Embryo development is impaired by sperm mitochondrial-derived ROS

Background Basal energetic metabolism in sperm, particularly oxidative phosphorylation, is known to condition not only their oocyte fertilising ability, but also the subsequent embryo development. While the molecular pathways underlying these events still need to be elucidated, reactive oxygen species (ROS) could have a relevant role. We, therefore, aimed to describe the mechanisms through which mitochondrial activity can influence the first stages of embryo development. Results We first show that embryo development is tightly influenced by both intracellular ROS and mitochondrial activity. In addition, we depict that the inhibition of mitochondrial activity dramatically decreases intracellular ROS levels. Finally, we also demonstrate that the inhibition of mitochondrial respiration positively influences sperm DNA integrity, most likely because of the depletion of intracellular ROS formation. Conclusion Collectively, the data presented in this work reveals that impairment of early embryo development may result from the accumulation of sperm DNA damage caused by mitochondrial-derived ROS.

Intoxicación por cianuro posterior a infusión prolongada de Nitroprusiato de sodio: a propósito de un caso / Cyanide poisoning following prolonged infusion of Sodium Nitroprusside: A case report / Intoxicação por cianeto após infusão prolongada de nitroprussiato de sódio: a propósito de um caso

El nitroprusiato de sodio fue aprobado desde 1974 como medicamento para las crisis hipertensivas. Desde entonces existe controversia sobre el potencial tóxico del mismo. La toxicidad por cianuro es una complicación poco común del nitroprusiato de sodio, que puede ser difícil de diagnosticar en pacientes críticamente enfermos. Dentro de los factores de riesgo está la utilización de infusiones prolongadas y dosis altas, por lo que se sugiere que la dosis no exceda 2 µg/kg/min. A continuación, se reporta el caso de un paciente de 49 años que ingresa a un servicio de urgencias después de un síndrome cerebrovascular y una emergencia hipertensiva que requirió infusión de nitroprusiato de sodio, seguida de signos y síntomas, los cuales sugirieron toxicidad. Entre dichos síntomas predominaban los cambios neurológicos, por lo cual fue manejado con hiposulfito de sodio y hubo una buena respuesta. El objetivo de este reporte de caso es sensibilizar al personal de salud sobre la necesidad de un uso adecuado del nitroprusiato y sobre la importancia de sospechar toxicidad en paciente con factores de riesgo.

Sodium Nitroprusside has been approved since 1974 as a medication for hypertensive crises. However, there has been ongoing controversy regarding its potential toxicity. Cyanide toxicity is a rare complication of Sodium Nitroprusside, which can be challenging to diagnose in critically ill patients. Risk factors include prolonged infusions and high doses; hence it is suggested that the dose should not exceed 2 µg/kg/min. Here, we present the case of a 49-year-old patient who entered the emergency department after a cerebrovascular event and a hypertensive emergency that required Sodium Nitroprusside infusion, followed by signs and symptoms suggestive of toxicity. Neurological changes were predominant among these symptoms, and the patient was managed with sodium thiosulfate, resulting in a good response. The objective of this case report is to raise awareness among healthcare professionals about the need for appropriate use of Sodium Nitroprusside and the importance of suspecting toxicity in patients with risk factors.

O nitroprussiato de sódio foi aprovado desde 1974 como medicamento para crises hipertensivas. Desde então tem havido controvérsia sobre o seu potencial tóxico. A toxicidade por cianeto é uma complicação rara do nitroprussiato de sódio, que pode ser difícil de diagnosticar em pacientes gravemente enfermos. Entre os fatores de risco está o uso de infusões prolongadas e altas doses, por isso sugere-se que a dose não ultrapasse 2 µg/kg/min. Relata-se a seguir o caso de uma paciente de 49 anos admitida em pronto-socorro após síndrome cerebrovascular e emergência hipertensiva com necessidade de infusão de nitroprussiato de sódio acompanhada de sinais e sintomas sugestivos de toxicidade. Dentre esses sintomas, predominaram as alterações neurológicas, para as quais foi tratado com hipossulfito de sódio e apresentou boa resposta. O objetivo deste relato de caso é sensibilizar os profissionais de saúde sobre a necessidade do uso adequado do nitroprussiato e sobre a importância de suspeitar de toxicidade em pacientes com fatores de risco.

Quantitative determination of cyanide residues in polysaccharide-protein conjugate vaccines based on flow injection analysis(FIA) with amperometric detector / 中国生物制品学杂志

@#Objective To develop and verify a method for determination of cyanide residues in polysaccharide-protein conjugate vaccines based on flow injection analysis(FIA)with amperometric detector.Methods After removing macromolecular substances in the samples by ultrafiltration,the cyanide residues were determined by a 3700 automatic chemical analyzer with the injection time of 35 s,injection volume of 200 μL,pump speed of 40%,sample cycle time of 140 s,ultraviolet wavelength of 312 nm and an Ampere detector.The developed method was verified for the specificity,matrix effect,linear range,limit of detection(LOD),limit of quantitation(LOQ),accuracy,presicion and stability.The cyanide residues in the polysaccharide derivative bulk(13 batches)and conjugate bulk(21 batches)of Haemophilus influenzae type b conjugate vaccine and group A and C meningococcal conjugate vaccine produced by 5 manufacturers were determined by the developed method.Results Blank samples showed no interference to detection;The recoveries of the matrix effector solution of Haemophilus influenzae type b conjugate vaccine polysaccharide derivative and conjugate bulk,group A and C meningococcal conjugate vaccine polysaccharide derivative and conjugate bulk were 97.4%,102.4%,96.8% and 99.8% respectively,with all CV values less than 15%;In the range of 0.312 5 ～ 80 ng/mL,cyanogroup concentration showed a good linear relationship with peak height with a regression equation of y = 133.13 x + 57.556,R2= 0.999 1;The LOD was 0.2 ng/mL,and the LOQ was 0.6 ng/mL.The average recoveries of the control solution were 108.9%,106.5%,103.5% respectively with RSD value of 6.4% in the groups added with 5,10 and 20 ng/mL of cyanogroup and the CV values of precision verification were all less than 15%;The average concentrations of the control solution injected 20 times continuously were 76,38,18and 5 ng/mL with all CV values less than 15%,when cyanogroup concentration was 80,40,20 and 5 ng/mL,respectively.Cyanide residues were detected in 13 batches of derivative bulk samples,while not in 2 batches of 21 batches of polysaccharide conjugate bulk samples,and were detected in the others.Conclusion The developed method had good accuracy,precision and stability,which might be applied to the quantitative determination of cyanide residues in polysaccharideprotein conjugate vaccines.

Determination of cyanide in whole seeds and brans of linseed (Linum usitatissimum Linn) by molecular spectrophotometry

Abstract The addition of linseed (Linum usitatissimum Linn) in the diet, as a functional food, has increased over the years. However, it possesses cyanogenic glycosides. This study aimed to quantify and compare cyanide concentration in whole seed and bran of brown and golden types to establish a safe limit of intake. Three commercial labels, from brown and golden whole seed types (Ab, Ag, Bb, Bg, Cb and Cg), and six commercial labels of brown and golden bran (1b, 2g, 3g, 4b, 5g, and 6b), were selected, totalizing twelve samples. Total cyanide concentration was quantified by a colorimetric method employing alkaline picrate, after acid hydrolysis. The whole seed cyanide values were between 348.4 and 473.20 µg/g and the bran cyanide values were between 459.53 and 639.35 µg/g. The analyzed bran presented increased cyanide concentrations than the whole seeds with no differences between brown and golden types. Food able to produce cyanide less than 90 µg/kg body weight, daily, is considered secure for consumption. Considering this limit and analyzed samples, it is safe to eat approximately two tablespoons of seeds or one tablespoon of bran. These results point out the importance of cyanide amount daily intake information to be in linseed packaging, to ensure secure consumption

Screening for neurocognitive deficits in adult populations reliant on toxic cassava as the main source of food

Context and objective. Chronic dietary reliance on improperly processed cyanogenic toxic cassava is widespread in sub-Saharan Africa. The objective of the present study was to screen for neurocognition impairments and daily-life functioning in adults with dietary dependency on cyanogenic cassava as the main source of food. Methods. A cross-sectional design enrolled heads of households (in couples) in the rural district of Kahemba, Democratic Republic of Congo. Participants were screened for neurocognitive impairments using the Community Screening Interview for Dementia (CSID). Detailed neuropsychiatric evaluations were performed and disease entities classified according to the Diagnostic and Statistical Manual of Mental Disorders (DSM IV) criteria when applicable. Cassava cyanogenic exposure was ascertained by urinary concentrations of thiocyanate (SCN). Regression models were used to identify predictors of CSID performance at the 0.05 significance level. Results. For hundred and six households (203 couples, mean age 38.4 ± 11. 4 years) were involved. One hundred thirty-six subjects (33.5 %) [69 women and 67 men, mean age 39 ± 14.4 years)] and 13 (3.2 %) [7 women and 6 men, mean age: 32 ± 2.6 years] fulfilled the criteria for mild cognitive impairment (MCI) and Major Neurocognitive disorder (MNCD), respectively. The overall mean urinary concentration of SCN was 949.5+518.3 mol/l after adjusting Context and objective. Chronic dietary reliance on improperly processed cyanogenic toxic cassava is widespread in sub-Saharan Africa. The objective of the present study was to screen for neurocognition impairments and daily-life functioning in adults with dietary dependency on cyanogenic cassava as the main source of food. Methods. A cross-sectional design enrolled heads of households (in couples) in the rural district of Kahemba, Democratic Republic of Congo. Participants were screened for neurocognitive impairments using the Community Screening Interview for Dementia (CSID). Detailed neuropsychiatric evaluations were performed, and disease entities classified according to the Diagnostic and Statistical Manual of Mental Disorders (DSM IV) criteria when applicable. Cassava cyanogenic exposure was ascertained by urinary concentrations of thiocyanate (SCN). Regression models were used to identify predictors of CSID performance at the 0.05 significance level. Results. For hundred and six households (203 couples, mean age 38.4 ± 11. 4 years) were involved. One hundred thirty-six subjects (33.5 %) [69 women and 67 men, mean age 39 ± 14.4 years)] and 13 (3.2 %) [7 women and 6 men, mean age: 32 ± 2.6 years] fulfilled the criteria for mild cognitive impairment (MCI) and Major Neurocognitive disorder (MNCD), respectively. The overall mean urinary concentration of SCN was . for age, gender, nutritional status, and history of konzo, neurocognition domain-specific deficits were independently associated with either hypertension or USCN (350mol / l incremental increase in excretion Functional impairments in daily-life activities increased as subjects poorly performed at the CSID screening (Spearman r = - .2, p < 0.01). Conclusion. Neurocognitive deficits in adults are common in Congolese adults relying on cyanogenic cassava as the main source of food. Our study findings warrant further studies to elucidate the overall lifespan brain/behavioral burden and mechanisms of cassava toxicity among adults with dietary dependency on cyanogenic cassava as the main source of food

Horacio Quiroga y su relación con los venenos / Horacio Quiroga and his relationship with poisons

Resumen La vida y obra del literato Horacio Quiroga ha estado signada por venenos en varias oportunidades. Profundo conocedor de la selva misionera, plasmó en sus personajes el conocimiento de ponzoñas y venenos que poseía su flora y su fauna. En cuanto a su vida personal y la de sus familiares, el veneno estuvo presente hasta sus últimos días. Se describen en este trabajo tanto los venenos detallados en su obra como también el desenlace de su suicidio con cianuro de potasio.

Abstract The life and the work of the writer Horacio Quiroga has been marked by poisons and venoms on several occasions. Connoisseur of the flora and fauna of the Misiones jungle, he reflected in his characters the knowledge of poisons and venoms that he possessed. As for his personal life and that of his family, the poison was present until his last days. Both the poisons and venoms detailed in his work as well as the outcome of his suicide with potassium cyanide are detailed in this work.

Studies on the enzymatic specificity of diguanylate cyclases domains and the production of new second messengers / Estudos na especificidade enzimática de diguanilato ciclases e na produção de novos segundos mensageiros

In the first chapter, studies on substrate recognition and enzymatic activity of GGDEF domains are presented. Many proteins containing GGDEF domains are diguanylate cyclases (DGCs, EC 2.7.7.65), enzymes that catalyze the conversion of 2 GTP molecules into the second messenger c-di-GMP in prokaryotes. This molecule is primarily implicated in the transition between motile and sessile lifestyles, as well several other phenotypes. Redundancy and diversity of GGDEF domain sequences in many bacterial genomes raises the possibility that other enzymatic functions may yet be discovered. To test this hypothesis, i) the effect of point mutations on the structure and enzymatic activity of GGDEF domains is analyzed, ii) the enzymatic specificity of wild-type GGDEF domains from different proteins is also tested, and iii) when non-canonical products are detected, enzymatic models are studied to understand its preferential production. The principal results obtained from these studies are as follows. Seven mutants of the DGC PleD (a GGDEF containing-protein from Caulobacter crescentus) were constructed and the crystallographic structure of two of them was solved, showing that they are unlikely to bind the guanine moiety in its active site. Additionally, five mutants of XAC0610, another DGC from Xanthomonas citr, were constructed and their substrate specificities were evaluated. None of those mutants were able to use ATP as a substrate. Finally, seven different GGDEF domain-containing DGCs from different sources were expressed and purified and their enzymatic specificities were tested with several nucleotide triphosphates. One enzyme, GSU1658 from Geobacter sulfurreducens was particularly promiscuous and shown to produce c-di-GMP, c-di-AMP, c-di-IMP, c-di-2´dGMP, cGAMP, c-GIMP, and c-AIMP. Interestingly, XAC0610 was able to recognize 2´dGTP as substrate. Analysis of enzyme kinetics of XAC0610 in presence of 2´dGTP and/or GTP showed the preferential formation of the hybrid linear product pppGp2´dG. The second chapter present studies on cyanide metabolism in Bacillus with focus on the cyanide dihydratase of Bacillus safensis. Cyanide is widely used in industries due to its high affinity for metals. This same ability confers potent toxicity to this compound. Thus, industries must reduce the cyanide concentration from wastewater before its final disposal. Physical, chemical, and biological methods have been developed to achieve this goal, but knowledge about metabolic pathways and the biology of enzymes involved in cyanide degradation is still scarce. Here, the isolation of a Bacillus safensis strain from mine tailings in Peru is described. Classification of this strain was done through a comparative analysis of 132 core genomes of strains from the Bacillus pumilus group. Sequence analysis determined that a cyanide dihydratase (CynD, EC 3.5.5.1)) encoded in the genome of the isolated strain was likely the enzyme responsible for cyanide degradation. Confirmation of the cyanide degrading activity of CynD from this strain was achieved by cloning, expression and purification of the enzyme and its enzymatic characterization. CynD from this strain was active up to pH 9 and oligomerization patterns analyzed by SEC-MALS and electron microscopy showed that the enzyme forms large helical structures at pH 8 and smaller structures at higher pHs. Finally, we show that CynD expression is strongly induced in the presence of cyanide. The last two years of graduate studies were carried out in the context of the COVID-19 pandemic. Thanks to the large amount of publicly available genomic data, we were able to carry out studies on the worldwide dynamics of the spread of SARS-CoV-2 mutants forms. In the first year of the pandemic, genomic classification of 171,461 genomes showed the presence of five major haplotypes based on nine mutations. The worldwide distribution and the temporal evolution of frequency of these haplotypes was carefully analyzed. All the haplotypes were identified in the six regions analyzed (South America, North America, Europe, Asia, Africa, and Oceania); however, the frequency of each of them was different in each of these regions. As of September 30, 2020, haplotype 3 (or operational taxonomic unit 3, OTU_3) was the most prevalent in four regions (South America, Asia, Africa, and Oceania). OTU_5 was the most prevalent in North America and OTU_2 in Europe. Temporal dynamics of the haplotypes showed that OTU_1 became nearly extinct after 8 months of pandemic (November 2020). Other OTUs are still present in different frequencies all around the world, while currently generating new variants. Based on their temporal dynamics, a classification scheme of 115 SARS-CoV-2 mutations identified from 1,058,020 SARS-COV-2 genomes was also performed. Three types of temporal dynamics of mutations were identified: i) High-Frequency mutations are characterized by a rapid increase in frequency upon its appearance, ii) medium and iii) low-frequency mutations maintain mid or low-frequencies for several months and can be region-specific. Finally, we performed a correlation analysis of the effective reproduction number (Rt) of SARS-CoV-2 harboring the high-frequency mutation N501Y with the level of control measures adopted in specific jurisdictions. We show that Rt is negatively correlated with the level of control measures in eight of the nine countries analyzed. This negative correlation was similar when we analyzed the Rt of SARS-CoV-2 not-harboring N501Y. Thus, the control measures likely diminish the Rt of both SARSCoV-2 wild-type and N501Y

O presente trabalho está dividido em três capítulos sobre linhas de pesquisa diferentes desenvolvidas pelo autor durante o período de doutorado No primeiro capítulo, são apresentados estudos relacionados ao reconhecimento estrutural de substratos e análise enzimática de domínios GGDEF com atividade diguanilato ciclase (EC 2.7.7.65). As proteínas contendo domínios GGDEF estão relacionados à produção enzimática do segundo mensageiro c-di-GMP, a partir de duas moléculas de GTP, em procariotos. Esta molécula está principalmente envolvida na transição entre os estilos de vida móveis e sésseis, bem como vários outros fenótipos. Redundância e diversidade de sequências de domínio GGDEF aumentam a possibilidade de que outras funções enzimáticas ainda possam ser descobertas. Para testar esta hipótese, i) o efeito de mutações pontuais na estrutura e atividade enzimática dos domínios GGDEF é analisado, ii) a especificidade enzimática de domínios GGDEF de enzimas diferentes também é testada e iii) quando produtos não canônicos são detectados, modelos enzimáticos são estudados para entender sua produção preferencial. Como resultados mais importantes, sete mutantes do PleD (uma proteína contendo GGDEF) foram construídos e a estrutura cristalográfica de dois delas foi resolvida, mostrando que é improvável que eles liguem à porção guanina em seu sítio ativo. Além disso, cinco mutantes da proteína XAC0610 de Xanthomonas citri foram construídos e sua capacidade de usar ATP ou GTP como substrato foi avaliada. Nenhum desses mutantes foi capaz de usar ATP como substrato. Finalmente, sete outras proteínas contendo GGDEF foram purificadas e sua especificidade enzimática foi avaliada com vários trifosfatos de nucleotídeos. Uma enzima promíscua chamada GSU1658 mostrou produzir c-di-GMP, c-di-AMP, c-di-IMP, c-di-2´dGMP, c-GAMP, cGIMP e c-AIMP. Curiosamente, o XAC0610 foi capaz de reconhecer 2´dGTP como substrato. A análise da cinética enzimática de XAC0610 na presença de 2´dGTP e GTP mostrou a formação preferencial do produto linear híbrido pppGp2´dG. O segundo capítulo aborda estudos sobre o metabolismo do cianeto em Bacillus com foco na cianeto dihidratase de Bacillus safensis. O cianeto é amplamente utilizado nas indústrias devido à sua alta afinidade com os metais. Esta mesma capacidade confere toxicidade potente a este composto. Assim, as indústrias têm que reduzir a concentração de cianeto das águas residuais antes de sua disposição final. Métodos físicos, químicos e biológicos têm sido desenvolvidos para atingir esse objetivo, mas o conhecimento sobre as vias metabólicas e a biologia das enzimas envolvidas na degradação do cianeto ainda é escasso. Aqui, é descrito o isolamento de uma cepa de Bacillus safensis de rejeitos de minas no Peru. A classificação desta cepa foi feita através de uma análise comparativa de 132 core genomes de cepas do grupo de Bacillus pumilus. Em seguida, determinamos que uma cianeto dihidratase (CynD, EC 3.5.5.1) codificada no genoma da cepa isolada era provavelmente a enzima responsável pela degradação do cianeto. A confirmação da atividade degradante de cianeto de CynD desta cepa foi feita por clonagem, expressão e purificação da enzima e realização de caracterização enzimática. O CynD desta cepa é ativo até pH 9 e os padrões de oligomerização analisados por SEC-MALS mostraram que a enzima forma longas estruturas helicoidais em pH 8 e estruturas menores enquanto o pH aumenta. Finalmente, foi demonstrado que a expressão de CynD é fortemente induzida na presença de cianeto. Os últimos dois anos do doutorado foram realizados no contexto da pandemia COVID- 19. Vários laboratórios se dedicaram a gerar conhecimento para ajudar no combate à pandemia. Nesta situação e graças à grande quantidade de dados genômicos disponíveis publicamente, estudos sobre a dinâmica das mutações do SARS-CoV-2 foram realizados. No primeiro ano da pandemia, a classificação genômica de 171.461 genomas mostrou a presença de cinco haplótipos principais com base em nove mutações. A distribuição mundial e a mudança de frequência desses haplótipos foram analisadas cuidadosamente. Todos os haplótipos foram identificados nas seis regiões analisadas (América do Sul, América do Norte, Europa, Ásia, África e Oceania); no entanto, a frequência de cada um deles foi diferente em cada uma dessas regiões. Em 30 de setembro de 2020, o haplótipo 3 (ou unidade taxonômica operacional 3, OTU_3) era o mais prevalente em quatro regiões (América do Sul, Ásia, África e Oceania). OTU_5 foi o mais prevalente na América do Norte e OTU_2 na Europa. A dinâmica temporal dos haplótipos mostrou que OTU_1 parece perto da extinção após 8 meses de pandemia (novembro de 2020). Outros OTUs ainda estão presentes em diferentes frequências em todo o mundo, mesmo atualmente gerando novas variantes. Com base em sua dinâmica temporal, um esquema de classificação de 115 mutações SARS-CoV-2 identificadas a partir de 1.058.020 genomas SARS-COV-2 também foi feito. Três tipos de dinâmica temporal de mutações foram identificados: i) Mutações de alta frequência, ii) mutações de média frequência e iii) mutações de baixa frequência. Finalmente, foi analisada a correlação do número de reprodução efetiva (Rt) do SARS-CoV-2 que contém a mutação de alta frequência N501Y com o nível de medidas de controle, mostrando que seu Rt está negativamente correlacionado com o nível de medidas de controle em oito dos nove países analisados. Esta correlação negativa foi semelhante quando foi analisado o Rt de SARS-CoV-2 sem a mutação N501Y. Assim, as medidas de controle provavelmente diminuirão o Rt de SARS-CoV-2 tipo selvagem e N501Y

Intoxicación por cianuro como método suicida: reporte de caso / Cyanide poisoning as a suicide method: case report

Resumen Las intoxicaciones derivan de la presencia en el organismo de un tóxico o veneno, la muerte por intoxicación es una muerte violenta y por tanto requiere de la realización de una autopsia medico legal, la misma puede darse en el contexto de una exposición accidental ya sea en el hogar o laboral o sucitada por un intento de autoeliminación. La intoxicación por cianuro puede ser intencional (suicidio u homicidio) o accidental, los hallazgos en la autopsia medico legal son inespecíficos por lo que son importantes los datos aportados en el informe sobre muerte en investigación, el informe del escenario de muerte en caso de que un médico forense se hiciera presente al mismo y el resultado de los análisis toxicológicos, los cuales actualmente no se realizan en la sección de toxicología del poder judicial. Se realizó una revisión bibliográfica en diferentes bases de datos, de los artículos publicados referentes al tema de los últimos cinco años, con el objetivo de revisar las caracteristicas del químico, el metabolismo y la intoxicación como tal, tanto por sus secuelas como por sus implicaciones letales. Se concluye que para mejorar la pericia médico legal ante casos de intoxicacion por cianuro es fundamental conocer el mecanismo de acción y los posibles hallazgos presentes tanto al examen externo como interno, así como implementar que dicho escrutinio se incluya dentro del listado de sustancias a analizar.

Abstract Poisoning derives from the presence of a toxic substance or poison in the body, death by poisoning is a violent death and requires a legal medical autopsy, it may occur in the context of an accidental exposure at home or work, or caused by an attempt of self-elimination. Cyanide poisoning can be intentional (suicide / homicide) or accidental, the findings in the autopsy are unspecific, so data provided in the report of death in investigation, the report of the death scene (in case a forensic doctor was present) and the result of the toxicological analyzes, which are not currently performed in the toxicology section of the judiciary, are important. A bibliographic review was carried out in different databases of articles published in the last five years on the subject, with the objective of reviewing the chemical characteristics, the metabolism and the intoxication, as well, including their sequels and lethal implications. It is concluded that to improve the medical legal expertise in cases of cyanide poisoning, it is essential to know the mechanism of action and the possible findings in the external and internal examination; and to implement such scrutiny in the list of substances to be analyzed.

Determination of cyanide in cassava starch by continuous flow online distillation / 公共卫生与预防医学

Objective To explore the determination of cyanide in cassava starch by a continuous flow online distillation method. Methods Cassava starch was extracted with sodium hydroxide solution, filtered and then quantitatively analyzed with a continuous flow analyzer. The effects of extraction concentration, ultrasonic time, sample size and separation method on cyanide determination were investigated. Results 2.0 g samples were directly weighed and added to 100 mL 2 g/L sodium hydroxide solution for ultrasonic extraction for 20 min, and filtered with qualitative filter paper. The filtrate was quantitatively determined with a continuous flow analyzer. The linear relationship between cyanide concentration of 0 mg/L and 0.20 mg/L was good, r ≥ 0.999. The detection limit of the method was 0.01 mg/kg, the limit of quantitation was 0.03 mg/kg, the relative standard deviation was 0.26%-1.52%, and the standard addition recovery rate was 90.36%-104.19%. Conclusion The continuous flow online distillation method has the advantages of low detection limit, good repeatability, automatic operation, reduction of manual errors, and high analysis speed, which is beneficial to improve work efficiency, and can be used for batch detection of cyanide in cassava starch.

Determination of Azide Ions in Blood by Pentafluorobenzyl Derivation Followed by GC-MS / 法医学杂志

Objective To establish a method for determination of the azide ions in blood by gas chromatography-mass spectrometry （GC-MS） following pentafluorobenzyl derivatization. Methods A blood sample of 0.2 mL was placed into a 10 mL glass test tube, and the internal standard sodium cyanide, derivatization reagent pentafluorobenzyl bromide and catalyst tetradecyl benzyl dimethyl ammonium chloride were added in turn. After vortex mixing, the mixture was heated with low-power microwave for 3 min. After centrifugation, the organic phase was taken for GC-MS analysis. Results The azide ions in blood had a good linear relationship in the mass concentration range of 0.5 to 20 μg/mL. The lowest detection limit was 0.25 μg/mL and the relative recovery was 91.36%-94.58%. The method was successfully applied to a case of death from sodium azide poisoning. The mass concentration of azide ions in the blood of the dead was 11.11 μg/mL. Conclusion The method developed in this paper has strong specificity and is easy to operate, which is suitable for the rapid detection of azide ions in blood.

Biodégradation of cyanide via recombinant cyanide dehydratase from Bacillus pumilus expressed heterologously in Escherichia coli / Biodegradación de cianuro por la enzima cianuro dihidratasa de Bacillus pumilus expresada heterologamente en Escherichia coli

ABSTRACT Despite its high toxicity, cyanide is used in several industrial processes, and as a result, large volumes of cyanide wastewater need to be treated prior to discharge. Enzymatic degradation of industrial cyanide wastewater by cyanide dihydratase, which is capable of converting cyanide to ammonia and formate, is an attractive alternative to conventional chemical methods of cyanide decontamination. However, the main impediment to the use of this enzyme for the biodegradation of cyanide is its intolerance to the alkaline pH at which cyanide waste is kept and its low thermoresistance. In the present study, the catalytic properties of whole Escherichia coli cells overexpressing a cyanide dihydratase gene from Bacillus pumilus were compared to those of the purified enzyme under conditions similar to those found in industrial cyanide wastewater. In addition, the capacity of whole cells to degrade free cyanide in contaminated wastewater resulting from the gold mining process was also determined. The characteristics of intracellular enzyme, relative to purified enzyme, included increased thermostability (>60% activity at 50°C), as well as greater tolerance to heavy metals, and to a lesser extent pH (20% activity remaining at pH 9.0) On the other hand, enzymatic degradation of 70% of free cyanide (initial concentration 528 mM) in the industrial sample was achieved only after dilution. Nevertheless, the increased thermostability observed for intracellular CynD suggest that whole cells of E. coli overexpressing CynD are suited for process that operate at elevated temperatures (50°C), a limitation observed for the purified enzyme.

RESUMEN A pesar de su alta toxicidad, el cianuro es usado en diversos procesos industriales, y como resultado, grandes volúmenes de aguas residuales de cianuro deben ser tratados antes de su descarga. Una alternativa atractiva a los métodos químicos convencionales de descontaminación es la degradación enzimática por la enzima cianuro dihidratasa, la cual es capaz de convertir cianuro en amonio y ácido fórmico. No obstante, la inactivación de esta enzima a pH superior a 8.5 y su poca termoestabilidad han sido el principal impedimento para la implementación exitosa de esta alternativa de biorremediación. En el presente estudio, las propiedades catalíticas de células completas de Escherichia coli que sobre expresan el gen de cianuro dihidratasa de Bacillus pumilus se estudian bajo condiciones similares a las encontradas en aguas residuales industriales de cianuro y los resultados se discuten en comparación con las de la enzima purificada. Además, se determinó la capacidad de las células completas para degradar el cianuro libre en aguas residuales resultantes del proceso de extracción de oro. Las características de la enzima intracelular, relativa a la enzima purificada, incluyeron un incremento en la termoestabilidad (>60% actividad a 50°C), así como mayor tolerancia a metales pesados y en menor medida al pH (20% actividad residual a pH 9.0). Por otra parte, la degradación enzimática del 70% del cianuro libre en la muestra industrial (concentración inicial 528 mM) se logró solo después de la dilución de la muestra. Sin embargo, el incremento en la termoestabilidad observado para CynD intracelular sugiere que las células completas de E. coli que sobre expresan CynD son adecuadas para procesos que operan a temperaturas elevadas (50°C), una limitación observada para la enzima purificada.

Intoxicación por cianuro, perspectiva desde urgencias: reporte de dos casos y revisión de la literatura / Cyanide poisoning, perspective from the emergency room: report of two cases and literature review / Intoxicação por cianeto, perspectiva desde urgências: relato de dois casos e revisão da literatura

Aunque poco común, la intoxicación por cianuro acarrea una mortalidad alta, sobre todo, cuando no se administra el tratamiento oportuno. La exposición a este tóxico puede darse por inhalación o por ingestión oral. Lo que se ha descrito sobre el manejo de esta intoxicación está basado en reportes de casos y pequeñas series, dado que en la literatura no se encuentran estudios que den un sustento claro. Dentro de estos reportes, y según los aspectos fisiopatológicos, son especialmente relevantes: el nitrito de sodio, el nitrito de amilo, el tiosulfito de sodio y la hidroxicobalamina, como arsenal terapéutico de esta intoxicación.

Cyanide intoxication is a rare condition. Yet, it leads to a high mortality, especially when proper treatment is not available. The exposure to this substance may occur via inhalation or oral ingestion. The basis of this intoxication´s treatment is on little case series and reports; this is due to the fact that the literature to support it remains scant and unclear. Based on physiopathological aspects, within this evidence the following are particularly relevant: sodium nitrile, amile nitrile, sodium tiosulphite, and hidroxicobalamin, as part of this intoxication´s treatment.

Embora pouco comum, a intoxicação por cianeto provoca uma mortalidade alta, sobre tudo, quando não se administra o tratamento oportuno. A exposição a este tóxico pode dar-se por inalação ou por ingestão oral. O que se há descrito sobre o manejo desta intoxicação está baseado em reportes de casos e pequenas séries, dado que na literatura não se encontram estudos que deem um sustento claro. Dentro destes relatos, e segundo os aspectos fisiopatológicos, são especialmente relevantes: o nitrito de sódio, o nitrito de amilo, o tiossulfato de sódio e a hidroxicobalamina, como arsenal terapêutico desta intoxicação.

Caracterización molecular de bacterias cultivables y no cultivables procedentes de pozas de lixiviación con cianuro / Molecular characterization of culturable and non-culturable bacteria from leaching pools with cyanide

En el presente estudio, las comunidades bacterianas en muestras de suelo y agua, procedentes de pozas artesanales de lixiviación con cianuro, fueron caracterizadas por análisis dependientes e independientes de cultivo. Para la caracterización de la comunidad bacteriana cultivable, se emplearon técnicas clásicas de microbiología hasta la obtención de cepas puras, las cuales fueron identificadas a nivel molecular. Por otro lado, las comunidades bacterianas no cultivables fueron caracterizadas por secuenciación de próxima generación del gen ARNr 16S. La comunidad bacteriana cultivable estaba principalmente representada por los géneros Pseudomonas, Bacillus y Acinetobacter; mientras que las comunidades no cultivables, predominantes en muestras de suelo, fueron los filos Proteobacteria (12.91%), Firmicutes (11.32%), Actinobacteria (11.25%) y Bacteroidetes (10.16%). Por otro lado, en muestras de agua predominaron los filos Firmicutes (59.16%) y Actinobacteria (38.99%).

In the present study, bacterial communities in soil and water samples, from artisanal leaching pools with cyanide, were characterized by dependent and independent culture analyzes. For the characterization of the culturable bacterial community, classical techniques of microbiology were used, until obtaining pure strains, which were identified at the molecular level. On the other hand, uncultured bacterial communities were characterized by next-generation sequencing of the 16S rRNA gene. The culturable bacterial community was mainly represented by the genera Pseudomonas, Bacillus and Acinetobacter; while the predominant uncultured communities, in soil samples, were the proteobacteria (12.91%), Firmicutes (11.32%), Actinobacteria (11.25%) and Bacteroidetes (10.16%). On the other hand, in water samples, the edges of Firmicutes (59.16%) and Actinobacteria (38.99%) predominated.

Antinutritional Factors Reduction from Cassava (Manihot esculenta Crantz) Roots by Grating or Chipping Processing Technique in Mtwara Tanzania

Regular intake of diets high in cyanogenic amount in iodine deficiency areas with limited sulfur supply diets have been associated with the development of hypothyroidism, goiter and cretinism in young and adult populations. This study aimed at the investigation of the suitable processing technique for the production of high-quality cassava flour (HQCF) with acceptable consumption level of cyanide residues from different cassava varieties available at Mmongo village in Mtwara region. Grating technique was used to produce wet and dried cassava grits, whereas chipping technique produced wet and dried cassava chips. The wet and dried cassava products were all produced in a single day and under the same environmental conditions. The results showed that the hydrogen cyanide (HCN) in fresh cassava roots was 62.18–139.28 mg HCN/kg. Upon processing, the cyanide was lower (P<0.05) in wet cassava grits 24.27–76.74 mg HCN/Kg but higher in wet cassava chips 44.81–92.57 mg HCN/kg. After sun drying, the HCN retention was higher in dried cassava chips (10.7–14.2%) and lower in dried cassava grits (7.3–9.1%). Grating, efficiently reduced HCN from bitter cassava roots to the level within the WHO/FAO recommended safe limit (10 mg HCN/kg) as chipping only suite for sweet cassava roots. Therefore, HQCF can be nutritionally strategic cassava product consumed regularly and during food shortage without causing potential health problems even in iodine deficiency areas.

Occupational acute mixed gas poisoning due to improper disposal of hazardous waste / 中华劳动卫生职业病杂志

Objective@#To investigate an occupational hazardous gas poisoning incident caused by gas leakage in the process of hazardous waste treatment.@*Methods@#An investigation was conducted on a case of occupational acute hazardous gas poisoning caused by waste treatment gas leakage in Shandong province in December 2017. Meanwhile, the clinical data of 5 cases of poisoning patients were analyzed, and the accident related poison test report and other relevant data were analyzed.@*Results@#The incident was caused by the toxic waste did not do labeling work, the workers' protection measures were not in place, the illegal operation and the blind rescue, resulting in a total of 5 people died on the spot, 12 people were hospitalized with poisoning. Among them, 5 patients admitted to our hospital showed varying degrees of damage to the nervous system and respiratory system. After active treatment, they all got better and were discharged.@*Conclusion@#The poisoning is mainly caused by hydrogen sulfide dichloromethane hydrogen cyanide gas leakage serious production liability accident, clinical main performance for the nervous system circulatory system respiratory system and other system damage.

Cyanide Toxicity of Freshly Prepared Smoothies and Juices Frequently Consumed

Aims: This study was conducted to detect the presence of cyanide in popular fruit and vegetable smoothies and juices marketed as raw and natural. Study Design: Eleven (11) popular varieties of drinks were analyzed for total cyanide (TCN). Drinks contained raw vegetables and fruits, flax seeds, whole apples with seeds, raw almond milk, and pasteurized almond milk as ingredients. Place and Study Duration: Samples were collected from health food eateries located within Las Vegas, Nevada (USA) during the summer of 2017. Methodology: Fifty milliliters (mL) of a homogenized smoothie and juice drink and 1 gram of flax seeds were subjected to the above-referenced methods for sample preparation per USEPA Methods 9012B (digestion) followed by USEPA method 9014 (colorimetry). Results: The highest TCN was detected in drinks containing raw flax seed followed by unpasteurized raw almond milk, then fresh whole apple juice. No TCN was observed in drinks that contained none of the above mentioned items (e.g. flax seed, raw almond milk) or those utilizing pasteurized ingredients. Conclusion: This study observed that TCN is present in smoothies and juices containing raw flax seeds, fresh whole apples, and/or unpasteurized almond milk. Concentrations were detected as high as 341 μg L-1 in commercially available smoothies containing vegetables, raw flax seeds, almond milk and fruits. Smoothies with vegetables, fruits, unpasteurized almond milk, and no flax seeds contained 41 ug L-1TCN, while similar smoothies with pasteurized almond milk contained negligible to 9.6 ug L-1 CN-. Unpasteurized almond milk and raw flax seeds were the major sources of TCN in drinks. With the increased demand for raw and natural foods, there is a potential sublethal exposure of TCN by consumers.

Attenuation of KCN-induced Neurotoxicity by Solvent Fractions of Antiaris africana Leaf

Antiaris africana belongs to the family Moraceae, it is commonly called “False Iroko” tree and one of the medicinal plants used in treatment of mental and nervous disorders in Nigeria. We have previously established the neuroprotective properties of crude extract of A. africana. The present study was thus designed to investigate the neuroprotective effect of different solvent fractions of A. africana against cyanide neurotoxicity in vitro. Cyanide induced a significant (P<0.01) inhibition of NADH succinate dehydrogenase, a key enzyme in mitochondria function as well as significant increase in oxidative stress as observed in the high level of malonedialdehyde (MDA), protein carbonyl (PC) and activity of monoamine oxidase (MAO) and decreased concentration of reduced glutathione (GSH) as compared to the control. Co-administration with different solvent fractions of A. africana (hexane fraction [HFA], dichloromethane fraction [DFA] and methanolic fraction [MFA]) significantly ameliorated the toxic effect of KCN as compared to the induced, untreated group (P<0.05). The results in this study showed that HFA (79.04% reversal of NSD inhibitory activity of KCN), DFA (63.68% and 72.6% activity against KCN induced LPO and PC respectively). However, MFA showed the best activity against GSH depletion caused by KCN (12.21%) and inhibition of MAO activity induced by KCN (94.63%). In conclusion, all the fractions possess neuroprotective activities at varying degrees against mitochondria damage by KCN. This result further substantiated the ethnomedicinal usage of A. africana and can provide novel compounds in the treatment of mitochondria-related neurodegenerative diseases.

A Novel Fluorescent Probe Based on Schiff Base for Dual Sensing of Zn2+and CN- / 分析化学

A novel Schiff base probe 4'-hydroxy-3'-((2-pyridin-2-ylmethylimino) methyl)-4-biphenyl carbonitrile (HPBC) for dual sensing of Zn2+and CN-was synthesized and characterized by various techniques such as UV-Vis, fluorescence, HRMS and NMR spectroscopy. HPBC was highly selective toward these two ions with different fluorescence signals in two different media. In EtOH-H2O(2:3,V/V;HEPES,pH 7.4),HPBC selectively bound Zn2+to form a 1:1 ligand/metal complex. Addition of Zn2+to the solution of HPBC resulted in a blue shift (△λ=15 nm) with a pronounced fluorescence enhancement at 468 nm, while there was no enhancement in the presence of other metal ions,especially Cd2+. HPBC displayed an"ON-OFF-ON"mode fluorescence change with alternative addition of Zn2+and EDTA. Hence, HPBC is a reversible and reusable sensor for Zn2+. The dynamic range of the assay is linear up to 4.0 μmol/L Zn2+ion and the limit of detection was 36.5 nmol/L,which was thousand fold lower than the WHO guideline (about 76 μmol/L) for drinking water. The fluorescence response of HPBC toward Zn2+was pH-dependent, and the maximal signal was observed at near neutral pH values, which makes it suitable for application in physiological conditions. Cell imaging studies demonstrate that this sensor is capable of sensing Zn2+in living cells. In DMSO-H2O (3:7,V/V) medium,addition of cyanide ion to HPBC led to deprotonation of the phenol hydrogen,resulting in a color change from colorless to pale yellow and a significant fluorescence enhancement at 510 nm. The probe exhibited high selectivity and sensitivity for CN- ion and the detection limit was 5.75×10-7mol/L. Finally,the use of a test strip of probe HPBC to detect cyanide was reported.

Quality Evaluation of Shuwei Mixture Based on Multi-index Constituents / 医药导报

Objective The quality evaluation of the Shuwei mixture was determined by the content of the six components of sinapine cyanide sulfonate, magnolol, honokiol, hesperidin, naringin and neohesperidin. Methods RP-HPLC method was used.The separation was performed on a Agilent ZORBAX Eclispse SB-C18column (4.6 mm×250 mm,5 mm),the mobile phase consisted of acetonitrile(A)-0.1% phosphoric acid with gradient elution at the flow rate of 1.0 mL·min-1.The detection wavelength was 326 nm ( sinapine cyanide sulfonate ), 294 nm ( magnolol, honokiol ) and 283 nm ( naringin, neohesperidin).The column temperature was kept at 30 ℃. Results The sinapine cyanide sulfonate, magnolol, honokiol, hesperidin,naringin and neohesperidin all had good linear relationship in the ranges of 0.049 6-1.24,0.048 2-1.205,0.060 5-1.512 5,0.187 2-4.68,0.131 6-3.29,0.197-4.925 μg.The average recoveries were 100.66%, 99.86%, 101.37%, 102.41%, 99.01%, 102.05%, respectively, RSD were 0.82%, 1.89%, 2.56 %, 0.74%, 1.54%, 0.99%, respectively. Conclusion The method is simple,accurate,reproducible and nice to the separation,and can be used for the quality evaluation of Shuwei mixture.

Combined effect of noise and hydrogen cyanide on noise-induced hearing loss in a metal electroplating enterprise / 中国职业医学

OBJECTIVE: To study the combined effect of noise and hydrogen cyanide on noise-induced hearing loss( NIHL)in a metal electroplating enterprise. METHODS: The judgment sampling method was used to select 663 workers in a largescale metal electroplating enterprise as the study subjects. Among them,186 workers exposed to noise alone were designated as noise group; 138 workers exposed to hydrogen cyanide alone were designated as hydrogen cyanide group; and161 workers exposed to noise and hydrogen cyanide were designated as combined effect group,and 178 workers without exposure from occupational disease risk factors were designated as control group. Questionnaires survey and pure tone audiometry were used for analyzing the effects of combined noise and hydrogen cyanide exposure on NIHL. RESULTS: The hearing loss detection rate of the study subjects was 40. 4%. The hearing loss detection rates in the control group,noise group,hydrogen cyanide group,and combined effect group were 17. 4%,47. 8%,32. 6% and 64. 0%,respectively. The detection rate of hearing loss in the control group was lower than that in the other three groups( P < 0. 008). The NIHL detection rates in the combined effect group and the noise group were higher than that in the hydrogen cyanide group( P <0. 008). The hearing loss detection rate of the combined effect group was higher than that of the noise group and the hydrogen cyanide group( P < 0. 008). Ordinal multi-categorical logistic regression model results showed that after adjusting confounding factors such as age,length of service,gender,marital status,smoking,alcohol drinking,we found hydrogen cyanide exposure,noise exposure,and combined exposure to hydrogen cyanide and noise had effects on workers' hearing(P < 0. 05). The risk of hearing loss in workers exposed to noise and hydrogen cyanide was higher than that of workers exposed to noise alone or hydrogen cyanide alone. CONCLUSION: There is a combined effect of noise and hydrogen cyanide in this metal electroplating enterprise,which can increase the risk of NIHL in workers.