Laboratory work

3.3.1. Foraminifera analysis

Foraminiferal analysis was carried out for both the contemporary samples and fossil samples from cores OB5 and DMC1. On return to the laboratory, the contemporary samples were stained using a rose bengal solution (Walton, 1952; Murray and Bowser, 2000). The solution was made with a 30% ethanol solution (300 ml Ethanol and 700 ml distilled water) to which rose bengal (1.5 g) and sodium bicarbonate (1.5 g) were added. The rose bengal solution was added to the samples within 24 hours of collection and shaken to ensure all foraminifera took up the stain and soaked for at least 24 hours following Gehrels (2002).

Both sets of samples (contemporary and fossil) then followed the same methodological procedure. The samples were sieved through a 500 µm sieve and collected in 63 µm sieve following the methods of Scott and Medioli (1980a). They were then washed into beakers and the decanted material of random samples were examined to check for any foraminifera before being discarded (Gehrels, 2002; Horton and Edwards 2006). A wet-splitter was used in order to divide the sample into equal amounts allowing the volume of the amount counted to be known. A wet-splitter maximises the number of samples that may be processed whilst minimising loss or damage to foraminiferal tests and maintains a representative sub-sample (Gehrels, 1994). It is the most accurate and time efficient

preparation method (Edwards et al., 2004). To test whether the 1/8th was representative of

the whole sample, 1/8th (one section of the splitter) of a randomly selected sample was

counted, as well as the whole sample. Similar assemblages (<5% difference) were found for

the two counts. Therefore it was deemed suitable to use only 1/8th of the samples if enough

total and dead tests were counted.

The samples were counted wet to assist in the detection of rose bengal-stained foraminifera for the contemporary samples following Scott and Hermelin (1993) Gehrels (1994, 2002), Edwards et al. (2004) and Horton and Edwards (2006). In addition, wet counting is also favoured over drying the sample as drying can cause many problems including: consolidation or ‘pancaking’ as a result of drying out of organic residue which is often irreversible (Scott and Medioli, 1980a; Scott, et al., 2001); identification problems due to organic matter adhering to the tests making them difficult to identify (Patterson et al., 2005); the clumping of sediment together, making specimen picking difficult; and finally the

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organic linings of foraminifera that mark the dissolution of calcareous foraminifera or the breakdown of agglutinated tests may be lost (Scott et al., 2001).

A manageable amount of sediment was then transferred to a counting tray and counted wet under a binocular microscope at typical magnifications of x50, using Haynes (1973) and

Murray (1971b, 1979) for identification. Species of Ammonia beccarii, Elphidium and

Quinqueloculina were combined into generic groups (Hayward et al., 2004a; Horton and Edwards, 2006). A known sample volume was counted fully and repeated until at least 150 individuals were identified as well as at least 100 dead foraminifera (Fatela and Taborda, 2002).

3.3.2. Salinity and pH analysis

Salinity and pH were analysed for the surface samples as part of the environmental variables analysed for the contemporary study. The variables were measured at the same time using a 1:2 soil to water mix, made using 20 ml of sample which was made up to 70 ml with double distilled water. The sample was stirred and left for one hour. Both variables were measured using a K&M 7002 conductivity and pH probe and was calibrated for both pH and salinity. Salinity was converted into ‰ salinity using a standard equation (Salinity ‰

= 0.6679 (conductivity in mS cm-1) – 0.1513) following Gehrels and Newman (2004).

3.3.3. Organic matter content analysis

Organic matter content was analysed for both the surface samples and the collected cores. The organic matter was calculated using the loss on ignition method (LOI). Sub-samples of the collected sediment were weighed and freeze-dried at 35° C until all moisture was removed. The samples were then re-weighed to determine soil moisture content excluding hydroscopic moisture. To remove all the moisture, the samples were further sub-sampled (0.5 g) into 10 ml porcelain crucibles, weighed, oven-dried at 105° C overnight, and then re- weighed to determine the amount of hydroscopic water present. The samples were then placed into the furnace at 450° C for 4 hours to remove all the organic matter present. The samples were then re-weighed and the percentage of organic matter was calculated using the soil moisture lost and the loss on ignition results.

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3.3.4. Dry bulk density analysis

Dry bulk density (DBD) was calculated for cores OB5 and DMC1 to be used for radionuclide determination as well as to establish whether compaction has occurred. To calculate DBD, a 1 ml volume of sediment was cut out of each slice, measuring the volume of the cube as accurately as possible. The sub-sample was then weighed, frozen, freeze-dried and then re- weighed. The DBD was calculated by;

DBD = Volume / Dry weight

3.3.5. Grain size analysis

Grain size analysis was carried out for the surface samples as well as the cores for fossil foraminifera analysis OB5 and DMC1. The grain size distributions of the samples were measured using a Coulter Laser Granulometer (LS200). As the samples were very rich in organic matter, this was removed by digestion before grain size analysis was carried out. Approximately 5 g of wet or dry sample was sieved through a 2 mm sieve into a beaker then 20% hydrogen peroxide was added. The samples were heated gently on a hot plate and any floating organic material removed with tweezers to quicken the process. The samples remained on the hot plate alternating between adding more 20% hydrogen peroxide and double distilled water until all the organic matter was removed. A small

amount of sediment (<1 cm2) from each of the digested sample was then mixed on a watch

glass with calgon to disaggregate the sediment. Once smooth, the mixture was added to the granulometer and the grain size measured when satisfactory obscuration was reached.

The data were then processed using the computer program GRADISTAT (Blott, 2000).

3.3.6. XRF analysis

XRF analysis was carried out for 6 cores from Oglet Bay and one core from Decoy Marsh. The metal concentrations were determined using a BRUKER S2 Ranger energy dispersive X- ray fluorescence (XRF) and Atomic Adsorption Analysis (AAS). This analysis was carried out on several cores for Oglet Bay as well as for OB5 and DMC1. A sub-sample of core material was freeze-dried at 35°C until all moisture was removed. The sediment was then disaggregated into powder form using a pestle and mortar. A thin layer of the resulting sample was placed and lightly pressed with a plunger into a 25 mm-deep polythene tube which had a polypropylene film stretched across the base. The samples were then analysed along with certified standard reference materials of Buffalo River Sediment (srm2704) and Stream Sediment (GBW07305) which were used for calibration at the beginning of the

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analysis. In addition, a standard silica disc (Baxs-S2) was measured along with each set of samples measured. These were measured in addition to the samples to ensure consistency between measurement runs. As the samples were organic-rich, each sample was adjusted for the organic matter content using the LOI data in order to calculate the mass attenuation correction which is applied using the theoretical coefficients of Theisen and Vollath (1969).

3.3.7. Hg analysis

Hg concentrations were carried out for cores OB1, OB4, OB5 and DMC1 on a sub-sample of the sliced cores. This was carried out for sediment from every other centimetre in the core.

The wet sub-sample was firstly dried in a drying oven at 60 °_{C. The samples were then put}

into tinfoil-made pots and covered with blue roll and left in the oven overnight along with three standards; River Sediment (2704), San Joaquin (2709) and Light Sandy Soil (7002). Once the samples were dry, they were sub-sampled further by weighing 0.25 g (to 4 decimal places) of each into acid-washed polythene tubes. 1 ml of concentrated AnalaR

nitric acid was added and then placed in a shallow water bath at 90 °_{C for one hour. Once}

removed and cooled, 7 ml of double distilled water was added to each sample and shaken thoroughly. The samples were then left to settle for 30 minutes, flicked twice to bring clean liquid to the top of the tube, and this was repeated three times. The samples were then centrifuged at 1200 rpm for 15 minutes and the supernatant decanted. Along with the samples and sediment standards, three acid standards were also made which contained only 1ml of concentrated AnalaR nitric acid and 7 ml of double distilled water. Once each

sample had been prepared, a standard of SnCl2.2H2O solution was made by placing 5 g of

SnCl2.2H2O in a conical flask with 20 ml of concentrated HCl and stirred. 80 ml of double

distilled water was added and the sample was stirred. The solution was placed on a magnetic stirrer for the rest of the laboratory analysis. A Hg standard reagent was made at a dilution of 1:1000. 5 ml of concentrated HCl was added to 100 µl of Hg stock in a 100 ml flask and made up to 100 ml with double distilled water.

The samples were measured using an AAS. The instrument was first calibrated by measuring several standards including a blank sample. To make a blank sample 40-50 ml of double distilled water was put in a conical flask and 2 ml of the stannous chloride solution was added and a stopper put in the flask until it was ready to be measured. The program was then run and after 10 seconds of measuring the Drechsel head was fitted to the flask. After 80 seconds of measuring the Drechsel head was removed and rinsed with double distilled water. The standard samples measured were 10, 20, 40, 100 and 200 ng. A 10 ng

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standard was made by adding 10 µl of Hg reagent to 40-50 ml of double distilled water and 2 ml of stannous chloride solution. The samples were prepared by adding 1-5 ml of the sample to 45-49 ml of double distilled water and 2 ml of stannous chloride solution added. If the sample concentration was below that of the blank, the sample was re-measured using more sample. If the concentration of Hg in the sample was higher than that of the standard, less sample was used or a new standard was made using a higher concentration of Hg reagent. All samples were measured, including the sediment standards and acid standards. Every 5 samples a QBlank and QCheck of a standard (20 ng) were measured. Once all were measured, further Hg standards and blanks were measured.

The peak absorption for each sample was read from the signal graph produced by the AAS. The height of the increase was measured from each graph, measuring from the baseline readings to the top of the peak where it levelled off. The data was then entered into EXCEL and a calibration curve made. The results were then adjusted for standard drift in the values between blank and standard checks.

The samples were then corrected for the dilution and weight and the results produced in mg/g as follows; the unknown concentrations were converted to ng per measurement vessel using the calibration curve then converted into ng per sample.

The used mass (g) = (original sample weight (gm) x used extractant volume)/ dissolution volume (ml).

The sample concentration in (ng/g) was then calculated by dividing the AAS measurement by the used mass (g). To convert to mg/g, the sample concentration was further divided by 1000.

3.3.8. Radionuclide analysis

210

Pb dating was commissioned from the Quaternary Research Association and the Department of Geography, and was carried out on the cores OB5 and DMC1. Radiometric analyses were carried out in the Environmental Radioactivity Research Centre (ERRRC),

University of Liverpool. 210Pb, 226Ra, 137Cs and 241Am were analysed by direct gamma assay

using Ortec HPGe GWL (well-type) and GMX series coaxial low background intrinsic

germanium detectors (Appleby et al., 1986). 210Pb was determined via its gamma emissions

at 46.5 keV, and 226Ra by the 295 keV and 352 keV γ-rays emitted by its daughter

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equilibration. 137Cs and 241 Amwere measured by their emissions at 662keV and 59.5 keV

respectively. To determine the absolute efficiencies of the detectors, calibrated sources and sediment samples of known activity were used and corrections were made for the

effect of self absorption of low energy γ-rays within the sample (Appleby et al., 1992). (See

appendix 4 for full dating report).

3.3.9. Statistical analysis

Statistical analysis was carried out for the different datasets individually. For clarity and ease of understanding, the statistical analysis methodology will not be included in this chapter but will precede the results in each of the chapters 4, 5 and 6.

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