Efficient acid hydrolysis for compound-specific δ15N analysis of amino acids for determining trophic positions.

Compound-specific isotope analysis of nitrogen in amino acids (CSIA-AA, δ15NAA) has gained increasing popularity for elucidating energy flow within food chains and determining the trophic positions of various organisms. However, there is a lack of research on the impact of hydrolysis conditions, such as HCl concentration and hydrolysis time, on δ15NAA analysis in biota samples. In this study, we investigated two HCl concentrations (6 M and 12 M) and four hydrolysis times (2 h, 6 h, 12 h, and 24 h) for hydrolyzing and derivatizing AAs in reference materials (Tuna) and biological samples of little egret (n = 4), night heron (n = 4), sharpbelly (n = 4) and Algae (n = 1) using the n-pivaloyl-iso-propyl (NPIP) ester approach. A Dowex cation exchange resin was used to purify amino acids before derivatization. We then determined δ15NAA values using gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). The results revealed no significant differences (p > 0.05) in δ15NAA values among samples treated with different HCl concentrations or hydrolysis times, particularly for δ15NGlx (range: 21.0-23.5‰) and δ15NPhe (range: 4.3-5.4‰) in Tuna (12 M). Trophic positions (TPs) calculated based on δ15NAA at 2 h (little egret: 2.9 ± 0.1, night heron: 2.8 ± 0.1, sharpbelly: 2.0 ± 0.1 and Algae: 1.3 ± 0.2) were consistent with those at 24 h (3.1 ± 0.1, 2.8 ± 0.1, 2.2 ± 0.1 and 1.1 ± 0.1, respectively), suggesting that a 2-h hydrolysis time and a 6 M HCl concentration are efficient pretreatment conditions for determining δ15NAA and estimating TP. Compared to the currently used hydrolysis conditions (24 h, 6 M), the proposed conditions (2 h, 6 M) accelerated the δ15NAA assay, making it faster, more convenient, and more efficient. Further research is needed to simplify the operational processes and reduce the time costs, enabling more efficient applications of CSIA-AA.

Using live videography observation and Bayesian isotope mixing model to identify food composition and dietary contribution to inorganic mercury and methylmercury intake by songbird nestlings.

Mercury (Hg) exposure is increasing in terrestrial birds; however, studies on its sources are scarce. In the present study, we elucidated the food composition of green-backed tit nestlings from three urban forest parks (CPL, AHL, and LCG) using live videography observation (LVO). Furthermore, the daily dietary intakes of inorganic Hg (IHg) (MDIIHg) and methylmercury (MeHg) (MDIMeHg) were determined using the Bayesian isotope mixing model (BIMM) to uncover the nestlings' specific dietary Hg contribution. Both LVO and BIMM indicated that Lepidoptera (primarily caterpillar) constituted the primary food source for the nestlings in the three forests, accounting for approximately 60% of their diet in all three forest parks. The estimated MDI of Hg revealed that lepidopterans and spiders primarily contributed to IHg exposure, with a co-contribution ratio of 71.8%-97.7%. Unexpectedly, dietary MeHg was mostly derived from spiders; the highest contribution ratio of 93.6% was recorded at CPL, followed by another peak ratio of 92.9% at LCG. However, the dietary exposure was primarily IHg, accounting for 69.8% (AHL), 62.0% (LCG), and 61.3% (CPL) of the nestlings. Our study findings highlight the importance of dietary IHg transfer in evaluating the effects of Hg in nestlings. LVO, coupled with BIMM, is an effective tool for determining the food compositions of songbird nestlings and estimating the contribution of specific diets.

Enhanced Mercury Accumulation in Riparian Spiders: An Evidence of Insects' Emergence Effect in Aquatic and Upland Terrestrial Crossed Habitat.

Many studies have focused on mercury (Hg) accumulation in both aquatic and terrestrial organisms, but the effects of aquatic Hg on terrestrial organisms have rarely been documented. Here we report the accumulation of Hg in two species of spiders, Argiope bruennichi, inhabiting paddy fields, and Nephila clavata, inhabiting small forests in the riparian zones of two hydroelectric reservoirs in Guiyang, southwest China. The mean concentration of total mercury (THg) was higher in N. clavata (0.38 mg kg-1) than in A. bruennichi (0.20 mg kg-1). The monthly average THg in N. clavata, collected consecutively from May to October, and the highest values for THg in June (1.2 mg kg-1) could be related to the emergence of aquatic insects during early summer, suggesting that emerging insects play a crucial role in the accumulation of Hg in riparian spiders. The high values could also be attributable to the different times of spider sampling or individual differences.

Distributions of Total Mercury and Methylmercury in Dragonflies from a Large, Abandoned Mercury Mining Region in China.

Dragonflies (Order Odonata) often are considered to be biosentinels of environmental contamination, e.g., heavy metals and/or persistent organic pollutants (POPs). Dragonflies (n = 439) belonging to 15 species of 8 genera were collected from an abandoned mercury (Hg) mining region in China to investigate the bioaccumulation of total Hg (THg) and methylmercury (MeHg). THg and MeHg concentrations in dragonflies varied widely within ranges of 0.06-19 mg/kg (average: 1.5 ± 2.2 mg/kg) and 0.02-5.7 mg/kg (average: 0.75 ± 0.65 mg/kg), respectively. THg and MeHg were positively correlated with bodyweight (THg: r2 = 0.10, P = 0.000; MeHg: r2 = 0.09, P = 0.000). Significant variations were observed among species, with the highest MeHg value (in Orthetrum triangulare) was fivefold higher than the lowest (in Pantala flavescens). These variations were consistent with those of nitrogen isotope (δ15N) values, indicating that increased δ15N, i.e., trophic levels, may reflect increased exposure and uptake of biomagnifying MeHg in dragonflies. A toxicological risk assessment found hazard quotients for specialist dragonfly-consuming birds of up to 7.2, which is 2.4 times greater than the permissible limit of 3, suggesting a potential toxicological risk of exposure.

Compound-specific nitrogen isotope of amino acids: Toward an improved understanding of mercury trophic transfer in different habitats.

Herein, we investigated the trophic transfer of mercury (Hg) through food chains in different habitats (namely aquatic, riparian, and terrestrial) through bulk stable isotope analysis of nitrogen (δ15Nbulk) and compound-specific isotope analysis of nitrogen in amino acids (δ15NAA) using bird feathers and their potential food sources from a Hg-contaminated site in southwest China. Results showed similar δ15Nphe for water birds (4.7 ± 2.6 ‰) and aquatic food sources (5.2 ± 2.1 ‰) and for land-based food sources (10.1 ± 0.4 ‰) and terrestrial birds (11.6 ± 3.0 ‰), verifying δ15Nphe as a potential discriminant indicator for different food sources. The trophic positions (TPs) of most organisms based on δ15Nbulk (TPbulk) tended to overestimate compared with those based on δ15NAA (TPAA), especially for predators (such as kingfisher: ΔTP = 1.3). Additionally, significant differences were observed in the aquatic, riparian, and terrestrial food webs between trophic magnification slope (TMS)bulk and TMSAA (p < 0.05). The trophic magnification factor (TMF)AA-multiple based on multiple-AAs in three food webs were higher than the TMFAA and TMFbulk, probably because of the greater variation of δ15Nbaseline, complex food sources or the notably different in individual organisms. Altogether, our results improve the understanding of Hg trophic transfer in aquatic, riparian, and terrestrial food webs.