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Isolation, Purification and Characterization of Phytotoxins Produced by Rhizoctonia solani AG-3, the Cause Agent of Potato Stem Canker

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Abstract

Studies were conducted to isolate and characterize the phytotoxins extracted from culture filtrates of Rhizoctonia solani isolate anastomosis group 3 (AG-3) using preparative thin layer chromatography (TLC) method. The culture filtrates of R. solani contain a toxic substance that causes canker on potato stems. Eight toxin fractions were collected by column layer chromatography but only four spots were obtained in the TLC analysis. All the four toxin fractions (third, fourth, sixth and seventh fractions) were recognised to have phytotoxic activity. Fourier-transform infrared spectroscopy (FTIR), high performance liquid chromatography (HPLC) and 1H and 13C NMR spectral techniques were used to characterize the phytotoxin. The major functional groups identified from FTIR spectrum includes 2962.1 cm−1 (−CH3), 1465.6 cm−1 (−CH2−), 1727.9 cm−1 (C = O), 1729.8 cm−1 (−COOH) and 1124.3 cm−1 (−OH). Result from HPLC, 1H NMR and 13C NMR showed phytotoxic fractions of 3-methylthiopropionic acid (3-MTPA) and 3-methylthioacrylic acid (3-MTAA). The results of this investigation add to the body of evidence that multiple related compounds are involved in R. solani disease development.

Resumen

Se desarrollaron estudios para aislar y caracterizar las fitotoxinas extraídas de filtrados de cultivo de un aislamiento de Rhizoctonia solani grupo de anastomosis 3 (AG-3), usando el método de cromatografía de capa fina (TLC). Los filtrados del cultivo de R. solani contienen una substancia tóxica que causa cáncer en los tallos de papa. Se colectaron ocho fracciones de la toxina por cromatografía de columna, pero solo se obtuvieron cuatro manchas en el análisis de TLC. A las cuatro fracciones (tercera, cuarta, sexta y séptima) se les reconoció que tenían actividad fitotóxica. Se usaron técnicas de espectrometría de espectroscopía de infrarrojos transformada de Fournier (FTIR), cromatografía de líquidos de alta resolución (HPLC), y 1H y 13C NMR, para caracterizar la fitotoxina. Los principales grupos funcionales identificados del espectro del FTIR incluyen 2962.1 cm−1 (−CH3), 1465.6 cm−1 (−CH2-), 1727.9 cm−1 (C = O), 1729.8 cm−1 (−COOH) y 1124.3 cm−1 (−OH). Los resultados del HPLC, 1HNMR y 13C NMR mostraron fracciones fitotóxicas de ácido 3-metiltiopropionico (3-MTPA) y ácido 3-metiltioacrilico (3-MTAA). Los resultados de esta investigación le agregan al cuerpo de evidencia que múltiples compuestos relacionados están involucrados en el desarrollo de la enfermedad por R. solani.

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Acknowledgments

This work was funded by the National Natural Science Foundation of China (31360500). We would like to thank Dr. Guorui Fu from the Department of Chemistry, Gansu Agriculture University, for his technical assistance obtaining FTIR and NMR data, respectively. The authors are also grateful to Ms. Xiaoqin Lu, from Gansu Provincial Key Lab of Aridland Crop Science, Gansu Agricultural University, for providing isolates of R. solani AG-3.

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Authors and Affiliations

  1. College of Resources and Environmental Sciences / Gansu Provincial Key Lab of Aridland Crop Science, Gansu Agricultural University, Lanzhou, 730070, China

    Frederick Kankam, Huizhen Qiu, Chunhong Zhang & Jing He

  2. Department of Chemistry, College of Sciences, Gansu Agriculture University, Lanzhou, 730070, China

    Lumei Pu, Haitao Long & Huixiu Zhang

  3. Department of Agronomy, University for Development Studies, P.O. Box 1882, Tamale, Ghana

    Frederick Kankam

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  1. Frederick Kankam
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  2. Huizhen Qiu
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Correspondence to Frederick Kankam or Huizhen Qiu.

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Kankam, F., Qiu, H., Pu, L. et al. Isolation, Purification and Characterization of Phytotoxins Produced by Rhizoctonia solani AG-3, the Cause Agent of Potato Stem Canker. Am. J. Potato Res. 93, 321–330 (2016). https://doi.org/10.1007/s12230-016-9506-8

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