Determination of trophic levels of marine fish in the Yellow Sea and northern East China Sea using nitrogen stable isotope (δ^(15)N) analysis of otoliths

Fish otolithδ^(15) N(δ^(15) N_(oto))is a demonstrated source of information of dietary history for marine fi sh as it is available iN_(oto)lith archives and sedimentary deposits unlike white muscle tissue(WMT).WMT and stomach content data are insufficient for trophic level(TL)data of past fi shes which is important for the changes of marine fi shery resources over long time scales.To determine the correlation betweenδ^(15) N_(oto) and fi sh WMTδ^(15) N(δ^(15) N_(wmt))and the feasibility of usingδ^(15) N_(oto) in characterizing the TLs of marine fi shes,we conducted nitrogen stable isotope analysis(SIA)in the otolith and WMT of 36 marine fi sh species sampled from the Yellow Sea and northern East China Sea in 2011-2014.Bothδ^(15) N_(oto) andδ^(15) N_(wmt) were analyzed using an elemental analyzer coupled with an isotope ratio mass spectrometer(EA-IRMS).Multiple otoliths were combined to make each otolith measurement and were analyzed as-is without a carbonate dissolution pre-processing step.δ^(15) N_(oto) andδ^(15) N_(wmt) comparisons for species in the Yellow Sea and northern East China Sea are currently lacking and would be helpful for both regional studies and for increasing the number of species for whichδ^(15) N_(oto) andδ^(15) N_(wmt) have been compared.Additionally,to determine the relative accuracy of trophic level calculated usingδ^(15) N_(oto),we compared TL calculated fromδ^(15) N_(oto) to traditional trophic level metrics calculated usingδ^(15) N_(wmt).The results showed a positive and highly signifi cant correlation(R=0.780,P<0.001)betweenδ^(15) N_(oto) andδ^(15) N_(wmt).Trophic level estimation using WMT(TL wmt)and otolith(TL oto)showed congruence in our study,which is not entirely surprising given thatδ^(15) N_(oto) was regressed againstδ^(15) N_(wmt) and the resulting regression coefficient was used to convertδ^(15) N_(oto) toδ^(15) N_(wmt) prior to calculating TL oto.This conversion was required in order to be consistent with previousδ^(15) N_(wmt)-based calculations of TL for comparison.TL oto calculations resulted in TL values that were largely within 5%-10%of TL values calculated withδ^(15) N_(wmt).Our fi ndings show thatδ^(15) N_(oto) is a feasible technique for characterizing the TLs of marine fi sh and can also assist in food web and marine ecosystem studies.

Stable Isotope Dilution Analysis of Gibberellin Residues in Tomato Paste by Liquid Chromatography-Tandem Mass Spectrometry

An accurate and sensitive method for the simultaneous determination of gibberellic acid（GA3）, gibberellin A4（GA4） and gibberellin A7（GA7） residues in tomato paste was developed by coupling solid phase extraction to high performance liquid chromatography-tandem mass spectrometry（LC-MS/MS） with electrospray ionization based stable isotope dilution analysis（SIDA）. The isotope labeled internal standard can compensate for the losses during the extraction and cleanup steps and for discrimination due to ion suppression. After extraction from methanol, hydrophile lipophilic balance（HLB） solid phase extraction（SPE） column was tested for the capacity of the cleanup of the tomato paste in compared with C18 SPE column which is the common way to the detection of GAs, and the former gained better result. Spiked experiments were performed in the non-contaminated tomato pastes and the recoveries of GA3, GA4 and GA7 were 42.6%―75.0% in external standard method（ESM） and 91.1%―103.8% in internal standard method（ISM） respectively. The validities of this method were investigated and good analytical performance for the three GAs was obtained, including low limits of method detection（2 ng/g for GA3 and GA4, 0.3 ng/g for GA7）, excellent linear dynamic ranges（5―500 ng/g for GA3 and GA4, 1―100 ng/g for GA7） and good relative standard deviation ranges（4.8%―9.4% for the intra-day test and 3.5%―11.9% for the inter-day test）.

Trophic ecology of small yellow croaker (Larimichthys polyactis Bleeker): stable carbon and nitrogen isotope evidence

The trophic ecology of the small yellow croaker （Larimichthys polyactis） was studied using stable isotope analyses. Samples were collected from July to September 2009 and 34 individuals from eight sites were examined for stable carbon and nitrogen isotopes. Stable carbon isotope ratios （~3C） ranged from -20.67 to -15.43, while stable nitrogen isotope ratios （~SN） ranged 9.18-12.23. The relationship between fi^3C and ~SN suggested high resource partitioning in the sampling area. Significant differences in stable isotope values among the eight sampling sites may be linked to environmental diversities involving various physical processes （such as ocean current, wind and tide） and different carbon sources. Furthermore, the stable isotope ratios may also explain the ontogenetic variability in diet and feeding, because δ13C and δ15N varied significantly with increasing body size. The findings are consistent with other studies on diet analyses in small yellow croaker. It was also demonstrated that stable isotope analysis could be used to estimate the trophic characters of small yellow croaker in feeding patterns and migrating habits.

Trophic Ecology of Sharks in the Mid-East Pacific Ocean Inferred from Stable Isotopes

As apex predators, sharks are of ecological and conservation importance in marine ecosystems. In this study, trophic positions of sharks were estimated using stable isotope ratios of carbon and nitrogen for five representative species caught by the Chinese longline fleet in the mid-east Pacific, i.e., the blue shark （Prionace glauca）, the bigeye thresher shark （Alopias superciliosus）, the silky shark （Carcharhinus falciformis）, the scalloped hammerhead （Sphyrna lewini）, and the oceanic whitetip shark （Car-charhinus longimanus）. Of these species, oceanic whitetip shark has the lowest trophic level and mean 815N value （3.9 and 14.93%o± 0.84%o）, whereas bigeye thresher shark has the highest level/values （4.5 and 17.02%o±1.21%o, respectively）. The bigeye thresher shark has significantly higher 515N value than other shark species, indicating its higher trophic position. The blue shark and oceanic whitetip shark has significantly higher 813C values than bigeye thresher shark, silky shark and scalloped hammerhead, possibly due to different diets and/or living habitats. The stable isotope data and stomach content data are highly consistent, suggesting that sta-ble isotope analysis supplements traditional feeding ecology study of sharks, and thus contributes to understanding their trophic linkage.

Effects of pretreatment procedures on fatty acid composition and stable carbon isotopes in the marine microalga Isochrysis zhanjiangenisis

This study aims to quantify the effects of different pretreatment methods on the stable carbon isotope values of fatty acids in marine microalgae（Isochrysis zhanjiangenisis）.To identify the effects of sample preparation on theδ^（13）C value and the fatty acid composition,we examined eight types of pretreatment methods including：（a） drying the sample followed by direct methyl esterification using HCl-CH3OH;（b） drying the sample followed by direct methyl esterification using H2SO4-CH3OH;（c） drying the sample by ultrasonic extraction and methylesterification using HCl-CH3OH;（d） drying the sample by ultrasonic extraction and methyl-esterification using H2SO4-CH3OH;（e） fresh sample followed by direct methyl-esterification using HCl-CH3OH;（f） fresh sample followed by direct methyl-esterification using H2SO4-CH3OH;（g） fresh sample with ultrasonic extraction followed by methyl-esterification using HCl-CH3OH,and（h） fresh sample with ultrasonic extraction followed by methylesterification using H2SO4-CH3 OH.The results show that the δ^（13）C values from Groups a-e,g and h fluctuated within 0.3‰,and the δ^（13）C values of Group f were approximately 0.7‰ lower than the other seven groups.Therefore,the different sample pretreatment methods used towards the extraction of fatty acids from marine microalgae may result in different results regarding the stable carbon isotope ratios,and if necessary a correction should be applied.

庙岛群岛毗邻海域秋季底栖食物网潜在碳来源贡献及对碳汇渔业的思考

陆架边缘海是全球海洋重要的碳汇区域,而近岸岛屿毗邻海域作为最具代表性的边缘海域,具有来自海洋与陆地的不同碳来源,在碳循环和海洋碳汇中发挥着重要作用。本研究基于碳氮稳定同位素方法,利用贝叶斯混合模型,分析2020年秋季庙岛群岛毗邻海域底栖食物网中不同碳来源(浮游植物、大型藻类、悬浮颗粒有机物和底质有机物)对主要消费者类群(水生底栖无脊椎动物、杂食性鱼类、底栖食性鱼类和肉食性鱼类)的相对贡献,并以此对海洋碳汇和碳汇渔业展开了讨论。结果显示,庙岛群岛毗邻海域秋季底栖食物网生物类群的潜在碳来源主要为藻类(包括浮游植物和大型藻类)和海底底质有机质(SOM),但悬浮颗粒有机物(POM)的贡献较低。碳来源主要以内源性碳(海源)为主。由于大型藻类对水生底栖无脊椎动物的贡献明显高于鱼类,在碳汇渔业的指导下,合理科学地进行藻类增养殖,并通过贝类或以贝藻为食的鱼类混和养殖可能增加碳汇能力,从而促进碳汇渔业的发展。

C and N stable isotope analysis of human and animal bones at the Beiqian site

The Beiqian site,located in Jimo city,Qingdao,went through three archaeological excavations,and a large number of human and animal bones in the Dawenkou culture and the Zhou Dynasty period were unearthed.By carbon and nitrogen stable isotope analysis,the diet of ancestors is expected to be recovered,and the domesticated plants and animals will be discussed.The investigated results showed that the food structure of ancestors in both periods included food crops,mainly C4 foods,and meat,mainly marine shellfish and livestock.For domestic pigs,they mainly eat C3 plants,and were affected by the lifestyle of ancestors.This study also showed that the diet of ancestors is the shellfish rather than marine fish despite the Beiqian site is located near the shoreline.We can conclude that the diet of ancestors in Dawenkou culture period included millet crops(C4plants),shellfish,marine food,and livestock,representing the lifestyle of farming,domestication,and fishing,supplemented by hunting.In the Zhou dynasty period the meat resource was enriched by domesticated animals and marine fisheries,and the living conditions were significantly improved.

Spatial and seasonal variations in the trophic spectrum of demersal fish assemblages in Jiaozhou Bay, China

Trophic structure of fish communities is fundamental for ecosystem-based fisheries management, and trophic spectrum classifies fishes by their positions in food web, which provides a simple summary on the trophic structure and ecosystem function. In this study, both fish biomass and abundance trophic spectra were constructed to study the spatial and seasonal variations in the trophic structure of demersal fish assemblages in Jiaozhou Bay, China. Data were collected from four seasonal bottom trawl surveys in Jiaozhou Bay from February to November in 2011. Trophic levels(TLs) of fishes were determined by nitrogen stable isotope analysis. This study indicated that most of these trophic spectra had a single peak at trophic level(TL) of 3.4–3.7, suggesting that demersal fish assemblages of Jiaozhou Bay were dominated by secondary consumers(eg. Pholis fangi and Amblychaeturichthys hexanema). The spatial and seasonal variations of trophic spectra of Jiaozhou Bay reflected the influence of fish reproduction, fishing pressure and migration of fishes. Two-way analysis of variance(ANOVA) showed that seasonal variations in trophic spectra in Jiaozhou Bay were significant(P <0.05), but variations among different areas were not significant( P >0.05). The trophic spectrum has been proved to be a useful tool to monitor the trophic structure of fish assemblages. This study highlighted the comprehensive application of fish biomass and abundance trophic spectra in the study on trophic structure of fish assemblages.

The importance of terrestrial carbon in supporting molluscs in the wetlands of Poyang Lake

Allochthonous organic matter plays an important role in nutrient cycling and energy mobilization in freshwater ecosystems. However, the subsidies of this carbon source in floodplain ecosystems have not yet well understood. We used a Bayesian mixing model and stable isotopes （613C and 815N） of primary food resources and dominant molluscs species, to estimate the relative importance of allochthonous carbon sources for consumers in a representative sub-lake of Poyang Lake during a prolonged dry season. Our study inferred that terrestrial-derived carbon from Carex spp. could be the primary contributor to snails and mussels in Dahuchi Lake. The mean percentage of allochthonous food resources accounted for 35%- 50% of the C incorporated by these consumers. Seston was another important energy sources for benthic consumers. However, during the winter and low water-level period, benthic algae and submerged vegetation contributed less carbon to benthic consumers. Our data highlighted the importance of terrestrial organic carbon to benthic consumers in the wetlands of Poyang Lake during the prolonged dry period. Further, our results provided a perspective that linkages between terrestrial and aquatic ecosystems might be facilitated by wintering geese via their droppings.

Recent applications of isotope analysis to forensic anthropology

Isotope analysis has become an increasingly valuable tool in forensic anthropology case-work over the past decade. Modern-day isotopic investigations on human remains have integrated the use of multi-isotope profiles (e.g. C, N, O, H, S, Sr, and Pb) as well as iso-topic landscapes ("isoscapes") from multiple body tissues (e.g. teeth, bone, hair, and nails) to predict possible region-of-origin of unidentified human remains. Together, data from various isotope analyses provide additional lines of evidence for human identification, including a decedent's possible region-of-birth, long-term adult residence, recent travel history, and dietary choices. Here, we present the basic principles of isotope analysis and provide a brief overview of instrumentation, analytical standards, sample selection, and sample quality measures. Finally, we present case studies that reflect the diverse applications of isotope analysis to the medicolegal system before describing some future research directions. As shown herein, isotope analysis is a flexible and powerful geolocation tool that can provide new investigative leads for unidentified human remains cases.

Extraction of soluble collagen and its feasibility in the palaeodietary research

In current palaeodietary research, gelatinization is the main method to extract insoluble collagen （ISC） from ancient bones. However, the degradation products of ISC, i.e., soluble collagen （SC）, is often neglected and abandoned. In this work, we try to separate the extracts of ancient bones using gel chromatography and compare the contents of carbon and nitrogen, atomic C/N ratio, and stable carbon and nitrogen isotopic values of the extracts from three peaks to determine which peak can be at- tributed to SC. At last, the potential application of SC in palaeodietary research is discussed based on the comparison of stable isotopic values between ISC and SC. Among the three peaks, the second with the retention time between 17.5 min and 27.5 min had the most broad peak shape, indicating that the molecular weights of proteins collected were most variable. Besides, the contents of carbon and nitrogen and atomic C/N ratio of extracts in this peak were closest to the corresponding ISC. Based on the above, we conclude that the extract in second peak is SC. More important, the δ13c and δ15N values of ISC and SC are very similar. For ISC and SC with atomic C/N ratios within the normal range （2.9-3.6）, the mean difference of δ13C value was only （0.3±0.2）%o （n=2） while δ15N value was （0.6±0.1）‰ （n=2）. Although the atomic C/N ratios of some SC are slightly be- yond the normal range, the mean differences of δ13C and δ15N values were still only （0.4±0.1）‰and （0.3±0）‰ （n=2） respec- tively. These isotopic differences are quite below the isotope fractionation in one trophic level δ13C values of 1‰-1.5‰ and 615N values of 3‰-5‰）, suggesting that SC had great application potentials in palaeodietary research.

Identification of a female murder victim found in Burgenland,Austria in 1993

In 1993,the skeletal remains of a female corpse were found in Burgenland,Austria.Initial identification of the approximately 25–35-year-old female appeared impossible,but the case was reopened 23 years later.By applying biogeochemical isotope methods to her body tissues,the geographical origin of the unknown corpse could be predicted.The results of the C,N,S,H,Sr,and Pb isotope analyses suggested that the female did not originate from Europe and most likely spent her youth in the northern Caribbean.Using these findings,the police were able to identify the woman within 2 weeks.The female came from the Dominican Republic and resided in Austria for only a short period before she was murdered.This case shows that isotope biogeochemistry investigations can provide the police with crucial information that enables unknown persons to be identified.