The understanding of analytical chemistry and its application in different areas of life science is something that should be encouraged for students studying chemistry, and also something that appeals to many students. Life science includes a vast variety of areas, out of which some are more easily approached in student projects. The chemical communication between organisms, such as plants, as described in chemical ecology is one example. It is therefore beneficial to offer students inquiry-based project work that allows them to explore, and gain a deeper understanding of, this discipline within the grand context that is chemistry. Hence, the present work describes the application of analytical chemistry using gas chromatography-mass spectrometry, including sample preparation for the study of chemical inhibitory (allelopathic) properties of compounds found in the plant Aegopodium podagraria. The questions posed to the students performing this project work were "What compounds can be found in the extracts of the plant?", "Are the extracts allelopathic?", and "Can you determine which compounds contribute to the possible allelopathic properties?". The reported results indicate that the extracts might have allelopathic properties and showed that the extracts contained compounds such as alpha-pinene and beta-caryophyllene. The identified terpenes were shown to have minor allelopathic properties by themselves but displayed an impact on the germination of seeds and length of the sprouts when applied as a blend. This project, and its results, proposes a framework for investigation of other plants and can be adapted to suit students at different academic levels.

An important aspect of understanding how plants effect their environmentis the study of volatile compounds and how these influence interactions between plants and other living organisms. The study of these secondary metabolites from plants can be used in practical applications, such as behaviour control of insects, and is an important part in the search for novel compounds that can be used to influence the behaviour of disease spreading vectors. The study of secondary metabolites can also be used as tools for introducing and teaching chemical ecology and analytical chemistry. The proper use of sample preparation methods and analysis has a key role in both these applications.

In this thesis, different sample collection methods are combined with gas chromatography-mass spectrometry to investigate the diversity of secondary metabolites emitted from plants. In paper I, two different sample collection methods, ultrasound assisted extraction and solid-phase microextraction, were used as a basis to collect six metabolites as a metric of comparison of the methods. The work compares the ultrasound assisted extraction and solid-phase microextraction of stored Cyperus rotundus, and also compares solid-phase microextraction collections from stored and live plant samples.

The static headspace solid-phase microextraction method used in paper I was then used to analyse the headspace samples of three other graminoid plants, namely Cynodon dactylon, Cyperus exaltatus and Panicum repens. In paper III, the identification of the headspace volatiles of the graminoid plants were determined, and the relative peak area percentages were analysed using one-way ANOVA. The result of this work allowed for a comparison between the three different plants to determine significant differences between the compounds detected based on relative peak area percentages.

Adding to the results of paper I and III, the results from paper II offers more insight into the chemicals present in the gas phase above the plants by employing dynamic headspace sampling from live plants. This adds another sampling method to the body of the work, which expands the knowledge of what volatile compounds are present around the plants. Further, in paper II, the use of two-port olfactometric and semi-field bioassays were used to compare which plants have the largest impact on the behaviour of the malaria vector Anopheles gambiae.

In Paper IV, the ultrasound assisted extraction method from paper I was used in order to evaluate the allelopathy of Aegopodium podagraria. This work was used as an inquiry-based learning project for Swedish highschool students, and the feedback of the students were collected for development of the project. The results of the work in paper IV demonstrated the implementation of the developed experimental methods as tools for teaching students of different academic levels about analytical chemistry and chemical ecology.

En viktig aspekt för förståelsen av hur växter påverkar sin omgivning är studien av flyktiga ämnen och hur dessa påverkar interaktionen mellan växter och andra levande organismer. Studier av sekundära metaboliter från växter kan användas i praktiska applikationer, så som beetendekontroll av insekter, till exempel myggor, där denna kunskap är en viktig del i sökandet efter nya ämnen som kan användas för att påverka beteendet hos smittbärande insekter. Studier av sekundära metaboliter kan också användas som ett redskap för att introducera och undervisa i kemisk ekologi och analytisk kemi. God användning av provpreparering och provanalys har en viktig roll i båda applikationerna.

I denna avhandling så används olika provberedningsmetoder, i kombination med gaskromatografi-masspektometri, för att undersöka den mångfald av sekundära metaboliter som utsöndras från växter. I paper I används två olika provuppsamlingsmetoder, ultraljudsassisterad extraktion och fastfas-mikroextraktion, som grunden för uppsamling av sex metaboliter som mått för jämförelse mellan metoderna. Detta arbete jämför provuppsamling med ultraljudsassisterad extraktion och fastfasmikroextraktion på gräs som har förvarats en tid innan analys, men jämför även fastfas-mikroextraktion uppsamlingar från levande gräs med det som legat på förvaring.

Den statiska gasfas fastfas-mikroextraktionsmetod som användes i paper I, användes sedan också för att analysera den kemiska gasfasen för tre andra gräs-liknande växter, nämligen Cynodon dactylon, Cyperus exaltatus och Panicum repens. I paper III så utförs identifiering av flyktiga ämnen från de gräs-liknande växternas gasfas, och deras relativa toppareaprocent analyseras med one-way ANOVA. Resultaten från detta arbete möjliggör en jämförelse mellan de tre växter för att avgöra om man kan se signifikanta skillnader för de ämnen som detekteras baserat på den relativa toppareaprocenten.

Vidare, tillför arbetet från paper II mer insikt om de kemiska föreningar som kan finnas i gasfasen kring växterna utöver det som har visats i paper I och paper III, genom användning av dynamisk gasfasuppsamling från levande växter. Detta bidrar med ytterligare en provuppsamlingsmetod,vilket vidare utökar informationen om vilka flyktiga ämnen som finns kring växterna. I paper II används två-ports olfaktometrisk-, samt semi-field bioanalyser för att jämföra vilken växt som har störst inverkan på malariavektorn Anopheles gambiae beteende.

I paper IV så används den ultraljudsassisterade extraktionsmetoden från paper I för att utvärdera den allelopatiska förmågan hos Aegopodium podagraria. Detta arbete användes som ett undersökande arbetsprojektför svenska gymnasieelever, och återkoppling från eleverna användes föratt utveckla projektet. Arbete från paper IV visar sig användbart som ett verktyg för utlärning, på olika akademiska nivåer, i ämnena kemisk ekologi samt analytisk kemi.

In recent work, it was shown that the graminoid plants Cynodon dactylon (Poaceae), Cyperus exaltatus (Cyperaceae), and Panicum repens (Poaceae) have an ovipositional effect on the malaria vector Anopheles gambiae in olfactometric bioassays. In order to get a view of the diversity of semiochemicals present in the environment of the vector during olfactometric trials, in the present work, the volatile profiles of these graminoid plants were analyzed using headspace solid-phase microextraction (HS-SPME) together with gas chromatography-mass spectrometry (GC-MS). In addition, one-way ANOVA comparison of compounds detected in two or more headspace samples are presented in order to provide a basis for comparison of compounds that could constitute a starting point for novel blends of volatile organic compounds to be tested as oviposition attractants.

Background: Understanding the ecology and behaviour of disease vectors, including the olfactory cues used to orient and select hosts and egg-laying sites, are essential for the development of novel, insecticide-free control tools. Selected graminoid plants have been shown to release volatile chemicals attracting malaria vectors; however, whether the attraction is selective to individual plants or more general across genera and families is still unclear. Methods: To contribute to the current evidence, we implemented bioassays in two-port airflow olfactometers and in large field cages with four live graminoid plant species commonly found associated with malaria vector breeding sites in western Kenya: Cyperus rotundus and C. exaltatus of the Cyperaceae family, and Panicum repens and Cynodon dactylon of the Poaceae family. Additionally, we tested one Poaceae species, Cenchrus setaceus, not usually associated with water. The volatile compounds released in the headspace of the plants were identified using gas chromatography/mass spectrometry. Results: All five plants attracted gravid vectors, with the odds of a mosquito orienting towards the choice-chamber with the plant in an olfactometer being 2-5 times higher than when no plant was present. This attraction was maintained when tested with free-flying mosquitoes over a longer distance in large field cages, though at lower strength, with the odds of attracting a female 1.5-2.5 times higher when live plants were present than when only water was present in the trap. Cyperus rotundus, previously implicated in connection with an oviposition attractant, consistently elicited the strongest response from gravid vectors. Volatiles regularly detected were limonene, beta-pinene, beta-elemene and beta-caryophyllene, among other common plant compounds previously described in association with odour-orientation of gravid and unfed malaria vectors. Conclusions: The present study confirms that gravid Anopheles gambiae sensu stricto use chemical cues released from graminoid plants to orientate. These cues are released from a variety of graminoid plant species in both the Cyperaceae and Poaceae family. Given the general nature of these cues, it appears unlikely that they are exclusively used for the location of suitable oviposition sites. The utilization of these chemical cues for attract-and-kill trapping strategies must be explored under natural conditions to investigate their efficiency when in competition with complex interacting natural cues.