Accumulation and transformation of inorganic and organic arsenic in rice and role of thiol-complexation to restrict their translocation to shoot

Mishra et al. 2017 — Thiol complexation restricts arsenic translocation from rice roots to shoots, but methylated arsenic is more mobile

This Scientific Reports paper investigated in planta accumulation, redox transformation and thiol-complexation of inorganic arsenate (AsV), methylarsonate (MAV) and dimethylarsinate (DMAV) in rice (cv. Triguna) under 7-day hydroponic exposure. Roots and shoots were analyzed by anion-exchange HPLC-ICP-MS (redox/species) and RP-HPLC-(ICP-MS)-(ESI-MS) in parallel (thiol complexes). AsV and MAV were efficiently reduced to AsIII and MAIII respectively in both roots and shoots; no trivalent DMA was detected. In roots, up to 48% of root As in AsV-exposed plants and 83% of root As in MAV-exposed plants was bound as thiol complexes covering up to 20 and 16 distinct As-species, including PC2, hm-PC2, des-Gly-PC2, hm-GSH and γ-EC homologues. Despite extensive complexation in roots, shoot transfer factor (TF, shoot/root As) ranked DMAV (4.91) > MAV (0.18) > AsV (0.033), and in shoots 78% of total As in MAV-exposed and 71% of As in AsV-exposed plants was present as weakly bound MAIII and AsIII respectively.

Key numbers

Plants were exposed to 10 µM AsV, 50 µM MAV, or 50 µM DMAV for 7 days; values are mean ± SD; analytical detection is ICP-MS at m/z 75 with HPLC speciation upstream. “µg/g FW” denotes µg arsenic per gram fresh weight of plant tissue.

Growth response (Fig 1; n=30):

• Pre-experiment: AsV at 20 µM caused 47 ± 8.3% inhibition of root length after 7 d (data not shown); MAV and DMAV were moderately toxic up to 50 µM. The 10 µM AsV vs 50 µM MAV/DMAV exposures used in the main experiment were chosen because total accumulation was comparable at those concentrations.
• Root length reduction at 7 days, exposure vs control (Fig 1A): AsV 16%, MAV 23%.
• Root and shoot weight and length significantly reduced for all three As species vs control (P ≤ 0.05); maximum reduction in root and shoot weight was in MAV-exposed plants followed by AsV.

Total As accumulation (Fig 2A; n=12):

• At equimolar exposure (10 µM AsV vs 10 µM MAV/DMAV in supplementary comparison): total accumulation in MAV- and DMAV-exposed plants was approximately 3- and 14-fold lower than in AsV-exposed plants.
• Shoot/root transfer factor (TF): AsV 0.033 ± 0.012; MAV 0.18 ± 0.05; DMAV 4.91 ± 0.72.
• AsV-exposed plants: ~96% of accumulated As retained in roots; ~4% translocated to shoots.
• MAV-exposed plants: ~85% in roots; ~15% in shoots.
• DMAV-exposed plants: ~19% in roots; ~81% in shoots.

Speciation by anion-exchange HPLC-ICP-MS (Figs 2B, 3):

• Extraction efficiency 91 ± 5.3% (n=32); chromatographic recovery 88 ± 7.9%.
• AsV-exposed plants: only AsV and AsIII were detected in roots and shoots; AsIII contributed up to ~84% of total As in roots and ~75% of total As in shoots.
• MAV-exposed plants: ~84–86% of As in roots and shoots was reduced to MAIII; ~12% remained as MAV; small additional quantities of AsIII (and small AsV/AsIII in shoots) detected.
• DMAV-exposed plants: ~85% of As in roots and ~98% in shoots remained as DMAV; small amounts of AsV, MAIII and MAV detected in roots; only AsV and AsIII detected in shoots alongside DMAV.

Thiol complexation by RP-HPLC-ICP-MS/ESI-MS (Figs 4–6):

• AsV-exposed roots: up to 20 distinct As-containing species detected; thiol-bound As accounted for 48 ± 6.4% of eluted As; 12 of 20 species identified as PC- and homologue-complexes (PC2 mixed-thiol complexes with Cys/GSH/hm-GSH/γ-EC plus PC3 and hm-PC3 homologues).
• Main AsV-root species at retention 15.2 and 15.6 min corresponded to hm-PC2-As-γEC (m/z 446) and hm-PC2-As-hm-GS (m/z 490, co-eluted with peak 7 and 8).
• AsV-exposed shoots: most As eluted unbound; only ~2–6% of total shoot As was thiol-complexed, mostly as hm-PC2-γEC, hm-PC2-As-hm-GS, hm-GS-As-GS and hm-PC3.
• MAV-exposed roots: up to 16 As-containing species; 11 identified as thiol-bound; peak 1 (unbound As) ~17 ± 4.3% of eluted As; of the total thiol-bound pool, ~78% was bound to GSH and homologues of GSH with hm-GS-As(CH3)-GS and (GS)2-As-CH3 as main species, and ~16% was bound to PC2 and PC2-homologues; no PC3 or hm-PC3 complex identified.
• MAV-exposed shoots: only an unidentified species at 2.9 min (containing 7.8 ± 2.6% of total shoot As) was observed alongside uncomplexed As.
• DMAV-exposed roots/shoots: most As eluted as DMAV; an unidentified shoot species at 3.9 min contained 2.3 ± 1.1% of shoot As; no DMA-thiol complex identified.

Free thiols (Fig 7; n=12):

• Control roots and shoots: GSH and hm-GSH were the main free thiols; small amounts of oxidized species also present.
• Maximum increase in free reduced thiols (mainly GSH and hm-GSH) was observed in MAV-exposed plants: 24-fold (roots) and 3-fold (shoots) over control.
• AsV-exposed: free reduced thiols ~5-fold (roots) and 2.8-fold (shoots) higher than control.
• DMAV-exposed: ~5.5-fold (roots) and 2.8-fold (shoots) higher than control.
• AsV-exposed plants showed a significant increase in free PC2 (mostly hm-PC2) in shoots vs roots (P < 0.001); roots mostly contained oxidized PCs.

Quality control:

• Standards used for As-species identification: sodium arsenite (0.05 M, Merck), arsenate (Arsenic standard 1000 mg As L−1 [As2O5 in water] Titrisol, Merck), dimethylarsinic acid (cacodylic salt, Alfa Aesar), methylarsonic acid (PAMOL, Tel-Aviv); MAIII synthesized in-house by the Cullen et al. (1989) method and confirmed by HPLC-ICP-MS-ESI-Q-TOF-MS.
• Total-As CRMs in solution: SLRS-5 (River Water Reference Material for Trace Metals, NRCC), SPSSW1 (Surface Water Level 1, Spectra Pure Standards), SRM 1643e (Trace Elements in Water, NIST).
• Plant-matrix CRM analyzed for sum-of-species QC: NIST 1568a Rice Flour, total sum of species 279 ± 3 µg/kg, with 32% iAs and 64% methylated As (matches certified speciation pattern).
• Internal standard: rhodium (4 µg/L) added to all samples.

Methods (brief)

Seeds of Oryza sativa L. subsp. indica cv. Triguna (purchased from an authenticated supplier in Kolkata, West Bengal) were surface-sterilized in 0.5% NaOCl for 15 min, rinsed, and germinated in moist Petri dishes for 5 days at 28 °C in the dark. Uniform seedlings were transferred to a black-lidded Araponics hydroponic growing system (Araponics SA, Liège, Belgium; 18 plants/tray, 2 L resident volume). The nutrient solution composition was: 2 mM Ca(NO3)2; 1 mM MgSO4·7H2O; 0.5 mM K2HPO4; 0.1 mM H3BO3; 10 µM KCl; 0.25 mM MnSO4·H2O; 0.2 µM Na2MoO4·2H2O; 0.5 µM NiSO4·6H2O; 0.5 µM CuSO4·5H2O; 0.5 µM ZnSO4; 50 µM Fe-EDTA; 50 µM FeCl3·6H2O. pH 5.8 by 0.1 M KOH or HCl; half the solution replaced every day. After 10 days of growth in controlled environment (14 h light at 250–300 µEm−2 s−1, 28–20 °C day/night), plants were exposed for 7 days to AsV (10 µM, Na2HAsO4·7H2O, Alfa Aesar), MAV (50 µM, (CH3)HAsNaO5, PAMOL) or DMAV (50 µM, (CH3)2NaAsO2·3H2O, Sigma); K2HPO4 and KCl reduced to 10 µM and 1 mM during exposure.

Harvested roots and shoots were rinsed in ice-cold 2 mM phosphate buffer (pH 6.0, 15 min), then twice in deionized water (2 min), separated, weighed, and frozen in liquid N2. Total As was measured after microwave digestion (HNO3-H2O2) on a Thermo Fisher Element XR sector-field ICP-MS, with rhodium (4 µg/L) as internal standard. As-speciation samples were ground under liquid N2, aliquoted (200–400 mg), and extracted in 1% degassed formic acid (90 min at 1 °C) for anion-exchange HPLC-ICP-MS speciation; a separate aliquot was extracted in degassed deionized water for 18 h on a tube rotator for As-thiol-complex analysis. All handling, extraction and analysis steps were under N2 in a glove box.

Speciation chromatography used a Dionex IonPac AS7 anion-exchange column on a 1100 Series HPLC (Agilent Technologies) coupled to a VG PQ EXCELL ICP-MS (Thermo Elemental), eluted with a 0.04 mM HNO3 → 50 mM HNO3 staged gradient. As-thiol-complex analysis used an Atlantis reversed-phase dC18 column (4.6 × 150 mm, 5 µm; Waters) on the same HPLC, with post-column 1:1 split to ESI-MS (6130 quadrupole LC/MSD; Agilent) in positive SIM mode (m/z 120–1200) and ICP-MS (7500ce; Agilent) in parallel. Identifications used characteristic retention times and m/z signals; quantification used ICP-MS m/z 75 data only, calibrated against DMA standard; ArCl+ and CaCl+ interferences were checked on first samples per batch. PC2, PC3, PC4, des-Gly-PC2 and des-Gly-PC3 standards (90–95% purity in 6.3 mM EDTA / 0.1% TFA) were procured from Clonestar Peptide Services; in-vitro As-thiol complexes were synthesized in 5 mM AsIII/MAIII + 200 µM PC + 2 mM GSH + 10 mM Cys in 0.1% degassed formic acid and stored overnight under N2.

Free thiols were quantified by HPLC-ESI-MS in single-ion-monitoring (SIM) mode using GSH for reduced-thiol calibration and GSSG for oxidized species. Statistics used SigmaPlot 11 (SPSS Science, USA), one-way ANOVA with Student–Newman–Keuls post-hoc all-pair-wise comparison, and Holm–Sidak post-hoc for two-way ANOVA, at α = 0.05.

Limitations the authors flag: (1) the 7-day exposure was short and may have constrained MAV reduction kinetics, possibly explaining persistent MAV-thiol complexation in roots; (2) the whole shoot (2 mm above root–shoot junction) was used for speciation, so vascular-tissue (xylem/phloem) species may dominate over mesophyll-cell species; (3) DMA-thiol complexes could not be identified, only an unidentified shoot species accounting for ~2.3% of shoot As was observed.

Implications

The source contributes mechanism evidence for rice arsenic accumulation: it characterizes in planta As-species redox transformations, thiol-complexation patterns and root-to-shoot translocation factors in a West-Bengal rice cultivar (Triguna) under controlled hydroponic exposure to AsV, MAV and DMAV. It does not measure rice grain or any rice food product, so it does not contribute occurrence data to any product-category page. Its routing role is upstream context for arsenic-inorganic discussions of why rice accumulates iAs preferentially in root tissue, why methylated As species (MAV, DMAV) translocate more efficiently to shoots than inorganic AsV does, and why MAIII (a known cytotoxic species more toxic than AsIII) is the predominant in-tissue species in MAV-exposed plants — a relevant consideration for soils where methylated As is generated by microbial action or where methylated arsenicals were historically applied as pesticides or herbicides.

Wiki pages this source may touch

• rice
• arsenic-inorganic
• arsenic-total

Verification notes

Fresh ingest 2026-05-28 from the source PDF at the verified raw_path. SHA-256 35b5c5c4… recorded.

• Scope: This is a mechanism / plant-physiology paper (hydroponic exposure, root and shoot tissues only); it does not measure rice grain or any market product. Frontmatter products: [] reflects this. matrices: [rice] is the closest broad matrix term but tags rice plant material rather than rice food matrix; downstream routing should treat this as a mechanism source, not an occurrence source.
• Metals vocabulary extension. MA and DMA are not in the system-prompt’s controlled metal abbreviation list (Pb, Cd, iAs, tAs, iHg, MeHg, tHg, Ni, Al, Cr, Cr-VI, Sn, Sb, U), but the paper centrally speciates them and reports finding-bearing results on them. They are included here following the precedent established in carey2015-rice-arsenic-percolating-cooking (which uses DMA). Flagged for taxonomy-vocab review.
• Jurisdictions. DE because the experimental work was performed at UFZ Helmholtz Centre for Environmental Research, Leipzig, Germany. IN because the rice cultivar (Triguna, subsp. indica) is a West Bengal variety from an authenticated Kolkata seed supplier, and the Discussion frames Indian Bengal soils as the disease-context motivation for the study.
• sample_n: 12 reflects the As-speciation/accumulation replicate count, which is the basis of the speciation findings reported in ## Key numbers. Growth measurements used n=30 plants per treatment (recorded in the inline text).
• Brand firewall (Part 12). The methods section names instrument vendors (Thermo Fisher Element XR, Agilent 1100 HPLC, VG PQ EXCELL, Agilent 6130 quadrupole, Agilent 7500ce), reference-material suppliers (NRCC SLRS-5, NIST 1643e, NIST 1568a), reagent suppliers (Alfa Aesar, Sigma, PAMOL, Merck, Tritisol), hydroponic-system supplier (Araponics SA), peptide supplier (Clonestar Peptide Services) and statistical software (SigmaPlot 11, SPSS Science). These are scientific-method vendor names permitted under Part 12 Exception 2 (locked 2026-05-17); they are not brand-by-brand contamination attributions and do not violate the brand firewall.
• No HMTc threshold work. This paper does not propose any contamination limit and is upstream of grain-occurrence data; it is not routed to any product page.

Audit subagent (2026-05-28) flagged the following; verified against source and applied or rejected:

• Root length reduction TRANSPOSED. Initial ingest reported AsV 23%, MAV 16% (max). Source p. 2 ¶3: “Maximum reduction in root length (16%) was in AsV exposed plants while in MAV exposed plant (23%)…” Verified — corrected so AsV = 16% and MAV = 23%.
• AsV pre-experiment concentration WRONG. Initial ingest gave the 47 ± 8.3% root-length-inhibition figure at 10 µM AsV. Source p. 2 ¶2: “The AsV at 20 µM caused severe toxicity after 7d (47 ± 8.3% inhibition in root length) (data not shown).” Verified — corrected to 20 µM and rephrased to note that 10 µM AsV vs 50 µM MAV/DMAV were chosen for the main experiment because total accumulation was comparable at those concentrations.
• Nutrient-solution composition transcription errors. Initial ingest had a row-shift error and wrong salt across several trace-element entries. Source p. 10 specifies: “2 mM Ca(NO3)2, 1 mM MgSO4 × 7H2O, 0.5 mM K2HPO4, 0.1 mM H3BO3, 10 µM KCL, 0.25 mM MnSO4 × H2O, 0.2 µM Na2MoO4 × 2H2O, 0.5 µM NiSO4 × 6H2O, 0.5 µM CuSO4 × 5H2O, 0.5 µM ZnSO4, 50 µM Fe-EDTA and 50 µM FeCl3 × 6H2O.” Verified — corrected KH2PO4 → K2HPO4, added missing KCl 10 µM, corrected MnSO4·H2O 10 µM → 0.25 mM, Na2MoO4·2H2O 0.25 µM → 0.2 µM, NiSO4·6H2O 0.2 µM → 0.5 µM.
• Arsenate standard chemistry wrong. Initial ingest wrote “As2O3 in water; Tritisol, Merck.” Source p. 11: “arsenate (Arsenic standard 1000 mg As L−1 [As2O5 in water] Titrisol, Merck).” Verified — As2O3 was incorrect (arsenic trioxide is AsIII; arsenate derives from As2O5, AsV). Corrected to As2O5, brand spelling corrected Tritisol → Titrisol, and the As-concentration of the standard (1000 mg As L−1) and the AsIII standard’s separate Merck citation (sodium arsenite 0.05 M) added for completeness.
• 78% denominator clarified. Initial ingest wrote “~78% of As in MAV-exposed plants bound to GSH and homologues of GSH.” Source p. 4: “Approximately 78% of total bound As in MAV exposed plant was bound to GSH and homologues of GSH…” Verified — the 78% is a fraction of the thiol-bound pool (which itself is ~83% of root As), not of total As. Corrected phrasing to “of the total thiol-bound pool.”
• “Of root As” precision clarified. Initial ingest opening said “up to 48% of AsV-exposed As and 83% of MAV-exposed As.” Source abstract: “up to 48 and 83% of root As in AsV and MAV exposed plants…” Verified — corrected to “up to 48% of root As in AsV-exposed plants and 83% of root As in MAV-exposed plants.”
• Extraction-efficiency sample size n=48 flagged by audit subagent as a transcription error. Source p. 2 ¶3 actually states “The extraction efficiency of As was 91 ± 5.3% (n = 32)”; Fig 3 caption does not mention n=48. Audit finding was a false positive — wiki value n=32 matches the source. No correction applied.

Page history

The five most recent substantive edits to this page. The full version history lives in git; when DOI minting comes online (see schema docs), each entry below will also link to a version-pinned DataCite DOI.

Commit	Date	Description
ce3e07c	2026-05-28	activation | Vercel DATACITE env slots set, curators.md filled with founder entry + six scoped reviewer invitations, peer-review onboarding playbook drafted
51400b9	2026-05-28	audit-queue: gasparik2017-wild-boar-slovakia-metals audited-revised