Organic Geochemistry 30 (1999) 477±483 Estimating the contribution of Spartina anglica biomass to salt-marsh sediments using compound speci®c stable carbon isotope measurements Ian D. Bull a, Pim F. van Bergen a, 1, Roland Bol b, Sue Brown c, Andrew R. Gledhill a, Alan J. Gray c, Douglas D. Harkness d, Simon E. Woodbury a, 2, Richard P. Evershed a,* aOrganic Geochemistry Unit, School of Chemistry, University of Bristol, Cantock's Close, Bristol BS8 1TS, UK bInstitute of Grassland and Environmental Research, North Wyke, Okehampton EX20 2SB, UK cInstitute of Terrestrial Ecology, Furzebrook Research Station, Wareham, Dorset BH20 5AS, UK dNERC Radiocarbon Laboratory, Scottish Enterprise Technology Park, East Kilbride G75 0QF, UK Received 23 November 1998; accepted 11 February 1999 (Returned to author for revision 17 December 1998) Abstract 13 Compound speci®c d C analyses were used to determine the relative input of a C4 temperate grass (Spartina anglica ) to primary biomass in a salt-marsh sediment. Lipid distributions revealed a C32 n-alkanol homologue as a characteristically dominant component of Spartina anglica whilst the cohabiting C3 species, Puccinellia maritima, exhibited a C26 maximum. The C32 n-alkanol component was used to create an isotopic mixing model, between organic matter derived from Spartina anglica and Puccinellia maritima, to estimate their relative contribution to the primary biomass input of salt-marsh sediments. The application of sedimentary lipid isotope data to the model gave values of Spartina anglica contributions ranging from 37 to 100%. This investigation represents the ®rst attempt to quantify inputs to sedimentary biomass based on compound speci®c stable carbon isotope techniques. # 1999 Elsevier Science Ltd. All rights reserved. 1. Introduction belonging to the family Poaceae and is a grass com- monly found in temperate salt-marshes (Vernberg, Spartina anglica is a monocotyledon angiosperm 1993). The origin of Spartina anglica can be traced to the accidental introduction of two species, Spartina maritima and Spartina alterni¯ora, to Britain in the early 19th century by shipping. Subsequent chromo- * Corresponding author. Tel.: +44-117-9287671; Fax: +44- some doubling of a sterile hybrid, Spartina townsendii, 117-9251295. produced the fertile amphidiploid hybrid, Spartina E-mail address: [email protected] (R.P. Evershed) anglica, which ``almost totally lacks in genetic vari- 1 Present address: Organic Geochemistry Group, Faculty of ation'' (Gray et al., 1991). As well as for its ability to Earth Sciences, Utrecht University, P.O. Box 80021, 3508 TA Utrecht, The Netherlands. stabilise/reclaim marsh-land, and the connected ben- 2 Present address: EKA Chemicals, 304 Worle Parkway, e®ts such as increased land for cattle grazing, Spartina Worle, Weston-super-Mare, Somerset BS22 6WA, UK. anglica has attracted additional attention. There has 0146-6380/99/$ - see front matter # 1999 Elsevier Science Ltd. All rights reserved. PII: S0146-6380(99)00022-4 478 I.D. Bull et al. / Organic Geochemistry 30 (1999) 477±483 been speculation about using certain species of 2. Experimental Spartina as an `environmentally friendly' fuel source by taking advantage of the C4 metabolism, inherent to 2.1. Sampling the genus, which enables high eciencies of radiant energy conversion to be realised (Long, 1987; Long et Table 1 summarises the sediment and vegetation al., 1989; Carruthers, 1994). Indeed, studies concerning samples collected for this study. The majority of Spartina anglica have demonstrated that this attribute samples were collected from a salt-marsh at Wytch is retained even in cool temperate climates with night Farm which is part of the intertidal environment of and day temperatures of 7 and 108C respectively caus- Poole Harbour, Dorset, UK. Two algal samples, Ulva ing little change in the photosynthetic rate of the plant lactata and Enteromorpha intestinalis, were collected at (Matsuba et al., 1997). Furthermore, if relative quan- Cleaval Point, a lower energy environment, located ap- tities of Spartina anglica primary biomass in sediment proximately one mile east of the Wytch Farm site. can be ascertained these results could be correlated Sediment samples were exposed using a spade followed with the extensive recorded chronology of Spartina by careful sub-sampling of the disturbed sediment. All anglica to provide a data set from which inferences samples were air dried at 608C then crushed with a about carbon ¯ows in salt-marshes and the eects of pestle and mortar. Vegetation was frozen with N2(l) to climatic change on this species might be made. facilitate crushing. Sediment samples were Soxhlet Bulk stable carbon isotope analyses have already extracted using a dichloromethane:acetone (9:1 v/v) been utilised in determining the dietary composition of solvent system as per Lehtonen and Ketola (1993). macro-invertebrates indigenous to salt-marshes Dried algae and ¯ora were extracted ultrasonically also (Jackson et al., 1986). However, analogous sedimen- using a dichloromethane:acetone (9:1 v/v) solvent sys- tary studies are hampered by the incorporation of iso- tem. Total lipid extracts (TLEs) were fractionated topically heavy organic matter from marine algae, e.g. according to compound class and derivatized for Ulva lactata (16.0-). Such values fall between the analysis as per Bull et al. (1998). range bounded by Spartina anglica (12.1-) and Puccinellia maritima (26.9-), a marsh-grass currently 2.2. Gas chromatographic (GC) and gas invading the environmental niches occupied by chromatographic±mass spectrometric (GC±MS) Spartina anglica (Gray et al., 1991; Bull, 1997). By analyses using gas chromatography combustion±isotope ratio monitoring mass spectrometry (GCC±IRMS) it is GC analyses were performed using a Hewlett± possible to perform d13C measurements on speci®c Packard 5890 series II gas chromatograph ®tted with a sedimentary lipids derived from the primary biomass fused silica capillary column coated with a 100% inputs of higher plants. This route of study circum- dimethylpolysiloxane stationary phase (Chrompack vents the problem of algal contribution and reliable CPSil-5 CB, 50 m  0.32 mm  0.12 mm, H2 carrier measurements pertaining to the relative incorporation gas). Derivatized samples were injected (1.0 ml) via an of Spartina anglica biomass into sediments may be on-column injector as solutions in hexane. The tem- made. perature was programmed from 408C (1 min) to 2008C Table 1 A summary of the samples collected Sample Description VegetationÐSpartina anglica Emergent aerial foliage obtained at the Wytch Farm site VegetationÐPuccinellia maritima Aerial foliage obtained at the Wytch Farm site AlgaeÐUlva lactata Whole algal sample obtained from a mud-¯at at Cleaval Point AlgaeÐEnteromorpha intestinalis Whole algal sample obtained from a mud-¯at at Cleaval Point AlgaeÐFilamentous (unidenti®ed) Thin mat present beneath and around Spartina anglica at Wytch Farm SedimentÐSpartina (oxic) Top (<7 cm) layer of brown sediment beneath Spartina anglica obtained at the Wytch Farm site SedimentÐSpartina (anoxic) Bottom (>7 cm) layer of grey sediment beneath Spartina anglica obtained at the Wytch Farm site SedimentÐPuccinellia Brown sediment beneath Puccinellia maritima obtained at the Wytch Farm site SedimentÐMud-¯at Grey sediment located 01 m away from the salt-marsh towards the water, obtained at the Wytch Farm site I.D. Bull et al. / Organic Geochemistry 30 (1999) 477±483 479 at a rate of 108C min1 then to 3008C at a rate of 38C ponents in each TLE are n-alkanols, dotriacontanol 1 min with an isothermal of 20 min (FID, 3008C). GC (C32: Spartina anglica ) and hexacosanol (C26: analyses of TLEs were made using an alternative col- Puccinellia maritima ), which have been established pre- umn capable of performing at higher temperatures viously as major lipid components in the epicuticular (J&W Scienti®c DB1, 15 m  0.32 mm  0.1 mm, H2 leaf waxes of higher plants (Tulloch, 1976, 1981). carrier gas). The temperature was programmed from GCC±IRMS analysis of n-alkanols is relatively facile 508C (2 min) to 3508C at a rate of 108C min1 with an and is improved by a simple chemical separation of isothermal of 10 min (FID, 3508C). these components prior to analysis thereby making GC±MS analyses were performed on a Carlo Erba them ideal compounds to be used in this study. 5160 GC, using the same capillary columns and tem- Fig. 2 summarises the abundance of n-alkanol com- perature programs reported above but employing ponents observed to occur in the separated n-alkanol helium as a carrier gas and on-column injection fractions of Spartina anglica and Puccinellia maritima. coupled, via a heated transfer line, to a Finnigan MAT Quite clearly the C26 n-alkanol homologue in 4500 quadrupole mass spectrometer scanning in the Puccinellia maritima is excessively abundant compared range of m/z 50 to 850 with a cycle time of 1.5 s. The with that in Spartina anglica rendering any isotopic current was maintained at 300 mA with an ion source results obtained from its analysis prone to `swamping' temperature of 1908C. The mass spectrometer was eects biased towards the C3 grass. However, the operated with an electron voltage of 70 eV. abundance of the C32 homologue in the two species is similar making it a suitable component to monitor 2.3. Stable carbon isotope analyses relative contributions of Spartina anglica to the pri- mary biomass of sediments. This may be done through Analyses were made on 1.0 ml sample aliquots using