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Soil Organic Carbon Fractions Under Different LandUses in Mardi Watershed of NepalZ. H. Yang a b , B. R. Singh a & B. K. Sitaula a ca Department of Plant and Environmental Sciences , Agricultural University of Norway ,Norwayb College of Resources and Environmental Sciences , Hunan Agricultural University ,Changsha, Chinac Centre for International Environment and Development Studies , Agricultural Universityof Norway , NorwayPublished online: 20 Aug 2006.

To cite this article: Z. H. Yang , B. R. Singh & B. K. Sitaula (2004) Soil Organic Carbon Fractions Under Different LandUses in Mardi Watershed of Nepal, Communications in Soil Science and Plant Analysis, 35:5-6, 615-629, DOI: 10.1081/CSS-120030347

To link to this article: http://dx.doi.org/10.1081/CSS-120030347

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COMMUNICATIONS IN SOIL SCIENCE AND PLANT ANALYSIS

Vol. 35, Nos. 5 & 6, pp. 615–629, 2004

Soil Organic Carbon Fractions Under Different

Land Uses in Mardi Watershed of Nepal

Z. H. Yang,# B. R. Singh,* and B. K. Sitaulaz

Department of Plant and Environmental Sciences, Agricultural

University of Norway, Norway

ABSTRACT

This study was conducted to investigate the organic carbon (C)

content and its chemical fractions in soils under forest, grassland, and

agricultural uses in the Mardi Watershed of Nepal. Surface soil

samples from 0 to 15 cm were collected. Classical extracting

procedure with alkali and acid solution was used to separate humic

acid (HA), fulvic acid (FA), and humin fractions. Hydrogen peroxide

was used to separate black carbon (BC) from humin. The results

showed that higher amounts of total soil organic carbon (SOC),

#Current address: Z. H. Yang, College of Resources and Environmental Sciences,

Hunan Agricultural University, Changsha, China.

*Correspondence: B. R. Singh, Department of Plant and Environmental Sciences,

Agricultural University of Norway, P.O. Box 5003, As N-1432, Norway;

Fax: 64948211; E-mail: balram.singh@ipm.nlh.no.zCurrent address: B. K. Sitaula, Centre for International Environment and

Development Studies, Agricultural University of Norway, Norway.

615

DOI: 10.1081/CSS-120030347 0010-3624 (Print); 1532-2416 (Online)

Copyright & 2004 by Marcel Dekker, Inc. www.dekker.com

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as well as of different SOC fractions, i.e., FA, HA, humin and BC,

existed in the forest soils compared to those from grassland and

agricultural areas. The carbon proportion of FA, HA, and humin

fractions accounted for 25, 29, and 37% of the total SOC in forest

soils. The corresponding values were 34, 27, and 36% in grassland,

and 28, 18, and 54% in agricultural land. The grassland soil showed a

higher FA fraction, while the agricultural soil contained lower HA

fraction and higher humin fraction as compared to the forest soil.

Forest soil showed higher HA/FA ratio and lower (HAþFA)/SOC

ratio, indicating high degree of humification and large humification

rate in this soil. Black carbon contents occupied 3.4, 3.4, and 11.5%

of SOC under forest, grass, and agricultural lands, respectively.

Key Words: Black carbon; Land use; Nepal; Organic carbon

fractions; Watershed.

INTRODUCTION

Soil-vegetation system can act as a sink or source of atmosphericcarbon dioxide (CO2), depending on its decomposition rate and the rateof SOC formation.[1] Since cultivation of virgin soils (native grassland orforest) usually elevates organic carbon (OC) turnover and increases CO2

emission, the soil after forest clearing can turn into a CO2 source.Cultivation of native grassland or forestland resulted in a decline in SOCcontent and the net release of CO2 to the atmosphere.[2] Detwiler[3]

estimated that the cropping in tropical forest soils reduced their Ccontent by 40%, whereas their use for pastures reduced it by about 20%.Data from Ellert and Gregorich[4] illustrated an overall decrease in SOCdue to the cultivation in 12 out of 15 sites in Ontario. The shift fromforest soil to agricultural soil would be associated with a change in thetypes of vegetation and subsequent changes in C input and magnitude ofvarious C pools. Meanwhile, conversion from forest soil to agriculturalsoil can also affect organic matter quality. Many studies revealed thatphysical fractionation and carbon distribution in different particle-sizeseparates are quite sensitive to cultivation.[5,6] For instance, particulateorganic matter and light fraction showed a large decrease due tocultivation.[7] Studies on physical fractionation of SOC revealed thatdeforestation and subsequent agricultural management increased theloss of labile SOC as well as stable SOC fractions, particularly SOCassociated with the silt-size separates.[8] The results on chemicalcomposition of SOC showed that the carbohydrates present in cultivatedsoils were richer than that of forest soils, while the phenolic compounds

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oxidized by CuO appeared to be richer in forest soil. At the same time,conversion of forest to agricultural land can also result in the depletion ofboth lipids and lignin component.[9,10]

Humic substances are relatively stable in soils, and are veryimportant in carbon sequestration and carbon cycling. A study onchemical fractionation of humic substances showed that the change ofland use, from primary forest to coffee or arable crops, led to a decreasein the proportion of HA and an increase in that of FA fraction of thetotal organic matter in the topsoil.[11] The decrease of HA/FA ratio aftercultivation as compared to that of uncultivated native grassland wasreported.[12] Although FA and HA are considered important to assess thequality of organic matter, humin, and inert carbon (black carbon)fractions have recently been observed to play important roles in C cyclingand sequestration.[13,14] However, very few studies on the effect of landuse practices on these fractions have been conducted. The present studywas aimed to investigate the chemical fractions of SOC under differentland uses in the Mardi Watershed of Annapurna Conservation Areain the western hills of Nepal.

METHODS AND MATERIALS

Site Description

The Mardi watershed covers an area of about 63 km2. The elevationranges from about 900m at the confluence of Mardi and Seti rivers nearHemja to as high as about 5400m. Annual precipitation exceeds 4000mmand is mainly concentrated during the months from June to September.Major forest types include mixed hardwood forest, oak forest, and thehigh mountain mixed forest. Above the tree line exist the alpinegrassland. Agricultural practices in the watershed extend from valleybottom to the hilltop. Therefore, we selected representative forest,grassland, and agricultural lands in this watershed to study the effectof land uses on total SOC and its chemical fractions.

Soil Sampling

Soils were collected from the topsoil horizon of 13 sites under forest,6 sites under grassland, and 7 sites under agricultural use (Table 1),located in the Mardi Watershed of Annapurna Conservation Area in thewestern hills of Nepal. In order to prevent fraction changes, soil sampleswere kept in airtight glass bottle in cool room (þ4�C) in moist condition.

Soil Organic Carbon Fractions 617

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Prior to analysis, soil samples were dried at 80�C in oven, ground, andpassed through a 2-mm sieve.

Determination of SOC and Its Fractions

Total soil carbon content was measured by dry combustion in EC12Carbon Analyzer (Model 752-100, LECO Corporation, USA). The totalcarbon content in these soils may generally represent SOC content due tovery low amount of inorganic carbon (pH< 4.5 in all soil samples). Soilorganic carbon was fractionated into FA, HA, humin, and black carbon

Table 1. Selected properties of the studied soils.

Land use

Range

Mean/SDaMin. Max.

pH (H2O)

Forest land 3.4 4.5 3.8±0.30

Grassland 2.8 3.8 3.5±0.41

Agricultural land 3.5 5.0 4.0±0.58

O.M. (kgm�2)

Forest land 2.3 15.7 5.7±3.8

Grassland 1.2 12.2 6.6±4.4

Agricultural land 1.8 6.1 4.0±2.0

N (kgm�2)

Forest land 0.12 0.74 0.33±0.19

Grassland 0.16 0.60 0.35±0.18

Agricultural land 0.16 0.25 0.19±0.03

P (gm�2)

Forest land 3.0 9.8 5.5±1.9

Grassland 1.6 16.2 6.9±5.5

Agricultural land 1.4 10.5 4.7±3.5

K (gm�2)

Forest land 19.9 119.2 63.9±29.7

Grassland 18.0 74.0 39.8±25.3

Agricultural land 19.9 89.4 36.2±26.4

Number of soil samples collected from forest, grassland, and agricultural land

were 13, 6, 7, respectively.aThe mean values of all properties in the same column were not statistically

different at P< 0.05. The values of all soil properties were extracted and modified

from Awasthi et al.[15]

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(BC) fractions using the procedures described by Wu et al.[15] In brief: 3 gsoil was suspended with 50mL of 0.5ML�1 NaOH under N2 in 250-mLpolyethylene centrifuge bottle. After 24 h, end over end shaking, thesuspension was centrifuged at 4000 rpm for 30min to separate the coloredsupernatant (FAþHA) fraction from the sediment. The supernatant wastransferred into 100-mL centrifuge glass tubes, acidified with 2.5ML�1

H2SO4 to pH 1.8 and allowed to stand at 25�C for 24 h. The suspensionwas centrifuged to separate the soluble material (FA fraction). Onemilliliter FA or 1mL FAþHA was taken into a tin foil cup and dried at60�C in oven. The sediment (humin) was washed with 50mL 0.1ML�1

Na2SO4, centrifuged at 4000 rpm, and finally dried at 80�C. One gram ofhumin was weighed in 100-mL centrifuge tube and suspended with 20mLof 30% H2O2. After 2 h oxidation at 60�C, the suspension was heatedto 90�C for 1 h to evaporate the remained H2O2. The solid residuecontaining black carbon was separated from the solution by centri-fugation as described above. The sediment was then acidified with0.025ML�1 H2SO4 to pH< 4 to decompose carbonates, centrifuged at4000 rpm, and dried at 105�C. The concentrations of the organic carbonin dried (FAþHA) and FA, humin, black carbon were measured by drycombustion in EC12 Carbon Analyzer.

Statistical Analyses

The data obtained were analyzed by using SAS procedure.[17]

Analysis of variance was tested for each variable, with means comparisonby least significant difference (LSD) method and Neman’s Kuel’s test(�¼ 0.05).

RESULTS AND DISCUSSION

Total Soil Organic Carbon

Total SOC content in the surface soil under forest varied from 1.7 to13.8 kgm�2, with mean value of 6.1 kgm�2. This value is about 20%more than the SOC found under grassland and 2 times higher than thatunder agricultural land. The difference between grassland and agricul-tural land was not statistically significant. Pardo et al.[18] estimated thatcultivated soil showed a significant loss of organic matter with regardsto the virgin soil (amounting to 60 and 50% in the surface layer andunderlying horizon, respectively). It was also reported that forest

Soil Organic Carbon Fractions 619

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clearing and continuous cultivation led to depletion of total organiccarbon by 55%.[8] The SOC losses under agricultural use in this studywere much larger than those reported above. It was likely due to higherdecomposition rate of SOC in the subtropical area of the present study.The shift from forest to agricultural uses would be associated withchanges in vegetation type and subsequent change in C input and magni-tude of various C pools. Cultivation may help to enhance decompositionby creating soil conditions (e.g., of moisture, aeration and temperature)that are more conducive to greater biological activity.[19] Soil disturbanceunder agricultural use caused a reduction in soil structural stabilityand the light fraction of SOC was readily decomposable by soilmicroorganisms.[20]

Humic Acid and Fulvic Acid

The carbon content of FA ranged from 0.4 to 2.4 kgm�2 in forestsoil, from 0.5 to 2.6 kgm�2 in grassland, and from 0.3 to 1.1 kgm�2 inagricultural soil (Table 2). The C content in FA of grassland wassignificantly higher than that of agricultural soil, but there was noobvious difference of FA between grassland and forest soil. The Ccontent in HA of forest soil was about 3 times larger than that ofagricultural soil, nevertheless small difference existed between forest soiland grassland.

The HA/FA ratio was termed humification degree.[21] The values ofHA/FA ratio were 1.26±0.37 (n¼ 13) in forest soils, 0.86±0.33 (n¼ 6)in grassland and 0.67±0.23 (n¼ 6) in agricultural soils. Humificationprocess of organic matter is closely related to climate and soil condition.Lower temperature and larger moisture content at higher elevation in theforest soils probably limit decomposition and result in a more favorablecondition for humic acid formation. A significantly positive correlationcoefficient (r¼ 0.507�, n¼ 26) between HA/FA ratio and elevation wasobserved in the present study (Table 3). Humic acid was considered to beof higher molecular weight and degree of polymerization than FA. Inturn, the polymerization is related to the degree of humification.[12]

Therefore, higher the HA/FA ratio, higher is the humification degree ofSOC. The present results indicated that forest soils showed a higherhumification degree than cultivated soils.

The ratio of HAþFA to SOC has been termed as the humificationratio.[22] The (HAþFA)/SOC ratio both in forest and agricultural soilswas significantly lower than that in grassland. However, there was nosignificant difference in this ratio between forest and agricultural soils

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(Fig. 1). Unlike HA/FA ratio, the correlation between (HAþFA)/SOCratio and elevation was poor (Table 3). In the present study, nonhumicsubstances, i.e., carbohydrates, proteins, peptides, fat, waxes, and otherlow-molecular-weight organic substances were not separated from humicsubstances during the extraction process. Although SOC showed asignificant correlation with elevation, the ratio of alkali-extractablehumic substances to SOC showed poor correlation with elevationprobably due to accumulation of non-humic substances in the soilsmainly affected by the types and quantity of plant material.[23] The ratioof HAþFA to SOC in this study is considerably lower than the value of

Table 2. Carbon content in different SOC fractions under different land uses.

Land use

Range

Mean Std.Min. Max.

SOC (kgCm�2)

Forest land 1.66 13.8 6.13 a 4.08

Grassland 1.25 8.50 5.03 b 2.73

Agricultural land 1.03 4.58 2.84 b 1.07

FA (kgCm�2)

Forest land 0.40 2.35 1.36 ab 0.64

Grassland 0.49 2.59 1.59 a 0.73

Agricultural land 0.33 1.14 0.80 b 0.32

HA (kgCm�2)

Forest land 0.47 4.03 1.82 a 1.25

Grassland 0.31 2.78 1.45 ab 0.98

Agricultural land 0.08 1.03 0.57 b 0.31

Humin (kgCm�2)

Forest land 0.60 5.89 2.25 a 1.58

Grassland 0.58 2.87 1.72 a 0.80

Agricultural land 0.73 2.20 1.44 a 0.45

BC (kgCm�2)

Forest land 0.03 0.56 0.21 a 0.17

Grassland 0.06 0.28 0.14 a 0.08

Agricultural land 0.10 0.78 0.28 a 0.24

Number of soil samples collected from forest, grassland, and agricultural land

were 13, 6, 7, respectively.

SOC¼ soil organic carbon; FA¼ fulvic acid; HA¼ humic acid; BC¼ black

carbon.

The mean values of SOC and each SOC fraction followed by different letters in

the same column were significantly different at P¼ 0.05.

Soil Organic Carbon Fractions 621

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A. HA/FA ratio

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

HA

/FA

a

b

b

B. (FA+HA)/SOC ratio

0

10

20

30

40

50

60

70

forest soil grassland agricultural soil

forest soil grassland agricultural soil

(FA

+H

A)/

SO

C (

%)

ab

b

Figure 1. HA/FA ratio and (HAþFA)/SOC ratio in soils under different land

uses. The value followed by different lower case letter in the same figure block are

significantly different at p¼ 0.05.

Table 3. The correlation matrix among organic C fractions and elevation.

SOC FA HA Humin BC HA/FA

(HAþFA)/

SOC

FA 0.878a

HA 0.980a 0.881a

Humin 0.955a 0.786a 0.908a

BC 0.405a 0.210 0.324 0.570a

HA/FA 0.766a 0.527a 0.815a 0.695a 0.174

(HAþFA)/SOC �0.079 0.244 0.031 �0.216 �0.62a 0.003

Elevation 0.659a 0.695a 0.676a 0.505a 0.046 0.507a 0.248

aSignificant at p< 0.05.

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9–10% reported by McCallister et al.[21] presumably due to differencein soil types and climate. Under subtropical environment in Nepal,intensive cultivation in agricultural soil may lead to higher decompositionrate resulting in low amount of HA and FA and relatively higher humin.The polymers in humic compounds other than FA and HA, for examplehumin, are generally stable and resistant to microbial attack.[23]

Therefore, a low (HAþFA)/SOC ratio usually indicates a higherdegree of humification. Humification ratio offers an alternative way toevaluate the quality and stability of humus.[12,24] In the present study, asignificantly higher HA/FA ratio and low (HAþFA)/SOC ratio in theforest soils may indicate that these soils had a higher humification degreethan the soils under other land uses.

Humin Contents

The C content of humin fraction in forest soil reached to 2.3 kgm�2,which was 24% and 36% higher than that of grassland and agriculturalsoil, respectively. In the biogeochemic C cycling, humin serves as theportal through which organic C produced in the biosphere is transferredto the geosphere.[13] In the forest ecosystems, more organic C as huminproduced by photosynthesis ultimately accumulated in soils. Meanwhile,humin has been considered a high molecular weight polymer, which hashigh resistances to microbial decomposition. Thus this study implied thatmore humin C was sequestered in forest soils than that in cultivated soils.

Distribution of Humic Substances

In forest soil, FA, HA, and humin accounted for 25, 29, and 37% ofSOC, respectively. The corresponding values in grassland were 34, 27,and 36% of SOC, and in agricultural soil 28, 18, and 54% of SOC(Fig. 2). Compared to forest soil, grassland showed a significantly higherFA, while the other two C fractions were not significantly different inthese soils. The agricultural soil contained lower HA fraction and higherhumin fraction as compared to forest soil. Under agricultural land use,the tropical to subtropical environment of this study may promote rapidorganic matter decomposition, reducing the accumulation of FA andHA, and resulting in relatively higher accumulation of humin.

In the present study, the sum of C content of FA, HA, and huminfractions contributed to 91% of SOC in the forest soils, while thecorresponding values for grassland and agricultural soils were obtained

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97%, and 100%, respectively. This indicated that forest soils had a largemass imbalance of C between SOC fractions and the total SOC.Generally, soil contains varying amounts of nonpolar compounds, suchas lipid. Soil lipids content varied from 4 to 8%,[25] and it can evencontribute to 42% in some organic soil.[26] This implies that organicmatter rich soils such as the forest soils in the present study may containhigher lipid substances and thus leading to larger imbalance in C contentof SOC and its fractions. Lipid could not be extracted with alkali andacid solutions and was thus retained in humin fraction. It is suspectedthat during the drying procedure of analysis, a part of lipid may bevolatilized.

Black Carbon

Soil commonly contains inert organic matter, which is also termed asBlack Carbon (BC).[27,28] In the present study, the BC content rangedfrom 0.03 to 0.56 kgm�2 in forest soil, from 0.06 to 0.28 kgm�2 ingrassland, and from 0.10 to 0.78 kgm�2 in agricultural soil. It occupied,on average, 3.4, 3.4, and 11.5% of SOC in forest, grassland andagricultural soils, respectively. Agricultural soils showed a significantlyhigher BC content as well as proportion in SOC than forest soil andgrassland. Black carbon originates from incomplete combustion of

0

10

20

30

40

50

60

70

FA HA Humin

C p

erce

nt (

%)

Forest soil

Grassland

Agricultural soil

a bb

a a

b

b

b

a

Figure 2. Distribution of different C fractions in soils under different land uses.

The value followed by different lower case letter in the same carbon fraction are

significantly different at p¼ 0.05.

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organic materials. Cultivation of forest soils into agricultural soil

inevitably resulted in burning of vegetation, which should increase BC

amount as observed in the present study (Fig. 3). It is reported that BC

content can reach up to 8 g kg�1, and it can contribute up to 40% of the

A. Black Carbon Content

0

0.1

0.2

0.3

0.4

0.5

0.6

Bla

ck C

arbo

n C

onte

nt (

kg m

-2)

a

a

a

B. Black Carbon percent of SOC

0

5

10

15

20

25

forest soil grassland agricultural soil

forest soil grassland agricultural soil

Pro

port

ion

of B

C in

SO

C (

%)

a

bb

Figure 3. Black carbon content and proportion of BC in SOC under different

land uses. The value followed by different lower case letter in the same figure

block are significantly different at p¼ 0.05.

Soil Organic Carbon Fractions 625

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total organic C.[29–31] Due to higher resistant ability to microbial attackthan the other organic carbon fractions,[32,33] BC gets accumulated inagricultural soils. Therefore, BC is thought to play an important role inthe composition and turnover of organic matter in the naturalenvironment since it should be a sink for biosphere C.

CONCLUSIONS

The forest soils contained higher SOC content and higher OCfractions, such as FA, HA, humin, and BC as compared to those fromgrassland and agricultural land. The grassland soil showed a significantlyhigher FA fraction, while the agricultural soil contained lower proportionin HA fraction and higher in humin fraction as compared to forest soil.The HA/FA ratio was higher and the (HAþFA)/SOC ratio lower in theforest soils as compared to grassland and agricultural land, indicating ahigher humification degree of organic matter existed in the former soils.A relatively higher proportion of BC fraction was obtained inagricultural soils than that in grassland and forest soils. Therefore,land uses obviously affected both the SOC content and its chemicalfractions in the Mardi Watershed of Nepal.

ACKNOWLEDGMENTS

The senior author is grateful to the Research Council of Norway(NFR) for providing fellowship during the period of this study. Thefinancial assistance for chemical analysis by the NFR funded project(project number 131692/730) is gratefully acknowledged.

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