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Objective:To establish a hydride generation atomic fluorescence method using ammonium persulfate as the digestion reagent for determination of arsenic in urine (hereinafter referred to as this method).Methods:The collected urine samples with ammonium persulfate were heated and digested on the tubular electric heating automatic control constant temperature digester (60 holes), with 5% hydrochloric acid solution as reaction medium and current carrier and 1.5% potassium borohydride solution as reducing agent. Arsenic content was determined with a four-channel atomic fluorescence spectrometer. The arsenic standard solution of 0 - 10 μg/L was prepared to determine the standard curve of this method, and the method was evaluated from the detection limit, linear range, correlation coefficient, precision, standard addition recovery experiment, and urine arsenic quality control sample detection. The standard method "Determination of Arsenic in Urine by Hydride Generation Atomic Fluorescence Spectrometry" (WS/T 474-2015, referred to as the standard method) was used for comparison experiments.Results:When the sampling volume was 1 ml, the detection limit of this method (digest with 1 ml 1.5 mol/L ammonium persulfate) was 0.03 μg/L. In the range of arsenic content from 0 - 10 μg/L, the linear relationship between arsenic content and fluorescence intensity was good, and the correlation coefficients ( r) were all 0.999 9. The relative standard deviations( RSD) of the three replicates of urine samples with different concentrations were 1.00%, 0.89% and 0.49%, respectively. Urine arsenic quality control samples were tested, and the test results were all within the range of public values; the overall average recovery was 102.29%, and the recovery range was 92.10% - 108.15%. Compared with the standard method in the determination results of 20 urine samples, the difference was not statistically significant ( t = - 0.40, P > 0.05). Conclusions:The hydride generation atomic fluorescence spectrometry using ammonium persulfate as digestion reagent for the determination of arsenic in urine has the advantages of low detection limit, good precision, high accuracy, small amount of sampling and digestion reagent, simple operation, and less harmful gas generation in sample pretreatment. It is suitable for rapid determination of arsenic in urine in large quantities.
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Objective:To determine the contents of inorganic arsenic（iAs），monomethylarsonic acid（MMA） and dimethylarsinic acid（DMA） in brain tissues and blood by using hydride generation-cold trap-atomic absorptionspectrometry（HG-CT-AAS）， and to explore the toxic effects of Realgar on central nervous system of rats. Method:The 96 Wistar rats were randomly divided into 4 groups：normal control group，0.3，0.9 and 2.7 g·kg<sup>-1</sup> Realgar groups. They then received intragastric administration for 14，28 and 42 days respectively， so a total of 12 groups were formed， with 8 animals in each group. The normal group was given the same dose of sodium carboxymethyl cellulose （CMC-Na） by gavage. The contents of iAs，MMA and DMA in blood and brain tissues were determined by HG-CT-AAS. The novel object recognition test was conducted to observe the learning and memory ability of rats. The changes of hippocampal neuron ultrastructure were observed by transmission electron microscopy. Result:There was no difference in the growth，weight and hippocampal coefficient of the experimental animals. The method of HG-CT-AAS showed a good linearity，precision，accuracy and recovery in content determination of arsenic （at various forms） in rat brain and blood. MMA and DMA were detected in the brain of realgar groups at time-dose-effect relationship. iAs，MMA and DMA were detected in the blood of Realgar groups. The nuclear membrane， mitochondria and endoplasmic reticulum in hippocampus neurons of rats were gradually damaged with the increase of Rhubarb exposure dose and time. After 14 days of exposure to Realgar，compared with the normal control group，there was no significant difference in the novel object recognition index among Realgar groups. After 28 days of exposure，only 2.7 g·kg<sup>-1</sup> Realgar group showed statistically significant difference with the control group （<italic>P</italic><0.05）. After 42 days of exposure， the novel object recognition index of 0.9 and 2.7 g·kg<sup>-1</sup> Realgar groups was significantly lower than that in normal control group（<italic>P</italic><0.05）. Conclusion:The metabolites of Realgar in rats are iAs，MMA and DMA. MMA and DMA can be accumulated in the brain tissue through the blood-brain barrier，causing the decline of the ability of learning and memory and leading to damage of hippocampal neurons.
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A sensitive and rapid method using liquid chromatography-hydride generation atomic fluorescence spectrometry (HPLC-HG-AFS) was developed for the simultaneous determination of seven arsenic species As3+, As5+,MMA, DMA, p-ASA, 4-OH and ROX in feeds. The isolation of the analytes from feed samples was accomplished using methanol water (1:1, V/V). The target compounds were separated on a PRP-X100 anion exchange column and then analyzed by HG-AFS. The mobile phase was 15 mmol/L (NH4)2HPO4and 10 mmol/L potassium acid phthalate. Good linearity was obtained for all of the seven arsenic species, with linear coefficients higher than 0.9964. The LODs of the seven arsenic species were between 5 and 30 μg/kg. Average recoveries for the seven analytes were in the ranges of 76.3%-108.1%, with intra- and inter-day repeatability lower than 7.7% and 17.4%,respectively. This validated method was successively applied to the determination of arsenic species in feed. This method was sensitive,simple,cheap and low operation cost,and could be used for the determination of the arsenicspecies in feeds.
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The purpose of this study was to evaluate the effect of acid-treatment conditions on the surface properties of the RBM (Resorbable Blast Media) treated titanium. Disk typed cp-titanium specimens were prepared and RBM treatments was performed with calcium phosphate ceramic powder. Acid solution was mixed using HCl, H2SO4 and deionized water with 4 different volume fraction. The RBM treated titanium was acid treated with different acid solutions at 3 different temperatures and for 3 different periods. After acid-treatments, samples were cleaned with 1 % Solujet solution for 30 min and deionized water for 30 min using ultrasonic cleanser, then dried in the electrical oven (37℃). Weight of samples before and after acid-treatment were measured using electric balance. Surface roughness was estimated using a confocal laser scanning microscopy, crystal phase in the surface of sample was analyzed using X-ray diffractometer. Surface morphology and components were evaluated using Scanning Electron Microscope (SEM) with Energy Dispersive X-ray spectroscopy (EDX) and X-ray Photoemission Spectroscopy (XPS). Values of the weight changes and surface roughness were statistically analyzed using Tukey-multiple comparison test (p=0.05). Weight change after acid treatments were significantly increased with increasing the concentration of H₂SO₄ and temperature of acid-solution. Acid-treatment conditions (concentration of H₂SO₄, temperature and time) did not produce consistent effects on the surface roughness, it showed the scattered results. From XRD analysis, formation of titanium hydrides in the titanium surface were observed in all specimens treated with acid-solutions. From XPS analysis, thin titanium oxide layer in the acid-treated specimens could be evaluated. Acid solution with 90℃ showed the strong effect on the titanium surface, it should be treated with caution to avoid the over-etching process.
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Calcio , Cerámica , Microscopía Confocal , Espectroscopía de Fotoelectrones , Espectrometría por Rayos X , Propiedades de Superficie , Titanio , Ultrasonido , AguaRESUMEN
Objective:To quantify the plasma concentrations of inorganic arsenic (As(III) and As(V)) and methylated metabo-lites ( MMA and DMA) , and to detect the total amount of arsenic in blood cells and plasma by high performance liquid chromatogra -phy-hydridegeneration-atomic fluorescence spectrometry ( HPLC-HG-AFS) and HG-AFS methods to clarify the arsenic species in acute promyelocytic leukemia (APL) patients.Methods:The blood cells and plasma were digested by the mixture of HNO 3-H2O2 and ana-lyzed by HG-AFS.For the arsenic species , the plasma samples were prepared with perchloric acid to precipitate protein .The superna-tant was separated on an anion-exchange column in 6 min with isocratic elution using 13 mmol · L-1 CH3 COONa, 3 mmol · L-1 NaH2 PO4 , 4 mmol· L-1 KNO3 and 0.2 mmol· L-1 EDTA-2Na.Results:The methods provided linear range of 0.2-20 ng· ml-1 for total arsenic and 2.0-50 ng· ml-1 for four arsenic species (r>0.9950).The spiked recoveries ranged from 81.2％to 108.6％, and the coefficients of variation for intra-and inter-batch precision were less than 9.3％and 12.5％, respectively.The developed methods were applied successfully in the assay of total arsenic and arsenic species in 5 APL patients.Conclusion:The method is simple, fast and accurate , which can be applied in the assay of arsenic compounds in plasma and blood cells in APL patients .
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OBJECTIVE:To establish a method for the contents determination of 5 heavy metals in Ershiwuwei shanhu pills, and to investigate the contents of heavy metals in Ershiwuwei shanhu pills produced by 5 manufacturers from different districts of Tibet. METHODS:Ershiwuwei shanhu pills were digested by HNO3-HClO4(4:1,V/V).The contents of 3 heavy metals as Cu,Pb, Cd in samples were measured by flame atomic absorption spectrometry(FAAS). The contents of 2 heavy metals as As,Hg in samples were measured by hydride generation atomic absorption spectrometry(HG-AAS). RESULTS:5 kinds of heavy metals have good linear relationship in the corresponding mass concentration range(all r≥0.999 1). The limits of detection of Cu,Pb, Cd were 0.001 6,0.041 2,0.036 3 mg/L,and the limits of quantitation were 0.005 3,0.137 3,0.121 0 mg/L;the limits of detection of As,Hg were 0.325 7,0.692 3 μg/L,and the limits of quantitation were 1.085 7,2.307 7 μg/L. RSDs of precision tests were ≤5.54%(n=6);RSDs of stability tests were all≤3.79%;RSDs of reproducibility tests were ≤3.72%. The average recovery rates were 91.34%-110.11%(RSDs were 0.66%-6.80%,n=6). Results of contents determination showed that the contents of Cu in samples from 5 manufacturers were not out of limits,but the contents of Cd and Hg were all out of limits; the contents of Pb in samples from 4 manufacturers were out of limits,and the contents of As in samples from 2 manufacturers were out of limits. CONCLUSIONS:The established method has good accuracy,sensitivity,stability and reproducibility,and it is suitable for contents determination of 5 heavy metals in Ershiwuwei shanhu pills. To some extent,there is a problem of excessive heavy metals in samples from 5 manufacturers.
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Objective To evaluate the method for detection of urinary mercury using a Zeeman atomic absorption mercury analyzer and to provide a reference for selecting a convenient method for mercury detection in experiments and clinical diagnoses. Methods Urinary mercury was detected by Zeeman atomic absorption spectroscopy (ZAAS) and hydride generation atomic absorption spectrometry (HG-AAS), and the detection limit, accuracy, precision and consistency of the two methods were compared. Results The Data collected by ZAAS and HG-AAS showed a good linear relationship in the range of 0 -1000 ng/mL (ZASS, R2 =1. 0000) and 0 -20 ng/mL (HG-AAS, R2 =0. 9990). The detection limits of ZAAS was 0. 156 ng/mL and that of HG-AAS was 1. 593 ng/mL, indicating that ZAAS is more sensitive. The recovery rate of standard addition of ZAAS was between 97. 5％ and 103. 2％, and that of HG-AAS was between 95. 6％ and 104. 5％. After measurement of 10 ng/mL and 100 ng/mL mercury standard solutions repeated for 10 times, the relative standard deviation (RSD) of ZAAS was 0. 30％ and 0. 36％ respectively, and the RSD of HG-AAS was 2. 82％ and 1. 11％, respectively. The accuracy and precision of both the two method met the standards of GBZ/T 210. 5-2008, and the precision of ZAAS was better. A total of 30 urine samples were measured by these two methods. The results were compared with paired-samples t-test and showed a non-significant difference (P ＞ 0. 05), indicating a high consistency of these two method (R2 =0. 9961). Conclusions ZAAS is a convenient and accurate method for the detection of urinary mercury, with a relatively low detection limit and better precision.
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Objective To establish a method for simultaneous determination of arsenic and selenium in hair by hydride generation atomic fluorescence spectrometry.Methods The A and B of the hair samples were digested via the wet method,and the arsenic and selenium contents were determined simultaneously with a dual channel atomic fluorescence spectrophotometer.Standard blank solution was continuously determined for 11 times,the detection limit was calculated according to the blank solution three times the standard deviation.Hair samples A and B were continuously determined for 11 times,and the precision was determined according to the coefficient of variation (CV).The hair samples C and D were did 11 times of standard addition experiment,the standard added was close to the sample low value.The accuracy was determined by the standard recovery rate.Results The lowest detection limits of arsenic and selenium were 0.081,0.036 μg/L,the precisions of arsenic and selenium in sample A were (0.694 9 ± 0.024 2) and (1.930 2 ± 0.098 5) μg/g,the CV were 3.54％ and 5.09％.The precisions of arsenic and selenium in sample B were (6.212 2 ± 0.137 3) and (8.282 2 ± 0.266 8) μg/g,the CV were 2.21％ and 3.22％;the recovery rates of sample C of arsenic and selenium were 96.30％ ～ 110.00％ and 94.80％ ～ 110.00％;the recovery rates of sample D of arsenic and selenium were 93.63％ ～ 108.20％ and 97.03％ ～ 108.34％.Conclusions The method using hydride atomic fluorescence for determination of arsenic and selenium in the hair has been successfully established.This method is simple and easy to operate,with high sensitivity and good accuracy and precision,high recovery,less matrix interference,and less reagent consumed etc.It is worth promoting in prevention and control of endemic diseases.
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Objective To establish a rapid, simple and accurate method for detection of selenium in grain that is suitable in Chinese situation. Methods Nitric acid and perchloric acid(7: 3, v/v) were used to digest the grain samples by heating on a hot plate. Selenium was determined with hydride generation atomic fluorescence spectrometry. Sample detection limit, precision, accuracy(recovery, method characteristics and method control) were studied. And the grain samples of Shandong Province were determined by this method. Results The lowest detection limit was 4 μg/kg. The coefficient of correlation of working curve was 0.999 9. Intra-day precision was 1.32%, day precision was 4.17%. The total average rate of recovery was 100.5% with a range of 96.7% - 105.5%, and the average rates of recovery were 104.0%, 99.0% and 98.4% (n = 6). The determination results of corn reference material [(0.022 ± 0.006) mg/kg] were in the standard value range [(0.021 ± 0.008) mg/kg]. The determination results of the samples [(0.424 ± 0.096) mg/kg] were consistent with the results of national standard fluorescence method [(0.406 ± 0.108) mg/kg]. The contents of selenium in wheat, maize and sweet potato samples from five regions of Shandong Province were:Shanting:(0.030 3 ± 0.025 2),(0.016 8 ± 0.013 5),(0.015 4 ± 0.002 9) mg/kg; Anqiu:(0.020 3 ± 0.000 1), (0.020 4 ± 0.009 9), (0.017 1 ± 0.007 5) mg/kg; Ju'nan:(0.021 3 ± 0.013 9), (0.018 5 ± 0.007 8),(0.019 9 ± 0.003 6)mg/kg;Yishui:(0.025 7 ± 0.006 2),(0.020 6 ± 0.003 2), (0.018 2 ± 0.003 2) mg/kg; Wulian:(0.020 3 ± 0.004 7), (0.020 1 ± 0.008 9), (0.018 4 ± 0.007 3) mg/kg. Conclusions The method has the advantages of higher precision and accuracy, less time, less pollution, less aciduse, easier operation and repeatability.It is very suitable for measuring selenium content in large amount of food samples.
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Objective To establish and evaluate a method for determination of total arsenic in urine by test-tube rapid digestion hydride generation atomic fluorescence spectrometry.Methods After digestion of urine samples using graduated test-tube and graphite digestion apparatus,arsenic content in urine was determined with atomic fluorescence spectrometer.Then the test results were evaluated by using quality control measures,such as precision and accuracy experiments,and the results between different laboratories were reviewed and compared.Results The urinary arsenic was in a linear range of 0-0.300 mg/L,correlation coefficient (r) ＞ 0.999 3,detection limit was 0.000 21 mg/L,relative standard deviation (RSD) ≤4.62％ and the recoveries of standard addition were 93.9％-104.3％.The value of standard reference material measured was within the allowable range.The blind sample of the national urinary arsenic was qualified.Conclusions This method is suitable for large scale determination of urinary arsenic for its micro sample amount needed,less interference and strong practicability.The error results are in a controlled range.
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Objective To apply hydride generation atomic fluorescence spectrophotometry (HG-AFS method) in urinary arsenic detection,and to provide a better,newer and more convenient detection method for quantitative analysis of urinary arsenic.Methods According to the Guide to Develop Biological Sample Inspection Method(WS/T 68-1996) and Guide for Establishing Occupational Health Standards-part 5:Determination Methods in Biological Materials (GB/T 210.5-2008),HG-AFS method was established to detect arsenic content in urine after modification of the method for sample pretreatment,and to verify the linear range of standard curve and linearity,detection limit,precision,accuracy,stability of the sample,and to compare the experimental results of HG-AFS method with those of standard methods of WS/T 28-1996 and Determination of Arsenic in Urine by Cyanide Generation Atomic Fluorescence Method (WS/T 474-2015).Results The HG-AFS method linear range was from 0-100 μg/L,the correlation coefficient r =0.999 9,the detection limit was 0.07 μg/L,the precision was 1.96％-3.97％,and the recovery rate was 95.1％-105.0％.There was no statistical significance between HG-AFS method,the standard of WS/T 28-1996 or WS/T 474-2015 methods (t =1.539,0.353,all P ＞ 0.05).Conclusion The new method is superior to the current detection method owing to its low detection limit,high precision,good accuracy,and wide linear range.
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Objective To establish a method for determination of arsenic in urine,using hydrogen peroxide as the main digestion reagent to digest urine,and using hydride generation atomic fluorescence spectrometry (HG-AFS) to determine arsenic (this method was referred to below),the feasibility of the application in the monitoring of endemic arsenic poisoning was discussed.Methods Temperature control instrument (60 holes) and supporting special calibration tube were used to digest urine.Digestion reagents was mainly hydrogen peroxide plus a small amount of nitric acid and sulfuric acid,HG-AFS was used to determinate.Based on standard curve to calculate linear relationship.Using this method to determinate detection limit,precision,accuracy and stability of samples,this method was compared with the national hygienic standard method (DDCAg method,WS/T 28-1996).Results The detection limit was 0.8 μg/L (1 ml of urine was tested),the correlation coefficient was larger than 0.999 5.Precision:3 samples were determined,the arsenic contents were (11.0 ± 0.6),(39.2 ± 1.0),(174.4 ± 3.8) μg/L and the relative standard deviations were 5.03％,2.59％ and 2.17％,respectively.Recovery rate:3 samples were determined for standard addition recovery test,the average recovery rate was 99.4％ and the recovery rate ranged between 94.0％-104.3％.Methods contrast:this method [(125.9 ± 61.6) μg/L] and DDCAg method [(121.3 ± 52.5) μg/L] were used to determinate 20 samples from endemic arsenic poisoning areas,respectively,the determination results of the two methods were not significantly different (t =1.22,P ＞ 0.05).Conclusions This method is built successfully,it has good precision and accuracy,it needs small amount of sample and reagent,the amount of harmful gases generated is greatly reduced,and it is easy to operate and beneficial to operator's health.Therefore,it is a good method to determine arsenic in urine,and it can be applied in prevention of endemic arsenism.
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El huevo es una fuente proteica de uso masivo, en consideración a su bajo costo, es consumido con mayor frecuencia por toda la población. Por esta razón, es de gran interés introducir al mercado un tipo de huevo "saludable", que disminuya los riesgos ligados a enfermedades cardiovasculares y que actúe como antioxidante. Por ello, se evaluó la incorporación del Selenio Orgánico (Sel-Plex®, Alltech Inc) en el programa de alimentación de gallinas ponedoras comerciales raza isa brown con el objeto de obtener huevos fortificados con selenio. Estos fueron evaluados empleando la técnica de espectroscopia analítica con generación de hidruros (HGAAS) y para llevar a cabo la digestión se empleó una mezcla de HNO3/H2O2. El intervalo lineal fue de 6,0 a 200,0 µg/L para el selenio. Con un límite detección de 0,11 µg/L. La precisión de 0,82% y la recuperación de 97-104,4% y una frecuencia de análisis de 24 muestras/hora. La principal ventaja de este método es la determinación de selenio en huevo completo. Los resultados indican que luego de un período de 12 semanas la concentración de selenio es de 62,45 de mg Selenio/huevo lo que muestra que estos proporcionan los requerimientos necesarios. Por lo tanto los huevos enriquecidos en Venezuela contribuirían a mejorar la dieta diaria de estos consumidores.
The egg is a protein source for mass use, considering its low cost, is most commonly consumed by the population. For this reason, it is of great interest to enter the market a type of egg "healthy" to decrease the risks associated with cardiovascular disease and act as an antioxidant. Therefore, it evaluated the incorporation of organic selenium (Sel-Plex ®, Alltech Inc.) in the feeding program commercial layers Isa Brown race in order to obtain eggs fortified with selenium. These were assessed using analytical technique spectroscopy with hydride generation (HGAAS) and to conduct the digestion was used a mixture of HNO3/H2O2. The linear range was from 6.0 to 200.0 mg / L for selenium. With detection limit of 0.11 mg / L 0.82% accuracy and recovery of 97 to 104.4% and a frequency analysis of 24 samples / hour. The main advantage of this method is the determination of selenium in whole egg. The results indicate that after a period of 12 weeks the concentration of selenium is 62.45 Selenium mg / egg showing that these provide the necessary requirements. Therefore enriched eggs in Venezuela would help improve the diet of these consumers.
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Humanos , Animales , Masculino , Femenino , Embrión de Pollo , Óvulo/metabolismo , Selenio/uso terapéutico , Espectrofotometría Atómica/análisis , Antioxidantes , Salud Pública , Compuestos Químicos/clasificaciónRESUMEN
A rapid and sensitive method has been developed for the simultaneous determination of four selenium species Se(Ⅵ), Se(Ⅵ), selenomethionine, and Se-methylselenocysteine in Se-enriched yeast by liquid chromatography-hydride generation atomic fluorescence spectrometry (HPLC-HG-AFS). The isolation of the analytes from yeast samples was accomplished by proteaseⅩⅣ and trypsin enzymatic digestion. The target compounds were separated on a PRP-X100 anion exchange column and analyzed by HG-AFS. The mobile phase was 20 mmol/L (NH4)2HPO4. Good linearity was obtained for all the selenium species, with linear correlation coefficients higher than 0. 9996. The LODs of the four species were between 0. 5 and 5. 0 μg/kg. Average recoveries for the four analytes were in the ranges of 82 . 5%-101 . 2%, with intra-and inter-day RSD lower than 8. 6% and 14. 5, respectively. The proposed analytical method is simple, sensitive, with low operation cost, making it applicable for the determination of the selenium species in Se-enriched feeds.
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Objective:To investigate the relationship between surface hydrogen form and the bioactivity of titanium.Methods:Sandblast titanium was etched with the combination of sulfuric and hydrochloric acids(SLA group,n=3 ).Then etched titanium was heat at 450 ℃in air(SLA+HT group,n=3).Surface topography,roughness,hydrophility,surface chemical texture were observed. Finally,the titanium samples were soaked in body simulate fluid for 3 days,the mineral deposition properties were observed by X-ray diffraction.Results:Titanium hydride was formed on the titanium surface after etching.After heat treatment,surface texture and roughness were not changed,titanium hydride decomposed and hydrophility increased.More hydroxyapatite was found on the surface of the samples treated by SLA+HT and followed by SBF.Conclusion:Titanium hydride can not improve the bioactivity of titanium, heat treatment may increase the mineralization.
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The build-in low-pressure monolithic column combined with hydride generation atomic fluorescence spectrometry ( HG-AFS ) was employed for speciation analysis of fish meat. The sample pretreatment and separation approach could be accomplished within 30 min. The proper amount of fish sample was weighed and smashed into puree. The extraction solution composed of 10% HCl, 1% thiourea, and 0. 15% KCl was added before loaded into the automatic temperature controlled vertex system with 2000 r/min. The sample solution was separated through Merck monolithic column, with 3% ( V/V) acetonitrile, 30 mmol/L amonium acetate and 0. 03%(V/V) 2-mercaptoethanol (2-ME) as the eluent. The after-column eluent was digested by novel UV digestion device with pipeline sintered into the lamp, and then detected by hydrid-generation AFS. The rapid LC separation enabled fast mercury speciation of fish sample within 10 min. The different UV lamp digestion effects, eluent components, carrier gas, shield gas, lamp current, as well as PMT working power was optimized. Under the optimal conditions, the robust system achieved detection limits (DL) of 0. 15 μg/L and 0. 14 μg/L for methylmercury and HgⅡ, respectively. The RSD (n=7) was less than 5%, the linear correlation coefficient was 0 . 999 , and the matrix spiked recovery was in the range of 85%-110% for Hg speciation. This method was used for the determination of Hg speciation in fish and soil samples, and was proofed to be a reliable, easy approach for daily inspection.
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An ultrasensitive immunoassay was developed based on As3+ and Hg2+ labeled SiO2 @ Au nanoparticles signal tags and hydride generation-atomic fluorescence spectrometry (HG-AFS) for the detection of carcinoembryonic antigen(CEA) and carbohydrate antigen 19-9 (CA 19-9) respectively. Firstly, amino SiO2@ Au NPs were synthesized for selective absorption of As3+ and Hg2+ ions respectively. Subsequently,the secondary antibody (Ab2) of CEA and CA 19-9 was respectively labeled on As3+ or Hg2+-SiO2 @ Au NPs to prepare the corresponding signal tags for CEA and CA 19-9. Based on the sandwich immunoassay scheme, the tags, two antigen and corresponding first antibodies were bio-conjugated on the bottom of 96-well plate at room temperature to form the immunocomplex. After it was dissolved in alkali solution, As3+ and Hg2+ ions were released in solution and detected by HG-AFS, which concentration was proportional with logarithms of CEA and CA 19-9. The reaction conditions were optimized and the tags were characterized. This assay was based on determination of the concentration of As3+ and Hg2+ for quantization of the corresponding CEA and CA 19-9 antigen. The assay showed a wide linear range from 0. 001 to 100. 0 μg / L for CEA and 0. 01-80 U/ mL for CA 19-9, and a lower detection limit of 0. 5 ng / L and 0. 005 U/ mL respectively. This proposed method was used in real serums samples, the results were consistence with that by ELISA. The immunoassay showed three orders of magnitude of sensitivity lower than that of ELISA, which provides a promising simultaneous immunoassay for the early diagnosis of cancer .
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Cytochrome c oxidase (COX) employs electrons obtained from cytochrome c to bring about the reduction of oxygen to water. It is known that the electrons originate from the haem edge of cytochrome c and enters bovine COX at Trp-104. It is also known that Tyr-105, Glu-198 and Asp-158 of COX subunit II play roles in the enzyme's catalysis but how these roles are linked to electron transfer remain unclear. Recently, we proposed that electrons travel from the haem edge of cytochrome c to CuA, the first metal redox centre of COX, by a hydrogen/hydride ion relay using six residues. Now using a similar computer assisted approach, we investigate the extent to which this hydride/hydrogen ion mechanism is common amongst oxidases. The crystal structures of COX from P denitrificans, R sphaeroides and T thermophilus and quinol oxidase from E coli were downloaded and their binding domains analysed. As with bovine, all four oxidases had only nine amino acid residues in that region and both the sequences and three-dimensional structures were highly conserved. We propose that these residues function as a hydrogen/hydride ion relay, participating directly in electron transfer to CuA. We further suggest that this electron transfer mechanism might be a common feature in oxidases.
La citocromo c oxidasa (COX) emplea electrones obtenidos del citocromo c para producir la reducción del oxígeno a agua. Se sabe que los electrones originan a partir del hemo del citocromo c, y entran en la COX bovina en Trp-104. También se conoce que Tyr-105, Glu-198 y Asp-158 de la subunidad II de COX, desempeñan papeles en la catálisis de la enzima, pero no hay todavía claridad en cuanto a cómo estos papeles se hallan vinculados con la transferencia de electrones. Recientemente, sugerimos que los electrones viajan del borde del hemo del citocromo c al CuA, el primer centro metálico de reacción redox de la COX, por un relé iónico hidrógeno-hidruro, usando seis residuos. Ahora, usando un enfoque similar computarizado, investigamos hasta que punto este mecanismo de iones hidrógeno/hidruro es común entre las oxidasas. Se bajaron y analizaron los dominios de unión de las estructuras cristalinas de la COX de P denitrificans, R sphaeroides, y T thermophilus, y de la quinol oxidasa de la E coli. Como en el caso de la bovina, las cuatro oxidasas tenían sólo nueve residuos de aminoácido en esa región, y tanto las secuencias como las estructuras tridimensionales presentaban un alto grado de conservación. Proponemos que estos residuos funcionan como un relé iónico hidrógeno-hidruro, participando directamente en una transferencia de electrones al CuA. Asimismo, sugerimos que este mecanismo de transferencia de electrones podría ser un rasgo común de las oxidasas.
Asunto(s)
Animales , Bovinos , Complejo IV de Transporte de Electrones/metabolismo , Citocromos c/metabolismo , Hemo/química , Hidrógeno/metabolismo , Oxidación-Reducción , Paracoccus denitrificans/enzimología , Protones , Rhodobacter sphaeroides/enzimología , Secuencia de Aminoácidos , Thermus thermophilus/enzimología , Escherichia coli/enzimologíaRESUMEN
La presencia de Arsenico (As) en aguas de consumo representa una problematica para la salud publica en muchas regiones del mundo, incluida la Argentina. La cuantificacion de arsenico en agua de bebida y en orina se utiliza para evaluar la exposicion a este contaminante. El presente trabajo tuvo como objetivo la validacion metodologica de una tecnica para la cuantificacion de especies del As [AsV + AsIII + acido monometilarsonico (MMA) + acido dimetilarsinico (DMA)] por inyeccion en flujo-generacion de hidruros-espectrometria de absorcion atomica (IF-GH-EAA), previa derivatizacion con L-cisteina. Los resultados fueron comparados con los obtenidos utilizando dos metodologias de referencia, generacion de hidruros-espectrometria de absorcion atomica (GH-EAA) para muestras de aguas y orina, y cromatografia de alta resolucion ¨Cgeneracion de hidruros-espectrometr¨ªa de absorcion atomica (HPLC-GH-EAA) para muestras de orina. Ademas, se evaluo la selectividad de la cuantificacion por IF-GH-EAA, en presencia de otras especies quimicas del As, provenientes del consumo de alimentos producto de la pesca, a traves de un ensayo biologico. Los niveles de As hallados en las muestras de agua y de orina utilizando las tecnicas de referencia presentaron un rango de 6 a 176 ¦Ìg/L y de 143 a 3312 ¦Ìg/g de creatinina, respectivamente. Los coeficientes de Pearson resultantes de la comparacion de los datos obtenidos por IF-GH-EAA, con los logrados por los metodos de referencia fueron r = 0,9976 y r = 0,9422, para agua y orina, respectivamente. Los resultados de la prueba biologica indicaron un mayor nivel de As, debido al consumo de alimentos producto de la pesca, cuando las muestras de orina fueron previamente mineralizadas (GH-EAA), con la consecuente sobreestimacion del contenido de As proveniente del consumo de As inorganico. Este aumento no se observo cuando estas fueron analizadas por IF-GH-EAA ...
The presence of arsenic (As) in drinking water is a public health concern in many regions of the world, including Argentina. Quantification of arsenic in drinking water and urine are used to assess exposure to this pollutant. This study aimed to validate a methodology for the quantification of As species [AsV + AsIII + acid monometilarsonico (MMA) + dimetilarsinico acid (DMA)] by flow injection-hydride generation-atomic absorption spectrometry (FI-HG-AAS), after derivatization with L-cysteine. The results were compared with those obtained using two methods of reference, hydride generation-atomic absorption spectrometry (HG-AAS) for water and urine samples, and high performance liquid chromatography-hydride generation-atomic absorption spectrometry (HPLC-HG-AAS) for urine samples. In addition, the selectivity of quantification by FI-HG-AAS in the presence of other chemical species of As, from fishery products intake, was evaluated through a biological assay.The As level found in water and urine samples, using the techniques of reference, showed a range from 6 to 176 ¦Ìg/L and from 143 to 3312 ¦Ìg/g creatinine, respectively. Pearson coefficients resulting from the comparison of data obtained by FI-HG-AAS with those achieved by the reference methods were r = 0.9976 and r = 0.9422 for water and urine, respectively. The results of the biological test showed a higher level of As, due to consumption of food fishery product, when urine samples were previously mineralized (HG-AAS), with consequent overestimation of the inorganic arsenic consumption. When these samples were analyzed by FI-HG-AAS this fact was not observed, and the values were comparable to baseline level prior to consumption ...
Asunto(s)
Animales , Arsénico/orina , Contaminantes Químicos del Agua/análisis , Contaminantes Químicos del Agua/orina , Exposición a Riesgos Ambientales , Espectrofotometría AtómicaRESUMEN
El arsénico (As) es un contaminante natural que afecta una amplia zona de Argentina. El nivel de As en agua de consumo es utilizado para evaluar la fuente de exposición y en orina para evaluar exposición a este tóxico. El presente trabajo tuvo como objetivo la optimización y validación metodológica de una técnica para la cuantificación de As [As suma = As inorgánico (AsI) + especies metiladas: ácido monometilarsónico (MMA) y ácido dimetilarsónico (DMA)], producto del metabolismo del AsI, por inyección en flujo- generación de hidruros- espectrometría de absorción atómica (IF-GH-EAA), previa derivatización con L-cisteína. La recuperación de las especies estudiadas: AsI (AsIII y AsV), MMA y DMA fue cercana al 100% en todos los casos. Los límites de detección y cuantificación encontrados fueron para agua y orina: 2 y 3 µg/L; 5 y 8 µg/L respectivamente y el rango dinámico de trabajo establecido fue desde 5 a 75 µg/L, permitiendo cuantificar As en muestras de agua cercanos a los estándares internacionales vigentes para valores máximos de As en agua de consumo y en orina en niveles comparables con los establecidos en población laboralmente no expuesta. Esta propuesta metodológica es una alternativa para evaluar la exposición al As en muestras de agua y orina, sin necesidad de utilizar prolongados pre-tratamientos de muestra, de forma más rápida y económica.
Arsenic (As) is a natural contaminant that affects a large area of Argentina. Quantification of As in drinking water has been used to evaluate the source of exposure and As in urine to assess exposure to this toxic. This study aimed to optimize and validate a methodological technique for the quantification of As [As sum = inorganic As (AsI) + methylated species: monometilarsonic acid (MMA) and dimetilarsinic acid (DMA)], product of AsI metabolism by flow injection hydridegeneration-atomic absorption spectrometry (FI-GH-AAS), after derivatization with L-cysteine. The recovery of the studied species: AsI (AsIII and AsV), MMA and DMA was close to 100% in all cases. The limits of detection and quantitation were foundfor water and urine: 2 and 3 µg/L; 5 and 8 µg/L respectively and a linear working range from 5 to 75 µg/L, allowing quantify As in water close to international standards of maximum As values for drinking water and urine samples with levels comparables with those found in people non exposed ocupacionally . This methodology is a valid alternative for assessing exposure to As in water and urine samples without the need of prolonged pre-treatment sample, more quickly and inexpensively.