Human scent has been the focal point of diverse scientific interests and research initiatives for the past several years. The knowledge gained about its composition has favored the advancement of multiple disciplines, and promoted the development of a wide variety of applications. Among these applications is the use of human scent as a resource for Forensic investigations, where scent profiles are often used as evidence to associate individuals to the scene of a crime. The characteristic nature of individual human scent has enabled this type of evidence to be used as a biometric tool for the differentiation of subjects. Nevertheless, the present study discusses a new perspective towards human scent's role and application in Forensic investigations. The foundation of this new perspective consists of employing human scent’s biometric quality to classify individuals using common traits. In this research study, underarm and hand odor samples were collected from Caucasian, Hispanic and East Asian individuals, of both genders. Subjects were also organized into 3 different age groups: 18-30, 35-50 and 55+ years. Headspace Solid Phase Micro-extraction Gas Chromatography Mass Spectrometry (HS-SPME-GC-MS) was used to create individual scent profiles for the evaluation of subject classification by age, gender and race/ethnicity. Individual classification was assessed through the identification of qualitative and quantitative patterns in the volatile organic compound (VOC) constituents that characterize human scent. Principal Component and Linear Discriminant analyses of the collected scent profiles, led to the identification and validation of characteristic VOC marker combinations for age, gender and race/ethnicity. Statistical analysis facilitated group classification and differentiation on the basis of these traits. Moreover, this study also evaluated the use of solvent extraction as a complementary technique to HS-SPME for human scent analysis. Findings from this assessment revealed that the simultaneous consideration of data from both extraction techniques favors an enhancement of the classification of subjects by means of human scent. The discoveries achieved in this study represent a significant step for human scent as a forensic tool. The outcome of this research has cleared a new path for further human scent investigation, and highlighted its further relevance to forensic applications.

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<p dir="ltr">Mass spectrometry (MS) is a powerful and versatile technique that is useful for addressing a wide range of complex analytical challenges. In this work, mass spectrometry-based assays were developed to address issues relating to environmental contamination and for detecting analytes of interest to the defense industry. Chapter one is an overview of the history of mass spectrometry, the fundamental operation of a mass spectrometer, as well as, advancements in chromatographic separation and ionization methods. Chapter two focuses on the development of an assay that uses blow flies as environmental sensors of chemical weapon release. In that work, a liquid chromatography – tandem mass spectrometry (LC-MS/MS) method was developed to detect chemical warfare agent simulants and chemical warfare agent hydrolysis products in flies exposed to the chemicals in controlled feeding experiments. The work in chapter three describes the development of a surface enhanced Raman spectroscopy assay coupled to paper spray mass spectrometry for a more fieldable and environmentally friendly approach to detect organophosphorus compounds. Chapter four describes the development of a paper spray mass spectrometry assay for the detection and semi-quantitation of per- and polyfluoroalkyl substances in whole blood without sample cleanup or chromatographic separations. This method would be useful in detecting high levels of these carcinogenic compounds in individuals highly exposed via their occupations. The final chapter (chapter five) returns to using blow flies as environmental sensors, but this time to detect insensitive munitions in the environment. The work focuses on the development of two different liquid chromatography mass spectrometry methods for the detection of insensitive munitions, which are less shock sensitive explosives, and their transformation products in the environment. Controlled feeding experiments were also performed where flies were exposed to contaminated soil and water sources to show the feasibility of this method in a more realistic scenario. The projects detailed herein show the extensive range with which mass spectrometry can be used for the detection of harmful chemistries of environmental concern.</p>

Com o intuito de avaliar os polimorfismos de mtDNA e a diversidade genética na população brasileira, foram estudados 84 indivíduos não aparentados: 49 caucasóides e 35 negróides da região sudeste do Brasil. As amostras de sangue obtidas por punção da polpa digital foram colocadas em papel de filtro e o mtDNA extraído pelo método orgânico. Foram amplificadas as regiões hipervariáveis HV1 a partir do nucleotídeo 16.051 até 16.365 e HV2 entre os nucleotídeos 71 e 340. O fragmento foi purificado e após a reação de seqüenciamento, as fitas sense e anti-sense foram alinhadas com a seqüência de Anderson e sobrepostas para confirmação. Foram encontradas 108 posições contendo transições, 15 transversões, 11 deleções e 4 inserções. Nove regiões apresentaram duas variantes de seqüência polimórfica. A maioria das seqüências foi observada uma única vez e em 84 indivíduos foram observados 80 haplótipos distintos. O índice de diversidade genética e a probabilidade de semelhança entre duas seqüências escolhidas ao acaso foram respectivamente de 0,9714 e 0,0286 para os negróides e de 0,9788 e 0,0212 para os caucasóides. Ao realizar análise pareada dos haplótipos, a média de nucleotídeos diferentes entre as seqüências foi de 14,36 &#177; 5,68 bases em um total de 3.486 pares comparados, indicando uma alta diversidade entre haplótipos nesta população. Os resultados demonstraram que o mtDNA é informativo para identificação humana e o conhecimento da freqüência dos haplótipos na população de interesse tem utilidade significativa na aplicação Forense, principalmente para se avaliar estatisticamente os resultados, fornecendo maior precisão, exatidão e dados contundentes nas investigações. / In order to evaluate the mtDNA polymorphism and demonstrate the sequence diversity in Brazilians, we established a database of 84 unrelated individuals, 49 Caucasians and 35 negroids from the southeast of Brazil. The samples were collected on a whatman paper by finger punction, the mtDNA was extracted by organic method and hipervariable regions, HV1 from 16.051 to 16.365 bp and HV2 from 71 to 340 bp were amplified. Both sense and anti-sense strands were sequenced. When compared to Anderson sequence, 108 transitions, 15 transversions, 11 deletions and 4 insertions were found. Nine positions exhibited two variants of polymorphic sequence. The majority of the sequences were observed once, and in 84 sequences, 80 haplotypes were observed. The sequence diversity and the probability for two individuals randomly matching over two hypervariable regions were 0.9714 and 0.0286 for persons of color and 0.9788 and 0.0212 for Caucasians. In pair wise comparison, the mean sequence difference of 14,36 &#177; 5,68 nucleotides over 3,486 pairs compared, indicates a high diversity in this population. The data demonstrates that mtDNA sequencing can be informative in forensic cases, but it is of crucial importance to know the frequency of the mtDNA haplotypes to statistically evaluate the results.

Com o intuito de avaliar os polimorfismos de mtDNA e a diversidade genética na população brasileira, foram estudados 84 indivíduos não aparentados: 49 caucasóides e 35 negróides da região sudeste do Brasil. As amostras de sangue obtidas por punção da polpa digital foram colocadas em papel de filtro e o mtDNA extraído pelo método orgânico. Foram amplificadas as regiões hipervariáveis HV1 a partir do nucleotídeo 16.051 até 16.365 e HV2 entre os nucleotídeos 71 e 340. O fragmento foi purificado e após a reação de seqüenciamento, as fitas sense e anti-sense foram alinhadas com a seqüência de Anderson e sobrepostas para confirmação. Foram encontradas 108 posições contendo transições, 15 transversões, 11 deleções e 4 inserções. Nove regiões apresentaram duas variantes de seqüência polimórfica. A maioria das seqüências foi observada uma única vez e em 84 indivíduos foram observados 80 haplótipos distintos. O índice de diversidade genética e a probabilidade de semelhança entre duas seqüências escolhidas ao acaso foram respectivamente de 0,9714 e 0,0286 para os negróides e de 0,9788 e 0,0212 para os caucasóides. Ao realizar análise pareada dos haplótipos, a média de nucleotídeos diferentes entre as seqüências foi de 14,36 &#177; 5,68 bases em um total de 3.486 pares comparados, indicando uma alta diversidade entre haplótipos nesta população. Os resultados demonstraram que o mtDNA é informativo para identificação humana e o conhecimento da freqüência dos haplótipos na população de interesse tem utilidade significativa na aplicação Forense, principalmente para se avaliar estatisticamente os resultados, fornecendo maior precisão, exatidão e dados contundentes nas investigações. / In order to evaluate the mtDNA polymorphism and demonstrate the sequence diversity in Brazilians, we established a database of 84 unrelated individuals, 49 Caucasians and 35 negroids from the southeast of Brazil. The samples were collected on a whatman paper by finger punction, the mtDNA was extracted by organic method and hipervariable regions, HV1 from 16.051 to 16.365 bp and HV2 from 71 to 340 bp were amplified. Both sense and anti-sense strands were sequenced. When compared to Anderson sequence, 108 transitions, 15 transversions, 11 deletions and 4 insertions were found. Nine positions exhibited two variants of polymorphic sequence. The majority of the sequences were observed once, and in 84 sequences, 80 haplotypes were observed. The sequence diversity and the probability for two individuals randomly matching over two hypervariable regions were 0.9714 and 0.0286 for persons of color and 0.9788 and 0.0212 for Caucasians. In pair wise comparison, the mean sequence difference of 14,36 &#177; 5,68 nucleotides over 3,486 pairs compared, indicates a high diversity in this population. The data demonstrates that mtDNA sequencing can be informative in forensic cases, but it is of crucial importance to know the frequency of the mtDNA haplotypes to statistically evaluate the results.

Drug facilitated assault (DFA) is an increasing problem in the UK. The crime often occurs through the surreptitious administration of a drug into a victims drink, rendering the victim unable to resist the assault. The detection of these drugs in a biological specimen from the victim is one of the most challenging facets of forensic chemistry. Drug concentrations can be very low, as often only a single dose is administered, and the pharmacodynamics of commonly employed drugs further hinders the testing process. The research presented in this work shows the development of several new assays for the detection of flunitrazepam, gamma-hydroxybutyrate (GHB) and ethyl glucuronide (EtG) in a variety of biological matrices. New methods of drug testing in blood and urine are demonstrated, as well as interesting developments in the field of hair testing. Using hair to detect drug exposure allows a much wider window of detection than the more traditional matrices of blood and urine. New methods are presented in this work using gas chromatography-tandem mass spectrometry (GCMS/MS) to detect drugs in hair. Validation data is presented along with the results of authentic DFA testing. All aspects of the drug testing procedure have been evaluated, from new extraction techniques utilising water instead of solvents, to novel clean up stages involving the unique combination of SFE and SPME. Several confirmation techniques are explored including single quadrupole, triple quadrupole and ion trap mass spectrometry. In addition to developing assays for DFA cases, the versatility of this type of analytical chemistry is explored in two population studies. The first study evaluates alcohol consumption between two groups; drugs users and non drug users in medico-legal cases. There is an anecdotal belief amongst drug clinic staff that alcohol use is lower in drugs users than it is in non drug users. This study presents the first scientific confirmation of this belief through EtG (an alcohol metabolite) testing in hair of the two groups. The second study investigates whether there is a correlation between EtG and cocaethylene (a metabolite of cocaine only produced in the presence of alcohol) in cocaine users. Results f this study suggest that there is no positive correlation between the two compounds. The research presented in this thesis aims to further the analytical science surrounding FA investigation and provide accurate, sensitive and reliable methodology for drug esting in blood, urine and hair.

Submitted by Liliane Ferreira (ljuvencia30@gmail.com) on 2018-11-27T10:54:13Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Tese - Thays Colletes de Carvalho - 2018.pdf: 10167238 bytes, checksum: e63cc8b9621f34c961582121789dd84c (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2018-11-27T11:56:05Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Tese - Thays Colletes de Carvalho - 2018.pdf: 10167238 bytes, checksum: e63cc8b9621f34c961582121789dd84c (MD5) / Made available in DSpace on 2018-11-27T11:56:05Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Tese - Thays Colletes de Carvalho - 2018.pdf: 10167238 bytes, checksum: e63cc8b9621f34c961582121789dd84c (MD5) Previous issue date: 2018-10-05 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Ionization is a crucial step in mass spectrometry (MS) and governs the versatility of this important analytical technique. Ionization is responsible for transferring atoms and molecules, in their respective ionized forms, into the high vacuum environment of mass spectrometers, where they are discriminated as a function of their m/z ratio. Among the several techniques of ionization of MS, the technique of ionization by Paper spray (PS) is one of the simplest, versatile and can be implemented easily in MS system. In this work, several PS-MS applications in the forensic area were developed, demonstrating their ability to screen new synthetic drugs as a complementary tool to the thin layer chromatography (TLC) method and as a tool to monitor Date-rape Drugs in blood. In the first application, PS-MS was shown to be an effective and rapid method for the identification of synthetic drugs lysergic acid diethylamide (LSD), 2,5-dimethoxy-4-chloroamphetamine (DOC), 2,5-dimethoxy-4-bromoamphetamine (DOB), 25C-NBOMe, 25B-NBOMe and 25I-NBOMe) in bottles, both by the isotopic profile of the substances and by the fragmentation presented in tandem mass spectrometry. In the second application, the TLC was coupled to the PS-MS in order to achieve greater reliability in the CCD results. In this way, a CCD method for the analysis of cocaine and adulterants (caffeine, benzocaine, lidocaine and phenacetin) was optimized by evaluating its sensitivity and selectivity. In order to improve the Detection and Quantification Limits for cocaine and adulterants, the spots were analyzed by PS-MS, obtaining improvements in the. Finally, in order to solve the low PS-MS selectivity, a new substrate was developed, especially when the analytical target is in complex mixtures, such as blood. It is a membrane coated with a molecularly printed polymer (MIP) with dual function: simultaneous extraction and ionization of targets in the same device. The developed membrane was applied in the determination of benzodiazepines in blood samples from alleged date rape- drugs victims. Blood is a complex sample containing several compounds that can suppress the analyte signal. With this modification, any suppression is avoided, obtaining excellent results, both qualitative and quantitative. In conclusion, PS-MS is a fast and low-cost technique that can replace or complement conventional analyzes in a laboratory of expertise, increasing the productivity of Brazilian justice. / A ionização é uma etapa crucial em espectrometria de massas (MS) e rege a versatilidade desta importante técnica analítica. A ionização é a responsável por transferir átomos e moléculas, em suas respectivas formas ionizadas, para o ambiente de alto vácuo dos espectrômetros de massas, onde são discriminados em função de sua razão massa sobre carga (m/z). Dentre as diversas técnicas de ionização de MS, a técnica de ionização por Paper spray (PS) é uma das mais simples, versátil e pode ser implementada facilmente em sistema de MS. Nesse trabalho diversas aplicações do PS-MS na área forense foram desenvolvidas, demonstrando sua capacidade para screening de novas drogas sintéticas, como ferramenta complementar ao método de cromatografia de camada delgada (TLC) e como ferramenta para monitorar “boa noite cinderela” em sangue. Na primeira aplicação, o PS-MS mostrou ser um método eficaz e rápido para identificação de drogas sintéticas dietilamida do ácido lisérgico (LSD), 2,5-dimetoxi-4-cloroanfetamina (DOC), 2,5-dimetoxi-4- bromoanfetamina (DOB), 25C-NBOMe, 25B- NBOMe e 25I-NBOMe) em selos, tanto pelo perfil isotópico das substâncias, quanto pela fragmentação apresentada na espectrometria de massas tandem. Já na segunda aplicação, acoplou-se TLC ao PS-MS visando alcançar maior confiabilidade nos resultados de TLC. Desta maneira, otimizou-se um método de TLC para análise de cocaína e adulterantes (cafeína, benzocaína, lidocaína e fenacetina) avaliando sua sensibilidade e seletividade. Com intuito de melhorar os Limites de Detecção e Quantificação para a cocaína e adulterantes, os spots foram analisados por PS-MS, obtendo melhorias nos resultados. Por fim, com o intuito de resolver a baixa seletividade do PS-MS, um novo substrato foi desenvolvido, principalmente quando o alvo analítico está em misturas complexas, como sangue. Trata-se de uma membrana recoberta com um polímero molecularmente impresso (MIP) com dupla função: extração e ionização simultânea de alvos em um mesmo dispositivo. A membrana desenvolvida foi aplicada na determinação de benzodiazepínicos em amostras de sangue de supostas vítimas de benzodiazepínicos. O sangue é uma amostra complexa que contém vários compostos que podem suprimir o sinal do analito. Com essa modificação qualquer supressão é evitada, conseguindo-se ótimos resultados, tanto qualitativos quanto quantitativos. Sendo assim, o PS-MS é uma técnica rápida e de baixo custo, que pode substituir ou complementar as análises convencionais em um laboratório de perícia, aumentando a produtividade da justiça brasileira.

Different sampling devices for paper spray - mass spectrometry (PS - MS) were investigated to improve the assay’s simplicity and sensitivity over traditional approaches. In the first one, pressure-sensitive adhesive paper was used as both sampling tool to collect drug residues on surfaces and paper substrate in PS - MS analysis. This method showed a significant improvement in drug collection on surfaces leading to low nano-gram level detection limits. Other sampling device being investigated was snap-in solid-phase extraction column, which demonstrated the ability to detect trace amounts of drugs in plasma while allowed easy transportation and the use of PS - MS automated system.

<p>Noble metal nanostructures display collective oscillation of the surface conduction electrons upon light irradiation as a form of localized surface plasmon resonance (LSPR) properties. Size, shape and the refractive index of surrounding environment are the key features that controls the LSPR properties. Surface passivating ligands have the ability to modify the charge density of nanostructures to allow resonant wavelength to match that of the incident light, a phenomenon called “plasmoelectric effect,”. According to the drude model Red and blue shifts of LSPR peak of nanostructures are observed in the event of reducing and increasing charge density, respectively. However, herein we report unusual LSPR properties of gold triangular nanoprisms (Au TNPs) upon functionalization with para-substituted thiophenols (X-Ph-SH, X = -NH₂, -OCH₃, -CH₃, -H, -Cl, -CF₃, and -NO₂). Accordingly, we hypothesized that an appropriate energy level alignment between the Au Fermi energy and the HOMO or LUMO of ligands allows delocalization of surface plasmon excitation at the hybrid inorganic-organic interface, and thus provides a thermodynamically driven plasmoelectric effect. We further validated our hypothesis by calculating the HOMO and LUMO levels and also work function changes of Au TNPs upon functionalization with para substituted thiol. We further utilized our unique finding to design ultrasensitive plasmonic substrate for biosensing of cancer microRNA in bladder cancer and owe to unpresidential sensitivity of the developed Au TNPs based LSPR sensor, for the first time we have been utilized to analysis the tumor suppressor microRNA for more accurate diagnosis of BC. Additionally, we have been advancing our sensing platform to mitigate the false positive and negative responses of the sensing platform using surface enhanced fluorescence technique. This noninvasive, highly sensitive, highly specific, also does not have false positives technique provide strong key to detect cancer at very early stage, hence increase the cancer survival rate. Moreover, the electromagnetic field enhancement of Surface-Enhanced Raman Scattering (SERS) and other related surface-enhanced spectroscopic processes resulted from the LSPR property. This dissertation describes the design and development of entirely new SERS nanosensors using flexible SERS substrate based on unique LSPR property of Au TNPs and developed sensors shows excellent SERS activity (enhancement factor = ~6.0 x 106) and limit of detection (as low as 56 parts-per-quadrillions) with high selectivity by chemometric analyses among three commonly used explosives (TNT, RDX, and PETN). Further we achieved the programable self-assembly of Au TNPs using molecular tailoring to form a 3D supper lattice array based on the substrate effect. Here we achieved the highest reported sensitivity for potent drug analysis, including opioids and synthetic cannabinoids from human plasma obtained from the emergency room. This exquisite sensitivity is mainly due to the two reasons, including molecular resonance of the adsorbate molecules and the plasmonic coupling among the nanoparticles. Altogether we are highly optimistic that our research will not only increase the patient survival rate through early detection of cancer but also help to battle the “war against drugs” that together is expected to enhance the quality of human life. </p> <p> </p>