Chemically stabilized soil organic carbon fractions in a reclaimed minesoil chronosequence: implications for soil carbon sequestration

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    Chemically stabilized soil organic carbon fractions in a reclaimedminesoil chronosequence: implications for soil carbonsequestration

    Sriroop Chaudhuri Louis M. McDonald

    Eugenia M. Pena-Yewtukhiw Jeff Skousen

    Mimi Roy

    Received: 21 March 2012 / Accepted: 15 January 2013

    Springer-Verlag Berlin Heidelberg 2013

    Abstract With adoption of appropriate reclamation

    strategies, minesoils can sequester significant amount of

    soil organic carbon (SOC). The objective of this study was

    to isolate different SOC fractions and coal-C in a reclaimed

    minesoil chronosequence and assess effects of increasing

    time since reclamation on each SOC fraction and selected

    soil properties. The chronosequence was comprised of four

    minesoils with time since reclamation ranging between 2

    and 22 years. Total SOC (TSOC, summation of all SOC

    fractions), ranged between 20 and 8 g kg-1, respectively,

    at the oldest (Mylan Park) and youngest (WVO1) minesite,

    indicating increasing SOC sequestration along the chron-

    osequence. The humin fraction accounted for about 43 and

    7 % of TSOC, respectively, at Mylan Park and WVO1,

    indicating increasing humification and biochemical stabil-

    ization of SOC with increasing time since reclamation. At

    WVO1,[60 % of TSOC was apportioned among the acid-hydrolysable (labile) and mineral-bound SOC fractions.

    Total soil carbon (TSC, TSOC ? coal-C) were signifi-

    cantly (p \ 0.05) related to the humin fraction in olderminesoils, whereas with the acid-hydrolysable (labile)

    fraction in the younger minesoils indicating that C stabil-

    ization mechanisms differed substantially along the

    chronosequence. Coal-C was unrelated to any SOC fraction

    at all minesites indicating that SOC sequestration estima-

    tions in this chronosequence was unaffected by coal-C. Soil

    cation exchange capacity and electrical conductivity were

    significantly (p \ 0.05) related to the humin fraction atMylan Park while to the acid-hydrolysable and mineral-

    bound SOC fractions at WVO1 indicating that the relative

    influences of different SOC fractions on soil quality indi-

    cators differed substantially along the chronosequence.

    Keywords Soil organic carbon sequestration Reclaimedminesoil Humin Coal Sequential fractionation Cationexchange capacity


    AHSOC 65 % HNO3 Hydrolysable soil organic carbon

    ARSC Residual soil carbon after 65 % HNO3hydrolysis

    BHSOC Base (0.5 M NaOH) hydrolysable soil organic


    CEC Cation exchange capacity

    EC Electrical conductivity

    FBHSOC Final base (0.5 M NaOH) hydrolysable soil

    organic carbon

    FBRSC Residual soil carbon after final base hydrolysis

    HFSOC 10 % HF Hydrolysable soil organic carbon

    HFRSC Residual soil carbon after 10 % HF hydrolysis

    RSC Residual soil carbon after each step of

    sequential fractionation

    SOC Soil organic carbon

    SOM Soil organic matter

    TSC Total soil carbon

    TSOC Total soil organic carbon

    S. Chaudhuri (&)Texas A&M AgriLife Research and Extension Center,

    P.O. Box 1658, Vernon, TX 76385, USA


    L. M. McDonald E. M. Pena-Yewtukhiw J. SkousenDivision of Plant and Soil Sciences, West Virginia University,

    1102 Agricultural Sciences Building, P.O. Box 6108,

    Morgantown, WV 26506-6108, USA

    M. Roy

    Department of Agronomy and Soils, Auburn University,

    Auburn, AL 36849, USA


    Environ Earth Sci

    DOI 10.1007/s12665-013-2256-8

  • Introduction

    Surface mining for coal in the Appalachian region leads to

    drastic perturbation of soil systems and alters ecosystem

    processes (Shrestha and Lal 2007; Chatterjee et al. 2009).

    Environmentally adverse consequences of mining include

    degradation of hydrologic systems and loss of aquatic

    habitat, destruction of soil structure, increased soil erosion,

    loss of soil organic matter (SOM), stunted plant growth,

    and restricted nutrient cycling (Johnson and Skousen 1995;

    Lal and Ussiri 2005). However, with the adoption of

    appropriate reclamation strategies, minesoils can sequester

    significant amounts of soil organic carbon (SOC) and

    contribute to terrestrial C sequestration efforts (Akala and

    Lal 2000, 2001; Lal 2004; Shukla et al. 2004, 2005;

    Shrestha and Lal 2006; Ussiri et al. 2006). Researches have

    shown that within 2050 years of reclamation, SOC can

    increase by 1050 % with substantial improvement in

    overall soil quality (Shukla et al. 2004; Ussiri et al. 2006;

    Shrestha and Lal 2007; Chatterjee et al. 2009). Significance

    of reclaimed minesoils in terrestrial greenhouse gas miti-

    gation effort was shown by a recent estimate that indicated

    that US minelands can sequester up 0.501.00 MgC

    ha-1 year-1 through reclamation which in turn could

    incorporate about 1.603.20 TgC year-1 into the soils and

    counterbalance about 5.811.2 Tg CO2 year-1 emerging

    from coal combustion (Lal 2004). Accumulation of SOC also

    has beneficiary effects on several agro-ecological and envi-

    ronmental processes (Stevenson 1994; Trumbore 1997).

    Soil organic carbon (SOC) sequestration ensues from

    SOC stabilization via: (i) soil aggregation (Six et al. 2004),

    (ii) organo-mineral complex formation (Guggenberger and

    Kaiser 2003) and (iii) inherent biochemical recalcitrance of

    SOC (Tan et al. 2004; Paul et al. 2006). A variety of

    chemical, isotopic, and molecular techniques have been

    used to elucidate SOC stabilization patterns. Chemical

    methods mainly include acid hydrolytic (Paul et al. 2006;

    Helfrich et al. 2007), oxidative (Eusterhues et al. 2003;

    Helfrich et al. 2007), and demineralization techniques

    (Eusterhues et al. 2007; Helfrich et al. 2007) to isolate

    different SOC fractions. Isotopic methods (14C) distinguish

    between SOC fractions based on their mean residence

    times (MRT) and turnover rates (Trumbore 2000; Bruun

    et al. 2005). Molecular methods rely upon the structural

    and compositional differences among the organic moieties

    associated with SOM (Kogel-Knabner 2000; Fontain et al.

    2007; Solomon et al. 2005, 2007).

    Owing to extreme biochemical heterogeneity of SOM,

    fractions isolated by different extractants, however, are

    considered operationally defined (Herbert and Bertsch

    1995). The dimensions and characteristics of SOC fractions

    vary widely depending on soil physico-chemical charac-

    teristics, biological processes, geographic location, climate,

    and land management practices. For example, base

    hydrolysis with 0.5 M NaOH fractionation can isolate up to

    80 % of SOM, with higher fractionation efficiencies gen-

    erally achieved for coarser textural classes (Stevenson

    1994). Helfrich et al. (2007) found that demineralization

    (10 % HF) followed by oxidation (sodium hypochlorite,

    NaOCl) can isolate about 4060 % of SOC. The oxidisable

    SOC fraction can account for as much as 72 % of total

    SOC (Siregar et al. 2005). Lee et al. (2009) found that

    organo-mineral association stabilized [70 % of total SOCwhile soil aggregation stabilized about 817 % of SOC.

    Zimmerman et al. (2007) showed that about 6391 and

    3566 % of total SOC, respectively, was isolated by oxi-

    dation with sodium hypochlorite (NaOCl) and acid

    hydrolysis (6 N HCl). Although no consensus exists

    regarding the amount of SOC that can be isolated by acid

    hydrolyses, in general, the acid-resistant (acid non-hydro-

    lysable, NHC) carbon constitutes about 3080 % of total

    SOC, depending upon soil type, texture, and land man-

    agement practices (Paul et al. 2006). The authors analyzed

    about 1,100 records obtained from literature and found that

    NHC accounted for about 48, 56, 55, and 56 % of total

    SOC, respectively, under conventional tillage, no tillage,

    forests, and grassland soils. Tan et al. (2004) found that

    NHC constituted about 53, 37, and 39 % of total SOC,

    respectively, in forest, conventional, and no till soils. Jen-

    kinson and Rayner (1977) have shown that NHC could

    represent 2050 % of total SOC in the upper horizons in

    the temperate zones. The NHC fraction in agricultural soils

    in the US Midwest accounted for about 3565 % of the

    total SOC and was about 1,3001,800 years older than the

    bulk soil (Leavitt et al. 1996; Paul et al. 1997, 2001).

    Unlike native soils, chemical isolation studies of different

    SOC fractions are relatively rare in reclaimed minesoils.

    Lorenz and Lal (2007) used oxidative (disodium perox-

    odisulphate, Na2S2O8) removal of carbon followed by

    demineralization (10 % HF) in minesoils reclaimed to

    forest and pasture ecosystem. Results indicated that the

    highest amounts of both SOC fractions were found at older

    minesites while the least at the youngest. However, the

    study did not account for coal-C and a rigorous chrono-

    sequence based approach was recommended to identify

    temporal changes in the dimensions of SOC fractions.

    Although reclaimed minesoils can sequester significant

    amount of SOC, quantification of SOC sequestration in

    these disturbed ecosystems, however, is challenged by

    presence of coal-C (Rumpel et al. 1998; Dick et al. 2006).

    Distinction between coal-C and SOC has been achieved by

    stable isotopic (d13C) (Ussiri and Lal 2007), radiocarbon(14C) (Rumpel et al. 2005; Ussiri and Lal 2007) and ther-

    mogravimetric (Siewert 2004; Maharaj et al. 2007a, b)

    methods. Such methods, however, are expensive and

    require significant amount of resources (Bruun et al. 2005).

    Environ Earth Sci


  • In addition, studies documenting the relative contribution

    of different SOC fractions to overall soil C budget and/or

    SOC dynamics and temporal changes therein are still

    lacking for reclaimed minesoils. The main objective of this

    study was to isolate and quantify different SOC fractions in

    a reclaimed minesoil chronosequence and assess their rel-

    ative effects on selected soil quality parameters. We used

    relatively inexpensive and easy-to-use chemical fraction-

    ation scheme to (i) isolate different SOC fractions and

    characterize temporal trends along a well-established

    chronosequence, (ii) quantify coal-C, and (iii) evaluate

    interrelationships between different SOC fractions and

    selected soil quality parameters such as soil cation

    exchange capacity (CEC) and electrical conductivity (EC).

    Materials and methods

    Study area

    A chronosequence comprising four reclaimed minesoil

    namely, Mylan Park (MP), New Hill (NH), WVSK, and

    WVO1, was identified in Monongalia County (393704500N,795702200W), West Virginia. Details of the minesoilcharacteristics and reclamation methods are described

    elsewhere (Chaudhuri et al. 2011, 2012a). The minesoils

    had similar soil forming conditions except for time since

    reclamation which ranged from 2 to 22 years along the

    chronosequence (Table 1). At the time of sampling mine-

    soil ages were 2, 4, 5 and 22 years for WV01, WVSK, New

    Hill, and Mylan Park, respectively. Initial reclamation

    work at WVSK was performed in the mid-1990s. However,

    in 2004, the topsoil (03 cm) was scraped and pushed back

    to the high wall situated along the rear margin which

    effectively established its time since reclamation as

    4 years. The minesoils were reclaimed to mixed grass

    legume pasture ecosystem. The predominant species were

    orchard grass (Dactylis glomerata), alfalfa (Medicago

    sativa), red clover (Trifolium pratense), white clover

    (Trifolium repens), timothy (Phleum pratense L.), tall

    fescue (Festuca arundinacea), and birds foot trefoil (Lotus

    corniculatus). The reclaimed sites were owned by the same

    company and were mined and reclaimed in similar manner

    (Table 1), following the protocols established by the 1977

    Surface Coal Mining Reclamation Act. Minesoils were

    compacted and graded to adjoining contours so as to merge

    uniformly with the regional topography. The regional

    geology consisted of interbedded, limy and acidic gray

    shale, siltstone, sandstone, coal, limestone, and thin beds of

    limy red shale. The region is characterized by temperate

    climate with average winter and summer temperatures

    around 0 and 22 C respectively. Thirty year averageannual precipitation amounts to about 104 cm.

    Soil sampling and analysis

    Each minesite was sampled over approximately 0.5 ha area

    in the early summer (MayJune) of 2008. Soil cores were

    collected from the upper 6 cm using site-specific irregular

    grids (7 9 5 m) based on the previous knowledge of spa-

    tial variability of SOC (Chaudhuri et al. 2011). A total of

    64, 83, 79, and 74 soil cores were collected from WVO1,

    WVSK, New Hill, and Mylan Park, respectively. Soils

    were air dried for 48 h, ground and passed through\2 mmsieve before performing chemical analysis.

    Before performing the SOC fractionation method, soil

    samples were treated with 1M HCl to remove inorganic

    carbonates (Midwood and Boutton 1998; Harris et al. 2000;

    Komada et al. 2008). In brief, 10 g of soil was shaken with

    40 mL of 1M HCL for 8 h (Midwood and Boutton 1998).

    Following acid treatment, the soil slurries were centrifuged

    and supernatants were decanted. Soil residues were rinsed

    twice with deionized water (DI), freeze-dried for 48 h, and

    gently ground to pass through \2 mm sieve. Soil carbonin the acid-treated and untreated soil was determined by

    complete combustion using an elemental analyzer unit. The

    difference between pre- and post-acid treatment yielded the

    amount of soil inorganic carbon. Residual carbon left in

    soil after inorganic carbonate removal represented organic

    carbon and was subjected to sequential fractionation using

    acid and base hydrolyses reaction followed by thermal

    oxidation to isolate different SOC fractions.

    The complete combustion was performed with LECO

    CNS-2000 analyzer, a non-dispersive, infrared, micro-

    computer-based facility designed to measure total carbon,

    nitrogen, and sulfur in solid samples (soil, plant tissue,

    fertilizers, meat products, dairy products, seeds, food, res-

    ins, and environmental wastes) with a nominal sample

    requirement (200 mg). The instrument has a detection limit

    of 0.02 mg C with a precision of about 0.4 % relative

    standard deviation (RSD). The combustion involves oxi-

    dation of SOM at 950 C, in the presence of ultra-high pure(UHP) helium, oxygen and low-moisture compressed air.

    Oxidation of SOM yields CO2 gas which is detected by an

    infrared sensor and expressed as %C on a soil dry weight

    basis. Before carbon analysis, 10 blanks and five desic-

    cated, pure primary standards were used to determine the

    calibration factor and correct for drift in SOC estimates.

    Quality control tests were performed by using (1) three

    replicates and (2) one each of a primary standard and blank

    after 15 unknowns (soil samples).

    The detail of the sequential fractionation method is

    available in Ussiri and Lal (2007). In brief, 2 g of car-

    bonate-free soil was subject to sequential treatment with

    0.5M NaOH (1:10 soil:solution ratio; 15 h shaking; twice),

    followed by 60 % HNO3 (1:10 soil:solution ratio shaking;

    15 min), 10 % HF (1:10 soil:solution ratio shaking; 4 h),

    Environ Earth Sci


  • 0.5M NaOH (1:10 soil:solution ratio; 30 min shaking;

    twice) and thermal oxidation in muffle furnace (340 C;3 h) (Fig. 1). Following each fractionation step, soil resi-

    dues were rinsed twice with DI, freeze-dried for 48 h, and

    gently ground to pass through \2 mm sieve. Residual soil

    carbon (RSC) in the soil residues after each fractionation

    step and after the final thermal oxidation was determined

    by complete combustion as outlined earlier. Hydrolysable

    soil organic carbon (HSOC) isolated at any particular

    fractionation step was determined by computing the dif-

    ference in the RSC between two successive steps as


    1. Base-hydrolysable fraction (BHSOC) = TSC RSC

    after base hydrolysis (BRSC).

    2. Acid-hydrolysable fraction (AHSOC) = BRSC RSC

    after acid hydrolysis (ARSC).

    3. HF-hydrolysable fraction (HFHSOC) = ARSC


    4. Final base-hydrolysable fraction (FBHSOC) =


    5. Coal-C = RSC after thermal oxidation.

    Where, BHSOC = Base (0.5 M NaOH) Hydrolysable soil

    organic carbon; AHSOC = 65 % HNO3 Hydrolysable soil

    organic carbon; HFSOC = 10 % HF hydrolysable carbon;

    FBHSOC = Final base (0.5 M NaOH) hydrolysable soil

    organic carbon; TSC = Total soil carbon; RSC = Resid-

    ual soil carbon after each step of sequential fractionation;

    ARSC = Residual soil carbon after 65 % HNO3 hydro-

    lysis; HFRSC = Residual soil carbon after 10 % HF

    hydrolysis; FBRSC = Residual soil carbon after final base


    Total soil organic carbon (TSOC) was determined by

    adding each HSOC fraction (BHSOC ? AHSOC ?

    HFHSOC ? FBHSOC). Each SOC fraction was presented

    as (1) absolute concentration (g kg-1) and (2) relative

    proportion (%). Relative proportions of each SOC fraction

    Table 1 General characteristics of the reclaimed minesites included in the study

    Minesoil characteristics WVO1 WVSK New Hill Mylan Park

    Time since reclamation (years) 2 4 5 22

    Coal type Waynesburg

    Mining method Contour mining, front end loaders

    Overburden 7080 % Sandstone, rest is shale

    Reclamation method Backfilled, 300 topsoil, gradedPost-reclamation land management practice Grass-legume pasture

    Soil C (g kg-1) 11.04 1.66c 11.96 3.84c 17.69 4.62b 21.73 5.97a

    Bulk density (Mg m-3) 2.01 0.33a 1.34 0.22c 1.27 0.18c 1.47 0.31b

    CEC (cmolc kg-1) 2.82 0.06d 4.75 0.07c 7.63 1.38b 11.78 2.35a

    EC (dS m-1) 0.05 0.001a 0.04 0.001b 0.04 0.001b 0.03 0.01c

    pH 6.68 0.11b 5.54 0.26c 6.67 0.72b 7.43 0.47a

    Clay (g kg-1) 17.7 2.52a 27.6 2.85a 16.3 2.69b 15.6 2.37c

    Sand (g kg-1) 29.5 4.63c 32.5 7.56b 36.7 5.27a 20.8 6.86d

    Samples collected 64 83 79 74

    Reported values for each soil property indicate mean standard deviation. Same letters for each soil property indicate no significant difference

    at a = 0.05

    Fig. 1 Sequential fractionation scheme to isolate SOC fractions andcoal-C (BHSOC base-hydrolysable SOC fraction, AHSOC Acid-hydro-lysable SOC fraction, HFHSOC 10 % HF-hydrolysable SOC fraction,FBHSOC final base-hydrolysable SOC fraction; TSOC = BHSOC ?AHSOC ? HFHSOC ? FBHSOC) (modified from Ussiri and Lal 2007)

    Environ Earth Sci


  • were expressed as percentages of the particular SOC

    fraction with respect to the TSOC. Total soil carbon (TSC)

    for each minesoil was determined by adding the TSOC

    (g kg-1) and coal-C (g kg-1).

    To test the effects of the chemical extractants on coal-C

    the entire fractionation scheme was applied on artificial

    coal:soil (w:w) mixture containing 100, 50, 10, and 5 %

    coal. No statistically significant (p \ 0.05) differenceswere observed in coal-C for any of the mixtures which

    indicated suitability of procedure in selectively extracting

    different soil C fractions without affecting coal-C.

    Soil C recovery by the fractionation scheme was

    assessed for each minesoil by computing the percentage of

    (TSOC ? coal-C), determined by the fractionation proce-

    dure with respect to the TSC (determined on un-fraction-

    ated whole soil). Soil-C recovery by this fractionation

    scheme ranged between 84 and 94 % among the minesoils

    (Table 2). Relatively better soil-C recovery was observed

    for the older minesoils. Lowest recovery was found at

    WVO1 (84 %) which probably was due lack of adequate

    time to initiate pedological processes that lead to accu-

    mulation of pedogenic C (SOC).

    Statistical analyses were performed with SAS v 9.1.3

    (SAS 1990). Individual SOC fractions were compared

    between minesoils by 1-way ANOVA using the MIXED

    model approach and Fishers least significant difference

    (LSD) at 95 % confidence level. Regression analyses were

    conducted to assess the influence of each SOC fraction on

    TSC and soil quality parameters.

    Results and discussion

    Distribution of total soil carbon (TSC; g kg-1)

    along the chronosequence

    Average TSC ranged from about 22 (Mylan Park) to

    11 g kg-1 (WVO1) and decreased significantly (p \ 0.05)along the chronosequence following the order Mylan

    Park [ New Hill [ WVSK & WVO1 (Fig. 2). AverageTSOC ranged from about 20 (Mylan Park) to 8 g kg-1

    (WVO1) and followed similar trend as TSC along the

    chronosequence indicating significantly (p \ 0.05) higherSOC sequestration in the older minesoils (Fig. 2). Time is a

    major soil forming factor and a key driver of all pedolog-

    ical processes. Owing to their well-defined ages (time since

    reclamation) reclaimed minesoils are suitable for deter-

    mining the rates and directions of changes in pedological

    processes, including changes in soil C and soil quality

    indicators (White et al. 2005). Overall, the results indicated

    a progressive SOC sequestration trend over time which

    corroborated with earlier findings (Chaudhuri et al. 2012a,

    b). Inorganic carbonates accounted for\4 % of TSC in thechronosequence, without statistically significant (p \ 0.05)differences among the minesoils which indicated that

    overall soil C budget in this chronosequence was dictated

    by changes in organic carbon (SOC and coal-C).

    Absolute concentrations (g kg-1) of SOC fractions

    along the chronosequence

    Average BHSOC fractions ranged between 4.9 (Mylan

    Park) and 2.3 g kg-1 (WVSK), decreasing significantly

    (p \ 0.05) following the order Mylan Park [ New Hill [WVO1 & WVSK along the chronosequence (Fig. 3a).Base hydrolysis (e.g. 0.5 M NaOH) removes the ester-

    bound biopolymers enriching the residual soil organic

    matter (SOM) with alkyl-C species, such as alkanols,

    alkanoic acids, hydroxyalkanoic acids (Nierop et al. 2003;

    Zegouagh et al. 2004; Rumpel et al. 2005). There are two

    main mechanisms involved in base hydrolysis reactions,

    (i) replacement of proton bridges within SOM by Na? ions,

    leading to stabilization and rearrangement of organic

    associations, and (ii) competition between the OH- groups

    of NaOH and other anionic moieties of the SOM for

    adsorption sites via pH-induced changes (Piccolo 2002).

    Table 2 Descriptive statistics for total C recovery (%) along thechronosequence

    Minesoil Mean Min Max

    Mylan Park (22 year) 93.04a 88.48 94.51

    New Hill (5 year) 89.98ab 81.01 92.07

    WVSK (4 year) 85.49b 78.10 96.69

    WVO1 (2 year) 83.77b 77.54 88.24

    Same letters indicate no significant difference at a = 0.05

    Fig. 2 Comparison of the average standard deviation values oftotal soil carbon (TSC, analyzed on the whole soil before sequential

    fractionation) and total soil organic carbon (TSOC, sum total of all

    hydrolysable SOC fractions) for each reclaimed minesoil across the

    chronosequence. Same letter for TSC and TSOC indicates nosignificant difference between the minesoils at a = 0.05. Years inthe parenthesis indicate time since reclamation. (MP Mylan Park,NH New Hill)

    Environ Earth Sci


  • Average acid-hydrolysable fraction (AHSOC) ranged

    from 4.9 (Mylan Park) to 2.2 g kg-1 (WVSK) and decreased

    significantly (p \ 0.05) along the chronosequence followingthe order Mylan Park & New Hill [ WVO1 & WVSK(Fig. 3a). Acid-hydrolysis isolates the labile SOM compo-

    nents such as proteins and polysaccharides (Barriuso et al.

    1987; Ostle et al. 1999), thus enriching the RSC with rela-

    tively recalcitrant biomolecules such as long-chain alkyls,

    waxes, lignin, and aromatic C species (Schnitzer and Preston

    1983; Haile-Mariam et al. 2000; Collins et al. 2000; Kiem

    et al. 2000; Paul et al. 2000, 2001, 2006). In addition, acid

    hydrolysis removes polyvalent cations which forms

    bridges between soil particles which leads to soil aggre-

    gation and in turn protect the organic molecules from

    microbial enzymatic degradation (Oades 1984). The SOC

    remaining after acid-hydrolysis have been related to rela-

    tively older and resistant passive SOC fractions (Leavitt

    et al. 1996; Paul et al. 1997).

    Average HFHSOC fractions ranged from 1.8 to

    1.2 g kg-1 with no statistically significant (p \ 0.05) dif-ference between the minesoils (Fig. 3a). Average FBHSOC

    fractions ranged between 9.2 (Mylan Park) and 0.6 g kg-1

    (WVO1), decreasing significantly (p \ 0.05) following theorder Mylan Park [ New Hill [ WVSK [ WVO1 along

    the chronosequence (Fig. 3a). The RSC after the final base

    hydrolysis (0.5M NaOH) step constituted the humin, the

    most stable and resistant SOC fraction resulting from

    extensive soil humification (Ussiri and Lal 2007).

    Relative proportions (%) of SOC fractions

    along the chronosequence

    Expressing SOC fractions as percentages of TSC eliminated

    the confounding effect arising from the differences in TSC

    concentrations between the minesoils and thus offered a

    better insight into relative contribution of each SOC fraction

    to the overall soil-C budget and SOC sequestration. Inter-

    estingly, contrasting trends were apparent when the per-

    centages of the SOC fractions were compared to their

    respective absolute concentrations (g kg-1). For example,

    the average BHSOC (%) ranged from about 31 to 22 % of

    TSOC and decreased significantly (p \ 0.05) following theorder WVSK & WVO1 [ Mylan Park & New Hill alongthe chronosequence which was opposite to what was

    observed for the absolute concentrations of the BHSOC

    (g kg-1) fraction (Fig. 3b). Within the chronosequence, the

    acid-hydrolysable fraction accounted for about 41 and 24 %

    of TSOC and decreased significantly (p \ 0.05) followingthe order WVO1 & New Hill [ Mylan Park & WVSKalong the chronosequence (Fig. 3b). Proteins and carbohy-

    drates have their origins in microbial and plant exudates

    (Spielvogel et al. 2007). With increasing time since recla-

    mation vegetative development and soil biological activi-

    ties were rejuvenated in the minesoils leading to increased

    production of amino acids, proteins, and sugars (Mummey

    et al. 2002). This accounted for higher absolute concentra-

    tion (g kg-1) of the labile SOM species (e.g. proteins and

    polysaccharides) in the older minesoils. However, higher

    relative proportions (%) of these labile fractions in younger

    minesoils (e.g. WVO1) indicated that with increasing age

    the relative contribution of labile fractions to the overall soil

    C budget had decreased.

    Although no significant (p \ 0.05) differences wereobserved in the absolute concentrations (g kg-1) of the

    HFHSOC fractions among the minesoils, relative propor-

    tions of HFHSOC (%) revealed a decreasing trend with

    increasing time since reclamation following the order

    WVO1 & WVSK [ New Hill & Mylan Park (Fig. 3b).Hydrofluoric acid (HF) selectively dissolves alluminosili-

    cate minerals (clays) thereby releasing the mineral-bound

    C (Dick et al. 2006; Eusterhues et al. 2007; Mikutta et al.

    2007). Organo-mineral associations reduce C degradation

    (Jones and Edwards 1998; Kalbitz et al. 2005) by microbial

    enzymatic reactions (Kaiser and Guggenberger 2007).

    Highest HFSOC (%) found at WVO1and WVSK indicated

    that organo-mineral complex formation was an important

    mode of SOC stabilization at the younger minesites. Higher

    Fig. 3 Comparison of the average standard deviation values fora individual SOC (g kg-1) fractions and b relative abundance of eachindividual SOC fractions (expressed as percentages of TSOC) for

    each reclaimed minesoil along the chronosequence. Same letter foreach individual TSOC fraction indicates no significant difference

    between the minesoils at a = 0.05. Years in the parenthesis indicatetime since reclamation. (BHSOC base hydrolysable SOC fraction,AHSOC acid hydrolysable SOC fraction, HFHSOC 10 % hydrofluoricacid hydrolysable SOC fraction and FBHSOC final base hydrolysableSOC fraction, MP Mylan Park, NH New Hill)

    Environ Earth Sci


  • mineral-bound SOC (%) at WVSK and WVO1 probably

    resulted from significantly (p \ 0.05) higher clay contentsat WVO1 and WVSK compared to the older minesoils

    (Mylan Park and New Hill) (Table 1).

    Relative proportions (%) of the humin fraction (FBH-

    SOC %) accounted for about 44 (Mylan Park) and 7 %

    (WVO1) of TSOC and followed similar trend as that of

    FBHSOC (g kg-1) (Fig. 3b). Humin represents a complex

    biochemical assemblage, remaining after various phases of

    C decomposition, constituting the most stable SOC frac-

    tions (Almendros et al. 1996; Augris et al. 1998; Piccolo

    et al. 2004). The presence of significantly (p \ 0.05) higherhumin fraction at Mylan Park indicated that with increasing

    time since reclamation biochemical stabilization of SOC

    became a key mode of soil C preservation and sequestra-

    tion in this minesoil chronosequence. This was in agree-

    ment with the previous molecular investigations performed

    in this minesoil chronosequence which indicated signifi-

    cantly (p \ 0.05) higher degree of SOC-humification atMylan Park, characterized by presence of more polycon-

    densed and polysubstituted aromatic-C species probably

    imparting additional biochemical stability to the organic

    molecules (Chaudhuri et al. 2012b).

    Distribution of coal-C along the chronosequence

    The average concentration (g kg-1) of coal-C ranged from

    1.76 to 0.6 g kg-1 along the chronosequence with no sta-

    tistically significant (p \ 0.05) differences observedbetween the minesoils (Table 3). Higher coal-C at WVSK

    probably resulted from the recent soil-scraping activities

    that exposed the subsoil having more coal fragments.

    Although no significant (p \ 0.05) differences wereobserved in the absolute coal-C concentrations (g kg-1)

    among the minesoils, relative proportions of coal-C (%),

    however, differed significantly (p \ 0.05) along thechronosequence. The highest and lowest coal-C (%) were

    observed at Mylan Park (2.5 %) and WVO1 (14 %),

    respectively, which suggested negligible contribution of

    coal-C to overall soil C dynamics in this chronosequence

    except at the youngest site (Table 2). In accordance with

    this finding, no significant (p \ 0.05) differences wereobserved between TSC (g kg-1) and TSOC (g kg-1) at any

    minesite, except for WVO1, which indicated that TSC can

    be considered a suitable surrogate for TSOC in this

    chronosequence. This was in agreement with Ussiri and Lal

    (2007) who noted that contribution of coal-C is negligible

    in the surface horizons (*10 cm depth) where appropriatereclamation strategies, such as topsoil application (similar

    to this chronosequence), have been implemented. Differ-

    ences in TSC and TSOC at WVOI probably resulted due to

    more recent disturbances and shorter time since reclama-

    tion as compared to the older minesites, inadequate to

    initiate pedological and biological processes required for

    soil profile development and SOC accumulation.

    Interrelationship between TSC and SOC fractions

    Significant (p \ 0.05) positive relationships were foundbetween TSC and the humin fractions (FBHSOC, g kg-1)

    in all minesoils. However, the r2 values decreased from the

    oldest to the youngest minesoil underscoring the varying

    degree of influence of humin on overall soil-C budget

    (Table 4). The highest r2 at Mylan Park indicated that as

    minesoils aged, humification processes began to set in,

    imparting biochemical resistance to the SOC mole-

    cules from degradation leading to soil C stabilization and

    long-term sequestration. In contrast, highest r2 values at

    WVO1 were found between TSC and acid-hydrolysable

    and mineral-bound SOC fractions, which indicated that soil

    C-stabilization mechanisms in younger minesoils differed

    substantially from the older minesoils (e.g. Mylan Park). At

    younger minesites SOC stabilization mainly resulted from

    formation of organo-mineral complexes. Lack of any sig-

    nificant (p \ 0.05) relationship (r2) between coal-C andTSC or any SOC fraction at any minesite indicated that the

    overall soil C dynamics in this minesoil chronosequence

    was unaffected by coal-C.

    Interrelationship between SOC fractions and soil

    quality parameters

    Our previous study revealed increasing CEC and decreas-

    ing EC with increasing time since reclamation along the

    chronosequence (Table 1) (Chaudhuri et al. 2012a). Higher

    CEC and coupled with lower EC at Mylan Park had

    favorable impacts on soil biological processes, such as root

    network development and microbial activities, which led to

    increased SOC accumulation. Higher SOC contents at

    Mylan Park, and New Hill, had positively influenced CEC

    Table 3 Descriptive summary for coal-C along the chronosequence

    Minesoil Mean Min Max

    Coal-C (g kg-1)

    Mylan Park (22 year) 0.59a 0.01 1.93

    New Hill (5 year) 1.76a 0.01 3.94

    WVSK (4 year) 1.68a 0.36 3.19

    WVO1 (2 year) 1.46a 0.02 3.05

    Coal-C (%)

    Mylan Park (22 year) 2.69c 0.06 6.47

    New Hill (5 year) 9.22b 0.09 9.22

    WVSK (4 year) 14.34a 4.80 18.94

    WVO1 (2 year) 13.45a 0.01 15.85

    Same letters indicate no significant difference at a = 0.05

    Environ Earth Sci


  • and EC which were more influenced by the clays at WVO1

    (Chaudhuri et al. 2012a). At Mylan Park and New Hill,

    both CEC and EC were strongly influenced by the stable

    humin fraction (Table 5). In contrast, at WVO1 soil CEC

    was more related with the labile acid-hydrolysable and the

    mineral-bound SOC fraction while no significant

    (p \ 0.05) effects of any SOC fraction found on EC(Table 5). The above observations indicated that different

    SOC fractions had varying effects on soil quality indicators

    and were time-dependent phenomenon.


    Due to well-defined ages minesoil chronosequences are

    ideal for identifying spatio-temporal changes in soil prop-

    erties including SOC sequestration. Intricate mixing of

    SOC with coal-C in reclaimed minesoils, however, hinders

    accurate estimation of soil carbon stocks and thus carbon

    sequestration potential. Methods to distinguish between

    coal-C and SOC in minesoils are often expensive and

    resource-intensive. In the present context, we used a

    relatively inexpensive, rapid, easy-to-use yet accurate

    chemi-thermal fractionation scheme to isolate coal-C in a

    chronosequence comprising four minesoils which differed

    only by their respective times since reclamation. The

    results indicated that contribution of coal-C to overall soil

    C budget was negligible. A relatively higher proportion of

    coal-C, however, was observed in the younger minesoils

    which probably was due to lack of adequate time to initiate

    pedologic processes at younger minesites that lead to SOC


    The fractionation scheme effectively isolated a number

    of chemically stabilized SOC fractions, each having a

    distinct temporal trend and varying effect on soil quality

    indicators. At the oldest site (Mylan Park), the soil-C

    budget was dominated by the humin fraction underscoring

    the importance of humification processes on carbon sta-

    bilization and overall soil-C dynamics. In contrast, at the

    younger minesites the labile and mineral-bound SOC

    fractions controlled the SOC dynamics. Soil CEC and EC

    were significantly (p \ 0.05) influenced by the SOC frac-tions with differing strengths (r2) and were a function of

    time since reclamation. Soil CEC and EC were influenced

    Table 4 Coefficient of determination (r2) between TSC, SOC frac-tions and coal-C along the minesoil chronosequence

    Minesoil TSC

    (g kg-1)


    (g kg-1)


    (g kg-1)


    (g kg-1)


    (g kg-1)

    BHSOC (g kg-1)

    MP 0.66*

    NH 0.57*

    WVSK 0.73*

    WV01 0.24

    AHSOC (g kg-1)

    MP 0.57* 0.44

    NH 0.60* 0.31

    WVSK 0.57* 0.54*

    WVO1 0.76** 0.15

    HFSOC (g kg-1)

    MP 0.12 0.03 0.01

    NH 0.46* 0.44 0.07

    WVSK 0.30 0.06 0.13

    WV01 0.61* 0.15 0.30

    Humin (g kg-1)

    MP 0.88** 0.70** 0.38 0.08

    NH 0.76** 0.35 0.26 0.24

    WVSK 0.66* 0.31 0.29 0.12

    WVO1 0.57* 0.11 0.40 0.29

    Coal-C (g kg-1)

    MP 0.00 0.03 0.05 0.16 0.00

    NH 0.34 0.03 0.07 0.13 0.14

    WVSK 0.30 0.09 0.06 0.00 0.12

    WVO1 0.01 0.34 0.00 0.07 0.04

    MP Mylan Park, NH New Hill

    ** Significant at p \ 0.01, * p \ 0.05

    Table 5 Coefficient of determination (r2) between different SOC fractions and soil properties including CEC and EC

    Soil property Minesoil name BHSOC (g kg-1) AHSOC (g kg-1) HFSOC (g kg-1) Humin (g kg-1)

    CEC (cmolc kg-1) Mylan Park 0.43 0.41 0.08 0.82**

    New Hill 0.44* 0.39 0.21 0.73*

    WVSK 0.39 0.28 0.50* 0.33

    WVO1 0.20 0.54* 0.51* 0.26

    EC (dS m-1) Mylan Park 0.39 0.38 0.03 0.75*

    New Hill 0.41 0.32 0.09 0.53*

    WVSK 0.20 0.32 0.26 0.37

    WV01 0.09 0.21 0.18 0.38

    ** Significant at p \ 0.01, * p \ 0.05

    Environ Earth Sci


  • more by the stable humin fraction at the oldest minesite,

    while more so by the acid-hydrolysable (labile) and min-

    eral-bound SOC fractions at the youngest site. Overall,

    appreciable soil-C recovery indicated that this fractionation

    procedure can aptly be used to characterize SOC dynamics

    in coal-C-contaminated soils.

    With emerging environmental issues emanating from

    progressive accumulation of CO2 and other greenhouse

    gases in the atmosphere, the approach outlined in this study

    will become increasingly relevant to assess and understand

    SOC dynamics in reclaimed minesoils. However, some

    effort to identifying the molecular properties and turnover

    times of different SOC fractions isolated at different steps

    of this sequential fractionation method will provide more

    insight into the overall SOC dynamics in these disturbed


    Acknowledgments The authors are thankful to the scientific con-tribution no. 3153 from the West Virginia Agricultural and Forestry

    Experiment Station, Morgantown, WV for supporting this research.


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    Chemically stabilized soil organic carbon fractions in a reclaimed minesoil chronosequence: implications for soil carbon sequestrationAbstractIntroductionMaterials and methodsStudy areaSoil sampling and analysis

    Results and discussionDistribution of total soil carbon (TSC; g kgminus1) along the chronosequenceAbsolute concentrations (g kgminus1) of SOC fractions along the chronosequenceRelative proportions (%) of SOC fractions along the chronosequenceDistribution of coal-C along the chronosequenceInterrelationship between TSC and SOC fractionsInterrelationship between SOC fractions and soil quality parameters



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