Os objetivos do presente estudo foram os de verificar a validade do método de reação em cadeia da polimerase em tempo real (qPCR) para detectar e quantificar o Staphylococcus aureus em amostras de leite conservadas com bronopol oriundas de quartos mamários bovinos subclinicamente infectados, e de avaliar os efeitos da presença e da quantidade de células da bactéria sobre a contagem de células somáticas (CCS), a composição do leite (lactose, gordura, proteína bruta, proteína verdadeira e caseína), e a produção de leite de quartos mamários bovinos subclinicamente infectados pelo patógeno. Para a quantificação do S. aureus e das células somáticas bovinas por meio do qPCR, foi utilizado leite cru bovino para o preparo dos padrões como meio de diluição da inoculação seriada de células somáticas e do S. aureus ATCC 29213, e construídas as equações log10UFC = 37,86 23,54 log10CtSAU e log10CCS = 49,3 - 34,0 log10CtBMCB, com base nos resultados obtidos pelas metodologias de referência para cada procedimento. Para testar a equivalência dessas equações aos respectivos métodos de referência, determinar a sensibilidade e especificidade analíticas e a repetibilidade do método proposto, foram coletadas amostras de leite dos quartos mamários de 60 animais de 2 rebanhos leiteiros da região de Pirassununga dos quais se determinou previamente a ocorrência de casos subclínicos de mastite por S. aureus. Dos quartos mamários também foram mensuradas as produções e coletadas amostras de leite para análise de composição, diagnóstico da mastite, e determinação da sensibilidade e especificidade diagnósticas do procedimento de qPCR estabelecido no estudo. Cada amostra foi submetida à análise de composição, CCS, cultura microbiológica, contagem em placas do S. aureus, processadas para a extração do DNA genômico bovino e do S. aureus, e submetida à reação de qPCR. Para análise da concordância entre os resultados obtidos pelos métodos de referência para o diagnóstico da mastite por S. aureus e o de qPCR foi utilizado o teste Kappa. Para avaliação da equivalência das contagens obtidas pelos métodos de referência do S. aureus e de células somáticas bovinas, foi utilizado o teste das diferenças de Bland-Altman. Para a identificação do efeito da infecção subclínica pelo S. aureus sobre a composição e produção de leite do quarto mamário afetado foi utilizada a análise da variância num delineamento em parcelas subdivididas em faixas. Para estimar o grau de relação entre as contagens de S. aureus, a CCS, produção e composição do leite produzido pelo quarto mamário afetado foi utilizado o coeficiente de correlação de Pearson. A correlação entre os resultados de contagem de células somáticas bovinas determinados pelos métodos de rotina e de qPCR para a quantificação de células somáticas apresentou coeficiente r = - 0,978 (P < 0,001). A correlação entre os resultados da contagem do S. aureus ATCC 29213 determinados pelos métodos de rotina e de qPCR para a quantificação do patógeno apresentou coeficiente r = - 0,989 (P < 0,001). A especificidade analítica do qPCR para a detecção do S. aureus em amostras de leite frente a Escherichia coli, Enterococcus sp., Pseudomonas aeruginosa, Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, os estafilococos coagulase-negativa, e as espécies coagulase-positiva Staphylococcus hyicus e Staphylococcus intermedius foi de 100%. O método de qPCR aplicado à detecção de Staphylococcus aureus ATCC 29213 em amostras de leite é replicável e apresentou sensibilidade analítica com limite de detecção para a faixa de 10 UFC/mL à 4,2 x 106 UFC/mL. Em amostras de leite conservadas com bronopol provenientes de quartos mamários subclinicamente infectados, o S. aureus pôde ser detectado, mas não pôde ser quantificado pelo método de qPCR. Nessas amostras, a CCS pôde ser determinada de forma equivalente ao método de rotina. A CCS independe da contagem de S. aureus viáveis, mas foi observada correlação linear e negativa entre o número total de células do patógeno e a CCS. A mastite subclínica pelo S. aureus aumentou a CCS nos quartos mamários, mas não alterou a composição do leite. A doença diminuiu a produção de leite e de gordura dos quartos mamários anteriores acometidos pela infeção, mas não se observou efeito da interação entre o posicionamento da glândula e a infecção sobre a produção de leite. Houve correlação entre as concentrações de lactose (r = 0,42; P = 0,0051), de gordura (r = 0,46; P = 0,0016), de produção de gordura (r = 0,49; P = 0,001), e de leite com produção ajustada para o teor de 3,5% de gordura (r = 0,41; P = 0,006), e o número de S. aureus presentes na amostra de leite. / The objectives of this study were to verify the validity of the real-time polymerase chain reaction (qPCR) to detect and quantify Staphylococcus aureus in bronopol-preserved milk samples from subclinically infected mammary quarters, and to assess the effects of the presence and amount of the pathogen on the somatic cell count (SCC), the composition of milk and milk yield of bovine mammary quarters subclinically infected by the pathogen. In order to quantify S. aureus and bovine somatic cells through qPCR, raw bovine milk was used as a means of serial inoculation media of somatic cells and S. aureus ATCC 29213. From that, equations based on the reference methods for each procedure were built, log10UFC = 37,86 23,54 log10CtSAU and log10CCS = 49,3 - 34,0 log10CtBMCB, respectively. To test their equivalence with the reference methods, determine the analytical sensitivity and specificity, and repeatability of the proposed method, milk was sampled from quarters of 60 animals from two dairy herds in Pirassununga, where subclinical S. aureus mastitis cases had been previously diagnosed. Also, quarter milk yield had been measured and samples collected for milk composition analysis, diagnosis of mastitis, sensitivity and specificity of the procedure established in the study had been determined. Each sample was subjected to composition analysis, SCC, microbiological culture, plate counting of S. aureus, DNA extraction, and subjected to qPCR reaction. Agreement between results from reference methods and qPCR for the diagnosis of mastitis by S. aureus was assessed by Kappa test. Equivalence between S. aureus, SCC scores obtained by reference and qPCR was assessed with Bland-Altman procedures. The effect of S. aureus subclinical infection on milk composition and milk yield of affected quarters was measured using a strip plot design. To estimate the degree of relationship between the counts of S. aureus, SCC, yield and composition of the milk from affected quarters was assessed by the Pearson Correlation. Correlation between SCC determined by routine methods and qPCR was r = - 0.978 (P <0.001). Correlation between S. aureus ATCC 29213 determined by routine methods and qPCR was r = - 0.989 (P <0.001). Analytical specificity of qPCR to detect S. aureus in milk samples against Escherichia coli, Enterococcus sp., Pseudomonas aeruginosa, Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, coagulase-negative staphylococci and coagulase-positive species, Staphylococcus hyicus and Staphylococcus intermedius was 100%. The use of the qPCR to detect S. aureus ATCC 29213 in milk samples is replicable. Analytical sensitivity detection limit of the method ranged from 10 CFU/mL to 4.2 x 106 CFU/mL. S. aureus could be detected, but not quantified by qPCR in bronopol-conserved milk samples from subclinically infected quarters. In these samples, SCC could be determined by qPCR as it had been done by routine method. SCC was not dependent on S. aureus viable cells, but a negative linear correlation between the total number of cells of the pathogen and SCC was observed. S. aureus subclinical mastitis increased quarters SCC, but did not change milk composition. The disease decreased quarter milk and fat yield, but no interaction effect was observed between the gland positioning and S. aureus subclinical infection on milk production. Correlations between lactose (r = 0.42, P = 0.0051), fat (r = 0.46, P = 0.0016), fat yield (r = 0.49, P = 0.001), and 3.5% fat adjusted milk yield (r = 0.41, P = 0.006), and the number of S. aureus present in the milk sample were observed.

Os objetivos do presente estudo foram os de verificar a validade do método de reação em cadeia da polimerase em tempo real (qPCR) para detectar e quantificar o Staphylococcus aureus em amostras de leite conservadas com bronopol oriundas de quartos mamários bovinos subclinicamente infectados, e de avaliar os efeitos da presença e da quantidade de células da bactéria sobre a contagem de células somáticas (CCS), a composição do leite (lactose, gordura, proteína bruta, proteína verdadeira e caseína), e a produção de leite de quartos mamários bovinos subclinicamente infectados pelo patógeno. Para a quantificação do S. aureus e das células somáticas bovinas por meio do qPCR, foi utilizado leite cru bovino para o preparo dos padrões como meio de diluição da inoculação seriada de células somáticas e do S. aureus ATCC 29213, e construídas as equações log10UFC = 37,86 23,54 log10CtSAU e log10CCS = 49,3 - 34,0 log10CtBMCB, com base nos resultados obtidos pelas metodologias de referência para cada procedimento. Para testar a equivalência dessas equações aos respectivos métodos de referência, determinar a sensibilidade e especificidade analíticas e a repetibilidade do método proposto, foram coletadas amostras de leite dos quartos mamários de 60 animais de 2 rebanhos leiteiros da região de Pirassununga dos quais se determinou previamente a ocorrência de casos subclínicos de mastite por S. aureus. Dos quartos mamários também foram mensuradas as produções e coletadas amostras de leite para análise de composição, diagnóstico da mastite, e determinação da sensibilidade e especificidade diagnósticas do procedimento de qPCR estabelecido no estudo. Cada amostra foi submetida à análise de composição, CCS, cultura microbiológica, contagem em placas do S. aureus, processadas para a extração do DNA genômico bovino e do S. aureus, e submetida à reação de qPCR. Para análise da concordância entre os resultados obtidos pelos métodos de referência para o diagnóstico da mastite por S. aureus e o de qPCR foi utilizado o teste Kappa. Para avaliação da equivalência das contagens obtidas pelos métodos de referência do S. aureus e de células somáticas bovinas, foi utilizado o teste das diferenças de Bland-Altman. Para a identificação do efeito da infecção subclínica pelo S. aureus sobre a composição e produção de leite do quarto mamário afetado foi utilizada a análise da variância num delineamento em parcelas subdivididas em faixas. Para estimar o grau de relação entre as contagens de S. aureus, a CCS, produção e composição do leite produzido pelo quarto mamário afetado foi utilizado o coeficiente de correlação de Pearson. A correlação entre os resultados de contagem de células somáticas bovinas determinados pelos métodos de rotina e de qPCR para a quantificação de células somáticas apresentou coeficiente r = - 0,978 (P < 0,001). A correlação entre os resultados da contagem do S. aureus ATCC 29213 determinados pelos métodos de rotina e de qPCR para a quantificação do patógeno apresentou coeficiente r = - 0,989 (P < 0,001). A especificidade analítica do qPCR para a detecção do S. aureus em amostras de leite frente a Escherichia coli, Enterococcus sp., Pseudomonas aeruginosa, Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, os estafilococos coagulase-negativa, e as espécies coagulase-positiva Staphylococcus hyicus e Staphylococcus intermedius foi de 100%. O método de qPCR aplicado à detecção de Staphylococcus aureus ATCC 29213 em amostras de leite é replicável e apresentou sensibilidade analítica com limite de detecção para a faixa de 10 UFC/mL à 4,2 x 106 UFC/mL. Em amostras de leite conservadas com bronopol provenientes de quartos mamários subclinicamente infectados, o S. aureus pôde ser detectado, mas não pôde ser quantificado pelo método de qPCR. Nessas amostras, a CCS pôde ser determinada de forma equivalente ao método de rotina. A CCS independe da contagem de S. aureus viáveis, mas foi observada correlação linear e negativa entre o número total de células do patógeno e a CCS. A mastite subclínica pelo S. aureus aumentou a CCS nos quartos mamários, mas não alterou a composição do leite. A doença diminuiu a produção de leite e de gordura dos quartos mamários anteriores acometidos pela infeção, mas não se observou efeito da interação entre o posicionamento da glândula e a infecção sobre a produção de leite. Houve correlação entre as concentrações de lactose (r = 0,42; P = 0,0051), de gordura (r = 0,46; P = 0,0016), de produção de gordura (r = 0,49; P = 0,001), e de leite com produção ajustada para o teor de 3,5% de gordura (r = 0,41; P = 0,006), e o número de S. aureus presentes na amostra de leite. / The objectives of this study were to verify the validity of the real-time polymerase chain reaction (qPCR) to detect and quantify Staphylococcus aureus in bronopol-preserved milk samples from subclinically infected mammary quarters, and to assess the effects of the presence and amount of the pathogen on the somatic cell count (SCC), the composition of milk and milk yield of bovine mammary quarters subclinically infected by the pathogen. In order to quantify S. aureus and bovine somatic cells through qPCR, raw bovine milk was used as a means of serial inoculation media of somatic cells and S. aureus ATCC 29213. From that, equations based on the reference methods for each procedure were built, log10UFC = 37,86 23,54 log10CtSAU and log10CCS = 49,3 - 34,0 log10CtBMCB, respectively. To test their equivalence with the reference methods, determine the analytical sensitivity and specificity, and repeatability of the proposed method, milk was sampled from quarters of 60 animals from two dairy herds in Pirassununga, where subclinical S. aureus mastitis cases had been previously diagnosed. Also, quarter milk yield had been measured and samples collected for milk composition analysis, diagnosis of mastitis, sensitivity and specificity of the procedure established in the study had been determined. Each sample was subjected to composition analysis, SCC, microbiological culture, plate counting of S. aureus, DNA extraction, and subjected to qPCR reaction. Agreement between results from reference methods and qPCR for the diagnosis of mastitis by S. aureus was assessed by Kappa test. Equivalence between S. aureus, SCC scores obtained by reference and qPCR was assessed with Bland-Altman procedures. The effect of S. aureus subclinical infection on milk composition and milk yield of affected quarters was measured using a strip plot design. To estimate the degree of relationship between the counts of S. aureus, SCC, yield and composition of the milk from affected quarters was assessed by the Pearson Correlation. Correlation between SCC determined by routine methods and qPCR was r = - 0.978 (P <0.001). Correlation between S. aureus ATCC 29213 determined by routine methods and qPCR was r = - 0.989 (P <0.001). Analytical specificity of qPCR to detect S. aureus in milk samples against Escherichia coli, Enterococcus sp., Pseudomonas aeruginosa, Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, coagulase-negative staphylococci and coagulase-positive species, Staphylococcus hyicus and Staphylococcus intermedius was 100%. The use of the qPCR to detect S. aureus ATCC 29213 in milk samples is replicable. Analytical sensitivity detection limit of the method ranged from 10 CFU/mL to 4.2 x 106 CFU/mL. S. aureus could be detected, but not quantified by qPCR in bronopol-conserved milk samples from subclinically infected quarters. In these samples, SCC could be determined by qPCR as it had been done by routine method. SCC was not dependent on S. aureus viable cells, but a negative linear correlation between the total number of cells of the pathogen and SCC was observed. S. aureus subclinical mastitis increased quarters SCC, but did not change milk composition. The disease decreased quarter milk and fat yield, but no interaction effect was observed between the gland positioning and S. aureus subclinical infection on milk production. Correlations between lactose (r = 0.42, P = 0.0051), fat (r = 0.46, P = 0.0016), fat yield (r = 0.49, P = 0.001), and 3.5% fat adjusted milk yield (r = 0.41, P = 0.006), and the number of S. aureus present in the milk sample were observed.

O crescimento da população mundial e a expansão industrial são fatores que atualmente desencadeiam um aumento no consumo de produtos e geração de resíduos. Frequentemente utilizados na medicina tradicional e veterinária, os antibióticos são classificados como uma classe emergente de poluentes, pois, devido à sua baixa taxa de degradação e à ineficácia dos sistemas de tratamentos de água, encontram-se disseminados no meio ambiente e nos alimentos. Em consequência desse fato, estudos para o desenvolvimento de novos materiais e métodos que removam estes contaminantes utilizando menor quantidade de consumíveis químicos e a baixo custo, têm sido um campo em desenvolvimento na química analítica. Inserido nesse contexto encontra-se o grafeno, que devido à sua estrutura de anéis aromáticos com elétrons &pi; deslocalizados e elevada área superficial (3600 m2/g), pode favorecer fenômenos de sorção, elevando a eficiência na extração de contaminantes presentes em matrizes complexas. O objetivo do projeto foi sintetizar e caracterizar novas fases sorventes à base de grafeno para atuar no preparo da amostra avaliando a eficácia destas na remoção de tetraciclinas presentes em leite. Para isso, utilizou-se da microextração por sorvente empacotado MEPS como método de preparo de amostra com posterior análise por cromatografia líquida acoplada a espectrometria de massas HPLC-MS/MS. A caracterização dos materiais foi feita através de microscopia eletrônica de varredura MEV e espectroscopia vibracional na região do infravermelho IV revelando eficácia na síntese das folhas de óxido de grafeno, assim como, no recobrimento das partículas de sílica com os novos materiais. Foram avaliadas as fases extratoras: G, GO-SIL, G-Sil e C18-G-Sil com relação a eficiência na extração das tetraciclinas. A fase G-Sil mostrou-se a mais apropriada para continuidade do estudo, pois apresentou os melhores resultados. Com o objetivo de otimizar a etapa de preparo de amostra, foi feito um planejamento fatorial fracionário 24-1 avaliando as variáveis químicas: solvente de eluição, força iônica, pH da solução de lavagem e inclusão de EDTA na solução tampão de Mcllvaine (pH 4.2). Posteriormente foi feito um planejamento fatorial completo 23 visando a otimização dos ciclos de extração da MEPS, possibilitando determinar a condição ótima de extração do método. Baseando-se no documento: ICH - Q2 (R1) guideline, a metodologia foi submetida a testes para avaliar as figuras de mérito referentes a validação analítica. O método apresentou faixa linear de 15 - 110 &micro;g/L, LQ entre 0,05 e 0,9 &micro;g/L e boa seletividade. A precisão foi avaliada intra e inter-dia apresentando desvio padrão relativo - RSD abaixo de 18%. Ao final do desenvolvimento do estudo foram analisadas onze amostras reais incluindo leite bovino, caprino, ovino e leite em pó. A aplicação em amostras de diferentes espécies reforça a versatilidade do método desenvolvido, além de evidenciar o elevado potencial dos novos materiais de grafeno para atuar como sorvente no preparo da amostra. / Nowadays, the rise of world population and industrial expansion are leading to an increase in consumption and consequently waste generation. Large quantities of chemical residues are released into the environment affecting humans and wildlife. Among these residues, antibiotics used in medicine and veterinary are classified as an emerging contaminant due to their low degradation rate, ineffectiveness of wastewater treatment and consequently water and food contamination. An effort has been made to develop microextraction techniques to remove these drugs using reduce volume of solvent and chemical consumable and researches of new sorbent phases are increasing due the high potential to improve these microextraction techniques. Recent studies pinpoint graphene (G) as an effectively sorbent phase to act in sample preparation. G has a structure composed by carbon nanosheets arranged in a honeycomb pattern with large surface area (3600 m2 /g) and delocalized &pi;-electrons system that suggests good interaction between them and contaminants compounds such as veterinary drugs, pesticides, personal care products etc. This study focuses to evaluate sorbent phases based upon graphene particle supported on silica to analyze Tetracycline residues present in milk. For this purpose, was used microextraction by packed sorbent - MEPS as sample preparation method with analysis by liquid chromatography and mass spectrometry. The characterization of synthesized materials was performed by scanning electron microscopy - SEM and vibrational spectroscopy in infrared region - FTIR. The results suggest that synthesis of graphene\'s particles as well as the coating of silica surface with these materials were both performed. To evaluate extraction efficiency each sorbent material: GO-Sil, G-Sil, G, C18-GO-Sil were tested in MEPS and confronted to commercial phases: C8 and C18. By the results, G-Sil was choose to application as sorbent phase. MEPS extractions were optimized and the effect of some factors was investigated by application of experimental design. Firstly, a factorial experimental design 2(4-1) was executed to evaluate chemistry variables such as elution solvent, washing solution, media ionic strength and inclusion of EDTA salt in Mcllvaine solution. After, an experimental design 23 was made in order to estimate the cycles of MEPS extraction like sampling, washing and elution steps. By the results, an optimized extraction conditional was achieved and evaluation of validation parameters was carried out on sequence based on the ICH - Q2 (R1) guide. The method showed linearity ranging from 15 - 110 &micro;g/L, LOQ values from 0,05 to 0,9 &micro;g/L and good selectivity. Precision showed relative standard deviation RSD less than 18% to intra and inter-day analysis. The developed method was applied in analysis of eleven milk samples including bovine, caprine and ovine milk and milk powder. Application in samples from different species enhances the versatility of this analytical method and show the great potential for graphene\'s particles to act as sorbent phase in sample preparation.

O crescimento da população mundial e a expansão industrial são fatores que atualmente desencadeiam um aumento no consumo de produtos e geração de resíduos. Frequentemente utilizados na medicina tradicional e veterinária, os antibióticos são classificados como uma classe emergente de poluentes, pois, devido à sua baixa taxa de degradação e à ineficácia dos sistemas de tratamentos de água, encontram-se disseminados no meio ambiente e nos alimentos. Em consequência desse fato, estudos para o desenvolvimento de novos materiais e métodos que removam estes contaminantes utilizando menor quantidade de consumíveis químicos e a baixo custo, têm sido um campo em desenvolvimento na química analítica. Inserido nesse contexto encontra-se o grafeno, que devido à sua estrutura de anéis aromáticos com elétrons &pi; deslocalizados e elevada área superficial (3600 m2/g), pode favorecer fenômenos de sorção, elevando a eficiência na extração de contaminantes presentes em matrizes complexas. O objetivo do projeto foi sintetizar e caracterizar novas fases sorventes à base de grafeno para atuar no preparo da amostra avaliando a eficácia destas na remoção de tetraciclinas presentes em leite. Para isso, utilizou-se da microextração por sorvente empacotado MEPS como método de preparo de amostra com posterior análise por cromatografia líquida acoplada a espectrometria de massas HPLC-MS/MS. A caracterização dos materiais foi feita através de microscopia eletrônica de varredura MEV e espectroscopia vibracional na região do infravermelho IV revelando eficácia na síntese das folhas de óxido de grafeno, assim como, no recobrimento das partículas de sílica com os novos materiais. Foram avaliadas as fases extratoras: G, GO-SIL, G-Sil e C18-G-Sil com relação a eficiência na extração das tetraciclinas. A fase G-Sil mostrou-se a mais apropriada para continuidade do estudo, pois apresentou os melhores resultados. Com o objetivo de otimizar a etapa de preparo de amostra, foi feito um planejamento fatorial fracionário 24-1 avaliando as variáveis químicas: solvente de eluição, força iônica, pH da solução de lavagem e inclusão de EDTA na solução tampão de Mcllvaine (pH 4.2). Posteriormente foi feito um planejamento fatorial completo 23 visando a otimização dos ciclos de extração da MEPS, possibilitando determinar a condição ótima de extração do método. Baseando-se no documento: ICH - Q2 (R1) guideline, a metodologia foi submetida a testes para avaliar as figuras de mérito referentes a validação analítica. O método apresentou faixa linear de 15 - 110 &micro;g/L, LQ entre 0,05 e 0,9 &micro;g/L e boa seletividade. A precisão foi avaliada intra e inter-dia apresentando desvio padrão relativo - RSD abaixo de 18%. Ao final do desenvolvimento do estudo foram analisadas onze amostras reais incluindo leite bovino, caprino, ovino e leite em pó. A aplicação em amostras de diferentes espécies reforça a versatilidade do método desenvolvido, além de evidenciar o elevado potencial dos novos materiais de grafeno para atuar como sorvente no preparo da amostra. / Nowadays, the rise of world population and industrial expansion are leading to an increase in consumption and consequently waste generation. Large quantities of chemical residues are released into the environment affecting humans and wildlife. Among these residues, antibiotics used in medicine and veterinary are classified as an emerging contaminant due to their low degradation rate, ineffectiveness of wastewater treatment and consequently water and food contamination. An effort has been made to develop microextraction techniques to remove these drugs using reduce volume of solvent and chemical consumable and researches of new sorbent phases are increasing due the high potential to improve these microextraction techniques. Recent studies pinpoint graphene (G) as an effectively sorbent phase to act in sample preparation. G has a structure composed by carbon nanosheets arranged in a honeycomb pattern with large surface area (3600 m2 /g) and delocalized &pi;-electrons system that suggests good interaction between them and contaminants compounds such as veterinary drugs, pesticides, personal care products etc. This study focuses to evaluate sorbent phases based upon graphene particle supported on silica to analyze Tetracycline residues present in milk. For this purpose, was used microextraction by packed sorbent - MEPS as sample preparation method with analysis by liquid chromatography and mass spectrometry. The characterization of synthesized materials was performed by scanning electron microscopy - SEM and vibrational spectroscopy in infrared region - FTIR. The results suggest that synthesis of graphene\'s particles as well as the coating of silica surface with these materials were both performed. To evaluate extraction efficiency each sorbent material: GO-Sil, G-Sil, G, C18-GO-Sil were tested in MEPS and confronted to commercial phases: C8 and C18. By the results, G-Sil was choose to application as sorbent phase. MEPS extractions were optimized and the effect of some factors was investigated by application of experimental design. Firstly, a factorial experimental design 2(4-1) was executed to evaluate chemistry variables such as elution solvent, washing solution, media ionic strength and inclusion of EDTA salt in Mcllvaine solution. After, an experimental design 23 was made in order to estimate the cycles of MEPS extraction like sampling, washing and elution steps. By the results, an optimized extraction conditional was achieved and evaluation of validation parameters was carried out on sequence based on the ICH - Q2 (R1) guide. The method showed linearity ranging from 15 - 110 &micro;g/L, LOQ values from 0,05 to 0,9 &micro;g/L and good selectivity. Precision showed relative standard deviation RSD less than 18% to intra and inter-day analysis. The developed method was applied in analysis of eleven milk samples including bovine, caprine and ovine milk and milk powder. Application in samples from different species enhances the versatility of this analytical method and show the great potential for graphene\'s particles to act as sorbent phase in sample preparation.

<p>Cilj&nbsp; ove&nbsp; doktorske&nbsp; disertacije&nbsp; je&nbsp; bio&nbsp; razvoj&nbsp; brzih&nbsp; i&nbsp; pouzdanih&nbsp; voltametrijskih&nbsp; metoda zasnovanih na primeni jednostavnih i savremenih elektroda/senzora na bazi ugljeničnih materijala (ugljenične&nbsp; paste&nbsp; napravljene&nbsp; od&nbsp; grafitnog&nbsp; praha&nbsp; i&nbsp; parafinskog&nbsp; ulja&nbsp; i&nbsp; &scaron;tampanih&nbsp; ugljeničnih elektroda)&nbsp; za&nbsp; određivanje&nbsp; H ₂O₂u&nbsp; odabranim&nbsp; složenim&nbsp; model&nbsp; i&nbsp; realnim&nbsp; uzorcima.&nbsp; U&nbsp; tu&nbsp; svrhu, ispitana je primenljivost različitih radnih elektroda. Amperometrijska&nbsp; metoda&nbsp; zasnovana&nbsp; na&nbsp; elektrodi&nbsp; od&nbsp; ugljenične&nbsp; paste&nbsp; (CPE)&nbsp; zapreminski modifikovane&nbsp; sa&nbsp; 5%&nbsp; (m:m)&nbsp; MnO2 je,&nbsp; pri&nbsp; optimizovanim&nbsp; uslovima&nbsp; i&nbsp; pri&nbsp; radnom&nbsp; potencijalu&nbsp; od 0,40 V&nbsp; u&nbsp; odnosu&nbsp; na&nbsp; zasićenu&nbsp; kalomelovu&nbsp; elektrodu&nbsp; (ZKE)&nbsp; u&nbsp; fosfatnom&nbsp; puferu&nbsp; pH&nbsp; 7,50&nbsp; kao pomoćnom elektrolitu, omogućila kvantifikaciju H ₂O₂ u opsegu koncentracija od 1,4 do 65 &micro;g mL -1 sa&nbsp; relativnom&nbsp; standardnom&nbsp; devijacijom&nbsp; (RSD)&nbsp; manjom&nbsp; od&nbsp; 10%.&nbsp; Ova&nbsp; metoda&nbsp; je&nbsp; primenjena&nbsp; za određivanje sadržaja H ₂O₂u uzorcima podzemne vode iz centralnog Banata (Pokrajina Vojvodina, Srbija)&nbsp; tretirane&nbsp; Fentonovim&nbsp; (Fe²⁺ i&nbsp; H ₂O₂ )&nbsp; i&nbsp; Fentonu-sličnim&nbsp; (Fe ³⁺ i&nbsp; H ₂O₂ )&nbsp; reagensima&nbsp; u&nbsp; cilju uklanjanja&nbsp; prirodnih&nbsp; organskih&nbsp; materija&nbsp; (POM)&nbsp; pri&nbsp; čemu&nbsp; su&nbsp; kori&scaron;ćene&nbsp; različite&nbsp; početne koncentracije&nbsp; gvožđa&nbsp; i&nbsp; različiti&nbsp; odnosi&nbsp; molarnih&nbsp; koncentracija&nbsp; gvožđa&nbsp; i&nbsp; H ₂O₂ .&nbsp; Utvrđeno&nbsp; je&nbsp; da oksidaciono&nbsp; stanje&nbsp; gvožđe&nbsp; (Fe²⁺ ili&nbsp; Fe ³⁺)&nbsp; i&nbsp; molarni&nbsp; odnos&nbsp; jona&nbsp; Fe&nbsp; i&nbsp; H₂O₂ utiču&nbsp; na&nbsp; stepen potro&scaron;nje/razgradnje&nbsp; H₂O₂<br />u&nbsp; podzemnoj&nbsp; vodi&nbsp; sa&nbsp; visokim&nbsp; sadržajem&nbsp; POM.&nbsp; Takođe,&nbsp; u&nbsp; slučaju Fentonu-sličnog&nbsp; procesa,&nbsp; za&nbsp; sve&nbsp; početne&nbsp; koncentracije&nbsp; Fe ³⁺ i&nbsp; H ₂O_{2 ,}&nbsp; signifikantna&nbsp; količina&nbsp; H ₂O₂ostaje&nbsp; neiskori&scaron;ćena,&nbsp; &scaron;to&nbsp; ukazuje&nbsp; na&nbsp; nižu&nbsp; efikasnost&nbsp; ovakvog&nbsp; sistema u&nbsp; poređenju&nbsp; sa&nbsp; Fentonovim procesom.&Scaron;tampana&nbsp; ugljenična&nbsp; elektroda&nbsp; (SPCE)&nbsp; zapreminski&nbsp; modifikovana&nbsp; sa&nbsp; MnO 2 kao medijatorom&nbsp; je&nbsp; primenjena&nbsp; za&nbsp; određivanje&nbsp; sadržaja&nbsp; H₂O₂ u&nbsp; toku&nbsp; Fentonovog&nbsp; (Fe ²⁺ ,&nbsp; H₂O₂ )&nbsp; i vidljivom&nbsp; svetlo&scaron;ću&nbsp; potpomognutog&nbsp; foto-Fentonovog&nbsp; (Fe²⁺,&nbsp; H ₂O₂,&nbsp; h&nu;)&nbsp; procesa&nbsp; uklanjanja neonikotinoidnog&nbsp; insekticida&nbsp; acetamiprida&nbsp; (ACT).&nbsp; Pri&nbsp; optimizovanim&nbsp; uslovima&nbsp; (radni&nbsp; potencijal 0,40&nbsp; V&nbsp; u&nbsp; odnosu&nbsp; na&nbsp; ZKE,&nbsp; fosfatni&nbsp; pufer&nbsp; pH&nbsp; 7,50&nbsp; kao&nbsp; pomoćni&nbsp; elektrolit)&nbsp; amperometrijskog određivanja&nbsp; H ₂O₂ ,&nbsp; postignuta&nbsp; je&nbsp; linearnost&nbsp; u&nbsp; opsegu&nbsp; koncentracija&nbsp; 0,01&ndash;1,24&nbsp; mmol&nbsp; L-1(0,34&ndash; 42,2 &micro;g mL -1<br />) i vrednost RSD nije prelazila 4,2%. U ispitivanimuzorcima (nakon odgovarajućeg<br />pode&scaron;avanja pH vrednosti od 2,8 do 7,5 odmah nakon&nbsp; uzorkovanja radi stopiranja ili maksimalnog usporavanja&nbsp; procesa&nbsp; oksidacije,&nbsp; filtriranja,&nbsp; zamrzavanja&nbsp; i&nbsp; odmrzavanja&nbsp; neposredno&nbsp; pre&nbsp; merenja) sadržaj&nbsp; H ₂O₂ je&nbsp; određen&nbsp; metodom&nbsp; standardnog&nbsp; dodatka&nbsp; analiziranjem&nbsp; odgovarajućih amperometrijskih&nbsp; krivi.&nbsp; Paralelna&nbsp; HPLC-DAD&nbsp; merenja&nbsp; su&nbsp; vr&scaron;ena&nbsp; u&nbsp; cilju&nbsp; praćenja koncentracije/uklanjanja ACT. U slučaju foto-Fentonovog procesa (početne koncentracije 0,31; 2,0 i 3,0 mmol L -1 (70,0; 111,7 i 102,1 &micro;g mL -1 ) za ACT, Fe<br />²⁺ i H ₂O₂, redom) nakon 10 min H ₂O₂je izreagovao, a može se smatrati da je ACT uklonjen nakon 5 min. U toku Fentonovog procesa ACT je&nbsp; uklonjen&nbsp; nakon&nbsp; 20&nbsp; min&nbsp; tretmana&nbsp; i&nbsp; oko&nbsp; 10%&nbsp; početne&nbsp; koncentracije&nbsp; H ₂O₂ je&nbsp; ostalo&nbsp; u&nbsp; sistemu<br />neiskori&scaron;ćeno.CPE&nbsp; je&nbsp; povr&scaron;inski&nbsp; modifikovane&nbsp; kompozitom&nbsp; na&nbsp; bazi&nbsp; nanočestica&nbsp; Pt&nbsp; (&lt;&nbsp; 5&nbsp; nm)&nbsp; i grafitizovanog&nbsp; ugljenika&nbsp; (Pt-C,&nbsp; 10%&nbsp; Pt&nbsp; na&nbsp; Vulkanu&nbsp; XC72)&nbsp;&nbsp; etodom&nbsp; nano&scaron;enja&nbsp; kapi. Nemodifikovana&nbsp; CPE&nbsp; i&nbsp; modifikovana&nbsp; (Pt-C/CPE)&nbsp; su&nbsp;&nbsp; okarakterisane&nbsp; primenom&nbsp; SEM/EDS&nbsp; i&nbsp; CV merenja.&nbsp; Pt-C/CPE&nbsp; je&nbsp; pokazala&nbsp; izuzetne&nbsp; elektrokatalitičke&nbsp; osobine&nbsp; u&nbsp; pogledu&nbsp; elektrohemijskeredoks&nbsp; reakcije&nbsp; H<br />₂O₂ u&nbsp; poređenju&nbsp; sa&nbsp; nemodifikovanom&nbsp; CPE&nbsp; u&nbsp; fosfatnom&nbsp; puferu&nbsp; (0,1 mol&nbsp; L -1 ;<br />pH 7,50),&nbsp; a&nbsp; takođe&nbsp; i&nbsp; u&nbsp; acetatnom&nbsp; puferu&nbsp; (0,1&nbsp; mol&nbsp; L -1 ;&nbsp; pH&nbsp; 4,50)&nbsp; kao&nbsp;&nbsp; pomoćnim&nbsp; elektrolitima. Prilikom&nbsp; amperometrijskog&nbsp; određivanja&nbsp; H ₂O₂ primenom&nbsp; Pt-C/CPE&nbsp; u&nbsp; model&nbsp; sistemima, zadovoljavajuća linearnost je postignuta u koncentracionom opseguH₂O₂od 0,15 do 1,45 &micro;g mL -1 ,dok su vrednosti GO iznosile 0,06 &micro;g mL -1 (pH 7,50, radni potencijal 0,20 V) i 0,10 &micro;g mL -1 (pH<br />4,50,&nbsp; radni&nbsp; potencijal&nbsp; 0,50&nbsp; V).&nbsp; Optimizovane&nbsp; analitičke&nbsp; metode&nbsp; su&nbsp; primenjene&nbsp; za&nbsp; određivanje sadržaja H ₂O₂ u komercijalno dostupnim proizvodima za ličnu negu: rastvoru za dezinfekciju (pH 7,50)&nbsp; i&nbsp; rastvoru&nbsp; za&nbsp; či&scaron;ćenje&nbsp; kontaktnih&nbsp; sočiva&nbsp; (pH&nbsp; 4,50).&nbsp; Amperometrijski&nbsp; dobijeni&nbsp; rezultati&nbsp; su&nbsp; u dobrom&nbsp; slaganju&nbsp; sa&nbsp; rezultatima&nbsp; dobijenim&nbsp; primenom&nbsp; tradicionalne&nbsp; spektrofotometrijske&nbsp; metode bazirane&nbsp; na&nbsp; titanijum-sulfatu&nbsp; kao&nbsp; reagensu&nbsp; sa&nbsp; određenim&nbsp; koncentracijama&nbsp; 2,91%&nbsp; i&nbsp; 2,94%&nbsp; za<br />dezinfekcioni rastvor i 3,04% i 3,17% za rastvor zakontaktna sočiva, redom. RSD je bila manja od 2%.&nbsp; Postignuti&nbsp; rezultati&nbsp; su&nbsp; u&nbsp; dobrom&nbsp; slaganju&nbsp; sa&nbsp; sadržajem&nbsp; H<br />₂O₂deklarisanim&nbsp; od&nbsp; strane proizvođača (3%) u oba ispitivana uzorka. Pt-C/CPE je takođe testirana za praćenje koncentracije H₂O₂ u rastvoru za kontaktna sočiva u toku procesa njegove neutralizacije/razgradnje. Nakon 6 h procesa neutralizacije, 24,68&nbsp; &micro;g mL -1 je bila određena koncentracija H ₂O_{2 ,} &scaron;to je ispod dozvoljene<br />H₂O₂koncentracije u rastvoru za kontaktna sočiva imajući u vidu granicu koja izaziva iritaciju oka. CPE&nbsp; je&nbsp; povr&scaron;inski&nbsp; modifikovana&nbsp; vi&scaron;ezidnim&nbsp; ugljeničnim&nbsp; nanocevima&nbsp; (MWCNT)&nbsp; i kompozitima MnO 2 -MWCNT ili Pt-MWCNT metodom nano&scaron;enja kapi radi pripreme jednostavnih, osetljivih i pouzdanih voltametrijskih senzora za&nbsp; određivanje H ₂O₂u odabranom uzorku. Rezultati SEM/EDS&nbsp; analize&nbsp; kompozitnih&nbsp; materijala&nbsp; su&nbsp; potvrdili da&nbsp; su&nbsp; medijatori,&nbsp; čestice&nbsp; MnO 2 i&nbsp; Pt, nasumično&nbsp; raspoređeni na povr&scaron;ini MWCNT i zastupljeni sa blizu 5% (m:m) u kompozitu izraženo<br />preko Mn i Pt. CV merenja su vr&scaron;ena sa pripremljenim radnim elektrodama u acetatnom (pH 4,50), fosfatnom&nbsp; (pH&nbsp; 7,50)&nbsp; i&nbsp; boratnom&nbsp; (pH&nbsp; 9,18)&nbsp; puferu&nbsp; kako&nbsp; bi&nbsp; se&nbsp; okarakterisalo&nbsp; osnovno elektrohemijsko&nbsp; pona&scaron;anje&nbsp; H ₂O₂i&nbsp; odabrali&nbsp; pogodni&nbsp; radni&nbsp; potencijali&nbsp; za&nbsp; amperometrijsko određivanje ovog ciljnog analita. Pt- WCNT/CPE je primenljiva za rad kako u fosfatnom puferu pH&nbsp; 7,50&nbsp; tako&nbsp; i&nbsp; u&nbsp; acetatnom&nbsp; puferu&nbsp; pH&nbsp; 4,50&nbsp; V&nbsp; kako&nbsp; pri&nbsp; negativnim&nbsp; tako&nbsp; i&nbsp; pri&nbsp; pozitivnim&nbsp; radnim<br />potencijalima, pri&nbsp; čemu su vrednosti RSD uglavnom ispod 2,5%. U slučaju MnO 2<br />-MWCNT/CPE, na&nbsp; potencijalu&nbsp; 0,30&nbsp; V&nbsp; i&nbsp; vi&scaron;im&nbsp; vrednostima,&nbsp; oksidacioni&nbsp; signali&nbsp; H<br />₂O₂ su&nbsp; signifikantni&nbsp; u&nbsp; blago alkalnoj sredini (pH 7,50), pri pH 4,50 ova elektroda pokazuje nezadovoljavajuće pona&scaron;anje, dok&nbsp; pri pH 9,18 ima prihvatljive performanse. Granice određivanja (GO) su bile u oblasti &micro;g mL -1 . H ₂O₂ je&nbsp; određen&nbsp; u&nbsp; spajkovanom&nbsp; uzorku&nbsp; mleka&nbsp; metodom&nbsp; standardnog&nbsp; dodatka&nbsp; nakon&nbsp; odgovarajuće pripreme&nbsp; uzorka&nbsp; (pH&nbsp; pode&scaron;avanje&nbsp; i&nbsp; centrifugiranje)&nbsp; i&nbsp; primenom&nbsp; optimizovane&nbsp; amperometrijske procedure (acetatni pufer pH 4,50, radni&nbsp; potencijal-0,75 V) koristeći Pt-MWCNT/CPE kao radnu elektrodu. RSD za tri&nbsp; ponovljena merenja je iznosila 2,5%, dok je prinos metode bio ne&scaron;to veći od 71%.<br />Kompozitni materijali koji se sastoje od MWCNT i&nbsp; čestica na bazi Pd (Pd-MWCNT) ili Pt (Pt-MWCNT) su primenjeni za pripremu zapreminski modifikovanih SPCE (Pd-MWCNT-SPCE i Pt-MWCNT-SPCE)&nbsp; i&nbsp; povr&scaron;inski&nbsp; modifikovane&nbsp; SPCE&nbsp; (Pd-MWCNT/SPCE).&nbsp; Ove&nbsp; elektrode,&nbsp; kao&nbsp; i nemodifikovana&nbsp; SPCE&nbsp; i&nbsp; MWCNT-SPCE,&nbsp; su&nbsp; okarakterisane primenom&nbsp; CV&nbsp; i&nbsp; amperometrije&nbsp; u fosfatnom puferu pH 7,50 radi određivanja H ₂O₂ . Pd-MWCNT-SPCE i Pd-MWCNT/SPCE su se pokazale&nbsp; pogodnim&nbsp; za&nbsp; određivanje&nbsp; H ₂O₂ na&nbsp; radnim&nbsp; potencijalima&nbsp; između&nbsp; -0,50&nbsp; i&nbsp; 0,50&nbsp; V,&nbsp; a&nbsp; PtMWCNT-SPCE na ispitivanim radnim potencijalima od -0,80 do 0,70 V. Ove&nbsp; elektrode su zatim modifikovane&nbsp; enzimom&nbsp; glukoza&nbsp; oksidazom&nbsp; (GOx)&nbsp; metodom&nbsp; nano&scaron;enja&nbsp; kapi&nbsp; rastvora&nbsp; GOx&nbsp; i Nafion &reg; -a&nbsp; na&nbsp; njihovu&nbsp; povr&scaron;inu,&nbsp; pri&nbsp; čemu&nbsp; je&nbsp; optimizovana&nbsp; količina&nbsp; nanetog&nbsp; biofilma.&nbsp; GOx/PdMWCNT-SPCE&nbsp; je&nbsp; pokazala&nbsp; bolje&nbsp; analitičke&nbsp; performanse&nbsp; za&nbsp; određivanje&nbsp; glukoze&nbsp; u&nbsp; poređenju&nbsp; sa GOx/Pd- WCNT/SPCE.&nbsp; Kao&nbsp; optimalan&nbsp; radni&nbsp; potencijal&nbsp; GOx/Pd-MWCNT-SPCE&nbsp; je&nbsp; odabrana<br />vrednost&nbsp; potencijala&nbsp; -0,40&nbsp; V&nbsp; u&nbsp; odnosu&nbsp; na&nbsp; ZKE,&nbsp; sa&nbsp; zadovoljavajućom&nbsp; linearno&scaron;ću&nbsp; u&nbsp; ispitivanom opsegu&nbsp; koncentracija&nbsp; glukoze&nbsp; od&nbsp; 0,16&nbsp; do&nbsp; 0,97&nbsp; mmol&nbsp; L -1 (od&nbsp; 29,1&nbsp; do&nbsp; 174&nbsp; &micro;g&nbsp; mL -1),&nbsp; dok&nbsp; je&nbsp; GO iznosila 0,14 mmol L -1 (25 &micro;g mL<br />-1 ). Optimizovana metoda zasnovana na GOx/Pd-MWCNT-SPCE je&nbsp; uspe&scaron;no&nbsp; primenjena&nbsp; za&nbsp; određivanje&nbsp; glukoze&nbsp; u&nbsp; uzorku&nbsp; livadskog&nbsp; meda.&nbsp; Dobijeni&nbsp; rezultati&nbsp; su&nbsp; u dobroj&nbsp; saglasnosti&nbsp; sa&nbsp; onima&nbsp; dobijenim&nbsp; primenom&nbsp; komercijalno&nbsp; dostupnog&nbsp; aparata&nbsp; za&nbsp; merenje glukoze. Pogodan radni potencijal za GOx/Pt-MWCNT-SPCE je bio -0,50 V u odnosu na ZKE, pri čemu je zadovoljavajuća linearnost postignuta u ispitivanom opsegu koncentracija glukoze od 65,8 do 260,6 &micro;g mL -1 , sa GO 35 &micro;g mL -1 . Optimizovana metoda zasnovana na GOx/Pt-MWCNT-SPCE je&nbsp; uspe&scaron;no&nbsp; primenjena&nbsp; za&nbsp; određivanje&nbsp; glukoze&nbsp; u&nbsp; u&nbsp; uzorku&nbsp; belog&nbsp; grožđa&nbsp; i&nbsp; uzorku&nbsp; tableta (Traubenzucker-bonbons),&nbsp; pri&nbsp; čemu&nbsp; su&nbsp; dobijeni&nbsp; rezultati&nbsp; u&nbsp; dobroj&nbsp; saglasnosti&nbsp; sa&nbsp; rezultatima dobijenim primenom Accu-Chek aparata.<br />Na&nbsp; osnovu&nbsp; dobijenih&nbsp; rezultata&nbsp; može&nbsp; se&nbsp; zaključiti&nbsp; da&nbsp; su&nbsp; razvijene&nbsp; analitičke&nbsp; metode&nbsp; pre svega jednostavne, pouzdane i pogodne za dobijanje brzih informacija o sadržaju H ₂O₂ u različitim tipovima&nbsp; uzoraka.&nbsp; Svakako&nbsp; odabir&nbsp; pogodne&nbsp; radne&nbsp; elektrode,&nbsp; kao&nbsp; i&nbsp; optimizacija&nbsp; eksperimentalnih uslova su ključni faktori za uspe&scaron;no određivanje H ₂O₂</p> / null / <p>The aim of this doctoral dissertation was the development of fast and reliable voltammetric methods&nbsp; based&nbsp; on&nbsp; the&nbsp; application&nbsp; of&nbsp; simple&nbsp; and&nbsp; contemporary&nbsp; electrodes/sensors&nbsp; based&nbsp; on carbonaceous materials (carbon paste made of graphite powder and paraffin oil and screen printed carbon electrodes) for the determination of H <span id="cke_bm_571S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_571E" style="display: none;">&nbsp;</span>O₂ in the selected complex model and real samples.For this purpose, applicability of different working electrodes was investigated.The&nbsp; amperometric&nbsp; method&nbsp; based&nbsp; on&nbsp; carbon&nbsp; paste&nbsp; electrode&nbsp; (CPE)&nbsp; bulk- modified&nbsp; &nbsp;&nbsp; with 5% (m:m) MnO 2 , under optimized conditions, with a working potential of 0.40 V vs. the saturated calomel&nbsp; elect<span id="cke_bm_572E" style="display: none;">&nbsp;</span>rode&nbsp; (SCE)&nbsp; and&nbsp; a&nbsp; phosphate&nbsp; buffer&nbsp; solution&nbsp; (pH&nbsp; 7.50)&nbsp; as&nbsp; supporting&nbsp; electrolyte, enabled the quantitation of H <span id="cke_bm_577S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_577E" style="display: none;">&nbsp;</span>O₂in the concentration interval from 1.4 to 65 &micro;g mL &minus;1 with a relative standard deviation (RSD) of less than 10%. This meth<span id="cke_bm_578E" style="display: none;">&nbsp;</span>od was applied for the determination of the H<span id="cke_bm_583S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_583E" style="display: none;">&nbsp;</span>O₂consumption&nbsp; in&nbsp; samples&nbsp; of&nbsp; groundwater&nbsp; fro<span id="cke_bm_584E" style="display: none;">&nbsp;</span>m&nbsp; the&nbsp; Central&nbsp; Banat&nbsp; region&nbsp; (Province&nbsp; of Vojvodina, Serbia) treated by the Fenton (Fe ²⁺and H ₂O₂ ) and Fenton-&nbsp; like (Fe ³⁺and H ₂O₂ ) reagents to remove natural organic matter (NOM) at&nbsp; differentinitial concentrations of iron species, and of their molar ratios to the initial concentration of H₂O₂ . It was found that the form of Fe (Fe ²⁺ or Fe ³⁺ )and the molar&nbsp; ratio to H ₂O₂influenced the degree of the H₂O₂ decomposition in the groundwater with high NOM content. Besides, in the case of the Fenton-like process, for all initial doses of Fe ³⁺ and H<span id="cke_bm_604S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_604E" style="display: none;">&nbsp;</span>O₂, a sign<span id="cke_bm_605E" style="display: none;">&nbsp;</span>ificant amount of H<span id="cke_bm_610S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_610E" style="display: none;">&nbsp;</span>O₂ remained unused, whi<span id="cke_bm_611E" style="display: none;">&nbsp;</span>ch also indicates a lower efficiency of such system compared to the Fenton process. Screen&nbsp; printed&nbsp; carbon&nbsp; electrode&nbsp; (SPCE)&nbsp; bulk-modified&nbsp; with&nbsp; MnO&nbsp;₂ as&nbsp; a&nbsp; mediator&nbsp; was applied&nbsp; for&nbsp; amperometric&nbsp; determination&nbsp; of&nbsp; the&nbsp; H <span id="cke_bm_617S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_617E" style="display: none;">&nbsp;</span>O₂ content&nbsp; during&nbsp; the&nbsp; Fenton&nbsp; (Fe ²⁺ ,&nbsp; H <span id="cke_bm_624S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_624E" style="display: none;">&nbsp;</span>O₂)&nbsp; and <span id="cke_bm_625E" style="display: none;">&nbsp;</span>visible&nbsp; light-assisted<span id="cke_bm_618E" style="display: none;">&nbsp;</span>,&nbsp; photo-Fenton&nbsp; (Fe ^2+&nbsp; ,&nbsp; H <span id="cke_bm_631S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_631E" style="display: none;">&nbsp;</span>O₂ ,&nbsp; h&nu;)<span id="cke_bm_632E" style="display: none;">&nbsp;</span>&nbsp; based&nbsp; removal&nbsp; of&nbsp; neonicotinoid&nbsp; insecticide acetamiprid (ACT). Under&nbsp; optimized conditions (working potential of 0.40 V vs. SCE, phosphate buffer&nbsp; pH&nbsp; 7.50&nbsp; as&nbsp; supporting&nbsp; electrolyte)&nbsp; amperometric&nbsp; determination&nbsp; of&nbsp; H <span id="cke_bm_637S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_637E" style="display: none;">&nbsp;</span>O₂showed&nbsp; a&nbsp; linear dynamic range from 0.01 to 1.24 mmol L -1 (from 0.34 to 42.2 &micro;g mL -1) and the RSD did not exceed 4.2%. In the investigated samples (after appropriate pH adjustment from 2.8 to 7.5 instantly after the sampling<span id="cke_bm_638E" style="display: none;">&nbsp;</span> to stop or maximum decelerate the oxidation processes, filtering, and storage of the deep- frozen sample with defrosting immediately before the measurements) the H <span id="cke_bm_643S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_643E" style="display: none;">&nbsp;</span>O₂&nbsp; contents were determined by the standard addition method by analyzing the corresponding amperometric curves. Parallel HPLC-DAD measurements were performed to monito<span id="cke_bm_644E" style="display: none;">&nbsp;</span>r the concentration/removal of ACT. In the case of the photo- Fenton process (initial concentrations: 0.31; 2.0 and 3.0 mmol L -1 (70.0; 111.7 and 102.1 &micro;g mL -1 ) of ACT, Fe²⁺ and H <span id="cke_bm_650S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_650E" style="display: none;">&nbsp;</span>O₂, respec<span id="cke_bm_651E" style="display: none;">&nbsp;</span>tively) after 10 min of irradiation H <span id="cke_bm_656S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_656E" style="display: none;">&nbsp;</span>O₂ was&nbsp; consumed&nbsp; and&nbsp; it&nbsp; can&nbsp; be&nbsp; consi<span id="cke_bm_657E" style="display: none;">&nbsp;</span>dered&nbsp; that&nbsp; ACT&nbsp; was&nbsp; removed&nbsp; after&nbsp; 5&nbsp; min.&nbsp; During&nbsp; the&nbsp; Fenton process ACT was removed after 20 min of treatment and around 10% of the initial concentration of the H 2O2 remained still unused.<br />CPE&nbsp; was&nbsp; surface&nbsp; modified&nbsp; with&nbsp; a&nbsp; composite&nbsp; of&nbsp; Pt&nbsp; nanoparticles&nbsp; (&lt;&nbsp; 5&nbsp; nm)&nbsp; on&nbsp; graphitized carbon (Pt-C, 10% Pt on Vulcan XC72) by simply dropcoating method. The unmodified CPE and the&nbsp; modified&nbsp; one&nbsp; (Pt-C/CPE)&nbsp; were&nbsp; characterized&nbsp; by&nbsp;&nbsp; EM/EDS&nbsp; and&nbsp; CV&nbsp; measurements.&nbsp; The&nbsp; PtC/CPE showed remarkable electrocatalytic propertiestoward the electrochemical redox reaction of H<span id="cke_bm_662S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_662E" style="display: none;">&nbsp;</span>O₂ compared to&nbsp; modified CPE in phosphate buffer (0.1 mol L -1 ; pH 7.50), as well in acetatebuffer&nbsp; (0.1 mol&nbsp; L -1 ; pH .50) supporting&nbsp; electrolytes. Amperometry of&nbsp; H2O2 in the concentration range from 0.15 to 1.45 &micro;g mL -1 with the Pt-C/CPE showed acceptable linearity, while the obtained values of LOQs were 0.06 &micro;g mL -1&nbsp; (pH 7.50, working potential 0.20 V) and 0.10 &micro;g mL -1 (pH 4<span id="cke_bm_663E" style="display: none;">&nbsp;</span>.50, working potential 0.50 V). The proposed analytical&nbsp; methods were applied to the determination of the H 2O2 content in commercially available personal care products; i.e., disinfection (pH 7.50) and contact lens cleaning solutions (pH 4.50). The obtained amperometric results are in good agreement with those measured by traditional titanium sulfatereagent based spectrophotometric method with determined concentrations as 2.91% and 2.94%&nbsp; for the disinfection product, and 3.04% and 3.17% for the contact lens solution, respectively. RSD was lower than 2%. The obtained results are in a good agreement with the amounts of the H<span id="cke_bm_668S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_668E" style="display: none;">&nbsp;</span>O₂declared by producers (3%) in the both investigated samples. The Pt-C/CPE was also tested for monitoring of the H<span id="cke_bm_674S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_674E" style="display: none;">&nbsp;</span>O₂ residual concentration in contact lens&nbsp; solution&nbsp; during&nbsp; its&nbsp; neutralization/decomposition&nbsp; rocess.&nbsp; At&nbsp; 6&nbsp; h&nbsp; of<span id="cke_bm_675E" style="display: none;">&nbsp;</span>&nbsp; neutralization&nbsp; treatment 24.68 &micro;g mL -1 of the H <span id="cke_bm_680S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_680E" style="display: none;">&nbsp;</span>O₂was&nbsp; determined which is almost half of the allowedH2<span id="cke_bm_681E" style="display: none;">&nbsp;</span>O2 concentration in the case of the contact lens solution concerningthe limit of eye irritation. CPE&nbsp; was&nbsp; surface&nbsp;&nbsp; modified&nbsp; with&nbsp; multiwalled&nbsp; carbon&nbsp; nanotubes&nbsp; (MWCNT)&nbsp; and&nbsp; with composites of MnO₂-MWCNT or Pt-MWCNT by drop coating method to prepare simply, sensitive and reliable volta<span id="cke_bm_669E" style="display: none;">&nbsp;</span>mmetric sensors for the determination of H <span id="cke_bm_687S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_687E" style="display: none;">&nbsp;</span>O₂in selected sample. The results of the&nbsp; SEM/<span id="cke_bm_688E" style="display: none;">&nbsp;</span>EDS&nbsp; analysis&nbsp; of&nbsp; composite&nbsp; materials&nbsp; have&nbsp; confirmed&nbsp; that&nbsp; the&nbsp; mediators,&nbsp; MnO ₂ and&nbsp; Pt&nbsp; articles, are randomly distributed on the surface of MWCNT and represent nearly 5% (m:m) of the composite expressed as Mn and Pt. CV measurements were performed&nbsp; with prepared electrodes in acetate&nbsp; (pH&nbsp; 4.50),&nbsp; phosphate&nbsp; (pH&nbsp; 7.50)&nbsp; and&nbsp; borate&nbsp; (pH&nbsp; 9.18)&nbsp; buffers&nbsp; to&nbsp; characterize&nbsp; the&nbsp; basic electrochemical&nbsp; behavior&nbsp; of&nbsp; H <span id="cke_bm_694S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_694E" style="display: none;">&nbsp;</span>O₂ and&nbsp; to&nbsp; select&nbsp; the&nbsp; working&nbsp; potentials&nbsp; suitable&nbsp; for&nbsp; amperometric determination&nbsp; of&nbsp; this&nbsp; target&nbsp; analyte.&nbsp; The&nbsp; Pt-MWCNT/CPE&nbsp; performs&nbsp; well&nbsp; in&nbsp; phosphate&nbsp; buffer pH .50 and acetate buffer solution pH 4.50 in the&nbsp; negative as well as in the positive polarization range with RSD mainly lower than 2.5%. In case of MnO₂-MWCNT/CPE at <span id="cke_bm_695E" style="display: none;">&nbsp;</span>0.30 V and above the H<span id="cke_bm_701S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_701E" style="display: none;">&nbsp;</span>O₂oxidation signal is rem<span id="cke_bm_702E" style="display: none;">&nbsp;</span>arkable in slightly alkaline media (pH 7.50), at pH 4.50 this electrode showed poor behavior and at pH 9.18 offered acceptable performance. LOQs were in the &micro;g mL -1 concentration&nbsp; range.&nbsp; H<span id="cke_bm_707S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_707E" style="display: none;">&nbsp;</span>O₂was&nbsp; determined&nbsp; in&nbsp; a&nbsp; spiked&nbsp; milk&nbsp; sample&nbsp; by&nbsp; standard addition&nbsp; method after&nbsp; appropriate&nbsp; sample&nbsp; preparation&nbsp; (pH&nbsp; adjustment and&nbsp; centrifugation)&nbsp; and&nbsp; using&nbsp; optimized amperometric p<span id="cke_bm_708E" style="display: none;">&nbsp;</span>rocedure (acetate buffer pH 4.50, working potential -0.75 V) by Pt-MWCNT/CPE as a working electrode. RSD for three repeated measurements was 2.5%, while the recovery of the method was a bit higher than 71%. The&nbsp; composite&nbsp; materials&nbsp; consisting&nbsp; of&nbsp; MWCNT&nbsp; and&nbsp; Pd&nbsp; (Pd-MWCNT)&nbsp; or&nbsp; Pt&nbsp; containing particles&nbsp; (Pt-WCNT)&nbsp; were&nbsp; applied&nbsp; to&nbsp; the&nbsp; preparation&nbsp; of&nbsp; bulk- modified&nbsp; SPCEs&nbsp; (Pd-MWCNTSPCE and Pt-MWCNT-SPCE) and surface modifiedSPCE (Pd- MWCNT/SPCE). These electrodes, as well as unmodified SPCE and MWCNT-SPCE,&nbsp; were characterized by CV and&nbsp; amperometry&nbsp; in phosphate&nbsp; buffer&nbsp; solution&nbsp; of&nbsp; pH&nbsp; 7.50&nbsp; for&nbsp; the&nbsp; H<span id="cke_bm_713S" style="display: none;">&nbsp;</span>₂<span id="cke_bm_713E" style="display: none;">&nbsp;</span>O₂determination.&nbsp; Pd-MWCNT-SPCE&nbsp; and&nbsp; PdMWCNT/SPCEare convenient for the etermination of H 2O2 at working potentials from -0.50 to 0.50 V, and Pt-MWCNT-SPCE at investigated working potentials in the range from -0.80 to 0.70 V. These electrodes were then modified with glucose&nbsp; oxidase (GOx) by drop coating a solution of GOxand Nafion &reg; on their surface, whereby the applied amount of biococktail was optimized. GOx/PdMWCNT-SPCE&nbsp; showed&nbsp; better&nbsp; analytical&nbsp; performance&nbsp; for&nbsp; glucose&nbsp; determination&nbsp; in&nbsp; comparison with&nbsp; GOx/Pd-MWCNT/SPCE.&nbsp; The&nbsp; optimal&nbsp; working&nbsp; potential&nbsp; for&nbsp; GOx/Pd-MWCNT- SPCE&nbsp; was -0.40 V vs. SCE and <span id="cke_bm_714E" style="display: none;">&nbsp;</span>satisfactory linearity was obtained in the investigated glucose concentration range from 0.16 to 0.97 mmol L -1 (from 29.1 to 174&nbsp; &micro;g mL -1 ), hile the LOQ was 0.14 mmol L -1 (25 &micro;g mL -1 ). The optimized method based on GOx/Pd-MWCNT-SPCE was successfully applied to the determination of glucose in multifloral honey sample.&nbsp; The results are in a good agreement with those&nbsp; obtained by commercially available equipment for determination of glucose. Optimal working potential&nbsp; for&nbsp; GOx/Pt-MWCNT-SPCE&nbsp; was&nbsp; -0.50&nbsp; V&nbsp; vs.&nbsp; SCE,&nbsp; and&nbsp; the&nbsp;&nbsp; satisfactory&nbsp; linearity&nbsp; was obtained in the investigated concentration range ofglucose from 65.8 to 260.6&nbsp; &micro;g mL -1 , with LOQ of 35&nbsp; &micro;g mL -1 . The optimized method based on GOx/Pt- MWCNT-SPCE was successfully applied for determination of glucose in white grape and glucose tablets (Traubenzucker-bonbons) samples, whereby&nbsp; the&nbsp; obtained&nbsp; results&nbsp; were&nbsp; in&nbsp; a&nbsp; good&nbsp; agreement&nbsp; with&nbsp; the&nbsp; results&nbsp; obtained&nbsp; by&nbsp; Accu-Chek device. Based on the results, the developed analytical methods are first of all simple, reliable and suitable&nbsp; for&nbsp; obtaining&nbsp; fast&nbsp; information&nbsp; about&nbsp; the&nbsp; content&nbsp; of&nbsp; H ₂O₂ in&nbsp; different&nbsp; types&nbsp; of&nbsp; samples. Certainly the selection of a suitable working electrode, as well as the optimization of experimental conditions are key factors for the successful determination of H₂O₂.</p>