Using biomarkers to determine dispersion and cycling of organic matter from the Sepik River, Papua New Guinea

In January 2000, two sediment trap arrays consisting of five stainless steel cylinders (1 m long and 10 cm diameter) fitted into a weighted aluminium frame were deployed for 48 hours on a free drifting mooring in the Sepik Canyon (SC) at depths of 100 and 260 m. During deployment, the mooring drifted 19.5 km north west from the Canyon axis, but remained within the area of the river plume. The mooring was retrieved in approximately 380 m of water and the contents of the sediment traps were drained into 4 L glass bottles. The bottles were shaken and sub-sampled for particulate carbon and the remaining samples were then filtered through pre-weighed 0.45 µm GFF filters, rinsed with MilliQ water to remove salt, and were wrapped in pre-combusted foil, then stored at -20°C. Water samples for lipid analysis were collected with Infiltrex II (TM) high volume samplers. Samplers were attached to the sediment trap mooring at depths of 55, 180, 200 and 220 m. Depths were chosen to target layers of turbidity as detected with the CTD instruments. Each sampler pumped 388- 432 L of seawater before the filters clogged and the pumps shut down. At other offshore stations an Infiltrix sampler was attached to the hydrowire and lowered to sample at 220 m within the undercurrent while the ship drifted. These deployments were short in duration and only sampled about 90 L each time. Samples were collected at Vitiaz Strait (Vitiaz), a region not influenced by the river, and another station (S+2) to the northwest of the river, which may be influenced by it, to determine dispersion and potential entrainment in the 200 m undercurrents. Samples were also collected upstream of the small village of Kopar (SR), at a depth of 1 m below the river surface, in 4 L glass bottles and pumped through an Infiltrex II (TM) sampler on the deck of the vessel. Upon retrieval of the samplers, the GFF filters were removed and packed in glass jars with foil lined lids and stored at -20°C. The XAD2 columns were removed, resealed and stored at 4°C. Samples for marine dissolved lignin analysis were collected using 10 L Niskin bottles attached to a CTD system. Specific depths were targeted, which offshore included the chlorophyll maximum and the depth of the New Guinea Coastal Undercurrent (NGCUC1, NGCUC2) or the Equatorial Undercurrent (EUC1, EUC2) at 220 m. Upon retrieval the Niskin bottles were clamped shut, pressurised with N2 gas and the water was filtered through GFF filters into 4 L glass bottles. The filtered seawater was acidified to pH 2 with concentrated HCl and poured back into the 10 L Niskins. The bottles were again clamped and pressurized and the water forced through prepared cartridges of C18 silica for solid phase extraction of lignin. After sampling, the cartridges were re-wrapped in cleaned foil and stored at 4°C. Riverine dissolved lignin samples were collected using GFF filtration into 1 L glass bottles and frozen at dry ice temperature. In the laboratory, the water was removed using rotary evaporation, leaving a powder for lignin analysis. Surface sediment samples for lignin analysis were collected from 19 sites on the coastal margin (around SC) by gravity and Kasten corers.Additional water samples were collected using Niskin bottles for dissolved and particulate carbon (DOC, POC), nutrient and stable isotope analyses. Riverine samples, collected for lignin analysis, were also analysed to determine organic carbon, total nitrogen, stable carbon isotope ratios and stable nitrogen isotope ratios. Samples collected for lipid analysis were extracted and a subsample taken for determination of total extractable organic matter (EOM). The remaining extracts were reduced and saponified and the neutral lipids extracted and then separated into hydrocarbon and esters plus alcohol/sterol fractions on silica gel. The remaining extract was acidified and the fatty acids were extracted.Alkanes in the hydrocarbon fractions were determined using a gas chromatograph fitted with a flame ionization detector (GC-FID). Polynuclear aromatic hydrocarbon (PAH) analysis was carried out using selected ion monitoring gas chromatography-mass spectroscopy (SIM GC-MS). Sterols and n-alcohols were converted to TMS esters and analysed using full scan gas chromatography/mass spectroscopy (GC-MS). The fatty acid fraction was converted to methyl esters and analysed using GC-FID.Dried riverine and surface sediment samples for lignin analysis were converted to phenols and the lignin-derived phenols separated using GC-MS. Quantification was achieved using SIM GC-MS, This research was undertaken to describe the fate and cycling of organic matter input from the Sepik River into the near coastal waters using molecular markers and to investigate stable biomarkers that might be useful for tracing long range transport. This work is part of Project TROPICS (Tropical River Ocean Processes in Coastal Settings), a joint coastal oceanographic study by Australia, Indonesia, Papua New Guinea, and the USA.

2000年1月，两套沉积物捕集器（sediment trap）阵列，每套由5根长1m、直径10cm的不锈钢圆柱组装于配重铝制框架而成，被部署于塞皮克峡谷（Sepik Canyon, SC）海域的自由漂流锚系上，布放水深分别为100m与260m，布放时长共计48小时。布放期间，锚系向西北方向漂移19.5km，偏离峡谷轴线，但仍处于河流羽状流影响范围内。最终在约380m水深处回收锚系，将沉积物捕集器内的样品转移至4L玻璃瓶中。
对玻璃瓶进行振荡后，分取子样用于颗粒碳分析；剩余样品经预先称重的0.45 µm GFF滤膜过滤，用密理博纯水（MilliQ water）冲洗以去除盐分，随后用预灼烧铝箔包裹，于-20℃低温保存。
用于脂质分析的水样采用Infiltrex II（商标）大体积采样器采集。采样器安装于沉积物捕集器锚系上，布放水深分别为55m、180m、200m与220m，所选水深对应温盐深仪（CTD instruments）探测到的浊度层。每台采样器在滤膜堵塞、泵体停机前，可累计抽取388~432L海水。
在其余近海站位，Infiltrix采样器安装于水文缆绳上，随船漂移过程中下放至220m水深的底流层开展采样。此类布放时长较短，单次采样量仅约90L。
研究团队分别在不受河流影响的维蒂亚兹海峡（Vitiaz Strait, Vitiaz），以及河流西北侧可能受其影响的S+2站位采集样品，以探究200m水深底流中的物质扩散与潜在夹带过程。此外，在科帕尔村（Kopar, SR）上游、距河面1m深度处，采用4L玻璃瓶采集水样，并在船载平台上通过Infiltrex II（商标）采样器完成抽滤。采样器回收后，取出GFF滤膜，封装于带有铝箔衬垫盖子的玻璃罐中，于-20℃保存；取出XAD-2树脂柱并重新密封，于4℃低温保存。
用于海洋溶解木质素（marine dissolved lignin）分析的样品，采用安装于温盐深（CTD）系统上的10L尼斯金采水器（Niskin bottle）采集。目标水深涵盖近海叶绿素最大值层、新几内亚沿岸底流（New Guinea Coastal Undercurrent, NGCUC1、NGCUC2）所在水深，以及220m处的赤道底流（Equatorial Undercurrent, EUC1、EUC2）所在水深。采样器回收后，将尼斯金采水器夹闭，用氮气（N2 gas）加压，将水样经GFF滤膜过滤至4L玻璃瓶中。将过滤后的海水用浓盐酸酸化至pH=2，倒回10L尼斯金采水器中，再次夹闭并加压，使水流通过制备好的C18硅胶固相萃取柱，以萃取木质素。采样完成后，将萃取柱用洁净铝箔重新包裹，于4℃低温保存。
河流溶解木质素样品经GFF滤膜过滤后收集于1L玻璃瓶中，置于干冰温度下冷冻保存。实验室阶段通过旋转蒸发（rotary evaporation）去除水分，得到用于木质素分析的粉末样品。
用于木质素分析的表层沉积物样品，采用重力取样器与凯斯顿采泥器，在塞皮克峡谷周边沿岸带的19个站位采集。
研究团队还额外采集水样，用于溶解态与颗粒态碳（dissolved and particulate carbon, DOC、POC）、营养盐及稳定同位素分析。针对木质素分析采集的河流样品，同时开展有机碳、总氮、稳定碳同位素比值及稳定氮同位素比值测定。
用于脂质分析的样品经萃取后，分取子样测定总可提取有机质（total extractable organic matter, EOM）。剩余萃取液经浓缩、皂化处理，提取中性脂质，再经硅胶柱分离为烃类组分与酯/醇/甾醇组分。剩余萃取液经酸化后，提取脂肪酸。
烃类组分中的烷烃采用配备火焰离子化检测器的气相色谱仪（GC-FID）测定；多环芳烃（PAH, Polynuclear aromatic hydrocarbon）分析采用选择离子监测气相色谱-质谱联用仪（SIM GC-MS）；甾醇与正构醇经衍生为三甲基硅基酯（TMS ester）后，采用全扫描气相色谱-质谱联用仪（GC-MS）分析；脂肪酸组分经衍生为甲酯后，采用GC-FID分析。
用于木质素分析的干燥河流与表层沉积物样品被转化为酚类物质，通过GC-MS分离木质素源酚类（lignin-derived phenols），采用SIM GC-MS完成定量。
本研究旨在利用分子标志物描述塞皮克河输入近岸海域的有机质归趋与循环过程，并探究可用于示踪长距离输运过程的稳定生物标志物。本研究属于TROPICS项目（热带河流-近海海岸带海洋过程，Tropical River Ocean Processes in Coastal Settings, Project TROPICS）的一部分，该项目是澳大利亚、印度尼西亚、巴布亚新几内亚与美国联合开展的近海海洋学研究。