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Purification and Characterization of Antioxidant Peptides from Leukocyte Extract of Crocodylus siamensis

The Protein Journal volume 33, pages 24–31 (2014)Cite this article

Abstract

Antioxidant peptides were isolated from the leukocyte extract of the Siamese crocodile, Crocodylus siamensis. Crocodile leukocyte was extracted by a combination of methods including freeze-thawing, acetic acid extraction and homogenization. The peptides in the leukocyte extract were purified by anion exchange chromatography and reversed phase-high performance liquid chromatography. The 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay was used to evaluate the antioxidant activity of the elution peaks at each purification step. As a result, there were two purified peptides exhibiting strong antioxidant activity in reducing free radicals on DPPH molecules. The amino acid sequences of these peptides were determined by LC-MS/MS as TDVLGLPAK (912.5 Da) and DPNAALPAGPR (1,148.6 Da), and their IC50 values were 153.4 and 95.7 μM, respectively. The results of this study therefore indicate that leukocyte extract of C. siamensis contains peptides with antioxidant activity which could be used as a novel antioxidant.

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Abbreviations

ACN:

Acetonitrile

DPPH:

1,1-Diphenyl-2-picrylhydrazyl

hRBCs:

Human red blood cells

IC50 :

Half maximal inhibitory concentration

TFA:

Trifluoroacetic acid

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Acknowledgments

This study was supported by the National Research University Project of Thailand, Office of the Higher Education Commission, through the Food and Functional Food Research Cluster of Khon Kaen University and Thailand Research Fund (RMU5380045). Finally, we wish to acknowledge the support of the Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Faculty of Science, Khon Kaen University, Thailand and to thank Sriracha Moda co., Ltd., Chon Buri, Thailand for supporting the project with crocodile blood samples.

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Affiliations

  1. Department of Biochemistry, Faculty of Science, Khon Kaen University, Khon Kaen, 40002, Thailand

    Tinnakorn Theansungnoen, Nualyai Yaraksa, Sakda Daduang & Sompong Thammasirirak

  2. Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Faculty of Science, Khon Kaen University, Khon Kaen, 40002, Thailand

    Tinnakorn Theansungnoen, Nualyai Yaraksa, Sakda Daduang & Sompong Thammasirirak

  3. Faculty of Humanities and Social Sciences, Khon Kaen University, Khon Kaen, 40002, Thailand

    Apisak Dhiravisit

Authors
  1. Tinnakorn Theansungnoen
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  2. Nualyai Yaraksa
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  3. Sakda Daduang
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  4. Apisak Dhiravisit
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  5. Sompong Thammasirirak
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Correspondence to Sompong Thammasirirak.

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Theansungnoen, T., Yaraksa, N., Daduang, S. et al. Purification and Characterization of Antioxidant Peptides from Leukocyte Extract of Crocodylus siamensis . Protein J 33, 24–31 (2014). https://doi.org/10.1007/s10930-013-9536-8

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  • DOI: https://doi.org/10.1007/s10930-013-9536-8

Keywords

  • Antioxidant
  • DPPH assay
  • White blood cell
  • Crocodile blood
  • Crocodile