以同位素標定法及二次離子質譜儀探討耳石氮硫之同化機制

Abstract

耳石（otolith）是硬骨魚類內耳中具代謝惰性的生物礦化結構，其有機基質中的穩定性氮與硫同位素具備重建魚類攝食生態的潛力。然而，受限於有機物含量低與整體分析（bulk analysis）空間解析度不足，由攝食端進入耳石的同化動力學與時間整合尺度仍缺乏實驗量化。本研究利用同位素標定餵食實驗結合奈米級二次離子質譜儀（NanoSIMS），探討莫三比克吳郭魚（Oreochromis mossambicus）幼魚耳石有機物中氮（δ15N值）與硫（δ34S值）同位素的置換動力學，並評估餌料品質與標定濃度的影響。實驗透過餵食富含 15N 的藻類與添加 L-Methionine-34S 的飼料建立同位素訊號，搭配茜素紅（alizarin red S, ARS）螢光標記進行時間定位。隨後利用 NanoSIMS 進行高空間解析度原位分析，重建標定訊號的時序變化，並以一階動力學模型估算其置換半衰期（t50%）。結果顯示，餌料基質顯著影響同化效率，攝食藻類基質的耳石標定硫貢獻度，顯著高於攝食鰻魚粉基質，即使兩者飼料端的標定34S比例相近。這顯示當天然含硫胺基酸受限時，魚體對外加L-Methionine-34S的利用率會提升。此外，耳石有機物同位素對餌料變化的反應極迅速，同化與代謝階段的 t50% 多落在 0.6 至 5 天內。而停止標定後耳石訊號迅速回落，顯示體內既存代謝庫的延遲效應甚微。本研究成功量化耳石有機同位素的同化時歷程，證實其能反映短期的攝食轉換，且訊號主要反映的是攝食來源，並且會受餌料成分的生物利用率所調控。未來若能增加實驗個體數以進一步釐清個體間的變異，將能強化該技術作為生態重建工具的應用深度。本研究成果為未來高解析度之魚類生活史重建提供了實驗性的學理基礎。

Otoliths are metabolically inert biomineralized structures in the inner ear of teleost fish, and the stable nitrogen and sulfur isotopes within their organic matrix hold potential for reconstructing fish feeding ecology. However, due to the low organic content of otoliths and the insufficient spatial resolution of conventional bulk analysis, the assimilation dynamics and temporal integration scales of isotopic signals from diet to otolith organic matter remain poorly quantified. This study employed a stable isotope labeling feeding experiment combined with Nanoscale Secondary Ion Mass Spectrometry (NanoSIMS) to investigate the turnover dynamics of nitrogen (δ15N) and sulfur (δ34S) isotopes in the otolith organic matrix of juvenile Mozambique tilapia (Oreochromis mossambicus), and to evaluate the effects of dietary quality and labeling dosage. Isotopic pulses were established by feeding fish 15N -enriched algae and diets supplemented with L-Methionine-34S, with alizarin red S, (ARS) fluorescent marking used for temporal referencing. NanoSIMS was then applied for high-resolution in situ analysis to reconstruct the temporal distribution of labeling signals, and a first-order kinetic model was used to estimate turnover half-lives (t50%). Results showed that dietary matrix significantly influenced assimilation efficiency: the labeled sulfur contribution in otoliths was markedly higher in fish fed algae-based diets than in those fed fishmeal-based diets, even when the proportion of labeled 34S in the diet was comparable between groups. This suggests that when natural sulfur-containing amino acids are limiting, fish preferentially utilize exogenous labeled methionine. Isotopic turnover was rapid, with t50% values of 0.6–5 days across assimilation and depuration phases, and the swift post-labeling signal decline indicated minimal delay effects from endogenous metabolic pools. These findings provide an experimental framework for high-resolution reconstruction of fish life history using otolith organic isotope chemistry.