Root-induced changes in rhizosphere pH and their effect on micronutrient availability in calcareous soils

Introduction

About 1.5 billion hectares of agricultural land worldwide contain free calcium carbonate, buffering soil pH above 7.5 and sharply reducing the solubility of iron, manganese, zinc, and copper [1,2] . On such soils, total micronutrient content may be adequate, yet plant-available fractions remain below critical thresholds because high pH drives precipitation and adsorption reactions [3] . Plant roots actively modify the chemistry of the narrow soil zone they inhabit. Proton release during cation uptake, organic acid exudation, and CO₂ from root respiration can lower rhizosphere pH by 0.5-2.0 units relative to bulk soil [4,5] . Legumes, which assimilate atmospheric N₂ and take up more cations than anions, tend to produce stronger acidification than cereals [6] . This localised pH drop increases the dissolution of carbonate-bound and oxide-bound micronutrients, raising their availability at the root surface [7] . Despite a considerable body of work in temperate calcareous soils, few controlled experiments have compared rhizosphere acidification intensity among tropical crops grown on the same calcareous material. This research used a rhizobox approach to quantify root-induced pH gradients and their consequences for DTPA-extractable Fe, Mn, Zn, and Cu across five crop species in a Thai calcareous Vertisol.

Research Area Description

The experiment was conducted in the net house and soil chemistry laboratory at the Faculty of Agriculture, Kasetsart University, Bangkhen Campus, Bangkok (13°50′N, 100°34′E). The calcareous soil was collected from Saraburi Province, approximately 110 km northeast of Bangkok, a region where limestone-derived Vertisols are common under sugarcane and upland crop cultivation.

Materials and Methods

Materials

Bulk soil (0-30 cm) was air-dried, sieved to 2 mm, and packed into rhizobox compartments.

Five crops were tested: rice (KDML 105), maize (Suwan 5), soybean (Chiang Mai 60), sunflower (Pacific 77), and mung bean (Kamphaeng Saen 2). Seeds were surface-sterilised in 1% NaOCl for 10 min, rinsed, and pre-germinated for 48 h before transplanting. Each rhizobox (20 × 15 × 25 cm) had a root compartment separated from soil compartments by 30 µm nylon mesh, permitting water and solute exchange but restricting root penetration [8] . Plants received a basal solution of 40 mg N, 20 mg P, and 20 mg K per kg soil; no micronutrient fertiliser was added.

Methods

The trial followed a completely randomised design with five crop treatments and four replications, conducted between January and March 2024. After 45 days of growth, soil was sampled at four distances from the root surface: 0-2, 2-4, 4-6, and 6-10 mm, plus bulk soil (>20 mm). Soil pH was measured in 1:2.5 soil-water suspension. DTPA-extractable Fe, Mn, Zn, and Cu were determined following the method of Lindsay and Norvell [9] . Organic acids in rhizosphere soil solution were analysed by HPLC [10] . Data were subjected to ANOVA and means separated by Duncan's multiple range test at p ≤ 0.05.

Soil Quality Analysis

The Saraburi Vertisol had pH 7.83, EC 0.38 dS m⁻¹, organic matter 1.92%, free CaCO₃ 12.4%, CEC 34.7 cmol kg⁻¹, and clay content 52.3%. Initial DTPA-extractable concentrations were Fe 3.87, Mn 6.21, Zn 0.62, and Cu 1.14 mg kg⁻¹-Fe and Zn both below established critical levels of 4.5 and 0.8 mg kg⁻¹ respectively [9] . All five crops significantly lowered rhizosphere pH compared with bulk soil (Table 1). Mung bean produced the strongest acidification (pH 6.42, a drop of 1.41 units), followed by soybean (6.53). DTPA-extractable Zn in the mung bean rhizosphere (1.09 mg kg⁻¹) was 76% higher than bulk soil, while Fe rose by 116%. Legumes exuded substantially more organic acids than cereals (Table 2). Mung bean had the highest total organic acid concentration (8.72 µmol g⁻¹), dominated by citric acid (3.89 µmol g⁻¹). Citric acid was the main organic acid across all species.

Results

The relative share of Zn among extracted micronutrients increased from 19.8% in bulk soil to 28.9% in the rhizosphere (Figure 1), indicating that Zn availability responded more strongly to pH reduction than Fe or Mn on a proportional basis. The pH gradient was steepest within the first 2 mm and flattened beyond 6 mm (Figure 2). Mung bean and soybean maintained acidified zones extending further from the root surface than rice or maize.

Comprehensive Interpretation

Pearson correlation between rhizosphere pH (0-2 mm) and DTPA-Zn yielded r = -0.91 (p < 0.01), confirming the strong inverse relationship. Total organic acid concentration correlated with the pH drop (r = 0.87, p < 0.01), supporting organic acid exudation as a primary acidification mechanism on this calcareous soil.

Discussion

The magnitude of rhizosphere acidification observed here-up to 1.41 pH units for mung bean-falls at the upper end of values reported in the literature. Hinsinger et al. [4] noted that legumes commonly lower rhizosphere pH by 0.5-1.5 units in calcareous soils due to excess cation uptake during N₂ fixation. The strong correlation between total organic acids and pH change supports the dual mechanism of proton extrusion plus organic acid chelation [11,12] . From a practical standpoint, these results reinforce the agronomic value of including legumes in rotations on calcareous soils. Residual acidification from a legume crop may persist long enough to benefit a succeeding cereal. Singh et al. [13] showed that rhizosphere pH effects lasted 15-20 days after legume harvest in a calcareous Indian soil, sufficient to overlap with early growth of a following crop. The disproportionately large response of Zn to pH reduction (47% increase for mung bean versus 38% for Mn) is consistent with the steep solubility curve of zinc carbonate between pH 7.0 and 8.0 [14] . This suggests that even modest pH reductions on calcareous soils can substantially improve Zn nutrition.

Conclusion

Root-induced acidification significantly lowered rhizosphere pH and improved DTPA-extractable Fe, Mn, Zn, and Cu in a calcareous Vertisol. Legumes, particularly mung bean and soybean, caused the greatest pH reductions through organic acid exudation and proton release. DTPA-extractable Zn increased by up to 76% in the mung bean rhizosphere relative to bulk soil. These findings support the inclusion of legumes in crop rotations on calcareous soils as a low-cost strategy to improve micronutrient availability for subsequent cereal crops.

References

1. Marschner H. Mineral nutrition of higher plants. London: Academic Press; 1995:1-889.
2. Mengel K, Kirkby EA. Principles of plant nutrition. Kluwer Academic Publishers; 2001:1-849.
3. Cakmak I. Enrichment of cereal grains with zinc: agronomic or genetic biofortification? Plant and Soil. 2008;302(1):1-17.
4. Hinsinger P, Plassard C, Tang C, Jaillard B. Origins of rootmediated pH changes in the rhizosphere and their responses to environmental constraints: a review. Plant and Soil. 2003;248(1):43-59.
5. Neumann G, Romheld V. Root excretion of carboxylic acids and protons in phosphorus-deficient plants. Plant and Soil. 1999;211(1):121-130.
6. Tang C, Barton L, Raphael C. Pasture legumes and grasses differ in their capacity to acidify a low-buffer soil. Australian Journal of Agricultural Research. 1998;49(1):5358.
7. Dinkelaker B, Romheld V, Marschner H. Citric acid excretion and precipitation of calcium citrate in the rhizosphere of white lupin. Plant, Cell and Environment. 1989;12(3):285-292.
8. Youssef RA, Chino M. Development of a new rhizobox system to study the nutrient status in the rhizosphere. Soil Science and Plant Nutrition. 1989;35(3):461-465.
9. Lindsay WL, Norvell WA. Development of a DTPA soil test for zinc, iron, manganese and copper. Soil Science Society of America Journal. 1978;42(3):421-428.
10. Jones DL. Organic acids in the rhizosphere: a critical review. Plant and Soil. 1998;205(1):25-44.
11. Dakora FD, Phillips DA. Root exudates as mediators of mineral acquisition in low-nutrient environments. Plant and Soil. 2002;245(1):35-47.
12. Lambers H, Shane MW, Cramer MD, Pearse SJ, Veneklaas EJ. Root structure and functioning for efficient acquisition of phosphorus: matching morphological and physiological traits. Annals of Botany. 2006;98(4):693-713.
13. Singh B, Natesan SKA, Singh BK, Usha K. Improving zinc efficiency of cereals under zinc deficiency. Current Science. 2003;88(1):36-44.
14. Alloway BJ. Zinc in soils and crop nutrition. Brussels: International Zinc Association; 2008:1-135.
15. Gao X, Hoffland E, Stomph T, Grant CA, Zou C, Zhang F. Improving zinc bioavailability in transition from flooded to aerobic rice. Agronomy for Sustainable Development. 2012;32(2):465-478.
16. Rengel Z. Availability of Mn, Zn and Fe in the rhizosphere. Journal of Soil Science and Plant Nutrition. 2015;15(2):397409.
17. Fageria NK, Baligar VC, Clark RB. Micronutrients in crop production. Advances in Agronomy. 2002;77:185-268.
18. Brennan RF, Armour JD, Reuter DJ. Diagnosis of zinc deficiency. In: Zinc in soils and plants. Springer; 1993. p. 167-181.
19. Loeppert RH, Suarez DL. Carbonate and gypsum. In: Methods of soil analysis part 3. SSSA; 1996. p. 437-474.