Ukr.Biochem.J. 2018; Volume 90, Issue 6, Nov-Dec, pp. 12-20


Phenylalanine ammonia-lyase activity and content of flavonoid compounds in wheat seedlings at the action of hypothermia and hydrogen sulfide donor

Yu. E. Kolupaev1,2, E. I. Horielova1, T. O. Yastreb1, Yu. V. Popov3, N. I. Ryabchun3

1Dokuchaev Kharkiv National Agrarian University, Ukraine;
2Karazin Kharkiv National University, Ukraine;
3Yuryev Рlant Production Institute, National Academy of Agrarian Sciences of Ukraine, Kharkiv

At present hydrogen sulfide (H2S) is considered as one of the signal mediators in plant cells. However, its role in formation of plant resistance to low temperatures and, in particular, in regulation of secondary metabolism under stress conditions remains poorly understood. The influence of H2S donor sodium hydrosulfide (NaHS) on phenylalanine ammonia-lyase (PAL) activity and content of flavonoids in wheat seedlings at normal temperature (21 °C) and under cold hardening conditions (7 days at 3 °C) was studied. After 2 days of the hardening temperature, a transient increase in PAL activity was noted. Also, activity of the enzyme was increased by treatment of plants with 0.1 or 0.5 mM NaHS under normal temperature conditions and especially at the background of cold hardening. By themselves, the cold hardening and the action of H2S donor caused an increase in total content of flavonoids and amount of anthocyanins. With the combination of hypothermia and treatment of seedlings with NaHS, this effect enlarged and the total content of flavonoids increased by 3.8, and anthocyanins increased by 1.8 times in comparison to the control. Treatment with the H2S donor caused a decrease in content of the lipid peroxidation product malonic dialdehyde in seedlings after the action of hardening temperature, and especially after their freezing at –5 °C. Also, under the influence of NaHS, survival of hardened and unhardened seedlings after cryostress increased. It was concluded that one of the mechanisms of the positive influence of the H2S donor on resistance of wheat seedlings to hypothermia is the PAL-dependent accumulation of flavonoid compounds, which have a high antioxidant activity, and a decrease in effects of secondary oxidative stress.

Keywords: , , , , ,


  1. Łowicka E, Bełtowski J. Hydrogen sulfide (H2S) – the third gas of interest for pharmacologists. Pharmacol Rep. 2007 Jan-Feb;59(1):4-24. PubMed
  2. Gadalla MM, Snyder SH. Hydrogen sulfide as a gasotransmitter. J Neurochem. 2010 Apr;113(1):14-26. PubMed, PubMedCentral, CrossRef
  3. Zaichko NV, Melnik AV, Yoltukhivskyy MM, Olhovskiy AS, Palamarchuk IV. Hydrogen sulfide: metabolism, biological and medical role. Ukr Biochem J. 2014 Sep-Oct;86(5):5-25. PubMed, CrossRef
  4. He H, He LF. Regulation of gaseous signaling molecules on proline metabolism in plants. Plant Cell Rep. 2018 Mar;37(3):387-392. PubMed, CrossRef
  5. Hancock JT, Whiteman M. Hydrogen sulfide signaling: interactions with nitric oxide and reactive oxygen species. Ann N Y Acad Sci. 2016 Feb;1365(1):5-14. PubMed, CrossRef
  6. Jin Z, Xue S, Luo Y, Tian B, Fang H, Li H, Pei Y. Hydrogen sulfide interacting with abscisic acid in stomatal regulation responses to drought stress in Arabidopsis. Plant Physiol Biochem. 2013 Jan;62:41-6. PubMed, CrossRef
  7. Lai D, Mao Y, Zhou H, Li F, Wu M, Zhang J, He Z, Cui W, Xie Y. Endogenous hydrogen sulfide enhances salt tolerance by coupling the reestablishment of redox homeostasis and preventing salt-induced K⁺ loss in seedlings of Medicago sativa. Plant Sci. 2014 Aug;225:117-29. PubMed, CrossRef
  8. Wang Y, Li L, Cui W, Xu S., Shen W, Wang R. Hydrogen sulfide enhances alfalfa (Medicago sativa) tolerance against salinity during seed germination by nitric oxide pathway. Plant Soil. 2012; 351(1-2): 107-119. CrossRef
  9. Li ZG. Hydrogen sulfide: a multifunctional gaseous molecule in plants. Russ J Plant Physiol. 2013; 60(6): 733-740. CrossRef
  10. Li ZG, Yi XY, Li YT. Effect of pretreatment with hydrogen sulfide donor sodium hydrosulfide on heat tolerance in relation to antioxidant system in maize (Zea mays) seedlings. Biologia. 2014; 69(8): 1001-1009. CrossRef
  11. Ding H, Han Q, Ma D, Hou J, Huang X, Wang C, Xie Y, Kang G, Guo T. Characterizing physiological and proteomic analysis of the action of H2S to mitigate drought stress in young seedling of wheat. Plant Mol Biol Rep. 2018; 36(1): 45-57. CrossRef
  12. Shi H, Ye T, Chan Z. Exogenous application of hydrogen sulfide donor sodium hydrosulfide enhanced multiple abiotic stress tolerance in bermudagrass (Cynodon dactylon (L). Pers.). Plant Physiol Biochem. 2013 Oct;71:226-34. PubMed, CrossRef
  13. Piotrovskii MS, Shevyreva TA, Zhestkova IM, Trofimova MS. Activation of plasmalemmal NADPH oxidase in etiolated maize seedlings exposed to chilling temperatures. Russ J Plant Physiol. 2011; 58(2): 290-298.  CrossRef
  14. Kolupaev YuE, Karpets YuV. Reactive oxygen species and stress signaling in plants. Ukr Biochem J. 2014 Jul-Aug;86(4):18-35. (In Russian). PubMed, CrossRef
  15. Janmohammadi M, Enayati V, Sabaghnia N. Impact of cold acclimation, de-acclimation and re-acclimation on carbohydrate content and antioxidant enzyme activities in spring and winter wheat. Icel Agric Sci. 2012. 25; 3-11.
  16. Christou A, Manganaris GA, Papadopoulos I, Fotopoulos V. Hydrogen sulfide induces systemic tolerance to salinity and non-ionic osmotic stress in strawberry plants through modification of reactive species biosynthesis and transcriptional regulation of multiple defence pathways. J Exp Bot. 2013 Apr;64(7):1953-66. PubMed, PubMedCentral, CrossRef
  17. Li SP, Hu KD, Hu LY, Li YH, Jiang AM, Xiao F, Han Y, Liu YS, Zhang H. Hydrogen sulfide alleviates postharvest senescence of broccoli by modulating antioxidant defense and senescence-related gene expression. J Agric Food Chem. 2014 Feb 5;62(5):1119-29. PubMed, CrossRef
  18. Kolupaev YuE, Fіrsova EN, Yastreb TO. Induction of plant cells heat resistance by hydrogen sulfide donor is mediated by H2O2 generation with participation of NADPH oxidase and superoxide dismutase. Ukr Biochem J. 2017; 89(4): 34-42.  CrossRef
  19. Khlestkina EK. The adaptive role of flavonoids: emphasis on cereals. Cereal Res Commun. 2013; 41(2): 185-198. CrossRef
  20. Neill SO, Gould KS. Anthocyanins in leaves: light attenuators or antioxidants? Funct Plant Biol. 2003; 30(8): 865-873.  CrossRef
  21. Olenichenko NA, Zagoskina NV, Astakhova NV, Trunova TI, Kuznetsov YuV. Primary and secondary metabolism of winter wheat under cold hardening and treatment with antioxidants. Appl Biochem Microbiol. 2008; 44(5): 535-540. PubMed, CrossRef
  22. Kolupaev YuE, Yastreb TO, Oboznyi AI, Ryabchun NI, Kirichenko VV. Constitutive and cold-induced resistance of rye and wheat seedlings to oxidative stress. Russ J Plant Physiol. 2016; 63(3): 326-337.  CrossRef
  23. Luo Z, Li D, Du R, Mou W. Hydrogen sulfide alleviates chilling injury of banana fruit by enhanced antioxidant system and proline content. Sci Horticult. 2015; 183: 144-151.  CrossRef
  24. Adamovskaya VG, Molodchenkova OO, Ciselskaya LY, Bezkrovnaya LA, Levitsky YuA. Peculiarities of phenylalanine ammonia-lyase, phenolic compounds and lignin accumulation under the action of Fusarium and salicylic acid in the seedlings of cereal crops. Fiziol Biokhim Kult Rast. 2007; 39(4): 353-361. (In Russian).
  25. Li ZG, Xie LR, Li XJ. Hydrogen sulfide acts as a downstream signal molecule in salicylic acid-induced heat tolerance in maize (Zea mays L.) seedlings. J Plant Physiol. 2015 Apr 1;177:121-127.  PubMed, CrossRef
  26. Kolupaev YuE, Ryabchun NI, Vayner AA, Yastreb TO, Oboznyi AI. Antioxidant enzyme activity and osmolyte content in winter cereal seedlings under hardening and cryostress. Russ J Plant Physiol. 2015; 62(4): 499-506.  CrossRef
  27. Zucker M. Induction of phenylalanine ammonia-lyase in Xanthium leaf disks. Photosynthetic requirement and effect of day length. Plant Physiol. 1969; 44(6): 912-922. CrossRef
  28. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72(1-2):248-54. PubMed, CrossRef
  29. Bulatov АV, Falkova МТ, Pushina МО, Moskvin LN, Alekseeva GM. Spectrophotometric determination of flavonoids in plant material. Analitika i Kontrol. 2012; 16(4): 358-362. (In Russian).
  30. Harvaux M, Kloppstech K. The protective functions of carotenoid and flavonoid pigments against excess visible radiation at chilling temperature investigated in Arabidopsis npq and tt mutants. Planta. 2001 Oct;213(6):953-66. PubMed, CrossRef
  31. Kolupaev YuE, Firsova KM, Shvidenko MV, Yastreb TO. Hydrogen sulfide donor influence on state of antioxidant system of wheat seedlings under osmotic stress. Fiziol Rast i Genetika. 2018; 50(1): 29-38. (In Ukrainian).
  32. Li Q, Wang Z, Zhao Y, Zhang X, Zhang S, Bo L, Wang Y, Ding Y, An L. Putrescine protects hulless barley from damage due to UV-B stress via H2S- and H2O2-mediated signaling pathways. Plant Cell Rep. 2016 May;35(5):1155-68. PubMed, CrossRef
  33. Aghdama MS, Mahmoudi R, Razavi F, Rabiei V, Soleimani A. Hydrogen sulfide treatment confers chilling tolerance in hawthorn fruit during cold storage by triggering endogenous H2S accumulation, enhancing antioxidant enzymes activity and promoting phenols accumulation. Sci Horticult. 2018; 238: 264-271. CrossRef
  34. Sun Y, Zhang W, Zeng T, Nie Q, Zhang F, Zhu L. Hydrogen sulfide inhibits enzymatic browning of fresh-cut lotus root slices by regulating phenolic metabolism. Food Chem. 2015 Jun 15;177:376-81. PubMed, CrossRef
  35. Liang X, Zhang L, Natarajan SK, Becker DF. Proline mechanisms of stress survival. Antioxid Redox Signal. 2013 Sep 20;19(9):998-1011. PubMed, PubMedCentral, CrossRef
  36. Hancock JT, Whiteman M. Hydrogen sulfide and cell signaling: team player or referee? Plant Physiol Biochem. 2014 May;78:37-42. PubMed, CrossRef

Creative CommonsThis work is licensed under a Creative Commons Attribution 4.0 International License.