13C values, indicating little or no effect of the acid treatments on the integrity of the organic C reservoir.
Prior to conducting our analytical work on the Site 1040 samples, we undertook a series of tests to (1) investigate the possible effects (e.g., decomposition) of long-term storage of seafloor sediments under refrigeration and therefore investigate our ability to exploit the extensive archives of sediment sections obtained from oceans and lakes with confidence that the organic C-N reservoirs have been preserved, (2) examine whether low-temperature heating during drying causes decomposition of organic matter affecting the C-N compositions, and (3) identify any possible effects of H2O rinsing and treatments with HCl on the organic and inorganic N reservoirs.
For determinations of TOC concentration and isotopic composition, each sample was divided into three parts and the splits were reacted overnight with 0.5-N, 2-N, or 6-N HCl. The residues were rinsed with distilled H2O, centrifuged three times, freeze-dried, and again homogenized. Approximately 100 mg of decalcified sediment sample was loaded with Cu-CuxOx reagent, sealed in 6-mm quartz tubes at high vacuum, and combusted at 910°C for 3 hr. TOC concentrations were calculated from CO2 yields determined using a Hg manometer.
All samples except one used for N concentration and isotopic composition analyses were Leg 205 sediments frozen on board ship. For the N analyses, each sample was divided into three parts. One split was reacted with 6-N HCl overnight, rinsed with distilled H2O, and centrifuged three times. A second split was only rinsed with distilled H2O and centrifuged three times. The third split did not receive any pretreatment. About 500–1000 mg of freeze-dried, homogenized sample rinsed with 6-N HCl or distilled H2O rinsing and mixed with Cu-CuxOx reagent was put into 9-mm quartz tubes, sealed at high-vacuum, and combusted at 910°C for 3 hr (see description of heating routines by Bebout and Fogel, 1992; Sadofsky and Bebout, 2000; Bebout and Sadofsky, 2004). Nitrogen concentrations were determined by measuring the m/z 28 signal for calibrated inlet volumes in the mass spectrometer.
Concentrations and isotopic ratios of TOC of samples treated with 0.5- to 6-N HCl acid and rinsed three times with H2O are listed in Table AT1. For all seven of the samples we tested, the three treatments yielded reasonably similar values in both TOC concentrations and 13C values, indicating little or no effect of the acid treatments on the integrity of the organic C reservoir.
15N
Nitrogen concentrations and 15N values of untreated sediment, H2O-rinsed sediment, and sediment treated with 6-N HCl and rinsed with H2O three times are listed in Table AT2. Relative to the untreated splits, the sample splits rinsed with H2O showed slight decreases in N concentrations but significant increases in 15N values. Sample splits treated with 6-N HCl then rinsed three times with H2O showed significant decreases in N concentrations but smaller increases in 15N values (Fig. AF1).
The decrease of N concentrations related to these treatments indicates that certain N species were removed by the H2O or HCl. In general, distilled H2O is expected to leach only the more soluble ions/cations, such as ammonium (NH4+) and nitrate (NO3–), and maybe some surface-absorbed material. Regarding the two soluble N species, the heavy isotope 15N is preferentially incorporated into nitrate, whereas the light isotope 14N is preferentially incorporated into ammonium (i.e., ammonium has lower 15N and nitrate has higher 15N values). The removal of a 14N-enriched N component with distilled H2O can explain the increase in 15N of the sediment N. Removal of the 15N-enriched portion would be expected to produce an increase in the sediment 15N values. In sediments from Legs 170 and 205, interstitial fluid contains hundreds to tens of thousands of parts per million NH4+ (Kimura, Silver, Blum, et al., 1997), but nitrate data are not yet available. If nitrate is not abundant, the dominant H2O-mobile N component in these sediments is ammonium, which is consistent with the observation of increase in 15N values with decrease of N concentrations after H2O rinsing.
The decrease in the N concentrations of the sediments after 6-N HCl treatment and rinsing with H2O indicates more dramatic removal than in the treatments with only H2O. Since there is little or no difference between the 15N values of the untreated samples and the 6-N HCl–rinsed samples, the N species removed by this treatment, but not the 15N-only treatments, presumably had heavier N isotopic ratios resulting in a removal of N with averaged 15N near that of the untreated sediment. Proteins and amino acids, which are characterized by more positive C and N isotopic compositions, could be candidates for these loosely bound but easily acid-decomposed N species. Decrease in 13C values or TOC concentrations, however, were not observed in our 0.5-N to 6-N HCl treatments (Table AT1), implying preferential removal of components with high N/C during the acid treatments. The decoupling of C and N isotopes during sediment decalcification was also documented by Krishnamurthy et al. (1999), and the mechanism remains unclear.
Based on the comparisons above, we used 1-N HCl rinsing pretreatment for TOC analyses and unpretreated samples for the TN analyses for all other sediments from Site 1040 and all but the three samples from Hole 1255A that were pretreated using distilled H2O or by 6-N HCl treatment followed by rinsing with H2O. The results are shown in Figures F2 and F3. On these figures, it is evident that the air-dried/stored samples from the Site 1040 sediments yielded C and N concentrations and isotopic compositions quite similar to those of the refrigerated or freeze-dried sediments from Site 1040 from similar depths. The samples are generally collected from several centimeters away (Table AT2), or at the least, falling within the range of natural scatter in the core. Furthermore, the data for Hole 1254A overlap well with those for Site 1040 in both TOC and TN concentrations and isotopic compositions but with a small number of outliers. The three samples from Hole 1255A show more variation in C and N concentrations and isotopic compositions, Variations for N are partly a result of the different pretreatments for the N determinations, but are possibly also related to the natural sediment heterogeneity. The consistency in C and N concentrations and isotopic compositions between these differentially preserved samples indicates that room-temperature-air dried sediments should be suitable for organic C-N geochemical study. This conclusion might also be extended to the low-temperature heated samples, which are dried in a drying oven at 40°–60°C (Minoura et al., 1997; Tolun et al., 2002; Sadofsky and Bebout, 2004).