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Molecular stable isotope ratios are a novel type of dietary biomarker with high sensitivity and specificity for certain foods. Among these, fatty acid carbon isotope ratios (CIRs) have strong potential but have not been investigated as dietary biomarkers.We evaluated whether fatty acid CIRs and mass proportions were associated with meat, fish and sugar sweetened beverage (SSB) intake.Thirty-two men (aged 46.2 ± 10.5 y; BMI: 27.2 ± 4.0 kg/m2) underwent a 12-wk inpatient dietary intervention at the National Institute of Diabetes and Digestive and Kidney Diseases in Phoenix, Arizona. Men were randomly assigned to one of eight dietary treatments varying the presence/absence of dietary meat, fish and SSBs in all combinations. Fatty acid CIRs and mass proportions were measured in fasting blood samples and adipose tissue biopsies that were collected pre- and post-intervention. Dietary effects were analyzed using multivariable regression and receiver operating characteristic AUCs were calculated using logistic regression.CIRs of the most abundant saturated (14:0, 16:0, 18:0) and mono-unsaturated (16:1n-7, 18:1) fatty acids and arachidonic acid (20:4n-6) in plasma were strongly associated with meat, as were a subset of these fatty acids in RBC. Effect sizes in plasma ranged from 1.01 to 1.93‰ and were similar but attenuated in RBC. Mass proportions of those fatty acids were not associated with diet. CIRs of plasma 20:3n-6 and adipose 16:0 were weakly associated with SSB. Mass proportions of plasma odd-chain fatty acids i-15:0, 17:0, and 17:1 were associated with meat, and mass proportions of plasma EPA and DHA (20:5n-3 and 22:6n-3) were associated with fish.CIRs of plasma saturated and mono-unsaturated fatty acids show promise as sensitive and specific measures of dietary meat. These provide different information to that provided by fatty acid mass proportions, and are informative where mass proportion is not.

Accurate information on diet composition is central to understanding and conserving carnivore populations. Quantitative fatty acid signature analysis (QFASA) has emerged as a powerful tool for estimating the diets of predators, but ambiguities remain about the timeframe of QFASA estimates and the need to account for species-specific patterns of metabolism. We conducted a series of feeding experiments with four juvenile male brown bears (Ursus arctos) to (1) track the timing of changes in adipose tissue composition and QFASA diet estimates in response to a change in diet and (2) quantify the relationship between consumer and diet FA composition (i.e., determine "calibration coefficients"). Bears were fed three compositionally distinct diets for 90-120 days each. Two marine-based diets were intended to approximate the lipid content and composition of the wild diet of polar bears (U. maritimus). Bear adipose tissue composition changed quickly in the direction of the diet and showed evidence of stabilization after 60 days. During hibernation, FA profiles were initially stable but diet estimates after 10 weeks were sensitive to calibration coefficients. Calibration coefficients derived from the marine-based diets were broadly similar to each other and to published values from marine-fed mink (Mustela vison), which have been used as a model for free-ranging polar bears. For growing bears on a high-fat diet, the temporal window for QFASA estimates was 30-90 days. Although our results reinforce the importance of accurate calibration, the similarities across taxa and diets suggest it may be feasible to develop a generalized QFASA approach for mammalian carnivores.

Young-of-the-year (YOY) striped bass (Morone saxatilis) suffer significant mortality during their first winter. While causes of this mortality are unclear, lipids may play role in adapting to winter stresses, including thermal change and food scarcity. To address this, YOY striped bass were placed in mesh cages in freshwater ponds in the fall (November) and were held until the end of winter, in March. Liver and white muscle tissue were sampled at the beginning and end of the study to compare concentrations of specific lipid classes and fatty acid composition. Muscle-tissue total lipid and triacylglycerol (TAG) was higher in March (late winter) samples. Additionally, concentrations of phosphatidylethanolamine (PE) were higher in the white muscle of striped bass sampled in March, this was accompanied by a decrease in proportions of 18:0 and 22:6n-3 in PE (from ~11 to 7% and 36 to 28%, respectively) and 18:1n-9 and 22:6n-3 in phosphatidylcholine (from ~15 to 10% and 24 to 18%, respectively). This suggests that these fish were not utilizing energy reserves in previously described ways and appear to rely more on other lipid classes or body tissues for overwinter survival than those analyzed in this study.

Linking production, transfer and subsequent bioavailability of nutritionally significant matter from phytoplankton to higher trophic levels is a fundamental aspect in understanding marine food webs. The plant–animal interface is of interest because of the highly variable transfer between producers and consumers, and the myriad of factors that influence it. Essential fatty acids (EFAs) are dietary nutrients that are necessary for normal function in all consumers, yet it remains unclear how efficiently they are transferred through marine food webs. We introduced a 13C-labelled carbon source to the cryptophyte Rhodomonas salina to quantify primary production of two omega-3 long-chain polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). We investigated transfer and assimilation efficiencies of these EFAs from phytoplankton to the calanoid copepod Calanus finmarchicus in an 8-day feeding experiment. We found low production of both EFAs in R. salina. Assimilation efficiencies of both EFAs ranged from 5 to 15% throughout the experiment, remaining slightly higher on average for DHA. This was mirrored in more efficient trophic transfer of DHA (up to 28%, compared to 13% for EPA). These results add to previously scarce experiments empirically quantifying the assimilation and transfer efficiency of EFAs in a basic marine planktonic food chain.

Copepods are an important link in marine food webs, transferring crucial nutrients such as essential fatty acids (EFA) from primary producers to higher consumers. Yet data on lipid dynamics in marine ecosystems associated with small lipid-poor copepods are scarce. In this study, we used the coastal calanoid copepod Eurytemora herdmani, which is an opportunistic generalist feeder, to assess EFA-specific gross growth efficiency (GGEEFA), defined as the fraction of ingested EFA retained in copepod tissue. We conducted two eight-day feeding experiments with 13C-labeled phytoplankton to quantify incorporation and GGEEFA of eicosapentaenoic acid (20:5n-3; EPA) and docosahexaenoic acid (22:6n-3; DHA) in stage CV E. herdmani copepodites consuming either the EPA-replete diatom Thalassiosira weissflogii or the DHA-replete mixotrophic dinoflagellate Heterocapsa triquetra. EFA incorporation was quantified using gas chromatography. We recorded rapid shifts (within 48 h) in copepod tissue fatty acid (FA) composition following diet change, but not directly mirroring dietary algal FA. We also found high GGEEFA, with maxima of 46% and 61% for EPA and DHA, respectively, in copepods ingesting T. weissflogii and 39% and 87%, respectively, in copepods ingesting H. triquetra. GGEDHA remained higher than GGEEPA throughout the experiments regardless of dietary algae. GGEEFA decreased overall when copepods matured and produced nauplii, indicating EFA allocation for reproductive purposes. GGEDHA values in non-reproducing E. herdmani copepodites were high, suggesting that under predicted shifts in copepod community size structure, trophic transfer by small copepods could help compensate for DHA reductions resulting from lower abundances of large, lipid-rich species.

Accurate information on diet composition is central to understanding and conserving carnivore populations. Quantitative fatty acid signature analysis (QFASA) has emerged as a powerful tool for estimating the diets of predators, but ambiguities remain about the timeframe of QFASA estimates and the need to account for species-specific patterns of metabolism. We conducted a series of feeding experiments with four juvenile male brown bears (Ursus arctos) to (1) track the timing of changes in adipose tissue composition and QFASA diet estimates in response to a change in diet and (2) quantify the relationship between consumer and diet FA composition (i.e., determine "calibration coefficients"). Bears were fed three compositionally distinct diets for 90-120 days each. Two marine-based diets were intended to approximate the lipid content and composition of the wild diet of polar bears (U. maritimus). Bear adipose tissue composition changed quickly in the direction of the diet and showed evidence of stabilization after 60 days. During hibernation, FA profiles were initially stable but diet estimates after 10 weeks were sensitive to calibration coefficients. Calibration coefficients derived from the marine-based diets were broadly similar to each other and to published values from marine-fed mink (Mustela vison), which have been used as a model for free-ranging polar bears. For growing bears on a high-fat diet, the temporal window for QFASA estimates was 30-90 days. Although our results reinforce the importance of accurate calibration, the similarities across taxa and diets suggest it may be feasible to develop a generalized QFASA approach for mammalian carnivores.

Baleen whales accumulate fat reserves during the summer to sustain reproduction while fasting in the winter. The southern right whale Eubalaena australis population that calves off Península Valdés, Argentina, experienced high calf mortality events from 2003 to 2013 and poor nutritional states of mothers could be a contributing cause. Previous studies found that the population’s reproductive success is influenced by prey availability. Mothers unable to build sufficient fat reserves or feeding on prey with different nutritional value may fail to meet the demands of lactation. Milk is the only source of nutrients and energy for calves at Valdés, so their fatty acids (FAs) and stable isotopes should reflect their mother’s diet and feeding-ground locations. Here, we compared FA profiles and C and N stable isotopes of dead calves with those of living calves to evaluate the potential impact of maternal nutrition on calf survival. We found no differences in the FA composition of blubber in dead and living calves, indicating similar maternal diets. Likewise, the isotopic values of living and dead calves imply that their mothers had similar foraging ranges. However, FA composition was greatly affected by calf length, indicating effects of calf age and duration of nursing. These findings suggest that mothers of dead calves did not feed on different diets or feeding grounds compared to mothers of living calves. Future research should further assess the overall health and body condition of the Valdés southern right whale calves. Fil: Marón, Carina Flavia. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Departamento de Diversidad Biológica y Ecológica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Conservación de Ballenas; Argentina Fil: Budge, Suzanne M.. Dalhousie University Halifax; Canadá Fil: Ward, Robert E.. Utah State University; Estados Unidos Fil: Valenzuela, Luciano Oscar. Universidad Nacional del Centro de la Provincia de Buenos Aires. Facultad de Ciencias Sociales. Departamento de Arqueología. Laboratorio de Ecología Evolutiva Humana (Sede Quequén); Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil; Argentina Fil: Di Martino, Matías. Programa de Monitoreo Sanitario Ballena Franca Austral; Argentina Fil: Ricciardi, Marcos. Instituto de Conservación de Ballenas; Argentina Fil: Sironi, Mariano. Instituto de Conservación de Ballenas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Departamento de Diversidad Biológica y Ecológica; Argentina. Programa de Monitoreo Sanitario Ballena Franca Austral; Argentina Fil: Uhart, Marcela. Programa de Monitoreo Sanitario Ballena Franca Austral; Argentina. University of California; Estados Unidos Fil: Seger, Jon. University Of Utah. Department Of Biology; Estados Unidos Fil: Rowntree, Victoria J.. University Of Utah. Department Of Biology; Estados Unidos. Instituto de Conservación de Ballenas; Argentina. Programa de Monitoreo Sanitario Ballena Franca Austral; Argentina. Whale Conservation Institute/Ocean Alliance; Estados Unidos

A key aspect in the use of fatty acids (FA) to estimate predator diets using quantitative FA signature analysis (QFASA) is the ability to account for FA assimilation through the use of calibration coefficients (CC). Here, we tested the assumption that CC are independent of dietary fat concentrations by feeding Atlantic pollock (Pollachius virens) three formulated diets with very similar FA proportions but different fat concentrations (5–9% of diet) for 20 weeks. CC calculated using FA profiles of diet and triacylglycerols in pollock liver were significantly different for the three diets. To test the robustness of diet estimates to these differences, we used the CC set derived from feeding the diet with the lowest fat concentration, published prey FA profiles and realistic diet estimates of pollock to construct ‘pseudo-predators'. Application of QFASA to each pseudo-predator using the three sets of CC and the same prey FA profiles resulted in diet estimate biases of twofold for major prey items and approximately fivefold for minor prey items. This work illustrates the importance of incorporating diets with fat concentrations that are similar to natural prey when conducting feeding experiments to calculate CC. This article is part of the theme ‘The next horizons for lipids as ‘trophic biomarkers': evidence and significance of consumer modification of dietary fatty acids'.

Fatty acids are commonly used as biomarkers for making inferences about trophic relationships in aquatic and soil food webs. However, researchers are often unaware of the physiological constraints within organisms on the trophic transfer and modification of dietary biomarkers in consumers. Fatty acids are bioactive molecules, which have diverse structures and functions that both complicate and enhance their value as trophic tracers. For instance, consumers may synthesize confounding non-dietary sourced markers from precursor molecules, and environmental conditions also affect fatty acid composition. There is a vital need for more research on the uptake and transfer of trophic biomarkers in individual organisms in order to advance the field and make meaningful use of these tools at the scale of populations or ecosystems. This special issue is focused on controlled feeding experiments on a diverse taxonomic breadth of model consumers from freshwater, marine and soil ecosystems with a goal of creating a more integrated understanding of the connection between consumer physiology and trophic ecology. This article is part of the theme issue ‘The next horizons for lipids as ‘trophic biomarkers’: evidence and significance of consumer modification of dietary fatty acids'.

The essential fatty acids (EFA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are critical nutrients for all organisms, and the temperature sensitivity of their trophic transfer in marine systems is of concern because of rising ocean temperatures. Laboratory-reared copepodites of the marine calanoid Calanus finmarchicus were used to test the effects of temperature (at 6°C, 12°C and increasing temperature stress) and prey type (the dinoflagellate Heterocapsa triquetra and the diatom Thalassiosira weissflogii ) on the extent and efficiency of dietary EPA and DHA incorporation from phytoplankton to copepods in a set of feeding experiments using 13 C labelling. Temperature was a significant determinant of C . finmarchicus copepodites' EFA incorporation and gross growth efficiency, defined as the fraction of ingested EFA retained in copepod tissue. Ingestion and incorporation of both EFA were higher at warmer temperature, except in the case of DHA in copepods feeding on diatoms. DHA-associated growth efficiency was higher at the higher temperature for copepodites consuming the dinoflagellate, but temperature-related variation in algal EFA content was also a predictive factor. Moreover, our results strongly suggest that copepodites are capable of synthesizing EPA when consuming an EPA-depleted diet. Our study implies that the copepod link of marine food webs is resilient in terms of EFA transfer when confronted with alterations of ambient temperature and prey type availability. Measurements presented here are critical for estimating how EFA transfer dynamics respond to intra- and interannual environmental variability. This article is part of the theme issue ‘The next horizons for lipids as ‘trophic biomarkers’: evidence and significance of consumer modification of dietary fatty acids’.

Epoxy and hydroxy fatty acids are important intermediates during lipid oxidation; quantification of both structures may help evaluate the extent of competition among various lipid oxidation pathways. This article describes a method to simultaneously determine saturated- and unsaturated- epoxy and hydroxy fatty acids derived from oxidation of vegetable oils. The experimental procedures employed transesterification with sodium methoxide, separation of epoxy and hydroxy fatty acid methyl esters (FAME) using solid-phase extraction (SPE), and trimethylsilyl (TMS) derivatization of hydroxy groups. GC–MS was used to identify the epoxy and hydroxy FAME in two different SPE fractions, while GC-flame ionization detection (GC-FID) was used to determine their quantities. Epoxy-octadecanoate/octadecenoate and hydroxy-octadecanoate/octadecenoate/octadecadienoate were determined as lipid oxidation products generated from oxidation of sunflower and canola oils. An isomer of methyl 13-hydroxyoctadeca-9,11-dienoate (13-HODE) TMS ether co-eluted with methyl 15-hydroxyoctadeca-9,12-dienoate TMS ether, which was only present in canola oil; thus, GC–MS-selected ion monitoring (GC–MS–SIM) was used to determine the concentration of 13-HODE. The proposed method has been successfully applied to monitor epoxy and hydroxy fatty acids in sunflower oil and canola oil oxidized at 40 °C.

Knowledge of predator diets, including how diets might change through time or differ among predators, provides essential insights into their ecology. Diet estimation therefore remains an active area of research within quantitative ecology. Quantitative fatty acid signature analysis (QFASA) is an increasingly common method of diet estimation. QFASA is based on a data library of prey signatures, which are vectors of proportions summarizing the fatty acid composition of lipids, and diet is estimated as the mixture of prey signatures that most closely approximates a predator’s signature. Diets are typically estimated using proportions from a subset of all fatty acids that are known to be solely or largely influenced by diet. Given the subset of fatty acids selected, the current practice is to scale their proportions to sum to 1.0. However, scaling signature proportions has the potential to distort the structural relationships within a prey library and between predators and prey. To investigate that possibility, we compared the practice of scaling proportions with two alternatives and found that the traditional scaling can meaningfully bias diet estimators under some conditions. Two aspects of the prey types that contributed to a predator’s diet influenced the magnitude of the bias: the degree to which the sums of unscaled proportions differed among prey types and the identifiability of prey types within the prey library. We caution investigators against the routine scaling of signature proportions in QFASA.

In recent years, 1H NMR has been used to study epoxides in lipid oxidation and industrial processes, but the peak assignments reported for monoepoxides and diepoxides have been inconsistent. Lack of clear assignments for chemical shifts of epoxides derived from polyunsaturated fatty acids (PUFA) has also limited the use of 1H NMR in detecting and quantifying these products during both oxidative degradation and industrial epoxidation. In this study, 1H NMR was used to characterize the epoxides synthesized from trilinolein, trilinolenin, canola oil, and fish oils by reaction with formic acid and hydrogen peroxide. Assignments for epoxides derived from PUFA in canola oil and fish oil were between 2.90–3.23 ppm and 2.90–3.28 ppm, distinct from other chemical groups in these oils. Chemical shifts of epoxy groups moved downfield with an increasing number of epoxy groups in the fatty acid chain. Hence, peaks for diepoxides appeared at 3.00, 3.09, and 3.14 ppm and for triepoxides at 3.00, 3.16, and 3.21 ppm. Results also suggested that stereoisomers of diepoxides and triepoxides were formed during the epoxidation process under the conditions of this study. These new assignments for di- and tri-epoxide stereoisomers were supported by GC–MS analysis of their methyl esters, H–H COSY experiments, and a re-evaluation of several previous epoxide-related studies.

Understanding the diet of deep-diving predators can provide essential insight to the trophic structure of the mesopelagic ecosystem. Comprehensive population-level diet estimates are exceptionally difficult to obtain for elusive marine predators due to the logistical challenges involved in observing their feeding behavior and collecting samples for traditional stomach content or fecal analyses. We used quantitative fatty acid signature analysis (QFASA) to estimate the diet composition of a wide-ranging mesopelagic predator, the northern elephant seal (Mirounga angustirostris), across five years. To implement QFASA, we first compiled a library of prey fatty acid profiles from the mesopelagic eastern North Pacific. Given the scarcity of a priori diet data for northern elephant seals, our prey library was necessarily large to encompass the range of potential prey in their foraging habitat. However, statistical constraints limit the number of prey species that can be included in the prey library to the number of dietary fatty acids in the analysis. Exceeding that limit could produce non-unique diet estimates (i.e. multiple diet estimates fit the data equally well). Consequently, we developed a novel ad-hoc method to identify which prey were unlikely to contribute to diet and could, therefore, be excluded from the final QFASA model. The model results suggest that seals predominantly consumed small mesopelagic fishes, including myctophids (lanternfishes) and bathylagids (deep sea smelts), while non-migrating mesopelagic squids comprised a third of their diet, substantially less than suggested by previous studies. Our results revealed that mesopelagic fishes, particularly energy-rich myctophids, were a critical prey resource, refuting the long-held view that elephant seals are squid specialists.

Summary Knowledge of animal diets can provide important insights into life history and ecology, relationships among species in a community and potential response to ecosystem change or perturbation. Quantitative fatty acid signature analysis (QFASA) is a method of estimating diets from data on the composition, or signature, of fatty acids stored in adipose tissue. Given data on signatures of potential prey, a predator diet is estimated by minimizing the distance between its signature and a mixture of prey signatures. Calibration coefficients, constants derived from feeding trials, are used to account for differential metabolism of individual fatty acids. QFASA has been widely applied since its introduction and several variants of the original estimator have appeared in the literature. However, work to compare the statistical properties of QFASA estimators has been limited. One important characteristic of an estimator is its robustness to violations of model assumptions. The primary assumptions of QFASA are that prey signature data contain representatives of all prey types consumed and the calibration coefficients are known without error. We investigated the robustness of two QFASA estimators to a range of violations of these assumptions using computer simulation and recorded the resulting bias in diet estimates. We found that the Aitchison distance measure was most robust to errors in the calibration coefficients. Conversely, the Kullback–Leibler distance measure was most robust to the consumption of prey without representation in the prey signature data. In most QFASA applications, investigators will generally have some knowledge of the prey available to predators and be able to assess the completeness of prey signature data and sample additional prey as necessary. Conversely, because calibration coefficients are derived from feeding trials with captive animals and their values may be sensitive to consumer physiology and nutritional status, their applicability to free-ranging animals is difficult to establish. We therefore recommend that investigators first make any improvements to the prey signature data that seem warranted and then base estimation on the Aitchison distance measure, as it appears to minimize risk from violations of the assumption that is most difficult to verify.

A new method to determine epoxide concentrations in oxidized oils was developed and validated using (1)H NMR. Epoxides derived from lipid oxidation gave signals between 2.90 and 3.24 ppm, well separated from the signals of other lipid oxidation products. To calibrate, soybean oils with a range of epoxide concentrations were synthesized and analyzed using (1)H NMR by taking the sn-1,3 glycerol protons (4.18, 4.33 ppm) as internal references. The (1)H NMR signals were compared to the epoxide content determined by titration with hydrogen bromide (HBr)-acetic acid solution. As expected, the signal response increased with concentration linearly (R(2) = 99.96%), and validation of the method gave results comparable to those of the HBr method. A study of the oxidative stability of soybean oil was performed by applying this method to monitor epoxides during thermal lipid oxidation. The epoxide content increased over time and showed a different trend compared to peroxide value (PV). A phenomenological model was suggested to model epoxides derived from lipid oxidation.

Quantitative fatty acid signature analysis has become an important method of diet estimation in ecology, especially marine ecology. Controlled feeding trials to validate the method and estimate the calibration coefficients necessary to account for differential metabolism of individual fatty acids have been conducted with several species from diverse taxa. However, research into potential refinements of the estimation method has been limited. We compared the performance of the original method of estimating diet composition with that of five variants based on different combinations of distance measures and calibration-coefficient transformations between prey and predator fatty acid signature spaces. Fatty acid signatures of pseudopredators were constructed using known diet mixtures of two prey data sets previously used to estimate the diets of polar bears Ursus maritimus and gray seals Halichoerus grypus, and their diets were then estimated using all six variants. In addition, previously published diets of Chukchi Sea polar bears were re-estimated using all six methods. Our findings reveal that the selection of an estimation method can meaningfully influence estimates of diet composition. Among the pseudopredator results, which allowed evaluation of bias and precision, differences in estimator performance were rarely large, and no one estimator was universally preferred, although estimators based on the Aitchison distance measure tended to have modestly superior properties compared to estimators based on the Kullback-Leibler distance measure. However, greater differences were observed among estimated polar bear diets, most likely due to differential estimator sensitivity to assumption violations. Our results, particularly the polar bear example, suggest that additional research into estimator performance and model diagnostics is warranted.

Fish oil is available primarily as triacylglyc- erols (TAG) or ethyl esters (EE). Anecdotal evidence sug- gests that TAG have superior bioavailability and oxidative stability compared to EE. In this work, peroxide value (PV) and p-anisidine value (AV) were used to monitor oxidation in commercially available TAG and EE fish oils incubated at temperatures from 5 to 60 °C. Pseudo first-order kinet- ics were assumed and rate constants were calculated for each temperature. At all temperatures, the rates of oxida- tion were higher for EE oils than TAG oils. For PV and AV measured in both oils, non-linear Arrhenius models were plotted, generating activation energies that ranged from 7 to 103 and 2 to 159 kJ/mol for PV and AV, respectively. Although TAG were more resistant to oxidation than EE, they had lower activation energies (E a ) at ≤15 °C for reac- tions measured with PV and AV. The Ea for EE was nega- tive at temperatures ≥45 °C, indicating that reaction rate was influenced by factors in addition to temperature.

In many branches of lipid science, researchers are interested in non-esterified fatty acid (NEFA) content and composition. For many years, diazomethane was the reagent of choice to selectively derivatize and then detect NEFA due to its highly specific methylation of the carboxylic acid functional group. While the activity of this derivatizing reagent is very defined, it is dangerous and can be difficult to obtain. In this brief review, we have compiled a collection of methods which allow for the detection of NEFA and hydroxy NEFA without the use of diazomethane. We have concentrated on methods that employ three distinct approaches of selective quantification/extraction, purification from total lipids and derivatization techniques. Chemical reactions that occur during selective extraction of NEFA using a quaternary ammonium salt (a) and the later pyrolytic derivatization when the mixture is placed in the hot injection port of a GC (b).

We analyzed the fatty acid (FA) composition and carbon stable isotope ratios of indi- vidual FAs (δ 13 CFA) of 3 zooplankton species (Themisto libellula, Calanus marshallae/glacialis, and Thysanoessa raschii) sampled from the Bering Sea during winter maximum ice extent, spring ice melt, and summer ice-free conditions in 2009 and 2010. Our goal was to assess diets of these eco- logically important species and estimate the proportional contribution of pelagic and sympagic carbon sources to their diets. FA profiles showed little variation in diet within species between ice conditions or years but revealed differences in diet among species. FA biomarkers confirmed that T. libellula was predominately carnivorous and that C. marshallae/glacialis and T. raschii were primarily herbivorous. Estimates from 4 stable isotope mixing models using combinations of δ 13 CFA values of diatom FA markers (16:1n-7, 20:5n-3), and a flagellate FA marker (22:6n-3) showed that substantial, albeit highly variable, proportions of these FAs originated from organic matter originating from sea ice algae (T. libellula 36 to 72%, C. marshallae/glacialis 27 to 63%, and T. raschii 39 to 71%). Our results suggest that ice algae may be an important food source for zooplankton when water column phytoplankton are not available during critical periods in their life history. Predicted increases in water column phytoplankton production in the Bering Sea may help offset the expected reduction in ice algal production and any detrimental effects that this might have on consumers such as zooplankton.

Knowledge of animal diets provides essential insights into their life history and ecology, although diet estimation is challenging and remains an active area of research. Quantitative fatty acid signature analysis (QFASA) has become a popular method of estimating diet composition, especially for marine species. A primary assumption of QFASA is that constants called calibration coefficients, which account for the differential metabolism of individual fatty acids, are known. In practice, however, calibration coefficients are not known, but rather have been estimated in feeding trials with captive animals of a limited number of model species. The impossibility of verifying the accuracy of feeding trial derived calibration coefficients to estimate the diets of wild animals is a foundational problem with QFASA that has generated considerable criticism. We present a new model that allows simultaneous estimation of diet composition and calibration coefficients based only on fatty acid signature samples from wild predators and potential prey. Our model performed almost flawlessly in four tests with constructed examples, estimating both diet proportions and calibration coefficients with essentially no error. We also applied the model to data from Chukchi Sea polar bears, obtaining diet estimates that were more diverse than estimates conditioned on feeding trial calibration coefficients. Our model avoids bias in diet estimates caused by conditioning on inaccurate calibration coefficients, invalidates the primary criticism of QFASA, eliminates the need to conduct feeding trials solely for diet estimation, and consequently expands the utility of fatty acid data to investigate aspects of ecology linked to animal diets.

Lipid oxidation can lead to flavor and safety issues in fat-containing foods. In order to measure the extent of lipid oxidation, hydroperoxides and their scission products are normally targeted for analytical purposes. In recent years, the formation of rarely monitored oxygenated products, including epoxides, alcohols, and ketones, has also raised concerns. These products are thought to form from alternative pathways that compete with chain scissions, and should not be neglected. In this review, a number of instrumental techniques and approaches to determine epoxides, alcohols, and ketones are discussed, with a focus on their selectivity and sensitivity in applications to food lipids and oils. Special attention is given to methods employing gas chromatography (GC), high-performance liquid chromatography (HPLC), and nuclear magnetic resonance (NMR). For characterization purposes, GC-mass spectrometry (GC-MS) provides valuable information regarding the structures of individual oxygenated fatty acids, typically as methyl esters, isolated from oxygenated triacylglycerols (TAGs), while the use of liquid chromatography-MS (LC-MS) techniques allows analysis of intact oxygenated TAGs and offers information about the position of the oxygenated acyl chain on the glycerol backbone. For quantitative purposes, traditional chromatography methods have exhibited excellent sensitivity, while spectroscopic methods, including NMR, are superior to chromatography for their rapid analytical cycles. Future studies should focus on the development of a routine quantitative method that is both selective and sensitive.

BACKGROUND Heightened awareness of the health benefits of fish oil consumption has led to a great increase in the number of fish oil supplements available to the consumer. Therefore manufacturers are continually looking for ways to distinguish their products from those of competitors. Minimally refined or virgin fish oils provide a unique feature; however, petroleum hydrocarbon contamination from oil spills is a reality in the world's oceans. The question arises whether oil produced from fish species caught in these polluted areas is free of petroleum hydrocarbons, with particular interest in unresolved complex mixtures (UCMs). This study investigates the presence of UCMs in commercially available fish oil supplements advertised as being virgin, as well as refined. RESULTS Weathered petroleum hydrocarbons in the form of a UCM were found at 523 µg g−1 in a virgin Alaskan salmon oil supplement. Supplements that were refined were free of this contamination. CONCLUSION Fish used in the production of fish oil supplements appear to have accumulated petrogenic hydrocarbons in their tissues which were not removed by minimal oil refining. Further study is required to determine if there are any health implications associated with long-term consumption of these contaminated supplements. © 2014 Society of Chemical Industry

Juvenile polar cod (Boreogadus saida) are often found in close association with sea ice and represent an important trophic link in the Arctic food web. However, the proportional contribution of sea ice algal production via the sympagic food web to the diet of polar cod is unknown. To estimate the proportional contribution of fatty acids (FAs) from sea ice-derived particulate organic matter (i-POM) to the diet of juvenile polar cod, we used FA profiling and compound-specific stable carbon isotope analysis of individual FAs from juvenile polar cod collected from three regions in the Beaufort Sea. The δ13C values of the FAs 14:0, 16:4n-1, 18:0, 20:5n-3 and 22:6n-3 in the polar cod were found to most strongly resemble pelagic POM rather than i-POM. Results from isotope-mixing models using diatom FA markers indicated that the proportional contribution of FAs from i-POM to juvenile polar cod was ≤2 %, which suggests that juvenile polar cod had not sourced their FAs from i-POM. Thus, changes in sea ice coverage due to environmental change may not affect juvenile polar cod in regard to nutrients such as FAs but may still affect their populations by reducing critical shelter from predators.

Phocids routinely fast for extended periods. During these fasts, energetic requirements are met primarily through the catabolism of blubber lipid. To assess whether fatty acid (FA) composition changes during the postweaning fast in northern elephant seals, blubber biopsies were acquired longitudinally from 43 pups at 2.3 ± 1.5 and 55.2 ± 3.7 days postweaning in 1999 and 2000. At weaning, short-chain monounsaturated FA (SC-MUFA, ≤18 carbons) dominated the blubber while saturated FA (SFA) were found in the next highest proportion. The major FA (all ≥1 % by mass) comprised approximately 91 % of total blubber FA. In both years, 18:1n-9 and 16:0 were the most prevalent FA. Major FA mobilized during the fast consisted of polyunsaturated FA (PUFA), SFA, and SC-MUFA. Long-chain MUFA (>18 carbons) tended to be conserved. The fractional mobilization value of 20:5n-3 was the highest, resulting in significant reductions of this PUFA. Although concentrations of some blubber FA changed significantly during the postweaning fast, the general FA signature of blubber was similar at weaning and near the end of the fast. Changes in some FA differed across years. For example, the concentration of 20:4n-6, a minor PUFA, was significantly reduced in 1999 but not in 2000. FA mobilization patterns in northern elephant seal pups are somewhat similar to those reported previously for other fasting phocids and terrestrial mammals, though there are some notable differences. Differences in FA mobilization patterns across mammalian species may be related to differences in diets, geographical distribution, environmental factors, physiological adaptations, and life history stage.

Despite the ubiquitous distribution of oxylipins in plants, animals, and microbes, and the application of numerous analytical techniques to study these molecules, 3-OH oxylipins have never been quantitatively assayed in yeasts. The formation of heptafluorobutyrate methyl ester derivatives and subsequent analysis with gas chromatography – negative chemical ionization – mass spectrometry allowed for the first determination of yeast 3-OH oxylipins. The concentration of 3-OH 10:0 (0.68–4.82 ng/mg dry cell mass) in the SMA strain of Saccharomyces pastorianus grown in laboratory-scale beverage fermentations was elevated relative to oxylipin concentrations in plant tissues and macroalgae. In fermenting yeasts, the onset of 3-OH oxylipin formation has been related to fermentation progression and flocculation initiation. When the SMA strain was grown in laboratory-scale fermentations, the maximal sugar consumption rate preceded the lowest concentration of 3-OH 10:0 by ∼4.5 h and a distinct increase in 3-OH 10:0 concentration by ∼16.5 h.

Climate-mediated changes in the phenology of Arctic sea ice and primary production may alter benthic food webs that sustain populations of Pacific walruses (Odobenus rosmarus divergens) and bearded seals (Erignathus barbatus). Interspecific resource competition could place an additional strain on ice-associated marine mammals already facing loss of sea ice habitat. Using fatty acid (FA) profiles, FA trophic markers, and FA stable carbon isotope analyses, we found that walruses and bearded seals partitioned food resources in 2009-2011. Interspecific differences in FA profiles were largely driven by variation in non-methylene FAs, which are markers of benthic invertebrate prey taxa, indicating varying consumption of specific benthic prey. We used Bayesian multi-source FA stable isotope mixing models to estimate the proportional contributions of particulate organic matter (POM) from sympagic (ice algal), pelagic, and benthic sources to these apex predators. Proportional contributions of FAs to walruses and bearded seals from benthic POM sources were high [44 (17-67)% and 62 (38-83)%, respectively] relative to other sources of POM. Walruses also obtained considerable contributions of FAs from pelagic POM sources [51 (32-73)%]. Comparison of δ13C values of algal FAs from walruses and bearded seals to those from benthic prey from different feeding groups from the Chukchi and Bering seas revealed that different trophic pathways sustained walruses and bearded seals. Our findings suggest that (1) resource partitioning may mitigate interspecific competition, and (2) climate change impacts on Arctic food webs may elicit species-specific responses in these high trophic level consumers.

Carnivora includes three independent evolutionary transitions to the marine environment: pinnipeds (seals, sea lions, and walruses), sea otters, and polar bears. Among these, only the pinnipeds have retained two forms of insulation, an external fur layer and an internal blubber layer for keeping warm in water. In this study we investigated key factors associated with the transition to the use of blubber, by comparing blubber characteristics among the pinnipeds. Characteristics included gross morphology (blubber thickness), fat composition (fatty acid profiles, percentage lipid, and water), and thermal conductivity. Sea lions, phocids, and walrus, which have lower fur densities than fur seals, have thicker blubber layers than fur seals (P

This study was performed to evaluate the stability of the fatty acid (FA) composition of blubber from Baltic grey seals Halichoerus grypus archived at −25°C in the Swedish Environmental Specimen Bank (SESB). Possible alterations of FA compositions with storage time and temperature were evaluated. Blubber from 5 seals was kept at +2°C and at −20°C and sampled for up to 6 mo and 33 mo, respectively. Furthermore, FA profile stability was evaluated in blubber stored in the SESB for 4 and 6 yr. This was done by re-sampling and re-analyzing blubber samples from SESB. In both parts of the study, a principal component analysis was performed on 48 FAs comprising 98 to 99% of the total FAs in blubber of Baltic grey seals. We found no indication of a critically altered FA composition, neither in blubber stored at +2°C for 6 mo nor in blubber stored at −20°C for 33 mo. The re-analysis of samples stored in SESB for ~4 to 6 yr only differed for a few FAs in some samples. The most remarkable differences were a slight increase in some long-chain unsaturated FAs (18:2n-6, 20:3n-3, 20:4n-3, 22:4n-3). As an increase in long-chain polyunsaturated FAs due to storage is highly unlikely, we believe that this is probably more due to the sampling technique than to a critically altered FA composition due to storage. Our results suggest that archived seal blubber may be safely used in retrospective analyses of diet using an FA signature analysis technique.