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Petroleum is one of the most important pollutants in the marine ecosystem, and oil spills have a long-term effect on the marine environment. After an oil spill, petroleum continues to diffuse, flowing to the intertidal zone where it accumulates. Ulva pertusa is the main macroalgae species in the intertidal zone of the ocean, and petroleum is toxic to marine macroalgae. Fatty acids, which are involved in various physiological activities, including metabolism, are indispensable substances in organisms. Thus, an organism's fatty acid composition reflects its physiological state. Because fatty acids are carbon-rich compounds, their stable carbon isotope composition can be measured to assess the state of the organism. In this study, the effects of the water accommodated fraction (WAF) of different concentrations of 180# fuel oil and 0# diesel oil on the chlorophyll a and fatty acid carbon stable isotope composition (δ13CFAs) of U. pertusa were assessed. The fatty acid carbon stable isotope value varied among the different petroleum treatments, and high WAF concentrations significantly inhibited the synthesis of unsaturated fatty acids. Thus, exposure to petroleum products affected the synthesis of fatty acids in U. pertusa and also caused carbon stable isotope fractionation during the fatty acid synthesis process. The results indicate that the saturated fatty acid δ13C16:0 and the unsaturated fatty acid δ13C18:1 can be used as biomarkers to evaluate the degree of petroleum-induced stress in U. pertusa and that carbon stable isotope analysis of U. pertusa can be used as an environmental risk assessment tool in the intertidal zone.

Marine microalgae, the most important primary producers in marine ecosystems, are susceptible to toxicants and usually selected as the promising organisms for chemical risk testing. The stable carbon isotope ratios (δ13C) of amino acids (AAs) were measured to determine if compound-specific stable isotope analysis could be used to provide insight into the toxic effects of oil spill on the marine microalgae. The water accommodated fractions (WAF) of #180 fuel oil were selected as the toxic targets with different concentrations in acute (96 h) toxic tests. Naphthalene, phenanthrene, anthracene, and fluoranthene were detected as the predominant congeners of polycyclic aromatic hydrocarbons in the WAF, and may be the main toxic components. At the same WAF concentration, the δ13C values of leucine in Nitzschia closterium increased with the culture time, whereas decreased in Heterosigma akashiwo. However, with the increasing WAF concentrations, the δ13C values of glutamic acid exhibited the increasing trend in both of the two kinds of microalgae at the same culture time. The similar phenomenon was also observed for histidine in H. akashiwo, while opposite in N. closterium, but both enriching in δ13C compared to the controlled experiments without the WAF. Under the different culture times and WAF concentrations, the δ13C values of alanine and methionine showed unexpected trend. These findings indicated that the synthesis pathways of AA in microalgae cells were affected by the WAF, thus leading to the carbon isotopic fractionation in AAs. The present study provide a novel approach to explore the toxicity mechanism of primary producers under oil spill stress, and monitor and evaluate the marine ecological risk of oil spill pollution.

Heterosigma akashiwo is an algal blooms species, and thus selected as the target microalgae in this work. This study attempted to investigate the influence of carbon limitation on the growth of H. akashiwo. Experiments were carried out in CO2-unlimited and CO2-limited systems (both include three nutritional groups). The stable isotope signatures and compound-specific stable three nutritional groups carbon isotopic composition of fatty acids were measured. Here we hypothesized that the carbon limitation could lead to the enriched of stable isotope ratios in the H. akashiwo. The results showed that carbon limitation made δ13C and δ13CFAs values more and more enriched in H. akashiwo. δ13C values were enriched in normal group of H. akashiwo within CO2-limited. δ15N values were enriched in nitrogen deficiency of H. akashiwo within both CO2-unlimited and CO2-limited. Furthermore, compared with the exponential phase, the enriched in δ13C was detected during the stationary phase in H. akashiwo within CO2-limited. A total of 8 major FAs were detected in H. akashiwo. Within CO2-unlimited, nitrogen deficiency promoted the synthesis of 4 FAs (14:1n-5c, 16:0, 18:0 and 18:3n-6c) in exponential phase. Within CO2-limited, nitrogen deficiency promoted the synthesis of FAs 14:0, 16:0 and 18:3n-3c, while phosphorus deficiency promoted the synthesis of all 6 FAs in exponential phase. δ13CFAs of H. akashiwo within CO2-limited showed the valley values and were significantly greater than those within CO2-unlimited. Furthermore, δ13CFAs of stationary phase were greater than those of exponential phase. δ13C14:1n-5c in nitrogen deficiency were the highest of the three nutrient groups within CO2-unlimited. This may point out that δ13CFAs could become an indicator of marine phytoplankton blooms. Overall, the present study may provide a novel approach to investigate the physiology and lipid metabolism of H. akashiwo blooms by using stable isotope ratios coupled with FAs profiles.