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Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes
Open accessAbstract
Lead (Pb) is a common toxic metal that causes severe health and environmental problems. However, the defensive role and underlying mechanism of combined biochar (BC) and melatonin (MT) against Pb stress are still unclear. Therefore, to fill this gap, this study investigated the impacts of BC and MT on rice growing in Pb-polluted soil. This study included different treatments: control, Pb stress (300 mg kg−1), Pb stress (300 mg kg−1) + BC (2%), Pb stress (300 mg kg−1) + MT (30 µM), and Pb stress (300 mg kg−1) + BC (2%) + MT (30 µM). Pb reduced rice growth and yield by hindering photosynthetic pigments, relative water contents (RWCs), osmolyte synthesis, nutrient uptake, increase in oxidative markers, and Pb accumulation. Biochar and MT increased rice productivity by increasing chlorophyll synthesis, osmolytes, and nutrient uptake and decreasing Pb accumulation. The co-application of BC and MT decreased Pb accumulation in the roots (30.40%) and shoots (72.79%), the translocation factor (30.01%), the biological accumulation coefficient (20.17%), and the soil Pb concentration (59.02%). The co-application of BC and MT enhanced proline (39.65%), soluble protein (47.09%), ascorbate peroxidase (APX; 26.47%), catalase (CAT; 65.51%), peroxidase (POD; 89.56%), and superoxide dismutase (SOD; 65.53%) activities, which ensured better productivity. Additionally, the BC+MT application increased the expression of antioxidant defense genes (OsAPX, OsCAT, OsPOX, and OsSOD) and decreased the expression of metal transporter genes (OsHMA9 and OsNRAMP5), which protected the rice plants from damage caused by Pb toxicity. These results suggested that BC+MT could be a promising strategy to mitigate Pb toxicity and maintain sustainable and safer food production.
Overview
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TL;DR
In a pot experiment with Pb-polluted soil, combined biochar (BC) and melatonin (MT) mitigated Pb toxicity in rice more effectively than either alone. The combined treatment reduced Pb accumulation and soil Pb while improving growth, yield, antioxidant defense, osmolyte levels, iron plaque formation, and expression of metal transporter genes.
Key findings
- Pb stress (300 mg kg−1) reduced rice growth and yield; co-application of BC (2%) + MT (30 µM) decreased root Pb by 30.40%, shoot Pb by 72.79%, translocation factor by 30.01%, biological accumulation coefficient by 20.17%, and soil Pb concentration by 59.02%.
- BC+MT increased proline by 39.65%, soluble protein by 47.09%, APX by 26.47%, CAT by 65.51%, POD by 89.56%, and SOD by 65.53%, and increased grain yield, biological yield, and harvest index by 54.39%, 33.30%, and 14.65%, respectively.
- BC+MT increased Fe concentration in root ACA extracts by 67.70% and decreased Pb concentration in ACA extracts by 50.14%; it also upregulated antioxidant genes OsAPX, OsCAT, OsPOX, and OsSOD and downregulated metal transporter genes OsHMA9 and OsNRAMP5 by 96.19% and 83%, respectively, compared with control.
Methods
- Open-greenhouse pot study at Jiangxi Agricultural University in 2024 using silty loam soil (pH 5.45); treatments were control, Pb stress (300 mg kg−1), Pb+BC (2%), Pb+MT (30 µM), and Pb+BC+MT, arranged in a completely randomized design with three replications.
- Rice straw biochar was pyrolyzed at 600°C for 8 hours; Pb was applied as PbCl2 and stabilized for 1 month; five 25-day-old rice seedlings were transplanted per pot, and MT was foliar-sprayed 20 days after transplanting.
- Measured traits included photosynthetic pigments, RWC, oxidative markers, osmolytes, antioxidant enzyme activities, nutrient and Pb concentrations in plant/soil, Pb fractions, iron plaque formation via ACA extraction, and gene expression by qRT-PCR; data were analyzed with one-way ANOVA and Tukey’s HSD, plus PCA and correlation analysis.
Limitations
- The paper states that more transcriptome and metabolomics studies are needed to validate the results, field studies in contaminated areas are needed before recommendation, and future work should test BC alone and MT alone in unpolluted soils.
- Evident from the text: findings come from a single greenhouse pot experiment with n = 3, one Pb concentration (300 mg kg−1), one soil type, and one rice cultivar/context, so field-scale and long-term effects are not established.
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In the paper
“The maximum Pb concentration in the roots (139.67 mg kg−1) and shoots (96.37 mg kg−1) was detected in Pb-contaminated soil.”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Pb and nutrient concentrations in plant parts
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In the paper
“The co-application of BC and MT decreased Pb accumulation in the roots (30.40%) and shoots (72.79%), the translocation factor (30.01%), the biological accumulation coefficient (20.17%), and the soil Pb concentration (59.02%).”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Abstract
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In the paper
“We also noted that BC alone increased TSP, FAA, and proline synthesis by 34.69%, 17.60%, and 17.71%, respectively, while MT alone increased TSP, FAA, and proline synthesis by 16.32%, 8.04%, and 0.37%, respectively ( Table 2 ).”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Oxidative markers and antioxidant activities
plainstates a cause- Asserted: passes — The authors put it forward as true.
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In the paper
“while MT alone enhanced soil, P, and K availability by 20.27%, 31.30%, and 21.55%, respectively ( Figure 4 ).”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Soil properties
plainstates a cause- Asserted: passes — The authors put it forward as true.
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In the paper
“Vector analysis revealed an inverse relationship between growth parameters and stress indicators, underscoring the effectiveness of BT+MT in alleviating Pb-induced stress and enhancing plant growth and physiological responses ( Figure 8 ).”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Principal component and correlation analyses
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Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
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Rated major: it would touch a result — a value a result rests on, reported and recomputed p-values on opposite sides of the significance level, two passages that conflict with no sign which is right, or a cited work retracted before this paper was published.
In the paper
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Concentrations of Pb and iron in root surface iron plaques and relative expression of antioxidant defense and metal transporter genes
“Pb toxicity decreased the expression of antioxidant defense genes; however, BC+MT significantly enhanced the expression of all the antioxidant genes”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Discussion
“Pb toxicity increased the expression of antioxidant defense genes, and BC and MT applications further increased the expression of antioxidant defense genes (OsAPX, OsCAT, OsPOX, and OsSOD).”
Rated minor: it would not touch a result — a value no result rests on, a p-value that stays on the same side of the significance level, or a conflict that is clerical, resolved elsewhere in the paper, or outside its own findings.
In the paper
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Discussion
“However, BC and MT significantly increased chlorophyll synthesis as compared to control.”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Chlorophyll and relative water contents
“| Control | 0.83a ± 0.040 | 0.67a ± 0.016 | 5.05a ± 0.057 | 84.40a ± 1.76 | | Pb stress | 0.53c ± 0.016 | 0.33d ± 0.094 | 3.43d ± 0082 | 63.23d ± 0.85 | | Pb stress+BC | 0.70b ± 0.017 | 0.52b ± 0.028 | 3.94b ± 0.086 | 71.57c ± 0.98 | | Pb stress+MT | 0.66b ± 0.020 | 0.43c ± 0.020 | 3.71c ± 0.065 | 68.37c ± 1.51 | | Pb stress+BC+MT | 0.73b ± 0.014 | 0.61a ± 0.041 | 4.46a ± 0.043 | 77.40b ± 1.60 |”
Rated minor: it would not touch a result — a value no result rests on, a p-value that stays on the same side of the significance level, or a conflict that is clerical, resolved elsewhere in the paper, or outside its own findings.
In the paper
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Abstract
“peroxidase (POD; 89.56%)”
§ Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes > Results > Oxidative markers and antioxidant activities
“The co-application of BC and MT enhanced APX, CAT, POD, and SOD activities by 26.47%, 65.51%, 93.10%, and 65.53%, respectively.”
Cited works that were retracted
Works this paper cites that were later retracted, from the retraction notices publishers deposit. Major when the notice came out before this paper was published.
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In the paper
“Fu, 2023, Recent progresses, challenges, and opportunities of carbon-based materials applied in heavy metal polluted soil remediation, Sci. Total Environ., 856, 10.1016/j.scitotenv.2022.158810”
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A note, not an issue: for example, a cited work that was retracted only after this paper was published, which its authors could not have known.
In the paper
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A note, not an issue: for example, a cited work that was retracted only after this paper was published, which its authors could not have known.
In the paper
“El-Naggar, 2020, Biochar-induced metal immobilization and soil biogeochemical process: an integrated mechanistic approach, Sci. Total Environ., 698, 10.1016/j.scitotenv.2019.134112”
Cited work 10.1016/j.scitotenv.2019.134112 · retracted 2026-08-26 · notice 10.1016/j.scitotenv.2026.182256 · this paper published 2025-07-03
Citations
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Cited by
Noor, M. A., Hassan, M., Khan, T. A., Zhou, B., & Guo-qin, H. (2025) · Plants · doi:10.3390/plants14152453 · Cited by 11
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Le, L. B., Nguyen, B. T., & Dinh, T. D. (2026) · Water Air & Soil Pollution · doi:10.1007/s11270-026-09109-9 · Cited by 3
Synergistic role of gamma-aminobutyric acid and melatonin in mitigating lead stress in rice
Wang, X., Chen, C., Sun, X., Zaman, Q., Wang, C., & Tang, H. (2025) · BMC Plant Biology · doi:10.1186/s12870-025-07774-2 · Cited by 2
Wang, X., Chen, C., Sun, X., Wang, C., & Tang, H. (2026) · Frontiers in Plant Science · doi:10.3389/fpls.2026.1778126 · Cited by 2
Metadata
- Authors
- Khan, T. A., Su, Q., Guoqin, H., Du, Z., Noor, M. A., Asseri, T. A. Y., & Hassan, M. U.
- Affiliations
- Jiangxi Agricultural University (CN)
- Jinggangshan University (CN)
- King Khalid University (SA)
- Corresponding author
- Su, Qitao
- Guoqin, Huang
- Venue
- Frontiers in Plant Science
- Publisher
- Frontiers Media SA
- Published
- 2025-07-03
- Volume
- 16
- Pages
- 1609825-1609825
- ISSN
- 1664-462X (ISSN-L)
- PMC
- PMC12267010
- OpenAlex
- W4411947837
- Semantic Scholar
- 34f0447b830ef128d883a05a7ed45fdf5450f333
- Cited by
- 9
- References
- 82
- Citations by year
- 2026: 7 · 2025: 2
- FWCI
- 1.84
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Frontiers in Plant Science (publishedVersion, cc-by)
- PubMed (publishedVersion)
- PubMed Central (publishedVersion, cc-by)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Funders
- National Natural Science Foundation of China (41661070)
- King Khalid University
- Topics
- Physical Sciences › Environmental Science › Pollution › Heavy metals in environment
- Life Sciences › Agricultural and Biological Sciences › Plant Science › Plant Stress Responses and Tolerance
- Life Sciences › Agricultural and Biological Sciences › Plant Science › Plant Micronutrient Interactions and Effects
- Keywords
- lead toxicity, biochar, melatonin, rice, heavy metal stress, antioxidant enzymes, OsNRAMP5, proline accumulation, photosynthetic pigments, translocation factor, metal accumulation, soil remediation
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- Khan, T. A., Su, Q., Guoqin, H., Du, Z., Noor, M. A., Asseri, T. A. Y., & Hassan, M. U. (2025). Integrative biochar and melatonin application mitigates lead toxicity in rice by modulating antioxidant activities and iron plaque formation and downregulating the expression of metal uptake genes. Frontiers in Plant Science. https://doi.org/10.3389/fpls.2025.1609825
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