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The FAME yield from microalgae of two in situ transesterification methods were compared to a typical Folch et al. (1957) extraction followed by transesterification using the Hilditch et al. (1964) procedure. A method based on Park & Goins (1994), utilizing 0.5 N NaOH in methanol, then 14 % BCl3 in methanol, was found to be superior to a method based on Lepage & Roy (1986), utilizing acetyl chloride in methanol. The Park & Goins (1994) method was equivalent to the traditional method and was, therefore, selected for further study. In establishing the parameters of the method, water contents up to 0.55 mL were not found to inhibit the reaction within the maximum lipid load, conservatively assessed at ~1 mg. The reaction time and temperature required to produce a maximum FAME yield was 10 min at 90 °C for the BCl3-catalyzed reaction, while the NaOH-catalyzed reaction happened instantaneously at ambient temperature.

<p dir="ltr">This thesis focuses on ion-ion reactions for making ions for lipid structural elucidation and chemical reactions.</p>

Bedrock features represent various economic, social, and symbolic aspects of past societies, but have historically received little study, particularly in North America. Fortunately, new techniques for analyzing spatial configurations, use-wear, and organic residues are beginning to unlock more of the interpretive potential of these features. Though preliminary in nature, the present study contributes to this trend by documenting an application of lipid analysis to bedrock features in a dry rockshelter. Results of this initial application indicate that bedrock features in dry rockshelters may provide especially favorable conditions for the preservation and interpretation of ancient organic residues. Abundant lipids, comparable to concentrations present in some pottery sherds, were extracted from a bedrock grinding surface at Gila Cliff Dwellings National Monument and analyzed using gas chromatography-mass spectrometry. Though the lipids were highly oxidized, degradation products indicative of former unsaturated fatty acids were retained. Comparisons to experimentally aged residues, and absence of a known biomarker for maize, indicate that the bulk of the lipids preserved in the milling surface probably derive from processing an oily nut or seed resource, and not from processing maize. Substantially lower amounts of lipids were recovered from a small, blackened cupule. It is hypothesized that some portion of the lipids in the blackened cupule was deposited from condensed smoke of cooking and heating fires in the caves. Potential for the preservation of organic residues in similar sheltered bedrock contexts is discussed, and a practical method for sampling bedrock features in the field is described.

Biofuel is a renewable and sustainable energy source with near-neutral carbon footprint. Algae are an ideal feedstock for biofuel production because they reproduce quickly and have high oil. Algae can be cultivated in non-arable land, and would not impact the food supply. Unfortunately, processing algae into biofuel is more expensive than land crops due to the large volumes of dilute algal suspension that must be harvested and concentrated. In order to improve algae-based biofuel economics, resins were developed that reduce costs associated with water pumping and transport. Hydrophobic resins were developed for binding oil out of an algal suspension so that the residual biomass could be recovered without solvent contamination. Binding behavior displayed lipid species specificity, and binding capacity was improved by ethanol treatment of the biomass. Algae was harvested by binding to anion exchange resin and directly converted into biodiesel. One-step, room temperature in situ transesterification of algae yielded nearly as much biodiesel as two-step, heated transesterification of dried biomass. Elution with transesterification reagent also regenerated the resin for subsequent algal binding. Functionalized resins were developed with high algal binding capacity at neutral pH. Binding was easily reversed, as treatment with buffer with pH higher than pKa of the resin functional group removed the algae and regenerated the resin for subsequent use. The resin bound 10% of its weight in algae and released it as a 100-fold concentrated suspension. The polymers developed can be scaled up for commercial processes and reduce algal harvesting and concentration costs. / text

Integral membrane proteins (IMPs) assume critical roles in cell biology and are key targets for drug discovery. Given their involvement in a wide range of diseases, the structural and functional characterization of IMPs are of significant importance. However, this remains notoriously challenging due to the difficulties of stably purifying membrane-bound, hydrophobic proteins. Compounding this, many diseases are caused by IMP mutations that further decrease their stability. One such example is cystic fibrosis (CF), which is caused by misfolding or dysfunction of the epithelial cell chloride channel cystic fibrosis transmembrane conductance regulator (CFTR). Roughly 70% of CF patients world-wide harbor the ΔF508-CFTR mutation, which interrupts CFTR’s folding, maturation, trafficking and function. No existing treatment sufficiently addresses the consequences of ΔF508, and the substantial instability that results from this mutation limits our ability to study ΔF508-CFTR in search of better treatments. To that end, my colleagues at Sanofi generated homology models of full-length wild-type and ΔF508-CFTR +/- second-site suppressor mutations (SSSMs) V510D and R1070W, and performed molecular dynamics (MD) simulations for each model. Using information obtained from this analysis, I tested several hypotheses on the mechanism by which ΔF508 destabilizes full-length CFTR and how SSSMs suppress this effect. Leveraging studies of the purified NBD1 subdomain and of full-length CFTR in a cellular context, I confirmed the prediction of a key salt-bridge interaction between V510D and K564 important to second-site suppression. Furthermore, I identified a novel class of SSSMs that support a key prediction from these analyses: that helical unraveling of TM10, within CFTR’s second transmembrane domain, is an important contributor to ΔF508-induced instability. In addition, I developed a detergent-free CFTR purification method using styrene-maleic acid (SMA) copolymer to extract the channel directly from its cell membrane along with the surrounding lipid content. The resulting particles were stable, monodisperse discs containing a single molecule of highly-purified CFTR. With this material, I optimized grid preparation techniques and carried out cryo-EM structural analysis of WT-hCFTR which resulted in 2D particle class averages which were consistent with an ABC transporter shape characteristic of CFTR, and a preliminary 3D reconstruction. This result establishes a foundation for future characterization of ΔF508-CFTR in its native state. I have also applied this SMA-based purification method to the facilitated glucose transporter GLUT1 (SLC2A1). SLC2A1 mutations contribute to a rare and developmentally debilitating disease called GLUT1-deficiency syndrome. Using SMA, I successfully extracted GLUT1 in its native state. With the application of this method, I was able to purify endogenous GLUT1 from erythrocytes, in complex with several associated proteins as well as the surrounding lipids, in its monomeric, dimeric and tetrameric forms without the use of cross-linking or chimeric mutations. These results point to the potential for studying isolated IMPs without the use of destabilizing detergents and thereby offer a pathway to analysis of wild-type and mutant membrane protein structure, function and pharmacodynamics.

The Early Iron Age period has been underexplored in the archaeology of West Africa, particularly Nigeria. Also, there is a lack of research and knowledge on organic residue analysis in Nigeria, particularly in the southwestern region of the country, despite the fact that other studies on archaeobotany and paleoenvironmental studies are well established. To address this gap, lipid analysis was conducted on 10 vessels from the Early Iron Age (490 B.C. to A.D. 40) site BSM6 in Bara, Old Oyo, Nigeria. The analysis revealed the plant and animal materials consumed by the Early Iron Age community. These findings provide vital insights into the dietary and subsistence practices of the period, illustrating continuities and changes in food practices from ancient to present times.

This paper is about food culture in Uppland under early iron age in Upplans, Sverige. Gas chromatography–mass spectrometry (GC-MS) was used to analysis the lipids that were extracted from archaeological potsherds from Påtåker Raä 62, Sollentuna, Uppland. The result of the lipid analysis shows content of aquatic animal products, terrestrial animal products, vegetables and indication of being heated. Based on these results and what is considered to be a normal diet during the Iron Age, it is possible to say that there are similarities. These results are also compared with three sites from Late Iron Age – Vendel 1:1, Vendel 28 and Tuna, to see if there are any similarities. The reason to choose three sites from Late Iron Age, and not Early Iron Age, is because there haven’t been done studies like this on material from the early Iron Age. Vendel 28 was the site that was most similar to Påtåker, when it comes to its enviorment with meadows and woods, and the ceramics application areas. / This study is part of the on going research of Påtåker Raä 62, Sollentuna, Oppland.

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