Simvastatin is a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor that is used for the treatment of hyperlipidemia. Simvastatin has recently been studied for its potential use in cancer therapy. In-vitro studies have shown that simvastatin displays anticancer activity, but at concentrations unlikely to be achieved in patients being receiving typical antihyperlipidemic treatment doses. Thus, several clinical trials were conducted to study the tolerability of high dose statins in cancer patients. The maximum tolerated dose of simvastatin was determined to be 15 mg/kg/day, 25-fold higher than a typical dose. However, it is not known if simvastatin plasma concentrations can reach those found to be effective in-vitro. In this context, we initiated a clinical study to determine the pharmacokinetics of high dose simvastatin in patients with chronic lymphocytic leukemia. For this purpose, an LC-MS/MS method was developed and validated for the quantitation of simvastatin and its acid form in plasma and peripheral blood mononuclear cells obtained from CLL patients. Results show that simvastatin concentrations were dose proportional relative to the antihyperlipidemic doses, but lower than those required for in-vitro cytotoxicity against cancer cells. These findings demonstrate that the in-vitro effective concentrations of simvastatin are not achievable clinically, which might explain the limited effectiveness of high dose simvastatin in this study and in previous clinical trials. In view of these data, the use of simvastatin as a sole therapy in cancer treatment was not encouraging and led us to examine the use in combination with other anticancer drugs. After screening several chemotherapeutic agents in combination with simvastatin, we showed that tipifarnib (a farnesyltransferase inhibitor) interacts synergistically in several leukemia cell lines. Mechanistically we showed that simvastatin augments the cytotoxicity of tipifarnib by disrupting the localization of RAS in the cell membrane and by subsequent deactivation of the ERK pathway. Consistent with this observation, drug treatment led to the induction of apoptosis through the caspase cascade activation and the cleaved PARP upregulation. Notably, this synergistic effect was observed at clinically achievable concentrations of simvastatin and tipifarnib. Thus, the effectiveness of this combination should be explored further in future clinical studies.

N-linked glycosylation is the most common post-translational modification (PTM) of proteins that exist in nature. N-glycosylation and change in cells serve as a criterion to monitor the activity of developmental stages and diseases severity. Currently, there is an increasing application of mass spectrometry on glycoprotein for malicious, chronic or acute diseases, such as cancers, rheumatoid arthritis (RA) or influenza. In this dissertation, several mass spectrometric assays have been utilized to, quantitatively and qualitatively, characterize protein N-glycosylation at the glycan, glycopeptide and peptide levels. The goals are to identify serum-based RA biomarker (Chapter 2), or to determine possible glycan structures from monoclonal antibody (Chapter 3), or comprehensively to study one influenza glycoprotein, hemagglutinin (Chapter 4). In Chapter 2, LC-MS/MS with CID as MS 2 is the primary technique that is applied to collect raw data for RA biomarker screening; western blot is the verification method for newfound biomarkers. This mass spectrometry based comparative analysis of N-glycoprotein in RA and healthy patients’ sera reveal 41 potential biomarkers for RA that can be applied in clinical research. Chapter 3 describes another LC-MS/MS based method developed for the structural analysis of N-glycan released from the monoclonal antibody, immunoglobin G. Higher-energy collision dissociation (HCD) was the surprior technique utilized to identify glycopeptide fragments. The results show that 19 and 23 N-glycan structures were determined from standard and modified mAb samples respectively by using SimGlycan software, while 38 and 35 glycan structures were recognized by manually mapping respectively. 13 N-glycoforms, out of 26 overlapped glycan structures, were identified with significant alterations by comparing standard sample (sample A) and modified mAb (sample B) utilizing our method. In Chapter 4, we comprehensively studied hemagglutinin by using LC-MS/MS and MALDI from both proteomic perspective and glycomics prospective. After confirmed and verified protein sequence and glycosylation sites, galactose-specific quantitation was performed with exoglycosidase digestion combined HPLC with fluorescence detection. The MALDI-MS/MS based method was utilized to confirm glycan structures. The results in this dissertation provide insights into the significance of protein glycosylation alterations as RA biomarkers, and these quantitative methods can be reapplied to any other disease biomarkers screening for clinical researchers.

Immunoassays have traditionally been employed for the determination of plasma concentration-time profiles for pharmacokinetic studies of therapeutic proteins and peptides. These ligand binding assays have high sensitivity but require significant time for antibody generation (1 to 2 years) for assay development. Despite high sensitivity, these assays suffer from cross-reactivity that can lead to inaccurate results. As an alternative to immunoassays, this dissertation was focused on the development and validation of assays that can be used for quantitative analysis of peptides or proteins in plasma using liquid chromatography tandem mass spectrometry (LC-MS/MS). Two approaches were considered for measurement of proteins and peptides fortified in plasma. The first approach involved employing signature peptides as quantitative surrogates of a target protein. This approach is a multistep process that includes: computer simulated (in silico) peptide predictions, protein purification, proteolytic digestion, peptide purification, and ultimately mass spectrometry. Signature peptides were determined through in silico peptide predictions and iterative tuning processes to represent Amevive® (Alefacept), a therapeutic for psoriasis, for quantification in human plasma. Horse heart myoglobin was chosen as a protein analogue internal standard to compensate for errors associated with matrix effects and to track recovery throughout the entire sample pretreatment process. Samples were prepared for analysis by selective precipitation of the target proteins with optimized pH and heat conditions followed by enzymatic digestion, dilution, and filtration. Combining selective precipitation and protein analogue internal standard lead to a method validated according to current FDA guidelines and achieved a linear range (250-10,000 ng/mL) suitable for monitoring the therapeutic levels of Alefacept (500 -6000 ng/mL) without the use of antibodies. A second approach exploited the mass spectrometric behavior of intact polypeptides. A polypeptide can exist in multiple charge states separated by mass to charge ratio (m/z). Herein, the charge state distribution and the formation of product ions to form selected reaction monitoring (SRM) transitions for intact polypeptide quantitative analysis was evaluated in plasma. Oxyntomodulin, a 37 amino acid anorectic peptide (4449 Da), was employed as a model for analysis in rat plasma. The +7 charge state form of OXM was used to form an SRM for quantitative analysis. Two-dimensional reversed phase ion pair chromatography, a modified solid phase extraction, and a multiply charged SRM of oxyntomodulin enabled a lower limit of quantification of 1 ng/mL. Following development of the LC-MS/MS method, a validation of this approach was performed according to FDA guidelines. Finally, to show further utility of LC-MS/MS, the validated oxyntomodulin method was used in a pharmacokinetic study with sprague-dawley rats. Rats were dosed with oxyntomodulin through intravenous or intratracheal instillation routes of administration. Plasma concentration-time profiles were determined. Using these profiles, noncompartmental parameters were determined for each dose and routes of administration.

Antipsychotics (AP) are used to treat psychotic symptoms in patients with e.g. schizophrenia or bipolar disorder. To find the right treatment may be difficult and demands carefully monitoring of the concentration in blood tests. In psychopharmacology therapeutic drug monitoring (TDM) of antipsychotics is an important tool for optimizing the treatment. With repeated blood tests it is possible to monitor the patient response, interactions and compliance. The aim of this project was to develop a method for sample preparation and analysis by UHPLC-MS/MS suitable for 12 antipsychotic drugs and 9 major metabolites in human plasma samples. The project was divided in to six different steps (1) Preparation of stock solutions and infusions of the analytes and internal standards. (2) Calculation of the monoisotopic molecular weight. (3) Optimization of the cone voltage and collision energy (4) Optimization of the chromatography by selecting the ideal mobile phase and column (5) Preparation of the calibration curve (5) Extraction with protein precipitation (6) Analysis on an Acquity I Class coupled to an Xevo TQ-XS MS and separation on a Cortects UPLC C18 (1.6 µm 2.1x50 mm) with gradient elution at 0.5 ml/min and a 5.20-min run-time. Detection was made with MRM, monitoring 2 ion transitions per compound. Isotope labeled internal standards were used for 9 of the compounds. In order for the method to be used in routine analysis, some work remains to be done regarding validating, optimizing the mass spectrometer's settings, testing for extraction yield, matrix effects and analysis of patient samples. / Antipsykotika är läkemedel som används för behandling av psykotiska symtom hos patienter som lider av t.ex. schizofreni eller bipolär sjukdom. Att hitta rätt behandling kan vara svårt och kräver att man noga övervakar koncentrationen i blodet. Med upprepade blodprov och så kallat terapeutiskt intervall (TDM) går det att optimera behandlingen, övervaka patientens respons på behandlingen samt eventuella interaktioner med andra läkemedel. Det går även att följa hur noga patienten är med att ta sina mediciner så kallat följsamhet eller compliance. Syftet med detta projekt har varit att utveckla en ny metod för att mäta 12 antipsykotiska läkemedel och 9 metaboliter i plasma med UHPLC-MS/MS. Det experimentella arbetet i detta projekt delades in i sex olika steg: (1) Beredning av stamlösningar och infusionslösningar av eftersökta substanser och deras internstandarder till rätt koncentration (2) Beräkning av molekylmassan för monoisotopen av varje substans. (3) Optimering av konvolt och kollisionsenergi (4) Optimering av kromatografin med val av lämplig mobilfas och kolonn (5) Beredning av kalibreringslösningar (6) Extraktion av substanser i plasma. Extraktionsmetoden som användes var proteinutfällning och analysen utfördes på Acquity I Class kopplat till Xevo TQ-XS MS. Separation gjordes med Cortects UPLC C18 (1,6 µm 2,1x50 mm) och gradienteluering med ett flöde på 0,5 ml/min och tiden för analys 5,20 min. Detektionen gjordes med MRM och två transitioner per substans. Isotop-inmärkt internstandard användes för 9 av substanserna. För att metoden ska kunna användas vid rutinanalys återstår en del arbete när det kommer till validering, optimering av masspektrometerns inställningar, test av extraktionsutbyte, matriseffekter och analys av patientprov.

A bioluminescência desperta o interesse humano há muitos séculos. Presente em quatro dos sete reinos taxonômicos, Monera, Chromista, Animalia e Fungi, cada um com mecanismos muito diferentes. Pode-se dizer que o estudo químico da bioluminescência começou com os experimentos de Dubois no século XIX, que cunhou os termos luciferina e luciferase, termos genéricos para o substrato e enzima envolvidos na reação, respectivamente. No caso específico de fungos, o envolvimento de enzimas foi debatido por quase cinco décadas, após a proposta enzimática de Airth e Foerster na década de 1960 e a não enzimática por Shimomura, em 1989. Somente em 2009 a hipótese de Airth e Foerster foi confirmada pelo nosso grupo, seguido da identificação da luciferina fúngica e o envolvimento de hispidina, como molécula precursora, em 2015 pelo grupo de Yampolsky. Para conseguir elucidar mecanismos químicos, a técnica de espectrometria de massas pode ser empregada para a identificação estrutural de reagentes e intermediários destas e de outras reações orgânicas. Após a confirmação do envolvimento de hispidina na bioluminescência de fungos, utilizou-se a técnica de cromatografia líquida acoplada a espectrometria de massas para identificar a presença de hispidina, seus derivados e intermediários precursores, em fungos e em algumas plantas. / Bioluminescence arises human interest for centuries. Occurring in four of seven taxonomical Kingdoms, Monera, Chromista, Animalia and Fungi, each of them with completely different mechanisms. The chemical study of bioluminescence starts in XIX century with Dubois, who coined the terms luciferin and luciferase, generic terms for substrate and enzyme involved in the bioluminescent reaction, respectively. In the specific case of fungi, enzyme involvement has been debated for almost five decades, after the enzymatic proposal by Airth and Foerster, during the 1960 decade, and the non-enzymatic proposal by Shimomura in 1989. It was only in 2009 when the proposal by Airth and Foerster was confirmed by our group, followed by the identification of the fungal luciferin and the involvement of hispidin, as the precursor molecule, in 2015 by Yampolskys group. To elucidate the chemical mechanisms, mass spectrometry can be employed to structural identification of reagents and intermediates on these and other organic reactions. After the confirmation of hispidin involvement in fungi bioluminescence, liquid chromatography coupled with mass spectrometry was uses do identify the presence of hispidin, its derivatives and precursor intermediates in fungi and selected plants.

A ifosfamida é um pró-fármaco que apresenta um átomo de fósforo quiral, disponível na clínica como mistura racêmica dos enantiômeros(+)-(R)-ifosfamida e (-)-(S)-ifosfamida para a utilização na quimioterapia. O objetivo do presente estudo foi o de avaliar a disposição cinética dos enantiômeros da ifosfamida em plasma de pacientes portadoras de câncer de colo do útero. As pacientes investigadas (n=6) receberam 2,5 g/m2 de ifosfamida racêmica administrada como infusão de 12 horas, sendo coletadas amostras de sangue imediatamente antes da administração e em 6, 10, 11, 12, 13, 14, 16, 18, 20 e 22 horas após a administração do fármaco. Os enantiômeros da ifosfamida foram quantificados por LC-MS/MS, sendo separados na coluna OD-R em aproximadamente 14 min empregando como fase móvel mistura de acetonitrila e água (20:80) adicionada de 0,2% de ácido fórmico. O método é linear no intervalo de 1-100 ?g de cada enantiômero/mL de plasma a partir de extrações de alíquotas de 25 ?L de plasma, compatíveis com a aplicação em farmacocinética de infusão de curta duração da ifosfamida em pacientes com câncer de colo do útero.A disposição cinética da ifosfamidaéenantiosseletiva, com observação de maiores valores de AUC (437,31 vs349,18 h.?g/mL) e menores valores de clearance(4,17 vs5,22 L/h) para o enantiômero(+)-(R)-ifosfamida. / The prodrugifosfamide has a chiral phosphorus atom, and is available clinically as a racemic mixture of the enantiomers (+)-(R)-ifosfamide and (-)-(S)-ifosfamide for use in chemotherapy. The aim of this study was to evaluate the kinetic disposition of the enantiomers of ifosfamide in plasma of patients with cancer of the cervix. The investigated patients (n = 6) received 2.5 g/m2 of racemic ifosfamide administered as infusion of 12 hours and blood samples were collected immediately before administration and at 6, 10, 11, 12, 13, 14, 16, 18, 20 and 22 hours after drug administration. The enantiomers of ifosfamide were quantified by LC-MS/MS and were separated in an OD-R column in about 14 min using as mobile phase a mixture of acetonitrile and water (20:80) plus 0.2% of formic acid. The method is linear within the range of 1-100 mg of each enantiomer/mL of plasma from extractions of 25 mL aliquots of plasma, suitable for the application in pharmacokinetics of short duration infusion of ifosfamide in patients with cervical cancer. The kineticdisposition of ifosfamide is enantioselective, with observation of higher values of AUC (437.31 vs 349.18 h.?g/mL) and lower values of clearance (4.17 vs 5.22 L/h) for the enantiomer (+)-(R)-ifosfamide.

Os agrotóxicos, quando presentes na água, são definidos como micropoluentes: mesmo em baixas concentrações, conferem à água características de toxicidade. Aponta-se o RS como o quarto estado do Brasil com maior volume de vendas anuais de agrotóxicos, chegando a mais de 50 mil toneladas por ano. Desde 2014 está em vigência no território gaúcho uma portaria estadual que acrescenta a exigência de 46 parâmetros de agrotóxicos no padrão de potabilidade da água, além dos 27 já exigidos pela portaria nacional. Neste trabalho, 89 pesticidas foram avaliados conforme três métodos teóricos de predição de risco de contaminação em mananciais subterrâneos e superficiais: índice Ground Ubiquity Score (GUS), método Screening da USEPA e método de GOSS, baseados nas propriedades físico-químicas dos pesticidas. Nos anos de 2015 e 2016, foram realizadas 143 coletas de água para consumo humano em 45 municípios da bacia hidrográfica do Alto Jacuí (G-50), a que possui a maior taxa de aplicação de agrotóxicos do estado, para análises de vigilância através de laboratório contratado, envolvendo os 89 pesticidas presentes na portaria nacional e estadual. Em paralelo, 183 pesticidas presentes em uma solução-padrão foram empregados no desenvolvimento de um novo método de análise multiresíduos, com etapas de pré-tratamento por filtração seguidas por extração em fase sólida e LC-MS/MS, aplicada para os três maiores municípios da G-50 (Carazinho, Soledade e Cruz Alta) em amostras de água bruta e tratada, durante quatro períodos de aplicação de agrotóxicos dos principais cultivos agrícolas da região. Dos pesticidas mencionados nas portarias nacional e estadual, 12 foram classificados com o maior risco de contaminação tanto em água superficial e subterrânea de acordo com os três métodos teóricos empregadas. Nas análises de vigilância foi detectado permetrina em Carazinho e alaclor em Espumoso. No método desenvolvido, 75 pesticidas foram validados de acordo com os critérios propostos e atingiram limites de detecção (LD) e limites de quantificação (LQ) que variaram de 10 a 300 ng L-1. Na aplicação do método nas coletas dos três municípios da G-50 não houve detecção de nenhum pesticida. / Agrochemicals, when present in water, are defined as micropollutants, thus giving the water toxic characteristics, even at low concentrations. The Rio Grande do Sul state in Brazil was found to rank fourth in annual agrochemical sales in the country, surpassing 50 thousand tons per year. A state regulation in effect in the RS state since 2014 requires the inclusion of 46 new agrochemical parameters concerning the standards for potable drinking water, in addition to 27 existing parameters mandated by national ordinance. Seventy-five pesticides were evaluated based on three theoretical methodologies of contamination risk prediction in underground and surface water sources, by measuring the physicochemical properties of pesticides: GUS index, USEPA screening method and Goss method. In 2015 and 2016, 143 water samples were collected from sources of potable water in 45 municipalities located in the Alto Jacuí river basin, a region which has the highest pesticide application rate in the RS state. A private laboratory analyzed samples from 89 pesticides present in the national and state regulation. Paralely, 183 pesticides were evaluated by a new multi-residue analysis method. Filtration was conducted in the pre-treatment steps, followed by a solid phase extraction liquid chromatography coupled with mass spectrometry analysis (SPE-LC-MS/MS) of raw and treated water samples from the three largest G-50 municipalities (Carazinho, Soledade and Cruz Alta), during the four pesticide application periods of the main crops cultivated in the region. Twelve pesticides were classified as of high risk in terms of contamination for both surface and groundwater, in accordance with the three theoretical methodologies implemented. During analysis of the surveillance data collected, the pesticides permethrin and alachlor were found in Carazinho and Espumoso, respectively. Through the methodology developed, 75 pesticides were evaluated according to the criteria proposed, reaching lower detection limit (LD) and quantification limit (LQ) ranging from 10 to 300 ng L-1, respectively. During the implementation of the methodology for sample collections in the three G-50 municipalities, no pesticides were detected.

Pluripotent embryonic stem cells (ESCs) possess an unlimited capacity for self-renewal. This property of ES cells is both defining and unique. Harnessing this potential of ESCs would provide tremendous opportunity in the field of regenerative medicine and its attempts to combat degenerative diseases such as Parkinson’s, muscular dystrophy, etc. In 2006, Shinya Yamanaka was able to demonstrate that the ectopic expression of four proteins could reverse the process of differentiation and provide somatic cells with the characteristics ESCs. One year later, James Thompson’s group proved the same feat could be accomplished in human somatic cells using a different set of four proteins, including Nanog. The prospect of converting one’s own cells into a stem cell which could subsequently differentiate and repopulate an area of the body afflicted by gross degeneration was revolutionary. In the years following Yamanaka’s and Thompson’s discoveries, however, there has been little insight gained into how these proteins are regulated post-translationally. In this study, four proteins which had previously been identified by Yamanaka as being ‘pluripotency factors’ were used as baits in order to ascertain a protein-protein interaction network. This network was subsequently interrogated using various chemical compounds and small molecules in order to dissect the signal transduction pathways feeding into pluripotency, as well as, post-translational modifications regulating the factors themselves. In this way, the chemical inhibitor H89 was found to decrease the presence of Nanog phosphorylation and possibly its dimerization resulting in the Nanog protein being destabilized and targeted for degradation. Inversely, the pan-cullin inhibitor MLN4924 was identified to increase the abundance of both phosphorylated Nanog and total Nanog protein. In an attempt to identify the Cullin Ring Ligase (CRL) responsible for the degradation of Nanog protein in ESCs, each cullin identified in the protein interaction network was inhibited using specific shRNAs. Quantitative fluorescence microscopy was performed and identified that inhibition of CUL3 increases Nanog protein levels, suggesting that a CUL3-based CRL may be responsible for the post-translation regulation of Nanog. Additionally, the quantitation of Sox2 protein levels in CUL4B shRNA cell line demonstrates that Sox2 protein levels may be regulated by a CUL4B-based CRL. Further studies will reveal whether or not CUL4A depletion also results in elevated Sox2 protein levels. If not, this would include the pluripotency factor Sox2 among the recently identified CUL4B-isoform-specific substrates for degradation and possibly provide the basis for a hypothesis of developmentally regulated substrate specificity. In addition to MLN4924, several other small molecules were identified as being able to increase phospho-Nanog protein levels in this study. Among them were the cell permeable peptides Ht-31 and PKI (14-22) amide. These peptides were found to both stabilize phospho-Nanog and produce ES cell colonies that uniformly express the Nanog protein. The development of a growth medium containing these peptides in order to maintain homogeneous pluripotent ES cells is currently in progress and received backing for a patent application by the University of Edinburgh on February 23, 2012.

A bioluminescência desperta o interesse humano há muitos séculos. Presente em quatro dos sete reinos taxonômicos, Monera, Chromista, Animalia e Fungi, cada um com mecanismos muito diferentes. Pode-se dizer que o estudo químico da bioluminescência começou com os experimentos de Dubois no século XIX, que cunhou os termos luciferina e luciferase, termos genéricos para o substrato e enzima envolvidos na reação, respectivamente. No caso específico de fungos, o envolvimento de enzimas foi debatido por quase cinco décadas, após a proposta enzimática de Airth e Foerster na década de 1960 e a não enzimática por Shimomura, em 1989. Somente em 2009 a hipótese de Airth e Foerster foi confirmada pelo nosso grupo, seguido da identificação da luciferina fúngica e o envolvimento de hispidina, como molécula precursora, em 2015 pelo grupo de Yampolsky. Para conseguir elucidar mecanismos químicos, a técnica de espectrometria de massas pode ser empregada para a identificação estrutural de reagentes e intermediários destas e de outras reações orgânicas. Após a confirmação do envolvimento de hispidina na bioluminescência de fungos, utilizou-se a técnica de cromatografia líquida acoplada a espectrometria de massas para identificar a presença de hispidina, seus derivados e intermediários precursores, em fungos e em algumas plantas. / Bioluminescence arises human interest for centuries. Occurring in four of seven taxonomical Kingdoms, Monera, Chromista, Animalia and Fungi, each of them with completely different mechanisms. The chemical study of bioluminescence starts in XIX century with Dubois, who coined the terms luciferin and luciferase, generic terms for substrate and enzyme involved in the bioluminescent reaction, respectively. In the specific case of fungi, enzyme involvement has been debated for almost five decades, after the enzymatic proposal by Airth and Foerster, during the 1960 decade, and the non-enzymatic proposal by Shimomura in 1989. It was only in 2009 when the proposal by Airth and Foerster was confirmed by our group, followed by the identification of the fungal luciferin and the involvement of hispidin, as the precursor molecule, in 2015 by Yampolskys group. To elucidate the chemical mechanisms, mass spectrometry can be employed to structural identification of reagents and intermediates on these and other organic reactions. After the confirmation of hispidin involvement in fungi bioluminescence, liquid chromatography coupled with mass spectrometry was uses do identify the presence of hispidin, its derivatives and precursor intermediates in fungi and selected plants.

Dietary intake is one of the major routes of human exposure to perfluoroalkyl and/or polyfluoroalkyl substances (PFAS). The objective of this study was to measure perfluorooctane sulfonic acid (PFOS), perfluorooctanoic acid (PFOA) and perfluorohexane sulfonic acid (PFHxS) in organic and conventional egg from Sweden (n=8, consisted of 4 pooled eggs and 4 individuals) and China (n=9, consisted of 4 pooled eggs and 5 individuals) and compare the concentrations of PFAS between the two categories (organic and conventional). Also, to evaluate if there was any difference in concentrations of PFAS between both countries. In the end, evaluation of tolerable weekly intake of PFOS and PFOA due to consumption of egg recommended by the European Food Safety Authority (EFSA) was conducted if consuming these eggs would cause any human health risk. Liquid chromatography-mass spectrometry (LC-MS/MS) was used to analyze PFOS, PFOA and PFHxS in the egg samples. In egg samples from China, PFOA was the predominant PFAS; in an organic egg sample from Shenzhen with concentration up to 2000 pg/g, making up to 86% of the 3 PFAS. In contrast, PFOS had the greatest concentration of all PFAS in egg samples from Sweden and was detected in organic egg sample with concentration up to 184 pg/g, making up to 78% of the 3 PFAS. PFOA in samples from China was 18 times higher compared to egg samples to Sweden; results showed no significant differences in PFAS concentrations in egg samples between Sweden and China. In samples from China, concentrations of PFAS had total mean of 50 pg/g for PFOS, 373 pg/g for PFOA and 13 pg/g for PFHxS. In Sweden, mean concentrations of PFOS, PFOA, and PFHxS were found to be 5, 2, and 1,5 times (respectively) higher in organic eggs when compared to conventional. However, significant difference was only observed for PFOS in Swedish organic eggs (p<0.05, t-7.96, df=6). The different concentrations of contamination between organic and conventional egg could be due to the fish powder in organic chicken feed and ingestion of soil through pecking. The result suggests that current concentrations of PFOS and PFOA in organic and conventional chicken eggs are unlikely to cause any immediate harm to Swedish populations. For Chinese population since the consumption of egg has a high risk of exceeding the TWI, the current concentration of PFOA in organic chicken eggs may cause harm to the population based on TWIs established by EFSA. Further investigation is needed with more samples to be analyzed to confirm this point.