Concerns about increased stratospheric ozone depletion increasing ambient levels of ultraviolet-B radiation (UV-B), and the fact that some ecosystems are naturally exposed to high levels, has resulted in an increased interest in the effects of UV-B on plant communities. Despite this, there has been a paucity of studies into its effects on plant competition. Artificial plant communities consisting of Lolium perenne and Lotus corniculatus and a sub-montane community consisting of Agrostis tenuis, Festuca ovina and Galium saxatile (also including different nitrogen levels) were created using the response surface approach. The long-term effects of UV-B were also studied on a natural sub-Arctic community in Abisko, Sweden. In addition, all plant samples were analysed by Fourier-Transform Infrared Spectroscopy (FT-IR) to obtain a ‘metabolic fingerprint’ which was used to detect chemical differences to the whole biochemical complement of the sample. The results showed that enhanced UV-B altered the competitive interaction of Lolium perenne and Lotus corniculatus in favour of Lolium perenne although ambient levels of UV-B did not elicit an effect in the sub-montane community. Only one dwarf shrub species in the sub-Arctic experiment, Vaccinium myrtillus, was negatively affected by UV-B. In most cases, elevated UV-B elicited a change in the metabolic fingerprint in the samples and in some cases an alteration in competitive stress altered the metabolome. This suggests that FT-IR can be used as a screening tool to detect for both abiotic stress and competitive biochemical alterations. In addition, this thesis proposes that the facilitative effect between the grass-legume mixture of Lolium perenne and Lotus corniculatus is not related to nitrogen fixation in the early stages of competition which has traditionally been believed.

Taxonomia vegetal é a ciência que trata da descrição, identificação, nomenclatura e classificação de plantas. O desenvolvimento de novas técnicas que podem ser aplicadas nesta área de conhecimento é essencial para dar suporte às decisões relacionadas a conservação de hotspots de biodiversidade. Nesta dissertação de mestrado foi desenvolvido um protocolo de metabolic fingerprinting utilizando MALDI-MS (matrix-assisted laser desorption/ionisation mass spectrometry) e subsequente análise multivariada utilizando scripts desenvolvidos para o pacote estatístico R. Foram classificadas, com base nos seus metabólitos detectados, 24 plantas de diferentes famílias vegetais, sendo todas elas coletadas em áreas da Savana Brasileira (Cerrado), que foi considerada um hotspot de biodiversidade. Metabolic fingerprinting compreende uma parte da Metabolômica, i.e., a ciência que objetiva analisar todos os metabólitos de um dado sistema (celula, tecído ou organismo) em uma dada condição. Comparada com outros métodos de estudo do metaboloma MALDI-MS apresenta a vantagem do rápido tempo de análise. A complexidade e importância da correta classificação taxonômica é ilustrada no exemplo do gênero Lychnophora, o qual teve diversas espécies incluídas neste estudo. No Brasil espécies deste gênero são popularmente conhecidas como \"arnica da serra\" ou \"falsa arnica\". Os resultados obtidos apontam similaridades entre a classificação proposta e a classificação taxonômica atual. No entanto ainda existe um longo caminho para que a técnica de metabolic fingerprinting possa ser utilizada como um procedimento padrão em taxonomia. Foram estudados e discutidos diversos fatores que afetaram os resultados como o preparo da amostra, as condições de análise por MALDI-MS e a análise de dados, os quais podem guiar futuros estudos nesta área de pesquisa. / Plant taxonomy is the science of description, identification, nomenclature and classification of plants. The development of new techniques that can be applied in this field of research are essential in order to assist informed and efficient decision-making about conservation of biodiversity hotspots. In this master\'s thesis a protocol for metabolic fingerprinting by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS) with subsequent multivariate data analysis by in-house algorithms in the R environment for the classification of 24 plant species from closely as well as from distantly related families and tribes was developed. Metabolic fingerprinting forms part of metabolomics, a research field, which aims to analyse all metabolites, i.e., the metabolome in a given system (cell, tissue, or organism) under a given set of conditions. Compared to other metabolomics techniques MALDI-MS shows potential advantages, mainly due to its rapid data acquisition. All analysed species were collected in areas of the Brazilian Savanna (Cerrado), which was classified as \"hotspot for conservation priority\". The complexity and importance of correct taxonomic classification is illustrated on the example of the genus Lychnophora, of which several species also have been included into analysis. In Brazil species of this genus are popularly known as \"arnica da serra\" or \"falsa arnica\". Similarities to taxonomic classification could be obtained by the proposed protocol and data analysis. However there is still a long way to go in making metabolic fingerprinting by MALDI-MS a standard procedure in taxonomic research. Several difficulties that are inherent to sample preparation, analysis of plant\'s metabolomes by MALDI-MS as well as data analysis are highlighted in this study and might serve as a basis for further research.

Taxonomia vegetal é a ciência que trata da descrição, identificação, nomenclatura e classificação de plantas. O desenvolvimento de novas técnicas que podem ser aplicadas nesta área de conhecimento é essencial para dar suporte às decisões relacionadas a conservação de hotspots de biodiversidade. Nesta dissertação de mestrado foi desenvolvido um protocolo de metabolic fingerprinting utilizando MALDI-MS (matrix-assisted laser desorption/ionisation mass spectrometry) e subsequente análise multivariada utilizando scripts desenvolvidos para o pacote estatístico R. Foram classificadas, com base nos seus metabólitos detectados, 24 plantas de diferentes famílias vegetais, sendo todas elas coletadas em áreas da Savana Brasileira (Cerrado), que foi considerada um hotspot de biodiversidade. Metabolic fingerprinting compreende uma parte da Metabolômica, i.e., a ciência que objetiva analisar todos os metabólitos de um dado sistema (celula, tecído ou organismo) em uma dada condição. Comparada com outros métodos de estudo do metaboloma MALDI-MS apresenta a vantagem do rápido tempo de análise. A complexidade e importância da correta classificação taxonômica é ilustrada no exemplo do gênero Lychnophora, o qual teve diversas espécies incluídas neste estudo. No Brasil espécies deste gênero são popularmente conhecidas como \"arnica da serra\" ou \"falsa arnica\". Os resultados obtidos apontam similaridades entre a classificação proposta e a classificação taxonômica atual. No entanto ainda existe um longo caminho para que a técnica de metabolic fingerprinting possa ser utilizada como um procedimento padrão em taxonomia. Foram estudados e discutidos diversos fatores que afetaram os resultados como o preparo da amostra, as condições de análise por MALDI-MS e a análise de dados, os quais podem guiar futuros estudos nesta área de pesquisa. / Plant taxonomy is the science of description, identification, nomenclature and classification of plants. The development of new techniques that can be applied in this field of research are essential in order to assist informed and efficient decision-making about conservation of biodiversity hotspots. In this master\'s thesis a protocol for metabolic fingerprinting by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS) with subsequent multivariate data analysis by in-house algorithms in the R environment for the classification of 24 plant species from closely as well as from distantly related families and tribes was developed. Metabolic fingerprinting forms part of metabolomics, a research field, which aims to analyse all metabolites, i.e., the metabolome in a given system (cell, tissue, or organism) under a given set of conditions. Compared to other metabolomics techniques MALDI-MS shows potential advantages, mainly due to its rapid data acquisition. All analysed species were collected in areas of the Brazilian Savanna (Cerrado), which was classified as \"hotspot for conservation priority\". The complexity and importance of correct taxonomic classification is illustrated on the example of the genus Lychnophora, of which several species also have been included into analysis. In Brazil species of this genus are popularly known as \"arnica da serra\" or \"falsa arnica\". Similarities to taxonomic classification could be obtained by the proposed protocol and data analysis. However there is still a long way to go in making metabolic fingerprinting by MALDI-MS a standard procedure in taxonomic research. Several difficulties that are inherent to sample preparation, analysis of plant\'s metabolomes by MALDI-MS as well as data analysis are highlighted in this study and might serve as a basis for further research.

Metabolomics approaches have recently been used to understand the complex molecular interactions of biological systems. One popular area in which these methods are being developed is to understand the biochemical changes during abiotic and biotic stresses; for example, how a cell may respond to a drug. Since metabolites are the end products of gene expression, these can be used to indicate the result of the activities and interaction of the cell or organism with its environment. The investigation of the level and compositional changes of metabolites against metabolic stresses such as chemotherapeutic treatment (drug exposure) are required to understand more fully abiotic perturbation to biological systems. The aim of this project was to understand the metabolic effect that the anti-viral drugs indinavir and lopinavir (currently used by HIV patients) have on HPV-related cervical cancer cell lines by measuring changes in metabolism using a wide range of analytical techniques; including Fourier transform infrared (FT-IR) and Raman spectroscopies, and gas and liquid chromatography-mass spectrometry (GC and LC-MS). The analyses and interpretation of the large volumes of complex multidimensional data generated by metabolomics approaches were performed with a combination of multivariate data analysis techniques such as principal components analysis (PCA) and canonical variates analysis (CVA), as well as univariate approaches such as N-Way analysis of variance (ANOVA). By combining biochemical imaging, metabolite fingerprinting and footprinting, and metabolite profiling, with multi- and uni-variate analyses, the actions and effects of the anti-viral drugs were investigated. FT-IR spectroscopy was initially used to generate global biochemical finger- and foot-prints, and Raman spectroscopy was employed to investigate intracellular distribution of metabolites, and other cellular species, as well as the localisation of drug molecules within cells. FT-IR spectroscopy ascertained that the intra- and extra-cellular metabolomes were being directly influenced in a fashion that correlated with increasing anti-viral dosing; these effects were phenotypic rather than measurements of the drug level. Raman imaging spectroscopy indicated that the indinavir but not lopinavir was being compartmentalised within the cell nucleus, but only in HPV early protein 6 (E6) expressing cells. This observation was further confirmed by fractionation of cell samples into nuclear and cytoplasmic fractions and assessing the indinavir concentrations via LC-MS. Finally, LC-MS and GC-MS metabolite profiling were employed to investigate changes in the intracellular metabolome in response to the anti-viral compounds across a range of physiologically relevant concentrations and in the presence and absence of the E6 oncoprotein. General effects of both anti-viral compounds included the regulation of metabolites such as glutathione, octenedionoic and octadecenoic acids, which may be involved in stress related responses, reduced levels of sugars and sugar-phosphates indicating a potential arrest of glycolysis, and reduced levels of malic acid indicating potential decreased flux into the TCA cycle; all indicating that central metabolism was being reduced. Finally, LC-MS based quantification indicated that in the presence of E6, lopinavir was actively removed from the cell, whereas the indinavir intracellular concentration increased concomitantly with the level of dosing. These investigations have revealed that metabolomics approaches are an apt tool for the study of anti-viral effects within cell cultures, but improvements need to be made with respect to the major limitation of metabolite identification.

Maize is the staple food for a majority of people in Southern Africa, but plant diseases are responsible for at least 10% of crop production losses. Bacterial leaf streak (BLS) of maize was first reported in South Africa in 1949 and has not been reported elsewhere. Very little is known about the pathogen involved and therefore it is deemed necessary to compile a characteristic profile for the pathogen to prevent the possibility of major crop losses as a result of this disease. This study aimed to use biochemical and phenotypic methods to determine the specific characteristics of the causal agent of BLS. Diseased plant material showing symptoms of BLS were collected during the maize production seasons of 2012 and 2013 within South Africa’s maize production regions namely the North West, Free State, Gauteng and Northern Cape provinces. To prevent contamination, maize leaves were surface sterilised thoroughly before bacterial isolation commenced. Sections of the infected maize leaves were placed on GYC agar plates on which yellow, mucoid bacterial colonies after incubation for 24 to 48 hrs. The isolated bacteria were purified and the molecular identification of the bacteria was conducted in a related study. Although literature indicates that Xanthomonas campestris pv. zeae is the causal agent of BLS, pure cultures obtained from maize leaves showing characteristic symptoms of BLS were identified as species of Xanthomonas, Pantoea, and Enterobacter. To elucidate the pathogenicity of the isolated strains, pathogenicity tests based on Koch’s postulates were performed. Results from the pathogenicity tests confirmed that only the isolate Xanthomonas species was capable of inducing the characteristic BLS symptoms when healthy maize plants were inoculated with the suspected pathogens. It is important to inoculate the maize seedlings at the correct age (four-leaf stage) and the spray method is recommended. Re-isolation was repeated from the same plant material used during the initial isolation process but the isolation method was amended. The optimised isolation method involved the use of a dilution range and spread plate method. Colonies from this isolation technique grew as bright yellow colonies that were identified as Xanthomonas spp. This outcome indicates the importance of surface sterilisation, pulverisation and subsequent dilution of plant materials for isolation of bacterial pathogens from diseases plants. These isolates were used to create protein profiles with SDS-PAGE electrophoresis and carbon utilisation patterns with the Biolog® GN2 system. Protein profiling banding patterns was assessed based on presence/absence criteria. Highly similar protein profiles were observed among the X. campestris pv. zeae isolates but groupings of different protein profiles were determined when minor differences in the protein profiles was taken into account. Xanthomonas campestris pv. zeae was successfully distinguished from the X. axonopodis pv. vasculorum reference strain through unique SDS banding patterns. Banding patterns obtained from cultures grown in a liquid medium (tryptic soy broth) were of a higher quality than the banding patterns obtained from bacteria harvested from solid media (CYG agar plates). Carbon source utilisation data was used to evaluate the average well colour development obtained from each isolate. Statistically significant differences were found among some of the isolates, with some isolates being metabolically more active than other isolates. Substrate utilisation patterns produced by the isolates corresponded to previously published studies on various Xanthomonas species. The cell count of the samples used during carbon utilisation patterns must be standardised in order to obtain reliable results. During this study, the application of Koch’s postulates and two inoculation techniques confirmed that Xanthomonas campestris pv. zeae is the pathogen responsible for bacterial leaf streak of maize. Members of the Pantoea and Enterobacter genera were found on the leaf surface of maize plants infected with BLS but inoculations of healthy maize plants with these bacteria did not result in bacterial leaf streak symptoms on the maize plants. These bacteria were not pathogenic and were considered endophytes. The identified pathogen was characterised through protein and metabolic profiling. The protein profiles of the pathogen obtained through analysis of the major bands of the SDS-PAGE gels were highly similar and distinguishable from the Xanthomonas reference culture. Groupings within the X. campestris pv. zeae group was found when major and minor bands were considered, this may however be altered when the intensities of the bands are used during analysis. Carbon utilisation patterns were assessed using Biolog® GN2 plates. A metabolic fingerprint was created for the pathogen of BLS, it was possible to distinguish between X. campestris pv. zeae and other Xanthomonas strains based on the fingerprint. This fingerprint could be used to identify the pathogen. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

Maize is the staple food for a majority of people in Southern Africa, but plant diseases are responsible for at least 10% of crop production losses. Bacterial leaf streak (BLS) of maize was first reported in South Africa in 1949 and has not been reported elsewhere. Very little is known about the pathogen involved and therefore it is deemed necessary to compile a characteristic profile for the pathogen to prevent the possibility of major crop losses as a result of this disease. This study aimed to use biochemical and phenotypic methods to determine the specific characteristics of the causal agent of BLS. Diseased plant material showing symptoms of BLS were collected during the maize production seasons of 2012 and 2013 within South Africa’s maize production regions namely the North West, Free State, Gauteng and Northern Cape provinces. To prevent contamination, maize leaves were surface sterilised thoroughly before bacterial isolation commenced. Sections of the infected maize leaves were placed on GYC agar plates on which yellow, mucoid bacterial colonies after incubation for 24 to 48 hrs. The isolated bacteria were purified and the molecular identification of the bacteria was conducted in a related study. Although literature indicates that Xanthomonas campestris pv. zeae is the causal agent of BLS, pure cultures obtained from maize leaves showing characteristic symptoms of BLS were identified as species of Xanthomonas, Pantoea, and Enterobacter. To elucidate the pathogenicity of the isolated strains, pathogenicity tests based on Koch’s postulates were performed. Results from the pathogenicity tests confirmed that only the isolate Xanthomonas species was capable of inducing the characteristic BLS symptoms when healthy maize plants were inoculated with the suspected pathogens. It is important to inoculate the maize seedlings at the correct age (four-leaf stage) and the spray method is recommended. Re-isolation was repeated from the same plant material used during the initial isolation process but the isolation method was amended. The optimised isolation method involved the use of a dilution range and spread plate method. Colonies from this isolation technique grew as bright yellow colonies that were identified as Xanthomonas spp. This outcome indicates the importance of surface sterilisation, pulverisation and subsequent dilution of plant materials for isolation of bacterial pathogens from diseases plants. These isolates were used to create protein profiles with SDS-PAGE electrophoresis and carbon utilisation patterns with the Biolog® GN2 system. Protein profiling banding patterns was assessed based on presence/absence criteria. Highly similar protein profiles were observed among the X. campestris pv. zeae isolates but groupings of different protein profiles were determined when minor differences in the protein profiles was taken into account. Xanthomonas campestris pv. zeae was successfully distinguished from the X. axonopodis pv. vasculorum reference strain through unique SDS banding patterns. Banding patterns obtained from cultures grown in a liquid medium (tryptic soy broth) were of a higher quality than the banding patterns obtained from bacteria harvested from solid media (CYG agar plates). Carbon source utilisation data was used to evaluate the average well colour development obtained from each isolate. Statistically significant differences were found among some of the isolates, with some isolates being metabolically more active than other isolates. Substrate utilisation patterns produced by the isolates corresponded to previously published studies on various Xanthomonas species. The cell count of the samples used during carbon utilisation patterns must be standardised in order to obtain reliable results. During this study, the application of Koch’s postulates and two inoculation techniques confirmed that Xanthomonas campestris pv. zeae is the pathogen responsible for bacterial leaf streak of maize. Members of the Pantoea and Enterobacter genera were found on the leaf surface of maize plants infected with BLS but inoculations of healthy maize plants with these bacteria did not result in bacterial leaf streak symptoms on the maize plants. These bacteria were not pathogenic and were considered endophytes. The identified pathogen was characterised through protein and metabolic profiling. The protein profiles of the pathogen obtained through analysis of the major bands of the SDS-PAGE gels were highly similar and distinguishable from the Xanthomonas reference culture. Groupings within the X. campestris pv. zeae group was found when major and minor bands were considered, this may however be altered when the intensities of the bands are used during analysis. Carbon utilisation patterns were assessed using Biolog® GN2 plates. A metabolic fingerprint was created for the pathogen of BLS, it was possible to distinguish between X. campestris pv. zeae and other Xanthomonas strains based on the fingerprint. This fingerprint could be used to identify the pathogen. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

No description available.

Le métabolisme secondaire joue un rôle écologique majeur dans les interactions des organismes avec leur environnement. Une étude intégrée de la biologie, de l’écologie des organismes et des variations de leur métabolisme est essentielle pour comprendre le rôle des métabolites secondaires au sein des écosystèmes. Or ce type d’approche est rare en milieu marin.Le petit clade d’éponges Homoscleromorpha constitue un vrai potentiel de découverte de nouvelles espèces et de molécules bioactives valorisables. Par ailleurs, leur position de dominance dans certaines communautés benthiques de Méditerranée en faisait un modèle de choix pour démarrer des recherches en écologie chimique marine.Ce travail a débuté avec la description d’une espèce du genre Oscarella, O. balibaloi ainsi que de nouvelles molécules produites par cet organisme. Cette nouvelle espèce avec deux autres espèces communes du même genre, O. tuberculata et O. lobularis constituent parfois de vrais faciès au sein des communautés du coralligène et des grottes semi-obscures. L’étude comparée du cycle de vie de ces Oscarella a montré dans tous les cas une reproduction saisonnière, avec des différences dans les périodes de gamétogénèse et d’émission des larves, et des sensibilités variables face aux changements des conditions de régime thermique. Deux composés majoritaires de type lysophospholipides ont été isolés et caractérisés pour la première fois dans O. tuberculata, et retrouvés dans O. lobularis. Leur rôle potentiel de médiateurs moléculaires impliqués dans le processus de reproduction (embryogenèse et développement) a été proposé, et devra être confirmé par des études expérimentales. Une nouvelle famille de sesterterpènes glycosylés (dénommés balibalosides) a été découverte dans O. balibaloi.Une étude pluriannuelle des variations du métabolisme de ces espèces a été réalisée à travers trois approches complémentaires permettant de tester les modalités d’allocation des ressources à la production de métabolites secondaires. Les patrons de variation des niveaux d’expression de métabolites ciblés, des signatures métaboliques et des bioactivités des extraits d’éponges ont montré une influence significative du cycle de reproduction sur le métabolisme secondaire. Les méthodes globales (métabolomique et bioactivité) ont permis de montrer que le cycle de variation du métabolisme secondaire était marqué par une modification importante de sa production accompagnée par une baisse de bioactivité pendant les périodes les plus coûteuses de la reproduction (reproduction asexuée, embryogenèse et développement larvaire). Ces résultats montrent un compromis d’allocation des ressources entre un métabolisme primaire (la reproduction) et la production des métabolites secondaires, et soutient ainsi la théorie de défense optimale.L’approche de métabolomique s’est avérée un bon indicateur de la chimio-diversité. Appliquée à l’étude des relations inter-spécifiques, cette méthode de chimio-systématique a permis de proposer une classification des espèces méditerranéennes d’Homoscleromorpha. Cette classification soutient les résultats les plus récents de phylogénie moléculaire et propose la restauration de deux anciens clades au sein des Homoscleromorpha: les Plakinidae, un groupe qui ne contient aujourd’hui que des espèces à squelette, et les Oscarellidae qui ne contient que des espèces sans squelette. Les approches développées au cours de cette thèse permettent de nombreuses perspectives en chimio-systématique et écologie chimique marine. L’utilisation des signatures métaboliques peut être transposée à d’autres questions de systématique, particulièrement pour démontrer l’existence d’espèces cryptiques, et pour soutenir des hypothèses phylogénétiques au sein d’autres clades problématiques [...] / Secondary metabolism plays a major ecological role in the interactions between the organisms and their environment. An integral study of the organisms’ biology and ecology and the variations of their metabolism is essential for understanding the role of secondary metabolites in the ecosystems. This kind of approach is rare in the marine environment. Small sponge clade Homoscleromorpha constitutes a real potential for the discovery of new species and potentially bioactive molecules. In addition, its dominance in some Mediterranean benthic communities makes it a good model in marine chemical ecology research. This work has started with a description of new species of Oscarella genus, O. balibaloi. This new species forms sometimes, with two other commun Oscarella species, O. tuberculata and O. lobularis, special facies within the coralligenous and semi-dark cave communities. All three Oscarella species are caraterized by a seasonal reproductive cycle with differences in the period of gametogenesis and larval emission as well as the variation in sensitivity facing the changes in thermal regime. Two major lysophospholipid compounds were isolated and caracterized for the first time in O. tuberculata and confirmed in O. lobularis. Their potential role as signal molecules in the reproduction process (embryogenesis and development) was proposed and should be confirmed by experimental studies. One new familly of glycosilated seterterpens (named balibalosides) was found in O. balibaloi. A pluriannual study of species metabolism was performed using three complementary approaches and enabled to test the models of resource allocation to secondary metabolite production. Variation patterns in the expression level of target metabolites, in the metabolic fingerprints and the bioactivities of sponge extracts reflected the significant influence of the reproductive cycle to the secondary metabolite production. Holisitic approaches (métabolomics and bioactivity) pointed out the important modification in the secondary metabolism variation pattern followed by the decrease in bioactivity during the costly period of reproduction (asexual reproduction, embryogenesis and larval development). These results highlight the trade-off in resource allocation between the primary (reproduction) and secondary metabolism and therefore support the Optimal Defense Theory. Metabolomic approach applied to the study of interspecific relations turned out as a good indicator of chemical diversity which allowed the classification of Mediterranean Homoscleromorpha sepcies. The obtained classification was congruent with recent molecular phylogeny results proposing the restauration of two ancient clades within Homoscleromorpha, the Plakinidae, a group of species possesing skeleton and the Oscarellidae, a group of species lacking skeleton. Approaches developed during my thesis opened a numerous perspectives in chemosystematics and marine chemical ecology. The use of metabolic fingerprints can be transposed to other questions in systematics, particularly to demonstrate the existance of cryptic species and to support phylogenetic hypothesis within other problematic clades. [...]

Fatty acids (FAs) represent an important class of metabolites, impacting on membrane building blocks and signaling compounds in cellular regulatory networks. In nature, prokaryotes are characterized with the most impressing FA structural diversity and the highest relative content of free fatty acids (FFAs). In this context, nitrogen-fixing bacteria (order Rhizobiales), the symbionts of legumes, are particularly interesting. Indeed, the FA profiles influence the structure of rhizobial nodulation factors, required for successful infection of plant root. Although FA patterns can be assessed by gas chromatography—(GC-) and liquid chromatography—mass spectrometry (LC-MS), sample preparation for these methods is time-consuming and quantification suffers from compromised sensitivity, low stability of derivatives and artifacts. In contrast, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) represents an excellent platform for high-efficient metabolite fingerprinting, also applicable to FFAs. Therefore, here we propose a simple and straightforward protocol for high-throughput relative quantification of FFAs in rhizobia by combination of Langmuir technology and MALDI-TOF-MS featuring a high sensitivity, accuracy and precision of quantification. We describe a step-by-step procedure comprising rhizobia culturing, pre-cleaning, extraction, sample preparation, mass spectrometric analysis, data processing and post-processing. As a case study, a comparison of the FFA metabolomes of two rhizobia species—Rhizobium leguminosarum and Sinorhizobium meliloti, demonstrates the analytical potential of the protocol.