50 de aproximadamente 1.7 mg/mL. A quercetina, flavonol que segundo a literatura está presente no chá verde e ausente nos chás de maçã e hortelã, pode ser um dos constituintes responsáveis por esta inibição. Desta forma, também foi avaliado o potencial anticolinesterásico da quercetina em cérebro canino, obtendo-se uma IC50 de 0,41 mM. Uma vez que a quercetina já tem sido utilizada na medicina veterinária para outros fins, o potencial anticolinesterásico deste flavonol em cérebro canino mostra-se de grande relevância para os estudos da DCC.(AU)

Canine Cognitive Dysfunction (CCD) is a neuodegenerative disease with very similar characteristics to humans Alzhemers disease. Currently, the increase of pets life expectancy leads to the more frequent appearance of neurodegenerative diseases in these species. Among the main hypotheses for the appearance of CCD has the reduction of acetylcholine levels in the synaptic cleft. This hypothesis justifies many researches that search CCD treatment based on acetylcholinesterase (AChE) inhibition, an acetylcholine degrading enzyme. In this work, inhibitory potential of some commecial teas (apple tea, peppermint tea and green tea) toward brain AChE activity of dogs was evaluated. Green tea is tea more effective to inhibit this enzyme, getting a IC 50 of approximately 1.7 mg / mL. Quercetin, flavonol which according to the literature is present in green tea and absent in apple and peppermint teas, may be one of the molecules that contribute to AChE inhibition. In this way, quercetn anticholinesterase potential in canine brain was also evaluated, obtaining an IC 50 of 0.41 mM. Since quercetin has already been used in veterinary medicine for other fins, the anticholinesterase potential of this flavonol toward canine brain is of great relevance for CCD studies.(AU)

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Se caracterizaron los extractos etanólicos de hojas y cortezas de 13 especies de la familia Lauraceae mediante cromatografía en capa fina de dos dimensiones (2D-CCD). Los datos posteriores se analizaron mediante técnicas de análisis estadístico multivariado (cluster y análisis de componentes principales (PCA)). Lo anterior permitió hacer una distinción entre los extractos obtenidos de diferentes partes de la planta (hojas y cortezas). Se observó, además, que la metodología usada es capaz de diferenciar entre extractos obtenidos a partir de especies de Lauraceae y los de otras familias de plantas.

Leaves and barks ethanolic extracts from 13 Lauraceae species were characterized through two-dimensional thin layer chromatography (2D-TLC). The subsequent data was analized through multivariate statistical analysis techniques (cluster analysis and principal components analysis (PCA)). This allowed to do a distinction between extracts obtained from different parts of the plant (leaves and bark). In addition, it was observed that the implemented methodology is able to differentiate between extracts obtained from Lauraceae species and some obtained from other plant families.

Caracterizaram-se os extratos etanólicos de folhas e casca obtidos a partir de espécies da família Lauraceae por cromatografia em camada fina em duas dimensões (2D-CCF). Os dados obtidos foram analisados utilizando técnicas de análise estatística multivariada tipo análise de cluster e análise de componentes principais (PCA). As técnicas estadísticas permitiram fazer uma distinção entre os extratos obtidos a partir de diferentes partes da planta (folhas e casca). Além disso, observou-se que o método utilização é capaz de diferenciar entre os extratos provenientes de espécies de Lauraceae daqueles obtidos a partir de outras famílias de plantas.

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The growing trade market in herbal medicines is aworldwide phenomenon due to several factors, fromthe high cost of manufactured drugs to mere fashion.In this study, we tested the quality of pharmaceuticalproducts based on Hypericum perforatum L. (Eng. ?StJohn?s Wort?) marketed in Divinópolis City, State ofMinas Gerais, Brazil. Samples of products labeled H.perforatum were purchased at three commercial stores,at various times during one year. We assessed thefollowing items: pharmacobotanical characteristics,humidity, total ash, hypericin content and thin layerchromatographic profile. Close inspection showed that4 of the 12 samples sold as H. perforatum products werefrom a different plant from that advertised, representinga forgery in which the true species was replaced byAgeratum sp. Besides, the samples did not entirely meetthe legal requirements for herbal medicine. Therefore,there is a need to strengthen pharmaceutical vigilance,to ensure that herbal products are suitable for publicuse.

O crescente comércio de plantas medicinais e fitoterápicos ocorre em todo o mundo em razão de diversos fatores, como o alto custo dos medicamentos industrializados ou modismo. Neste trabalho verificou-se a qualidade de produtos farmacêuticos contendo Hypericum perforatum L. comercializados em Divinópolis, Minas Gerais, Brasil. Adquiriram-se amostras de H. perforatum em três estabelecimentos distintos em diferentes épocas ao longo de um ano. Avaliaram-se os seguintes itens: aspectos farmacobotânicos, umidade, cinzas totais, doseamento de hipericina e cromatografia em camada delgada. Os resultados demonstraram que quatro amostras comercializadas como H. perforatum eram produtos diferentes daqueles anunciados, caracterizando uma falsificação e substituição da espécie verdadeira por Ageratum sp. Além disso, as amostras não cumpriram totalmente os requisitos exigidos pela legislação. Portanto, faz-se necessário reforçar a vigilância farmacêutica, buscando garantir produtos de qualidade adequados ao uso da população.

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Various cultivation parameters were optimized for the production of extra cellular protease by Brevibacterium linens DSM 20158 grown in solid state fermentation conditions using statistical approach. The cultivation variables were screened by the Plackett-Burman design and four significant variables (soybean meal, wheat bran, (NH4)2SO4 and inoculum size were further optimized via central composite design (CCD) using a response surface methodological approach. Using the optimal factors (soybean meal 12.0g, wheat bran 8.50g, (NH4)2SO4) 0.45g and inoculum size 3.50%), the rate of protease production was found to be twofold higher in the optimized medium as compared to the unoptimized reference medium.

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Various cultivation parameters were optimized for the production of extra cellular protease by Brevibacterium linens DSM 20158 grown in solid state fermentation conditions using statistical approach. The cultivation variables were screened by the Plackett-Burman design and four significant variables (soybean meal, wheat bran, (NH4)2SO4 and inoculum size were further optimized via central composite design (CCD) using a response surface methodological approach. Using the optimal factors (soybean meal 12.0g, wheat bran 8.50g, (NH4)2SO4) 0.45g and inoculum size 3.50%), the rate of protease production was found to be twofold higher in the optimized medium as compared to the unoptimized reference medium.

RESUMO

Various cultivation parameters were optimized for the production of extra cellular protease by Brevibacterium linens DSM 20158 grown in solid state fermentation conditions using statistical approach. The cultivation variables were screened by the Plackett-Burman design and four significant variables (soybean meal, wheat bran, (NH4)2SO4 and inoculum size were further optimized via central composite design (CCD) using a response surface methodological approach. Using the optimal factors (soybean meal 12.0g, wheat bran 8.50g, (NH4)2SO4) 0.45g and inoculum size 3.50%), the rate of protease production was found to be twofold higher in the optimized medium as compared to the unoptimized reference medium.

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Plantas medicinais são utilizadas mundialmente como uma das principais formas de cuidado primário de saúde. No entanto, a literatura indica que muitas espécies podem apresentar composição química variável, toxicidade ou difícil identificação. O objetivo do presente estudo foi obter critérios úteis para o controle de qualidade farmacognóstico das principais espécies vegetais de uso popular no estado do Rio Grande do Norte, utilizando metodologias de análise botânica (morfodiagnose macro e microscópica) e química (triagem fitoquímica e cromatografia em camada delgada), evitando assim adulterações ou uso inadequado dessas plantas medicinais no estado. No total, sete espécies foram analisadas Acmella oleracea, Chenopodium ambrosioides, Lippia alba, Mentha piperita, Ocimum gratissimum, Peumus boldus e Rosmarinus officinalis. Diversos marcadores botânicos e fitoquímicos foram identificados, contribuindo dessa forma para a correta identificação destas espécies de plantas medicinais importantes no estado do Rio Grande do Norte.

Medicinal plants are worldwide used as one of the main forms of primary healthcare. However, the literature indicates that many species may have variable chemical composition, toxicity, or even difficult identification. The aim of this study was to obtain useful criteria for pharmacognostic quality control of the main plant species of popular use in the state of Rio Grande do Norte, using methods of botanical (macro and microscopic morphodiagnosis) and chemical (phytochemical screening and thin-layer chromatography) analysis, thus preventing adulteration or inappropriate use of these medicinal plants in the state. In total, seven species were analyzed Acmella oleracea, Chenopodium ambrosioides, Lippia alba, Mentha piperita, Ocimum gratissimum, Peumus boldus and Rosmarinus officinalis. Several botanical and phytochemical markers were identified, thereby contributing to the correct identification of these important medicinal plant species in the state of Rio Grande do Norte.

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Se realizó la validación de un método analítico con vistas a su aplicación en los estudios de estabilidad de las futuras formulaciones de supositorios de naproxeno para uso infantil y adulto. Se determinaron los factores que más influyeron en la estabilidad del naproxeno; la mayor degradación ocurrió en el medio ácido, oxidante y por acción de la luz. Se determinó la posibilidad de formación de ésteres entre el grupo carboxilo libre presente en el naproxeno y el monoestearato de glicerilo presente en la base como una de las vías de degradación en la nueva formulación; se obuvieron resultados satisfactorios. Se desarrolló un método por cromatografía en capa delgada y se seleccionaron las mejores condiciones cromatográficas. Se emplearon placas de sílica gel GF254 y revelador ultravioleta a 254 nm. Se evaluaron 3 sistemas de disolventes entre los cuales el A, compuesto por: acético glacial:tetrahidrofurano:tolueno (3:9:90 v/v/v), permitió una adecuada resolución entre el analito y los posibles productos de degradación, con un límite de detección de 1 µg. La aplicación del método propuesto se limitó a la identificación de los posibles productos de degradación solo con fines cualitativos y no como prueba límite. El método fue suficientemente sensible y selectivo para aplicarlo con el objetivo para el cual fue diseñado, según los resultados obtenidos en la validación

The validation of an analytical method was carried out to be applied to the stability studies of the future formulations of naproxen suppositories for infant and adult use. The factors which mostly influenced in the naproxen stability were determined, the major degradation occurred in oxidizing acid medium and by action of light. The possible formation of esters between the free carboxyl group present in naproxen and the glyceryl monoestereate present in the base was identified as one of the degradation paths in the new formulation. The results were satisfactory. A thin-layer chromatography-based method was developed as well as the best chromatographic conditions were selected. GF254 silica gel plates and ultraviolet developer at 254 nm were employed. Three solvent systems were evaluated of which A made up of glacial acetic: tetrahydrofurane:toluene (3:9:90 v/v/v)allowed adequate resolution between the analyte and the possible degradation products, with detection limit of 1 µg. The use of the suggested method was restricted to the identification of possible degradation products just for qualitative purposes and not as final test. The method proved to be sensitive and selective enough to be applied for the stated objective, according to the validation results

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Cleidocranial dysplasia (CCD) is an autosomal dominant human skeletal disorder comprising hypoplastic clavicles, wide cranial sutures, supernumerary teeth, short stature, and other skeletal abnormalities. It is known that mutations in the human RUNX2 gene mapped at 6p21 are responsible for CCD. We analyzed the mutation patterns of the RUNX2 gene by direct sequencing in six Taiwanese index cases with typical CCD. One of the patients was a familial case and the others were sporadic cases. Sequencing identified four mutations. Three were caused by single nucleotide substitutions, which created a nonsense (p.R391X), two were missense mutations (p.R190W, p.R225Q), and the forth was a novel mutation (c.1119delC), a one-base deletion. Real time quantitative PCR adapted to determine copy numbers of the promoter, all exons and the 3'UTR region of the RUNX2 gene detected the deletion of a single allele in a sporadic case. The results extend the spectrum of RUNX2 mutations in CCD patients and indicate that complete deletions of the RUNX2 gene should be considered in those CCD patients lacking a point mutation detected by direct sequencing.