The waxy coating that protects the leaves and other soft tissues of plants includes n-alkane and nalkanoic acid compounds that are commonly used as biomarkers to reconstruct past environment.
Plant waxes have geological relevance given their persistence in soils and paleosols, as well as in lake and marine sediments, yet diagenesis may alter their molecular and isotopic signatures from synthesis to deposition. This study seeks to understand the fate of plant wax biomarkers in soils after leaf-fall as characterized by a series of tropical soil profiles. We investigate the changes in abundance, molecular distributions, and hydrogen (δD) and carbon isotopic compositions (δ 13 C) of plant waxes (n-alkanes and n-alkanoic acids) in six litter-to-soil profiles along a 2740 m elevation transect from the eastern flank of the Andes mountains down to the lowland Amazon floodplain in Peru. From litter to soil, we find acid/alkane ratios increase, while absolute abundances decrease. In contrast, within each soil, acid/alkane ratios are roughly constant and we find an equivalent exponential decline in concentration in both compound classes with depth; with molecular distributions indicating some new production. We observe a 4 -6‰ 13 Cenrichment from litter to deeper soils for both C 29 n-alkanes and C 30 n-alkanoic acids; of which the Suess effect accounts for ≤ 2‰. We infer that microbial degradation and production (or 'turnover') processes influence the δ 13 C of plant waxes that survive in soils; in contrast, no systematic change in δD values is observed. The plant wax signal in soils includes averaging of inputs and diagenetic effects, so this signature is particularly relevant for the interpretation of plant waxes archives in paleosols and the plant waxes eroded from soils and exported to downstream sedimentary archives. We show that soils represent the major stock of plant wax under living ecosystems, suggesting that soils may be a quantitatively-important source of plant waxes available for fluvial erosion, with implications for studies of carbon cycling and 38 paleoenvironmental reconstructions from downstream geological archives.
Plant wax biomarkers are commonly used to reconstruct past environments based upon the carbon and hydrogen isotopic compositions that reflect aspects of vegetation and climate (Eglinton & Eglinton, 2008). Geological applications focus on sedimentary deposits that archive the spatial and temporal record of these molecular fossils, and plant waxes have been found preserved in paleosols (e.g., Magill et al., 2016), lake sediments (e.g., Fornace et al., 2014) and marine sediments (e.g., Tipple and Pagani, 2010). In order to calibrate how the plant wax proxy records aspects of vegetation and climate, many studies have sampled leaves from living vegetation, including studies of temperate forests (Sachse et al., 2006), arid ecosystems (Feakins & Sessions, 2010), tropical forests (Vogts et al., 2009) and high latitude ecosystems (Wilkie et al., 2013). Modern lake sediments (Sachse et al., 2004) and marine core tops (Rommerskirchen et al., 2003) have been used to study plant wax delivered by wind and water transport. Soils have also been surveyed to characterize plant wax variations along altitudinal transects (Jia et al., 2008;Bai et al., 2011), latitudinal transects (Bush and McInerney, 2015;Bakkelund et al., 2018) and aridity gradients (Schwab et al., 2015).
Given the ~2000 Pg of organic carbon stored in soils globally (Batjes, 1996;Jobbágy & Jackson, 2000), soils are a major source of the organic carbon (including plant waxes) eroded from the continents to lake and ocean sediments (Blair et al., 2004). Soils are a particularly important storage step (Blair et al., 2004) between new plant production and erosion by rivers given the age of plant waxes transported by rivers revealed by compound specific radiocarbon (Kusch et al., 2010;French et al., 2018), that suggests storage from decades to thousands of years.
Soils can be sampled as an archive of environmental information in situ integrating the time of soil formation, and given requisite burial or protection from erosion soils may be preserved in the form of paleosols, yielding information based on pedogenic structures and thicknesses (e.g., Retallack, 2013) as well as pedogenic carbonate nodules (e.g., Cerling andQuade, 1989, Quade et al., 2013) and more recently plant waxes (e.g. Magill et al., 2016).
When interpreting plant waxes stored in paleosols or derived from soil erosion, we need to understand how plant wax biomarkers are incorporated into soils and how diagenesis may alter their molecular and isotopic signatures from synthesis to deposition. Once a leaf falls from the canopy it forms the litter layer on top of the soil, with leaves comprising the majority, often >60%, of litterfall (Kögel-Knabner & Amelung, 2014). Removal processes associated with herbivory and microbial degradation (and/or runoff erosion on steep slopes) may be considerable, but litter represents the input of organic matter at the top of the soil profile and can contribute to the upward accumulation of soils. In lower layers of the soil, weathering of parent rock may deepen the soil downwards (Amundson, 2014). Over time, soil erosion by water or wind may remove surficial layers of the soil, or landsliding may remove forest, soil and rock such as in the steep-sided Andes (Clark et al., 2016) or the banks of meandering lowland rivers (Torres et al., 2017). The residence time of soil is therefore controlled by the balance of additions from above and below, and removal processes of degradation and erosion (Heimsath et al., 1997). The persistence of soil organic matter, and its individual compounds, is decoupled from the intrinsic thermodynamic stability expected based on molecular structure: many compounds persist decades beyond their expected residence time, reflecting the importance of packaging within soil aggregates and adsorption to minerals (Schmidt et al., 2011). If the persistence of soil organic carbon is an "ecosystem property" (Schmidt et al., 2011), then more work needs to be done to characterize the fate of individual compounds in a range of ecosystems and terrains for carbon cycle applications, as well as for paleoclimate reconstructions using biomarkers.
Previous research to understand the effect of early diagenesis on plant wax biomarkers has included field studies in low-diversity temperate ecosystems, comparing fresh leaves with litter and soil (Nguyen Tu et al., 2004;Chikaraishi & Naraoka, 2006;Zhang et al., 2017), and monitoring changes with time in litterbag experiments (Huang et al., 1997;Nguyen Tu et al., 2017, 2011;Zech et al., 2011;Wang et al., 2014;Li et al., 2017). Most have studied n-alkanes only, with the exception of Chikaraishi & Naraoka (2006) who studied a suite of lipids including n-alkanes and n-alkanoic acids.
Study of hydrogen isotope effects associated with diagenesis is limited. In a litter bag experiment of three broadleaf tree species, Zech et al. (2011) found seasonal variations of 10 -20 ‰ in nalkane δD values that were attributed to microbial n-alkane production, but they found no systematic overall trend across the 2 year study. In contrast, a study of a soil profile in a Japanese maple forest found D-depletion (by ~50‰) in both n-alkanes and n-alkanoic acids from leaf to soil, suggesting a significant hydrogen isotope effect during early diagenesis in soils (Chikaraishi & Naraoka, 2006).
In contrast, the carbon isotopic effect associated with plant wax degradation is relatively wellknown. Prior studies have reported an increase (~1 -2‰) in plant wax δ 13 C during early diagenesis, as reflected in differences between fresh leaves and leaf litter, and also seen in changes during 1-3 yrs of litter decomposition in experiments (Nguyen Tu et al., 2004;Chikaraishi & Naraoka, 2006;Wang et al., 2014;Li et al., 2017;Zhang et al., 2017), although two shrub species showed no temporal change in δ 13 C (Huang et al., 1997;Li et al., 2017).
Considering the diversity in species (including maple, ginkgo, bamboo, C3 and C4 grasses, moss) and sites studied so far, a 1 -2‰ 13 C-enrichment appears to be a widespread signature associated with degradation.
1.2 Tropical soils in an Andes-Amazon transect A litter translocation experiment across the Andes-Amazon transition in Peru has found a strong dependence of litter degradation on soil temperatures, with ~3-fold higher degradation rates at lowland sites of 24°C compared to upland sites of 12°C mean annual soil temperature (Salinas et al., 2010). The dependence of degradation rates on temperature also leads to thicker soils and higher soil organic carbon (OC) contents in the colder montane cloud forests compared to lowland tropical rainforests (Whitaker et al., 2014). Microbial community also changes in response to elevation, with increased microbial biomass and fungi relative to bacteria with increasing altitude, which affects soil respiration rates (Whitaker et al., 2014). These environmental controls and microbial processes not only determine the fate of bulk OC as a whole, but also may have different influence on various types of organic compounds, including plant wax n-alkanes and n-alkanoic acids.
Here, we study plant waxes in leaf litters and soils from a series of soil pits under tropical forests at contrasting altitudes, spanning sites from the eastern flank of the Peruvian Andes to the Amazon floodplain. We quantify how bulk organic carbon and plant wax molecular and isotopic signatures vary during soil storage by sampling the progression from leaf litter down through the soil profile, with detailed sampling of soil pits dug along the elevation transect across a wide range of temperatures and soil organic layer thickness. We study molecular abundances and C and H isotope compositions of both n-alkanes and n-alkanoic acids, aiming towards a more comprehensive understanding of the preservation/alteration of plant wax biomarkers from plants to soils.
This study adds to prior work on plant waxes in the Madre de Dios region of Peru including canopy surveys of leaf wax n-alkane molecular abundance distributions and productivity (Feakins et al., 2016a); canopy bulk leaf and leaf wax carbon isotopic composition (Wu et al., 2017); and canopy leaf wax hydrogen isotopic composition together with plant ecohydrology (Feakins et al., 2016b), as well as river export of plant waxes n-alkanoic acids related to soil mineral horizon hydrogen isotopic composition (Ponton et al., 2014) and dual isotope comparison of plant wax n-alkanes and n-alkanoic acids in soils and river (Feakins et al., 2018).
Those studies found a linear trend in both δ 13 C and δD values of both n-alkanes and n-alkanoic acids in canopy leaves and soils with elevation, supporting the use of these metrics as proxies for elevation for paleoaltimetry for ancient deposits and to indicate sourcing-elevation of plant waxes exported by rivers within the catchment. The latter study also found an isotopic offset in δ 13 C values of the C 29 n-alkane between canopy and soils (Feakins et al., 2018) that we investigate further here.
We collected samples from four sites across our study area located in the Madre de Dios region of Peru, spanning elevations from 286 m in the Amazon floodplain to 3025 m along the eastern flank of the Andes (Fig. 1, Table 1). The region receives 1560 -5300 mm mean annual precipitation (MAP) and is fully forested (tropical montane cloud forest to lowland rainforest).
The sample sites span a temperature range of 11.1 -24.4 °C. All sites are primary forests with one secondary growth forest site in the foothills, at Villa Carmen (VC), previously logged and now dominated by bamboos. The primary forest sites are highly-biodiverse. Tree species with high abundance include Weinmannia crassifolia, Clusia alata cf., and Hesperomeles ferruginea at Wayqecha (WAY), as well as Alchornea latifolia, Tachigali setifera, and Tapirira obtuse at San Pedro (SP). The lowland tropical rainforest (TR) is characterized by even higher biodiversity, but abundant Amazonian lineages include Inga, Swartzia, Protieae, and Guatteria including presence of species of those genera at the Los Amigos (LA) site (Dexter et al., 2017).
Soil types include Umbrisol at WAY, Cambisol at SP (Whitaker et al., 2014), and Ultisol LA (Pittman et al., 2001), but have not been previously classified at our VC site. Soil organic layer thickness varies from 1 to 26 cm with a tendency towards increasing thickness at higher altitudes (Table 1). Along this transect are a series of permanent forest plots that are part of the Global Ecosystems Monitoring Network (GEM; http://gem.tropicalforests.ox.ac.uk/projects/aberg), where canopy leaf wax has been studied before (Feakins et al., 2016a,b;Wu et al., 2017), and where aggregate soil organic (O) and mineral (M) samples have been collected and studied by amalgamating soils from five locations at each plot (Ponton et al., 2014;Nottingham et al., 2015;Feng et al., 2016;Feakins et al., 2018). Here we study individual vertical soil profiles, sampling within a single pit to investigate degradation processes and transformation of plant wax signatures during soil storage at each of 4 sites along the elevation transect. Although this region experiences landslides in areas of steep relief (Clark et al., 2016), we selected soil pits at locations where the surface did not appear to be disturbed. Within the soil pits, examination of the color, texture, and structure of the soil profiles suggested that the soils had formed from downward weathering and upward accumulation of leaf litter, without sedimentary structures indicative of disturbance by erosional reworking. At two sites (VC and LA), an additional site was sampled to contrast hillslope setting, by digging one pit at the ridgetop and one at the slope base. In total we present data for 6 pits (Table 1). This soil profile study overlaps with the prior plant wax study of soil O and M layers (Feakins et al., 2018) at WAY and SP where we can make direct comparisons. In addition, we can compare the litter-soil profile at SP with data from canopy leaves from previous studies (Feakins et al., 2016a,b;Wu et al., 2017). The two soil profiles at the high elevation sites at Wayqecha (WAY) and San Pedro (SP) are located under tropical montane cloud forest and correspond to the RAINFOR sites of the same names, with the SP site equivalent to RAINFOR SP-1500 (www.rainfor.org). The Villa Carmen (VC) plot is under a secondary forest in the foothills of Andes, and Los Amigos (LA) is located in the tropical rain forest of the Amazon floodplain. Two soil pits were dug each at VC and LA, with one located at a slope base (VC2 and LA5) and the other located on top of a nearby ridge (VC3 and LA4).
We collected leaf litter at WAY, SP, VC2 and LA4, with the litter at WAY (the upper site) divided into top, middle, and bottom litter because of its thickness (~12 cm). Soil pits were dug to ~90 -150 cm depth, and 3 -4 samples (integrating 5 -50 cm of soil vertically) were taken at each pit based on the soil profile characterization (based on color and physical properties). We also sampled roots at SP and LA4. Samples were stored under cool conditions in the field until transport back to the laboratory where they were stored in a freezer at -20°C, before being freeze-dried. As rock fragments were present in many soil samples, clasts >2 mm were removed by sieving. The soil samples were then ground in a pestle and mortar to homogenize for geochemical analyses.
Aliquots of the soil samples were taken for total organic carbon (TOC) and bulk organic carbon isotope (δ 13 C OC ) analysis. The samples were heated in dilute (10%) HCl to 70°C in a water bath for 1 h to remove carbonates. The decarbonated samples were then rinsed three times with deionized water, and dried in an oven at 56°C. The dried samples were analyzed for TOC and δ 13 C OC using a Costech Elemental Combustion System (EA 4010) connected via a Picarro Liaison (A0301) to a Picarro cavity ring down spectrometer (G2131-i). A USGS-40 standard (Glutamic Acid with δ 13 C OC = -26.6‰ in VPBD scale) was run with replicates at different weights at the beginning and end of the sequence to provide a calibration curve for the measured TOC, as well as an assessment of the precision in δ 13 C OC measurements (determined to be better than 0.2‰).
Total lipid extracts (TLE) were extracted from freeze-dried samples with 9:1 dichloromethane (DCM) to methane (MeOH) using an Accelerated Solvent Extraction system (ASE 350, Dionex) at 100°C and 1500 psi for 2 cycles of 15 mins. The TLE was separated into neutral (FN; containing n-alkanes) and acid (FA; containing n-alkanoic acids) fractions by eluting 2:1 DCM to isopropanol and 4% formic acid in ethyl ether respectively through a column of LC-NH 2 gel.
The n-alkanes were then further separated from the FN fraction by eluting with hexane through a silica gel column. The FA fraction was methylated in 5% HCl in MeOH of known isotopic compositions at 70°C overnight, during which the n-alkanoic acids were reacted into fatty acid methyl esters (FAMEs). The product was diluted with milliQ water and partitioned in hexane using liquid-liquid extraction. The extract was further separated by eluting through a silica gel column using hexane and DCM, with the DCM fraction carrying the FAMEs.
Samples were dissolved in hexane ready for compound identification and quantification using a gas chromatograph (Agilent 6890) coupled with a mass spectrometer (Agilent 5973) and flame ionization detector (GC-MS/FID). Compound identification was based on retention time and mass spectra of target peaks. Absolute abundance was calculated from peak area response on the FID, based on a calibration curve of an in-house standard mixture of n-alkanes and n-alkanoic acids of known abundance. We recorded the abundance of n-alkanes (C 23 -C 33 ) and n-alkanoic acids (C 22 -C 32 ), individual homologues conventionally considered terrestrial plant-derived (G. Eglinton & Hamilton, 1967), and calculated their total abundance on a µg g -1 dry weight basis (∑alk and ∑acid) as well as normalized to TOC, i.e., µg g OC -1 (Λalk and Λacid). To represent the molecular distributions of the plant wax homologues, we also calculated the average chain length (ACL) and carbon preference index (CPI) using the following equations:
where n indicates the chain length (n = 23 -33 for n-alkanes and n = 22 -32 for n-alkanoic acids), and [C n ] indicates the abundance of that chain length.
The compound-specific carbon and hydrogen isotopic compositions (δD and δ 13 C) were measured by gas chromatography -isotopic ratio mass spectrometry (GC-IRMS) using a Thermo Scientific Trace gas chromatograph connected to a Delta V Plus mass spectrometer via an Isolink pyrolysis furnace at 1400°C for δD, and a combustion furnace at 1000°C for δ 13 C. We monitored the linearity of isotopic determinations across 1-7 V peak amplitude daily and only accepted measurements from peaks with amplitude within the range of acceptable linearity. δD
Here we report total abundance of organic carbon (TOC, mass C per gram sediment), and plant diamond; soil: circle) of overlapping sites at WAY and SP (Feakins et al. 2016a,b;Feakins et al., 2018). Open shows exceptionally high C 23 and C 25 that is distinct from litter and soil distributions at this site (Fig. 3g).
n-Alkanoic acids exhibit an even-over-odd preference from C 22 -C 32 , with C 30 or C 32 being the dominant compound in litter, whereas soil shows a more 'flat' distribution across chain lengths owing to an increase in the abundance of mid-chains C 22-26 (Fig. 3). Similar to n-alkanes, we also observe a trend towards longer chain lengths (higher C 32 /C 30 ratio) towards lower-elevation sites.
Roots show distinctively different distributions with dominance by mid-chains in both sites (Fig. 3d,h).
Carbon preference index (CPI) and average chain length (ACL) calculations provide more quantitative comparisons of molecular distributions between samples (Fig. 4). We find that CPI of n-alkanes exhibits a wide range from ~4 -16, with decreasing values from litter to soil at all sites. Lower n-alkane CPI values are found in roots (CPI = 4.1-7.1) compared to litter (CPI = 6.6-16.1). In contrast, CPI of n-alkanoic acids shows a relatively invariant vertical profile among litter, roots, and soil, with only the top litter at WAY and the canopy leaves at SP being exceptions with slightly elevated CPI.
ACL of both n-alkanes and n-alkanoic acids shows a decrease (by c.1) from litter to soil at WAY, VC and LA, but a relatively straight profile at SP. Litter shows a trend towards higher ACL at lower-elevation sites. Roots exhibit lower ACL compared to litter and soil, related to high abundance of mid-chain length compounds (Fig. 3). Overall, the soil CPI and ACL results from this study match well with data from previous soil studies (Fig. 4
We report δD and δ 13 C values of C 27-31 odd-chain n-alkanes and C 22-32 even-chain n-alkanoic acids when reliable isotopic measurements could be made on these samples (Appendix A). We focus our attention on the most dominant chain length of each compound class, C 29 n-alkane and C 30 n-alkanoic acid. Although we do not show data from all the chain lengths on Fig. 5, the general isotopic patterns described below are shared among the homologues of each compound class, as reported in Appendix A. We also report bulk δ 13 C OC values to compare with the plant wax data.
The hydrogen isotopic composition of C 29 n-alkane (δD 29alk ) and C 30 n-alkanoic acid (δD 30acid )
ranges from -173 to -210‰ and -158 to -207‰ respectively across all sites, with a general trend towards more enriched values at lower-elevation sites (Fig. 5). C 30 n-alkanoic acid is generally about 5 -20‰ enriched relative to C 29 n-alkane in the same samples. Both δD 29alk and δD 30acid
show relatively small (<20‰) variations within the soil profiles. However we see no systematic patterns in δD with depth across the four sites: a 5 -20‰ decreasing trend at WAY and VC slope base for C 29 n-alkane, and at WAY, SP, VC slope base and LA ridge top for C 30 n-alkanoic acid; a ~5‰ increase for C 30 n-alkanoic acid at LA slope base; a ~10‰ increase towards ~40cm depth followed by a 5‰ decrease below at VC ridgetop for both compounds; and no trend for C 29 n-alkane at SP and LA (Fig. 5). We find a much depleted δD 30acid value (-206‰) for the root sample from LA compared to soil at this site (ranging ~160 -170‰), though abundance of C 29 n-alkane was insufficient for δD analysis. For the root sample from SP (10 -20g mass), neither compound was sufficiently abundant for δD analysis.
The carbon isotopic compositions of C 29 n-alkane (δ 13 C 29alk ), C 30 n-alkanoic acid (δ 13 C 30acid ), and bulk OC (δ 13 C bulk ) range from -32.4 to -42.9‰, -31.5 to -40.4‰, and -24.5 to -33‰ respectively across all sites (Fig. 5). C 29 n-alkanes are depleted relative to C 30 n-alkanoic acids (by ~2‰) which are in turn depleted by ~6‰ from bulk OC in the same samples. We find consistent patterns of δ 13 C with depth for bulk OC and both plant wax compounds across all four sites: a trend of c. 4 -6‰ enrichment from litter to soil at depth, which is a combination of c. 2 -4‰ enrichment between litter and top-layer soil, and c. 2‰ gradual enrichment down the soil profile (Fig. 5). Roots yield δ 13 C values that are similar to litter or top-layer soil but more depleted than soils at depth (Fig. 5f,h). Across the elevation transect, we find a general trend towards more depleted δ 13 C values in lower-elevation sites.
When comparing soil data from this study (Fig. 5, squares) to that at WAY and SP from previous studies (Fig. 5, circles; data from Feakins et al., 2018) in which isotopic data were measured at two depth ranges (organic and mineral horizons), we find results are consistent, with both sets of data showing no systematic patterns in δD with depth, and a 13 C-enrichment in the deeper soil.
We find the vertical profiles of δD (no consistent trend) and δ 13 C (deeper layers are more enriched) are consistent among all chain lengths of n-alkanes and n-alkanoic acids (Appendix A).
We contrasted slope base and ridgetop settings by digging two soil pits each at VC and LA, to reveal possible difference at locations that are less well-drained with thicker O-layer (slope base)
and more well-drained with thin O-layer (ridgetop). While n-alkane δD at LA has limited data to allow comparison, we find that in general, the ridgetop shows enrichment in both H (by ~10-20‰) and C (by ~1-3‰) isotopes relative to slope base. At VC, while the uppermost soil samples show similar δD and δ 13 C values, the ridgetop location appears to be more enriched in both isotopes at deeper depths. At LA, there is consistently isotopic enrichment from the top soil to deeper soil.
Numerous plant-based surveys have characterized how plant waxes record environmental variables, providing a basis for interpreting the chemical fingerprints in these biomarkers as climate proxies in sedimentary archives. But an unresolved question relates to possible changes in plant wax signatures between plant and sediment which may compromise the environmental information they carry. In this section, we evaluate the changes observed in abundance, molecular distributions, and isotopic compositions of plant wax from litter to soil in the Peru transect studied here, and we discuss the processes that may lead to these changes.
Sampling of thick leaf litter accumulations at WAY and SP reveals substantial loss of plant waxes within the leaf litter, in contrast to a limited OC loss (~2 -5%). We find a decrease in concentrations (in terms of both per gram dry weight and OC-normalized) by ~77 -87% for nalkanes and ~10 -45% for n-alkanoic acids between top litter (large litter) and bottom litter (litter debris) at both sites. We note that there is an increase in ∑acid in the middle litter layer at WAY, which may imply new additions (perhaps by microbial productions during litter diagenesis) or simply heterogeneity within the coarse debris. The overall significant decrease in n-alkane abundance within litter suggests rapid degradation of these molecules during early diagenesis. Such rapid loss via degradation has also been observed from litterbag experiments that show >80% loss of plant waxes within 1-3 years, as a result of microbial degradation and perhaps also consumption by herbivores such as mesofauna (Zech et al., 2011;Li et al., 2017;Nguyen Tu et al., 2011, 2017). Our field data corroborate these experimental observations.
Although both compound classes lose concentrations within litter, there appears a better preservation for n-alkanoic acids, as shown by an increase of their abundance relative to nalkanes from top (large) to lower (small) litter at WAY and SP (Fig. 2i-l). The better preservation for n-alkanoic acids is also evidenced by generally higher litter-to-soil Λacid in contrast to the decrease in Λalk, indicating a greater portion of n-alkanoic acids survive litter degradation and enter the soil (Fig. 2e-h
Below the litter layer, we find further significant drop in plant wax concentrations in soils with depth (by ~65 -99% from top soil to 50 cm depth for both compounds, similar to the decline in TOC by ~57 -90% from top to 50 cm; Fig. 2). Since the absolute concentrations at different sites significantly vary, in order to compare sites, we first calculated fractional concentrations relative to the top sample (within O horizon) at each site, and then characterized the rate of loss (k z ; depth-dependent decay rate) by fitting an exponential decay function. We find exponential loss in concentrations in soils with k z ranging from 2.1±0.9 to 6.2±1.4 m -1 for OC, from 2.3±0.6
to 16.1±1.2 m -1 for n-alkanes, and from 2.9±0.3 to 15.2±0.7 m -1 for n-alkanoic acids (Fig. 6).
In contrast to the greater loss of n-alkanes relative to n-alkanoic acids observed in litter as described in section 4.1.1, the two compound classes appear to drop in concentration with depth at the same rate (no significant difference in k z values) except at WAY where the k z value of nalkanes is about double that of n-alkanoic acids. This distinction between plant wax loss in litter and within soil profiles soil implies different mechanisms governing the resilience of plant waxes in litter vs. soils. Within soils, physical protection in soil aggregates and absorption to soil minerals is known to play an important role in the stability of soil organic matter (Schmidt et al., 2011). Previous soil studies have found turnover times for both n-alkanes and n-alkanoic acids also to be similar and on the order of several decades (Wiesenberg et al. 2004;Schmidt et al., 2011).
We cannot infer turnover times from the k z values calculated based on the decreases in concentration within depth in our soil profiles, because we lack chronological information for the soils in this study. Further, we note that the exponential decline in concentrations with depth observed in this study may be affected by downward mobilization in addition to decomposition, although downward transport would not be expected to produce a carbon isotope fractionation with depth. Overall, the exponential decrease in plant wax concentrations is probably determined by a combination of accumulation of plant wax inputs on the soil top, downward-transportation by mesofauna such as earthworms (Oades et al., 1993) and decomposition of plant waxes within the soil over time. Across the four sites, we observe up to six-fold difference in the rate of loss with soil depth, with increase in k z values from VC, to WAY, SP and LA (except for OC at LA). What determines the difference among sites? The tendency is for an increase in k z as elevation decreases and temperature increases, which is a known factor for determining rates of respiration. Across the same Andes-Amazon transect, a litterbag translocation experiment which tracked litter decomposition of 15 species over 1.2 yr (Salinas et al., 2010) found that while species type has a large influence on the decomposition rate (k), soil temperature stands out as the main control when averaging all species, such that a five-fold increase in the k value is observed from the high Andes (WAY at 3025 m, 11.1°C) to the lowland Amazon (Tambopata at 210 m, 23.9°C; similar elevation to LA site in this study). This temperature-sensitivity for litter decomposition rates (k; Salinas et al., 2010) is reflected in the different k z values of plant waxes at WAY, SP and LA (Fig. 7), which increase progressively from high to low elevation, implying that temperature may also be the main control on the rate of plant wax loss. The k z values of OC at WAY and SP also follow the same trend as plant waxes, but the apparent lower rate of OC loss at LA is surprising (Fig. 6a; Fig. 7). It is possible that the OC left in these deeper soils at LA is relatively recalcitrant, if the majority of labile OC have already been degraded near the top of the soil due to the warm temperature at this site. This change in recalcitrance would not be expected to be seen in the plant waxes, perhaps explaining with the trends in k z for waxes are more systematically related to temperature than bulk OK. Overall, we infer that temperature is a primary control on the rate of decline of plant wax concentration and that decomposition likely dominates the depth-decay of plant wax concentration profiles.
An exception to the overall temperature trend for the k z of the plant waxes is found at VC, which
shows much slower loss (lower k z ) than predicted for its elevation (Fig. 7). We note that while the other three sites are in primary tropical forests, VC is in a secondary forest previously logged for timber (Table 1), with secondary-growth bamboos dominating the canopy. Bamboos are known as one of the fastest growing plants on earth, with relatively slow litter decomposition rates (Liu et al., 2010) partly due to the abundance of phytoliths (Piperno and Pearsall, 1998), hence having significant implications for carbon accumulation and storage in soils (Zhou et al., 2005). However, little plant wax research has been done on bamboo leaves (Li et al., 2012(Li et al., , 2016)), and the preservation of plant waxes in soils of bamboo forests remains unknown. Here, we find a lower than expected k z which may imply a greater accumulation of soil organic matter in this presently bamboo-dominated forest, or some other aspect of the landscape disturbance, such as any use of fire which may add to soil organic carbon and slow decomposition. This exception is a useful reminder that while temperature may be a major control on the rate of organic carbon and plant wax loss in well-drained soils, other factors like waterlogging, disturbance and species succession may also modify soil organic properties. n-alkanes relative to total (Fig. 8), together with a decrease in CPI (Fig. 4), is apparent down profile in our study, consistent with the occurrence of microbial degradation of n-alkanes. The soil profiles suggest that microbial degradation affects plant waxes from the very earliest stages of diagenesis; for example the biggest drop in n-alkane CPI happens at the litter-soil interface (Fig. 4). Apart from degradation, microbial activities can also affect plant wax signatures by contributing n-alkanes and n-alkanoic acids to the soil pool. Though commonly assumed to be dominated by terrestrial vascular plant sources in paleo reconstructions, long-chain n-alkanes (>C 27 ) and n-alkanoic acids (>C 28 ) could also be produced by microbes as previous studies have shown (Nguyen Tu et al., 2011;Summons et al., 2013;Makou et al., 2018). It is possible that both degradation and addition of these compound classes may occur in soils, and this can be detected by examination of molecular abundance distributions. In a 1.5-year soil incubation experiment, researchers detected microbial production of long-chain (C 27-31 ) n-alkanes with an estimated turnover rate of ~0.1% per year (for n-C 29 ) under aerobic conditions, though no significant production was detected under anaerobic conditions (Li et al., 2018). n-Alkane-degrading microbes can convert these molecules into n-alkanoic acids following identified aerobic and anaerobic degradation pathways involving alkane hydroxylases (Ji et al., 2013), leading to accumulation of n-alkanoic acids relative to n-alkanes, which is favored in soils with low pH ~3.8 as observed in a soil experiment, whereas higher pH ~7.3 favors higher abundance of nalkanes (Bull et al., 2000). We note that soils throughout the Peruvian Andes-Amazon transect have low pH values ~4 (Whitaker et al., 2014), which based on these experimental results may enhance the accumulation of n-alkanoic acids relative to n-alkanes as we see in our soils (Fig. 2).
4.1.4 Are root and fungal contributions of plant waxes significant?
Root-derived organic carbon (OC) represents a significant source of soil organic matter, in part due to enhanced protection mechanisms of root-derived versus shoot-derived OC in soils, e.g. as root-hairs can burrow inside of soil aggregates providing physical protection of root-derived OC (Rasse et al., 2005). In terms of plant waxes, grass roots have been found to produce long-chain n-alkanes with high odd-over-even preference and C 31 dominance (Marseille et al., 1999), as well as n-alkanoic acids with a distinct chain length distribution (C 22-24 dominance) compared to leaves and stems (C 28 and C 30 dominance) (Wiesenberg et al., 2012). From a litter-soil profile in a grass-dominated landscape, Naafs et al. (2004) deduced substantial root input of lipids, including long-chain n-alkanes and n-alkanoic acids, into soils. It is unclear how much roots contribute to plant waxes in soils in natural tropical forests such as in our study area, as such data are lacking perhaps in part due to the difficulties in field collection and identification of entangled roots from diverse tree species, especially in the context of very high biodiversity in the western Amazon and Andes (Silman, 2014). However, given the anatomy of plants, rootderived OC is presumably less important in forests than in grasslands (Oades, 1993), given the higher above-ground biomass of trees relative to grasses.
Although we only have studied two available root samples from SP and LA, the results provide some clues to whether root inputs of n-alkanes and n-alkanoic acids are important in these soils.
These two roots show very low plant wax abundance, with <1% n-alkane and 6-16% n-alkanoic acid concentrations compared to litter (Fig. 2b,d). Given the lower net primary productivity allocated to roots than canopy across the same Andean transect (Malhi et al., 2016), a low plant wax concentration in roots suggests a tiny root contribution to soil on a biomass basis, although these may be overrepresented given the greater preservation potential previously noted. Another line of evidence comes from the molecular distributions. If plant waxes in soils are mainly derived from roots, we would expect to see molecular distributions in soils that are more similar to roots compared to leaf litter. While the n-alkane molecular distributions are similar between litter and roots at SP (Fig. 3c), confounding any separation on this basis, the molecular distributions of n-alkanes at LA (Fig. 3g) and n-alkanoic acids at both SP and LA (Fig. 3d,h) are distinct in roots versus litter. In general, we do not find evidence of significant root inputs of nalkanes and n-alkanoic acids, as soils show more similar molecular distributions to that of litter (C 29 and C 31 n-alkane, and C 30 and C 32 n-alkanoic acid dominance), and lack the distinct signatures shown in roots (C 23 and C 25 n-alkane dominance at LA, C 22-26 n-alkanoic acid dominance with low even-over-odd preference in that range). Moreover, the one available δD measurement of root n-alkanoic acid at LA is significantly D-depleted (by ~40‰) relative to the adjacent soil profile (Fig. 5d), further supporting a minor influence of root inputs to soil plant waxes.
Fungi have been reported to contain n-alkanes often with C 27 , C 29 and C 31 dominance similar to that of vascular plants (Weete, 1972), but not long-chain (>C 28 ) n-alkanoic acids (Weete, 1972;Madan et al., 2002). Other studies have found increases in C 25 and C 27 n-alkanes in sub-surface horizons and attributed these to fungal production (Huang et al., 1996;Marseille et al., 1999). It may be difficult to use n-alkane molecular distributions to detect fungal inputs given the wide diversity of fungi, poor characterization of n-alkane production (only few species characterized), and possible confounded distributions with vascular plants (Weete, 1972). A sub-surface increase in abundance of total n-alkanes or particular chain lengths such as C 27 is expected if fungal input is substantial as in previous reports (Huang et al., 1996;Marseille et al., 1999), and we do not observe any such feature in our profiles. However, we note that in those prior fungal studies, the sub-surface increase in fungal n-alkanes occurs within a discrete, thin layer a few to ten centimeters down from soil tops, perhaps guided by visual evidence during sampling. If any fungal n-alkane production happens at such shallow depth within our soils, this production would not be observed by our relatively coarse profile sampling, and no fungal evidence was observed in the field.
The hydrogen isotopic compositions of both C 29 n-alkane and C 30 n-alkanoic acid (the dominant chain length of each compound class) show minor variations (<20‰) down profile with no systematic pattern observed across sites (Fig. 5). The lack of systematic trend in plant wax δD values within the litter layer and soil profiles in this study means we have no evidence for any isotopic effect, whether via new inputs or below ground processes such as degradation and remobilization, during soil formation. One possibility is that downward-transport of plant waxes (such as by mesofauna), may have homogenized plant wax characteristics; however the different patterns of δD and δ 13 C values with depth, measured on the same molecules, do not support mixing as a major process for these soils. Concentration data have been interpreted as indicating microbial decomposition within litter and soil during the timescales of soil formation. As we do not find a systematic change in δD values, we infer no evidence for any consistent hydrogen isotope fractionation effects associated with early diagenesis here.
This interpretation is consistent with a 27-month litterbag degradation study conducted on three higher plant species in a German spruce forest (Zech et al., 2011). In that study, researchers found no overall trend in C 27-31 n-alkane δD over the course of the study. They suggested minor (~10 -20‰) fluctuations were linked to seasonal variations of soil water δD on the microbial community, but no systematic change in n-alkane δD was observed in their 2-year litterbag experiment. Our study extends from leaf litter to consider the soil profile and finds that there is no change in plant wax δD during the timescale of soil formation in this system.
In contrast, a leaf-litter-soil profile in a maple forest in Japan (Chikaraishi & Naraoka, 2006) found D-depletion by 33 -77‰ for both compound classes. Most of that D-depletion occurred between canopy and leaves on the ground (litter), whereas the D-depletion within the litter-soil profile was only ~5 -20‰. We find no such systematic directional change, and at SP, we observe that litter is 5 -15‰ depleted for C 29 n-alkane, but 20 -30‰ enriched for C 30 nalkanoic acid relative to average canopy (Fig. 5b). Although it is hard to reconcile the different findings in a Japanese temperate maple forest dominated by just two species (Acer argutum and Acer carpinifolium) and the Peruvian tropical high biodiversity forest sites spanning an altitude range (this study), one possibility is that there has been a directional change in the hydroclimate at the Japanese location during the time of soil formation. Another possibility is that high biodiversity at our Peruvian sites masks any diagenetic changes, with variability down-profile driven primarily by different species inputs over time. Future work might study the isotope effects down-profile in a wider range of soil types and ecosystems in order to better constrain plant wax δD values in soil archives.
4.1.6 A systematic shift in plant wax δ 13 C across between canopy, litter and soil
This study was motivated by the observation of plant wax δ 13 C offsets between canopy leaves and soils (Feakins et al., 2018). In the current detailed study of leaf litter and soil profiles we confirm that offset and study the progression via more detailed sampling within soil pits. We find a 4 -6‰ enrichment in both plant wax compounds from litter to deeper soils (Fig. 5). The larger enrichment step happens between the litter and top soil (~2 -4‰) followed by a smaller change (~2‰) deeper in the soil, and the profiles in plant waxes mirror that of bulk OC. The 13 Cdepletion in litter relative to canopy leaves at SP (~1‰ for C 29 n-alkane and ~3‰ for C 30 nalkanoic acid, Fig. 5f) may indicate the addition of relatively 13 C-depleted understory leaves (Wu et al., 2017). Up to 2‰ of the down-profile 13 C-enrichment may be explained by the more enriched pre-industrial atmospheric CO 2 compared to today due to the Suess effect (Francey et al., 2002; Scripps CO 2 program), if the plant waxes in the deeper soils were entirely preindustrial. Root inputs cannot explain the enrichment in soils, as the roots are 2 -4‰ depleted relative to soils (Fig. 5f,h). Plant wax δ 13 C entering the soils could have shifted through time if there was a directional change in plant type/composition over the timescale of decades; however this is very unlikely in these pristine highly biodiverse tropical forests (except VC) where no single tree species dominate the landscape. Hence, after accounting for Suess effect, we infer at least ~2 -4‰ of post-mortem 13 C-enrichment of plant waxes within soils which is likely a result of diagenesis.
Our profiles corroborate previous studies of a leaf-litter-soil sequence in a Japanese maple forest that found ~2.5 -4‰ 13 C-enrichment between leaf and surface soil (Chikaraishi and Naraoka, 2006), and depth profiles of three types of tundra-covered British acid upland soils that found ~2 -4‰ 13 C-enrichment of C 27-31 n-alkanes downwards (Huang et al., 1996). Considering very different settings among the three studies (and the continued Suess effect over recent decades), the 13 C-enrichment of plant waxes during soil storage appears to be a common feature across a range of environmental and soil conditions. 13 C-enrichment of n-alkanes during early diagenesis has also been observed in several litterbag experiments in mid-latitude temperate forests, mostly
showing 1 -2‰ enrichment within 1 -3 years, likely a result of microbial processes (Nguyen Tu et al., 2004;Wang et al., 2014;Li et al., 2017;Zhang et al., 2017). Our study confirms that direction and trend, and reveals additional change down profile, which could be due to the much longer time scale of soil formation compared to the litterbag experiments. Not only do we find this result for the n-alkanes, we also confirm this transition in litter and soil profiles for the nalkanoic acids, which have been less often reported in litterbag degradation experiments.
4.2 Implications for plant wax calibration studies for paleoclimate applications
Much attention for modern plant wax studies has focused on leaves from living plants (as summarized in review papers by Sachse et al., 2012;Diefendorf & Freimuth, 2017), but soilbased studies (e.g. Jia et al., 2008) provide integrated records of multi-species plant inputs and post-mortem soil processes that may affect plant wax signatures in the transition from leaf to soil (Nguyen Tu et al., 2004;Chikaraishi & Naraoka, 2006). Several studies have surveyed soils across environmental transects using soils to understand molecular abundance distribution (Bush & McInerney, 2015), carbon isotopic composition (Wei & Jia, 2009;Schwab et al., 2015;Feakins et al., 2018) and hydrogen isotopic composition (e.g. Jia et al., 2008;Bai et al., 2011;Zhang & Liu, 2011;Ernst et al., 2013;Ponton et al., 2014;Zhuang et al., 2015;Nieto-Moreno et al., 2016;Wang et al., 2017;Feakins et al., 2018), almost all of which studied n-alkanes, with only few exceptions that have studied n-alkanoic acids (Ponton et al., 2014;Feakins et al., 2018;Bakkelund et al., 2018). Although both plant and soil-based approaches have merits, plant-based calibrations include significant scatter among individual plants associated with differences in plant type, species, biosynthetic processes, seasonality and microclimate, whereas soils provide an average of plant inputs and reveal how environmental controls are represented in the soil archive. For example, along a slope of Mount Taibai in China, soil n-alkane δD values capture the altitudinal gradient in source water composition which was not observed in plant measurements due to significant scatter among individuals, especially between woody plants and grasses (Zhang & Liu, 2011).
Regions of high biodiversity, such as tropical forests, pose even bigger challenges for plantbased calibrations, as large-quantity sampling and knowledge of species dominance may be required to adequately capture the ecosystem-scale average signatures. Recent surveys of plant wax δD and δ 13 C in canopy leaves in the same region as this study, along a 3320 m elevation transect in the highly-biodiverse tropical forests of the Peruvian Andes, sampled at an unprecedented scale (>300 samples) and revealed significant scatter among individual tree leaves and species. Despite the scatter these studies could identify a robust altitudinal trend (Feakins et al., 2016a,b;Wu et al., 2017), but one that would have been difficult to reveal without substantial sampling of leaves and sites as demonstrated by Monte Carlo simulations (Wu et al., 2017). In contrast, a relatively small number of soil samples may be needed to calibrate the archived proxy across an environmental transect (e.g., as shown for this region in Ponton et al., 2014 andFeakins et al., 2018). In a series of studies in this region, we have both constrained the isotopic signal fixed in the plant canopy (Feakins et al., 2016a,b;Wu et al., 2017), the processes of alteration down profile (this study) and the archived proxy in soils (Ponton et al., 2014;Feakins et al., 2018).
Another advantage of soil-based calibrations is that these capture the post-mortem alterations to plant wax signatures during residence in soil, which may modify the environmental information being recorded from time of synthesis. While we find that δD does not systematically vary, we find a 4 -6‰ 13 C-enrichment in both plant wax compounds from litter to deeper soils (Fig. 5).
The larger enrichment step happens between the litter and top soil (~2 -4‰) followed by a smaller change (~2‰) deeper in the soil, and the profiles in plant waxes mirror that of bulk OC.
Knowing this, what are the implications for application of plant-based calibrations to the sedimentary record? Corrections for the changing δ 13 C values of atmospheric CO 2 (Suess effect) based on the year of plant collection can be readily applied, when using modern plant-based calibrations for interpretations of the pre-industrial geologic record. Corrections associated with diagenetic processes in soils will be harder to quantify and we anticipate that the diagenetic effect will likely vary with climate, soil type and microbial community. Based on this study of tropical forests, a 2 -4‰ diagenetic correction may be relevant for vegetation reconstructions in tropical settings based on soils, paleosols, and sedimentary archives where plant waxes are mainly derived from pre-aged plant waxes that have been diagenetically altered in soils. In contrast, archives that mainly integrate leaves that did not go through soil storage (e.g. swamps, some lakes) may not experience such diagenetic effects.
Without a diagenetic correction for soil-stored plant waxes, the reconstruction of vegetation composition using soil-derived plant wax δ 13 C may be subject to bias relative to calibrations based on living plants. For example, a common tropical application of carbon isotopic analyses is to estimate the proportion of plants using the C4 pathway (e.g., Schefuß et al., 2003, Castaneda et al., 2009). Based on our C3 tropical forest soils, a 3‰ post-mortem diagenetic enrichment would lead to a ~20% overestimation of C4 coverage (based on a 14‰ difference between C3 and C4 end-members in Cerling et al., 1997) We posit that soils are the dominant plant wax stock relative to plant biomass in these tropical forest ecosystems. This inference comes from our estimates of the stock of plant waxes in plants vs soils (Appendix A), explained as follows and calculated along a series of tropical rainforest (TR) and tropical montane cloud forest (TMCF) sites across the same Andes-Amazon transect that were previously studied for plant wax work (Ponton et al., 2014;Feakins et al., 2016a,b;Wu et al., 2017) and are analogous to those studied in more detailed here.
To estimate plant wax stock in the leaves of living trees, we take the OC-normalized plant wax concentration data from prior studies (Feakins et al., 2016a,b) We estimate plant wax stock in soils for the top 30 cm of soils only, using a two-layer (organic and mineral layer) approach, based on OC-normalized plant wax concentration data (Feakins et al., 2018) and soil OC stock estimates (Girardin et al., 2014). We find an estimated 0.6 -3.9 Mg km -2 for n-alkanes and 3.2 -21.5 Mg km -2 for n-alkanoic acids for the top 30 cm of soils again with a tendency towards bigger stocks in the TMCF. Together these results show a much bigger plant wax stock in soils compared to canopy, which accounts for only 17% and 23% of total nalkane stock, and 1% and 5% of total n-alkanoic acid stock, on average for TR and TMCF sites respectively (Fig. 9). from soil organic and mineral layers (Feakins et al., 2018) and OC stock estimates for the top 30cm of soils (Girardin et al., 2014). The average of TR and TMCF sites is shown in the summary, with numbers in green indicating the fraction of plant wax stock allocated to the leaves. Crosses denote where leaf or soil estimate is unavailable for the site. Readers are referred to previous publications (Wu et al., 2017, Feakins et al., 2018) for site information.
While these estimates are subject to caveats such as not accounting for non-leaf plant waxes in living biomass, and simplification of the two-layer model for soil (top 30 cm) estimates, they depict an overall picture that the vast majority of terrestrial plant waxes is stored in the soils rather than in the living biomass. Moreover, the soil top 30 cm stock represents an underestimation of the overall soil stock, especially in the higher-elevation sites where the organic layers are deeper (Table 1) and plant wax loss with depth is gentler (Fig. 7). Although it has long been known that soils are important archives of OC (Blair et al., 2004), the effort to quantify plant wax production (Feakins et al., 2016a) and here to quantify and compare plant waxes stocks above and below ground, is a new contribution. It would be interesting for carbon cycle quantification and tracking to see this biomarker approach to stocks and fluxes expanded to more climates and ecosystems.
Given the much greater stock of plant waxes in soils relative to living plants (Fig. 9), soils are the likely source for the majority of riverine-erosion and export of plant waxes used in studies of catchment sourcing (e.g. Ponton et al., 2014;Häggi et al., 2016;Hemingway et al., 2016), and thus for the plant waxes deposited downstream in sedimentary repositories used for paleoclimate reconstructions (e.g., Tipple and Pagani, 2010;Schefuß et al., 2011;Hein et al., 2017). We acknowledge that the actual sourcing from plants vs soils depends not just on the stock, but also on the erosional processes. For example, it has been suggested that landsliding plays an important role in soil OC sourcing to rivers in the Peruvian Andes, stripping 80% of the OC from soils and 20% from vegetation (Clark et al., 2016). Landslides would enhance supply of soil plant waxes from deeper depths (beyond 30 cm), as well as the plant waxes directly from the living biomass. While estimating the exact living vegetation vs soil plant wax sourcing is beyond the scope of this study, we suggest that the stock estimates gives us a first-order view on the relative importance of these two pools, such that soil-based calibrations carry merit of likely being the pool from which most riverine plant waxes are sourced. In terms of parsing fluvial sourcing proportions between living plants and soil stocks of plant wax, compound specific radiocarbon analysis is needed (French et al., 2018).
The idea that soil is the major source of sedimentary and riverine plant waxes has important implications especially for elevation-sourcing studies. For example, Feakins et al. (2018) evaluated fluvial sourcing of plant waxes within the Andes-Amazon Madre de Dios catchment based on plant wax isotopic gradients in soils. If the δ 13 C gradient in canopy leaves (which is c. -1 and -2‰ offset from soil organic and mineral layer respectively) were to be used instead, this would result in an overestimate of the average sourcing elevation by more than 700 m (taking c.1.5‰ km -1 altitudinal gradient for C 29 n-alkane; Feakins et al., 2018). The degree of litter-tosoil 13 C-enrichment, however, appears similar across the four sites that span a range in elevation (286 -3025 m), forest structure (from montane cloud forests to lowland tropical rain forests), and soil organic content (soil organic layer 1 -26 cm thick, thinner towards lower elevation), such that the δ 13 C altitudinal gradients are kept nearly constant between plant and soil, though their values may be offset (Feakins et al., 2018). This implies that when relative isotopic changes are interpreted (e.g., as relative shifts in C3/C4 coverage, or when applying δ 13 C in plant waxes for paleoaltimetry reconstruction) instead of the absolute values, the problem caused by 13 Cenrichment within soils may be avoided.
Here we studied plant wax biomarkers (n-alkanes and n-alkanoic acids) from four locations (six soil pits) along a 2740 m elevation transect in the Andes-Amazon. We measured plant wax concentrations, molecular distributions, and hydrogen and carbon isotopic compositions, as well as bulk organic carbon and carbon isotopic composition, in these litter-soil profiles. Based on the observations within these profiles we draw inferences about inputs, degradation processes and alteration of plant wax properties within these tropical soil profiles. Within leaf litter, although both compound classes decline in absolute abundance, we find that n-alkanes are lost relative to n-alkanoic acids, and it is this greater loss of n-alkanes (rather than in-soil inputs of n-alkanoic acids) which leads to an increase in n-alkanoic acid relative abundance from litter to soil. Within the soil profiles, concentrations of both compound classes decline with depth (k z ) at similar rates within a site. Between sites, k z decreases with elevation, such that k z is smaller at higher elevation (colder) sites and larger at lowland (warmer) sites. The only exception to this trend is at VC, the only soil sampled under a secondary growth forest, now dominated by bamboo, and aspects of the disturbance history or bamboo regrowth may explain the lower than expected k z .
We find signs of microbial activities altering molecular distributions of plant waxes, but no evidence of root and fungal contributions being quantitatively important. Across the litter-soil profiles, we find no systematic change in δD values, but a consistent 4 -6‰ increase in δ 13 C down-profile, which is attributed to a combination of Suess effect (≤2‰) and diagenetic processes (2 -4‰), corroborating results from previous litter degradation experiments. With these observations, we suggest that soil-based calibrations carry considerable merit as integrated recorder of plant signals, and this approach is especially relevant in high biodiversity ecosystems where it reduces the number of samples needed to adequately characterize the system. Further, soil-based calibrations capture the post-mortem diagenetic processes that affect plant wax. It is important to characterize plant waxes in soils for a range of applications. Most obviously, surveying plant waxes within modern soil profiles is important for calibration of the recorded signals that may inform applications to paleosol archives of the plant wax proxy. As we show the below-ground stock of plant wax is much greater than that of the living forest here, further quantification and characterization of the soil stock of plant waxes in a range of environments and ecosystems would be informative for carbon cycle and sourcing studies. While not all soils are connected to fluvial systems, some soils are episodically eroded in this system by landslides in high relief areas and by migrating river meanders in lowland systems. Overall, river studies of inverted triangle; soil: square), as well as from previous studies (canopy: diamond; soil: circle) of overlapping sites at WAY and SP (Feakins et al. 2016a,b;Feakins et al., 2018). Open symbols at LA and VC denote additional pits at these sites at the ridge top, with closed symbols representing slope base. Vertical bars indicate the depth range from which the soil profile samples were taken.
Horizontal bars of canopy data at SP represent standard error of the site means (n = 39). Note the change in x-axes for soil data on the left panels. showing data from this study (litter: triangle; root: inverted triangle; soil: square). Open symbols at LA and VC denote additional pits at these sites at the ridge top, with closed symbols representing slope base. Also shown are canopy (diamond) and soil (circle) data from previous studies of overlapping sites at WAY and SP (Feakins et al. 2016a,b;Feakins et al., 2018).
Vertical bars indicate the depth range from which the soil profile samples were taken. Horizontal error bars of canopy data at SP represent standard error of the site means (n = 39). shown are canopy (diamond) and soil (circle) data from previous studies of overlapping sites at WAY and SP (Feakins et al. 2016a;Wu et al. 2017;Feakins et al., 2018)