Distribution of inorganic phosphorus in profiles and particle ‐ size fractions across 1 an established riparian buffer and adjacent cropped area at the Dian lake 2

an established riparian buffer and adjacent cropped area at the Dian lake 2 G. S. Zhang∗ and J. C. Li 3 Environmental Science and Ecological Rehabilitation Institute of Yunnan University, 4 Kunming 650091,China 5 6 Abstract 7 Riparian buffer can trap sediment and nutrients sourced from upper cropland and 8 minimizing eutrophication risk of water quality. This study aimed to investigate the 9 distributions of soil inorganic phosphorus (Pi) forms among profile and particle-size 10 fractions in an established riparian buffer and adjacent cropped area at the Dian lake, 11 Southwestern China. The Ca-bound fraction (62%) was the major proportion of the Pi in 12 the riparian soils. Buffer rehabilitation from cropped area had a limited impact on total 13 phosphorus (TP) concentrations after 3 years, but has contributed to a change in Pi forms. 14 At 0-20 cm soil layer, levels of the Olsen-P, nonoccluded, Ca-bound and total Pi were 15 lower in the buffer than the cropped area; however, the Pi distribution between the 16 cropped area and the buffer did not differ significantly as depth increased. The clay 17 fraction corresponded to 57% of TP and seemed to be both a sink for highly recalcitrant 18 Pi and a source for labile Pi. The lower concentration of Pi forms in the silt and sand 19 particle fraction in the surface soil was observed in the buffer area, which indicating that 20 the Pi distribution in coarse particle fraction has sensitively responded to land-use 21 changes. 22


Introduction 26
Eutrophication of surface water has been linked to runoff of excess nutrients from 27 agricultural soils in many parts of the world. The improper management or use of 28 phosphorus (P) fertilizer during cultivation enhances the P transport affecting the water 29 quality, leading to alterations in water ecosystems (Sharpley et al. 2003;Troitiño et al. 30 2008). Riparian buffer is an efficient and economical tool to reduce agricultural non-point 31 source pollution (Correll, 2000). Because of the filtering function of soil, the effectiveness 32 of riparian buffers in reducing sediment and nutrient loading in surface and subsurface 33 flows from cropland has been shown across many geographic regions (Dosskey et al.,34 2010; Keesstra et al. 2012). 35 In the basin of the Dian lake, Southwestern China, agriculture development from 36 conventional field crop to intensive horticultural crop has occurred for more than 20 37 years. Due to severe eutrophication of the Dian lake, a conservation program of Dian lake 38 was established in 2009. Since its inception, thousands of hectare of buffers have been 39 established by converting strips of cropland around the lake riparian zones to permanent 40 vegetation. It is critical to know how the change of land-use affects soil P and its various 41 forms which in turn affect its potential as a pollutant. 42 Much experimental evidence from research on both arable and buffer soils indicates that 43 P are transported from soil to water with eroded soil (Abrams and Jarrell, 1995). 44 Minimizing the risk of P enrichment of surface water bodies with P transported from 45 agricultural soils will require specific attention to forms of P in soils. Mooer and Reddy 46 the organic forms of P. P distribution in chemical fractions can vary among land uses. 48 The clay fraction (< 2 μm) was freeze-dried after separating it from the fine sand and the 106 silt by repeated centrifugation until the supernatant was clear. The fine sand (250-50 μm) 107 was obtained by passing through a 50 μm sieve, and dried at 40 °C, whereas the 108 remaining silt (2-50 μm) in the remaining suspension was freeze-dried. Due to the low 109 recovery of sand fractions, it was necessary to combine coarse-sand and fine-sand 110 fractions to give one sample for chemical analyses. 111 Total soil P was determined using wet oxidation (Shi, 1998). Olsen P was determined by 112 adding 20 ml of 0.5 M NaHCO , pH 8.5 extractant to 1.00 g of soil, shaking for 30 min, 113 and filtering through Whatman 42 filter paper (Kuo, 1996). Inorganic P fractions were 114 conducted by Kuo fractionation schemes (Zhang and Kovar, 2000). A 0.5 g soil sample 115 was placed in a 50 ml centrifuge tube and sequentially extracted with 25 ml each of 1.0 116 order. Each extraction was performed for 0.1, 1, and 17 h using a horizontal shaker 118 followed by centrifugation at 3300 rpm for 15 min, respectively. The occluded Pi was 119 subsequently extracted by adding 20 ml of 0.3 M Na3C6H5O7 , 2.5 ml of 1 M NaHCO3 , 120 and 0.5 g Na2S2O4 to the residue in each tube and heating for 15 min at 85°C, followed 121 by centrifugation at 3300 rpm for 15 min. The calcium-bound Pi was subsequently 122 extracted by adding 25 ml of 0.25 M H2SO4 followed by centrifugation at 3300 rpm for 123 15 min. All the measurements were triplicated. 124

Statistical analysis 125
All data was submitted to analysis of variance (ANOVA) and treatment means were crop and the buffer soils were less significant as depth increased at the HG and XL sites 150 but at the NL sites. The organic-rich subsurface soil in the buffer area at the NL sites may 151 been due to the buried river channel sediment or peat. 152 The higher pH of surface soil (0-20 cm) in the cropped area reflect the regular fertilization 153 increasing salt concentration in the soil solution (Godsey et al, 2007). In addition, the pH 154 of the NL soils was 0.2 to 0.5 higher than the pH of the XL and HG soils. Compared as 155 the cropped soils, the lower pH of subsurface buffer soils in the NL sites may be partly 156 explained as the higher soil organic matter which can be a potential source of soil acidity 157 (Coleman and Thomas, 1967). Soil particle density (ρd) of the NL sites was significantly 158 lower than that of the XL and HG sites. 159

Land-use effects on P fractions in whole soil samples 160
Fertilizer application has a significant effect on increasing the available P forms in the 161 cropped soils (Figure 4). Comparison of Olsen-P levels in the crop and buffer soils 162 indicated that higher amount of available P (99.3 mg/kg) was accumulated in topsoil (0-163 20 cm) in crop areas. The higher extractable P in the cropped soil implicated that it would 164 be more easily depleted by movement from the soil surface by erosion and leaching 165 (Sharpley et al. 2003). Also, non-occluded, Ca-bound and total inorganic P (Pi) levels at 166 0-20 cm layer were higher in the cropped soil than in the buffer soil. This may be due to 167 the regular fertilization in surface soil for crop growth. Sharply and Smith (1985)  Additionally, the percentage of loosely-bound, non-occluded Pi also decreased as 183 sampling depth increased except occluded and Ca-bound Pi (Figure 4). 184 Distribution of total Pi in the soils was, on the average, 5% loosely-bound, 9% non-185 occluded, 24% occluded and 62% Ca-bound. High levels of Ca-P in these soils suggested 186 that at least some of the soil's alluvial parent material probably originated from the 187 limestone bedrock in the area above the catchment of the Dian lake (Schroeder and Kovar, 188 2006). Except for the occluded Pi in 0-20cm layer, the percentages of Pi fractions did not 189 differ significantly between the cropped and the buffer soils. Compared with the cropped 190 area topsoils (0-20 cm), the higher percentages of occluded Pi was found in the buffer 191 topsoils (Table 1). In general, this observation suggests the increasing possibility of 192 occlusion of P with Fe/Al hydrous oxides or formation of insoluble Al/Fe phosphates in 193 buffer soils relative to cropped soils. 194 Although subsoil Ca-P (below 0-20 cm) alone did not differ among those areas, the others 195 Pi fractions were greater in the XL soils than in the HG and NL soils (Figure 4). The NL 196 soil had the lowest loosely-bound, non-occluded, occluded and total Pi (Figure 4). 197 However, total P concentration did not differ significantly among those areas. Soil TP 198 concentration was positively correlated with clay content (r=0.508, n=30, P<0.01). 199 However, there was not a significant relationship between Pi fractions and clay content. 200 Occluded Pi was negatively correlated with soil pH (r=-0.621, n=30, P<0.01). 201 Furthermore, there were strong correlations among the P fractions. 202

Land-use effects on inorganic phosphorus fractions in particle-size fractions 203
Although the error of individual fractions was up to 12%, the sum of all Pi fractions in 204 the particle-size fractions was similar to Pi in the whole soil (Table 2). In these soils, the 205 proportion of P froms decreased continuously with increasing particle size of the 206 fractions. The high proportion of P froms in the clay fraction reflected that the clay 207 fraction not only seemed to be a sink for highly recalcitrant Pi but also a source for labile 208 Pi forms (Neufeldt et al., 2000;Suñer and Galantini, 2015). The concentrations of Pi 209 forms in the sand fraction were both significantly lower than in the clay and silt fraction, 210 whereas the concentrations of Pi forms did not differ significantly between the clay and 211 silt fraction. The higher concentration of TP in the clay fraction could be attributed to the 212 enrichment of organic phosphorus in the finer particle size class. Christensen (2001) reported that in arable soils, clay-sized complexes (< 2μm) have the largest concentrations 214 of OM, silt-sized (2-20μm) particles are less enriched, and size separates > 20μm usually 215 contain little OM. 216 Figure 5 shows the complex distribution of surface soil (0-20 cm) Pi fractions in the 217 particle-size fractions under different land-use types. Buffer rehabilitation had no effect 218 on the concentration of Pi fractions within clay fraction, but the coarse particle fraction 219 exhibited an decrease in the concentration of loosely-bound, nonoccluded, occluded (not 220 in the silt fraction) and Ca-bound Pi. Chen et al. (2015) also reported that topsoil nutrients 221 stored in coarse particle fractions were more sensitive than those stored in the fine fraction 222 to soil recovery. However, Suñer et al. (2014) reported that the coarse fraction of the 223 cultivated field had low levels of Pi as a consequence of the particulate organic matter 224 decomposition and coarse mineral particle weathering. The reduction of Pi in coarse 225 particle fractions in the buffer soil could have a positive influence on the function of the 226 buffer to act as a sink for P. In the cropped soil, however, the accumulation of labile Pi in 227 coarse particle fractions was considered to be more susceptible to lose P via leaching. 228 Zheng et al. (2003) reported that soil particles containing high amounts of extractable P 229 suggested to have a higher P release potential. The higher nonoccluded and occluded Pi 230 in the sand and silt fractions of the cropped soil indicated that the dissoluted fertilizer P 231 was easy transformed into more stable Pi forms (Neufeldt et al., 2000).The significant 232 accumulation of Ca-bound Pi in the sand and silt fractions of the cropped soils as 233 compared to the buffer soils could be explained by the presence of undissolved P fertilizer 234 granules in these fractions, because of rock phosphate had been often used together with 235 soluble P fertilizers in these areas. Tiessen et al. (1983) also observed a similar 236 enrichment of recalcitrant P in the coarse silt fraction of a cropped soil. Below 20 cm, the 237 concentrations of Pi fractions in the particle-size fractions did not differ significantly 238 between the crop and buffer soil. 239 To enable a better understanding of P transformations in those riparian soils, regardless 240 of land uses, the average proportions of the P fractions in the particle size fractions are 241 presented in Figure 6. Compared with the sand and silt fractions, the proportions of 242 occluded Pi was increased in the clay fraction, which should be related to the increasing 243 adsorption to Fe/Al hydrous oxides with finer particle size classes. Agbenin and Tiessen 244 (1995) and Neufeldt et al. (2000) also reported a similar change of the proportions in 245 entisols, inceptisols and oxisols from Brazil. The proportions of Ca-bound Pi in the clay 246 and silt sand fraction were significantly lower than that in the sand fractions, which 247 suggested that the increase in occluded Pi with finer particle size fractions may be at the 248 expense of Ca-bound Pi. The occluded Pi, which consisted of insoluble Al/Fe phosphates, 249 are progressively sequestrated in finer particle size fractions and therefore more difficult 250 to extract. 251 The proportion of nonoccluded Pi was nearly twice as high in the silt and clay fractions 252 as compared to the sand fractions, which could be attributed to their comparatively higher 253 amounts of discrete Fe/Al hydrous oxides. However, Neufeldt et al. (2000) reported that 254 the proportions of NaOH-Pi showed no consistent trends between the particle-size 255 classes. In contrast, the proportions of loosely-bound Pi did not differ significantly 256 between the particle-size classes, which suggested the potential of labile Pi lose would 257 not decrease with decreasing the particle-size fractions. 258

Conclusion 259
Inorganic phosphorus forms and their distribution in particle-size fractions were different 260 between the conservation buffer and the continuously cropped area. Amounts of Pi increased in the surface soil of crop field associated with fertilizer P application, with 262 nonoccluded and Ca-bound Pi constituting the major proportion of the change. Lower 263 concentration of Pi forms in the silt and sand particle fraction was found in the surface 264 soil of the buffer, suggesting that P in these fractions had a lower release potential. The 265 enrichment of labile Pi in the coarse particle fraction of cropped soil implicated that it 266 would be more easily depleted by movement from the soil surface by leaching. More 267 important, the application of particle-size separation of P forms determination can be 268 represented to a better understanding of soil P distribution between different land uses.   For each phosphorus form, data followed by the same capital case letter indicate that the P concentrations between particle size fractions were