Metabolic stimulation of phenolic biosynthesis and antioxidant enzyme response in dark germinated barley (Hordeum vulgare L.) sprouts using bioprocessed elicitors

  • Ramnarain Ramakrishna
  • Dipayan Sarkar
  • Kalidas ShettyEmail author


Sprouting and seed elicitor treatments stimulate the biosynthesis of health relevant phenolic bioactives in plants partly by upregulating proline-associated pentose phosphate pathway (PAPPP). This study investigated the upregulation of PAPPP-linked and antioxidant enzyme associated metabolic responses in elicitor-treated barley (Hordeum vulgare L.) sprouts previously established with stimulation of health relevant phenolic bioactives. Barley seeds were treated with bioprocessed elicitors marine protein hydrolysates (GroPro®, GP) and soluble chitosan oligosaccharide and germinated under dark conditions. Upregulation of PAPPP and subsequent stimulation of phenolic biosynthesis and antioxidant enzyme responses were monitored at day 2, 4, and 6 of sprouting. High PAPPP-linked antioxidant enzyme responses were observed at early stages of germination with selected doses of GP treatments, especially in cv. Pinnacle. Total soluble phenolic content remained at higher level, while guaiacol peroxidase activity increased over the course of sprouting indicating increased phenolic polymerization to support structural needs of sprouts.


Antioxidant enzymes Elicitors Pentose phosphate pathway Phenolics Proline 



We thank Dr. Richard Horsley, the coordinator of the North Dakota Malting Barley Improvement Program at North Dakota State University, Fargo, ND, USA, for providing malting barley seeds for this experiment.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.


  1. Andarwulan N, Shetty K. Improvement of pea (Pisum sativum) seed vigour response by fish protein hydrolysates in combination with acetyl salicylic acid. Process Biochem. 35: 159–165 (1999)CrossRefGoogle Scholar
  2. Baik BK, Ullrich SE. Barley for food: characteristics, improvement, and renewed interest. J. Cereal Sci. 48: 233–242 (2008)CrossRefGoogle Scholar
  3. Beers RF, Sizer IW. A spectrophotometric method for measuring the breakdown of hydrogen peroxide by catalase. J. Biol. Chem. 195: 133–140 (1952)PubMedGoogle Scholar
  4. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254 (1976)CrossRefGoogle Scholar
  5. Bregman AA. Laboratory investigations in cell biology. 2nd ed. Wiley, Hoboken, NJ, USA (1987)Google Scholar
  6. Costilow RN, Cooper D. Identity of proline dehydrogenase and delta 1-pyrroline-5-carboxylic acid reductase in Clostridium sporogenes. J. Bacteriol. 134: 139–146 (1978)PubMedPubMedCentralGoogle Scholar
  7. Crozier A, Clifford MN, Ashihara H. Plant secondary metabolites: occurrence, structure and role in the human diet. Wiley, Hoboken, NJ, USA (2008)Google Scholar
  8. Deutsch J. Methods of enzymatic analysis. 3rd ed. Verlag Chemie Academic Press, Deerfield Beach, FL, USA (1983)Google Scholar
  9. Hare PD, Cress WA. Metabolic implications of stress-induced proline accumulation in plants. Plant Growth Regul. 21: 79–102 (1997)CrossRefGoogle Scholar
  10. Huang S, Millar HA. Succinate dehydrogenase: the complex roles of a simple enzyme. Curr. Opin. Plant Biol. 16: 344–349 (2013)CrossRefGoogle Scholar
  11. Idehen E, Tang Y, Sang S. Bioactive phytochemicals in barley. J. Food Drug Anal. 25: 148–161 (2017)CrossRefGoogle Scholar
  12. Laloue H, Weber-Lofti F, Lucau-Danila A, Guillemaut P. Identification of ascorbate and guaiacol peroxidase in needle chloroplasts of spruce trees. Plant Physiol. Biochem. 35: 341–346 (1997)Google Scholar
  13. McCue P, Shetty K. Clonal herbal extracts as elicitors of phenolic synthesis in dark-germinated mung beans for improving nutritional value with implications for food safety. J. Food Biochem. 26: 209–232 (2002)CrossRefGoogle Scholar
  14. Morales M, Barceló AR. A basic peroxidase isoenzyme from vacuoles and cell walls of Vitis vinifera. Phytochemistry. 45: 229–232 (1997)CrossRefGoogle Scholar
  15. Naczk M, Shahidi F. Extraction and analysis of phenolics in food. J. Chromatogr. A. 1054: 95–111 (2004)CrossRefGoogle Scholar
  16. Oberley LW, Spitz DR. Assay of SOD activity in tumor tissue. Method Enzymol. 105: 457–461 (1984)CrossRefGoogle Scholar
  17. Orwat J (2016) Phenolic antioxidant-linked bioactive enrichment in black beans (Phaseolus vulgaris L.) to screen for health benefits and enhancement of salinity resilience. M.S. thesis, North Dakota State University, Fargo, USA (2016)Google Scholar
  18. Ramakrishna R, Sarkar D, Manduri A, Iyer SG, Shetty K. Improving phenolic bioactive-linked anti-hyperglycemic functions of dark germinated barley sprouts (Hordeum vulgare L.) using seed elicitation strategy. J. Food Sci. Technol. Mysore. 54: 1–13 (2017)CrossRefGoogle Scholar
  19. Randhir R, Shetty K. Light-mediated fava bean (Vicia faba) response to phytochemical and protein elicitors and consequences on nutraceutical enhancement and seed vigour. Process Biochem. 38: 945–952 (2003)CrossRefGoogle Scholar
  20. Randhir R, Shetty K. Developmental stimulation of total phenolics and related antioxidant activity in light- and dark-germinated corn by natural elicitors. Process Biochem. 40: 1721–1732 (2005)CrossRefGoogle Scholar
  21. Randhir R, Shetty P, Shetty K. L-DOPA and total phenolic stimulation in dark germinated fava bean in response to peptide and phytochemical elicitors. Process Biochem. 37: 1247–1256 (2002)CrossRefGoogle Scholar
  22. Randhir R, Lin YT, Shetty K. Stimulation of phenolics, antioxidant and antimicrobial activities in dark germinated mung bean sprouts in response to peptide and phytochemical elicitors. Process Biochem. 39: 637–646 (2004)CrossRefGoogle Scholar
  23. Randhir R, Kwon YI, Shetty K. Improved health-relevant functionality in dark germinated Mucuna pruriens sprouts by elicitation with peptide and phytochemical elicitors. Bioresour. Technol. 100: 4507–4514 (2009)CrossRefGoogle Scholar
  24. Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice-Evans C. Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radical BioMed. 26: 1231–1237 (1999)CrossRefGoogle Scholar
  25. Rice-Evans C, Miller N, Paganga G. Antioxidant properties of phenolic compounds. Trends Plant Sci. 2: 152–159 (1997)CrossRefGoogle Scholar
  26. Sarkar D, Shetty K. Metabolic stimulation of plant phenolics for food preservation and health. Annu. Rev. Food Sci. Technol. 5: 395–413 (2014)CrossRefGoogle Scholar
  27. Sarkar D, Bhowmik PC, In-Kwon Y, Shetty K. Cold acclimation responses of three cool-season turfgrasses and the role of proline-associated pentose phosphate pathway. J. Am. Soc. Hortic. Sci. 134: 210–220 (2009)Google Scholar
  28. Sarkar D, Bhowmik PC, In-Kwon Y, Shetty, K. The role of proline-associated pentose phosphate pathway in cool-season turfgrasses after UV-B exposure. Environ. Exp. Bot. 70: 251–258 (2011)CrossRefGoogle Scholar
  29. Shetty K. Biotechnology to harness the benefits of dietary phenolics: focus on Lamiaceae. Asia Pac. J. Clin. Nutr. 6: 162–171 (1997)PubMedGoogle Scholar
  30. Shetty K. Role of proline-linked pentose phosphate pathway in biosynthesis of plant phenolics for functional food and environmental applications: a review. Process Biochem. 39: 789–804 (2004)CrossRefGoogle Scholar
  31. Shetty K, McCue P. Phenolic antioxidant biosynthesis in plants for functional food application: integration of systems biology and biotechnological approaches. Food Biotechnol. 17: 67–97 (2003)CrossRefGoogle Scholar
  32. Shetty K, Wahlqvist M. A model for the role of the proline-linked pentose-phosphate pathway in phenolic phytochemical bio-synthesis and mechanism of action for human health and environmental applications. Asia Pac. J. Clin. Nutr. 13: 1–24 (2004)PubMedGoogle Scholar
  33. Shetty K, Curtis OF, Levin RE, Witkowsky R, Ang W. Prevention of vitrification associated with in vitro shoot culture of oregano (Origanum vulgare) by Pseudomonas spp. J. Plant Physiol. 147: 447–451 (1995)CrossRefGoogle Scholar
  34. Singh A, Sharma S. Bioactive components and functional properties of biologically activated cereal grains: a bibliographic review. Crit. Rev. Food Sci. 57: 3051–3071 (2017)CrossRefGoogle Scholar

Copyright information

© The Korean Society of Food Science and Technology 2018

Authors and Affiliations

  1. 1.Department of Plant SciencesNorth Dakota State UniversityFargoUSA

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