TEXTURAL, MINERALOGICAL AND STRUCTURAL CONTROLS ON SOIL

ORGANIC CARBON RETENTION IN THE BRAZILIAN CERRADOS

DISSERTATION

Presented in Partial Fulfillment of the Requirements for

the Degree Doctor in Philosophy in the Graduate

School of The Ohio State University

By

Yuri Lopes Zinn, M. S.

The Ohio State University

2005

Dissertation Committee: Approved by

Professor Rattan Lal, Adviser

Professor Jerry Bigham Adviser

Professor Warren Dick Soil Science Graduate Program

Professor Andrew Ward

ABSTRACT

The retention of soil organic carbon (SOC) is commonly considered the result of climate, vegetation, internal drainage and management interactions. However, a sparse but considerable set of evidence in the literature suggests that soil texture, mineralogy and aggregation also affect SOC retention. The objective of this research was to qualitatively and quantitatively assess the controls that these three soil properties exert on SOC levels in the Brazilian Savanna (Cerrado) region, in tropical humid South America. Thus, three soils under similar climate and slope, but of contrasting texture and under native vegetation and Eucalyptus plantation, were sampled in triplicate to 1 m depth, and characterized by physical, chemical, mineralogical, Yoder wet sieving, and microscopic analyses. Total C and N concentration were determined in bulk soils, particle size separates (clay, silt, sand) and water-stable aggregates (WSA). A basic assumption was made that the SOC particle size is inherently associated to its retention mechanism: colloidal and soluble forms are retained by sorption to clays; particulate organic matter (POM, >20 μm) are retained outside (free-POM) or inside aggregates (occluded-POM), and silt-sized SOC has intermediate properties. The rationale is that these mechanisms are necessarily determined by soil texture, mineralogy and structure, which then control SOC retention. The three soils were highly weathered and composed mostly of quartz and kaolinite, with minor and variable contents of Fe-Al oxides, hydroxyl-interlayered vermiculite and illite. These soils were classified as clayey Haplustox, loamy Haplustox and sandy soils (one Quartzipsamment and two sandy Haplustoxes). In all soils, Eucalyptus plantation increased the C/N ratio and decreased aggregation and aggregate- occluded POM in the top 10 cm layer in relation to native vegetation, but bulk SOC

ii concentrations and stocks were not affected. In the sandy soils only, the relative content of SOC in the sand fraction was enriched, whereas it decreased in the clay fraction. Soil texture and depth strongly affected bulk SOC concentrations, which could be credibly estimated as a function of clay+silt contents and depth, by means of a novel mathematical model (using data from samples under the two land uses). The specific surface area (SSA) of soil<2 mm under native Cerrado was modeled as a function of clay, silt and SOC contents but not depth. This suggests that SOC levels increase with higher SSA associated with higher clay contents. In a single soil profile, however, SSA decreases near the soil surface because of slightly lower clay contents but also because higher SOC levels promote clay flocculation and aggregation. The SOC concentration in particle size separates was inversely related to the proportion of that size fraction in soil (SOC dilution effect), which precluded its use in modeling SOC size partition. However, the calculation of relative amounts of total SOC in each fraction allowed for prediction of clay-sized SOC (as percent of total SOC), based on clay contents and depth. A quantitative assessment of clay mineralogy showed that, for the bulk soil, SOC concentrations were better correlated with contents of crystalline and amorphous Fe- oxides in surface layers and amorphous Al oxides in the subsoil, with higher coefficients of determination than those of SOC vs. clay+silt. These trends were even stronger when the clay-sized SOC pool was correlated to the same mineral phases. Aggregation, as indicated by the mean weight diameter (MWD) and percent of WSA>2 mm, was strongly correlated with clay+silt contents, but bulk SOC was poorly correlated with MWD and WSA>2 mm except for the 0-5 cm depth. The fraction of POM occluded inside aggregates was strongly affected by soil texture, varying from ca. 25% in the sandy soils to ca. 50% in the clayey Haplustox. The activity of soil fauna resulted in three types of zoogenic aggregates (fecal pellets, cocoons and aggrotubules), the latter more common and SOC-enriched in soils of loamy and coarser texture. Because texture directly affects the contents of Fe and Al oxides and the protection of POM within aggregates, SOC retention in comparable aerobic Cerrado soils is controlled, in decreasing order of importance, by: 1) texture, 2) mineralogy, and 3) structure (including pedogenic and faunal peds).

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DEDICATION

This work is dedicated to my wife Alba who gave me, more than strength, a reason to go on.

iv

ACKNOWLEDGMENTS

I am deeply indebted to the Capes Foundation within the Ministry of Education of Brazil, which gave the generous financial support that made this doctorate possible. I also thank the Graduate School of the Ohio State University for the award of a Presidential Fellowship, which allowed an essential extra time to complete this work. I thank Dr. Rattan Lal for many things, but especially for: accepting to supervise my doctoral studies; selecting an outstanding graduate committee; fast and efficient review of all submitted manuscripts; and the nomination to the Presidential Fellowship. The operational support that made possible the sampling travels was kindly provided by the Embrapa Cerrados, in the person of Dr. Dimas V.S. Resck, and to V & M Tubes Florestal Co., in the person of Dr. Hélder B. Andrade. My deep thanks are also due to Dr. Jerry M. Bigham and Franklin “Sandy” Jones, for their constant and fundamental support with the soil characterization, particle-size fractionation, mineralogy and micromorphology parts of this work. I must thank the following faculty, who provided very kind letters of recommendation for the Presidential Fellowship and other awards: within OSU, Drs. Jerry Bigham, Frank Calhoun, Warren Dick, Brian Slater (SNR), Andy Ward (FABE), Garry McKenzie (GS), and of course Dr. Rattan Lal. Outside OSU, I thank Drs. Wolfgang Zech (Univ. Bayreuth, Germany) and Dimas Resck (Embrapa Cerrados, Brazil).

v A great number of people provided help with other fundamental phases of this work or my stau in the U.S., from which I specifically thank: Drs. Steve Saint-Martin and Bert Bishop (OARDC), and Antônio C. Gomes (Embrapa Cerrados), for help with the experimental design and statistical analyses; Mrs. Jesuíno S. Caldas and Wantuir C. Vieira (Embrapa Cerrados), for field and laboratory support in Brazil; Mrs. Yogendra Raut and Tony Karcher (SNR) for laboratory support; Mss. Pat Polczinsky and René Johnston for help with many SNR office issues; Dr. Neil Smeck for helping classify soils according to the U.S. Soil Taxonomy; and Dr. E. Bonnelo and his students of Dept. of Plant Pathology-OSU, for the use of their binocular microscope and digital camera. I cannot forget to thank my wife Alba who helped me many times, with data typing and relevant suggestions. Finally, I thank my parents, Jorge A. Zinn and Vera Lucia Lopes, for their support (personal and official), and all the other members of my family, in special to vó Maria Jacy and mana Lara, who supported me in their small but important ways.

vi

VITA

November 29, 1970…………..… Born – Brasília-DF, Brazil 1994…………………………….. B.S. in Forestry, Federal University of Viçosa 1998………………………….…. M.S. Agronomy/Soil Science, University of Brasília 1997-2000……………………… Analyst of Science and Technology, Capes Foundation, Ministry of Education of Brazil 2001-2004……………………… Ph.D. student in Soil Science, the Ohio State University, with a grant from the Capes Foundation 2005-present…………………… Presidential Fellow, the Ohio State University

PUBLICATIONS

1. Zinn, Y.L., D.V.S. Resck, and J.E. Silva. 2002. Soil organic carbon as affected by afforestation with Eucalyptus and Pinus in the Cerrado region of Brazil. For. Ecol. Manage. 166: 285-294. 2. Zinn, Y.L., R. Lal, and D.V.S. Resck. 2005. Changes in soil organic carbon stocks through agriculture in Brazil. Soil Till. Res. 84:28-40. 3. Zinn, Y.L.; R. Lal, and D.V.S. Resck. 2005. Texture and organic carbon relation described by a profile pedotransfer function in Brazilian Cerrado soils. Geoderma 127:168-173.

FIELDS OF STUDY

Major field: Soil Science

vii

TABLE OF CONTENTS

Page Abstract…………………………………………………………………………….. ii Dedication…………………………………………………………………….…….. iv Acknowledgments………………………………………………………………….. v Vita…………………………………………………………………………………. vii List of Tables……………………………………………………………………….. x List of Figures………………………………………………………………………. xi

1. Introduction……………………………………………………………………… 1

2. Literature review………………………………………………………………… 3 2.1 Soil organic carbon (SOC)……………..……………………………………. 3 2.1.1 Textural controls on SOC storage and dynamics: proposed mechanisms………………………………………………………….……………... 4 2.1.2 Mechanisms of SOC storage: role of soil structure ……...…………… 8 2.1.3 Mechanisms of SOC storage: role of soil mineralogy.….……………. 12 2.2 The Brazilian Cerrados – environment, soils and land use………………….. 16 2.3 Hypotheses…………………………………………………………………... 19

3. Materials and Methods…………………………………………………….……. 20 3.1 Sampling areas……………………………………………………………….. 20 3.2 Experimental design and sampling…………………………………….…….. 23

3.3 Soil analyses…………………………………………………………………. 25

viii 3.3.1 Physical properties…………………………………………………….. 25 3.3.2 Chemical properties…………………………………………………… 26 3.3.3 Soil mineralogy …………………………………….…………………. 27 3.3.4 Mechanisms of SOC retention ………………………………………… 31 3.3.5 Soil micromorphology ………………………………………………… 36

4. Results and Discussion…………………………………………………………... 37 4.1 Soil characterization…………………………………………………………. 37 4.2 Effect of Eucalyptus plantation on SOC and other properties………………. 48 4.3 Textural control on SOC retention………….……………………………….. 68 4.4 Mineralogical control on SOC retention…..………………………………… 86 4.5 Structural control on SOC retention: POM-occlusion and fauna……………. 99

5. Conclusions……………………………………………………………………… 127

Bibliography………………………………………………………………….……. 129

Appendix A. Sampling locations…………………………………………………… 146 Appendix B. Aspects from fractionations and separates ………………….……….. 151 Appendix C. Soil thin sections…….……………………………………………….. 157 Appendix D. Original data (excerpts)……………………………………………… 171

ix

LIST OF TABLES

Number Page 3.1 Climatic data from the sampled areas…………………….……….. 21 3.2 Geographical location and silvicultural practices…………………. 24 4.1 Physical characterization of native soils…………………………... 38 4.2 Chemical characterization of native soils…………………………. 41 4.3 Munsell colors of selected depths of sampled soils……………….. 47 4.4 Summary of ANOVAS for soil chemical characterization………... 49 4.5 Summary of ANOVAS for SOC and other properties in bulk soil... 50 4.6 Summary of ANOVAS for SOC in particle size separates………... 54 4.7 Summary of ANOVAS for SOC and C/N in aggregates………….. 57 4.8 Summary of ANOVAS for free- and occluded-POM…………….. 63 4.9 Summary of ANOVAS for SOC stocks and litter layer…………… 65 4.10 Yields and SOC contents of particle size separates……………….. 78 4.11 Quantitative mineralogy of clay fractions…………………………. 87 4.12 Correlations between mineralogy and other properties in clay……. 89 4.13 Linear relations between aggregation and texture…………………. 99 4.14 Linear relations between SOC and aggregation …………………... 101 4.15 Mean SOC and C/N of faunal aggregates…………………………. 124 D.1 C and N, textural and bulk density data…………………………… 172-5 D.2 Yield, SOC and C/N of particle-size fractions……………………. 176 D.3 Specific surface area data of bulk soil, clay and silt fractions…….. 177 D.4 Data from quantitative clay mineralogy…………………………… 178 D.5 WSA and respective SOC and C/N data from 0-5 cm…………….. 179-81 D.6 WSA and respective SOC and C/N data from 30-40, 90-100 cm…. 181-3

x

LIST OF FIGURES

Number Page 2.1 Ideal partition of SOC in a hypothetical soil………………………… 7 2.2 Spatial distribution of the Cerrados in Central Brazil……………….. 16 3.1 Altimetric map of study area………………………………………… 20 3.2 Flow chart of mineralogical analyses……………………………….. 30 3.3 Flow chart of analyses for mechanisms of soil retention……………. 35 4.1 pF curves of selected depths of native soils…………………………. 40 4.2 X-ray diffractograms of silt and clay, selected depths……………… 43-45 4.3 Munsell colors of clay fractions…………………………………….. 46 4.4 C/N ratios of native soils (all depths) and as affected by land use….. 51 4.5 Profiles of mean weighed diameter and SOC concentration………… 53 4.6 C/N ratio of particle size fractions, 0-5 cm depth…………………… 54 4.7 SOC partition in particle size pools, 0-5 cm depth…….……………. 55 4.8 Size distribution and sand content of WSA, 0-5 cm depth………….. 58 4.9 Mean SOC and C/N ratios of WSA, 0-5 cm depth………………….. 60 4.10 Percent and C/N ratios of occluded-POM, 0-5 cm depth…………… 61 4.11 Sand-corrected SOC concentration of WSA, 0-5 cm depth…………. 62 4.12 Overall free- and occluded POM 0-5 cm depth……………………... 64 4.13 SOC stocks and litter layer as affected by land use…………………. 65 4.14 Linear relation between SOC and texture, all depths……………….. 69 4.15 Mathematical functions and fit of the textural control of SOC ……... 71 4.16 Specific surface area of <2 mm soil, selected depths……………….. 73 4.17 Linear relations between surface area, texture and SOC……………. 74 4.18 Fit of model for specific surface area as a function of texture-SOC… 75

xi 4.19 Graphical view of the SOC dilution effect…………………………... 79 4.20 Percent distribution of SOC through particle size pools……………. 81 4.21 Mathematical functions and model fit of clay-sized SOC pool……... 83 4.22 C/N ratios of particle size separates of native soils, selected depths... 85 4.23 Specific surface area of clay fractions, selected depths……………... 88 4.24 Linear relations between bulk SOC and Fe/Al oxides………………. 91 4.25 Linear relations between bulk SOC and gibbsite, kaolinite, illite…… 92 4.26 Linear relations between clay-bound SOC and Fe/Al oxides……….. 93 4.27 Linear relations between clay-SOC and gibbsite, kaolinite, illite…… 94 4.28 Linear relations between texture and aggregation…………………… 100 4.29 Linear relations between SOC and aggregation……………………... 102 4.30 Size distribution of WSA in native soils, all depths………………… 104 4.31 SOC concentration of WSA in native soils, selected depths………… 105 4.32 Sand content of WSA in native soils, selected depths………………. 106 4.33 % Occluded POM of WSA in native soils, selected depths…………. 108 4.34 Sand-corrected SOC concentration of WSA, selected depths………. 109 4.35 C/N ratio of WSA in native soils, selected depths…………………... 111 4.36 C/N ratio of occluded POM in native soils, selected depths………… 112 4.37 Total free- and occluded POM in native soils, selected depths……... 114 4.38 Linear relation between % free-POM and clay content……………... 115 Figures in Appendixes A.1 Map of sampling locations in Unaí, MG…………………………….. 147 A.2 Landscape view of sampling locations in Unaí, MG………………... 147 A.3 Eucalyptus and Cerrado plots on clayey Halustox in Unaí, MG……. 148 A.4 Sampling of litter layer………………………………………………. 148 A.5 Map of sampling locations in João Pinheiro, MG…………………… 149 A.6 Landscape view of sampling locations in João Pinheiro, MG………. 149 A.7 Eucalyptus and Cerrado plots on loamy Haplustox , J. Pinheiro, MG. 150 A.8 Eucalyptus and Cerrado plots in Quartzipsamment, J. Pinheiro, MG.. 150 B.1 Aspects of water-stable aggregates …………………………………. 152

xii B.2 Microaggregates of Quartzipsamment………………………………. 153 B.3 Microaggregates of loamy Haplustox……………………………….. 153 B.4 Microaggregates of clayey Haplustox……………………………….. 154 B.5 Silt fraction of loamy Haplustox…………………………………….. 154 B.6 Sand and occluded-POM from WSA of sandy Haplustox…………... 155 B.7 Sand and occluded-POM from WSA of clayey Haplustox………….. 155 B.8 Common and faunal WSA>2 mm…………………………………… 156 C.1 Semi-thin sections of common WSA>2 mm………………………… 158 C.2 Semi-thin sections of biogenic WSA>2 mm and tunnels…………… 159 C.3 S-matrix of Quartzipsamment ………………………………………. 160 C.4 S-matrix of sandy Haplustox………………………………………… 160 C.5 S-matrix of loamy Haplustox...……………………………………… 161 C.6 S-matrix of clayey Haplustox……………………………………….. 161 C.7 S-matrix of Quartzipsamment ……………………………………… 162 C.8 Runiquartz in loamy Haplustox……………………………………… 162 C.9 Fe-oxide nodule in clayey Haplustox………………………………... 163 C.10 Mn-oxide nodule in clayey Haplustox………………………………. 163 C.11 Mn-oxide coating in clayey Haplustox……………………………… 164 C.12 Unknown mineral in clayey Haplustox……………………………… 164 C.13 Possible aggrotubule in common ped of clayey Haplustox…………. 165 C.14 Planar void in loamy Haplustox…………………………………….. 165 C.15 Unknown mineral in aggrotubule of clayey Haplustox……………... 166 C.16 Runiquartz in aggrotubule of loamy Haplustox……………………... 166 C.17 Aggrotubule (detail) in Quartzipsamment ………………………….. 167 C.18 Occluded-POM in aggrotubule of loamy Haplustox………………… 167 C.19 Fecal pellet in loamy Haplustox……………………………………... 168 C.20 Occluded-POM in fecal pellet from clayey Haplustox……………… 168 C.21 Aspects of charcoal occluded in a WSA>2 mm, loamy Haplustox…. 169 C.22 Root remains occluded in a WSA>2 mm, Loamy Haplustox………. 170

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1. INTRODUCTION

Soil organic carbon (SOC) has long been studied because of its important and beneficial influence on soil fertility, water retention, and other properties of agricultural or environmental significance. The apparently infinite chemical forms and diverse physical states in which SOC occurs is an inherent consequence of the complexity of organic chemistry and biochemistry that have inspired decades of intensive research worldwide. With the increasing concern about anthropogenic global warming mostly due to CO2 emissions, the storage and dynamics of SOC under different land uses has received even more attention due to the significant potential of soils to act as a sink or

source of atmospheric CO2. Texture, or size distribution of primary particles, is perhaps the most influential characteristic of a soil, directly or indirectly affecting practically all its physical, chemical and biological properties and processes. The quantitative distribution of the primary particle sizes, namely sand, silt and clay, and their respective mineralogical constitution, are indicative of the primordial soil-forming factor, the parent material, and the main pedogenetic processes and intensity. Consequently, the relation between soil texture and its quality for a specific purpose has been known by land users throughout human history. Sandy soils are widely known by their usually low fertility and low water retention, while very clayey soils may pose problems of slow internal drainage and poor trafficability. The mechanisms by which texture affects soil physical properties and fertility have been extensively studied and are satisfactorily explained, but not in relation to SOC pools and dynamics. The global, regional and even local variability in the five factors of soil formation allied to the multiplicity of physical and chemical SOC forms has rendered it difficult to formulate a comprehensive theory of SOC formation, partition, dynamics and management, such as the well-explained behavior of plant macronutrients in different-

1 textured soils. Although the effect of soil texture is generally recognized as a direct relation between clay and SOC contents, the exact mechanisms responsible for it are neither clearly understood nor widely quantified in the literature. The same can be said about the other key soil property, mineralogy, which refers to the type of minerals comprising the different particle size fractions (especially clay). The effect of clay mineralogy on soil chemistry and physics is adequately established, but its influence on SOC is often contradictory in the literature and not fully understood. Soil structure refers to the organization of primary particles into secondary particles, known as aggregates or peds. It depends on texture per se and is closely related to SOC, in such a way that a study of the texture-SOC relation is certainly incomplete without considering soil structure. In highly weathered soils of the humid tropics, soil structure is considerably different from that in soils of temperate regions. The most plausible cause of this discrepancy is the mineralogy of the clay fraction, commonly dominated by low-activity kaolinite and various oxides of Al and Fe. The relatively low number of studies on the mineralogy/structure of tropical soils has posed an additional degree of difficulty to the understanding of the SOC dynamics. Last but not least, most studies assessing the relations between soil texture, mineralogy, structure and SOC are restricted to the upper or arable layer of soil, although most of the SOC stock is contained in subsoil layers, that are rarely sampled and studied. Tropical savannas comprise a large fraction of the world land area and play an important role in regional agricultural production. In Brazil, the neotropical savanna formations are called Cerrados. Because of its immense area and central geographical location, the Cerrados share border7s with all other major Brazilian biomes and include many variants of savanna vegetation. Although currently responsible for a significant part of national food production, much remains to be learned about the impact of agriculture and forestry on soil properties in the Cerrados, especially SOC levels and dynamics. The objective of this research is to mechanistically investigate the textural, structural and mineralogical controls on SOC storage and dynamics in the profiles of three different soils developed under similar climate and slope, and under two land uses (native vegetation and Eucalyptus afforestation) in the Brazilian Cerrados.

2

2. LITERATURE REVIEW

2.1 Soil organic carbon

Soil organic carbon (SOC) can be considered as the C part of soil organic matter (SOM), which is the remnants of plant, microbial or faunal biomass that were produced or introduced into the soil. More specifically, SOC is a continuum of biochemical compounds in variable degrees of decomposition, ranging from comminuted residues of recognizable origin, to simple or polymeric substances representing an intermediate decomposition status, and finally to highly changed, large molecules of complex composition, the humic compounds (Anderson and Ingram, 1992; Gregorich et al., 1996). Many workers have extensively studied the composition of different components of SOC. For a comprehensive review on SOM components, refer to Beyer (1996). For humic substances, refer to Schulten and Leinweber (2000) and to a special issue of Soil Science (v.166, n. 11, 2001). The resistance to decay and residence time of SOC usually increases with the degree of alteration: relatively fresh organic residues are easily decomposed by fauna and microbes, resulting in net mineralization of nutrients and C. On the other hand, humic substances are resistant to decomposition and thus little energy or nutrients are available from them, although carboxyl and other pH-dependent functional groups in their structure adsorb or chelate mineral nutrients and chemical compounds in large amounts. Therefore, SOC in the broad sense can be responsible for a considerable part of nutrient stocks and cation exchange capacity (CEC) (Dick and Gregorich, 2004), especially in highly weathered soils. Because of the beneficial impact of SOC on soil quality, SOC sequestration was suggested by Lal (2004) as a mitigation strategy for anthropogenic global warming, as an example of a win-win situation. The most studied abiotic factors affecting SOC concentrations are climate, particularly temperature and precipitation (Jenny, 1941; Parton et al., 1987; Burke et al., 3 1989). Within the same climatic zone, the dominant effect on SOC concentration is that of aquic soil moisture regimes (Davidson and Lefebvre, 1993) or poor drainage (Tan et al., 2004), followed by soil texture.

2.1.1 Textural controls on SOC retention and dynamics: proposed mechanisms

The relationship between SOC concentration and soil texture, usually described by the clay content, is widely recognized. Most commonly, studies of local or regional catenas or non-related but neighboring soils show that coarser-textured soils contain less SOC than finer-textured soils. Such relationship has been validated for soils of temperate (e.g. Brown, 1936, cited by Jenny, 1941; Bauer and Black, 1992; Arrouays et al., 1995; Franzluebbers and Arshad, 1997; Kay et al., 1997; Hassink and Whitmore, 1997; Konen et al., 2003; Tan et al., 2004, and many others) and tropical-subtropical regions (Spain, 1990; Feller et al., 1991, 1996; Mendham et al., 2002; Bird et al., 2003, Galantini et al., 2004). In Brazil, this relation has been documented by Lepsch et al. (1994), Silva and Resck (1997), Tognon et al. (1998), Neufeldt et al. (1999), Bernoux et al. (2002), Williams et al. (2002), Zinn et al. (2002), Telles et al. (2003) and Marques et al. (2004). However, comparisons across broad geographic or climatic regions may not show a direct relation between texture and SOC concentration (McDaniel and Munn, 1985; Lugo and Brown, 1993; Hassink, 1997; Rühlmann, 1999; Percival et al., 2000; Müller and Höper, 2004). This apparent contradiction can be ascribed to a wide range of diverse and interactive factors including soil climate, vegetation and drainage, and the use of only clay content as the measure of texture. For tropical soils under a broad range of annual precipitation but with similar drainage and erosion, Feller et al. (1991) found a strong relation between SOC concentration and the fraction <20 μm, but not with precipitation. A very different cause of the invalidity of the positive clay-SOC relation is the occurrence of low-clay, high-SOC Spodosols, as reported in Maine by Davidson and Lefebvre (1993), by Silver et al. (2000) in the Amazon forest, and in a more extreme case, by Vejre et al. (2003) for 1-m depth SOC stocks of 140 Danish soils. The clay-SOC relation may also not be valid in some recently glaciated areas (Schjønning et al., 1999).

4 Despite the importance of the relation between texture and SOC, a considerable uncertainty exists about the exact mechanisms by which clay content affects SOC retention, and the relative importance of each mechanism among different soils. Clayey soils are more efficient than sandy soils in adsorbing and inactivating C-degrading enzymes (Bremer, 1965), which would hypothetically result in higher equilibrium SOC concentrations. Munn et al. (1978) concluded that sandy soils in sub-humid areas had lower water retention and consequently lower biomass input and SOC concentration than finer-textured soils. Comparing different uncultivated tropical soils, Feller et al. (1996) concluded that total clay content is directly related to SOC (“texture effect”) for all soil particle-size separates (clay, silt, sand), but it was not clear if this trend was caused by the higher biomass input or the higher aggregation of clayey soils in relation to coarser ones. Hassink and Whitmore (1997) concluded that interactions between clay and microbial biomass and products are important controls of SOC dynamics. Indeed, Müller and Höper (2004) found a positive correlation between clay content and microbial biomass. Oades (1988) opined that since clay content is related to so many properties, it is difficult to prove that the clay vs. SOC relation is causative. However, he suggested that finer soils have more cation bridges (Ca in alkaline, Fe and Al in acidic soils) to bind organic molecules to clay particles. This idea implies an assumption that all or most organic and mineral surfaces are negatively charged, which is not necessarily true since variable charges occur in both organic and mineral (e.g., kaolinite edges and Fe and Al oxides) colloids. In his seminal review on physical SOC fractionation, Christensen (1992, p. 61) concluded that the major effect of high clay and silt contents is to stabilize SOC compounds formed through microbial decomposition of organic debris, which in sandy soils would be readily lost to leaching or further decay. Most likely, many of those processes occur simultaneously in soils and isolating a specific one can be challenging, due to complex, non-linear variability involved in sampling a broad range of soil textures. Probably as a direct consequence of the textural effect on SOC retention, the dynamics of SOC turnover under natural vegetation or cultivation is also strongly affected by the clay content. The CENTURY model assumes that SOM decomposition decreases, and stabilization of SOC increases with increasing contents of clay+silt

5 (Parton et al., 1987). Virgin soils with lower clay contents are usually associated with more rapid or strong decline in SOC upon conversion to agriculture or intensive forestry in the tropics (Dalal and Mayer, 1986; Feller et al., 1991, Spacinni et al., 2001) and sub- tropics (Oades, 1988; Dominy et al., 2002). In Brazil, stronger SOC losses in cultivated coarse-textured soils were documented by Lepsch et al. (1994), Silva et al. (1994), and Zinn et al. (2002), among others. Chan et al. (2003) noted that light-textured, hard-setting Australian soils continue to lose SOC even under conservation tillage. This agrees with the meta-analyses by Zinn et al. (2005a), who pointed out that Brazilian soils with <200 g clay kg-1 soil lose ca. 20% of their SOC stock (0-40 cm) even under land uses without annual tillage such as pastures and plantation forestry. As a result of the clay-mediated SOC stabilization, clayey soils tend to have a higher δ13C signal than sandy soils (Bird et al., 2003). In the Amazon, Telles et al. (2003) found that SOC in clayey Oxisols had a much higher turnover time (700-400 yr) than in Ultisols and Spodosols (300-100 yr) in the 0-40 cm layer. For short-term SOM decomposition, the protective effect of clay seems to occur for relatively more stable SOC forms rather than for more labile forms, as -1 shown by Wang et al. (2003) with a range of soil textures: CO2/SOC (mg g in 10 days)= 12-0.0085*clay (g kg-1). Müller and Höper (2004) reported a negative correlation

between clay content and metabolic quotient (indicated by CO2 emission). For a climatic- textural range of Great Plains soils, Kaye et al. (2002) found that fine-textured soils have a much higher pool of stable SOC, calculated as the % of SOC remaining after 1 yr of in vitro incubation at 350C. The texture effect on SOC retention and dynamics can perhaps be better explained conceptually from the following perspective. The continuum nature of SOC (Kogel- Knabner, 1993) implies that non-living C forms range from comminuted but recognizable debris to colloidal components of ordered or complex structure (e.g., celluloses and humic substances, respectively) and simple compounds that are more or less water- soluble (e.g., sugars, acids). Therefore, if the physical states of SOC are solid, colloidal and soluble, one can simplify that non-living C exists (i.e., is retained) in soils as in Fig. 2.1.

6 Total SOC stock

Solid (particles) Colloidal and Soluble

a) Outside b) Within c) Sorbed d) Sorbed e) Free aggregates aggregates on mineral on organic in soil (free) (occluded) particles particles solution

Figure 2.1. Ideal partition of total SOC (excluding biota) in a hypothetical soil.

Mechanisms a, b and c are generally responsible for storing much more SOC than d and e, not treated in this dissertation (refer to the reviews by Kalbitz et al., 2000, and to a special issue of Geoderma in v. 113, 2002). Therefore, the main SOC retention mechanisms are: a) particulation, consisting of organic debris and particles mostly disassociated from mineral soil particles, named free-POM by Besnard et al. (1996); b) particle-occlusion, sand-sized organic debris and particles (usually sand-sized) contained within aggregates, named occluded-POM by Besnard et al. (1996), c) sorption of colloidal-size organic matter to exposed or occluded mineral particles (mostly colloidal), now simply called sorption. (Kaiser and Guggenberger, 2003).

Additionally, the three main C retention mechanisms are temporally and spatially variable. For example, an organic molecule sorbed on an organic particle can be “freed” and then sorbed on a mineral surface, or occluded-POM can become free by aggregate disruption. The relative importance of each of the SOC retention mechanisms is likely to

7 differ with soil forming factors, and with texture and depth. The origin and fate of SOC in subsoil layers is poorly known (Swift, 2001), and it is likely that C retention processes differ from those in surface layers. The sorption process is likely to predominate in subsoil layers (Eusterhues et al., 2004), since the amount of POM tends to decrease with depth (Roscoe et al., 2001). Within the same soil type, different land use and soil tillage practices may also quantitatively alter the relative importance of each mechanism. Obviously, the model presented in Figure 2.1 is an oversimplification, since sharp boundaries between mechanisms or size limits are unrealistic because of their inherent complexity. However, the model can be useful since it considers SOC forms that are easily assessed by particle-size fractionation and aggregate size distribution (so it can be used to interpret data from diverse sources), and does not require information about the almost infinite chemical forms, particle size, density, “lability” and other SOC properties.

2.1.2 Mechanisms of SOC retention: role of soil structure

Factors affecting soil aggregation vary according to soil order and type, but SOC is a common one (Bronick and Lal, 2005). The relation between soil structure and SOC retention mechanisms is clearly inseparable from the discussion of the texture-SOC effect, as implicated by the particulation and POM-occlusion mechanisms outlined in Figure 2.1. As early as the 1940’s, Jenny (1941) wrote “soil structure and organic matter… are properties that often change simultaneously”. More recently, Carter (1996) didactically affirmed that “aggregation and organic matter accumulation are interrelated: organic matter or fractions thereof are basic to the aggregation process, while organic matter sequestered within aggregates is protected against decomposition”. Therefore, the retention of organic C in soils is affected by soil texture directly through sorption, and indirectly through aggregation. Baver (1934) elegantly explained how the degree of aggregation is affected by the contents of silt and clay, and Jenny (1941) affirmed that the amount and type of colloidal clay is intrinsically related to soil structure. The effect of soil texture on structure is obvious: aggregate size distribution and stability depend on the ratio of plasma (mostly clay and silt) to skeletal (mostly sand and

8 gravel) constituents (Buol et al., 1997). While sandy soils have weak or single-grain structure, the size and stability of aggregates increase with increasing clay contents, except if dispersing conditions such as Na saturation occur. SOC stabilization through aggregation is unlikely in sandy soils (Eusterhues et al., 2004), since the low clay contents do not allow significant POM-occlusion and sorption. Thus, SOC in sandy soils is likely to be stored mainly by particulation. Consequently, cultivation of these soils results in more rapid and heavy loss of the unprotected organic particles than observed for fine-textured soils, since higher clay contents allow for considerably higher aggregate stability and sorption. Evidences of these dynamics for tropical soils are presented by Feller et al. (1996), who noted that most SOC lost with cultivation of sandy soils was mainly plant debris in the 20-2000 μm size class, whereas in clayey soils the losses occurred in the <2 μm size class, with no effect of soil mineralogy. Feller and colleagues also reported that after adoption of fallow or meadow, SOC gains occur in the sand-size pool in sandy soils, and in sand- and clay-size pools in clayey soils. There exists well-documented evidence in the literature about the mechanisms of POM occlusion within aggregates, including images obtained by optical or electronic microscope photography of disturbed samples. Images of microaggregates (<250 μm) consisting of mineral particles (mostly clay and silt) total or partially occluding decayed organic particles were presented by Oades and Waters (1991), and Angers and Chenu (1998). Macroaggregates (>250 μm) consisting of organic particles, such as roots and straw, encapsulated by mineral particles are illustrated by Oades and Waters (1991), Golchin et al. (1994), Angers and Chenu (1998), and Golchin et al. (1998). The latter describe the transient process by which POM becomes coated with mineral particles as adsorption, although adhesion is a more appropriate term, as used by Besnard et al. (1996). The fungal and bacterial colonization of fresh POM produces mucilage and hyphae that enmesh and aggregate mineral particles; therefore, less decomposable materials would have a lesser aggregation effect (Lynch and Bragg, 1985; Angers et Chenu, 1998). Some workers propose that POM-occlusion is a major storage mechanism in temperate soils. Nevertheless, Besnard et al. (1996) showed that for silty soils in

9 France, most POM (78% in forested and 58% in arable soils) occurs free rather than occluded in aggregates, which also shows the effect of tillage in degrading free-POM. In some cases, SOC retention through aggregation can be confused with sorption. Tisdall (1996) suggested a different type of C occlusion, i.e. organic compounds diffusing and being deposited within pores small enough to prevent microbial attack. These SOC forms are likely to be identified as sorbed C or clay-bound C in most fractionation procedures. In their review, Jastrow and Miller (1998) described the sorption process as being mediated by polyvalent cations, H-bonds, van der Walls forces and interaction with hydrous oxides. Occlusion within expansive clays may also occur. Therefore, the stabilization of altered organic matter by clays occurs by sorption but also by aggregation, with no clear distinction between both processes (Guggenberger and Kaiser, 2003). Imaging of sorption or occlusion of colloidal-size SOC within aggregates poses much more difficulty than imaging of occluded-POM, and in thin sections this colloidal SOC appears as amorphous and isotropic dark impregnations within the soil matrix. The role of aggregation in C dynamics is by no means homogeneous in all soils. Feller et al. (1996) affirmed that the “literature is rich in conflicting results of organic C” and aggregate-size fractions. Indeed, some authors report that macroaggregates are SOC- rich (Elliott, 1986, Cambardella and Elliott, 1993; Puget et al., 1995) while others point to microaggregates (Bossuyt et al., 2002). This contradiction, also mentioned by Angers and Chenu (1998) for soils in temperate regions, is obviously due to widely different environmental conditions and experimental procedures. For example, when Feller et al. (1996) and Jastrow (1996) applied the correction for sand content and sand-sized debris suggested by Elliott et al. (1991) to aggregates of tropical and temperate soils, all size fractions exhibited similar C contents. This observation suggests that SOC sorption is similar among different aggregate sizes, and that SOC differences are due to sand and POM-occlusion. Also, the separation of free-POM by flotation in water or denser solutions is critical to interpreting such data, especially for the microaggregates in which free-POM can be counted as occluded, especially if only dry sieving is used. Thus, the interpretation of independent works describing SOC partition through aggregate sizes

10 must first consider different climate, drainage, soil texture and mineralogy. Furthermore, one must also consider the following experimental procedures: 1) dry or wet sieving; 2) degree of slaking for wet-sieving; 3) post-sieving separation of free (interaggregate) POM by flotation or hand-picking; and 4) correction for sand and sand-size SOC. The study of the texture-structure-SOC relation in tropical soils is further complicated by the fact that these soils may not conform to what is known for their more intensively studied temperate counterparts. A widely used model of soil aggregation states that highly stable microaggregates (<250 μm diameter) bind into macroaggregates (>250 μm), which are less stable and subject to disruption by tillage (Edwards and Bremer, 1967). This model was proven adequate by many authors for Alfisols and Mollisols (Tisdall, 1996). Oades and Waters (1991) consolidated the concept of aggregate hierarchy and concluded that, unlike Alfisols and Mollisols, tropical Oxisols do not follow this response. However, this conclusion is undermined in part by the fact that they compared aggregate stability of very clayey Oxisols with that of much coarser soils of other orders. This is also the case in the report by Denef et al. (2002). The study of aggregation and SOC dynamics in soils of different taxonomic order or mineralogy must consider a priori the textural effect on these properties, which is not always the case. Nevertheless, it is commonly accepted that the concept of macroaggregate and microaggregate and their hierarchy may not be fully applicable for highly-weathered soils, or at least to clayey Oxisols. Other physical properties related to the structure of different-textured soils also play an important role in SOC retention and dynamics. High sand contents are associated with low water retention, resulting in poor plant growth (i.e., low C accretion) and low microbial activity (reducing SOM decomposition). In vitro incubations with simulated optimal temperature and moisture conditions can mask the effect of different soil textures in the decomposition of SOC or litter, which could otherwise be detected by in situ trials where soil texture affects temperature and moisture variations. Perhaps because of this, Giardina et al. (2001) found no effect of substrate quality (conifer/broadleaf residues) and clay content (70 to 390 g kg-1) on C mineralization in vitro. The role of soil mineralogy and SOC as aggregation factors is discussed in the following section.

11 2.1.3 Mechanisms of SOC retention: role of soil mineralogy

Soil mineralogy is a major determinant of physical and chemical properties, mainly due to the large differences in surface area and charge of minerals in the clay fraction. Thus, as stated earlier, clay minerals affect SOC retention and dynamics by particle-occlusion and sorption processes. Nevertheless, the effect of clay type on SOC retention and dynamics is much less clear than of total clay content. Although studies with highly crystalline clays often indicate stronger, intralamellar sorption to smectites (Varadachari et al., 1991, 1995), field data indicate that the activity of non-allophanic1 soil clays, as determined by surface area and charge, is not directly related to the SOC retention capacity. Hassink (1997) did not observe any effect of clay mineralogy on SOC retention, but his data set contained much non-discriminated variability since it comprised a review of independent works in 4 continents. Moraes et al. (1995) estimated similar SOC stocks to 1-m depth for Vertisols and most Oxisols in the Brazilian Amazon. Comparing six kaolinitic and six smectitic tropical soils under natural savanna vegetation, Wattel-Koekkoek et al. (2001) found no difference in SOC concentrations in bulk soil and clay-sized separates. For three clayey soils of southern Queensland, Krull and Skjemstad (2003) observed that the Oxisol contained much more total SOC to 140 cm depth than the Vertisols. For the same region, Dalal and Mayer (1986) found similar SOC stocks (0-10 cm depth) for 5 clayey Vertisols and one sandy loam Alfisol. However, they attributed the similar SOC stocks to a 5 to 20-fold higher biomass production on the Alfisol, and the C turnover period for that soil was also 5 to 20–fold faster. Gonzalez (2002) studied the dynamics of 14C-marked residues added to a Hapludoll, and detected no difference in C contents between the coarse clay fraction (mostly illite and feldspars) and the smectitic fine clay (<0.2 μm), which nevertheless was the most rapid in sequestering newly formed SOC compounds. In Mozambique, Wattel-Koekkoek et al. (2004) noted that upland, well-drained kaolinitic soils had much higher SOC and a

1 The presence of non-crystalline clay minerals such as allophane in volcanic soils is associated with very high SOC concentrations (Motavalli et al., 1994, Torn et al., 1997). 12 slightly higher mean C residence time (14C analysis) than poorly-drained lowland Vertisols that would supposedly contain even more SOC. Perhaps as a partial consequence of the weak relation between clay mineralogy and SOC, and despite the faster organic matter decomposition expected in the tropics, earlier literature suggests that tropical soils dominated by low-activity clays have SOC concentrations similar to temperate, less-weathered soils. Sanchez (1976) reported that tropical Oxisols and temperate Mollisols have similar SOC concentrations up to 1 m depth, although in the 0-15 cm depth Mollisols contain ~20% more SOC than Oxisols. Sanchez et al. (1982, cited by Sanchez and Logan, 1992) found no differences in mean SOC concentration (0-1 m) in 61 tropical and 45 temperate soils, as well as among tropical Oxisols-Alfisols-Ultisols and temperate Mollisols-Alfisols-Ultisols. Hydrated polyvalent cations such as Fe3+ and Al3+ promote flocculation of negatively-charged clays (Tisdall, 1996). The pedogenic polymerization of these Fe and Al species results in the formation of amorphous and crystalline oxides, hydroxides and oxi-hydroxides (herein to referred simply as oxides). Such oxides occur in many soils of the world, more commonly in the form of goethite and gibbsite, respectively (Bigham et al., 2002, Huang et al., 2002). Due to their net positive charge at the pH values of most soils, these oxides may also enhance flocculation of phyllosilicates and soil aggregation, which can have an impact on SOC retention. Highly weathered tropical soils can have high contents of Fe/Al oxides in the clay and silt fractions, which can impart physical and chemical properties that are not common in phyllosilicate-dominated soils. The effects on soil properties are determined by the ratio of Fe/Al oxides to phyllosilicates in the clay fraction, and also by the dominant oxide (Fe or Al). In some Oxisols of Southeastern Brazil, gibbsite plays a more important structural role than Fe-oxides (Ferreira et al., 1999ab), since it comprises most of the oxide fraction. However, some Oxisols are dominated by Fe- rather than by Al-oxides. Fe oxides can be studied more or less easily by selective dissolution, magnetic susceptibility, and densitometry (Jaynes and Bigham, 1986). Ammonium oxalate is used for selective extraction of amorphous Fe oxide forms, whereas citrate-bicarbonate-dithionite (CBD) extracts both amorphous and crystalline forms. Amorphous Al can be estimated in the oxalate extracts and gibbsite can be

13 determined by thermal analysis, although its study is more difficult than that of Fe oxides due to its high chemical stability (Bigham, J.M., 2003, personal comm.), low density and weak magnetic susceptibility. It is generally believed that soil structure and aggregate stability are strongly affected by high contents of Fe/Al oxides (Oades and Waters, 1991; Feller et al., 1996). Oades and Waters (1991) concluded that these oxides are responsible for the high stability and absence of aggregate hierarchy in Oxisols (see discussion in 2.1.2). The shape of aggregates is also affected by the content of oxides, thereby impacting other physical properties. Studying the B horizons of 7 clayey Oxisols, Ferreira et al. (1999a; b) noted that soils dominated by gibbsite tended to have small granular structure, whereas considerable amounts of kaolinite caused the development of small subangular blocky structure. For kaolinite contents >800 g kg-1 clay, soils were characterized by much higher bulk densities and lower macroporosities and hydraulic conductivities. Similar trends were reported by Schaeffer et al. (2004). Accordingly, intensive cultivation seems to cause more compaction of kaolinitic than gibbsitic soils. Souza et al. (2003) observed higher bulk density (~1.6 g cm-3) and lower macroporosity for a kaolinitic, loamy Oxisol than for a gibbsitic, clayey Oxisol (0-40 cm), both under >30-yr sugarcane cultivation. While SOC data were not presented, SOC stocks are likely to differ in the kaolinite- or gibbsite-dominated Oxisols because of such wide differences in structure and density. The specific role of Fe or Al oxides in SOC retention in the tropics has also been investigated by more direct approaches. In spite of, or even because of a weak relation between clay activity and SOC, there is some indication of enhanced SOC retention by oxides. The technique of high gradient magnetic separation has been used to separate the ferrimagnetic, iron-enriched fraction from the iron-depleted, paramagnetic fraction of clay separates (Schulze and Dixon, 1979). Hughes (1982) determined SOC and oxalate- Fe/Al in clays of tropical soils, and reported that the magnetic separates were richer in Fe-oxides and organic C than the tailings, while the Al-rich separates contained less SOC. Studying an Alfisol and an Oxisol of similar clay contents (180 and 190 g kg-1) in the semi-arid area of Northeastern Brazil, Shang and Tiessen (1998) found that the fractions with magnetic susceptibility >1.38 Tesla (T) comprised about 50% of the total

14 SOC in the clay fractions of both soils. The authors discussed the role of Fe oxide crystallinity, size and association to kaolinite, and concluded that more C is retained by the kaolinitic clay in the Oxisol than by the K-feldspar-dominated clay in the Alfisol. Further study of the same Oxisol (Shang and Tiessen, 2000) showed that the intermediately magnetic fractions contained ca. 50% more SOC than the <0.25 and >1.38 T fractions. The kaolinitic soils studied by Wattel-Koekkoek et al. (2004) probably

contained more SOC than the Vertisols also because of the much higher contents of Fecbd. There seems to exist today two opposing schools of thought, one suggesting that aggregation is controlled by texture and mineralogy (including Fe/Al oxides). The other argues that SOC and biotic activity are the main aggregation factors. For Oxisols, some propose that termite or ant activity through geologic time is the major factor in aggregation (Schaeffer, 2001;Van Breemen and Buurman, 2002). Both views are probably right for specific cases but wrong for general situations. Traditionally, these ideas were based on correlations between aggregate size distribution and data such as

clay, SOC, Fecbd contents (e.g. Chesters et al., 1957, and many others), and on micromorphology. Although a proper discussion is given in section 4.5, a critical issue for selective extractions is discussed now. Studying 14 Oxisol profiles in Minas Gerais, Muggler et al. (1999) found that Fe oxides occur as: a) impregnations (red or yellow background color), b) isolated hematite droplets of 1 to several μm, and c) as coatings around pores. This suggests a ubiquitous presence of Fe oxides in the soil fabric.

However, by particle-size determination after H2O2 and H2O2-CBD treatments, the authors concluded that organic matter plays a larger role than Fe oxides in the aggregation of most soils. Since the CBD extraction for that study was done after SOM removal, it is possible that some aggregation caused by Fe oxides-SOC complexes was not detected, i.e. part of the Fe could have been simultaneously removed. Shaw et al.

(2003) used H2O2 and CBD treatments independently and concluded that Fe-oxides cause more aggregation than SOC in Piedmont Rhodic Paleudults. Simply put, when SOC is removed, some Fe/Al oxides (esp. amorphous) are also extracted; when Fe oxides are extracted with CBD, SOC is also extracted, which must always be considered in planning and interpreting this kind of experiment.

15 2.2 The Brazilian Cerrados – environment, soils and land use

Neotropical savannas occupy large areas of many countries in South and Central America. In the Northern hemisphere, they constitute the Llanos of Colombia and Venezuela, the coastal savannas of the Guianas, Colombia and Atlantic Central America, and sparse Amazonic savannas (Sarmiento, 1984). However, the largest neotropical savanna occurs in Central Brazil, mostly south of the Equator, where it is called the Cerrado or collectively Cerrados. Its central geographical location means that this biome shares boundaries and transitions with other dominant ecosystems in Brazil, namely the Amazonian forest, the hypoxerophytic Caatinga, the Atlantic rainforest and the subtropical, Araucaria-dominated forests (Figure 1.).

Figure 2.2. Spatial distribution of the Cerrados in Central Brazil. Area in detail shown in Figure 3.1. Map courtesy of Embrapa Cerrados. 16 The Cerrados occupy mainly the relatively high Central Brazilian Shield (Pre- Cambrian granite) and parts of the lower, adjacent Paraná, São Francisco and Amazonian basins (Salamuni and Bigarella, 1967). The specific tectonic events responsible for this uplift range from Pre-Brazilian (unknown age) on the Amazonian fringe to the Brazilian cycle (900-500 M yrs) in the southeast (Ferreira, 1971). In many areas, the shield is at the surface or under non-conformed strata, and the important tectonic fold belt (600 M yrs) near Brasília result in the highest altitudes of the Cerrado biome, of ca. 1200 m a.s.l. (Trompette, 1994). The climate in most of the Cerrados is tropical warm and humid with mean precipitation about 1,000-1,500 mm/yr, with a well-defined dry season in winter (Prance, 1987). In terms of plant physiognomy, the word Cerrado is as imprecise as sabana, since both include a range from pure grass cover to dry forests, most commonly an intermediary form (Sarmiento, 1984), and many different types of riverine formations. The many different tree physiognomies and their spatial distribution in the Cerrados are reviewed by Oliveira-Filho and Ratter (2002). The number of woody species in the Cerrado exceeds 500, much higher than in the Venezuelan and African savannas (Franco, 2002), and a much higher number of herbaceous species also occur, most of them perennials (Filgueiras, 2002). The high biodiversity of fauna and flora in the Cerrados, and other neotropical formations, is commonly explained by the Theory of Refugia, synthesized by Ab’Saber (1992) as follows: during the Quaternary glacial ages, temperature in the neotropics decreased little but precipitation was drastically affected, causing flora and fauna to retract to moister “refugia” (usually dissection lines), where speciation was accelerated due to the environmental stress. When the climate became more favorable (interglacial periods), the refugia expanded and coalesced, and the numerous newly evolved species in different refugia expanded their distribution. In this dissertation, the term Cerrado implies a savanna woodland with variable but discontinuous cover of grass and trees, the latter often with twisted stems (see Appendix A, Figures 3 and 6-8). Cerrado trees usually are evergreen, with deep taproots allowing some transpiration even during the dry season (Franco, 2002). The main factors involved in the evolution and characteristics of the Cerrado vegetation are probably the

17 high precipitation with pronounced dry season and recurrent fires. Grass charcoal in deep soils has been dated to the Tertiary, and palynological evidence of simultaneous maximal occurrence of grass and trees (Byrsonima and Curatella sp.) during the Pliocene and Miocene point to the existence of savanna vegetation over the last 3 M yrs (van der Hammen, 1983). The highly weathered Cerrado soils, mostly oligotrophic, acidic and Al- saturated also played a role in the evolution and morphology of the Cerrado vegetation. Many native species accumulate Al in leaves without apparent negative effects (Franco, 2002), and the failure of exotic crops under these unfavorable conditions has inhibited the agricultural development of the region. Cerrado soils are in general more acidic, but also more SOC-rich than soils in African and Venezuelan savannas, and although greater biomass occur in more fertile Cerrado soils, that is not necessarily a cause-effect relation (Montgomery and Askew, 1983). Soil texture is considered by Lopes (1983) as the foremost physical property of Cerrado soils, closely related to water retention, CEC, and phosphorus fixation. Batista and Couto (1992) reported that the sand content is strongly and negatively correlated with the distribution and individual size of the three most important tree species (Qualea sp., Stryphnodendron sp., and Anadenanthera sp.) in a Cerrado gradient in Northern São Paulo State. The dominant soils in the region are Ustic Oxisols (Brazilian Latossolos, occupying 46 % of the total Cerrado area), followed by Ustic Ultisols (Argissolos) and Quartzipsamments (Neossolos), each covering ca. 15% of the area (Adámoli et al., 1986). Lal (2004) suggested that the Cerrados offer a large potential for SOC sequestration, an idea supported by the meta-analyses of Zinn et al. (2005a) for soils with >200 g clay kg-1. Land use in the Cerrados has always been determined by its isolated, central location in South America and by climatic/edaphic factors. The history of human occupation in the Cerrados was reviewed by Klink and Moreira (2002). The first hunther- gatherers of the Itaparica culture flourished approximately 9,000 yrs before present (YBP), and by 6,500 YBP they were replaced by modern indigenous populations. In the early 18th century, Portuguese colonization was motivated by gold mining, resulting in the foundation of sparse villages (including Paracatu) that later shifted their activity to extensive cattle grazing. Grazing escalated from 1920-30, and after 1960 cash crops were

18 finally introduced. The population in the Cerrados has grown to ca. 18 M in 2001, greatly stimulated by the construction of the new capital Brasília in 1960. It is estimated that about 1/3 (672,000 km2) of the Cerrados area of ca. 2,000,000 km2 has been cleared, mostly for planted pastures (453,000 km2). Currently, the Cerrados are a major agricultural region, responsible for the production of 40% and 22% of the nation’s soybean and corn, respectively, and ca. 33% of the bovine herd. The steel industry was introduced during the 1940-1950’s in the State of Minas Gerais, initially consuming wood charcoal from native Cerrado and forest vegetation and, after 1980, from extensive Eucalyptus plantations. In 1998, the area under Eucalyptus plantations in that State was approximately 1.52 M ha, slightly more than half of the nation’s total, and which in great parts supplies the State’s demand for ca. 73% of the national charcoal production (Waniez et al., 2000). A large part of these plantations are located in the Cerrados of Minas Gerais.

2.3 Hypotheses

As stated in p. 2, this dissertation aims to assess the textural, mineralogical and structural controls on SOC retention. The following hypotheses were formulated: a) in tropical savanna soils under similar climate, vegetation and slope, SOC levels are controlled by texture, mineralogy and structure; b) the textural, mineralogical and structural controls on SOC levels can be mathematically described and modeled; c) the relative importance of the three major mechanisms of SOC retention (sorption, particulation, POM-occlusion) is controlled by texture, mineralogy and structure; d) depth is a major factor affecting the textural, mineralogical and structural controls on SOC retention and its mechanisms, in a predictable way; e) Eucalyptus afforestation is a non-intensive land use system that can preserve antecedent SOC levels or even sequester SOC.

19

3. MATERIALS AND METHODS

3.1 Sampling areas

The study areas are located near the geographical center of the Cerrado region, in the districts of João Pinheiro and Unaí, Northwest of the State of Minas Gerais, near the border with the State of Goiás and the Federal District (Figs. 3.1 below, and 2.2).

Figure 3.1. Altimetric map of the study area. Approximate sampling areas circled in black. From www.multimaps.com.

20

Some climatologic data from the stations of J. Pinheiro and Paracatu, located ca. 50 km south from both sampled areas (Figure 3.1), are shown in Table 3.1. Both areas are characterized by the Aw climate of Köppen, i.e. tropical humid, megathermic (Epamig/Embrapa, 1998).

Station and period / Annual June December Monthly Monthly Variable mean mean mean mean min. mean max. J. Pinheiro (1961-1990) Temperature (oC) 22.5 19.9 22.8 19.9 (Jul) 23.8 (Oct) Precipitation (mm) 1441.5 5.2 280.5 5.2 (Jun) 280.5 (Dec) Total evaporation (mm) 1518.2 129.4 90.4 74.4 (Mar) 201.5 (Aug) Air relative humidity (%) 70.1 67.3 78.9 58.4 (Jul) 78.9 (Dec) Sunshine (hrs) 2596.1 254 114.1 114.4 (Dec) 272.9 (Jul) Paracatu (1974-1990) Temperature (oC) 22.6 19.4 23.2 19.2 (Jul) 24.2 (Out) Precipitation (mm) 1438.7 6.7 324.1 6.7 (Jun) 324.1 (Dec) Total evaporation (mm) 1314.3 103.7 77.8 77.8 (Dec) 162.9 (Sep) Air relative humidity (%) 74.2 73.9 81.6 63.0 (Sep) 88.7 (Jan) Sunshine (hrs) 2106.8 192.2 135.6 135.6 (Dec) 222.1 (Jul)

Table 3.1. Climatic data from the sampled areas (MARA/SNI/DNMet, 1992).

21 All soil and litter samples were collected from plantation and preservation areas owned by V&M Florestal Co. The native vegetation areas consist of 25 m-wide strips between Eucalyptus stands, originally intended to provide inocula of predator insects against potential pests to the plantations. In the Nova Esperança Farm, located in Unaí, the eucalypt/native vegetation pairs were located on a very clayey red/ yellow reddish soil (Appendix A, Figures 1-3). In the Chapadinha Farm, J. Pinheiro, the eucalypt (500 x 500 m) and native Cerrado (500 x 25 m) plots were located on two soil types, along a ca. 3 km-long transition from a coarse-textured, gray soil to a loamy red soil (Appendix A, Figures 5-8). All soil types are located on interfluves, with plain to gently rolling slopes. The soil formation factors resulting in the three different soil types are, therefore, parent material and pedologic time, because topography, climate and vegetation are similar. A 1:5,000,000 tectonic map (Ferreira, 1971) locates both areas within the Brazilian folding (recent Brasilide, ca. 500 M yrs.) and describes the cover as post-Triassic sediments of unknown depth. In J. Pinheiro, the presence of different soil types along short distances is due to a relatively complex lithography. The geologic map at the scale of 1:1,000,000 (MME/DNPM, 1978) indicates that the area of João Pinheiro is characterized by associations of the Três Marias formation (Pre-Cambrian shales and arkoses of the Bambuí Group) and Quaternary detrital layers. A more recent and detailed survey indicates that the region of João Pinheiro also contains discontinuous, irregular Cretaceous sandstones and slates (Areado formation), well-distributed detrital Tertiary coverage (clay, silt, sand) and Quaternary alluvial deposits (clay, silt, sand), from which quartzipsamments, Oxisols and Ultisols are developed (Epamig/Embrapa, 1998). Also according to this source, the area of Unaí is mapped as LVa (yellow-reddish alic Oxisol) developed on clayey/lateritic Cenozoic sediments. Given the altitude and proximity of both areas to the toposequences studied by Marques et al. (2004), the sites in J. Pinheiro almost certainly occur on the transitions of Velhas I to II denudational surfaces, i.e., lower parts or pediments of the Areado sandstone that give origin to the Psamment and loamy Oxisol. According to this geomorphic configuration, the soil from Unaí probably is on or below the South American denudation surface (clayey sediments).

22 The Eucalyptus stands sampled are representative of the silvicultural systems usually employed in the Cerrado region in the decades of 1980 and 1990. All sampled sites were planted to Eucalyptus camaldulensis Dehnh from seeds (current practice involves the use of improved clones obtained by vegetative reproduction). The geographical location shown in Table 3.2 corresponds to the central point of a 3-point (replicates) transect (see Appendix A, Figures 1 and 5) at each site. In J. Pinheiro, the Cerrado is partially dominated by the 70-200 cm tall, bamboo-like grass Actinocladum verticillatum (Filgueiras, 2002), which does no occur in Unaí.

3.2 Experimental design and sampling

A factorial design was adopted, in which the main factor was soil texture at 3 levels (sandy, loamy, clayey) and the secondary was land use at 2 levels (Cerrado and Eucalyptus), with triplicate samples (n=3). This design provides N=18 samples per depth, and the following degrees of freedom: soil: 2, land use: 1, interaction soil vs. land use: 2, and residual: 12. Means were compared by the least significant difference obtained from least squares means analysis. This design was applied only for the part of this work related to land use effects, and not for soil characterization and edaphical SOC controls. Soil samples were obtained during the dry winter season (July 2003), when the clayey soils were dry and hard and the coarse soils loose. To facilitate sampling, the locations within the replicates described in section 3.1 were wetted with abundant water one day before sampling. Undisturbed soil samples were collected with metal cores of 5.0 cm diameter x 5.1 cm height inserted using a 1 m-long auger probe, sampling the depths of 0-5, 5-10, 10-20, 20-30, 30-40, 50-60 and 90-100 cm. A composite, disturbed sample was bulked from samples collected at the same depths with a dutch auger near the first probing hole and in four other points located ca. 2 m distant from the center, in a cross shape. The total number of core and composite soil samples obtained was 3 soils x 3 replicates x 2 land uses x 7 depths =126. The litter layer was sampled in two sub-samples for replication with a 15 cm diameter metallic cylinder (Appendix A, Figure 4). The material was cut with a knife, dried at 60 oC and weighed.

23

Variable João Pinheiro J. Pinheiro Unaí (sandy soil) (loamy soil) (clayey soil) Latitude1, 17o25’36” S 17o24’32” S 17o00’27” S Longitude, 46o03’39” W 46o03’02” W 46o48’27” W Altitude 554 m (±7m) 573 m (±7m) 585 m (±7m) Date of planting November 1989 Dec. 1990 Oct. 1989 Soil preparation Heavy disk harrow, light disk Same Same for planting harrow and chisel on tree strip Fertilization at 600 kg ha-1 of rock phosphate; Same Same planting 100g superphosphate/ seedling Tree spacing 3 X 2 m 3 X 2 m 3 X 2 m After-planting 90 kg.ha-1 of NPK 7-0-32 + Same Same fertilization 1.3% boron at age 1 yr; 3.4 g boron/tree at age 2 yr

Wood production 93 m3 ha-1 103.5 m3.ha-1 143.6 m3.ha-1 (1st cycle, 1997) Mean wood production 61.9 m3.ha-1 52.5 m3.ha-1 (2nd cycle, 2003) 1Topographic data of central point of sampling obtained from GPS on Aug. 25-28, 2003. Silvicultural and production data kindly provided by V&M Florestal Co.

Table 3.2. Geographical location and silvicultural practices of the Eucalyptus stands sampled. Note increasing wood yield in the 1st cycle with increasing clay content.

24 3.3 Soil analyses

3.3.1 Physical properties2

Undisturbed samples. The soil cores were wetted from below and saturated with water for 24 h. Water retention was estimated by centrifugation of cores, based on the principle that “at hydrostatic equilibrium the soil matric potential is equal and opposite to the 2 2 2 2 -1 centrifugal force”, expressed as ψ=[(ρw ω )/2](r – r0 ), where ω is the rotation in rad s (Nimmo et al., 2002). This method has the advantage of requiring a much shorter equilibration time than the tension table or hyperbaric chamber. For the centrifuges used, the water potentials correspond to the following centrifugal accelerations: 540 rpm = 6.1 kPa; 700 rpm = 10.3 kPa; 1300 rpm = 35.4 kPa; 1700 rpm = 60.6 kPa; 2200 rpm = 101.5 kPa; 8500 rpm = 1.51 MPa. After a rotation time of 30 minutes at each potential, the weight of the cores was measured. The log of water columns equivalent to the tension was calculated to obtain a pF curve. After determining water retention at 1.5 MPa, the cores were oven-dried at 105 oC and the bulk density determined by dividing soil weight by the core volume of 100.1 cm3.

Disturbed samples. Size distribution of water-stable aggregates (WSA) was measured by the Yoder method (1936). Disturbed soil samples were air dried in the shade and sieved <8mm, of which 100 g were placed on the top of a 5-sieve set (2, 1, 0.5, 0.25, and 0.106 mm), rapidly wetted and subjected to 30 min. of approximately 2 cm vertical oscillation (16 cycles per minute). The WSA separates were dried at 40 oC with forced air circulation and weighed for calculations of WSA size distribution, WSA 2-8 mm and mean weighted diameter (MWD). In the clayey soil, different types of concretions and nodules were abundant, and were separated from WSA fractions by hand picking with a magnet and tweezers (WSA>1mm) and magnet (WSA 1-0.5 mm). The weight of nodules/concretions and their mean density of 2.68 g cm-3 were then used to correct the size distribution parameters. Since very few WSA>1 mm existed in the two coarser soils,

2 Conducted at Embrapa Cerrados, Planaltina-DF, Brazil. 25 part of the samples <8mm was re-sieved and the material >2 mm was then subjected to wet-sieving, in order to provide more sample of WSA>1 mm for further analyses. After separating soil for the wet-sieving described above, fine earth was obtained by passing the <8 mm samples through a 2 mm-sieve, with the help of mortar and pestle when necessary. Primary particle size (texture) was determined by dispersion of 20 g of the fine earth with 100 g NaOH 0.1 M and shaking for 3 hours, with no other pre- treatments. Coarse and fine sand fractions were separated by sieving, whereas suspended silt (2-20 μm) + clay and suspended clay were obtained with the pipette method after different decantation times according to Stoke’s law, and oven dried at 105oC, (Embrapa Solos, 1997). Particle density was determined by dividing a known mass of fine earth by the volume of solids as determined by addition of absolute ethanol in a 50 ml volumetric flask (Embrapa Solos, 1997).

3.3.2 Chemical properties

Soil chemical characterization was conducted on the fine earth obtained as above. The procedures and extractors used follow the standard for routine analysis in Brazil, as prescribed in Embrapa Solos (1997), and briefly explained bellow. Soil pH in water and KCl was measured after brief agitation of 5 g soil in 12.5 ml of deionized (d.i.) water and KCl 1 M. Exchangeable aluminum, calcium and magnesium were extracted with KCl 1 M (2 g soil in 20 ml solution) and determined by atomic absorption with acethylene gas. Available potassium was extracted with 2 g fine earth in 20 ml of Mehlich-I -1 -1 solution (H2SO4 0.025mol L +HCl 0.05mol.L ), and quantified by flame photometry. Al+H was extracted with 1 mol L-1 calcium acetate buffered at pH 7 (1 g soil in 20 ml solution) and titrated with NaOH 0.1 mol L-1, using phenolphthalein as the indicator and correcting with a blank titration.

26 3.3.3 Soil mineralogy

Soil mineralogy for three selected depths (0-5, 30-40, 90-100 cm) was assessed individually for the primary particle-size fractions. Preparation of primary particles. For the 0-5 cm depth, 18 samples were used (3 soils x 3 replicates x 2 land uses). For the two lower depths, 18 samples (3 soils x 3 replicates x 2 depths) were obtained by a 50:50 mixing of the two Eucalyptus / Cerrado samples. Enough fine earth to produce ca. 10 g of clay was obtained for each of the three replicates, and dispersed with NaOH 0.1 M as explained in sections 3.3.1 and 3.3.4. No pre-treatment was used to remove SOC, since the fractionation used here was also part of the SOC fractionation procedure (see 3.3.4). In the near-neutral pH, clayey Oxisols (the other soils are acidic), carbonates were not detected by effervescence with HCl. The dispersed samples were placed on a set of sieves (500/ 250/ 106/ 53 μm) and gently sieved using water spray and rubber spatula. The clean sand >53 μm retained was oven- dried and weighed, whereas the material <53 μm was placed in an automatic fractionator (Rutledge et al., 1967) to separate the <2, 2-20 and >20 μm fractions by stirring and decantation. The times of decantation for a 10 cm depth, according to Stoke’s Law, were 7 h 30 min and 4 min 35 s for particles >2 and >20 μm, respectively. Clay and silt fractions were sequentially separated by near- surface siphoning and transferred to 20 l buckets. Siphoned silt and residual fine sand were transferred to beakers and oven-dried (fine sand was added to the rest of the sand fraction).

To the large volume of suspended clay in the buckets, MgCl2 0.5 M was added in order to flocculate the clay. Since SOC was not oxidized, flocculation of the SOC-rich samples (0-5 cm) was not achieved before addition of 10% HCl to a pH of around 2. The flocculated clay was then transferred to 250 ml plastic centrifuge bottles, an excess of

MgCl2 was added, and the clay samples were successively stirred-shaken-centrifuged (3- 4 times) to wash and remove excessive salt. Partially suspended clay was flocculated by centrifugation with a drop of 10% HCl, and a complete wash was determined when no precipitate formed after a drop of AgCl was added to the clear suspensions. The Mg-clay

27 was transferred with a spatula to a 150-ml cup, redispersed by sonication (40 W) and a

drop of NaOH 0.5 M, fast-frozen by immersion in liquid N2, and freeze-dried.

Mineralogy of primary particles. The sequence of mineralogical analyses is shown in Figure 3.2. The sand and silt fractions were analyzed by optical microscopy. The silt was also analyzed by powder X-ray diffraction, as described below for the clay fraction. The particle-size fractionation was conducted without having pretreated the soil by removal of SOC and iron oxides, both of major interest in this research. Qualitative and quantitative clay mineralogy analyses would suffer undesired interference with the o presence of SOC in the clay. Therefore, SOM was oxidized with 30% H2O2 at 80 C for 4 h using 3 g of intact freeze-dried Mg-clay. The SOC-free, Mg-clay was then dispersed by ultra-sound, quick-frozen with liquid N2, and again freeze-dried. X-ray diffraction (XRD) patterns were obtained with oriented clay slides. Approximately 0.45 g of SOC-free, Mg-clay was re-suspended with ultrasound in 30 ml of d.i. water, from which 4 ml (60 mg clay) were passed through a Millipore® nitrocellulose filter (0.22 μm, since a 0.44 μm filter allowed Fe-oxides to pass) under vacuum. The oriented air-dried films were then transferred, with help of a water spray,

from the membrane filters to Buehler® glass slides, air-dried and stored in a CaCl2 desicator. CuΚα X-rays were produced by a Philips® XRG-3100 generator (35kV, 20mA), and XRD patterns were obtained with a Philips goniometer at step intervals of 0.050 2θ for 4 s, from 2 to 450 2θ. Clays with patterns containing peaks at 1.4 nm were solvated with ethylene glycol, x-rayed, heated at 350 oC for 4 h, and x-rayed again to determine the nature of the 2:1 phyllosilicates. XRD patterns were stored in digital file in a MDI databox and edited with the software Jade 3.1. Thermogravimetry (TG) was used to quantitatively determine gibbsite, goethite and kaolinite in the peroxide-treated clay, according to Karathanasis and Harris (1994). 0 -1 -1 Briefly, a 15-30 mg sample was heated at a rate of 20 C min under 200 ml N2 min to 650 0C in a Seiko TG/DTA 200 (Torrance, CA), while the weight loss and thermal events were recorded. The calculations used standard weight losses for endothermic

28 dehydroxylation of gibbsite (312 g kg-1, 220-2900C range), goethite (106 g kg-1, 300- 3700C) and kaolinite (140 g kg-1, 400-5900C). Amorphous Fe/Al oxides contents were determined according to Schwertmann (1964). Approximately 1 g of intact (non-peroxide treated) Mg-clay was shaken in 50 ml of NH4-oxalate 0.2 M + oxalic acid 0.2 M (pH 2.92) for 2 h in the dark. Total Fe and Al in the extracts were read with atomic absorption following a 1:5 dilution. Total Fe-oxides (amorphous + crystalline) were determined from the intact (non- peroxide treated) Mg-clay using the citrate-bicarbonate-dithionite (CBD) method adapted from Mehra and Jackson (1960). To ca. 1.0 g of Mg-clay, 40 ml of citrate-bicarbonate buffer was added, followed by immersion in a 80oC hot bath, and then 1 g of Na- dithionite was added to reduce and dissolve the Fe-oxides, with complexation by the citrate. After 10 min. of occasional stirring, another 1 g of Na-dithionite was added, and after another 10 min. the suspension was cooled and centrifuged at 1800 rpm. The clear supernatant was put aside in a 250-ml flask, and the sample stirred and treated again with a second cycle, now using only 20 ml of buffer and no dithionite. After centrifugation, the supernatant was added to the flask, and the volume completed with water. Total Fe and Al in the extract were measured by atomic absorption after a 1:20 dilution, necessary to overcome undesired effects of the CB buffer on the flame. The Fe-extracted, Mg-clay

was re-suspended by ultra-sound, quick-frozen with liquid N2, freeze-dried and stored. Total K in the peroxide treated, Mg-clay was determined by flame emission. A 50 mg sample was totally dissolved in 5ml HF 48% + 0.5 ml aqua regia (3:1 concentrated 0 HCl/HNO3) at 110 C for 40 min. in a pressurized Teflon decomposition vessel, as described by Sawhney and Stilwell (1994). For analysis, 2.8 g of boric acid was added to the extract and the volume completed to 100 ml total with d.i. water. The content of illite

was calculated assuming a total K2O content of 10%. Specific surface area was determined by the BET technique (Brunauer et al., 1938) in a Micromeritics® Flowsorb 2300 (Norcross, GA), using up to 150 mg of freeze- dried Mg- and fine earth, both without peroxide removal of SOC. Briefly, the sample was 0 de-aired and exposed to a flow of He and N2 at –198 C, allowing for maximum N2 condensation and monolayer sorption on the solid. After fast warming to room

29 temperature, the amount of desorbed N2 was measured, and the surface area calculated from the molecular cross-section area calibrated for experimental temperature and pressure. The surface area was then divided by the sample weight to compute the specific surface area.

Bulk samples <2mm (0-5, 30-40, 90-100cm, n = 3 soils x 3 rep x 3 depths = 27)1

Dispersion in 0.1 M NaOH, shaking, sieving, decantation

Sand separates Silt separates Clay separates (>20μm, n=27) (n=27) (n=27)

-Naked eye -Microscopy -30%H2O2, -Total Fe-oxides CBD -Microscopy -powder XRD then XRD -NH4oxal. Fe/Al oxide -C/N analyzer -C/N analyzer and TG - C/N analyzer

Information provided: - Qualitative/quantitative mineralogy of sand, silt and clay fractions - Role of clay mineralogy in SOC and specific surface area

1 For the 0-5 cm depth, samples of Eucalyptus and Cerrado were fractionated separately (for use in 3.3.4); then size fractions were combined for mineralogical analyses. For the other depths, samples of Eucalyptus and Cerrado were combined before fractionation. XRD – X-ray diffraction; TG – thermogravimetry.

Figure 3.2. Flow chart of mineralogical analyses.

30 3.3.4. Mechanisms of SOC retention

Wet or dry combustion is the standard method for quantifying total C in bulk soil samples (Nelson and Sommers, 1996). However, the results thus obtained are of little or no value for a qualitative description of SOM. For this purpose, a variety of chemical and physical fractionations and derivate analyses are available in the literature, each with their own pros and cons. Chemical methods are more commonly used for studying “humic” or specific organic compounds, and are not discussed here. The most authoritative review on physical methods for soil and SOC fractionation is the one by Christensen (1992), summarized below. Chemical dispersion procedures (the standard method for textural analyses) are not appropriate to obtain intact primary organomineral complexes, assumed to represent the actual association of SOC and individual mineral particles. Ultrasonic dispersion causes little chemical alteration of SOC compounds and can be calibrated (for each soil type) to produce results similar to standard chemical dispersion methods. However, sonication can abrade organomineral complexes, mineral particles and, most likely, POM. After dispersion, different organomineral complexes are usually separated by density or particle-size fractionations. Density procedures are based on the continuum of SOC-mineral bonding strength, varying from the “light” (non-complexed POM) to “heavy” (SOC strongly bound to minerals, i.e. clay) fractions. However, the density limits for light and heavy organomineral complexes vary confusingly in the literature from 1.6 to 2.8 g cm-3, in such a way that Christensen concluded that density fractionations have a number of unsettled problems on their basic conception and experimental procedures, especially the very high densities ascribed to non-complexed POM. Particle-size fractionations are based on wet-sieving of the dispersed sample (to remove sand-sized minerals and POM) and sedimentation based on Stoke’s Law (to separate clay from silt). This principle is conceptually sound compared to density separations, despite the partial validity for Stoke’s Law assumptions, and the common result that the silt fraction is often found to be the most SOC-rich fraction.

31 Although in some cases attributed to incomplete dispersion, the high-SOC content in silt is physically and chemically difficult to explain when the silt fraction consists of chemically inert primary minerals such as quartz with low surface area and charge density, not likely to bond with SOC. Another common cause for this probable artifact is the ultrasonic disintegration of POM by high-input ultrasound dispersion and its subsequent accumulation in the silt fraction (Feller et al., 1991, 1996; Tisdall, 1996). Indeed, higher C concentration in the silt-sized fraction was verified by Zinn et al. (2002) for the two soils in J. Pinheiro after ultrasonic dispersion, although these soils contain negligible silt as determined by standard dispersion in NaOH0.1 N. Other conceptual problems are related to the density and particle-size fractionations described above, especially in assigning certain biochemical properties for specific fractions. The assumption that “light” or coarse SOC is more readily decomposable than “heavy” or clay-sized SOC is often wrong due to the presence of recalcitrant components in the light/coarse fractions (peat, charcoal, conifer needles etc.) and to significant loss of clay-sized SOC upon cultivation (Christensen, 1992). Considering all points discussed above, particle-size fractionation by sieving and Stoke’s Law sedimentation was chosen as the most appropriate procedure for this research. However, because of the plausible comminution of POM through ultrasound treatment, it was desirable to use an alternative dispersion procedure. The option was to disperse the samples with NaOH 0.1 M, and shaking for 3 h without sonication, for the following reasons: 1) this is the standard procedure used for textural analysis in Brazil, so the results obtained here are directly applicable to Brazilian soils and literature; 2) the objective of this research is the quantitative determination of SOC with no chemical characterization other than C/N ratios; 3) total dispersion of oxide-rich Oxisols may not be achieved with ultrasound (Oorts et al, 2005); and 4) the samples thus prepared can also be simultaneously used for mineralogical analyses. Therefore, the dispersion of bulk samples was conducted according to section 3.3.3, using individual samples for Eucalyptus and Cerrado for the 0-5 cm depth (where SOC changes are likely to occur) and a 50:50 mix of Eucalyptus/Cerrado samples for the 30-40 and 90-100 cm (where

32 SOC changes are slight). The sand and silt fractions thus obtained were dried at 50 0C, whereas the clay fraction was quick-frozen and freeze-dried (see 3.3.3). Another objective of this fractionation was to quantify the role of POM-occlusion within water-stable aggregates (WSA) as a mechanism for SOC retention. The aggregation of mineral particles around decomposing POM has been documented for Mollisols and Alfisols in the temperate region (see 2.1.1), but little data exists on its importance for soils of the humid tropics. The method suggested for addressing this issue is similar to the procedure by Beare et al. (1994ab), which in its turn is an adaptation of that by Cambardella and Elliott (1993). In the latter, soil samples dispersed in NaHMP were sieved <53 μm, and SOC was measured on the evaporated slurry and compared with SOC in non-dispersed soil (the difference assumed to be equal to POM). Similarly, Beare and associates sieved dispersed and non-dispersed samples, but using WSA size fractions from conventional tillage and no-till plots on a Piedmont Rhodic Kanhapludult. It is important to note that, in the present study, the free-POM within WSA fractions was not representative of the free-POM in the <2 mm soil because: a) the wet- sieving operation per se causes POM flotation on the water surface and this material was removed by hand; and b) dry-sieving the two coarser-textured soils in order to obtain more samples of WSA >1 mm artificially increased free-POM in the samples before wet- sieving. In other words, free-POM in these WSA fractions cannot be considered as an accurate measure. Therefore, free-POM was estimated by the difference between the total sand-sized SOC in bulk soil <2mm (obtained as in the Soil mineralogy section - 3.3.3) and occluded-POM (weighted sum of sand-sized SOC inside WSA size fractions), previously separated from free-POM. Thus, it is critical to separate all free-POM from the WSA fractions obtained according to section 3.3.1. For WSA >1 mm, this can be fully accomplished by spreading the sample on a petri dish and hand picking the free-POM with tweezers. For WSA<1 mm, separation of free-POM was done by a sequence of electrostatic adhesion and flotation-decantation in water. Electrostatic adhesion was conducted by rubbing the plastic bag containing the sample and allowing the excess free-POM from a scoop of sample to adhere to the plastic bag. Then, 3-4 g of sample were placed in a 250 ml

33 beaker, slowly sprayed with water to avoid slaking, and adding ca. 100 ml water. To allow free-POM to disentangle from overlaying aggregates and float, the beakers were gently shaken, and the WSA were stirred with water pumped with a 20 ml polyethylene transfer pipette. Floating and slow-settling free-POM was removed by decantation and siphoning with the transfer pipette, and the WSA samples were then oven-dried at 500C. Some aspects of the fractionation procedure and occluded-POM separates are shown in Appendix B. Flotation in water is not a common way to separate “light” POM (Christensen, 1992) because not all fresh residues and POM are less dense than water. However, the method described was effective for the samples used: upon observation of dried WSA samples in a light microscope (X20 magnification), the estimated removal efficacy of free-POM was 99% for WSA 0.5-1 mm, and 97-98% for WSA <0.25 mm (Appendix B, Figures 2-4). From the WSA fractions thus obtained, a 1 g sub-sample was separated for C/N analyses. The remainder of the samples were dispersed in NaOH according to 3.3.3, and wet-sieved (20 μm) with a water jet and rubber spatula. The >20 μm fraction, assumed to represent total occluded-POM, was collected and oven-dried at 1050C. Free-POM and occluded-POM were estimated as follows:

j Occluded-POM C = Σi (wsasand w x wsasand-SOC), and Free POM-C = sand-sized SOC in bulk soil – occluded-POM C,

where wsasand w and wsasand-SOC are the total weight and SOC content of particles > 20 μm in the i th WSA size class (i varying from 1 to 5, or >2mm to 0.25-0.11 mm).

Total SOC of ground, 1-g samples of bulk soil and particle-size separates were determined by dry combustion in a Variomax CNHOS analyzer (Hanau, Germany). Total N was simultaneously determined in the same apparatus by Dumas combustion. Figure 3.3 shows the sequence of all analyses for identifying the mechanisms of SOC retention.

34

126 composite soil samples (0-5, 5-10, 10-20, 20-30, 30-40, 50-60, 90-100 cm)

126 bulk soil samples, sieved <8mm

Bulk samples, ground Yoder WSA fractions: 8-2, 2-1, 1-0.5, 0.5-0.25, <2mm 0.25-0.106 mm (n=126*5=630 samples)

Finely ground - Dispersion NaOH 0.1N Separation from POM C/N analyzer Sand, silt, clay fractions (hand pick / shaking in water) (all depths, (0-5, 30-40, 90-100cm, (0-5, 30-40, 90-100cm, n=126) n=1081) n=1802)

C/N Dispersion NaOH analyzer sand fractions n=180

Data provided: Data provided: C/N analyzer - SOC % and stock - Specific effect of forest - Relation to texture, plantation on SOC size aggregation etc. - SOC partition among size Data provided: - Overall effect of classes Free- and occluded forest plantation - Overall C-sorbed to clay POM-C (>20 μm) - Total POM-C (>20 μm)

1Assumes Eucalyptus and Cerrado do not differ in SOC and aggregation in the two lower depths, so samples can be mixed in each replication, resulting in n = (3 soils x 2 l.u. x 3 rep. x 1 depth x 3 fractions) +(3 soils x 3 rep. x 2 depths x 3 fractions) =108. 2 Same assumption: n =(3 soils x 2 l.u. x 3 rep. x 1 depth x 5 fractions) + (3 soils x 3 rep. x 2 depths x 5 fractions) =180.

Figure 3.3. Flow chart of analyses for mechanisms of SOC retention.

35 3.3.5 Soil micromorphology

During the hand separation of concretions from WSA of the clayey Oxisol (see 3.3.1), it was observed that the predominant type of soil structure was subangular blocky. However, a significant part of the 2-8 mm WSA in the 0-5 cm depth showed very different morphological features, probably derived from faunal activity (Appendix B). The WSA 2-8 mm samples from sandy soils and loamy Oxisol were then carefully examined and both showed the same type of faunal peds. Therefore, 2-8 mm WSA from the 0-5 cm depths of the three soil types, representing the biological features were manually separated from those with subangular blocky structure. These two types of WSA, along with subangular blocky peds from the 30-40 and 90-100 cm depths, were prepared for micromorphology and CN analyses. An epoxy electric resin (Scotchcast® No. 3, 2A/1B), prepared and de-aerated under vacuum, was then used to impregnate oven-dried (105 oC), 2-8 mm WSA placed in a 50 ml cup. The impregnated samples were evacuated for 3 days to remove all air from pores in the WSA and allow full resin penetration. The cups were then heated at 105 oC to cure and harden the resin. The hardened resin blocks were sanded and polished on a kerosene-lubricated lap wheel with a series of Buehler® silicon carbide abrasive powders until reaching an adequate section plan for study. After polishing on a water-lubricated, 30 μm spinning diamond saw, the resin blocks were rapidly mounted on warm (50 oC) glass slides with de-aerated Hillquist® epoxy resin 7A/3B, heated for 1 min. at 105 oC, and cured at room temperature (heat after warming causes differential expansion and damages the mounts). After curing, the excess resin was removed with a diamond saw to a thickness of 40-60 μm and then polished with a 600 grit Buehler® silicon carbide powder. Low magnification (8-30 X) images of the whole WSA were obtained with a Nikon® SMZ645 binocular microscope coupled to a Nikon® Coolpix 990 digital camera. The samples were further lapped to ca. 30 μm, and polished with 400 and 600 grit Buehler® silicon carbide powder. High magnification images were obtained with a Nikon® UFX-II petrologic microscope coupled to a Nikon® FX-35A camera.

36

4. RESULTS AND DISCUSSION

4.1 Soil characterization

The characterization of soils under native Cerrado is given here as an initial description the studied area (i.e., not considering land use and the factorial design at this time). Table 4.1 shows some physical properties for selected depths. As expected, there is considerable difference in texture among the three soil types, with little variation within the same type, as indicated by the standard errors. However, within the sandy soil type, the clay content is always less than 150 g kg-1 for replicate 1, but that value is exceeded in subsurface of replicates 2 and 3. This is important since 150 g clay kg-1 is the limit for the distinction between a Quartzipsamment and an Oxisol, and thus these data were presented separately in Table 4.1. To understand the difference in clay content among the replicates of sandy soils, consider that replication 1 is the southernmost pair of plots on the toposequence in J. Pinheiro (Appendix A, Figure 5), and represents the typical Quartzipsamment. Replicates 2 and 3 represent an intermediate with the loamy red soils, but are texturally much more similar to the Quartzipsamment and always contain <200 g clay kg-1, a critical limit deduced by Zinn et al. (2005a) for coarse-textured Brazilian soils that are highly susceptible to SOC loss upon cultivation. Clay contents also tend to increase with depth for all soils. Soils were not sampled by horizon so it is not possible to detect a Bt horizon; however, textural horizons are unlikely since the increases in clay content are gradual. Therefore, the higher clay contents in subsoil layers are more likely to result from appauvrissement than by lessivage processes, as described by van Wambeke (1992, p. 104-106). Silt contents are below the detection level for the sandy soils and are only significant in the clayey soils. Because of the negligible porosity of the soil skeletal components, bulk density increases directly with the sand content.

37

Depth Clay Silt Fine sand Coarse Bulk Particle Porosity (cm) sand density density (g kg-1) (g kg-1) (g kg-1) (%) (g cm-3) (g kg-1) (g cm-3) Sandy soil, rep. 1 0-5 70.0 0.0 620.0 310.0 1.24 2.76 55.1 10-20 90.0 0.0 660.0 250.0 1.41 2.83 50.2 30-40 80.0 0.0 640.0 280.0 1.24 2.91 57.4 90-100 100.0 0.0 570.0 330.0 1.24 2.76 55.1

Sandy soil rep. 2-3 0-5 130.0(10.0) 0.0 530.0(40.0) 340.0(30.0) 1.15(0.06) 2.79(0) 58.8(2.1) 10-20 130.0(10.0) 0.0 555.0(55.0) 315.0(45.0) 1.20(0.08) 2.87(0) 58.2(2.8) 30-40 160.0(0) 0.0 430.0(10.0) 410.0(10.0) 1.30(0.11) 2.94(0.09) 55.6(4.7) 90-100 155.0(25.0) 10.0(10.0) 410.0(50.0) 425.0(35.0) 1.34(0.11) 2.98(0.09) 54.9(4.6)

Loamy soil 0-5 296.7(14.5) 13.3(3.3) 236.7(14.5) 453.3(21.9) 1.03(0.05) 2.78(0.02) 62.8(1.9) 10-20 333.3(8.8) 13.3(6.7) 196.7(14.5) 456.7(8.8) 1.05(0.05) 2.86(0.02) 63.3(1.5) 30-40 370.0(5.8) 6.7(3.3) 183.3(8.8) 440.0(10.0) 1.09(0.03) 2.93(0.07) 62.8(0.5) 90-100 386.7(3.3) 0.0 173.3(12.0) 440.0(11.5) 1.24(0.06) 2.95(0.08) 57.8(2.4) Clayey soil 0-5 406.7(38.4) 213.3(8.8) 93.3(20.3) 286.7(14.5) 1.09(0.11) 2.78(0.02) 60.9(4.2) 10-20 496.7(18.6) 200.0(15.3) 70.0(15.3) 233.3(27.3) 1.07(0.04) 2.86(0.02) 62.4(1.4) 30-40 493.3(23.3) 166.7(6.7) 103.3(31.8) 236.7(39.3) 1.26(0.08) 2.93(0.07) 56.8(2.8) 90-100 466.7(49.1) 150.0(15.3) 123.3(54.6) 260.0(10.0) 1.17(0.11) 2.95(0.08) 60.6(3.2)

Table 4.1. Physical characterization of soils (<2 mm) under native Cerrado (mean of 3 replicates, except for sandy soils), with standard errors shown in parentheses.

38 The volumetric moisture content at different pF values for selected depths is shown in Figure 4.1. Moisture retention is generally proportional to the clay content and the curves for each soil type are distinct, although the loamy and clayey soils had similar moisture contents at pF>2.5 for the 90-100 cm depth. Table 4.2 shows chemical properties of soils under native Cerrado. Similar to Table 4.1, the results for replicate 1 of the sandy soils are shown separately for comparative purpose. The sandy and loamy soils are characterized by similar pH (in water) values indicating a strongly acid reaction, whereas the clayey soil had pH values above 5.0 for all depths. In all cases, pH in KCl is less than in water, indicating that an acric character (predominance of surface positive charges) does not occur in any soil at any depth. Nutrient status and CEC are low and representative of Cerrado highly weathered soils. Replication 1 of the sandy soils had slightly higher exchangeable bases than replications 2 and 3, which in conjunction with lower CEC resulted in a higher base saturation. The loamy soils had slightly higher base saturation and CEC than sandy soils, whereas the clayey soils had the highest CEC and base saturation. There is a clear trend of increase in CEC with increasing clay content. Exchangeable acidity (Al3+ and Al3+ + H+) also tended to increase with clay content, and contributed to most of the CEC for all soils and depths. In all cases, the main component of the CEC is H+ (Al3+ subtracted from Al3++H+), suggesting that hydroxyl and carboxyl groups of soil organic matter are responsible for most of the exchange complex in Cerrado soils, as reported by Silva et al. (1994), and many other authors. The mineralogy of particle-size separates permitted a more complete soil characterization. Gravel (2-8 mm) occurred only in the clayey soils, in the form of nodules and concretions, some magnetic, composed of secondary minerals in which Fe, Al, and Mn oxides are the cementing agents. These materials also comprise a variable part of the sand fraction in the clayey soils, although quartz is the main component. The sand fractions of the sandy and loamy soils are mostly quartz with variable but small amounts of accessory minerals. The reader is referred to Appendixes B and C for some microscopic aspects of the sand and gravel fractions.

39 0.6 0-5 cm 0.5 )

-3 0.4 Sandy, Cer. m 3 Loamy, Cer. 0.3 Clayey, Cer. 0.2 Theta (m 0.1

0.0 0.00 1.78 2.00 2.52 2.78 3.00 4.18 pF

0.6 30-40 cm

0.5 )

-3 0.4

m Sandy, Cer. 3 0.3 Loamy, Cer. 0.2 Clayey, Cer. Theta (m 0.1

0.0 0.00 1.78 2.00 2.52 2.78 3.00 4.18 pF

0.6 90-100 cm

0.5 )

-3 0.4

m Sandy, Cer. 3 0.3 Loamy, Cer. 0.2 Clayey, Cer. Theta (m 0.1

0.0 0.00 1.78 2.00 2.52 2.78 3.00 4.18 pF

Figure 4.1. Moisture retention curves of selected depths of soils under native Cerrado. Bars represent standard error.

40

Depth pH pH Ca2+ Mg2+ K+ Al3+ Al3++ CEC Base

(cm) + H2O KCl H Sat. Sandy soil, rep. 1 ------cmol(+) kg-1------% 0-5 4.67 4.22 0.96 0.44 0.22 0.63 3.01 4.63 34.9 10-20 4.62 4.1 0.33 0.10 0.08 0.59 1.33 1.84 27.9 30-40 4.59 4.13 0.26 0.05 0.04 0.58 0.93 1.28 27.6 90-100 4.7 4.15 0.14 0.04 0.07 0.61 1.32 1.55 15.4 Sandy soils rep. 2-3 0-5 4.5(.08) 3.9(.01) 0.6(.02) 0.3(.02) 0.13(.01) 1.1(.24) 5.8(1.0) 6.8(1.1) 14.5(1.5) 10-20 4.3(.08) 4.0(.01) 0.4(.22) 0.1(.06) 0.10(.01) 1.2(.03) 3.6(.56) 4.1(.84) 12.5(4.5) 30-40 4.4(.04) 4.0(.01) 0.3(.17) 0.1(.06) 0.09(0) 1.0(.01) 2.5(.38) 3.0(.61) 16.5(4.5) 90-100 4.6(.03) 4.1(.01) 0.2(.01) 0.1(0) 0.07(.01) 0.8(.08) 1.8(.04) 2.2(.06) 16.0(0) Loamy soil 0-5 4.6(.01) 3.9(.03) 0.6(.15) 0.5(.05) 0.17(.01) 1.6(.07) 7.9(.50) 9.3(.67) 14.7(0.9) 10-20 4.6(.02) 4.0(.01) 0.1(.07) 0.1(.03) 0.11(.01) 1.0(.48) 4.9(.17) 5.2(.20) 6.0(2.0) 30-40 4.7(.03) 4.0(.01) 0.4(.04) 0.1(.03) 0.10(.01) 1.3(.03) 4.1(.04) 4.7(.02) 12.7(0.3) 90-100 4.6(.04) 4.1(.02) 0.2(.03) 0.0(.02) 0.07(.01) 1.2(.11) 2.7(.04) 3.0(.06) 10.0(1.2) Clayey soil 0-5 5.3(.14) 4.2(.20) 3.6(1.2) 2.9(.56) 0.58(.02) 0.9(.32) 8.0(.70) 15.0(1.0) 45.7(7.8) 10-20 5.1(.08) 4.0(.13) 1.1(.43) 0.8(.19) 0.42(.05) 1.6(.29) 7.0(.22) 9.3(.60) 24.3(5.5) 30-40 5.1(.06) 4.0(.12) 0.8(.29) 0.5(.1) 0.30(.01) 1.6(.35) 6.2(.51) 7.8(.28) 20.3(5.2) 90-100 5.2(.14) 4.2(.22) 0.4(.1) 0.3(.04) 0.27(.09) 1.5(.78) 4.6(1.2) 5.5(1.2) 19.0(6.1)

Table 4.2. Chemical characterization of soils (<2 mm) under native Cerrado (mean of 3 replicates, except for sandy soils), with standard errors shown in parentheses.

41 The powder X-ray diffraction patterns (XRD) indicate that the silt fractions of all soils are composed mostly of resistant primary minerals, especially quartz, but also rutile (Figure 4.2a, b). Secondary minerals such as anatase are also common, and kaolinite and gibbsite also occur, especially in the loamy Oxisols that also include goethite. The XRD patterns of oriented clay fractions of replicate 1 of the sandy soil and the patterns for the sandy (rep. 3), loamy and clayey soils (rep. 2) are shown in Figure 4.2c to 4.2f. The clay fraction of all soils is dominantly kaolinitic, as seen by the three strong, well-defined peaks for the d=7.2, 3.6 and 2.4 nm Å. However, accessory minerals occur in all soils and become relatively more important with increasingly finer texture, which is clearly demonstrated for the toposequence in J. Pinheiro. Clay in the sandy soil, replicate 1 (Fig. 4.2c) is mostly kaolinite with very small peaks for gibbsite, goethite, quartz, feldspar and aluminum-interlayered vermiculite (AIV, d=14 Å). Replication 3 of the sandy soil show enhanced peaks for AIV, gibbsite, goethite and a weak peak for hematite (Fig. 4.2d). In the loamy soils (Fig. 4.2e), these trends are reinforced in the form of strong peaks for all the accessory minerals, particularly hematite; additionally, a small peak of illite (clay mica) occurred. Identification of AIV as the 14 Å peak was confirmed by non-expansion upon glycolation (i.e., negative for smectite) and its gradual collapse upon heating to 350 0C, indicating the presence of interlayer Al (Fig. 4.2e). In the geographically unrelated clayey soils of Unaí, all of the same accessory minerals occur, but illite is clearly the most important among them (Fig. 4.2f). The maximum content of weatherable primary minerals in the sand fraction is 10% for the order Oxisols and 4% (6% for muscovite) for the order Latossolos (Embrapa, 1999). There is no such limit for the clay fraction, but high illite concentration in the clayey soils (see also 4.5) is unexpected for Oxisols. It is possible that mica domains became resistant to chemical weathering by partial or total coating with Fe/Al/Mn oxides. Also, the slow natural drainage of these clayey soils can result in a relatively weaker leaching and desilication, resulting in further preservation of micaceous clay.

42 d=3.34 q d=4.26 q d=2.46 d=2.28 d=2.23 d=2.13 q q q q d=3.52 d=3.24 d=7.17 d=2.38 d=3.58 d=2.57 d=4.48 d=2.69 d=2.50

a d=2.35 k r a k k go k k a,k Clayey soil

Loamy s oil deg. 10 15 20 25 30 35 40 45 2θ a) d=1.82 q d=1.54 q d=1.37

d=1.38 q d=2.00 d=1.67

q d=1.26 d=1.29 d=1.20 d=1.42 d=1.45 d=7.17 q q q q d=1.23 q q q

q d=1.69 q

d=1.70 a r Clayey soil

Loamy s oil deg. 45 50 55 60 65 70 75 80 2θ b) k= kaolinite; q=quartz, go=goethite; h= hematite; a=anatase; r=rutile. d expressed in Å.

Figure 4.2a,b. X-ray diffraction patterns (CuΚα, powder) of silt fractions (30-40 cm depth) from clayey and loamy soils.

43 d=7.16 d=3.58 k k d=2.38 k d=13.91 d=4.84 d=4.38 v gi gi d=3.38 d=3.25 q f 0-5 cm

30-40 cm

90-100 cm deg. 2 7 12 17 22 27 32 37 42 2θ c) d=7.15 d=3.58 k k d=2.38 k d=13.98 d=4.34 d=4.13 d=2.70 d=3.38 d=3.25 d=2.50 v d=4.79 g g go q h h 0-5 cm

30-40 cm

90-100 cm deg. 2 7 12 17 22 27 32 37 42 2θ d) v=hydroxil-interlayered vermiculite; i=illite; k= kaolinite; g=gibbsite; go=goethite; h= hematite; q=quartz, f=feldspar. d expressed in Å

Figure 4.2 (cont.). X-ray diffraction patterns (CuΚα, Mg-saturated at 250C) of oriented clay fractions of selected depths from a) sandy soil rep. 1; b) sandy soil rep. 3.

44 k d=7.16 d=3.57 k d=2.38 d=10.10 d=14.23 k v i d=4.37 d=4.15 d=3.34 d=3.25 d=2.69 d=2.51 d=4.83 350C g g go d=4.97 q f h h 0-5 cm eg i

350C 30-40 cm

90-100 cm 350C deg. 27 2θ e) d=3.57

d=7.16 k k d=2.38

d=3.32 k d=13.93 d=9.93 i d=4.37 v i d=4.15 d=2.69 d=2.51 d=4.83 d=4.97 i g g go h h 0-5 cm

30-40 cm

90-100 cm deg. 2 7 12 17 22 27 32 37 42 2θ f) v=hydroxil-interlayered vermiculite; i=illite; k= kaolinite; g=gibbsite; go=goethite; h= hematite; q=quartz, f=feldspar; eg=ethylene glycol solvated. d expressed in Å

Figure 4.2 (cont.). X-ray diffraction patterns of oriented clay fractions of selected depths: e) loamy soil (CuΚα, Mg-saturated at 250C); f) clayey soil.

45

Figure 4.3. Munsell colors of the clay fraction.

The Munsell colors of peroxide-treated clay in Figure 4.3 indicate that the triplicates of sandy and clayey soils differ in total and relative amounts of goethite and hematite (see quantitative mineralogy in Table 4.11), but not the loamy soils which have a homogeneous color. However, clays from different depths of the same replicate had the same color. As seen in the XRD patterns, this indicates that clay mineralogy varies little within the 1 m depth of all soil types. Soil color results from the clay mineralogy but also from SOC contents, resulting in different colors down the profile. Table 4.3 shows how dry color of the fine earth varies among and within the three soil types, and with depth. As shown in Figure 4.3, colors are the same among replicates of the loamy soils, but vary in the sandy and clayey soils mostly as a consequence of the ratios of goethite to hematite in the clay fraction.

46

Soil type Replication 0-5 cm 30-40 cm 90-100 cm Sandy 1 10YR 6/2 10YR 7/2 10YR 7/3 2 10YR 5/2 10YR 6/3 10YR 7/3 3 7.5YR 5/3 7.5YR 6/4 7.5YR 6/6 Loamy 1 5YR 4/4 5YR 4/6 2.5YR 4/6 2 5YR 4/4 5YR 4/6 2.5YR 4/6 3 5YR 4/4 5YR 4/6 2.5YR 4/6 Clayey 1 10YR 5/4 7.5YR 5/6 7.5YR 6/6 2 10YR 5/4 7.5YR 5/6 7.5YR 5/6 3 10YR 4/4 5YR 4/4 5YR 4/6

Table 4.3. Soil colors (Munsell, dry) according to soil type, replicate and depth.

Physical, chemical and mineralogical properties indicate that all soils are highly weathered, in consistence with the latitude and climate. Their classification according to the Soil Taxonomy is attempted below. The clayey and loamy soils can be classified as a Haplustox, and will be from now on identified as clayey Haplustox1 and loamy Haplustox, respectively. Replicate 1 of the sandy soils fits in the great group of Quartzipsamment, but not replicates 2 and 3 which can be classified as Oxisols. In most of the following results and discussion, the sandy soils will be statistically treated as a homogeneous group, following the original experimental design. However, to better illustrate the effect of clay content on some properties such as quantitative mineralogy and surface area, the mean of replicates 2 and 3 will be shown separately as in Tables 4.1 and 4.2, and the soils named sandy Haplustox. According to the Brazilian system (Embrapa, 1999) the Quartzipsamment correspond to Neossolo Quartzarênico. Classification of the Haplustoxes at the great group level is not possible without a full description and characterization of the A and B horizons, not conducted in this study. Thus, these soils fall in the order Latossolos, either in the suborders Vermelho (red) or Vermelho-Amarelo (yellow-reddish).

1 Although illite contents in this soil are unusually high for Oxisols (see Fig. 4.2 and Table4.11), the low CEC in subsurface is consistent with a diagnostic (oxic) B horizon. 47 4.2 Effect of Eucalyptus plantation on SOC and other properties

Eucalyptus afforestation has been reported to change soil properties, often deleteriously, all around the world. In his comprehensive review, Lima (1996, pp. 139- 168) concluded, however, that sometimes the impacts on soils can be positive, and remarked that very often the reported soil degradation resulted from poor soil management and fertilization rather than to Eucalyptus growth per se. The time frame for this type of comparison is also important, since the rapid growth of the plantations can result in significant temporal changes in soil properties, as discussed below. Table 4.4 shows summaries of the Analysis of Variance (ANOVA) for chemical properties at selected soil depths. In all cases, all significant differences are due to soil type only and no effect of Eucalyptus plantation is detected. These data contrast with the earlier results by Zinn (1998), who studied the same sandy and loamy soils shortly before the first timber harvest, when the Eucalyptus stands were 7 yr old. At that time, the loamy Haplustox had significantly lower available K at 0-5 cm depth and underwent acidification up to 60 cm depth (as indicated by pH and exchangeable Al and Al+H) compared to the Cerrado, whereas there were no change in the Quartzipsamment. It is possible that the acidification and K-depletion were reversed after the first harvest, when a large amount of wood and leaf debris was left on the ground to decompose, resulting in a gradual release of mineral nutrients that would replenish base levels and decrease Al saturation and pH. Similar results of K replenishment were reported by Mendham et al. (2003) for second-rotation Eucalyptus plantation in Australian sandy soils. Therefore, after the coppice growth that constituted the second rotation of Eucalyptus production, the natural fertility of the soil was restored. Considering that shortly after the sampling in 2003 the coppice forest was harvested for the second and last time, another mass of above- and belowground organic debris was left to decompose on the ground. The decomposition of these residues can eventually result in an even higher nutrient status than under native Cerrado, as a consequence of the fertilizer applied and the efficient nutrient uptake and cycling by the plantation forest.

48

Source of pH pH Ca2+ Mg2+ K+ Al3+ Al3++ CEC Base + variance H2O KCl H Satur. 0-5 cm Soil *** n.s. n.s. *** *** * *** ** ** Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 10-20 cm Soil *** n.s. n.s. *** *** n.s. *** *** * Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 30-40 cm Soil *** n.s. * ** *** ** *** *** n.s. Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 90-100 cm Soil *** n.s. *** *** *** n.s. *** *** n.s. Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s.

Table 4.4. Levels of significance from the ANOVA for soil chemical characterization. * significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant.

Table 4.5 summarizes the ANOVAs for SOC, N and other soil properties at all sampled depths. As for Table 4.4, most significant effects are due to soil type, including bulk density and porosity as discussed earlier in 4.1. It is notable that SOC and N contents were not significantly affected by the Eucalyptus plantation, contrasting with the SOC depletion reported by the end of the first productive cycle in 1997 by Zinn et al. (2002), for the sandy and loamy soils at the 0-5 cm depth. This suggests a recovery to pre-cultivation levels after the second rotation for the same reasons discussed on the preceding page, this time for SOC and N contents. Despite the fertilizer applied, N levels did not change probably due to the strong uptake by the plantation and little return via litterfall, as discussed earlier for exchangeable bases. 49

Source of SOC N C/N MWD WSA Bulk Part. Total >2mm dens. dens. por. variance 0-5 cm Soil *** *** ** *** *** n.s. n.s. n.s. Land use n.s. n.s. *** ** ** n.s. n.s. n.s. Soil*L. use n.s. n.s. * n.s. n.s. n.s. n.s. n.s. 5-10 cm Soil ** *** n.s. *** *** ** n.s. ** Land use n.s. n.s. n.s *** *** n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 10-20 cm Soil *** *** n.s. *** *** ** n.s. ** Land use n.s. n.s. n.s n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 20-30 cm Soil *** *** n.s. *** *** * n.s. * Land use n.s. n.s. n.s n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 30-40 cm Soil *** *** n.s. *** *** n.s. n.s. n.s. Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 50-60 cm Soil *** *** n.s. *** *** * n.s. * Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 90-100 cm Soil *** *** * *** *** n.s. n.s. n.s. Land use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s.

Table 4.5. Levels of significance from the ANOVA for SOC, N, C/N, mean weight diameter (MWD), water stable aggregates (WSA)>2 mm, bulk and particle densities, and total porosity. * significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant.

50 9 101112131415 C/N ratio 20 (bulk soil) 0 10 15 20 Sandy a a 30 10 40 Loamy b b

50 C/N ratio 5 60 c Clayey c 70 0 80 Cer. Euc. Cer. Euc. Cer. Euc. 90 0-5 cm (cm) 100 Sandy Sandy Loamy Loamy Clayey Clayey a) b)

Figure 4.4. Bulk soil mean C/N ratios: a) profiles under Cerrado, and b) as affected by land use (0-5 cm depth). Means with the same letter are not significantly different by the least squares means test of least significant difference (LSMeans). Bars represent standard error.

Despite the non-significant effect on SOC and N, Table 4.5 shows that the C/N ratio increased under Eucalyptus at 0-5 cm depth, contrasting with the decrease reported by Ashagrie et al. (2005) in Ethiopia. The significant soil vs. land use interaction reflected a high initial C/N values and increase in the sandy soils (Figure 4.4b). Table 4.5 shows that soil aggregation, as indicated by the mean weight diameter (MWD) and percent of water-stable aggregates (WSA)>2 mm, decreased under Eucalyptus for the two top depths. This decline was most likely caused by pre-planting cultivation with a heavy disk harrow, and suggests that, although strong aggregation is characteristic of highly weathered soils, recovery of this structure after cultivation is slower than for SOC content. This decrease in macroaggregation was not detected by Zinn (1998), since at that time only soil samples 2-8 mm were wet-sieved, contrasting with the use of soil <8 mm in this study. This result suggests that the pre-planting cultivation for Eucalyptus plantation disrupted large (>2 mm) WSA, but the stability of remaining WSA was not affected. 51 Figure 4.5 a and b show respectively the mean SOC concentration (g kg-1) and the MWD (mm) of WSA along the profile. As expected, there is a distinct and positive effect of clay content on SOC and aggregation, discussed in detail in sub-chapters 4.3 and 4.5. SOC levels are generally distinct for each soil type, and decrease considerably with depth. For the sandy soils at 0-5 cm depth, the mean SOC under Eucalyptus is higher than under Cerrado and their error bars do not intercept. Although this difference is not significant at P<0.05, the actual P=0.14 was much lower than for the loamy and clayey Haplustoxes (P>0.7), suggesting that if a slightly higher replication was used (e.g., n=4), a significant effect could have been observed. Thus, pre-cultivated Eucalyptus plantations may sequester SOC above native levels in sandy soils, which are also the most likely to lose SOC even under land use systems without annual tillage (Zinn et al., 2002, 2005a). The significant changes due to afforestation in MWD in the top 10 cm layer are clearly seen in Figure 4.5b. Below that depth, the loamy and sandy soils had very similar MWDs, whereas the clayey Haplustox had much higher MWDs. The strong decline in MWD below the top 10 cm for all soils contrasts sharply with the more gradual decline of SOC levels with depth (Figure 4.5a). This asymmetry may suggest that aggregation is not necessarily the major control or the best predictor of SOC levels (see further discussion later in this sub-chapter and in sub-chapter 4.5). Table 4.6 summarizes the results of the ANOVAs for SOC concentration and C/N ratios of the different size separates, as well as the particle size SOC pools (percent of total SOC, calculated as the product of the yield and SOC concentration of each fraction divided by the sum of products of the three fractions) for the 0-5 cm depth. As for bulk soil, there was no effect of afforestation on SOC concentrations in particle-size fractions, thus the actual mean values are shown in Table 4.10. However, Eucalyptus plantation significantly increased the C/N ratio in clay and silt separates, as specifically shown in Figure 4.6. Although the ANOVA indicated a significant land use effect for C/N ratio in the sand fraction, this was not shown in the test of means because of the pooled variance used. In the sandy soils, sand C/N under Eucalyptus is different from that under Cerrado at P<0.0537 (very close to the usual limit of 0.05). Also, standard errors show that C/N ratios in the sand are much more variable than those in the two other size fractions.

52 SOC (g kg-1) 2 4 6 8 10 12 14 16 18 20 22 24 26 0 10 20 30 40 50 60 70 80 90 100 Depth, cm Sandy, Cer. Sandy, Euc. Loamy, Cer. Loamy, Euc. Clayey, Cer. Clayey, Euc. a)

MWD (mm) 0.5 1.0 1.5 2.0 2.5 3.0 3.5 0 10 20 30 40 50 60 70 80 90 100 Sandy, Cer. Sandy, Euc. Loamy, Cer. Loamy, Euc. Depth, cm Clayey, Cer. Clayey, Euc. b)

Figure 4.5. Depth distribution of a) mean SOC content, and b) mean weight diameter (MWD) of water-stable aggregates (WSA). Bars indicate standard error.

53 Source of SOC concentration C/N ratio % of total SOC variance 0-5 cm clay silt sand clay silt sand clay silt sand Soil *** *** ** ** *** n.s. *** n.s. *** Land use n.s. n.s. n.s. ** ** * n.s. * * Soil*L. use n.s. n.s. n.s. n.s. ** n.s. n.s. n.s. n.s.

Table 4.6. Levels of significance from the ANOVA for SOC in particle size separates. * significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant.

30 0-5 cm 25 depth

20 a Sand ab ab ab 15 Silt a C/N ratio b a b Clay 10 a a b b 5 ab bc c b b b 0 Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey

Note: despite silt contents were mostly below detection limits of textural analysis in sandy and loamy soils (Table 4.1), silt samples fpr C/N analyses were still obtained from 30-100 g soil samples (see values in Table 4.10).

Figure 4.6. Mean C/N ratios in particle size fractions as affected by land use. For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error.

54 70 0-5 cm . 60 depth a a a a 50 40 a b Sand 30 Silt b c Clay 20 c c c c 10 a

SOC size pools (% of total) abc ab abc bc 0 c Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey

Figure 4.7. Percent of total SOC in particle size fractions as affected by soil and land use. For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error.

Table 4.6 also shows that Eucalyptus afforestation changed the sand and silt-sized SOC pools at the 0-5 cm depth (but not clay). However, the details presented in Figure 4.7 indicate that SOC partition through size fractions did not change under Eucalyptus for the loamy and clayey Haplustoxes. In reality, for the sandy soils, Eucalyptus plantation increased the sand-sized SOC pool by 40% (from 40 to 56% of total SOC, P=0.004) mostly at the expense of the clay-sized SOC pool, which declined from 48 to 37% (P=0.015). Comparing the test of means with the ANOVA shows the following: the change in sand-size SOC pool of sandy soils is so high that it caused an overall significance for land use. Simultaneously, the change in the clay-sized SOC pool for sandy soils was not detected by the ANOVA, because of the high and similar clay-sized SOC pools in the two finer-textured soils. Also, significant effects of land use on the silt- sized SOC pool indicated by the ANOVA were only detected by the test of means at p=0.0715. 55 Based on the SOC and C/N data for bulk soil and particle size fractions shown in the preceding pages, the introductory description of aggregation by MWD and WSA>2 mm is hereby extended to a detailed view of WSA size distribution and properties in the 0-5 cm depth. Table 4.7 summarizes the ANOVAs for SOC concentration and C/N ratios of the different WSA size classes, as well as for the sand fraction from dispersed WSA, occluded-POM (sand-sized SOC within WSA) as percent of SOC in each WSA class, and SOC concentration corrected for sand and occluded-POM. The ANOVA results will be discussed separately and with reference to Figures 4.8-4.11. Pre-planting for Eucalyptus afforestation markedly decreased MWD in all soils in the top 10 cm, as discussed earlier. According to Table 4.7 and Figure 4.8a, this decrease occurred mostly at the expenses of WSA>2 mm class for the clayey and loamy Haplustoxes, and the soil material released was transferred mostly to WSA<1 mm and 0.5 mm of each soil respectively. These trends suggest that the aggregate hierarchy of Oxisols differs with texture, since disruption of large macroaggregates from the clayey Haplustox releases larger fragments than in the loamy Haplustox. For the sandy soils, WSA>2 mm also tended to decrease, although no significance was observed because of the large difference from the two other soils. The sand content in WSA size classes roughly resembles that of bulk soils, but some important differences exist. Figure 4.8b shows that, for the sandy and loamy soils, WSA>1 mm contained much less sand than the smaller WSA. For the clayey Haplustox, sand contents vary little among WSA>0.25 mm, but are higher below that limit. This suggests that in Oxisols <350 g clay kg-1, large aggregates contain more clay, which is probably the main cementing agent for this size, whereas smaller sizes are mostly skeletal grains coated with clay and SOC, of variable extent and thickness (Appendix B, Figures b2-3). In clayey Oxisols, cementation by clay does not change with WSA size, and microaggregates (<0.25 mm) contain more sand because of individual grains (Appendix B, Figure b.4). Although Table 4.7 indicates an effect of land use on the total sand (>20 μm) content in each WSA size class, Figure 4.8b shows that in fact the mean values are very similar and the significant differences are due to extremely low variation among the replicates, so the test of means was not shown.

56

Source of % of SOC C/N % sand Sand Sand % Oc- Sand- total in WSA SOC C/N POM corr. C variance 2-8 mm Soil *** n.s. *** *** ** n.s. * ** Land use ** n.s. n.s. n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 1-2 mm Soil *** n.s. ** *** * n.s. n.s. *** Land use n.s. n.s ** * n.s. ** n.s. n.s. Soil*L. use n.s. n.s. ** n.s. n.s. n.s. n.s. n.s. 0.5-1 mm Soil *** *** ** *** ** * ** *** Land use n.s. ** n.s * n.s. * * * Soil*L. use n.s. n.s. n.s. n.s. n.s. * n.s. n.s. 0.25-0.5 mm Soil *** *** n.s. *** *** n.s. n.s. *** Land use * * * n.s. n.s. n.s. n.s. * Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 0.1-0.25 mm Soil * *** n.s. *** *** n.s. n.s. *** Land use ** n.s. ** n.s. n.s. n.s. n.s. n.s. Soil*L. use n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s.

Table 4.7. Levels of significance from the ANOVA for percent from total, SOC content, and C/N ratio of total WSA and sand fraction within WSA, and sand- corrected SOC content, by WSA size class. * significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant. 0-5 cm depth only.

57 50 0-5 cm ab a depth 40 a a bc 2-8mm 30 a a 1-2mm cd b b b a b 0.5-1mm d b 20 b bc c 0.25-0.5mm b bc 0.11-0.25mm d Percent of soil<8mm . 10 a c a b b b c c 0 c Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey a)

100 90 0-5 cm 80 depth 70 60 2-8mm 50 40 1-2mm 30 0.5-1mm 20 0.25-0.5mm 10 0.11-0.25mm

% sand in WSA size class 0 Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey b)

Figure 4.8. Percent of a) WSA size classes in soil <8 mm, and b) sand in each WSA size class. For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error.

58 According to Table 4.7, the mean SOC concentration changed under Eucalyptus plantation only for the WSA 0.25-1 mm (reminder: free-POM was removed from all WSA samples by flotation in water, and no correction for sand is done at this time). Figure 4.9a shows that these changes actually occurred only in the clayey Haplustox. Along with the transfer of soil material from WSA> 2 mm to WSA<1 mm discussed earlier, these changes suggest that, upon macroaggregate disruption, SOC decomposition occurs mainly in the smaller released soil fragments rather than within partially disrupted large macroaggregates. Table 4.7 indicates that the C/N ratio of three WSA size classes changed with Eucalyptus afforestation, and Figure 4.9b shows these changes in more detail. In the sandy soils, C/N ratio increased in all WSA sizes, whereas in the loamy Haplustox the increase occurred in WSA<2 mm. These results are consistent with the results for bulk soil and particle size fractions (Figures 4.4 and 4.6) and suggest that native Cerrado SOC was replaced by less humified material from the plantations. However, in the clayey Haplustox the only change in C/N ratio was a decrease for WSA 0.5-1 mm, inconsistent with the increase in bulk soil and size fractions. Sand-sized SOC inside the WSA may indicate the role of POM occlusion in stabilization of aggregates. Figure 4.10a and b show occluded-POM C (percent of sand- sized SOC from total SOC in WSA size class) and C/N ratios of the sand retained after dispersion and sieving of the WSA samples. Percent occluded-POM decreased with increasing clay content (see sub-chapter 4.5). Figure 4.10a indicates that land use changed the percent occluded-POM only for WSA 0.25-0.5 mm in the sandy soils, again suggesting a preferential SOM decomposition within fragments released from disrupted large macroaggregates, agreeing with the mechanism proposed by Ashman et al. (2003). The C/N ratio of occluded-POM also changed under Eucalyptus for WSA classes 0.5-2 mm (Table 4.7). Figure 4.10b shows higher values than for intact WSA (Figure 4.9b), and very high and variable ratios for WSA<0.25 mm for sandy and loamy soils. These trends may suggest that occluded-POM is partly composed of charcoal (see also section 3). C/N ratios in occluded-POM increased under Eucalyptus in the sandy soils (WSA>0.5 mm) and loamy Haplustox (WSA 0.5-2 mm), but not in the clayey Haplustox.

59 30 0-5 cm a ) ab a

-1 depth 25 ab ab a ab ab ab ab 20 b b a a b b a 15 bc 2-8mm b b c b 10 c b c b b b 1-2mm 0.5-1mm 5 c c 0.25-0.5mm SOC concentration (g kg 0 0.11-0.25mm Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey a)

19 a a 0-5 cm 18 depth b b 17 bc a aba 16 abc a ab a bcd a cc a abc 2-8mm 15 bc c cdc d 1-2mm b b b 14 b b b 0.5-1mm C/N ratio 13 c 0.25-0.5mm 0.11-0.25mm 12 Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey

b)

Figure 4.9. a) SOC concentration in WSA size classes, and b) C/N ratios in WSA size classes. For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error.

60 20 a a 0-5 cm ab depth 16 a ab abc a bc 2-8mm bc ab ab c 12 1-2mm a bcc bc ab a b a 0.5-1mm a a c a 0.25-0.5mm 8 aaa a a a 0.11-0.25mm

4

0

Occluded-POM (% of total SOC) . Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey a)

90 a 0-5 cm 80 a a depth 70

60 2-8mm ab a 1-2mm 50 ab a a 0.5-1mm 40 a a b a a a 0.25-0.5mm a a a a b ab b 30 bc abc b c 0.11-0.25mm

C/N ratio of occluded POM C/N 20 b b b b b 10 Cer. Euc. Cer. Euc. Cer. Euc. Sandy Sandy Loamy Loamy Clayey Clayey b)

Figure 4.10. a) percent of occluded-POM from total SOC within WSA size classes, and b) C/N ratios of occluded-POM. For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error.

61 80 0-5 cm a a 70 ab depth ) a

-1 a 60 a a a 50 bb 2-8mm bc b 1-2mm 40 bc bc b bc cd bc cd 0.5-1mm c bc 30 cd bc cd bc c 0.25-0.5mm c c 20 d d 0.11-0.25mm

10

Sand-corrected SOC (g kg Cer. Euc. Cer. Euc. Cer. Euc.

Sandy Sandy Loamy Loamy Clayey Clayey

Figure 4.11. SOC concentration of WSA size classes corrected for sand and occluded- POM contents. For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars are standard error.

Figure 4.11 above shows the calculated SOC concentration in WSA size classes after correction for the sand fraction (mineral and organic). This correction was used by Elliott et al. (1991) as an alternative to compensate for the wide variability in SOC levels in aggregate fractions among different soils and land uses, mostly due to varying sand contents. Table 4.7 shows that changes due to land use occurred in WSA 0.25-1 mm, and Figure 4.11 shows more specifically that the changes were restricted to a decrease under Eucalyptus in the sandy soil. Since the sand-corrected SOC reflects the concentration in the clay+silt fractions, this is consistent with the decrease in the percent of clay-sized SOC in that soil (Figure 4.7). The sand-corrected SOC values shown above also corroborate the SOC dilution effect discussed in the next sub-chapter.

62

Source of Free-POM Free-POM Occl.-POM Occl.-POM

variance g kg-1 % g kg-1 % 0-5 cm Soil ** *** * n.s. Land use *** ** ** *** Soil*L. use * * n.s. n.s.

Table 4.8. Levels of significance from the ANOVA for overall free- and occluded-POM. * significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant.

The weighed sum of occluded-POM C within all WSA size classes was subtracted from total POM (i.e., sand-sized SOC pool in the bulk soil), to estimate the general partition of POM outside and inside WSA. Table 4.8 summarizes the results of the ANOVAs for occluded-POM C in absolute (g kg-1) and relative (%) units, and Figure 4.12a and b show the LSMean tests. The sandy soils have considerably more POM C than the other two soils, as noted earlier (Figure 4.12a is in fact a partition of the sand fraction in Figure 4.7). Table 4.8 indicates that the partition of POM C changed consistently with land use. Occluded-POM C as a percent of total SOC decreased under Eucalyptus in all soils, in accord with the decrease in WSA>1 mm to 10 cm depth. However, when absolute concentrations of occluded-POM C are considered, the significant effect occurred only for the clayey Haplustox. Finally, Table 4.9 and Figure 4.13 synthesize the ANOVAs and test of means, respectively, for the litter layer and SOC stocks of three selected depths, in Mg per ha. The average litter layer was not affected by soil type, and in the clayey Haplustox the Eucalyptus forest produced a significantly higher litter stock than native Cerrado. On the other hand, SOC stocks increased substantially with clay content for each depth, with no effect of afforestation, in consonance with the SOC concentration and bulk density values presented earlier.

63 60

. b 50

40 a a 30 a b a b 20 b 10 c c POM (% of total SOC) c c 0 Occluded Cer. Euc. Cer. Euc. Cer. Euc.

Free Sandy Sandy Loamy Loamy Clayey Clayey a)

10 c 8 soil .

-1 6 bc bc a ab bc 4 a b g POM-C kg 2 b b b b

0 Occluded Cer. Euc. Cer. Euc. Cer. Euc. Free Sandy Sandy Loamy Loamy Clayey Clayey b)

Figure 4.12. Overall free- and occluded-POM C in 0-5 cm depth as a) percent of total SOC, and b) absolute values (g kg-1 soil). For the same fraction, means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error.

64

Source of SOC stock SOC stock SOC stock Litter layer

variance 0-20 cm 0-40 cm 0-100 cm Soil ** *** *** n.s. Land use n.s. n.s. n.s. * Soil*L. use n.s. n.s. n.s. n.s.

Table 4.9. Levels of significance from ANOVA for SOC stocks and litter layer (Mg ha-1). * significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant.

100 90 a a ab bc 80 bc c 70 Litter a 60 a 0-20cm -1 0-40cm 50 ab bc bc 0-100cm Mg ha 40 c a a a ab 30 b b 20 a 10 b ab b ab b 0 Cer. Euc. Cer. Euc. Cer. Euc. Sandy Sandy Loamy Loamy Clayey Clayey

Figure 4.13. Litter layer and SOC stocks of selected soil depths (Mg ha-1). Means with the same letter are not significantly different by the LSMeans test (P<0.05). Bars represent standard error. 65 As reviewed by Zinn et al. (2002), the literature shows disparate effects of Eucalyptus afforestation on SOC levels and stocks that can be in most part attributed to: a) different experimental and soil/forest management conditions, and b) control soils under very different native vegetation or degradation status. Both sources of variances were avoided by the experimental design of the present work. When data on SOC concentration and stocks in the sandy and loamy soils in this study are compared to those in Zinn et al. (2002), the importance of stand age affecting SOC levels and stocks is clear: SOC losses in bulk samples and sand/clay fractions for the end of the first cycle (7 yrs) were recovered at the end of the second cycle (14 yrs). This effect of Eucalyptus stand age in re-sequestering bulk SOC concurs with the literature (e.g., Bernhard-Reversat et al., 1996; Paul et al., 2002; Davis and Condron, 2002; Ashagrie et al. 2005), although the latter reported a strong decrease in SOC concentration in size fractions >clay for a clayey, Rhodic Nitisol in Ethiopia. Sand-sized SOC is depleted with agricultural management in 90% of the cases (Hassink, 1997); however, the present work has shown that the sand pool can also be the most likely to recover or sequester C in sandy soils, as reported earlier by Feller et al. (1996). The effects of Eucalyptus plantation on aggregation have seldom been studied. Resck et al. (2000) found that 16 yr-old Eucalyptus stands planted on clayey Cerrado Oxisols had aggregation (WSA>2 mm) levels similar to native vegetation and higher than pastureland. Comparing a native Podocarpus forest with wide-spaced Eucalyptus stands (21-yr) subjected to cattle grazing, Ashagrie et al. (2005) found no differences in size distribution and SOC concentration (not sand-corrected, free-POM separated at 1.85 g cm-3) of WSA, but free-POM and POM occluded in macro- and microaggregates were strongly decreased under Eucalyptus. Those data suggesting similar aggregation between Eucalyptus and native vegetation contrast with the data presented here and can be caused by a much lower intensity of soil preparation before planting in that experiment. In any case, the data suggest that strip tillage or less intensive soil cultivation for Eucalyptus plantations would better preserve soil structure and perhaps result in higher SOC levels.

66 In summary:

For bulk soil at all depths, Eucalyptus plantation did not change chemical and most physical properties, SOC and total N levels, but increased the C/N ratio for all soils at the 0-5 cm depth. The SOC data contrast with the previous decline in SOC concentration (0-5 cm depth) reported for the sandy and loamy soils (Zinn et al., 2002). Previous and new analysis of particle size fractions from 0-5 cm depth showed that, for the sandy soils only, Eucalyptus plantation initially reduced SOC levels in all size fractions, but by the end of the second cycle SOC was re-sequestered in the sand fraction only. Soil macroaggregation (WSA>2 mm) decreased in the surface 10 cm of all soils, probably because of the pre-planting heavy harrow tillage. In consequence, the percent occluded-POM C declined, probably because of intense decomposition of SOC in the soil fragments released from the macroaggregates. Widely variable C/N ratios in occluded- POM C suggested the presence of charcoal in the sandy and loamy soils. SOC stocks (0- 0.2, 0-0.40 and 0-1 m depth) depended strongly on soil texture, and were not affected by Eucalyptus plantation.

67 4.3 Textural control on SOC retention

Many authors have tried to model the effect of texture on SOC concentrations in a range of topsoils or within the profile of one or a few soils. Clay+silt, or separate clay and silt contents, were the best parameters describing soil texture in the models by Parton et al. (1987) and Burke et al. (1989) for prediction of SOC stocks (mass/area) over wide climatic gradients. Bosatta and Ägren (1996) included clay content in their depth modeling of SOC and N using a limited textural range (180-300 g clay kg-1). For temperate forest soils, Arrouays and Pélissier (1994) concluded that the decline of SOC concentration with depth is better explained by an exponential function, a conclusion supported by Bernoux et al. (1998) for a wide textural range in the Amazonian forest. Implicit limitations for predicting SOC come from the use of soil texture as classes (e.g., sandy, fine soils) rather than as a continuous variable. Simultaneously, studies of a textural effect on SOC are commonly restricted to surface layers, where strong spatial variability occurs due to management, bioturbation etc., and most SOC is in a POM form that is less likely to fully interact with mineral surfaces. The effect of depth on the complex and poorly understood relationship between soil texture and SOC concentration has not been described. As discussed in sub-chapter 4.1, the three soil types differed widely in texture, without marked clay illuviation, and within each soil type, no significant differences (P<0.05) were observed in bulk soil texture and SOC concentration between the two land uses. Therefore, these data were pooled for purposes of modeling in the first part of this sub-chapter (Figures 4.14 and 4.15). In other cases where significant effects of land use occur at the 0-5 cm depth, such as sand-size SOC pool in sandy soils, only the data from the Cerrado plots were used (reminder: for the 30-40 and 90-100 cm depths, samples from Cerrado and Eucalyptus plots were mixed 50:50). No linear relations between SOC and clay contents were observed. However, significant linear relations between SOC and combined clay+silt (0-20 μm) contents were observed for each depth (Figure 4.14), with relatively poorer fits for the three uppermost layers, especially for the 5-10 cm.

68 30

25 ) -1 20 2.5 cm, SOC=13.52+0.0178x, R2 = 0.77

7.5 cm, SOC=9.07+0.0126x, R2 = 0.53 15 15 cm, SOC=5.1+0.0117x, R2 = 0.77

25 cm, SOC=4.08+0.0109x, R2 = 0.84 10 35 cm, SOC=3.04+0.0103x, R2 = 0.91 Soil organic carbon (g kg 55 cm, SOC=2.68+0.0066x, R2 = 0.80 5 95 cm, SOC=1.75+0.0053x, R2 = 0.85

0 0 200 400 600 800 1000 Clay + silt content (g kg-1)

Figure 4.14. Linear relationship of SOC (g kg-1) vs. clay+silt (<20 μm, g kg-1) content for each depth (middle point of layer). All relations statistically significant for P<0.0001, except for 7.5 cm (P<0.001).

69 Similarly, Feller and Beare (1997) found SOC=0.037*(clay+silt) + 0.69 (n=65, R2 =0.76) in unrelated kaolinitic tropical soils (0-20 cm). For temperate, cultivated topsoils, Konen et al. (2003) estimated R2’s about 0.7 for SOC vs. clay and sand, whereas Carter et al (2003) found r=0.63 for SOC vs. clay+silt in Canada. Galantini et al. (2004) reported excellent correlations between clay+silt and bulk SOC, particulate and mineral-bound C, and humic-humin fractions in Argentina. Data in Figure 4.14 also show that both the intercept and slope of the SOC vs. (clay+silt) relations decrease with increasing soil depth (as middle point of the layer). Figure 4.15a shows the plot of the intercept and slope vs. depth, which follow respectively power and logarithmic functions. This result suggests that the intercept and slope parameters of the depth-specific relation SOC (g kg-1)= a + b*(clay+silt, g kg-1) can be estimated for use in a depth-weighted equation for the 0-1.0 m depth interval as follows:

SOC(d) = 24.847d-0.5715 + [(-0.0032*ln d + 0.0204)*(clay+silt)] (Eq. 1) where d is depth in cm.

Figure 4.15b shows the plot of measured SOC concentrations vs. values estimated with the above equation. The model explained the SOC variation very satisfactorily:

Estimated SOC = 0.447n.s. + 0.939(measured SOC), P<0.0001, R2=0.924, n= 126 (Eq. 2)

This model has a better fit for SOC values below 10 g kg-1, mostly associated with subsurface layers, where POM and spatial variability are lower than in topsoil. A revised version of this model, using a sum of exponentials function for the intercepts, has been recently published (Zinn et al., 2005b). In that article, Eq. 1 was applied to published data on other Cerrado soils and produced similarly high R2’s, however with intercepts and slopes for Eq. 2 significantly different from 0 and 1. This indicates that the textural control on SOC along profiles of Cerrado soils is mathematically the same, and simple calibration is required to predict SOC levels accurately for other localities in the biome.

70 20 0.02 Intercept

a 16 0.016 slope Slope b

Intercept 12 0.012 Intercept a= 24.85(d)-0.57 P<0.0001, R2 = 0.99 8 0.008 Slope b =0.0204 - 0.0032 ln(d) 2 4 0.004 P<0.0002, R = 0.95

0 0 0 20406080100Soil depth (cm)

a) Estimated vs. measured SOC (g/kg) 30

25

20

15

10 Estimated SOC (g/kg) 5

0 0 5 10 15 20 25 30 Measured SOC (g/kg)

Linear Fit

b)

Figure 4.15. a) Decrease of intercept a and slope b (from linear relations in Figure 4.14) with depth; and b) Graphical relationship between measured and estimated SOC (dotted line represents 95% confidence interval).

71 The strong relation between SOC vs. (clay + silt) concentrations reported here implies an equal but negative SOC vs. sand (20-2000 μm) relation, as reported by McDaniel and Munn (1985) for mesic Mollisols. Additionally, total N and SOC concentrations (g kg-1) were strongly correlated: N=0.143+0.063*SOC, (P<0.0001, R2=0.96, n=126). Thus, in unamended, highly weathered tropical soils where N is mostly associated with organic compounds, total N concentrations can also be predicted based on texture of subsoil layers. Naturally, this depth-weighted model must be calibrated for different soil-specific, regional and local conditions, or alternatively include climate, plant biomass and other environmental factors as model parameters. Finally, the relation SOC vs. clay+silt is not necessarily linear: although Carter et al. (2003) found significant linear functions in eastern Canada, their data seemed rather to indicate a maximum SOC for 700 g clay+silt kg-1 (for total N, at 600 g kg-1).

Texture, surface area and SOC

The ability of soils to retain SOC is most likely limited by the surface area available for sorption (Kaiser and Guggenberger, 2003). Therefore, part of the textural control on SOC retention demonstrated before must be due to higher surface areas with increasingly finer texture. Figure 4.16 shows that specific surface area (SSA, by the BET- N2 method) increased in soils of finer texture, and tended to increase with depth for the same soil (see discussion below). Little variability occurred among replicates of the same soil/depth combination, and higher variability in the clayey Haplustox is attributed to irregular distribution of nodules/concretions. Figure 4.17a shows SSA as a function of clay content. The same slopes and similar intercepts for the three depths indicate that SSA is mostly controlled by texture with little or no effect of depth, i.e. the increased SSA with depth results from higher clay contents (Table 4.2). On the other hand, SOC concentration was linearly correlated to SSA but with a pronounced effect of depth (Figure 4.17b).

72 0 10203040m2 g-1 soil

2.21 Specific surface area ) 5.65 0-5 (soil <2 mm) 13.95 28.58 Quartzipsamment (n=1)

Depth (cm 4.10 Sandy Haplustox (n=2) 8.02 30-40 Loamy Haplustox (n=3) 20.25 Clayey Haplustox (n=3) 34.54

5.69 10.74 90-100 20.30 37.36

2 -1 Figure 4.16. BET-N2 specific surface area (m g ) of fine earth fraction from Cerrado soils at different depths. Bars show standard error.

A multivariate model of SSA as a response variable to textural (clay, silt, fine- coarse sand), SOC and depth data was computed. The model obtained showed that SSA was strongly dependent on the contents of clay, silt and SOC, but not on depth, fine and coarse sand contents. Figure 4.18 shows the fit of the model

BET-SSA (m2 g-1) = 0.0581(clay****) + 0.0594(silt****) – 0.271 (SOC***) (Eq. 3) where clay, silt and SOC contents are expressed in g kg-1, (R2=0.97, n=27). Significance level for each parameter: ***= P<0.001, ****=P<0.0001.

73

45 40 0-5 cm: SSA = 0.074clay - 4.38 2 35 R = 0.89 ) -1 30 30-40 cm: SSA = 0.074clay - 3.97 g 2 2 0-5 cm 25 R = 0.95 20 30-40 cm

SSA (m 15 90-100 cm 2 10 90-100 cm: SSA = 0.073clay - 1.75

BET-N 5 2 R = 0.82 0 Clay -1 0 100 200 300 400 500 600 (g kg ) a)

30 SOC = 0.46SSA + 12.36 R2 = 0.85 )

-1 20 0-5 cm SOC = 0.19SSA + 3.45 2 30-40 cm R = 0.89 10 90-100 cm SOC (g kg

SOC = 0.07SSA + 2.12, R2 = 0.71 0 SSA 2 -1 0 10203040(m g ) b)

Figure 4.17. Linear relations between a) specific surface area (SSA, BET-N2) and clay content, and b) SOC concentrations and SSA for 3 depths (fine earth).

74 Estimated SSA(m2/g) By actual SSA(m2/g) 40 35

30

25

20

15

10 Pred Formula SSA(m2/g) 5

0 0 5 10 15 20 25 30 35 40 SSA(m2/g)

Linear Fit

Figure 4.18. Fit of the model describing SSA as a function of clay, silt and SOC contents. Dotted lines mark the 95% confidence interval.

In Eq. 3, it is notable that the SSA estimator for the silt fraction is similar to that for clay. When the SSA of selected silt samples was independently measured, it varied from 6.4 m2 g-1 (clayey Haplustox, 90-100 cm) to 11.5 m2 g-1 (loamy Haplustox, 0-5 cm), values that are much lower than SSA of the clay fractions (see 4.4). Additionally, silt SSA was directly related to its SOC content: this indicates that any aggregation of near- spherical silt grains promoted by SOC does not result in occlusion of mineral surfaces, as is the case for the clay fraction, mostly composed of platy particles (see 4.4). Consequently, silt SSA can be estimated simply as a function of its SOC content, which in its turn is determined by depth and the dilution effect (see next item), as follows:

75 Silt BET-SSA (m2 g-1) = 1.42Ln(SOC) + 5.03 (Eq. 4) where SOC is expressed as g SOC kg-1 silt (R2 = 0.99, n = 6).

Interpretation of clay and silt SSA data indicates that the estimators of clay and silt in Equation 3 do not reflect actual SSA values, but are simply the best mathematical descriptors for the multivariate regression for those soils. Thus, the SSA equations above can be applied only to soils within the restricted range of silt content in this study, which are however typical of highly weathered tropical soils. Although SSA tended to increase with depth for each soil, depth was not a significant component of the models. This indicates that depth effects on SSA of these soils are due to different clay and SOC contents and/or higher aggregation in the topsoil. Also, the model shows SSA negatively correlated to SOC contents (see discussion below). In consequence, Eq. 3 is valid for any point in the 0-100 cm interval, contrasting to the preceding SOC vs. (clay+silt) equation (Eq. 1). The apparently contrasting effect of depth in the textural control on bulk SOC and SSA can be interpreted in the following way. For a textural range of soils, SOC retention for a specific depth is proportional to the <20 μm fraction, since a finer texture necessarily provides a higher SSA, and promotes higher sorption of colloidal SOC and greater aggregate size/stability. The fact that SOC concentrations decrease with depth, while SSA increases slightly, indicates that the local input of organic matter (mostly from root remains and exudates) is the main determinant of SOC retention in a given layer, followed then by total (clay+silt) contents. However, in any soil profile and irrespective of its texture, the higher SOC levels near the soil surface simultaneously promote aggregation of clay, resulting in lower SSA compared to the subsurface, where SOC is lower (see also Christensen, 1992, p. 49-50). This effect is also apparent in the SSA of the SOC-containing clay fractions (see 4.4). It is almost a paradox that, for a range of soil textures, SOC concentration is directly related to and dependent on SSA, and at the same time, in a single soil profile, SSA is inversely related to and dependent on SOC concentration.

76 Particle-size fractions

The partition of SOC by particle sizes is in theory a indicator of its quality: SOC in coarse fractions is commonly considered a less decomposed material with high C/N ratios, whereas SOC sorbed to clay are mostly humic substances with low C/N (Christensen, 1992, p. 43-47). As expected, textural differences in the studied soils also affected the partition of total SOC through the sand, silt and clay particle-size pools (Table 4.10 and discussion below). The yields of each fraction were similar to those from the textural analysis (Table 4.1), and SOC contents also decreased significantly with depth. Mean recovery of SOC was 80% of total SOC for the two coarser soils and 75% for the clayey Haplustox, in all cases approaching 95% for the 90-100 cm layer. SOC contents for the same particle size fraction vary considerably with soil texture. More specifically, SOC contents for a particle size at the same depth tend to be inversely related to the yield of that fraction. Similar results have been reported earlier: Christensen (1992, p.41) and more recently Schulten and Leinweber (2000) compiled literature showing that the SOC enrichment factors for clay and silt (but not sand) fractions were exponentially but inversely related to each fraction’s yield for many different soils. For grassland topsoils (0-10 cm) in North America, Amelung et al. (1998) found similar tendencies for the sand and clay fractions (but not silt), ascribing it to a “simple dilution effect”, a phrase that will be used hereafter. This effect can be understood as follows: upon the particle size fractionation of any soil, the total amount of SOC (which occurs throughout a continuum of coarse to colloidal size) is distributed within the size fractions (clay, silt, sand) by sieving and sedimentation techniques. Assuming that a soil of any texture contains SOC in all size fractions and in relatively constant proportions (a valid assumption for the studied soils, as seem next), a certain particle size fraction is more or less SOC-enriched if its yield is less or more abundant, respectively (see also explanation by Christensen, 1992, p. 41). The SOC dilution effect is better visualized graphically, as in Figure 4.19. Moreover, it is satisfactorily described by simple exponential or logarithmical functions based on the yield of each fraction, most decidedly for the SOC-rich surface layer.

77

Soil / Sand Silt Clay Sand Silt Clay

Depth (cm) (20-2000μm) (2-20μm) (<2μm) SOC SOC SOC -1 Sandy Soils g kg 0-5 869.55 7.90 122.55 5.18 139.01 44.21 (n=6) (14.08) (0.55) (13.95) (1.56) (3.86) (3.56) 30-40 839.06 6.26 154.68 0.95 53.93 13.76 (n=3) (23.24) (1.03) (22.32) (0.11) (2.11) (0.70) 90-100 824.04 8.40 167.56 0.77 22.92 8.86 (n=3) (26.82) (0.34) (26.77) (0.03) (0.30) (0.57) Loamy Haplustox 0-5 633.57 17.51 348.92 6.30 86.59 28.55 (n=6) (8.31) (1.62) (9.01) (0.61) (4.02) (1.19) 30-40 596.94 15.21 387.85 1.35 28.90 10.63 (n=3) (14.02) (1.43) (13.17) (0.18) (3.32) (0.25) 90-100 576.70 14.96 408.34 0.75 18.75 6.17 (n=3) (9.43) (0.46) (9.24) (0.08) (3.34) (0.01) Clayey Haplustox 0-5 374.56 140.42 485.02 12.66 14.70 22.07 (n=6) (33.06) (14.96) (24.84) (1.58) (1.54) (1.21) 30-40 332.96 114.57 552.47 1.98 5.60 10.07 (n=3) (28.92) (2.01) (30.34) (0.10) (0.55) (0.25) 90-100 334.08 109.03 556.89 1.28 3.04 6.48 (n=3) (26.32) (3.89) (28.80) (0.30) (0.39) (0.37) Obs.: n=3 for SOC in sand fractions of Sandy Soils 0-5 cm.

Table 4.10. Yields and SOC contents for each particle size fraction for 3 depths of the soils studied. Numbers in parenthesis are standard error.

78 25 0-5cm: y = -10.3Ln(x) + 73.8, R2 = 0.80

20 30-40 cm: y = -1.1Ln(x) + 8.3, R2 = 0.83 0-5 cm 90-100cm: y = -0.5Ln(x) + 4.1, 30-40 cm 15 R2 = 0.31

sand 90-100 cm -1 10

5 g SOC kg

0 g sand -1 0 200 400 600 800 1000 kg soil

180 160 0-5cm: y = 715.2x-0.78, R2 = 0.93

. 140 30-40 cm: y = 214.7x-0.76, R2 = 0.95 120 0-5 cm silt 100 90-100 cm: y = 147.4x-0.82, R2 = 0.95

-1 30-40 cm 80 90-100 cm 60

g SOC kg 40 20 0 g silt -1 0 50 100 150 200 kg soil

60 0-5 cm: y = -15.3Ln(x) + 117.1, R2 = 0.84 50 0-5 cm 30-40 cm: y = 42.95x-0.23, R2 = 0.81 30-40 cm 40 0.0007x 90-100 cm: y = 4.8035e 90-100 cm

clay . 2 30 R = 0.6064 -1 20

g SOC kg 10

0 g clay -1 0 200 400 600 800 1000 kg soil

Note: in the regression equations, y and x correspond to variables in y and x axis.

Figure 4.19. Graphical view of the SOC dilution effect for the sand, silt and clay. 79 Since the SOC dilution effect was detected in the literature of temperate soils and in this study, it probably occurs in most textural ranges in the world. Also, the method for soil dispersion and fractionation has little importance: in this study, SOC concentrations in particle size fractions of the two coarser soils were remarkably similar to those obtained for the same soils sampled in 1997, but dispersed by sonication (Zinn et al., 2002). As a result of its ubiquity, the dilution effect has caused appreciable confusion in the literature. The most common misunderstanding resulting from the dilution effect is the silt fraction being SOC-richer than the clay, which is hard to explain since it supposedly has much lower SSA and surface charge than clay. Table 4.10 shows that the sandy and loamy soils had a very low silt yield (<2%), and this fraction is much richer in SOC than the clay. Conversely, for the clayey Haplustox SOC with a significant (>10%) silt yield, the clay fraction was higher in SOC. The high, relatively homogeneous silt content in soils studied by Amelung et al. (1998) is probably the reason why the dilution effect was not significant in that fraction. The dilution effect is of critical importance in modeling the control of soil texture on SOC size distribution. The functions in Figure 4.19 describe SOC concentrations in each fraction and can be adjusted to different depths, but a more practical model would predict the total SOC pool in each size fraction and depth. The SOC size pools shown in Figure 4.20 were calculated by multiplying the yield and SOC contents of each size fraction, and by dividing this product by the sum of fractions for each soil and depth combination. Considering all soil textures and depths, the overall trends are: 1) the clay fraction is the main SOC pool, even in sandy topsoils, in accord with the findings of Christensen (1992, p. 42); 2) sand-sized SOC pool is more important in coarser soils, in accord to Kay (1998) and Bird et al. (2003). Sand-sized SOC is also greater in the surface layers of all soils because of abundant fresh, undecomposed residues. Sand- sized SOC decreases with depth, where C inputs are lower and the relative importance of clay-SOC increases, in consonance with Bird et al. (2003); 3) silt-sized SOC pool is remarkably constant (7-12% of total SOC) throughout the profiles, with a tendency to decrease slightly with depth.

80 0% 20% 40% 60% 80% 100% Particle size 11.14 SOC pools: 40.30 48.56 0-5 sandy soils (n=3)

10.24

30-40 24.87 64.88 % SOC in sand

8.46 % SOC in silt 90-100 28.33 63.21 % SOC in clay Depth (cm)

0% 20% 40% 60% 80% 100% Particle size 9.84 SOC pools: 25.66 64.50 0-5 loamy Haplustox (n=3; n=6 at 0-5cm) 8.19

30-40 14.79 77.02 % SOC in sand 8.58 % SOC in silt 90-100 13.37 78.05 % SOC in clay Depth (cm)

0% 20% 40% 60% 80% 100% Particle size 11.81 SOC pools: 0-5 26.48 61.71 clayey Haplustox (n=3; n=6 at 0-5cm) 9.28

30-40 80.96 9.77 % SOC in sand 7.52 % SOC in silt 82.74 90-100 % SOC in clay Depth (cm) 9.73

Figure 4.20. Percent distribution of SOC size pools for three selected depths. Actual means are shown within the chart and bars represent standard error.

81 The successful modeling of SOC, textural and depth data for bulk soils shown earlier suggested the possibility of modeling the relative SOC partition through the different particle sizes (i.e., percent of total SOC contained in each size pool) of profiles of different texture. The modeling process was similar to the one developed for the bulk SOC vs. (clay+silt). Therefore, it was first attempted to plot the percent SOC size pool as a function of the respective contents of each fractions in the soils. No significant relations ocurred for the silt, whereas linear functions were the best descriptors for the sand and logarithmical relations explained better the behavior for the clay+silt and clay. In all cases, the functions show a better fit for subsurface layers, which again indicates higher variability in topsoil. The final model was significant for the clay and sand pools, but since the fit for the sand was poorer (R2=0.70, n=33) and for simplicity, only the model for clay-SOC will be discussed here. Figure 4.21a shows the plot of percent of total SOC in the clay fraction vs. clay content for the three depths. The intercepts and logarithm operators were plotted against depth, and the best descriptors for their change were linear and quadratic functions, respectively (Figure 4.21b). These functions were then used to parameterize a model of the type: clay-sized SOC pool (% of total SOC) = a + Ln(clay, g kg-1) as follows: y = (5.43 + 0.059d - 0.0035d2) + (0.074d + 9.47)*Ln(clay), (Eq. 5) where d is depth in cm. Figure 4.21c shows the plot and the 95% confidence intervals (dotted line) of values measured vs. estimated by that equation, resulting in the following fit:

Estimated = 10.90 + 0.841*Measured (R2=0.87, n=33). (Eq. 6) Although the determination coefficient is reasonable, Eq. 5 is not accurate since the intercept and slopes are respectively different from 0 and 1 in Eq. 6. Even though Eq. 5 did not predict adequately the clay-sized SOC pool, it may still be useful since the estimated values are linearly correlated to the observed ones. Considering that the silt- sized SOC pool is constant through the range of depth and texture, sand-sized SOC pool can be estimated by difference from the values modeled for the clay-sized SOC pool.

82 100 90-100 cm: y = 16.4Ln(x) - 20.6, R2 = 0.95 30-40 cm: y=12.3Ln(x) 0-5 cm 80 2 +3.2, R = 0.90 30-40 cm 60 90-100 cm

2

(% of total) 0-5 cm: y = 9.5Ln(x) + 5.56, R = 0.57 40 g clay -1 clay-sized SOC pool 0 200 400 600 800 kg soil a) 20

10 Ln op. = 0.074d + 9.47 R2 = 0.996 0 ln operator Intercept -10

-20 interc. =5.43+0.059d - 0.0035d2 R2 = 0.999 -30 0 20406080100Depth (cm) b) Estimated vs. Measured clay-sized SOC pool (% of total SOC) 100 90 80 70 60 50

Estimated 40 30 20 10 0 0 10 20 30 40 50 60 70 80 90 100 Measured

Linear Fit

c)

Figure 4.21. a) Clay-sized SOC pool as a function of clay content throughout the profiles; b) Plot of intercepts and logarithm operators (from a) vs. depth; c) Plot of estimated vs. measured percent of clay-sized SOC pool.

83 Gill and Burke (2002) reviewed that SOC contents and decomposition rates decrease consistently but not equally with depth, and concluded that the mechanisms involved in root litter dynamics and SOC storage in deeper layers are not well understood. The C/N ratios of size separates of deep layers can give an idea of the decomposition status of organic matter stored there, assumed to be mostly derived from roots. Figure 4.22 shows the C/N ratios of different particle sizes for selected depths. The overall values decline with decreasing particle size, as reviewed previously by Christensen (1992). More specific tendencies are: 1) the C/N ratio is about 10 for all clay fractions, regardless of soil texture and depth, indicating a high and homogeneous degree of humification; 2) the sand fractions had a minimal C/N ratio of 20, indicating as expected an intermediate degree of alteration. However, there is a strong increase in C/N with depth in the two coarser soils, reaching 75 for the loamy Haplustox at 90-100 cm. The most plausible cause for this is the dominant presence of sand-sized charcoal fragments, macroscopically visible in the sand separates and also in the water- stable aggregates (Appendix B, Fig. 21) as reported by Schaeffer et al. (2004). Teixeira et al. (2002) ascribed the presence of charcoal fragments to past fire events, obviously more ancient with increasing soil depth. 3) for the surface layers, C/N ratios for the silt fraction are intermediate between the clay and sand; for deeper layers, it is similar to that of clay.

The C/N ratios indicate that, at least for the surface layers, SOC in silt fractions differs from that in clay. While the latter is most likely composed by sorbed compounds of colloidal or smaller size, silt-sized SOC is less decomposed and probably not retained by sorption, as also indicated by the SSA function (Eq. 6). Based on bulk SOC vs. clay+silt relations similar to those described here, Hassink (1997) and Six et al. (2002) suggested a “silt+clay associated or protected SOC” pool that, according to data presented here, is in fact a heterogeneous mixture. Thus, any SOC protective effect is likely caused solely by sorption to clay, whereas silt-associated SOC is rather a mass of organic micro-debris or colloidal clots loosely bound to the mineral component.

84 C / N ratios of particle size separates Sandy soils

80

70 Y

60 Sand

50 Silt

40 Clay

C / N ratio 30 Mean(Sand)

20 Mean(Silt)

10 Mean(Clay)

0

2.5 35 95

Depth (cm)

Loamy Haplustox Clayey Haplustox

80 80

70 70

60 60

50 50

Y 40 Y 40

30 30

20 20

10 10

0 0

2.5 35 95 2.5 35 95

Depth (cm) Depth (cm)

Figure 4.22. Patterns of C/N ratios in particle-size separates throughout the profiles. Dots represent actual values and lines connect the mean values.

85 4.4 Mineralogical control on SOC retention

The sand and silt particle size fractions of any soil are characterized by much lower SSA and surface charges than the clay, which means that SOC occurring in those fractions must either be POM or SOC sorbed to non-dispersed aggregates of clay or secondary minerals. Therefore, a basic assumption to study the mineralogical control on SOC retention is that it is totally or mostly due to the clay fraction, to which the results and correlations on this sub-chapter refer. Table 4.11 shows the quantitative mineralogical analyses of the studied soils. The sum of mineral fractions does not attain the total of 1,000 g kg-1 due to the impossibility of quantifying aluminum-interlayer vermiculite (AIV), residual water (retained at energies higher than removable at 1050C) and other amorphous compounds not measured. Additionally, quantification of goethite by thermogravimetry was not feasible due to interference of AIV in the same dehydration range (300-400 0C); thence, the crystalline Fe oxides represent goethite+hematite. However, in all cases the mineral composition was determined for about 80 % of the clay fraction, and the results are consistent with the qualitative data from the X-ray diffractograms presented in section 4.1. The clay mineralogy is almost totally kaolinitic in the Quartzipsamment, with small amounts of accessory minerals including AIV, gibbsite and goethite. The importance of the accessory minerals, including hematite and illite (clay mica) increases continually with soil clay content until reaching a kaolinitic-illitic character, with significant amounts of Fe/Al oxides, in the clayey Haplustox. Amorphous Fe-oxides, as determined in the acid oxalate extracts, are very low (<8 %) in relation to total Fe-oxides determined by Na citrate-bicarbonate-dithionite (CBD) extraction. This is not the case for amorphous Al- oxides, which occur as ca. 20% of the gibbsite content. There is little deviation in concentrations of minerals within each soil type as indicated by the standard deviations, except for amorphous Fe and Al oxides. Differences in the relative amounts of minerals within the 1 m depth of each soil are also low.

86

Soil / Fe O Al O Cryst. 2 3 2 3 Depth (cm) Kaolinite Fe2O3 Ox. Gibbsite Ox. Illite Total Quartzipsamment (n=1) g kg-1 0-5 809.9 13.7 1.1 17.6 2.3 n.d. 844.6 30-40 799.5 16.5 1.2 12.3 4.8 n.d. 834.4 90-100 802.9 13.9 0.8 15.2 3.2 n.d. 835.9 Sandy Haplustox (n=2) 0-5 742.4 48.4 1.7 20.5 2.8 16.0 828.6 (0.51) (2.19) (0.50) (5.63) (0.57) (1.1) 30-40 727.3 38.7 1.3 16.1 5.0 16.6 801.5 (13.62) (4.84) (0.31) (1.52) (0.00) (1.2) 90-100 742.2 39.0 0.6 13.6 4.6 20.8 816.5 (23.00) (0.81) (0.14) (0.73) (0.75) (1.7) Loamy Haplustox (n=3) 0-5 609.0 68.3 2.8 65.5 5.8 40.2 784.9 (7.42) (1.57) (0.09) (9.98) (1.21) (2.9) 30-40 618.6 75.9 1.7 52.2 6.7 39.2 787.8 (15.50) (2.05) (0.13) (7.27) (0.58) (4.8) 90-100 635.6 70.0 1.7 52.8 6.2 36.6 796.9 (4.85) (7.48) (0.60) (4.72) (0.19) (4.5) Clayey Haplustox (n=3) 0-5 464.9 68.1 2.6 53.7 7.3 204.5 766.9 (11.96) (7.90) (0.45) (13.67) (0.15) (16.1) 30-40 497.3 75.2 1.6 45.2 8.3 198.6 793.0 (43.87) (9.48) (0.25) (10.20) (0.55) (18.6) 90-100 506.8 63.9 1.1 48.9 9.2 203.6 799.6 (13.14) (5.47) (0.25) (12.36) (0.61) (17.90)

Table 4.11. Quantitative mineralogy of clay fractions from different depths of the studied

soils. Numbers in parenthesis are standard errors. Crystalline Fe2O3 (goethite + hematite) is the difference between CBD and oxalate extracts.

87 0 102030405060m2 g-1 clay

36.99

) Specific surface area 0-5 39.14 (clay with SOC) 41.96 46.02 Depth (cm 40.77 Quartzipsamment (n=1) 44.42 Sandy Haplustox (n=2) 30-40 44.11 Loamy Haplustox (n=3) 49.45 Clayey Haplustox (n=3) 41.55 44.86 90-100 42.63 50.92

2 -1 Figure 4.23. BET-N2 specific surface area (m g ) of clay fractions of different depths of the studied soils. Bars show standard error.

The BET-N2 SSA of the non-peroxide treated, Mg-clay of the studied soils is shown in Figure 4.23. The values for the clayey Haplustox are similar to those determined by Feller et al. (1992) in another area of the Cerrado. There is an increase in SSA with soil depth (except for the loamy Haplustox), which may be explained by the concurrent decrease in SOC concentration (see also Figure 4.16) and/or preferential illuviation of fine clay. Sorption of C to different artificial and natural clay minerals was shown to decrease SSA heterogeneously in sorption studies, a phenomenon attributed to organic colloids coating mineral micropores and surface (Kaiser and Guggenberger, 2003). Dispersion of non-peroxide treated soil to obtain clay fractions, as done in this study, would have a similar effect. Table 4.12 shows the correlation between contents of different minerals and SSA, SOC, total N and C/N ratios of the clay fractions. The 88 negative correlation between SOC and SSA reinforces this conclusion, especially for the SOC-rich 0-5 cm depth. Additionally, the relative increase of Fe/Al-oxides affected positively the SSA of the clay fractions, as reported earlier by Feller et al. (1992) for different Cerrado soils. SSA is best correlated (negatively) with kaolinite contents, probably because of the relatively larger size of kaolinite crystallites in comparison to the accessory minerals. Table 4.12 also shows, however, that contents of SOC, and consequently total N, are negatively correlated with Fe/Al oxides in the clay fractions. This result was unexpected since there is literature demonstrating that these oxides adsorb more SOC than phyllosilicates (see item 2.1.3 and following discussion).

0-5 cm Fecbd Feox Alox Fecbd - ox Gibbsite Kaolinite SSA N SSA 0.66 0.56 0.87 0.66 0.51 -0.93 N -0.86 -0.77 -0.91 -0.86 -0.71 0.88 -0.89 C -0.81 -0.7 -0.9 -0.81 -0.66 0.88 -0.91 0.99 CN 0.81 0.84 0.64 0.8 0.74 -0.56 0.46 -0.65 30-40 cm SSA 0.51 0.13 0.63 0.51 0.37 -0.83 N -0.83 -0.28 -0.72 -0.83 -0.78 0.66 -0.5 C -0.8 -0.25 -0.75 -0.81 -0.75 0.73 -0.56 0.99 CN 0.63 0.37 0.22 0.63 0.69 0 -0.05 -0.64 90-100 cm SSA 0.34 -0.15 0.75 0.36 0.15 -0.81 N -0.86 -0.55 -0.46 -0.86 -0.76 0.46 -0.09 C -0.93 -0.41 -0.68 -0.94 -0.79 0.73 -0.35 0.89 CN -0.5 0.1 -0.68 -0.51 -0.4 0.8 -0.62 0.19

Table 4.12. Correlations between selected mineral contents and SSA, SOC, total N and C/N ratios in clay fractions at different depths.

89 Clays with higher SSA supposedly adsorb more soluble C compounds (Kahle et al., 2004), in contrast to the negative SSA vs. SOC correlation shown above. To understand these correlations, one must consider the SOC dilution effect shown in Figure 4.19 and discussed in sub-chapter 4.3. In the 0-5 and 30-40 cm depths, the SOC dilution effect for the clay fraction is clear. Therefore, correlations between SOC and mineralogy can be flawed if calculated for SOC concentrations in the clay fraction. Schulten and Leinweber (2000), although aware of the dilution effect, compared SOC concentrations in clay fractions from independent works in the literature (i.e., different soils, climate, land use and depths) and concluded without much basis that kaolinitic clays were “as expected” poorer in SOC than those dominated by 2:1 minerals. To avoid the confusion caused by the SOC dilution effect, an alternative is to correlate contents in bulk soil each mineral (calculated from results on the clay fractions) with bulk SOC and clay-sized SOC pool. Figures 4.24 and 4.25 show that bulk SOC concentration correlates relatively well with contents of individual minerals in the bulk soil. However, this is partially a consequence of the correlation between SOC and clay+silt (Figure 4.14), thus only correlations better than those will be considered. There is a clear effect of depth on the determination coefficients: crystalline Fe-oxides were closely related to SOC (R2=0.91) in the 0-5 cm, but not in the 90-100 cm depth 2 (R =0.66). Conversely, contents of Alox (amorphous Al-oxides) were well correlated to SOC in the 90-100 cm (R2=0.90), but more weakly at 0-5 cm depth (R2=0.75). Correlations for the 30-40 cm depth were intermediate, with SOC correlating equally well with crystalline Fe and amorphous Al oxides. Amorphous Fe-oxides showed a poorer correlation than the crystalline, which can be in part caused by their relatively low concentration in the soils studied. When the sand- and silt-sized SOC are removed from the calculations, the correlations with SOC improved. Figures 4.26 and 4.27 show the correlations of mineral contents with the clay-sized SOC pool in the bulk soil. The trends are exactly the same as discussed above, but with even better coefficients of determination, especially for crystalline Fe-oxides and Alox below the 0-5 cm depth.

90 30 y = 0.31x + 13.5

) 25 2 -1 R = 0.91 0-5 cm 20 30-40 cm y = 0.14x + 3.6 15 R2 = 0.90 90-100 cm 10

Bulk SOC (g kg y = 0.06x + 2.4 5 R2 = 0.66 0 g cryst. Fe2O3 -1 0 204060 kg soil

30 y = 7.69x + 13.73 2 ) 25 R = 0.87 -1 20 0-5 cm 15 y = 6.64x + 3.24 2 30-40 cm 10 R = 0.84 90-100 cm Bulk SOC (g kg 5 y = 1.37x + 3.2 R2 = 0.20 0 g Fe2O3-oxal. 0.0 0.5 1.0 1.5 2.0 kg-1 soil

30 y = 2.59x + 14.8 2 ) 25 R = 0.75 -1 0-5 cm 20 y = 1.33x + 3.6 30-40 cm 2 15 R = 0.90 90-100 cm 10

Bulk SOC (g kg y = 0.54x + 2.4 5 2 R = 0.90

0 g Al2O3-Oxal. 0.0 2.0 4.0 6.0 8.0 kg-1 soil

Note: in the regression equations, y and x correspond to variables in y and x axis.

Figure 4.24. Relation between bulk SOC with crystalline Fe- and amorphous Fe/Al oxides.

91 30 y = 0.26x + 15.4

) 25 2

-1 R = 0.71 0-5 cm 20 30-40 cm 15 y = 0.17x + 4.4 90-100 cm R2 = 0.71 10

Bulk SOC (g kg y = 0.059x + 2.9 5 R2 = 0.51 0 g gibbsite 0 10203040 kg-1 soil

30 y = 0.054x + 10.5

) 25 2 -1 R = 0.70 0-5 cm 20 30-40 cm 15 y = 0.027x + 1.64 90-100 cm 2 10 R = 0.69

Bulk SOC (g kg y = 0.012x + 1.22 5 R2 = 0.70 0 g kaolinite 50 150 250 350 kg-1 soil

30 y = 0.08x + 16.9

) 25 2 -1 R = 0.69 0-5 cm 20 30-40 cm 15 y = 0.043x + 5.34 90-100 cm 2 10 R = 0.76 Bulk SOC (g kg 5 y = 0.02x + 3.03, R2 = 0.88 0 g illite 0 50 100 150 kg-1 soil

Note: in the regression equations, y and x correspond to variables in y and x axis.

Figure 4.25. Relation between bulk SOC with crystalline minerals.

92 14

l 12 y = 0.20x + 4.38 2 soi R = 0.91 -1 10 0-5 cm 8 y = 0.086x + 1.67 30-40 cm R2 = 0.95 6 90-100 cm 4 g clay-SOC kg 2 y = 0.057x + 1.16 R2 = 0.80 0 g cryst. Fe2O3 0204060 kg-1 soil

16

l 14 y = 4.98x + 4.51 2 soi 12 R = 0.88 -1 0-5 cm 10 30-40 cm 8 y = 4.25x + 1.39 2 90-100 cm 6 R = 0.92 4

g clay-SOC kg g clay-SOC y = 1.56x + 1.75 2 R2 = 0.38 0 g Fe2O3-Oxal. 0.0 0.5 1.0 1.5 2.0 kg-1 soil

14

l 12 y = 1.52x + 5.51

soi 2

-1 10 R = 0.62 0-5 cm 8 30-40 cm y = 0.84x + 1.66 6 90-100 cm R2 = 0.95 4 g clay-SOC kg 2 y = 0.46x + 1.19 R2 = 0.97 0 g Al2O3-Oxal. 0.0 2.0 4.0 6.0 8.0 kg-1 soil

Note: in the regression equations, y and x correspond to variables in y and x axis.

Figure 4.26. Relation between clay-SOC with crystalline Fe- and amorphous Fe/Al oxides.

93 14 y = 0.18x + 5.4

l 12 R2 = 0.83 soi

-1 10 0-5 cm 8 y = 0.074x + 3.1 30-40 cm 2 6 R = 0.45 90-100 cm 4 g clay-SOC kg 2 y = 0.06x + 1.45 2 0 R = 0.75 g gibbsite 010203040 kg-1 soil

14

l 12 y = 0.038x + 1.82 2 soi R = 0.85 0-5 cm -1 10 8 y = 0.018x + 0.22 30-40 cm R2 = 0.82 6 90-100 cm 4 g clay-SOC kg 2 y = 0.011x + 0.067 R2 = 0.83 0 g kaolinite 0 100 200 300 400 kg-1 soil

14

l y = 0.039x + 7.05 12 2

soi R = 0.39

-1 10 0-5 cm 8 30-40 cm y = 0.024x + 2.86, R2 = 0.67 6 90-100 cm 4

g clay-SOC kg 2 y = 0.015x + 1.81, R2 = 0.7723 0 g illite 0 50 100 150 kg-1 soil

Note: in the regression equations, y and x correspond to variables in y and x axis.

Figure 4.27. Relation between clay-SOC with crystalline minerals.

94 The dilution effect has also interfered in other comparative studies. Wattel- Koekkoek et al. (2001) found that mean SOC concentrations in the clay fraction were the same for 6 kaolinitic and 6 smectitic tropical soils, and concluded that clay-sized SOC in soils is independent of mineralogy. However, since all their smectitic soils were very clayey and all but one of the kaolinitic soils had <300 g clay kg-1 soil, the dilution effect in their data is clear: g SOC kg-1 clay = 40.97e-0.0013x (R2 = 0.62), where x is g clay kg-1. soil. Therefore, the role of clay mineralogy on SOC retention in that study could not be properly assessed. The correlation of SOC with mineral contents for whole soils instead of clay fractions can help explain the role of Fe/Al oxides in other studies where the dilution effect occurs. As observed in this study, a negative correlation between SOC and Fecbd concentrations in clay fractions was found by Kahle et al. (2002). These authors studied long-cultivated, thick Ap horizons of seven illitic soils within a narrow range of soil -1 texture (all >70% silt) and Fe-oxide content (22.5-26.3 g Fecbd kg of clay), with 30-55 g SOC kg-1 of clay. For these relatively narrow experimental conditions, they concluded that “accumulation and decline of organic matter in soils is not regulated by clay mineralogy”, and the negative correlation of Fecbd with SOC was unexplained. However, when their data for whole soils are correlated, SOC varies directly although weakly with 2 2 clay (R =0.50) and Fecbd (R =0.28) contents. Better correlations could probably be obtained if data were available below the Ap horizon, since SOC concentrations in topsoil are more likely to be determined by management than in subsoil layers. Wisemann and Puttmann (2004) studied bulk soil samples of five German soils within broad textural and mineralogical ranges and concluded that SOC was best correlated with oxalate-Fe and –Al contents. However, their interpretation of the data was flawed because: a) they computed correlations for individual 1-m soil profiles, disregarding SOC decline with depth; b) Feox and Alox decreased concurrently with depth in each soil. Although the simultaneous decrease in SOC and amorphous oxide resulted in significant although weak correlations, it cannot be assumed that the depth-wise decrease of amorphous Fe and Al is due to binding to SOC and not to other pedogenic factors. Additionally, they inserted undesirable variability by comparing an arable soil, a forested

95 Gley and 3 well-drained forest soils. If correlations are recalculated for 40-110 cm depth 2 excluding the Gley and arable soils, the correlations between SOC and Feox (R =0.65, 2 n=9) and Alox (R =0.88, n=9) become evident and well based. For the 10-40 cm depth, 2 2 SOC is better correlated with Feox (R =0.85, n=6) than with Alox (R =0.65, n=6). Therefore, the reinterpretation of these two works produced correlations which support the conclusions drawn here, including the idea that the SOC-stabilizing role of Fe and Al oxides is respectively more important in surface and subsoil layers. The literature provides enough evidence indicating that sorption of SOC to soil mineral colloids differs considerably from that of exchangeable cations. Approximately 85% of the total humic acids sorbed by kaolinite (but not illite and smectite) are located on its variable-charge edges via bivalent cations (Varadachari et al., 1995). Moreover, the preferential sorption of organic C to Fe/Al oxide surfaces rather than phyllosilicates has been demonstrated and explained independently by many workers. Jardine et al. (1989) reported that after dithionite extraction an Ultisol from Tennessee adsorbed considerably less dissolved organic carbon (DOC). In vitro assays of natural DOC sorption to clays (monomineralic or from 5 German soils, at neutral pH) showed that DOC sorption was directly related to Fecbd and SSA, and strongly reduced after pre-sorption with phosphate (Kahle et al., 2004). These authors concluded then that DOC sorption is more common and strong on reactive, positively-charged hydroxyl groups (edges of phyllosilicates and Fe/Al oxide surfaces) than on negatively-charged siloxane surfaces (responsible for permanent CEC on phyllosilicates). This mechanism is also responsible for organic matter sorption temporarily inhibiting phosphate sorption in Cerrado soils (Afif et al., 1995). Hydrophobic rather than hydrophilic DOC is adsorbed to artificial goethite and amorphous Al oxides (Kaiser and Zech, 1997). These two oxides adsorb similar quantities of whole DOC, and the strength of sorption is such that desorption of hydrophobic compounds is only significant with relatively concentrated phosphate or NaOH solutions, whereas hydrophilic DOC is easily removed (Kaiser and Zech, 1999). Kalbitz et al. (2000) reviewed DOC dynamics and concurred with the idea of dominant sorption of organic compounds (esp. hydrophobic) on Fe/Al oxides, with limited sorption sites and an apparently higher sorption at low pH. Kaiser and Guggenberger (2003)

96 reviewed many works that suggest a rather fast and stable sorption of DOC to Fe/Al oxides. Kaiser and Guggenberger (2003) found that DOC sorption is directly related to Fe-oxide contents in soil clay fractions (47 soil horizons from 4 continents). For monomineral clays, DOC sorption was an order of magnitude higher for amorphous Al and goethite than for illite, kaolinite and gibbsite, with hematite in an intermediary situation. However, more DOC was found to be sorbed by hematite than to other oxides (Varadachari et al, 1997), although (Tombácz et al., 2004) reported a higher sorption on magnetite than on hematite. It is likely that there is a limit to Fe-oxide mediated SOC retention. For a range of basalt-derived, tropical forest soils in Australia, characterized by very high contents of clay (>50%), SOC (4-9%) and Fecbd (8-18%), Spain (1990) found negative correlations between SOC and clay contents. This suggests a saturation of the SOC-binding capacity of clay and Fe-oxides to a point where other factors better explain SOC contents, because correlations were positive for soils derived from other parent materials and with lower contents of clay, SOC and Fecbd. Schulten and Leinweber (2000) used computer models to visually describe the stable interaction of SOC to Fe oxides as an organo-mineral macromolecule, which only secondarily relates to phyllosilicates in soils. Crystalline Fe and Al oxides adsorb SOC via cation bridges and coordination mechanisms, that differ in intensity for each mineral (Varadachari et al., 2000). According to these authors, hematite and goethite sorb SOC by both mechanisms, whereas gibbsite does it only by coordination. Considering that the soils studied here contain predominantly well-crystallized oxides and very few polyvalent, free cations other than Al3+ and Fe3+, the aforementioned sorption mechanisms help explain the good correlations of SOC with crystalline Fe-oxides and the weak correlation with gibbsite. On the other hand, SOC was well correlated in subsurface horizons with amorphous Al oxides, that supposedly occur in equilibrium with the concentration of free (“exchangeable”) Al3+. This polycation promotes SOC sorption to 2:1 and 1:1 phyllosilicates, and synthetic crystalline Fe/Al oxides (Varadachari et al., 1991, 1997, respectively). Also, Al complexation by SOC is known to reduce its mineralization rate (Sollins et al. 1996; Schwesig et al., 2003). Thus, at subsoil depths of the soils studied

97 here, most SOC is probably stabilized by amorphous Al oxides, which can result in a high level of protection. This effect would still occur in surface layers, but is obscured by sorption of much higher amounts of clay-sized SOC to hematite/goethite, especially by cation-bridging (cations are more abundant at the surface). More research is needed to address this depth effect on SOC sorption, including sorption and C characterization studies. Guggenberger and Kaiser (2003) considered that, although SOC sorption by Fe/Al oxides and other mineral surfaces is important, it does not imply a permanent stabilization, especially for surface horizons characterized by weathered mineral grains that are nearly saturated with SOC. Since microbial populations are much higher on organo-mineral surfaces than in soil solutions, sorption of fresh DOC actually increases its decomposition, although the attendant generation of organic metabolites would also result in additional, more resistant coatings of those surfaces. The data and correlations presented here and interpreted in view of the cited literature suggest that total SOC retention occurs significantly in the form of organic coatings of mineral surfaces with pH-dependent or positive charges. This also provides an explanation of why soils with high activity clays tend not to differ from those with low-activity clay in regard to SOC retention, especially under acidic soil conditions when DOC sorption to Fe/Al oxides is favored (Varadachari et al., 1997).

98 4.5 SOC retention as affected by aggregation

Before discussing the aggregation control of SOC, it is necessary to note that soil texture at all depths is strongly related to aggregation as indicated by the mean weight diameter (MWD, in mm) and percent of water-stable aggregates (WSA)> 2mm. Table 4.13 shows the parameters of linear fits of MWD and WSA>2 mm = a + b*(clay+silt) in bulk soil, and Figure 4.28 shows the respective plots. Because there was a significant effect of Eucalyptus afforestation on soil aggregation at the 0-10 cm depth (sub-chapter 4.2), the results in this section are discussed for soils under native Cerrado only. Although correlations with clay contents were also found, they were weaker than with clay+silt and thus were not shown. Contrasting to the case of texture-SOC, the texture- aggregation relations were better in the top 20 cm layers, where most aggregation factors (e.g. SOC, biota, wetting/drying cycles etc) are more active.

Depth MWD (mm) as function of %WSA>2mm as function of -1 -1 clay+silt (g kg clay+silt (g kg ) ) (cm) a b R2 a b R2 0-5 1.09 0.00328 0.84*** 11.77 0.057 0.72** 5-10 0.89 0.00383 0.90*** 6.8 0.071 0.84*** 10-20 0.56 0.00127 0.90*** 1.24 0.014 0.66** 20-30 0.52 0.00110 0.84*** 0.75 0.010 0.74** 30-40 0.47 0.00130 0.73** -0.6 0.014 0.86*** 50-60 0.48 0.0010 0.53* -0.55 0.011 0.78** 90-100 0.36 0.0011 0.60* -1.1 0.0098 0.94***

Table 4.13. Parameters for the best linear relations found between aggregation and texture. * Significant at P<0.05, ** at P<0.01, *** at P<0.001 (n=9). 99 4

0-5cm 3 5-10cm 10-20cm

2 20-30cm 30-40cm

MWD (mm) 50-60cm 1 90-100cm

0 clay+silt -1 0 200 400 600 800 (g kg ) a)

60 0-5cm 50 5-10cm 40 10-20cm 20-30cm 30 30-40cm 20 50-60cm

WSA>2mm (%) . 90-100cm 10

0 clay+silt -1 0 200 400 600 800 (g kg ) b)

Figure 4.28. Linear relations between clay+silt (g kg-1) and a) MWD (mm) and b) percent of WSA>2 mm throughout the soil profiles.

100 As a consequence of the texture-aggregation relationship, MWD and % WSA>2 mm were also related to SOC concentrations, since the latter were well correlated with clay+silt contents (sub-chapter 4.3). Table 4.14 summarizes the parameters of linear fits of SOC = a + b*(MWD or WSA>2 mm), and Figure 4.29 shows the respective plots. The relations were excellent for the 0-5 cm depth only; with depth they were not always valid and the R2’s declined considerably. Also, their intercept and slopes vary irregularly, contrasting with the constant declines shown in Figure 4.15a. Aside from overall lower correlations than those in Table 4.13, this is an initial but strong indication that the structural controls on SOC levels are weaker than for soil texture and clay mineralogy for the soils studied. Consequently, a mathematical modeling of SOC based on aggregation such as shown in section 4.3 would be neither useful nor accurate.

Depth SOC (g kg-1) as function of SOC (g kg-1) as function of

MWD (mm) %WSA>2mm (cm) a b R2 a b R2 0-5 5.60 6.26 0.91*** 9.04 0.333 0.90*** 5-10 6.60 3.41 0.52* 8.28 0.188 0.59* 10-20 4.53 4.88 0.34n.s. 6.61 0.46 0.49* 20-30 1.83 7.12 0.50* 4.84 0.79 0.59* 30-40 2.30 5.2 0.54* 4.39 0.59 0.73** 50-60 3.26 2.45 0.18 n.s. 3.87 0.41 0.43 n.s. 90-100 1.81 2.5 0.46* 2.53 0.45 0.82***

Table 4.14. Linear relations between SOC levels and aggregation. * Significant at P<0.05, ** at P<0.01, *** at P<0.001, n.s. non-significant (n=9).

101 30

25 0-5cm 20 5-10cm 10-20cm 15 20-30cm ) -1 10 30-40cm 50-60cm 5 90-100cm

SOC (g kg 0 MWD 01234(mm) a)

30 0-5cm 25 5-10cm 20 10-20cm 20-30cm 15 30-40cm )

-1 10 50-60cm 90-100cm 5

SOC (g kg 0 WSA>2 mm 0 10203040506070 (%) b)

Figure 4.29. Linear relations between SOC (g kg-1) and a) MWD (mm) and b) percent of WSA>2 mm throughout the soil profiles.

102 For a better appreciation of the structural control on SOC, the WSA separates must be evaluated separately. Figure 4.30 shows the percent of each WSA size class through the whole profile of each soil type under native Cerrado. Increasingly finer texture resulted in larger proportions of WSA>2 mm for all depths, although overall WSA>2 mm declined markedly with depth. For sandy soils, the percent of WSA>2 mm decreased from >10% for the 0-10 cm depth to approximately 1% at 1 m depth; for the loamy Haplustox this decrease was from >20% in the topsoil to ca. 3% at 1 m. For the clayey Haplustox, WSA>2 mm comprised about half of the total soil sample at 0-10 cm, declining to <10% below that depth. The dominant size classes were: a) for the sandy soils, WSA 0.25-0.5 mm for all depths; b) for the Loamy Haplustox, WSA>2 mm in the top 10 cm and WSA<0.25 mm below; c) for the Clayey Haplustox, WSA>2 mm in the top 10 cm, and WSA<1 mm below, with a nearly homogeneous distribution. Figure 4.31 shows how soil texture affected the SOC contents of intact WSA size fractions for selected depths. As expected, SOC levels in WSA fractions decline with depth. In the sandy soils, WSA>1 mm had SOC concentrations about twice as high as the smaller size fractions. The same occurred in the loamy Haplustox, but with a smaller relative difference between WSA>1 mm and the smaller WSA classes. In the clayey Haplustox, SOC levels are more homogeneous throughout the different size classes. It is notable that only WSA>1 mm of both the sandy and loamy soils reached SOC levels comparable to smaller WSA of the clayey Haplustox. These results suggest that SOC plays a more important role in stabilizing large aggregates in surface loamy and coarser Oxisols, but not in clayey Oxisols where abundant clay promotes optimal cementation. The higher SOC levels in WSA<1 mm in the sandy and loamy soils are partially explained by their lower sand content (Figure 4.32), as noted also by Lima and Anderson (1997) in two clayey Cerrado Oxisols. Single and thin-coated grains comprise a significant part of WSA<1 mm in sandy and loamy soils. For WSA> 0.25 mm in all soils, sand contents do not vary substantially with depth, whereas microaggregates (<0.25 mm) of loamy and clayey Haplustox tend to comprise more single grains in the 0-5 cm depth (Appendix B, Figures 3 and 4) and more clay clusters in the lower depths. This pattern suggests that the role of clay as a cementing agent in each soil does not vary with depth.

103 % 0 102030405060

Sandy soils 0-5cm (n=3) 5-10cm 2-8mm 10-20cm 1-2mm 20-30cm 0.5-1.0mm 0.25-0.5mm 30-40cm 0.11-0.25mm 50-60cm

90-100cm

% 0 102030405060 % 0 102030405060

0-5cm 0-5cm

5-10cm 5-10cm

10-20cm 10-20cm

20-30cm 20-30cm

30-40cm 30-40cm

50-60cm 50-60cm

90-100cm 90-100cm

Loamy Haplustox (n=3) Clayey Haplustox (n=3)

Figure 4.30. WSA size classes as percent of total soil <8 mm in the different soil depth layers. Bars represent standard error.

104 0 5 10 15 20 25 SOC (g kg-1)

0-5 sandy soils (n=3)

2-8mm 30-40 1-2mm

Depth (cm) 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 5 10 15 20 25 SOC (g kg-1)

0-5 loamy Haplustox (n=3)

2-8mm 30-40 1-2mm

Depth (cm) Depth 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 5 10 15 20 25 SOC (g kg-1)

0-5 clayey Haplustox (n=3)

2-8mm 30-40 1-2mm Depth (cm) Depth 0.5-1.0mm

90-100 0.25-0.5mm 0.11-0.25mm

Figure 4.31. Mean SOC (g kg-1) of WSA size fractions of selected soil depths. Bars represent standard error.

105 0 20406080100Sand content (%)

0-5 sandy soils (n=3) Depth (cm) Depth 2-8mm 30-40 1-2mm 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 20406080100Sand content (%)

0-5 Loamy Haplustox (n=3) Depth (cm) 2-8mm 30-40 1-2mm 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 20406080100 Sand content (%)

0-5 Clayey Haplustox (n=3) Depth (cm) 2-8mm 30-40 1-2mm 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

Figure 4.32. Mean sand content (%) of WSA size classes of selected soil depths. Bars represent standard error.

106 The SOC concentration of the sand fraction within the WSA size classes was used to calculate occluded-POM C as a percent of total SOC within the respective WSA. Figure 4.33 shows that the sandy soil contained relatively more occluded-POM C than the loamy and clayey Oxisols, for the 0-40 cm depth. The sandy soils also had more variability, although the clayey Haplustox was also highly variable at 90-100 cm depth. The sand-corrected, mean SOC concentrations of the WSA size classes are shown in Figure 4.34. As mentioned in sub-chapter 4.2, it estimates SOC concentration in the clay+silt fraction of the WSA size classes. As expected, sand-corrected SOC decreased with depth for all soils and WSA sizes, and these decreases were stronger in coarser soils. There was little variation among WSA size classes for the same soil and depth, indicating that the clay fraction sorbed similar amounts of SOC, in accord with the literature (Feller et al., 1996; Jastrow, 1996, and others). For the same depth, sand-corrected SOC decreased with increasing soil clay content, as a consequence of the SOC dilution effect discussed in sections 4.2 to 4.4. More importantly, it proves that the SOC dilution effect in particle size fractions occurs in natural soil environments and is not merely an artifact from soil dispersion and SOC redistribution. Consequently, it suggests that the SOC- sorbing capacity of the clay fraction is much closer to saturation in sandy than in clayey soils. This does not necessarily mean that clayey soils offer an immense, unexplored SOC sequestration potential by sorption, since it would require exposing closely-arranged clay domains to organic colloids/solutes, probably requiring disruption of the soil matrix. Figures 4.31 to 4.34 indicate that large macroaggregates (>1 mm) in the sandy and loamy soils always had higher concentrations of clay and total SOC, but not sand- corrected SOC, than smaller WSA and the surrounding soil. Also, WSA>0.5 mm had relatively more occluded-POM in the sandy than in the finer-textured soils. This suggests that formation and stabilization of large macroaggregates in sandy and loamy Oxisols depend strongly on the relative accumulation of clay (by any mechanisms) that simultaneously imply relatively higher total SOC retention, and that any aggregation induced by coating and occlusion of labile POM is significant only in coarse-textured soils (<200 g clay kg-1).

107 0 5 10 15 20 occluded-POM (% of total C in WSA)

0-5 sandy soils

Depth (cm) (n=3) 2-8mm 30-40 1-2mm 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 5 10 15 20 occluded-POM (% of total C in WSA)

0-5 Loamy Haplustox

Depth (cm) (n=3) 2-8mm 30-40 1-2mm 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 5 10 15 20 occluded-POM (% of total C in WSA)

0-5 Clayey Haplustox (n=3) Depth (cm)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

Figure 4.33. Mean occluded-POM C as percent of total SOC content in WSA size classes of selected soil depths. Bars represent standard error.

108 0 1020304050607080Corrected SOC (g kg-1)

0-5 sandy soils Depth (cm) (n=3)

2-8mm 30-40 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

0 1020304050607080 Corrected SOC (g kg-1)

0-5 Loamy Haplustox (n=3) Depth (cm)

2-8mm 30-40 1-2mm 0.5-1.0mm 0.25-0.5mm 90-100 0.11-0.25mm

0 1020304050607080 Corrected SOC -1 (g kg )

0-5 Clayey Haplustox Depth (cm) (n=3)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

Figure 4.34. SOC concentration (g kg-1) corrected for sand and sand-sized SOC content of WSA size classes of selected soil depths. Bars represent standard error.

109

The C/N ratios vary little among WSA size classes (Figure 4.35), almost no effect of soil texture occurs, and the C/N ratios decline with depth only in the clayey Haplustox. However, the C/N ratios of occluded-POM in the WSA size classes (i.e., sand-sized SOC) followed a very different pattern. As seem in Figure 4.36, only in the 0-5 cm depth are the C/N ratios comparable those in intact WSA, and even at that depth, C/N ratios of WSA<0.25 mm are much higher than those of intact WSA. For the 30-40 and 90-100 cm depths, the C/N ratios of occluded-POM in all soils are extremely high and variable, especially in the sandy and loamy soils. It is interesting to compare the C/N values of WSA and occluded-POM with those of bulk soil and particle size separates. In the sandy and loamy soils, bulk C/N ratios decreased sharply with depth, but even at the 0-5 cm layer are lower than those of corresponding WSA (Figure 4.4a). The C/N ratio of the total POM (sand-sized SOC) in these two soils is >20 in the 0-5 cm layer and increase with depth (Figure 4.22); this trend also occurs for occluded-POM but the increases with depth are even stronger, reaching almost 200. Considering that C/N ratios of silt and clay fractions are <15 and vary little with depth (Figure 4.22), this proves that total POM causes most of the higher bulk C/N ratios at or near the surface of the sandy and loamy soils. For the lower depths, the extreme C/N ratios of occluded-POM compared to total POM may indicate occlusion of: a) root tissues with initially low C/N ratios, that upon pedological time became devoid of N by mineralization and leaching/ uptake, and/or b) charcoal, as discussed in section 4.3.

The same applies to the clayey Haplustox, although the contribution of high C/N occluded-POM C in subsoil layers was much smaller than for the other soils.

110 0 5 10 15 20 25 C/N ratio

Depth (cm) 0-5 sandy soils (n=3)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

0 5 10 15 20 25 C/N ratio

0-5 loamy Haplustox

Depth (cm) (n=3)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

0 5 10 15 20 25 C/N ratio

0-5 clayey Haplustox Depth (cm) (n=3)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

Figure 4.35. Mean C/N ratio of intact WSA size classes of selected soil depths. Bars represent standard error.

111 0 20 40 60 80 100 120 140 160 180 200 C/N ratio of occluded-POM

0-5 sandy soils Depth (cm) (n=3)

2-8mm 30-40 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

0 20 40 60 80 100 120 140 160 180 200 C/N ratio of occluded-POM

0-5 loamy Haplustox Depth (cm) (n=3)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

0 20 40 60 80 100 120 140 160 180 200 C/N ratio of occluded-POM

0-5 clayey Haplustox

Depth (cm) (n=3)

30-40 2-8mm 1-2mm 0.5-1.0mm 90-100 0.25-0.5mm 0.11-0.25mm

Figure 4.36. Mean C/N ratio of sand fractions from dispersed WSA size classes of selected soil depths. Bars represent standard error.

112 Based on the total contents of POM C (sand-sized SOC pool, Figure 4.20), and WSA size distribution and SOC concentration, the partition of total POM-C outside and inside aggregates is shown in Figure 4.37a. As discussed in section 4.2, at 0-5 cm depth the sandy soils store ca. 40% of their total SOC in the sand fraction, whereas the loamy and clayey Haplustox retain a remarkably similar 25%. For the two lower depths, the percent of total POM C decreases constantly with increasing clay content. Conversely, the relative POM protection in aggregates increases with clay content: in the clayey Haplustox, occluded-POM comprises about half of total POM, whereas in the loamy Haplustox and sandy soils occluded-POM is roughly 33% and 25%, respectively, of total POM. Balesdent et al. (2000) reviewed the literature and reached similar conclusion. For clayey Cerrado Oxisols, Roscoe et al. (2001) found much lower occluded/free-POM ratio, perhaps because they used soil <2 mm instead of WSA, releasing occluded-POM. In terms of absolute units (g kg-1, Figure 4.37b), these trends for total POM are less clear for all depths, although they persist in the case of relative occlusion for the two lower depths. However, the most important notion is that the absolute contents of POM C are remarkably similar among the different soil textures: for 0-5 cm depth, between 5 and 6 g kg-1; for the 30-40 cm, about 1 g kg-1; and for 90-100 cm, about 0.5 g kg-1. These data may suggest that, in spite of the strong differences in SOC concentration and storage due to texture, throughout the textural range the absolute concentration of total POM-C reaches similar equilibrium levels, probably determined by the balance between input and decomposition of coarse residues, which in its turn depends mostly on soil depth. A mathematical model to predict the partition of POM-C throughout free and occluded forms as functions of texture and depth, such as the models obtained in sub- chapter 4.3, was attempted. The only correlations valid for all depths were found for % free-POM with clay and clay+silt. For each depth, % free-POM was best correlated with clay content (R2=0.72-0.90), but since the intercept and slopes of those functions vary irregularly with depth, a profile function was not possible. Figure 4.38 shows the plot of the linear relation % free-POM = 32.03 – 0.0487*(clay, g kg-1) for the combined 0-5, 30- 40 and 90-100 depths. Unfortunately, the wide 95% confidence interval shows that the predictive value of this equation is very low.

113 45 . 40 35 30 25 Occluded 20 Free 15 10

POM (% of total SOC) 5 0 2.5cm 35cm 95cm 2.5cm 35cm 95cm 2.5cm 35cm 95cm

Sandy Sandy Sandy LoamyLoamyLoamyClayeyClayeyClayey a)

7 6

. 5 Occluded 4 Free

soil 3 -1 2 1

g POM kg 0 2.5cm 35cm 95cm 2.5cm 35cm 95cm 2.5cm 35cm 95cm

Sandy Sandy SandyLoamyLoamyLoamyClayeyClayeyClayey

b)

Figure 4.37. Total free- and occluded-POM C of selected soil depths as a) percent of total SOC, and b) absolute units (g kg-1). Bars represent standard error.

114 Free-POM vs. clay content 40

35

30

25

20

15

10

Free-POM (% of total SOC) 5

0 0 100 200 300 400 500 600 700 g clay/ kg soil

Linear Fit

Figure 4.38. Linear relation between percent free-POM C and clay content (g kg-1) for the combined 0-5, 30-40 and 90-100 cm depths (R2=0.74, n=27). Dotted line is 95% confidence interval.

Before discussing the structure control on SOC retention, a general appreciation of aggregation factors in the studied soils is necessary. The results presented in this section lead to conclude that clay is the major aggregation factor in ustic Oxisols, because: a) the clayey Haplustox has consistently higher aggregation (no slaking, higher MWD and WSA>1 mm) than the two other soils throughout their profile; sand content within WSA changes little with depth and size classes, and total and sand-corrected SOC concentrations vary little among WSA size classes at the same depth; b) compared to the clayey Haplustox, soils of loamy and coarser texture undergo slaking (pronounced in the sandy soils), and are characterized by much fewer WSA>1 mm. These WSA>1 mm always contain higher clay contents than smaller aggregates and bulk soil, which in part cause a higher total SOC levels. Nevertheless, relatively higher occluded-POM C in WSA>0.5 mm in the sandy soils suggests that POM-

115 occlusion plays a role in the formation and stabilization of macroaggregates only in coarse-textured soils (see further discussion later in this section).

To detect the role of clay as the major aggregation factor, it seems critical to sample soils along a broad textural range, with ceteris paribus. As reviewed in section 2.3, the effect of different aggregation factors on soil structure has long been investigated by means of mathematical correlations and selective extractions, but with little uniformity of methods for obtaining and treating the aggregates (Ashman et al., 2003; Loveland and Webb, 2003). Consequently, the results have often lead to disparate conclusions, especially with respect to SOC partitioning through macro- and microaggregates, as reviewed by Feller et al. (1996), Loveland and Webb (2003), and Blanco-Canqui and Lal (2004). The interaction of soil climate, physical properties, cultivation and C input add further complications to comparison purposes (Balesdent et al., 2000). Additionally, the data on profiles of different-textured soils presented here strongly suggest that contradictory results found in the literature are also caused, at least partially, by comparing SOC and aggregation throughout a single profile, or in soils within a narrow textural ranges, or by lack of comparability between soils of disparate texture and mineralogy. Although quantitative mineralogy of WSA fractions was not determined, one has no reason to suppose that it would differ considerably from that of bulk soil. In the studied soils, the clay content is also related to total content of Fe-oxides in bulk soil, as shown in section 4.4. Even for soils in temperate regions, Fe-oxides can play an important role in aggregation. In Wisconsin, Chesters et al. (1957) found that the best correlations between percent of WSA sizes were with microbial gums; Fe oxides were also important (esp. for WSA<0.1 mm), but clay content was important only when >30%, and total SOC showed little relation. For three soils in Spain, Barral et al. (1998) concluded that concentrations of Fe-oxides and SOC were directly related to aggregate stability. In tropical soils, Fe-oxides become even more important, with little apparent effect of texture. In the Brazilian Cerrados, Neufeldt et al. (1999) found that the percent of macroaggregates in a clayey Acrustox was only correlated to Fe-oxalate and POM

116 (r=0.66 and 0.61, respectively). For a sandy Haplustox, macroaggregation was best correlated to cellulose, Fe-oxalate and SOC contents (r=0.83, 0.79, 0.78, respectively). Denef et al. (2002) incubated soils ground to <0.25 mm from temperate and tropical regions, and reported that a Brazilian Hapludox was the most efficient in re-forming macroaggregates which, despite the different clay and SOC contents among soils, suggests that aggregation in Oxisols is more related to Fe/Al oxides than SOC. The role of SOC as an aggregation factor must be assessed with caution. For all soils, the depth-wise decrease in bulk SOC concentrations is much more gradual than that of MWD and % WSA>2 mm (Figure 4.5), and only for the 0-5 cm depth does SOC correlate well with aggregation indicators (Figure 4.29). Although higher aggregation generally follows higher SOC levels, one cannot attribute the higher aggregation in topsoils to SOC alone, since sand-corrected SOC and % occluded-POM C vary little among WSA size classes in the top 5 cm. Other aggregation factors are also much more active in the surface layers, such as biologic activity (see next section), and swelling/shrinking cycles caused by moisture and temperature fluctuations (Bronick and Lal, 2005). Dissimilar variations in SOC and aggregation in native and cultivated soils have been noticed earlier (Haynes and Swift, 1990). Investigating a chronosequence of prairie restorations in Mollisols, Jastrow (1996) reported that after 10 yrs the percent of macroaggregates (>212 μm) equaled that of virgin soils, whereas bulk, sand-corrected SOC concentration was only <50% of virgin soils and still increasing after 12 yrs. Increased macroaggregation also enhanced total free-POM, occluded-POM and sorbed SOC (different names were used), while total SOC in microaggregates decreased. She concluded that formation of macroaggregates occurs before microaggregates, which was reinforced by better correlations with mycorrhizal hyphae and fine roots than with total SOC (Jastrow and Miller, 1998). Without discussing the time of formation, one can argue that this interpretation is based solely on a sharp limit of micro- and macroaggregate size using the aggregate hierarchy concept (Edwards and Bremer, 1967; Oades and Waters, 1991). However, WSA form a size continuum from small platelets to clods and even if the hierarchy works for all soils (even with variable “macro/micro” limits), using MWD

117 would better indicate the aggregation status. However, at least for Oxisols, MWD is also not strongly correlated with SOC, as demonstrated here and by Zotarelli et al. (2005), who reported that the decline of SOC concentration (sand-corrected, no free-POM separation) upon cultivation was much stronger than that of MWD. Loveland and Webb (2003) reviewed other examples of weak correlations between SOC and MWD. Tisdall and Oades (1982) reviewed many relations between aggregate stability and SOC concentrations in widely different soils, and concluded that SOC is not the major binding agent, not all SOC takes part in the stabilization, and there is a SOC level beyond which there is no more aggregation effect. The data presented in this section and interpretation of other literature confirms that soil aggregation is related to SOC, but not strongly enough to be considered causative and to allow predictive functions such as for texture. As reviewed by Balesdent et al. (2000), SOC contributes to its own preservation by enhancing aggregate stability. This feedback nature of the SOC-aggregation relation renders an appraisal of the structural control of SOC retention more difficult. In other words, the difference between “SOC as an aggregation factor” and “structural control of SOC retention” is a chicken-and-egg discussion. Also, it is important to stress that the protection given by aggregation is partial, since it does not stop decomposition but rather reduces its rate (Baldock and Skjemstad, 2000). The structural control on SOC can better be appreciated for POM, although it is known that easily dispersible clay is SOC- impoverished compared to hard-to-disperse (i.e., aggregated) clay (Nelson et al., 1999). The synthetic data in Figures 4.37 and 4.38 show that in most cases, most POM-C pool is not protected by soil aggregation, which agrees to data from a cultivated sand clay loam in Georgia,USA (Beare et al., 1994) and from silty forest soils in France (Besnard et al., 1996). Also, this aggregate protection increases with clay content to a maximum of 50% of total POM. Thus, the structural control on SOC retention is restricted to part of the POM pool, and clearly secondary and dependent on the textural control. Nonetheless, this does not mean that the structural control is not important, since aggregation reduces SOC accessibility to decomposers (Sollins et al, 1996). On the other hand, there is little doubt that POM-occlusion is an effective aggregation agent in coarse- textured soils, where microbial activity depends strongly on POM. For silty soils, Puget

118 et al. (1995) concluded that only SOC-rich macroaggregates resist slaking. Short-term aggregation induced by glucose and N amendments was more pronounced in sandy than in finer soils, and in soils with high initial microbial biomass (Kiem and Kandeler, 1997). For coarse-textured soils of the Niger floodplain, Igwe and Stahr (2004) found that SOC was directly related to WSA>2 mm, reinforcing the idea that SOC is a major factor for stabilization of large aggregates if low clay contents limit cementation.

Micromorphology of WSA 2-8 mm: common and faunal peds and SOC

Micromorphology is the only direct assessment of soil aggregation in actual field conditions, with no disturbances such as dry disruption or slaking on quick wetting. Conventional soil micromorphology usually describes undisturbed soil samples or clods with known orientation. However, a few authors have prepared thin sections of various WSA size classes, including the classical paper of Tisdall and Oades (1982) and Chotte et al. (1994). The thin sections of WSA described here can only provide information comparable to the basic structure of Brewer and Sleeman (1960), which defines the size, shape and arrangement of skeleton, plasma and voids within single peds (aggregates), a concept later named s-matrix (Brewer, 1964). No information such as interpedal arrangement/porosity or free-POM location is available through this approach. However, the identification of peds with different origin (abiotic or faunal) within the disturbed soil sample and their resistance to slaking and water-sieving is greatly facilitated in relation to the conventional micromorphology of undisturbed samples. This is clearly shown in the low-magnification Figures c.1 and c.2 in Appendix C. In order to better illustrate the differences between the two major types of WSA and their pedogenetic implications, their micromorphologies are described separately. All micrographs referred in the following text by consecutively numbered figure captions are contained in Appendix C.

Subangular blocks Most of WSA 2-8 mm in the profile of all soil types appear to be subangular blocky (Figure c.1), although no systematic soil and structure description was conducted in the

119 field. The 2-8 mm blocky peds are more common in the top 10 cm layer but occur at all depths, including the sandy soils which mostly have a single-grain structure. The c/f (coarse to fine) distribution of Stoops and Jongerius (1975) is a description of the proportion and arrangement of skeletal grains (e.g., gravel and sand) and plasmic (e.g., clay and colloidal SOC) components of the soil fabric, and will be described here according to the more intuitive nomenclature by Eswaran and Baños (1976). The WSA 2- 8 mm in the Quartzipsamment (ca. 100 g clay kg-1 bulk soil) show mostly a close-spaced dermatic (plasma coatings around grains) to intertextic (plasma bridges among grains) character, as shown in Figure c.3. In the sandy Haplustox (ca. 170 g clay kg-1 bulk soil), the higher clay content results in a pronounced intertextic c/f distribution (Figure c.4). In the blocky WSA 2-8 mm from all depths of the clayey and loamy Haplustoxes, the c/f distribution is porphyric, i.e. coarse grains within a dense groundmass of finer material, which is generally observed in Oxisols (Stoops and Buol, 1985). The different texture of the Oxisols is evident in the thin sections: it is close-spaced (skeletal grains in close contact) in the Loamy Haplustox (Figure c.5), and open-spaced (skeletal grains distant from each other) in the Clayey Haplustox (Figure c.6). The more rare, congelic c/f distribution was observed in some sparse whitish peds within the Quartzipsamment, consisting of plasma as in silt-sized aggregates isolated from skeletal grains, which seem to indicate little surface affinity with the grains (Figure c.7). The strong intertextic c/f distribution of the sandy Haplustox can represent a transition between the Quartzipsamment and loamy Haplustox, forming a pseudo- or sub-porphyric pattern. The differences in c/f distribution within the Quartzipsamment-Loamy Haplustox toposequence shown here are similar to those demonstrated by Bravard and Righi (1988) for a Spodosol-Oxisol toposequence in Amazon. In the sandy and loamy soils, skeletal grains are exclusively well-rounded quartz, coarser and less spherical in the sandy soils (Figure c.3-5), which may indicate limited sand transport in sandy soils and reinforces their origin from the Areado sandstone, whereas it may also point to some transport/ sorting in the loamy Haplustox, perhaps under the influence of dissection lines. In many cases, the quartz grains show irregular fractures or etchings impregnated with plasma (Figure c.3, c.8, also c.16), most likely Fe

120 oxides. These etched grains are an indicator of extreme weathering and are called runiquartz by Eswaran et al. (1975, cited by Stoops, 1983). Some features also resemble the solution lines (Figure c.3, c.8) and triangular/rhomboidal etch pits, often coated by Si, Al and Fe, described by Newsome and Ladd (1999) in the Australian sand plains. The clayey Haplustox has a very different skeletal fraction, whose most distinctive skeletal type are hard, opaque nodules of fine sand to gravel size, mostly of Fe and Mn oxides. The opaque Fe nodules almost always contain quartz and other minerals (Figure c.2i, c.6, c.9) and are remarkably similar to those found in Thai Ultisol laterites (Suddhiprakarn and Kheoruenromne, 1994) and Australian Plinthustalfs (Mucher and Coventry, 1994). According to Brewer (1964), these plasma concentrations can be of a mixed sesquioxidic/ manganiferous nature, round or irregular in shape, and with sharp or diffuse outline, and are often of a cellular type (Figure c.10 and c.11, Fitzpatrick, 1993, p. 201). Manganese coatings (neomangans) also occur in the clayey Haplustox (Figure c.11). Nodules and coatings probably result from spatially variable soil aeration due to imperfect drainage, since both often occur on the aerobic microsites of ped surfaces or intrapedal voids where soluble, reduced forms are oxidized and precipitate. Quartz in the clayey Haplustox is mostly small, sub-rounded and angular, with some runiquartz, and suggest little or no transport and a relatively stable pedoenvironment. Gravel-sized quartz and euhedral (perfectly-shaped) crystals and also occur, probably composed by the extremely resistant minerals zircon or rutile (Figure c.15). Pores and voids can play a major role in intrapedal processes and are described separately. Although pore spaces are identifiable and measurable in thin sections, their micromorphologic study is not a quantitative technique such as conventional water retention curves, due to the small and planar fraction of soil volume effectively studied. Nevertheless, their qualitative description according to Buol et al. (1997) provides additional information on how texture affects soil structure in the studied soils. Pores within WSA 2-8 mm of sandy soils are mostly packing voids, although vughs (irregular- shaped and walled voids) and joint planes (cracks) occur where plasma is more concentrated (Figure c.3). The loamy and clayey Haplustoxes are dominated by vughs, craze planes (irregular-shape, elongated voids) and some joint planes (Figures c.1, c.5,

121 c.6, c.8, c.10-14). Some vesicles (round, smooth-outline pores) are visible in the loamy Haplustox (Figures c.5, c.14). Other notable types of voids are wide channels or tunnels that do not disrupt, or perhaps help stabilizing, WSA 2-8 mm (Figure c.2g-i). They are more common in the 0-5 cm layer and almost certainly are of a biological origin, probably from root growth or burrowing activity of fauna, and are not plasma-enriched in relation to other blocky peds. Plasma separations were not observed in thin sections, which may be due either to their absence (asepic s-matrix) or because of the ubiquity of Fe-oxides and SOC in the matrix, resulting in an isotropic fabric that precludes visualization of plasma separations (Brewer, 1964).

Faunal or zoogenic peds

Micromorphology indicates that the characteristic external properties of faunal peds are maintained in their interior, indicating a different genesis rather than surface reworking of subangular blocks. The occurrence of zoogenetic or zoogenic micropeds as irregular patches in Oxisol surface horizons was briefly described by Stoops and Buol (1985), although the significant role of fauna in the structure of these soils has been suspected and shown by many authors (see the review by Schaeffer, 2001). In the soils studied here, zoogenic peds occur as: a) mostly aggrotubules; b) fecal pellets; and c) cocoons. Aggrotubules are defined by Brewer (1964) as a special type of pedotubules, in which the infilling of faunal tunnels and chambers was actively carried out by the organisms rather than passively by gravity after abandonment. Aggrotubules are the most striking and abundant of the zoogenic peds: they comprise 12% of the weight of WSA>2 mm in the Quartzipsamment, and 6, 11 and 26 % in the sandy, loamy and clayey Haplustoxes, respectively. Although varying somewhat in color and size, they are basically clusters (1.2 cm max. diameter), built of smooth-outlined ellipsoidal crumbs of 0.5-1 mm diameter (Figure c.2a-c). The crumbs are mutually welded, with a moderate to large packing porosity that often includes quartz grains and POM (Figures c.2a-b, c.15-

122 18). Their porphyric s-matrix is very different from that of the subangular blocks, with much smaller and fewer skeletal grains (as often reported for zoogenic peds) and no apparent internal voids. This concentration of plasma is partially responsible for the higher C and N contents in the sandy soils and Loamy Haplustox (Table 4.15). They fit the variety 3 aggrotubule described by Brewer (1964), which is mostly composed of reworked soil material and morphologically very different from surrounding peds. In contrast to fecal pellets, no visibly shrunk organic particles exist in their interior, suggesting that most of their inner organic matter is colloidal or soluble. However, the round, smooth outline of individual grains in aggrotubules suggests that they were egested in a semi-liquid state rather than reworked in solid state. Little is known about aggrotubules and not many thin sections are available in published literature other than in Brewer (1964), and rarely under the name of aggrotubules. Examples are the “granular infilling” in a Ustoxic Dystropept of Zaire described by Stoops et al. (1994) and in coarse-textured soils near Sydney, Australia by Humphreys (1994). This author estimated that about 8% of the A and E horizons are faunal-produced channels, chambers and pedotubules, a number similar to the percent of aggrotubules observed in the WSA 2-8 mm from the sandy soils in the present study. However, aggrotubules tend to be more common in finer-textured soils, which indirectly agrees to the higher frequency and surface area of termite mounds in clayey soils (Sys, 1955, cited by Lal, 1987). Balbino et al. (2002) presented micrographs of Cerrado soil showing granular aggregates in biological channels and vughs that correspond to the aggrotubules described here, and correctly identified its faunal origin. Gomes et al. (2004) identified aggrotubules in B horizons of different Cerrado soils. The amount of aggrotubules in all soils indicates that their builders are widely distributed and a major player in the formation of structure in neotropical Savanna soils. Most likely they were social arthropods such as termites or ants, since both are more common than earthworms in the Cerrado, and aggrotubules are very different from worm casts in thin sections (Lavelle et al., 1992). Indeed, mound termites and leaf-cutting ants are common in the sampled areas, whereas no earthworms were noted.

123

Fecal Cocoons Bulk Soil Aggro- Subang. tubules pellets Blocks Soil (0-5 cm depth) ------SOC g kg-1 ------(C/N) ------Quartzipsamment 74.4 30.3 27.5 17.6 10.7 (15.8) (26.5) (16.1) (22.6) (12.4) Sandy Haplustox 25.1 30.3 18.1 15.2 (16.8) (26.5) - (23.3) (13.9) Loamy Haplustox 22.6 32.2 17.0 20.3 (14.5) (15.8) - (15.9) (13.4) Clayey Haplustox 22.7 33.3 22.1 24.6 (15.4) (17.1) - (15.0) (14.5)

Table 4.15. SOC concentration and C/N ratios (in parenthesis) of faunal peds, blocky WSA>2mm, and bulk soil under Cerrado.

It is not clear if aggrotubules are built exclusively by termites, since relatively less is known about ant effects in tropical soils (Alvarado et al. 1981, Lal, 1987). Decaëns et al. (2001) ascribe the presence of aggrotubules in the Colombian Llanos to termites. In termite nests or mounds, soil plays a structural role as a finely packed and cemented material (Matthews, 1977) molded as cohesive mound (Lal, 1987). Termites can also use and produce granular aggregates, and instant production of aggrotubules by unidentified African termites has been visually documented (National Geographic Society, 2001). However, thin sections of soil material in Macrotermes nests by Jungerius et al. (1999) in Kenya show little similarity with the aggrotubules described here. Conversely, even when forming mounds, tropical ants transport and use single-grained, loosely-packed or –piled soil with low bulk density and high permeability (Lal, 1987, p. 432, Decaëns et al., 2001). Weber (1972) presented many pictures of soil in Attine (leaf-cutting, fungus-gardening ants) nests as reworked, round grains and crumbs, loosely piled where structural role is

124 not required such as in craters and the bulk of the nest. However, in places where loads are supported against gravity such as chambers, galleries and turrets, individual crumbs would have necessarily to be welded to act structurally, and aggrotubules would be a remnant of those materials. Eschenbrenner et al. (1994) described aggrotubules formed by backfilling of channels and chambers within nests built by minor Attine ants in Andosols of Martinique, similar to those described here except for the smaller size. Jungerius et al. (1999) reported welding of soil grains of similar size as described here, as the result of ant activity on abandoned termite mounds. The amount of soil disturbed by Attine ants also agrees with the percentage of aggrotubules reported here. Alvarado et al. (1981) reported that 39 and 85% of the surface and belowground of some Costa Rican Andosols show pedoturbation by Atta cephalotes, although only 1% of the topsoil were active, live nests or mounds. The authors even describe that normal A horizons are overlain by an entomogenic Ai, and that “filled-in chambers” are the more common structures in relict nests. In Brazil, Coutinho (1984) affirmed that Atta are the main primary consumer in the Cerrados and their massive belowground nests can have fungus chambers up to 3 m deep, while refuse deposits reach deeper than 7 m. Although remains of heads of soldier ants were found in the WSA 2-8 mm from the loamy Haplustox (Atta sexdens var. rubropilosa, a common pest in Eucalyptus plantations), no conclusion about the actual origin of aggrotubules can be drawn from the descriptions in this study. The fate of the aggrotubules after their formation apparently differs among soil type: in the loamy and clayey Haplustoxes, some of the WSA classified as subangular blocks appear to result from the compaction of aggrotubules forming a block with a faintly granular surface, and others with a regular surface but showing internal grain welding (Figure c.13). This does not occur in the sandy soils. However, aggrotubules may also break into smaller clusters or individual crumbs which would occur in the smaller WSA fractions, which can be seen in the micrographs of sandy and clayey Haplustoxes (75-80 cm) of Balbino et al. (2002). For deep B horizons of Brazilian clayey Oxisols, Schaeffer (2001) found many structures which are clearly aggrotubule remnants, and based on a comprehensive literature review, constructed an interesting theory about the granular aggregation of those soils resulting from the interaction of termites,

125 mineralogy, and evolution of angiosperms through geological history of tropical America and Africa. The relatively large fecal pellets (Figures c.2e,f) are mostly mineral, have an ellipsoidal, smooth outline and comprise <0.1% of the mass of WSA 2-8 mm of all soil types. As for the aggrotubules, their porphyric s-matrix contains less and smaller skeletal grains than the blocky peds. However, they contain numerous organic particles surrounded by voids caused by their preferential shrinking during drying at 105oC (Figures c.19-20). These fecal pellets resemble those produced by Coleopterae larvae (Fitzpatrick, 1992, p.141), also characterized by a high content of mineral material but smaller than those shown here. Mostly mineral fecal pellets ca. 5 mm long are produced by large beetle larvae feeding on decaying termite mounds in the Cerrados (Matthews, 1977). Another possibility is that those pellets are produced by cicada larvae, common in the Cerrados. Cicada pellets in Australia are 4-6 mm in size (Humphreys, 1994), similar to those described here. Fecal pellets were the only type of WSA 2-8 mm which always contained visible POM and their C content varies little with texture of the surrounding soil (Table 4.15). Cocoons were observed in all soil types, but are much more common and larger in the sandy soils. They consist of soil tubes, hollow or backfilled, 2-3 mm in diameter and up to 1 cm long (Figure c.2d). They are strongly concentrated in plasma, compared to the surrounding soil and WSA with tunnel/channels (Figures c.2g, h, i). In thin cross- sections, they appear almost opaque, probably because of high humus content. However, their SOC content is lower than that of aggrotubules and fecal pellets (Table 4.15), probably because of a significant content of coarse quartz. In the soils studied here, few occluded-POM were visible (Figures c.5, c.17). Nevertheless, a dark blocky ped from the loamy Haplustox showed many occluded charcoal fragments (Figure c.21), ascribed to the wildfires which regularly occur during the dry season. Figure c.22 is a detail of Figure c.5 showing a root that underwent internal decomposition but retained its outer, more recalcitrant part, to which Fe-impregnated clay adhered to, probably conferring additional resistance to microbial attack.

126

5. CONCLUSIONS

™ All soils studied are typical of the Cerrado region, being highly weathered and composed mostly of quartz (sand and silt fractions) and kaolinite (clay fraction). The clay fraction also comprised minor and variable contents of Fe-Al oxides, hydroxyl-interlayer vermiculite and illite. Base saturation, CEC and pH are low compared to soils of temperate climates but consistent with other Cerrado soils. The three soil types were classified as clayey-skeletal Haplustox, loamy Haplustox and sandy soils (replication 1 is a Quartzipsamment and reps. 2 and 3 a sandy Haplustox). ™ After two cycles of Eucalyptus growth (14 yrs), there were no significant changes in chemical properties, but in the surface 10 cm the C/N ratio increased, and aggregation as indicated by mean weight diameter (MWD) and percent of water- stable aggregates (WSA) >2 mm decreased, probably because of the use of heavy disk harrow before planting the forests. Bulk soil organic carbon (SOC) concentration and stocks did not change under afforestation, confirming hypothesis e (p. 19). However, SOC size partition and relative protection in aggregates was affected at the 0-5 cm depth: SOC was enriched in the sand fraction and depleted in the clay fraction of the sandy soils, and in all soils the decreased aggregation resulted in lower relative occlusion of particulate organic matter (POM) inside aggregates. ™ SOC concentration and stocks are strongly affected by soil texture and depth. A mathematical model based on depth and clay+silt content was developed to predict SOC concentration in bulk soils, and further extended to predict the relative partition of SOC to the clay fraction based on clay contents and depth.

127 ™ Soil specific surface (SSA) area was modeled as function of clay, silt and SOC concentrations but not depth. Aside from the texture-SOC models described above and SSA of the clay fractions, these data suggest that, for a range of soils at a same depth, SOC levels increase with increasing SSA caused by higher clay contents. However, in a single soil profile, SOC promotes clay flocculation and aggregation, so SSA increases with decreasing SOC contents with depth. ™ Quantitative mineralogy of the clay fraction showed that SOC concentrations in bulk soils and the clay fraction are best correlated by crystalline Fe-oxides in surface and amorphous Al oxides in subsoil, with even better determination coefficients than those for SOC vs. clay+silt. ™ Aggregation (MWD and WSA>2 mm) is, like SOC concentration, strongly correlated to clay+silt contents, but SOC is weakly correlated with MWD and WSA>2 mm except for the 0-5 cm depth. The percentage of POM occlusion inside aggregates is strongly affected by soil texture, varying from ca. 25% in the sandy soils to ca. 50% in the clayey Haplustox. ™ Activity of soil fauna results in three types of zoogenic aggregates (aggrotubules, fecal pellets and cocoons), which are SOC-enriched in soils of loamy and coarser texture. ™ Therefore, hypotheses a to d (p. 19) are confirmed, but not to the same degree for the three controls: although SOC is better correlated with Fe and Al oxides than with clay+silt, the contents of those minerals in soils depend directly on clay content. Likewise, the protection of POM inside aggregates also depends strongly on clay content. Considering the subjacent and inseparable effect of texture, SOC retention in aerobic Cerrado soils is controlled, in decreasing order of importance by: 1) texture, 2) mineralogy, and 3) structure.

128

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145

APPENDIX A

SAMPLING LOCATIONS

146

Figure A.1. Map of Nova Esperança Farm in Unaí-MG and detail with sampling sites on Eucalyptus stands and Cerrado strips (red dots). Courtesy of V & M Florestal Inc.

Figure A.2. Aerial view (20-m observation deck), looking North, of the Clayey Haplustox landscape with Eucalyptus stands (native Cerrado not visible).

147

Figure A.3. Eucalyptus and native Cerrado plots on clayey Haplustox, Unaí-MG.

Figure A.4. Sampling of Cerrado litter layer, Chapadinha Farm, Unaí-MG.

148

Figure A.5. Map and detail of Chapadinha Farm in J. Pinheiro-MG, with sampling sites on the toposequence from Loamy Haplustox (3 upper pairs of red dots) to Quartzipsamment (3 lower pairs). Courtesy of V & M Florestal Inc.

Figure A.6. Aerial view of transition Loamy Haplustox (red brown) to Quartzipsamment (gray) looking South. Strip (25 m width) in the middle is native vegetation among Eucalyptus. Voz Florestal 1, 2003 (V&M Florestal Newsletter).

149

Figure A.7. Eucalyptus and native Cerrado plots on Loamy Haplustox, J. Pinheiro.

Figure A.8. Eucalyptus and native Cerrado plots on Quartzipsamment, J. Pinheiro. 150

APPENDIX B

ASPECTS FROM FRACTIONATIONS AND SEPARATES

151

Figure B.1. Water-stable aggregates (WSA, 0-5 cm depth, under Cerrado) before flotation of POM. From left to right: 2-8, 1-2, 0.5-1, 0.25-0.5, 0.11-0.25 mm size classes. Lower row: Quartzipsamment, Middle row- Loamy Haplustox, Upper row –clayey Haplustox.

152

Figure B.2. WSA<0.25 mm (Quartzipsamment, 0-5 cm depth) after flotation of free- POM. Note dominance of single grains and absence of POM. Scale in mm.

Figure B.3. WSA<0.25 mm (Loamy Haplustox, 0-5 cm depth) after flotation of free- POM. Note lower amount of single grains and absence of POM. Scale in mm.

153

Figure B.4. WSA<0.25 mm (Clayey Haplustox, 0-5 cm depth) after POM flotation. Minor presence of quartz / opaque grains, absence of free-POM. Scale in mm.

Figure B.5. Silt fraction from dispersion of bulk sample (Loamy Haplustox, 30-40 cm depth). Dominance of quartz, dark grains probably free-POM and Fe-oxides (macroscopically, the sample has an overall light red color). Scale in mm.

154

Figure B.6. Dispersed (NaOH 0.1M), sieved (>20μm) WSA from Sandy Haplustox (0-5 cm, under Cerrado) showing occluded-POM deposited over sand. From left to right: 0-5, 1-2 and 2-8 mm sizes. 12 h after sieving.

Figure B.7. Dispersed WSA<0.25mm from clayey Oxisol (0-5 cm) before and after sieving. Most SOC is humic (dark suspension), contrasting with sandy soils. 155 a)

b)

c)

Figure B.8. Selected WSA>2mm (0-5 cm depth) of pedogenic and faunal origin: a) Clayey Oxisol, subangular blocks (note roots) contrasting with granular cluster on left; b) Quartzipsamment, cocoons; c) all soils, fecal pellets.

156

APPENDIX C

SOIL THIN SECTIONS

157 Quartzipsamment Loamy Haplustox Clayey Haplustox

0-5 cm

30-40 cm

90-100 cm

Figure C.1. Low magnification (optical microscope) overviews of semi-thin sections of subangular blocky, water-stable aggregates (WSA) 2-8 mm (scale in mm).

158 Quartzipsamment Loamy Haplustox Clayey Haplustox Aggrotubules

a) b) c) Cocoon / Fecal pellets

d) e) f) Tunnels and channels within WSA>2mm

g) h) i)

Figure C.2. Low magnification overviews of semi-thin sections of biogenic WSA 2-8 mm or with biogenic tunnels / channels (0-5 cm depth, scale in mm).

159

Q Q V Q V Q V Q

V Q V Q

V Q Q

Q Q

Figure C.3. Quartzipsamment, block, 0-5 cm: close-spaced, dermatic/intertextic s-matrix. Note large packing voids and round, some etched, quartz grains (BCP1).

V

Q

Q Q

Q

Q Q

Figure C.4. Sandy Haplustox, block, 0-5 cm: close-spaced sub-porphyric s-matrix. Note dark, isotropic plasma and well-rounded skeleton grains (BCP).

1 BCP = between crossed polars; Q = quartz, V = voids. 160 V

V

Q Q

Figure C.5. Loamy Haplustox, block 0-5 cm: open-spaced porphyric s-matrix. Note hollow organic structure and surrounding void in upper right corner (BCP).

V

V V

V

Figure C.6. Clayey Haplustox, block, 0-5 cm: open-spaced porphyric s-matrix. Note opaque nodules (PPL2).

2 PPL = plane polarized light; BCP = between crossed polars; Q = quartz, V = voids. 161

Q

Q Q

V Q

Figure C.7. Quartzipsamment, light-colored block, 0-5 cm: congelic s-matrix. (BCP3).

Q V

Q Q Q

Q Q Q

V

Figure C.8. Loamy Haplustox, block, 0-5 cm: runiquartz (BCP).

3 BCP = between crossed polars; Q = quartz, V = voids. 162

V Q

Q

Figure C.9. Clayey Haplustox, block, 30-40 cm: a Fe oxide nodule (non-effervescent with 4 30% H2O2) including quartz grains (BCP ).

V V V

V

Figure C.10. Clayey Haplustox, block, 0-5 cm: cellular nodule of Mn oxide, effervescent with 30% H2O2 (PPL).

4 BCP = between crossed polars; PPL = plane polarized light; Q = quartz, V = voids. 163

V

Figure C.11. Clayey Haplustox, block, 0-5 cm: neomangan on surface of craze plane and 5 cellular nodule, both effervescent with 30% H2O2 (PPL ).

V

Figure C.12. Clayey Haplustox, block, 30-40 cm: unknown mineral in a vugh. The light- colored, concentric area results from poor resin impregnation (BCP and PPL).

5 PPL = plane polarized light; BCP = between crossed polars; Q = quartz, V = voids. 164

V

V

V

V

Figure C.13. Clayey Haplustox, block, 90-100 cm: a former aggrotubule? (PPL6).

V

V

Figure C.14. Loamy Haplustox, block, 90-100 cm: craze plane in the porphyric s-matrix (BCP).

6 PPL = plane polarized light; BCP = between crossed polars; Q = quartz, V = voids. 165

V

V

Figure C.15. Clayey Haplustox, aggrotubule, 0-5 cm, unknown euhedral crystal (BCP7).

Q V

Q

Figure C.16. Loamy Haplustox, aggrotubule, 0-5 cm with runiquartz (BCP).

7 PPL = plane polarized light; BCP = between crossed polars; Q = quartz, V = voids. 166

Q Q

V

Figure C.17. Quartzipsamment, aggrotubule, 0-5 cm, near welding of two grains (BCP8).

V

Figure C.18. Loamy Haplustox, aggrotubule, 0-5 cm, welded grains and POM (BCP).

8 BCP = between crossed polars; POM: particulate organic matter, Q = quartz, V = voids. 167

V

Figure C.19. Loamy Haplustox, fecal pellet, 0-5 cm, with many shrunken POM (BCP9).

V

Figure C.20. Clayey Haplustox, fecal pellet, 0-5 cm, with large POM, detail of C.2f (PPL).

9 PPL = plane polarized light; BCP = between crossed polars; POM: particulate organic matter; V = voids. 168

V

V

Figure C.21. Loamy Haplustox, subangular block 0-5 cm, showing charcoal with cell 10 wall structure (non-effervescent with H2O2, PPL ).

10 PPL = plane polarized light; V = voids. 169

V

V

Figure C.22. Loamy Haplustox, subangular block 0-5 cm (detail of Figure C.5), showing root with decomposed core but preserved epidermis to which Fe-impregnated clay adhered (PPL11).

11 PPL = plane polarized light; V = voids. 170

APPENDIX D

ORIGINAL DATA (EXCERPTS)

171

Soil Land Depth Rep Total Total Fine Coarse Bulk use N C Clay Silt sand sand dens. (cm) ------g kg-1 soil------g cm-3 sandy Cer. 2.5 1 0.86 10.69 70 0 620 310 1.24 sandy Cer. 7.5 1 0.47 5.04 70 0 610 320 1.35 sandy Cer. 15 1 0.42 4.38 90 0 660 250 1.41 sandy Cer. 25 1 0.34 3.29 90 0 600 310 1.37 sandy Cer. 35 1 0.30 2.80 80 0 640 280 1.24 sandy Cer. 55 1 0.24 2.21 70 0 580 350 1.30 sandy Cer. 95 1 0.24 2.11 100 0 570 330 1.39 sandy Euc. 2.5 1 0.95 16.72 100 0 560 340 1.00 sandy Euc. 7.5 1 0.65 9.91 90 0 600 310 1.35 sandy Euc. 15 1 0.35 4.48 90 0 520 390 1.48 sandy Euc. 25 1 0.42 5.08 110 0 430 460 1.21 sandy Euc. 35 1 0.26 3.17 100 0 550 350 1.34 sandy Euc. 55 1 0.30 3.61 110 0 460 430 1.29 sandy Euc. 95 1 0.19 2.34 130 0 470 400 1.47 sandy Cer. 2.5 2 1.21 17.18 140 0 490 370 1.09 sandy Cer. 7.5 2 0.95 13.45 130 0 470 400 1.23 sandy Cer. 15 2 0.60 7.96 140 0 500 360 1.28 sandy Cer. 25 2 0.57 6.82 150 0 370 480 1.39 sandy Cer. 35 2 0.53 6.79 160 0 440 400 1.41 sandy Cer. 55 2 0.34 4.47 170 0 380 450 1.17 sandy Cer. 95 2 0.27 3.32 180 0 360 460 1.45 sandy Euc. 2.5 2 1.09 18.90 150 0 510 340 1.04 sandy Euc. 7.5 2 0.60 8.22 110 0 510 380 1.21 sandy Euc. 15 2 0.41 5.09 90 0 430 440 1.18 sandy Euc. 25 2 0.40 4.93 170 0 440 390 1.17 sandy Euc. 35 2 0.32 4.36 180 0 360 460 1.28 sandy Euc. 55 2 0.27 3.36 210 0 340 450 1.25 sandy Euc. 95 2 0.23 2.71 190 0 410 400 1.11

Continued

Table D.1. C and N, textural and bulk density data. Depth refers to center of layer.

172 Table D.1 continued.

Soil Land Depth Rep Total Total Fine Coarse Bulk use N C Clay Silt sand sand dens. (cm) ------g kg-1 soil------g cm-3 sandy Cer. 2.5 3 0.97 13.20 120 0 570 310 1.21 sandy Cer. 7.5 3 0.77 10.21 120 0 470 410 1.25 sandy Cer. 15 3 0.60 7.46 120 0 610 270 1.12 sandy Cer. 25 3 0.58 6.72 120 0 470 410 1.24 sandy Cer. 35 3 0.42 4.85 160 0 420 420 1.19 sandy Cer. 55 3 0.40 4.39 180 0 440 380 1.47 sandy Cer. 95 3 0.26 3.03 130 20 460 390 1.23 sandy Euc. 2.5 3 0.90 15.61 130 0 510 360 1.23 sandy Euc. 7.5 3 0.57 8.10 120 0 500 380 1.32 sandy Euc. 15 3 0.55 6.86 130 0 480 390 1.18 sandy Euc. 25 3 0.45 5.69 130 0 390 480 1.43 sandy Euc. 35 3 0.41 4.99 140 0 470 390 1.30 sandy Euc. 55 3 0.32 3.74 190 0 330 480 1.47 sandy Euc. 95 3 0.29 3.00 190 0 400 410 1.52 loamy Cer. 2.5 1 1.40 18.60 300 10 210 480 1.09 loamy Cer. 7.5 1 1.29 16.99 300 20 210 470 0.95 loamy Cer. 15 1 1.01 12.36 350 20 170 460 1.02 loamy Cer. 25 1 0.90 10.49 350 10 180 460 0.94 loamy Cer. 35 1 0.55 7.14 370 10 170 450 1.10 loamy Cer. 55 1 0.63 7.29 370 10 180 440 1.02 loamy Cer. 95 1 0.35 3.78 390 0 150 460 1.35 loamy Euc. 2.5 1 1.40 20.82 310 30 200 460 1.10 loamy Euc. 7.5 1 1.14 16.38 310 20 190 480 1.09 loamy Euc. 15 1 0.80 9.84 340 0 190 470 1.03 loamy Euc. 25 1 0.77 9.12 360 20 190 430 1.04 loamy Euc. 35 1 0.59 6.83 340 10 170 480 1.21 loamy Euc. 55 1 0.49 5.76 370 20 150 460 1.03 loamy Euc. 95 1 0.36 3.82 380 10 150 460 1.30 loamy Cer. 2.5 2 1.57 21.35 320 10 260 410 1.08 loamy Cer. 7.5 2 1.35 17.65 290 0 230 480 1.02 loamy Cer. 15 2 0.86 10.40 320 20 220 440 1.14 loamy Cer. 25 2 0.66 8.08 370 10 200 420 1.16

Continued 173 Table D.1 continued

Soil Land Depth Rep Total Total Fine Coarse Bulk use N C Clay Silt sand sand dens. (cm) ------g kg-1 soil------g cm-3 loamy Cer. 35 2 0.64 7.27 380 0 200 420 1.04 loamy Cer. 55 2 0.46 5.22 370 10 180 440 1.11 loamy Cer. 95 2 0.34 3.71 390 0 190 420 1.22 loamy Euc. 2.5 2 1.17 18.73 340 10 270 380 1.10 loamy Euc. 7.5 2 1.07 15.34 350 10 210 430 1.03 loamy Euc. 15 2 0.70 8.96 380 20 200 400 1.02 loamy Euc. 25 2 0.58 6.98 400 0 190 410 1.07 loamy Euc. 35 2 0.54 6.57 400 0 190 410 1.15 loamy Euc. 55 2 0.42 4.63 410 0 170 420 1.11 loamy Euc. 95 2 0.33 3.68 420 0 180 400 1.06 loamy Cer. 2.5 3 1.57 20.97 270 20 240 470 0.93 loamy Cer. 7.5 3 1.09 14.13 280 10 220 490 0.98 loamy Cer. 15 3 0.76 9.29 330 0 200 470 0.99 loamy Cer. 25 3 0.61 7.29 330 0 180 490 1.14 loamy Cer. 35 3 0.61 7.38 360 10 180 450 1.13 loamy Cer. 55 3 0.40 4.50 340 20 190 450 1.14 loamy Cer. 95 3 0.33 3.40 380 0 180 440 1.16 loamy Euc. 2.5 3 1.25 19.18 300 10 240 450 1.00 loamy Euc. 7.5 3 0.96 12.87 300 20 240 440 1.05 loamy Euc. 15 3 0.67 8.33 310 10 190 490 1.13 loamy Euc. 25 3 0.62 7.39 290 60 190 460 1.10 loamy Euc. 35 3 0.58 6.44 350 10 170 470 0.97 loamy Euc. 55 3 0.41 4.56 380 0 190 430 1.18 loamy Euc. 95 3 0.35 3.68 380 0 170 450 1.12 clayey Cer. 2.5 1 1.50 21.89 330 230 130 310 1.31 clayey Cer. 7.5 1 1.10 13.97 310 220 150 320 1.20 clayey Cer. 15 1 0.84 9.53 460 210 80 250 1.09 clayey Cer. 25 1 0.84 9.57 440 170 90 300 1.42 clayey Cer. 35 1 0.79 9.20 450 160 100 290 1.41 clayey Cer. 55 1 0.59 6.10 460 130 110 300 1.14 clayey Cer. 95 1 0.59 5.48 470 170 90 270 1.09 clayey Euc. 2.5 1 1.25 21.56 380 190 110 320 1.07

Continued 174 Table D.1 continued

Soil Land Depth Rep Total Total Fine Coarse Bulk use N C Clay Silt sand sand dens. (cm) ------g kg-1 soil------g cm-3 clayey Euc. 7.5 1 0.99 12.30 410 170 80 340 1.28 clayey Euc. 15 1 0.92 11.10 430 150 90 330 1.08 clayey Euc. 25 1 0.92 10.85 450 160 100 290 1.06 clayey Euc. 35 1 0.77 8.45 440 170 100 290 1.49 clayey Euc. 55 1 0.66 6.98 450 160 80 310 1.04 clayey Euc. 95 1 0.57 5.32 450 150 80 320 1.21 clayey Cer. 2.5 2 1.69 24.37 440 210 60 290 0.94 clayey Cer. 7.5 2 1.21 16.20 470 180 70 280 1.03 clayey Cer. 15 2 0.96 11.80 520 170 40 270 1.13 clayey Cer. 25 2 0.92 11.15 540 160 40 260 1.12 clayey Cer. 35 2 0.84 10.21 530 160 50 260 1.12 clayey Cer. 55 2 0.64 7.21 560 170 50 220 1.10 clayey Cer. 95 2 0.55 5.39 550 160 50 240 1.02 clayey Euc. 2.5 2 1.32 21.96 490 180 50 280 0.91 clayey Euc. 7.5 2 1.11 14.74 470 180 60 290 0.92 clayey Euc. 15 2 1.00 11.59 530 150 50 270 1.00 clayey Euc. 25 2 0.89 11.04 540 180 50 230 1.09 clayey Euc. 35 2 0.80 9.74 510 180 40 270 1.23 clayey Euc. 55 2 0.62 6.65 520 160 40 280 1.13 clayey Euc. 95 2 0.47 4.51 560 130 40 270 0.93 clayey Cer. 2.5 3 1.88 27.57 450 200 90 260 1.01 clayey Cer. 7.5 3 1.46 19.86 510 190 90 210 1.01 clayey Cer. 15 3 1.09 13.76 510 220 90 180 1.00 clayey Cer. 25 3 1.06 12.79 510 170 140 180 1.15 clayey Cer. 35 3 0.85 10.35 500 180 160 160 1.26 clayey Cer. 55 3 0.60 6.79 390 120 290 200 1.42 clayey Cer. 95 3 0.34 3.41 380 120 230 270 1.39 clayey Euc. 2.5 3 1.84 28.81 470 240 80 210 1.11 clayey Euc. 7.5 3 1.35 18.74 490 240 80 190 1.13 clayey Euc. 15 3 1.18 14.87 550 230 60 160 1.13 clayey Euc. 25 3 1.09 12.76 520 210 90 180 1.11 clayey Euc. 35 3 1.04 11.45 520 240 90 150 0.97 clayey Euc. 55 3 0.78 8.40 650 190 20 140 1.07 clayey Euc. 95 3 0.71 7.19 670 170 30 130 1.09

175

Land Sand Clay Sand Silt Clay Soil Depth use Rep. Sand Silt Clay SOC Silt SOC SOC CN CN C/N cm ---g fraction kg-1 soil--- --g SOC kg-1 fraction-- Sandy 2.5 Euc. 1 889.19 7.60 103.21 10.39 150.88 49.35 24.46 18.28 11.30 Sandy 2.5 Euc. 2 843.28 6.36 150.36 10.29 141.34 39.91 22.50 17.74 11.76 Sandy 2.5 Euc. 3 850.25 7.61 142.13 7.05 128.74 30.96 27.75 20.29 10.45 Sandy 2.5 Cer. 1 931.42 7.22 61.36 3.84 128.58 54.39 16.04 15.99 10.41 Sandy 2.5 Cer. 2 848.13 10.32 141.55 8.28 148.02 50.53 21.22 16.23 11.16 Sandy 2.5 Cer. 3 855.02 8.28 136.70 3.41 136.50 40.14 23.39 16.67 11.17 Loamy 2.5 Euc. 1 610.26 19.52 370.21 8.18 70.00 28.25 25.43 18.61 13.23 Loamy 2.5 Euc. 2 607.35 14.79 377.87 7.07 87.57 25.86 30.49 19.97 13.34 Loamy 2.5 Euc. 3 639.36 12.82 347.83 5.10 89.11 32.25 24.43 18.01 12.63 Loamy 2.5 Cer. 1 658.67 23.00 318.33 5.26 81.58 24.87 23.04 16.57 12.65 Loamy 2.5 Cer. 2 644.23 20.19 335.58 7.59 93.12 28.72 25.15 16.74 11.39 Loamy 2.5 Cer. 3 641.55 14.74 343.71 4.63 98.15 31.34 20.27 16.09 11.66 Clayey 2.5 Euc. 1 466.04 115.76 418.20 7.94 9.69 19.87 23.14 14.78 12.73 Clayey 2.5 Euc. 2 340.00 110.00 550.00 13.88 12.54 17.81 23.59 14.27 12.33 Clayey 2.5 Euc. 3 282.77 201.24 516.00 19.23 16.54 25.48 20.38 15.10 14.01 Clayey 2.5 Cer. 1 477.77 122.10 400.13 9.97 12.33 22.13 22.33 15.51 10.60 Clayey 2.5 Cer. 2 370.45 123.60 505.95 11.86 17.32 22.02 19.86 15.24 11.08 Clayey 2.5 Cer. 3 310.31 169.84 519.85 13.07 19.78 25.11 19.51 15.09 12.49 Sandy 35 Cer. 1 884.33 4.25 111.42 0.80 56.66 13.49 18.78 10.79 Sandy 35 Cer. 2 807.29 6.88 185.83 1.16 49.77 12.71 21.93 14.18 10.97 Sandy 35 Cer. 3 825.57 7.64 166.79 0.89 55.35 15.09 33.15 13.90 10.76 Loamy 35 Cer. 1 601.00 17.13 381.87 1.60 29.76 11.12 15.60 11.37 Loamy 35 Cer. 2 570.87 16.07 413.05 1.46 34.16 10.48 39.61 13.88 11.46 Loamy 35 Cer. 3 618.94 12.42 368.64 0.99 22.77 10.30 36.48 13.93 11.68 Clayey 35 Cer. 1 390.21 116.88 492.90 1.96 4.53 9.70 30.26 12.23 10.94 Clayey 35 Cer. 2 297.18 110.57 592.24 1.82 5.91 9.95 21.81 12.18 10.99 Clayey 35 Cer. 3 311.50 116.25 572.25 2.18 6.35 10.55 16.00 11.66 10.98 Sandy 95 Cer. 1 877.43 8.22 114.34 0.83 22.72 9.96 37.59 17.01 11.66 Sandy 95 Cer. 2 792.86 7.90 199.24 0.78 22.53 8.05 32.87 14.42 12.18 Sandy 95 Cer. 3 801.83 9.06 189.11 0.71 23.51 8.56 31.01 14.18 11.10 Loamy 95 Cer. 1 591.15 15.17 393.67 0.71 13.09 6.15 14.52 9.84 Loamy 95 Cer. 2 558.97 15.63 425.40 0.90 18.51 6.17 74.34 14.44 10.75 Loamy 95 Cer. 3 579.98 14.08 405.94 0.64 24.67 6.19 77.00 14.96 11.41 Clayey 95 Cer. 1 375.26 116.54 508.20 1.18 2.72 6.54 18.57 10.02 9.31 Clayey 95 Cer. 2 285.08 107.03 607.89 0.82 2.59 5.83 19.59 10.30 10.24 Clayey 95 Cer. 3 341.91 103.53 554.57 1.83 3.81 7.09 24.88 10.75 10.19

Table D.2. Data from particle-size fractionation, and SOC concentration and C/N ratio of obtained fractions. Depth refers to center of layer.

176

Soil Depth rep Soil<2 mm Clay Silt cm ------m2 g-1 ------Sandy 2.5 1 2.21 36.99 Sandy 35 1 4.1 40.77 Sandy 95 1 5.69 41.55 Sandy 2.5 2 5.35 38.87 Sandy 35 2 8.32 43.8 Sandy 95 2 10.89 44.44 Sandy 2.5 3 5.95 39.41 Sandy 35 3 7.71 45.04 Sandy 95 3 10.58 45.28 Loamy 2.5 1 11.78 42.24 Loamy 35 1 19.51 42.34 Loamy 95 1 20.24 44.13 Loamy 2.5 2 14.98 43.53 11.46 Loamy 35 2 19.65 42.29 10.17 Loamy 95 2 20.74 40.13 8.56 Loamy 2.5 3 15.09 40.12 Loamy 35 3 21.59 47.7 Loamy 95 3 19.91 43.63 Clayey 2.5 1 26.09 47.79 Clayey 35 1 31.7 50.29 Clayey 95 1 35.91 51.84 Clayey 2.5 2 29.11 45.11 8.96 Clayey 35 2 35.93 49.33 7.61 Clayey 95 2 41.1 51.51 6.37 Clayey 2.5 3 30.53 45.15 Clayey 35 3 36 48.72 Clayey 95 3 35.07 49.42

Table D.3. Specific surface area data of bulk soil, clay and silt fractions. Depth refers to center of layer.

177

Fe2O3 Al2O3 Fe2O3 Al2O3 Soil Depth rep CBD CBD NH4Ox. NH4Ox. Gibbsite Kaolinite Illite cm ------g mineral kg-1 clay ------Sandy 2.5 1 14.83 5.77 1.13 2.33 16.22 742.72 0.00 Sandy 2.5 2 47.39 22.28 1.20 2.19 7.36 694.83 0.00 Sandy 2.5 3 52.76 26.30 2.20 3.33 18.61 695.85 13.00 Sandy 35 1 17.67 3.89 1.17 2.90 10.81 784.85 0.00 Sandy 35 2 44.55 26.18 0.99 5.03 11.83 698.47 0.00 Sandy 35 3 35.48 13.80 1.61 5.02 14.86 671.24 13.87 Sandy 95 1 14.66 3.63 0.85 3.16 13.01 765.31 0.00 Sandy 95 2 38.89 11.29 0.71 5.37 15.60 733.76 0.00 Sandy 95 3 40.24 17.74 0.44 3.87 14.14 687.77 17.34 Loamy 2.5 1 68.29 12.90 2.88 8.06 51.94 612.35 38.30 Loamy 2.5 2 73.65 8.35 2.84 5.54 84.93 594.86 31.97 Loamy 2.5 3 71.25 6.47 2.58 3.91 59.48 619.93 30.20 Loamy 35 1 75.13 15.78 1.98 7.69 37.69 587.91 40.09 Loamy 35 2 76.23 12.02 1.58 6.70 60.06 637.50 26.49 Loamy 35 3 81.53 7.63 1.62 5.68 58.86 630.50 31.37 Loamy 95 1 74.03 12.93 1.15 6.30 44.45 625.97 37.83 Loamy 95 2 56.98 6.48 1.10 6.30 60.80 641.56 25.47 Loamy 95 3 84.23 6.72 2.92 5.72 53.22 639.17 28.29 Clayey 2.5 1 58.99 27.53 2.22 7.45 26.99 484.42 195.27 Clayey 2.5 2 66.12 29.14 1.99 7.54 61.84 467.00 149.35 Clayey 2.5 3 86.72 22.56 3.45 7.05 72.17 443.15 166.66 Clayey 35 1 60.02 29.53 1.19 7.55 25.07 355.49 200.04 Clayey 35 2 76.78 33.51 1.54 9.36 56.57 484.37 131.91 Clayey 35 3 93.72 25.78 2.04 7.90 54.67 489.68 164.44 Clayey 95 1 57.69 30.70 0.69 8.28 24.21 531.10 198.77 Clayey 95 2 75.64 32.70 0.97 8.95 60.27 485.94 149.57 Clayey 95 3 61.45 15.14 1.53 10.35 62.25 503.49 160.71

Table D.4. Data from quantitative clay mineralogy. Depth refers to center of layer.

178

Sand Land WSA WSA Total WSA Total WSA in WSA sand WSA sand Soil use Rep size dist. C/N SOC WSA C/N SOC % g kg-1 % g kg-1 Sandy Cer. 1 5 5.8 15.91 17.63 78.76 22.65 4.49 Sandy Cer. 1 1.5 3.5 14.60 26.40 76.93 18.57 8.47 Sandy Cer. 1 0.75 28.4 15.54 5.56 94.08 31.06 1.06 Sandy Cer. 1 0.38 38 13.70 3.84 95.81 38.04 0.53 Sandy Cer. 1 0.18 24.3 12.81 3.63 50.11 62.95 0.47 Sandy Euc. 1 5 8.5 19.27 21.35 68.60 30.81 4.29 Sandy Euc. 1 1.5 6.3 18.28 35.02 66.62 27.24 7.53 Sandy Euc. 1 0.75 30 18.52 5.47 91.14 98.04 1.20 Sandy Euc. 1 0.38 34.9 15.54 4.18 92.66 223.25 0.49 Sandy Euc. 1 0.18 20.3 15.00 5.28 91.53 124.61 0.49 Sandy Cer. 2 5 21.9 17.38 22.76 93.29 21.36 3.56 Sandy Cer. 2 1.5 10 14.96 22.61 75.15 23.31 2.44 Sandy Cer. 2 0.75 26.2 15.47 9.13 86.11 22.46 2.26 Sandy Cer. 2 0.38 24 14.42 7.60 87.96 30.18 1.34 Sandy Cer. 2 0.18 17.9 14.16 7.87 87.84 18.52 0.71 Sandy Euc. 2 5 7.8 18.00 16.32 69.47 29.43 3.66 Sandy Euc. 2 1.5 5.7 16.01 23.73 66.77 36.13 3.63 Sandy Euc. 2 0.75 26.8 16.00 7.35 85.78 51.82 1.14 Sandy Euc. 2 0.38 33 15.16 5.66 88.18 27.11 0.53 Sandy Euc. 2 0.18 26.7 16.46 8.77 85.79 67.14 0.60 Sandy Cer. 3 5 17 16.75 13.50 60.45 25.26 2.41 Sandy Cer. 3 1.5 6.2 14.48 17.47 76.42 24.31 2.59 Sandy Cer. 3 0.75 23.7 15.16 6.91 88.22 23.18 1.46 Sandy Cer. 3 0.38 31.5 13.89 5.29 90.51 49.67 0.66 Sandy Cer. 3 0.18 21.6 13.67 7.29 86.85 53.44 0.93 Sandy Euc. 3 5 10 17.22 15.61 71.69 34.39 2.00 Sandy Euc. 3 1.5 6.3 16.49 20.00 70.25 28.33 2.21 Sandy Euc. 3 0.75 25.3 15.65 6.27 85.38 50.89 0.96 Sandy Euc. 3 0.38 36.7 14.56 4.51 89.69 142.05 0.31 Sandy Euc. 3 0.18 21.7 15.04 5.22 89.42 80.04 0.63 Loamy Cer. 1 5 25.4 15.09 16.74 57.58 25.70 3.85 Loamy Cer. 1 1.5 11.4 13.49 19.43 55.50 20.03 4.38 Loamy Cer. 1 0.75 19.8 13.76 14.89 62.41 27.54 2.69 Loamy Cer. 1 0.38 19.8 13.51 8.88 74.41 34.52 1.65 Loamy Cer. 1 0.18 23.6 13.56 8.69 77.07 122.52 1.02 Loamy Euc. 1 5 19.9 16.40 16.54 53.95 24.09 3.32 Loamy Euc. 1 1.5 7.9 15.63 18.55 52.17 26.31 3.59 Loamy Euc. 1 0.75 15.5 15.53 11.03 63.16 27.00 1.86 Loamy Euc. 1 0.38 22 15.42 6.26 74.62 36.99 1.06 Loamy Euc. 1 0.18 34.7 14.99 8.66 69.14 42.87 1.22 Continued

Table D.5. Quantitative, SOC and C/N data of total and sand (>20 μm) fraction of water- stable aggregates (WSA) for depth 0-5 cm. 179 Table D.5 continued

Land WSA WSA Total WSA Total WSA Sand in WSA sand WSA sand Soil use Rep size dist. C/N SOC WSA C/N SOC % g kg-1 % g kg-1 Loamy Cer. 2 5 33.5 15.42 18.66 54.26 28.02 4.35 Loamy Cer. 2 1.5 10.9 14.09 20.88 53.02 25.38 5.20 Loamy Cer. 2 0.75 17.7 14.30 15.27 66.08 27.22 2.47 Loamy Cer. 2 0.38 17.5 14.34 9.88 75.51 22.87 1.73 Loamy Cer. 2 0.18 20.4 13.52 7.95 76.81 43.90 1.23 Loamy Euc. 2 5 14 16.63 13.41 51.24 30.28 2.65 Loamy Euc. 2 1.5 7.4 14.76 16.36 48.42 33.62 2.87 Loamy Euc. 2 0.75 24 15.53 13.25 58.22 23.96 1.85 Loamy Euc. 2 0.38 24.5 15.79 9.07 73.70 35.04 1.43 Loamy Euc. 2 0.18 34.2 15.26 7.87 73.20 69.40 0.71 Loamy Cer. 3 5 45.2 17.22 15.63 57.17 27.11 2.97 Loamy Cer. 3 1.5 11.7 13.93 20.53 55.68 22.22 3.19 Loamy Cer. 3 0.75 16.9 14.26 15.92 66.51 20.50 3.10 Loamy Cer. 3 0.38 12.5 14.11 10.68 75.71 32.12 1.40 Loamy Cer. 3 0.18 13.7 13.44 8.36 75.84 52.93 1.10 Loamy Euc. 3 5 24.7 16.10 16.24 56.80 28.20 2.26 Loamy Euc. 3 1.5 8 15.00 17.89 52.95 28.32 3.01 Loamy Euc. 3 0.75 19.8 15.65 12.67 63.15 32.11 2.16 Loamy Euc. 3 0.38 21.5 15.67 9.20 75.44 52.60 1.12 Loamy Euc. 3 0.18 26 14.90 7.90 74.70 26.76 0.90 Clayey Cer. 1 5 42.4 15.38 20.23 33.92 27.93 6.78 Clayey Cer. 1 1.5 16.6 14.26 18.52 33.42 25.29 7.45 Clayey Cer. 1 0.75 16.6 14.29 19.02 32.91 26.29 7.16 Clayey Cer. 1 0.38 7 14.42 19.10 36.74 26.18 6.46 Clayey Cer. 1 0.18 17.4 14.03 16.07 43.61 33.37 3.69 Clayey Euc. 1 5 36.6 14.33 13.05 30.97 38.45 4.36 Clayey Euc. 1 1.5 16.4 14.41 13.74 31.51 34.75 3.43 Clayey Euc. 1 0.75 18 13.27 10.77 14.47 20.03 2.69 Clayey Euc. 1 0.38 9.41 13.52 9.83 34.53 27.31 2.54 Clayey Euc. 1 0.18 19.5 14.90 9.66 44.17 33.74 1.79 Clayey Cer. 2 5 40.2 14.87 22.56 26.31 25.74 8.41 Clayey Cer. 2 1.5 16 15.20 21.58 33.00 23.49 11.12 Clayey Cer. 2 0.75 17.2 14.52 20.43 29.68 24.43 9.04 Clayey Cer. 2 0.38 8.95 14.50 18.99 30.64 26.95 6.71 Clayey Cer. 2 0.18 17.6 14.67 16.32 38.05 32.52 4.51 Clayey Euc. 2 5 30.8 14.00 12.75 25.07 19.95 3.31 Clayey Euc. 2 1.5 18 13.72 13.18 24.93 25.99 4.34 Clayey Euc. 2 0.75 23.5 13.65 12.69 26.68 26.50 4.16 Clayey Euc. 2 0.38 9.75 13.93 12.58 28.76 28.13 3.67 Clayey Euc. 2 0.18 17.9 14.62 13.38 29.96 33.89 3.09

Continued

180 Table D.5 continued

Sand Land WSA WSA Total WSA Total WSA in WSA sand WSA sand Soil use Rep size dist. C/N SOC WSA C/N SOC % g kg-1 % g kg-1 Clayey Cer. 3 5 52.3 14.88 25.60 20.93 24.50 10.24 Clayey Cer. 3 1.5 18.1 14.86 24.64 19.85 25.78 10.83 Clayey Cer. 3 0.75 12.8 15.40 24.27 23.06 24.16 13.39 Clayey Cer. 3 0.38 4.41 15.36 23.79 29.63 25.83 6.00 Clayey Cer. 3 0.18 12.3 15.98 20.03 43.13 24.93 3.58 Clayey Euc. 3 5 20.4 15.56 25.19 16.91 28.48 8.55 Clayey Euc. 3 1.5 16 14.75 20.37 19.32 26.31 14.57 Clayey Euc. 3 0.75 21 14.91 19.10 20.36 25.75 12.99 Clayey Euc. 3 0.38 16 15.13 19.73 23.53 27.62 10.03 Clayey Euc. 3 0.18 26.6 15.26 19.44 29.62 30.00 6.73

WSA WSA Total WSA Total WSA Sand in WSA sand WSA sand Soil Depth Rep size dist. C/N SOC WSA C/N SOC % g kg-1 % g kg-1 Sandy 35 1 5 0.45 22.00 4.41 80.92 122.60 0.61 Sandy 35 1 1.5 2.45 16.65 3.45 86.85 33.18 0.31 Sandy 35 1 0.75 25.70 16.15 1.14 95.90 6.09 0.14 Sandy 35 1 0.375 47.55 14.02 1.42 93.49 24.45 0.20 Sandy 35 1 0.178 23.85 14.50 2.93 85.40 20.32 0.17 Sandy 95 1 5 0.35 16.40 5.76 76.16 75.16 0.57 Sandy 95 1 1.5 1.90 15.95 2.21 88.80 21.04 0.28 Sandy 95 1 0.75 20.50 27.31 0.60 96.35 14.97 0.09 Sandy 95 1 0.375 42.30 16.20 0.71 94.19 13.21 0.07 Sandy 95 1 0.178 34.95 14.72 2.33 84.29 23.86 0.14 Sandy 35 2 5 1.25 16.33 9.83 61.12 32.30 1.66 Sandy 35 2 1.5 2.10 13.38 7.07 64.78 74.56 0.54 Sandy 35 2 0.75 19.10 16.05 1.62 91.00 48.88 0.17 Sandy 35 2 0.375 39.90 13.60 2.47 87.87 31.88 0.12 Sandy 35 2 0.178 37.65 13.54 4.10 79.35 35.26 0.15 Sandy 95 2 5 0.30 16.62 6.17 58.51 111.17 0.63 Sandy 95 2 1.5 1.55 14.80 3.02 71.67 109.17 0.40 Sandy 95 2 0.75 16.25 19.43 0.80 94.06 38.35 0.11 Sandy 95 2 0.375 35.60 16.10 1.16 91.31 25.29 0.12 Sandy 95 2 0.178 46.30 14.27 2.76 77.38 7.12 0.13

Continued

Table D.6. Quantitative, SOC and C/N data of total and sand (>20 μm) fraction of water- stable aggregates (WSA).

181 Table D.6 continued

WSA WSA Total WSA Total WSA Sand in WSA sand WSA sand Soil Depth Rep size dist. C/N SOC WSA C/N SOC % g kg-1 % g kg-1 Sandy 35 3 5 1.20 15.11 6.82 65.89 44.35 1.82 Sandy 35 3 1.5 1.90 12.55 6.88 71.57 114.32 0.70 Sandy 35 3 0.75 16.85 15.33 1.77 93.00 64.04 0.21 Sandy 35 3 0.375 45.85 14.46 1.86 91.50 133.85 0.22 Sandy 35 3 0.178 34.20 12.96 5.15 78.75 83.56 0.31 Sandy 95 3 5 0.35 14.65 7.05 62.47 95.40 0.61 Sandy 95 3 1.5 1.40 13.63 4.47 68.90 146.39 0.55 Sandy 95 3 0.75 13.90 21.16 0.98 93.96 9.69 0.15 Sandy 95 3 0.375 36.20 15.63 1.27 92.14 62.38 0.14 Sandy 95 3 0.178 48.15 13.68 2.74 77.76 112.72 0.29 Loamy 35 1 5 4.55 14.62 8.82 53.59 177.45 1.72 Loamy 35 1 1.5 5.15 12.81 7.24 52.67 64.00 1.36 Loamy 35 1 0.75 14.15 13.51 4.79 66.98 136.41 0.60 Loamy 35 1 0.375 25.55 13.39 4.01 73.55 95.35 0.39 Loamy 35 1 0.178 50.60 12.71 4.89 63.70 101.34 0.52 Loamy 95 1 5 2.75 14.70 3.99 52.17 148.14 0.59 Loamy 95 1 1.5 2.75 13.03 3.99 52.49 113.80 0.71 Loamy 95 1 0.75 9.65 14.37 2.14 74.42 89.20 0.25 Loamy 95 1 0.375 23.80 18.67 1.83 74.68 94.73 0.27 Loamy 95 1 0.178 61.05 12.25 2.61 63.50 118.17 0.32 Loamy 35 2 5 4.50 14.67 8.12 49.41 48.50 1.33 Loamy 35 2 1.5 4.55 12.97 8.26 48.79 76.46 1.06 Loamy 35 2 0.75 15.20 13.57 4.88 65.99 101.94 0.47 Loamy 35 2 0.375 27.80 13.24 4.19 72.10 18.71 0.29 Loamy 35 2 0.178 47.95 12.88 5.61 61.03 149.84 0.38 Loamy 95 2 5 2.30 15.66 5.04 47.45 191.45 0.56 Loamy 95 2 1.5 2.85 13.79 4.54 48.34 86.49 0.65 Loamy 95 2 0.75 11.40 15.27 2.40 74.22 18.21 0.29 Loamy 95 2 0.375 26.25 14.54 2.24 74.98 86.59 0.18 Loamy 95 2 0.178 57.20 12.88 3.19 58.28 141.36 0.27 Loamy 35 3 5 4.75 15.02 9.22 54.80 111.22 0.99 Loamy 35 3 1.5 4.90 13.04 7.59 52.38 93.05 0.90 Loamy 35 3 0.75 14.90 13.80 4.43 69.63 189.16 0.37 Loamy 35 3 0.375 26.15 13.90 3.79 72.75 262.36 0.42 Loamy 35 3 0.178 49.30 13.01 5.15 62.85 237.27 0.43 Loamy 95 3 5 2.35 15.56 4.23 51.75 45.84 0.59 Loamy 95 3 1.5 2.80 13.97 4.18 52.31 380.19 0.62 Loamy 95 3 0.75 10.55 15.46 1.85 75.35 73.31 0.24 Loamy 95 3 0.375 24.40 14.12 2.12 74.93 55.20 0.15 Loamy 95 3 0.178 59.90 12.42 2.48 62.83 98.08 0.27

Continued 182 Table D.6 continued

WSA WSA Total WSA Total WSA Sand in WSA sand WSA sand Soil Depth Rep size dist. C/N SOC WSA C/N SOC % g kg-1 % g kg-1 Clayey 35 1 5 5.69 13.04 9.12 26.90 69.30 1.66 Clayey 35 1 1.5 12.26 11.97 8.66 26.08 24.30 1.70 Clayey 35 1 0.75 25.69 11.91 8.06 29.31 22.81 2.05 Clayey 35 1 0.375 24.57 11.81 8.02 27.53 36.35 1.58 Clayey 35 1 0.178 31.79 11.74 7.81 30.35 66.10 0.97 Clayey 95 1 5 4.24 10.16 6.48 26.23 219.09 1.39 Clayey 95 1 1.5 6.73 11.47 5.40 27.74 66.01 1.28 Clayey 95 1 0.75 19.13 11.06 5.34 28.94 34.74 1.38 Clayey 95 1 0.375 23.17 10.66 5.12 24.73 31.34 0.96 Clayey 95 1 0.178 46.75 10.37 4.86 25.37 96.72 0.63 Clayey 35 2 5 10.43 13.41 9.64 22.13 49.41 2.36 Clayey 35 2 1.5 12.91 12.48 9.25 21.71 40.07 2.03 Clayey 35 2 0.75 23.34 12.55 9.09 23.69 27.26 1.93 Clayey 35 2 0.375 20.86 12.46 9.16 23.49 32.79 1.63 Clayey 35 2 0.178 32.45 12.35 8.95 24.60 50.35 1.15 Clayey 95 2 5 6.78 8.02 5.03 22.30 70.44 1.06 Clayey 95 2 1.5 9.59 12.75 4.90 21.87 117.90 1.18 Clayey 95 2 0.75 19.98 11.81 4.60 23.43 61.22 0.94 Clayey 95 2 0.375 19.14 11.46 4.67 21.82 39.64 0.86 Clayey 95 2 0.178 44.51 10.94 4.42 22.60 70.58 0.66 Clayey 35 3 5 7.64 12.51 11.27 14.22 42.15 4.00 Clayey 35 3 1.5 12.00 12.36 11.04 16.71 23.32 4.43 Clayey 35 3 0.75 22.76 12.50 10.20 23.77 19.96 3.59 Clayey 35 3 0.375 21.83 12.22 10.28 20.25 22.37 3.02 Clayey 35 3 0.178 35.77 12.25 10.11 21.40 235.00 2.35 Clayey 95 3 5 4.40 7.99 7.44 18.35 49.25 2.37 Clayey 95 3 1.5 8.71 12.19 5.20 37.00 21.26 2.21 Clayey 95 3 0.75 19.78 11.66 4.82 41.72 28.40 1.72 Clayey 95 3 0.375 21.27 11.11 5.66 25.02 26.36 1.55 Clayey 95 3 0.178 45.86 10.82 5.41 26.35 33.16 1.40

183