Microbial lipids from a nearshore sediment core from Bowling Green Bay, North Queensland

Sediment samples were collected from 2 sites in Bowling Green Bay, a shallow coastal embayment located between Townsville and the Burdekin River delta in the central Great Barrier Reef province. A sediment core was collected at Station U for use in a depth profile study. The core was collected using a plastic, 51 mm diameter, core tube. The core was returned to the laboratory within 3 hours of collection, subsectioned and transferred to centrifuge tubes for pore water isolation. A surface sediment sample, collected at nearby Station 11 was used for an intact lipid study. The surface sediment (0-1 cm depth) was collected using a frame supported grab sampler and immediately frozen.All sediment samples were freeze dried and ground to < 100 µm in a stainless steel ring mill. Total carbon was measured using a LECO CHN Analyser. Total phosphorus was determined by ICPES after nitric acid/perchloric acid digestion. Pore waters were analysed for ammonium, phosphate, nitrite and nitrate using an Autoanalyser system.The results of pore water analyses indicated that bacterial activity responsible for the mineralization of the nutrients was confined to the upper 15-20 cm of the core. On this basis, samples from five depths were selected for lipid analysis (0-1 cm, 3-4 cm, 7-9 cm, 11-13 cm and for comparison 30-35 cm). Sediment samples (1-5 g dry wt) were extracted (x2) with hexane:isopropanol:water (HIP) 30:20:1.25. An aliquot of the extract was saponified and the resultant fatty acids were isolated and esterified to methyl esters. Lipid phosphate was determined by P analysis of an aliquot of the TSE after combustion and acid hydrolysis.An aliquot of the lipid extract from the surface sediment sample was separated into fractions by silica column chromatography. Four fractions were obtained. Each fraction was evaporated just to dryness under nitrogen and then transesterified. Non-ionic compounds were extracted from the resultant mixture with toluene. The toluene extract was evaporated to a small volume, treated and analysed by capillary GC and GC-MS. This research was undertaken to:1. examine the results of direct application of the total solvent extract (TSE) fatty acid based method to estimate bacterial biomass in a sediment core from an environment with low bacterial populations and significant relict sediment input2. examine methods which make the lipid phosphate and fatty acid procedures more reliable. Sedimentary monoenoic fatty acids have been shown to be potentially able to indicate the microbial abundance and community structure in diverse marine sediments. This procedure has been extended to enable detailed examination of bacterial community structure changes in surficial mangrove-associated sediments based solely on total solvent extract (TSE) fatty acid compositions. However, in environments where relict organic material contributes to the extractable fatty acid pool, or where bacterial populations are relatively low, the TSE fatty acids will include free fatty acids derived from dead and for decomposing organic matter. In such circumstances, biomass estimates based on TSE fatty acids can potentially lead to gross overestimations.

沉积物样本采集于鲍灵格林湾（Bowling Green Bay）的两个站点，该湾是位于汤斯维尔（Townsville）与伯德金河（Burdekin River）三角洲之间的浅海沿岸海湾，地处大堡礁（Great Barrier Reef）中部区域。

在U站点采集了一根沉积物岩心，用于深度剖面研究。岩心采用直径51毫米的塑料岩心管采集，采集后3小时内返回实验室，进行分段并转移至离心管以分离孔隙水。附近11号站点采集的表层沉积物样本用于完整脂质研究。表层沉积物（0-1厘米深度）采用框架支撑式抓斗采样器采集，并立即冷冻。

所有沉积物样本经冷冻干燥后，在不锈钢环磨机中研磨至粒径小于100微米。总碳含量采用LECO CHN分析仪测定；总磷含量经硝酸/高氯酸消解后，通过电感耦合等离子体发射光谱仪（ICPES）测定。孔隙水的铵、磷酸盐、亚硝酸盐及硝酸盐含量采用自动分析仪系统测定。

孔隙水分析结果表明，参与营养盐矿化的细菌活性局限于岩心上部15-20厘米区域。基于此，选取五个深度的样本进行脂质分析（0-1厘米、3-4厘米、7-9厘米、11-13厘米，以及用于对照的30-35厘米）。沉积物样本（干重1-5克）采用己烷:异丙醇:水（HIP，比例30:20:1.25）提取两次。取部分提取物进行皂化，所得脂肪酸经分离后酯化生成甲酯。脂质磷酸盐含量通过总溶剂提取物（TSE）部分样品的燃烧及酸水解后磷分析测定。

表层沉积物样本的部分脂质提取物通过硅胶柱层析分离为不同组分，共得到四个组分。每个组分在氮气保护下蒸发至近干，随后进行酯交换反应。从反应混合物中用甲苯萃取非离子化合物，萃取液蒸发至小体积，经处理后通过毛细管气相色谱（GC）及气相色谱-质谱联用（GC-MS）分析。

本研究旨在：
1. 探讨直接应用基于总溶剂提取物（TSE）脂肪酸的方法，对细菌种群数量低且存在大量遗留沉积物输入的环境中沉积物岩心的细菌生物量进行估算的结果；
2. 探索提高脂质磷酸盐及脂肪酸分析方法可靠性的途径。

沉积环境中的单不饱和脂肪酸已被证明可潜在指示不同海洋沉积物中的微生物丰度及群落结构。该方法已被拓展至仅基于总溶剂提取物（TSE）脂肪酸组成，即可详细分析表层红树林相关沉积物中细菌群落结构的变化。然而，在遗留有机物质对可提取脂肪酸库有贡献，或细菌种群相对较低的环境中，总溶剂提取物（TSE）脂肪酸将包含来自死亡及分解中有机物质的游离脂肪酸。在此情况下，基于TSE脂肪酸的生物量估算可能导致严重高估。